Induction if immunological tolerance to the major antigenic determinant of penicillin: a therapeutic approach to penicillin allergy. High titered IgE, IgG, and IgM antibody responses to the major antigenic determinant of penicillin, the benzylpenicilloyl hapten, were elicited by the intraperitoneal injection of the hapten coupled to keyhole limpet hemocyanin mixed with the appropriate adjuvant. However, treatment of such mice with the benzylpenicilloyl derivatized synthetic copolymer of D-glutamic acid and D-lysine, either before or after primary immunization, resulted in significant suppression of the subsequent anti-benzylpenicilloyl antibody responses of the IgE and IgG classes, as measured at the humoral and cellular levels. The state of tolerance induced by benzylpenicili-poly(DGlu, Lys) was highly specific, of long duration, and could be induced in a manner that would be appropriate for clinical use. These results provide a direct demonstration of the potential application of the poly(DGlu, Lys) immunotherapeutic approach to penicillin allergy in humans.


Primate type-C virus nucleic acid sequences (woolly monkey and baboon types) in tissues from a patient with acute myelogenous leukemia and in viruses isolated from cultured cells of the same patient. Cultured peripheral blood leukocytes from a woman (patient HL23) with acute myelogenous leukemia produced type-C RNA tumor viruses (HL23V). The viruses were analyzed by molecular hybridization experiments after transmission to five secondary cell culture lines. Using the criteria of molecular hybridization, we concluded that all of the transmitted virus isolates have nucleotide sequences related to the genome of simian sarcoma virus (SiSV). In addition, in agreement with data reported elsewhere, some of the transmitted viruses also have nucleotide sequences related to those of the baboon endogenous virus (BaEV). We also used molecular hybridization to ascertain whether both viruses could have originated from the patient HL23. Utilizing [3H] cDNA complementary to RNA from the separated BaEV-related component of HL23V and hybridizing this cDNA to DNA from tissues of the patient, we detected sequences related to BaEV in DNA obtained from the patient's spleen. These BaEV DNA sequences were also detectable when 125I-labeled RNA from BaEV was used as a probe. In agreement with earlier results, however, no SiSV-related sequences were detectable in the DNA of her tissues. Cytoplasmic viral-like particles, which had a buoyant density of 1.15-1.2 g/ml and were capable of synthesizing cDNA in association with a 35S RNA in vitro, were also found in the patient's fresh uncultured leukemic blood cells. cDNA synthesized by the cytoplasmic particles contained some sequences that hybridized to RNA from SiSV and, in addition, some that hybridized to RNA from BaEV. The cDNA also hybridized significantly to DNA isolated from the spleen of patient HL23 and to cytoplasmic RNA from the patient's leukocytes. These molecular hybridization results with nucleic acids obtained from the fresh blood cells of the patient, combined with the repeated isolation of similar viruses from different blood and bone marrow samples from the same patient, suggest that the virus come directly from the leukemic cell samples. The finding of BaEV-related DNA proviral sequences in the spleen of the patient strongly supports this interpretation. The failure so far to find a complete SiSV-related provirus is perplexing, but could be attributable to the existence of such a provirus in DNA of only a small population of cells in most leukemic patient.


[Triiodothyronine thyrotoxicosis in nodular toxic goiter]. Serum protein-bound iodine (PBI), total blood thyrotoxin (T4), triiodthyronine (T3), coefficient of thyrotoxin efficacy (CTE), serum thyrotoxin-binding capacity (TBC) or some of these indices were determined in 44 patients with nodular toxic goiter with thyrotoxemia. An increase in T3 level was revealed in 27 of 35 of the patients; T3-thyrotoxicosis was diagnosed in 18 of these in whom an increase of the T3 level was combined with normal T4 or normal CTE content and an unchanged TBC. The symptoms of T3-thyrotoxicosis displayed no significant differences from the symptoms of thyrotoxicosis caused by the excess of thyroxin. In many of the patients T3-thyrotoxicosis coursed for a long time with but a few symptoms. Loss of weight in this form of thyrotoxicosis was much less frequent than in "thyrotoxin'' thyrotoxicosis. PBI was elevated in 12 of 44 patients, and total T4 and CTE -- in 10 of 35. These methods do not permit to diagnose hyperthyroidism in many of the patients with nodular toxic goiter in which T3-thyrotoxicosis is of frequent occurrence.


[Remote results of long-term mercazolyl treatment combined with irradiation of the the hypothalamo-hypophyseal region in patients with diffuse toxic goiter]. Prolonged mercazolyl therapy (for from 9 months to 10 years) in combination with X-irradiation of the hypothalamo-hypophysial area (sum total dose of 5000 to 15000 rad) led only in 19 of 54 patients with diffuse toxic goiter to normalization of I131 uptake by the thyroid gland and depression of I131 uptake by the thyroid gland in a test with T3, which could indicate recovery from the disease. Despite a stable euthyroid state in 34 of 54 patients there persisted a hyperthyroid type of the curve of I131 uptake by the thyroid gland, and no depression of I131 uptake by the thyroid gland occurred in a test with T3. Thyrotoxicosis relapses were observed in 15 of 34 of the patients of this group. Development of thyrotoxicosis relapses only in the patients in whom pathological tests persisted pointed to the significance of disturbed interrelations in the hypothalamus-hypophysis-thyroid gland system in the genesis of thyrotoxicosis relapses.


The purification of calcium-binding protein from the uterus of the laying hen. Calcium-binding proteins from the chick intestine and uterus of the laying hen (Gallus domesticus) were isolated and purified by identical methods. Proteins from each source were shown to exhibit identical electrophoretic mobility and were immunologically identical. Estimation of molecular size by gel filtration indicated a value of approximately 28,000 daltons for both the chick intestinal and hen uterine calcium-binding proteins. The amino acid compositions of chick intestinal and hen uterine calcium-binding proteins were essentially identical.


Effects of calcium and phosphatidyl serine in rat mast cell reaction to dextran. Histamine release from Sprague-Dawley rat mast cells by dextran was completely inhibited by the absence of exogenous Ca2+ (in contrast to release from the same cells by antigen). Also, spontaneous leakage of histamine from the cells increased in the absence of Ca2+, and cell responsiveness was not completely restored by readding Ca2+. We found no effective substitute for Ca2+ in the release reaction. Ca2+ was not maximally effective immediately when added back to Ca-deficient cells, but almost the full effect of diluting Ca2+ in the medium (which decreased release) and of adding PS (which increased release) were very rapidly established, suggesting that both Ca2+ and PS might act (in part) at superficial cell sites. Release from activated cells could be stopped short by adding glucose or by diluting the cell-dextran mixture with normal buffer, as well as by adding EDTA, which deserves further study.


Sinusitis.

Sinusitis. The treatment of acute bacterial sinusitis is medical, the regimen including oral and nasal decongestants and antibiotics. Progression to chronic sinusitis can sometimes be prevented by periodic sinus washings until the acute process subsides. Sinus x-ray examinations are important in follow-up when symptoms or signs persist.


Histofluorescence in the unperfused CNS by cryostat and glyoxylic acid: a preliminary report. This paper describes an adaptation of glyoxylic acid histofluorescence to cryostat sections. The technique utilizes unperfused frozen brain, cryostat sectioning, immersion in 2% glyoxylic acid solution, warm-air drying, and exposure to hot glyoxylic acid gas. Fine, well-localized catecholamine histofluorescence is produced in cells, axons, and terminals. Both the anatomical localization and pharmacological specificity of the fluorescence conform to traditional catecholamine literature. The technique has the advantage of overcoming preservation and sectioning problems associated with the Vibratome. Because unperfused brain is used, alternate sections can be prepared for a variety of anatomical demonstrations or biochemical assays.


An immunoenzymatic method for improving the sensitivity of antigen measurement by electroimmunodiffusion techniques. Application to the quantification of human alpha-fetoprotein. A double antibody technique of electroimmunodiffusion, which uses glucose oxidase-labelled sheep antibodies to rabbit immunoglobulins as second antibody, is described. Primary antigen-antibody reaction is carried out with a rabbit antiserum by electroimmundiffusion. The glucose oxidase-labelled immunoreagent, being of general application, can serve for the quantification of different antigens and is here used for measurement of low levels of human alpha-fetoprotein. Reproducible results in the range of 50-800 ng/ml were obtained with a variation coefficient of 5 to 10%.


Chemical mediators of anaphylaxis (histamine, 5-HT, and SRS-A) released from horse lung and leukocytes in vitro. Horses were sensitized to bovine plasma in Freund's complete adjuvant. Leukocytes, separated from venous blood, yielded histamine upon incubation with bovine plasma. Ioslated lung fragments incubated with bovine plasma liberated histamine and 5-HT, but not SRS-A. Pulmonary veins obtained from the same animals contracted to histamine, 5-HT and to antigen (Schultz-Dale reaction). Histamine and 5-HT probably contribute to immediate-type hypersensitivity in horses whereas the role of SRS-A is not proved.


Hormone binding by cells and cell fragments as visualized by fluorescence microscopy. Fluorescent-labeled hormones offer an alternative approach to radio-labeling in studying the binding of hormones to intact cells or cell fragments. The binding of fluorescent-labeled hormones may be followed quantitatively by measurement of the polarization or the binding may be directly visualized in the fluorescence microscope. The binding of both fluorescein labeled prolactin and estradiol to a variety of whole cells or to microsomal fragments has been observed by fluorescence microscopy. No staining was observed with fresh cells whereas all cell types investigated, after freeze-thawing, stained at physiological levels (10-9M) of either hormone. Microsomal preparations from the mammary tissue of mid-pregnant rabbits likewise stained at low levels of prolactin. Inhibition of staining was not produced even by 10-6 M unlabeled hormone.


Protection of cattle against experimental haemorrhagic septicaemia by the capsular antigens of Pasteurella multocida, types B and E. Aluminum hydroxide adjuvant vaccines containing endotoxin-free capsular antigens of Pasteurella multocida, types B and E, were administered to cattle. Dose dependent serological responses were observed which were similar for both antigens. The immunised cattle were subjected to intravenous challenge by a virulent type E strain. All animals which received the highest vaccine dose survived and all unimmunised control animals died and a vaccine dose-response relationship was obtained. The results of passive mouse protection and indirect haemagglutination tests (type E) on the sera of immunised cattle corresponded with the degree of protection against challenge of the cattle.


