Mitochondrial and nuclear glutamate dehydrogenases in Chinese hamster ovary cells in culture. Nuclear glutamate dehydrogenase (EC 1.4.1.3) activity has been demonstrated in Chinese hamster ovary cells. Some characteristics of this enzyme have been examined and compared with those of the mitochondrial glutamate dehydrogenase from the same source. Differences were detected in the extent of the activation by inorganic phosphate, in the pH versus activity curves, in the affinity of the two enzymes for the cofactor NAD+ and in the electrophosretic mobility. A different rate of decay of the two enzymes has been observed in cells grown in the presence of chloramphenicol. Immunological studies show that, as in ox liver, the nuclear enzyme has specific antigenic determinants besides those in common with mitochondrial glutamate dehydrogenase. Finally, experiments of thermal inactivation indicate a higher stability of the mitochondrial enzyme.


Current questions on the tumour-associated antigens of chemically-induced tumours II. In search of practical uses. With special reference to tumours which were chemically-induced in animals, we present a brief review on the applicability of the results obtained, as to immunoprophylaxis and to immunotherapy. A search is made onto what are the mostly representative animal models for human studies. From the present available data, experimental chemically-induced bladder and colonic tumours seem to be the "best" models for human counterparts.


Diatom diversity as a function of insecticidal treatment with a controlled-release formulation of chlorpyrifos. Following treatment with a controlled-release formulation of chlorpyrifos, no substantial differences between diversity estimates were evident for 6 post-treatment weeks for both treated and control plots. By post-treatment week 12, significant decreases in diversity estimates occurred in treated plots, suggesting that a directly proportional relationship exists between chlorpyrifos concentrations and reduced diatom colonization through time. Comparatively, diatom colonization progress "normally" and populations proceeded toward "maturity" in the control plots as indicated by the increase in the diversity estimates between post-treatment weeks 6 and 12. Any adverse effects due to the insecticidal treatment, however, were not considered to be environmentally deleterious in light of the restricted type of habitat (rice culture) in which the chlorpyrifos was used.


Volume estimation of the seminal vesicles by means of transrectal ultrasonotomography: a preliminary report. The volume of the seminal vesicles was estimated in 27 cases by means of transrectal ultrasonotomography. The seminal vesicles of normal subjects were found to be generally larger than in cases of prostatic hypertrophy or cancer. The volume of the seminal vesicles in Klinefelter's syndrome was remarkably small. The correlation between the volume of the seminal vesicles and of the prostate is given.


The effect of bromocriptine in patients with benign prostatic hypertrophy. A double blind trial of bromocriptine in the treatment of 14 patients with the symptoms of benign prostatic hypertrophy is reported. 5 patients did not complete the trial mainly because of side-effects due to an overdosage of bromocriptine. In the remaining 9 patients there was symptomatic improvement in 6, with objective evidence of improvement in 3 patients on flow rate measurements. The possible modes of action of bromocriptine in this condition are discussed.


Immunological studies in pre-eclamptic toxaemia. Although five patients with severe pre-eclamptic toxaemia (PET) had increased anticomplementary activity in their serum, there was no evidence of complement activation in the plasma of four of the five patients. These results are not implicated in the pathogenesis of PET. No significant correlation was found between anticomplementary activity and pregnancy-associated alpha2-glycoprotein.


Polyarthritis in adults with hypogammaglobulinaemia and its rapid response to immunoglobulin treatment. Five patients with primary hypogammaglobulinaemia developed a severe polyarthritis that had some features in common with rheumatoid arthritis. Their joint disease could be distinguished from rheumatoid arthritis, however, by the dramatic improvement after gammaglobulin treatment. The arthritis of hypogammaglobulinaemia can, therefore, be included among the few potentially curable polyarthritides.


Axonal transport in serotonin neurons of the midbrain raphe. The projections of serotonin-containing neurons of the midbrain raphe nuclei (nucleus raphe dorsalis, nucleus centralis superior) are studied by analysis of axonal transport of labeled amino acids. These results are correlated with regional alterations of serotonin content following midbrain raphe lesions which produce significant serotonin depletion in nearly all regions of the central nervous system. Twenty-four hours following injection of 100 muCi [3H]proline, raphe neurons have taken up labeled material and transported it, presumably as protein, to telencephalon, diencephalon, brain stem, the cerebellum and the spinal cord. This transport appears to take place predominantly in serotonin neurons. After injection of 100 muCi [3H]5-HTP into nucleus raphe dorsalis or nucleus centralis superior, the pattern of regional distribution of transported material is very similar to that obtained with tritiated proline. Selective lesions of serotonin terminals with 5.6-DHT result in greatly diminished axonal transport of proteins to all telencephalic, diencephalic and mesencephalic areas as well as to cerebellum, pons-medulla and spinal cord. Unilateral destruction of the medial forebrain bundle results in significant reduction in axonal transport of labeled material to ipsilateral telencehalon and thalamus. These results provide further support for the view that serotonin neurons of the midbrain raphe nuclei project widely throughout the neuraxis to telencephalon, diencephalon, brain stem, cerebellum and spinal cord.


