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Potentials and limitations of molecular diagnostic methods in food safety
Mariani, Paola O.
Molecular methods allow the detection of pathogen nucleic acids (DNA and RNA) and, therefore, the detection of contamination in food is carried out with high selectivity and rapidity. In the last 2 decades molecular methods have accompanied traditional diagnostic methods in routine pathogen detection, and might replace them in the upcoming future. In this review the implementation in diagnostics of four of the most used molecular techniques (PCR, NASBA, microarray, LDR) are described and compared, highlighting advantages and limitations of each of them. Drawbacks of molecular methods with regard to traditional ones and the difficulties encountered in pathogen detection from food or clinical specimen are also discussed. Moreover, criteria for the choice of the target sequence for a secure detection and classification of pathogens and possible developments in molecular diagnostics are also proposed. PMID:19067016
Molecular Diagnostic Methods for Detection and Characterization of Human Noroviruses.
Chen, Haifeng; Hu, Yuan
Human noroviruses are a group of viral agents that afflict people of all age groups. The viruses are now recognized as the most common causative agent of nonbacterial acute gastroenteritis and foodborne viral illness worldwide. However, they have been considered to play insignificant roles in the disease burden of acute gastroenteritis for the past decades until the recent advent of new and more sensitive molecular diagnostic methods. The availability and application of the molecular diagnostic methods have led to enhanced detection of noroviruses in clinical, food and environmental samples, significantly increasing the recognition of noroviruses as an etiologic agent of epidemic and sporadic acute gastroenteritis. This article aims to summarize recent efforts made for the development of molecular methods for the detection and characterization of human noroviruses. PMID:27335620
Molecular Diagnostic Methods for Detection and Characterization of Human Noroviruses
Chen, Haifeng; Hu, Yuan
Human noroviruses are a group of viral agents that afflict people of all age groups. The viruses are now recognized as the most common causative agent of nonbacterial acute gastroenteritis and foodborne viral illness worldwide. However, they have been considered to play insignificant roles in the disease burden of acute gastroenteritis for the past decades until the recent advent of new and more sensitive molecular diagnostic methods. The availability and application of the molecular diagnostic methods have led to enhanced detection of noroviruses in clinical, food and environmental samples, significantly increasing the recognition of noroviruses as an etiologic agent of epidemic and sporadic acute gastroenteritis. This article aims to summarize recent efforts made for the development of molecular methods for the detection and characterization of human noroviruses. PMID:27335620
Molecular and Nonmolecular Diagnostic Methods for Invasive Fungal Infections
Arvanitis, Marios; Anagnostou, Theodora; Fuchs, Beth Burgwyn; Caliendo, Angela M.
SUMMARY Invasive fungal infections constitute a serious threat to an ever-growing population of immunocompromised individuals and other individuals at risk. Traditional diagnostic methods, such as histopathology and culture, which are still considered the gold standards, have low sensitivity, which underscores the need for the development of new means of detecting fungal infectious agents. Indeed, novel serologic and molecular techniques have been developed and are currently under clinical evaluation. Tests like the galactomannan antigen test for aspergillosis and the β-glucan test for invasive Candida spp. and molds, as well as other antigen and antibody tests, for Cryptococcus spp., Pneumocystis spp., and dimorphic fungi, have already been established as important diagnostic approaches and are implemented in routine clinical practice. On the other hand, PCR and other molecular approaches, such as matrix-assisted laser desorption ionization (MALDI) and fluorescence in situ hybridization (FISH), have proved promising in clinical trials but still need to undergo standardization before their clinical use can become widespread. The purpose of this review is to highlight the different diagnostic approaches that are currently utilized or under development for invasive fungal infections and to identify their performance characteristics and the challenges associated with their use. PMID:24982319
Molecular Beacons in Diagnostics
Kramer, Fred Russell
Recent technical advances have begun to realize the potential of molecular beacons to test for diverse infections in clinical diagnostic laboratories. These include the ability to test for, and quantify, multiple pathogens in the same clinical sample, and to detect antibiotic resistant strains within hours. The design principles of molecular beacons have also spawned a variety of allied technologies. PMID:22619695
Combined Use of Cytogenetic and Molecular Methods in Prenatal Diagnostics of Chromosomal Abnormalities
Stomornjak-Vukadin, Meliha; Kurtovic-Basic, Ilvana; Mehinovic, Lejla; Konjhodzic, Rijad
Aim: The aim of prenatal diagnostics is to provide information of the genetic abnormalities of the fetus early enough for the termination of pregnancy to be possible. Chromosomal abnormalities can be detected in an unborn child through the use of cytogenetic, molecular- cytogenetic and molecular methods. In between them, central spot is still occupied by cytogenetic methods. In cases where use of such methods is not informative enough, one or more molecular cytogenetic methods can be used for further clarification. Combined use of the mentioned methods improves the quality of the final findings in the diagnostics of chromosomal abnormalities, with classical cytogenetic methods still occupying the central spot. Material and methods: Conducted research represent retrospective-prospective study of a four year period, from 2008 through 2011. In the period stated, 1319 karyotyping from amniotic fluid were conducted, along with 146 FISH analysis. Results: Karyotyping had detected 20 numerical and 18 structural aberrations in that period. Most common observed numerical aberration were Down syndrome (75%), Klinefelter syndrome (10%), Edwards syndrome, double Y syndrome and triploidy (5% each). Within observed structural aberrations more common were balanced chromosomal aberrations then non balanced ones. Most common balanced structural aberrations were as follows: reciprocal translocations (60%), Robertson translocations (13.3%), chromosomal inversions, duplications and balanced de novo chromosomal rearrangements (6.6% each). Conclusion: With non- balanced aberrations observed in the samples of amniotic fluid, non- balanced translocations, deletions and derived chromosomes were equally represented. Number of detected aneuploidies with FISH, prior to obtaining results with karyotyping, were 6. PMID:26005269
Molecular diagnostics of neurodegenerative disorders
Agrawal, Megha; Biswas, Abhijit
Molecular diagnostics provide a powerful method to detect and diagnose various neurological diseases such as Alzheimer's and Parkinson's disease. The confirmation of such diagnosis allows early detection and subsequent medical counseling that help specific patients to undergo clinically important drug trials. This provides a medical pathway to have better insight of neurogenesis and eventual cure of the neurodegenerative diseases. In this short review, we present recent advances in molecular diagnostics especially biomarkers and imaging spectroscopy for neurological diseases. We describe advances made in Alzheimer's disease (AD), Parkinson's disease (PD), Amyotrophic lateral sclerosis (ALS) and Huntington's disease (HD), and finally present a perspective on the future directions to provide a framework for further developments and refinements of molecular diagnostics to combat neurodegenerative disorders. PMID:26442283
Molecular diagnostics for tuberculosis.
Noor, K M; Shephard, L; Bastian, I
The phenotypic methods of smear microscopy, culture and indirect drug susceptibility testing (DST) remain the 'gold standard' diagnostics for tuberculosis (TB) in 2015. However, this review demonstrates that genotypic methods are in the ascendancy. Current-generation nucleic acid amplification tests (NAATs) are important supplementary tests for the rapid direct detection of (multidrug-resistant) TB in specific clinical settings. Genotypic detection is already the preferred method of detecting rifampicin and pyrazinamide resistance. Next-generation NAATs able to detect about 10 colony forming units/mL of sputum could replace culture as the initial test for detecting TB. Whole genome sequencing could also plausibly replace phenotypic DST but much work is required in method standardisation, database development and elucidation of all resistance gene determinants. The challenge then will be to rollout these increasingly complex and expensive diagnostics in the low-income countries where TB is prevalent. PMID:25719854
Molecular diagnostics and parasitic disease.
Vasoo, Shawn; Pritt, Bobbi S
Molecular parasitology represents an emerging field in microbiology diagnostics. Although most assays use nonstandardized, laboratory-developed methods, a few commercial systems have recently become available and are slowly being introduced into larger laboratories. In addition, a few methodologies show promise for use in field settings in which parasitic infections are endemic. This article reviews the available techniques and their applications to major parasitic diseases such as malaria, leishmaniasis, and trichomoniasis. PMID:23931835
Methods for improved selectivity in photo-activation and detection of molecular diagnostic agents
Wachter, Eric A.; Fisher, Walter G.; Dees, H. Craig
A method for the imaging of a particular volume of plant or animal tissue, wherein the plant or animal tissue contains at least one photo-active molecular agent. The method comprises the steps of treating the particular volume of the plant or animal tissue with light sufficient to promote a simultaneous two-photon excitation of the photo-active molecular agent contained in the particular volume of the plant or animal tissue, photo-activating at least one of the at least one photo-active molecular agent in the particular volume of the plant or animal tissue, thereby producing at least one photo-activated molecular agent, wherein the at least one photo-activated molecular agent emits energy, detecting the energy emitted by the at least one photo-activated molecular agent, and producing a detected energy signal which is characteristic of the particular volume of plant or animal tissue. The present invention also provides a method for the imaging of a particular volume of material, wherein the material contains at least one photo-active molecular agent.
Method for improved selectivity in photo-activation and detection of molecular diagnostic agents
Wachter, E.A.; Fisher, W.G.; Dees, H.C.
A method for the imaging of a particular volume of plant or animal tissue, wherein the plant or animal tissue contains at least one photo-active molecular agent. The method includes the steps of treating the particular volume of the plant or animal tissue with light sufficient to promote a simultaneous two-photon excitation of the photo-active molecular agent contained in the particular volume of the plant or animal tissue, photo-activating at least one of the at least one photo-active molecular agent in the particular volume of the plant or animal tissue, thereby producing at least one photo-activated molecular agent, wherein the at least one photo-activated molecular agent emits energy, detecting the energy emitted by the at least one photo-activated molecular agent, and producing a detected energy signal which is characteristic of the particular volume of plant or animal tissue. The present invention is also a method for the imaging of a particular volume of material, wherein the material contains at least one photo-active molecular agent. 13 figs.
Method for improved selectivity in photo-activation and detection of molecular diagnostic agents
Wachter, Eric A.; Fisher, Walter G.; Dees, H. Craig
A method for the imaging of a particular volume of plant or animal tissue, wherein the plant or animal tissue contains at least one photo-active molecular agent. The method includes the steps of treating the particular volume of the plant or animal tissue with light sufficient to promote a simultaneous two-photon excitation of the photo-active molecular agent contained in the particular volume of the plant or animal tissue, photo-activating at least one of the at least one photo-active molecular agent in the particular volume of the plant or animal tissue, thereby producing at least one photo-activated molecular agent, wherein the at least one photo-activated molecular agent emits energy, detecting the energy emitted by the at least one photo-activated molecular agent, and producing a detected energy signal which is characteristic of the particular volume of plant or animal tissue. The present invention is also a method for the imaging of a particular volume of material, wherein the material contains at least one photo-active molecular agent.
Molecular IR Spectroscopy: New Trends and Methods of Noninvasive Diagnostics of Tissue IN VIVO
NASA Astrophysics Data System (ADS)
Afanasyeva, Natalia; Bruch, Reinhard
Fiberoptic evanescent wave Fourier transform infrared (FEW-FTIR) spectroscopy using fiberoptic sensors operated in the attenuated total reflection (ATR) regime in the middle infrared (IR) region of the spectrum (850-1850 cm-1) has recently been applied to the diagnostics of tissues. The method is suitable for noninvasive and rapid (seconds) direct measurements of the spectra of normal and pathological tissues in vitro, ex vivo and in vivo. The aim of our studies is the express testing of various tumor tissues at the early stages of their development. The method is expected to be further developed for endoscopic and biopsy applications. We measured the normal skin and malignant tissues in vivo on the surface (directly on patients) in various cases of basaloma, melanoma and nevus. The experiments were performed in the operating room to measure the skin in the depth (under/in the layers of epidermis) of human breast, stomach, lung, and kidney tissues. The breast and skin tissues at different stages of tumor or cancer were distinguished very clearly in spectra of amide, side cyclic and noncyclic hydrogen bonded fragments of aminoacid residuals, phosphate groups and sugars. Computer monitoring is being developed for diagnostics.
[Molecular diagnostic methods of respiratory infections. Has the scheme diagnosis changed?].
Vila Estapé, Jordi; Zboromyrska, Yuliya; Vergara Gómez, Andrea; Alejo Cancho, Izaskun; Rubio García, Elisa; Álvarez-Martínez, Miriam José; la Bellacasa Brugada, Jorge Puig de; Marcos Maeso, M Ángeles
Lower respiratory tract infections remain one of the most common causes of mortality worldwide, which is why early diagnosis is crucial. Traditionally the microbiological diagnosis of these infections has been based on conventional methods including culture on artificial media for isolation of bacteria and fungi and cell cultures for virus and antibody or antigen detection using antigen-antibody reactions. The main drawback of the above mentioned methods is the time needed for an etiological diagnosis of the infection. The techniques based on molecular biology have drawn much attention in recent decades as tools for rapid diagnosis of infections. Some techniques are very expensive, especially those that can detect various microorganisms in the same reaction, therefore the question that arises is whether the cost of such testing is justified by the information obtained and by the clinical impact that its implementation will determine. In this article we make a review of the various techniques of molecular biology applied to the diagnosis of pneumonia and focus primarily on analysing the impact they may have on the management of patients with acute respiratory tract infections. PMID:27474246
Molecular diagnostic tests for microsporidia.
Ghosh, Kaya; Weiss, Louis M
The Microsporidia are a ubiquitous group of eukaryotic obligate intracellular parasites which were recognized over 100 years ago with the description of Nosema bombycis, a parasite of silkworms. It is now appreciated that these organisms are related to the Fungi. Microsporidia infect all major animal groups most often as gastrointestinal pathogens; however they have been reported from every tissue and organ, and their spores are common in environmental sources such as ditch water. Several different genera of these organisms infect humans, but the majority of infections are due to either Enterocytozoon bieneusi or Encephalitozoon species. These pathogens can be difficult to diagnose, but significant progress has been made in the last decade in the development of molecular diagnostic reagents for these organisms. This report reviews the molecular diagnostic tests that have been described for the identification of the microsporidia that infect humans. PMID:19657457
Molecular Diagnostic Tests for Microsporidia
Ghosh, Kaya; Weiss, Louis M.
The Microsporidia are a ubiquitous group of eukaryotic obligate intracellular parasites which were recognized over 100 years ago with the description of Nosema bombycis, a parasite of silkworms. It is now appreciated that these organisms are related to the Fungi. Microsporidia infect all major animal groups most often as gastrointestinal pathogens; however they have been reported from every tissue and organ, and their spores are common in environmental sources such as ditch water. Several different genera of these organisms infect humans, but the majority of infections are due to either Enterocytozoon bieneusi or Encephalitozoon species. These pathogens can be difficult to diagnose, but significant progress has been made in the last decade in the development of molecular diagnostic reagents for these organisms. This report reviews the molecular diagnostic tests that have been described for the identification of the microsporidia that infect humans. PMID:19657457
Molecular method for the detection of Andes hantavirus infection: validation for clinical diagnostics.
Vial, Cecilia; Martinez-Valdebenito, Constanza; Rios, Susana; Martinez, Jessica; Vial, Pablo A; Ferres, Marcela; Rivera, Juan C; Perez, Ruth; Valdivieso, Francisca
Hantavirus cardiopulmonary syndrome is a severe disease caused by exposure to New World hantaviruses. Early diagnosis is difficult due to the lack of specific initial symptoms. Antihantavirus antibodies are usually negative until late in the febrile prodrome or the beginning of cardiopulmonary phase, while Andes hantavirus (ANDV) RNA genome can be detected before symptoms onset. We analyzed the effectiveness of quantitative reverse transcription polymerase chain reaction (RT-qPCR) as a diagnostic tool detecting ANDV-Sout genome in peripheral blood cells from 78 confirmed hantavirus patients and 166 negative controls. Our results indicate that RT-qPCR had a low detection limit (~10 copies), with a specificity of 100% and a sensitivity of 94.9%. This suggests the potential for establishing RT-qPCR as the assay of choice for early diagnosis, promoting early effective care of patients, and improving other important aspects of ANDV infection management, such as compliance of biosafety recommendations for health personnel in order to avoid nosocomial transmission. PMID:26508102
Novel readout method for molecular diagnostic assays based on optical measurements of magnetic nanobead dynamics.
Donolato, Marco; Antunes, Paula; Bejhed, Rebecca S; Zardán Gómez de la Torre, Teresa; Østerberg, Frederik W; Strömberg, Mattias; Nilsson, Mats; Strømme, Maria; Svedlindh, Peter; Hansen, Mikkel F; Vavassori, Paolo
We demonstrate detection of DNA coils formed from a Vibrio cholerae DNA target at picomolar concentrations using a novel optomagnetic approach exploiting the dynamic behavior and optical anisotropy of magnetic nanobead (MNB) assemblies. We establish that the complex second harmonic optical transmission spectra of MNB suspensions measured upon application of a weak uniaxial AC magnetic field correlate well with the rotation dynamics of the individual MNBs. Adding a target analyte to the solution leads to the formation of permanent MNB clusters, namely, to the suppression of the dynamic MNB behavior. We prove that the optical transmission spectra are highly sensitive to the formation of permanent MNB clusters and, thereby to the target analyte concentration. As a specific clinically relevant diagnostic case, we detect DNA coils formed via padlock probe recognition and isothermal rolling circle amplification and benchmark against a commercial equipment. The results demonstrate the fast optomagnetic readout of rolling circle products from bacterial DNA utilizing the dynamic properties of MNBs in a miniaturized and low-cost platform requiring only a transparent window in the chip. PMID:25539065
Huntington Disease: Molecular Diagnostics Approach.
Bastepe, Murat; Xin, Winnie
Huntington disease (HD) is caused by expansion of a CAG trinucleotide repeat in the first exon of the Huntingtin (HTT) gene. Molecular testing of Huntington disease for diagnostic confirmation and disease prediction requires detection of the CAG repeat expansion. There are three main types of HD genetic testing: (1) diagnostic testing to confirm or rule out disease, (2) presymptomatic testing to determine whether an at-risk individual inherited the expanded allele, and (3) prenatal testing to determine whether the fetus has inherited the expanded allele. This unit includes protocols that describe the complementary use of polymerase chain reactions (PCR) and Southern blot hybridization to accurately measure the CAG trinucleotide repeat size and interpret the test results. In addition, an indirect linkage analysis that does not reveal the unwanted parental HD status in a prenatal testing will also be discussed. PMID:26439718
Molecular diagnostics of myeloproliferative neoplasms.
Langabeer, Stephen E; Andrikovics, Hajnalka; Asp, Julia; Bellosillo, Beatriz; Carillo, Serge; Haslam, Karl; Kjaer, Lasse; Lippert, Eric; Mansier, Olivier; Oppliger Leibundgut, Elisabeth; Percy, Melanie J; Porret, Naomi; Palmqvist, Lars; Schwarz, Jiri; McMullin, Mary F; Schnittger, Susanne; Pallisgaard, Niels; Hermouet, Sylvie
Since the discovery of the JAK2 V617F mutation in the majority of the myeloproliferative neoplasms (MPN) of polycythemia vera, essential thrombocythemia and primary myelofibrosis ten years ago, further MPN-specific mutational events, notably in JAK2 exon 12, MPL exon 10 and CALR exon 9 have been identified. These discoveries have been rapidly incorporated into evolving molecular diagnostic algorithms. Whilst many of these mutations appear to have prognostic implications, establishing MPN diagnosis is of immediate clinical importance with selection, implementation and the continual evaluation of the appropriate laboratory methodology to achieve this diagnosis similarly vital. The advantages and limitations of these approaches in identifying and quantitating the common MPN-associated mutations are considered herein with particular regard to their clinical utility. The evolution of molecular diagnostic applications and platforms has occurred in parallel with the discovery of MPN-associated mutations, and it therefore appears likely that emerging technologies such as next-generation sequencing and digital PCR will in the future play an increasing role in the molecular diagnosis of MPN. PMID:25951317
Companion Diagnostics and Molecular Imaging.
Puranik, Ameya D; Kulkarni, Harshad R; Baum, Richard P
Companion diagnostics (CDx) is a positive attempt in the direction of improving the drug development process, especially in the field of oncology, with the advent of newer targeted therapies. It helps the oncologist in deciding the choice of treatment for the individual patient. The role of CDx assays has attracted the attention of regulators, and especially the US Food and Drug Administration developed regulatory strategies for CDx and the drug-diagnostic codevelopment project. For an increasing number of cancer patients, the treatment selection will depend on the result generated by a CDx assay, and consequently this type of assay has become critical for the care and safety of the patients. In addition to the assay-based approach, molecular imaging with its ability to image at the genetic and receptor level has made foray into the field of drug development and personalized medicine. We shall review these aspects of CDx, with special focus on molecular imaging and the upcoming concept of Theranostics. PMID:26049701
Fundamentals of quality assessment of molecular amplification methods in clinical diagnostics. International Federation of Clinical Chemistry Scientific Division Committee on Molecular Biology Techniques.
Neumaier, M; Braun, A; Wagener, C
The increasing interest in molecular biology diagnostics is a result of the tremendous gain of scientific knowledge in genetics, made possible especially since the introduction of amplification techniques. High expectations have been placed on genetic testing, and the number of laboratories now using the relevant technology is rapidly increasing--resulting in an obvious need for standardization and definition of laboratory organization. This communication is an effort towards that end. We address aspects that should be considered when structuring a new molecular diagnostic laboratory, and we discuss individual preanalytical and analytical procedures, from sampling to evaluation of assay results. In addition, different means of controlling contamination are discussed. Because the methodology is in constant change, no general standards can be defined. Accordingly, this publication is intended to serve as a recommendation for good laboratory practice and internal quality control and as a guide to troubleshooting, primarily in amplification techniques. PMID:9550553
Molecular diagnostics for low resource settings
NASA Astrophysics Data System (ADS)
Weigl, Bernhard H.
As traditional high quality diagnostic laboratories are not widely available or affordable in developing country health care settings, microfluidics-based point-of-care diagnostics may be able to address the need to perform complex assays in under-resourced areas. Many instrument-based as well as non-instrumented microfluidic prototype diagnostics are currently being developed. In addition to various engineering challenges, the greatest remaining issue is the search for truly low-cost disposable manufacturing methods. Diagnostics for global health, and specifically microfluidics and molecular-based low resource diagnostics, have become a very active research area over the last five years, thanks in part to new funding that became available from the Bill and Melinda Gates Foundation, the National Institutes of Health, and other sources. This has led to a number of interesting prototype devices that are now in advanced development or clinical validation. These devices include disposables and instruments that perform multiplexed PCR-based lab-on-a-chips for enteric, febrile, and vaginal diseases, as well as immunoassays for diseases such as malaria, HIV, and various sexually transmitted diseases. More recently, instrument-free diagnostic disposables based on isothermal nucleic acid amplification have been developed as well. Regardless of platform, however, the search for truly low-cost manufacturing methods that would result in cost of goods per disposable of around US1/unit at volume remains a big challenge. This talk will give an overview over existing platform development efforts as well as present some original research in this area at PATH.
Molecular Diagnostics of β-Thalassemia
Atanasovska, B; Bozhinovski, G; Chakalova, L; Kocheva, S; Karanfilski, O; Plaseska-Karanfiska, D
A high-quality hemoglobinopathy diagnosis is based on the results of a number of tests including assays for molecular identification of causative mutations. We describe the current diagnostic strategy for the identification of β-thalassemias and hemoglobin (Hb) variants at the International Reference Laboratory for Haemoglobinopathies, Research Centre for Genetic Engineering and Biotechnology (RCGEB) "Georgi D. Efremov," Skopje, Republic of Macedonia. Our overall approach and most of the methods we use for detection of mutations are designed for the specific target population. We discuss new technical improvements that have allowed us to substantially reduce the average time necessary for reaching a conclusive diagnosis. PMID:24052746
Next-generation molecular diagnostics.
Aldape, Kenneth; Pfister, Stefan M
The classification of brain tumors is based on the time-honored tradition of histologic examination, coupled with clinicopathologic correlation, and is based on the fundamental importance of microscopic morphologic interpretation. Supplementation by immunohistochemical markers is of substantial value to distinguish related entities and to confirm morphologic impressions. The use of techniques such as fluorescent in situ hybridization (FISH) is also critical in specific situations. However, with these practices, it is clear that the use of state-of-the-art molecular techniques has great promise to add to classification to (1) reduce the subjectivity inherent in interobserver discordance, particularly with specific entities; and (2) elucidate the biologic diversity of entities that are not resolvable by routine methods. In this chapter, we discuss these possibilities, focusing on several tumor types affecting the central nervous system, including diffuse glioma and ependymoma. PMID:26948351
Comparison of molecular diagnostic methods for the detection of Acanthamoeba spp. from clinical specimens submitted for keratitis.
Khairnar, Krishna; Tamber, Gurdip S; Ralevski, Filip; Pillai, Dylan R
Acanthamoeba spp. are responsible for a significant annual number of keratitis (AK) cases leading to vision-threatening disease worldwide. Current methods rely on direct examination of specimens by microscopy and/or culture. The former lacks sensitivity and the latter suffers from a poor turnaround time. We undertook a comparison of all published molecular methods, evaluating performance characteristics such as analytical sensitivity, specificity, limit of detection (LOD), reproducibility, accuracy, and cost of test. The study population comprised 128 patients. Eligible specimens were tested prospectively between April 2007 and May 2010 by microscopy and/or culture. Eleven different specimen types were used including corneal scrapings (51.5%), corneal swab (17.9%), and contact lens material (10.9%). Results of 2 published gel-based polymerase chain reaction (PCR) and 2 published real-time quantitative (Q) PCR methods were compared in a blinded manner to direct microscopic examination and/or culture for the detection of Acanthamoeba in clinical specimens. QPCR (Riviere method) had the highest sensitivity at 89.3%, excellent accuracy using ROC analysis (AUC ∼0.90), lowest LOD down to 0.1 organism per microliter, and superior linear correlation with parasite density (R(2) = 0.9965) when compared with microscopy, culture, and other molecular methods. Phylogenetic analysis using a sequence-based typing method revealed that clinical isolates in this population with AK were genetically distinct from granulomatous amebic encephalitis or environmental isolates. The QPCR method was more expensive ($14.80) than traditional methods such as culture ($2.50) or microscopy ($2.50). However, 13 culture- and microscopy-negative specimens were positive by QPCR during the study period, suggesting that detection using QPCR may result in reduced complications and health care costs associated with misdiagnosed AK. PMID:21658877
Molecular Diagnostics for Soil-Transmitted Helminths
O'Connell, Elise M.; Nutman, Thomas B.
Historically, the diagnosis of soil-transmitted helminths (STHs) (e.g., Strongyloides stercoralis, Trichuris trichiura, Ancylostoma duodenale, Necator americanus, and Ascaris lumbricoides) has relied on often-insensitive microscopy techniques. Over the past several years, there has been an effort to use molecular diagnostics, particularly quantitative polymerase chain reaction (qPCR), to detect intestinal pathogens. While some platforms have been approved by regulatory bodies (e.g., Food and Drug Administration) to detect intestinal bacteria, viruses, and protozoa, there are no approved tests currently available for STH. Although studies comparing qPCR to microscopy methods for STH are imperfect, due in large part to a lack of a sufficient gold standard, they do show a significant increase in sensitivity and specificity of qPCR compared with microscopic techniques. These studies, as well as the advantages and disadvantages of using qPCR for STH diagnosis, are discussed. Guidelines for those designing future studies utilizing qPCR are proposed for optimizing results, as is the proposition for using standardized molecular diagnostics routinely for STH in clinical laboratories and for field-based studies when possible. PMID:27481053
Molecular Diagnostics for Soil-Transmitted Helminths.
O'Connell, Elise M; Nutman, Thomas B
Historically, the diagnosis of soil-transmitted helminths (STHs) (e.g., Strongyloides stercoralis, Trichuris trichiura, Ancylostoma duodenale, Necator americanus, and Ascaris lumbricoides) has relied on often-insensitive microscopy techniques. Over the past several years, there has been an effort to use molecular diagnostics, particularly quantitative polymerase chain reaction (qPCR), to detect intestinal pathogens. While some platforms have been approved by regulatory bodies (e.g., Food and Drug Administration) to detect intestinal bacteria, viruses, and protozoa, there are no approved tests currently available for STH. Although studies comparing qPCR to microscopy methods for STH are imperfect, due in large part to a lack of a sufficient gold standard, they do show a significant increase in sensitivity and specificity of qPCR compared with microscopic techniques. These studies, as well as the advantages and disadvantages of using qPCR for STH diagnosis, are discussed. Guidelines for those designing future studies utilizing qPCR are proposed for optimizing results, as is the proposition for using standardized molecular diagnostics routinely for STH in clinical laboratories and for field-based studies when possible. PMID:27481053
Laboratory Information Systems in Molecular Diagnostics: Why Molecular Diagnostics Data are Different.
Lee, Roy E; Henricks, Walter H; Sirintrapun, Sahussapont J
Molecular diagnostic testing presents new challenges to information management that are yet to be sufficiently addressed by currently available information systems for the molecular laboratory. These challenges relate to unique aspects of molecular genetic testing: molecular test ordering, informed consent issues, diverse specimen types that encompass the full breadth of specimens handled by traditional anatomic and clinical pathology information systems, data structures and data elements specific to molecular testing, varied testing workflows and protocols, diverse instrument outputs, unique needs and requirements of molecular test reporting, and nuances related to the dissemination of molecular pathology test reports. By satisfactorily addressing these needs in molecular test data management, a laboratory information system designed for the unique needs of molecular diagnostics presents a compelling reason to migrate away from the current paper and spreadsheet information management that many molecular laboratories currently use. This paper reviews the issues and challenges of information management in the molecular diagnostics laboratory. PMID:26849819
Molecular Diagnostic Applications in Colorectal Cancer
Huth, Laura; Jäkel, Jörg; Dahl, Edgar
Colorectal cancer, a clinically diverse disease, is a leading cause of cancer-related death worldwide. Application of novel molecular diagnostic tests, which are summarized in this article, may lead to an improved survival of colorectal cancer patients. Distinction of these applications is based on the different molecular principles found in colorectal cancer (CRC). Strategies for molecular analysis of single genes (as KRAS or TP53) as well as microarray based techniques are discussed. Moreover, in addition to the fecal occult blood testing (FOBT) and colonoscopy some novel assays offer approaches for early detection of colorectal cancer like the multitarget stool DNA test or the blood-based Septin 9 DNA methylation test. Liquid biopsy analysis may also exhibit great diagnostic potential in CRC for monitoring developing resistance to treatment. These new diagnostic tools and the definition of molecular biomarkers in CRC will improve early detection and targeted therapy of colorectal cancer.
Modeling Complex Workflow in Molecular Diagnostics
Gomah, Mohamed E.; Turley, James P.; Lu, Huimin; Jones, Dan
One of the hurdles to achieving personalized medicine has been implementing the laboratory processes for performing and reporting complex molecular tests. The rapidly changing test rosters and complex analysis platforms in molecular diagnostics have meant that many clinical laboratories still use labor-intensive manual processing and testing without the level of automation seen in high-volume chemistry and hematology testing. We provide here a discussion of design requirements and the results of implementation of a suite of lab management tools that incorporate the many elements required for use of molecular diagnostics in personalized medicine, particularly in cancer. These applications provide the functionality required for sample accessioning and tracking, material generation, and testing that are particular to the evolving needs of individualized molecular diagnostics. On implementation, the applications described here resulted in improvements in the turn-around time for reporting of more complex molecular test sets, and significant changes in the workflow. Therefore, careful mapping of workflow can permit design of software applications that simplify even the complex demands of specialized molecular testing. By incorporating design features for order review, software tools can permit a more personalized approach to sample handling and test selection without compromising efficiency. PMID:20007844
Molecular diagnostics: future probe-based strategies.
Marsh, Peter; Cardy, Donald L N
Nucleic acid amplification technologies (NAATs) represent powerful tools in clinical microbiology, particularly in areas where traditional culture-based methods alone prove insufficient. A notable advantage is in reducing the time from taking samples to reporting results. This, and the specificity and sensitivity imparted by NAATs, can help to improve patient care. Both thermal and isothermal NAATs have been adapted to aid diagnosis in clinical laboratories. Current molecular diagnostic assays are generally high-tech, and are expensive to buy and perform. Easy-to-use NAATs are beginning to appear, not only facilitating acceptable throughput in clinical laboratories, but also allowing tests to move out of the laboratory, closer to the point of care. Demand for simpler, miniaturized equipment and assays, and the trend toward personalized medicine, is leading towards the development of fully integrated automation and home-use kits. The integration of diverse disciplines, such as genomics, molecular biology, microelectromechanical systems, microfluidics, microfabrication, and organic chemistry, is behind the emerging DNA microarray technology. Development of DNA microchips allows the simultaneous detection of potentially thousands of target sequences, not only favoring high throughput, but also the potential for genotyping patient subsets with respect to their response to particular drug types (pharmakogenomics). It is envisaged that the future of probe-based technologies will see the development of fully integrated assays and devices suitable for nonskilled users. PMID:15148419
Towards in vitro molecular diagnostics using nanostructures.
Kurkina, Tetiana; Balasubramanian, Kannan
Nanostructures appear to be promising for a number of applications in molecular diagnostics, mainly due to the increased surface-to-volume ratio they can offer, the very low limit of detection achievable, and the possibility to fabricate point-of-care diagnostic devices. In this paper, we review examples of the use of nanostructures as diagnostic tools that bring in marked improvements over prevalent classical assays. The focus is laid on the various sensing paradigms that possess the potential or have demonstrated the capability to replace or augment current analytical strategies. We start with a brief introduction of the various types of nanostructures and their physical properties that determine the transduction principle. This is followed by a concise collection of various functionalization protocols used to immobilize biomolecules on the nanostructure surface. The sensing paradigms are discussed in two contexts: the nanostructure acting as a label for detection, or the nanostructure acting as a support upon which the molecular recognition events take place. In order to be successful in the field of molecular diagnostics, it is important that the nanoanalytical tools be evaluated in the appropriate biological environment. The final section of the review compiles such examples, where the nanostructure-based diagnostic tools have been tested on realistic samples such as serum, demonstrating their analytical power even in the presence of complex matrix effects. The ability of nanodiagnostic tools to detect ultralow concentrations of one or more analytes coupled with portability and the use of low sample volumes is expected to have a broad impact in the field of molecular diagnostics. PMID:22009454
Advances in molecular diagnostics for Mycobacterium bovis.
Collins, Desmond M
The two most important molecular diagnostic techniques for bovine tuberculosis are the polymerase chain reaction (PCR) because of its rapid determination of infection, and DNA strain typing because of its ability to answer important epidemiological questions. PCR tests for Mycobacterium bovis have been improved through recent advances in PCR technology, but still lack the sensitivity of good culture methods, and in some situations are susceptible to giving both false negative and false positive results. Therefore, PCR does not usually replace the need for culture, but is used to provide fast preliminary results. DNA typing of M. bovis isolates by restriction endonuclease analysis (REA) was developed 25 years ago in New Zealand, and remains an important tool in the New Zealand control scheme, where the typing results are combined with other information to determine large and expensive possum poisoning operations. A range of other DNA typing systems developed for M. bovis in the 1990 s have assisted epidemiological investigations in some countries but are now less commonly used. Variable number tandem repeat (VNTR) typing and spoligotyping, either alone or together, have now become the preferred approaches as they are robust and amenable to electronic analysis and comparison. Spoligotyping gives only moderate discrimination but can be easily applied to large numbers of isolates, and VNTR typing provides better discrimination than all other methods except for REA. While the current typing techniques are sufficient for most epidemiological purposes, more discriminating methods are likely to become available in the near future. PMID:21420257
Trends in Laboratory Diagnostic Methods in Periodontology.
Bolerázska, Beáta; Mareková, Mária; Markovská, Neda
This work presents a summary of current knowledge on the laboratory diagnosis of periodontitis. It focuses on the theoretical foundations and is supplemented with new knowledge. It subsequently describes specifically the laboratory diagnosis methods of periodontitis: the protein expression of inflammation, oral microbiology and molecular diagnostics. Periodontitis is a serious disease worldwide and its confirmed association with systemic diseases means its severity is increasing. Its laboratory diagnosis has the potential to rise to the level of clinical and diagnostic imaging. The transfer of diagnostic methods from laboratory to clinical use is increasingly used in the prevention and monitoring of the exacerbation and treatment of periodontal disease, as well as of its impact on systemic disease. PMID:27131349
Intelligent medical diagnostics via molecular logic
Konry, Tania; Walt, David R.
In this communication, we describe the integration of microarray sensor technology with logic capability for screening combinations of proteins and DNA in a biological sample. In this system, we have demonstrated the use of a single platform amenable to both protein detection and protein-DNA detection using molecular logic gates. The pattern of protein and DNA inputs results in fluorescence outputs according to a truth table for AND and INHIBIT gates, thereby demonstrating the feasibility of performing medical diagnostics using a logic gate design. One possible application of this technique would be for the direct screening of various medical conditions that are dependent on combinations of diagnostic markers. PMID:19715272
"Paper Machine" for Molecular Diagnostics.
Connelly, John T; Rolland, Jason P; Whitesides, George M
Clinical tests based on primer-initiated amplification of specific nucleic acid sequences achieve high levels of sensitivity and specificity. Despite these desirable characteristics, these tests have not reached their full potential because their complexity and expense limit their usefulness to centralized laboratories. This paper describes a device that integrates sample preparation and loop-mediated isothermal amplification (LAMP) with end point detection using a hand-held UV source and camera phone. The prototype device integrates paper microfluidics (to enable fluid handling) and a multilayer structure, or a "paper machine", that allows a central patterned paper strip to slide in and out of fluidic path and thus allows introduction of sample, wash buffers, amplification master mix, and detection reagents with minimal pipetting, in a hand-held, disposable device intended for point-of-care use in resource-limited environments. This device creates a dynamic seal that prevents evaporation during incubation at 65 °C for 1 h. This interval is sufficient to allow a LAMP reaction for the Escherichia coli malB gene to proceed with an analytical sensitivity of 1 double-stranded DNA target copy. Starting with human plasma spiked with whole, live E. coli cells, this paper demonstrates full integration of sample preparation with LAMP amplification and end point detection with a limit of detection of 5 cells. Further, it shows that the method used to prepare sample enables concentration of DNA from sample volumes commonly available from fingerstick blood draw. PMID:26104869
Whole Genome Sequencing Increases Molecular Diagnostic Yield Compared with Current Diagnostic Testing for Inherited Retinal Disease
Ellingford, Jamie M.; Barton, Stephanie; Bhaskar, Sanjeev; Williams, Simon G.; Sergouniotis, Panagiotis I.; O'Sullivan, James; Lamb, Janine A.; Perveen, Rahat; Hall, Georgina; Newman, William G.; Bishop, Paul N.; Roberts, Stephen A.; Leach, Rick; Tearle, Rick; Bayliss, Stuart; Ramsden, Simon C.; Nemeth, Andrea H.; Black, Graeme C.M.
Purpose To compare the efficacy of whole genome sequencing (WGS) with targeted next-generation sequencing (NGS) in the diagnosis of inherited retinal disease (IRD). Design Case series. Participants A total of 562 patients diagnosed with IRD. Methods We performed a direct comparative analysis of current molecular diagnostics with WGS. We retrospectively reviewed the findings from a diagnostic NGS DNA test for 562 patients with IRD. A subset of 46 of 562 patients (encompassing potential clinical outcomes of diagnostic analysis) also underwent WGS, and we compared mutation detection rates and molecular diagnostic yields. In addition, we compared the sensitivity and specificity of the 2 techniques to identify known single nucleotide variants (SNVs) using 6 control samples with publically available genotype data. Main Outcome Measures Diagnostic yield of genomic testing. Results Across known disease-causing genes, targeted NGS and WGS achieved similar levels of sensitivity and specificity for SNV detection. However, WGS also identified 14 clinically relevant genetic variants through WGS that had not been identified by NGS diagnostic testing for the 46 individuals with IRD. These variants included large deletions and variants in noncoding regions of the genome. Identification of these variants confirmed a molecular diagnosis of IRD for 11 of the 33 individuals referred for WGS who had not obtained a molecular diagnosis through targeted NGS testing. Weighted estimates, accounting for population structure, suggest that WGS methods could result in an overall 29% (95% confidence interval, 15–45) uplift in diagnostic yield. Conclusions We show that WGS methods can detect disease-causing genetic variants missed by current NGS diagnostic methodologies for IRD and thereby demonstrate the clinical utility and additional value of WGS. PMID:26872967
[Practical aspects of molecular diagnostics in genodermatoses].
Has, C; He, Y
Genodermatoses are rare genetic disorders with a broad spectrum of cutaneous and extracutaneous manifestations that have a genetic background. A thorough clinical examination, laboratory workup and morphological analyses of the skin remain crucial for the diagnosis in the era of next generation sequencing (NGS). The diagnostic algorithm depends on the clinical and molecular heterogeneity and should be adapted for each group of genodermatoses. In cases with uncharacteristic phenotypes which cannot be classified, NGS-based testing accelerates the time to diagnosis and leads to the identification of new disorders and new disease-associated genes. The new knowledge on genotype-phenotype correlations should enable revision of the classification of genodermatoses on a molecular basis. PMID:26537890
Myasthenia Gravis: Tests and Diagnostic Methods
... Affiliations Foundation Focus Newsletter E-Update Test & Diagnostic methods In addition to a complete medical and neurological ... How can I help? About MGFA Test & Diagnostic methods Treatment for MG FAQ's Upcoming Events Spring 2016 ...
The development of a post-baccalaureate certificate program in molecular diagnostics.
Williams, G S; Brown, J D; Keagle, M B
A post-baccalaureate certificate program in diagnostic molecular sciences was created in 1995 by the Diagnostic Genetic Sciences Program in the School of Allied Health at the University of Connecticut. The required on-campus lecture and laboratory courses include basic laboratory techniques, health care issues, cell biology, immunology, human genetics, research, management, and molecular diagnostic techniques and laboratory in molecular diagnostics. These courses precede a 6-month, full-time practicum at an affiliated full-service molecular laboratory. The practicum includes amplification and blotting methods, a research project, and a choice of specialized electives including DNA sequencing, mutagenesis, in situ hybridization methods, or molecular diagnostic applications in microbiology. Graduates of the program are immediately eligible to sit for the National Credentialing Agency examination in molecular biology to obtain the credential Clinical Laboratory Specialist in Molecular Biology (CLSp(MB). This description of the University of Connecticut program may assist other laboratory science programs in creating similar curricula. PMID:11232107
Genetic Diagnostic Methods for Inherited Eye Diseases
Gabriel, Luis A. R.; Traboulsi, Elias I.
Accurate molecular diagnosis of genetic eye diseases has proven to be of great importance because of the prognostic and therapeutic value of an accurate ascertainment of the underlying genetic mutation. Efforts continue in diagnostic laboratories to develop strategies that allow the discovery of responsible gene/mutations in the individual patient using the least number of assays and economizing on the expenses and time involved in the process. Once the ophthalmologist has made the best possible clinical diagnosis, blood samples are obtained for genetic testing. In this paper we will review the basic laboratory methods utilized to identify the chromosomal or mutational etiology of genetic diseases that affect the eye. PMID:21572730
Accelerated molecular dynamics methods
Perez, Danny
The molecular dynamics method, although extremely powerful for materials simulations, is limited to times scales of roughly one microsecond or less. On longer time scales, dynamical evolution typically consists of infrequent events, which are usually activated processes. This course is focused on understanding infrequent-event dynamics, on methods for characterizing infrequent-event mechanisms and rate constants, and on methods for simulating long time scales in infrequent-event systems, emphasizing the recently developed accelerated molecular dynamics methods (hyperdynamics, parallel replica dynamics, and temperature accelerated dynamics). Some familiarity with basic statistical mechanics and molecular dynamics methods will be assumed.
[Molecular markers of carcinogenesis in the diagnostics of cervical cancer].
Bedkowska, Grazyna Ewa; Ławicki, Sławomir; Szmitkowski, Maciej
Cervical carcinoma is the most frequent disease of the reproductive organ and is the second most common cancer in women after breast cancer. As it is characterized by high mortality, new diagnostic methods are needed, for example tumor markers, enabling earlier diagnosis and rapid detection of recurrence after therapy. Different tumor markers may be useful in the diagnostics of cervical cancer, for example squamous cell carcinoma antigen (SCC-Ag), tissue polypeptide antigen (TPA), and CYFRA 21-1, as well as some cytokines such as vascular endothelial growth factor (VEGF), granulocyte colony-stimulating factor, and macrophage colony-stimulating factor (M-CSF). About 150 genes connected with the carcinogenesis of cervical carcinoma have been identified. This paper is devoted to evaluating the diagnostic usefulness of molecular markers of carcinogenesis, especially P53, Bcl-2, Brn-3a, and MCM, and comparing the results with those of typical tumor markers or cytokines useful in diagnosing this type of cancer. It was shown that telomerase and Brn-3a proteins demonstrate usefulness in screening examination, P53 in monitoring the effectiveness of therapy, and Bcl-2 as a survival prognostic factor. In summary, it is evident that molecular makers of carcinogenesis are helpful in the diagnostics of cervical cancer, but further investigation and confirmation by a prospective study is necessary. PMID:19252468
Supramolecular Nanoparticles for Molecular Diagnostics and Therapeutics
NASA Astrophysics Data System (ADS)
Chen, Kuan-Ju
single SNP for both diagnosis and therapy were generated. The results show that this type of theranostic SNPs may have a great contribution in the optimization of therapeutic efficacy for individual patients in clinical translation in the near future. It is anticipated that our supramolecular synthetic approach could be adopted to assemble various SNP-based delivery agents for molecular diagnostics and therapeutics that pave the way toward personalized medicine.
Molecular profiling of single circulating tumor cells with diagnostic intention
Polzer, Bernhard; Medoro, Gianni; Pasch, Sophie; Fontana, Francesca; Zorzino, Laura; Pestka, Aurelia; Andergassen, Ulrich; Meier-Stiegen, Franziska; Czyz, Zbigniew T; Alberter, Barbara; Treitschke, Steffi; Schamberger, Thomas; Sergio, Maximilian; Bregola, Giulia; Doffini, Anna; Gianni, Stefano; Calanca, Alex; Signorini, Giulio; Bolognesi, Chiara; Hartmann, Arndt; Fasching, Peter A; Sandri, Maria T; Rack, Brigitte; Fehm, Tanja; Giorgini, Giuseppe; Manaresi, Nicolò; Klein, Christoph A
Several hundred clinical trials currently explore the role of circulating tumor cell (CTC) analysis for therapy decisions, but assays are lacking for comprehensive molecular characterization of CTCs with diagnostic precision. We therefore combined a workflow for enrichment and isolation of pure CTCs with a non-random whole genome amplification method for single cells and applied it to 510 single CTCs and 189 leukocytes of 66 CTC-positive breast cancer patients. We defined a genome integrity index (GII) to identify single cells suited for molecular characterization by different molecular assays, such as diagnostic profiling of point mutations, gene amplifications and whole genomes of single cells. The reliability of > 90% for successful molecular analysis of high-quality clinical samples selected by the GII enabled assessing the molecular heterogeneity of single CTCs of metastatic breast cancer patients. We readily identified genomic disparity of potentially high relevance between primary tumors and CTCs. Microheterogeneity analysis among individual CTCs uncovered pre-existing cells resistant to ERBB2-targeted therapies suggesting ongoing microevolution at late-stage disease whose exploration may provide essential information for personalized treatment decisions and shed light into mechanisms of acquired drug resistance. PMID:25358515
Review of rapid molecular diagnostic tools for avian influenza
Technology Transfer Automated Retrieval System (TEKTRAN)
Molecular diagnostics tests are commonly used to diagnose avian influenza virus (AIV) because they are sensitive, can be performed rapidly, with high throughput, and at a moderate cost. Molecular diagnostic tests have recently proven themselves to be invaluable in controlling disease outbreaks arou...
Molecular diagnostics in tuberculosis: basis and implications for therapy.
Balasingham, Seetha V; Davidsen, Tonje; Szpinda, Irena; Frye, Stephan A; Tønjum, Tone
The processing of clinical specimens in the mycobacterial diagnostic laboratory has undergone remarkable improvements during the last decade. While microscopy and culture are still the major backbone for laboratory diagnosis of tuberculosis on a worldwide basis, new methods including molecular diagnostic tests have evolved over the last two decades. The majority of molecular tests have been focused on (i) detection of nucleic acids, both DNA and RNA, that are specific to Mycobacterium tuberculosis, by amplification techniques such as polymerase chain reaction (PCR); and (ii) detection of mutations in the genes that are associated with resistance to antituberculosis drugs by sequencing or nucleic acid hybridization. Recent developments in direct and rapid detection of mycobacteria, with emphasis on M. tuberculosis species identification by 16S rRNA gene sequence analysis or oligohybridization and strain typing, as well as detection of drug susceptibility patterns, all contribute to these advances. Generally, the balance between genome instability and genome maintenance as the basis for evolutionary development, strain diversification and resistance development is important, because it cradles the resulting M. tuberculosis phenotype. At the same time, semi-automated culture systems have contributed greatly to the increased sensitivity and reduced turnaround time in the mycobacterial analysis of clinical specimens. Collectively, these advances are particularly important for establishing the diagnosis of tuberculosis in children. More basic and operational research to appraise the impact and cost effectiveness of new diagnostic technologies must, however, be carried out. Furthermore, the design and quality of clinical trials evaluating new diagnostics must be improved to allow clinical and laboratory services that would provide rapid response to test results. Thus, important work remains before the new diagnostic tools can be meaningfully integrated into national
Polymerase chain reaction: A molecular diagnostic tool in periodontology.
Maheaswari, Rajendran; Kshirsagar, Jaishree Tukaram; Lavanya, Nallasivam
This review discusses the principles of polymerase chain reaction (PCR) and its application as a diagnostic tool in periodontology. The relevant MEDLINE and PubMed indexed journals were searched manually and electronically by typing PCR, applications of PCR, PCR in periodontics, polymorphism studies in periodontitis, and molecular techniques in periodontology. The searches were limited to articles in English language and the articles describing PCR process and its relation to periodontology were collected and used to prepare a concise review. PCR has now become a standard diagnostic and research tool in periodontology. Various studies reveal that its sensitivity and specificity allow it as a rapid, efficient method of detecting, identifying, and quantifying organism. Different immune and inflammatory markers can be identified at the mRNA expression level, and also the determination of genetic polymorphisms, thus providing the deeper insight into the mechanisms underlying the periodontal disease. PMID:27143822
Polymerase chain reaction: A molecular diagnostic tool in periodontology
Maheaswari, Rajendran; Kshirsagar, Jaishree Tukaram; Lavanya, Nallasivam
This review discusses the principles of polymerase chain reaction (PCR) and its application as a diagnostic tool in periodontology. The relevant MEDLINE and PubMed indexed journals were searched manually and electronically by typing PCR, applications of PCR, PCR in periodontics, polymorphism studies in periodontitis, and molecular techniques in periodontology. The searches were limited to articles in English language and the articles describing PCR process and its relation to periodontology were collected and used to prepare a concise review. PCR has now become a standard diagnostic and research tool in periodontology. Various studies reveal that its sensitivity and specificity allow it as a rapid, efficient method of detecting, identifying, and quantifying organism. Different immune and inflammatory markers can be identified at the mRNA expression level, and also the determination of genetic polymorphisms, thus providing the deeper insight into the mechanisms underlying the periodontal disease. PMID:27143822
Multispectral optical tweezers for molecular diagnostics of single biological cells
NASA Astrophysics Data System (ADS)
Butler, Corey; Fardad, Shima; Sincore, Alex; Vangheluwe, Marie; Baudelet, Matthieu; Richardson, Martin
Optical trapping of single biological cells has become an established technique for controlling and studying fundamental behavior of single cells with their environment without having "many-body" interference. The development of such an instrument for optical diagnostics (including Raman and fluorescence for molecular diagnostics) via laser spectroscopy with either the "trapping" beam or secondary beams is still in progress. This paper shows the development of modular multi-spectral imaging optical tweezers combining Raman and Fluorescence diagnostics of biological cells.
Molecular diagnostic assays for infectious diseases in cats.
Veir, Julia K; Lappin, Michael R
With the advent of more accessible polymerase chain reaction panels, the use of molecular techniques for the detection of infectious organisms has become more routine in veterinary medicine. The use of molecular diagnostics is best reserved for the detection of organisms that are difficult to detect or identify expediently. In this article, the fundamentals of molecular techniques are reviewed along with an examination of specific feline infectious diseases in which diagnosis via molecular techniques is advantageous. PMID:20933144
Malignant Catarrhal Fever: Understanding Molecular Diagnostics in Context of Epidemiology
Li, Hong; Cunha, Cristina W.; Taus, Naomi S.
Malignant catarrhal fever (MCF) is a frequently fatal disease, primarily of ruminants, caused by a group of gammaherpesviruses. Due to complexities of pathogenesis and epidemiology in various species, which are either clinically-susceptible or reservoir hosts, veterinary clinicians face significant challenges in laboratory diagnostics. The recent development of specific assays for viral DNA and antibodies has expanded and improved the inventory of laboratory tests and opened new opportunities for use of MCF diagnostics. Issues related to understanding and implementing appropriate assays for specific diagnostic needs must be addressed in order to take advantage of molecular diagnostics in the laboratory. PMID:22072925
The Application of Molecular Diagnostics to Stained Cytology Smears.
Oktay, Maja H; Adler, Esther; Hakima, Laleh; Grunblatt, Eli; Pieri, Evan; Seymour, Andrew; Khader, Samer; Cajigas, Antonio; Suhrland, Mark; Goswami, Sumanta
Detection of mutational alterations is important for guiding treatment decisions of lung non-small-cell carcinomas and thyroid nodules with atypical cytologic findings. Inoperable lung tumors requiring further testing for staging and thyroid lesions often are diagnosed using only cytology material. Molecular diagnostic tests of these samples typically are performed on cell blocks; however, insufficient cellularity of cell blocks is a limitation for test performance. In addition, some of the fixatives used while preparing cell blocks often introduces artifacts for mutation detection. Here, we applied qClamp xenonucleic technology and quantitative RT-PCR to cells microdissected directly from stained cytology smears to detect common alterations including mutations and translocations in non-small-cell carcinomas and thyroid lesions. By using this approach, we achieved a 1% molecular alteration detection rate from as few as 50 cells. Ultrasensitive methods of molecular alteration detection similar to the one described here will be increasingly important for the evaluation of molecular alterations in clinical scenarios when only tissue samples that are small are available. PMID:26921541
Methods in Molecular Biophysics
NASA Astrophysics Data System (ADS)
Serdyuk, Igor N.; Zaccai, Nathan R.; Zaccai, Joseph
Our knowledge of biological macromolecules and their interactions is based on the application of physical methods, ranging from classical thermodynamics to recently developed techniques for the detection and manipulation of single molecules. These methods, which include mass spectrometry, hydrodynamics, microscopy, diffraction and crystallography, electron microscopy, molecular dynamics simulations, and nuclear magnetic resonance, are complementary; each has its specific advantages and limitations. Organised by method, this textbook provides descriptions and examples of applications for the key physical methods in modern biology. It is an invaluable resource for undergraduate and graduate students of molecular biophysics in science and medical schools, as well as research scientists looking for an introduction to techniques beyond their specialty. As appropriate for this interdisciplinary field, the book includes short asides to explain physics aspects to biologists and biology aspects to physicists. Comprehensive coverage and up-to-date treatment of the latest physical methods in modern biology Each method includes a brief historical introduction, theoretical principles, applications, advantages and limitations, and concludes with a checklist of key ideas Interdisciplinary and accessible to biologists, physicists, and those with medical backgrounds
Nutritional Lipidomics: Molecular Metabolism, Analytics, and Diagnostics
Smilowitz, Jennifer T.; Zivkovic, Angela M.; Wan, Yu-Jui Yvonne; Watkins, Steve M.; Nording, Malin L.; Hammock, Bruce D.; German, J. Bruce
The field of lipidomics is providing nutritional science a more comprehensive view of lipid intermediates. Lipidomics research takes advantage of the increase in accuracy and sensitivity of mass detection of mass spectrometry with new bioinformatics toolsets to characterize the structures and abundances of complex lipids. Yet, translating lipidomics to practice via nutritional interventions is still in its infancy. No single instrumentation platform is able to solve the varying analytical challenges of the different molecular lipid species. Biochemical pathways of lipid metabolism remain incomplete and the tools to map lipid compositional data to pathways are still being assembled. Biology itself is dauntingly complex and simply separating biological structures remains a key challenge to lipidomics. Nonetheless, the strategy of combining tandem analytical methods to perform the sensitive, high-throughput, quantitative and comprehensive analysis of lipid metabolites of very large numbers of molecules is poised to drive the field forward rapidly. Among the next steps for nutrition to understand the changes in structures, compositions and function of lipid biomolecules in response to diet is to describe their distribution within discrete functional compartments-lipoproteins. Additionally, lipidomics must tackle the task of assigning the functions of lipids as signaling molecules, nutrient sensors, and intermediates of metabolic pathways. PMID:23818328
Tomographic methods in flow diagnostics
NASA Technical Reports Server (NTRS)
Decker, Arthur J.
This report presents a viewpoint of tomography that should be well adapted to currently available optical measurement technology as well as the needs of computational and experimental fluid dynamists. The goals in mind are to record data with the fastest optical array sensors; process the data with the fastest parallel processing technology available for small computers; and generate results for both experimental and theoretical data. An in-depth example treats interferometric data as it might be recorded in an aeronautics test facility, but the results are applicable whenever fluid properties are to be measured or applied from projections of those properties. The paper discusses both computed and neural net calibration tomography. The report also contains an overview of key definitions and computational methods, key references, computational problems such as ill-posedness, artifacts, missing data, and some possible and current research topics.
[Syndrome of diabetic foot: modern diagnostic methods].
Plekhanov, A N; Markevich, P S
We summarize the literature data on diagnostics of diabetic foot syndrome including clinical examination and special invasive and non-invasive studies of the vascular system. The main methods are ultrasound dopplerography, X-ray contrast angiography, and ultrasound duplex scanning. Special attention is given to instrumental diagnostics of diabetic neuropathies. The golden standard for the evaluation of the function of the peripheral nervous system is electroneuromyography. Methods for the study of diabetic foot complications, such as osteoarthropathy and trophic ulcers, are discussed. PMID:25782303
Dental diagnostics: molecular analysis of oral biofilms.
Hiyari, Sarah; Bennett, Katie M
Dental biofilms are complex, multi-species bacterial communities that colonize the mouth in the form of plaque and are known to cause dental caries and periodontal disease. Biofilms are unique from planktonic bacteria in that they are mutualistic communities with a 3-dimensional structure and complex nutritional and communication pathways. The homeostasis within the biofilm colony can be disrupted, causing a shift in the bacterial composition of the colony and resulting in proliferation of pathogenic species. Because of this dynamic lifestyle, traditional microbiological techniques are inadequate for the study of biofilms. Many of the bacteria present in the oral cavity are viable but not culturable, which severely limits laboratory analysis. However, with the advent of new molecular techniques, the microbial makeup of oral biofilms can be better identified. Some of these techniques include DNA-DNA hybridization, 16S rRNA gene sequencing, denaturing gradient gel electrophoresis, terminal restriction fragment length polymorphism, denaturing high-performance liquid chromatography and pyrosequencing. This review provides an overview of biofilm formation and examines the major molecular techniques currently used in oral biofilm analysis. Future applications of the molecular analysis of oral biofilms in the diagnosis and treatment of caries and periodontal disease are also discussed. PMID:22309866
Capillary electrophoresis of DNA for molecular diagnostics.
Righetti, P G; Gelfi, C
A number of applications of capillary zone electrophoresis (CZE) in sieving liquid polymers (notably linear polyacrylamides and cellulose) for the analysis of polymerase chain reaction (PCR) products of clinically relevant, diagnostic DNA, are reviewed. The fields covered are: human genetics, quantitative gene dosage, microbiology and virology, forensic medicine and therapeutic DNA (notably, antisense nucleotides). Some unique, novel developments are highlighted, such as: (i) nonisocratic CZE, i.e., temperature-programmed CZE for detection of DNA point mutations; (ii) the synthesis of novel N-substituted acrylamides, offering extreme resistance to alkaline hydrolysis coupled to high hydrophilicity. In the field of denaturing gradient gel electrophoresis (DGGE), as routinely performed in gel slabs, a novel methodology is described in CZE: double-gradient DGGE. In this technique, two gradients are simultaneously applied along the migration direction: a chemical (or thermal) denaturing gradient, for partially unwinding homo- and hetero-duplexes of DNA, and a porosity gradient, for recompacting diffuse bands melting over a broader range of denaturing conditions. It is thus demonstrated that chemical gradients, in addition to temperature gradients, can be easily implemented even in a capillary format. PMID:9372261
A Complete Molecular Diagnostic Procedure for Applications in Surveillance and Subtyping of Avian Influenza Virus
Tseng, Chun-Hsien; Tsai, Hsiang-Jung; Chang, Chung-Ming
Introduction. The following complete molecular diagnostic procedure we developed, based on real-time quantitative PCR and traditional PCR, is effective for avian influenza surveillance, virus subtyping, and viral genome sequencing. Method. This study provides a specific and sensitive step-by-step procedure for efficient avian influenza identification of 16 hemagglutinin and 9 neuraminidase avian influenza subtypes. Result and Conclusion. This diagnostic procedure may prove exceedingly useful for virological and ecological advancements in global avian influenza research. PMID:25057497
Personalized Cancer Medicine: Molecular Diagnostics, Predictive biomarkers, and Drug Resistance
Gonzalez de Castro, D; Clarke, P A; Al-Lazikani, B; Workman, P
The progressive elucidation of the molecular pathogenesis of cancer has fueled the rational development of targeted drugs for patient populations stratified by genetic characteristics. Here we discuss general challenges relating to molecular diagnostics and describe predictive biomarkers for personalized cancer medicine. We also highlight resistance mechanisms for epidermal growth factor receptor (EGFR) kinase inhibitors in lung cancer. We envisage a future requiring the use of longitudinal genome sequencing and other omics technologies alongside combinatorial treatment to overcome cellular and molecular heterogeneity and prevent resistance caused by clonal evolution. PMID:23361103
The impact of new trends in POCTs for companion diagnostics, non-invasive testing and molecular diagnostics.
Huckle, David
Point-of-care diagnostics have been slowly developing over several decades and have taken on a new importance in current healthcare delivery for both diagnostics and development of new drugs. Molecular diagnostics have become a key driver of technology change and opened up new areas in companion diagnostics for use alongside pharmaceuticals and in new clinical approaches such as non-invasive testing. Future areas involving smartphone and other information technology advances, together with new developments in molecular biology, microfluidics and surface chemistry are adding to advances in the market. The focus for point-of-care tests with molecular diagnostic technologies is focused on advancing effective applications. PMID:25990929
Intelligent DNA-based molecular diagnostics using linked genetic markers
Pathak, D.K.; Perlin, M.W.; Hoffman, E.P.
This paper describes a knowledge-based system for molecular diagnostics, and its application to fully automated diagnosis of X-linked genetic disorders. Molecular diagnostic information is used in clinical practice for determining genetic risks, such as carrier determination and prenatal diagnosis. Initially, blood samples are obtained from related individuals, and PCR amplification is performed. Linkage-based molecular diagnosis then entails three data analysis steps. First, for every individual, the alleles (i.e., DNA composition) are determined at specified chromosomal locations. Second, the flow of genetic material among the individuals is established. Third, the probability that a given individual is either a carrier of the disease or affected by the disease is determined. The current practice is to perform each of these three steps manually, which is costly, time consuming, labor-intensive, and error-prone. As such, the knowledge-intensive data analysis and interpretation supersede the actual experimentation effort as the major bottleneck in molecular diagnostics. By examining the human problem solving for the task, we have designed and implemented a prototype knowledge-based system capable of fully automating linkage-based molecular diagnostics in X-linked genetic disorders, including Duchenne Muscular Dystrophy (DMD). Our system uses knowledge-based interpretation of gel electrophoresis images to determine individual DNA marker labels, a constraint satisfaction search for consistent genetic flow among individuals, and a blackboard-style problem solver for risk assessment. We describe the system`s successful diagnosis of DMD carrier and affected individuals from raw clinical data.
Something old and something new about molecular diagnostics in gliomas
Horbinski, Craig
Synopsis Progress in our understanding of the molecular biology of neoplasms has been driven by remarkable improvements in molecular biology techniques. This has created a rapidly moving field in which even subspecialists struggle to keep abreast of the current literature. Nowhere is this more clearly demonstrated than in neuro-oncology, wherein molecular diagnostics can now wring more clinically useful information out of very small biopsies than ever before. Herein the biologic and practical aspects of four key molecular biomarkers in gliomas are discussed, including two that have been known for some time (1p/19q codeletion and EGFR amplification) as well as two whose relevance was discovered via advanced whole-genome assays (IDH1/2 mutations and BRAF alterations). PMID:23459421
A new molecular diagnostic tool for quantitatively detecting and genotyping "Candidatus Liberibacter species"
Technology Transfer Automated Retrieval System (TEKTRAN)
A new molecular diagnostic method was developed for quantitative detection of "Candidatus Liberibacter" species associated with citrus Huanglongbing ("Ca. Liberibacter asiaticus", "Ca. Liberibacter africanus" and "Ca. Liberibacter americanus") and potato zebra chip disorder ("Ca. Liberibacter solana...
Non-invasive diagnostic methods in dentistry
NASA Astrophysics Data System (ADS)
Todea, Carmen
The paper, will present the most important non-invasive methods for diagnostic, in different fields of dentistry. Moreover, the laser-based methods will be emphasis. In orthodontics, 3D laser scanners are increasingly being used to establish database for normative population and cross-sectional growth changes but also to asses clinical outcomes in orthognatic surgical and non-surgical treatments. In prevention the main methods for diagnostic of demineralization and caries detection in early stages are represented by laser fluorescence - Quantitative Light Florescence (QLF); DiagnoDent-system-655nm; FOTI-Fiberoptic transillumination; DIFOTI-Digital Imaging Fiberoptic transillumination; and Optical Coherence Tomography (OCT). In odontology, Laser Doppler Flowmetry (LDF) is a noninvasive real time method used for determining the tooth vitality by monitoring the pulp microcirculation in traumatized teeth, fractured teeth, and teeth undergoing different conservative treatments. In periodontology, recently study shows the ability of LDF to evaluate the health of gingival tissue in periodontal tissue diseases but also after different periodontal treatments.
Paper-based sample-to-answer molecular diagnostic platform for point-of-care diagnostics.
Choi, Jane Ru; Tang, Ruihua; Wang, ShuQi; Wan Abas, Wan Abu Bakar; Pingguan-Murphy, Belinda; Xu, Feng
Nucleic acid testing (NAT), as a molecular diagnostic technique, including nucleic acid extraction, amplification and detection, plays a fundamental role in medical diagnosis for timely medical treatment. However, current NAT technologies require relatively high-end instrumentation, skilled personnel, and are time-consuming. These drawbacks mean conventional NAT becomes impractical in many resource-limited disease-endemic settings, leading to an urgent need to develop a fast and portable NAT diagnostic tool. Paper-based devices are typically robust, cost-effective and user-friendly, holding a great potential for NAT at the point of care. In view of the escalating demand for the low cost diagnostic devices, we highlight the beneficial use of paper as a platform for NAT, the current state of its development, and the existing challenges preventing its widespread use. We suggest a strategy involving integrating all three steps of NAT into one single paper-based sample-to-answer diagnostic device for rapid medical diagnostics in the near future. PMID:26164488
Molecular Pathogenesis and Diagnostic, Prognostic and Predictive Molecular Markers in Sarcoma.
Mariño-Enríquez, Adrián; Bovée, Judith V M G
Sarcomas are infrequent mesenchymal neoplasms characterized by notable morphological and molecular heterogeneity. Molecular studies in sarcoma provide refinements to morphologic classification, and contribute diagnostic information (frequently), prognostic stratification (rarely) and predict therapeutic response (occasionally). Herein, we summarize the major molecular mechanisms underlying sarcoma pathogenesis and present clinically useful diagnostic, prognostic and predictive molecular markers for sarcoma. Five major molecular alterations are discussed, illustrated with representative sarcoma types, including 1. the presence of chimeric transcription factors, in vascular tumors; 2. abnormal kinase signaling, in gastrointestinal stromal tumor; 3. epigenetic deregulation, in chondrosarcoma, chondroblastoma, and other tumors; 4. deregulated cell survival and proliferation, due to focal copy number alterations, in dedifferentiated liposarcoma; 5. extreme genomic instability, in conventional osteosarcoma as a representative example of sarcomas with highly complex karyotype. PMID:27523972
Early Identification and Treatment of Pathogens in Sepsis: Molecular Diagnostics and Antibiotic Choice.
Riedel, Stefan; Carroll, Karen C
Sepsis and septic shock are serious conditions associated with high morbidity and mortality. Rapid molecular methods for detection of microorganisms and antimicrobial resistance genes from positive blood cultures or whole blood have evolved over the past 10 years. Such diagnostic methods coupled with therapeutic interventional programs are desirable to improve the overall clinical outcome and mortality. This article discusses the usefulness of current molecular test methods for the diagnosis of sepsis and their potential to enhance the success of antimicrobial stewardship programs. Clinicians and laboratories alike must appreciate key factors influencing the appropriate use and potential impact of these methods. PMID:27229637
New Approaches to Sepsis: Molecular Diagnostics and Biomarkers
Bauer, Michael; Riedemann, Niels C.; Hartog, Christiane S.
Summary: Sepsis is among the most common causes of death in hospitals. It arises from the host response to infection. Currently, diagnosis relies on nonspecific physiological criteria and culture-based pathogen detection. This results in diagnostic uncertainty, therapeutic delays, the mis- and overuse of antibiotics, and the failure to identify patients who might benefit from immunomodulatory therapies. There is a need for new sepsis biomarkers that can aid in therapeutic decision making and add information about screening, diagnosis, risk stratification, and monitoring of the response to therapy. The host response involves hundreds of mediators and single molecules, many of which have been proposed as biomarkers. It is, however, unlikely that one single biomarker is able to satisfy all the needs and expectations for sepsis research and management. Among biomarkers that are measurable by assays approved for clinical use, procalcitonin (PCT) has shown some usefulness as an infection marker and for antibiotic stewardship. Other possible new approaches consist of molecular strategies to improve pathogen detection and molecular diagnostics and prognostics based on transcriptomic, proteomic, or metabolic profiling. Novel approaches to sepsis promise to transform sepsis from a physiologic syndrome into a group of distinct biochemical disorders and help in the development of better diagnostic tools and effective adjunctive sepsis therapies. PMID:23034322
New approaches to sepsis: molecular diagnostics and biomarkers.