[New findings on an anti-proteinase in human plasma: l'inter-alpha-trypsin inhibitor]. Inter-alpha-trypsin inhibitor (I alpha I) has been purified from C.N.T.S. fraction III as starting material. The purification procedure includes D.E.A.E. cellulose chromatography and gel filtration on G 150 Sephadex in the presence of EDTA. The purified protein gives one precipitation line in immunoelectrophoresis against anti-whole human sérum. It reacts only with an anti I alpha I immune serum and possesses a strong antitryptic activity. When studied in starch or polyacrylamide gel electrophoresis 2 components are observed, each of them having the same antigenic structure and the same antitryptic activity as the crude preparation. The slower and less important component is dissociated by 0,1% SDS. The molecular weight estimation of I alpha I BY PAA/SDS is about 180,000. This result is not modified by the presence of 1% beta mercaptoethanol indicating that I alpha I consists of one polypeptide chain. Crude preparation reveals under the same electrophoretical conditions small amounts of low molecular weight components (135,000 52,000 and 26,000) which can be due to a proteolytic action on I alpha I. Indeed plasmin is able to produce such fragments having an antitryptic activity as shown by fibrin/polyacrylamide gel electrophoresis. The relationship between small molecular weight inhibitors of human serum and bronchial secretions and those obtained after degradation of I alpha I by plasmin is discussed.


Multi-variate analysis of serum protein rheumatoid arthritis. The content of IgG, IgA, IgM, transferrin, haptoglobin, alpha2 macroglobulin, alpha1 acid glycoprotein, alpha1 antitrypsin, beta1C/1A globulin and beta1E globulin in the sera of 172 female rheumatoid patients and 132 female non-rheumatoid subjects was measured by the single radial immunodiffusion method. Rheumatoid patients were classified into 3 groups according to their stage and class. Canonical discriminant analysis revealed that the alpha1 acid glycoprotein and beta1C/1A globulin was of assistance in differentiating the rheumatoid and non-rheumatoid groups. There is no significant difference in serum protein levels between rheumatoid factor positive and negative groups, whereas IgA and beta1E globulin levels exhibited certain differences between these two groups. Among the results of other clinical tests, the correlation observed between ESR and serum protein levels in rheumatoid arthritis should be mentioned.


Type C virogenes: genetic transfer and interspecies transfer. Somatic cells of vertebrates contain gene sequences which are an integral part of chromosomal DNA and which code for the production of complete type C RNA viruses. These virogenes are genetically transmitted from parent to progeny along with other cellular genes (virogene-oncogene hypothesis). Activation of this endogenous virogene information from a normally repressed state, rather than infection by exogenous oncogenic viruses, has been proposed as the most common mechanism of cancer causation in animals, including man. Recent isolates of baboon type C RNA viruses, while related morphologically and biochemically to other mammalian type C RNA viruses, can be distinguished by nucleic acid hybridization and immunologic criteria. Within primates, type C virogenes have evolved as the species have evolved; virogenes from closely related genera and families have the closest gene sequence homology. Endogenous viruses from one species may infect animals of a distantly related species and become incorporated into their germ line. Genomes of exogenous viruses, such as the murine leukemia viruses, which are infectious from animal to animal within the same species, evolve more rapidly than the endogenous virogenes which replicate solely as cellular genes. A major viral structural protein of baboon type C RNA viruses, p30 was detected by radioimmunoassay in normal primate tissues. Radioimmunoassays have also detected p30 antigen in human tissues which appear to be immunologically related to primate viral p30. Hybridization experiments have confirmed that type C viral sequences are also present in the human genome.


Prostatic tissue cell growth and assessment. The potential for use of cell and organ culture systems in the study of prostatic tumors is high. The nature of this organ and our lack of basic information about it limit the usefulness of in vitro techniques at present but these shortcomings can be overcome by concerted effort of interested investigators. At the present time organ culture methods are available which can yield valuable information. To achieve notable success requires considerable understanding of the method in terms of principles, detail of technique, and interpretation. Several important questions must be resolved in order for cell culture methods to achieve their potential: (1) effect of disaggregation procedures on prostatic tissue, (2) appropriate selection of specimens, particularly with regard to controls, (3) ability to clearly identify and select cell types, and (4) how to overcome the variable of serum as a source of hormones. Considering the fact that less work has been done on the prostate than most tissues, recent developments in the area of cell and organ culture seem most encouraging and the future looks bright indeed. Two continuously propagated cell lines of pupative prostatic origin have been described. One of these developed in culture apparently as the result of a spontaneous transformation while the other was propagated from an adenocarcinoma. If these cell lines can be conclusively proved to be representative of the tissues of origin we already are well on the way to the application of in vitro systems to study of the prostate.


[Rosenthal fibres and vascular anomalies in cerebral gangliocytomas (author's transl)]. The Authors describe three cases of gangliogliomas histologically studied among 3100 neoplasias of the CNS. In one case the study was supported by the cytological observations on the in vitro cultures as performed with the roll technique. The tumors differed in location and clinical behaviour, but showed the same histological characteristics of mixed neoplasias. In fact in all cases a component of mature ganglion cells together with glial proliferation of spongioblastic type was identified. Particular attention was given to the vascular stroma and to the morphological aspects of the glial proliferation. All tumors presented a large amount of anomalous vessels frequently of angiomatous type. Beside neuronal cells, a neoplastic glial cell proliferation was identified as of spongioblastic nature by means of in vitro culture and on the basis of particular degenerative changes typical of the spongioblastic elements, such as Rosenthal fibres. According to the Authors these data support the hamarto-blastomatous origin of the oncotype.


Cancer of the esophagus: ten-year experience. A critical analysis of 52 patients with cancer of the esophagus indicates the continuing challenge that this disease entity presents to the surgeon and radiotherapist. Difficulty of cure or palliation is further compounded by the many significant associated medical problems found in certain patient populations, such as that characteristic of our VA Hospital. Although cure is a rarity in cancer of the esophagus, continued palliative efforts must be exerted for these otherwise hopeless patients.


Palliative treatment of carcinoma of the esophagus. In a ten-year period 139 patients with epidermoid carcinoma of the esophagus were treated by palliative feeding procedures, palliative radiation therapy, radiation therapy for cure, or resection. Analysis of the results of these modes of therapy indicates that long-term palliation is best achieved when resection is the primary method of therapy. Radiation therapy improved survival in the first 12 months of disease; however, no patients treated with radiation therapy alone were cured. In five cases resection resulted in apparent cure. For this reason we suggest that resection for palliation is indicated, and in an occasional patient it will result in a cure.


Squamous cell carcinoma of the anus.

Squamous cell carcinoma of the anus. The institution of proper therapy for squamous cell carcinoma of the anus requires an intimate knowledge of the histology and anatomy of this region. It also requires an awareness of the disease entity by the physician and a realization that everything that bleeds around the anus is not due to hemorrhoidal disease. Once the diagnosis and extent of the disease are established, several known facts can be applied in the decision for specific therapy. For small lesions (less than 3 cm in diameter) located in the perianal skin, it is now clear that wide local excision, usually with skin graft, is a safe, reliable method of treatment. However, if these lesions are invading deeply, one should resort to a more radical form of therapy. The combined abdominoperineal resection is the definitive treatment for lesions in the anal canal. In this way, local recurrence can be prevented insofar as possible, invasive lesions can be completely eradicated, and patients with lymph node involvement will be given the greatest opportunity for survival. As is the case in all radical types of therapy, the anal sphincter mechanism of a few patients will be sacrificed unnecessarily. If a lesion is located in the anal canal and does not invade beyond the submucosa, such as those lesions discovered during routine hemorrhoidectomy, it might be safe to undertake wide local excision of the region. Precise pathologic study must be available, however, and the patient must be apprised of this compromise in management and be willing to be examined frequently for evidence of recurrent disease. With present knowledge and development of the operation, radical removal of inguinal lymph nodes should be undertaken when the nodes are thought to contain tumor. It must be realized, however, that the overall salvage rate in this situation is low. Finally, the treatment of recurrent and metastatic squamous cell carcinoma of the anus remains a great dilemma. The first choice of treatment for local recurrence should be excision whenever possible. Radiation may have some value, but excellent responses are rare. Lastly, several chemotherapeutic regimens are available for use in specific cases.


Alien cancers of the duodenum.

Alien cancers of the duodenum. Invasion of the duodenum by contiguous cancer or an isolated metastasis can manifest itself with the signs and symptoms of primary cancer of the duodenum. Fourteen patients were identified with this type of diagnostic confusion. All of their tumors were in the second or third portion of the duodenum. Although the median length of survival after excision of the tumor or palliative bypass operations in these patients was nine months, considerably longer survival times have also been described. Barium enema examinations and excretory urograms should be considered as diagnostic tests for any patient with a tumor of the duodenum to identify the contiguous carcinomas of the colon and kidney subject to long palliation.


[Bronchial carcinoma, dependence of operationsuccess (author's transl)]. From 1958 to 1973, 3792 cases of bronchial carcinoma were admitted in our department. 1577 of them could be operated radically. The influence of various factors on the result of the operation is discussed, considering eventual changes in the course of the reported period.


Permanent lymphoid lines from genetically marked lymphocytes: success with lymphocytes recovered from frozen storage. Permanent human lymphoid cell lines were established successfully from peripheral blood lymphocytes which had been separated for HL-A typing and stored in liquid nitrogen for two years. Frozen lymphocytes were chosen from two siblings who were homozygous at the LA and FOUR HL-A loci. Thawed lymphocytes were transformed with EB virus produced by the marmoset lymphoid line B95-8. No chromosome abnormalities were seen on karyotypes prepared on cells from the established human lymphoid lines using G and Q banding techniques. HL-A typing showed the expected HL-A antigens plus a considerable number of additional reactions. Separation of lymphocytes and freezing them for possible future use requires a relatively small investment. This method of preserving cells can be applied to patients with interesting genetic disorders or other biochemical markers to provide cells which can be transformed and propagated years later.


Recognitive specificity of human cytotoxic T lymphocytes. II. The non-recognition of antigens controlled outside the major histocompatibility complex. In vitro sensitized cytotoxic T lymphocytes (CTLs) primarily detect antigens controlled within the major histocompatibility complex (MHC) that are associated with the serologically detected (SD) loci LA and FOUR. Frequently CTLs sensitized to cells from one individual kill target cells of a third party that shares no SD cross-reactivities by serological criteria with the initial stimulating cells. We have studied the possibility that this cross-killing is due to the recognition of non-MHC controlled antigens. Allogeneic tissue from MHC identical siblings is rejected in the absence of immunosuppression, demonstrating the importance of non-MHC antigens. Nevertheless, CTLs sensitized simultaneously with mitomycin-C treated cells from an MHC identical sibling and from an unrelated individual were only able to mediate cytotoxicity on the unrelated's target cells. No significant cytotoxicity was observed in any combination on target cells differing only for non-MHC antigens.