Effect of bleomycin on deoxynucleotide-polymerizing enzymes from human cells. DNA polymerases alpha and beta from Molt-4 cells are inhibited by bleomycin, whereas DNA polymerase gamma assayed with poly-(A)-(dT)12-18 as the template primer or terminal deoxynucleotidyl transferase assayed with activated DNA, poly(dA), (dG)12-18 or (dA)12-18 as the initiator are not inhibited by this antibiotic. Inhibition by bleomycin increased the Km for template DNA but not that for dTTP. Increasing amounts of bleomycin did not affect the Vmax for DNA polymerase alpha or beta when the amount of template DNA was varied but it reduced the Vmax for these enzymes when dTTP was varied. Moreover, the addition of extra template reversed the bleomycin inhibition but the addition of extra enzyme did not. Although dithiothreitol was required for bleomycin inhibition of DNA polymerase activity, bleomycin preincubated with dithiothreitol (or beta-mercaptoethanol) at pH 6.5 to 9.0 lost its inhibitory activity. This was not the case when DNA was also included in the preincubation mixture. The results obtained in this study indicate that bleomycin inhibits DNA polymerases alpha and beta by a thiol reagent-dependent interaction with the template. Thus, the antitumor activity of bleomycin may be greatly influenced by the concentration of sulfhydryl compounds and their proximity to DNA in the target cells.


Shared antigens between bacteria and guinea pig line 10 hepatocarcinoma cells. This study was undertaken to investigate the possibility that Listeria monocytogenes, Brucella abortus, and Salmonella typhimurium share antigenic components with guinea pig line 10 hepatocarcinoma cells. Rabbits were immunized with sonicates of these bacteria or line 10 tumor cells. Other rabbits were immunized with line 1 cells, a tumor with antigenic characteristics different from those of line 10. The binding of antibodies to radiolabeled antigens prepared from extracts of bacteria and line 10 cells was studied by precipitation of radiolabeled antigen-antibody complexes with anti-rabbit immunoglobulin. Antibodies in sera from rabbits immunized with these bacteria and line 10 cells bound both the labeled bacteria and line 10 antigens. Antibodies in sera from rabbits immunized with line 1 cells did not bind the bacterial antigens. Inhibition studies involving reactions between radiolabeled Listeria and line 10 antigens and antibodies to Listeria and line 10 cells confirmed that the binding reactions were specific and that line 10 cells shared antigens with Listeria cells. The possibility that B. abortus and S. typhimurium also shared antigens with line 10 cells was suggested. Whether antigens shared by these bacteria and line 10 cells are identical with tumor-specific antigens was not determined.


Characterization of carcinoembryonic antigen fractionated by concanavalin A chromatography. A small percentage (15%) of particles morphologically distinct from carcinoembryonic antigen (CEA) cruller-like particles (averaging 9 x 40 nm) as determined by electron microscopy, and previously presumed to be impurities, has been removed from CEA preparations by concanavalin A (Con A) affinity chromatography. The first of four affinity fractions constituted 45% of the recovered weight; the last, which constituted 17% of the recovered weight, was obtained only after a 2-day soak in 20% alpha-D-mannopyranoside. The affinity fractions were of essentially equivalent specific activity, morphological appearance, and composition. They all demonstrated markedly different chemical compositions and an approximately 100-fold higher specific activity than the bulk of the nonaffinity material. Thus, the bulk of the Con A-nonbound fractions apparently is not CEA but contaminating particles of several different varieties. Compositional analyses further indicated that the mannose content of various CEA fractions was directly correlated with Con A binding affinity, amino acid content, morphological type, and CEA specific activity. This led to the conclusion that the protein chain of the various CEA preparations fractionated by us is characteristic. An attempt to fractionate very impure crude CEA by Con A affinity chromatography indicated that this scheme cannot by itself produce as pure CEA as standard methods. Thus, for the first time the effectiveness of the purification of a glycoprotein has been successfully monitored by electron microscopy, demonstrating that the cruller-like appearance of the CEA particle is closely related to antigenic specificity. Whether antisera to our affinity fractions will improve the specificity and sensitivity of the clinical assay is currently being investigated.