Reinhart, Konrad; Bauer, Michael; Riedemann, Niels C; Hartog, Christiane S
Sepsis is among the most common causes of death in hospitals. It arises from the host response to infection. Currently, diagnosis relies on nonspecific physiological criteria and culture-based pathogen detection. This results in diagnostic uncertainty, therapeutic delays, the mis- and overuse of antibiotics, and the failure to identify patients who might benefit from immunomodulatory therapies. There is a need for new sepsis biomarkers that can aid in therapeutic decision making and add information about screening, diagnosis, risk stratification, and monitoring of the response to therapy. The host response involves hundreds of mediators and single molecules, many of which have been proposed as biomarkers. It is, however, unlikely that one single biomarker is able to satisfy all the needs and expectations for sepsis research and management. Among biomarkers that are measurable by assays approved for clinical use, procalcitonin (PCT) has shown some usefulness as an infection marker and for antibiotic stewardship. Other possible new approaches consist of molecular strategies to improve pathogen detection and molecular diagnostics and prognostics based on transcriptomic, proteomic, or metabolic profiling. Novel approaches to sepsis promise to transform sepsis from a physiologic syndrome into a group of distinct biochemical disorders and help in the development of better diagnostic tools and effective adjunctive sepsis therapies. PMID:23034322
Ebola Check: Delivering molecular diagnostics at the point of need.
Moschos, Sterghios A
The 2013-5 global outbreak of Ebolavirus disease brought to sharp focus the need for diagnostic capacity to be equitably available on a global scale: from the most under-developed areas of resource-limited countries in West Africa to high volume international travel hubs in Europe and the USA. Quick detection of the causal agent of disease is pivotal to containment, contact tracing and clinical action to protect healthcare workers, communities and patients. Nucleic acid testing (NAT) by real time reverse transcription quantitative polymerase chain reaction (RT-PCR) has emerged as the preferred method for reliable patient status confirmation. Presently, this is served through advanced clinical molecular laboratory testing in a <8hr manual process that requires 3.5ml venous blood samples. To meet the demand in West Africa, this has necessitated large-scale mobile laboratory and volunteer biomedical scientist deployment: a solution that has proven eventually adequate, albeit temporary against future re-emergence of this and other haemorrhagic fever disease agents prevalent in the region. The EbolaCheck consortium was formed in August 2014 to address the need for delivering NAT at the point of care. We have developed a novel platform technology that can QUantitatively, RAPidly IDentify (QuRapID) known RNA or DNA targets in viruses, bacteria, or eukaryotic cells directly in crude biofluids, including whole blood, in under 40min using a 5 microliter sample. The portable, battery-operated system lacks microfluidics, pumps or other sensitive/high cost parts making it suitable for the environmental and economic challenges of resource-limited countries. The simple, safe, 5-step sample-to-answer process requires minimal training and informs frontline healthcare workers of diagnostic status, whilst reporting remotely epidemiologically relevant results. Data on biosafety level 2 surrogate Ebolavirus templates presented in encapsulated or enveloped viruses indicate performance
Diagnostic gold standard for soft tissue tumours: morphology or molecular genetics?
Pfeifer, J D; Hill, D A; O'Sullivan, M J; Dehner, L P
The recognition of recurrent genetic alterations in specific tumour types has provided the basis for the reclassification of certain soft tissue neoplasms, and molecular analysis of patient material has the potential to provide both diagnostic and prognostic information. In this review, we evaluate the role of molecular genetic testing as the prospective 'gold standard' for sarcoma diagnosis. Molecular genetic testing, as with every new method, promises to improve accuracy and to be more sensitive and less subjective, claims that have been made previously by histochemistry, electron microscopy and immunohistochemistry. Technical limitations in molecular assays, as well as more general specificity issues, decrease the clinical usefulness of molecular pathological testing significantly and suggest that, at present, molecular evaluation is best considered an ancillary technique that neither supersedes other ancillary techniques nor eclipses traditional pathological examination. PMID:11122430
Three-Gene Molecular Diagnostic Model for Thyroid Cancer
Kowalski, Jeanne; Tsai, Hua-Ling; Talbot, Kristin; Somervell, Helina; Kouniavsky, Guennadi; Wang, Yongchun; Dackiw, Alan P.B.; Westra, William H.; Clark, Douglas P.; Libutti, Steven K.; Umbricht, Christopher B.
Background The preoperative diagnosis of thyroid nodules primarily depends upon fine needle aspiration (FNA) cytology. However, up to 25% of FNA samples have associated "suspicious or indeterminate", but not diagnostic cytologic reports, resulting in difficulty deciding appropriate clinical management for these patients. We hypothesize that the use of molecular markers as an adjunct to FNA cytology can improve the distinction of benign from malignant nodules that have associated suspicious or indeterminate cytology. Methods Using microarray analysis, we previously identified and reported on 75 genes useful in the distinction of benign versus malignant thyroid nodules. In the present study, we have further validated the expression of 14 of these markers in a large number of thyroid samples by immunohistochemistry (IHC) analysis of 154 thyroid tumors and quantitative real-time RT-PCR (QRT-PCR) analysis of 95 FNA samples. Of the 154 tumors analyzed by IHC, 44 samples (29%) had associated suspicious or indeterminate FNA cytology. Results Receiver operating characteristic using three-gene model, (HMGA2, MRC2, and SFN) analysis for the detection of malignant nodules resulted in areas under the curve (AUCs) of≥0.95 (80% sensitivity; 100% specificity) and≥0.84 (71% sensitivity; 84% specificity) for the IHC data in tumors, and QRT-PCR data in FNA samples, respectively. Conclusions Our results suggest that a three-gene model for the cytological diagnosis of indeterminate thyroid nodules is both feasible and promising. Implementation of this as an adjunct to thyroid cytology may significantly impact the clinical management of patients with suspicious or indeterminate thyroid FNA nodules. PMID:22280184
Molecular Diagnostic Experience of Whole-Exome Sequencing in Adult Patients
Posey, Jennifer E.; Rosenfeld, Jill A.; James, Regis A.; Bainbridge, Matthew; Niu, Zhiyv; Wang, Xia; Dhar, Shweta; Wiszniewski, Wojciech; Akdemir, Zeynep H.C.; Gambin, Tomasz; Xia, Fan; Person, Richard E.; Walkiewicz, Magdalena; Shaw, Chad A.; Sutton, V. Reid; Beaudet, Arthur L.; Muzny, Donna; Eng, Christine M.; Yang, Yaping; Gibbs, Richard A.; Lupski, James R.; Boerwinkle, Eric; Plon, Sharon E.
Purpose Whole exome sequencing (WES) is increasingly used as a diagnostic tool in medicine, but prior reports focus on predominantly pediatric cohorts with neurologic or developmental disorders. We describe the diagnostic yield and characteristics of whole exome sequencing in adults. Methods We performed a retrospective analysis of consecutive WES reports for adults from a diagnostic laboratory. Phenotype composition was determined using Human Phenotype Ontology terms. Results Molecular diagnoses were reported for 17.5% (85/486) of adults, lower than a primarily pediatric population (25.2%; p=0.0003); the diagnostic rate was higher (23.9%) in those 18–30 years of age compared to patients over 30 years (10.4%; p=0.0001). Dual Mendelian diagnoses contributed to 7% of diagnoses, revealing blended phenotypes. Diagnoses were more frequent among individuals with abnormalities of the nervous system, skeletal system, head/neck, and growth. Diagnostic rate was independent of family history information, and de novo mutations contributed to 61.4% of autosomal dominant diagnoses. Conclusion Early WES experience in adults demonstrates molecular diagnoses in a substantial proportion of patients, informing clinical management, recurrence risk and recommendations for relatives. A positive family history was not predictive, consistent with molecular diagnoses often revealed by de novo events, informing the Mendelian basis of genetic disease in adults. PMID:26633545
Recent advances in the molecular diagnostics of gastric cancer
Kanda, Mitsuro; Kodera, Yasuhiro
Gastric cancer (GC) is the third most common cause of cancer-related death in the world, representing a major global health issue. Although the incidence of GC is declining, the outcomes for GC patients remain dismal because of the lack of effective biomarkers to detect early GC and predict both recurrence and chemosensitivity. Current tumor markers for GC, including serum carcinoembryonic antigen and carbohydrate antigen 19-9, are not ideal due to their relatively low sensitivity and specificity. Recent improvements in molecular techniques are better able to identify aberrant expression of GC-related molecules, including oncogenes, tumor suppressor genes, microRNAs and long non-coding RNAs, and DNA methylation, as novel molecular markers, although the molecular pathogenesis of GC is complicated by tumor heterogeneity. Detection of genetic and epigenetic alterations from gastric tissue or blood samples has diagnostic value in the management of GC. There are high expectations for molecular markers that can be used as new screening tools for early detection of GC as well as for patient stratification towards personalized treatment of GC through prediction of prognosis and drug-sensitivity. In this review, the studies of potential molecular biomarkers for GC that have been reported in the publicly available literature between 2012 and 2015 are reviewed and summarized, and certain highlighted papers are examined. PMID:26379391
A technological update of molecular diagnostics for infectious diseases
Liu, Yu-Tsueng
Identification of a causative pathogen is essential for the choice of treatment for most infectious diseases. Many FDA approved molecular assays; usually more sensitive and specific compared to traditional tests, have been developed in the last decade. A new trend of high throughput and multiplexing assays are emerging thanks to technological developments for the human genome sequencing project. The applications of microarray and ultra high throughput sequencing technologies for diagnostic microbiology are reviewed. The race for the $1000 genome technology by 2014 will have a profound impact in diagnosis and treatment of infectious diseases in the near future. PMID:18782035
Current status of diagnostic methods for henipavirus.
Tamin, A; Rota, P A
Hendra virus (HeV) and Nipah virus (NiV) are the causative agents of emerging transboundary animal disease in pigs and horses. They also cause fatal disease in humans. NiV has a case fatality rate of 40 - 100%. In the initial NiV outbreak in Malaysia in 1999, about 1.1 million pigs had to be culled. The economic impact was estimated to be approximately US$450 million. Worldwide, HeV has caused more than 60 deaths in horses with 7 human cases and 4 deaths. Since the initial outbreak, HeV spillovers from Pteropus bats to horses and humans continue. This article presents a brief review on the currently available diagnostic methods for henipavirus infections, including advances achieved since the initial outbreak, and a gap analysis of areas needing improvement. PMID:23689891
Molecular diagnostics and the public health management of legionellosis
Yates, Tom A; Bruin, Jacob P; Harrison, Timothy G; Mannes, Trish
In 2009–2010, we investigated four legionella cases notified over an 8-month period in two adjacent villages in South East England. Molecular techniques enabled us to conclude that three of the cases had distinct infections. The absence of an adequate respiratory sample in one case necessitated epidemiological investigations to exclude a potential common environmental source of further infections. One of the cases had spent a part of their incubation period in a country in South East Asia. DNA-sequence-based typing of their isolate showed it to be of the Legionella pneumophila serogroup 1 (LP1) DNA-sequence type (ST) 481. Intriguingly, the only other two ST 481 isolates in the European Working Group for Legionella Infections database were among Dutch travellers to the same country in 2003 and 2006. This case makes clear the value of molecular diagnostics and the importance of obtaining adequate clinical specimens. The potential future uses for typing data are discussed. PMID:23606383
Molecular diagnostics and the public health management of legionellosis.
Yates, Tom A; Bruin, Jacob P; Harrison, Timothy G; Mannes, Trish
In 2009-2010, we investigated four legionella cases notified over an 8-month period in two adjacent villages in South East England. Molecular techniques enabled us to conclude that three of the cases had distinct infections. The absence of an adequate respiratory sample in one case necessitated epidemiological investigations to exclude a potential common environmental source of further infections. One of the cases had spent a part of their incubation period in a country in South East Asia. DNA-sequence-based typing of their isolate showed it to be of the Legionella pneumophila serogroup 1 (LP1) DNA-sequence type (ST) 481. Intriguingly, the only other two ST 481 isolates in the European Working Group for Legionella Infections database were among Dutch travellers to the same country in 2003 and 2006. This case makes clear the value of molecular diagnostics and the importance of obtaining adequate clinical specimens. The potential future uses for typing data are discussed. PMID:23606383
Sudden unexpected death in epilepsy genetics: Molecular diagnostics and prevention.
Goldman, Alica M; Behr, Elijah R; Semsarian, Christopher; Bagnall, Richard D; Sisodiya, Sanjay; Cooper, Paul N
Epidemiologic studies clearly document the public health burden of sudden unexpected death in epilepsy (SUDEP). Clinical and experimental studies have uncovered dynamic cardiorespiratory dysfunction, both interictally and at the time of sudden death due to epilepsy. Genetic analyses in humans and in model systems have facilitated our current molecular understanding of SUDEP. Many discoveries have been informed by progress in the field of sudden cardiac death and sudden infant death syndrome. It is becoming apparent that SUDEP genomic complexity parallels that of sudden cardiac death, and that there is a pauci1ty of analytically useful postmortem material. Because many challenges remain, future progress in SUDEP research, molecular diagnostics, and prevention rests in international, collaborative, and transdisciplinary dialogue in human and experimental translational research of sudden death. PMID:26749013
Molecular Diagnostic and Drug Delivery Agents based on Aptamer-Nanomaterial Conjugates
Lee, Jung Heon; Yigit, Mehmet V.; Mazumdar, Debapriya; Lu, Yi
Recent progress in an emerging area of designing aptamer and nanomaterial conjugates as molecular diagnostic and drug delivery agents in biomedical applications is summarized. Aptamers specific for a wide range of targets are first introduced and compared to antibodies. Methods of integrating these aptamers with a variety of nanomaterials, such as gold nanoparticles, quantum dots, carbon nanotubes, and superparamagnetic iron oxide nanoparticles, each with unique optical, magnetic, and electrochemical properties, are reviewed. Applications of these systems as fluorescent, colorimetric, magnetic resonance imaging, and electrochemical sensors in medical diagnostics are given, along with new applications as smart drug delivery agents. PMID:20338204
Diagnostic methods for invasive fungal diseases in patients with hematologic malignancies
Riwes, Mary Mansour; Wingard, John R
Invasive fungal disease is associated with increased morbidity and mortality in hematologic malignancy patients and hematopoietic stem cell transplant recipients. Timely recognition and treatment of invasive fungal diseases in these patients are essential and decrease mortality. However, conventional definitive diagnostic methods are difficult and time consuming. While conventional microbiological and histopathological methods are still needed for a definitive diagnosis of invasive fungal disease, new noninvasive diagnostic methods including serologic and molecular biomarkers are now available. These new diagnostic methods facilitate an early diagnosis of invasive fungal disease and allow for utilization of a pre-emptive treatment approach, which may ultimately lead to improved treatment outcomes and reduced toxicity. PMID:23216596
Ultrasensitive microanalytical diagnostic methods for rickettsial pathogens
Hatch, A. V.
A strategic CRADA was established between Sandia National Laboratories (SNL) and the University of Texas Medical Branch (UTMB) at Galveston to address pressing needs for US protection against biological weapons of mass destruction (WMD) and emerging infectious diseases. The combination of unique expertise and facilities at UTMB and SNL enabled interdisciplinary research efforts in the development of rapid and accurate diagnostic methods for early detection of trace priority pathogen levels. Outstanding postdoctoral students were also trained at both institutions to help enable the next generation of scientists to tackle the challenging interdisciplinary problems in the area of biodefense and emerging infectious diseases. Novel approaches to diagnostics were developed and the both the speed of assays as well as the detection sensitivity were improved by over an order of magnitude compared to traditional methods. This is a significant step toward more timely and specific detection of dangerous infections. We developed in situ polymerized porous polymer monoliths that can be used as (1) size exclusion elements for capture and processing of rickettsial infected cells from a sample, (2) photopatternable framework for grafting high densities of functionalized antibodies/fluorescent particles using novel monolith chemistry. Grafting affinity reagents specific to rickettsial particles enables rapid, ultra-sensitive assays by overcoming transport limitations of traditional planar assay approaches. We have selectively trapped particles and bacteria at the cell trap and have also detected picomolar mouse IL-6 captured with only 20 minutes total incubation times using the densely patterned monolith framework. As predicted, the monolith exhibits >10x improvements in both capture speed and capture density compared to traditional planar approaches. The most significant advancements as part of this CRADA is the optimization of techniques allowing the detection of <10 rickettsial
Comparison of Diagnostic Methods for Asperger Syndrome
ERIC Educational Resources Information Center
Kopra, Kristiina; von Wendt, Lennart; Nieminen-von Wendt, Taina; Paavonen, E. Julia
Several different diagnostic sets of criteria exist for Asperger syndrome (AS), but there is no agreement on a gold standard. The aim of this study was to compare four diagnostic sets of criteria for AS: the ICD-10, the DSM-IV, the Gillberg & Gillberg, and the Szatmari criteria. The series consists of 36 children who had been referred to two…
Diagnostic Methods in Primary Ciliary Dyskinesia.
Lucas, Jane S; Paff, Tamara; Goggin, Patricia; Haarman, Eric
Diagnosing primary ciliary dyskinesia is difficult. With no reference standard, a combination of tests is needed; most tests require expensive equipment and specialist scientists. We review the advances in diagnostic testing over the past hundred years, with emphasis on recent advances. We particularly focus on use of high-speed video analysis, transmission electron microscopy, nasal nitric oxide and genetic testing. We discuss the international efforts that are in place to advance the evidence base for diagnostic tests. PMID:26362507
Applications of Molecular Diagnostic Testing in Food Allergy.
Hoffmann-Sommergruber, Karin; Pfeifer, Sabine; Bublin, Merima
IgE-mediated food allergy is a relevant health problem inducing symptoms ranging from mild local reactions up to severe life-threatening situations. Currently, no immunotherapy is available and avoidance of the incriminating food is the method of choice. Therefore, reliable diagnostic tools to formulate dietary recommendations and to avoid unnecessary exclusion diets for the individual patient are urgently needed. This review provides an update on the current knowledge on food allergens and their application in various diagnostic approaches such as skin prick test, basophil activation test, and serum IgE testing. Furthermore, these new approaches are discussed and compared to conventional extract-based assays and correlated to the gold standard of food allergy diagnosis, the double-blind placebo-controlled food challenge. Finally, the application of food allergens for preventive measurements such as allergen detection assays and the determination of threshold levels for allergen levels are discussed. PMID:26233429
Cell and molecular biology for diagnostic and therapeutic technology
NASA Astrophysics Data System (ADS)
Tan, M. I.
Our body contains 100 trillion cells. However, each cell has certain function and structure. For maintaining their integrity, cells will be collaborating with each other and with extracellular matrix surround them to form a tissue. These interactions effect internally on many networks or pathway such as signalling pathway, metabolic pathway and transport network in the cell. These networks interact with each other to maintain cell survival, cell structure and function and moreover the tissue as well as the organ which the cells built. Therefore, as part of a tissue, genetic and epigenetic abnormality of a cell can also alter these networks, and moreover disturb the function of the tissue itself. Hence, condition of genetic and epigenetic of the cell may affect other conditions in omics level such as transcriptomic, proteomic, metabolomics characteristics which can be differentiated by a particular unique molecular profile from each level, which can be used for diagnostic as well as for targeted therapy.
Application of molecular diagnostics for the detection of Lynch syndrome
Pino, Maria S; Chung, Daniel C
Lynch syndrome (LS), or hereditary nonpolyposis colorectal cancer, is the most common hereditary colorectal cancer (CRC) syndrome, accounting for approximately 2–5% of all newly diagnosed cases of CRC. Patients with LS have an increased lifetime risk of colorectal (52.2% in women and 68.7% in men) and endometrial cancer (15–70%), as well as certain extra-colonic cancers. Germline mutations in one of several DNA mismatch repair genes underlie LS. Molecular testing has emerged as an indispensable strategy for the diagnosis of LS. The diagnostic work-up of at-risk individuals includes a careful family history evaluation, microsatellite instability, immunohistochemistry and germline DNA analysis. A positive test result can guide clinicians in formulating the appropriate screening, surveillance and management strategies. However, because of the absence of an overt phenotype, such as a diffuse polyposis, it is not always straightforward to recognize LS clinically. PMID:20629513
[Molecular biology methods in immunohematology].
Tournamille, C
The molecular basis of almost all antigens of the 33 blood group systems are known. These knowledge and the advent of the PCR technology have allowed the DNA-based genotyping in order to predict the presence or absence of a blood group antigen on the cell membrane of red blood cells. DNA genotyping is required in cases where red blood cells patient cannot be used for serological typing either after a recent transfusion or because of the presence of autoantibodies on the red blood cells. Numerous DNA assays are available to detect any nucleotide polymorphism on the genes encoding blood group antigens. The technologies have improved to answer quickly to any case of transfusion emergency and to limit the risk of DNA contamination in a molecular diagnostic laboratory. Some technologies are ready for high-throughput blood group genotyping. They will be used in the future to obtain a fully typed blood group card of each donor but also to detect blood donors with rare phenotypes to register them to the Banque Nationale de Sang de Phénotype Rare (BNSPR). PMID:23587623
Platform technologies for molecular diagnostics near the patient's bedside.
Schumacher, Soeren; Lüdecke, Christine; Ehrentreich-Förster, Eva; Bier, Frank F
It is believed Lab-on-Chip systems will become a mainstream technology within the next centuries. Especially because of new findings in molecular medicine and global trends such as the changing global population in third world countries and an ageing population in industrial countries, the need for fast and reliable diagnostics is rising tremendously. Hence, diagnostics have to become more frequently and more easily available. In this regard, technologies have to be found that enable the cost-effective production and hence an affordable price. In a joint-project between seven Fraunhofer institutes a Lab-on-Chip system was developed which consists of a credit-card-sized cartridge and a base station. The cartridges contain besides the reagents necessary for a specific assay also functionalities such as pumping or heating enabling a self-contained system without any fluidic interfaces, which tend to be error-prone. Because of the modularity of the system it is possible to integrate an optical sensor as well an electrochemical sensor into the cartridge. Hence, the system can be customized to serve the needs of the particular assays. This chapter will describe the system including generic design rules for such Lab-on-Chip systems, the development of these rules into a modular Lab-on-Chip system, the integration of biomedical assays, and the production possibility of this system. PMID:23192590
Recombinase polymerase amplification: Emergence as a critical molecular technology for rapid, low-resource diagnostics.
James, Ameh; Macdonald, Joanne
Isothermal molecular diagnostics are bridging the technology gap between traditional diagnostics and polymerase chain reaction-based methods. These new techniques enable timely and accurate testing, especially in settings where there is a lack of infrastructure to support polymerase chain reaction facilities. Despite this, there is a significant lack of uptake of these technologies in developing countries where they are highly needed. Among these novel isothermal technologies, recombinase polymerase amplification (RPA) holds particular potential for use in developing countries. This rapid nucleic acid amplification approach is fast, highly sensitive and specific, and amenable to countries with a high burden of infectious diseases. Implementation of RPA technology in developing countries is critically required to assess limitations and potentials of the diagnosis of infectious disease, and may help identify impediments that prevent adoption of new molecular technologies in low resource- and low skill settings. This review focuses on approaching diagnosis of infectious disease with RPA. PMID:26517245
A molecular diagnostic approach able to detect the recurrent genetic prognostic factors typical of presenting myeloma
Boyle, Eileen M; Proszek, Paula Z; Kaiser, Martin F; Begum, Dil; Dahir, Nasrin; Savola, Suvi; Wardell, Christopher P; Leleu, Xavier; Ross, Fiona M; Chiecchio, Laura; Cook, Gordon; Drayson, Mark T; Owen, Richard G; Ashcroft, John M; Jackson, Graham H; Anthony Child, James; Davies, Faith E; Walker, Brian A; Morgan, Gareth J
Risk stratification in myeloma requires an accurate assessment of the presence of a range of molecular abnormalities including the differing IGH translocations and the recurrent copy number abnormalities that can impact clinical behavior. Currently, interphase fluorescence in situ hybridization is used to detect these abnormalities. High failure rates, slow turnaround, cost, and labor intensiveness make it difficult and expensive to use in routine clinical practice. Multiplex ligation-dependent probe amplification (MLPA), a molecular approach based on a multiplex polymerase chain reaction method, offers an alternative for the assessment of copy number changes present in the myeloma genome. Here, we provide evidence showing that MLPA is a powerful tool for the efficient detection of copy number abnormalities and when combined with expression assays, MLPA can detect all of the prognostically relevant molecular events which characterize presenting myeloma. This approach opens the way for a molecular diagnostic strategy that is efficient, high throughput, and cost effective. PMID:25287954
Electron-Beam Diagnostic Methods for Hypersonic Flow Diagnostics
NASA Technical Reports Server (NTRS)
The purpose of this work was the evaluation of the use of electron-bean fluorescence for flow measurements during hypersonic flight. Both analytical and numerical models were developed in this investigation to evaluate quantitatively flow field imaging concepts based upon the electron beam fluorescence technique for use in flight research and wind tunnel applications. Specific models were developed for: (1) fluorescence excitation/emission for nitrogen, (2) rotational fluorescence spectrum for nitrogen, (3) single and multiple scattering of electrons in a variable density medium, (4) spatial and spectral distribution of fluorescence, (5) measurement of rotational temperature and density, (6) optical filter design for fluorescence imaging, and (7) temperature accuracy and signal acquisition time requirements. Application of these models to a typical hypersonic wind tunnel flow is presented. In particular, the capability of simulating the fluorescence resulting from electron impact ionization in a variable density nitrogen or air flow provides the capability to evaluate the design of imaging instruments for flow field mapping. The result of this analysis is a recommendation that quantitative measurements of hypersonic flow fields using electron-bean fluorescence is a tractable method with electron beam energies of 100 keV. With lower electron energies, electron scattering increases with significant beam divergence which makes quantitative imaging difficult. The potential application of the analytical and numerical models developed in this work is in the design of a flow field imaging instrument for use in hypersonic wind tunnels or onboard a flight research vehicle.
Molecular separation method and apparatus
Villa-Aleman, E.
A method and apparatus are disclosed for separating a gaseous mixture of chemically identical but physically different molecules based on their polarities. The gaseous mixture of molecules is introduced in discrete quantities into the proximal end of a porous glass molecular sieve. The molecular sieve is exposed to microwaves to excite the molecules to a higher energy state from a lower energy state, those having a higher dipole moment being excited more than those with a lower energy state. The temperature of the sieve kept cold by a flow of liquid nitrogen through a cooling jacket so that the heat generated by the molecules colliding with the material is transferred away from the material. The molecules thus alternate between a higher energy state and a lower one, with the portion of molecules having the higher dipole moment favored over the others. The former portion can then be extracted separately from the distal end of the molecular sieve. 2 figs.
Molecular separation method and apparatus
Villa-Aleman, Eliel
A method and apparatus for separating a gaseous mixture of chemically identical but physically different molecules based on their polarities. The gaseous mixture of molecules is introduced in discrete quantities into the proximal end of a porous glass molecular. The molecular sieve is exposed to microwaves to excite the molecules to a higher energy state from a lower energy state, those having a higher dipole moment being excited more than those with a lower energy state. The temperature of the sieve kept cold by a flow of liquid nitrogen through a cooling jacket so that the heat generated by the molecules colliding with the material is transferred away from the material. The molecules thus alternate between a higher energy state and a lower one, with the portion of molecules having the higher dipole moment favored over the others. The former portion can then be extracted separately from the distal end of the molecular sieve.
Molecular filter-based diagnostics in high speed flows
NASA Technical Reports Server (NTRS)
Elliott, Gregory S.; Samimy, MO; Arnette, Stephen A.
The use of iodine molecular filters in nonintrusive planar velocimetry methods is examined. Detailed absorption profiles are obtained to highlight the effects that determine the profile shape. It is shown that pressure broadening induced by the presence of a nonabsorbing vapor can be utilized to significantly change the slopes bounding the absorbing region while remaining in the optically-thick regime.
Simultaneous Extraction of Viral and Bacterial Nucleic Acids for Molecular Diagnostic Applications
Kajiura, Lauren N.; Stewart, Scott D.; Dresios, John; Uyehara, Catherine F. T.
Molecular detection of microbial pathogens in clinical samples requires the application of efficient sample lysis protocols and subsequent extraction and isolation of their nucleic acids. Here, we describe a simple and time-efficient method for simultaneous extraction of genomic DNA from gram-positive and -negative bacteria, as well as RNA from viral agents present in a sample. This method compared well with existing bacterial- and viral-specialized extraction protocols, worked reliably on clinical samples, and was not pathogen specific. This method may be used to extract DNA and RNA concurrently from viral and bacterial pathogens present in a sample and effectively detect coinfections in routine clinical diagnostics. PMID:26543438
Diffusion of Molecular Diagnostic Lung Cancer Tests: A Survey of German Oncologists
Steffen, Julius Alexander
This study was aimed at examining the diffusion of diagnostic lung cancer tests in Germany. It was motivated by the high potential of detecting and targeting oncogenic drivers. Recognizing that the diffusion of diagnostic tests is a conditio sine qua non for the success of personalized lung cancer therapies, this study analyzed the diffusion of epidermal growth factor receptor (EGFR) and anaplastic lymphoma kinase (ALK) tests in Germany. Qualitative and quantitative research strategies were combined in a mixed-method design. A literature review and subsequent Key Opinion Leader interviews identified a set of qualitative factors driving the diffusion process, which were then translated into an online survey. The survey was conducted among a sample of 961 oncologists (11.34% response rate). The responses were analyzed in a multiple linear regression which identified six statistically significant factors driving the diffusion of molecular diagnostic lung cancer tests: reimbursement, attitude towards R&D, information self-assessment, perceived attitudes of colleagues, age and test-pathway strategies. Besides the important role of adequate reimbursement and relevant guidelines, the results of this study suggest that an increasing usage of test-pathway strategies, especially in an office-based setting, can increase the diffusion of molecular diagnostic lung cancer tests in the future. PMID:25562146
Method and apparatus for holographic wavefront diagnostics
Toeppen, John S.
A wavefront diagnostic apparatus has an optic and a measuring system. The optic forms a holographic image in response to a beam of light striking a hologram formed on a surface of the optic. The measuring system detects the position of the array of holographic images and compares the positions of the array of holographic images to a reference holographic image.
Method and apparatus for holographic wavefront diagnostics
Toeppen, J.S.
A wavefront diagnostic apparatus has an optic and a measuring system. The optic forms a holographic image in response to a beam of light striking a hologram formed on a surface of the optic. The measuring system detects the position of the array of holographic images and compares the positions of the array of holographic images to a reference holographic image. 3 figs.
How can molecular diagnostics contribute to the elimination of human African trypanosomiasis?
Büscher, Philippe; Deborggraeve, Stijn
A variety of molecular diagnostic tests for human African trypanosomiasis (HAT) (sleeping sickness) has been developed. Some are effectively used for research and confirmation diagnosis in travel medicine, usually following non-standardized protocols. Others have become commercially available as diagnostic kits. WHO aims to eliminate HAT as a public health problem by the year 2020, and zero transmission by the year 2030. This article gives an overview of the recent progress in molecular diagnostics for sleeping sickness, including the most recent data on test performances. Also discussed is how molecular diagnostics can play an important role in the process toward the elimination of HAT. PMID:25786994
Aircraft Engine Gas Path Diagnostic Methods: Public Benchmarking Results
NASA Technical Reports Server (NTRS)
Simon, Donald L.; Borguet, Sebastien; Leonard, Olivier; Zhang, Xiaodong (Frank)
Recent technology reviews have identified the need for objective assessments of aircraft engine health management (EHM) technologies. To help address this issue, a gas path diagnostic benchmark problem has been created and made publicly available. This software tool, referred to as the Propulsion Diagnostic Method Evaluation Strategy (ProDiMES), has been constructed based on feedback provided by the aircraft EHM community. It provides a standard benchmark problem enabling users to develop, evaluate and compare diagnostic methods. This paper will present an overview of ProDiMES along with a description of four gas path diagnostic methods developed and applied to the problem. These methods, which include analytical and empirical diagnostic techniques, will be described and associated blind-test-case metric results will be presented and compared. Lessons learned along with recommendations for improving the public benchmarking processes will also be presented and discussed.
[Diagnostic methods of nasal respiratory function].
Mlynski, G; Beule, A
Objective assessment of nasal obstruction may help with preoperative planning for rhinosurgery and indicate different aspects of endonasal pathology. To improve quality control, preoperative and postoperative objective assessment is desirable. This review presents objective functional diagnostic tools and explains their appropriate uses, the information obtained, and their limitations. An algorithm is presented for analysing nasal obstruction by means of objective functional assessment. Examples illustrate how to use this information for preoperative planning in rhinosurgery. PMID:18210011
[Hereditary stomatocytoses--diagnostic problems and their molecular basis].
Bogusławska, Dzamila M; Machnicka, Beata; Sikorski, Aleksander F
Hereditary stomatocytosis (HSt) is a group of haemolytic anaemias in which the common symptom is an increased permeability of the red cell membrane for monovalent cations. HSt is diagnosed really seldom and the difficulties in diagnosing are connected to the fact that the clinical presentation of individual subtypes of HSt is very diverse. Many cases are characterised by unique phenotypes. Nevertheless, the number of diagnosed HSt cases is increasing each year. The aim of this review was the presentation of current information and an attempt to systematize it, what might be helpful in clinical diagnostic of the new cases of this anaemia. The most frequent mistake is to classify a case of HSt as the most common haemolytic anaemia--hereditary spherocytosis (HS), in which to improve patient condition a splenectomy is often recommended. Most cases of HSt no positive response to splenectomy and often thromboembolic complications are observed. It is interesting that commonly present in blood film stomatocytes and in many cases absent or severely reduced stomatin in HSt red cell membrane are not correlated with nucleotide sequence changes of the gene encoding stomatin. Many diagnosed cases are related to mutations in SLC4A1 and RHAG genes. Extensive research carried out on HSt in the entire world will certainly permit to know the molecular basis of the disease, the diversify of its subtypes and to estimate the real incidence of HSt. PMID:20842826
The Far Infrared Lines of OH as Molecular Cloud Diagnostics
NASA Astrophysics Data System (ADS)
Smith, Howard A.; Ashby, Matt; Fischer, Jackie; Gonzales, Eduardo; Spinoglio, Luigi; Dudley, Chris
There are 15 far-IR lines arising between the lowest eight rotational levels of OH and ISO detected nine of them. Furthermore ISO found the OH lines sometimes in emission and sometimes in absorption in a wide variety of galactic and extragalactic objects ranging from AGB stars to molecular clouds to active galactic nuclei and ultra-luminous IR galaxies. This set of OH lines provides a uniquely valuable diagnostic for many reasons: the lines span a wide wavelength range (28.9μm to 163.2μm); the transitions have fast radiative rates; the abundance of the species is relatively high; the IR continuum plays an important role as a pump; the contribution from shocks is relatively minor; and not least the powerful centimeter-wave radiation from OH allows comparison with radio and VLBI datasets. The problem is that the large number of sensitive free parameters and the large optical depths of the strongest lines make modeling the full set a difficult job. We have used a modified SWAS montecarlo radiative transfer code to analyze the ISO/LWS spectra of a number of objects including in both the lines and the FIR continuum; the DUSTY radiative transfer code was used to insure a self-consistent continuum.
Gold-based hybrid nanomaterials for biosensing and molecular diagnostic applications.
Kim, Jung Eun; Choi, Ji Hye; Colas, Marion; Kim, Dong Ha; Lee, Hyukjin
The properties of gold nanomaterials are particularly of interest to many researchers, since they show unique physiochemical properties such as optical adsorption of specific wavelength of light, high electrical conductance with rich surface electrons, and facile surface modification with sulfhydryl groups. These properties have facilitated the use of gold nanomaterials in the development of various hybrid systems for biosensors and molecular diagnostics. Combined with various synthetic materials such as fluorescence dyes, polymers, oligonucleotides, graphene oxides (GO), and quantum dots (QDs), the gold-based hybrid nanomaterials offer multi-functionalities in molecular detection with high specificity and sensitivity. These two aspects result in the increase of detection speed as well as the lower detection limits, having shown that this diagnosis method is more effective than other conventional ones. In this review, we have highlighted various examples of nanomaterials for biosensing and molecular diagnostics. The gold-based hybrid systems are categorized by three distinct detection approaches, in which include (1) optical, such as surface plasmon resonance (SPR), RAMAN, and surface-enhanced Raman scattering (SERS), (2) fluorescence, such as förster resonance energy transfer (FRET) and nanomaterial surface energy transfer (NSET), and (3) electrochemical, such as potentiometic, amperometric, and conductometric. Each example provides the detailed mechanism of molecular detection as well as the supporting experimental result with the limit of detection (LOD). Lastly, future perspective on novel development of gold-based hybrid nanomaterials is discussed as well as their challenges. PMID:26894985
Diagnostic methods for CW laser damage testing
NASA Astrophysics Data System (ADS)
Stewart, Alan F.; Shah, Rashmi S.
High performance optical coatings are an enabling technology for many applications - navigation systems, telecom, fusion, advanced measurement systems of many types as well as directed energy weapons. The results of recent testing of superior optical coatings conducted at high flux levels will be presented. The diagnostics used in this type of nondestructive testing and the analysis of the data demonstrates the evolution of test methodology. Comparison of performance data under load to the predictions of thermal and optical models shows excellent agreement. These tests serve to anchor the models and validate the performance of the materials and coatings.
Pertussis: the disease and new diagnostic methods.
Friedman, R L
Bordetella pertussis, the causative agent of whooping cough, produces an acute and chronic respiratory infection in infants and young children. B. pertussis is still a major health problem of young children throughout the world even though effective immunization against whooping cough is available. While predominantly a childhood disease, it has been reported also to be a cause of persistent cough in adults. This review discusses the numerous bacterial virulence factors that may play roles in the pathogenesis of pertussis and in immunity to infection. The present problems with pertussis diagnosis, recent advances, and future prospects for new and improved rapid diagnostics tests also are explored. PMID:2906814
Numerical methods for molecular dynamics
Skeel, R.D.
This report summarizes our research progress to date on the use of multigrid methods for three-dimensional elliptic partial differential equations, with particular emphasis on application to the Poisson-Boltzmann equation of molecular biophysics. This research is motivated by the need for fast and accurate numerical solution techniques for three-dimensional problems arising in physics and engineering. In many applications these problems must be solved repeatedly, and the extremely large number of discrete unknowns required to accurately approximate solutions to partial differential equations in three-dimensional regions necessitates the use of efficient solution methods. This situation makes clear the importance of developing methods which are of optimal order (or nearly so), meaning that the number of operations required to solve the discrete problem is on the order of the number of discrete unknowns. Multigrid methods are generally regarded as being in this class of methods, and are in fact provably optimal order for an increasingly large class of problems. The fundamental goal of this research is to develop a fast and accurate numerical technique, based on multi-level principles, for the solutions of the Poisson-Boltzmann equation of molecular biophysics and similar equations occurring in other applications. An outline of the report is as follows. We first present some background material, followed by a survey of the literature on the use of multigrid methods for solving problems similar to the Poisson-Boltzmann equation. A short description of the software we have developed so far is then given, and numerical results are discussed. Finally, our research plans for the coming year are presented.
The Far Infrared Lines of OH as Molecular Cloud Diagnostics
NASA Technical Reports Server (NTRS)
Smith, Howard A.
Future IR missions should give some priority to high resolution spectroscopic observations of the set of far-IR transitions of OH. There are 15 far-IR lines arising between the lowest eight rotational levels of OH, and ISO detected nine of them. Furthermore, ISO found the OH lines, sometimes in emission and sometimes in absorption, in a wide variety of galactic and extragalactic objects ranging from AGB stars to molecular clouds to active galactic nuclei and ultra-luminous IR galaxies. The ISO/LWS Fabry-Perot resolved the 119 m doublet line in a few of the strong sources. This set of OH lines provides a uniquely important diagnostic for many reasons: the lines span a wide wavelength range (28.9 m to 163.2 m); the transitions have fast radiative rates; the abundance of the species is relatively high; the IR continuum plays an important role as a pump; the contribution from shocks is relatively minor; and, not least, the powerful centimeter-wave radiation from OH allows comparison with radio and VLBI datasets. The problem is that the large number of sensitive free parameters, and the large optical depths of the strongest lines, make modeling the full set a difficult job. The SWAS montecarlo radiative transfer code has been used to analyze the ISO/LWS spectra of a number of objects with good success, including in both the lines and the FIR continuum; the DUSTY radiative transfer code was used to insure a self-consistent continuum. Other far IR lines including those from H2O, CO, and [OI] are also in the code. The OH lines all show features which future FIR spectrometers should be able to resolve, and which will enable further refinements in the details of each cloud's structure. Some examples are given, including the case of S140, for which independent SWAS data found evidence for bulk flows.
[Diagnosis and prognosis of gliomas--current prospects of molecular diagnostics].
Haapasalo, Joonas; Hyartt, Antti; Salmi, Minja; Nordfors, Kristiina; Lahtela, Sirpa-Liisa; Kähkönen, Marketta; Helén, Pauli; Haapasalo, Hannu
Gliomas are tumors of the support cells of the brain and the most common of the primary brain tumors. Treatment of diffuse gliomas is based on surgical excision of the tumor and on radiotherapy and chemotherapy. The diagnosis is made in histopathological examination of the tumor, which today can be complemented with examinations involving molecular diagnostics. The most important new methods predicting the prognosis of glioma patients include demonstrations of the IDH mutation and the 1p/19q co-deletion. Profiling of gliomas may in the future allow tailoring of therapy in a patient-specific manner. PMID:24881141
An improved molecular diagnostic assay for canine and feline dermatophytosis.
Cafarchia, Claudia; Gasser, Robin B; Figueredo, Luciana A; Weigl, Stefania; Danesi, Patrizia; Capelli, Gioia; Otranto, Domenico
The few studies attempting to specifically characterize dermatophytes from hair samples of dogs and cats using PCR-based methodology relied on sequence-based analysis of selected genetic markers. The aim of the present investigation was to establish and evaluate a PCR-based approach employing genetic markers of nuclear DNA for the specific detection of dermatophytes on such specimens. Using 183 hair samples, we directly compared the test results of our one-step and nested-PCR assays with those based on conventional microscopy and in vitro culture techniques (using the latter as the reference method). The one step-PCR was highly accurate (AUC > 90) for the testing of samples from dogs, but only moderately accurate (AUC = 78.6) for cats. A nested-PCR was accurate (AUC = 93.6) for samples from cats, and achieved higher specificity (94.1 and 94.4%) and sensitivity (100 and 94.9%) for samples from dogs and cats, respectively. In addition, the nested-PCR allowed the differentiation of Microsporum canis from Trichophyton interdigitale (zoophilic) and geophilic dermatophytes (i.e., Microsporum gypseum or Trichophyton terrestre), which was not possible using the one step-assay. The PCRs evaluated here provide practical tools for diagnostic applications to support clinicians in initiating prompt and targeted chemotherapy of dermatophytoses. PMID:22686247
Method and system for diagnostics of apparatus
NASA Technical Reports Server (NTRS)
Gorinevsky, Dimitry (Inventor)
Proposed is a method, implemented in software, for estimating fault state of an apparatus outfitted with sensors. At each execution period the method processes sensor data from the apparatus to obtain a set of parity parameters, which are further used for estimating fault state. The estimation method formulates a convex optimization problem for each fault hypothesis and employs a convex solver to compute fault parameter estimates and fault likelihoods for each fault hypothesis. The highest likelihoods and corresponding parameter estimates are transmitted to a display device or an automated decision and control system. The obtained accurate estimate of fault state can be used to improve safety, performance, or maintenance processes for the apparatus.
Gold nanoprobe-based non-crosslinking hybridization for molecular diagnostics.
Larguinho, Miguel; Canto, Rafaela; Cordeiro, Milton; Pedrosa, Pedro; Fortuna, Andreia; Vinhas, Raquel; Baptista, Pedro V
Non-crosslinking (NCL) approaches using DNA-modified gold nanoparticles for molecular detection constitute powerful tools with potential implications in clinical diagnostics and tailored medicine. From detection of pathogenic agents to identification of specific point mutations associated with health conditions, these methods have shown remarkable versatility and simplicity. Herein, the NCL hybridization assay is broken down to the fundamentals behind its assembly and detection principle. Gold nanoparticle synthesis and derivatization is addressed, emphasizing optimal size homogeneity and conditions for maximum surface coverage, with direct implications in downstream detection. The detection principle is discussed and the advantages and drawbacks of different NCL approaches are discussed. Finally, NCL-based applications for molecular detection of clinically relevant loci and potential integration into more complex biosensing platforms, projecting miniaturization and portability are addressed. PMID:26292557
Molecular Pathology: Prognostic and Diagnostic Genomic Markers for Myeloid Neoplasms.
Kuo, Frank C
Application of next-generation sequencing (NGS) on myeloid neoplasms has expanded our knowledge of genomic alterations in this group of diseases. Genomic alterations in myeloid neoplasms are complex, heterogeneous, and not specific to a disease entity. NGS-based panel testing of myeloid neoplasms can complement existing diagnostic modalities and is gaining acceptance in the clinics and diagnostic laboratories. Prospective, randomized trials to evaluate the prognostic significance of genomic markers in myeloid neoplasms are under way in academic medical centers. PMID:27523973
Diagnostic Value of Halitosis Examination Methods.
Aydin, Murat; Bollen, Curd M L; Özen, Murat Eren
There are many methods and varied protocols for examining halitosis. Chemical and enzymatic tests determine the presence of bacterial species and their metabolic products or enzymes in the mouth, while halitometers precisely quantify gases but not halitosis itself. Examinations by the human nose (ie, self assessment, feedback from others, or organoleptic test by an examiner) directly target halitosis, however organoleptic examination alone is insufficient for a definitive diagnosis when the individual has no complaints about halitosis. The underlying reasons why patients seek consultation concerning halitosis are usually based on their own assessment and the opinion of others, even if those assessments are not correlated with oral odorous gas measurements. This article seeks to summarize findings and review methods of examining halitosis to determine their usefulness. PMID:26977897
Signal processing methods for MFE plasma diagnostics
Candy, J.V.; Casper, T.; Kane, R.
The application of various signal processing methods to extract energy storage information from plasma diamagnetism sensors occurring during physics experiments on the Tandom Mirror Experiment-Upgrade (TMX-U) is discussed. We show how these processing techniques can be used to decrease the uncertainty in the corresponding sensor measurements. The algorithms suggested are implemented using SIG, an interactive signal processing package developed at LLNL.
Sodium and T1ρ MRI for molecular and diagnostic imaging of articular cartilage†
Borthakur, Arijitt; Mellon, Eric; Niyogi, Sampreet; Witschey, Walter; Kneeland, J. Bruce; Reddy, Ravinder
In this article, both sodium magnetic resonance (MR) and T1ρ relaxation mapping aimed at measuring molecular changes in cartilage for the diagnostic imaging of osteoarthritis are reviewed. First, an introduction to structure of cartilage, its degeneration in osteoarthritis (OA) and an outline of diagnostic imaging methods in quantifying molecular changes and early diagnostic aspects of cartilage degeneration are described. The sodium MRI section begins with a brief overview of the theory of sodium NMR of biological tissues and is followed by a section on multiple quantum filters that can be used to quantify both bi-exponential relaxation and residual quadrupolar interaction. Specifically, (i) the rationale behind the use of sodium MRI in quantifying proteoglycan (PG) changes, (ii) validation studies using biochemical assays, (iii) studies on human OA specimens, (iv) results on animal models and (v) clinical imaging protocols are reviewed. Results demonstrating the feasibility of quantifying PG in OA patients and comparison with that in healthy subjects are also presented. The section concludes with the discussion of advantages and potential issues with sodium MRI and the impact of new technological advancements (e.g. ultra-high field scanners and parallel imaging methods). In the theory section on T1ρ, a brief description of (i) principles of measuring T1ρ relaxation, (ii) pulse sequences for computing T1ρ relaxation maps, (iii) issues regarding radio frequency power deposition, (iv) mechanisms that contribute to T1ρ in biological tissues and (v) effects of exchange and dipolar interaction on T1ρ dispersion are discussed. Correlation of T1ρ relaxation rate with macromolecular content and biomechanical properties in cartilage specimens subjected to trypsin and cytokine-induced glycosaminoglycan depletion and validation against biochemical assay and histopathology are presented. Experimental T1ρ data from osteoarthritic specimens, animal models, healthy human
Differential temperature integrating diagnostic method and apparatus
Doss, James D.; McCabe, Charles W.
A method and device for detecting the presence of breast cancer in women by integrating the temperature difference between the temperature of a normal breast and that of a breast having a malignant tumor. The breast-receiving cups of a brassiere are each provided with thermally conductive material next to the skin, with a thermistor attached to the thermally conductive material in each cup. The thermistors are connected to adjacent arms of a Wheatstone bridge. Unbalance currents in the bridge are integrated with respect to time by means of an electrochemical integrator. In the absence of a tumor, both breasts maintain substantially the same temperature, and the bridge remains balanced. If the tumor is present in one breast, a higher temperature in that breast unbalances the bridge and the electrochemical cells integrate the temperature difference with respect to time.
Examinations of electron temperature calculation methods in Thomson scattering diagnostics
Oh, Seungtae; Lee, Jong Ha; Wi, Hanmin
Electron temperature from Thomson scattering diagnostic is derived through indirect calculation based on theoretical model. {chi}-square test is commonly used in the calculation, and the reliability of the calculation method highly depends on the noise level of input signals. In the simulations, noise effects of the {chi}-square test are examined and scale factor test is proposed as an alternative method.
The Evolution of Advanced Molecular Diagnostics for the Detection and Characterization of Mycoplasma pneumoniae.
Diaz, Maureen H; Winchell, Jonas M
Over the past decade there have been significant advancements in the methods used for detecting and characterizing Mycoplasma pneumoniae, a common cause of respiratory illness and community-acquired pneumonia worldwide. The repertoire of available molecular diagnostics has greatly expanded from nucleic acid amplification techniques (NAATs) that encompass a variety of chemistries used for detection, to more sophisticated characterizing methods such as multi-locus variable-number tandem-repeat analysis (MLVA), Multi-locus sequence typing (MLST), matrix-assisted laser desorption ionization-time-of-flight mass spectrometry (MALDI-TOF MS), single nucleotide polymorphism typing, and numerous macrolide susceptibility profiling methods, among others. These many molecular-based approaches have been developed and employed to continually increase the level of discrimination and characterization in order to better understand the epidemiology and biology of M. pneumoniae. This review will summarize recent molecular techniques and procedures and lend perspective to how each has enhanced the current understanding of this organism and will emphasize how Next Generation Sequencing may serve as a resource for researchers to gain a more comprehensive understanding of the genomic complexities of this insidious pathogen. PMID:27014191
The Evolution of Advanced Molecular Diagnostics for the Detection and Characterization of Mycoplasma pneumoniae
Diaz, Maureen H.; Winchell, Jonas M.
Over the past decade there have been significant advancements in the methods used for detecting and characterizing Mycoplasma pneumoniae, a common cause of respiratory illness and community-acquired pneumonia worldwide. The repertoire of available molecular diagnostics has greatly expanded from nucleic acid amplification techniques (NAATs) that encompass a variety of chemistries used for detection, to more sophisticated characterizing methods such as multi-locus variable-number tandem-repeat analysis (MLVA), Multi-locus sequence typing (MLST), matrix-assisted laser desorption ionization–time-of-flight mass spectrometry (MALDI-TOF MS), single nucleotide polymorphism typing, and numerous macrolide susceptibility profiling methods, among others. These many molecular-based approaches have been developed and employed to continually increase the level of discrimination and characterization in order to better understand the epidemiology and biology of M. pneumoniae. This review will summarize recent molecular techniques and procedures and lend perspective to how each has enhanced the current understanding of this organism and will emphasize how Next Generation Sequencing may serve as a resource for researchers to gain a more comprehensive understanding of the genomic complexities of this insidious pathogen. PMID:27014191
Companion diagnostics and molecular imaging-enhanced approaches for oncology clinical trials
Van Heertum, Ronald L; Scarimbolo, Robert; Ford, Robert; Berdougo, Eli; O'Neal, Michael
In the era of personalized medicine, diagnostic approaches are helping pharmaceutical and biotechnology sponsors streamline the clinical trial process. Molecular assays and diagnostic imaging are routinely being used to stratify patients for treatment, monitor disease, and provide reliable early clinical phase assessments. The importance of diagnostic approaches in drug development is highlighted by the rapidly expanding global cancer diagnostics market and the emergent attention of regulatory agencies worldwide, who are beginning to offer more structured platforms and guidance for this area. In this paper, we highlight the key benefits of using companion diagnostics and diagnostic imaging with a focus on oncology clinical trials. Nuclear imaging using widely available radiopharmaceuticals in conjunction with molecular imaging of oncology targets has opened the door to more accurate disease assessment and the modernization of standard criteria for the evaluation, staging, and treatment responses of cancer patients. Furthermore, the introduction and validation of quantitative molecular imaging continues to drive and optimize the field of oncology diagnostics. Given their pivotal role in disease assessment and treatment, the validation and commercialization of diagnostic tools will continue to advance oncology clinical trials, support new oncology drugs, and promote better patient outcomes. PMID:26392755
A Diagnostic Assessment for Introductory Molecular and Cell Biology
ERIC Educational Resources Information Center
Shi, Jia; Wood, William B.; Martin, Jennifer M.; Guild, Nancy A.; Vicens, Quentin; Knight, Jennifer K.
We have developed and validated a tool for assessing understanding of a selection of fundamental concepts and basic knowledge in undergraduate introductory molecular and cell biology, focusing on areas in which students often have misconceptions. This multiple-choice Introductory Molecular and Cell Biology Assessment (IMCA) instrument is designed…
Molecular biology offers sensitive and expedient tools for the detection of exposure to environmental stressors. Molecular approaches provide the means for detection of the "first cellular event(s)" in response to environmental changes-specifically, immediate changes in gene expr...
Diagnostic utility of molecular investigation in extraskeletal myxoid chondrosarcoma.
Benini, Stefania; Cocchi, Stefania; Gamberi, Gabriella; Magagnoli, Giovanna; Vogel, Daniela; Ghinelli, Cristina; Righi, Alberto; Picci, Piero; Alberghini, Marco; Gambarotti, Marco
Extraskeletal myxoid chondrosarcoma is characterized by the reciprocal chromosomal translocation t(9;22) and the resultant fused gene EWS RNA-binding protein 1 and nuclear receptor subfamily 4, group A, member 3 (EWSR1-NR4A3). A second cytogenetic rearrangement t(9;17) involves the genes NR4A3 and TAF 15 RNA polymerase II, TATA box binding protein (TBP)-associated factor (TAF15). Less frequent fusion transcript variants of the NR4A3 gene, transcription factor 12 (TCF12)-NR4A3 and TRK-fused gene (TFG)-NR4A3, are associated with t(9;15) and t(9;3) respectively. The samples from 42 patients with extraskeletal myxoid chondrosarcoma were examined for the presence of EWSR1-NR4A3, TAF15-NR4A3, TCF12-NR4A3, and TFG-NR4A3 fusion transcripts by using RT-PCR. Fluorescence in situ hybridization was performed to analyze the status of EWSR1 and NR4A3 genes. The fusion transcripts were detected in 34 of 42 samples (81%); the presence of an EWSR1 or NR4A3 gene rearrangements were detected in 8 of 42 samples (19%) which had tested negative for all fusion transcripts detected by RT-PCR. Of the 34 samples evaluable for fusion transcripts, 23 yielded positive results for EWSR1-NR4A3, 10 for TAF15-NR4A3, and 1 for TCF12-NR4A3. The combination of RT-PCR and fluorescence in situ hybridization on frozen and paraffin-embedded tissue is a sensitive and specific method for molecular detection of recurrent translocations and is an important ancillary method to establish the diagnosis of extraskeletal myxoid chondrosarcoma. PMID:24508382
Are the classic diagnostic methods in mycology still state of the art?
Wiegand, Cornelia; Bauer, Andrea; Brasch, Jochen; Nenoff, Pietro; Schaller, Martin; Mayser, Peter; Hipler, Uta-Christina; Elsner, Peter
The diagnostic workup of cutaneous fungal infections is traditionally based on microscopic KOH preparations as well as culturing of the causative organism from sample material. Another possible option is the detection of fungal elements by dermatohistology. If performed correctly, these methods are generally suitable for the diagnosis of mycoses. However, the advent of personalized medicine and the tasks arising therefrom require new procedures marked by simplicity, specificity, and swiftness. The additional use of DNA-based molecular techniques further enhances sensitivity and diagnostic specificity, and reduces the diagnostic interval to 24-48 hours, compared to weeks required for conventional mycological methods. Given the steady evolution in the field of personalized medicine, simple analytical PCR-based systems are conceivable, which allow for instant diagnosis of dermatophytes in the dermatology office (point-of-care tests). PMID:27119470
Toward molecular parasitologic diagnosis: enhanced diagnostic sensitivity for filarial infections in mobile populations.
Fink, Doran L; Fahle, Gary A; Fischer, Steven; Fedorko, Daniel F; Nutman, Thomas B
The diagnosis of filarial infections among individuals residing in areas where the disease is not endemic requires both strong clinical suspicion and expert training in infrequently practiced parasitological methods. Recently developed filarial molecular diagnostic assays are highly sensitive and specific but have limited availability and have not been closely evaluated for clinical use outside populations residing in areas of endemicity. In this study, we assessed the performance of a panel of real-time PCR assays for the four most common human filarial pathogens among blood and tissue samples collected from a cohort of patients undergoing evaluation for suspected filarial infections. Compared to blood filtration, real-time PCR was equally sensitive for the detection of microfilaremia due to Wuchereria bancrofti (2 of 46 samples positive by both blood filtration and PCR with no discordant results) and Loa loa (24 of 208 samples positive by both blood filtration and PCR, 4 samples positive by PCR only, and 3 samples positive by blood filtration only). Real-time PCR of skin snip samples was significantly more sensitive than microscopic examination for the detection of Onchocerca volvulus microfiladermia (2 of 218 samples positive by both microscopy and PCR and 12 samples positive by PCR only). The molecular assays required smaller amounts of blood and tissue than conventional methods and could be performed by laboratory personnel without specialized parasitology training. Taken together, these data demonstrate the utility of the molecular diagnosis of filarial infections in mobile populations. PMID:20980560
Comprehensive Carrier Screening and Molecular Diagnostic Testing for Recessive Childhood Diseases
Kingsmore, Stephen
Of 7,028 disorders with suspected Mendelian inheritance, 1,139 are recessive and have an established molecular basis. Although individually uncommon, Mendelian diseases collectively account for ~20% of infant mortality and ~18% of pediatric hospitalizations. Molecular diagnostic testing is currently available for only ~300 recessive disorders. Preconception screening, together with genetic counseling of carriers, has resulted in remarkable declines in the incidence of several severe recessive diseases including Tay-Sachs disease and cystic fibrosis. However, extension of preconception screening and molecular diagnostic testing to most recessive disease genes has hitherto been impractical. Recently, we reported a preconception carrier screen / molecular diagnostic test for 448 recessive childhood diseases. The current status of this test is reviewed here. Currently, this reports analytical validity of the comprehensive carrier test. As the clinical validity and clinical utility in the contexts described is ascertained, this article will be updated. PMID:22872815
Comprehensive carrier screening and molecular diagnostic testing for recessive childhood diseases.
Kingsmore, Stephen
Of 7,028 disorders with suspected Mendelian inheritance, 1,139 are recessive and have an established molecular basis. Although individually uncommon, Mendelian diseases collectively account for ~20% of infant mortality and ~18% of pediatric hospitalizations. Molecular diagnostic testing is currently available for only ~300 recessive disorders. Preconception screening, together with genetic counseling of carriers, has resulted in remarkable declines in the incidence of several severe recessive diseases including Tay-Sachs disease and cystic fibrosis. However, extension of preconception screening and molecular diagnostic testing to most recessive disease genes has hitherto been impractical. Recently, we reported a preconception carrier screen / molecular diagnostic test for 448 recessive childhood diseases. The current status of this test is reviewed here. Currently, this reports analytical validity of the comprehensive carrier test. As the clinical validity and clinical utility in the contexts described is ascertained, this article will be updated. PMID:22872815
A method for knowledge acquisition in diagnostic expert system.
Li, Weishi; Li, Aiping; Li, Shudong
Knowledge acquisition plays very important role in the diagnostic expert system. It usually takes a long period to acquire disease knowledge using the traditional methods. To solve this problem, this paper describes relations between rough set theory and rule-based description of diseases, which corresponds to the process of knowledge acquisition of diagnostic expert system. Then the exclusive rules, inclusive rules and disease images of disease are built based on the PDES diagnosis model, and the definition of probability rule is put forward. At last, the paper presents the rule-based automated induction reasoning method, including exhaustive search, post-processing procedure, estimation for statistic test and the bootstrap and resampling methods. We also introduce automated induction of the rule-based description, which is used in our diseases diagnostic expert system. The experimental results not only show that rough set theory gives a very suitable framework to represent processes of uncertain knowledge extraction, but also that this method induces diagnostic rules correctly. This method can act as the assistant tool for development of diagnosis expert system, and has an extensive application in intelligent information systems. PMID:26410329
Method to directly radiolabel antibodies for diagnostic imaging and therapy
Thakur, M.L.
This patent describes a method for directly labeling proteins with radionuclides for use in diagnostic imaging and therapy. It comprises: the steps of incubating a protein-containing solution with a solution of sodium ascorbate; adding a required quantity of reduced radionuclide to the incubated protein-containing solution and incubating.
Method of azimuthally stable Mueller-matrix diagnostics of blood plasma polycrystalline films in cancer diagnostics
NASA Astrophysics Data System (ADS)
Ushenko, Yu. A.; Prysyazhnyuk, V. P.; Gavrylyak, M. S.; Gorsky, M. P.; Bachinskiy, V. T.; Vanchuliak, O. Ya.
A new information optical technique of diagnostics of the structure of polycrystalline films of blood plasma is proposed. The model of Mueller-matrix description of mechanisms of optical anisotropy of such objects as optical activity, birefringence, as well as linear and circular dichroism is suggested. The ensemble of informationally topical azimuthally stable Mueller-matrix invariants is determined. Within the statistical analysis of such parameters distributions the objective criteria of differentiation of films of blood plasma taken from healthy and patients with liver cirrhosis were determined. From the point of view of probative medicine the operational characteristics (sensitivity, specificity and accuracy) of the information-optical method of Mueller-matrix mapping of polycrystalline films of blood plasma were found and its efficiency in diagnostics of liver cirrhosis was demonstrated. Prospects of application of the method in experimental medicine to differentiate postmortem changes of the myocardial tissue was examined.
Molecular Pathology: Predictive, Prognostic, and Diagnostic Markers in Lymphoid Neoplasms.
Ho, Caleb; Kluk, Michael J
Lymphoid neoplasms show great diversity in morphology, immunophenotypic profile, and postulated cells of origin, which also reflects the variety of genetic alterations within this group of tumors. This review discusses many of the currently known genetic alterations in selected mature B-cell and T-cell lymphoid neoplasms, and their significance as diagnostic, prognostic, and therapeutic markers. Given the rapidly increasing number of genetic alterations that have been described in this group of tumors, and that the clinical significance of many is still being studied, this is not an entirely exhaustive review of all of the genetic alterations that have been reported. PMID:27523974
Integration methods for molecular dynamics
Leimkuhler, B.J.; Reich, S.; Skeel, R.D.
Classical molecular dynamics simulation of a macromolecule requires the use of an efficient time-stepping scheme that can faithfully approximate the dynamics over many thousands of timesteps. Because these problems are highly nonlinear, accurate approximation of a particular solution trajectory on meaningful time intervals is neither obtainable nor desired, but some restrictions, such as symplecticness, can be imposed on the discretization which tend to imply good long term behavior. The presence of a variety of types and strengths of interatom potentials in standard molecular models places severe restrictions on the timestep for numerical integration used in explicit integration schemes, so much recent research has concentrated on the search for alternatives that possess (1) proper dynamical properties, and (2) a relative insensitivity to the fastest components of the dynamics. We survey several recent approaches. 48 refs., 2 figs.
Toward Integrated Molecular Diagnostic System (iMDx): Principles and Applications
Park, Seung-min; Sabour, Andrew F.; Son, Jun Ho; Lee, Sang Hun
Integrated molecular diagnostic systems (iMDx), which are automated, sensitive, specific, user-friendly, robust, rapid, easy-to-use, and portable, can revolutionize future medicine. This review will first focus on the components of sample extraction, preservation, and filtration necessary for all point-of-care devices to include for practical use. Subsequently, we will look for low-powered and precise methods for both sample amplification and signal transduction, going in-depth to the details behind their principles. The final field of total device integration and its application to the clinical field will also be addressed to discuss the practicality for future patient care. We envision that microfluidic systems hold the potential to breakthrough the number of problems brought into the field of medical diagnosis today. PMID:24759281
Method to directly radiolabel antibodies for diagnostic imaging and therapy
Thakur, Mathew L.
The invention is a novel method and kit for directly radiolabeling proteins such as antibodies or antibody fragments for diagnostic and therapeutic purposes. The method comprises incubating a protein-containing solution with a solution of sodium ascorbate; adding a required quantity of reduced radionuclide to the incubated protein. A kit is also provided wherein the protein and/or reducing agents may be in lyophilized form.
Method to directly radiolabel antibodies for diagnostic imaging and therapy
Thakur, Mathew L.
The invention is a novel method and kit for directly radiolabeling proteins such as antibodies or antibody fragments for diagnostic and therapeutic purposes. The method comprises incubating a protein-containing solution with a solution of sodium ascorbate; adding a required quantity of reduced radionuclide to the incubated protein. A kit is also provided wherein the protein and/or reducing agents may be in lyophilized form.
The Utilization of Cytologic Fine-Needle Aspirates of Lung Cancer for Molecular Diagnostic Testing
Roh, Michael H.
In this era of precision medicine, our understanding and knowledge of the molecular landscape associated with lung cancer pathogenesis continues to evolve. This information is being increasingly exploited to treat advanced stage lung cancer patients with tailored, targeted therapy. During the management of these patients, minimally invasive procedures to obtain samples for tissue diagnoses are desirable. Cytologic fine-needle aspirates are often utilized for this purpose and are important not only for rendering diagnoses to subtype patients' lung cancers, but also for ascertaining molecular diagnostic information for treatment purposes. Thus, cytologic fine-needle aspirates must be utilized and triaged judiciously to achieve both objectives. In this review, strategies in utilizing fine-needle aspirates will be discussed in the context of our current understanding of the clinically actionable molecular aberrations underlying non-small cell lung cancer and the molecular assays applied to these samples in order to obtain treatment-relevant molecular diagnostic information. PMID:26076721
Disorders of sex development: effect of molecular diagnostics.
Achermann, John C; Domenice, Sorahia; Bachega, Tania A S S; Nishi, Mirian Y; Mendonca, Berenice B
Disorders of sex development (DSDs) are a diverse group of conditions that can be challenging to diagnose accurately using standard phenotypic and biochemical approaches. Obtaining a specific diagnosis can be important for identifying potentially life-threatening associated disorders, as well as providing information to guide parents in deciding on the most appropriate management for their child. Within the past 5 years, advances in molecular methodologies have helped to identify several novel causes of DSDs; molecular tests to aid diagnosis and genetic counselling have now been adopted into clinical practice. Occasionally, genetic profiling of embryos prior to implantation as an adjunct to assisted reproduction, prenatal diagnosis of at-risk pregnancies and confirmatory testing of positive results found during newborn biochemical screening are performed. Of the available genetic tests, the candidate gene approach is the most popular. New high-throughput DNA analysis could enable a genetic diagnosis to be made when the aetiology is unknown or many differential diagnoses are possible. Nonetheless, concerns exist about the use of genetic tests. For instance, a diagnosis is not always possible even using new molecular approaches (which can be worrying for the parents) and incidental information obtained during the test might cause anxiety. Careful selection of the genetic test indicated for each condition remains important for good clinical practice. The purpose of this Review is to describe advances in molecular biological techniques for diagnosing DSDs. PMID:25942653
Workshop on molecular methods for genetic diagnosis. Final technical report
Rinchik, E.M.
The Sarah Lawrence College Human Genetics Program received Department of Energy funding to offer a continuing medical education workshop for genetic counselors in the New York metropolitan area. According to statistics from the National Society of Genetic Counselors, there are approximately 160 genetic counselors working in the tri-state area (New York, New Jersey, and Connecticut), and many of them had been working in the field for more than 10 years. Thus, there was a real need to offer these counselors an in-depth opportunity to learn the specifics of the major advances in molecular genetics, and, in particular, the new approaches to diagnostic testing for genetic disease. As a result of the DOE Award DE-FG02-95ER62048 ($20,583), in July 1995 we offered the {open_quotes}Workshop on Molecular Methods for Genetic Diagnosis{close_quotes} for 24 genetic counselors in the New York metropolitan area. The workshop included an initial review session on the basics of molecular biology, lectures and discussions on past and current topics in molecular genetics and diagnostic procedures, and, importantly, daily laboratory exercises. Each counselor gained not only background, but also firsthand experience, in the major techniques of biochemical and molecular methods for diagnosing genetic diseases as well as in mathematical and computational techniques involved in human genetics analyses. Our goal in offering this workshop was not to make genetic counselors experts in these laboratory diagnostic techniques, but to acquaint them, by hands-on experience, about some of the techniques currently in use. We also wanted to provide them a technical foundation upon which they can understand and appreciate new technical developments arising in the near future.
[The comparative characteristic of methods of laboratory diagnostic of opisthorchiasis].