Alloantigens defined by cytotoxic isoantisera prepared against peripheral rabbit lymphocytes. Seven lymphocytotoxic antisera were produced in rabbits by means of isoimmunization with purified peripheral lymphocytes. The phenotypic profile of the defined lymphocyte antigens was determined in a panel of 97 rabbits. The results indicated the existence of at least two major antigenic determinants. Further typing and cross-absorptions were performed with several of the lymphocytotoxic antisera in order to demonstrate monospecificity. The anti-3 antiserum was used to determine the distribution of the respective antigens in various rabbit organs. The highest concentration of the alloantigen was contained in the spleen, followed in descending order by the lymph nodes, lung, skin, kidney, heart and liver. Fat and red blood cells (rbc) contained no detectable antigen. A two-stage physical adherence column method was used to separate T-derived from B-derived lymphocytes and each fraction was tested for the alloantigen. Alloantigen was detected on both cell types. The fact that the anti-3 antiserum never killed more than 30% of the cells suggests that it could define a subpopulation of lymphocytes.


Linkage of GLO with HLA and Bf. Effect of population and sex on recombination frequency. Further data on the linkage relationships of red cell glyoxalase I(GLO) with HLA and Bf are reported. The most likely order of loci is GLO: Bf: HLA-B: HLA-A. No sex difference in the frequency of recombination between GLO and HLA was noted, but recombination was more frequent for both males and females in the American black population than in the white population.


Cell mediated lympholysis in man. The impact of HLA-C antigens. From a population of individuals, all HLA-A, B, and C tissue typed in relation to the Sixth International Histocompatibility Workshop, an experimental investigation has been performed to study the influence of the HLA-Cwl, w2, w3, w4, and w5 antigens in the Cell Mediated Lympholysis (CML) test. The average cytolysis obtained due to allogenic attack of one HLA-C antigen equals 12.6%, but like the HLA-A and B antigens, HLA-C antigens exhibit differences with regard to sensitizing and target potential. This indicates either a heterogeneity of these antigens or the existence of separate CML determinants. It is concluded that the HLA-C antigens may account for the cytolysis observed in some of the combinations exhibiting cytolysis which cannot be explained by the HLA-A and B antigens.


A rapid method for demonstrating skeletal muscle motor innervation in frozen sections. Longitudinal 50-100 mum-thick frozen sections of muscle are picked up on slides coated with 3% EDTA and after drying are incubated to demonstrate acetylcholinesterase. Subsequent incubation in 0.5% K3Fe(CN)6 is followed by fixation for 30 minutes in formol-calcium or formol-saline. After washing, the slides are incubated in 20% aqueous AgNO3 containing 0.1% CuSO4 for 2-30 minutes at 37 C. Following development in a 1% solution of quinol (w/v) 5% with respect to NaSO3 (w/v), axons and subneural apparatus stain dark brown to black in contrast to the less well stained muscle fibers and nuclei. This procedure permits study of the pattern of neuromuscular innervation in skeletal muscle 3 1/2-4 hours after receipt of a sample, and makes possible determination of the terminal innervation ratio.


A Golgi-electron microscope method for insect nervous tissue. Golgi's light microscope method of selective silver impregnation for nervous tissue combined with electron microscopy appears to offer a promising method for working out the detailed anatomy of individual neurons and their connections. Insect nervous tissue is fixed in a mixture of 2% paraformaldehyde and 2 1/2% glutaraldehyde in Millonig's buffer (pH 7.2) before postfixation for 12 hours in a solution brought to pH 7.2 with KOH containing 2% potassium dichromate, 1% osmium tetroxide and 2% D-glucose. The tissue is then transferred to a solution of 4% potassium dichromate for 1 day; and for 1-2 days to a 0.75% silver nitrate solution. After dehydration and embedding in Araldite, 50 mum sections are made. Areas of interest are cut from these sections and re-embedded in silicone molds. Ultrathin sections are then cut and stained with uranyl acetate and lead citrate. The Golgi method described here gives good results at the level of both light and electron microscopy.


A method for the preparation of serial thick sections of plastic-embedded plant tissues. A procedure is described in which thick sections (2-10 mu or more) of plastic-embedded plant tissues are mounted in serial order on slides for use in routine light microscopy. Sections are cut with a steel knife on a rotary microtome while the block and blade are bathed with 40% alcohol. The cut sections are placed, in order, in 50% alcohol in the small wells of modified plastic trays where they become flat, pliable and suitable for subsequent handling. Sections remain separate and in correct order in the trays while they are stained, washed, and prepared for final mounting on slides. Mounting involves a simple and rapid procedure of transferring the sections to a slide and heating first on a 70-75 C hot plate (to slowly evaporate the water around the section and to partially affix the section) and then on a C hot plate. This second heating ensures adhesion when xylene-base mounting media, which tend to loosen weakly adhered plastic from the slides, are used. The technique of staining the sections loose provides the following advantages: (1) the problems of section loss and entrapment of stain between section and slide during staining are eliminated, (2) relatively high staining temperature, alkalinity, and alcohol concentration of the stain solvent (all of which promote loosening of pre-affixed sections from slides during staining) is allowed, and (3) staining is more even and selective. The procedure has been found to be reliable and fast enough to be of value in a significant variety of routine light microscope studies.


Hematoxylin substitutes: a survey of mordant dyes tested and consideration of the relation of their structure to performance as nuclear stains. In the search for hematoxylin substitutes 26 dyes were more or less extensively tested for performance as nuclear stains, usually in combination with aluminum, chronic, ferrous and ferric salts. Reports from the literature on hematoxylin substitutes were also considered, and efforts were made to obtain samples of favorably reported dyes and test them. The reports on anthocyanins include isolated reports on several berry juices and a considerable number of studies on Sambucus niger and Vaccinium myrtillus. None of these have so far been tested by us. Otherwise favorable reports have appeared on eleven synthetic dyes and on carmine, brazilin, and hematin. Except for one of the synthetics, naphthazarin, which is no longer fractured, we had samples of all of these. In addition, more or less unsuccessful trials were made on twelve dyestuffs, some of which were new syntheses designed to combine chelating capacity with nucleophilia. Following Fyg's report of blue nuclear staining with chrome alum carmine, trial was made to change the red nuclear stain of kernechtrot by altering the metal mordant. The most successful dyes were phenocyanin TC, gallein, fluorone black, alizarin cyanin BB and alizarin blue S. Celestin blue B with an iron mordant is quite successful if properly handled to prevent gelling of solutions.


A method for staining infection hyphae in pine leaves. Fungus-inoculated Pinus radiata leaves were fixed and then stained with periodic acid-Schiff reagent. Pieces of leaf with fungal material on the surface were removed. These pieces were stained in lactophenol cotton blue for a few minutes and then mounted in dilute lactophenol cotton blue. Microscopic examination of fungal material inside and outside the mounted leaf pieces revealed the following: condidia and germ tubes on the leaf surface were red, appressoria remained unstained, and infection hyphae within the leaf were stained blue. This differential staining method was particularly useful for distinguishing germ tubes from infection hyphae arising from appressoria.


New source for positive controls used for the identification of spirochaetes in tissue sections. Lyophilized T. pallidum is recontituted or purchased as a suspension, evenly dispersed, made into smears using alcohol-cleaned slides and a bacteriological platinum wire loop. Smears are air dried for 15 minutes and fixed in concentrated formaldehyde fumes for 30 minutes and stored at room temperature. These preparations are used as positive controls for conventional silver impregnation techniques recommended for spirochaete detection in tissue sections.


The use of tannic acid-glutaraldehyde in the study of elastic and elastic-related fibers. Fixation with tannic acid-glutaraldehyde permits distinction of oxytalan, elaunin and elastic fibers in the electron microscope. The results obtained using tannic acid at concentrations of 1.0%, 0.5% and 0.25% in 3% glutaraldehyde were compared. The 0.25% concentration is recommended for studying fine details of connective fibrils and for regular staining of elastin.


A hybrid technique for staining meiotic nuclei of Basidiomycetes. A hybrid nuclear staining procedure used on the basidiomycete Schizophyllum commune produced squash preparations of sufficient density for effective brightfield photomicrography. Tissure is fixed in Newcomer's solution, rehydrated in a graded ethanol series, rinsed, hydrolyzed in 1 N HC1 at 60 C, washed, mordanted in 4% iron alum, rinsed again, and stained with aceto-iron-hematoxylin prior to mounting and squashing.


Hydroxyethyl lactamide, a dye solvent useful in vital staining. In a search for new vital stains to reveal the microanatomy of nudibranch mollusks, the slow or very low solubility of many dyes in sea water posed a serious problem. Preliminary dissolution in tap water proved impractical. Hydroxyethyl lactamide, an odorless liquid and dye solvent was found ideal since it permits immediate attainment of saturated solutions of dyes in sea water. Since hydroxyethyl lactamide passed the severe "eolid nudibranch test" and has been found nonirritating for the very sensitive rhinophorial structures, and furthermore since it has been used by the pharmaceutical industry as a vehicle in antibiotic preparations, it appears to be an ideal universal dye solvent for general use in vital staining. It has been used extensively in unpublished research by the writer on vital staining of nudibranchs. It has a low order of physiological activity and can be regarded an essentially inert when used in vital staining.


Methods for studying the distribution of pigment in malpighian tubules of Drosophila melanogaster. The pigmented cells in Malpighian tubules of Drosophila melanogaster contain yellow globules which are blackened by osmium tetroxide. Permanent preparations of the tubules showing pigmentation can thus be made. Following prestaining acid hydrolysis, osmium-fixed tubules can be stained with orcein. For observations on pigment distribution and Feulgen-DNA content in the same preparation, tubules are fixed in mercuric chloride. Color remains visible for several hours in unfixed tubules mounted in modified buffered Ringer solution.


A procedure for differential staining of cartilage and bone in whole formalin-fixed vertebrates. This paper describes a modification of the Simons and Van Horn (1971) procedure for rendering cartilage blue, bone red, and soft tissue translucent or transparent in whole vertebrate specimens. Alcian blue and alizarin red S are used to stain cartilage and bone respectively. In our procedure formalin is used as a fixative. This is a significant modification because formalin is the common fixative for museum specimens. This clearing and staining procedure is thus readily applicable to comparative studies in anatomy, embryology and systematic zoology.