Structural and immunological relationships of isoferritins in normal and malignant cells. Ferritins from normal adult human liver and heart were compared with ferritins from a lung carcinoma metastatic to liver and from HeLa cells on the basis of their isoferritin profiles, subunit composition, and immunological relationships. Each ferritin preparation gave different isoferritin profiles, but several contained common isoferritins. All of the tumor isoferritins had counterparts in the normal tissues. All ferritins contained similar subunits but in different proportions. Qualitative differences were demonstrable in some ferritins with antibodies to different tissue ferritins. These differences correlated with the subunit composition of the ferritins. By appropriate absorption, an antibody population was obtained that was apparently specific for one subunit type. Heart ferritin gave lines of apparent identity with the tumor ferritins with these antibodies. It is concluded that tumor ferritins are not tumor-specific antigens but correspond to isoferritins in normal adult heart.


Erythroid differentiation in cultured Friend leukemia cells treated with metabolic inhibitors. The induction of erythroid differentiation in the T3-C12 clone of Friend leukemia cells by dimethyl sulfoxide is accompanied by reduction in viral RNA-dependent DNA polymerase activity with increased cellular delta-aminolevulinic acid synthetase activity and hemoglobin synthesis. These cells were treated with a variety of compounds to determine whether other durgs are capable on inducing erythroid differentiation. While several hormones, inhibitors of RNA synthesis, organic solvents, inhibitors of DNA polymerase, sulfhydryl inhibitors, and inducers of delta-aminolevulinic acid synthetase administered singly did not stimulate hemoglobin synthesis like dimethyl sulfoxide, inhibitors of DNA and RNA synthesis such as adriamycin, mitomycin C, and hydroxyurea:mithramycin were synergistic in stimulating erythroid differentiation.


Isolation of a novel glycoprotein (EDC1) from the urine of a patient with acute myelocytic leukemia. Gel filtration of urine from Patient ED with acute myelocytic leukemia showed a prominent protein peak with elution position corresponding to molecular weights of 20,000 to 35,000. The protein (EDC1) was isolated in pure form by sequential gel filtration and ion-exchange chromatography. Molecular weight of purified EDC1 was 27,000; it contained 27% carbohydrate and was rich in half-cystine (5% of residues). EDC1 was antigenically and chemically distinct from the recognized glycoproteins of normal plasma. With a specific rabbit antiserum and 125l-labeled EDC1, a radioimmunoassay for the glycoprotein was developed. Both noncancer and cancer plasmas contained immunoreactive material. In noncancer plasma, all the immunoreactivity was eluted from Sephadex G-75 and G-200 in position corresponding to molecular weights of 60,000 to 100,000 (Peak 1). In cancer plasma, an additional peak of immunoreactivity was eluted in the position corresponding to EDC1 (M.W., 20,000 to 30,000; Peak 2). Eighty-six % of urines from patients without clinical cancer were nonreactive in radioimmunoassay (less than 0.1 microgram immunoreactive EDC1 per ml); 11 and 3%, respectively, contained immunoreactivity equivalent to 0.1 to 0.9 and 1 to 9 microgram EDC1 per mi, entirely of Peak 1 type. Ninety-one % of urines from patients with disseminated cancer contained immunoreactivity equivalent to 10 to 9,999 microgram EDC1 per ml, primarily of Peak 2 type.


Uridine 5'-diphosphate-galactose:glycoprotein galactosyltransferase activity in the ovarian cancer patient. Uridine 5'-diphosphate-galactose:glycoprotein galactosyltransferase activity was demonstrated in homogenates of normal ovary and ovarian epithelial adenocarcinomas. The specific activity of the enzyme in ovarian tumors was 3 to 5 times higher than in normal ovaries when the enzyme was assayed under identical conditions. The glycoprotein fetuin, from which terminal sialic acid and penultimate galactose were removed (fetuin minus N-acetylneuraminis acid and galactose), acted as an excellent exogenous acceptor. Galactosyltransferase from normal ovary and ovarian tumor cells had similar properties. Both required Mn2+ and Triton X-100 and had broad pH optima between 5.5 and 7. Galactosyltransferase activity was also measured in serum samples from ovarian cancer patients and normal healthy individuals in the presence of fetuin minus N-acetylneuraminic acid and galactose as exogenous acceptor. The enzyme levels were significantly elevated in the sera of ovarian cancer patients as compared to normal controls. The differences in the levels of this enzyme in the tissues and sera of normal individuals and ovarian cancer patients were not due to differential levels of the degrading enzymes such as uridine 5'-diphosphate-galactose pyrophosphatase or beta-D-galactosidase. Serial determinations were carried out on the sera of 5 ovarian cancer patients over a long period of time. The serum level of galactosyltransferase activity appeared to correlate with tumor volume as well as with the clinical status of the patient, which suggests possible leakage of the tumor enzyme into the host sera. Serial determination of this enzyme level in ovarian cancer patients seems promising in measuring tumor progression or success of therapeutic approaches.