Starostina, O Iu; Paniushkina, I I
The analysis of samples of blood serums and copromaterial from patients was carried out to evaluate effectiveness of diagnostic of opisthorchiasis invasion. The Kato-Miura technique of thick film under cellophane layer and sedimentation technique of acetic etheric precipitation were applied for parasitologic diagnostic. The technique of immunoenzymometric analysis was applied for serological diagnostic. The analysis of detection rate of eggs of opistorchis in copromaterial and level of anti-opistorchis antibodies in samples of blood serum of patients demonstrated the presence of strong direct correlation relationship between these indicators. It is demonstrated that for laboratory diagnostic of opisthorchiasis it is impossible to limit oneself to some single technique. It is established that on the territories with middle and low level of population infection rate of opistorchis it is appropriate to apply complex of methods including analysis of blood serum for presence of specific immunoglobulins to antigens of opistorchis and twice or thrice analysis of feces with copro-ovoscopic methods. PMID:25080800
Role of Serologic and Molecular Diagnostic Assays in Identification and Management of Hepatitis C Virus Infection
Talal, Andrew; Coller, Kelly; Steinhart, Corklin; Hackett, John; Dawson, George; Rockstroh, Juergen; Feld, Jordan
The drugs available for the treatment of hepatitis C virus (HCV) have evolved to provide shorter treatment duration and higher rates of sustained virologic response (SVR), and the role of HCV infection diagnostic tests has had to evolve in order to meet changing clinical needs. This review gives an overview on the role of HCV infection diagnostic testing (molecular and serological tools) used in the diagnosis and management of HCV infection. All of this critical information guides physician decisions to optimize patient clinical outcomes. Also discussed is the future direction of diagnostic testing in the context of further advances in drug development. PMID:26659219
Management of polysensitized patient: from molecular diagnostics to biomolecular immunotherapy.
Ciprandi, Giorgio; Incorvaia, Cristoforo; Frati, Franco
A panel of Italian allergists gathered to discuss the issue concerning the management of polysensitized patients. The main conclusions were as follows: polysensitization is a relevant clinical characteristic as it affects about 70-80% of the global allergic population; the diagnostic pathway needs the use of an adequate and thorough methodology, based on the demonstration of consistency between history and documented sensitization; polysensitization and polyallergy are not synonymous: true allergy should always be demonstrated; polysensitization does not constitute a limitation to allergen immunotherapy prescription, as 1-2 allergen extracts could be effective in polysensitized patients; the allergen immunotherapy product characteristics should include the following: high efficacy and optimal safety profile, standardized production, and documented presence and titration of the major allergen. PMID:26144241
Application of statistical process control to qualitative molecular diagnostic assays.
O'Brien, Cathal P; Finn, Stephen P
Modern pathology laboratories and in particular high throughput laboratories such as clinical chemistry have developed a reliable system for statistical process control (SPC). Such a system is absent from the majority of molecular laboratories and where present is confined to quantitative assays. As the inability to apply SPC to an assay is an obvious disadvantage this study aimed to solve this problem by using a frequency estimate coupled with a confidence interval calculation to detect deviations from an expected mutation frequency. The results of this study demonstrate the strengths and weaknesses of this approach and highlight minimum sample number requirements. Notably, assays with low mutation frequencies and detection of small deviations from an expected value require greater sample numbers to mitigate a protracted time to detection. Modeled laboratory data was also used to highlight how this approach might be applied in a routine molecular laboratory. This article is the first to describe the application of SPC to qualitative laboratory data. PMID:25988159
Technology Transfer Automated Retrieval System (TEKTRAN)
In addition to the 'one-step' species-specific molecular diagnostic ISSR-PCR DNA fingerprinting method, we developed an additional 'one-step' molecular diagnostic marker 'gmtx' toward Gonatocerus morrilli (Howard) designed toward the ribosomal internal transcribed spacer region 2 (ITS2) to aid in mo...
Computational methods for molecular docking
Klebe, G.; Lengauer, T.
This tutorial was one of eight tutorials selected to be presented at the Third International Conference on Intelligent Systems for Molecular Biology which was held in the United Kingdom from July 16 to 19, 1995. Recently, it has been demonstrated that the knowledge of the three-dimensional structure of the protein can be used to derive new protein ligands with improved binding properties. This tutorial focuses on the following questions: What is its binding affinity toward a particular receptor? What are putative conformations of a ligand at the binding site? What are the similarities of different ligands in terms of their recognition capabilities? Where and in which orientation will a ligand bind to the active site? How is a new putative protein ligand selected? An overview is presented of the algorithms which are presently used to handle and predict protein-ligand interactions and to dock small molecule ligands into proteins.
Patenting genetic diagnostic methods: NGS, GWAS, SNPs and patents.
Lawson, Charles
This article reviews the problems posed by patent claims to genetic diagnostic methods associated with genome-wide association studies (GWAS) adopting methodologies using next-generation sequencing (NGS) and single nucleotide polymorphisms (SNP). These problems are essentially about experimental reproducibility and the credibility and veracity of reported developments. An analysis of the relevant law demonstrates that the current Australian and United States laws about suitable patentable subject matter differ, and that the current reproducibility (sufficiency, enablement and inutility) standards are unlikely to address these problems. The article concludes that following the United States approach excluding these genetic diagnostic method claims from patenting is one solution. Failing this, improving analysis and quality controls that are now being adopted in the basic research will reduce the nature of the problems, although this will remain problematic for patent examiners and the broader public. PMID:26349382
Molecular Diagnostics in the Evaluation of Pancreatic Cysts.
Theisen, Brian K; Wald, Abigail I; Singhi, Aatur D
Within the past few decades, there has been a dramatic increase in the detection of incidental pancreatic cysts. It is reported a pancreatic cyst is identified in up to 2.6% of abdominal scans. Many of these cysts, including serous cystadenomas and pseudocysts, are benign and can be monitored clinically. In contrast, mucinous cysts, which include intraductal papillary mucinous neoplasms and mucinous cystic neoplasms, have the potential to progress to pancreatic adenocarcinoma. In this review, we discuss the current management guidelines for pancreatic cysts, their underlying genetics, and the integration of molecular testing in cyst classification and prognostication. PMID:27523971
Molecular subtyping methods for campylobacter
Technology Transfer Automated Retrieval System (TEKTRAN)
Campylobacter jejuni is a major cause worldwide of foodborne bacterial gastroenteritis. The continued development of more effective and informative typing methods is necessary to improve our understanding of the epidemiology and population dynamics of this important pathogen. Comparative genome ind...
Optical diagnostics for condensed-phase shock-compressed molecular systems
Schmidt, S.C.; Moore, D.S.; Shaner, J.W.
Experimental techniques capable of obtaining information about the molecular phenomenology in the region through and immediately behind the shockfront during the shock-compression of condensed-phase molecular systems are discussed and compared. Difficulties associated with performing measurements in this region are briefly reviewed. Some concomitant static experiments that can be used to complement the dynamic measurements are suggested. Developments and advances expected in diagnostic techniques during the next few years are summarized.
Optimising molecular diagnostic capacity for effective control of tuberculosis in high-burden settings.
Sabiiti, W; Mtafya, B; Kuchaka, D; Azam, K; Viegas, S; Mdolo, A; Farmer, E C W; Khonga, M; Evangelopoulos, D; Honeyborne, I; Rachow, A; Heinrich, N; Ntinginya, N E; Bhatt, N; Davies, G R; Jani, I V; McHugh, T D; Kibiki, G; Hoelscher, M; Gillespie, S H
The World Health Organization's 2035 vision is to reduce tuberculosis (TB) associated mortality by 95%. While low-burden, well-equipped industrialised economies can expect to see this goal achieved, it is challenging in the low- and middle-income countries that bear the highest burden of TB. Inadequate diagnosis leads to inappropriate treatment and poor clinical outcomes. The roll-out of the Xpert(®) MTB/RIF assay has demonstrated that molecular diagnostics can produce rapid diagnosis and treatment initiation. Strong molecular services are still limited to regional or national centres. The delay in implementation is due partly to resources, and partly to the suggestion that such techniques are too challenging for widespread implementation. We have successfully implemented a molecular tool for rapid monitoring of patient treatment response to anti-tuberculosis treatment in three high TB burden countries in Africa. We discuss here the challenges facing TB diagnosis and treatment monitoring, and draw from our experience in establishing molecular treatment monitoring platforms to provide practical insights into successful optimisation of molecular diagnostic capacity in resource-constrained, high TB burden settings. We recommend a holistic health system-wide approach for molecular diagnostic capacity development, addressing human resource training, institutional capacity development, streamlined procurement systems, and engagement with the public, policy makers and implementers of TB control programmes. PMID:27393531
Alternative Confidence Interval Methods Used in the Diagnostic Accuracy Studies
Gülhan, Orekıcı Temel
Background/Aim. It is necessary to decide whether the newly improved methods are better than the standard or reference test or not. To decide whether the new diagnostics test is better than the gold standard test/imperfect standard test, the differences of estimated sensitivity/specificity are calculated with the help of information obtained from samples. However, to generalize this value to the population, it should be given with the confidence intervals. The aim of this study is to evaluate the confidence interval methods developed for the differences between the two dependent sensitivity/specificity values on a clinical application. Materials and Methods. In this study, confidence interval methods like Asymptotic Intervals, Conditional Intervals, Unconditional Interval, Score Intervals, and Nonparametric Methods Based on Relative Effects Intervals are used. Besides, as clinical application, data used in diagnostics study by Dickel et al. (2010) has been taken as a sample. Results. The results belonging to the alternative confidence interval methods for Nickel Sulfate, Potassium Dichromate, and Lanolin Alcohol are given as a table. Conclusion. While preferring the confidence interval methods, the researchers have to consider whether the case to be compared is single ratio or dependent binary ratio differences, the correlation coefficient between the rates in two dependent ratios and the sample sizes. PMID:27478491
Approaches to uncovering cancer diagnostic and prognostic molecular signatures
Hong, Shengjun; Huang, Yi; Cao, Yaqiang; Chen, Xingwei; Han, Jing-Dong J
The recent rapid development of high-throughput technology enables the study of molecular signatures for cancer diagnosis and prognosis at multiple levels, from genomic and epigenomic to transcriptomic. These unbiased large-scale scans provide important insights into the detection of cancer-related signatures. In addition to single-layer signatures, such as gene expression and somatic mutations, integrating data from multiple heterogeneous platforms using a systematic approach has been proven to be particularly effective for the identification of classification markers. This approach not only helps to uncover essential driver genes and pathways in the cancer network that are responsible for the mechanisms of cancer development, but will also lead us closer to the ultimate goal of personalized cancer therapy. PMID:27308330
Digital diffraction analysis enables low-cost molecular diagnostics on a smartphone
Im, Hyungsoon; Castro, Cesar M.; Shao, Huilin; Liong, Monty; Song, Jun; Pathania, Divya; Fexon, Lioubov; Min, Changwook; Avila-Wallace, Maria; Zurkiya, Omar; Rho, Junsung; Magaoay, Brady; Tambouret, Rosemary H.; Pivovarov, Misha; Weissleder, Ralph; Lee, Hakho
The widespread distribution of smartphones, with their integrated sensors and communication capabilities, makes them an ideal platform for point-of-care (POC) diagnosis, especially in resource-limited settings. Molecular diagnostics, however, have been difficult to implement in smartphones. We herein report a diffraction-based approach that enables molecular and cellular diagnostics. The D3 (digital diffraction diagnosis) system uses microbeads to generate unique diffraction patterns which can be acquired by smartphones and processed by a remote server. We applied the D3 platform to screen for precancerous or cancerous cells in cervical specimens and to detect human papillomavirus (HPV) DNA. The D3 assay generated readouts within 45 min and showed excellent agreement with gold-standard pathology or HPV testing, respectively. This approach could have favorable global health applications where medical access is limited or when pathology bottlenecks challenge prompt diagnostic readouts. PMID:25870273
78 FR 21128 - Molecular Diagnostic Instruments With Combined Functions; Draft Guidance for Industry and Food...
Federal Register 2010, 2011, 2012, 2013, 2014
... draft guidance document provides industry and Agency staff with FDA's current thinking on regulation of...-resistant Staphylococcus aureus, Hepatitis C virus, and genetic markers of cystic fibrosis. These types of... meant to communicate FDA's policy regarding molecular diagnostic instruments with combined...
Pathogen Inactivating Properties and Increased Sensitivity in Molecular Diagnostics by PAXgene, a Novel Non-Crosslinking Tissue Fixative
Loibner, Martina; Buzina, Walter; Viertler, Christian; Groelz, Daniel; Hausleitner, Anja; Siaulyte, Gintare; Kufferath, Iris; Kölli, Bettina; Zatloukal, Kurt
Background Requirements on tissue fixatives are getting more demanding as molecular analysis becomes increasingly relevant for routine diagnostics. Buffered formaldehyde in pathology laboratories for tissue fixation is known to cause chemical modifications of biomolecules which affect molecular testing. A novel non-crosslinking tissue preservation technology, PAXgene Tissue (PAXgene), was developed to preserve the integrity of nucleic acids in a comparable way to cryopreservation and also to preserve morphological features comparable to those of formalin fixed samples. Methods Because of the excellent preservation of biomolecules by PAXgene we investigated its pathogen inactivation ability and biosafety in comparison to formalin by in-vitro testing of bacteria, human relevant fungi and human cytomegalovirus (CMV). Guidelines for testing disinfectants served as reference for inactivation assays. Furthermore, we tested the properties of PAXgene for detection of pathogens by PCR based assays. Results All microorganisms tested were similarly inactivated by PAXgene and formalin except Clostridium sporogenes, which remained viable in seven out of ten assays after PAXgene treatment and in three out of ten assays after formalin fixation. The findings suggest that similar biosafety measures can be applied for PAXgene and formalin fixed samples. Detection of pathogens in PCR-based diagnostics using two CMV assays resulted in a reduction of four to ten quantification cycles of PAXgene treated samples which is a remarkable increase of sensitivity. Conclusion PAXgene fixation might be superior to formalin fixation when molecular diagnostics and highly sensitive detection of pathogens is required in parallel to morphology assessment. PMID:26974150
New diagnostic methods for laser plasma- and microwave-enhanced combustion.
Miles, Richard B; Michael, James B; Limbach, Christopher M; McGuire, Sean D; Chng, Tat Loon; Edwards, Matthew R; DeLuca, Nicholas J; Shneider, Mikhail N; Dogariu, Arthur
The study of pulsed laser- and microwave-induced plasma interactions with atmospheric and higher pressure combusting gases requires rapid diagnostic methods that are capable of determining the mechanisms by which these interactions are taking place. New rapid diagnostics are presented here extending the capabilities of Rayleigh and Thomson scattering and resonance-enhanced multi-photon ionization (REMPI) detection and introducing femtosecond laser-induced velocity and temperature profile imaging. Spectrally filtered Rayleigh scattering provides a method for the planar imaging of temperature fields for constant pressure interactions and line imaging of velocity, temperature and density profiles. Depolarization of Rayleigh scattering provides a measure of the dissociation fraction, and multi-wavelength line imaging enables the separation of Thomson scattering from Rayleigh scattering. Radar REMPI takes advantage of high-frequency microwave scattering from the region of laser-selected species ionization to extend REMPI to atmospheric pressures and implement it as a stand-off detection method for atomic and molecular species in combusting environments. Femtosecond laser electronic excitation tagging (FLEET) generates highly excited molecular species and dissociation through the focal zone of the laser. The prompt fluorescence from excited molecular species yields temperature profiles, and the delayed fluorescence from recombining atomic fragments yields velocity profiles. PMID:26170432
New diagnostic methods for laser plasma- and microwave-enhanced combustion
Miles, Richard B; Michael, James B; Limbach, Christopher M; McGuire, Sean D; Chng, Tat Loon; Edwards, Matthew R; DeLuca, Nicholas J; Shneider, Mikhail N; Dogariu, Arthur
The study of pulsed laser- and microwave-induced plasma interactions with atmospheric and higher pressure combusting gases requires rapid diagnostic methods that are capable of determining the mechanisms by which these interactions are taking place. New rapid diagnostics are presented here extending the capabilities of Rayleigh and Thomson scattering and resonance-enhanced multi-photon ionization (REMPI) detection and introducing femtosecond laser-induced velocity and temperature profile imaging. Spectrally filtered Rayleigh scattering provides a method for the planar imaging of temperature fields for constant pressure interactions and line imaging of velocity, temperature and density profiles. Depolarization of Rayleigh scattering provides a measure of the dissociation fraction, and multi-wavelength line imaging enables the separation of Thomson scattering from Rayleigh scattering. Radar REMPI takes advantage of high-frequency microwave scattering from the region of laser-selected species ionization to extend REMPI to atmospheric pressures and implement it as a stand-off detection method for atomic and molecular species in combusting environments. Femtosecond laser electronic excitation tagging (FLEET) generates highly excited molecular species and dissociation through the focal zone of the laser. The prompt fluorescence from excited molecular species yields temperature profiles, and the delayed fluorescence from recombining atomic fragments yields velocity profiles. PMID:26170432
[Clinical importance and diagnostic methods of minimal hepatic encephalopathy].
Stawicka, Agnieszka; Zbrzeźniak, Justyna; Świderska, Aleksandra; Kilisińska, Natalia; Świderska, Magdalena; Jaroszewicz, Jerzy; Flisiak, Robert
Minimal hepatic encephalopathy (MHE) encompasses a number of neuropsychological and neurophysiological disorders in patients suffering from liver cirrhosis, who do not display abnormalities during a medical interview or physical examination. A negative influence of MHE on the quality of life of patients suffering from liver cirrhosis was confirmed, which include retardation of ability of operating motor vehicles and disruption of multiple health-related areas, as well as functioning in the society. The data on frequency of traffic offences and accidents amongst patients diagnosed with MHE in comparison to patients diagnosed with liver cirrhosis without MHE, as well as healthy persons is alarming. Those patients are unaware of their disorder and retardation of their ability to operate vehicles, therefore it is of utmost importance to define this group. The term minimal hepatic encephalopathy (formerly "subclinical" encephalopathy) erroneously suggested the unnecessity of diagnostic and therapeutic procedures in patients with liver cirrhosis. Diagnosing MHE is an important predictive factor for occurrence of overt encephalopathy - more than 50% of patients with this diagnosis develop overt encephalopathy during a period of 30 months after. Early diagnosing MHE gives a chance to implement proper treatment which can be a prevention of overt encephalopathy. Due to continuing lack of clinical research there exist no commonly agreed-upon standards for definition, diagnostics, classification and treatment of hepatic encephalopathy. This article introduces the newest findings regarding the importance of MHE, scientific recommendations and provides detailed descriptions of the most valuable diagnostic methods. PMID:27000818
Post- and prenatal diagnostic methods for the homocystinurias.
Fowler, B; Jakobs, C
Diagnosis of the homozygous homocystinurias can be performed by investigations at the metabolite, enzyme and DNA level. The existence of variant forms due to the wide range of genetic variation may result in only small differences in various parameters between controls and affected subjects. 1. Sulphur amino acid concentrations in plasma, especially total homocysteine, are useful in first line diagnostic investigations. 2. Cystathionine-beta-synthase (CBS), methylenetetrahydrofolate reductase (MTHFR) and methylfolate homocysteine methyltransferase (MFMT) can be directly assayed in many tissues including fibroblasts (each) and blood cells (except CBS). Indirect whole cell assays which measure pathway activity dependent on a particular enzyme can provide useful diagnostic information. 3. Direct analysis of mutations is available for CBS, MTHFR and recently also for MFMT deficiencies. However the existence of a larger number of very rare, often private, mutations limits the usefulness of this approach in routine diagnosis. The above diagnostic approaches can generally be applied to prenatal diagnosis. Measurement of methylmalonic acid and other metabolites in amniotic fluid by stable isotope dilution / gas chromatography-mass spectrometry is well established for the methylmalonic acidurias. This method has also been applied to combined homocystinuria/methylmalonic aciduria supported by enzyme assays in cultured cells. Total homocysteine measurement in cell free amniotic fluid is also possible, performed so far in 14 cases with two affected fetuses. The indirect assay of methionine formation from [14C] labelled formate in intact cultured amniotic fluid cells has been for prenatal diagnosis of the remethylation defects. PMID:9587033
Nonparametric Methods in Molecular Biology
Wittkowski, Knut M.; Song, Tingting
In 2003, the completion of the Human Genome Project[1] together with advances in computational resources[2] were expected to launch an era where the genetic and genomic contributions to many common diseases would be found. In the years following, however, researchers became increasingly frustrated as most reported 'findings' could not be replicated in independent studies[3]. To improve the signal/noise ratio, it was suggested to increase the number of cases to be included to tens of thousands[4], a requirement that would dramatically restrict the scope of personalized medicine. Similarly, there was little success in elucidating the gene–gene interactions involved in complex diseases or even in developing criteria for assessing their phenotypes. As a partial solution to these enigmata, we here introduce a class of statistical methods as the 'missing link' between advances in genetics and informatics. As a first step, we provide a unifying view of a plethora of non-parametric tests developed mainly in the 1940s, all of which can be expressed as u-statistics. Then, we will extend this approach to reflect categorical and ordinal relationships between variables, resulting in a flexible and powerful approach to deal with the impact of (1) multi-allelic genetic loci, (2) poly-locus genetic regions, and (3) oligo-genetic and oligo-genomic collaborative interactions on complex phenotypes. PMID:20652502
[Comparative diagnostic value of Helicobacter pylori infection testing methods].
Girdalidze, A M; Elisabedashvili, G V; Sharvadze, L G; Dzhorbenadze, T A
In 213 patients with gastric and duodenal pathology, including received surgery, comparative estimation of results of Helicobacter pylori (Hp) infection testing with invasive and noninvasive methods, were performed. Material for invasive endoscopic biopsy test (EBT), including rapid urease test (RUT) for rapid Hp identification by determination of urease activity, smear cytology and histology was extracted on endoscopy or intraoperationally. RUT was carried out with the help of URE-HP test kit. Serological test for Hp antibodies IgG and IgA class was performed by IFA using kit ELISA. 13С urea breath test (UBT) was made by determination of 13/12CO2 in breath samples on infrared spectroscope. Based on 5 different methods of Hp infection testing Hp positivity in 172 (80,8%) and Hp negativity in 41 (19,2%) from 213 examined patients was revealed. 13С-UBT revealed the highest diagnostic value (accuracy-97,5%, sensibility-97,0%, specificity-100%) in Hp infection diagnostics. In treatment efficiency control this parameters of 13С-UBT are also much high (96,7%, 90,0% and 100% respectively). In spite of high sensitivity of serological test (100%), it had comparative low specificity (71,0%) with high probability of false positive results in treated patients (antibodies titer to Hp after eradication retains for a long time). Thought, this test may be successfully used only in primary patients and in epidemiological studies. Among three methods of EBT, Hp infection detection with RUT revealed the best results (accuracy-94,8%, sensibility-95,0%, specificity-100%). Correlation of RUT and UBT results and much higher diagnostic value of UBT, necessitate RUT with histological study of stomach body mucosa to perform in patients over 45 year, with prolonged anamnesis and dangerous symptoms of disease. In Hp positive patients correlation of index DOB‰ of breath test with results of RUT was revealed. This can serve the index DOB‰ of 13С-UBT as a marker of Hp infection rate. The
Real-time caries diagnostics by optical PNC method
NASA Astrophysics Data System (ADS)
Masychev, Victor I.; Alexandrov, Michail T.
The results of hard tooth tissues research by the optical PNC- method in experimental and clinical conditions are presented. In the experiment under 90 test-sample of tooth slices with thickness about 1mm (enamel, dentine and cement) were researched. The results of the experiment were processed by the method of correlation analyze. Clinical researches were executed on teeth of 210 patients. The regions of tooth tissue diseases with initial, moderate and deep caries were investigated. Spectral characteristics of intact and pathologically changed tooth tissues are presented and their peculiar features are discussed. The results the optical PNC-method application while processing tooth carious cavities are presented in order to estimate efficiency of the mechanical and antiseptic processing of teeth. It is revealed that the PNC-method can be sued as for differential diagnostics of a degree dental carious stage, as for estimating of carefulness of tooth cavity processing before filling.
The Changing Landscape of Molecular Diagnostic Testing: Implications for Academic Medical Centers
Rehm, Heidi L.; Hynes, Elizabeth; Funke, Birgit H.
Over the last decade, the field of molecular diagnostics has undergone tremendous transformation, catalyzed by the clinical implementation of next generation sequencing (NGS). As technical capabilities are enhanced and current limitations are addressed, NGS is increasingly capable of detecting most variant types and will therefore continue to consolidate and simplify diagnostic testing. It is likely that genome sequencing will eventually serve as a universal first line test for disorders with a suspected genetic origin. Academic Medical Centers (AMCs), which have been at the forefront of this paradigm shift are now presented with challenges to keep up with increasing technical, bioinformatic and interpretive complexity of NGS-based tests in a highly competitive market. Additional complexity may arise from altered regulatory oversight, also triggered by the unprecedented scope of NGS-based testing, which requires new approaches. However, these challenges are balanced by unique opportunities, particularly at the interface between clinical and research operations, where AMCs can capitalize on access to cutting edge research environments and establish collaborations to facilitate rapid diagnostic innovation. This article reviews present and future challenges and opportunities for AMC associated molecular diagnostic laboratories from the perspective of the Partners HealthCare Laboratory for Molecular Medicine (LMM). PMID:26828522
Assessing value of innovative molecular diagnostic tests in the concept of predictive, preventive, and personalized medicine.
Akhmetov, Ildar; Bubnov, Rostyslav V
Molecular diagnostic tests drive the scientific and technological uplift in the field of predictive, preventive, and personalized medicine offering invaluable clinical and socioeconomic benefits to the key stakeholders. Although the results of diagnostic tests are immensely influential, molecular diagnostic tests (MDx) are still grudgingly reimbursed by payers and amount for less than 5 % of the overall healthcare costs. This paper aims at defining the value of molecular diagnostic test and outlining the most important components of "value" from miscellaneous assessment frameworks, which go beyond accuracy and feasibility and impact the clinical adoption, informing healthcare resource allocation decisions. The authors suggest that the industry should facilitate discussions with various stakeholders throughout the entire assessment process in order to arrive at a consensus about the depth of evidence required for positive marketing authorization or reimbursement decisions. In light of the evolving "value-based healthcare" delivery practices, it is also recommended to account for social and ethical parameters of value, since these are anticipated to become as critical for reimbursement decisions and test acceptance as economic and clinical criteria. PMID:26425215
Cystic Fibrosis: A Review of Associated Phenotypes, Use of Molecular Diagnostic Approaches, Genetic Characteristics, Progress, and Dilemmas.
Brennan, Marie-Luise; Schrijver, Iris
Cystic fibrosis (CF) is an autosomal recessive disease with significant associated morbidity and mortality. It is now appreciated that the broad phenotypic CF spectrum is not explained by obvious genotype-phenotype correlations, suggesting that CF transmembrane conductance regulator (CFTR)-related disease may occur because of multiple additive effects. These contributing effects include complex CFTR alleles, modifier genes, mutations in alternative genes that produce CF-like phenotypes, epigenetic factors, and environmental influences. Most patients in the United States are now diagnosed through newborn screening and use of molecular testing methods. We review the molecular testing approaches and laboratory guidelines for carrier screening, prenatal testing, newborn screening, and clinical diagnostic testing, as well as recent developments in CF treatment, and reasons for the lack of a molecular diagnosis in some patients. PMID:26631874
DNA markers in molecular diagnostics for hepatocellular carcinoma
Su, Ying-Hsiu; Lin, Selena Y; Song, Wei; Jain, Surbhi
Hepatocellular carcinoma (HCC) is the one of the leading causes of cancer mortality in the world, mainly due to the difficulty of early detection and limited therapeutic options. The implementation of HCC surveillance programs in well-defined, high-risk populations were only able to detect about 40–50% of HCC at curative stages (Barcelona Clinic Liver Cancer stages 0 & 1) due to the low sensitivities of the current screening methods. The advance of sequencing technologies has identified numerous modifications as potential candidate DNA markers for diagnosis/surveillance. Here we aim to provide an overview of the DNA alterations that result in activation of cancer pathways known to potentially drive HCC carcinogenesis and to summarize performance characteristics of each DNA marker in the periphery (blood or urine) for HCC screening. PMID:25098554
Array-on-a-disk? How Blu-ray technology can be applied to molecular diagnostics.
Morais, Sergi; Tortajada-Genaro, Luis; Maquieira, Angel
This editorial comments on the balance and perspectives of compact disk technology applied to molecular diagnostics. The development of sensitive, rapid and multiplex assays using Blu-ray technology for the determination of biomarkers, drug allergens, pathogens and detection of infections would have a direct impact on diagnostics. Effective tests for use in clinical, environmental and food applications require versatile and low-cost platforms as well as cost-effective detectors. Blu-ray technology accomplishes those requirements and advances on the concept of high density arrays for massive screening to achieve the demands of point of care or in situ analysis. PMID:24946682
Evaluating the accuracy of molecular diagnostic testing for canine visceral leishmaniasis using latent class analysis.
Solcà, Manuela da Silva; Bastos, Leila Andrade; Guedes, Carlos Eduardo Sampaio; Bordoni, Marcelo; Borja, Lairton Souza; Larangeira, Daniela Farias; da Silva Estrela Tuy, Pétala Gardênia; Amorim, Leila Denise Alves Ferreira; Nascimento, Eliane Gomes; de Sá Oliveira, Geraldo Gileno; dos-Santos, Washington Luis Conrado; Fraga, Deborah Bittencourt Mothé; Veras, Patrícia Sampaio Tavares
Host tissues affected by Leishmania infantum have differing degrees of parasitism. Previously, the use of different biological tissues to detect L. infantum DNA in dogs has provided variable results. The present study was conducted to evaluate the accuracy of molecular diagnostic testing (qPCR) in dogs from an endemic area for canine visceral leishmaniasis (CVL) by determining which tissue type provided the highest rate of parasite DNA detection. Fifty-one symptomatic dogs were tested for CVL using serological, parasitological and molecular methods. Latent class analysis (LCA) was performed for accuracy evaluation of these methods. qPCR detected parasite DNA in 100% of these animals from at least one of the following tissues: splenic and bone marrow aspirates, lymph node and skin fragments, blood and conjunctival swabs. Using latent variable as gold standard, the qPCR achieved a sensitivity of 95.8% (CI 90.4-100) in splenic aspirate; 79.2% (CI 68-90.3) in lymph nodes; 77.3% (CI 64.5-90.1) in skin; 75% (CI 63.1-86.9) in blood; 50% (CI 30-70) in bone marrow; 37.5% (CI 24.2-50.8) in left-eye; and 29.2% (CI 16.7-41.6) in right-eye conjunctival swabs. The accuracy of qPCR using splenic aspirates was further evaluated in a random larger sample (n = 800), collected from dogs during a prevalence study. The specificity achieved by qPCR was 76.7% (CI 73.7-79.6) for splenic aspirates obtained from the greater sample. The sensitivity accomplished by this technique was 95% (CI 93.5-96.5) that was higher than those obtained for the other diagnostic tests and was similar to that observed in the smaller sampling study. This confirms that the splenic aspirate is the most effective type of tissue for detecting L. infantum infection. Additionally, we demonstrated that LCA could be used to generate a suitable gold standard for comparative CVL testing. PMID:25076494
Evaluating the Accuracy of Molecular Diagnostic Testing for Canine Visceral Leishmaniasis Using Latent Class Analysis
Solcà, Manuela da Silva; Bastos, Leila Andrade; Guedes, Carlos Eduardo Sampaio; Bordoni, Marcelo; Borja, Lairton Souza; Larangeira, Daniela Farias; da Silva Estrela Tuy, Pétala Gardênia; Amorim, Leila Denise Alves Ferreira; Nascimento, Eliane Gomes; de Sá Oliveira, Geraldo Gileno; dos-Santos, Washington Luis Conrado; Fraga, Deborah Bittencourt Mothé; Veras, Patrícia Sampaio Tavares
Host tissues affected by Leishmania infantum have differing degrees of parasitism. Previously, the use of different biological tissues to detect L. infantum DNA in dogs has provided variable results. The present study was conducted to evaluate the accuracy of molecular diagnostic testing (qPCR) in dogs from an endemic area for canine visceral leishmaniasis (CVL) by determining which tissue type provided the highest rate of parasite DNA detection. Fifty-one symptomatic dogs were tested for CVL using serological, parasitological and molecular methods. Latent class analysis (LCA) was performed for accuracy evaluation of these methods. qPCR detected parasite DNA in 100% of these animals from at least one of the following tissues: splenic and bone marrow aspirates, lymph node and skin fragments, blood and conjunctival swabs. Using latent variable as gold standard, the qPCR achieved a sensitivity of 95.8% (CI 90.4–100) in splenic aspirate; 79.2% (CI 68–90.3) in lymph nodes; 77.3% (CI 64.5–90.1) in skin; 75% (CI 63.1–86.9) in blood; 50% (CI 30–70) in bone marrow; 37.5% (CI 24.2–50.8) in left-eye; and 29.2% (CI 16.7–41.6) in right-eye conjunctival swabs. The accuracy of qPCR using splenic aspirates was further evaluated in a random larger sample (n = 800), collected from dogs during a prevalence study. The specificity achieved by qPCR was 76.7% (CI 73.7–79.6) for splenic aspirates obtained from the greater sample. The sensitivity accomplished by this technique was 95% (CI 93.5–96.5) that was higher than those obtained for the other diagnostic tests and was similar to that observed in the smaller sampling study. This confirms that the splenic aspirate is the most effective type of tissue for detecting L. infantum infection. Additionally, we demonstrated that LCA could be used to generate a suitable gold standard for comparative CVL testing. PMID:25076494
[Beta-thalassemias: molecular, epidemiological, diagnostical and clinical aspects].
Joly, Philippe; Pondarre, Corinne; Badens, Catherine
Beta-thalassemia is one of most common autosomal recessive disorders worldwide. In France, 5 to 10 new major or intermedia forms are diagnosed annually and the global prevalence is about 500 cases. Since 20 years and thanks to the generalization of iron chelator treatments, the life expectancy has dramatically increased. Nearly 90% of the β-thalassemic alleles are point mutations easily identified by Sanger sequencing or dedicated methods. The remaining 10% are deletions detectable by MLPA or CGH Array. The alpha-globin genotype is also essential in the exploration of beta-thalassemia because an alpha-thalassemia improves the clinical state whereas an alpha triplication worsens it. The additional genotyping of a few HbF inducer polymorphisms allows to predict the age of the first transfusion, thanks to a recent dedicated algorithm, making beta-thalassemia one of the first potential application of predictive medicine. Gene therapy, pre-implantatory diagnosis and new drugs (Sotatercept®, hepcidin-like molecules) have also recently contributed to make beta-thalassemia a main scientific topic again. PMID:25486662
Methods in virus diagnostics: from ELISA to next generation sequencing.
Boonham, Neil; Kreuze, Jan; Winter, Stephan; van der Vlugt, René; Bergervoet, Jan; Tomlinson, Jenny; Mumford, Rick
Despite the seemingly continuous development of newer and ever more elaborate methods for detecting and identifying viruses, very few of these new methods get adopted for routine use in testing laboratories, often despite the many and varied claimed advantages they possess. To understand why the rate of uptake of new technologies is so low, requires a strong understanding of what makes a good routine diagnostic tool to begin. This can be done by looking at the two most successfully established plant virus detection methods: enzyme-linked immunosorbant assay (ELISA) and more recently introduced real-time polymerase chain reaction (PCR). By examining the characteristics of this pair of technologies, it becomes clear that they share many benefits, such as an industry standard format and high levels of repeatability and reproducibility. These combine to make methods that are accessible to testing labs, which are easy to establish and robust in their use, even with new and inexperienced users. Hence, to ensure the establishment of new techniques it is necessary to not only provide benefits not found with ELISA or real-time PCR, but also to provide a platform that is easy to establish and use. In plant virus diagnostics, recent developments can be clustered into three core areas: (1) techniques that can be performed in the field or resource poor locations (e.g., loop-mediated isothermal amplification LAMP); (2) multiplex methods that are able to detect many viruses in a single test (e.g., Luminex bead arrays); and (3) methods suited to virus discovery (e.g., next generation sequencing, NGS). Field based methods are not new, with Lateral Flow Devices (LFDs) for the detection being available for a number of years now. However, the widespread uptake of this technology remains poor. LAMP does offer significant advantages over LFDs, in terms of sensitivity and generic application, but still faces challenges in terms of establishment. It is likely that the main barrier to the
A composite likelihood method for bivariate meta-analysis in diagnostic systematic reviews
Liu, Yulun; Ning, Jing; Nie, Lei; Zhu, Hongjian; Chu, Haitao
Diagnostic systematic review is a vital step in the evaluation of diagnostic technologies. In many applications, it involves pooling pairs of sensitivity and specificity of a dichotomized diagnostic test from multiple studies. We propose a composite likelihood method for bivariate meta-analysis in diagnostic systematic reviews. This method provides an alternative way to make inference on diagnostic measures such as sensitivity, specificity, likelihood ratios and diagnostic odds ratio. Its main advantages over the standard likelihood method are the avoidance of the non-convergence problem, which is non-trivial when the number of studies are relatively small, the computational simplicity and some robustness to model mis-specifications. Simulation studies show that the composite likelihood method maintains high relative efficiency compared to that of the standard likelihood method. We illustrate our method in a diagnostic review of the performance of contemporary diagnostic imaging technologies for detecting metastases in patients with melanoma. PMID:25512146
Diagnostic methods of solar cells in dependence on temperature
NASA Astrophysics Data System (ADS)
Dolensky, J.; Vesely, A.; Vanek, J.; Hrozek, J.
This study is focused on testing methods determining quality of solar cells. Nowadays the development of solar cells is much faster and there is still necessary to increase their quality by removing causes of materials defects and also defects in a process of their production. Non-destructive methods are used for correct determination of defects by using of recombination effect of charge carrier in PN junction. Due to these methods can be the solar cell diagnosed and described. By using of various temperatures during the testing we can receive more objective results thanks to simulated operation conditions. Peltier cells are used for graditional change of temperature. Cooling system with liquid nitro - LN2 is used to reach the very low temperature. Diagnostic and testing methods described in this study are based on emission of light and the recombination processes in PN junction. It is especially electroluminescence and photoluminescence method. For comparison it is used the observation of emitted light from microplasma method. Described methods detect materials and process defects due to use of lownoise and very sensitive CCD camera.
New Methods and Transducer Designs for Ultrasonic Diagnostics and Therapy
NASA Astrophysics Data System (ADS)
Rybyanets, A. N.; Naumenko, A. A.; Sapozhnikov, O. A.; Khokhlova, V. A.
Recent advances in the field of physical acoustics, imaging technologies, piezoelectric materials, and ultrasonic transducer design have led to emerging of novel methods and apparatus for ultrasonic diagnostics, therapy and body aesthetics. The paper presents the results on development and experimental study of different high intensity focused ultrasound (HIFU) transducers. Technological peculiarities of the HIFU transducer design as well as theoretical and numerical models of such transducers and the corresponding HIFU fields are discussed. Several HIFU transducers of different design have been fabricated using different advanced piezoelectric materials. Acoustic field measurements for those transducers have been performed using a calibrated fiber optic hydrophone and an ultrasonic measurement system (UMS). The results of ex vivo experiments with different tissues as well as in vivo experiments with blood vessels are presented that prove the efficacy, safety and selectivity of the developed HIFU transducers and methods.
A WAO - ARIA - GA²LEN consensus document on molecular-based allergy diagnostics
Molecular-based allergy (MA) diagnostics is an approach used to map the allergen sensitization of a patient at a molecular level, using purified natural or recombinant allergenic molecules (allergen components) instead of allergen extracts. Since its introduction, MA diagnostics has increasingly entered routine care, with currently more than 130 allergenic molecules commercially available for in vitro specific IgE (sIgE) testing. MA diagnostics allows for an increased accuracy in allergy diagnosis and prognosis and plays an important role in three key aspects of allergy diagnosis: (1) resolving genuine versus cross-reactive sensitization in poly-sensitized patients, thereby improving the understanding of triggering allergens; (2) assessing, in selected cases, the risk of severe, systemic versus mild, local reactions in food allergy, thereby reducing unnecessary anxiety for the patient and the need for food challenge testing; and (3) identifying patients and triggering allergens for specific immunotherapy (SIT). Singleplex and multiplex measurement platforms are available for MA diagnostics. The Immuno-Solid phase Allergen Chip (ISAC) is the most comprehensive platform currently available, which involves a biochip technology to measure sIgE antibodies against more than one hundred allergenic molecules in a single assay. As the field of MA diagnostics advances, future work needs to focus on large-scale, population-based studies involving practical applications, elucidation and expansion of additional allergenic molecules, and support for appropriate test interpretation. With the rapidly expanding evidence-base for MA diagnosis, there is a need for allergists to keep abreast of the latest information. The aim of this consensus document is to provide a practical guide for the indications, determination, and interpretation of MA diagnostics for clinicians trained in allergology. PMID:24090398
Molecular energies from an incremental fragmentation method
NASA Astrophysics Data System (ADS)
Meitei, Oinam Romesh; Heßelmann, Andreas
The systematic molecular fragmentation method by Collins and Deev [J. Chem. Phys. 125, 104104 (2006)] has been used to calculate total energies and relative conformational energies for a number of small and extended molecular systems. In contrast to the original approach by Collins, we have tested the accuracy of the fragmentation method by utilising an incremental scheme in which the energies at the lowest level of the fragmentation are calculated on an accurate quantum chemistry level while lower-cost methods are used to correct the low-level energies through a high-level fragmentation. In this work, the fragment energies at the lowest level of fragmentation were calculated using the random-phase approximation (RPA) and two recently developed extensions to the RPA while the incremental corrections at higher levels of the fragmentation were calculated using standard density functional theory (DFT) methods. The complete incremental fragmentation method has been shown to reproduce the supermolecule results with a very good accuracy, almost independent on the molecular type, size, or type of decomposition. The fragmentation method has also been used in conjunction with the DFT-SAPT (symmetry-adapted perturbation theory) method which enables a breakdown of the total nonbonding energy contributions into individual interaction energy terms. Finally, the potential problems of the method connected with the use of capping hydrogen atoms are analysed and two possible solutions are supplied.
Plasma DNA integrity index as a potential molecular diagnostic marker for breast cancer.
Kamel, Azza M; Teama, Salwa; Fawzy, Amal; El Deftar, Mervat
Plasma DNA integrity index is increased in various malignancies including breast cancer, the most common cancer in women worldwide; early detection is crucial for successful treatment. Current screening methods fail to detect many cases of breast cancer at an early stage. In this study, we evaluated the level of plasma DNA integrity index in 260 females (95 with breast cancer, 95 with benign breast lesions, and 70 healthy controls) to verify its potential value in discriminating malignant from benign breast lesions. The criteria of the American Joint Committee on Cancer were used for staging of breast cancer patients. DNA integrity index was measured by real-time PCR. DNA integrity index was significantly higher in breast cancer than in benign breast patients and healthy subjects (P = <0.001). DNA integrity index is correlated with TNM stage. Given 100 % specificity, the highest sensitivity achieved in detecting cancer group was 85.3 % at 0.55 DNA integrity index cutoff. In conclusion, the plasma DNA integrity index may be a promising molecular diagnostic marker of malignancy in breast lesions. PMID:26684805
Assessing the necessity of confirmatory testing for exome sequencing results in a clinical molecular diagnostic laboratory
Strom, Samuel P.; Lee, Hane; Das, Kingshuk; Vilain, Eric; Nelson, Stanley F.; Grody, Wayne W.; Deignan, Joshua L.
Purpose Sanger sequencing is currently considered the gold standard methodology for clinical molecular diagnostic testing. However, next generation sequencing (NGS) has already emerged as a much more efficient means to identify genetic variants within gene panels, the exome, or the genome. We sought to assess the accuracy of NGS variant identification in our clinical genomics laboratory with the goal of establishing a quality score threshold for confirmatory Sanger-based testing. Methods Confirmation data for reported results from 144 sequential clinical exome sequencing cases (94 unique variants) and an additional set of 16 variants from comparable research samples were analyzed. Results 103 of 110 total SNVs analyzed had a quality score ≥Q500, 103 (100%) of which were confirmed by Sanger sequencing. Of the remaining 7 variants with quality scores
An Analysis of Inhalation Injury Diagnostic Methods and Patient Outcomes.
Ching, Jessica A; Ching, Yiu-Hei; Shivers, Steven C; Karlnoski, Rachel A; Payne, Wyatt G; Smith, David J
The purpose of this study was to compare patient outcomes according to the method of diagnosing burn inhalation injury. After approval from the American Burn Association, the National Burn Repository Dataset Version 8.0 was queried for patients with a diagnosis of burn inhalation injury. Subgroups were analyzed by diagnostic method as defined by the National Burn Repository. All diagnostic methods listed for each patient were included, comparing mortality, hospital days, intensive care unit (ICU) days, and ventilator days (VDs). Z-tests, t-tests, and linear regression were used with a statistical significance of P value of less than .05. The database query yielded 9775 patients diagnosed with inhalation injury. The greatest increase in mortality was associated with diagnosis by bronchoscopy or carbon monoxide poisoning. A relative increase in hospital days was noted with diagnosis by bronchoscopy (9 days) or history (2 days). A relative increase in ICU days was associated with diagnosis according to bronchoscopy (8 days), clinical findings (2 days), or history (2 days). A relative increase in VDs was associated with diagnosis by bronchoscopy (6 days) or carbon monoxide poisoning (3 days). The combination of diagnosis by bronchoscopy and clinical findings increased the relative difference across all comparison measures. The combination of diagnosis by bronchoscopy and carbon monoxide poisoning exhibited decreased relative differences when compared with bronchoscopy alone. Diagnosis by laryngoscopy showed no mortality or association with poor outcomes. Bronchoscopic evidence of inhalation injury proved most useful, predicting increased mortality, hospital, ICU, and VDs. A combined diagnosis determined by clinical findings and bronchoscopy should be considered for clinical practice. PMID:26594867
Helicobacter pylori identification: a diagnostic/confirmatory method for evaluation.
Mesquita, B; Gonçalves, M J; Pacheco, P; Lopes, J; Salazar, F; Relvas, M; Coelho, C; Pacheco, J J; Velazco, C
The Helicobacter pylori extra gastric reservoir is probably the oral cavity. In order to evaluate the presence of this bacterium in patients with periodontitis and suspicious microbial cultures, saliva was collected from these and non-periodontitis subjects. PCRs targeting 16S rRNA gene and a 860 bp specific region were performed, and digested with the restriction enzyme DdeI. We observed that the PCR-RFLP approach augments the accuracy from 26.2 % (16/61), found in the PCR-based results, to 42.6 % (26/61), which is an excellent indicator for the establishment of this low-cost procedure as a diagnostic/confirmatory method for H. pylori evaluation. PMID:24715050
Novel scanner characterization method for color measurement and diagnostics applications
NASA Astrophysics Data System (ADS)
Lee, Bong-Sun; Bala, Raja; Sharma, Gaurav
We propose a novel scanner characterization approach for applications requiring color measurement of hardcopy output in printer calibration, characterization, and diagnostic applications. It is assumed that a typical printed medium comprises the three basic colorants C, M, Y. The proposed method is particularly advantageous when additional colorants are used in the print (e.g. black (K)). A family of scanner characterization targets is constructed, each varying in C, M, Y and at a fixed level of K. A corresponding family of 3-D scanner characterizations is derived, one for each level of K. Each characterization maps scanner RGB to a colorimetric representation such as CIELAB, using standard characterization techniques. These are then combined into a single 4-D characterization mapping RGBK to CIELAB. A refinement of the technique improves performance significantly by using a function of the scanned values for K (e.g. the scanner's green channel response to printed K) instead of the digital K value directly. This makes this new approach more robust with respect to variations in printed K over time. Secondly it enables, with a single scanner characterization, accurate color measurement of prints from different printers within the same family. Results show that the 4-D characterization technique can significantly outperform standard 3-D approaches especially in cases where the image being scanned is a patch target made up of unconstrained CMYK combinations. Thus the algorithm finds particular use in printer characterization and diagnostic applications. The method readily generalizes to printed media containing other (e.g "hi-fi") colorants, and also to other image capture devices such as digital cameras.
Diagnostic Methods for Platelet Bacteria Screening: Current Status and Developments
Störmer, Melanie; Vollmer, Tanja
Summary Bacterial contamination of blood components and the prevention of transfusion-associated bacterial infection still remains a major challenge in transfusion medicine. Over the past few decades, a significant reduction in the transmission of viral infections has been achieved due to the introduction of mandatory virus screening. Platelet concentrates (PCs) represent one of the highest risks for bacterial infection. This is due to the required storage conditions for PCs in gas-permeable containers at room temperature with constant agitation, which support bacterial proliferation from low contamination levels to high titers. In contrast to virus screening, since 1997 in Germany bacterial testing of PCs is only performed as a routine quality control or, since 2008, to prolong the shelf life to 5 days. In general, bacterial screening of PCs by cultivation methods is implemented by the various blood services. Although these culturing systems will remain the gold standard, the significance of rapid methods for screening for bacterial contamination has increased over the last few years. These new methods provide powerful tools for increasing the bacterial safety of blood components. This article summarizes the course of policies and provisions introduced to increase bacterial safety of blood components in Germany. Furthermore, we give an overview of the different diagnostic methods for bacterial screening of PCs and their current applicability in routine screening processes. PMID:24659944
Molecular Biological Methods in Environmental Engineering.
Zhang, Guocai; Wei, Li; Chang, Chein-Chi; Zhang, Yuhua; Wei, Dong
Bacteria, acting as catalysts, perform the function of degrading pollutants. Molecular biological techniques play an important role in research on the community analysis, the composition and the functions of complex microbial communities. The development of secondary high-throughput pyrosequencing techiniques enhances the understanding of the composition of the microbial community. The literatures of 2015 indicated that 16S rDNA gene as genetic tag is still the important method for bacteria identification and classification. 454 high throughput sequencing and Illumina MiSeq sequencing have been the primary and widely recognized methods to analyze the microbial. This review will provide environmental engineers and microbiologists an overview of important advancements in molecular techniques and highlight the application of these methods in diverse environments. PMID:27620079
Optical biosensor technologies for molecular diagnostics at the point-of-care
NASA Astrophysics Data System (ADS)
Schotter, Joerg; Schrittwieser, Stefan; Muellner, Paul; Melnik, Eva; Hainberger, Rainer; Koppitsch, Guenther; Schrank, Franz; Soulantika, Katerina; Lentijo-Mozo, Sergio; Pelaz, Beatriz; Parak, Wolfgang; Ludwig, Frank; Dieckhoff, Jan
Label-free optical schemes for molecular biosensing hold a strong promise for point-of-care applications in medical research and diagnostics. Apart from diagnostic requirements in terms of sensitivity, specificity, and multiplexing capability, also other aspects such as ease of use and manufacturability have to be considered in order to pave the way to a practical implementation. We present integrated optical waveguide as well as magnetic nanoparticle based molecular biosensor concepts that address these aspects. The integrated optical waveguide devices are based on low-loss photonic wires made of silicon nitride deposited by a CMOS compatible plasma-enhanced chemical vapor deposition (PECVD) process that allows for backend integration of waveguides on optoelectronic CMOS chips. The molecular detection principle relies on evanescent wave sensing in the 0.85 μm wavelength regime by means of Mach-Zehnder interferometers, which enables on-chip integration of silicon photodiodes and, thus, the realization of system-on-chip solutions. Our nanoparticle-based approach is based on optical observation of the dynamic response of functionalized magneticcore/ noble-metal-shell nanorods (`nanoprobes') to an externally applied time-varying magnetic field. As target molecules specifically bind to the surface of the nanoprobes, the observed dynamics of the nanoprobes changes, and the concentration of target molecules in the sample solution can be quantified. This approach is suitable for dynamic real-time measurements and only requires minimal sample preparation, thus presenting a highly promising point-of-care diagnostic system. In this paper, we present a prototype of a diagnostic device suitable for highly automated sample analysis by our nanoparticle-based approach.
Molecular and Biological Diagnostic Tests for Monitoring Benzimidazole Resistance in Human Soil-Transmitted Helminths
Diawara, Aïssatou; Schwenkenbecher, Jan M.; Kaplan, Ray M.; Prichard, Roger K.
In endemic countries with soil-transmitted helminths mass drug administration with albendazole or mebendazole are being implemented as a control strategy. However, it is well known in veterinary helminths that the use of the same benzimidazole drugs can place selection on the β-tubulin gene, leading to resistance. Given the concern that resistance could arise in human soil-transmitted helminths, there is an urgent need to develop accurate diagnostic tools for monitoring resistance. In this study, we developed molecular assays to detect putative resistance genetic changes in Ascaris lumbricoides, Trichuris trichiura, and hookworms, and we optimized an egg hatch assay for the canine hookworm Ancylostoma caninum and applied it to Necator americanus. Both assays were tested on field samples. The molecular assays demonstrated their reproducibility and capacity to detect the presence of worms carrying putative resistance-associated genetic changes. However, further investigations are needed to validate our molecular and biological tests on additional field isolates. PMID:23458960
Molecular and biological diagnostic tests for monitoring benzimidazole resistance in human soil-transmitted helminths.
Diawara, Aïssatou; Schwenkenbecher, Jan M; Kaplan, Ray M; Prichard, Roger K
In endemic countries with soil-transmitted helminths mass drug administration with albendazole or mebendazole are being implemented as a control strategy. However, it is well known in veterinary helminths that the use of the same benzimidazole drugs can place selection on the β-tubulin gene, leading to resistance. Given the concern that resistance could arise in human soil-transmitted helminths, there is an urgent need to develop accurate diagnostic tools for monitoring resistance. In this study, we developed molecular assays to detect putative resistance genetic changes in Ascaris lumbricoides, Trichuris trichiura, and hookworms, and we optimized an egg hatch assay for the canine hookworm Ancylostoma caninum and applied it to Necator americanus. Both assays were tested on field samples. The molecular assays demonstrated their reproducibility and capacity to detect the presence of worms carrying putative resistance-associated genetic changes. However, further investigations are needed to validate our molecular and biological tests on additional field isolates. PMID:23458960
Diagnostic potential of ancillary molecular testing in differentiation of benign and malignant thyroid nodules.
Bhatia, Parisha; Deniwar, Ahmed; Friedlander, Paul; Aslam, Rizwan; Kandil, Emad
Fine needle aspiration (FNA) cytology, being the mainstay to diagnose thyroid nodules, does not provide definitive results in a subset of patients. The use of molecular markers testing has been described as a useful aid in differentiation of thyroid nodules that present with an indeterminate cytodiagnosis. Molecular tests, such as the Afirma gene classifier, mutational assay and immunohistochemical markers have been increasingly used to further increase the accuracy and defer unnecessary surgeries for benign thyroid nodules. However, in light of the current literature, their emerging roles in clinical practice are limited due to financial and technical limitations. Nevertheless, their synergistic implementation can predict the risk of malignancy and yield an accurate diagnosis. This review discusses the clinical utility of various molecular tests done on FNA indeterminate nodules to avoid diagnostic thyroidectomies and warrant the need of future multi-Institutional studies. PMID:25750270
Diagnostic methods to assess inspiratory and expiratory muscle strength*
Caruso, Pedro; de Albuquerque, André Luis Pereira; Santana, Pauliane Vieira; Cardenas, Leticia Zumpano; Ferreira, Jeferson George; Prina, Elena; Trevizan, Patrícia Fernandes; Pereira, Mayra Caleffi; Iamonti, Vinicius; Pletsch, Renata; Macchione, Marcelo Ceneviva; Carvalho, Carlos Roberto Ribeiro
Impairment of (inspiratory and expiratory) respiratory muscles is a common clinical finding, not only in patients with neuromuscular disease but also in patients with primary disease of the lung parenchyma or airways. Although such impairment is common, its recognition is usually delayed because its signs and symptoms are nonspecific and late. This delayed recognition, or even the lack thereof, occurs because the diagnostic tests used in the assessment of respiratory muscle strength are not widely known and available. There are various methods of assessing respiratory muscle strength during the inspiratory and expiratory phases. These methods are divided into two categories: volitional tests (which require patient understanding and cooperation); and non-volitional tests. Volitional tests, such as those that measure maximal inspiratory and expiratory pressures, are the most commonly used because they are readily available. Non-volitional tests depend on magnetic stimulation of the phrenic nerve accompanied by the measurement of inspiratory mouth pressure, inspiratory esophageal pressure, or inspiratory transdiaphragmatic pressure. Another method that has come to be widely used is ultrasound imaging of the diaphragm. We believe that pulmonologists involved in the care of patients with respiratory diseases should be familiar with the tests used in order to assess respiratory muscle function.Therefore, the aim of the present article is to describe the advantages, disadvantages, procedures, and clinical applicability of the main tests used in the assessment of respiratory muscle strength. PMID:25972965
Vaccines and diagnostic methods for bovine mastitis: fact and fiction.
Yancey, R J
A number of problems are uniquely associated with vaccination of dairy cows for mastitis. One of these is that the number of mastitis pathogens is numerous and heterogeneous. Vaccine efforts have concentrated mainly on the major mastitis pathogens. While at least one S. aureus bacterin has been commercially available for a number of years, no large-scale, independent field trials have been published in refereed journals which support the efficacy of this vaccine. Experimental vaccines for S. aureus composed of pseudocapsule-enriched bacterins supplemented with alpha- and/or beta-toxoids appear promising, but none of these has been commercialized. With S. uberis, some protection against homologous strain challenges was reported recently with a live strain and a bacterin, but other data from the same laboratory showed this vaccine would not protect against heterologous challenge strains. At this time there is only one highly effective vaccine for mastitis, the core-antigen vaccine for coliform mastitis. All of the commercially available vaccines for this indication are bacterins of rough mutants of E. coli strain J5 or Salmonella spp. Preliminary success with an experimental vaccine based on the plasminogen activator of S. uberis is a very different approach for a mastitis vaccine. Little success has been reported with vaccination against other mastitis pathogens. For diagnostic methods, the high somatic cell count, as measured by direct count or indirect assays, remains the cornerstone of mastitis diagnosis. However, for subclinical mastitis, bacterial cell culture is a reliable diagnostic method. Pathogen identification may rely on older biochemical testing methods or newer commercial identification systems, depending on the laboratory budget. ELISA assays also have been used to assess herd infection status. Epidemiologic studies have used DNA fingerprinting and ribotyping, but none of these methods has yet produced an easily utilized commercial format. Within the
The development, evaluation and performance of molecular diagnostics for detection of Mycobacterium tuberculosis.
Bates, Matthew; Zumla, Alimuddin
The unique pathogenesis of tuberculosis (TB) poses several barriers to the development of accurate diagnostics: a) the establishment of life-long latency by Mycobacterium tuberculosis (M.tb) after primary infection confounds the development of classical antibody or antigen based assays; b) our poor understanding of the molecular pathways that influence progression from latent to active disease; c) the intracellular nature of M.tb infection in tissues means that M.tb and/or its components, are not readily detectable in peripheral specimens; and d) the variable presence of M.tb bacilli in specimens from patients with extrapulmonary TB or children. The literature on the current portfolio of molecular diagnostics tests for TB is reviewed here and the developmental pipeline is summarized. Also reviewed are data from recently published operational research on the GeneXpert MTB/RIF assay and discussed are the lessons that can be taken forward for the design of studies to evaluate the impact of TB diagnostics. PMID:26735769
Computer methods for ITER-like materials LIBS diagnostics
NASA Astrophysics Data System (ADS)
Łepek, Michał; GÄ sior, Paweł
Recent development of Laser-Induced Breakdown Spectroscopy (LIBS) caused that this method is considered as the most promising for future diagnostic applications for characterization of the deposited materials in the International Thermonuclear Experimental Reactor (ITER), which is currently under construction. In this article the basics of LIBS are shortly discussed and the software for spectra analyzing is presented. The main software function is to analyze measured spectra with respect to the certain element lines presence. Some program operation results are presented. Correct results for graphite and aluminum are obtained although identification of tungsten lines is a problem. The reason for this is low tungsten lines intensity, and thus low signal to noise ratio of the measured signal. In the second part artificial neural networks (ANNs) as the next step for LIBS spectra analyzing are proposed. The idea is focused on multilayer perceptron network (MLP) with backpropagation learning method. The potential of ANNs for data processing was proved through application in several LIBS-related domains, e.g. differentiating ancient Greek ceramics (discussed). The idea is to apply an ANN for determination of W, Al, C presence on ITER-like plasma-facing materials.
Using prior knowledge from cellular pathways and molecular networks for diagnostic specimen classification
For many complex diseases, an earlier and more reliable diagnosis is considered a key prerequisite for developing more effective therapies to prevent or delay disease progression. Classical statistical learning approaches for specimen classification using omics data, however, often cannot provide diagnostic models with sufficient accuracy and robustness for heterogeneous diseases like cancers or neurodegenerative disorders. In recent years, new approaches for building multivariate biomarker models on omics data have been proposed, which exploit prior biological knowledge from molecular networks and cellular pathways to address these limitations. This survey provides an overview of these recent developments and compares pathway- and network-based specimen classification approaches in terms of their utility for improving model robustness, accuracy and biological interpretability. Different routes to translate omics-based multifactorial biomarker models into clinical diagnostic tests are discussed, and a previous study is presented as example. PMID:26141830
[Application of molecular diagnostic techniques in precision medicine of personalized treatment for colorectal cancer].
Fu, Ji; Lin, Guole
Precision medicine is to customize the treatment options for individual patient based on the personal genome information. Colorectal cancer (CRC) is one of the most common cancer worldwide. Molecular heterogeneity of CRC, which includes the MSI phenotype, hypermutation phenotype, and their relationship with clinical preferences, is believed to be one of the main factors responsible for the considerable variability in treatment response. The development of powerful next-generation sequencing (NGS) technologies allows us to further understand the biological behavior of colorectal cancer, and to analyze the prognosis and chemotherapeutic drug reactions by molecular diagnostic techniques, which can guide the clinical treatment. This paper will introduce the new findings in this field. Meanwhile we integrate the new progress of key pathways including EGFR, RAS, PI3K/AKT and VEGF, and the experience in selective patients through associated molecular diagnostic screening who gain better efficacy after target therapy. The technique for detecting circulating tumor DNA (ctDNA) is introduced here as well, which can identify patients with high risk for recurrence, and demonstrate the risk of chemotherapy resistance. Mechanism of tumor drug resistance may be revealed by dynamic observation of gene alteration during treatment. PMID:26797832
Molecular pathways of human adrenocortical carcinoma - translating cell signalling knowledge into diagnostic and treatment options.
Szyszka, Paulina; Grossman, Ashley B; Diaz-Cano, Salvador; Sworczak, Krzysztof; Dworakowska, Dorota
Adrenocortical carcinoma is associated with a low cure rate and a high recurrence rate. The prognosis is poor, and at diagnosis 30-40% of cases are already metastatic. The current therapeutic options (surgical resection, followed by adjuvant mitotane treatment +/- chemotherapy) are limited, and the results remain unsatisfactory. Key molecular events that contribute to formation of adrenocortical cancer are IGF2 overexpression, TP53-inactivating mutations, and constitutive activation of the Wnt/b-catenin signalling pathway via activating mutations of the b-catenin gene. The underlying genetic causes of inherited tumour syndromes have provided insights into molecular pathogenesis. The increased occurrence of adrenocortical tumours in Li-Fraumeni and Beckwith-Wiedemann syndromes, and Carney complex, has highlighted the roles of specific susceptibility genes: TP53, IGF2, and PRKAR1A, respectively. Further studies have confirmed that these genes are also involved in sporadic tumour cases. Crucially, transcriptome-wide studies have determined the differences between malignant and benign adrenocortical tumours, providing potential diagnostic tools. In conclusion, enhancing our understanding of the molecular events of adrenocortical tumourigenesis, especially with regard to the signalling pathways that may be disrupted, will greatly contribute to improving a range of available diagnostic, prognostic, and treatment approaches. (Endokrynol Pol 2016; 67 (4): 427-440). PMID:27387247
Studying the Impact of Spaceflight Environment on Immune Functions Using New Molecular Diagnostics System
NASA Astrophysics Data System (ADS)
Cohen, Luchino
Immune functions are altered during space flights. Latent virus reactivation, reduction in the number of immune cells, decreased cell activation and increased sensitivity of astronauts to infections following their return on Earth demonstrate that the immune system is less efficient during space flight. The causes of this immune deficiency are not fully understood and this dysfunction during long-term missions could result in the appearance of opportunistic infections or a decrease in the immuno-surveillance mechanisms that eradicate cancer cells. Therefore, the immune functions of astronauts will have to be monitored continuously during long-term missions in space, using miniature and semi-automated diagnostic systems. The objectives of this project are to study the causes of space-related immunodeficiency, to develop countermeasures to maintain an optimal immune function and to improve our capacity to detect infectious diseases during space missions through the monitoring of astronauts' immune system. In order to achieve these objectives, an Immune Function Diagnostic System (IFDS) will be designed to perform a set of immunological assays on board spacecrafts or on planet-bound bases. Through flow cytometric assays and molecular biology analyses, this diagnostic system could improve medical surveillance of astronauts and could be used to test countermeasures aimed at preventing immune deficiency during space missions. The capacity of the instrument to assess cellular fluorescence and to quantify the presence of soluble molecules in biological samples would support advanced molecular studies in space life sciences. Finally, such diagnostic system could also be used on Earth in remote areas or in mobile hospitals following natural disasters to fight against infectious diseases and other pathologies.
CpG methylation analysis--current status of clinical assays and potential applications in molecular diagnostics: a report of the Association for Molecular Pathology.
Sepulveda, Antonia R; Jones, Dan; Ogino, Shuji; Samowitz, Wade; Gulley, Margaret L; Edwards, Robin; Levenson, Victor; Pratt, Victoria M; Yang, Bin; Nafa, Khedoudja; Yan, Liying; Vitazka, Patrick
Methylation of CpG islands in gene promoter regions is a major molecular mechanism of gene silencing and underlies both cancer development and progression. In molecular oncology, testing for the CpG methylation of tissue DNA has emerged as a clinically useful tool for tumor detection, outcome prediction, and treatment selection, as well as for assessing the efficacy of treatment with the use of demethylating agents and monitoring for tumor recurrence. In addition, because CpG methylation occurs early in pre-neoplastic tissues, methylation tests may be useful as markers of cancer risk in patients with either infectious or inflammatory conditions. The Methylation Working Group of the Clinical Practice Committee of the Association of Molecular Pathology has reviewed the current state of clinical testing in this area. We report here our summary of both the advantages and disadvantages of various methods, as well as the needs for standardization and reporting. We then conclude by summarizing the most promising areas for future clinical testing in cancer molecular diagnostics. PMID:19541921
Development of semiclassical molecular dynamics simulation method.
Nakamura, Hiroki; Nanbu, Shinkoh; Teranishi, Yoshiaki; Ohta, Ayumi
Various quantum mechanical effects such as nonadiabatic transitions, quantum mechanical tunneling and coherence play crucial roles in a variety of chemical and biological systems. In this paper, we propose a method to incorporate tunneling effects into the molecular dynamics (MD) method, which is purely based on classical mechanics. Caustics, which define the boundary between classically allowed and forbidden regions, are detected along classical trajectories and the optimal tunneling path with minimum action is determined by starting from each appropriate caustic. The real phase associated with tunneling can also be estimated. Numerical demonstration with use of a simple collinear chemical reaction O + HCl → OH + Cl is presented in order to help the reader to well comprehend the method proposed here. Generalization to the on-the-fly ab initio version is rather straightforward. By treating the nonadiabatic transitions at conical intersections by the Zhu-Nakamura theory, new semiclassical MD methods can be developed. PMID:27067383
Diagnostic Methods for Bile Acid Malabsorption in Clinical Practice
Vijayvargiya, Priya; Camilleri, Michael; Shin, Andrea; Saenger, Amy
Altered bile acid (BA) concentrations in the colon may cause diarrhea or constipation. BA malabsorption (BAM) accounts for >25% of patients with irritable bowel syndrome (IBS) with diarrhea and chronic diarrhea in Western countries. As BAM is increasingly recognized, proper diagnostic methods are desired in clinical practice to help direct the most effective treatment course for the chronic bowel dysfunction. This review appraises the methodology, advantages and disadvantages of 4 tools that directly measure BAM: 14C-glycocholate breath and stool test, 75Selenium HomotauroCholic Acid Test (SeHCAT), 7 α-hydroxy-4-cholesten-3-one (C4) and fecal BAs. 14C-glycocholate is a laborious test no longer widely utilized. 75SeHCAT is validated, but not available in the United States. Serum C4 is a simple, accurate method that is applicable to a majority of patients, but requires further clinical validation. Fecal measurements to quantify total and individual fecal BAs are technically cumbersome and not widely available. Regrettably, none of these tests are routinely available in the U.S., and a therapeutic trial with a BA binder is used as a surrogate for diagnosis of BAM. Recent data suggest there is an advantage to studying fecal excretion of the individual BAs and their role in BAM; this may constitute a significant advantage of the fecal BA method over the other tests. Fecal BA test could become a routine addition to fecal fat measurement in patients with unexplained diarrhea. In summary, availability determines the choice of test among C4, SeHCAT and fecal BA; more widespread availability of such tests would enhance clinical management of these patients. PMID:23644387
Nanomechanical transduction of molecular interactions on microcantilevers for biochemical detection and diagnostics
NASA Astrophysics Data System (ADS)
Tark, Soo-Hyun
There is a strong demand for a reliable detection platform that can provide the benefits of enhanced sensitivity and selectivity with greater simplicity and cost-effectiveness. The new generations of biosensors also require microfabricated platforms with integrated biologically sensitive components for specific and quantitative detection of analytes in a miniaturized format as well as capabilities for label-free detection and massive parallelization. It has been unambiguously demonstrated that the molecular binding-induced surface stress can be used to monitor specific biochemical binding events and kinetics in real-time with high sensitivity, representing the promising potential for nanomechanical sensors. The fundamental validation of the receptor immobilization and target binding as well as the transduction and quantitative detection of such molecular recognition events taking place on microcantilevers are demonstrated utilizing the optical approach for monitoring the cantilever deflection. The label-free detection of cholera toxin using microcantilevers functionalized with ganglioside-Nanodiscs is demonstrated as a new strategy for immobilizing receptors on microcantilevers. The microcantilever-based sensors, however, require a new paradigm for signal transduction and detection beyond the optical method that supports the unique multiplexing capability by operating a large array of cantilevers with means for simple and accurate readout. Hence, a new electrical readout mechanism comprising a microcantilever array with MOSFETs embedded in the high stress region of the microcantilevers is developed, which provides label- and optics-free signal transduction mechanism. In this work, significant strides have been made towards the MOSFET-microcantilever detection approach. The process and device simulations for embedded-MOSFETs are performed to optimize process parameters and establish guidelines for device design and fabrication. Various designs of MOSFET
Dental and dental hygiene students' diagnostic accuracy in oral radiology: effect of diagnostic strategy and instructional method.