Science and art in preparing tissues embedded in plastic for light microscopy, with special reference to glycol methacrylate, glass knives and simple stains. Plastic embedding preserves tissue structure much more faithfully than does paraffin. Acrylic polymerization is innocuous to dye-binding groups in sections. The water solubility of glycol methacrylate monomer and the hydrophilic properties of the polymer allow for convenience in dehydration and for versatility in staining sections. Five years of experience with glycol methacrylate (GMA) embedding for light microscopy is summarized. Methods for purifying GMA monomer are cited. Procedures for fixing, dehydrating, embedding, polymerizing, sectioning and staining, using GMA, are explained. A method is provided for making glass knives long enough to cut large blocks. Simple, reliable, quick staining methods are outlined. When compared with paraffin, GMA offers opportunities for simpler, quicker procedures and yields sections of superior quality, greater information content, and less distortion.


Identification of functional tumor vessels by resorcin-crystal violet stain infusion. A resorcin-crystal violet solution of low viscosity injected into the circulatory plexus supplying a tumor is used to identify and characterize functional tumor vessels. Unstained sections of tumor tissue demonstrate heavily stained vascular endothelium with no leakage of stain to extravascular tissue for intact vessels, and little or no background staining. The method is simple to apply for tumor vessel morphology and morphometry.


Improved microradiographic contrast for bone stain-historadiography. Microradiographs of 5-micron sections of methyl methacrylate embedded undemineralized bone show poor resolution, but prestaining with silver nitrate increases the radioopacity of the calcified tissues to soft x-rays without masking regional differences in microscopic mineralization. Identical results are achieved using aqueous (pH 5.8 and 7.5) or ammoniacal solutions (pH 7.9). Atomic absorption spectrometry detected no loss of calcium from the sections during staining. Osteoid in plastic-embedded bone is not rendered radiopaque by this technique even after prolonged staining times (5 min-2 hr).


An improved polychrome staining method for thick epoxy sections. Improved polychrome staining of 1-1.5 mum epoxy sections is achieved with sequential applications of a single basic fuchsin-methylene blue mixture at two different pH values. The dye solution is applied for 2-3 min at 50-52 C first at pH 7.9, then at pH 6.7. In sections of mouse mammary tissue, epithelial cells are stained deep blue, connective tissue pink, and fat cells bright olive-green. This simple technique consistently yields uniform, vivid, contrasting colors that sharply delineate the elements of the complex glandular architecture of the mammary gland. Similar polychromatic effects are obtained in applications to other tissues, such as stomach, adrenal gland, mammary tumor and artery.


A combined silver and acetylcholinesterase method for staining intramuscular innervation. A combined acetylcholinesterase and silver stain for demonstrating the intramuscular innervation of fresh frozen tissue is described. Intramuscular nerves, subterminal axons, and motor end plates are simultaneously stained brown or black with minimal staining of connective tissue and muscle fibers in longitudinal sections 30-100 mu thick. The method has been applied to fetal and adult rat, porcine, and bovine skeletal muscle. Antemortem and postmortem tissue samples stained equally well. The method facilitates simultaneous appreciation of morphological alterations in nervous and muscular tissues; in clinical and research laboratories alike it is of value when muscle abnormalities which may be related to disorders of nervous origin are studied. Compared with other published procedures this method has shorter time requirements, uses fresh frozen tissue, and displays superior staining characteristics.


Improved staining procedures for semithin epoxy sections of plant tissues. Improved and reliable methods are described for staining semithin sections of plant materials fixed in glutaraldehyde-osmium and embedded in epoxy resins. One-micron sections are fixed to slides, stained with a two-solution hematoxylin procedure or with a methylene blue-azure A combination, counterstained in aqueous safranin O, cleared, and mounted permanently. Basophilic tissue components are stained gray to black by the hematoxylin and blue or purple by the methylene blue-azure A combination; cell wall structures are colored by the safranin. With the procedures recommended, stains are sharp and intense, sections are flat, wrinkling and loss are held to a minimum, and unsightly precipitates do not form.


Iron and aluminum lakes of Gallo blue E as nuclear and metachromatic mucin stains. Gallo blue E, C. I. No. 51040, Mordant Violet 54, furnishes a blue black nuclear stain when applied to tissue sections in the form of its moderately stable iron lakes. This coloring combined well with such counterstains as orange G and eosin B. The Van Gieson stain tends to decolorize mucins, cartilage, and mast cells previously stained with this dye. Its aluminum lake solutions tend to gel in a few minutes to 24 hours depending on the solvent used and the amount of Al3+ present. Aluminum lake solutions give a moderately good blue to dark blue nuclear stain and a brilliant purplish red to dark purple stain to a variety of epithelial and connective tissue mucins. Acid dye counterstains are poorly tolerated. With either lake, nuclear staining is abolished by deoxyribonuclease digestion or relatively short mineral acid extraction of DNA.


Grid staining--a useful device.

Grid staining--a useful device. A simple and disposable holder for electron microscope grids can be constructed from discs of silicon rubber. The discs (10 mm thick X 25 mm diameter) are scored 0.4 mm deep with a razor. Flexing the discs opens the scores for insertion of grids. Staining wells can be made from polyethylene snap-in vial tops by cutting away the nipple-like portion. After impaling the holders on the tips of a pair of forceps the grids can be washed by immersion. This holder is rigid, inert and inexpensive. It is of particular advantage for handling single-slotted grids.


[Therapy of vulvar carcinoma].

[Therapy of vulvar carcinoma]. Thirty-five of totally 50 patients with carcinoma vulvae were treated with bleomycin. Most of the patients were older people and had more advanced disease. They were grouped according to the TNM system and the age. Bleomycin alone (2 X 15 mg/m2 weekly; 390 and 420 mg total dose) was given to two patients. One of the patients had a complete remission and the other one a partial remission with reduction in tumor size of more than 50%. Nineteen patients received bleomycin (2 X 10 up to 2 X 15 mg/m2 weekly: 200-300 mg total dose) in combination with radiation therapy using fast electrons (betatron; 3000-5000 R). In this group 21,1% of the patients had complete remission and 31,5% of patients had a partial remission of more than 50% tumor size reduction (objective remission rate 52,6%). Bleomycin was also beneficial when combined with surgery plus irradiation. Palliative irradiation was used in four patients with a very advanced disease. Eleven patients were irradiated postoperatively with fast electrons (4000-6000 R). In this group, 65% of the patients were free from recurrences more than one year after the beginning of the treatment. Our results indicate that combined therapies using bleomycin, surgery and radiation therapy were more effective in the treatment of vulvar carcinoma than single treatments alone. It should be emphasized that bleomycin is effective as a palliative treatment of squamous cell carcinoma of the vulva.


Occurrence of S-antigens in serum in Plasmodium falciparum infections in man. Gambians with severe Plasmodium falciparum infections frequently have malarial S-antigens in their serum or plasma. Detection of these with the available antisera which are from immune adults is complicated because there are many different S-antigens and none of the antisera contains antibodies to them all. Despite this we found S-antigens in about 60% of a large series of malarious children and demonstrated that this was a considerable underestimate of the true incidence. We also showed that the prevalence of S-antigens was related to the malaria season and to the intensity of infection.


The effect of neuraminidase on the sensitivity of tumor cells toward lysis by antibody and complement or by sensitized lymphocytes. The effect of neuraminidase treatment on the antigenicity of sarcoma I (Sal) and mastocytoma P-815-X2 cells was compared, both in terms of their reaction with various antisera (C57BL/6 anti-SaI, C57BL/6 anti-mastocytoma, rabbit and horse ALS) and complements (guinea pig, rabbit) or with sensitized lymphocytes (C57BL/6 anti-SaI, C57BL/6 anti-mastocytoma, A anti-mastocytoma). With sensitized lymphocytes the reactivity of neuraminidase-treated tumor cells was similar to that of nontreated cells. In contrast, after neuraminidase treatment, the tumor cells showed increased specific lysis with the antisera and complement. Rabbit complement was highly cytotoxic to the neuraminidase-treated cells but this nonspecific activity could be removed by absorption with agar or agarose. The results of this study would strongly suggest that neuraminidase treatment does not change the antigenic determinants of tumor cells but increases their interaction with complement after removal of negatively charged sialic acids.


Further evidence of heterogeneity within the Kidd blood group system. Serological studies on blood of selected individuals show that red blood cells from individuals genetically JkaJk and JkbJk give single dose agglutination reactions with anti-Jka and anti-Jkb respectively, but react as strongly with anti-JkaJkb as do cells from random individuals. A Jk(a+b-) Oriental person has been found whose red blood cells give a single dose reaction with anti-Jka, but react more weakly than do cells from random people in tests with anti-JkaJkb. The results support a conclusion that the antigen recognized by anti-JkaJkb is a distinct and separate antigen of the Kidd blood group system.


Leukapheresis of patients with chronic granulocytic leukemia (CGL), using the Haemonetics blood processor. By the use of citrated hydroxyethyl starch (HES) as anticoagulant, the Haemonetics blood processor can be used to obtain large numbers of granulocytes from patients with CGL. This report is of 67 leukaphereses on 11 different patients. A median of 1.14 x 10(11) granulocytes was obtained per 6-cycle pheresis (3.4 x 10(10) per liter of blood processed), or eight to ten times the number obtained from comparable leukaphereses of normal donors. High yields of platelets were also obtained, although not in proportion to granulocytes, since some of the patients used as donors have normal or even low platelet counts. The patients tolerated the procedure well, and no adverse reactions to HES were observed. The patients experienced a mean 35 per cent drop in the postleukapheresis WBC count, but in no case was this drop sustained for more than a few days, and no lasting effect on the disease process was observed.


Eluate analysis of anti-Bg a associated renal allograft rejection. Renal allograft rejection, following development of a positive indirect antiglobulin reaction, occurred in a multiply transfused recipient. Panel techniques confirmed both anti-Bga and anti-Kell antibodies. Recognition of the concordant antigenic specificity of the Bga and HL-A7 antigens led to an investigation of the potential role of this antibody in the rejection phenomenon. In the absence of serological cytotoxicity, a modified elution technique was used to directly obtain immunoglobulin from the rejected allograft. The eluate obtained displayed specificity for the Bga red blood cell antigen. The described technique affords an additional approach to the documentation of immunologically mediated graft rejection and obviates the limitations imposed by the absence of serological cytotoxicity. Emphasis is placed on the need for recognition of the relationship between red blood cell and HL-A antigens.