On the occurrence of argyrophil cells in salivary glands. Salivary glands (parotid, submandibular and sublingual glands) of nine mammalian species were investigated with respect to presence and localization of argyrophil and argentaffin cells. With the exception of the parotid gland of the rat, no positive staining was observed within the examined glands. In the rat parotid distinctly argyrophil cells could be demonstrated in the intercalated ducts. Histochemical studies of the cells, ultrastructural analysis of their cytoplasmic granules as well as their reactions to certain drugs indicate that these cells are of exocrine rather than of endocrine nature. After a subcutaneous injection of pilocarpine, the intensity of the argyrophil staining was markedly reduced. No specific catecholamine fluorescence could be detected within the cells, not even after pretreatment of the animals with high doses of L-DOPA. The membrane-bounded cytoplasmic granules of the intercalated duct cells furthermore displayed a strong positive staining reaction after treatment of ultrathin Vestopal sections with the periodic acid-chromic acid-silver technique of Rambourg et al. (1969).


Visualization of the retino-hypothalamic projection in the rat by cobalt precipitation. The technique of cobalt sulfide precipitation combined with Timm's sulfide-silver method for intensification of heavy metals was used to delineate the retino-hypothalamic projection of the rat. Freshly isolated rat brains were dissected and a solution of cobaltous chloride was applied to one of the cut optic nerves. Sixteen hours later, after cobalt ions had passed into the brain along the entire length of the optic fibers, the preparation was treated with ammonium sulfide to precipitate the cobalt as cobalt sulfide. In thick light microscopic sections, cobalt-filled axons were visualized as black fibers against a light gold background. Such fibers were observed to leave the posterior medial portion of the optic chiasm and, after arching dorsally, to project into the posterior fifth of the suprachiasmatic nucleus (SCN), as well as into the rostral part of the arcuate nucleus. Neither bifurcation of these axons nor looping of the axons back to the chiasm was seen. Most fibers projected to the SCN contralateral to the filled nerve, but the projection represented less than 0.1% of the total number of fibers in one optic nerve. These observations are considered to be graphic evidence of a retino-hypothalamic projection. The interpretation of the cobalt method is discussed, as are the functions of the connections that have been observed.


On argyrophil reactions of endocrine cells in the human fetal pancreas: a light and electron microscopic study. The endocrine cells in the pancreas of five human fetuses with gestational ages of 18-20 weeks were examined by light and electron microscopy with special regard to argyrophil reactions. B-cells and typical A- and D-cells were easily identified electron microscopically on the basis of their typical secretory granules. In the Grimelius argyrophil silver stain, a concentration of silver grains over the less electron dense peripheral mantle of the A-cell secretory granules was observed by electron microscopy. In the Hellerström and Hellman modification of the argyrophil Davenport alcoholic silver stain, silver grains were concentrated over the internal structures of the D-cell secretory granules. With this stain an accumulation of silver grains was also seen at the surface of the A-cell secretory granules. The argyrophil reaction of the A-granules was less pronounced than in the D-cells. In addition to B-cells and A- and D-cells, two other types of endocrine cell were observed by electron microscopy. These cells were argyrophil with the silver impregnation method of Grimelius. The electron microscopic findings at least partly explain the frequent overlapping between the two staining methods observed at the light microscope level.


Purification and radioimmunoassay of human alpha-1-fetoprotein: the effect of aggregates on the radioimmunoassay. A simple method for the extraction of alpha1-fetoprotein from fetal serum in high yield and of sufficient purity for use as radio-iodinated tracer in a radioimmunoassay is described. The purified alpha1-fetoprotein formed aggregates during storage. The presence of 125I-labelled aggregates affected the double antibody radioimmunoassay by increasing the non-specific bound, resulting in decreases assay sensitivity. Freshly prepared 125I-labelled monomer alpha1-fetoprotein is prepared for each assay. The radioimmunoassay described is sensitive to between 100 and 200 pg per tube. Normal adult serum levels were found to be in the range from less than 1 mug/l to 16 mug/l.


Immune complexes in skin of NZB/NZW mice. Skin biopsies were performed on female (NZB X NZW)F1 mice (B/W) at ages ranging from 1 to 12 months. The control strain was the C57B1/6. Immunoglobulin G and beta 1C globulin deposition was sought using the indirect immunofluorescence method. Both globulins were detected consistently at the dermal-epidermal junction from the age of 6 months. The pattern of staining was granular, and progressed from focal deposition to confluent and diffuse involvement of the basement membrane at 9-10 months of age. No staining was observed in any of the C57B1/6 mice up to 14 months of age. These findings increase the resemblance of B/W disease to human systemic lupus erythematosus.


Drawings as a therapeutic medium. The treatment of separation anxiety in a 4-year-old boy. This paper describes a 4-year-old boy whose presenting problem was anxiety. The treatment used is of particular interest because of the graphic exposition of the development of the therapeutic alliance, the immediacy of the Oedipal issues, and the usefulness of child's drawings as a therapeutic medium. The phases of therapy are reflected in the drawings, as is identification with the therapist and the integration of loss when patient and therapist terminate.