Baghdady, Mariam T; Carnahan, Heather; Lam, Ernest W N; Woods, Nicole N
There has been much debate surrounding diagnostic strategies and the most appropriate training models for novices in oral radiology. It has been argued that an analytic approach, using a step-by-step analysis of the radiographic features of an abnormality, is ideal. Alternative research suggests that novices can successfully employ non-analytic reasoning. Many of these studies do not take instructional methodology into account. This study evaluated the effectiveness of non-analytic and analytic strategies in radiographic interpretation and explored the relationship between instructional methodology and diagnostic strategy. Second-year dental and dental hygiene students were taught four radiographic abnormalities using basic science instructions or a step-by-step algorithm. The students were tested on diagnostic accuracy and memory immediately after learning and one week later. A total of seventy-three students completed both immediate and delayed sessions and were included in the analysis. Students were randomly divided into two instructional conditions: one group provided a diagnostic hypothesis for the image and then identified specific features to support it, while the other group first identified features and then provided a diagnosis. Participants in the diagnosis-first condition (non-analytic reasoning) had higher diagnostic accuracy then those in the features-first condition (analytic reasoning), regardless of their learning condition. No main effect of learning condition or interaction with diagnostic strategy was observed. Educators should be mindful of the potential influence of analytic and non-analytic approaches on the effectiveness of the instructional method. PMID:25179924
CMOS Time-Resolved, Contact, and Multispectral Fluorescence Imaging for DNA Molecular Diagnostics
Guo, Nan; Cheung, Ka Wai; Wong, Hiu Tung; Ho, Derek
Instrumental limitations such as bulkiness and high cost prevent the fluorescence technique from becoming ubiquitous for point-of-care deoxyribonucleic acid (DNA) detection and other in-field molecular diagnostics applications. The complimentary metal-oxide-semiconductor (CMOS) technology, as benefited from process scaling, provides several advanced capabilities such as high integration density, high-resolution signal processing, and low power consumption, enabling sensitive, integrated, and low-cost fluorescence analytical platforms. In this paper, CMOS time-resolved, contact, and multispectral imaging are reviewed. Recently reported CMOS fluorescence analysis microsystem prototypes are surveyed to highlight the present state of the art. PMID:25365460
Diagnostic Methods for Detection of Blood-Borne Candidiasis.
Clancy, Cornelius J; Nguyen, M Hong
β-D-glucan (Fungitell) and polymerase chain reaction-based (T2Candida) assays of blood samples are FDA-approved adjuncts to cultures for diagnosing candidemia and other types of invasive candidiasis, but their clinical roles are unclear. In this chapter, we describe laboratory protocols for performing Fungitell and T2Candida assays. We then discuss step-by-step methods for interpreting test results at the bedside using a Bayesian framework, and for incorporating assays into rational patient management strategies. Prior to interpreting results, clinicians must recognize that test performance varies based on the type of invasive candidiasis being diagnosed. In general, the type of invasive candidiasis that is most likely in a given patient can be identified, and the pretest likelihood of disease estimated. From there, positive and negative predictive values (PPV, NPV) for an assay can be calculated. At a population level, tests can be incorporated into screening strategies for antifungal treatment. NPV and PPV thresholds can be defined for discontinuing antifungal prophylaxis or initiating preemptive treatment, respectively. Using the thresholds, it is possible to assign windows of pretest likelihood for invasive candidiasis (and corresponding patient populations) in which tests are most likely to valuable. At the individual patient level, tests may be useful outside of the windows proposed for screening populations. The interpretive and clinical decision-making processes we discuss will be applicable to other diagnostic assays as they enter the clinic, and to existing assays as more data emerge from various populations. PMID:26519076
CpG Methylation Analysis—Current Status of Clinical Assays and Potential Applications in Molecular Diagnostics
Sepulveda, Antonia R.; Jones, Dan; Ogino, Shuji; Samowitz, Wade; Gulley, Margaret L.; Edwards, Robin; Levenson, Victor; Pratt, Victoria M.; Yang, Bin; Nafa, Khedoudja; Yan, Liying; Vitazka, Patrick
Methylation of CpG islands in gene promoter regions is a major molecular mechanism of gene silencing and underlies both cancer development and progression. In molecular oncology, testing for the CpG methylation of tissue DNA has emerged as a clinically useful tool for tumor detection, outcome prediction, and treatment selection, as well as for assessing the efficacy of treatment with the use of demethylating agents and monitoring for tumor recurrence. In addition, because CpG methylation occurs early in pre-neoplastic tissues, methylation tests may be useful as markers of cancer risk in patients with either infectious or inflammatory conditions. The Methylation Working Group of the Clinical Practice Committee of the Association of Molecular Pathology has reviewed the current state of clinical testing in this area. We report here our summary of both the advantages and disadvantages of various methods, as well as the needs for standardization and reporting. We then conclude by summarizing the most promising areas for future clinical testing in cancer molecular diagnostics. PMID:19541921
Optical caries diagnostics: comparison of laser spectroscopic PNC method with method of laser integral fluorescence
NASA Astrophysics Data System (ADS)
Masychev, Victor I.
In this research we present the results of approbation of two methods of optical caries diagnostics: PNC-spectral diagnostics and caries detection by laser integral fluorescence. The research was conducted in a dental clinic. PNC-method analyses parameters of probing laser radiation and PNC-spectrums of stimulated secondary radiations: backscattering and endogenous fluorescence of caries-involved bacterias. He-Ne-laser ((lambda) =632,8 nm, 1-2mW) was used as a source of probing (stimulated) radiation. For registration of signals, received from intact and pathological teeth PDA-detector was applied. PNC-spectrums were processed by special algorithms, and were displayed on PC monitor. The method of laser integral fluorescence was used for comparison. In this case integral power of fluorescence of human teeth was measured. As a source of probing (stimulated) radiation diode lasers ((lambda) =655 nm, 0.1 mW and 630nm, 1mW) and He-Ne laser were applied. For registration of signals Si-photodetector was used. Integral power was shown in a digital indicator. Advantages and disadvantages of these methods are described in this research. It is disclosed that the method of laser integral power of fluorescence has the following characteristics: simplicity of construction and schema-technical decisions. However the method of PNC-spectral diagnostics are characterized by considerably more sensitivity in diagnostics of initial caries and capability to differentiate pathologies of various stages (for example, calculus/initial caries). Estimation of spectral characteristics of PNC-signals allows eliminating a number of drawbacks, which are character for detection by method of laser integral fluorescence (for instance, detection of fluorescent fillings, plagues, calculus, discolorations generally, amalgam, gold fillings as if it were caries.
Computational methods for optical molecular imaging
Chen, Duan; Wei, Guo-Wei; Cong, Wen-Xiang; Wang, Ge
Summary A new computational technique, the matched interface and boundary (MIB) method, is presented to model the photon propagation in biological tissue for the optical molecular imaging. Optical properties have significant differences in different organs of small animals, resulting in discontinuous coefficients in the diffusion equation model. Complex organ shape of small animal induces singularities of the geometric model as well. The MIB method is designed as a dimension splitting approach to decompose a multidimensional interface problem into one-dimensional ones. The methodology simplifies the topological relation near an interface and is able to handle discontinuous coefficients and complex interfaces with geometric singularities. In the present MIB method, both the interface jump condition and the photon flux jump conditions are rigorously enforced at the interface location by using only the lowest-order jump conditions. This solution near the interface is smoothly extended across the interface so that central finite difference schemes can be employed without the loss of accuracy. A wide range of numerical experiments are carried out to validate the proposed MIB method. The second-order convergence is maintained in all benchmark problems. The fourth-order convergence is also demonstrated for some three-dimensional problems. The robustness of the proposed method over the variable strength of the linear term of the diffusion equation is also examined. The performance of the present approach is compared with that of the standard finite element method. The numerical study indicates that the proposed method is a potentially efficient and robust approach for the optical molecular imaging. PMID:20485461
Chronic intraoral pain--assessment of diagnostic methods and prognosis.
Pigg, Maria
The overall goal of this thesis was to broaden our knowledge of chronic intraoral pain. The research questions were: What methods can be used to differentiate inflammatory, odontogenic tooth pain from pain that presents as toothache but is non-odontogenic in origin? What is the prognosis of chronic tooth pain of non-odontogenic origin, and which factors affect the prognosis? Atypical odontalgia (AO) is a relatively rare but severe and chronic pain condition affecting the dentoalveolar region. Recent research indicates that the origin is peripheral nerve damage: neuropathic pain. The condition presents as tooth pain and is challenging to dentists because it is difficult to distinguish from ordinary toothache due to inflammation or infection. AO is of interest to the pain community because it shares many characteristics with other chronic pain conditions, and pain perpetuation mechanisms are likely to be similar. An AO diagnosis is made after a comprehensive examination and assessment of patients' self-reported characteristics: the pain history. Traditional dental diagnostic methods do not appear to suffice, since many patients report repeated care-seeking and numerous treatment efforts with little or no pain relief. Developing methods that are useful in the clinical setting is a prerequisite for a correct diagnosis and adequate treatment decisions. Quantitative sensory testing (QST) is used to assess sensory function on skin when nerve damage or disease is suspected. A variety of stimuli has been used to examine the perception of, for example, touch, temperature (painful and non-painful), vibration, pinprick pain, and pressure pain. To detect sensory abnormalities and nerve damage in the oral cavity, the same methods may be possible to use. Study I examined properties of thermal thresholds in and around the mouth in 30 pain-free subjects: the influence of measurement location and stimulation area size on threshold levels, and time variability of thresholds
Molecular Methods and Platforms for Infectious Diseases Testing
Emmadi, Rajyasree; Boonyaratanakornkit, Jerry B.; Selvarangan, Rangaraj; Shyamala, Venkatakrishna; Zimmer, Barbara L.; Williams, Laurina; Bryant, Bonita; Schutzbank, Ted; Schoonmaker, Michele M.; Amos Wilson, Jean A.; Hall, Leslie; Pancholi, Preeti; Bernard, Kathryn
The superior sensitivity and specificity associated with the use of molecular assays has greatly improved the field of infectious disease diagnostics by providing clinicians with results that are both accurate and rapidly obtained. Herein, we review molecularly based infectious disease diagnostic tests that are Food and Drug Administration approved or cleared and commercially available in the United States as of December 31, 2010. We describe specific assays and their performance, as stated in the Food and Drug Administration's Summary of Safety and Effectiveness Data or the Office of In Vitro Diagnostic Device Evaluation and Safety's decision summaries, product inserts, or peer-reviewed literature. We summarize indications for testing, limitations, and challenges related to implementation in a clinical laboratory setting for a wide variety of common pathogens. The information presented in this review will be particularly useful for laboratories that plan to implement or expand their molecular offerings in the near term. PMID:21871973
Ruling in or ruling out thyroid malignancy by molecular diagnostics of thyroid nodules.
Eszlinger, Markus; Hegedüs, László; Paschke, Ralf
Routine morphologic cytology is the basis for any kind of (integrated) molecular FNA diagnostics. The rule out (gene expression classifier) approach requires confirmation by independent studies, whereas the rule in approach (detection of BRAF, NRAS, HRAS, and KRAS and PAX8/PPARG- and RET/PTC rearrangements) has been investigated by several groups with overall reproducible results. Moreover, molecular screening for point mutations and rearrangements is feasible in routine air-dried FNA smears, offering several advantages over obtaining additional fresh FNA material. The current panel of somatic mutations (rule in approach) for indeterminate FNAs clarifies only a subgroup of indeterminate FNAs. Therefore, further markers are urgently needed that can reliably identify the malignant, but mutation negative and especially the many benign nodules, among the indeterminate FNAs. miRNA markers and the targeted next generation sequencing (NGS) technology do have the potential to identify those nodules that are mutation negative by current approaches. PMID:25047205
Challenges of bringing next generation sequencing technologies to clinical molecular diagnostic laboratories.
Wong, Lee-Jun C
Molecular diagnosis of complex dual genome mitochondrial disorders is a challenge. It requires the identification of deleterious mutations in one of the ~1,500 nuclear genes and the mitochondrial genome. If the molecular defect is in the mitochondrial genome, quantification of degree of mutation load (heteroplasmy) in affected tissues is important. Due to the extreme clinical and genetic heterogeneity, conventional sequence analysis of the candidate genes one-by-one is impractical, if not impossible. The newly developed massively parallel next generation sequencing (NGS) technique, that allows simultaneous sequence analysis of multiple target genes, when appropriately validated with deep coverage and proper quality controls, can be used as an effective comprehensive diagnostic approach in CLIA certified clinical laboratories. PMID:23269496
Genetic diversity in Treponema pallidum: implications for pathogenesis, evolution and molecular diagnostics of syphilis and yaws
Šmajs, David; Norris, Steven J.; Weinstock, George M.
Pathogenic uncultivable treponemes, similar to syphilis-causing Treponema pallidum subspecies pallidum, include T. pallidum ssp. pertenue, T. pallidum ssp. endemicum and Treponema carateum, which cause yaws, bejel and pinta, respectively. Genetic analyses of these pathogens revealed striking similarity among these bacteria and also a high degree of similarity to the rabbit pathogen, T. paraluiscuniculi, a treponeme not infectious to humans. Genome comparisons between pallidum and non-pallidum treponemes revealed genes with potential involvement in human infectivity, whereas comparisons between pallidum and pertenue treponemes identified genes possibly involved in the high invasivity of syphilis treponemes. Genetic variability within syphilis strains is considered as the basis of syphilis molecular epidemiology with potential to detect more virulent strains, whereas genetic variability within a single strain is related to its ability to elude the immune system of the host. Genome analyses also shed light on treponemal evolution and on chromosomal targets for molecular diagnostics of treponemal infections. PMID:22198325
Genetic diversity in Treponema pallidum: implications for pathogenesis, evolution and molecular diagnostics of syphilis and yaws.
Smajs, David; Norris, Steven J; Weinstock, George M
Pathogenic uncultivable treponemes, similar to syphilis-causing Treponema pallidum subspecies pallidum, include T. pallidum ssp. pertenue, T. pallidum ssp. endemicum and Treponema carateum, which cause yaws, bejel and pinta, respectively. Genetic analyses of these pathogens revealed striking similarity among these bacteria and also a high degree of similarity to the rabbit pathogen, Treponema paraluiscuniculi, a treponeme not infectious to humans. Genome comparisons between pallidum and non-pallidum treponemes revealed genes with potential involvement in human infectivity, whereas comparisons between pallidum and pertenue treponemes identified genes possibly involved in the high invasivity of syphilis treponemes. Genetic variability within syphilis strains is considered as the basis of syphilis molecular epidemiology with potential to detect more virulent strains, whereas genetic variability within a single strain is related to its ability to elude the immune system of the host. Genome analyses also shed light on treponemal evolution and on chromosomal targets for molecular diagnostics of treponemal infections. PMID:22198325
Advanced Optical Diagnostic Methods for Describing Fuel Injection and Combustion Flowfield Phenomena
NASA Technical Reports Server (NTRS)
Locke, Randy J.; Hicks, Yolanda R.; Anderson, Robert C.
Over the past decade advanced optical diagnostic techniques have evolved and matured to a point where they are now widely applied in the interrogation of high pressure combusting flows. At NASA Glenn Research Center (GRC), imaging techniques have been used successfully in on-going work to develop the next generation of commercial aircraft gas turbine combustors. This work has centered on providing a means by which researchers and designers can obtain direct visual observation and measurements of the fuel injection/mixing/combustion processes and combustor flowfield in two- and three-dimensional views at actual operational conditions. Obtaining a thorough understanding of the chemical and physical processes at the extreme operating conditions of the next generation of combustors is critical to reducing emissions and increasing fuel efficiency. To accomplish this and other tasks, the diagnostic team at GRC has designed and constructed optically accessible, high pressurer high temperature flame tubes and sectar rigs capable of optically probing the 20-60 atm flowfields of these aero-combustors. Among the techniques employed at GRC are planar laser-induced fluorescence (PLIF) for imaging molecular species as well as liquid and gaseous fuel; planar light scattering (PLS) for imaging fuel sprays and droplets; and spontaneous Raman scattering for species and temperature measurement. Using these techniques, optical measurements never before possible have been made in the actual environments of liquid fueled gas turbines. 2-D mapping of such parameters as species (e.g. OH-, NO and kerosene-based jet fuel) distribution, injector spray angle, and fuel/air distribution are just some of the measurements that are now routinely made. Optical imaging has also provided prompt feedback to researchers regarding the effects of changes in the fuel injector configuration on both combustor performance and flowfield character. Several injector design modifications and improvements have
Numerical methods for molecular dynamics. Progress report
Skeel, R.D.
This report summarizes our research progress to date on the use of multigrid methods for three-dimensional elliptic partial differential equations, with particular emphasis on application to the Poisson-Boltzmann equation of molecular biophysics. This research is motivated by the need for fast and accurate numerical solution techniques for three-dimensional problems arising in physics and engineering. In many applications these problems must be solved repeatedly, and the extremely large number of discrete unknowns required to accurately approximate solutions to partial differential equations in three-dimensional regions necessitates the use of efficient solution methods. This situation makes clear the importance of developing methods which are of optimal order (or nearly so), meaning that the number of operations required to solve the discrete problem is on the order of the number of discrete unknowns. Multigrid methods are generally regarded as being in this class of methods, and are in fact provably optimal order for an increasingly large class of problems. The fundamental goal of this research is to develop a fast and accurate numerical technique, based on multi-level principles, for the solutions of the Poisson-Boltzmann equation of molecular biophysics and similar equations occurring in other applications. An outline of the report is as follows. We first present some background material, followed by a survey of the literature on the use of multigrid methods for solving problems similar to the Poisson-Boltzmann equation. A short description of the software we have developed so far is then given, and numerical results are discussed. Finally, our research plans for the coming year are presented.
Conventional, molecular methods and biomarkers molecules in detection of septicemia
Arabestani, Mohammad Reza; Rastiany, Sahar; Kazemi, Sima; Mousavi, Seyed Masoud
Sepsis is a leading cause of morbidity and mortality in hospitalized patients worldwide and based on studies, 30–40% of all cases of severe sepsis and septic shock results from the blood stream infections (BSIs). Identifying of the disease, performing laboratory tests, and consequently treatment are factors that required for optimum management of BSIs. In addition, applying precise and immediate identification of the etiologic agent is a prerequisite for specific antibiotic therapy of pathogen and thereby decreasing mortality rates. The diagnosis of sepsis is difficult because clinical signs of sepsis often overlap with other noninfectious cases of systemic inflammation. BSIs are usually diagnosed by performing a series of techniques such as blood cultures, polymerase chain reaction-based methods, and biomarkers of sepsis. Extremely time-consuming even to take up to several days is a major limitation of conventional methods. In addition, yielding false-negative results due to fastidious and slow-growing microorganisms and also in case of antibiotic pretreated samples are other limitations. In comparison, molecular methods are capable of examining a blood sample obtained from suspicious patient with BSI and gave the all required information to prescribing antimicrobial therapy for detected bacterial or fungal infections immediately. Because of an emergency of sepsis, new methods are being developed. In this review, we discussed about the most important sepsis diagnostic methods and numbered the advantage and disadvantage of the methods in detail. PMID:26261822
Conventional, molecular methods and biomarkers molecules in detection of septicemia.
Arabestani, Mohammad Reza; Rastiany, Sahar; Kazemi, Sima; Mousavi, Seyed Masoud
Sepsis is a leading cause of morbidity and mortality in hospitalized patients worldwide and based on studies, 30-40% of all cases of severe sepsis and septic shock results from the blood stream infections (BSIs). Identifying of the disease, performing laboratory tests, and consequently treatment are factors that required for optimum management of BSIs. In addition, applying precise and immediate identification of the etiologic agent is a prerequisite for specific antibiotic therapy of pathogen and thereby decreasing mortality rates. The diagnosis of sepsis is difficult because clinical signs of sepsis often overlap with other noninfectious cases of systemic inflammation. BSIs are usually diagnosed by performing a series of techniques such as blood cultures, polymerase chain reaction-based methods, and biomarkers of sepsis. Extremely time-consuming even to take up to several days is a major limitation of conventional methods. In addition, yielding false-negative results due to fastidious and slow-growing microorganisms and also in case of antibiotic pretreated samples are other limitations. In comparison, molecular methods are capable of examining a blood sample obtained from suspicious patient with BSI and gave the all required information to prescribing antimicrobial therapy for detected bacterial or fungal infections immediately. Because of an emergency of sepsis, new methods are being developed. In this review, we discussed about the most important sepsis diagnostic methods and numbered the advantage and disadvantage of the methods in detail. PMID:26261822
JAK2 V617F in Myeloid Disorders: Molecular Diagnostic Techniques and Their Clinical Utility
Steensma, David P.
In early 2005, several groups of investigators studying myeloid malignancies described a novel somatic point mutation (V617F) in the conserved autoinhibitory pseudokinase domain of the Janus kinase 2 (JAK2) protein, which plays an important role in normal hematopoietic growth factor signaling. The V617F mutation is present in blood and marrow from a large proportion of patients with classic BCR/ABL-negative chronic myeloproliferative disorders and of a few patients with other clonal hematological diseases such as myelodysplastic syndrome, atypical myeloproliferative disorders, and acute myeloid leukemia. The JAK2 V617F mutation causes constitutive activation of the kinase, with deregulated intracellular signaling that mimics continuous hematopoietic growth factor stimulation. Within 7 months of the first electronic publication describing this new mutation, clinical molecular diagnostic laboratories in the United States and Europe began offering JAK2 mutation testing on a fee-for-service basis. Here, I review the various techniques used by research groups and clinical laboratories to detect the genetic mutation underlying JAK2 V617F, including fluorescent dye chemistry sequencing, allele-specific polymerase chain reaction (PCR), real-time PCR, DNA-melting curve analysis, pyrosequencing, and others. I also discuss diagnostic sensitivity, performance, and other practical concerns relevant to the clinical laboratorian in addition to the potential diagnostic utility of JAK2 mutation tests. PMID:16931578
Infectious Disease Management through Point-of-Care Personalized Medicine Molecular Diagnostic Technologies
Bissonnette, Luc; Bergeron, Michel G.
Infectious disease management essentially consists in identifying the microbial cause(s) of an infection, initiating if necessary antimicrobial therapy against microbes, and controlling host reactions to infection. In clinical microbiology, the turnaround time of the diagnostic cycle (>24 hours) often leads to unnecessary suffering and deaths; approaches to relieve this burden include rapid diagnostic procedures and more efficient transmission or interpretation of molecular microbiology results. Although rapid nucleic acid-based diagnostic testing has demonstrated that it can impact on the transmission of hospital-acquired infections, we believe that such life-saving procedures should be performed closer to the patient, in dedicated 24/7 laboratories of healthcare institutions, or ideally at point of care. While personalized medicine generally aims at interrogating the genomic information of a patient, drug metabolism polymorphisms, for example, to guide drug choice and dosage, personalized medicine concepts are applicable in infectious diseases for the (rapid) identification of a disease-causing microbe and determination of its antimicrobial resistance profile, to guide an appropriate antimicrobial treatment for the proper management of the patient. The implementation of point-of-care testing for infectious diseases will require acceptance by medical authorities, new technological and communication platforms, as well as reimbursement practices such that time- and life-saving procedures become available to the largest number of patients. PMID:25562799
Electronic detection of nucleic acids: a versatile platform for molecular diagnostics.
Umek, R M; Lin, S W; Vielmetter, J; Terbrueggen, R H; Irvine, B; Yu, C J; Kayyem, J F; Yowanto, H; Blackburn, G F; Farkas, D H; Chen, Y P
A novel platform for the electronic detection of nucleic acids on microarrays is introduced and shown to perform well as a selective detection system for applications in molecular diagnostics. A gold electrode in a printed circuit board is coated with a self-assembled monolayer (SAM) containing DNA capture probes. Unlabeled nucleic acid targets are immobilized on the surface of the SAM through sequence-specific hybridization with the DNA capture probe. A separate signaling probe, containing ferrocene-modified nucleotides and complementary to the target in the region adjoining the capture probe binding site, is held in close proximity to the SAM in a sandwich complex. The SAM allows electron transfer between the immobilized ferrocenes and the gold, while insulating the electrode from soluble redox species, including unbound signaling probes. Here, we demonstrate sequence-specific detection of amplicons after simple dilution of the reaction product into hybridization buffer. In addition, single nucleotide polymorphism discrimination is shown. A genotyping chip for the C282Y single nucleotide polymorphism associated with hereditary hemochromatosis is used to confirm the genotype of six patients' DNA. In addition, a gene expression-monitoring chip is described that surveys five genes that are differentially regulated in the cellular apoptosis response. Finally, custom modification of individual electrodes through sequence-specific hybridization demonstrates the potential of this system for infectious disease diagnostics. The versatility of the electronic detection platform makes it suitable for multiple applications in diagnostics and pharmacogenetics. PMID:11333303
[Clinical usefulness of diagnostic methods for human papilloma virus dependent lesions].
Suwalska, Anna; Owczarek, Witold; Fiedor, Piotr
Persistent infection of Human Papilloma Virus (HPV) is confirmed necessary factor for development of cervical cancer and anogenital neoplasia. DNA HPV is detected in 96% of cervical cancer, 40% of vulvar and vaginal cancer, 90% of anal cancer and 26% of oral cavity cancer cases in general population. The most common high-risk HPV types observed in anogenital intraepithelial neoplasia or anogenital cancer are HPV 16, 18 and 45. Numerous diagnostic methods of detection of HPV infection and lesions causes by persistent HPV infection are widely used. Epidemiological data reveals correlation of incidence and mortality reduction due to cervical cancer and consequent prosecution and improvement of screening programmes based on morphological assessment of exfoliative smears. In last decade some limitations of conventional smear method were pointed out and a new diagnostic techniques were introduced: liquid-based cytology and HPV DNA testing. Combination of cytological examination and HPV DNA testing seems to be optimal solution to be introduced in large population because of combining high sensitivity of molecular test with high specificity of cytological smear. PMID:24720112
Scratching the surface of tomorrow's diagnostics: the Editor-in-Chief's opinion at the 15th year of Expert Review of Molecular Diagnostics.
Lorincz, Attila; Raison, Claire
Interview with Attila Lorincz by Claire Raison (Commissioning Editor) To mark the beginning of the 15th year of Expert Review of Molecular Diagnostics, the journal's Editor-in-Chief shares his expert knowledge on translational diagnostics, his opinion on recent controversies and his predictions for molecular diagnostics in 2015 and beyond. Attila Lorincz received his doctorate from Trinity College, Dublin, Republic of Ireland, and went on to become a research fellow at the University of California, Santa Barbara, CA, USA. During Professor Lorincz's research on human papillomavirus (HPV), he found several important and novel carcinogenic HPV types and pioneered the use of HPV DNA testing for clinical diagnostics. In 1988, Professor Lorincz's team produced the first HPV test to be FDA-approved for patients and in 2003, for general population cervical precancer screening. Now Professor of Molecular Epidemiology at the Centre for Cancer Prevention, Queen Mary University of London, UK, he and his team are furthering translational research into DNA methylation assays for cancer risk prediction. PMID:25537568
Molecular Gas Kinematics and Line Diagnostics in Early-type Galaxies: NGC 4710 & NGC 5866
NASA Astrophysics Data System (ADS)
Topal, Selçuk; Bureau, Martin; Davis, Timothy A.; Krips, Melanie; Young, Lisa M.; Crocker, Alison F.
We present interferometric observations of CO lines (12CO(1-0, 2-1) and 13CO(1-0, 2-1)) and dense gas tracers (HCN(1-0), HCO+(1-0), HNC(1-0) and HNCO(4-3)) in two nearby edge-on barred lenticular galaxies, NGC 4710 and NGC 5866, with most of the gas concentrated in a nuclear disc and an inner ring in each galaxy. We probe the physical conditions of a two-component molecular interstellar medium in each galaxy and each kinematic component by using molecular line ratio diagnostics in three complementary ways. First, we measure the ratios of the position-velocity diagrams of different lines, second we measure the ratios of each kinematic component's integrated line intensities as a function of projected position, and third we model these line ratios using a non-local thermodynamic equilibrium radiative transfer code. Overall, the nuclear discs appear to have a tenuous molecular gas component that is hotter, optically thinner and with a larger dense gas fraction than that in the inner rings, suggesting more dense clumps immersed in a hotter more diffuse molecular medium. This is consistent with evidence that the physical conditions in the nuclear discs are similar to those in photo-dissociation regions. A similar picture emerges when comparing the observed molecular line ratios with those of other galaxy types. The physical conditions of the molecular gas in the nuclear discs of NGC 4710 and NGC 5866 thus appear intermediate between those of spiral galaxies and starbursts, while the star formation in their inner rings is even milder.
[Cognitive functions, their development and modern diagnostic methods].
Klasik, Adam; Janas-Kozik, Małgorzata; Krupka-Matuszczyk, Irena; Augustyniak, Ewa
provided a theory. The psychometric approach concentrates on studying the differences in intelligence. The aim of this approach is to test intelligence by means of standardized tests (e.g. WISC-R, WAIS-R) used to show the individual differences among humans. Human cognitive functions determine individuals' adaptation capabilities and disturbances in this area indicate a number of psychopathological changes and are a symptom enabling to differentiate or diagnose one with a disorder. That is why the psychological assessment of cognitive functions is an important part of patients' diagnosis. Contemporary neuropsychological studies are to a great extent based computer tests. The use of computer methods has a number of measurement-related advantages. It allows for standardized testing environment, increasing therefore its reliability and standardizes the patient assessment process. Special attention should be paid to the neuropsychological tests included in the Vienna Test System (Cognitron, SIGNAL, RT, VIGIL, DAUF), which are used to assess the operational memory span, learning processes, reaction time, attention selective function, attention continuity as well as attention interference resistance. It also seems justified to present the CPT id test (Continuous Performance Test) as well as Free Recall. CPT is a diagnostic tool used to assess the attention selective function, attention continuity of attention, attention interference resistance as well as attention alertness. The Free Recall test is used in the memory processes diagnostics to assess patients' operational memory as well as the information organization degree in operational memory. The above mentioned neuropsychological tests are tools used in clinical assessment of cognitive function disorders. PMID:17471820
Use of Molecular Diagnostic Tools for the Identification of Species Responsible for Snakebite in Nepal: A Pilot Study
Sharma, Sanjib Kumar; Kuch, Ulrich; Höde, Patrick; Bruhse, Laura; Pandey, Deb P.; Ghimire, Anup; Chappuis, François; Alirol, Emilie
Snakebite is an important medical emergency in rural Nepal. Correct identification of the biting species is crucial for clinicians to choose appropriate treatment and anticipate complications. This is particularly important for neurotoxic envenoming which, depending on the snake species involved, may not respond to available antivenoms. Adequate species identification tools are lacking. This study used a combination of morphological and molecular approaches (PCR-aided DNA sequencing from swabs of bite sites) to determine the contribution of venomous and non-venomous species to the snakebite burden in southern Nepal. Out of 749 patients admitted with a history of snakebite to one of three study centres, the biting species could be identified in 194 (25.9%). Out of these, 87 had been bitten by a venomous snake, most commonly the Indian spectacled cobra (Naja naja; n = 42) and the common krait (Bungarus caeruleus; n = 22). When both morphological identification and PCR/sequencing results were available, a 100% agreement was noted. The probability of a positive PCR result was significantly lower among patients who had used inadequate "first aid" measures (e.g. tourniquets or local application of remedies). This study is the first to report the use of forensic genetics methods for snake species identification in a prospective clinical study. If high diagnostic accuracy is confirmed in larger cohorts, this method will be a very useful reference diagnostic tool for epidemiological investigations and clinical studies. PMID:27105074
Use of Molecular Diagnostic Tools for the Identification of Species Responsible for Snakebite in Nepal: A Pilot Study.
Sharma, Sanjib Kumar; Kuch, Ulrich; Höde, Patrick; Bruhse, Laura; Pandey, Deb P; Ghimire, Anup; Chappuis, François; Alirol, Emilie
Snakebite is an important medical emergency in rural Nepal. Correct identification of the biting species is crucial for clinicians to choose appropriate treatment and anticipate complications. This is particularly important for neurotoxic envenoming which, depending on the snake species involved, may not respond to available antivenoms. Adequate species identification tools are lacking. This study used a combination of morphological and molecular approaches (PCR-aided DNA sequencing from swabs of bite sites) to determine the contribution of venomous and non-venomous species to the snakebite burden in southern Nepal. Out of 749 patients admitted with a history of snakebite to one of three study centres, the biting species could be identified in 194 (25.9%). Out of these, 87 had been bitten by a venomous snake, most commonly the Indian spectacled cobra (Naja naja; n = 42) and the common krait (Bungarus caeruleus; n = 22). When both morphological identification and PCR/sequencing results were available, a 100% agreement was noted. The probability of a positive PCR result was significantly lower among patients who had used inadequate "first aid" measures (e.g. tourniquets or local application of remedies). This study is the first to report the use of forensic genetics methods for snake species identification in a prospective clinical study. If high diagnostic accuracy is confirmed in larger cohorts, this method will be a very useful reference diagnostic tool for epidemiological investigations and clinical studies. PMID:27105074
Electrically heated particulate filter diagnostic systems and methods
Gonze, Eugene V [Pinckney, MI
A system that diagnoses regeneration of an electrically heated particulate filter is provided. The system generally includes a grid module that diagnoses a fault of the grid based on at least one of a current signal and a voltage signal. A diagnostic module at least one of sets a fault status and generates a warning signal based on the fault of the grid.
Comparison of Self-Instruction Methods for Teaching Diagnostic Testing.
ERIC Educational Resources Information Center
Puskas, Jane C.
Self-teaching booklets and computer media were evaluated for teaching diagnostic testing with first (n=49), second (n=41) and third year (n=71) dental students as a foundation for further development of clinical decision-making skills. Results found the media more effective than no instruction and equally effective to the traditional lecture…
Sherlock Holmes' methods of deductive reasoning applied to medical diagnostics.
Miller, L
Having patterned the character of Sherlock Holmes after one of his professors, Sir Arthur Conan Doyle, himself a physician, incorporated many of the didactic qualities of the 19th century medical diagnostician into the character of Holmes. In this paper I explore Holmes's techniques of deductive reasoning and their basis in 19th and 20th century medical diagnostics. PMID:3887762
Sherlock Holmes's Methods of Deductive Reasoning Applied to Medical Diagnostics
Miller, Larry
Having patterned the character of Sherlock Holmes after one of his professors, Sir Arthur Conan Doyle, himself a physician, incorporated many of the didactic qualities of the 19th century medical diagnostician into the character of Holmes. In this paper I explore Holmes's techniques of deductive reasoning and their basis in 19th and 20th century medical diagnostics. PMID:3887762
Enterovirus infections in England and Wales, 2000-2011: the impact of increased molecular diagnostics.
Kadambari, S; Bukasa, A; Okike, I O; Pebody, R; Brown, D; Gallimore, C; Xerry, J; Sharland, M; Ladhani, S N
There have recently been significant changes in diagnostic practices for detecting enterovirus (EV) infections across England and Wales. Reports of laboratory-confirmed EV infections submitted by National Health Service (NHS) hospital laboratories to Public Health England (PHE) over a 12-year period (2000-2011) were analysed. Additionally, the PHE Virus Reference Department (VRD) electronic database containing molecular typing data from 2004 onwards was interrogated. Of the 13,901 reports, there was a decline from a peak of 2254 in 2001 to 589 in 2006, and then an increase year-on-year to 1634 in 2011. This increase coincided with increasing PCR-based laboratory diagnosis, which accounted for 36% of reported cases in 2000 and 92% in 2011. The estimated annual incidence in 2011 was 3.9/100,000 overall and 238/100,000 in those aged <3 months, who accounted for almost one-quarter of reported cases (n = 2993, 23%). During 2004-2011, 2770 strains were submitted for molecular typing to the VRD, who found no evidence for a predominance of any particular strain. Thus, the recent increase in reported cases closely reflects the increase in PCR testing by NHS hospitals, but is associated with a lower proportion of samples being submitted for molecular typing. The high EV rate in young infants merits further investigation to inform evidence-based management guidance. PMID:25039903
Coccidia of turkey: from isolation, characterisation and comparison to molecular phylogeny and molecular diagnostics.
Vrba, Vladimir; Pakandl, Michal
Coccidiosis is a disease caused by apicomplexan parasites of the genus Eimeria, which has a significant economic impact on poultry production. Multiple species infecting the turkey have been described; however, due to the general lack of unambiguous description, their identification and taxonomy is debatable. In this work, a systematic approach was taken to isolate, characterise and compare coccidian species in the turkey. Individual species were tracked according to their unique 18S ribosomal DNA sequence. The single-oocyst isolation technique and passaging of mixed species field isolates in selectively immunised birds enabled the derivation of pure species. Six distinct strains representing five eimerian species that infect the turkey were obtained. It appears highly probable that these species represent all species described in the past with the exception of Eimeria subrotunda. The species were analysed using both traditional methods and DNA sequencing. For each strain the oocyst morphology, prepatent period, gross pathology, pathogenicity, host specificity and endogenous cycle were studied. Antigenic similarity was investigated in multiple cross-immunity experiments. For identification and quantification of each individual species or strain, quantitative real-time PCR markers were also developed. Parallel characterisation of pure strains allowed comprehensive comparison with the original descriptions and assignment of correct species names. The species Eimeria meleagridis, Eimeria dispersa, Eimeria gallopavonis, Eimeria meleagrimitis and Eimeria innocua were identified. Comparison of our data with those of previous studies indicates that Eimeria adenoeides is most probably a synonym for either E. meleagridis or E. gallopavonis, or a description based on a mixture of these species, and thus nomen dubium. The species E. dispersa and E. innocua were also found to infect Bobwhite Quail. Phylogenetic reconstruction based on 18S rDNA and cytochrome c oxidase subunit
Method of making molecularly doped composite polymer material
Affinito, John D [Tucson, AZ; Martin, Peter M [Kennewick, WA; Graff, Gordon L [West Richland, WA; Burrows, Paul E [Kennewick, WA; Gross, Mark E. , Sapochak, Linda S.
A method of making a composite polymer of a molecularly doped polymer. The method includes mixing a liquid polymer precursor with molecular dopant forming a molecularly doped polymer precursor mixture. The molecularly doped polymer precursor mixture is flash evaporated forming a composite vapor. The composite vapor is cryocondensed on a cool substrate forming a composite molecularly doped polymer precursor layer, and the cryocondensed composite molecularly doped polymer precursor layer is cross linked thereby forming a layer of the composite polymer layer of the molecularly doped polymer.
[Survey of studies on adjuvant diagnostic method of stimulating auricular points at home and abroad].
Chen, Gong-Sun; Hu, Zhi-Hui; Zhu, Bing
The auricular (including auricular acupoints) adjuvant diagnostic method, besides inspection (including dyeing method), palpation (including thermometric method), tenderness method (including impressing method) and electrical detection, includes the adjuvant diagnostic method of stimulating auricular points as well, it has been mostly studied and used by specialists of western medicine or doctors of integrated Chinese and western medicine. But it hasn't been introduced in the published Chinese monographs of auricular acupuncture yet. This article briefly introduces the adjuvant diagnostic method of stimulating auricular points combined with X-ray radiography; application in fetal heart electronic monitoring and fetal biophysical monitoring; and diagnostic methods of auricle reflex, vascular autonomous signals, and auricle and somatic 7 frequency response regions, which began to be researched abroad 35 years ago. The authors hope it will give some invigoration or illumination to my colleagues in acupuncture, especially those who are interested in auricular acupuncture. PMID:18271243
Speckle methods for diagnostics of the human oral cavity
NASA Astrophysics Data System (ADS)
Kharish, Natalia A.; Sedykh, Alexey V.; Ulyanov, Sergey S.; Lepilin, Alexander V.; Tuchin, Valery V.
Possibility of application of speckle interferometry for diagnostics in dentistry has been analyzed. Problem of standardization of the measuring procedure has been studied. Deviation of output characteristics of Doppler system for blood microcirculation measurements has been investigated. Dependence of form of Doppler spectrum on the degree of seriousness of diseases has been studied in experiments in vivo. Behavior of spectral moments of measuring signal during the treatment of parodontitis has been analyzed.
Molecular tumor markers for asbestos-related mesothelioma: serum diagnostic markers.
Maeda, Masahiro; Hino, Okio
Mesothelioma is an aggressive tumor arising from the mesothelium, and is usually associated with previous exposure to asbestos. The incubation period of the tumor may be described as 30-40 years, and the prognosis is dismal. In addition to immunohistochemical markers, recently, serum markers for the diagnosis of mesothelioma have been reported as candidates. In contrast, the expression in renal carcinoma (ERC) gene has been discovered in the Eker rat model (Tsc2 gene mutant), which is a homolog of the human mesothelin/megakaryocyte potentiating factor gene, and a novel ELISA system (N-ERC/mesothelin) has been developed. It has also been found that N-ERC/mesothelin is very stable and plentiful in the blood. In the present paper the potential utility of molecular diagnostic markers is reviewed, including ELISA systems for asbestos-related mesothelioma. PMID:17040286
Fluorescence-Raman Dual Modal Endoscopic System for Multiplexed Molecular Diagnostics
NASA Astrophysics Data System (ADS)
Jeong, Sinyoung; Kim, Yong-Il; Kang, Homan; Kim, Gunsung; Cha, Myeong Geun; Chang, Hyejin; Jung, Kyung Oh; Kim, Young-Hwa; Jun, Bong-Hyun; Hwang, Do Won; Lee, Yun-Sang; Youn, Hyewon; Lee, Yoon-Sik; Kang, Keon Wook; Lee, Dong Soo; Jeong, Dae Hong
Optical endoscopic imaging, which was recently equipped with bioluminescence, fluorescence, and Raman scattering, allows minimally invasive real-time detection of pathologies on the surface of hollow organs. To characterize pathologic lesions in a multiplexed way, we developed a dual modal fluorescence-Raman endomicroscopic system (FRES), which used fluorescence and surface-enhanced Raman scattering nanoprobes (F-SERS dots). Real-time, in vivo, and multiple target detection of a specific cancer was successful, based on the fast imaging capability of fluorescence signals and the multiplex capability of simultaneously detected SERS signals using an optical fiber bundle for intraoperative endoscopic system. Human epidermal growth factor receptor 2 (HER2) and epidermal growth factor receptor (EGFR) on the breast cancer xenografts in a mouse orthotopic model were successfully detected in a multiplexed way, illustrating the potential of FRES as a molecular diagnostic instrument that enables real-time tumor characterization of receptors during routine endoscopic procedures.
Fluorescence-Raman Dual Modal Endoscopic System for Multiplexed Molecular Diagnostics
Jeong, Sinyoung; Kim, Yong-il; Kang, Homan; Kim, Gunsung; Cha, Myeong Geun; Chang, Hyejin; Jung, Kyung Oh; Kim, Young-Hwa; Jun, Bong-Hyun; Hwang, Do Won; Lee, Yun-Sang; Youn, Hyewon; Lee, Yoon-Sik; Kang, Keon Wook; Lee, Dong Soo; Jeong, Dae Hong
Optical endoscopic imaging, which was recently equipped with bioluminescence, fluorescence, and Raman scattering, allows minimally invasive real-time detection of pathologies on the surface of hollow organs. To characterize pathologic lesions in a multiplexed way, we developed a dual modal fluorescence-Raman endomicroscopic system (FRES), which used fluorescence and surface-enhanced Raman scattering nanoprobes (F-SERS dots). Real-time, in vivo, and multiple target detection of a specific cancer was successful, based on the fast imaging capability of fluorescence signals and the multiplex capability of simultaneously detected SERS signals using an optical fiber bundle for intraoperative endoscopic system. Human epidermal growth factor receptor 2 (HER2) and epidermal growth factor receptor (EGFR) on the breast cancer xenografts in a mouse orthotopic model were successfully detected in a multiplexed way, illustrating the potential of FRES as a molecular diagnostic instrument that enables real-time tumor characterization of receptors during routine endoscopic procedures. PMID:25820115
Carbon Nanotube Biosensors for Space Molecule Detection and Clinical Molecular Diagnostics
NASA Technical Reports Server (NTRS)
Han, Jie
Both space molecule detection and clinical molecule diagnostics need to develop ultra sensitive biosensors for detection of less than attomole molecules such as amino acids for DNA. However all the electrode sensor systems including those fabricated from the existing carbon nanotubes, have a background level of nA (nanoAmp). This has limited DNA or other molecule detection to nA level or molecules whose concentration is, much higher than attomole level. A program has been created by NASA and NCI (National Cancer Institute) to exploit the possibility of carbon nanotube based biosensors to solve this problem for both's interest. In this talk, I will present our effort on the evaluation and novel design of carbon nanotubes as electrode biosensors with strategies to minimize background currents while maximizing signal intensity.The fabrication of nanotube electrode arrays, immobilization of molecular probes on nanotube electrodes and in vitro biosensor testing will also be discussed.
Plekhanova, O S; Naumova, E V; Lugovskaya, S A; Potchtar, M E; Bugrov, I Yu; Dolgov, V V
The article presents diagnostic of night paroxysmal hemoglobinuria. The night paroxysmal hemoglobinuria is an orphan disease characterized by absence of GPI-anchor on blood cells as a result of mutation of PIG-A gene on the short arm of X-chromosome. The particular proteins bounded with GPI-anchor implement function of defense from activation of components of complement and development of membrane-attacking complex. The erythrocytes exposed to destruction in bloodstream are among the most impacted. Therefore, one of the main signs of night paroxysmal hemoglobinuria is complement-depending intravascular hemolysis which indicators for a long time played a key role in diagnostic of night paroxysmal hemoglobinuria. The actual technique of diagnostic of night paroxysmal hemoglobinuria is flow cytometry. The analysis of night paroxysmal hemoglobinuria clone is recommended to patients with hemolysis of unclear genesis, thrombosis of cerebral and abdominal veins, thrombocytopenia and macrocytosis and also patients with AA, myelodysplastic syndrome, myelofibrosis. The international protocol recommended by the International Society of Clinical Cytometry (2010) is implemented to diagnose night paroxysmal hemoglobinuria. The original technique of evaluation of reticulocytes was developed with purpose to detect night paroxysmal hemoglobinuria clone. The high correlation was substantiated between size of night paroxysmal hemoglobinuria clone measured among reticulocytes according to proposed mode and night paroxysmal hemoglobinuria clone measured among granulocytes and monocytes detected according international standardized approach. PMID:27506106
Next-Generation Sequencing in Clinical Molecular Diagnostics of Cancer: Advantages and Challenges
Luthra, Rajyalakshmi; Chen, Hui; Roy-Chowdhuri, Sinchita; Singh, R. Rajesh
The application of next-generation sequencing (NGS) to characterize cancer genomes has resulted in the discovery of numerous genetic markers. Consequently, the number of markers that warrant routine screening in molecular diagnostic laboratories, often from limited tumor material, has increased. This increased demand has been difficult to manage by traditional low- and/or medium-throughput sequencing platforms. Massively parallel sequencing capabilities of NGS provide a much-needed alternative for mutation screening in multiple genes with a single low investment of DNA. However, implementation of NGS technologies, most of which are for research use only (RUO), in a diagnostic laboratory, needs extensive validation in order to establish Clinical Laboratory Improvement Amendments (CLIA) and College of American Pathologists (CAP)-compliant performance characteristics. Here, we have reviewed approaches for validation of NGS technology for routine screening of tumors. We discuss the criteria for selecting gene markers to include in the NGS panel and the deciding factors for selecting target capture approaches and sequencing platforms. We also discuss challenges in result reporting, storage and retrieval of the voluminous sequencing data and the future potential of clinical NGS. PMID:26473927
In search of a potential diagnostic tool for molecular characterization of lymphatic filariasis.
Saeed, Mohd; Adnan, Mohd; Khan, Saif; Al-Shammari, Eyad; Mustafa, Huma
Lymphatic filariasis (LF) is a chronic disease and is caused by the parasites Wuchereria bancrofti (W. bancrofti), Brugia malayi (B. malayi) and Brugia timori (B. timori). In the present study, Setaria cervi (S. cervi), a bovine filarial parasite has been used. Previously, it has been reported that the S. cervi shares some common proteins and antigenic determinants with that of human filarial parasite. The larval stages of filarial species usually cannot be identified by classical morphology. Hence, molecular characterization allows the identification of the parasites throughout all their developmental stages. The genomic DNA of S. cervi adult were isolated and estimated spectrophotometrically for the quantitative presence of DNA content. Screening of DNA sequences from filarial DNA GenBank and Expressed Sequence Tags (EST's) were performed for homologous sequences and then multiple sequence alignment was executed. The conserved sequences from multiple sequence alignment were used for In Silico primer designing. The successfully designed primers were used further in PCR amplifications. Therefore, in search of a promising diagnostic tool few genes were identified to be conserved in the human and bovine filariasis and these novel primers deigned may help to develop a promising diagnostic tool for identification of lymphatic filariasis. PMID:26751881
Next-Generation Sequencing in Clinical Molecular Diagnostics of Cancer: Advantages and Challenges.
Luthra, Rajyalakshmi; Chen, Hui; Roy-Chowdhuri, Sinchita; Singh, R Rajesh
The application of next-generation sequencing (NGS) to characterize cancer genomes has resulted in the discovery of numerous genetic markers. Consequently, the number of markers that warrant routine screening in molecular diagnostic laboratories, often from limited tumor material, has increased. This increased demand has been difficult to manage by traditional low- and/or medium-throughput sequencing platforms. Massively parallel sequencing capabilities of NGS provide a much-needed alternative for mutation screening in multiple genes with a single low investment of DNA. However, implementation of NGS technologies, most of which are for research use only (RUO), in a diagnostic laboratory, needs extensive validation in order to establish Clinical Laboratory Improvement Amendments (CLIA) and College of American Pathologists (CAP)-compliant performance characteristics. Here, we have reviewed approaches for validation of NGS technology for routine screening of tumors. We discuss the criteria for selecting gene markers to include in the NGS panel and the deciding factors for selecting target capture approaches and sequencing platforms. We also discuss challenges in result reporting, storage and retrieval of the voluminous sequencing data and the future potential of clinical NGS. PMID:26473927
Hepatocellular Carcinoma, Fibrolamellar Variant: Diagnostic Pathologic Criteria and Molecular Pathology Update. A Primer
Sergi, Consolato M.
Fibrolamellar hepatocellular carcinoma (FL-HCC) is generally a fairly rare event in routine pathology practice. This variant of hepatocellular carcinoma (HCC) is peculiarly intriguing and,in addition, poorly understood. Young people or children are often the target individuals with this type of cancer. Previously, I highlighted some pathology aspects of FL-HCC, but in this review, the distinctive clinico-pathologic features of FL-HCC and the diagnostic pathologic criteria of FL-HCC are fractionally reviewed and expanded upon. Further, molecular genetics update data with reference to this specific tumor are particularly highlighted as a primer for general pathologists and pediatric histopathologists. FL-HCC may present with metastases, and regional lymph nodes may be sites of metastatic spread. However, peritoneal and pulmonary metastatic foci have also been reported. To the best of our knowledge, FL-HCC was initially considered having an indolent course, but survival outcomes have recently been updated reconsidering the prognosis of this tumor. Patients seem to respond well to surgical resection, but recurrences are common. Thus, alternative therapies, such as chemotherapy and radiation, are ongoing. Overall, it seems that this aspect has not been well-studied for this variant of HCC and should be considered as target for future clinical trials. Remarkably, FL-HCC data seem to point to a liver neoplasm of uncertain origin and unveiled outcome. A functional chimeric transcript incorporating DNAJB1 and PRKACA was recently added to FL-HCC. This sensational result may give remarkable insights into the understanding of this rare disease and potentially provide the basis for its specific diagnostic marker. Detection of DNAJB1-PRKACA seems to be, indeed, a very sensitive and specific finding in supporting the diagnosis of FL-HCC. In a quite diffuse opinion, prognosis of this tumor should be reconsidered following the potentially mandatory application of new molecular
Expression profiling shows differential molecular pathways and provides potential new diagnostic biomarkers for colorectal serrated adenocarcinoma.
Conesa-Zamora, Pablo; García-Solano, José; García-García, Francisco; Turpin, María Del Carmen; Trujillo-Santos, Javier; Torres-Moreno, Daniel; Oviedo-Ramírez, Isabel; Carbonell-Muñoz, Rosa; Muñoz-Delgado, Encarnación; Rodriguez-Braun, Edith; Conesa, Ana; Pérez-Guillermo, Miguel
Serrated adenocarcinoma (SAC) is a recently recognized colorectal cancer (CRC) subtype accounting for 7.5 to 8.7% of CRCs. It has been shown that SAC has a poorer prognosis and has different molecular and immunohistochemical features compared with conventional carcinoma (CC) but, to date, only one previous study has analyzed its mRNA expression profile by microarray. Using a different microarray platform, we have studied the molecular signature of 11 SACs and compared it with that of 15 matched CC with the aim of discerning the functions which characterize SAC biology and validating, at the mRNA and protein level, the most differentially expressed genes which were also tested using a validation set of 70 SACs and 70 CCs to assess their diagnostic and prognostic values. Microarray data showed a higher representation of morphogenesis-, hypoxia-, cytoskeleton- and vesicle transport-related functions and also an overexpression of fascin1 (actin-bundling protein associated with invasion) and the antiapoptotic gene hippocalcin in SAC all of which were validated both by quantitative real-time PCR (qPCR) and immunohistochemistry. Fascin1 expression was statistically associated with KRAS mutation with 88.6% sensitivity and 85.7% specificity for SAC diagnosis and the positivity of fascin1 or hippocalcin was highly suggestive of SAC diagnosis (sensitivity = 100%). Evaluation of these markers in CRCs showing histological and molecular characteristics of high-level microsatellite instability (MSI-H) also helped to distinguish SACs from MSI-H CRCs. Molecular profiling demonstrates that SAC shows activation of distinct signaling pathways and that immunohistochemical fascin1 and hippocalcin expression can be reliably used for its differentiation from other CRC subtypes. PMID:22696308
C3H2 observations as a diagnostic probe for molecular clouds
NASA Technical Reports Server (NTRS)
Avery, L. W.
Recently the three-membered ring molecule, cyclopropenylidene, C3H2, has been identified in the laboratory and detected in molecular clouds by Thaddeus, Vrtilek and Gottlieb (1985). This molecule is wide-spread throughout the Galaxy and has been detected in 25 separate sources including cold dust clouds, circumstellar envelopes, HII regions, and the spiral arms observed against the Cas supernova remnant. In order to evaluate the potential of C3H2 as a diagnostic probe for molecular clouds, and to attempt to identify the most useful transitions, statistical equilibrium calculations were carried out for the lowest 24 levels of the ortho species and the lowest 10 levels of the para species. Many of the sources observed by Matthews and Irvine (1985) show evidence of being optically thick in the 1(10)-1(01) line. Consequently, the effects of radiative trapping should be incorporated into the equilibrium calculations. This was done using the Large Velocity Gradient approximation for a spherical cloud of uniform density. Some results of the calculations for T(K)=10K are given. Figures are presented which show contours of the logarithm of the ratio of peak line brightness temperatures for ortho-para pairs of lines at similar frequencies. It appears that the widespread nature of C3H2, the relatively large strength of its spectral lines, and their sensitivity to density and molecular abundance combine to make this a useful molecule for probing physical conditions in molecular clouds. The 1(10)-1(01) and 2(20)-2(11) K-band lines may be especially useful in this regard because of the ease with which they are observed and their unusual density-dependent emission/absorption properties.
Method matters: Understanding diagnostic reliability in DSM-IV and DSM-5.
Chmielewski, Michael; Clark, Lee Anna; Bagby, R Michael; Watson, David
Diagnostic reliability is essential for the science and practice of psychology, in part because reliability is necessary for validity. Recently, the DSM-5 field trials documented lower diagnostic reliability than past field trials and the general research literature, resulting in substantial criticism of the DSM-5 diagnostic criteria. Rather than indicating specific problems with DSM-5, however, the field trials may have revealed long-standing diagnostic issues that have been hidden due to a reliance on audio/video recordings for estimating reliability. We estimated the reliability of DSM-IV diagnoses using both the standard audio-recording method and the test-retest method used in the DSM-5 field trials, in which different clinicians conduct separate interviews. Psychiatric patients (N = 339) were diagnosed using the SCID-I/P; 218 were diagnosed a second time by an independent interviewer. Diagnostic reliability using the audio-recording method (N = 49) was "good" to "excellent" (M κ = .80) and comparable to the DSM-IV field trials estimates. Reliability using the test-retest method (N = 218) was "poor" to "fair" (M κ = .47) and similar to DSM-5 field-trials' estimates. Despite low test-retest diagnostic reliability, self-reported symptoms were highly stable. Moreover, there was no association between change in self-report and change in diagnostic status. These results demonstrate the influence of method on estimates of diagnostic reliability. PMID:26098046
Raman spectroscopy of saliva as a perspective method for periodontitis diagnostics Raman spectroscopy of saliva
NASA Astrophysics Data System (ADS)
Gonchukov, S.; Sukhinina, A.; Bakhmutov, D.; Minaeva, S.
In view of its potential for biological tissues analyses at a molecular level, Raman spectroscopy in optical range has been the object of biomedical research for the last years. The main aim of this work is the development of Raman spectroscopy for organic content identifying and determination of biomarkers of saliva at a molecular level for periodontitis diagnostics. Four spectral regions were determined: 1155 and 1525 cm-1, 1033 and 1611 cm-1, which can be used as biomarkers of this widespread disease.
Molecular method for determining sex of walruses
Fischbach, A.S.; Jay, C.V.; Jackson, J.V.; Andersen, L.W.; Sage, G.K.; Talbot, S.L.
We evaluated the ability of a set of published trans-species molecular sexing primers and a set of walrus-specific primers, which we developed, to accurately identify sex of 235 Pacific walruses (Odobenus rosmarus divergens). The trans-species primers were developed for mammals and targeted the X- and Y-gametologs of the zinc finger protein genes (ZFX, ZFY). We extended this method by using these primers to obtain sequence from Pacific and Atlantic walrus (0. r. rosmarus) ZFX and ZFY genes to develop new walrus-specific primers, which yield polymerase chain reaction products of distinct lengths (327 and 288 base pairs from the X- and Y-chromosome, respectively), allowing them to be used for sex determination. Both methods yielded a determination of sex in all but 1-2% of samples with an accuracy of 99.6-100%. Our walrus-specific primers offer the advantage of small fragment size and facile application to automated electrophoresis and visualization.
An objective method and measuring equipment for noise control and acoustic diagnostics of motorcars. [acoustic diagnostics on automobile engines
NASA Technical Reports Server (NTRS)
Kacprowski, J.; Motylewski, J.; Miazga, J.
An objective method and apparatus for noise control and acoustic diagnostics of motorcar engines are reported. The method and apparatus let us know whether the noisiness of the vehicle under test exceeds the admissible threshold levels given by appropriate standards and if so what is the main source of the excessive noise. The method consists in measuring both the overall noise level and the sound pressure levels in definite frequency bands while the engine speed is controlled as well and may be fixed at prescribed values. Whenever the individually adjusted threshold level has been exceeded in any frequency band, a self-sustaining control signal is sent.
Jet Noise Diagnostics Supporting Statistical Noise Prediction Methods
NASA Technical Reports Server (NTRS)
Bridges, James E.
The primary focus of my presentation is the development of the jet noise prediction code JeNo with most examples coming from the experimental work that drove the theoretical development and validation. JeNo is a statistical jet noise prediction code, based upon the Lilley acoustic analogy. Our approach uses time-average 2-D or 3-D mean and turbulent statistics of the flow as input. The output is source distributions and spectral directivity. NASA has been investing in development of statistical jet noise prediction tools because these seem to fit the middle ground that allows enough flexibility and fidelity for jet noise source diagnostics while having reasonable computational requirements. These tools rely on Reynolds-averaged Navier-Stokes (RANS) computational fluid dynamics (CFD) solutions as input for computing far-field spectral directivity using an acoustic analogy. There are many ways acoustic analogies can be created, each with a series of assumptions and models, many often taken unknowingly. And the resulting prediction can be easily reverse-engineered by altering the models contained within. However, only an approach which is mathematically sound, with assumptions validated and modeled quantities checked against direct measurement will give consistently correct answers. Many quantities are modeled in acoustic analogies precisely because they have been impossible to measure or calculate, making this requirement a difficult task. The NASA team has spent considerable effort identifying all the assumptions and models used to take the Navier-Stokes equations to the point of a statistical calculation via an acoustic analogy very similar to that proposed by Lilley. Assumptions have been identified and experiments have been developed to test these assumptions. In some cases this has resulted in assumptions being changed. Beginning with the CFD used as input to the acoustic analogy, models for turbulence closure used in RANS CFD codes have been explored and
The Revised Research Diagnostic Criteria for Temporomandibular Disorders: Methods used to Establish and Validate Revised Axis I Diagnostic Algorithms
Schiffman, Eric L.; Ohrbach, Richard; Truelove, Edmond L.; Feng, Tai; Anderson, Gary C.; Pan, Wei; Gonzalez, Yoly M.; John, Mike T.; Sommers, Earl; List, Thomas; Velly, Ana M.; Kang, Wenjun; Look, John O.
AIMS To derive reliable and valid revised Research Diagnostic Criteria for Temporomandibular Disorders (RDC/TMD) Axis I diagnostic algorithms for clinical TMD diagnoses. METHODS The multi-site RDC/TMD Validation Project's dataset (614 TMD community and clinic cases, and 91 controls) was used to derive revised algorithms for Axis I TMD diagnoses. Validity of diagnostic algorithms was assessed relative to reference standards, the latter based on consensus diagnoses rendered by 2 TMD experts using criterion examination data, including temporomandibular joint imaging. Cut-offs for target validity were sensitivity ≥ 0.70 and specificity ≥ 0.95. Reliability of revised algorithms was assessed in 27 study participants. RESULTS Revised algorithm sensitivity and specificity exceeded the target levels for myofascial pain (0.82, 0.99, respectively) and myofascial pain with limited opening (0.93, 0.97). Combining diagnoses for any myofascial pain showed sensitivity of 0.91 and specificity of 1.00. For joint pain, target sensitivity and specificity were observed (0.92, 0.96) when arthralgia and osteoarthritis were combined as "any joint pain." Disc displacement without reduction with limited opening demonstrated target sensitivity and specificity (0.80, 0.97). For the other Group II disc displacements and Group III osteoarthritis and osteoarthrosis, sensitivity was below target (0.35 to 0.53), and specificity ranged from 0.80 to meeting target. Kappa for revised algorithm diagnostic reliability was ≥ 0.63. CONCLUSION Revised RDC/TMD Axis I TMD diagnostic algorithms are recommended for myofascial pain and joint pain as reliable and valid. However, revised clinical criteria alone, without recourse to imaging, are inadequate for valid diagnosis of two of the three disc displacements and osteoarthritis/osteoarthrosis. PMID:20213032
Diagnostic efficacy of in vitro methods vs. skin testing in patients with inhalant allergies
Corey, J.P.; Liudahl, J.J.; Young, S.A.; Rodman, S.M. )
The purpose of our study was to investigate the diagnostic efficacy of two selected methods of in vitro allergy testing. Specifically, the PRIST/modified RAST I125 isotope systems and the Quantizyme/modified EAST alkaline phosphatase method were compared. The time, expense, convenience, and diagnostic efficacy of the two procedures are discussed. Special attention is given to the practicality of each method for the practicing physician.
Benchmarking Gas Path Diagnostic Methods: A Public Approach
NASA Technical Reports Server (NTRS)
Simon, Donald L.; Bird, Jeff; Davison, Craig; Volponi, Al; Iverson, R. Eugene
Recent technology reviews have identified the need for objective assessments of engine health management (EHM) technology. The need is two-fold: technology developers require relevant data and problems to design and validate new algorithms and techniques while engine system integrators and operators need practical tools to direct development and then evaluate the effectiveness of proposed solutions. This paper presents a publicly available gas path diagnostic benchmark problem that has been developed by the Propulsion and Power Systems Panel of The Technical Cooperation Program (TTCP) to help address these needs. The problem is coded in MATLAB (The MathWorks, Inc.) and coupled with a non-linear turbofan engine simulation to produce "snap-shot" measurements, with relevant noise levels, as if collected from a fleet of engines over their lifetime of use. Each engine within the fleet will experience unique operating and deterioration profiles, and may encounter randomly occurring relevant gas path faults including sensor, actuator and component faults. The challenge to the EHM community is to develop gas path diagnostic algorithms to reliably perform fault detection and isolation. An example solution to the benchmark problem is provided along with associated evaluation metrics. A plan is presented to disseminate this benchmark problem to the engine health management technical community and invite technology solutions.
Development of wide area environment accelerator operation and diagnostics method
NASA Astrophysics Data System (ADS)
Uchiyama, Akito; Furukawa, Kazuro
Remote operation and diagnostic systems for particle accelerators have been developed for beam operation and maintenance in various situations. Even though fully remote experiments are not necessary, the remote diagnosis and maintenance of the accelerator is required. Considering remote-operation operator interfaces (OPIs), the use of standard protocols such as the hypertext transfer protocol (HTTP) is advantageous, because system-dependent protocols are unnecessary between the remote client and the on-site server. Here, we have developed a client system based on WebSocket, which is a new protocol provided by the Internet Engineering Task Force for Web-based systems, as a next-generation Web-based OPI using the Experimental Physics and Industrial Control System Channel Access protocol. As a result of this implementation, WebSocket-based client systems have become available for remote operation. Also, as regards practical application, the remote operation of an accelerator via a wide area network (WAN) faces a number of challenges, e.g., the accelerator has both experimental device and radiation generator characteristics. Any error in remote control system operation could result in an immediate breakdown. Therefore, we propose the implementation of an operator intervention system for remote accelerator diagnostics and support that can obviate any differences between the local control room and remote locations. Here, remote-operation Web-based OPIs, which resolve security issues, are developed.
Application of modern diagnostic methods to environmental improvement. Annual progress report, October 1994--September 1995
Shepard, W.S.
The Diagnostic Instrumentation and Analysis Laboratory (DIAL), an interdisciplinary research department in the College of Engineering at Mississippi State University (MSU), is under contract with the US Department of Energy (DOE) to develop and apply advanced diagnostic instrumentation and analysis techniques to aid in solving DOE`s nuclear waste problem. The program is a comprehensive effort which includes five focus areas: advanced diagnostic systems; development/application; torch operation and test facilities; process development; on-site field measurement and analysis; technology transfer/commercialization. As part of this program, diagnostic methods will be developed and evaluated for characterization, monitoring and process control. Also, the measured parameters, will be employed to improve, optimize and control the operation of the plasma torch and the overall plasma treatment process. Moreover, on-site field measurements at various DOE facilities are carried out to aid in the rapid demonstration and implementation of modern fieldable diagnostic methods. Such efforts also provide a basis for technology transfer.
Combat-Related Pythium aphanidermatum Invasive Wound Infection: Case Report and Discussion of Utility of Molecular Diagnostics
Murray, Clinton K.; Driscoll, Ian R.; Wickes, Brian L.; Wiederhold, Nathan; Sutton, Deanna A.; Sanders, Carmita; Mende, Katrin; Enniss, Brent; Feig, James; Ganesan, Anuradha; Rini, Elizabeth A.; Vento, Todd J.
We describe a 22-year-old soldier with 19% total body surface area burns, polytrauma, and sequence- and culture-confirmed Pythium aphanidermatum wound infection. Antemortem histopathology suggested disseminated Pythium infection, including brain involvement; however, postmortem PCR revealed Cunninghamella elegans, Lichtheimia corymbifera, and Saksenaea vasiformis coinfection. The utility of molecular diagnostics in invasive fungal infections is discussed. PMID:25832301
Combat-Related Pythium aphanidermatum Invasive Wound Infection: Case Report and Discussion of Utility of Molecular Diagnostics.
Farmer, Aaron R; Murray, Clinton K; Driscoll, Ian R; Wickes, Brian L; Wiederhold, Nathan; Sutton, Deanna A; Sanders, Carmita; Mende, Katrin; Enniss, Brent; Feig, James; Ganesan, Anuradha; Rini, Elizabeth A; Vento, Todd J
We describe a 22-year-old soldier with 19% total body surface area burns, polytrauma, and sequence- and culture-confirmed Pythium aphanidermatum wound infection. Antemortem histopathology suggested disseminated Pythium infection, including brain involvement; however, postmortem PCR revealed Cunninghamella elegans, Lichtheimia corymbifera, and Saksenaea vasiformis coinfection. The utility of molecular diagnostics in invasive fungal infections is discussed. PMID:25832301
Molecular Phylogenetic Analysis of Enterobius vermicularis and Development of an 18S Ribosomal DNA-Targeted Diagnostic PCR▿
Zelck, Ulrike E.; Bialek, Ralf; Weiß, Michael
We genetically characterized pinworms obtained from 37 children from different regions of Germany and established new species-specific molecular diagnostic tools. No ribosomal DNA diversity was found; the phylogenetic position of Enterobius vermicularis within the Oxyurida order and its close relationship to the Ascaridida and Spirurida orders was confirmed. PMID:21248085
The Rapid-Heat LAMPellet Method: A Potential Diagnostic Method for Human Urogenital Schistosomiasis
Carranza-Rodríguez, Cristina; Pérez-Arellano, José Luis; Vicente, Belén; López-Abán, Julio; Muro, Antonio
Background Urogenital schistosomiasis due to Schistosoma haematobium is a serious underestimated public health problem affecting 112 million people - particularly in sub-Saharan Africa. Microscopic examination of urine samples to detect parasite eggs still remains as definitive diagnosis. This work was focussed on developing a novel loop-mediated isothermal amplification (LAMP) assay for detection of S. haematobium DNA in human urine samples as a high-throughput, simple, accurate and affordable diagnostic tool to use in diagnosis of urogenital schistosomiasis. Methodology/Principal Findings A LAMP assay targeting a species specific sequence of S. haematobium ribosomal intergenic spacer was designed. The effectiveness of our LAMP was assessed in a number of patients´ urine samples with microscopy confirmed S. haematobium infection. For potentially large-scale application in field conditions, different DNA extraction methods, including a commercial kit, a modified NaOH extraction method and a rapid heating method were tested using small volumes of urine fractions (whole urine, supernatants and pellets). The heating of pellets from clinical samples was the most efficient method to obtain good-quality DNA detectable by LAMP. The detection limit of our LAMP was 1 fg/µL of S. haematobium DNA in urine samples. When testing all patients´ urine samples included in our study, diagnostic parameters for sensitivity and specificity were calculated for LAMP assay, 100% sensitivity (95% CI: 81.32%-100%) and 86.67% specificity (95% CI: 75.40%-94.05%), and also for microscopy detection of eggs in urine samples, 69.23% sensitivity (95% CI: 48.21% -85.63%) and 100% specificity (95% CI: 93.08%-100%). Conclusions/Significance We have developed and evaluated, for the first time, a LAMP assay for detection of S. haematobium DNA in heated pellets from patients´ urine samples using no complicated requirement procedure for DNA extraction. The procedure has been named the Rapid
Non-Intrusive Optical Diagnostic Methods for Flowfield Characterization
NASA Technical Reports Server (NTRS)
Tabibi, Bagher M.; Terrell, Charles A.; Spraggins, Darrell; Lee, Ja. H.; Weinstein, Leonard M.