Tuberculosis in Tanzania: a national sampling survey of drug resistance and other factors. This survey was conducted in 1969/70 in a random sample of 15 of the 61 administrative districts in Tanzania. It included clinics with a long established tuberculosis service (A group), those with a tuberculons service of recent inception (B group) and those with no specialised tuberculosis service (C group), and 3 additional centres of special interest. The aim was to obtain, for tuberculous patients newly registered for treatment during a specified 6-month period, information on: a) the proportions of patients with pulmonary and/or extra-pulmonary tuberculosis; b) the history of previous chemotherapy with antituberculosis drugs: c) the prevalence of bacteriologically-positive pulmonary tuberculosis; d) the prevalence of initial and acquired resistance to the standard antituberculosis drugs; e) the radiographic extent and type of disease and of cavitation. Of 1884 patients in the random sample, 87.4 per cent had pulmonary tuberculosis only, 2.5 per cent had pulmonary and extra-pulmonary tuberculosis and 10.1 per cent had extra-pulmonary tuberculosis only. Although there were differences in the proportions with extra-pulmonary disease in individual districts in each group there were no important differences in the average levels for the A, B and C groups. Of 256 extra-pulmonary tuberculous lesions in 237 patients, 58 per cent were lymph node, 26 per cent bone and joint and 12 per cent pleural, pericardial or peritoneal. Of 1694 patients with pulmonary tuberculosis with or without extra-pulmonary tuberculosis, 96.3 per cent gave no history of previous chemotherapy and 3.6 per cent a definite history. A sputum specimen from each of 1338 patients with pulmonary tuberculosis was examined by direct smear; 675 (50 per cent) were positive as were 694 (55 per cent) of 1257 cultures. For the patients with no history of previous chemotherapy the positivity rates were 59 per cent on smear and 66 per cent on culture for the A group, 53 per cent and 57 per cent for the B group and 35 per cent and 38 per cent for the C group. Of 636 patients with no history of previous chemotherapy and sensitivity test results, 9 per cent had a strain resistant to isoniazid and/or streptomycin. The total prevalence of resistance to isoniazid was 6 per cent and to streptomycin 4 per cent. Of 1278 patients aged 5 or more with a postero-anterior chest radiography available and a diagnosis of intra-thoracic tuberculosis in Tanzania, 86 per cent were reported at an independent assessment in London as having a lung lesion. Among the 1171 patients of all ages reported as having a lung lesion, gross extensive or moderate disease was present in 73 per cent of the radiographs and cavitation in 66 per cent. The type of radiographic disease was assessed as acute in 36 per cent, mixed-acute in 40 per cent, mixed in 19 per cent and chronic in 5 per cent. There was no evidence of associations between drug resistance and the extent or type of disease of presence of cavitation...


Microbiology of cutaneous tuberculosis.

Microbiology of cutaneous tuberculosis. Detailed bacteriological characteristics including drug sensitivity of 12 mycobacteria isolated from 51 cases of different types of cutaneous tuberculosis were studied; no mycobacterium could be isolated from remaining 39 cases. Nine were identified as M. tuberculosis and 3 as 'anonymous' mycobacteria. Acquired resistance to one or more than one drug (streptomycin, isoniazid, PAS and thiacetazone) was observed in 3 out of 9 strains of M. tuberculosis. All the 'anonymous' strains were resistant to all the drugs except ethionamide and ethambutol.


Radiotherapy: definitive, integrated and palliative therapy. Bladder cancer which invades the muscle is a serious problem which usually requires extensive therapy.  However, conservative therapy is often prescribed with the subsequent course all too freqently being relentlessly progressive.  Procrastination with aggressive therapy often results in the patient being unsuitable for such therapy when it is seriously contemplated...


[Transcervical adenomectomy with prior hemostasis (author's transl)]. After giving a brief description of the anatomy of the prostate, with special emphasis on the irrigation of same, the authors discuss 226 cases of adenoma of the prostate operated on transcervically with prior hemostasis, the transfusion consisting of a total of 111 of blood. In 68% of the cases, the urine became clear within 24 h after the operation. The probe was removed within the first 5 days in 70% of the cases. The complications found included: 9 cases of epididymitis (keeping in mind that deferent ducts are never linked), 4 cases of postoperative incontinence (in 3 cases the incontinence was temporary and disappeared within 3 months), and 3 deaths 2 weeks after the operation (1.3%) due to ictus, bronchopneumonia, and pneumonia, respectively. The hospital stay lasted 9 days, on the average. There were no cases of late hemorrhaging, fistula, or vesicle atony.


[A second bladder-neck intervention following open transvesical prostatectomy (author's transl)]. Over a 10-year period 27 patients required a second transurethral revision operation after previous open transvesical prostatectomy. The so-called sphincter-sclerosis, residual and recidivous adenoma, and prostatic cancer were the causes of repeated bladder neck obstruction. Prostatic cancer after prostatectomy seems to be more benign clinically, more differentiated histologically and is diagnosed sooner because it presents with earlier symptoms than does primary prostatic cancer.


Feline infectious peritonitis. Proteins of plasma and ascitic fluid. Electrophoreses of sera, plasma and ascitic fluids of cats with natural or experimental infectious peritonitis show important modifications. Special stainings of electrophoreses and chromatographic and immunoelectrophoretic technics characterized some of the modified proteins. In the experimental disease, fibrinogen, haptoglobin, transferrin, and probably orosomucoid are increased; in the natural disease, in addition to these modifications, the gamma-globulins are strongly increased; the immunoglobulins found in the often abundant ascitic fluid belong to the IgG class. Increased proteins such as fibrinogen, haptoglobin and orosomucoid and decreased albumin are aspecific aspects of inflammatory processes, whereas hypergammaglobulinemia appears in the course of immunological response. The rapid evolution of the experimental disease explains the fact that immunoglobulins do not increase.


Pulmonary hyalinosis in dogs.

Pulmonary hyalinosis in dogs. Pulmonary hyalinosis occurred in Beagles exposed to radon daughters with uranium ore dust. The lesion was composed of alveolar cells distended with material positive for periodic acid-Schiff (PAS) and oil red O that ultrastructurally consisted of a whorled arrangement of lamellar membranes suggestive of a storage disease. The high incidence in exposed dogs and the ultrastructural appearance suggested the material originated endogenously as a degenerative response to injury.


Progestogen-related gross and microscopic changes in female Beagles. Long-term studies of megestrol acetate and chlormadinone acetate in 100 female dogs are in progress. Doses of zero, one, 10 and 25 times the expected human dose of megestrol acetate and 25 times the expected human dose of chlormadinone acetate (on a milligram per kilogram body weight basis) are being given daily. During the first 4 years, eight dogs from each of the five groups were killed. The principal gross findings included enlarged uteri with mucoid material in the lumina, mammary development in dogs given middle and high doses of megestrol acetate and chlormadinone acetate, and thickened gallbladder walls in dogs given high doses of each. Histologic evaluation showed inhibition of ovulation for progestogen-treated dogs and suppression of ovarian follicular development with the high doses. Cystic endometrial hyperplasia was slight in the low-dose dogs and moderate to severe in most of the high-dose dogs; a few also had ulcerative endometritis and pyometra. The mammary glands of dogs given the middle and high doses produced lobules, acini, and secretion exceeding natural metestrus. Slight to marked cystic mucinous hyperplasia occurred in the gallbladders of most dogs given the high doses. Tow high-dose megestrol dogs had clinical signs and microscopic pancreatic, renal, and ocular changes indicative of diabetes mellitus.


Acquired B antigen disappearance by in vitro acetylation associated with A1 activity restoration. The chemical acetylation of RBC bearing the acquired B antigen led to the disappearance of the agglutinability by anti-B and restored the A1 specificity. The same results are obtained using RBC transformed in vitro by a Clostridium Tertium filtrate, where a deacetylase was reported.


[Improvement of parenteral feeding through the additional administration of a serum protein solution]. On 7 days - 5 postoperative - patients undergoing Billroth I or Billroth II were continuously infused apparatively witn an amino acid mixture of 100% and a glucose-xylitol-solution of 25% (group I). One group (group II) additionally received a serum protein solution. From the first preoperative day to the fifth day of the postoperative period we determined individual parameters in blood and urine. A decrease of serum phosphate level beginning on the first day after starting parenteral nutrition was observed. In the nitrogen balances, in total protein and albumin concentration there were no differences, but serum gamma-globulin concentration of group II was very low at the end of the parenteral nutrition period, a point which at present cannot be explained.


[Significance of metabolic parameters in Hodgkin's disease (author's transl)]. In 56 patients with Hodgkin's disease, the following bloodtests were carried out: erythrocyte sedimentation rate (ESR), fibrinogen, alpha2-globuline, serium iron concentrations and alkaline phosphatase activity. In some patients we additionally measured alkaline leucocyte phosphatase and serum ribonuclease activity. In our series ESR, serum iron and alpha2-globuline concentrations were the most sensitive metabolic parameters. A rise in fibrinogen concentration, alkaline phosphatase and serum ribonclease activity seems to indicate extensive disease. It is not possible, however, to discern between a state of remission and stage I by means of these parameters. ESR, serum iron and alpha2-globuline concentrations might be either elevated or normal in both instances. These parameters seem important in order to distinguish between a remission or stage I on the one hand and extensive disease in stage III and IV on the other hand. Concomitant findings of ESR above 40 mmh, elevated concentrations of fibrinogen and alpha2-globuline, as well as elevated alkaline phosphatase and serum and serum ribonuclease activity mostly indicate stage III or IV.


[Comparative studies on influences of physical and psychological strain on the ECG in coronary disease patients]. In 7 patients suffering from heart infarction the electrocardiogram was telemetrically registered during telecast of the world championship in football 1974. The same patients were tested on a bicycle ergometer with standardized loads of 50, 75, 100 and 125 Watt for 3 minutes. Heart rate under psychial strain did not exceed the rate of 50 Watt on bicycle ergometer. Only 1 patient showed some extrasystoles during ergometry. Looking at television all patients developed extrasystoles, in one case 122. If the FRG took part in the game, ventricular extrasystoles were two times, auricular extrasystoles were four times more frequent than during games between foreign countries. It is concluded that psychical strain is able to produce dangerous arrhythmias in patients suffering from coronary heart disease. This danger cannot be proved by bicycle ergometry.