[Cardiac arrhythmias in the chronic phase of myocardial infarction (author's transl)]. Incidence and type of arrhythmia, using continuous ECG monitoring, were examined in 79 patients six weeks to 24 months after acute myocardial infarction, the findings being compared with arrhythmias at rest and on exercise, and correlated with signs of heart failure and coronary insufficiency. Arrhythmias were revealed in 16% (n = 13 at rest) and 19% (n = 10 of 52) on exercise. But continuous ECG monitoring revealed it in 86% (n = 68). On the other hand, arrhythmias may occur in the resting ECG but be rare on continuous monitoring. Ventricular extrasystoles were the most common arrhythmia, often multifocal and giving rise to bigeminy, only rarely as a result of exercise. Such extrasystoles usually showed right or right and left bundle conduction delays. Their frequency increased with increasing age of the patient. They were particularly frequent in patient who already had cardiac symptoms at rest, in patients with a large heart, and those who could not easily be exercised. Arrhythmias were more frequent a year after infarction than shortly before discharge from hospital or six months after infarction.


Epileptogenesis of human limbic neurons in psychomotor epileptics. Different stages of epileptogenesis of neurons in deep temporal lobe structures have been studied with fine wire microelectrodes chronically implanted in patients with drug-refractory psychomotor epilepsy. The interictal firing patterns of single neurons ipsilateral to the focus (identified by EEG seizure onset and/or neuropathology studies and seizure reduction following anterior temporal lobectomy) often exhibited burst when contralateral neurons did not. The intraburst sequence of action potentials was not organized or reliable except in one focal hippocampal neuron. Bursts of action potentials often occurred in the absence of regional sharp waves recorded with the same microelectrode; however, sharp waves with fast rise times were almost always associated with action potentials from nearby neurons. During sub-clinical EEG seizures, when EEG abnormalities did not propagate contralaterally, neurons were activated in rough proportion to the intensity of EEG activation and extent of spread of seizure activity to ipsilateral temporal lobe structures. During clinical seizures, involving both hemispheres, firing rates of neurons near the focus increased during the small amplitude high-frequency EEG phase and decreased as this high-frequency rhythmical waveform increased in amplitude. Variable firing rates followed until the clonic EEG phase, where a reliable excitation of neurons occurred during the sharp waves and strong inhibition during the following slow waves. Between these sharp--slow wave events many neurons were inhibited, but to a lesser degree and for longer than the post-excitation inhibition. These neurophysiological phenomena are discussed in relation to the literature on cellular mechanisms of epileptogenesis in experimental epilepsy.


The EEG response to repetitive photic stimulation in various regions of the chicken brain. The EEG response in the chicken to repetitive photic stimulation was studied by frequency analysis and by the averaged response. Evoked responses were observable not only in stations along the visual pathway but also in broad areas apart from the visual pathway. The electrical activity in the archistriatum showed a marked response to flickering stimuli, indicating that this area is involved in the visual function in the chicken. In other telencephalic areas, photically evoked potentials could not be clearly demonstrated in the EEG records. In the hypothalamus and the nucleus rotundus of the diencephalon and in the nucleus reticularis superior of the mesencephalon, sinusoidal waves appeared during stimulation at 8-13/sec. No rhythmic after-discharge was observed following termination of photic stimulation. These finding are indicative of the difference of the visual response in the chicken from previously reported responses in other species.


Recording of the cochlear microphonic potential with surface electrodes. The cochlear microphonic potential was recorded in human subjects with surface electrodes (earlobe clip and scalp vertex disc) and an averaging procedure. Special precautions were taken to identify and separate artefactual, neural and microphonic components. These included shielding of the earphone, a rubber tube to introduce a time delay between artefact and biological response and white noise to mask the neural component. The cochlear microphonic potential was larger in amplitude in response to low frequency sounds and had a high threshold. Two clinical cases of cochlear hearing loss are presented, both lacking neural responses. The cochlear microphonic potential was present in one of them (i.e., neural hearing loss) and absent in the other (i.e., sensory hearing loss).


Excitability of human motoneurones after discharge in a conditioning reflex. A new method of exploring the H reflex excitability cycle is proposed which permits the exploration of motoneurones that discharged in response to the conditioning shock. This is made possible by using a test stimulus supramaximal for the alpha motor fibres of the nerve, which causes a collision in the alpha fibres between the orthodromic conditioning reflex volley and the antidromic motor volley. As a result of this collision, a test reflex contraction appears in the EMG, due to motoneurones that have already discharged in the conditioning response. This method permits studies in human subjects of the poststimulus refractory period of motoneurones as well as the early inhibitory phenomena related to the conditioning stimulus and response. It is shown that during the initial period of the classical H reflex excitability cycle, the test shock explores only alpha motoneurones corresponding to a fringe of cells subliminally excited by the conditioning stimulus. The difference between the populations of motoneurones tested by the two methods explains the differences between the features of the cycles obtained.