Non-intrusive optical diagnostic techniques such as Electron Beam Fluorescence (EBF), Laser-Induced Fluorescence (LIF), and Focusing Schlieren (FS) have been setup for high-speed flow characterization and large flowfield visualization, respectively. Fluorescence emission from the First Negative band of N2(+) with the (0,0) vibration transition (at lambda =391.44 nm) was obtained using the EBF technique and a quenching rate of N2(+)* molecules by argon gas was reported. A very high sensitivity FS system was built and applied in the High-Speed Flow Generator (HFG) at NASA LaRC. A LIF system is available at the Advanced Propulsion Laboratory (APL) on campus and a plume exhaust velocity measurement, measuring the Doppler shift from lambda = 728.7 nm of argon gas, is under way.
Molecular signature of salivary gland tumors: potential use as diagnostic and prognostic marker.
Fonseca, Felipe Paiva; Sena Filho, Marcondes; Altemani, Albina; Speight, Paul M; Vargas, Pablo Agustin
Salivary gland tumors are a highly heterogeneous group of lesions with diverse microscopic appearances and variable clinical behavior. The use of clinical and histological parameters to predict patient prognosis and survival rates has been of limited utility, and the search for new biomarkers that could not only aid in a better understanding of their pathogenesis but also be reliable auxiliaries for prognostic determination and useful diagnostic tools has been performed in the last decades with very exciting results. Hence, gene rearrangements such as CRTC1-MAML2 in mucoepidermoid carcinomas have shown excellent specificity, and more than that, it has been strongly correlated with low-grade tumors and consequently with an increased survival rate and better prognosis of patients affected by neoplasms carrying this translocation. Moreover, MYB-NFIB and EWSR1-ATF1 gene fusions were shown to be specifically found in cases of adenoid cystic carcinomas and hyalinizing clear cell carcinomas, respectively, in the context of salivary gland tumors, becoming reliable diagnostic tools for these entities and potential therapeutic targets for future therapeutic protocols. Finally, the identification of ETV6-NTRK3 in cases previously diagnosed as uncommon acinic cell carcinomas, cystadenocarcinomas, and adenocarcinomas not otherwise specified led to the characterization of a completely new and now widely accepted entity, including, therefore, mammary analogue secretory carcinoma in the list of well-recognized salivary gland carcinomas. Thus, further molecular investigations of salivary gland tumors are warranted, and the recognition of other genetic abnormalities can lead to the acknowledgment of new entities and the acquirement of reliable biomarkers. PMID:25990369
Hydroxymethyl phosphine compounds for use as diagnostic and therapeutic pharmaceuticals and method of making same
Katti, Kattesh V.; Karra, Srinivasa Rao; Berning, Douglas E.; Smith, C. Jeffrey; Volkert, Wynn A.; Ketring, Alan R.
A compound and method of making a compound for use as a diagnostic or therapeutic pharmaceutical comprises at least one functionalized hydroxyalkyl phosphine donor group and one or more sulfur or nitrogen donor and a metal combined with the ligand.
Hydroxymethyl phosphine compounds for use as diagnostic and therapeutic pharmaceuticals and method of making same
Katti, Kattesh V.; Karra, Srinivasa Rao; Berning, Douglas E.; Smith, C. Jeffrey; Volkert, Wynn A.; Ketring, Alan R.
A compound and method of making a compound for use as a diagnostic or therapeutic pharmaceutical comprises at least one functionalized hydroxyalkyl phosphine donor group and one or more sulfur or nitrogen donor and a metal combined with the ligand.
Hb H disease with various β hemoglobinopathies: molecular, hematological and diagnostic aspects.
Fucharoen, Supan; Fucharoen, Goonnapa
We report the molecular, hematological and diagnostic features of three Thai patients with Hb H (β4) disease in combination with different β hemoglobinopathies. The first case was a boy with a hitherto undescribed condition of Hb H-Hb Tak (HBB:c.441_442insAC) syndrome. The second case was an adult male with the Hb H-Hb Hope (HBB:c.410G>A) disease. The last case was a male patient with a combination of Hb H-Hb Constant Spring (Hb CS) (HBA2:c.427T>C) and a high Hbs A(2)/F β(0)-thalassemia (β-thal) trait (the 3.4 kb deletion). In all cases, no Hb H was detected and Hb H inclusion was absent, accurate diagnosis could be obtained after DNA analysis. All of them had mild hypochromic microcytic anemia with hemoglobin (Hb) values ranging from 8.8-10.4 g/dL, PCV 0.31-0.36 L/L and MCV 57.6-66.9 fL and none had ever received a blood transfusion. The levels of Hb Tak (7.8%) and Hb Hope (21.6%), respectively, observed in the first two cases were relatively lower than values found in heterozygotes for these two variants. In the last case, the level of Hb A(2) (4.8%) was within the diagnostic range for a β-thal trait. The Hb F level was not elevated but a small amount of Hb Bart's was observed. Hematological findings and diagnostics using combined Hb and polymerase chain reaction (PCR) analyses are presented and compared with those of the other Thai patients with Hb H-β(0)-thal, Hb H-Hb CS-Hb J-Bangkok (HBB:c.170G>A) and Hb H-Hb CS-Hb Pyrgos (HBB:c.251G>A) previously observed in our series. PMID:22145566
CFTR mutations spectrum and the efficiency of molecular diagnostics in Polish cystic fibrosis patients.
Ziętkiewicz, Ewa; Rutkiewicz, Ewa; Pogorzelski, Andrzej; Klimek, Barbara; Voelkel, Katarzyna; Witt, Michał
Cystic fibrosis (CF) is caused by mutations in the cystic fibrosis transmembrane regulator gene (CFTR). In light of the strong allelic heterogeneity and regional specificity of the mutation spectrum, the strategy of molecular diagnostics and counseling in CF requires genetic tests to reflect the frequency profile characteristic for a given population. The goal of the study was to provide an updated comprehensive estimation of the distribution of CFTR mutations in Polish CF patients and to assess the effectiveness of INNOLiPA_CFTR tests in Polish population. The analyzed cohort consisted of 738 patients with the clinically confirmed CF diagnosis, prescreened for molecular defects using INNOLiPA_CFTR panels from Innogenetics. A combined efficiency of INNOLiPA CFTR_19 and CFTR_17_TnUpdate tests was 75.5%; both mutations were detected in 68.2%, and one mutation in 14.8% of the affected individuals. The group composed of all the patients with only one or with no mutation detected (109 and 126 individuals, respectively) was analyzed further using a mutation screening approach, i.e. SSCP/HD (single strand conformational polymorphism/heteroduplex) analysis of PCR products followed by sequencing of the coding sequence. As a result, 53 more mutations were found in 97 patients. The overall efficiency of the CF allele detection was 82.5% (7.0% increase compared to INNOLiPA tests alone). The distribution of the most frequent mutations in Poland was assessed. Most of the mutations repetitively found in Polish patients had been previously described in other European populations. The most frequent mutated allele, F508del, represented 54.5% of Polish CF chromosomes. Another eight mutations had frequencies over 1%, 24 had frequencies between 1 and 0.1%; c.2052-2053insA and c.3468+2_3468+3insT were the most frequent non-INNOLiPA mutations. Mutation distribution described herein is also relevant to the Polish diaspora. Our study also demonstrates that the reported efficiency of
Biomarkers and Molecular Analysis to Improve Bloodstream Infection Diagnostics in an Emergency Care Unit
Loonen, Anne J. M.; de Jager, Cornelis P. C.; Tosserams, Janna; Kusters, Ron; Hilbink, Mirrian; Wever, Peter C.; van den Brule, Adriaan J. C.
subsequent pathogen identification using molecular diagnostics. PMID:24475269
Gene dosage methods as diagnostic tools for the identification of chromosome abnormalities.
Gouas, L; Goumy, C; Véronèse, L; Tchirkov, A; Vago, P
Cytogenetics is the part of genetics that deals with chromosomes, particularly with numerical and structural chromosome abnormalities, and their implications in congenital or acquired genetic disorders. Standard karyotyping, successfully used for the last 50 years in investigating the chromosome etiology in patients with infertility, fetal abnormalities and congenital disorders, is constrained by the limits of microscopic resolution and is not suited for the detection of subtle chromosome abnormalities. The ability to detect submicroscopic chromosomal rearrangements that lead to copy-number changes has escalated progressively in recent years with the advent of molecular cytogenetic techniques. Here, we review various gene dosage methods such as FISH, PCR-based approaches (MLPA, QF-PCR, QMPSF and real time PCR), CGH and array-CGH, that can be used for the identification and delineation of copy-number changes for diagnostic purposes. Besides comparing their relative strength and weakness, we will discuss the impact that these detection methods have on our understanding of copy number variations in the human genome and their implications in genetic counseling. PMID:18513889
Field and laboratory comparative evaluation of rapid malaria diagnostic tests versus traditional and molecular techniques in India
Background Malaria presents a diagnostic challenge in most tropical countries. Microscopy remains the gold standard for diagnosing malaria infections in clinical practice and research. However, microscopy is labour intensive, requires significant skills and time, which causes therapeutic delays. The objective of obtaining result quickly from the examination of blood samples from patients with suspected malaria is now made possible with the introduction of rapid malaria diagnostic tests (RDTs). Several RDTs are available, which are fast, reliable and simple to use and can detect Plasmodium falciparum and non-falciparum infections or both. A study was conducted in tribal areas of central India to measure the overall performance of several RDTs for diagnosis of P. falciparum and non-falciparum infections in comparison with traditional and molecular techniques. Such data will be used to guide procurement decisions of policy makers and programme managers. Methods Five commercially available RDTs were tested simultaneously in field in parallel with peripheral blood smears in outbreak-affected areas. The evaluation is designed to provide comparative data on the performance of each RDT. In addition, molecular method i.e. polymerase chain reaction (PCR) was also carried out to compare all three methods. Results A total of 372 patients with a clinical suspicion of malaria from Bajag Primary Health Centre (PHC) of district Dindori and Satanwada PHC of district Shivpuri attending the field clinics of Regional Medical Research Centre were included in the study. The analysis revealed that the First Response Malaria Antigen pLDH/HRP2 combo test was 94.7% sensitive (95% CI 89.5-97.7) and 69.9% specific (95% CI 63.6-75.6) for P. falciparum. However, for non-falciparum infections (Plasmodium vivax) the test was 84.2% sensitive (95% CI 72.1-92.5) and 96.5% specific (95% CI 93.8-98.2). The Parascreen represented a good alternative. All other RDTs were relatively less sensitive for
The SUCCESS model for laboratory performance and execution of rapid molecular diagnostics in patients with sepsis.
Dekmezian, Mhair; Beal, Stacy G; Damashek, Mary Jane; Benavides, Raul; Dhiman, Neelam
Successful performance and execution of rapid diagnostics in a clinical laboratory hinges heavily on careful validation, accurate and timely communication of results, and real-time quality monitoring. Laboratories must develop strategies to integrate diagnostics with stewardship and evidence-based clinical practice guidelines. We present a collaborative SUCCESS model for execution and monitoring of rapid sepsis diagnostics to facilitate timely treatment. Six months after execution of the Verigene Gram-Positive Blood Culture (BC-GP) and the AdvanDx PNA-FISH assays, data were collected on 579 and 28 episodes of bacteremia and fungemia, respectively. Clinical testing was executed using a SUCCESS model comprising the following components: stewardship, utilization of resources, core strategies, concierge services, education, support, and surveillance. Stewardship needs were identified by evaluating the specialty services benefiting from new testing. Utilization of resources was optimized by reviewing current treatment strategies and antibiogram and formulary options. Core strategies consisted of input from infectious disease leadership, pharmacy, and laboratory staff. Concierge services included automated Micro-eUpdate and physician-friendly actionable reports. Education modules were user-specific, and support was provided through a dedicated 24/7 microbiology hotline. Surveillance was performed by daily audit by the director. Using the SUCCESS model, the turnaround time for the detailed report with actionable guidelines to the physician was ∼3 hours from the time of culture positivity. The overall correlation between rapid methods and culture was 94% (546/579). Discrepant results were predominantly contaminants such as a coagulase-negative staphylococci or viridans streptococci in mixed cultures. SUCCESS is a cost-effective and easily adaptable model for clinical laboratories with limited stewardship resources. PMID:25829640
Frequency-Domain Methods for Characterization of Pulsed Power Diagnostics
White, A D; Anderson, R A; Ferriera, T J; Goerz, D A
This paper discusses methods of frequency-domain characterization of pulsed power sensors using vector network analyzer and spectrum analyzer techniques that offer significant simplification over time-domain methods, while mitigating or minimizing the effect of the difficulties present in time domain characterization. These methods are applicable to characterization of a wide variety of sensors.
Reevaluation of an Acanthamoeba Molecular Diagnostic Algorithm following an Atypical Case of Amoebic Keratitis
Lau, Rachel; Cunanan, Marlou; Jackson, Jonathan; Ali, Ibne Karim M.; Chong-Kit, Ann; Gasgas, Jason; Tian, Jinfang; Ralevski, Filip
Amoebic keratitis (AK) is a potentially blinding infection, the prompt diagnosis of which is essential for limiting ocular morbidity. We undertook a quality improvement initiative with respect to the molecular detection of acanthamoebae in our laboratory because of an unusual case of discordance. Nine ATCC strains of Acanthamoeba and 40 delinked, biobanked, surplus corneal scraping specimens were analyzed for the presence of acanthamoebae with four separate real-time PCR assays. The assay used by the Free-Living and Intestinal Amebas Laboratory of the CDC was considered the reference standard, and the performance characteristics of each individual assay and pairs of assays were calculated. Outcome measures were sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV). Of 49 included specimens, 14 (28.6%) were positive by the gold standard assay, and 35 (71.4%) were negative. The sensitivities of the individual assays ranged from 64.3% to 92.9%, compared to the gold standard, while the specificities ranged from 88.6% to 91.4%. The PPVs and NPVs ranged from 69.2% to 78.6% and from 86.1% to 96.9%, respectively. Combinations of assay pairs led to improved performance, with sensitivities ranging from 92.9% to 100% and specificities ranging from 97.1% to 100%. ATCC and clinical strains of Acanthamoeba that failed to be detected by certain individual assays included Acanthamoeba castellanii, Acanthamoeba culbertsoni, and Acanthamoeba lenticulata. For three clinical specimens, false negativity of the gold standard assay could not be excluded. Molecular diagnostic approaches, especially combinations of highly sensitive and specific assays, offer a reasonably performing, operator-independent, rapid strategy for the detection of acanthamoebae in clinical specimens and are likely to be more practical than either culture or direct microscopic detection. PMID:26202123
Propulsion Diagnostic Method Evaluation Strategy (ProDiMES) User's Guide
NASA Technical Reports Server (NTRS)
Simon, Donald L.
This report is a User's Guide for the Propulsion Diagnostic Method Evaluation Strategy (ProDiMES). ProDiMES is a standard benchmarking problem and a set of evaluation metrics to enable the comparison of candidate aircraft engine gas path diagnostic methods. This Matlab (The Mathworks, Inc.) based software tool enables users to independently develop and evaluate diagnostic methods. Additionally, a set of blind test case data is also distributed as part of the software. This will enable the side-by-side comparison of diagnostic approaches developed by multiple users. The Users Guide describes the various components of ProDiMES, and provides instructions for the installation and operation of the tool.
Applications of Replicating-Competent Reporter-Expressing Viruses in Diagnostic and Molecular Virology
Li, Yongfeng; Li, Lian-Feng; Yu, Shaoxiong; Wang, Xiao; Zhang, Lingkai; Yu, Jiahui; Xie, Libao; Li, Weike; Ali, Razim; Qiu, Hua-Ji
Commonly used tests based on wild-type viruses, such as immunostaining, cannot meet the demands for rapid detection of viral replication, high-throughput screening for antivirals, as well as for tracking viral proteins or virus transport in real time. Notably, the development of replicating-competent reporter-expressing viruses (RCREVs) has provided an excellent option to detect directly viral replication without the use of secondary labeling, which represents a significant advance in virology. This article reviews the applications of RCREVs in diagnostic and molecular virology, including rapid neutralization tests, high-throughput screening systems, identification of viral receptors and virus-host interactions, dynamics of viral infections in vitro and in vivo, vaccination approaches and others. However, there remain various challenges associated with RCREVs, including pathogenicity alterations due to the insertion of a reporter gene, instability or loss of the reporter gene expression, or attenuation of reporter signals in vivo. Despite all these limitations, RCREVs have become powerful tools for both basic and applied virology with the development of new technologies for generating RCREVs, the inventions of novel reporters and the better understanding of regulation of viral replication. PMID:27164126
Applications of Replicating-Competent Reporter-Expressing Viruses in Diagnostic and Molecular Virology.
Li, Yongfeng; Li, Lian-Feng; Yu, Shaoxiong; Wang, Xiao; Zhang, Lingkai; Yu, Jiahui; Xie, Libao; Li, Weike; Ali, Razim; Qiu, Hua-Ji
Commonly used tests based on wild-type viruses, such as immunostaining, cannot meet the demands for rapid detection of viral replication, high-throughput screening for antivirals, as well as for tracking viral proteins or virus transport in real time. Notably, the development of replicating-competent reporter-expressing viruses (RCREVs) has provided an excellent option to detect directly viral replication without the use of secondary labeling, which represents a significant advance in virology. This article reviews the applications of RCREVs in diagnostic and molecular virology, including rapid neutralization tests, high-throughput screening systems, identification of viral receptors and virus-host interactions, dynamics of viral infections in vitro and in vivo, vaccination approaches and others. However, there remain various challenges associated with RCREVs, including pathogenicity alterations due to the insertion of a reporter gene, instability or loss of the reporter gene expression, or attenuation of reporter signals in vivo. Despite all these limitations, RCREVs have become powerful tools for both basic and applied virology with the development of new technologies for generating RCREVs, the inventions of novel reporters and the better understanding of regulation of viral replication. PMID:27164126
Molecular investigations to improve diagnostic accuracy in patients with ARC syndrome.
Cullinane, Andrew R; Straatman-Iwanowska, Anna; Seo, Jeong K; Ko, Jae S; Song, Kyung S; Gizewska, Maria; Gruszfeld, Dariusz; Gliwicz, Dorota; Tuysuz, Beyhan; Erdemir, Gulin; Sougrat, Rachid; Wakabayashi, Yoshiyuki; Hinds, Rupert; Barnicoat, Angela; Mandel, Hanna; Chitayat, David; Fischler, Björn; Garcia-Cazorla, Angels; Knisely, A S; Kelly, Deirdre A; Maher, Eamonn R; Gissen, Paul
Arthrogryposis, Renal dysfunction and Cholestasis (ARC) syndrome is a multi-system autosomal recessive disorder caused by germline mutations in VPS33B. The detection of germline VPS33B mutations removes the need for diagnostic organ biopsies (these carry a>50% risk of life-threatening haemorrhage due to platelet dysfunction); however, VPS33B mutations are not detectable in approximately 25% of patients. In order further to define the molecular basis of ARC we performed mutation analysis and mRNA and protein studies in patients with a clinical diagnosis of ARC. Here we report novel mutations in VPS33B in patients from Eastern Europe and South East Asia. One of the mutations was present in 7 unrelated Korean patients. Reduced expression of VPS33B and cellular phenotype was detected in fibroblasts from patients clinically diagnosed with ARC with and without known VPS33B mutations. One mutation-negative patient was found to have normal mRNA and protein levels. This patient's clinical condition improved and he is alive at the age of 2.5 years. Thus we show that all patients with a classical clinical course of ARC had decreased expression of VPS33B whereas normal VPS33B expression was associated with good prognosis despite initial diagnosis of ARC. PMID:18853461
Molecular Investigations to Improve Diagnostic Accuracy in Patients With ARC Syndrome
Cullinane, Andrew R.; Straatman-Iwanowska, Anna; Seo, Jeong K.; Ko, Jae S.; Song, Kyung S.; Gizewska, Maria; Gruszfeld, Dariusz; Gliwicz, Dorota; Tuysuz, Beyhan; Erdemir, Gulin; Sougrat, Rachid; Wakabayashi, Yoshiyuki; Hinds, Rupert; Barnicoat, Angela; Mandel, Hanna; Chitayat, David; Fischler, Björn; Garcia-Cazorla, Angels; Knisely, A. S.; Kelly, Deirdre A.; Maher, Eamonn R.; Gissen, Paul
Arthrogryposis, Renal dysfunction and Cholestasis (ARC) syndrome is a multi-system autosomal recessive disorder caused by germline mutations in VPS33B. The detection of germline VPS33B mutations removes the need for diagnostic organ biopsies (these carry a >50% risk of life-threatening haemorrhage due to platelet dysfunction); however, VPS33B mutations are not detectable in ∼25% of patients. In order further to define the molecular basis of ARC we performed mutation analysis and mRNA and protein studies in patients with a clinical diagnosis of ARC. Here we report novel mutations in VPS33B in patients from Eastern Europe and South East Asia. One of the mutations was present in 7 unrelated Korean patients. Reduced expression of VPS33B and cellular phenotype was detected in fibroblasts from patients clinically diagnosed with ARC with and without known VPS33B mutations. One mutation-negative patient was found to have normal mRNA and protein levels. This patient's clinical condition improved and he is alive at the age of 2.5 years. Thus we show that all patients with a classical clinical course of ARC had decreased expression of VPS33B whereas normal VPS33B expression was associated with good prognosis despite initial diagnosis of ARC. PMID:18853461
Costs of genetic testing: Supporting Brazilian Public Policies for the incorporating of molecular diagnostic technologies
Schlatter, Rosane Paixão; Matte, Ursula; Polanczyk, Carisi Anne; Koehler-Santos, Patrícia; Ashton-Prolla, Patricia
This study identifies and describes the operating costs associated with the molecular diagnosis of diseases, such as hereditary cancer. To approximate the costs associated with these tests, data informed by Standard Operating Procedures for various techniques was collected from hospital software and a survey of market prices. Costs were established for four scenarios of capacity utilization to represent the possibility of suboptimal use in research laboratories. Cost description was based on a single site. The results show that only one technique was not impacted by rising costs due to underutilized capacity. Several common techniques were considerably more expensive at 30% capacity, including polymerase chain reaction (180%), microsatellite instability analysis (181%), gene rearrangement analysis by multiplex ligation probe amplification (412%), non-labeled sequencing (173%), and quantitation of nucleic acids (169%). These findings should be relevant for the definition of public policies and suggest that investment of public funds in the establishment of centralized diagnostic research centers would reduce costs to the Public Health System. PMID:26500437
Update in salivary gland cytopathology: Recent molecular advances and diagnostic applications.
Pusztaszeri, Marc P; Faquin, William C
Salivary gland tumors (SGT) are notorious for their extraordinary diversity and for the morphological overlap that exists between many of these entities. Fine-needle aspiration biopsy (FNAB) has a well-established role in the evaluation of patients with a salivary gland lesion, helping to guide clinical management. However, salivary gland FNAB has several limitations and does not allow for a specific diagnosis in some cases. For these reasons, salivary gland FNAB is considered one of the most challenging areas in cytopathology. Over the last decade, new salivary gland entities have been recognized, enlarging SGT diversity and complexity even more. In addition, a subset of SGT, including common entities such as pleomorphic adenoma and uncommon new entities such as mammary analog secretory carcinoma, have been characterized cytogenetically by the presence of specific translocations. The molecular consequences of these translocations and their potential prognostic and therapeutic values are not yet well characterized. However, these translocations and their resulting fusion oncogenes and oncoproteins can be used as diagnostic clues in salivary gland FNAB material in order to overcome the limitations of cytomorphological evaluation alone. In this review, we focus on SGTs currently known to harbor translocations and fusion genes, including uncommon and recently recognized entities, and discuss their potential application to salivary gland FNAB. PMID:25613003
Oral biofilms: molecular analysis, challenges, and future prospects in dental diagnostics
Do, Thuy; Devine, Deirdre; Marsh, Philip D
Oral biofilms are functionally and structurally organized polymicrobial communities that are embedded in an extracellular matrix of exopolymers on mucosal and dental surfaces. These biofilms are found naturally in health, and provide benefits to the host. However, this relationship can break down, and disease can occur; disease is associated with a shift in the balance of the species within these biofilms. Simple diagnostic tests have been developed that involve the culture of selected bacteria, eg, those implicated in dental caries, facilitating an assessment of risk of further disease in individual patients. However, oral diseases have a complex etiology, and because only around 50% of oral biofilm can be grown at present, culture-independent molecular-based approaches are being developed that give a more comprehensive assessment of the presence of a range of putative pathogens in samples. The diversity of these biofilms creates challenges in the interpretation of findings, and future work is investigating the ability of novel techniques to detect biological activity and function in oral biofilms, rather than simply providing a catalogue of microbial names. PMID:23674928
Tokamak physics studies using x-ray diagnostic methods
Hill, K.W.; Bitter, M.; von Goeler, S.; Beiersdorfer, P.; Fredrickson, E.; Hsuan, H.; McGuire, K.; Sauthoff, N.R.; Sesnic, S.; Stevens, J.E.
X-ray diagnostic measurements have been used in a number of experiments to improve our understanding of important tokamak physics issues. The impurity content in TFTR plasmas, its sources and control have been clarified through soft x-ray pulse-height analysis (PHA) measurements. The dependence of intrinsic impurity concentrations and Z/sub eff/ on electron density, plasma current, limiter material and conditioning, and neutral-beam power have shown that the limiter is an important source of metal impurities. Neoclassical-like impurity peaking following hydrogen pellet injection into Alcator C and a strong effect of impurities on sawtooth behavior were demonstrated by x-ray imaging (XIS) measurements. Rapid inward motion of impurities and continuation of m = 1 activity following an internal disruption were demonstrated with XIS measurements on PLT using injected aluminum to enhance the signals. Ion temperatures up to 12 keV and a toroidal plasma rotation velocity up to 6 x 10/sup 5/ m/s have been measured by an x-ray crystal spectrometer (XCS) with up to 13 MW of 85-keV neutral-beam injection in TFTR. Precise wavelengths and relative intensities of x-ray lines in several helium-like ions and neon-like ions of silver have been measured in TFTR and PLT by the XCS. The data help to identify the important excitation processes predicted in atomic physics. Wavelengths of n = 3 to 2 silver lines of interest for x-ray lasers were measured, and precise instrument calibration techniques were developed. Electron thermal conductivity and sawtooth dynamics have been studied through XIS measurements on TFTR of heat-pulse propagation and compound sawteeth. A non-Maxwellian electron distribution function has been measured, and evidence of the Parail-Pogutse instability identified by hard x-ray PHA measurements on PLT during lower-hybrid current-drive experiments.
A Simplified Diagnostic Method for Elastomer Bond Durability
NASA Technical Reports Server (NTRS)
White, Paul
A simplified method has been developed for determining bond durability under exposure to water or high humidity conditions. It uses a small number of test specimens with relatively short times of water exposure at elevated temperature. The method is also gravimetric; the only equipment being required is an oven, specimen jars, and a conventional laboratory balance.
Impact of Molecular Diagnostics for Tuberculosis on Patient-Important Outcomes: A Systematic Review of Study Methodologies
Schumacher, Samuel G.; Sohn, Hojoon; Qin, Zhi Zhen; Gore, Genevieve; Davis, J. Lucian; Denkinger, Claudia M.; Pai, Madhukar
Background Several reviews on the accuracy of Tuberculosis (TB) Nucleic Acid Amplification Tests (NAATs) have been performed but the evidence on their impact on patient-important outcomes has not been systematically reviewed. Given the recent increase in research evaluating such outcomes and the growing list of TB NAATs that will reach the market over the coming years, there is a need to bring together the existing evidence on impact, rather than accuracy. We aimed to assess the approaches that have been employed to measure the impact of TB NAATs on patient-important outcomes in adults with possible pulmonary TB and/or drug-resistant TB. Methods We first develop a conceptual framework to clarify through which mechanisms the improved technical performance of a novel TB test may lead to improved patient outcomes and outline which designs may be used to measure them. We then systematically review the literature on studies attempting to assess the impact of molecular TB diagnostics on such outcomes and provide a narrative synthesis of designs used, outcomes assessed and risk of bias across different study designs. Results We found 25 eligible studies that assessed a wide range of outcomes and utilized a variety of experimental and observational study designs. Many potentially strong design options have never been used. We found that much of the available evidence on patient-important outcomes comes from a small number of settings with particular epidemiological and operational context and that confounding, time trends and incomplete outcome data receive insufficient attention. Conclusions A broader range of designs should be considered when designing studies to assess the impact of TB diagnostics on patient outcomes and more attention needs to be paid to the analysis as concerns about confounding and selection bias become relevant in addition to those on measurement that are of greatest concern in accuracy studies. PMID:26954678
Magnetic capture from blood rescues molecular motor function in diagnostic nanodevices
Background Introduction of effective point-of-care devices for use in medical diagnostics is part of strategies to combat accelerating health-care costs. Molecular motor driven nanodevices have unique potentials in this regard due to unprecedented level of miniaturization and independence of external pumps. However motor function has been found to be inhibited by body fluids. Results We report here that a unique procedure, combining separation steps that rely on antibody-antigen interactions, magnetic forces applied to magnetic nanoparticles (MPs) and the specificity of the actomyosin bond, can circumvent the deleterious effects of body fluids (e.g. blood serum). The procedure encompasses the following steps: (i) capture of analyte molecules from serum by MP-antibody conjugates, (ii) pelleting of MP-antibody-analyte complexes, using a magnetic field, followed by exchange of serum for optimized biological buffer, (iii) mixing of MP-antibody-analyte complexes with actin filaments conjugated with same polyclonal antibodies as the magnetic nanoparticles. This causes complex formation: MP-antibody-analyte-antibody-actin, and magnetic separation is used to enrich the complexes. Finally (iv) the complexes are introduced into a nanodevice for specific binding via actin filaments to surface adsorbed molecular motors (heavy meromyosin). The number of actin filaments bound to the motors in the latter step was significantly increased above the control value if protein analyte (50–60 nM) was present in serum (in step i) suggesting appreciable formation and enrichment of the MP-antibody-analyte-antibody-actin complexes. Furthermore, addition of ATP demonstrated maintained heavy meromyosin driven propulsion of actin filaments showing that the serum induced inhibition was alleviated. Detailed analysis of the procedure i-iv, using fluorescence microscopy and spectroscopy identified main targets for future optimization. Conclusion The results demonstrate a promising approach for
Restrictive Stochastic Item Selection Methods in Cognitive Diagnostic Computerized Adaptive Testing
ERIC Educational Resources Information Center
Wang, Chun; Chang, Hua-Hua; Huebner, Alan
This paper proposes two new item selection methods for cognitive diagnostic computerized adaptive testing: the restrictive progressive method and the restrictive threshold method. They are built upon the posterior weighted Kullback-Leibler (KL) information index but include additional stochastic components either in the item selection index or in…
Molecular diagnostics via mass spectrometry of PCR-amplified DNA products
Buchanan, M.; Doktycz, M.; Hurst, G.
Identifying the presence of a specific DNA fragment is becoming increasingly critical to many applications in medical, clinical, forensic and other research laboratories. At present, regions of interest in DNA are amplified using the Polymerase Chain Reaction (PCR) or other reactions to produce fragments containing a specific number of nucleotide units that are diagnostic for the targeted genetic disease, person, or pathogen. These fragments are then typically analyzed by slab gel electrophoresis. Mass spectrometry has the potential of characterizing the DNA fragments faster and more confidently than chromatography-based methods. The authors have evaluated matrix assisted laser desorption (MALDI) time-of-flight (TOF) and electrospray (ES) quadrupole ion trap (QIT) mass spectrometry for the rapid analysis of PCR fragments.
AB029. Clinical utility of a molecular diagnostic in evaluation of interstitial lung disease
Wu, Xiaoping; Rosenbluth, Michael; Choi, Yoonha; Danese, Sherry; Bianchi, Pauline; Flaherty, Kevin; Martinez, Fernando
Background The accurate diagnosis of idiopathic pulmonary fibrosis (IPF) continues to be challenging due to its overlapping features with other interstitial lung diseases (ILDs). With the approval of pirfenidone and nintedanib for treatment of IPF, there is greater urgency to identify patients with IPF without requiring surgical lung biopsy (SLB). In this study, we evaluated the clinical utility of a genomic classifier under development to identify usual interstitial pneumonia (UIP), the pathology pattern associated with IPF, using bronchoscopically collected samples. Methods A national survey was conducted from March 17–20, 2015 among 76 pulmonologists from ILD centers and non-specialty clinics. The survey described a genomic test with high precision for detecting UIP pattern using bronchoscopically collected samples. Physicians were asked about diagnostic/treatment next steps on four ILD patient cases representing confident UIP, possible UIP, possible UIP vs. hypersensitivity pneumonitis (HP), and connective tissue disease related ILD and how the genomic test would alter management. Results Physicians' likelihood of using the genomic test varied with the method of sampling. Ninety-one percent of physicians reported they would likely use the test if it required bronchoalveolar lavage (BAL), compared with 85% for transbronchial biopsy (TBB), and 63% for SLB. Across four clinical scenarios, a positive genomic test result significantly increased treatment with pirfenidone/nintedanib and reduced biopsies. The largest impact occurred in the possible UIP cases with an increase in treatment from 11% to 46% (P<0.001) and a decrease in biopsies from 59% to 26% (P<0.001). A positive test in the setting of confident UIP raised treatment recommendation from 47% to 70% (P<0.001) and decreased biopsies from 42% to 18% (P<0.001), suggesting its utility even in cases with high pre-test probability for UIP. A negative test result was less impactful on management than a
A commentary on the role of molecular technology and automation in clinical diagnostics
O'Connor, Ciara; Fitzgibbon, Marie; Powell, James; Barron, Denis; O'Mahony, Jim; Power, Lorraine; O'Connell, Nuala H; Dunne, Colum
Historically, the identification of bacterial or yeast isolates has been based on phenotypic characteristics such as growth on defined media, colony morphology, Gram stain, and various biochemical reactions, with significant delay in diagnosis. Clinical microbiology as a medical specialty has embraced advances in molecular technology for rapid species identification with broad-range 16S rDNA polymerase chain reaction (PCR) and matrix-assisted laser desorption and/or ionization time of flight (MALDI-TOF) mass spectrometry demonstrated as accurate, rapid, and cost-effective methods for the identification of most, but not all, bacteria and yeasts. Protracted conventional incubation times previously necessary to identify certain species have been mitigated, affording patients quicker diagnosis with associated reduction in exposure to empiric broad-spectrum antimicrobial therapy and shortened hospital stay. This short commentary details such molecular advances and their implications in the clinical microbiology setting. PMID:24658184
Molecular Photoionization Calculations Using the Complex Basis Function Method.
NASA Astrophysics Data System (ADS)
Yu, Chin-Hui
The complex basis function method (CBF) using both real and complex basis functions has been applied to the calculation of photoionization cross sections. The CBF method requires less computational resources than rigorous full-scattering methods and is effective for the evaluation of shape-resonance features. Neither the number of electrons in the system nor the molecular geometry is restricted. Moreover, the cross section obtained by the CBF method satisfies a variational principle and provides a practical diagnostic tool for the calculation of cross sections. The photoionization cross sections of H _sp{2}{+}, H _2, N_2, CO _2, and SF_6 have been computed using the CBF method. The computed partial cross sections for linear molecules agreed fairly well with other theoretical and experimental values. Particularly encouraging is the nearly perfect agreement of the CBF results with the results by rigorous full-scattering methods in the regions of sharp resonance features such as the K-shell ionization of N_2 and the 4sigma_{rm g} --> ksigma_ {rm u} transition of CO _2. The effect of averaging over all vibrational modes on the ionization cross sections for the 4 sigma_{rm g} orbital in CO_2 has also been studied for the first time. The resonance peak in the totally vibrationally averaged cross sections was reduced by 20%, but still represents a feature which has not yet been detected experimentally. The photoionization of SF_6 valence shells, 1t_{1rm g} , 5t_{1rm u}, 1t_{2rm u}, 3e _{rm g}, 1t_ {2rm g}, 4t_{1 rm u}, and 5a_{1 rm g}, has also been studied for the continuum symmetries a_{1rm g }, t_{1rm u} , e_{rm g}, and t_{2rm g}. The CBF results of SF_6 are numerically stable and essentially approach the static-exchange limit. These static-exchange partial cross sections, however, do not compare well with the experimental measurements. The discrepancy may be attributed to the physical approximations made in the theoretical model and to the quality of the ground -state
Methods for molecular surveillance of influenza
Wang, Ruixue; Taubenberger, Jeffery K
Molecular-based techniques for detecting influenza viruses have become an integral component of human and animal surveillance programs in the last two decades. The recent pandemic of the swine-origin influenza A virus (H1N1) and the continuing circulation of highly pathogenic avian influenza A virus (H5N1) further stress the need for rapid and accurate identification and subtyping of influenza viruses for surveillance, outbreak management, diagnosis and treatment. There has been remarkable progress on the detection and molecular characterization of influenza virus infections in clinical, mammalian, domestic poultry and wild bird samples in recent years. The application of these techniques, including reverse transcriptase-PCR, real-time PCR, microarrays and other nucleic acid sequencing-based amplifications, have greatly enhanced the capability for surveillance and characterization of influenza viruses. PMID:20455681
Nanoscale molecularly imprinted polymers and method thereof
Hart, Bradley R.; Talley, Chad E.
Nanoscale molecularly imprinted polymers (MIP) having polymer features wherein the size, shape and position are predetermined can be fabricated using an xy piezo stage mounted on an inverted microscope and a laser. Using an AMF controller, a solution containing polymer precursors and a photo initiator are positioned on the xy piezo and hit with a laser beam. The thickness of the polymeric features can be varied from a few nanometers to over a micron.
Optical methods for diagnostic of cell-tissue grafts
NASA Astrophysics Data System (ADS)
Timchenko, P. E.; Timchenko, E. V.; Volova, L. T.; Boltovskaya, V. V.; Zherdeva, L. A.; Belousov, N. V.; Pershutkina, S. V.
In this work the results of cell-tissue grafts research with a complex of optical methods - confocal fluorescent microscopy and Raman spectroscopy are presented. It was established that coefficient M scatter is related to irregularity of demineralization process. It was microscopically shown that the quantity of integrated cells into these types of transplants amounts to 20% of its surface.
Calreticulin Mutations in Myeloproliferative Neoplasms: Comparison of Three Diagnostic Methods
Park, Ji-Hye; Sevin, Margaux; Ramla, Selim; Truffot, Aurélie; Verrier, Tiffany; Bouchot, Dominique; Courtois, Martine; Bas, Mathilde; Benali, Sonia; Bailly, François; Favre, Bernardine; Guy, Julien; Martin, Laurent; Maynadié, Marc; Carillo, Serge; Girodon, François
Calreticulin (CALR) mutations have recently been reported in 70–84% of JAK2V617F-negative myeloproliferative neoplasms (MPN), and this detection has become necessary to improve the diagnosis of MPN. In a large single-centre cohort of 298 patients suffering from Essential Thrombocythemia (ET), the JAK2V617F, CALR and MPL mutations were noted in 179 (60%), 56 (18.5%) and 13 (4.5%) respectively. For the detection of the CALR mutations, three methods were compared in parallel: high-resolution melting-curve analysis (HRM), product-sizing analysis and Sanger sequencing. The sensitivity for the HRM, product-sizing analysis and Sanger sequencing was 96.4%, 98.2% and 89.3% respectively, whereas the specificity was 96.3%, 100% and 100%. In our cohort, the product-sizing analysis was the most sensitive method and was the easiest to interpret, while the HRM was sometimes difficult to interpret. In contrast, when large series of samples were tested, HRM provided results more quickly than did the other methods, which required more time. Finally, the sequencing method, which is the reference method, had the lowest sensitivity but can be used to describe the type of mutation precisely. Altogether, our results suggest that in routine laboratory practice, product-sizing analysis is globally similar to HRM for the detection of CALR mutations, and that both may be used as first-line screening tests. If the results are positive, Sanger sequencing can be used to confirm the mutation and to determine its type. Product-sizing analysis provides sensitive and specific results, moreover, with the quantitative measurement of CALR, which might be useful to monitor specific treatments. PMID:26501981
A novel and rapid diagnostic method for discriminating between feces of sika deer and Japanese serow by loop-mediated isothermal amplification.
Aikawa, T; Horino, S; Ichihara, Y
Severe damages to natural vegetation, agriculture, and forestry caused by overpopulation of sika deer (Cervus nippon) have markedly increased in Japan in recent years. To devise a population management plan of sika deer, information on the distribution and population size of the animal in each region is indispensable. An easy and effective method to obtain this information is to count the fecal pellets in the field. However, the habitat of sika deer in Japan overlaps that of Japanese serow (Capricornis crispus). Additionally, it is difficult to discriminate between the feces of both animals. Here, we present a rapid and precise diagnostic method for discriminating between the feces of sika deer and Japanese serow using loop-mediated isothermal amplification (LAMP) targeting cytochrome b gene in the mitochondrial DNA. Our results showed that the LAMP can discriminate between the feces of sika deer and Japanese serow, and the method is simpler and more sensitive than the conventional molecular diagnostic method. Since LAMP method does not require special skills for molecular biology techniques, even the field researchers who have never done a molecular experiment can easily carry out the protocol. In addition, the entire protocol, from DNA extraction from fecal pellet to identification of species, takes only about 75 min and does not require expensive equipment. Hence, this diagnostic method is simple, fast, and accessible to anyone. As such, the method can be a useful tool to estimate distribution and population size of sika deer. PMID:26084704
Fraley's syndrome: case report and update on current diagnostic methods
Zuckier, L.S.; Patel, Y.D.; Fine, E.J.; Koenigsberg, M.
A 38-year-old woman presented with fever, right flank pain, and a clinical diagnosis of pyelonephritis. Work-up revealed the presence of a crossing arterial branch causing obstruction of the superior infundibulum of the right kidney, which is an uncommon cause of nephralgia and urinary infection initially described by Fraley in 1966. Intravenous urography, retrograde pyelography, and angiography remain the mainstay of diagnosis, much as in the initial descriptions of this entity. (/sup 131/I)Hippuran imaging, with analysis of the upper and lower pole regions of interest, provides a simple yet powerful method of evaluating functional and excretory changes in the superior infundibulum, and has proved more efficacious than previously reported whole-kidney renograms. Renal scintigraphy represents a relatively noninvasive method of serial functional examination in this disorder. Ultrasound imaging, by monitoring upper-pole dilatation, may provide complementary morphologic information important for long-term follow-up.
Multiple light scattering methods for multiphase flow diagnostics
NASA Astrophysics Data System (ADS)
Estevadeordal, Jordi
Multiphase flows of gases and liquids containing droplets, bubbles, or particulates present light scattering imaging challenges due to the interference from each phase, such as secondary reflections, extinctions, absorptions, and refractions. These factors often prevent the unambiguous detection of each phase and also produce undesired beam steering. The effects can be especially complex in presence of dense phases, multispecies flows, and high pressure environments. This investigation reports new methods for overcoming these effects for quantitative measurements of velocity, density, and temperature fields. The methods are based on light scattering techniques combining Mie and filtered Rayleigh scattering and light extinction analyses and measurements. The optical layout is designed to perform multiple property measurements with improved signal from each phase via laser spectral and polarization characterization, etalon decontamination, and use of multiple wavelengths and imaging detectors.
Apparatus and method for monitoring breath acetone and diabetic diagnostics
Duan, Yixiang; Cao, Wenqing
An apparatus and method for monitoring diabetes through breath acetone detection and quantitation employs a microplasma source in combination with a spectrometer. The microplasma source provides sufficient energy to produce excited acetone fragments from the breath gas that emit light. The emitted light is sent to the spectrometer, which generates an emission spectrum that is used to detect and quantify acetone in the breath gas.
An expanded age range for meningococcal meningitis: molecular diagnostic evidence from population-based surveillance in Asia
Background To understand epidemiologic patterns of meningococcal disease in Asia, we performed a retrospective molecular analysis of cerebrospinal fluid (CSF) specimens collected in prospective surveillance among children aged < 5 years of age in China, South Korea, and Vietnam. Methods A total of 295 isolates and 2,302 CSFs were tested by a meningococcal species- and serogroup-specific polymerase chain reaction (PCR) assay targeting the Neisseria meningitidis (Nm) ctrA gene. Multi-locus sequence typing (MLST) was performed in Nm gene amplification analysis and incidence rates for meningococcal meningitis were estimated. Results Among 295 isolates tested, 10 specimens from Vietnam were confirmed as serogroup B and all were Sequence Type (ST) 1576 by MLST. Among the 2,032 CSF specimen tested, 284 (14%) were confirmed by PCR (ctrA gene), including 67 (23.6%) from China, 92 (32.4%) from Korea, and 125 (44.0%) from Vietnam. Neonates and infants aged < 6 months of age accounted for more than 50% of Nm-PCR positive CSF. Two CSF specimens from Vietnam were identified as serogroup B using MLST. In addition, 44 specimens underwent sequencing to confirm meningococcal serogroup; of these, 21 (48%) were serogroup C, 12 (27%) were serogroup X, 9 (20%) were serogroup Y and 2 (5%) were serogroup B. The incidence rates of meningococcal meningitis among children < 5 years of age was highest in Vietnam (7.4/100,000 [95% CI, 3.6—15.3] followed by Korea (6.8/100,000 [95% CI, 3.5-13.5] and China (2.1/100,000) [95% CI, 0.7-6.2]). Conclusions These results suggest that there is a previously undetected, yet substantial burden of meningococcal meningitis among infants and young children. Standardized, sensitive and specific molecular diagnostic assays with Nm serogrouping capacity are needed throughout Asia to understand the true burden of N. meningitidis disease. PMID:23164061
Assessing molecular line diagnostics of triggered star formation using synthetic observations
NASA Astrophysics Data System (ADS)
Haworth, Thomas J.; Harries, Tim J.; Acreman, David M.; Rundle, David A.
We investigate observational signatures of triggered star formation in bright rimmed clouds (BRCs) by using molecular line transfer calculations based on radiation-hydrodynamic radiatively driven implosion models. We find that for BRCs the separation in velocity between the line profile peak of an optically thick and an optically thin line is determined by both the observer viewing angle and the density of the shell driving into the cloud. In agreement with observations, we find that most BRC line profiles are symmetric and that asymmetries can be either red or blue, in contrast to the blue dominance expected for a collapsing cloud. Asymmetries in the line profiles arise when an optically thick line is dominated by the shell and an optically thin line is dominated by the cloud interior to the shell. The asymmetries are red or blue depending on whether the shell is moving towards or away from the observer, respectively. Using the known motions of the molecular gas in our models we rule out the `envelope expansion with core collapse' mechanism as the cause of the lack of blue-asymmetry in our simulated observations. We show that the absence of a strong photon-dominated region (PDR) around a BRC may not rule out the presence of triggered star formation: if the BRC line profile has a strong blue component then the shell is expected to be driving towards the observer, suggesting that the cloud is being viewed from behind and the PDR is obstructed. This could explain why BRCs such as SFO 80, 81 and 86 have a blue secondary peak and only a weak PDR inferred at 8 μm. Finally we also test the use of 12CO, 13CO and C18O as diagnostics of cloud mass, temperature and column density. We find that the inferred conditions are in reasonable agreement with those from the models. Calculating the cloud mass assuming spherical symmetry is shown to introduce an error of an order of magnitude whereas integrating the column density over a given region is found to introduce an error of
Molecular epidemiological analysis of Mycoplasma bovis isolates from the Pennsylvania Animal Diagnostic Laboratory showing genetic diversity.
Soehnlen, M K; Kariyawasam, S; Lumadue, J A; Pierre, T A; Wolfgang, D R; Jayarao, B M
We have examined the genetic variability of Mycoplasma bovis strains submitted to the Pennsylvania Animal Diagnostics Laboratory, University Park (PA-ADL), between December 2007 and December 2008. Of 4,868 total samples submitted for Mycoplasma testing, 302 were determined to be culture positive. Mycoplasma bovis (63.6%), Mycoplasma californicum (7.3%), Mycoplasma bovirhinis (2.7%), Mycoplasma bovigenitalium (0.7%), Mycoplasma alkalescens (4.9%), Mycoplasma putrefaciens (0.3%), and Mycoplasma dispar (1.3%) and unidentified Mycoplasma sp. (19.2%) were identified using PCR. Mycoplasma bovis represented the largest portion of the positive samples submitted. Each of the 192 M. bovis isolates was examined for variations in the BglII and MfeI restriction sites of the DNA using amplified fragment length polymorphism fingerprinting and subsequently compared with the M. bovis type strain PG45 (ATCC 25523). Similarity between strains was calculated using the Dice similarity coefficient, which ranged from approximately 0.7 to 1.0. When clustering the isolates at greater than 95% similarity, it was determined that 11 distinct clusters were present. The results are consistent with the existence of at least 2 clonally distinct groups. No clear geographical, month of isolation, or source origination relationship was identified, indicating that a currently unclassified characteristic is responsible for the strain heterogeneity. These data indicate strong heterogeneity of M. bovis isolates submitted to PA-ADL. Additionally, multiple sites throughout Pennsylvania had isolates of separate clonal lineages present concomitantly, indicating the ability of multiple overlapping outbreaks to occur at a single location. Mycoplasma bovis represents the largest portion of Mycoplasma species isolated from PA-ADL samples. We propose that amplified fragment length polymorphism may serve as a valuable tool for molecular characterization of M. bovis strains from the United States. PMID:21426978
Assessment of diagnostic methods for solenoid-operated valves
NASA Astrophysics Data System (ADS)
Kryter, R. C.; Farmer, W. S.
Solenoid-operated valves (SOV's) were studied at Oak Ridge National Laboratory as part of the USNRC Nuclear Plant Aging Research (NPAR) Program. The primary objective of the study was to identify, evaluate, and recommend methods for inspection, surveillance, monitoring, and maintenance of SOV's that can help ensure their operational readiness - that is, their ability to perform required safety functions under all anticipated operating conditions, since failure of one of these small and relatively inexpensive devices could have serious consequences under certain circumstances. Intrusive techniques requiring the addition of magnetic or acoustic sensors or the application of special test signals were investigated briefly, but major emphasis was placed on the examination of condition-indicating techniques that can be applied with minimal cost and impact on plant operation. These include monitoring coil mean temperature remotely by means of coil dc resistance or ac impedance, determining valve plunger position by means of coil ac impedance, verifying unrestricted SOV plunger movement by measuring current and voltage at their critical bistable (pull-in and drop-out) values, and detecting the presence of shorted turns or insulation breakdown within the solenoid coil using interrupted-current test methods. Experimental results are presented that demonstrate the technical feasibility and practicality of the monitoring techniques assessed in the study, and recommendations for further work are provided.
eyeGENE®: a vision community resource facilitating patient care and paving the path for research through molecular diagnostic testing
Blain, D; Goetz, KE; Ayyagari, R; Tumminia, SJ
Molecular genetics and genomics are revolutionizing the study and treatment of inherited eye diseases. In recognition of the impact of molecular genetics on vision and ophthalmology, the National Eye Institute established the National Ophthalmic Disease Genotyping and Phenotyping Network (eyeGENE®) as a multidirectional research initiative whereby a clinical component for patients diagnosed with inherited eye disease fosters research into the causes and mechanisms of these ophthalmic diseases. This is accomplished by broadening access to genetic diagnostic testing and maintaining a repository of DNA samples from clinically characterized individuals and their families to allow investigations of the causes, interventions, and management of genetic eye disorders. The eyeGENE® Network currently includes Clinical Laboratory Improvement Amendments (CLIA)-certified diagnostic laboratory partners, over 270 registered clinical organizations with 500 registered users from around the United States and Canada, and is now testing approximately 100 genes representing 35 inherited eye diseases. To date, the Network has received 4400 samples from individuals with rare inherited eye diseases, which are available for access by the vision research community. eyeGENE® is a model partnership between the U.S. federal government, eye health care providers, CLIA-approved molecular diagnostic laboratories, private industry, and scientists who represent a broad research constituency. PMID:23662816
Method for coupling a hydrocarbon containing molecular species
Lingwood, C.A.
A method is described of covalently coupling two molecular species comprising: (a) combining (i) a first molecular species having a functionality reactive with hydrocarbon when photo-activated; and (ii) a solution of at least one, hydrocarbon containing, molecular species in the absence of photo-radiation to which the functionality is sensitive; (b) removing the solvent; (c) irradiating the mixture with photo-radiation to which the functionality is photosensitive.
Making Diagnostic Inferences about Cognitive Attributes Using the Rule-Space Model and Attribute Hierarchy Method
ERIC Educational Resources Information Center
Gierl, Mark J.
The purpose of this paper is to describe the logic and identify key assumptions associated with making cognitive inferences using two attribute-based psychometric methods. The first method is Kikumi Tatsuoka's rule-space model. This model provides a strong point of reference for studying the nature of diagnostic inferences because it is important…
Method of hepatitis diagnostics of changes in human skin diffuse reflectivity
NASA Astrophysics Data System (ADS)
Kirsh, M. L.; Sokol, A. M.; Lomanets, V. S.; Gayka, O. R.
The results on the study of influence of bilirubinum concentration in a human blood on the spectrum of a diffuse reflectivity of his skin are represented. On this basis, the method for hepatitis diagnostics has been developed, and the laboratory device implementing this method has been designed.
Using the Attribute Hierarchy Method to Make Diagnostic Inferences about Examinees' Cognitive Skills in Critical Reading
ERIC Educational Resources Information Center
Wang, Changjiang; Gierl, Mark J.
The purpose of this study is to apply the attribute hierarchy method (AHM) to a subset of SAT critical reading items and illustrate how the method can be used to promote cognitive diagnostic inferences. The AHM is a psychometric procedure for classifying examinees' test item responses into a set of attribute mastery patterns associated with…
[Molecular methods for authentication of Chinese medicinal materials].
Wang, Chuanyi; Guo, Baolin; Xiao, Peigen
The resource authentication is required for quality assurance and control of Chinese medicine. This review provides an informative introduction to molecular methods used for authentication of Chinese medicinal materials. The technical features of the methods based on sequencing, polymerase chain reaction (PCR) and hybridization are described, merits and demerits and development of the molecular methods in identification of Chinese medicinal materials are discussed. PMID:21585017
Exploring chemistry with the fragment molecular orbital method.
Fedorov, Dmitri G; Nagata, Takeshi; Kitaura, Kazuo
The fragment molecular orbital (FMO) method makes possible nearly linear scaling calculations of large molecular systems, such as water clusters, proteins and DNA. In particular, FMO has been widely used in biochemical applications involving protein-ligand binding and drug design. The method has been efficiently parallelized suitable for petascale computing. Many commonly used wave functions and solvent models have been interfaced with FMO. We review the historical background of FMO, and summarize its method development and applications. PMID:22410762
Methods in molecular cardiology: in silico cloning
Passier, R.; Doevendans, P.A.
Advancements in sequencing technology have made it possible to obtain more information about the DNA sequence, structure and the transcript products of the genome from different species. This information is collected in DNA databases. These databases contain many genes of which the functions have not yet been discovered. By using online biotechnology tools novel genes and their transcripts can be identified. The identification of novel genes using DNA database analysis is referred to as in silico cloning. In silico cloning may not only provide new information on genes and their biological function, it may also lead to identification of molecular targets for drug discovery activities. In this review we describe the process of in silico cloning and its application in biomedical research. ImagesFigure 1Figure 3 PMID:25696371
Thomson scattering as a method for laser plasma diagnostics
Alayi, Y.
The Thomson scattering has been used to determine the density and temperature of an inhomogeneous nonstationary plasma. A common method to calibrate the Thomson scattering device consists in replacing the plasma by a gas and measuring the Rayleigh scattering cross section. The angular distribution of the scattered light in Argon is measured, the incident light is a ruby laser with ..delta..t = 30ns and lambda = 6943nm and vertically polarized. We have found that angular distribution is strongly favored in the forward direction (30/sup 0/, 45/sup 0/, 60/sup 0/) and defavored for backward direction (90/sup 0/, 120/sup 0/, 135/sup 0/, 150/sup 0/) in agreement with the results of George, et al, but in disagreement with the Rayleigh theory which assumes a uniform distribution. Our results may be related to the form of the scattered light spectrum which undergoes a dramatic change through the kinetic-hydrodynamic transition. The general form of the spectrum is determined by the parameter y = 1/Kl (where K = 4..pi.. sin (theta/2)/lambda, theta is the scattering angle and l is the free path path), which increases in the direction of the hydrodynamic regime (small angles). By analogy, the Thomson scattering presents the same aspects with ..cap alpha.. = 1/Klambda /SUB D/ (where lambda /SUB D/ is the Debye length). The deviation from the uniform distribution provides the possibility to determine the plasma turbulence spectrum from the scattered light.
Characterization and diagnostic methods for geomagnetic auroral infrasound waves
NASA Astrophysics Data System (ADS)
Oldham, Justin J.
Infrasonic perturbations resulting from auroral activity have been observed since the 1950's. In the last decade advances in infrasonic microphone sensitivity, high latitude sensor coverage, time series analysis methods and computational efficiency have elucidated new types of auroral infrasound. Persistent periods of infrasonic activity associated with geomagnetic sub-storms have been termed geomagnetic auroral infrasound waves [GAIW]. We consider 63 GAIW events recorded by the Fairbanks, AK infrasonic array I53US ranging from 2003 to 2014 and encompassing a complete solar cycle. We make observations of the acoustic features of these events alongside magnetometer, riometer, and all-sky camera data in an effort to quantify the ionospheric conditions suitable for infrasound generation. We find that, on average, the generation mechanism for GAIW is confined to a region centered about ~60 0 longitude east of the anti-Sun-Earth line and at ~770 North latitude. We note furthermore that in all cases considered wherein imaging riometer data are available, that dynamic regions of heightened ionospheric conductivity periodically cross the overhead zenith. Consistent features in concurrent magnetometer conditions are also noted, with irregular oscillations in the horizontal component of the field ubiquitous in all cases. In an effort to produce ionosphere based infrasound free from the clutter and unknowns typical of geophysical observations, an experiment was undertaken at the High Frequency Active Auroral Research Program [HAARP] facility in 2012. Infrasonic signals appearing to originate from a source region overhead were observed briefly on 9 August 2012. The signals were observed during a period when an electrojet current was presumed to have passed overhead and while the facilities radio transmitter was periodically heating the lower ionosphere. Our results suggest dynamic auroral electrojet currents as primary sources of much of the observed infrasound, with
Molecular Mechanics: The Method and Its Underlying Philosophy.
ERIC Educational Resources Information Center
Boyd, Donald B.; Lipkowitz, Kenny B.
Molecular mechanics is a nonquantum mechanical method for solving problems concerning molecular geometries and energy. Methodology based on: the principle of combining potential energy functions of all structural features of a particular molecule into a total force field; derivation of basic equations; and use of available computer programs is…
Diagnostic test accuracy: methods for systematic review and meta-analysis.
Campbell, Jared M; Klugar, Miloslav; Ding, Sandrine; Carmody, Dennis P; Hakonsen, Sasja J; Jadotte, Yuri T; White, Sarahlouise; Munn, Zachary
Systematic reviews are carried out to provide an answer to a clinical question based on all available evidence (published and unpublished), to critically appraise the quality of studies, and account for and explain variations between the results of studies. The Joanna Briggs Institute specializes in providing methodological guidance for the conduct of systematic reviews and has developed methods and guidance for reviewers conducting systematic reviews of studies of diagnostic test accuracy. Diagnostic tests are used to identify the presence or absence of a condition for the purpose of developing an appropriate treatment plan. Owing to demands for improvements in speed, cost, ease of performance, patient safety, and accuracy, new diagnostic tests are continuously developed, and there are often several tests available for the diagnosis of a particular condition. In order to provide the evidence necessary for clinicians and other healthcare professionals to make informed decisions regarding the optimum test to use, primary studies need to be carried out on the accuracy of diagnostic tests and the results of these studies synthesized through systematic review. The Joanna Briggs Institute and its international collaboration have updated, revised, and developed new guidance for systematic reviews, including systematic reviews of diagnostic test accuracy. This methodological article summarizes that guidance and provides detailed advice on the effective conduct of systematic reviews of diagnostic test accuracy. PMID:26355602
New method of acne disease fluorescent diagnostics in natural and fluorescent light and treatment control
NASA Astrophysics Data System (ADS)
Karimova, L. N.; Berezin, A. N.; Shevchik, S. A.; Kharnas, S. S.; Kusmin, S. G.; Loschenov, V. B.
In the given research the new method of fluorescent diagnostics (FD) and photodynamic therapy (PDT) control of acne disease is submitted. Method is based on simultaneous diagnostics in natural and fluorescent light. PDT was based on using 5-ALA (5- aminolevulinic acid) preparation and 600-730 nanometers radiation. If the examined site of a skin possessed a high endogenous porphyrin fluorescence level, PDT was carried out without 5-ALA. For FD and treatment control a dot spectroscopy and the fluorescent imaging of the affected skin were used.
Accurate prediction of lattice energies and structures of molecular crystals with molecular quantum chemistry methods.
Fang, Tao; Li, Wei; Gu, Fangwei; Li, Shuhua
We extend the generalized energy-based fragmentation (GEBF) approach to molecular crystals under periodic boundary conditions (PBC), and we demonstrate the performance of the method for a variety of molecular crystals. With this approach, the lattice energy of a molecular crystal can be obtained from the energies of a series of embedded subsystems, which can be computed with existing advanced molecular quantum chemistry methods. The use of the field compensation method allows the method to take long-range electrostatic interaction of the infinite crystal environment into account and make the method almost translationally invariant. The computational cost of the present method scales linearly with the number of molecules in the unit cell. Illustrative applications demonstrate that the PBC-GEBF method with explicitly correlated quantum chemistry methods is capable of providing accurate descriptions on the lattice energies and structures for various types of molecular crystals. In addition, this approach can be employed to quantify the contributions of various intermolecular interactions to the theoretical lattice energy. Such qualitative understanding is very useful for rational design of molecular crystals. PMID:26574207
Damage diagnosis for SHM of existing civil structure with statistical diagnostic method
NASA Astrophysics Data System (ADS)
Iwasaki, Atsushi; Todoroki, Akira; Sugiya, Tsuneya; Sakai, Shinsuke
The present research proposes a new automatic damage diagnostic method that does not require data of damaged state. Structural health monitoring is a noticeable technology for civil structures. Multiple damage diagnostic method for has been proposed, and most of them employ parametric method based on modeling or non-parametric method such as artificial neural networks. These methods demand much costs, and first of all, it is impossible to obtain data for training of damaged existing structures. That causes importance of development of the method, which diagnoses damage just from data of the intact state structure for existing structures. Therefore we purpose new statistical diagnostic method for structural damage detection. In the present method, system identification using a response surface is performed and damage is diagnosed by testing the change of this identified system by statistical test. The new method requires data of non-damaged state and does not require the complicated modeling and data of damaged state structure. As an example, the present study deals damage diagnosis of a jet-fan which installed to a tunnel on an expressway as a ventilator fan. Damages are detected from load of turnbuckles. As a result, the damage is successfully diagnosed with the method.
A multivariate method for meta-analysis and comparison of diagnostic tests.
Dimou, Niki L; Adam, Maria; Bagos, Pantelis G
We present here an extension of the classic bivariate random effects meta-analysis for the log-transformed sensitivity and specificity that can be applied for two or more diagnostic tests. The advantage of this method is that a closed-form expression is derived for the calculation of the within-studies covariances. The method allows the direct calculation of sensitivity and specificity, as well as, the diagnostic odds ratio, the area under curve and the parameters of the summary receiver operator's characteristic curve, along with the means for a formal comparison of these quantities for different tests. There is no need for individual patient data or the simultaneous evaluation of both diagnostic tests in all studies. The method is simple and fast; it can be extended for several diagnostic tests and can be fitted in nearly all statistical packages. The method was evaluated in simulations and applied in a meta-analysis for the comparison of anti-cyclic citrullinated peptide antibody and rheumatoid factor for discriminating patients with rheumatoid arthritis, with encouraging results. Simulations suggest that the method is robust and more powerful compared with the standard bivariate approach that ignores the correlation between tests. Copyright © 2016 John Wiley & Sons, Ltd. PMID:26940666
Molecular Methods for the Detection of Mycoplasma and Ureaplasma Infections in Humans
Waites, Ken B.; Xiao, Li; Paralanov, Vanya; Viscardi, Rose M.; Glass, John I.
Mycoplasma and Ureaplasma species are well-known human pathogens responsible for a broad array of inflammatory conditions involving the respiratory and urogenital tracts of neonates, children, and adults. Greater attention is being given to these organisms in diagnostic microbiology, largely as a result of improved methods for their laboratory detection, made possible by powerful molecular-based techniques that can be used for primary detection in clinical specimens. For slow-growing species, such as Mycoplasma pneumoniae and Mycoplasma genitalium, molecular-based detection is the only practical means for rapid microbiological diagnosis. Most molecular-based methods used for detection and characterization of conventional bacteria have been applied to these organisms. A complete genome sequence is available for one or more strains of all of the important human pathogens in the Mycoplasma and Ureaplasma genera. Information gained from genome analyses and improvements in efficiency of DNA sequencing are expected to significantly advance the field of molecular detection and genotyping during the next few years. This review provides a summary and critical review of methods suitable for detection and characterization of mycoplasmas and ureaplasmas of humans, with emphasis on molecular genotypic techniques. PMID:22819362
Expanding Cancer Detection Using Molecular Imprinting for a Novel Point-of-Care Diagnostic Device
NASA Astrophysics Data System (ADS)
Yu, Yingjie; Rafailovich, Miriam; Wang, Yantian; Ranjbaran, Alina; Wang, Tom; Nam, David
We propose the use of a potentiometric biosensor that incorporates the efficient and specific molecular imprinting (MI) method with a self-assembled monolayer (SAM). We first tested the biosensor using carcinoembryonic antigen, CEA, a biomarker associated with pancreatic cancer. No change in detection efficiency was observed when detection was performed in the presence of 100% serum albumin, indicating that the sensor is able to discriminate for the template analyte even in concentrated solution of similar substances. Computer simulations of the protein structure were performed in order to estimate the changes in morphology and determine the sensitivity of the biosensor to conformational changes in the proteins. We found that even small changes in PH can generate rotation of the surface functional groups, without significant change in the morphology. Yet, the results show that only when the detection and imprinting conditions are similar, robust signals occurs. Hence we concluded that both morphology and surface chemistry play a role in the recognition.
Efficient molecular surface generation using level-set methods.
Can, Tolga; Chen, Chao-I; Wang, Yuan-Fang
Molecules interact through their surface residues. Calculation of the molecular surface of a protein structure is thus an important step for a detailed functional analysis. One of the main considerations in comparing existing methods for molecular surface computations is their speed. Most of the methods that produce satisfying results for small molecules fail to do so for large complexes. In this article, we present a level-set-based approach to compute and visualize a molecular surface at a desired resolution. The emerging level-set methods have been used for computing evolving boundaries in several application areas from fluid mechanics to computer vision. Our method provides a uniform framework for computing solvent-accessible, solvent-excluded surfaces and interior cavities. The computation is carried out very efficiently even for very large molecular complexes with tens of thousands of atoms. We compared our method to some of the most widely used molecular visualization tools (Swiss-PDBViewer, PyMol, and Chimera) and our results show that we can calculate and display a molecular surface 1.5-3.14 times faster on average than all three of the compared programs. Furthermore, we demonstrate that our method is able to detect all of the interior inaccessible cavities that can accommodate one or more water molecules. PMID:16621636
Method of deposition by molecular beam epitaxy
Chalmers, S.A.; Killeen, K.P.; Lear, K.L.
A method is described for reproducibly controlling layer thickness and varying layer composition in an MBE deposition process. In particular, the present invention includes epitaxially depositing a plurality of layers of material on a substrate with a plurality of growth cycles whereby the average of the instantaneous growth rates for each growth cycle and from one growth cycle to the next remains substantially constant as a function of time. 9 figures.
Method of deposition by molecular beam epitaxy
Chalmers, Scott A.; Killeen, Kevin P.; Lear, Kevin L.
A method is described for reproducibly controlling layer thickness and varying layer composition in an MBE deposition process. In particular, the present invention includes epitaxially depositing a plurality of layers of material on a substrate with a plurality of growth cycles whereby the average of the instantaneous growth rates for each growth cycle and from one growth cycle to the next remains substantially constant as a function of time.
Molecular diagnostic procedures for production of pathogen-free propagation material.
Manulis, Shulamit; Chalupowicz, Laura; Dror, Orit; Kleitman, Frida
Production of disease-free propagation material is a major means of controlling most bacterial diseases of plants, particularly when neither resistant clones nor effective chemical treatments are available. For this purpose sensitive, specific and rapid detection methods are required. The advent of molecular biology and, in particular, the polymerase chain reaction (PCR) has opened new ways for the characterization and identification of plant pathogens and the development of disease-management strategies. PCR-based detection methods rely on the development of primers for the specific detection of the pathogen. The use of pathogenicity genes as targets for primer design is the preferred procedure for obtaining specific primers but other procedures may also be useful for this purpose. In the present review we describe four examples of procedures for detecting four important bacterial pathogens in Israel: Erwinia herbicola pv gypsophilae in gypsophila, Xanthomonas campestris pv pelargonii in geranium, Agrobacterium tumefaciens in asters and roses, and Xanthomonas campestris pv campestris in crucifers. Procedures for constructing specific PCR primers for each bacterium are illustrated and discussed as well as the combination of PCR with other methods. PMID:12449531
Hydroxyalkyl phosphine gold complexes for use as diagnostic and therapeutic pharmaceuticals and method of making same
Katti, K.V.; Berning, D.E.; Volkert, W.A.; Ketring, A.R.
A complex and method for making a diagnostic or therapeutic pharmaceutical includes a ligand comprising at least one hydroxyalkyl phosphine donor group bound to a gold atom to form a gold-ligand complex that is stable in aqueous solutions containing oxygen, serum and other body fluids. 20 figs.
Hydroxyalkyl phosphine gold complexes for use as diagnostic and therapeutic pharmaceuticals and method of making same
Katti, Kattesh V.; Berning, Douglas E.; Volkert, Wynn A.; Ketring, Alan R.