Assessment of the mid systolic click late systolic murmur syndrome by noninvasive methods. In patients with the mid systolic click/late systolic murmur syndrome (MSC/LSM Sy) a ballooning, billowing or aneurysmal protrusion of one or both mitral leaflets has previously been described in angiographic studies. It is accepted that besides angiography noninvasive methods such as dynamic electrocardiography, phonocardiography, apexcardiography and echocardiography can provide valuable information in the diagnosis of this syndrome. Little is known however about diagnostic value and sensitivity of these methods. The dynamic Ecg, recorded during an average of 10 hours, showed in 8/20 patients (40%) one or several significant arrhythmias such as frequent ventricular premature contractions (VPCs) (greater than 12/min), multifocal VPCs, VPCs in a row or ventricular tachycardia, or 2 degrees SA-block. 12/20 patients (60%) showed in the phonocardiogram a movement of the click toward the first heart sound when the patients' position was changed from supine to sitting. In 40% of the patients a double or triple click was recorded. A late systolic bulge in the apexcardiogram was seen in 13/20 patients (65%). In one patient the recording was not successful. Echocardiographic signs of abnormal posterior movement (prolapse) of one or both mitral leaflets were recorded in 15/20 patients (75%). Five echocardiograms did not show any abnormality, but all these cases demonstrated in their apexcardiogram a late systolic bulge. We believe that in patients with the MSC/LSM Sy a mitral valve abnormality can be documented adequately by noninvasive methods.


[Changes of electrocardiography in salmonelloses and shigelloses]. In the period 1964-1973 there was done routing-like Ecg in 89 patients with typhoid with 24,7% pathologic findings, in 233 cases of paratyphus (13,3% Ecg abnormalities), in 91 patients with other forms of salmonellosis, mainly salmonella-enteritis (8,7% Ecg changes), and in 367 cases of dysentery from whom 12,5% had pathologic electrocardiograms. In all these patients the main changes represented low till negative T-waves in standard and precordial leads, sometimes combined with depressed ST-segment. In dysentery the loss of water and electrolytes played an important role in the genesis of Ecg changes, also for the prolongation of QT-interval in 2 cases. In 4 cases high and spiked P2 and P3 were present. The so much mentioned bradycardia was not much marked in out patients with typhoid, but a sinus bradycardia of 36/min was found in convalescence of a case of salmonella-enteritis where a salmonella from the group B was isolated from stool.


[Variable stainability of Purkinje cells]. The author refers about different staining of the Purkinje-cells with luxol-fast-blue, gallocyanin, thionin and toluidin blue, chrom-alum-hematoxylin-phloxin, impregnation according to Palmgren, lithium and iron-hematoxylin, combination of the staining with phloxin and the Palmgren-impregnation and about the different activity on the acid phosphatase. The phenomenon that in the same histological specimen the positive (dark, chromophile) and negative (light, chromophobe) cells are situated beside, is true for normal animals too, but the number of the dark Purkinje cells is conspicuous higher after stress situations (96-h. immobilisation, intermittent hypoxia). This finding interprets the author by the occurence of phospholipids by binding on the granulated endoplasmatic reticulum, but also as a property of the neuroplasm. The author emphasizes that the staining dualism "light -- dark" of the ganglion cells does not refer only to the ganglion cells of the spinal ganglions (et on some epithelial cells), but also on the Purkinje cells.


[Problems and experience in the light optical and histological presentation of glia cells]. 1. 8 histological techniques and 13 modifications derived from those were tested on usefulness for the demonstration of glial cells in the adult rat brain. From these methods the impregnation techniques of Golgi-Kopsch, Valenzuela y Chacón and Rio del Hortega were modified according to a scheme of variance to find out the optimal variants. 2. The impregnation quality depends on the animal species, the animal age, the health of brains, the brain area, the balanced proportion of the treatment stages and the biochemical state of the glial cells. 3. The silver impregnation techniques are not so specific that only one glial type is stained, but one type prevails. The silver carbonate procedure according to Hortega allows to impregnate oligodendrocytes, microglial cells and astrocytes in frozen as well as in paraffin sections. The method of Golgi-Kopsch is more suited for oligodendrocytes and microglial cells than for astrocytes. Following the procedure of Valenzuela y Chacón especially astrocytes, but also microglial cells allow impregnation in both frozen and paraffin sections. 4. The different demonstration qualities of the proved methods call for critical examination of absolute measurements of cell size, length of processes and ramification density. 5. The presence of cell groups of different disposition towards impregnation within a glial type speaks for a biochemical inhomogeneity of the glial types.


Laboratory procedures in adenoviruses. III. Sensitivity and specificity of serological tests (author's transl). 84 paired sera from patients with adenoviral infections were tested in neutralization and hemagglutination-inhibition (HI) against the homologous as well as against heterologous virus types of the respective subgroup. Only titer rises are considered as positive reactions. Sera of patients being infected with adenoviruses of subgroup I viruses (52 pairs, types 7, 3, 4, 14) exhibit high rates of positive reactions in the group-specific complementfixation (CF) and in homologous neutralization, lower rates in homologous HI. On the other hand, heterologous reactions are seen more frequently in neutralization than in HI. Patients with type 4 infections often show antibody responses with viruses of subgroup III (types 1, 2, 5, 6). In infections with these virus types, homologous antibody response is seen in approximately half of the cases only; heterologous reactions are observed rarely. Patients with type 8-infections (25 pairs) almost invariably show positive homologous neutralization, while HI or CF tests often fail to show rising titers. These patients, as well as 2 patients with type 19-infection, exhibit a remarcable high rate and range of cross-reactions in neutralization as well as in HI with numerous types if subgroup II and, to a lesser extent, with viruses outside of subgroup II. Although the relatively late appearance of type-specific antibodies (usually between the 10 th and 14 th days after onset of disease) is advantageous for the demonstration of antibody rises, the type-specific diagnosis of adenoviral infections should preferably be performed by isolation and typing of the virus. The heterologous reactions appear to be based on the presence of overlapping intertypic antigenic determinants rather than on an anamnestic antibody response.


DNA polymerases of murine LBN/b leukemic cells. In the post-mitochondrial fraction of murine LBN/b leukemic cells, four fractions with DNA polymerase activity (I, II, III, IV) were found. On the basis of ion exchanger affinity and poly(A), poly(C) and poly(Cm) replication ability, fraction I was classified as RNA-directed DNA polymerase of viral origin. On the basis of the differences in the ion exchanger affinity, molecular weight, template requirement, pH-dependence of enzymatic activity and NaCl concentration, divalent ion requirements and susceptibility to N-ethylmaleimide inhibition, fractions II, III and IV were classified as DNA-directed DNA polymerases beta, alpha and gamma, respectively. Three fractions, i.e. reverse transcriptase, and DNA-directed DNA polymerases beta and gamma,  were found to incorporate dTMP on a poly(A)-oligo(dT) template-primer. Despite the similarity of the reaction of DNA polymerases beta and gamma with poly(A)-oligo(dT), some other properties of these enzymes suggest that they represent distinct enzymatic entities.


Histogenesis of mucin in follicular mucinosis. An electron microscopic study. Many dilated cystic ergastoplasms, including their granular, filamentous contents, the developed Golgi apparatuses, and vesicles were observed in the cytoplasm of affected root sheath cells, which were accompanied by much amorphous agglutination of fine granular or flocculent materials in the interstices. These extracellular substances were stained with ruthenium red and seemed to correspond to increased metachromatic materials, probably composed of hyaluronic acid. Possible structures for hyaluronic acid were discussed. Mucin in follicular mucinosis seemed to be produced by secretory function of follicle cells.


Ultrastructure of localized myxedema.

Ultrastructure of localized myxedema. Five biopsies from three patients with localized myxedema were studied by electron microscopy. The dermis showed an overwhelming accumulation of microfibrils with knobs (acid glycosaminoglycans) and an amorphous material containing glycoprotein. Furthermore, evidence of degradation without new formation of collagen fibrils was found. Considerable numbers of mast cells with various types of granules, fibroblasts with dilated granular endoplasmic reticulum, and macrophages were seen in the mucinous areas.


Apoptosis in lichen planus and several other dermatoses. Intra-epidermal cell death with filamentous degeneration. In hyperplastic conditions of the skin, eosinophilic bodies are often observed in the epidermis and upper dermis. These have been named variously Civatte body of lichen planus and dyskeratotic cell of actinic keratosis. In the present studies, it was found that (i) these cells commonly contain whorls of distinct filaments (60-80 A), which may be attached to desmosomes; (ii) unlike keratinized cells, the cellular envelope was not thickened by the formation of the marginal band; (iii) the cytoplasm contains a large number of vacuoles; (iv) the nucleus is lost by condensation or diffuse disintegration; and (v) these cells could be dropped into the upper dermis and filamentous contents could be released to dermal phagocytes. From these observations it was concluded that Civatte bodies and other eosinophilic bodies of neoplastic and hyperplastic epidermis represent "filamentous degeneration" or premature keratinization of keratinocytes.


Cytophotometric studies of the DNA, nucleic acid and protein content of human liver cell nuclei. Cytophotometry was performed on human liver cell nuclei obtained from liver biopsies in 18 patients with normal or practically normal liver histology as judged by light microscopy. Imprints of liver cells and liver cells or liver cell nuclei obtained by different isolation procedures were studied. The nuclear DNA, total nucleic acid and protein content was evaluated after Feulgen, gallocyanin and naphtol-yellow staining and by UV-spectrophotometry. The nuclear area was obtained during the cytophotometric scanning procedure (Zeiss UMSP I). A total of 2,330 nuclei were investigated and approximately 80 per cent of these were diploid. The diploid value was confirmed by UV-spectrophotometry where a total nucleic acid content of approximately 7 pg was found. The nuclei could be grouped in classes corresponding to di-, tetra- and octaploid nuclei, according to their contents of DNA, total nucleic acid and protein and according to their size. The variation in nuclear contents was lowest for DNA with a coefficient of variation of approximately five percent, and highest for the protein content (15 per cent). Within diploid nuclei, insignificant as well as significant correlations between DNA content and size were found, but taken all together a weak positive correlation is likely. Higher correlations were found between nuclear nucleic acid content and size and between nuclear protein content and nuclear size.


Immunofluorescent studies of the skin in mixed cryoglobulinaemia and Schönlein-Henoch purpura. Immunofluorescence investigations in patients with mixed cryoglobulinaemia, Schönlein-Henoch purpura and purpuric allergic reaction with disseminated intravascular coagulation have been performed. In the lesional skin from 8 patients with mixed cryoglobulinaemia, granular vascular deposits of immunoglobulins of the same classes as those of the circulating cryoglobulins were detectable in all cases; complement and fibrinogen were concomitantly present. In the unaffected skin, IgM and complement were detected in the walls of the capillaries. In the early purpuric lesions from 6 patients with Schönlein-Henoch syndrome and from a patient with diseeminated vascular coagulation from acute allergic reaction to phenylbutazone, deposits of fibrinogen occurred mainly in the vessel walls.