Regulation of slow potential shifts in nucleus reticularis thalami by the mesencephalic reticular formation and the frontal granular cortex. Novel stimuli or electric stimulation of the mesencephalic reticular formation (MRF) produced large positive slow potentials (SPs) in rostral nucleus reticularis thalami (RVA) that accompanied the negative SPs known to occur in frontal cortex. SP durations (20-30 sec) were similar to the periods of unit inhibition that occur in RVA following MRF stimulation. Trains of 8 c/sec medial thalamic stimuli produced phasic negative SPs in RVA similar in duration to the intervals of unit excitation that follow each stimulus pulse. These results suggest that the polarity and duration of the SPs in RVA reflect changes in excitation of the underlying neurons. Direct activation of a specific region of RVA produced complete inhibition of visual cortex responses evoked by optic tract stimuli, a finding which suggests that RVA has an inhibitory action on the thalamus. A tone reinforced by electric shock also elicited SPs in frontal cortex (negative) and RVA (positive). In contrast to the long duration of the MRF- or novelty-elicited SPs, the durations of the conditioned SPs were phasic and were regulated by the tone--shock interval. Bilateral cryogenic blockade of the interconnections between the frontal cortex and medial thalamus abolished SPs of all origins in the frontal cortex. The blockade also abolished conditioned SPs in RVA, but did not affect the MRF-elicited ones. Thus, the subcortical SPs that accompany orienting to novel stimuli are distinct from those which occur during the higher cognitive process of conditioned expectancy and require the integrity of the mediothalamic-frontocortical system.


The relation between monosynaptic spinal reflex amplitudes and some EEG alpha activity parameters. Some investigators have found that in normal human subjects the amplitudes of repetitively evoked tendon reflexes decrease as the number of evoked reflexes progresses. The question whether this decrement is a spinal phenomenon or that it can be ascribed to supraspinal influences that are related to the degree of cortical activation was investigated in two experiments designed to test whether a relation exists between the Hoffmann (H) and Achilles tendon (T) reflex amplitudes and EEG alpha activity parameters during a rest condition. The principal results can be summarized as follows: 1. A constant alpha index was accompanied by stable reflex amplitudes. 2. A decreasing alpha index was accompanied by decreasing reflex amplitudes. 3. A positive relation was found between H and T reflex amplitudes. 4. A positive relation was found between alpha index and alpha amplitude. 5. No circumscribed relation was found between the alpha wave period on the one side and the alpha index and amplitude on the other side. Apparently these relations are dependent on the state of activation of the subject. 6. The relations between the reflex amplitudes and the alpha parameters showed great interindividual differences.


A proportional controlled thermoregulator circuit of simple design. This paper describes a proportional controlled thermoregulator circuit. The instrument controls D.C. current in the heating device, precluding the introduction of radiated interference by the heater. This circuit is capable of regulating the temperature of a typical decerebrated cat to within 0.1 degrees C. A dual channel temperature regulator can be constructed for about $ 300.


Ultrastructural visualization of Cabot rings in pernicious anemia. Using the ammoniacal silver stain, Cabot rings were identified in peripheral blood erythrocytes from patients with severe untreated pernicious anemia. Ultrastructural studies of these erythrocytes showed silver deposits in partial loops and figure-eight forms, indicating that arginine rich histone may be a prominent component of the Cabot ring.


Structural studies of insulin and insulin derivatives using various immunologic indicators and antibody populations. These studies suggest that the immunologic indicator in the radioimmune assay, 125I-iodoinsulin, selects antibody populations from within the antiserum that interact with determinants distant from the solvent surface on the insulin molecule to which iodine is substituted. Evidence is presented that the connecting peptide of proinsulin is in close proximity to regions on the solvent surface of the A-chain of insulin that include the tyrosyl residues at A-14 and A-19. A marked immunologic cross-reaction between derivatives of insulin with perturbations in the regions of tyrosyl A-14 and A-19 was noted in the radioimmune assay employing desalanine-(B-30)-desasparagine-(A-21)-insulin antiserum. This observation is consistent with the presence of a restricted population of antibodies in such antisera that is directed toward immunologic determinants in or near the insulin dimer site. The apparent immunologic activity of insulin derivatives depends on which antibody populations from the antiserum pool can react with the immunologic indicatory employed on the one hand and on the composition of antibodies in that antiserum on the other. These studies indicate that the specificity of antibody populations in a given antiserum can be identified and their levels quantitated with several assay systems, each employing one of a variety of indicators.