A complex and method for making same for use as a diagnostic or therapeutic pharmaceutical includes a ligand comprising at least one hydroxyalkyl phosphine donor group bound to a gold atom to form a gold-ligand complex that is stable in aqueous solutions containing oxygen, serum and other body fluids.
Hydroxymethyl phosphine compounds for use as diagnostic and therapeutic pharmaceuticals and method of making same
Katti, K.V.; Karra, S.R.; Berning, D.E.; Smith, C.J.; Volkert, W.A.; Ketring, A.R.
A compound and method of making a compound for use as a diagnostic or therapeutic pharmaceutical comprises at least one functionalized hydroxyalkyl phosphine donor group and one or more sulfur or nitrogen donor and a metal combined with the ligand. 21 figs.
Validation of a Theoretical Model of Diagnostic Classroom Assessment: A Mixed Methods Study
ERIC Educational Resources Information Center
Koh, Nancy
The purpose of the study was to validate a theoretical model of diagnostic, formative classroom assessment called, "Proximal Assessment for Learner Diagnosis" (PALD). To achieve its purpose, the study employed a two-stage, mixed-methods design. The study utilized multiple data sources from 11 elementary level mathematics teachers who…
Patents on diagnostic methods in Europe under the European Patent Convention (EPC).
Barba, Michelangelo
This article reviews the provisions of the European Patent Convention with regard to diagnostic methods practiced on the human or animal body. Moreover, it is also discussed the relevant jurisprudence (Case Law) interpreting the legal provisions. Some examples based on real cases are also presented and discussed. PMID:21095714
Depression and Spinal Cord Injury: A Review of Diagnostic Methods for Depression, 1985 to 2000.
ERIC Educational Resources Information Center
Skinner, Amy L.; Armstrong, Kevin J.; Rich, John
Studies of depression in individuals with spinal cord injuries (SCI) over a 15-year period were examined to determine if researchers used consistent diagnostic measures. The Beck Depression Inventory was the most frequently used instrument, but there was inconsistency among methods employed and disagreement regarding the inclusion of somatic…
Study on Fault Diagnostics of a Turboprop Engine Using Inverse Performance Model and Artificial Intelligent Methods
NASA Astrophysics Data System (ADS)
Kong, Changduk; Lim, Semyeong
Recently, the health monitoring system of major gas path components of gas turbine uses mostly the model based method like the Gas Path Analysis (GPA). This method is to find quantity changes of component performance characteristic parameters such as isentropic efficiency and mass flow parameter by comparing between measured engine performance parameters such as temperatures, pressures, rotational speeds, fuel consumption, etc. and clean engine performance parameters without any engine faults which are calculated by the base engine performance model. Currently, the expert engine diagnostic systems using the artificial intelligent methods such as Neural Networks (NNs), Fuzzy Logic and Genetic Algorithms (GAs) have been studied to improve the model based method. Among them the NNs are mostly used to the engine fault diagnostic system due to its good learning performance, but it has a drawback due to low accuracy and long learning time to build learning data base if there are large amount of learning data. In addition, it has a very complex structure for finding effectively single type faults or multiple type faults of gas path components. This work builds inversely a base performance model of a turboprop engine to be used for a high altitude operation UAV using measured performance data, and proposes a fault diagnostic system using the base engine performance model and the artificial intelligent methods such as Fuzzy logic and Neural Network. The proposed diagnostic system isolates firstly the faulted components using Fuzzy Logic, then quantifies faults of the identified components using the NN leaned by fault learning data base, which are obtained from the developed base performance model. In leaning the NN, the Feed Forward Back Propagation (FFBP) method is used. Finally, it is verified through several test examples that the component faults implanted arbitrarily in the engine are well isolated and quantified by the proposed diagnostic system.
Estimation of diagnostic test accuracy without full verification: a review of latent class methods
Collins, John; Huynh, Minh
The performance of a diagnostic test is best evaluated against a reference test that is without error. For many diseases, this is not possible, and an imperfect reference test must be used. However, diagnostic accuracy estimates may be biased if inaccurately verified status is used as the truth. Statistical models have been developed to handle this situation by treating disease as a latent variable. In this paper, we conduct a systematized review of statistical methods using latent class models for estimating test accuracy and disease prevalence in the absence of complete verification. PMID:24910172
BRAF mutation testing with a rapid, fully integrated molecular diagnostics system
Huang, Helen J.; Falchook, Gerald S.; Devogelaere, Benoit; Kockx, Mark; Bempt, Isabelle Vanden; Reijans, Martin; Naing, Aung; Fu, Siqing; Piha-Paul, Sarina A.; Hong, David S.; Holley, Veronica R.; Tsimberidou, Apostolia M.; Stepanek, Vanda M.; Patel, Sapna P.; Kopetz, E. Scott; Subbiah, Vivek; Wheler, Jennifer J.; Zinner, Ralph G.; Karp, Daniel D.; Luthra, Rajyalakshmi; Roy-Chowdhuri, Sinchita; Sablon, Erwin; Meric-Bernstam, Funda; Maertens, Geert; Kurzrock, Razelle
Fast and accurate diagnostic systems are needed for further implementation of precision therapy of BRAF-mutant and other cancers. The novel IdyllaTM BRAF Mutation Test has high sensitivity and shorter turnaround times compared to other methods. We used Idylla to detect BRAF V600 mutations in archived formalin-fixed paraffin-embedded (FFPE) tumor samples and compared these results with those obtained using the cobas 4800 BRAF V600 Mutation Test or MiSeq deep sequencing system and with those obtained by a Clinical Laboratory Improvement Amendments (CLIA)-certified laboratory employing polymerase chain reaction–based sequencing, mass spectrometric detection, or next-generation sequencing. In one set of 60 FFPE tumor samples (15 with BRAF mutations per Idylla), the Idylla and cobas results had an agreement of 97%. Idylla detected BRAF V600 mutations in two additional samples. The Idylla and MiSeq results had 100% concordance. In a separate set of 100 FFPE tumor samples (64 with BRAF mutation per Idylla), the Idylla and CLIA-certified laboratory results demonstrated an agreement of 96% even though the tests were not performed simultaneously and different FFPE blocks had to be used for 9 cases. The IdyllaTM BRAF Mutation Test produced results quickly (sample to results time was about 90 minutes with about 2 minutes of hands on time) and the closed nature of the cartridge eliminates the risk of PCR contamination. In conclusion, our observations demonstrate that the Idylla test is rapid and has high concordance with other routinely used but more complex BRAF mutation–detecting tests. PMID:26330075
BRAF mutation testing with a rapid, fully integrated molecular diagnostics system.
Janku, Filip; Claes, Bart; Huang, Helen J; Falchook, Gerald S; Devogelaere, Benoit; Kockx, Mark; Bempt, Isabelle Vanden; Reijans, Martin; Naing, Aung; Fu, Siqing; Piha-Paul, Sarina A; Hong, David S; Holley, Veronica R; Tsimberidou, Apostolia M; Stepanek, Vanda M; Patel, Sapna P; Kopetz, E Scott; Subbiah, Vivek; Wheler, Jennifer J; Zinner, Ralph G; Karp, Daniel D; Luthra, Rajyalakshmi; Roy-Chowdhuri, Sinchita; Sablon, Erwin; Meric-Bernstam, Funda; Maertens, Geert; Kurzrock, Razelle
Fast and accurate diagnostic systems are needed for further implementation of precision therapy of BRAF-mutant and other cancers. The novel IdyllaTMBRAF Mutation Test has high sensitivity and shorter turnaround times compared to other methods. We used Idylla to detect BRAF V600 mutations in archived formalin-fixed paraffin-embedded (FFPE) tumor samples and compared these results with those obtained using the cobas 4800 BRAF V600 Mutation Test or MiSeq deep sequencing system and with those obtained by a Clinical Laboratory Improvement Amendments (CLIA)-certified laboratory employing polymerase chain reaction-based sequencing, mass spectrometric detection, or next-generation sequencing. In one set of 60 FFPE tumor samples (15 with BRAF mutations per Idylla), the Idylla and cobas results had an agreement of 97%. Idylla detected BRAF V600 mutations in two additional samples. The Idylla and MiSeq results had 100% concordance. In a separate set of 100 FFPE tumor samples (64 with BRAF mutation per Idylla), the Idylla and CLIA-certified laboratory results demonstrated an agreement of 96% even though the tests were not performed simultaneously and different FFPE blocks had to be used for 9 cases. The IdyllaTMBRAF Mutation Test produced results quickly (sample to results time was about 90 minutes with about 2 minutes of hands on time) and the closed nature of the cartridge eliminates the risk of PCR contamination. In conclusion, our observations demonstrate that the Idylla test is rapid and has high concordance with other routinely used but more complex BRAF mutation-detecting tests. PMID:26330075
Surveillance of Acute Respiratory Infections Using Community-Submitted Symptoms and Specimens for Molecular Diagnostic Testing
Goff, Jennifer; Rowe, Aaron; Brownstein, John S.; Chunara, Rumi
Participatory systems for surveillance of acute respiratory infection give real-time information about infections circulating in the community, yet to-date are limited to self-reported syndromic information only and lacking methods of linking symptom reports to infection types. We developed the GoViral platform to evaluate whether a cohort of lay volunteers could, and would find it useful to, contribute self-reported symptoms online and to compare specimen types for self-collected diagnostic information of sufficient quality for respiratory infection surveillance. Volunteers were recruited, given a kit (collection materials and customized instructions), instructed to report their symptoms weekly, and when sick with cold or flu-like symptoms, requested to collect specimens (saliva and nasal swab). We compared specimen types for respiratory virus detection sensitivity (via polymerase-chain-reaction) and ease of collection. Participants were surveyed to determine receptivity to participating when sick, to receiving information on the type of pathogen causing their infection and types circulating near them. Between December 1 2013 and March 1 2014, 295 participants enrolled in the study and received a kit. Of those who reported symptoms, half (71) collected and sent specimens for analysis. Participants submitted kits on average 2.30 days (95 CI: 1.65 to 2.96) after symptoms began. We found good concordance between nasal and saliva specimens for multiple pathogens, with few discrepancies. Individuals report that saliva collection is easiest and report that receiving information about what pathogen they, and those near them, have is valued and can shape public health behaviors. Community-submitted specimens can be used for the detection of acute respiratory infection with individuals showing receptivity for participating and interest in a real-time picture of respiratory pathogens near them. PMID:26075141
Expanding the clinical spectrum of the 'HDAC8-phenotype' - implications for molecular diagnostics, counseling and risk prediction.
Parenti, I; Gervasini, C; Pozojevic, J; Wendt, K S; Watrin, E; Azzollini, J; Braunholz, D; Buiting, K; Cereda, A; Engels, H; Garavelli, L; Glazar, R; Graffmann, B; Larizza, L; Lüdecke, H J; Mariani, M; Masciadri, M; Pié, J; Ramos, F J; Russo, S; Selicorni, A; Stefanova, M; Strom, T M; Werner, R; Wierzba, J; Zampino, G; Gillessen-Kaesbach, G; Wieczorek, D; Kaiser, F J
Cornelia de Lange syndrome (CdLS) is a clinically heterogeneous disorder characterized by typical facial dysmorphism, cognitive impairment and multiple congenital anomalies. Approximately 75% of patients carry a variant in one of the five cohesin-related genes NIPBL, SMC1A, SMC3, RAD21 and HDAC8. Herein we report on the clinical and molecular characterization of 11 patients carrying 10 distinct variants in HDAC8. Given the high number of variants identified so far, we advise sequencing of HDAC8 as an indispensable part of the routine molecular diagnostic for patients with CdLS or CdLS-overlapping features. The phenotype of our patients is very broad, whereas males tend to be more severely affected than females, who instead often present with less canonical CdLS features. The extensive clinical variability observed in the heterozygous females might be at least partially associated with a completely skewed X-inactivation, observed in seven out of eight female patients. Our cohort also includes two affected siblings whose unaffected mother was found to be mosaic for the causative mutation inherited to both affected children. This further supports the urgent need for an integration of highly sensitive sequencing technology to allow an appropriate molecular diagnostic, genetic counseling and risk prediction. PMID:26671848
Development of Companion Diagnostics
Mankoff, David A.; Edmonds, Christine E.; Farwell, Michael D.; Pryma, Daniel A.
The goal of individualized and targeted treatment and precision medicine requires the assessment of potential therapeutic targets to direct treatment selection. The biomarkers used to direct precision medicine, often termed companion diagnostics, for highly targeted drugs have thus far been almost entirely based on in vitro assay of biopsy material. Molecular imaging companion diagnostics offer a number of features complementary to those from in vitro assay, including the ability to measure the heterogeneity of each patient's cancer across the entire disease burden and to measure early changes in response to treatment. We discuss the use of molecular imaging methods as companion diagnostics for cancer therapy with the goal of predicting response to targeted therapy and measuring early (pharmacodynamic) response as an indication of whether the treatment has "hit" the target. We also discuss considerations for probe development for molecular imaging companion diagnostics, including both small-molecule probes and larger molecules such as labeled antibodies and related constructs. We then describe two examples where both predictive and pharmacodynamic molecular imaging markers have been tested in humans: endocrine therapy for breast cancer and human epidermal growth factor receptor type 2–targeted therapy. The review closes with a summary of the items needed to move molecular imaging companion diagnostics from early studies into multicenter trials and into the clinic. PMID:26687857
External Quality Assessment of MERS-CoV Molecular Diagnostics During the 2015 Korean Outbreak
Seong, Moon-Woo; Lee, Seung Jun; Cho, Sung Im; Ko, Kyungphil; Kim, Mi-Na; Sung, Heungsub; Kim, Jae-Seok; Ahn, Ji Soo; Yu, Byung Su; Kim, Taek Soo; Kim, Eui Chong
Background The largest outbreak of Middle East respiratory syndrome coronavirus (MERS-CoV) infection outside Middle East Asia in 2015 has necessitated the rapid expansion of laboratories that conduct MERS-CoV molecular testing in Korea, together with external quality assessment (EQA) to evaluate the assays used. Methods The EQA program consisted of two phases; self-validation and blind assessment. For the first EQA phase, in vitro transcribed upstream region of the envelope gene (upE) and the open reading frame (ORF)1a RNAs were used at a concentration of 1,000 copies/µL. The test panel for the second EQA phase consisted of RNA extracts from three samples, which were obtained from two MERS-CoV positive patients and one MERS-CoV negative patient. Results The first EQA phase results for 46 participants showed a linear relationship between the threshold cycle (CT) values of RNA materials and the logarithmic concentrations for both upE and ORF1a gene targets (R2=0.73 and 0.75, respectively). The mean CT value for each concentration was different depending on which commercial kit was used for the assay. Among the three commonly used kits, PowerChek MERS Real-Time PCR kit (KogeneBiotech, Korea) showed the lowest CT values at all concentrations of upE and most concentrations of ORF1a. The second EQA phase results for 47 participants were 100% correct for all tested samples. Conclusions This EQA survey demonstrates that the MERS-CoV molecular testing performed in Korea during the 2015 outbreak is of robust capability. However, careful establishment and validation of a cut-off value are recommended to ensure good analytical sensitivity. PMID:26915611
Comparing implementations of magnetic-resonance-guided fluorescence molecular tomography for diagnostic classification of brain tumors
NASA Astrophysics Data System (ADS)
Davis, Scott C.; Samkoe, Kimberley S.; O'Hara, Julia A.; Gibbs-Strauss, Summer L.; Paulsen, Keith D.; Pogue, Brian W.
Fluorescence molecular tomography (FMT) systems coupled to conventional imaging modalities such as magnetic resonance imaging (MRI) and computed tomography provide unique opportunities to combine data sets and improve image quality and content. Yet, the ideal approach to combine these complementary data is still not obvious. This preclinical study compares several methods for incorporating MRI spatial prior information into FMT imaging algorithms in the context of in vivo tissue diagnosis. Populations of mice inoculated with brain tumors that expressed either high or low levels of epidermal growth factor receptor (EGFR) were imaged using an EGF-bound near-infrared dye and a spectrometer-based MRI-FMT scanner. All data were spectrally unmixed to extract the dye fluorescence from the tissue autofluorescence. Methods to combine the two data sets were compared using student's t-tests and receiver operating characteristic analysis. Bulk fluorescence measurements that made up the optical imaging data set were also considered in the comparison. While most techniques were able to distinguish EGFR(+) tumors from EGFR(-) tumors and control animals, with area-under-the-curve values=1, only a handful were able to distinguish EGFR(-) tumors from controls. Bulk fluorescence spectroscopy techniques performed as well as most imaging techniques, suggesting that complex imaging algorithms may be unnecessary to diagnose EGFR status in these tissue volumes.
A hierarchical method for molecular docking using cloud computing.
Kang, Ling; Guo, Quan; Wang, Xicheng
Discovering small molecules that interact with protein targets will be a key part of future drug discovery efforts. Molecular docking of drug-like molecules is likely to be valuable in this field; however, the great number of such molecules makes the potential size of this task enormous. In this paper, a method to screen small molecular databases using cloud computing is proposed. This method is called the hierarchical method for molecular docking and can be completed in a relatively short period of time. In this method, the optimization of molecular docking is divided into two subproblems based on the different effects on the protein-ligand interaction energy. An adaptive genetic algorithm is developed to solve the optimization problem and a new docking program (FlexGAsDock) based on the hierarchical docking method has been developed. The implementation of docking on a cloud computing platform is then discussed. The docking results show that this method can be conveniently used for the efficient molecular design of drugs. PMID:23017886
Comparison of laser spectroscopic PNC method with laser integral fluorescence in optical caries diagnostics
NASA Astrophysics Data System (ADS)
Masychev, Victor I.
In this research we represent the results of approbation of two methods of optical caries diagnostics: PNC-spectral diagnostics and caries detection by laser integral fluorescence. The research was conducted in a dental clinic. PNC-method analyzes parameters of probing laser radiation and PNC-spectrums of stimulated secondary radiations: backscattering and endogenous fluorescence of caries- involved bacteria. Ia-Ne laser ((lambda) equals632.8 nm, 1-2 mW) was used as a source of probing (stimulated) radiation. For registration of signals, received from intact and pathological teeth PDA-detector was applied. PNC-spectrums were processed by special algorithms, and were displayed on PC monitor. The method of laser integral fluorescence was used for comparison. In this case integral power of fluorescence of human teeth was measured. As a source of probing (stimulated) radiation diode lasers ((lambda) equals655 nm, 0.1 mW and 630 nm, 1 mW) and Ia-Na laser were applied. For registration of signals Si-photodetector was used. Integral power was shown in a digital indicator. Advantages and disadvantages of these methods are described in this research. It is disclosed that the method of laser integral power of fluorescence has the following characteristics: simplicity of construction and schema-technical decisions. However the method of PNC-spectral diagnostics are characterized by considerably more sensitivity in diagnostics of initial caries and capability to differentiate pathologies of various stages (for example, calculus/initial caries). Estimation of spectral characteristics of PNC-signals allows eliminating a number of drawbacks, which are character for detection by method of laser integral fluorescence (for instance, detection of fluorescent fillings, plagues, calculus, discolorations generally, amalgam, gold fillings as if it were caries).
Application of modern diagnostic methods to environmental improvement. Annual progress report, January--October 1994
Shepard, W.S.
The Diagnostic Instrumentation and Analysis Laboratory (DIAL), a research department in the College of Engineering at Mississippi State University (MSU), is under contract with the US Department of Energy (DOE) to develop and apply advanced diagnostic instrumentation and analysis techniques to real world processes; measurements are made in hot, highly corrosive atmospheres in which conventional measurement devices are ineffective. Task 1 of this agreement is concerned with the development and application of various diagnostic methods to characterize the plasma properties, the melt properties and the downstream emissions from a plasma torch facility designed to vitrify mixed waste. Correlation of the measured properties with the operating parameters of the torch will be sought to improve, optimize and control the overall operation of the plasma treatment process. As part of this program, diagnostic methods will be developed and evaluated for characterization, monitoring and control purposes of treatment processes in general. Task 2 of this agreement is concerned with the development of a system to monitor and control the combustion stoichiometry in real time in order to minimize environmental impact and maximize process efficiency. Staged fuel injection is also being studied to minimize NO{sub x} formation.
Molecular assay optimized by Taguchi experimental design method for venous thromboembolism investigation.
Celani de Souza, Helder Jose; Moyses, Cinthia B; Pontes, Fabrício J; Duarte, Roberto N; Sanches da Silva, Carlos Eduardo; Alberto, Fernando Lopes; Ferreira, Ubirajara R; Silva, Messias Borges
Two mutations - Factor V Leiden (1691G > A) and the 20210G > A on the Prothrombin gene - are key risk factors for a frequent and potentially fatal disorder called Venous Thromboembolism. These molecular alterations can be investigated using real-time Polymerase Chain Reaction (PCR) with Fluorescence Resonance Energy Transfer (FRET) probes and distinct DNA pools for both factors. The objective of this paper is to present an application of Taguchi Experimental Design Method to determine the best parameters adjustment of a Molecular Assays Process in order to obtain the best diagnostic result for Venous Thromboembolism investigation. The complete process contains six three-level factors which usually demands 729 experiments to obtain the final result, if using a Full Factorial Array. In this research, a Taguchi L27 Orthogonal Array is chosen to optimize the analysis and reduce the number of experiments to 27 without degrading the final result accuracy. The application of this method can lessen the time and cost necessary to achieve the best operation condition for a required performance. The results is proven in practice and confirmed that the Taguchi method can really offer a good approach for clinical assay efficiency and effectiveness improvement even though the clinical diagnostics can be based on the use of qualitative techniques. PMID:21867748
Instrumentation for noninvasive express-diagnostics bacteriophages and viruses by optical method
NASA Astrophysics Data System (ADS)
Moguilnaia, Tatiana A.; Andreev, Gleb I.; Agibalov, Andrey A.; Botikov, Andrey G.; Kosenkov, Evgeniy; Saguitova, Elena
The theoretical and the experimental researches of spectra of absent-minded radiation in medium containing viruses were carried out. The information on spectra luminescence 31 viruses was written down.The new method the express - analysis of viruses in organism of the man was developed. It shall be mentioned that the proposed method of express diagnostics allows detection of infection agent in the organism several hours after infection. It makes it suitable for high efficient testing in blood services for detection and rejection of potential donors infected with such viruses as hepatitis, herpes, Epstein-Barre, cytomegalovirus, and immunodeficiency. Methods of serum diagnostics used for that purpose can detect antibodies to virus only 1-3 months after the person has been infected. The device for the express analysis of 31 viruses of the man was created.
Solar thermal polymerase chain reaction for smartphone-assisted molecular diagnostics.
Jiang, Li; Mancuso, Matthew; Lu, Zhengda; Akar, Gunkut; Cesarman, Ethel; Erickson, David
Nucleic acid-based diagnostic techniques such as polymerase chain reaction (PCR) are used extensively in medical diagnostics due to their high sensitivity, specificity and quantification capability. In settings with limited infrastructure and unreliable electricity, however, access to such devices is often limited due to the highly specialized and energy-intensive nature of the thermal cycling process required for nucleic acid amplification. Here we integrate solar heating with microfluidics to eliminate thermal cycling power requirements as well as create a simple device infrastructure for PCR. Tests are completed in less than 30 min, and power consumption is reduced to 80 mW, enabling a standard 5.5 Wh iPhone battery to provide 70 h of power to this system. Additionally, we demonstrate a complete sample-to-answer diagnostic strategy by analyzing human skin biopsies infected with Kaposi's Sarcoma herpesvirus (KSHV/HHV-8) through the combination of solar thermal PCR, HotSHOT DNA extraction and smartphone-based fluorescence detection. We believe that exploiting the ubiquity of solar thermal energy as demonstrated here could facilitate broad availability of nucleic acid-based diagnostics in resource-limited areas. PMID:24553130
Solar thermal polymerase chain reaction for smartphone-assisted molecular diagnostics
NASA Astrophysics Data System (ADS)
Jiang, Li; Mancuso, Matthew; Lu, Zhengda; Akar, Gunkut; Cesarman, Ethel; Erickson, David
Nucleic acid-based diagnostic techniques such as polymerase chain reaction (PCR) are used extensively in medical diagnostics due to their high sensitivity, specificity and quantification capability. In settings with limited infrastructure and unreliable electricity, however, access to such devices is often limited due to the highly specialized and energy-intensive nature of the thermal cycling process required for nucleic acid amplification. Here we integrate solar heating with microfluidics to eliminate thermal cycling power requirements as well as create a simple device infrastructure for PCR. Tests are completed in less than 30 min, and power consumption is reduced to 80 mW, enabling a standard 5.5 Wh iPhone battery to provide 70 h of power to this system. Additionally, we demonstrate a complete sample-to-answer diagnostic strategy by analyzing human skin biopsies infected with Kaposi's Sarcoma herpesvirus (KSHV/HHV-8) through the combination of solar thermal PCR, HotSHOT DNA extraction and smartphone-based fluorescence detection. We believe that exploiting the ubiquity of solar thermal energy as demonstrated here could facilitate broad availability of nucleic acid-based diagnostics in resource-limited areas.
Expanding Cancer Detection Using Molecular Imprinting for a Novel Point-of-Care Diagnostic Device
NASA Astrophysics Data System (ADS)
Yu, Yingjie; Rafailovich, Miriam; Wang, Yantian; Kang, Yeona; Zhang, Lingxi; Rigas, Basil; Division of Gastroenterology, School of Medicine Team
We propose the use of a potentiometric biosensor that incorporates the efficient and specific molecular imprinting (MI) method with a self-assembled monolayer (SAM). We first tested the biosensor using carcinoembryonic antigen, CEA, a biomarker associated with pancreatic cancer. No change in detection efficiency was observed, indicating that the sensor is able to discriminate for the template analyte even in concentrated solution of similar substances. In addition, we use biosensor to discriminate normal fibrinogen and damaged fibrinogen, which is critical for the detection of bleeding disorder. Computer simulations of the protein structure were performed in order to estimate the changes in morphology and determine the sensitivity of the biosensor to conformational changes in the proteins. We found that even small changes in PH can generate rotation of the surface functional groups. Yet, the results show that only when the detection and imprinting conditions are similar, robust signals occurs. Hence we concluded that both morphology and surface chemistry play a role in the recognition.
The Diagnostic Use of Immunohistochemical Surrogates for Signature Molecular Genetic Alterations in Gliomas.
Tanboon, Jantima; Williams, Erik A; Louis, David N
A number of key mutations that affect treatment and prognosis have been identified in human gliomas. Two major ways to identify these mutations in a tumor sample are direct interrogation of the mutated DNA itself and immunohistochemistry to assess the effects of the mutated genes on proteins. Immunohistochemistry is an affordable, robust, and widely available technology that has been in place for decades. For this reason, the use of immunohistochemical approaches to assess molecular genetic changes has become an essential component of state-of-the-art practice. In contrast, even though DNA sequencing technologies are undergoing rapid development, many medical centers do not have access to such methodologies and may be thwarted by the relatively high costs of sending out such tests to reference laboratories. This review summarizes the current experience using immunohistochemistry of glioma samples to identify mutations in IDH1, TP53, ATRX, histone H3 genes, BRAF, EGFR, MGMT, CIC, and FUBP1 as well as guidelines for prudent use of DNA sequencing as a supplemental method. PMID:26671986
DNA Variations in Oculocutaneous Albinism: An Updated Mutation List and Current Outstanding Issues in Molecular Diagnostics
Simeonov, Dimitre R.; Wang, Xinjing; Wang, Chen; Sergeev, Yuri; Dolinska, Monika; Bower, Matthew; Fischer, Roxanne; Winer, David; Dubrovsky, Genia; Balog, Joan Z.; Huizing, Marjan; Hart, Rachel; Zein, Wadih M.; Gahl, William A.; Brooks, Brian P.; Adams, David R.
Oculocutaneous albinism (OCA) is a rare genetic disorder of melanin synthesis that results in hypopigmented hair, skin, and eyes. There are four types of OCA, caused by mutations in TYR (OCA-1), OCA2 (OCA-2), TYRP1 (OCA-3), or SLC45A2 (OCA-4). Here we report 22 novel mutations; 14 from a cohort of 61 patients seen as part of the NIH OCA Natural History Study and 8 from a prior study at the University of Minnesota. We also include a comprehensive list of almost 600 previously reported OCA mutations, along with ethnicity information, carrier frequencies, and in silico pathogenicity predictions. In addition to discussing the clinical and molecular features of OCA, we address the cases of apparent missing heritability. In our cohort, 25% of patients did not have two mutations in a single OCA gene. We demonstrate the utility of multiple detection methods to reveal mutations missed by Sanger sequencing. Finally, we review the TYR p.R402Q temperature sensitive variant and confirm its association with cases of albinism with only one identifiable TYR mutation. PMID:23504663
DNA variations in oculocutaneous albinism: an updated mutation list and current outstanding issues in molecular diagnostics.
Simeonov, Dimitre R; Wang, Xinjing; Wang, Chen; Sergeev, Yuri; Dolinska, Monika; Bower, Matthew; Fischer, Roxanne; Winer, David; Dubrovsky, Genia; Balog, Joan Z; Huizing, Marjan; Hart, Rachel; Zein, Wadih M; Gahl, William A; Brooks, Brian P; Adams, David R
Oculocutaneous albinism (OCA) is a rare genetic disorder of melanin synthesis that results in hypopigmented hair, skin, and eyes. There are four types of OCA caused by mutations in TYR (OCA-1), OCA2 (OCA-2), TYRP1 (OCA-3), or SLC45A2 (OCA-4). Here we report 22 novel mutations in the OCA genes; 14 from a cohort of 61 patients seen as part of the NIH OCA Natural History Study and eight from a prior study at the University of Minnesota. We also include a comprehensive list of almost 600 previously reported OCA mutations along with ethnicity information, carrier frequencies, and in silico pathogenicity predictions as a supplement. In addition to discussing the clinical and molecular features of OCA, we address the cases of apparent missing heritability. In our cohort, 26% of patients did not have two mutations in a single OCA gene. We demonstrate the utility of multiple detection methods to reveal mutations missed by Sanger sequencing. Finally, we review the TYR p.R402Q temperature-sensitive variant and confirm its association with cases of albinism with only one identifiable TYR mutation. PMID:23504663
Diagnostics of the molecular component of photon-dominated regions with mechanical heating
NASA Astrophysics Data System (ADS)
Kazandjian, M. V.; Meijerink, R.; Pelupessy, I.; Israel, F. P.; Spaans, M.
Context. Multitransition CO observations of galaxy centers have revealed that significant fractions of the dense circumnuclear gas have high kinetic temperatures, which are hard to explain by pure photon excitation, but may be caused by dissipation of turbulent energy. Aims: We aim to determine to what extent mechanical heating should be taken into account while modeling PDRs. To this end, the effect of dissipated turbulence on the thermal and chemical properties of PDRs is explored. Methods: Clouds are modeled as 1D semi-infinite slabs whose thermal and chemical equilibrium is solved for using the Leiden PDR-XDR code, where mechanical heating is added as a constant term throughout the cloud. An extensive parameter space in hydrogen gas density, FUV radiation field and mechanical heating rate is considered, covering almost all possible cases for the ISM relevant to the conditions that are encountered in galaxies. Effects of mechanical heating on the temperature profiles, column densities of CO and H2O and column density ratios of HNC, HCN and HCO+ are discussed. Results: In a steady-state treatment, mechanical heating seems to play an important role in determining the kinetic temperature of the gas in molecular clouds. Particularly in high-energy environments such as starburst galaxies and galaxy centers, model gas temperatures are underestimated by at least a factor of two if mechanical heating is ignored. The models also show that CO, HCN and H2O column densities increase as a function of mechanical heating. The HNC/HCN integrated column density ratio shows a decrease by a factor of at least two in high density regions with n ~ 105 cm-3, whereas that of HCN/HCO+ shows a strong dependence on mechanical heating for this same density range, with boosts of up to three orders of magnitude. Conclusions: The effects of mechanical heating cannot be ignored in studies of the molecular gas excitation whenever the ratio of the star formation rate to the gas density (SFR/n3
Methods for Subtyping and Molecular Comparison of Human Viral Genomes
Arens, Max
The development over the past two decades of molecular methods for manipulation of RNA and DNA has afforded molecular virologists the ability to study viral genomes in detail that has heretofore not been possible. There are many molecular techniques now available for typing and subtyping of viruses. The available methods include restriction fragment length polymorphism analysis, Southern blot analysis, oligonucleotide fingerprint analysis, reverse hybridization, DNA enzyme immunoassay, RNase protection analysis, single-strand conformation polymorphism analysis, heteroduplex mobility assay, nucleotide sequencing, and genome segment length polymorphism analysis. The methods have certain advantages and disadvantages which should be considered in their application to specific viruses or for specific purposes. These techniques are likely to become more widely used in the future for epidemiologic studies and for investigations into the pathophysiology of virus infections. PMID:10515905
Using the Attribute Hierarchy Method to Make Diagnostic Inferences about Examinees' Knowledge and Skills in Mathematics: An Operational Implementation of Cognitive Diagnostic Assessment
ERIC Educational Resources Information Center
Gierl, Mark J.; Alves, Cecilia; Majeau, Renate Taylor
The purpose of this study is to apply the attribute hierarchy method in an operational diagnostic mathematics program at Grades 3 and 6 to promote cognitive inferences about students' problem-solving skills. The attribute hierarchy method is a psychometric procedure for classifying examinees' test item responses into a set of structured attribute…
Advances in molecular imaging: targeted optical contrast agents for cancer diagnostics
Hellebust, Anne; Richards-Kortum, Rebecca
Over the last three decades, our understanding of the molecular changes associated with cancer development and progression has advanced greatly. This has led to new cancer therapeutics targeted against specific molecular pathways; such therapies show great promise to reduce mortality, in part by enabling physicians to tailor therapy for patients based on a molecular profile of their tumor. Unfortunately, the tools for definitive cancer diagnosis – light microscopic examination of biopsied tissue stained with nonspecific dyes – remain focused on the analysis of tissue ex vivo. There is an important need for new clinical tools to support the molecular diagnosis of cancer. Optical molecular imaging is emerging as a technique to help meet this need. Targeted, optically active contrast agents can specifically label extra-and intracellular biomarkers of cancer. Optical images can be acquired in real time with high spatial resolution to image-specific molecular targets, while still providing morphologic context. This article reviews recent advances in optical molecular imaging, highlighting the advances in technology required to improve early cancer detection, guide selection of targeted therapy and rapidly evaluate therapeutic efficacy. PMID:22385200
Clinical and Demographic Stratification of Test Performance: A Pooled Analysis of Five Laboratory Diagnostic Methods for American Cutaneous Leishmaniasis
Boggild, Andrea K.; Ramos, Ana P.; Espinosa, Diego; Valencia, Braulio M.; Veland, Nicolas; Miranda-Verastegui, Cesar; Arevalo, Jorge; Low, Donald E.; Llanos-Cuentas, Alejandro
We evaluated performance characteristics of five diagnostic methods for cutaneous leishmaniasis. Patients who came to the Leishmania Clinic of Hospital Nacional Cayetano Heredia in Lima, Peru, were enrolled in the study. Lesion smears, culture, microculture, polymerase chain reaction (PCR), and leishmanin skin test (LST) were performed. A total of 145 patients with 202 lesions were enrolled: 114 patients with 161 lesions fulfilled criteria for cutaneous leishmaniasis. Sensitivity and specificity were 57.8% (95% confidence interval [CI] = 50.2–65.4%) and 100.0% for culture, 78.3% (95% CI = 71.9–84.7%) and 100.0% for microculture, 71.4% (95% CI = 64.4–78.4%) and 100.0% for smears, 78.2% (95% CI = 70.6–85.8%) and 77.4% (95% CI = 62.7–92.1%) for LST, and 96.9% (95% CI = 94.2–99.6%) and 65.9% (95% CI = 51.4–80.4%) for PCR. PCR was more sensitive than the other assays (P < 0.001). Sensitivities of culture, smears, and LST varied by lesion duration and appearance. PCR offers performance advantages over other assays, irrespective of patient age, sex, lesion duration, or appearance. That clinical factors influence performance of non-molecular assays offers clinicians a patient-focused approach to diagnostic test selection. PMID:20682880
Nanomedicine: nanoparticles, molecular biosensors, and targeted gene/drug delivery for combined single-cell diagnostics and therapeutics
NASA Astrophysics Data System (ADS)
Prow, Tarl W.; Salazar, Jose H.; Rose, William A.; Smith, Jacob N.; Reece, Lisa; Fontenot, Andrea A.; Wang, Nan A.; Lloyd, R. Stephen; Leary, James F.
Next generation nanomedicine technologies are being developed to provide for continuous and linked molecular diagnostics and therapeutics. Research is being performed to develop "sentinel nanoparticles" which will seek out diseased (e.g. cancerous) cells, enter those living cells, and either perform repairs or induce those cells to die through apoptosis. These nanoparticles are envisioned as multifunctional "smart drug delivery systems". The nanosystems are being developed as multilayered nanoparticles (nanocrystals, nanocapsules) containing cell targeting molecules, intracellular re-targeting molecules, molecular biosensor molecules, and drugs/enzymes/gene therapy. These "nanomedicine systems" are being constructed to be autonomous, much like present-day vaccines, but will have sophisticated targeting, sensing, and feedback control systems-much more sophisticated than conventional antibody-based therapies. The fundamental concept of nanomedicine is to not to just kill all aberrant cells by surgery, radiation therapy, or chemotherapy. Rather it is to fix cells, when appropriate, one cell-at-a-time, to preserve and re-build organ systems. When cells should not be fixed, such as in cases where an improperly repaired cell might give rise to cancer cells, the nanomedical therapy would be to induce apoptosis in those cells to eliminate them without the damagin bystander effects of the inflammatory immune response system reacting to necrotic cells or those which have died from trauma or injury. The ultimate aim of nanomedicine is to combine diagnostics and therapeutics into "real-time medicine", using where possible in-vivo cytometry techniques for diagnostics and therapeutics. A number of individual components of these multi-component nanoparticles are already working in in-vitro and ex-vivo cell and tissue systems. Work has begun on construction of integrated nanomedical systems.
Efficient Molecular Dynamics Simulations of Multiple Radical Center Systems Based on the Fragment Molecular Orbital Method
Nakata, Hiroya; Schmidt, Michael W; Fedorov, Dmitri G; Kitaura, Kazuo; Nakamura, Shinichiro; Gordon, Mark S
The fully analytic energy gradient has been developed and implemented for the restricted open-shell Hartree–Fock (ROHF) method based on the fragment molecular orbital (FMO) theory for systems that have multiple open-shell molecules. The accuracy of the analytic ROHF energy gradient is compared with the corresponding numerical gradient, illustrating the accuracy of the analytic gradient. The ROHF analytic gradient is used to perform molecular dynamics simulations of an unusual open-shell system, liquid oxygen, and mixtures of oxygen and nitrogen. These molecular dynamics simulations provide some insight about how triplet oxygen molecules interact with each other. Timings reveal that the method can calculate the energy gradient for a system containing 4000 atoms in only 6 h. Therefore, it is concluded that the FMO-ROHF method will be useful for investigating systems with multiple open shells.
Disseminated histoplasmosis and AIDS: clinical aspects and diagnostic methods for early detection.
Corti, M E; Cendoya, C A; Soto, I; Esquivel, P; Trione, N; Villafañe, M F; Corbera, K M; Helou, S; Negroni, R
Disseminated histoplasmosis in AIDS patients is the focus of this paper. Cutaneous lesions are reported as a frequent clinical sign. Bone marrow aspiration and biopsy, blood cultures (lysis-centrifugation technique), bronchoalveolar lavage, and skin lesion scrapings are the most effective diagnostic methods. The identification of a specific antigen in blood and urine may be a rapid means of evaluation and follow-up of patients with this disease. PMID:10763544
Electromagnetism, Optics and Lasers: Handbook of Coherent Domain Optical Methods, Biomedical Diagnostics, Environment and Material Science
NASA Astrophysics Data System (ADS)
Tuchin, Valery V.
For the first time in one set of books, coherent-domain optical methods are discussed in the framework of various applications, which are characterized by a strong light scattering. A few chapters describe basic research c ontaining the updated results on coherent and polarized light non-destructive interactions with a scattering medium, in particular, diffraction, interference, and speckle formation at multiple scattering. These chapters allow for understanding coherent-domain diagnostic techniques presented in later chapters.
Microfluidic purification and concentration of malignant pleural effusions for improved molecular and cytomorphological diagnostics.
Che, James; Mach, Albert J; Go, Derek E; Talati, Ish; Ying, Yong; Rao, Jianyu; Kulkarni, Rajan P; Di Carlo, Dino
Evaluation of pleural fluids for metastatic cells is a key component of diagnostic cytopathology. However, a large background of smaller leukocytes and/or erythrocytes can make accurate diagnosis difficult and reduce specificity in identification of mutations of interest for targeted anti-cancer therapies. Here, we describe an automated microfluidic system (Centrifuge Chip) which employs microscale vortices for the size-based isolation and concentration of cancer cells and mesothelial cells from a background of blood cells. We are able to process non-diluted pleural fluids at 6 mL/min and enrich target cells significantly over the background; we achieved improved purity in all patient samples analyzed. The resulting isolated and viable cells are readily available for immunostaining, cytological analysis, and detection of gene mutations. To demonstrate the utility towards aiding companion diagnostics, we also show improved detection accuracy of KRAS gene mutations in lung cancer cells processed using the Centrifuge Chip, leading to an increase in the area under the curve (AUC) of the receiver operating characteristic from 0.90 to 0.99. The Centrifuge Chip allows for rapid concentration and processing of large volumes of bodily fluid samples for improved cytological diagnosis and purification of cells of interest for genetic testing, which will be helpful for enhancing diagnostic accuracy. PMID:24205153
Adaptation of LASCA method for diagnostics of malignant tumours in laboratory animals
Ul'yanov, S S; Laskavyi, V N; Glova, Alina B; Polyanina, T I; Ul'yanova, O V; Fedorova, V A; Ul'yanov, A S
The LASCA method is adapted for diagnostics of malignant neoplasms in laboratory animals. Tumours are studied in mice of Balb/c inbred line after inoculation of cells of syngeneic myeloma cell line Sp.2/0 Ag.8. The appropriateness of using the tLASCA method in tumour investigations is substantiated; its advantages in comparison with the sLASCA method are demonstrated. It is found that the most informative characteristic, indicating the presence of a tumour, is the fractal dimension of LASCA images.
Adaptation of LASCA method for diagnostics of malignant tumours in laboratory animals
NASA Astrophysics Data System (ADS)
Ul'yanov, S. S.; Laskavyi, V. N.; Glova, Alina B.; Polyanina, T. I.; Ul'yanova, O. V.; Fedorova, V. A.; Ul'yanov, A. S.
The LASCA method is adapted for diagnostics of malignant neoplasms in laboratory animals. Tumours are studied in mice of Balb/c inbred line after inoculation of cells of syngeneic myeloma cell line Sp.2/0 — Ag.8. The appropriateness of using the tLASCA method in tumour investigations is substantiated; its advantages in comparison with the sLASCA method are demonstrated. It is found that the most informative characteristic, indicating the presence of a tumour, is the fractal dimension of LASCA images.
Evaluation of molecular and culture methods to determine the optimum testing strategy for verotoxigenic Escherichia coli in faecal specimens.
Carroll, Anne M; Cobban, Emma; McNamara, Eleanor B
We evaluated 5 methods for the detection of verotoxogenic Escherichia coli (VTEC) from faecal specimens to determine the most sensitive and specific method(s) and to advise an optimum testing strategy. A total of 681 stool specimens were examined using up to 5 diagnostic molecular and phenotypic methods that are used routinely in the VTEC Reference laboratory, Dublin. A testing strategy incorporating a 2-step approach that included a single Real Time-PCR and 1 culture-based method yielded the highest sensitivity, specificity, positive predictive value, and negative predictive value of 98.21%, 100%, 100%, and 99.43%, respectively. PMID:26830271
[Value of molecular biology methods for diagnosis in bacteriology].
Piémont, Y; Jaulhac, B
Progress in molecular biology has led to the development of new tools for bacteriological diagnosis. Sporadic genes coding for virulence factors can be detected with highly specific genetic probes applied to cultured bacteria. Such genetic probes can also be used to specifically identified cultured bacteria whose general taxonomic classification is known. Another advantage of molecular genetics is the possibility that the cell culture step may not be needed, bacteria being identified directly in the sample specimen. Such techniques are particularly interesting to identify bacteria which are difficult to culture (for example: Borrelia burgdorferi, Chlamydia trachomatis) or which grow slowly (mycobacteria). The bacterial DNA must be isolated and amplified with an enzyme reaction. This is a critical step in the method: several positive and negative controls are required. When performed under optimal conditions, amplification techniques are excellent methods which can offer results similar to culture methods in culturable bacteria. Finally, molecular biology can be used to identify previously cultured bacteria for which there is no taxonomic orientation. Here the ribosome 165 DNA must be amplified and sequenced. The sequence is then compared with a data bank allowing classification. One could image future techniques applied to certain pathology samples for the detection and identification of bacteria without need for a culture step. However, direct microscope examination and bacterial culture remain the basic methods for bacteriologic diagnosis, the advantages and disadvantages of molecular biology leading to its use a complementary method for improving the quality of the diagnosis. PMID:8526414
Molecular characterization of porphyrias in Italy: a diagnostic flow-chart.
Martinez di Montemuros, F; Di Pierro, E; Patti, E; Tavazzi, D; Danielli, M G; Biolcati, G; Rocchi, E; Cappellini, M D
The porphyrias are disorders associated with inherited or acquired enzyme deficiencies in the heme biosynthetic pathway. The differential diagnosis is often difficult since the phenotype is very similar in some forms and the biochemical tests are not commonly available. Here we provide an update on the molecular diagnosis of porphyrias in Italy and a flow-chart to facilitate the identification of mutations in heme biosynthetic genes. The molecular analysis has allowed us to identify the molecular defect underlying the disease in 66 probands with different porphyrias [acute intermittent porphyria (AIP), variegate porphyria (VP), porphyria cutanea tarda (PCT), erythropoietic protoporphyria (EPP)]. No Italian patients with defects in coproporphyrinogen oxidise (CPOX) gene, responsible for hereditary coproporphyria (HCP), have been detected. The molecular characterization has been extended to 115 relatives with the identification of 55 asymptomatic mutation carriers and 60 normal subjects. We have so far identified 50 different mutations among 4 genes associated with the most common porphyrias showing a high molecular heterogeneity: 22 in the hydroxymethylbilane synthase (HMBS) gene (AIP), 7 in the protoporphyrinogen oxidase (PPOX) gene (VP), 16 in the uroporphyrinogen decarboxylase (UROD) gene (PCT) and 5 in the ferrochelatase (FECH) gene (EPP). Among the 50 molecular defects, 29 seem to be restricted to the Italian population. PMID:12699245
Bayesian and maximum entropy methods for fusion diagnostic measurements with compact neutron spectrometers
NASA Astrophysics Data System (ADS)
Reginatto, Marcel; Zimbal, Andreas
In applications of neutron spectrometry to fusion diagnostics, it is advantageous to use methods of data analysis which can extract information from the spectrum that is directly related to the parameters of interest that describe the plasma. We present here methods of data analysis which were developed with this goal in mind, and which were applied to spectrometric measurements made with an organic liquid scintillation detector (type NE213). In our approach, we combine Bayesian parameter estimation methods and unfolding methods based on the maximum entropy principle. This two-step method allows us to optimize the analysis of the data depending on the type of information that we want to extract from the measurements. To illustrate these methods, we analyze neutron measurements made at the PTB accelerator under controlled conditions, using accelerator-produced neutron beams. Although the methods have been chosen with a specific application in mind, they are general enough to be useful for many other types of measurements.
Molecular methods for microbiological quality control of meat and meat products: a review.
Gokulakrishnan, P; Vergis, Jess
Achieving food safety is a global health goal and the food-borne diseases take a major check on global health. Therefore, detection of microbial pathogens in food is the solution to the prevention and recognition of problems related to health and safety. Conventional and standard bacterial detection methods such as culture and colony counting methods and immunology-based methods may take up to several hours or even a few days to yield a result. Obviously, this is inadequate, and recently many researchers are focusing towards the progress of rapid diagnostic methods. The advent of molecular techniques has led to the development of a diverse array of assay for quality control of meat and meat products. Rapid analysis using DNA hybridization and amplification techniques offer more sensitivity and specificity to get results than culture based methods as well as dramatic reduction in the time to get results. Many methods have also achieved the high level automation, facilitating their application as routine sample screening assays. This review is intended to provide an overview of the molecular methods for microbiological quality control of meat and meat products. PMID:24915322
A large number of human enteric viruses are known to cause gastrointestinal illness and waterborne outbreaks. Many of these are emerging viruses that do not grow or grow poorly in cell culture and so molecular detectoin methods based on the polymerase chain reaction (PCR) are be...
Fecal contamination is widespread in the waterways of the United States. Both to correct the problem, and to estimate public health risk, it is necessary to identify the source of the contamination. Several culture-independent molecular methods for fecal source identification hav...
The Henry Ford Production System: LEAN Process Redesign Improves Service in the Molecular Diagnostic Laboratory
Cankovic, Milena; Varney, Ruan C.; Whiteley, Lisa; Brown, Ron; D'Angelo, Rita; Chitale, Dhananjay; Zarbo, Richard J.
Accurate and timely molecular test results play an important role in patient management; consequently, there is a customer expectation of short testing turnaround times. Baseline data analysis revealed that the greatest challenge to timely result generation occurred in the preanalytic phase of specimen collection and transport. Here, we describe our efforts to improve molecular testing turnaround times by focusing primarily on redesign of preanalytic processes using the principles of LEAN production. Our goal was to complete greater than 90% of the molecular tests in less than 3 days. The project required cooperation from different laboratory disciplines as well as individuals outside of the laboratory. The redesigned processes involved defining and standardizing the protocols and approaching blood and tissue specimens as analytes for molecular testing. The LEAN process resulted in fewer steps, approaching the ideal of a one-piece flow for specimens through collection/retrieval, transport, and different aspects of the testing process. The outcome of introducing the LEAN process has been a 44% reduction in molecular test turnaround time for tissue specimens, from an average of 2.7 to 1.5 days. In addition, extending LEAN work principles to the clinician suppliers has resulted in a markedly increased number of properly collected and shipped blood specimens (from 50 to 87%). These continuous quality improvements were accomplished by empowered workers in a blame-free environment and are now being sustained with minimal management involvement. PMID:19661386
The role of ultrasound and nuclear medicine methods in the preoperative diagnostics of primary hyperparathyroidism
Cacko, Marek; Królicki, Leszek
Primary hyperparathyroidism (PH) represents one of the most common endocrine diseases. In most cases, the disorder is caused by parathyroid adenomas. Bilateral neck exploration has been a widely used treatment method for adenomas since the 20's of the twentieth century. In the last decade, however, it has been increasingly replaced by a minimally invasive surgical treatment. Smaller extent, shorter duration and lower complication rate of such a procedure are emphasized. Its efficacy depends on a precise location of parathyroid tissue during the preoperative imaging. Scintigraphy and ultrasound play a major role in the diagnostic algorithms. The efficacy of both methods has been repeatedly verified and compared. The still-current guidelines of the European Association of Nuclear Medicine (2009) emphasize the complementary role of scintigraphy and ultrasonography in the preoperative diagnostics in patients with primary hyperparathyroidism. At the same time, attempts are made to improve both these techniques by implementing new study protocols or innovative technologies. Publications have emerged in the recent years in the field of ultrasonography, whose authors pointed out the usefulness of elastography and contrast media. Nuclear medicine studies, on the other hand, focus mainly on the assessment of new radiotracers used in the positron emission tomography (PET). The aim of this article is to present, based on literature data, the possibilities of ultrasound and scintigraphy in the preoperative diagnostics in patients with primary hyperparathyroidism. Furthermore, the main directions in the development of imaging techniques in PH patients were evaluated. PMID:26807297
The role of ultrasound and nuclear medicine methods in the preoperative diagnostics of primary hyperparathyroidism.
Nieciecki, Michał; Cacko, Marek; Królicki, Leszek
Primary hyperparathyroidism (PH) represents one of the most common endocrine diseases. In most cases, the disorder is caused by parathyroid adenomas. Bilateral neck exploration has been a widely used treatment method for adenomas since the 20's of the twentieth century. In the last decade, however, it has been increasingly replaced by a minimally invasive surgical treatment. Smaller extent, shorter duration and lower complication rate of such a procedure are emphasized. Its efficacy depends on a precise location of parathyroid tissue during the preoperative imaging. Scintigraphy and ultrasound play a major role in the diagnostic algorithms. The efficacy of both methods has been repeatedly verified and compared. The still-current guidelines of the European Association of Nuclear Medicine (2009) emphasize the complementary role of scintigraphy and ultrasonography in the preoperative diagnostics in patients with primary hyperparathyroidism. At the same time, attempts are made to improve both these techniques by implementing new study protocols or innovative technologies. Publications have emerged in the recent years in the field of ultrasonography, whose authors pointed out the usefulness of elastography and contrast media. Nuclear medicine studies, on the other hand, focus mainly on the assessment of new radiotracers used in the positron emission tomography (PET). The aim of this article is to present, based on literature data, the possibilities of ultrasound and scintigraphy in the preoperative diagnostics in patients with primary hyperparathyroidism. Furthermore, the main directions in the development of imaging techniques in PH patients were evaluated. PMID:26807297
Lab-on-a-CD: A Fully Integrated Molecular Diagnostic System.
Kong, Ling X; Perebikovsky, Alexandra; Moebius, Jacob; Kulinsky, Lawrence; Madou, Marc
The field of centrifugal microfluidics has experienced tremendous growth during the past 15 years, especially in applications such as lab-on-a-disc (LoD) diagnostics. The strength of LoD systems lies in its potential for development into fully integrated sample-to-answer analysis systems. This review highlights the technologies necessary to develop the next generation of these systems. In addition to outlining valving and other fluid-handling operations, we discuss the recent advances and future outlook in four categories of LoD processes: reagent storage, sample preparation, nucleic acid amplification, and analyte detection strategies. PMID:26082453
Novel molecular diagnostic tools for malaria elimination: a review of options from the point of view of high-throughput and applicability in resource limited settings.
Britton, Sumudu; Cheng, Qin; McCarthy, James S
As malaria transmission continues to decrease, an increasing number of countries will enter pre-elimination and elimination. To interrupt transmission, changes in control strategies are likely to require more accurate identification of all carriers of Plasmodium parasites, both symptomatic and asymptomatic, using diagnostic tools that are highly sensitive, high throughput and with fast turnaround times preferably performed in local health service settings. Currently available immunochromatographic lateral flow rapid diagnostic tests and field microscopy are unlikely to consistently detect infections at parasite densities less than 100 parasites/µL making them insufficiently sensitive for detecting all carriers. Molecular diagnostic platforms, such as PCR and LAMP, are currently available in reference laboratories, but at a cost both financially and in turnaround time. This review describes the recent progress in developing molecular diagnostic tools in terms of their capacity for high throughput and potential for performance in non-reference laboratories for malaria elimination. PMID:26879936
[Forensic medical diagnostics of intra-vitality of the strangulation mark by morphological methods].
Bogomolov, D V; Zbrueva, Yu V; Putintsev, V A; Denisova, O P
The objective of the present study WaS to overview the current domestic and foreign literature concerning the up-to-date methods employed for the expert evaluation of intra-vitality of the strangulation mark. The secondary objective was to propose the new approaches for addressing this problem. The methods of expert diagnostics with a view to determining the time of infliction of injuries as exemplified by mechanical asphyxia are discussed. It is concluded that immunohistochemical and morphometric studies provide the most promising tools for the evaluation of intra-vitality of the strangulation mark for the purpose of forensic medical expertise. PMID:27358932
Method of assembly of molecular-sized nets and scaffolding
Michl, Josef; Magnera, Thomas F.; David, Donald E.; Harrison, Robin M.
The present invention relates to methods and starting materials for forming molecular-sized grids or nets, or other structures based on such grids and nets, by creating molecular links between elementary molecular modules constrained to move in only two directions on an interface or surface by adhesion or bonding to that interface or surface. In the methods of this invention, monomers are employed as the building blocks of grids and more complex structures. Monomers are introduced onto and allowed to adhere or bond to an interface. The connector groups of adjacent adhered monomers are then polymerized with each other to form a regular grid in two dimensions above the interface. Modules that are not bound or adhered to the interface are removed prior to reaction of the connector groups to avoid undesired three-dimensional cross-linking and the formation of non-grid structures. Grids formed by the methods of this invention are useful in a variety of applications, including among others, for separations technology, as masks for forming regular surface structures (i.e., metal deposition) and as templates for three-dimensional molecular-sized structures.
Method of assembly of molecular-sized nets and scaffolding
Michl, J.; Magnera, T.F.; David, D.E.; Harrison, R.M.
The present invention relates to methods and starting materials for forming molecular-sized grids or nets, or other structures based on such grids and nets, by creating molecular links between elementary molecular modules constrained to move in only two directions on an interface or surface by adhesion or bonding to that interface or surface. In the methods of this invention, monomers are employed as the building blocks of grids and more complex structures. Monomers are introduced onto and allowed to adhere or bond to an interface. The connector groups of adjacent adhered monomers are then polymerized with each other to form a regular grid in two dimensions above the interface. Modules that are not bound or adhered to the interface are removed prior to reaction of the connector groups to avoid undesired three-dimensional cross-linking and the formation of non-grid structures. Grids formed by the methods of this invention are useful in a variety of applications, including among others, for separations technology, as masks for forming regular surface structures (i.e., metal deposition) and as templates for three-dimensional molecular-sized structures. 9 figs.
Experimental methods of molecular matter-wave optics.
Juffmann, Thomas; Ulbricht, Hendrik; Arndt, Markus
We describe the state of the art in preparing, manipulating and detecting coherent molecular matter. We focus on experimental methods for handling the quantum motion of compound systems from diatomic molecules to clusters or biomolecules.Molecular quantum optics offers many challenges and innovative prospects: already the combination of two atoms into one molecule takes several well-established methods from atomic physics, such as for instance laser cooling, to their limits. The enormous internal complexity that arises when hundreds or thousands of atoms are bound in a single organic molecule, cluster or nanocrystal provides a richness that can only be tackled by combining methods from atomic physics, chemistry, cluster physics, nanotechnology and the life sciences.We review various molecular beam sources and their suitability for matter-wave experiments. We discuss numerous molecular detection schemes and give an overview over diffraction and interference experiments that have already been performed with molecules or clusters.Applications of de Broglie studies with composite systems range from fundamental tests of physics up to quantum-enhanced metrology in physical chemistry, biophysics and the surface sciences.Nanoparticle quantum optics is a growing field, which will intrigue researchers still for many years to come. This review can, therefore, only be a snapshot of a very dynamical process. PMID:23907707
Translating biomedical science into clinical practice: Molecular diagnostics and the determination of malignancy.
Cox, Helen; Webster, Andrew
The identification of new biomarkers that help understand the epidemiological basis of patterns of malignancy at a population level is reshaping conceptions of health, disease and normality. These developments create new challenges for clinicians and the ways in which they work with scientists and engage with patients. Bioclinical collectives, an assemblage of laboratory and clinical evidence and practice, comprise different expert groups of scientists and clinicians who typically enact their expertise through boundary work to establish some degree of jurisdictional authority over their practice. Serra (2010) has argued for the existence of 'medical technocracies' wherein each speciality involved defines the boundaries between themselves in daily medical practices and use technology as a resource to construct their particular strategies. In this article we explore these two aspects of biomedical expertise - the collective and the boundaried domains of diagnostic practice (especially in regard to clinical utility) - to understand how haematological malignancy and disease are perceived and managed. The empirical data for the article are based on extensive primary research in hospitals based in the north of the UK, and among clinicians and laboratory scientists working in haematological malignancies. Our chosen field of inquiry - a haematological malignancy diagnostic service in the UK - is a particularly rich site through which to explore these twin aspects. PMID:23074299
Adoption of Lean Principles in a High-Volume Molecular Diagnostic Microbiology Laboratory
Mitchell, P. Shawn; Mandrekar, Jayawant N.
Clinical laboratories are constantly facing challenges to do more with less, enhance quality, improve test turnaround time, and reduce operational expenses. Experience with adopting and applying lean concepts and tools used extensively in the manufacturing industry is described for a high-volume clinical molecular microbiology laboratory, illustrating how operational success and benefits can be achieved. PMID:24829247
Adoption of lean principles in a high-volume molecular diagnostic microbiology laboratory.
Mitchell, P Shawn; Mandrekar, Jayawant N; Yao, Joseph D C
Clinical laboratories are constantly facing challenges to do more with less, enhance quality, improve test turnaround time, and reduce operational expenses. Experience with adopting and applying lean concepts and tools used extensively in the manufacturing industry is described for a high-volume clinical molecular microbiology laboratory, illustrating how operational success and benefits can be achieved. PMID:24829247
New insights into molecular diagnostic pathology of primary liver cancer: Advances and challenges.
Cong, Wen-Ming; Wu, Meng-Chao
Primary liver cancer (PLC) is one of the most common malignancies worldwide with increasing incidence and accounts for the third leading cause of cancer-related mortality. Traditional morphopathology primarily emphasizes qualitative diagnosis of PLC, which is not sufficient to resolve the major concern of increasing the long-term treatment efficacy of PLC in clinical management for the modern era. Since the beginning of the 21st century, molecular pathology has played an active role in the investigation of the evaluation of the metastatic potential of PLC, detection of drug targets, prediction of recurrence risks, analysis of clonal origins, evaluation of the malignancy trend of precancerous lesions, and determination of clinical prognosis. As a result, many new progresses have been obtained, and new strategies of molecular-pathological diagnosis have been formed. Moreover, the new types of pathobiological diagnosis indicator systems for PLC have been preliminarily established. These achievements provide valuable molecular pathology-based guide for clinical formulation of individualized therapy programs for PLC. This review article briefly summarizes some relevant progresses of molecular-pathological diagnosis of PLC from the perspective of clinical translational application other than basic experimental studies. PMID:26276723
Nonholonomic Hamiltonian Method for Molecular Dynamics Simulations of Reacting Shocks
NASA Astrophysics Data System (ADS)
Fahrenthold, Eric; Bass, Joseph
Conventional molecular dynamics simulations of reacting shocks employ a holonomic Hamiltonian formulation: the breaking and forming of covalent bonds is described by potential functions. In general these potential functions: (a) are algebraically complex, (b) must satisfy strict smoothness requirements, and (c) contain many fitted parameters. In recent research the authors have developed a new noholonomic formulation of reacting molecular dynamics. In this formulation bond orders are determined by rate equations and the bonding-debonding process need not be described by differentiable functions. This simplifies the representation of complex chemistry and reduces the number of fitted model parameters. Example applications of the method show molecular level shock to detonation simulations in nitromethane and RDX. Research supported by the Defense Threat Reduction Agency.
Inversion methods for the measurements of MHD-like density fluctuations by Heavy Ion Beam Diagnostic
NASA Astrophysics Data System (ADS)
Malaquias, A.; Henriques, R. B.; Nedzelsky, I. S.
We report here on the recent developments in the deconvolution of the path integral effects for the study of MHD pressure-like fluctuations measured by Heavy Ion Beam Diagnostic. In particular, we develop improved methods to account for and remove the path integral effect on the determination of the ionization generation factors, including the double ionization of the primary beam. We test the method using the HIBD simulation code which computes the real beam trajectories and attenuations due to electron impact ionization for any selected synthetic profiles of plasma current, plasma potential, electron temperature and density. Simulations have shown the numerical method to be highly effective in ISTTOK within an overall accuracy of a few percent (< 3%). The method here presented can effectively reduce the path integral effects and may serve as the basis to develop improved retrieving techniques for plasma devices working even in higher density ranges. The method is applied to retrieve the time evolution and spatial structure of m=1 and m=2 modes. The 2D MHD mode-like structure is reconstructed by means of a spatial projection of all 1D measurements obtained during one full rotation of the mode. A shorter version of this contribution is due to be published in PoS at: 1st EPS conference on Plasma Diagnostics
Personality Assessment in the Diagnostic Manuals: On Mindfulness, Multiple Methods, and Test Score Discontinuities
Bornstein, Robert F.
Recent controversies have illuminated the strengths and limitations of different frameworks for conceptualizing personality pathology (e.g., trait perspectives, categorical models), and stimulated debate regarding how best to diagnose personality disorders (PDs) in DSM-5, and in other diagnostic systems (i.e., the International Classification of Diseases, the Psychodynamic Diagnostic Manual). In this article I argue that regardless of how PDs are conceptualized and which diagnostic system is employed, multi-method assessment must play a central role in PD diagnosis. By complementing self-reports with evidence from other domains (e.g., performance-based tests), a broader range of psychological processes are engaged in the patient, and the impact of self-perception and self-presentation biases may be better understood. By providing the assessor with evidence drawn from multiple modalities, some of which provide converging patterns and some of which yield divergent results, the assessor is compelled to engage this evidence more deeply. The mindful processing that ensues can help minimize the deleterious impact of naturally occurring information processing bias and distortion on the part of the clinician (e.g., heuristics, attribution errors), bringing greater clarity to the synthesis and integration of assessment data. PMID:25856565
Molecular Diagnostics for Lassa Fever at Irrua Specialist Teaching Hospital, Nigeria: Lessons Learnt from Two Years of Laboratory Operation
Hass, Meike; Gabriel, Martin; Ölschläger, Stephan; Becker-Ziaja, Beate; Folarin, Onikepe; Phelan, Eric; Ehiane, Philomena E.; Ifeh, Veritas E.; Uyigue, Eghosasere A.; Oladapo, Yemisi T.; Muoebonam, Ekene B.; Osunde, Osagie; Dongo, Andrew; Okokhere, Peter O.; Okogbenin, Sylvanus A.; Momoh, Mojeed; Alikah, Sylvester O.; Akhuemokhan, Odigie C.; Imomeh, Peter; Odike, Maxy A. C.; Gire, Stephen; Andersen, Kristian; Sabeti, Pardis C.; Happi, Christian T.; Akpede, George O.; Günther, Stephan
Background Lassa fever is a viral hemorrhagic fever endemic in West Africa. However, none of the hospitals in the endemic areas of Nigeria has the capacity to perform Lassa virus diagnostics. Case identification and management solely relies on non-specific clinical criteria. The Irrua Specialist Teaching Hospital (ISTH) in the central senatorial district of Edo State struggled with this challenge for many years. Methodology/Principal Findings A laboratory for molecular diagnosis of Lassa fever, complying with basic standards of diagnostic PCR facilities, was established at ISTH in 2008. During 2009 through 2010, samples of 1,650 suspected cases were processed, of which 198 (12%) tested positive by Lassa virus RT-PCR. No remarkable demographic differences were observed between PCR-positive and negative patients. The case fatality rate for Lassa fever was 31%. Nearly two thirds of confirmed cases attended the emergency departments of ISTH. The time window for therapeutic intervention was extremely short, as 50% of the fatal cases died within 2 days of hospitalization—often before ribavirin treatment could be commenced. Fatal Lassa fever cases were older (p = 0.005), had lower body temperature (p<0.0001), and had higher creatinine (p<0.0001) and blood urea levels (p<0.0001) than survivors. Lassa fever incidence in the hospital followed a seasonal pattern with a peak between November and March. Lassa virus sequences obtained from the patients originating from Edo State formed—within lineage II—a separate clade that could be further subdivided into three clusters. Conclusions/Significance Lassa fever case management was improved at a tertiary health institution in Nigeria through establishment of a laboratory for routine diagnostics of Lassa virus. Data collected in two years of operation demonstrate that Lassa fever is a serious public health problem in Edo State and reveal new insights into the disease in hospitalized patients. PMID:23029594
The Long and Short of Circulating Cell-Free DNA and the Ins and Outs of Molecular Diagnostics.
Jiang, Peiyong; Lo, Y M Dennis
The discovery of cell-free tumor and fetal DNA molecules in the plasma of cancer patients and pregnant women, respectively, has opened up exciting opportunities in molecular diagnosis. The understanding of the biological properties of circulating cell-free DNA (cfDNA) molecules would be essential for us to make the best use of such molecules in different clinical settings. In this review we start by exploring the technologies that have been used for analyzing the size profiles of cfDNA in plasma. We then review the size profiles of cfDNA in different clinical scenarios, including cancer, pregnancy, transplantation, and autoimmune diseases. Finally, we discuss the potential diagnostic applications of plasma DNA size profiling. PMID:27129983
Evolving molecular diagnostics for familial cardiomyopathies: at the heart of it all
Callis, Thomas E; Jensen, Brian C; Weck, Karen E; Willis, Monte S
Cardiomyopathies are an important and heterogeneous group of common cardiac diseases. An increasing number of cardiomyopathies are now recognized to have familial forms, which result from single-gene mutations that render a Mendelian inheritance pattern, including hypertrophic cardiomyopathy, dilated cardiomyopathy, restrictive cardiomyopathy, arrhythmogenic right ventricular cardiomyopathy and left ventricular noncompaction cardiomyopathy. Recently, clinical genetic tests for familial cardiomyopathies have become available for clinicians evaluating and treating patients with these diseases, making it necessary to understand the current progress and challenges in cardiomyopathy genetics and diagnostics. In this review, we summarize the genetic basis of selected cardiomyopathies, describe the clinical utility of genetic testing for cardiomyopathies and outline the current challenges and emerging developments. PMID:20370590
Tsarev, V N; Nikolaeva, E N; Iagodina, E V; Trefilova, Yu A; Ippolitov, E V
The examination was carried out in the Moscow clinical infectious hospital No 2 concerning 102 patients with verified diagnosis "AIDS-infection" and seropositive according results of detection of anti-HIV-antibodies in blood serum. The study was organized to analyze rate ofcolonization of gums with virulent anaerobic bacteria in HIV-infected (polymerase chain reaction) and antibodies to HIV in gingival fluid (enzyme-linked immunosorbent assay). It is established that in HIV-infected patients, in scrape from gingival sulcus dominate anaerobic bacteria P. gigngivalis and A. ctinomycetemcomitans and in case of periodontitis--P. gingivalis and T. forsythia. The received data permits recommending the test-system "Multident-5" for polymerase chain reaction diagnostic. The reagents kit "Calypte®HIV-1/2"--for enzyme-linked immunosorbent assay gingival fluid. The results of polymerase chain reaction and enzyme-linked immunosorbent assay have no impact of concomitant stomatological (periodontitis, gingivitis) and somatic pathology. PMID:27183732
A Molecular Selection Index Method Based on Eigenanalysis
Cerón-Rojas, J. Jesús; Castillo-González, Fernando; Sahagún-Castellanos, Jaime; Santacruz-Varela, Amalio; Benítez-Riquelme, Ignacio; Crossa, José
The traditional molecular selection index (MSI) employed in marker-assisted selection maximizes the selection response by combining information on molecular markers linked to quantitative trait loci (QTL) and phenotypic values of the traits of the individuals of interest. This study proposes an MSI based on an eigenanalysis method (molecular eigen selection index method, MESIM), where the first eigenvector is used as a selection index criterion, and its elements determine the proportion of the trait's contribution to the selection index. This article develops the theoretical framework of MESIM. Simulation results show that the genotypic means and the expected selection response from MESIM for each trait are equal to or greater than those from the traditional MSI. When several traits are simultaneously selected, MESIM performs well for traits with relatively low heritability. The main advantages of MESIM over the traditional molecular selection index are that its statistical sampling properties are known and that it does not require economic weights and thus can be used in practical applications when all or some of the traits need to be improved simultaneously. PMID:18716338
A reduced basis method for molecular dynamics simulation
NASA Astrophysics Data System (ADS)
Vincent-Finley, Rachel Elisabeth
In this dissertation, we develop a method for molecular simulation based on principal component analysis (PCA) of a molecular dynamics trajectory and least squares approximation of a potential energy function. Molecular dynamics (MD) simulation is a computational tool used to study molecular systems as they evolve through time. With respect to protein dynamics, local motions, such as bond stretching, occur within femtoseconds, while rigid body and large-scale motions, occur within a range of nanoseconds to seconds. To capture motion at all levels, time steps on the order of a femtosecond are employed when solving the equations of motion and simulations must continue long enough to capture the desired large-scale motion. To date, simulations of solvated proteins on the order of nanoseconds have been reported. It is typically the case that simulations of a few nanoseconds do not provide adequate information for the study of large-scale motions. Thus, the development of techniques that allow longer simulation times can advance the study of protein function and dynamics. In this dissertation we use principal component analysis (PCA) to identify the dominant characteristics of an MD trajectory and to represent the coordinates with respect to these characteristics. We augment PCA with an updating scheme based on a reduced representation of a molecule and consider equations of motion with respect to the reduced representation. We apply our method to butane and BPTI and compare the results to standard MD simulations of these molecules. Our results indicate that the molecular activity with respect to our simulation method is analogous to that observed in the standard MD simulation with simulations on the order of picoseconds.