The cutaneous reactions to kallikrein, prostaglandin and thurfyl nicotinate in chronic urticaria and the effect of polyphloretin phosphate. The erythema and wealing resulting from the application of thurfyl nicotinate ointment (Trafuril) and from the inoculation of kallikrein has been studied in patients with chronic urticaria and normal controls. Polyphloretin phosphate (PPP) suppressed the reaction in controls but in patients with urticaria it increased the reactions to Trafuril and had little effect on the kallikrein reaction. PPP also suppressed the PGE2-induced erythma in normal controls but not in urticaria patients. In a separate study using fibrinolysis autography, prostaglandin (PG) E2 and PGF2alpha depressed fibrinolysis in the skin of two pigs and both kallikrein and Trafuril suppressed fibrinolysis in human skin. It is suggested that the inflammatory reaction induced by thurfyl nicotinate and kallikrein is mediated in part by a prostaglandin-like action. Several anomalies in the action of Trafuril in skin diseases can be explained if such prostaglandin-like activity is mediated in part through inhibition of fibrinolysis.


Effects of dibutyryl adenosine 3',5'-cyclic monophosphate on human melanocytes in vitro. The effects of cyclic AMP and its analogue, dibutyryl cyclic AMP (DBcAMP) on the pigmentary system were studied by using human epidermal melanocytes in culture. The melanocytes responded to 1 mM DBcAMP with an increase in number, length, and complexity of dendritic processes. The effect of DBcAMP on the dendritogenesis was reversible. Melanin synthesis, as indicated by the uptake of tyrosine in the presence of an inhibitor of protein synthesis, was significantly stimulated by DBcAMP. The maximum stimulation was observed at concentrations of 0.5 mM, and 1.0 mM. The melanin synthesis increased after 12-hour treatment with DBcAMP and continued to increase with the prolonged treatment. Cyclic AMP, theophylline, sodium butyrate, or 5'-AMP at a concentration of 1 mM did not have any remarkable effect on the morphology or the melanin synthesis of the melanocyte. The results of this investigation indicate the possible role of the MSH-cyclic AMP system in the melanin pigmentation of human skin and represent a system for further study of the pathobiology of human melanocytes.


Phagocytic activity of polymorphonuclear leukocytes in generalized pustular psoriasis. The phagocytic activity of polymorphonuclear leukocytes from 10 patients suffering from generalized pustular psoriasis (GPP) has been studied. In all patients intracellular iodination capacity was measured, and 6 were studied regarding hexose monophosphate shunt activity (HMS) and bactericidal capacity. In 2 patients leukocytes from pustules were tested by the iodination reaction. Sera from 7 patients were incubated with normal leukocytes in a bactericidal test to evaluate the opsonizing capacity, which was found to be normal. Leukocytes from patients with GPP after 120 min of incubation with bacteria exhibited significantly impaired lethality compared with the controls, but no significant difference was found after 60 min. Iodination capacity and HMS activity were normal in all patients except in one, who was found to be deficient in myeloperoxidase. The possible significance of the impaired lethability of bacteria in the pathogenesis of GPP is discussed.


The relation of HL-A antigens to liver histology in methotrexate-treated psoriatics. A total of 45 patients with severe psoriasis vulgaris treated with Methylaminopterin NFN (Methotrexate Lederle) were HL-A typed in order to seek a possible correlation between HL-A antigens and the histological abnormalities in post-Methotrexate (MTX) liver biopsies. Highly significantly increased frequencies of w17 and TY (P less than 0.0001) were found, while HL-A13 Was only slightly increased. No statistically significant correlations were found between HL-A antigens and liver histology. HL-A typing does not seem to be of any predictive value in the pre-MTX screening of psoriatics in order to rule out patients with a high risk of liver damage during long-term MTX therapy.


Aggravation of psoriasis and occurrence of psoriasiform cutaneous eruptions induced by practolol (Eraldin). Twelve patients with cutaneous and/or eye symptoms caused by practolol have been studied. The morphological appearance of the skin eruptions was psoriasiform in 5, vesiculo-papular in one, and exfoliating dermatitis was seen in 2 patients. Keratoconjunctivitis sicca was the only adverse reaction to practolol in one and was present in 3 additional patients. Practolol was found to aggravate existing psoriasis in 3 patients and render it extremely refractory to therapy.


The spontaneous regression of keratoacanthoma in man. This study was undertaken to search for immune mediated mechanisms which could possibly be involved in the spontaneous regression of keratoacanthomas. In eleven patients with solitary keratoacanthoma, immunofluorescence studies did not reveal significant staining patterns, while skin tests, using autologous extracts of keratoacanthomas, were not compatible with true delayed cutaneous hypersensitivity reactions.


Oral trimethylpsoralen in the treatment of vitiligo. One hundred and 18 children with various clinical types of vitiligo, were treated with oral trimethylpsoralen (Trisoralen, P. B. Elder Co.) followed by exposure to sunlight or an artificial ultraviolet radiation source. In 61 of the adult patients, more than 80% of the vitiliginous areas became repigmented, 18 patients re-pigmented 50-80%, 13 repigmented less than 50% and 8 patients failed to repigment at all. In all of the children, more than 80% of the treated vitiliginous patches became repigmented. No side effects were observed. A follow-up study of the retention of the new pigment revealed that 72 adults (out of 76 followed-up) and 10 children (out of 11) without any treatment had retained 95% or more of the new pigment, 18 months after the completion of treatment. Oral trimethylpsoralen is therefore suggested for the treatment of vitiligo.


[Erythrocyte freezing in the presence of hydroxyethyl starch]. Biochemical properties of erythrocytes frozen in the presence of hydroxyethyl starch (HES) as preserving substance produced in this country were assessed. Erythrocytes were frozen adding HES in 4 different concentrations. The most favourable results were obtained with erythrocytes subjected to the action of HES in 30% and 35% concentrations. The erythrocyte 2,3-DPG level during freezing with 35% HES was within normal limits while it decreased by 10% after defreezing when 30% HES had been used. The ATP concentration in erythrocytes fell by 50% in relation to the initial value. A slight fall of P50 was observed and erythrocyte loss after defreezing did not exceed 10%. The 35% HES solution in a 0.15 mol/l sodium chloride solution was most favourable for freezing erythrocytes in liquid nitrogen. A simple method of preparation of defrozen erythrocytes for transfusion was elaborated.


Placento-thyroidal relationship in normal pregnancy. Estimations of serum HCT, HTSH, T4, T3, PBI, ETR, Triosorb, TBG-binding capacity, BMR and urinary total estrogen were made simultaneously in 160 women in normal pregnancy. TRH stimulation tests were made in 20 cases in each trimester of pregnancy. HCT was detectable even in early pregnancy, tending to increase gradually toward the terminal stage of pregnancy as serum thyrotrophin bioactivity showed. On the other hand, serum TSH level measured by radio-immunoassay remained essentially the same throughout the course of pregnancy as in the nonpregnant state, moreover, it was suggested by the TRH stimulation test that pituitary TSH secreting function of pregnant women was similar to that of the non-pregnant. These findings suggest that thyroid hyperfunction during pregnancy which is shown by progressively increased T3, T4, and PBI may not be due to high estrogen-high TBG binding capacity-low free thyroxinenegative feed back-high TSH secretion but to HCT originating from placenta. In spite of thyroid hormone increase, it is true that the clinical picture of hyperthyroidism is not manifest among normal pregnant women, and ETR remained within the non-pregnant range throughout the course of pregnancy. We have also demonstrated that Triosorb decreased progressively. This may be interpreted to be due to the increase of TBG binding capacity which is increased progressively and binds more of free thyroxine during pregnancy. Such a change in TBG binding capacity is well known to be caused by the effect of estrogen which is progressively increased during pregnancy. In a word, it is possible to say that there is a placento-thyroidal system in pregnancy; HCT elevates thyroid function and TBG increased by estrogen carries thyroid hormone to target organ.


The product albumin by alpha-globulin in the assessment of protein nutritional status. The product albumin by alpha-globulins (A X alpha) was determined in the serum of two hundred preschool children with protein-calorie malnutrition grouped according to clinical-anthropometric categories, and the results were compared with those obtained in a group of one hundred well-nourished children of the same age range. Though values obtained in malnourished children showed significant differences from those obtained in the controls, this parameter does not allow a biochemical distinction among the four groups studied.


The effect of different oral anticoagulants on diphenylhydantoin (DPH) and tolbutamide metabolism. The effect of bishydroxycoumarin, phenprocoumon, warfarin and phenindione on the metabolism of diphenylhydantoin (DPH) and tolbutamide has been studied in 54 patients. The half-lives of DPH and tolbutamide in blood following i.v. injections were studied in 33 patients before and after one week of anticoagulant treatment. Bishydroxycoumarin increased the mean half-life values of DPH from 8.8 to 37.4 hours and of tolbutamide from 4.9 to 17.5. Phenprocoumon prolonged DPH half-life from a mean value of 9.9 to 14.0 hours but did not change the tolbutamide half-life. Warfarin and phenindione did not affect DPH or tolbutamide half-lives. Steady state concentration studies in 21 patients showed a rise in serum DPH during bishydroxycoumarin and phenprocoumon treatment but not during treatment with warfarin and phenindione. A rise in serum tolbutamide was noted during treatment with bishydroxycoumarin. These findings suggest that bishydroxycoumarin inhibits the betabolism of DPH and tolbutamide and that phenprocoumon inhibits DPH metabolism. No effect on DPH and tolbutamide metabolism could be demonstrated following administration of warfarin and phenindione.


Specificity of antigenic fractions of tuberculin. Purified protein derivative from Mycobacterium bovis was separated into 6 fractions by electrophoresis at 1500 V/40 mA in pH 4.2 acetic acid-pyridine buffer. Further purification of one of the fractions on Sephadex G 25 column and by acid hydrolysis yielded antigen "PS" eliciting tuberculin reaction only in animals vaccinated with M. bovis BCG but not in those sensitized with M. avium and some fast-growing mycobacteria. The specificity of antigen "PS" was confirmed in vitro: the antigen induced blastic transformation only in lymphocytes of guinea pigs sensitized with M. bovis BCG.


Non-specific antimicrobial cellular resistance in sensitized guinea pigs. Specific delayed-type hypersensitivity was induced in guinea pigs with bovine albumin + complete Freund adjuvant, bovine gamma globulin + complete Freund adjuvant and BCG vaccine. The animals were subsequently tested for nonspecific antimicrobial resistance. Sensitized and control groups were challenged intraperitoneally with Listeria monocytogenes 2 hr after reinjection with the sensitizing antigen. The listeria content of the spleens was determined 1 or 5 days after the infection. The number of organisms recovered from the spleen one day after infection was significantly less in guinea pigs sensitized with bovine gamma globulin and BCG than in the control group; after 5 days no such difference was recorded. There was no difference between the bovine albumin sensitized and the control group 1 day after infection, while on the 5th postinfection day listeria counts were higher in the sensitized than in the control animals.