Combined effect of X-ray and bleomycin on cultured mammalian cells. Investigations were carried out with cultured mammalian cell lines to assess the effect of combination of radiation and Bleomycin. When cells were treated with the antibiotic before and during exposure to X-rays, a slight potentiating effect was consistently found. The magnitude of potentiation of the radiation effect appeared to depend on the concentration of Bleomycin. When the cells were treated pulsewise at various periods after irradiation, the potentiating effect was found only during the first 2 hr. Simultaneous application of X-rays and Bleomycin provided the greatest effect.


Establishment of a cell line and its clonal sublines from a patient with hepatoblastoma. By cloning from cultivation in vitro of a human hepatoblastoma 3 epithelial cell lines were established, all of which produced alpha-fetoprotein and one of which did albumin in culture. Observation with electron microscopy demonstrated desmosomes and glycogen granules in the cytoplasm of these cells. The morphology, growth characteristics, alpha-fetoprotein production, heterotransplantability, and karyology of the cloned cells suggested that these clonal lines were derived from the tumor cells and retained their malignant characteristics.


Alpha-fetoprotein and albumin produced by subclonal cell population of the ascites hepatoma AH-66 in vitro. Subclonal cell populations were prepared from a clonal line of the rat ascites hepatoma AH-66 in vitro, using the agar plate culture method. alpha-Fetoprotein and albumin concentrations in culture media of these subclones were determined by 125I-radioimmunoassay and single radial immunodiffusion method, respectively. Results demonstrated that the AH-66 clone was a complex of cells with varying producibility of alpha-fetoprotein and albumin. All the subclones showed varied and distinct production of alpha-fetoprotein, but not all the subclones produced detectable levels of albumin.


Beta-glucuronidase activity of Yoshida ascites hepatomas of different drug-sensitivity and its change after treatment of host rats with various anticancer agents. Change in beta-glucuronidase activity of six Yoshida ascites hepatomas was examined after treatment of host rats with one of 12 anticancer agents. The hepatomas, AH-66F, AH-130, AH-109A, AH-60C, and AH-44, in decreasing order showed more or less distinct increase in beta-glucuronidase activity after treatment of the rats with Nitromin, Endoxan, 864-T, Carbazilquinone, Mitomycin-C, Toyomycin, Daunomycin, Neocarzinostatin, vincristine sulfate, 5-fluorouracil, or cytosine arabinoside only when the cytological effect was positive. Moreover, degree of the increase was generally correlated with that of cytological effect. Bleomycin was ineffective either enzymically or cytologically. AH-66 was insensitive to any of the agents tested in increasing beta-glucuronidase activity and showed only a very slight cytological response to some of the agents. Acid deoxyribonuclease behaved like beta-glucuronidase but to a lesser extent. The above order of drug sensitivity of the hepatomas was not in parallel with that of normal beta-glucuronidase level, which also did not correlate with the life span of host rats.


Comparative autosomal linkage in mammals: genetics of esterases in Mus musculus and Rattus norvegicus. Recombination between Esterase-4 and Esterase-2 in the rat was not observed in 278 backcross offspring. Es-4 is thus included within the "esterase cluster" in Linkage group V. A new map of this region was constructed and the relationship of the four esterase loci was found to be: Es-4-(9.6+/-1.6 cM)-Es-2, Es-4-(1.5+/-0.7cM)-Es-3. Homology of this region with a region of Linkage Group XVIII (Chromosome 8) of the mouse was proposed on the basis of tissue distribution, subcellular localization and response of enzymes to inhibiotrs. Specifically, rat Es-1 was suggested as the homolog of mouse Es-6. An autosomal segment comprising at least 15cM of the rat and mouse genomes appears to have remained relatively intact with respect to genetic content during rodent speciation. In addition, a new polymorphism for mouse esterase was described. The locus was designated Esterase-10 (Es-10) and proposed as the mouse homolog of human Esterase D. Linkage of Es-10 with nucleoside phosphorylase-1-(Np-1) on Chromosome 14 was established.


[A study of two non-complementary H-2 mutations in mice using skin transplantation and serologic analysis]. Hz1 and M505 are two mutants affected the Kend of H-2 major histocompatibility system in mouse. Inability of M505 to complement Hz1 is considered as an evidence that the same gene is altered by both mutations. The gained antigens of two mutants can cross-react in vivo, and the lost antigenic determinants are not identical as revealed in skin grafting tests with presensitized recipients. The antiserum ASY-0,15, (d X a) anti-i, could differentiate between normal and mutant cells as well as between both mutants in absorption tests. These results show that some haptenic determinants are changed in the Hz1 and M505 mutations together with changes of histocompatibility determinants.