A Fully Integrated Paperfluidic Molecular Diagnostic Chip for the Extraction, Amplification, and Detection of Nucleic Acids from Clinical Samples
Rodriguez, Natalia M.; Wong, Winnie S.; Liu, Lena; Dewar, Rajan; Klapperich, Catherine M.
Paper diagnostics have successfully been employed to detect the presence of antigens or small molecules in clinical samples through immunoassays; however, the detection of many disease targets relies on the much higher sensitivity and specificity achieved via nucleic acid amplification tests (NAAT). The steps involved in NAAT have recently begun to be explored in paper matrices, and our group, among others, has reported on paper-based extraction, amplification, and detection of DNA and RNA targets. Here, we integrate these paper-based NAAT steps onto a single paperfluidic chip in a modular, foldable system that allows for fully integrated fluidic handling from sample to result. We showcase the functionality of the chip by combining nucleic acid isolation, isothermal amplification, and lateral flow detection of human papillomavirus (HPV) 16 DNA directly from crude cervical specimens in under 1 hour for rapid, early detection of cervical cancer. The chip is made entirely of paper and adhesive sheets, making it low-cost, portable, and disposable, and offering the potential for a point-of-care molecular diagnostic platform even in remote and resource-limited settings. PMID:26785636
Cellular physiological assessment of bivalves after chronic exposure to spilled Exxon Valdez crude oil using a novel molecular diagnostic biotechnology.
Downs, Craig A; Shigenaka, Gary; Fauth, John E; Robinson, Charles E; Huang, Arnold
The objective of this study was to determine the cellular physiological status of the bivalves Mya arenaria and Mytilus trossulus in an area experiencing a 10-yr chronic exposure of spilled Exxon Valdez crude oil in Prince William Sound. Bivalves were collected from well-characterized oiled and unoiled sites. We used a novel biotechnology (Environmental Cellular Diagnostic System) to determine (i) if bivalves were physiologically stressed, (ii) the nature of the altered physiological state, and (iii) whether the bivalves were responding to an exposure of polyaromatic hydrocarbons (PAH). Molecular diagnostic analysis indicated that bivalves at the oiled site were experiencing both oxidative and xenobiotic stress, resulting in increased protein turnover and chaperone activity. Bivalves from the impacted area were responding specifically to a PAH-xenobiotic exposure and accumulating protein-PAH adducts. Finally, species-specific responses were observed that could be related to the habitat preferences of each species. We conclude that bivalves inhabiting a site impacted by crude oil from the 1989 Exxon Valdez spill showed clear indications of cellular physiological stress. PMID:12144276
A fully integrated paperfluidic molecular diagnostic chip for the extraction, amplification, and detection of nucleic acids from clinical samples.
Rodriguez, Natalia M; Wong, Winnie S; Liu, Lena; Dewar, Rajan; Klapperich, Catherine M
Paper diagnostics have successfully been employed to detect the presence of antigens or small molecules in clinical samples through immunoassays; however, the detection of many disease targets relies on the much higher sensitivity and specificity achieved via nucleic acid amplification tests (NAAT). The steps involved in NAAT have recently begun to be explored in paper matrices, and our group, among others, has reported on paper-based extraction, amplification, and detection of DNA and RNA targets. Here, we integrate these paper-based NAAT steps into a single paperfluidic chip in a modular, foldable system that allows for fully integrated fluidic handling from sample to result. We showcase the functionality of the chip by combining nucleic acid isolation, isothermal amplification, and lateral flow detection of human papillomavirus (HPV) 16 DNA directly from crude cervical specimens in less than 1 hour for rapid, early detection of cervical cancer. The chip is made entirely of paper and adhesive sheets, making it low-cost, portable, and disposable, and offering the potential for a point-of-care molecular diagnostic platform even in remote and resource-limited settings. PMID:26785636
Risk of Misdiagnosis Due to Allele Dropout and False-Positive PCR Artifacts in Molecular Diagnostics: Analysis of 30,769 Genotypes.
Blais, Jonatan; Lavoie, Sébastien B; Giroux, Sylvie; Bussières, Johanne; Lindsay, Carmen; Dionne, Jacqueline; Laroche, Mélissa; Giguère, Yves; Rousseau, François
Quality control is a complex issue for clinical molecular diagnostic applications. In the case of genotyping assays, artifacts such as allele dropout represent a risk of misdiagnosis for amplification-based methods. However, its frequency of occurrence in PCR-based diagnostic assays remains unknown. To maximize the likelihood of detecting allele dropout, our clinical genotyping PCR-based assays are designed with two independent assays for each allele (nonoverlapping primers on each DNA strand). To estimate the incidence of allelic dropout, we took advantage of the capacity of our clinical assays to detect such events. We retrospectively studied their occurrence in the initial PCR assay for 30,769 patient reports for mutations involved in four diseases produced over 8 years. Ninety-three allele dropout events were detected and all were solved before reporting. In addition, 42 cases of artifacts caused by amplification of an allele ultimately confirmed to not be part of the genotype (drop-in events) were detected and solved. These artifacts affected 1:227 genotypes, 94% of which were due to nonreproducible PCR failures rather than sequence variants interfering with the assay, suggesting that careful primer design cannot prevent most of these errors. This provides a quantitative estimate for clinical laboratories to take this phenomenon into account in quality management and to favor assay designs that can detect (and minimize) occurrence of these artifacts in routine clinical use. PMID:26146130
Evaluation of Molecular Methods for Identification of Salmonella Serovars.
Yoshida, Catherine; Gurnik, Simone; Ahmad, Aaminah; Blimkie, Travis; Murphy, Stephanie A; Kropinski, Andrew M; Nash, John H E
Classification by serotyping is the essential first step in the characterization of Salmonella isolates and is important for surveillance, source tracking, and outbreak detection. To improve detection and reduce the burden of salmonellosis, several rapid and high-throughput molecular Salmonella serotyping methods have been developed.The aim of this study was to compare three commercial kits, Salm SeroGen (Salm Sero-Genotyping AS-1 kit), Check&Trace (Check-Points), and xMAP (xMAP Salmonella serotyping assay), to the Salmonella genoserotyping array (SGSA) developed by our laboratory. They were assessed using a panel of 321 isolates that represent commonly reported serovars from human and nonhuman sources globally. The four methods correctly identified 73.8% to 94.7% of the isolates tested. The methods correctly identified 85% and 98% of the clinically important Salmonella serovars Enteritidis and Typhimurium, respectively. The methods correctly identified 75% to 100% of the nontyphoidal, broad host range Salmonella serovars, including Heidelberg, Hadar, Infantis, Kentucky, Montevideo, Newport, and Virchow. The sensitivity and specificity of Salmonella serovars Typhimurium and Enteritidis ranged from 85% to 100% and 99% to 100%, respectively.It is anticipated that whole-genome sequencing will replace serotyping in public health laboratories in the future. However, at present, it is approximately three times more expensive than molecular methods. Until consistent standards and methodologies are deployed for whole-genome sequencing, data analysis and interlaboratory comparability remain a challenge. The use of molecular serotyping will provide a valuable high-throughput alternative to traditional serotyping. This comprehensive analysis provides a detailed comparison of commercial kits available for the molecular serotyping of Salmonella. PMID:27194688
Preliminary Evidence on the Diagnostic and Molecular Role of Circulating Soluble EGFR in Non-Small Cell Lung Cancer
Lococo, Filippo; Paci, Massimiliano; Rapicetta, Cristian; Rossi, Teresa; Sancisi, Valentina; Braglia, Luca; Cavuto, Silvio; Bisagni, Alessandra; Bongarzone, Italia; Noonan, Douglas M.; Albini, Adriana; Maramotti, Sally
Assessment of biological diagnostic factors providing clinically-relevant information to guide physician decision-making are still needed for diseases with poor outcomes, such as non-small cell lung cancer (NSCLC). Epidermal growth factor receptor (EGFR) is a promising molecule in the clinical management of NSCLC. While the EGFR transmembrane form has been extensively investigated in large clinical trials, the soluble, circulating EGFR isoform (sEGFR), which may have a potential clinical use, has rarely been considered. This study investigates the use of sEGFR as a potential diagnostic biomarker for NSCLC and also characterizes the biological function of sEGFR to clarify the molecular mechanisms involved in the course of action of this protein. Plasma sEGFR levels from a heterogeneous cohort of 37 non-advanced NSCLC patients and 54 healthy subjects were analyzed by using an enzyme-linked immunosorbent assay. The biological function of sEGFR was analyzed in vitro using NSCLC cell lines, investigating effects on cell proliferation and migration. We found that plasma sEGFR was significantly decreased in the NSCLC patient group as compared to the control group (median value: 48.6 vs. 55.6 ng/mL respectively; p = 0.0002). Moreover, we demonstrated that sEGFR inhibits growth and migration of NSCLC cells in vitro through molecular mechanisms that included perturbation of EGF/EGFR cell signaling and holoreceptor internalization. These data show that sEGFR is a potential circulating biomarker with a physiological protective role, providing a first approach to the functional role of the soluble isoform of EGFR. However, the impact of these data on daily clinical practice needs to be further investigated in larger prospective studies. PMID:26295387
Express diagnostics of intact and pathological dental hard tissues by optical PNC method
NASA Astrophysics Data System (ADS)
Masychev, Victor I.; Alexandrov, Michail T.
The results of hard tooth tissues research by the optical PNC- method in experimental and clinical conditions are presented. In the experiment under 90 test-sample of tooth slices with thickness about 1 mm (enamel, dentine and cement) were researched. The results of the experiment were processed by the method of correlation analyze. Clinical researches were executed on teeth of 210 patients. The regions of tooth tissue diseases with initial, moderate and deep caries were investigated. Spectral characteristics of intact and pathologically changed tooth tissues are presented and their peculiar features are discussed. The results the optical PNC- method application while processing tooth carious cavities are presented in order to estimate efficiency of the mechanical and antiseptic processing of teeth. It is revealed that the PNC-method can be used as for differential diagnostics of a degree dental carious stage, as for estimating of carefulness of tooth cavity processing before filling.
Application of Molecular and Serological Methods for Rapid Detection of Mycoplasma gallisepticum Infection (Avian mycoplasmosis).
Qasem, Jafar A; Al-Mouqati, Salwa A; Al-Ali, Ebtesam M; Ben-Haji, Ahmad
Mycoplasma infection is a major problem in veterinary medicine and in poultry production. The pathogen has many strains, so that diagnosis of the disease using culture method is not effective. The objective of this work was to evaluate the prevalence of Mycoplasma gallisepticum (MG) in Kuwait poultry farms using serology and molecular methods in comparison to the culture under specific conditions. A total of 50 swab samples from choanal cleft and tracheal samples and blood samples were obtained from three different local farms, the blood samples were processed for an Enzyme Linked Immunosorbent Assay (ELISA) detection and the swab samples for Polymerase Chain Reaction (PCR) and culture methods detection. A PCR diagnostic kit (VenoMGs) and ELISA diagnostic kit (ProFLOK), were used in comparison to the traditional culture method, to study the spread of this disease in samples from broiler and layer flocks. Fifty chicken samples were tested for mycoplasmosis, samples tested with ELISA gave 24 positive (48%) and 29 were positive by PCR (58%) and only seven (14%) were positive with culture methods. Swab samples obtained from the choanal cleft gave more positive (60%) with PCR than tracheal samples (56.6%). The culture gave 20 and 5% positive, respectively for tracheal and choanal samples. The methods reported here are of high sensitivity and specificity for Mycoplasma. Both the PCR and ELISA methods are superior to culture method for detection of avian mycoplasmosis. This study showed that MG infection is prevalent in commercial broiler and layer chickens in Kuwait poultry farms. The use of these methods for surveillance of the disease will establish data concerning the predominant Mycoplasmosis diseases in Kuwait if done on a large scale. PMID:26364358
Triangulated manifold meshing method preserving molecular surface topology.
Chen, Minxin; Tu, Bin; Lu, Benzhuo
Generation of manifold mesh is an urgent issue in mathematical simulations of biomolecule using boundary element methods (BEM) or finite element method (FEM). Defects, such as not closed mesh, intersection of elements and missing of small structures, exist in surface meshes generated by most of the current meshing method. Usually the molecular surface meshes produced by existing methods need to be revised carefully by third party software to ensure the surface represents a continuous manifold before being used in a BEM and FEM calculations. Based on the trace technique proposed in our previous work, in this paper, we present an improved meshing method to avoid intersections and preserve the topology of the molecular Gaussian surface. The new method divides the whole Gaussian surface into single valued pieces along each of x, y, z directions by tracing the extreme points along the fold curves on the surface. Numerical test results show that the surface meshes produced by the new method are manifolds and preserve surface topologies. The result surface mesh can also be directly used in surface conforming volume mesh generation for FEM type simulation. PMID:23117290
Conformational analysis of methylphenidate: comparison of molecular orbital and molecular mechanics methods.
Gilbert, Kathleen M; Skawinski, William J; Misra, Milind; Paris, Kristina A; Naik, Neelam H; Buono, Ronald A; Deutsch, Howard M; Venanzi, Carol A
Methylphenidate (MP) binds to the cocaine binding site on the dopamine transporter and inhibits reuptake of dopamine, but does not appear to have the same abuse potential as cocaine. This study, part of a comprehensive effort to identify a drug treatment for cocaine abuse, investigates the effect of choice of calculation technique and of solvent model on the conformational potential energy surface (PES) of MP and a rigid methylphenidate (RMP) analogue which exhibits the same dopamine transporter binding affinity as MP. Conformational analysis was carried out by the AM1 and AM1/SM5.4 semiempirical molecular orbital methods, a molecular mechanics method (Tripos force field with the dielectric set equal to that of vacuum or water) and the HF/6-31G* molecular orbital method in vacuum phase. Although all three methods differ somewhat in the local details of the PES, the general trends are the same for neutral and protonated MP. In vacuum phase, protonation has a distinctive effect in decreasing the regions of space available to the local conformational minima. Solvent has little effect on the PES of the neutral molecule and tends to stabilize the protonated species. The random search (RS) conformational analysis technique using the Tripos force field was found to be capable of locating the minima found by the molecular orbital methods using systematic grid search. This suggests that the RS/Tripos force field/vacuum phase protocol is a reasonable choice for locating the local minima of MP. However, the Tripos force field gave significantly larger phenyl ring rotational barriers than the molecular orbital methods for MP and RMP. For both the neutral and protonated cases, all three methods found the phenyl ring rotational barriers for the RMP conformers/invertamers (denoted as cte, tte, and cta) to be: cte, tte > MP > cta. Solvation has negligible effect on the phenyl ring rotational barrier of RMP. The B3LYP/6-31G* density functional method was used to calculate the
Impact of gene patents on diagnostic testing: a new patent landscaping method applied to spinocerebellar ataxia
Berthels, Nele; Matthijs, Gert; Van Overwalle, Geertrui
Recent reports in Europe and the United States raise concern about the potential negative impact of gene patents on the freedom to operate of diagnosticians and on the access of patients to genetic diagnostic services. Patents, historically seen as legal instruments to trigger innovation, could cause undesired side effects in the public health domain. Clear empirical evidence on the alleged hindering effect of gene patents is still scarce. We therefore developed a patent categorization method to determine which gene patents could indeed be problematic. The method is applied to patents relevant for genetic testing of spinocerebellar ataxia (SCA). The SCA test is probably the most widely used DNA test in (adult) neurology, as well as one of the most challenging due to the heterogeneity of the disease. Typically tested as a gene panel covering the five common SCA subtypes, we show that the patenting of SCA genes and testing methods and the associated licensing conditions could have far-reaching consequences on legitimate access to this gene panel. Moreover, with genetic testing being increasingly standardized, simply ignoring patents is unlikely to hold out indefinitely. This paper aims to differentiate among so-called 'gene patents' by lifting out the truly problematic ones. In doing so, awareness is raised among all stakeholders in the genetic diagnostics field who are not necessarily familiar with the ins and outs of patenting and licensing. PMID:21811306
White-Nose Syndrome Disease Severity and a Comparison of Diagnostic Methods.
McGuire, Liam P; Turner, James M; Warnecke, Lisa; McGregor, Glenna; Bollinger, Trent K; Misra, Vikram; Foster, Jeffrey T; Frick, Winifred F; Kilpatrick, A Marm; Willis, Craig K R
White-nose syndrome is caused by the fungus Pseudogymnoascus destructans and has killed millions of hibernating bats in North America but the pathophysiology of the disease remains poorly understood. Our objectives were to (1) assess non-destructive diagnostic methods for P. destructans infection compared to histopathology, the current gold-standard, and (2) to evaluate potential metrics of disease severity. We used data from three captive inoculation experiments involving 181 little brown bats (Myotis lucifugus) to compare histopathology, quantitative PCR (qPCR), and ultraviolet fluorescence as diagnostic methods of P. destructans infection. To assess disease severity, we considered two histology metrics (wing area with fungal hyphae, area of dermal necrosis), P. destructans fungal load (qPCR), ultraviolet fluorescence, and blood chemistry (hematocrit, sodium, glucose, pCO2, and bicarbonate). Quantitative PCR was most effective for early detection of P. destructans, while all three methods were comparable in severe infections. Correlations among hyphae and necrosis scores, qPCR, ultraviolet fluorescence, blood chemistry, and hibernation duration indicate a multi-stage pattern of disease. Disruptions of homeostasis occurred rapidly in late hibernation. Our results provide valuable information about the use of non-destructive techniques for monitoring, and provide novel insight into the pathophysiology of white-nose syndrome, with implications for developing and implementing potential mitigation strategies. PMID:26957435
Epidemiology, molecular virology and diagnostics of Schmallenberg virus, an emerging orthobunyavirus in Europe
After the unexpected emergence of Bluetongue virus serotype 8 (BTV-8) in northern Europe in 2006, another arbovirus, Schmallenberg virus (SBV), emerged in Europe in 2011 causing a new economically important disease in ruminants. The virus, belonging to the Orthobunyavirus genus in the Bunyaviridae family, was first detected in Germany, in The Netherlands and in Belgium in 2011 and soon after in the United Kingdom, France, Italy, Luxembourg, Spain, Denmark and Switzerland. This review describes the current knowledge on the emergence, epidemiology, clinical signs, molecular virology and diagnosis of SBV infection. PMID:23675914
A Parallel Iterative Method for Computing Molecular Absorption Spectra.
Koval, Peter; Foerster, Dietrich; Coulaud, Olivier
We describe a fast parallel iterative method for computing molecular absorption spectra within TDDFT linear response and using the LCAO method. We use a local basis of "dominant products" to parametrize the space of orbital products that occur in the LCAO approach. In this basis, the dynamic polarizability is computed iteratively within an appropriate Krylov subspace. The iterative procedure uses a matrix-free GMRES method to determine the (interacting) density response. The resulting code is about 1 order of magnitude faster than our previous full-matrix method. This acceleration makes the speed of our TDDFT code comparable with codes based on Casida's equation. The implementation of our method uses hybrid MPI and OpenMP parallelization in which load balancing and memory access are optimized. To validate our approach and to establish benchmarks, we compute spectra of large molecules on various types of parallel machines. The methods developed here are fairly general, and we believe they will find useful applications in molecular physics/chemistry, even for problems that are beyond TDDFT, such as organic semiconductors, particularly in photovoltaics. PMID:26616067
This is Part 2 of "Diagnostic Evaluation of Air Quality Models Using Advanced Methods with Specialized Observations of Selected Ambient Species". A limited field campaign to make specialized observations of selected ambient species using advanced and innovative instrumentation f...
Evaluation of the Effect of Fluctuation of Absolute Value for Diagnostic Accuracy of Fatigue Crack Monitoring Via Statistical Diagnostic Method Using Correlation between Sensors
NASA Astrophysics Data System (ADS)
Iwasaki, Atsushi; Morimoto, Akihiro; Yatomi, Masataka; Kimura, Tadashi
This research is about improvement of the diagnostic accuracy of the fatigue crack monitoring via the statistical diagnostic method. Our research group proposes an unsupervised damage diagnostic method named SI-F method which diagnoses the damage from detecting the change of correlation between sensors caused by the initiation or propagation of the damage via the statistical evaluation. By the method, correlation between sensors is identified by using the response surface and the change of them is statistically investigated with the F-test. To identify the crack length by the method, identification about the relation between the crack length and the F0 statistic is required. Then in this research, to evaluate effect of the regression error, the noise magnitude and the fluctuation of the external force to the relation, numerical simulation was conducted. For the simulation, two sets of data, one with constant load and one with variable load, are generated and compared. And the applicability of the result of the simulation is experimentally investigated. Finally, the results indicate that the F0 affected by the regression error and the noise magnitude but not affected by the external force.
Efficacy Assessment of Nucleic Acid Decontamination Reagents Used in Molecular Diagnostic Laboratories.
Fischer, Melina; Renevey, Nathalie; Thür, Barbara; Hoffmann, Donata; Beer, Martin; Hoffmann, Bernd
The occurrence of nucleic acid cross contamination in the laboratory resulting in false positive results of diagnostic samples is seriously problematic. Despite precautions to minimize or even avoid nucleic acid cross contaminations, it may appear anyway. Until now, no standardized strategy is available to evaluate the efficacy of commercially offered decontamination reagents. Therefore, a protocol for the reliable determination of nucleic acid decontamination efficacy using highly standardized solution and surface tests was established and validated. All tested sodium hypochlorite-based reagents proved to be highly efficient in nucleic acid decontamination even after short reaction times. For DNA Away, a sodium hydroxide-based decontamination product, dose- and time-dependent effectiveness was ascertained. For two other commercial decontamination reagents, the phosphoric acid-based DNA Remover and the non-enzymatic reagent DNA-ExitusPlus™ IF, no reduction of amplifiable DNA/RNA was observed. In conclusion, a simple test procedure for evaluation of the elimination efficacy of decontamination reagents against amplifiable nucleic acid is presented. PMID:27410228
Efficacy Assessment of Nucleic Acid Decontamination Reagents Used in Molecular Diagnostic Laboratories
Fischer, Melina; Renevey, Nathalie; Thür, Barbara; Hoffmann, Donata; Beer, Martin; Hoffmann, Bernd
The occurrence of nucleic acid cross contamination in the laboratory resulting in false positive results of diagnostic samples is seriously problematic. Despite precautions to minimize or even avoid nucleic acid cross contaminations, it may appear anyway. Until now, no standardized strategy is available to evaluate the efficacy of commercially offered decontamination reagents. Therefore, a protocol for the reliable determination of nucleic acid decontamination efficacy using highly standardized solution and surface tests was established and validated. All tested sodium hypochlorite-based reagents proved to be highly efficient in nucleic acid decontamination even after short reaction times. For DNA Away, a sodium hydroxide-based decontamination product, dose- and time-dependent effectiveness was ascertained. For two other commercial decontamination reagents, the phosphoric acid-based DNA Remover and the non-enzymatic reagent DNA-ExitusPlus™ IF, no reduction of amplifiable DNA/RNA was observed. In conclusion, a simple test procedure for evaluation of the elimination efficacy of decontamination reagents against amplifiable nucleic acid is presented. PMID:27410228
Genomics, Molecular Epidemiology and Diagnostics of Infectious hypodermal and hematopoietic necrosis virus.
Rai, Praveen; Safeena, Muhammed P; Krabsetsve, Kjersti; La Fauce, Kathy; Owens, Leigh; Karunasagar, Indrani
Infectious hypodermal and hematopoietic necrosis virus (IHHNV) is one of the major viral pathogens of penaeid shrimps worldwide, which has resulted in severe mortalities of up to 90 % in cultured Penaeus (Litopenaeus) stylirostris from Hawaii and hence designated Penaeus stylirostris densovirus (PstDNV). IHHNV is distributed in shrimp culture facilities worldwide. It causes large economic loss to the shrimp farming industry. Our knowledge about the natural reservoirs of IHHNV is still scarce. Recent studies suggest that there is sufficient sequence variation among the isolates from different locations in Asia, suggesting multiple geographical strains of the virus. Four complete genomes and several partial sequences of the virus are available in the GenBank. Complete genome information would be useful for assessing the specificity of diagnostics for viruses from different geographical areas. Comparisons of complete genome sequences will help us gain insights into point mutations that can affect virulence of the virus. In addition, because of unavailability of shrimp cell lines for culturing IHHNV in vitro, quantification of virus is difficult. The recent progress in research regarding clinical signs, geographical distribution, complete genome sequence and genetic variation, transmission has made it possible to obtain information on IHHNV. A comprehensive understanding of IHHNV infection process, pathogenesis, structural proteins and replication is essential for developing prevention measures. To date, no effective prophylactic measure for IHHNV infection is available for shrimp to reduce its impact. This review provides an overview of key issues regarding IHHNV infection and disease in commercially important shrimp species. PMID:23997444
Diagnostic/prognostic molecular cytogenetic follow-up applied in satellited marker cases
Papenhausen, P.R.; Anderson, S.
Special caution needs to be exercised in offering a good prognosis in Prader-Willi probe negative 15-derived marker cases, since it is clear that phenotypic effects can still be associated with the apparent presence of proximal sequences. We have had two postnatal cases in this category, one which was inherited from an unaffected paternal (non-mosaic) carrier, possibly demonstrating imprinting effects. Familial studies are continuing in this case. Although the D22/S9 locus appears diagnostic of cateye syndrome (CES), the dual specificity of the 14/22 centromeric probe leaves the possibility of a poor prognosis 14 derivation when the CES probe is negative. Therefore, it is imperative that proximal long arm 13, 14, 21 and more proximal 15 FISH probes be implemented so that a phenotypically correlated database may indicate the proper FISH probes necessary for accurate prognosis. Bisatellited markers is which a bipartite centromeric probe signal was found were considered to be higher risk than those with the single signal in counseling.
NIR-Cyanine Dye Linker: a Promising Candidate for Isochronic Fluorescence Imaging in Molecular Cancer Diagnostics and Therapy Monitoring
Komljenovic, Dorde; Wiessler, Manfred; Waldeck, Waldemar; Ehemann, Volker; Pipkorn, Ruediger; Schrenk, Hans-Hermann; Debus, Jürgen; Braun, Klaus
Personalized anti-cancer medicine is boosted by the recent development of molecular diagnostics and molecularly targeted drugs requiring rapid and efficient ligation routes. Here, we present a novel approach to synthetize a conjugate able to act simultaneously as an imaging and as a chemotherapeutic agent by coupling functional peptides employing solid phase peptide synthesis technologies. Development and the first synthesis of a fluorescent dye with similarity in the polymethine part of the Cy7 molecule whose indolenine-N residues were substituted with a propylene linker are described. Methylating agent temozolomide is functionalized with a tetrazine as a diene component whereas Cy7-cell penetrating peptide conjugate acts as a dienophilic reaction partner for the inverse Diels-Alder click chemistry-mediated ligation route yielding a theranostic conjugate, 3-mercapto-propionic-cyclohexenyl-Cy7-bis-temozolomide-bromide-cell penetrating peptide. Synthesis route described here may facilitate targeted delivery of the therapeutic compound to achieve sufficient local concentrations at the target site or tissue. Its versatility allows a choice of adequate imaging tags applicable in e.g. PET, SPECT, CT, near-infrared imaging, and therapeutic substances including cytotoxic agents. Imaging tags and therapeutics may be simultaneously bound to the conjugate applying click chemistry. Theranostic compound presented here offers a solid basis for a further improvement of cancer management in a precise, patient-specific manner. PMID:26722379
NIR-Cyanine Dye Linker: a Promising Candidate for Isochronic Fluorescence Imaging in Molecular Cancer Diagnostics and Therapy Monitoring.
Komljenovic, Dorde; Wiessler, Manfred; Waldeck, Waldemar; Ehemann, Volker; Pipkorn, Ruediger; Schrenk, Hans-Hermann; Debus, Jürgen; Braun, Klaus
Personalized anti-cancer medicine is boosted by the recent development of molecular diagnostics and molecularly targeted drugs requiring rapid and efficient ligation routes. Here, we present a novel approach to synthetize a conjugate able to act simultaneously as an imaging and as a chemotherapeutic agent by coupling functional peptides employing solid phase peptide synthesis technologies. Development and the first synthesis of a fluorescent dye with similarity in the polymethine part of the Cy7 molecule whose indolenine-N residues were substituted with a propylene linker are described. Methylating agent temozolomide is functionalized with a tetrazine as a diene component whereas Cy7-cell penetrating peptide conjugate acts as a dienophilic reaction partner for the inverse Diels-Alder click chemistry-mediated ligation route yielding a theranostic conjugate, 3-mercapto-propionic-cyclohexenyl-Cy7-bis-temozolomide-bromide-cell penetrating peptide. Synthesis route described here may facilitate targeted delivery of the therapeutic compound to achieve sufficient local concentrations at the target site or tissue. Its versatility allows a choice of adequate imaging tags applicable in e.g. PET, SPECT, CT, near-infrared imaging, and therapeutic substances including cytotoxic agents. Imaging tags and therapeutics may be simultaneously bound to the conjugate applying click chemistry. Theranostic compound presented here offers a solid basis for a further improvement of cancer management in a precise, patient-specific manner. PMID:26722379
A Portable, Pressure Driven, Room Temperature Nucleic Acid Extraction and Storage System for Point of Care Molecular Diagnostics
Byrnes, Samantha; Fan, Andy; Trueb, Jacob; Jareczek, Francis; Mazzochette, Mark; Sharon, Andre; Sauer-Budge, Alexis F.; Klapperich, Catherine M.
Many new and exciting portable HIV viral load testing technologies are emerging for use in global medicine. While the potential to provide fast, isothermal, and quantitative molecular diagnostic information to clinicians in the field will soon be a reality, many of these technologies lack a robust front end for sample clean up and nucleic acid preparation. Such a technology would enable many different downstream molecular assays. Here, we present a portable system for centrifuge-free room temperature nucleic acid extraction from small volumes of whole blood (70 µL), using only thermally stable reagents compatible with storage and transport in low resource settings. Quantitative reverse transcriptase polymerase chain reaction (qRT-PCR) analysis of simulated samples demonstrate a lower limit of detection of 1000 copies/ml, with the ability to detect differences in viral load across four orders of magnitude. The system can also be used to store extracted RNA on detachable cartridges for up to one week at ambient temperature, and can be operated using only hand generated air pressure. PMID:23914255
Combining simple patient-oriented tests with state-of-the-art molecular diagnostics for early diagnosis of cancer
Early diagnosis is an important strategy to improve outcomes from cancer. Oesophageal adenocarcinoma is an example of a cancer that presents late, with very poor outcomes, and for which the presence of the precursor lesion Barrett's oesophagus provides the opportunity to intervene at an early stage. In this review, I describe the challenges in the field and the work that we have done to devise a conceptually novel approach to early diagnosis, using a cell collection device (Cytosponge), coupled with molecular assays. This is a personal perspective in which I also describe the career pathway that led me into academic gastroenterology, and the rewards and challenges of translational research in molecular diagnostics. There are fantastic opportunities for clinicians wishing to pursue academic medicine, because it is a time when massive strides are being made in a whole number of areas; for example: imaging, sequencing technology and targeted therapies. Clinicians who can straddle the laboratory and the clinic are essential, to maximise the progress that can be made for the benefit of patients. PMID:26137297
Investigation of opportunities of the optical non-invasive diagnostics method for the blood sugar control
NASA Astrophysics Data System (ADS)
Lastovskaia, Elena A.; Gorbunova, Elena V.; Chertov, Aleksandr N.; Korotaev, Valery V.
The relevance of noninvasive method for determining the blood sugar is caused by necessity of regular monitoring of glucose levels in diabetic patients blood. Traditional invasive method is painful, because it requires a finger pricking. Despite the active studies in the field of non-invasive medical diagnostics, to date the painless and inexpensive instrument for blood sugar control for personal use doesn't exist. It's possible to measure the concentration of glucose in the blood with help of spectrophotometry method. It consists of registering and analyzing the spectral characteristics of the radiation which missed, reflected or absorbed by the object. The authors proposed a measuring scheme for studying the spectral characteristics of the radiation, missed by earlobe. Ultra-violet, visible and near infrared spectral ranges are considered. The paper presents the description of construction and working principles of the proposed special retaining clip and results of experiment with real patient.
Application of Diagnostic/Prognostic Methods to Critical Equipment for the Spent Nuclear Fuel Cleanup Program
Casazza, Lawrence O.; Jarrell, Donald B.; Koehler, Theresa M.; Meador, Richard J.; Wallace, Dale E.
The management of the Spent Nuclear Fuel (SNF) project at the Hanford K-Basin in the 100 N Area has successfully restructured the preventive maintenance, spare parts inventory requirements, and the operator rounds data requirements. In this investigation, they continue to examine the different facets of the operations and maintenance (O&M) of the K-Basin cleanup project in search of additional reliability and cost savings. This report focuses on the initial findings of a team of PNNL engineers engaged to identify potential opportunities for reducing the cost of O&M through the application of advanced diagnostics (fault determination) and prognostics (residual life/reliability determination). The objective is to introduce predictive technologies to eliminate or reduce high impact equipment failures. The PNNL team in conjunction with the SNF engineers found the following major opportunities for cost reduction and/or enhancing reliability: (1) Provide data routing and automated analysis from existing detection systems to a display center that will engage the operations and engineering team. This display will be operator intuitive with system alarms and integrated diagnostic capability. (2) Change operating methods to reduce major transients induced in critical equipment. This would reduce stress levels on critical equipment. (3) Install a limited sensor set on failure prone critical equipment to allow degradation or stressor levels to be monitored and alarmed. This would provide operators and engineers with advance guidance and warning of failure events. Specific methods for implementation of the above improvement opportunities are provided in the recommendations. They include an Integrated Water Treatment System (IWTS) decision support system, introduction of variable frequency drives on certain pump motors, and the addition of limited diagnostic instrumentation on specified critical equipment.
Fluorescent-spectroscopic and imaging methods of investigations for diagnostics of head and neck tumors and control of PDT
NASA Astrophysics Data System (ADS)
Edinak, N. E.; Chental, Victor V.; Komov, D.; Vaculovskaya, E.; Tabolinovskaya, T. D.; Abdullin, N. A.; Pustynsky, I.; Chatikhin, V.; Loschenov, Victor B.; Meerovich, Gennady A.; Stratonnikov, A. A.; Linkov, Kirill G.; Agafonov, Vladimir I.; Zuravleva, V.; Lukjanets, E.
Methodics of PDT control and fluorescent-spectroscopic diagnostic of head and neck tumors and mammary gland cancer (nodular) with the use of Kr, He-Ne and semiconductor lasers and photosensitizer (PS) -- Al phtalocyanin (Photosense) are discussed. The results show that applied diagnostic methods permit us not only to identify the topology and malignancy of a tumor but also to correct PDT process directly during irradiation.
Mammary carcinoma – current diagnostic methods and symptomatology in imaging studies
Popiel, Monika; Mróz-Klimas, Danuta; Kasprzak, Renata; Furmanek, Mariusz
Summary Breast cancer is the most common neoplasm of the female population and its incidence is constantly rising. Social campaigns educating the public about the importance of the problem have been conducted for the past several years. Women are encouraged to self-examine on a monthly basis. Women aged 50–69 years can have an x-ray mammography performed once every 2 years as part of a prophylactic screening program. Ultrasound studies or MR mammography are adjuvant or, in some cases, alternative to x-ray mammography. Nuclear medicine techniques with application of oncophilic markers and receptor studies (this publication will not cover nuclear medicine methods) are not routinely used. Other techniques, such as computed tomography and conventional radiography are of no significance in the diagnostics of mammary cancer. However, together with isotopic methods, they are helpful in staging of the disease. X-ray mammography is, up to date, the only method with proven value in decreasing mortality. It is also the best available method for visualization of microcalicifications. Ultrasound examination is complementary to x-ray mammography as it is a cheap, easily available method of imaging mammary glands with higher glandular tissue content. It is also the most commonly used modality aiding in targeted biopsy of mammary gland. To date, MR mammography, characterized by the highest sensitivity in cancer diagnostics, remained a method reserved for "special tasks". MR is used for prophylaxis mainly in a population of women with particularly high risk of the disease and in cases where x-ray and ultrasound examinations are insufficient. Picture of mammary carcinoma in imaging studies is heterogeneous. However, it most often presents as an irregularly demarcated mass. Moreover, each modality can aid in visualization of additional features of a lesion such as typical shape of microcalcifications in x-ray mammography, characteristic pattern of contrast enhancement in MR
Evaluation of Three Rapid Diagnostic Methods for Direct Identification of Microorganisms in Positive Blood Cultures
Martinez, Raquel M.; Bauerle, Elizabeth R.; Fang, Ferric C.
The identification of organisms from positive blood cultures generally takes several days. However, recently developed rapid diagnostic methods offer the potential for organism identification within only a few hours of blood culture positivity. In this study, we evaluated the performance of three commercial methods to rapidly identify organisms directly from positive blood cultures: QuickFISH (AdvanDx, Wolburn, MA), Verigene Gram-Positive Blood Culture (BC-GP; Nanosphere, Northbrook, IL), and matrix-assisted laser desorption ionization–time of flight mass spectrometry (MALDI-TOF MS) with Sepsityper processing (Bruker Daltonics, Billerica, MA). A total of 159 blood cultures (VersaTREK Trek Diagnostic Systems, Cleveland, OH) positive for Gram-positive and Gram-negative bacteria as well as yeast were analyzed with QuickFISH and MALDI-TOF MS. In all, 102 blood cultures were analyzed using the BC-GP assay. For monomicrobial cultures, we observed 98.0% concordance with routine methods for both QuickFISH (143/146) and the BC-GP assay (93/95). MALDI-TOF MS demonstrated 80.1% (117/146) and 87.7% (128/146) concordance with routine methods to the genus and species levels, respectively. None of the methods tested were capable of consistently identifying polymicrobial cultures in their entirety or reliably differentiating Streptococcus pneumoniae from viridans streptococci. Nevertheless, the methods evaluated in this study are convenient and accurate for the most commonly encountered pathogens and have the potential to dramatically reduce turnaround time for the provision of results to the treating physician. PMID:24808235
Beyond Standard Molecular Dynamics: Investigating the Molecular Mechanisms of G Protein-Coupled Receptors with Enhanced Molecular Dynamics Methods
Johnston, Jennifer M.
The majority of biological processes mediated by G Protein-Coupled Receptors (GPCRs) take place on timescales that are not conveniently accessible to standard molecular dynamics (MD) approaches, notwithstanding the current availability of specialized parallel computer architectures, and efficient simulation algorithms. Enhanced MD-based methods have started to assume an important role in the study of the rugged energy landscape of GPCRs by providing mechanistic details of complex receptor processes such as ligand recognition, activation, and oligomerization. We provide here an overview of these methods in their most recent application to the field. PMID:24158803
Substructure synthesis method for simulating large molecular complexes
Ming, Dengming; Kong, Yifei; Wu, Yinghao; Ma, Jianpeng
This paper reports a computational method for describing the conformational flexibility of very large biomolecular complexes using a reduced number of degrees of freedom. It is called the substructure synthesis method, and the basic concept is to treat the motions of a given structure as a collection of those of an assemblage of substructures. The choice of substructures is arbitrary and sometimes quite natural, such as domains, subunits, or even large segments of biomolecular complexes. To start, a group of low-frequency substructure modes is determined, for instance by normal mode analysis, to represent the motions of the substructure. Next, a desired number of substructures are joined together by a set of constraints to enforce geometric compatibility at the interface of adjacent substructures, and the modes for the assembled structure can then be synthesized from the substructure modes by applying the Rayleigh–Ritz principle. Such a procedure is computationally much more desirable than solving the full eigenvalue problem for the whole assembled structure. Furthermore, to show the applicability to biomolecular complexes, the method is used to study F-actin, a large filamentous molecular complex involved in many cellular functions. The results demonstrate that the method is capable of studying the motions of very large molecular complexes that are otherwise completely beyond the reach of any conventional methods. PMID:12518058
Comparative genomics of Neisseria meningitidis strains: new targets for molecular diagnostics.
Diene, S M; Bertelli, C; Pillonel, T; Jacquier, N; Croxatto, A; Jaton, K; Greub, G
In 2010, Jaton et al. (False-negative PCR result due to gene polymorphism: the example of Neisseria meningitidis. J Clin Microbiol 2010;48:4590-2) reported an isolate of Neisseria meningitidis serogroup B that was not detected by the ctrA quantitative real-time PCR (qRT-PCR) used in our diagnostic laboratory. Sequence analysis of ctrA revealed several single nucleotide polymorphisms responsible for the negative qRT-PCR. Therefore, we sequenced the genome of this isolate and performed comparative genomics to propose new gene targets for the specific detection of N. meningitidis from clinical specimens. We identified 11 genes as specific to N. meningitidis genomes and common to at least 177 (97%) of the 183 genomes available. Among them, three genes (metA, tauE and shlA) were selected to develop new qRT-PCRs for the detection of N. meningitidis DNA. The three qRT-PCRs were highly sensitive and specific, and they exhibited a good reproducibility when tested on plasmidic positive controls and genomic DNA extracted from strains of N. meningitidis and other relevant bacterial species. The clinical sensitivity and specificity of metA and tauE qRT-PCRs were both 100% based on a testing of cerebrospinal fluid samples positive for N. meningitidis or other clinically relevant bacteria. Despite a 100% specificity, the sensitivity of the shlA qRT-PCR was only 70%. We thus recommend using the metA and/or tauE qRT-PCRs developed here. To prevent PCR failure in the presence of new polymorphic strains, the detection of dual targets by duplex qRT-PCR would be more accurate and suitable for the diagnosis of N. meningitidis from clinical specimens. PMID:27085725
A model for rewarding professional growth in the diagnostic molecular oncology laboratory.
Lennon, P A; Deleon, E; Reynolds, A; Pulido, L; Lewing, C; Mehta, P; Biscanin, S; Hai, S; Bowman, C; Galbincea, J; Jakacky, J; Hu, P
The University of Texas M.D. Anderson Cancer Center (UTMDACC), Department of Pathology and Laboratory Medicine is committed to the endless pursuit of innovative research, education, training and administration for the prevention, diagnosis and clinical management of cancer and associated diseases. The molecular genetic technology professional development model promotes personal development, recognizes increased competencies, and sets high standards for all skills and services provided. There are four competency levels that comprise our Professional Development Model (PDM): Discovery, Application, Maturation, and Expert. The skill, knowledge, education, and certification requirements for each level are defined based on the business needs of each lab. When a genetic technologist successfully completes all skills, knowledge, proficiency, education and certification requirements within the appropriate time frame for a particular competency level, his/her salary would be adjusted to the entry point for the competency level he/she has completed. PMID:21654071
Reconfigurable hybrid interface for molecular marker diagnostics and in-situ reporting.
Ehrhardt, Kristina; Guinn, Michael T; Quarton, Tyler; Zhang, Michael Q; Bleris, Leonidas
Combinations of molecular signals such as transcription factors and microRNAs in cells are a reliable indicator of multi-gene disorders. A system capable of detecting these conditions in-situ may be used as a tool for diagnosis and monitoring of disease. Here, we engineer genetic circuits that sense endogenous levels of the androgen receptor (AR), the glucocorticoid receptor (GR), and the microRNA hsa-miR-21 (miR-21) in cervical cancer cells (HeLa). Furthermore, using the mediator molecule human chorionic gonadotropin (hCG), we interface the intracellular information to enzyme-linked immunosorbent assay (ELISA) test strips. We demonstrate that this hybrid genetic circuit and test-strip interface can accommodate combinatorial, low-cost, and in-situ reporting, a versatile profiling tool. PMID:26210472
a Novel Method to Measure Spectra of Cold Molecular Ions
NASA Astrophysics Data System (ADS)
Chakrabarty, Satrajit; Holz, Mathias; Campbell, Ewen; Banerjee, Agniva; Gerlich, Dieter; Maier, John P.
A universal method has been developed in our group for measuring the spectra of molecular ions in a 22-pole radio frequency trap at low temperatures. It is based on laser induced inhibition of complex growth (LIICG)1. At low temperatures and high number densities of buffer gas, helium attaches to ions via ternary association. The formation of these weakly bound complexes, however, is inhibited following resonant absorption of the bare molecular ion. The first successful measurements have been demonstrated on the A 2Π_u ← X ^2Σ_g^+ electronic transition of N_2^+, with some thousand N_2^+ ions, helium densities of 1015 cm-3, and storage times of 1 s. The reduction in the number of N_2+-He complexes is the result of an interplay between excitation, radiative and collisional cooling, ternary association, and collision induced dissociation, and is explained using a kinetic model. The method is also applicable to larger molecular species. In this case internal conversion following electronic excitation produces internally "hot" ions, reducing the attachment of helium. The technique is universal because complex formation can be impeded over a wide wavelength range. [1] S. Chakrbarty, M. Holz, E. K. Campbell, A. Banerjee, D. Gerlich, and J. P. Maier, J. Phys. Chem. Lett. 2013, 4, 4051.
Molecular understanding of mutagenicity using potential energy methods
Broyde, S.; Shapiro, R.
Our objective, has been to elucidate on a molecular level, at atomic resolution, the structures of DNAs modified by 2-aminofluorene and its N-acetyl derivative, 2-acetylaminofluorene (AAF). The underlying hypothesis is that DNA replicates with reduced fidelity when its normal right-handed B-structure is altered, and one result is a higher mutation rate. This change in structure may occur normally at a low incidence, for example by the formation of hairpin loops in appropriate sequences, but it may be enhanced greatly after covalent modification by a mutagenic substance. We use computational methods and have been able to incorporate the first data from NMR studies in our calculations. Computational approaches are important because x-ray and spectroscopic studies have not succeeded in producing atomic resolution views of mutagen and carcinogen-oligonucleotide adducts. The specific methods that we employ are minimized potential energy calculations using the torsion angle space molecular mechanics program DUPLEX to yield static views. Molecular dynamics simulations, with full solvent and salt, of the important static structures are carried out with the program AMBER; this yields mobile views in a medium that mimics the natural aqueous environment of the cell as well as can be done with current available computing resources.
BREAST: a novel method to improve the diagnostic efficacy of mammography
NASA Astrophysics Data System (ADS)
Brennan, P. C.; Tapia, K.; Ryan, J.; Lee, W.
High quality breast imaging and accurate image assessment are critical to the early diagnoses, treatment and management of women with breast cancer. Breast Screen Reader Assessment Strategy (BREAST) provides a platform, accessible by researchers and clinicians world-wide, which will contain image data bases, algorithms to assess reader performance and on-line systems for image evaluation. The platform will contribute to the diagnostic efficacy of breast imaging in Australia and beyond on two fronts: reducing errors in mammography, and transforming our assessment of novel technologies and techniques. Mammography is the primary diagnostic tool for detecting breast cancer with over 800,000 women X-rayed each year in Australia, however, it fails to detect 30% of breast cancers with a number of missed cancers being visible on the image [1-6]. BREAST will monitor the mistakes, identify reasons for mammographic errors, and facilitate innovative solutions to reduce error rates. The BREAST platform has the potential to enable expert assessment of breast imaging innovations, anywhere in the world where experts or innovations are located. Currently, innovations are often being assessed by limited numbers of individuals who happen to be geographically located close to the innovation, resulting in equivocal studies with low statistical power. BREAST will transform this current paradigm by enabling large numbers of experts to assess any new method or technology using our embedded evaluation methods. We are confident that this world-first system will play an important part in the future efficacy of breast imaging.
An improved method for estimating the entrance exposure in diagnostic radiographic examinations.
Zamenhof, R G; Shahabi, S; Morgan, H T
There is currently a widespread consensus on the importance of monitoring patient radiation exposures during radiographic examinations. Diagnostic facilities under federal jurisdiction already legislate maximum patient exposure limits for various diagnostic radiologic examinations, while an increasing number of state legislatures have instituted such regulations. Compliance requires that institutions be capable of assessing each patient's entrance exposures. A method is proposed that would facilitate the acquisition of such patient exposure information in a relatively straightforward and accurate manner, requiring a minimum number of measurements and access to a suitable programmable calculator. A standardized set of exposure measurements obtained on an accurately calibrated three-phase radiographic unit has been fitted by an analytic function. The average accuracy of the fit between the limits of 40-140 kVp and 2.5- to 6.0-mm aluminum filtration was 0.3%. The concept of linear scaling was employed to allow the analytic function to accurately reproduce the exposure outputs of different radiographic units. Validation experiments on patients indicated that an overall accuracy of 10% can be expected when using well-calibrated radiographic equipment. The method described permits institutions to verify their compliance with federal and/or state regulations and to confirm that their radiation exposures are consistent with national averages. PMID:3497556
Role of Molecular Dynamics and Related Methods in Drug Discovery.
De Vivo, Marco; Masetti, Matteo; Bottegoni, Giovanni; Cavalli, Andrea
Molecular dynamics (MD) and related methods are close to becoming routine computational tools for drug discovery. Their main advantage is in explicitly treating structural flexibility and entropic effects. This allows a more accurate estimate of the thermodynamics and kinetics associated with drug-target recognition and binding, as better algorithms and hardware architectures increase their use. Here, we review the theoretical background of MD and enhanced sampling methods, focusing on free-energy perturbation, metadynamics, steered MD, and other methods most consistently used to study drug-target binding. We discuss unbiased MD simulations that nowadays allow the observation of unsupervised ligand-target binding, assessing how these approaches help optimizing target affinity and drug residence time toward improved drug efficacy. Further issues discussed include allosteric modulation and the role of water molecules in ligand binding and optimization. We conclude by calling for more prospective studies to attest to these methods' utility in discovering novel drug candidates. PMID:26807648
Elucidating diversity of exosomes: biophysical and molecular characterization methods.
Khatun, Zamila; Bhat, Anjali; Sharma, Shivani; Sharma, Aman
Exosomes are cell-secreted nanovesicles present in biological fluids in normal and diseased conditions. Owing to their seminal role in cell-cell communication, emerging evidences suggest that exosomes are fundamental regulators of various diseases. Due to their potential usefulness in disease diagnosis, robust isolation and characterization of exosomes is critical in developing exosome-based assays. In the last few years, different exosome characterization methods, both biophysical and molecular, have been developed to characterize these tiny vesicles. Here, in this review we summarize: first, biophysical techniques based on spectroscopy (e.g., Raman spectroscopy, dynamic light scattering) and other principles, for example, scanning electron microscopy, atomic force microscopy; second, antibody-based molecular techniques including flow cytometry, transmission electron microscopy and third, nanotechnology-dependent exosome characterization methodologies. PMID:27488053
Molecular Pathology and Personalized Medicine: The Dawn of a New Era in Companion Diagnostics—Practical Considerations about Companion Diagnostics for Non-Small-Cell-Lung-Cancer
Plönes, Till; Engel-Riedel, Walburga; Stoelben, Erich; Limmroth, Christina; Schildgen, Oliver; Schildgen, Verena
Companion diagnostics (CDx) have become a major tool in molecular pathology and assist in therapy decisions in an increasing number of various cancers. Particularly, the developments in lung cancer have been most impressing in the last decade and consequently lung cancer mutation testing and molecular profiling has become a major business of diagnostic laboratories. However, it has become difficult to decide which biomarkers are currently relevant for therapy decisions, as many of the new biomarkers are not yet approved as therapy targets, remain in the status of clinical studies, or still have not left the experimental phase. The current review is focussed on those markers that do have current therapy implications, practical implications arising from the respective companion diagnostics, and thus is focused on daily practice. PMID:26784235
Bayesian and maximum entropy methods for fusion diagnostic measurements with compact neutron spectrometers.
Reginatto, Marcel; Zimbal, Andreas
In applications of neutron spectrometry to fusion diagnostics, it is advantageous to use methods of data analysis which can extract information from the spectrum that is directly related to the parameters of interest that describe the plasma. We present here methods of data analysis which were developed with this goal in mind, and which were applied to spectrometric measurements made with an organic liquid scintillation detector (type NE213). In our approach, we combine Bayesian parameter estimation methods and unfolding methods based on the maximum entropy principle. This two-step method allows us to optimize the analysis of the data depending on the type of information that we want to extract from the measurements. To illustrate these methods, we analyze neutron measurements made at the PTB accelerator under controlled conditions, using accelerator-produced neutron beams. Although the methods have been chosen with a specific application in mind, they are general enough to be useful for many other types of measurements. PMID:18315297
A Review of the Giant Protein Titin in Clinical Molecular Diagnostics of Cardiomyopathies
Gigli, Marta; Begay, Rene L.; Morea, Gaetano; Graw, Sharon L.; Sinagra, Gianfranco; Taylor, Matthew R. G.; Granzier, Henk; Mestroni, Luisa
Titin (TTN) is known as the largest sarcomeric protein that resides within the heart muscle. Due to alternative splicing of TTN, the heart expresses two major isoforms (N2B and N2BA) that incorporate four distinct regions termed the Z-line, I-band, A-band, and M-line. Next-generation sequencing allows a large number of genes to be sequenced simultaneously and provides the opportunity to easily analyze giant genes such as TTN. Mutations in the TTN gene can cause cardiomyopathies, in particular dilated cardiomyopathy (DCM). DCM is the most common form of cardiomyopathy, and it is characterized by systolic dysfunction and dilation of the left ventricle. TTN truncating variants have been described as the most common cause of DCM, while the real impact of TTN missense variants in the pathogenesis of DCM is still unclear. In a recent population screening study, rare missense variants potentially pathogenic based on bioinformatic filtering represented only 12.6% of the several hundred rare TTN missense variants found, suggesting that missense variants are very common in TTN and are frequently benign. The aim of this review is to understand the clinical role of TTN mutations in DCM and in other cardiomyopathies. Whereas TTN truncations are common in DCM, there is evidence that TTN truncations are rare in the hypertrophic cardiomyopathy (HCM) phenotype. Furthermore, TTN mutations can also cause arrhythmogenic right ventricular cardiomyopathy (ARVC) with distinct clinical features and outcomes. Finally, the identification of a rare TTN missense variant cosegregating with the restrictive cardiomyopathy (RCM) phenotype suggests that TTN is a novel disease-causing gene in this disease. Clinical diagnostic testing is currently able to analyze over 100 cardiomyopathy genes, including TTN; however, the size and presence of extensive genetic variation in TTN presents clinical challenges in determining significant disease-causing mutations. This review discusses the current
Crystallo-optic diagnostics method of the soft laser-induced effects in biological fluids
NASA Astrophysics Data System (ADS)
Skopinov, S. A.; Yakovleva, S. V.
Presently, it is well known that individual cells"2 and higher organisms3'4 exhibit a marked response to soft laser irradiation in certain parts of the visible and near infrared spectral ranges. Broad clinical applications of laser therapy and slow progress in understanding of the physical, chemical and biological mechanisms of this phenomenon make the task to search new methods of objectivisation of laser-induces bioeffects very insistent. In this paper we give a short review of the methods of structural-optical diagnostics of the soft laser-induced effects in biofluids (blood and its fractions, saliva, juices, mucuses, exudations, etc.) and suggest their applications in experimental and clinical studies of the soft laser bioeffects.
A nondestructive method for diagnostic of insulated building walls using infrared thermography
NASA Astrophysics Data System (ADS)
Larbi Youcef, Mohamed H. A.; Mazioud, Atef; Bremond, Pierre; Ibos, Laurent; Candau, Yves; Piro, Michel; Filloux, Alain
This work deals with the development of an experimental protocol for the diagnostic of multi-layered insulated building walls. First, a test bench is set up in order to measure front and back sides temperatures of standard panels. The panels considered have insulation thicknesses of 2, 6 and 10cm. The front side is heated by two halogen lamps of 500W. A CEDIP Jade Long wave infrared camera and thermocouples are used to carry out temperature measurements. In a second time, a one dimensional model based on thermal quadruples and Laplace transforms was developped under Matlab environment. Also, we developped a three dimensional model based on finite volumes using Fluent computational code. Finally, a method of identification of physical parameters is implemented by performing least square minimization based on Levenberg-Marquardt method. The experimental measurements are compared to theoretical results and by minimization we obtain thermal conductivity and diffusivity as well as thickness of the two layers.
Diagnostic radiology peer review: a method inclusive of all interpreters of radiographic examinations regardless of specialty.
Hopper, K D; Rosetti, G F; Edmiston, R B; Madewell, J E; Beam, L M; Landis, J R; Miller, K L; Ricci, J A; McCauslin, M A
A proposed method of assessing the quality of diagnostic radiographic examinations includes peer review designed to evaluate physicians, including nonradiologists, involved in the performance and interpretation of such examinations. A pilot project evaluated this system with randomly selected Pennsylvania Blue Shield data files of 10 providers billing for chest radiography interpretations during the second quarter of 1989. Of the 98 chest radiographs reviewed blindly, all inadequately marked radiographs and incomplete written reports were produced by nonradiologists. Technical quality of images obtained by radiologists did not significantly differ from that of images obtained by nonradiologists (P = .189). All five interpretive errors that could have seriously affected the patient's health care were produced by nonradiologists (P = .019). Four of these serious errors were made by providers billing for fewer than 25 radiographs. While administrative and time cost limitations are obvious, this method of peer review encompasses all physicians billing for a particular radiographic service, irrespective of specialty. PMID:2068327
Molecular Rayleigh Scattering Diagnostic for Dynamic Temperature, Velocity, and Density Measurements
NASA Technical Reports Server (NTRS)
Mielke, Amy R.; Elam, Kristie A.; Sung, Chi-Jen
A molecular Rayleigh scattering technique is developed to measure dynamic gas temperature, velocity, and density in unseeded turbulent flows at sampling rates up to 16 kHz. A high power CW laser beam is focused at a point in an air jet plume and Rayleigh scattered light is collected and spectrally resolved. The spectrum of the light, which contains information about the temperature and velocity of the flow, is analyzed using a Fabry-Perot interferometer. The circular interference fringe pattern is divided into four concentric regions and sampled at 1 and 16 kHz using photon counting electronics. Monitoring the relative change in intensity within each region allows for measurement of gas temperature and velocity. Independently monitoring the total scattered light intensity provides a measure of gas density. A low speed heated jet is used to validate the measurement of temperature fluctuations and an acoustically excited nozzle flow is studied to validate velocity fluctuation measurements. Power spectral density calculations of the property fluctuations, as well as mean and fluctuating quantities are presented. Temperature fluctuation results are compared with constant current anemometry measurements and velocity fluctuation results are compared with constant temperature anemometry measurements at the same locations.
Bovine viral diarrhea virus: molecular cloning of genomic RNA and its diagnostic application
Brock, K.V.
Molecular cloning of a field isolate of bovine viral diarrhea virus (BVDV) strain 72 RNA was done in this study. The sensitivity and specificity of cloned cDNA sequences in hybridization assays with various BVDV strains were determined. cDNA was synthesized from polyadenylated BVDV RNA templates with oligo-dT primers, reverse transcriptase, and DNA polymerase I. The newly synthesized double-stranded BVDV cDNA was C-tailed with terminal deoxytransferase and annealed into G-tailed, Pst-1-cut pUC9 plasmid. Escherichia coli was transformed with the recombinant plasmids and a library of approximately 200 BVDV specific cDNA clones varying in length from 0.5 to 2.6 kilobases were isolated. The sensitivity and specificity of hybridization between the labelled cDNA and BVDV target sequences were determined. Cloned BVDV sequences were isolated from pUC9 plasmid DNA and labelled with /sup 32/P by nick translation. The detection limit by dot blot hybridization assay was 20 pg of purified genomic BVDV RNA. cDNA hybridization probes were specific for all strains of BVDV tested, regardless of whether they were noncytopathic and cytopathic, but did not hybridize with heterologous bovine viruses tested. Probes did not hybridize with uninfected cell culture or cellular RNA. Hybridization probes were at least as sensitive as infectivity assays in detecting homologous virus.
[Multiple endocrine neoplasia type 1 and 2. 1997 diagnostic guidelines and molecular pathology].
Komminoth, P
Multiple endocrine neoplasia syndromes (MEN) encompass autosomal dominantly inherited diseases which are characterized by the syn- or metachrone development of neoplastic and hyperplastic neuroendocrine lesions in several glands of an affected patient. In MEN type 1 the parathyroids, endocrine pancreas and duodenum and the pituitary and in MEN type 2 the thyroid C-cells, adrenal medulla and parathyroids are involved. Due to the recent identification of the mu gene and RET protooncogene as MEN-1 and MEN-2, respectively, and the elucidation of the genetic defects in affected patients, direct mutational analysis of germline DNA allows for the unambiguous identification of gene carriers and therefore the discrimination of MEN-associated and sporadically occurring neuroendocrine tumors. This is especially helpful in the context of the fairly high de novo mutation rates in MEN, since the discrimination of familial and sporadic neuroendocrine lesions by conventional and immunohistochemical analyses is rather unreliable. While the development of neuroendocrine lesions in young patients, bilateral or multicentricer tumors and the combination of hyperplastic and neoplastic lesions are indicative for a MEN syndrome, such constellations may also occur coincidentally or in association with other inherited diseases. In this overview, most recent findings concerning pathogenesis, molecular features, clinics and therapeutic concepts of MEN-1 and 2 are summarized and discussed. PMID:9380604
Diagnostic Molecular Mycobacteriology in Regions With Low Tuberculosis Endemicity: Combining Real-time PCR Assays for Detection of Multiple Mycobacterial Pathogens With Line Probe Assays for Identification of Resistance Mutations.
Deggim-Messmer, Vanessa; Bloemberg, Guido V; Ritter, Claudia; Voit, Antje; Hömke, Rico; Keller, Peter M; Böttger, Erik C
Molecular assays have not yet been able to replace time-consuming culture-based methods in clinical mycobacteriology. Using 6875 clinical samples and a study period of 35months we evaluated the use of PCR-based assays to establish a diagnostic workflow with a fast time-to-result of 1-2days, for 1. detection of Mycobacterium tuberculosis complex (MTB), 2. detection and identification of nontuberculous mycobacteria (NTM), and 3. identification of drug susceptible MTB. MTB molecular-based detection and culture gave concordant results for 97.7% of the specimens. NTM PCR-based detection and culture gave concordant results for 97.0% of the specimens. Defining specimens on the basis of combined laboratory data as true positives or negatives with discrepant results resolved by clinical chart reviews, we calculated sensitivity, specificity, PPV and NPV for PCR-based MTB detection as 84.7%, 100%, 100%, and 98.7%; the corresponding values for culture-based MTB detection were 86.3%, 100%, 100%, and 98.8%. PCR-based detection of NTM had a sensitivity of 84.7% compared to 78.0% of that of culture-based NTM detection. Molecular drug susceptibility testing (DST) by line-probe assay was found to predict phenotypic DST results in MTB with excellent accuracy. Our findings suggest a diagnostic algorithm to largely replace lengthy culture-based techniques by rapid molecular-based methods. PMID:27333026
Jakob, F; Genest, F; Seefried, L; Tsourdi, E; Lapa, C; Hofbauer, L C
Clinical diagnostics in metabolic bone diseases cover a broad spectrum of conventional and state of the art methods ranging from the medical history and clinical examination to molecular imaging. Patient treatment is carried out in an interdisciplinary team due to the multiple interactions of bone with other organ systems. Diagnosis of osteoporosis is supported by high level national guidelines. A paradigm shift concerning the clinical relevance of bone mineral density measurement renders this now to be a strong risk factor rather than a diagnostic parameter, while strengthening the value of other clinical factors for risk assessment. The impact of parameters for muscle mass, structure and function is steadily increasing in all age groups. In order to identify underlying diseases that influence bone metabolism a panel of general laboratory diagnostic parameters is recommended. Markers for bone formation and resorption and specific parameters for the regulation of calcium and phosphate metabolism should be evaluated by specialists because they require diligence in preanalytics and experience in interpretation. Genetic diagnosis is well established for rare bone diseases while diagnostic panels are not yet available for routine diagnostics in polygenetic diseases such as osteoporosis. Conventional radiology is still very important to identify, e. g. fractures, osteolytic and osteoblastic lesions and extraosseous calcifications; however tomography-based methods which combine, e. g. scintigraphy or positron emission technologies with anatomical imaging are of increasing significance. Clinical diagnostics in osteology require profound knowledge and are subject to a dynamic evolution. PMID:27307159
Diagnostic methods to cutaneous leishmaniasis detection in domestic dogs and cats.
Trevisan, Daliah Alves Coelho; Lonardoni, Maria Valdrinez Campana; Demarchi, Izabel Galhardo
Cutaneous leishmaniasis is caused by different species of Leishmania. In domestic animals such as dogs and cats, the diagnostic consists of clinical, epidemiological and serological tests, which changes among countries all around the world. Because of this diversity in the methods selected, we propose this systematic literature review to identify the methods of laboratory diagnosis used to detect cutaneous leishmaniasis in domestic dogs and cats in the Americas. Articles published in the last 5 years were searched in PubMed, ISI Web of Science, LILACS and Scielo, and we selected 10 papers about cutaneous leishmaniasis in dogs and cats in the Americas. In Brazil, often the indirect immunofluorescence and enzyme immunoassay (ELISA) have been applied. Other countries like United States and Mexico have been using antigenic fractions for antibodies detections by Western blot. ELISA and Western blot showed a higher sensitivity and efficacy in the detection of leishmaniasis. Analysis of sensibility and specificity of the methods was rarely used. Although confirmatory to leishmaniasis, direct methods for parasites detection and polymerase chain reaction showed low positivity in disease detection. We suggested that more than one method should be used for the detection of feline and canine leishmaniasis. Serological methods such as Western blot and enzyme immunoassay have a high efficacy in the diagnosis of this disease. PMID:26734869
Diagnostic methods to cutaneous leishmaniasis detection in domestic dogs and cats*
Trevisan, Daliah Alves Coelho; Lonardoni, Maria Valdrinez Campana; Demarchi, Izabel Galhardo
Cutaneous leishmaniasis is caused by different species of Leishmania. In domestic animals such as dogs and cats, the diagnostic consists of clinical, epidemiological and serological tests, which changes among countries all around the world. Because of this diversity in the methods selected, we propose this systematic literature review to identify the methods of laboratory diagnosis used to detect cutaneous leishmaniasis in domestic dogs and cats in the Americas. Articles published in the last 5 years were searched in PubMed, ISI Web of Science, LILACS and Scielo, and we selected 10 papers about cutaneous leishmaniasis in dogs and cats in the Americas. In Brazil, often the indirect immunofluorescence and enzyme immunoassay (ELISA) have been applied. Other countries like United States and Mexico have been using antigenic fractions for antibodies detections by Western blot. ELISA and Western blot showed a higher sensitivity and efficacy in the detection of leishmaniasis. Analysis of sensibility and specificity of the methods was rarely used. Although confirmatory to leishmaniasis, direct methods for parasites detection and polymerase chain reaction showed low positivity in disease detection. We suggested that more than one method should be used for the detection of feline and canine leishmaniasis. Serological methods such as Western blot and enzyme immunoassay have a high efficacy in the diagnosis of this disease. PMID:26734869
ICALEO '90 - Optical methods in flow and particle diagnostics; Proceedings of the Meeting, Boston, MA, Nov. 4-9, 1990
Not Available
Attention is given to multiple species CARS in turbulent jet flames, simultaneous measurements of temperature and density in air flows using UV laser spectroscopy, a combination of multispecies Raman scattering with molecular fluorescence, planar laser-induced fluorescence diagnostics for large scale test facilities, evidence of local stagnation in supersonic mixing layers using 1D laser Rayleigh and Raman scattering, vorticity field measurements using laser induced photochemical anemometry, and combustion diagnostics by 2D laser induced fluorescence using tunable excimer lasers. Attention is also given to a single laser apparatus for writing patterns into unseeded air, single exposure double frame particle image velocimeters, holographic recording of 3D flow configurations for particle image velocimetry, and flow field diagnostics by spectrally filtered Rayleigh scattering.