Antigenic relationship between Morganella morganii and Yersinia enterocolitica. Two new O antigens have been described for the Morganella morganii antigenic schema. O antigens of the strains representing the new serotypes (O43 :H2 and O44 : H19, 37) are related to Yersinia enterocolitica O9 and O17, respectively.


Molecular order of carbohydrate components in cell walls of bacteria, fungi and algae according to the topo-optical reaction of the vicinal OH groups. Selective topo-optical staining of vicinal OH groups with aldehyde-bisulphite-toluidine blue (ABT) has been used for studying the molecular structural order of polysaccharide components in microbial cell walls and capsules. (i) By intensive metachromatic staining in the light microscope, the method demonstrates carbohydrates, containing free vicinal OH or acylaminohydroxyl groups. (ii) The birefringence induced by oriented toluidine blue binding of the ABT reaction, provides information about the linear order of the reacting carbohydrate chains, a possibility not offered by other ultrastructural methods. Furthermore the different optical character of birefringence induced by ABT is indicative of the presence of (a) tangentially oriented polysaccharide chains in the cell wall of the molds and bacterial capsules; or (b) radially oriented OH groups suggesting helical glycan chains of peptidoglucomannane and mucopeptide in the yeasts and bacterial cell walls.


Cell wall lipopolysaccharide response to the ColIb plasmid mutants. Mutants of ColIb plasmid affected the synthesis of O-side chains of lipopolysaccharides (LPS) in Salmonella. The plasmid srd 25 (defective in colicin synthesis) caused a significant decline of rhamnose and mannose content and lack of abequose in LPS of S. typhimurium. The number of repeating units in O-side chains was decreased after the indroduction of srd 25. Cultures of S. typhimurium and S. enteritidis harboring drd2 (derepressed in colicin production) polymerised dideoxyhexose-defective O-side chains i.e. deprived of abequose and tyvelose, respectively. In dideoxyhexoseless S. meleagridis the content of rhamnose and mannose were reduced. The information for the alterations of Salmonella LPS was contained in the plasmid genome. In the wild-type plasmids the genes controlling the O-antigen changes were not expressed.


Amino acid metabolism and protein synthesis in streptomycin resistant Vibrio El Tor. Metabolic activities in relation to protein synthesis and amino acid utilization are altered in Vibrio El Tor after development of resistance towards streptomycin. Efficiency of in vivo and in vitro protein synthesis is markedly reduced in streptomycin resistant Vibrio El Tor. The rate of incorporation of 14C-amino acids into protein, uptake of 14C-valine and oxidation of certain amino acids are also altered.


The effect of rhizobiophages on the effectiveness of Rhizobium meliloti in symbiosis with lucerne. I. Effect of various strains of R. meliloti and rhizobiophages on the yield and nitrogen fixation in lucerne. Pot and test-tube experiments were carried out on the effect of phages on yield and nitrogen fixation in lucerne inoculated with effective and non-effective strains of Rhizobium meliloti. It was found that some strains of R. meliloti became activated by the phages, which was reflected in an increase in yield and in crude protein content in the plants, while others lost their effectiveness. The results of pot experiments were in line with those of the test-tube experiments and have demonstrated that the effect of phages on the effectiveness of Rhizobium is not simply negative or positive, but depends on the properties of the bacterial strains used.


The effect of rhizobiophages on the effectiveness of Rhizobium meliloti in symbiosis with lucerne. II. Effects of phages on the R. meliloti strains during passing them through the plant. Studies were carried out of the effectiveness of Rhizobium meliloti strains reisolated from the roots of lucerne inoculated with effective and non-effective strains in the presence or absence of rhizobiophages. Estimates were made of the changes in phage-sensitivity and lysogeny found in these strains compared with the parent strains. It was found that the character of the reisolated strains depended on the changes brought about by passing through the plant and on the activity of the phages.


RS virus components. I. Isolation and purification. RS virus was centrifuged in zonal rotor on 55% sucrose cushion. Three layers were collected: light (RS-LL) containing complement-fixing antigen, medium (RS-ML) containing both complement-fixing and virus particle antigens, and heavy (RS-HL) containing antigen associated with the virus particle. RS-LL was chromatographed on Sephadex G-200 column. Two peaks were obtained, containing complement-fixing activity (RS-LL-1 and RS-LL-2). After sonication (20 Hz, 30 min) RS-ML and RS-HL also were chromatographed on Sephadex G-200 column. Two protein peaks were obtained from each layer (RS-ML-1 and RS-ML-2 from medium, and RS-HL-1 and RS-HL-2 from the heavy layer), corresponding to RS virus proteins.


RS virus components. II. Antigenicity.

RS virus components. II. Antigenicity. Rabbits were immunized with RS virus components, isolated as described previously. The sera were tested by neutralization, double diffusion and complement-fixation. RS virus components induced low titers of precipitating and complement-fixing antibodies, and failed to stimulate the neutralizing antibodies.


Specificity of the agar gel microimmunodiffusion test in rabies diagnostics. The agar-gel microimmunodiffusion test (MIDT) with commercially available antirabies sera from different sources was applied to evaluation of rabies infection at about 500 brains from suspected animals. The high nonspecificity of the test and false positive results with nonvirulent materials were stated when compared with the histopathological, biological and FA tests. For evaluation of the nonspecificity and its cause, different antirabies sera and brain antigens from noninfected and rabid animals were used. Absorption of sera with tissue powders or immuno absorbent had a little influence on test specificity. The main causes of nonspecificity was the presence of antibrain antibodies in sera of producers-animals hiperimmunized by brain and spinal cord tissue vaccines. Application of the test to rabies diagnostics without purified control antigens and highly specific sera seems to be unjustified.


Laboratory investigations on "postvaccinall rabies" caused by live sheep-brain vaccine. The viral ethiology of postvaccinal complications among 30 dogs vaccinated by live antirabies vaccine (Umeno-Doi type, sheep brain vaccine) was fully confirmed. Three lots of virulent vaccine were inoculated subcutaneously into groups of "Wistar" rats according to the different schemes. Between the 1st and 12th day after the end of the vaccination there were no isolations of fixed virus in direct and blind i.c. passages of suspensions made from the thalamus area on succkling mice and rats. Also the viral antigen in the CNS of vaccinated but apparently healthy rats was undetectable. The "postvaccinal rabies" with the next isolation of fixed r.v. in the CNS was developed experimentally in rats only following the subcutaneous injection of the crude sheep-brain's and spinal cord's suspensions--composing the materials to production of antirabies vaccine.


The role of polymerase III in conjugation between E. coli K12 donor and recipient strains carrying dnaE ts mutation. The possible role of DNA polimerase III in conjugation was studied in a series of mutants temperature-sensitive for DNA polymerase III synthesis. The temperature-sensitive DNA mutation called dnaE 486 (ts) prohibits vegetative DNA replication at 41-45 degrees. Transfer of episome and chromosome from temperature-sensitive donor, carrying dnaE mutation to wild-type recipient strains, revertants and dnaE recipients was investigated. In the first two cases the number of Lac+ sexductants being even slightly higher at 43 degrees. Conjugational synthesis accompanying transfer involving the combination of dnaE (ts) thymine dependent and thymine independent donor and recipient strains measured by incorporation of 14C thymine was observed at the restrictive temperature. In the case of conjugation with temperaturesensitive recipient strains a drop of Lac+ sexductants and Pro+ recombinants may be as a result of disturbances in the synthesis of complementary strand in recipient, known to be dependent on pol III. However, the episome investigated by centrifugation in neutral CsC1 gradient after its transfer to the recipient with faulty polymerase III was double stranded (replicated) at the restrictive temperature.


Effect of dicumarol on the growth of some soil microorganisms. The effect of dicumarol on growth of selected soil bacteria: Azotobacter chroococcum, Arthrobacter globiformis, A. citreus and Bacillus megaterium was studied. The following minimum concentrations were inhibitory in vitro: Arthrobacter citreus--20 mug/ml., Bacillus megaterium--40 mug/ml., Azotobacter chroococcum--40 mug/ml. Arthrobacter globiformis--70 mug/ml. Cells of all microorganisms studied grown in the presence of dicumarol developed aberrant morphological forms.


Antigenic differences among bovine and porcine Factor VIII concentrates. Three anti-Factor-VIII antibodies from hemophiliacs were reacted with samples of batches of Maws commerical bovine and porcine Factor VIII concentrates manufactured over a 12-year period. The apparent antibody concentrations varied widely with the different batches of concentrates. The variations are probably due to intrinsic differences in the antigenic nature of the Factor VIII in the different preparations. With the older porcine concentrates, the low apparent concentrations may be in part due to the reaction of antibody with inactive Factor VIII. When animal Factor VIII concentrates are used in treating hemophiliac patients who have anti-Factor-VIII antibodies, the least reactive batch should be chosen. Random batches of animal concentrates are not suited as a standard for measuring anti-Factor-VIII antibody concentration.


Pancarditis in Whipple's disease: electronmicroscopic demonstration of intracardiac bacillary bodies. The advent of electron microscopy has repeatedly confirmed Whipple's original postulate that bacterial infestation might be the cause of intestinal lipodystrophy (Whipple's disease). We have recently studied two patients, a 67-year-old man and a 38-year-old woman, who died of untreated Whipple's disease, and both were found to have clinically unrecognized pancarditis. Histologically, PAS-positive histiocytes in foci of chronic inflammation were demonstrable in several organs, including the heart. Electron microscopy of autopsy tissues showed numerous intracellular and extracellular rod-shaped bacillary bodies and serpiginous membranes. The bacillary bodies, some sectioned transversely and others longitudinally, were about 0.2 mum wide and 2 mum long; each had a double-layered cell wall. These bacillary bodies have not been previously identified in the heart, and may be casually related to cardiac lesions occurring in many untreated cases of Whipple's disease.


Nonfunctioning argyrophilic tumor (APUDoma) of the hepatic duct: simplified methods of detecting biogenic amines in tissue. This report documents a foregut endocrine neoplasm in an unusual location, the main hepatic bile duct, and describes simplified methods of detecting similar endocrine tumors. Formaldehyde fume-induced fluorescence (FIF) is a quick, specific method to demonstrate biogenic amines. FIF combined with histochemical procedures showing abundant alpha-glycerol phosphate dehydrogenase and esterases indicates the neural crest origin of these and related neoplasms. Amine precursor uptake and decarboxylation (APUD) is a significant marker of these cells and gives them their name.