Essential veterinary education in modern molecular tools for the detection of disease: what veterinarians will need to know about genomics and molecular biology and diagnostics (including bioterrorist weapons) in 2025.
de Lamballerie, X
Future veterinary education programmes in microbiology will undoubtedly include an increasing emphasis on new and existing molecular tools. They should also, however, provide veterinarians with a comprehensive and clear understanding of the types of results that can be obtained using a particular approach (for example, specific diagnostic procedures as against open diagnostic procedures, phenotypic versus genotypic characterisation, etc.). Furthermore, students should gain a sound knowledge of which type of test is the most appropriate in a given clinical or epidemiological situation, and what conclusions can or cannot be drawn from the results. Consequently, each veterinary curriculum should focus on the following items: the principles of molecular biology and genomics; the detection of disease and characteristics of molecular tests; the principles of micro-organism taxonomy, sequence comparison and molecular epidemiology and their applications (such as: taxonomic identification, epidemiological survey, genetic evolution and the traceability of strains); and the role of the veterinarian in the field of zoonoses and human public health. PMID:20128476
Methods of Genome Engineering: a New Era of Molecular Biology.
Chugunova, A A; Dontsova, O A; Sergiev, P V
Genome sequencing now progressing much faster than our understanding of the majority of gene functions. Studies of physiological functions of various genes would not be possible without the ability to manipulate the genome. Methods of genome engineering can now be used to inactivate a gene to study consequences, introduce heterologous genes into the genome for scientific and biotechnology applications, create genes coding for fusion proteins to study gene expression, protein localization, and molecular interactions, and to develop animal models of human diseases to find appropriate treatment. Finally, genome engineering might present the possibility to cure hereditary diseases. In this review, we discuss and compare the most important methods for gene inactivation and editing, as well as methods for incorporation of heterologous genes into the genome. PMID:27449613
Applications of the molecular dynamics flexible fitting method.
Trabuco, Leonardo G; Schreiner, Eduard; Gumbart, James; Hsin, Jen; Villa, Elizabeth; Schulten, Klaus
In recent years, cryo-electron microscopy (cryo-EM) has established itself as a key method in structural biology, permitting the structural characterization of large biomolecular complexes in various functional states. The data obtained through single-particle cryo-EM has recently seen a leap in resolution thanks to landmark advances in experimental and computational techniques, resulting in sub-nanometer resolution structures being obtained routinely. The remaining gap between these data and revealing the mechanisms of molecular function can be closed through hybrid modeling tools that incorporate known atomic structures into the cryo-EM data. One such tool, molecular dynamics flexible fitting (MDFF), uses molecular dynamics simulations to combine structures from X-ray crystallography with cryo-EM density maps to derive atomic models of large biomolecular complexes. The structures furnished by MDFF can be used subsequently in computational investigations aimed at revealing the dynamics of the complexes under study. In the present work, recent applications of MDFF are presented, including the interpretation of cryo-EM data of the ribosome at different stages of translation and the structure of a membrane-curvature-inducing photosynthetic complex. PMID:20932910
Interobserver Reliability of Four Diagnostic Methods Using Traditional Korean Medicine for Stroke Patients
Lee, Ju Ah; Kang, Byoung-Kab; Alraek, Terje
Objective. The aim of this study is to evaluate the consistency of pattern identification (PI), a set of diagnostic indicators used by traditional Korean medicine (TKM) clinicians. Methods. A total of 168 stroke patients who were admitted into oriental medical university hospitals from June 2012 through January 2013 were included in the study. Using the PI indicators, each patient was independently diagnosed by two experts from the same department. Interobserver consistency was assessed by simple percentage agreement as well as by kappa and AC1 statistics. Results. Interobserver agreement on the PI indicators (for all patients) was generally high: pulse diagnosis signs (AC1 = 0.66–0.89); inspection signs (AC1 = 0.66–0.95); listening/smelling signs (AC1 = 0.67–0.88); and inquiry signs (AC1 = 0.62–0.94). Conclusion. In four examinations, there was moderate agreement between the clinicians on the PI indicators. To improve clinician consistency (e.g., in the diagnostic criteria used), it is necessary to analyze the reasons for inconsistency and to improve clinician training. PMID:25574181
Molecular Rayleigh Scattering Diagnostic for Measurement of High Frequency Temperature Fluctuations
NASA Technical Reports Server (NTRS)
Mielke, Amy F.; Elam, Kristie A.
A novel technique for measurement of high frequency temperature fluctuations in unseeded gas flows using molecular Rayleigh scattering is investigated. The spectrum of laser light scattered from molecules in a gas flow is resolved using a Fabry-Perot interferometer. The width of the spectral peak is broadened by thermal motion of the molecules and hence is related to gas temperature. The interference fringe pattern containing spectral information is divided into four concentric regions using a series of mirrors angled with respect to one another. Light from each of these regions is directed towards photomultiplier tubes and sampled at 10 kHz using photon counting electronics. Monitoring the relative change in intensity within each region allows measurement of gas temperature. Independently monitoring the total scattered intensity provides a measure of gas density. This technique also has the potential to simultaneously measure a single component of flow velocity by monitoring the spectral peak location. Measurements of gas temperature and density are demonstrated using a low speed heated air jet surrounded by an unheated air co-flow. Mean values of temperature and density are shown for radial scans across the jet flow at a fixed axial distance from the jet exit plane. Power spectra of temperature and density fluctuations at several locations in the jet are also shown. The instantaneous measurements have fairly high uncertainty; however, long data records provide highly accurate statistically quantities, which include power spectra. Mean temperatures are compared with thermocouple measurements as well as the temperatures derived from independent density measurements. The accuracy for mean temperature measurements was +/- 7 K.
Diagnostic and molecular evaluation of three iridovirus-associated salamander mortality events
Docherty, D.E.; Meteyer, C.U.; Wang, Jingyuan; Mao, J.; Case, S.T.; Chinchar, V.G.
In 1998 viruses were isolated from tiger salamander larvae (Ambystoma tigrinum diaboli and A. tigrinum melanostictum) involved in North Dakota and Utah (USA) mortality events and spotted salamander (A. maculatum) larvae in a third event in Maine (USA). Although sympatric caudates and anurans were present at all three sites only ambystomid larvae appeared to be affected. Mortality at the North Dakota site was in the thousands while at the Utah and Maine sites mortality was in the hundreds. Sick larvae were lethargic and slow moving. They swam in circles with obvious buoyancy problems and were unable to remain upright. On the ventral surface, near the gills and hind limbs, red spots or swollen areas were noted. Necropsy findings included: hemorrhages and ulceration of the skin, subcutaneous and intramuscular edema, swollen and pale livers with multifocal hemorrhage, and distended fluid-filled intestines with areas of hemorrhage. Light microscopy revealed intracytoplasmic inclusions, suggestive of a viral infection, in a variety of organs. Electron microscopy of ultra thin sections of the same tissues revealed iridovirus-like particles within the inclusions. These viruses were isolated from a variety of organs, indicating a systemic infection. Representative viral isolates from the three mortality events were characterized using molecular assays. Characterization confirmed that the viral isolates were iridoviruses and that the two tiger salamander isolates were similar and could be distinguished from the spotted salamander isolate. The spotted salamander isolate was similar to frog virus 3, the type species of the genus Ranavirus, while the tiger salamander isolates were not. These data indicate that different species of salamanders can become infected and die in association with different iridoviruses. Challenge assays are required to determine the fish and amphibian host range of these isolates and to assess the susceptibility of tiger and spotted salamanders to
A diagnostic for determining the quality of single-reference electron correlation methods
NASA Technical Reports Server (NTRS)
Lee, Timothy J.; Taylor, Peter R.
It was recently proposed that the Euclidian norm of the t(sub 1) vector of the coupled cluster wave function (normalized by the number of electrons included in the correlation procedure) could be used to determine whether a single-reference-based electron correlation procedure is appopriate. This diagnostic, T(sub 1) is defined for use with self-consistent-field molecular orbitals and is invariant to the same orbital rotations as the coupled cluster energy. T(sub 1) is investigated for several different chemical systems which exhibit a range of multireference behavior, and is shown to be an excellent measure of the importance of non-dynamical electron correlation and is far superior to C(sub 0) from a singles and doubles configuration interaction wave function. It is further suggested that when the aim is to recover a large fraction of the dynamical electron correlation energy, a large T(sub 1) (i.e., greater than 0.02) probably indicates the need for a multireference electron correlation procedure.
A diagnostic for determining the quality of single-reference electron correlation methods
NASA Technical Reports Server (NTRS)
Lee, Timothy J.; Taylor, Peter R.
It was recently proposed that the Euclidian norm of the t sub 1 vector of the coupled cluster wave function (normalized by the number of electrons included in the correlation procedure) could be used to determine whether a single-reference-based electron correlation procedure is appropriate. This diagnostic, T sub 1, is defined for use with self consistent field molecular orbitals and is invariant to the same orbital rotations as the coupled cluster energy. T sub 1 is investigated for several different chemical systems which exhibit a range of multireference behavior, and is shown to be an excellent measure of the importance of nondynamical electron correlation and is far superior to C sub 0 from a singles and doubles configuration interaction wave function. It is further suggested that when the aim is to recover a large fraction of the dynamical electron correlation energy, a large T sub 1 (i.e., greater than 0.02) probably indicates the need for a multireference electron correlation procedure.
Short-time Chebyshev wave packet method for molecular photoionization
NASA Astrophysics Data System (ADS)
Sun, Zhaopeng; Zheng, Yujun
In this letter we present the extended usage of short-time Chebyshev wave packet method in the laser induced molecular photoionization dynamics. In our extension, the polynomial expansion of the exponential in the time evolution operator, the Hamiltonian operator can act on the wave packet directly which neatly avoids the matrix diagonalization. This propagation scheme is of obvious advantages when the dynamical system has large Hamiltonian matrix. Computational simulations are performed for the calculation of photoelectronic distributions from intense short pulse ionization of K2 and NaI which represent the Born-Oppenheimer (BO) model and Non-BO one, respectively.
Application of Molecular Diagnostics in Primary Detection of ESBL Directly from Clinical Specimens.
Sittová, Martina; Röderová, Magdaléna; Dendis, Miloš; Hricová, Kristýna; Pudová, Vendula; Horváth, Radek; Růžička, Filip; Dosoudilová, Šárka; Kolář, Milan
The infections caused by extended-spectrum β-lactamase (ESBL)-producing organisms are associated with increased mortality. The real-time polymerase chain reaction (PCR) method, which enables detection of ESBLs directly from patients' clinical material, was developed. This study focused on blaCTX-M and blaSHV determination in endotracheal aspirates. Each sample was identified with standard microbiological procedures and simultaneously analyzed for the presence of nucleic acids, which encode CTX-M and SHV ESBL enzymes using real-time PCR. A total of 341 samples were investigated. In the set, 27 ESBL-positive samples were identified by phenotypic methods, while 60 positive samples were identified by the PCR method. Of the 60 PCR-positive samples, 58 were positive for the blaCTX-M. In two samples, the ESBL blaSHV-ESBL gene was detected. One phenotypically positive sample was PCR negative. The real-time PCR assay does not require a cultivation step and therefore enables detection of ESBL in 6 hours. The rapid method is necessary for early and adequate antimicrobial treatment. PMID:25588196
[Applications of the Fragment Molecular Orbital Method in Drug Discovery].
Ishikawa, Takeshi
Recently, ab initio quantum mechanical calculations have been applied to large molecules, including biomolecular systems. The fragment molecular orbital (FMO) method is one of the most efficient approaches for the quantum mechanical investigation of such molecules. In the FMO method, dividing a target molecule into small fragments reduces computational effort. The clear definition of inter-fragment interaction energy (IFIE) as an expression of total energy is another valuable feature of the FMO method because it provides the ability to analyze interactions in biomolecules. Thus, the FMO method is expected to be useful for drug discovery. This study demonstrates applications of the FMO method related to drug discovery. First, IFIE, according to FMO calculations, was used in the optimization of drug candidates for the development of anti-prion compounds. The second example involved interaction analysis of the human immunodeficiency virus type 1 (HIV-1) protease and a drug compound that used a novel analytical method for dispersion interaction, i.e., fragment interaction analysis based on LMP2 (FILM). PMID:26725679
[On the method of express-diagnostics of thyroid gland dysfunctions].
Abazova, Z Kh; Baĭsiev, A Kh; Kumykov, V K; Efendieva, M K
The method of express-diagnostics of thyroid diseases on a degree of moisture of the skin integument which is one of clinical attributes of hypothyroidism (a skin is dry, shelled, with sites of keratinization) and hyperthyroidism at which the return picture is observed, i.e. the (increased humidity of a skin is offered. At the same time as a parameter describing a degree of moisture of skin is a relative humidity of the air environment which are taking place above the skin integument in conditions of thermodynamic equilibrium. The instrument is a hermetic glass in which the sensor of humidity is mounted. Studies on the definition of threshold levels of parameter for several groups of patients with clinically confirmed diagnoses of diseases of a thyroid are carried out. PMID:16106951
Microvascular resistance in essential hypertension and flowmetry as a diagnostic method
NASA Astrophysics Data System (ADS)
Lukjanov, Valdimir F.
New Doppler-Laser flowmetry diagnostic test of functional condition of microcirculation was worked out of find precapillar and postcapillar resistance. Flowmetry was used to measure vasomotion and blood flow after arterial compression, decompression and venous hyperemia were held. Patients of essential hypertension were examined with the help of Doppler-Laser Flowmetry, optical photometry (540 nm). Precapillar resistance included next basis parameters: vasomotion with high frequency (10-16 per/min) and low amplitude, latent time after decompression, large postocclusive reactive hyperemia, absent venous hyperemia. Postcapillar resistance included next basis parameters: vasomotion with low frequency (4-8 per/min) and high amplitude, paradoxical hyperemia in arterial compression, little or absent postocclusive reactive hyperemia, large venous hyperemia. This test-method was applied to select patogenetic treatment of essential hypertension.
Embedded diagnostic, prognostic, and health management system and method for a humanoid robot
NASA Technical Reports Server (NTRS)
Barajas, Leandro G. (Inventor); Sanders, Adam M (Inventor); Reiland, Matthew J (Inventor); Strawser, Philip A (Inventor)
A robotic system includes a humanoid robot with multiple compliant joints, each moveable using one or more of the actuators, and having sensors for measuring control and feedback data. A distributed controller controls the joints and other integrated system components over multiple high-speed communication networks. Diagnostic, prognostic, and health management (DPHM) modules are embedded within the robot at the various control levels. Each DPHM module measures, controls, and records DPHM data for the respective control level/connected device in a location that is accessible over the networks or via an external device. A method of controlling the robot includes embedding a plurality of the DPHM modules within multiple control levels of the distributed controller, using the DPHM modules to measure DPHM data within each of the control levels, and recording the DPHM data in a location that is accessible over at least one of the high-speed communication networks.
The Scientific Method, Diagnostic Bayes, and How to Detect Epistemic Errors
NASA Astrophysics Data System (ADS)
Vrugt, J. A.
In the past decades, Bayesian methods have found widespread application and use in environmental systems modeling. Bayes theorem states that the posterior probability, P(H|D) of a hypothesis, H is proportional to the product of the prior probability, P(H) of this hypothesis and the likelihood, L(H|hat{D}) of the same hypothesis given the new/incoming observations, &hat; {D}. In science and engineering, H often constitutes some numerical simulation model, D = F(x,.) which summarizes using algebraic, empirical, and differential equations, state variables and fluxes, all our theoretical and/or practical knowledge of the system of interest, and x are the d unknown parameters which are subject to inference using some data, &hat; {D} of the observed system response. The Bayesian approach is intimately related to the scientific method and uses an iterative cycle of hypothesis formulation (model), experimentation and data collection, and theory/hypothesis refinement to elucidate the rules that govern the natural world. Unfortunately, model refinement has proven to be very difficult in large part because of the poor diagnostic power of residual based likelihood functions tep{gupta2008}. This has inspired te{vrugt2013} to advocate the use of 'likelihood-free' inference using approximate Bayesian computation (ABC). This approach uses one or more summary statistics, S(&hat; {D}) of the original data, &hat; {D} designed ideally to be sensitive only to one particular process in the model. Any mismatch between the observed and simulated summary metrics is then easily linked to a specific model component. A recurrent issue with the application of ABC is self-sufficiency of the summary statistics. In theory, S(.) should contain as much information as the original data itself, yet complex systems rarely admit sufficient statistics. In this article, we propose to combine the ideas of ABC and regular Bayesian inference to guarantee that no information is lost in diagnostic model
Steered Molecular Dynamics Methods Applied to Enzyme Mechanism and Energetics.
Ramírez, C L; Martí, M A; Roitberg, A E
One of the main goals of chemistry is to understand the underlying principles of chemical reactions, in terms of both its reaction mechanism and the thermodynamics that govern it. Using hybrid quantum mechanics/molecular mechanics (QM/MM)-based methods in combination with a biased sampling scheme, it is possible to simulate chemical reactions occurring inside complex environments such as an enzyme, or aqueous solution, and determining the corresponding free energy profile, which provides direct comparison with experimental determined kinetic and equilibrium parameters. Among the most promising biasing schemes is the multiple steered molecular dynamics method, which in combination with Jarzynski's Relationship (JR) allows obtaining the equilibrium free energy profile, from a finite set of nonequilibrium reactive trajectories by exponentially averaging the individual work profiles. However, obtaining statistically converged and accurate profiles is far from easy and may result in increased computational cost if the selected steering speed and number of trajectories are inappropriately chosen. In this small review, using the extensively studied chorismate to prephenate conversion reaction, we first present a systematic study of how key parameters such as pulling speed, number of trajectories, and reaction progress are related to the resulting work distributions and in turn the accuracy of the free energy obtained with JR. Second, and in the context of QM/MM strategies, we introduce the Hybrid Differential Relaxation Algorithm, and show how it allows obtaining more accurate free energy profiles using faster pulling speeds and smaller number of trajectories and thus smaller computational cost. PMID:27497165
Biogeographical characterization of Saccharomyces cerevisiae wine yeast by molecular methods
Tofalo, Rosanna; Perpetuini, Giorgia; Schirone, Maria; Fasoli, Giuseppe; Aguzzi, Irene; Corsetti, Aldo; Suzzi, Giovanna
Biogeography is the descriptive and explanatory study of spatial patterns and processes involved in the distribution of biodiversity. Without biogeography, it would be difficult to study the diversity of microorganisms because there would be no way to visualize patterns in variation. Saccharomyces cerevisiae, "the wine yeast," is the most important species involved in alcoholic fermentation, and in vineyard ecosystems, it follows the principle of "everything is everywhere." Agricultural practices such as farming (organic versus conventional) and floor management systems have selected different populations within this species that are phylogenetically distinct. In fact, recent ecological and geographic studies highlighted that unique strains are associated with particular grape varieties in specific geographical locations. These studies also highlighted that significant diversity and regional character, or 'terroir,' have been introduced into the winemaking process via this association. This diversity of wild strains preserves typicity, the high quality, and the unique flavor of wines. Recently, different molecular methods were developed to study population dynamics of S. cerevisiae strains in both vineyards and wineries. In this review, we will provide an update on the current molecular methods used to reveal the geographical distribution of S. cerevisiae wine yeast. PMID:23805132
Molecular Dynamics and Energy Minimization Based on Embedded Atom Method
This program performs atomic scale computer simulations of the structure and dynamics of metallic system using energetices based on the Embedded Atom Method. The program performs two types of calculations. First, it performs local energy minimization of all atomic positions to determine ground state and saddle point energies and structures. Second, it performs molecular dynamics simulations to determine thermodynamics or miscroscopic dynamics of the system. In both cases, various constraints can be applied to themore » system. The volume of the system can be varied automatically to achieve any desired external pressure. The temperature in molecular dynamics simulations can be controlled by a variety of methods. Further, the temperature control can be applied either to the entire system or just a subset of the atoms that would act as a thermal source/sink. The motion of one or more of the atoms can be constrained to either simulate the effects of bulk boundary conditions or to facilitate the determination of saddle point configurations. The simulations are performed with periodic boundary conditions.« less
Unveiling PHA-storing populations using molecular methods.
Queirós, Diogo; Lemos, Paulo C; Rossetti, Simona; Serafim, Luísa S
Enrichment of mixed microbial cultures (MMCs) in polyhydroxyalkanoate (PHA)-storing microorganisms must take place to develop a successful PHA production process. Moreover, throughout the operational period of a MMC system, the population needs to be checked in order to understand the changes in the performance that eventually occurred. For these reasons, it is necessary to monitor the population evolution, in order to identify the different groups of microorganisms and relate them with the storage capacity and kinetics of the MMC. Regarding this particular process, several culture-independent molecular techniques were already applied, with the use of hybridization techniques such fluorescence in situ hybridization and also PCR-based methods like denaturing gradient gel electrophoresis, terminal restriction fragment length polymorphism, pyrosequencing, and quantitative PCR standing out. This review intends, thus, to look at the molecular methods currently applied in monitoring the PHA-storing population evolution and how they can be combined with the evolutionary engineering step in order to optimize the overall process. PMID:26428237
(Molecular understanding of mutagenicity using potential energy methods)
Broyde, S.
The objective of our work has been, for many year, to elucidate on a molecular level at atomic resolution the structures of DNAs modified by highly mutagenic polycyclic aromatic amines and hydrocarbons, and their less mutagenic chemically related analogs and unmodified DNAs, as controls. The ultimate purpose of this undertaking is to obtain an understanding of the relationship DNA structures and mutagenicity. Our methods for elucidating structures are computational, but we keep in close contact with experimental developments, and have, very recently, been able to incorporate the first experimental information from NMR studies by other workers in our calculations. The specific computational methods we employ are minimized potential energy calculations using the torsion angle space program DUPLEX, developed and written by Dr. Brain Hingerty to yield static views. Molecular dynamics simulations of the important static structures with full solvation and salt are carried out with the program AMBER; this yields mobile views in a milieu that best mimics the natural environment of the cell. In addition, we have been developing new strategies for searching conformation space and building DNA duplexes from favored subunit structures. 30 refs., 12 figs.
eSensor: an electrochemical detection-based DNA microarray technology enabling sample-to-answer molecular diagnostics
NASA Astrophysics Data System (ADS)
Liu, Robin H.; Longiaru, Mathew
DNA microarrays are becoming a widespread tool used in life science and drug screening due to its many benefits of miniaturization and integration. Microarrays permit a highly multiplexed DNA analysis. Recently, the development of new detection methods and simplified methodologies has rapidly expanded the use of microarray technologies from predominantly gene expression analysis into the arena of diagnostics. Osmetech's eSensor® is an electrochemical detection platform based on a low-to- medium density DNA hybridization array on a cost-effective printed circuit board substrate. eSensor® has been cleared by FDA for Warfarin sensitivity test and Cystic Fibrosis Carrier Detection. Other genetic-based diagnostic and infectious disease detection tests are under development. The eSensor® platform eliminates the need for an expensive laser-based optical system and fluorescent reagents. It allows one to perform hybridization and detection in a single and small instrument without any fluidic processing and handling. Furthermore, the eSensor® platform is readily adaptable to on-chip sample-to-answer genetic analyses using microfluidics technology. The eSensor® platform provides a cost-effective solution to direct sample-to-answer genetic analysis, and thus have a potential impact in the fields of point-of-care genetic analysis, environmental testing, and biological warfare agent detection.
A Sweet Spot for Molecular Diagnostics: Coupling Isothermal Amplification and Strand Exchange Circuits to Glucometers.
Du, Yan; Hughes, Randall A; Bhadra, Sanchita; Jiang, Yu Sherry; Ellington, Andrew D; Li, Bingling
Strand exchange nucleic acid circuitry can be used to transduce isothermal nucleic acid amplification products into signals that can be readable on an off-the-shelf glucometer. Loop-mediated isothermal amplification (LAMP) is limited by the accumulation of non-specific products, but nucleic acid circuitry can be used to probe and distinguish specific amplicons. By combining this high temperature isothermal amplification method with a thermostable invertase, we can directly transduce Middle-East respiratory syndrome coronavirus and Zaire Ebolavirus templates into glucose signals, with a sensitivity as low as 20-100 copies/μl, equating to atto-molar (or low zepto-mole). Virus from cell lysates and synthetic templates could be readily amplified and detected even in sputum or saliva. An OR gate that coordinately triggered on viral amplicons further guaranteed fail-safe virus detection. The method describes has potential for accelerating point-of-care applications, in that biological samples could be applied to a transducer that would then directly interface with an off-the-shelf, approved medical device. PMID:26050646
A Sweet Spot for Molecular Diagnostics: Coupling Isothermal Amplification and Strand Exchange Circuits to Glucometers
NASA Astrophysics Data System (ADS)
Du, Yan; Hughes, Randall A.; Bhadra, Sanchita; Jiang, Yu Sherry; Ellington, Andrew D.; Li, Bingling
Strand exchange nucleic acid circuitry can be used to transduce isothermal nucleic acid amplification products into signals that can be readable on an off-the-shelf glucometer. Loop-mediated isothermal amplification (LAMP) is limited by the accumulation of non-specific products, but nucleic acid circuitry can be used to probe and distinguish specific amplicons. By combining this high temperature isothermal amplification method with a thermostable invertase, we can directly transduce Middle-East respiratory syndrome coronavirus and Zaire Ebolavirus templates into glucose signals, with a sensitivity as low as 20-100 copies/μl, equating to atto-molar (or low zepto-mole). Virus from cell lysates and synthetic templates could be readily amplified and detected even in sputum or saliva. An OR gate that coordinately triggered on viral amplicons further guaranteed fail-safe virus detection. The method describes has potential for accelerating point-of-care applications, in that biological samples could be applied to a transducer that would then directly interface with an off-the-shelf, approved medical device.
A Sweet Spot for Molecular Diagnostics: Coupling Isothermal Amplification and Strand Exchange Circuits to Glucometers
Du, Yan; Hughes, Randall A.; Bhadra, Sanchita; Jiang, Yu Sherry; Ellington, Andrew D.; Li, Bingling
Strand exchange nucleic acid circuitry can be used to transduce isothermal nucleic acid amplification products into signals that can be readable on an off-the-shelf glucometer. Loop-mediated isothermal amplification (LAMP) is limited by the accumulation of non-specific products, but nucleic acid circuitry can be used to probe and distinguish specific amplicons. By combining this high temperature isothermal amplification method with a thermostable invertase, we can directly transduce Middle-East respiratory syndrome coronavirus and Zaire Ebolavirus templates into glucose signals, with a sensitivity as low as 20–100 copies/μl, equating to atto-molar (or low zepto-mole). Virus from cell lysates and synthetic templates could be readily amplified and detected even in sputum or saliva. An OR gate that coordinately triggered on viral amplicons further guaranteed fail-safe virus detection. The method describes has potential for accelerating point-of-care applications, in that biological samples could be applied to a transducer that would then directly interface with an off-the-shelf, approved medical device. PMID:26050646
Modular development of a prototype point of care molecular diagnostic platform for sexually transmitted infections.
Branavan, Manoharanehru; Mackay, Ruth E; Craw, Pascal; Naveenathayalan, Angel; Ahern, Jeremy C; Sivanesan, Tulasi; Hudson, Chris; Stead, Thomas; Kremer, Jessica; Garg, Neha; Baker, Mark; Sadiq, Syed T; Balachandran, Wamadeva
This paper presents the design of a modular point of care test platform that integrates a proprietary sample collection device directly with a microfluidic cartridge. Cell lysis, within the cartridge, is conducted using a chemical method and nucleic acid purification is done on an activated cellulose membrane. The microfluidic device incorporates passive mixing of the lysis-binding buffers and sample using a serpentine channel. Results have shown extraction efficiencies for this new membrane of 69% and 57% compared to the commercial Qiagen extraction method of 85% and 59.4% for 0.1ng/µL and 100ng/µL salmon sperm DNA respectively spiked in phosphate buffered solution. Extraction experiments using the serpentine passive mixer cartridges incorporating lysis and nucleic acid purification showed extraction efficiency around 80% of the commercial Qiagen kit. Isothermal amplification was conducted using thermophillic helicase dependant amplification and recombinase polymerase amplification. A low cost benchtop real-time isothermal amplification platform has been developed capable of running six amplifications simultaneously. Results show that the platform is capable of detecting 1.32×10(6) of sample DNA through thermophillic helicase dependant amplification and 1×10(5) copy numbers Chlamydia trachomatis genomic DNA within 10min through recombinase polymerase nucleic acid amplification tests. PMID:27238759
A new method for tracking organ motion on diagnostic ultrasound images
Kubota, Yoshiki Matsumura, Akihiko; Fukahori, Mai; Minohara, Shin-ichi; Yasuda, Shigeo; Nagahashi, Hiroshi
Purpose: Respiratory-gated irradiation is effective in reducing the margins of a target in the case of abdominal organs, such as the liver, that change their position as a result of respiratory motion. However, existing technologies are incapable of directly measuring organ motion in real-time during radiation beam delivery. Hence, the authors proposed a novel quantitative organ motion tracking method involving the use of diagnostic ultrasound images; it is noninvasive and does not entail radiation exposure. In the present study, the authors have prospectively evaluated this proposed method. Methods: The method involved real-time processing of clinical ultrasound imaging data rather than organ monitoring; it comprised a three-dimensional ultrasound device, a respiratory sensing system, and two PCs for data storage and analysis. The study was designed to evaluate the effectiveness of the proposed method by tracking the gallbladder in one subject and a liver vein in another subject. To track a moving target organ, the method involved the control of a region of interest (ROI) that delineated the target. A tracking algorithm was used to control the ROI, and a large number of feature points and an error correction algorithm were used to achieve long-term tracking of the target. Tracking accuracy was assessed in terms of how well the ROI matched the center of the target. Results: The effectiveness of using a large number of feature points and the error correction algorithm in the proposed method was verified by comparing it with two simple tracking methods. The ROI could capture the center of the target for about 5 min in a cross-sectional image with changing position. Indeed, using the proposed method, it was possible to accurately track a target with a center deviation of 1.54 ± 0.9 mm. The computing time for one frame image using our proposed method was 8 ms. It is expected that it would be possible to track any soft-tissue organ or tumor with large deformations and
A two year BTV-8 vaccination follow up: molecular diagnostics and assessment of humoral and cellular immune reactions.
Hund, Alexandra; Gollnick, Nicole; Sauter-Louis, Carola; Neubauer-Juric, Antonie; Lahm, Harald; Büttner, Mathias
The compulsory vaccination campaign against Bluetongue virus serotype eight (BTV-8) in Germany was exercised in the state of Bavaria using three commercial monovalent inactivated vaccines given provisional marketing authorisation for emergency use. In eleven Bavarian farms representing a cross sectional area of the state the immune reactions of sheep and cattle were followed over a two year period (2008-2009) using cELISA, a serum neutralisation test (SNT) and interferon gamma (IFN-γ) ELISPOT. For molecular diagnostics of BTV genome presence two recommended real time quantitative RT-PCR protocols were applied. The recommended vaccination scheme led to low or even undetectable antibody titers (ELISA) in serum samples of both cattle and sheep. A fourfold increase of the vaccine dose in cattle, however, induced higher ELISA titers and virus neutralising antibodies. Accordingly, repeated vaccination in sheep caused an increase in ELISA-antibody titers. BTV-8 neutralising antibodies occurred in most animals only after multiple vaccinations in the second year of the campaign. The secretion of interferon gamma (IFN-γ) in ELISPOT after in vitro re-stimulation of PBMC of BTV-8 vaccinated animals with BTV was evaluated in the field for the first time. Sera of BTV-8 infected or vaccinated animals neutralising BTV-8 could also neutralise an Italian BTV serotype 1 cell culture adapted strain and PBMC of such animals secreted IFN-γ when stimulated with BTV-1. PMID:21840139
Diagnostic Methods of Helicobacter pylori Infection for Epidemiological Studies: Critical Importance of Indirect Test Validation.
Miftahussurur, Muhammad; Yamaoka, Yoshio
Among the methods developed to detect H. pylori infection, determining the gold standard remains debatable, especially for epidemiological studies. Due to the decreasing sensitivity of direct diagnostic tests (histopathology and/or immunohistochemistry [IHC], rapid urease test [RUT], and culture), several indirect tests, including antibody-based tests (serology and urine test), urea breath test (UBT), and stool antigen test (SAT) have been developed to diagnose H. pylori infection. Among the indirect tests, UBT and SAT became the best methods to determine active infection. While antibody-based tests, especially serology, are widely available and relatively sensitive, their specificity is low. Guidelines indicated that no single test can be considered as the gold standard for the diagnosis of H. pylori infection and that one should consider the method's advantages and disadvantages. Based on four epidemiological studies, culture and RUT present a sensitivity of 74.2-90.8% and 83.3-86.9% and a specificity of 97.7-98.8% and 95.1-97.2%, respectively, when using IHC as a gold standard. The sensitivity of serology is quite high, but that of the urine test was lower compared with that of the other methods. Thus, indirect test validation is important although some commercial kits propose universal cut-off values. PMID:26904678
Hybrid digital signal processing and neural networks for automated diagnostics using NDE methods
Upadhyaya, B.R.; Yan, W.
The primary purpose of the current research was to develop an integrated approach by combining information compression methods and artificial neural networks for the monitoring of plant components using nondestructive examination data. Specifically, data from eddy current inspection of heat exchanger tubing were utilized to evaluate this technology. The focus of the research was to develop and test various data compression methods (for eddy current data) and the performance of different neural network paradigms for defect classification and defect parameter estimation. Feedforward, fully-connected neural networks, that use the back-propagation algorithm for network training, were implemented for defect classification and defect parameter estimation using a modular network architecture. A large eddy current tube inspection database was acquired from the Metals and Ceramics Division of ORNL. These data were used to study the performance of artificial neural networks for defect type classification and for estimating defect parameters. A PC-based data preprocessing and display program was also developed as part of an expert system for data management and decision making. The results of the analysis showed that for effective (low-error) defect classification and estimation of parameters, it is necessary to identify proper feature vectors using different data representation methods. The integration of data compression and artificial neural networks for information processing was established as an effective technique for automation of diagnostics using nondestructive examination methods.
Diagnostic Methods of Helicobacter pylori Infection for Epidemiological Studies: Critical Importance of Indirect Test Validation
Miftahussurur, Muhammad; Yamaoka, Yoshio
Among the methods developed to detect H. pylori infection, determining the gold standard remains debatable, especially for epidemiological studies. Due to the decreasing sensitivity of direct diagnostic tests (histopathology and/or immunohistochemistry [IHC], rapid urease test [RUT], and culture), several indirect tests, including antibody-based tests (serology and urine test), urea breath test (UBT), and stool antigen test (SAT) have been developed to diagnose H. pylori infection. Among the indirect tests, UBT and SAT became the best methods to determine active infection. While antibody-based tests, especially serology, are widely available and relatively sensitive, their specificity is low. Guidelines indicated that no single test can be considered as the gold standard for the diagnosis of H. pylori infection and that one should consider the method's advantages and disadvantages. Based on four epidemiological studies, culture and RUT present a sensitivity of 74.2–90.8% and 83.3–86.9% and a specificity of 97.7–98.8% and 95.1–97.2%, respectively, when using IHC as a gold standard. The sensitivity of serology is quite high, but that of the urine test was lower compared with that of the other methods. Thus, indirect test validation is important although some commercial kits propose universal cut-off values. PMID:26904678
The state of cell block variation and satisfaction in the era of molecular diagnostics and personalized medicine
Crapanzano, John P.; Heymann, Jonas J.; Monaco, Sara; Nassar, Aziza; Saqi, Anjali
Background: In the recent past, algorithms and recommendations to standardize the morphological, immunohistochemical and molecular classification of lung cancers on cytology specimens have been proposed, and several organizations have recommended cell blocks (CBs) as the preferred modality for molecular testing. Based on the literature, there are several different techniques available for CB preparation-suggesting that there is no standard. The aim of this study was to conduct a survey of CB preparation techniques utilized in various practice settings and analyze current issues, if any. Materials and Methods: A single E-mail with a link to an electronic survey was distributed to members of the American Society of Cytopathology and other pathologists. Questions pertaining to the participants' practice setting and CBs-volume, method, quality and satisfaction-were included. Results: Of 95 respondents, 90/95 (94%) completed the survey and comprise the study group. Most participants practice in a community hospital/private practice (44%) or academic center (41%). On average, 14 CBs (range 0-50; median 10) are prepared by a laboratory daily. Over 10 methods are utilized: Plasma thrombin (33%), HistoGel (27%), Cellient automated cell block system (8%) and others (31%) respectively. Forty of 90 (44%) respondents are either unsatisfied or sometimes satisfied with their CB quality, with low-cellular yield being the leading cause of dissatisfaction. There was no statistical significance between the three most common CB preparation methods and satisfaction with quality. Discussion: Many are dissatisfied with their current method of CB preparation, and there is no consistent method to prepare CBs. In today's era of personalized medicine with an increasing array of molecular tests being applied to cytological specimens, there is a need for a standardized protocol for CB optimization to enhance cellularity. PMID:24799951
A rapid and precise diagnostic method for detecting the Pinewood nematode Bursaphelenchus xylophilus by loop-mediated isothermal amplification.
Kikuchi, Taisei; Aikawa, Takuya; Oeda, Yuka; Karim, Nurul; Kanzaki, Natsumi
ABSTRACT Bursaphelenchus xylophilus is the causal agent of pine wilt disease, which is a major forest disease in Japan, Korea, China, Taiwan, and Portugal. A diagnostic method which is rapid, precise, and simple could greatly help the proper management of this disease. Here, we present a novel detection method using loop-mediated isothermal amplification (LAMP) targeting the internal transcribed spacer region of ribosomal DNA of the nematode. Specificity of the primers and LAMP was confirmed using DNA from various nematode species related to B. xylophilus. Our experimental results suggest that LAMP can detect B. xylophilus faster and with higher sensitivity than the traditional diagnostic method. Moreover, because it does not require expensive equipment or specialized techniques, this LAMP-based diagnostic method has the potential to be used under field conditions. PMID:19900002
SAS molecular tests Salmonella detection kit. Performance tested method 021202.
Bapanpally, Chandra; Montier, Laura; Khan, Shah; Kasra, Akif; Brunelle, Sharon L
The SAS Molecular tests Salmonella Detection method, a Loop-mediated Isothermal Amplification method, performed as well as or better than the U.S. Department of Agriculture-Food Safety Inspection Service Microbiology Laboratory Guidebook and the U.S. Food and Drug Administration Bacteriological Analytical Manual reference methods for ground beef, beef trim, ground turkey, chicken carcass rinses, bagged mixed lettuce, and fresh spinach. The ground beef (30% fat, 25 g test portion), poultry matrixes and leafy greens were validated in a 6-7 h enrichment, and ground beef (30% fat, 375 g composite test portion) and beef trim (375 g composite test portion) were validated in a 16-20 h enrichment. The method performance for meat and leafy green matrixes was shown to be acceptable under conditions of co-enrichment with Escherichia coli 0157. Thus, after a short 6-7 h co-enrichment step, ground beef, beef trim, lettuce, and spinach can be tested for both Salmonella and E. coli O157. Inclusivity and exclusivity testing revealed no false negatives and no false positives among the 100 Salmonella serovars and 30 non-Salmonella species examined. The method was shown to be robust when enrichment time, DNA extract hold time, and DNA volume were varied. PMID:25051629
Smart point-of-care systems for molecular diagnostics based on nanotechnology: whole blood glucose analysis
NASA Astrophysics Data System (ADS)
Devadhasan, Jasmine P.; Kim, Sanghyo
Complementary metal oxide semiconductor (CMOS) image sensors are received great attention for their high efficiency in biological applications. The present work describes a CMOS image sensor-based whole blood glucose monitoring system through a point-of-care (POC) approach. A simple poly-ethylene terephthalate (PET) film chip was developed to carry out the enzyme kinetic reaction at various concentrations of blood glucose. In this technique, assay reagent was adsorbed onto amine functionalized silica (AFSiO2) nanoparticles in order to achieve glucose oxidation on the PET film chip. The AFSiO2 nanoparticles can immobilize the assay reagent with an electrostatic attraction and eased to develop the opaque platform which was technically suitable chip to analyze by the camera module. The oxidized glucose then produces a green color according to the glucose concentration and is analyzed by the camera module as a photon detection technique. The photon number decreases with increasing glucose concentration. The simple sensing approach, utilizing enzyme immobilized AFSiO2 nanoparticle chip and assay detection method was developed for quantitative glucose measurement.
Kozel, Thomas R.; Wickes, Brian
Early diagnosis of fungal infection is critical to effective treatment. There are many impediments to diagnosis such as a diminishing number of clinical mycologists, cost, time to result, and requirements for sensitivity and specificity. In addition, fungal diagnostics must meet the contrasting needs presented by the increasing diversity of fungi found in association with the use of immunosuppressive agents in countries with high levels of medical care and the need for diagnostics in resource-limited countries where large numbers of opportunistic infections occur in patients with AIDS. Traditional approaches to diagnosis include direct microscopic examination of clinical samples, histopathology, culture, and serology. Emerging technologies include molecular diagnostics and antigen detection in clinical samples. Innovative new technologies that use molecular and immunoassay platforms have the potential to meet the needs of both resource-rich and resource-limited clinical environments. PMID:24692193
Intelligent Condition Diagnosis Method Based on Adaptive Statistic Test Filter and Diagnostic Bayesian Network
Li, Ke; Zhang, Qiuju; Wang, Kun; Chen, Peng; Wang, Huaqing
A new fault diagnosis method for rotating machinery based on adaptive statistic test filter (ASTF) and Diagnostic Bayesian Network (DBN) is presented in this paper. ASTF is proposed to obtain weak fault features under background noise, ASTF is based on statistic hypothesis testing in the frequency domain to evaluate similarity between reference signal (noise signal) and original signal, and remove the component of high similarity. The optimal level of significance α is obtained using particle swarm optimization (PSO). To evaluate the performance of the ASTF, evaluation factor Ipq is also defined. In addition, a simulation experiment is designed to verify the effectiveness and robustness of ASTF. A sensitive evaluation method using principal component analysis (PCA) is proposed to evaluate the sensitiveness of symptom parameters (SPs) for condition diagnosis. By this way, the good SPs that have high sensitiveness for condition diagnosis can be selected. A three-layer DBN is developed to identify condition of rotation machinery based on the Bayesian Belief Network (BBN) theory. Condition diagnosis experiment for rolling element bearings demonstrates the effectiveness of the proposed method. PMID:26761006
Intelligent Condition Diagnosis Method Based on Adaptive Statistic Test Filter and Diagnostic Bayesian Network.
Li, Ke; Zhang, Qiuju; Wang, Kun; Chen, Peng; Wang, Huaqing
A new fault diagnosis method for rotating machinery based on adaptive statistic test filter (ASTF) and Diagnostic Bayesian Network (DBN) is presented in this paper. ASTF is proposed to obtain weak fault features under background noise, ASTF is based on statistic hypothesis testing in the frequency domain to evaluate similarity between reference signal (noise signal) and original signal, and remove the component of high similarity. The optimal level of significance α is obtained using particle swarm optimization (PSO). To evaluate the performance of the ASTF, evaluation factor Ipq is also defined. In addition, a simulation experiment is designed to verify the effectiveness and robustness of ASTF. A sensitive evaluation method using principal component analysis (PCA) is proposed to evaluate the sensitiveness of symptom parameters (SPs) for condition diagnosis. By this way, the good SPs that have high sensitiveness for condition diagnosis can be selected. A three-layer DBN is developed to identify condition of rotation machinery based on the Bayesian Belief Network (BBN) theory. Condition diagnosis experiment for rolling element bearings demonstrates the effectiveness of the proposed method. PMID:26761006
HEART Pathway Accelerated Diagnostic Protocol Implementation: Prospective Pre-Post Interrupted Time Series Design and Methods
Wells, Brian J
Background Most patients presenting to US Emergency Departments (ED) with chest pain are hospitalized for comprehensive testing. These evaluations cost the US health system >$10 billion annually, but have a diagnostic yield for acute coronary syndrome (ACS) of <10%. The history/ECG/age/risk factors/troponin (HEART) Pathway is an accelerated diagnostic protocol (ADP), designed to improve care for patients with acute chest pain by identifying patients for early ED discharge. Prior efficacy studies demonstrate that the HEART Pathway safely reduces cardiac testing, while maintaining an acceptably low adverse event rate. Objective The purpose of this study is to determine the effectiveness of HEART Pathway ADP implementation within a health system. Methods This controlled before-after study will accrue adult patients with acute chest pain, but without ST-segment elevation myocardial infarction on electrocardiogram for two years and is expected to include approximately 10,000 patients. Outcomes measures include hospitalization rate, objective cardiac testing rates (stress testing and angiography), length of stay, and rates of recurrent cardiac care for participants. Results In pilot data, the HEART Pathway decreased hospitalizations by 21%, decreased hospital length (median of 12 hour reduction), without increasing adverse events or recurrent care. At the writing of this paper, data has been collected on >5000 patient encounters. The HEART Pathway has been fully integrated into health system electronic medical records, providing real-time decision support to our providers. Conclusions We hypothesize that the HEART Pathway will safely reduce healthcare utilization. This study could provide a model for delivering high-value care to the 8-10 million US ED patients with acute chest pain each year. ClinicalTrial Clinicaltrials.gov NCT02056964; https://clinicaltrials.gov/ct2/show/NCT02056964 (Archived by WebCite at http://www.webcitation.org/6ccajsgyu) PMID:26800789
Multicenter Evaluation of Clinical Diagnostic Methods for Detection and Isolation of Campylobacter spp. from Stool.
Fitzgerald, Collette; Patrick, Mary; Gonzalez, Anthony; Akin, Joshua; Polage, Christopher R; Wymore, Kate; Gillim-Ross, Laura; Xavier, Karen; Sadlowski, Jennifer; Monahan, Jan; Hurd, Sharon; Dahlberg, Suzanne; Jerris, Robert; Watson, Renee; Santovenia, Monica; Mitchell, David; Harrison, Cassandra; Tobin-D'Angelo, Melissa; DeMartino, Mary; Pentella, Michael; Razeq, Jafar; Leonard, Celere; Jung, Carrianne; Achong-Bowe, Ria; Evans, Yaaqobah; Jain, Damini; Juni, Billie; Leano, Fe; Robinson, Trisha; Smith, Kirk; Gittelman, Rachel M; Garrigan, Charles; Nachamkin, Irving
The use of culture-independent diagnostic tests (CIDTs), such as stool antigen tests, as standalone tests for the detection of Campylobacter in stool is increasing. We conducted a prospective, multicenter study to evaluate the performance of stool antigen CIDTs compared to culture and PCR for Campylobacter detection. Between July and October 2010, we tested 2,767 stool specimens from patients with gastrointestinal illness with the following methods: four types of Campylobacter selective media, four commercial stool antigen assays, and a commercial PCR assay. Illnesses from which specimens were positive by one or more culture media or at least one CIDT and PCR were designated "cases." A total of 95 specimens (3.4%) met the case definition. The stool antigen CIDTs ranged from 79.6% to 87.6% in sensitivity, 95.9 to 99.5% in specificity, and 41.3 to 84.3% in positive predictive value. Culture alone detected 80/89 (89.9% sensitivity) Campylobacter jejuni/Campylobacter coli-positive cases. Of the 209 noncases that were positive by at least one CIDT, only one (0.48%) was positive by all four stool antigen tests, and 73% were positive by just one stool antigen test. The questionable relevance of unconfirmed positive stool antigen CIDT results was supported by the finding that noncases were less likely than cases to have gastrointestinal symptoms. Thus, while the tests were convenient to use, the sensitivity, specificity, and positive predictive value of Campylobacter stool antigen tests were highly variable. Given the relatively low incidence of Campylobacter disease and the generally poor diagnostic test characteristics, this study calls into question the use of commercially available stool antigen CIDTs as standalone tests for direct detection of Campylobacter in stool. PMID:26962088
Accurate Evaluation Method of Molecular Binding Affinity from Fluctuation Frequency
NASA Astrophysics Data System (ADS)
Hoshino, Tyuji; Iwamoto, Koji; Ode, Hirotaka; Ohdomari, Iwao
Exact estimation of the molecular binding affinity is significantly important for drug discovery. The energy calculation is a direct method to compute the strength of the interaction between two molecules. This energetic approach is, however, not accurate enough to evaluate a slight difference in binding affinity when distinguishing a prospective substance from dozens of candidates for medicine. Hence more accurate estimation of drug efficacy in a computer is currently demanded. Previously we proposed a concept of estimating molecular binding affinity, focusing on the fluctuation at an interface between two molecules. The aim of this paper is to demonstrate the compatibility between the proposed computational technique and experimental measurements, through several examples for computer simulations of an association of human immunodeficiency virus type-1 (HIV-1) protease and its inhibitor (an example for a drug-enzyme binding), a complexation of an antigen and its antibody (an example for a protein-protein binding), and a combination of estrogen receptor and its ligand chemicals (an example for a ligand-receptor binding). The proposed affinity estimation has proven to be a promising technique in the advanced stage of the discovery and the design of drugs.
Diagnostic imaging methods applied in long-term surveillance after EVAR. Will computed tomography angiography be replaced by other methods?
Stefaniak, Karolina; Gabriel, Marcin; Oszkinis, Grzegorz
Endovascular implantation of a stent graft in the abdominal aorta (endovascular aneurysm repair – EVAR) is a widely accepted alternative to open surgery in treatment of abdominal aortic aneurysms. Although EVAR is connected with a significant reduction in the risk of peri- and post-operative complications, it does not eliminate them totally. Long-term surveillance of post-EVAR patients is aimed at early detection of and fast reaction to a group of complications called endovascular leaks. Currently, the gold standard in leak diagnostics is computed tomography angiography (CTA). The other methods are ultrasonography, magnetic resonance (MR) angiography, intra-aneurysm sac pressure measurement, X-ray, and digital subtraction angiography (DSA). Despite many analyses based on long-term research, emphasising the high value and competitiveness of less invasive tests such as US or X-ray compared to CTA, it is still difficult for them to win the trust and acceptance of clinicians. The persisting view is that computed tomography is the test that finally resolves any inaccuracies. Consequently, a patient with a number of concurrent diseases is subject to absurdly high radiation exposure and effects of a radiocontrast agent within a short time. It is therefore logical to acknowledge that the EVAR-related risk is catching up with the open surgery risk, while the endovascular procedure is much more costly. Nevertheless, the status of CTA as the gold standard ultimately seems to be unthreatened. This paper presents a description of the diagnostic imaging tests that make it possible to detect any vascular leaks and to develop strategies for therapeutic processes. PMID:26966443
Accelerated molecular dynamics methods: introduction and recent developments
Uberuaga, Blas Pedro; Voter, Arthur F; Perez, Danny; Shim, Y; Amar, J G
A long-standing limitation in the use of molecular dynamics (MD) simulation is that it can only be applied directly to processes that take place on very short timescales: nanoseconds if empirical potentials are employed, or picoseconds if we rely on electronic structure methods. Many processes of interest in chemistry, biochemistry, and materials science require study over microseconds and beyond, due either to the natural timescale for the evolution or to the duration of the experiment of interest. Ignoring the case of liquids xxx, the dynamics on these time scales is typically characterized by infrequent-event transitions, from state to state, usually involving an energy barrier. There is a long and venerable tradition in chemistry of using transition state theory (TST) [10, 19, 23] to directly compute rate constants for these kinds of activated processes. If needed dynamical corrections to the TST rate, and even quantum corrections, can be computed to achieve an accuracy suitable for the problem at hand. These rate constants then allow them to understand the system behavior on longer time scales than we can directly reach with MD. For complex systems with many reaction paths, the TST rates can be fed into a stochastic simulation procedure such as kinetic Monte Carlo xxx, and a direct simulation of the advance of the system through its possible states can be obtained in a probabilistically exact way. A problem that has become more evident in recent years, however, is that for many systems of interest there is a complexity that makes it difficult, if not impossible, to determine all the relevant reaction paths to which TST should be applied. This is a serious issue, as omitted transition pathways can have uncontrollable consequences on the simulated long-time kinetics. Over the last decade or so, we have been developing a new class of methods for treating the long-time dynamics in these complex, infrequent-event systems. Rather than trying to guess in advance what
A Novel Method for Determining the Phase of T-Wave Alternans: Diagnostic and Therapeutic Implications
Sayadi, Omid; Merchant, Faisal M.; Puppala, Dheeraj; Mela, Theofanie; Singh, Jagmeet P.; Heist, E. Kevin; Owen, Chris; Armoundas, Antonis A.
Background T-wave alternans (TWA) has been implicated in the pathogenesis of ventricular arrhythmias and sudden cardiac death (SCD). However, in order to effectively estimate and suppress TWA, the phase of TWA must be accurately determined. Methods and Results We developed a method that computes the beat-by-beat integral of the T-wave morphology, over time points within the T-wave with positive alternans. Then, we estimated the signed derivative of the T-wave integral sequence which allows the classification of each beat to a binary phase index. In animal studies, we found that this method was able to accurately identify the T-wave phase in artificially induced alternans (p<0.0001). The coherence of the phase increased consistently after acute ischemia induction in all body-surface and intracardiac leads (p<0.0001). Also, we developed a phase resetting detection algorithm that enhances the diagnostic utility of TWA. We further established an algorithm that employs the phase of TWA in order to deliver appropriate polarity pacing pulses (all interventions compared to baseline, p<0.0001 for alternans voltage; p<0.0001 for Kscore), to suppress TWA. Finally, we demonstrated that using the phase of TWA we can suppress spontaneous TWA during acute ischemia; 77.6% for alternans voltage (p<0.0001) and 92.5% for Kscore (p<0.0001). Conclusions We developed a method to quantify the temporal variability of the TWA phase. This method is expected to enhance the utility of TWA in predicting ventricular arrhythmias and SCD and raises the possibility of using upstream therapies to abort a ventricular tachyarrhythmia prior to its onset. PMID:23884196
The Accuracy of Diagnostic Methods for Diabetic Retinopathy: A Systematic Review and Meta-Analysis
Martínez-Vizcaíno, Vicente; Cavero-Redondo, Iván; Álvarez-Bueno, Celia; Rodríguez-Artalejo, Fernando
Objective The objective of this study was to evaluate the accuracy of the recommended glycemic measures for diagnosing diabetic retinopathy. Methods We systematically searched MEDLINE, EMBASE, the Cochrane Library, and the Web of Science databases from inception to July 2015 for observational studies comparing the diagnostic accuracy of glycated hemoglobin (HbA1c), fasting plasma glucose (FPG), and 2-hour plasma glucose (2h-PG). Random effects models for the diagnostic odds ratio (dOR) value computed by Moses' constant for a linear model and 95% CIs were used to calculate the accuracy of the test. Hierarchical summary receiver operating characteristic curves (HSROC) were used to summarize the overall test performance. Results Eleven published studies were included in the meta-analysis. The pooled dOR values for the diagnosis of retinopathy were 16.32 (95% CI 13.86–19.22) for HbA1c and 4.87 (95% CI 4.39–5.40) for FPG. The area under the HSROC was 0.837 (95% CI 0.781–0.892) for HbA1c and 0.735 (95% CI 0.657–0.813) for FPG. The 95% confidence region for the point that summarizes the overall test performance of the included studies occurs where the cut-offs ranged from 6.1% (43.2 mmol/mol) to 7.8% (61.7 mmol/mol) for HbA1c and from 7.8 to 9.3 mmol/L for FPG. In the four studies that provided information regarding 2h-PG, the pooled accuracy estimates for HbA1c were similar to those of 2h-PG; the overall performance for HbA1c was superior to that for FPG. Conclusions The three recommended tests for the diagnosis of type 2 diabetes in nonpregnant adults showed sufficient accuracy for their use in clinical settings, although the overall accuracy for the diagnosis of retinopathy was similar for HbA1c and 2h-PG, which were both more accurate than for FPG. Due to the variability and inconveniences of the glucose level-based methods, HbA1c appears to be the most appropriate method for the diagnosis diabetic retinopathy. PMID:27123641
Perspective ground-based method for diagnostics of the lower ionosphere and the neutral atmosphere
NASA Astrophysics Data System (ADS)
Bakhmetieva, N. V.; Grigoriev, G. I.; Tolmacheva, A. V.
We present a new perspective ground-based method for diagnostics of the ionosphere and atmosphere parameters. The method uses one of the numerous physical phenomena observed in the ionosphere illuminated by high-power radio waves. It is a generation of the artificial periodic irregularities (APIs) in the ionospheric plasma. The APIs were found while studying the effects of ionospheric high-power HF modification. It was established that the APIs are formed by a standing wave that occurs due to interference between the upwardly radiated radio wave and its reflection off the ionosphere. The API studies are based upon observation of the Bragg backscatter of the pulsed probe radio wave from the artificial periodic structure. Bragg backscatter occurs if the spatial period of the irregularities is equal to half a wavelength of the probe signal. The API techniques makes it possible to obtain the following information: the profiles of electron density from the lower D-region up to the maximum of the F-layer; the irregular structure of the ionosphere including split of the regular E-layer, the sporadic layers; the vertical velocities in the D- and E-regions of the ionosphere; the turbulent velocities, turbulent diffusion coefficients and the turbopause altitude; the neutral temperatures and densities at the E-region altitudes; the parameters of the internal gravity waves and their spectral characteristics; the relative concentration of negative oxygen ions in the D-region. Some new results obtained by the API technique are discussed .
Rapid diagnostic methods for influenza virus in clinical specimens - A comparative study
NASA Technical Reports Server (NTRS)
Evans, A. S.; Olson, B.
A comparison of five rapid viral diagnostic techniques for identifying influenza virus in nasopharyngeal aspirates has been made on patients with influenza-like illnesses. Initial results with immune electron microscopy were positive in only one of 11 specimens from which virus was isolated and further work abandoned. Four other rapid tests were carried out on 39 specimens from which influenza virus had been isolated in tissue culture in 28. Of these 28 specimens yielding virus, 24 (85.7 percent) were positive by an indirect fluorescent antibody test (IFAT) on nasopharyngeal cells, 18 (64.3 percent) by enzyme-linked immunosorbent assay (ELISA), 19 (67.8 percent) by enzyme-linked fluorescent assay (ELFA), and 26 (92.8 percent) by a rapid tissue culture amplification method (TCA) in a continuous Rhesus monkey kidney line (LLC-MK2) with identification of virus by fluorescent antibody. In terms of sensitivity, simplicity, and rapidity, a combination of the IFAT and TCA methods seems to be very useful.
The ONIOM molecular dynamics method for biochemical applications: cytidine deaminase
Matsubara, Toshiaki; Dupuis, Michel; Aida, Misako
Abstract We derived and implemented the ONIOM-molecular dynamics (MD) method for biochemical applications. The implementation allows the characterization of the functions of the real enzymes taking account of their thermal motion. In this method, the direct MD is performed by calculating the ONIOM energy and gradients of the system on the fly. We describe the first application of this ONOM-MD method to cytidine deaminase. The environmental effects on the substrate in the active site are examined. The ONIOM-MD simulations show that the product uridine is strongly perturbed by the thermal motion of the environment and dissociates easily from the active site. TM and MA were supported in part by grants from the Ministry of Education, Culture, Sports, Science and Technology of Japan. MD was supported by the Division of Chemical Sciences, Office of Basic Energy Sciences, and by the Office of Biological and Environmental Research of the U.S. Department of Energy DOE. Battelle operates Pacific Northwest National Laboratory for DOE.
Authentication of animal signatures in traditional Chinese medicine of Lingyang Qingfei Wan using routine molecular diagnostic assays.
Cao, Meng; Wang, Jikun; Yao, Lu; Xie, Suhua; Du, Jing; Zhao, Xingbo
Lingyang Qingfei Wan produced by Beijing TongRenTang is a long-standing and popular medicine in China and international pharmaceutical markets. Concerns continue to be raised about the legality of usage of saiga antelope, which was defined as endangered species by Convention on International Trade in Endangered Species of Wild Fauna and Flora legislation and internal legislation in China. Therefore, the alternative pill in which substitutes saiga antelope with goat in the formula of Lingyang Qingfei Wan was developed. In order to authenticate the origin of animal contents in Lingyang Qingfei Wan and its alternative pill, molecular diagnostic assay was utilized by mtDNA polymorphism analysis. Four universal primer pairs containing mtDNA 12SrRNA, 16SrRNA, cytochrome b gene and cytochrome oxidase I were employed to obtain species-specific sequences of saiga antelope and goat, and multiple species-specific primer pairs for saiga antelope and goat were used to identify the animal origin in patent pills according to nucleotide polymorphisms between the two species. In additions, alternative techniques were attempted surrounding dilemmas of low concentration of target DNAs and presence of PCR-inhibitory substances in organic ingredients within complex pill. Results revealed that all species-specific primers could be successfully used for authentication of animal origin within complex pill, and sample preprocessing was critical during experimental manipulation. Internal positive control was an efficient and cost-effective way to assist in monitoring the potential interference from inhibitory substances which existed in the highly processed pills. PMID:24445529
Voltage-Gated Potassium Channels Kv1.3--Potentially New Molecular Target in Cancer Diagnostics and Therapy.
Teisseyre, Andrzej; Gąsiorowska, Justyna; Michalak, Krystyna
Voltage-gated potassium channels, Kv1.3, which were discovered in 1984, are integral membrane proteins which are activated ("open") upon change of the cell membrane potential, enabling a passive flux of potassium ions across the cell membrane. The channels are expressed in many different tissues, both normal and cancer. Since 2005 it has been known that the channels are expressed not only in the plasma membrane, but also in the inner mitochondrial membrane. The activity of Kv1.3 channels plays an important role, among others, in setting the cell resting membrane potential, cell proliferation, apoptosis and volume regulation. For some years, these channels have been considered a potentially new molecular target in both the diagnostics and therapy of some cancer diseases. This review article focuses on: 1) changes of expression of the channels in cancer disorders with special regard to correlations between the channels' expression and stage of the disease, 2) influence of inhibitors of Kv1.3 channels on proliferation and apoptosis of cancer cells, 3) possible future applications of Kv1.3 channels' inhibitors in therapy of some cancer diseases. In the last section, the results of studies performed in our Laboratory of Bioelectricity on the influence of selected biologically active plant-derived compounds from the groups of flavonoids and stilbenes and their natural and synthetic derivatives on the activity of Kv1.3 channels in normal and cancer cells are reviewed. A possible application of some compounds from these groups to support therapy of cancer diseases, such as breast, colon and lymph node cancer, and melanoma or chronic lymphocytic leukemia (B-CLL), is announced. PMID:26467143
Comparative evaluation of molecular diagnostic tests for Nucleospora salmonis and prevalence in migrating juvenile salmonids from the Snake River, USA
Badil, Samantha; Elliott, Diane G.; Kurobe, Tomofumi; Hedrick, Ronald P.; Clemens, Kathy; Blair, Marilyn; Purcell, Maureen K.
Nucleospora salmonis is an intranuclear microsporidian that primarily infects lymphoblast cells and contributes to chronic lymphoblastosis and a leukemia-like condition in a range of salmonid species. The primary goal of this study was to evaluate the prevalence of N. salmonis in out-migrating juvenile hatchery and wild Chinook salmon Oncorhynchus tshawytscha and steelhead O. mykiss from the Snake River in the U.S. Pacific Northwest. To achieve this goal, we first addressed the following concerns about current molecular diagnostic tests for N. salmonis: (1) nonspecific amplification patterns by the published nested polymerase chain reaction (nPCR) test, (2) incomplete validation of the published quantitative PCR (qPCR) test, and (3) whether N. salmonis can be detected reliably from nonlethal samples. Here, we present an optimized nPCR protocol that eliminates nonspecific amplification. During validation of the published qPCR test, our laboratory developed a second qPCR test that targeted a different gene sequence and used different probe chemistry for comparison purposes. We simultaneously evaluated the two different qPCR tests for N. salmonis and found that both assays were highly specific, sensitive, and repeatable. The nPCR and qPCR tests had good overall concordance when DNA samples derived from both apparently healthy and clinically diseased hatchery rainbow trout were tested. Finally, we demonstrated that gill snips were a suitable tissue for nonlethal detection of N. salmonis DNA in juvenile salmonids. Monitoring of juvenile salmonid fish in the Snake River over a 3-year period revealed low prevalence of N. salmonis in hatchery and wild Chinook salmon and wild steelhead but significantly higher prevalence in hatchery-derived steelhead. Routine monitoring of N. salmonis is not performed for all hatchery steelhead populations. At present, the possible contribution of this pathogen to delayed mortality of steelhead has not been determined.
Comparison of diagnostic methods for the detection and quantification of the four sympatric Plasmodium species in field samples from Papua New Guinea
Background Accurate diagnosis of Plasmodium infections is essential for malaria morbidity and mortality reduction in tropical areas. Despite great advantages of light microscopy (LM) for malaria diagnosis, its limited sensitivity is a critical shortfall for epidemiological studies. Robust molecular diagnostics tools are thus needed. Methods The present study describes the development of a duplex quantitative real time PCR (qPCR) assay, which specifically detects and quantifies the four human Plasmodium species. Performance of this method was compared to PCR-ligase detection reaction-fluorescent microsphere assay (PCR_LDR_FMA), nested PCR (nPCR) and LM, using field samples collected from 452 children one to five years of age from the Sepik area in Papua New Guinea. Agreement between diagnostic methods was calcualted using kappa statistics. Results The agreement of qPCR with other molecular diagnostic methods was substantial for the detection of P. falciparum, but was moderate for the detection of P. vivax, P. malariae and P. ovale. P. falciparum and P. vivax prevalence by qPCR was 40.9% and 65.7% respectively. This compares to 43.8% and 73.2% by nPCR and 47.1% and 67.5% by PCR_LDR_FMA. P. malariae and P. ovale prevalence was 4.7% and 7.3% by qPCR, 3.3% and 3.8% by nPCR, and 7.7% and 4.4% by PCR_LDR_FMA. Prevalence by LM was lower for all four species, being 25.4% for P. falciparum, 54.9% for P. vivax, 2.4% for P. malariae and 0.0% for P. ovale. The quantification by qPCR closely correlated with microscopic quantification for P. falciparum and P. vivax samples (R2 = 0.825 and R2 = 0.505, respectively). The low prevalence of P. malariae and P. ovale did not permit a solid comparative analysis of quantification for these species. Conclusions The qPCR assay developed proved optimal for detection of all four Plasmodium species. Densities by LM were well reflected in quantification results by qPCR, whereby congruence was better for P. falciparum than for P. vivax. This
Absolute calibration method for laser megajoule neutron yield measurement by activation diagnostics.
Landoas, Olivier; Glebov, Vladimir Yu; Rossé, Bertrand; Briat, Michelle; Disdier, Laurent; Sangster, Thomas C; Duffy, Tim; Marmouget, Jean Gabriel; Varignon, Cyril; Ledoux, Xavier; Caillaud, Tony; Thfoin, Isabelle; Bourgade, Jean-Luc
The laser megajoule (LMJ) and the National Ignition Facility (NIF) plan to demonstrate thermonuclear ignition using inertial confinement fusion (ICF). The neutron yield is one of the most important parameters to characterize ICF experiment performance. For decades, the activation diagnostic was chosen as a reference at ICF facilities and is now planned to be the first nuclear diagnostic on LMJ, measuring both 2.45 MeV and 14.1 MeV neutron yields. Challenges for the activation diagnostic development are absolute calibration, accuracy, range requirement, and harsh environment. At this time, copper and zirconium material are identified for 14.1 MeV neutron yield measurement and indium material for 2.45 MeV neutrons. A series of calibrations were performed at Commissariat à l'Energie Atomique (CEA) on a Van de Graff facility to determine activation diagnostics efficiencies and to compare them with results from calculations. The CEA copper activation diagnostic was tested on the OMEGA facility during DT implosion. Experiments showed that CEA and Laboratory for Laser Energetics (LLE) diagnostics agree to better than 1% on the neutron yield measurement, with an independent calibration for each system. Also, experimental sensitivities are in good agreement with simulations and allow us to scale activation diagnostics for the LMJ measurement range. PMID:21806179
Intraoperative Imaging-Guided Cancer Surgery: From Current Fluorescence Molecular Imaging Methods to Future Multi-Modality Imaging Technology
Chi, Chongwei; Du, Yang; Ye, Jinzuo; Kou, Deqiang; Qiu, Jingdan; Wang, Jiandong; Tian, Jie; Chen, Xiaoyuan
Cancer is a major threat to human health. Diagnosis and treatment using precision medicine is expected to be an effective method for preventing the initiation and progression of cancer. Although anatomical and functional imaging techniques such as radiography, computed tomography (CT), magnetic resonance imaging (MRI) and positron emission tomography (PET) have played an important role for accurate preoperative diagnostics, for the most part these techniques cannot be applied intraoperatively. Optical molecular imaging is a promising technique that provides a high degree of sensitivity and specificity in tumor margin detection. Furthermore, existing clinical applications have proven that optical molecular imaging is a powerful intraoperative tool for guiding surgeons performing precision procedures, thus enabling radical resection and improved survival rates. However, detection depth limitation exists in optical molecular imaging methods and further breakthroughs from optical to multi-modality intraoperative imaging methods are needed to develop more extensive and comprehensive intraoperative applications. Here, we review the current intraoperative optical molecular imaging technologies, focusing on contrast agents and surgical navigation systems, and then discuss the future prospects of multi-modality imaging technology for intraoperative imaging-guided cancer surgery. PMID:25250092
An In Vitro Comparison of Different Diagnostic Methods in Detection of Residual Dentinal Caries
Unlu, Nimet; Ermis, Rabia Banu; Sener, Sevgi; Kucukyilmaz, Ebru; Cetin, Ali Riza
The aim of this study was to investigate the efficiency of different diagnostic methods in detection of residual dentinal caries in excavated cavities. Fifty extracted molar with deep dentinal carious lesions were excavated using a slow-speed handpiece. All cavities were assessed by laser fluorescence(LF) device, electronic caries monitor(ECM), and caries detector dye(CDD) by three independent observers blindly. The measurements were repeated after two weeks. Specimens containing dentin slices 150 μm in thickness were prepared for histological analyses. The existence and absence of carious dentin was determined using a lightmicroscope. The average intraobserver accuracy was 1.00 (perfect agreement) for CDD, 0.86 (excellent agreement) for ECM, and 0.50 (good agreement) for LF. The average interobserver accuracy values were 0.92 (excellent agreement), (0.36 marginal agreement) and 0.48 (good agreement), for CDD, ECM, and LF, respectively. The average specificity was 0.60 for CDD, 73% for ECM, and 0.50 for LF. The average sensitivity was 0.55 for CDD, 0.85 for LF, and 0.47 for ECM. The average accuracy values were 0.53, 0.51, and 0.81 for CDD, ECM, and LF, respectively. LF had the greatest sensitivity and accuracy values of any of the methods tested. As a conclusion, LF device is appeared to most reliable method in detection of remain caries in cavity. However, because of its technical sensitivity it may susceptible to variations in measurements. To pay attention to the rule of usage and repeated measurements can minimize such variations in clinical practice. It was concluded that LF is an improvement on the currently available aids for residual caries detection. PMID:20613961