[Identification of simian chromosomes by a method of differential staining with Romanovskii--giemsa solution. II. Papio hamadryas]. By means of differential staining wiyh the Romanovsky-- Gimsa dye the distribution of G-bands was studied in chromosomes of bone marrow cells and of cultured peripheral blood lymphocytes in Papio hamadryas. All the homologous chromosome pairs were identified. It was shown, as compared Papio hamadryas karyotoypes with that of Macaca mulatta, that patterns of most chromosomes differentiation were similar. Certain differences between these species were observed in the character of differentiation of chromosomes 1 and 5 and also in parameters of chromosomes 2 and 21 (X).


Detection of HBSAG in fixed liver tissue - use of a modified immunofluorescent technique and comparison with histochemical methods. A modified immunofluorescent technique was used for the detection of HBSAg in formalin-fixed liver tissue, thereby allowing retrospective examination of paraffin sections and avoiding the need to split the sample at the time of biopsy. Comparison with two other methods, involving either orcein staining or standard haematoxylin and eosin (H and E) preparation for ground glass hepatocytes, showed slightly fewer positive hepatocytes in individual biopsies with the latter stain, but the specificity of both methods was high. In a series of 146 seropositive and 74 seronegative patients with a variety of liver disorders, hepatocytes positive for HBSAg were found in only one of 55 patients with acute hepatitis type B, whereas large numbers of positive cells were seen in all 22 healthy carriers of the surface antigen. In the 69 patients with chronic persistent or chronic aggressive hepatitis, the frequency of positive biopsies was 86% and 85% respectively. The positive cells in these cases were, in comparison with healthy carriers of HBSAg, much fewer in number and were scattered in random fashion throughout the lobule rather than occurring in discrete clumps or sheets. This variation in the intrahepatic expression of HBSAg may reflect differences in the immune response to hepatitis B viral antigens.


Histochemical studies on mucosubstances in the prostate gland of the pulmonate snail Semperula maculata in the annual breeding-aestivation cycle. Prostate gland of the pulmonate (terrestrial) snail Semperula maculata was studied histochemically for the presence of polysaccharides. PAS technique and differential staining methods employing Alcian blue, aldehyde fuchsin and Azure A at different pH values and combined with enzyme digestion test and lipid extraction were used. Two histochemically different cell types were found in the prostate gland: the type 1 cells which elaborate neutral mucosubstances and supposedly also phospholipoproteins, and type 2 cells producing glycogen. Both these cell types show seasonal fluctuations of the stores of materials secreted by them. Thus in type 1 cells the weakest histochemical reactivity indicating the lowest concentration of their secretion appears just after cessation of the snails' breeding activity (September) and persists throughout the post-breeding and early pre-breeding seasons which both together constitute the aestivation period. During the rest of the pre-breeding season a strong reactivity gradually develops and culminates in May i.e. immediately before the onset of the breeding activities which in turn initiate a decline of the reactivity. The type 2 cells become most reactive in the middle of the pre-breeding season (May) and decrease their reactivity after cessation of the breeding season. The number of both types of cells parallels the reactivity changes.


Cytochemical studies on the PAS-positive and metachromatic substances in the cells of enteric epithelium of Achoea janata (Noctuidae: Lepidoptera). Of the different polysaccharides present in the cells of insects the PAS-positive and metachromatic substances were studied in the cells of the enteric epithelium of the moth Achoea janata during its larval period and following metamorphosis. The amounts of both the substances progressively decrease in sequential pupal stages. The reaction intensity at the cell apex is higher than at the cell periphery. Reduction in the amount of carbohydrate material in the epithelial cells of the mid-gut is associated with change in intracellular localization (by vanishing from definite sites, not by migration). The loss of metachromatic substances is, however, more rapid than that of PAS-positive ones in the cells of the same region. It appears likely that there are simultaneous conversions of both kinds of substances into chitin precursors, independent of each other. No movement of these substances for the chemical change to be accomplished at specific sites in the cell is found. Association of PAS-positive substances with the formation of peritrophic membrane has been discussed.


Further studies on the possibility of differential demonstration of radicals of sulphur-containing amino-acids. The authors showed that in rat liver and brain sections with blocked SH groups and split S-S bonds mercury orange stains some tissue structures after treating the sections with BAL. Considering that the blockade of SH groups with N-ethylmaleimide in is stable the authors set forth the hypothesis that the use of BAL may enable the demonstration of a sulphur-containing amino-acid which is devoid of SH or SS groups but acquires them only when acted upon by BAL. The in vitro studies demonstrated that the effect of BAL makes it possible to stain methionine with mercury orange which otherwise does not stain this amino-acid.


Histochemical analysis of mucosubstances of the mid-gut gland in the mussel Lamellidens consobrinus Lea. The mid-gut gland of the fresh water mussel Lamellidens consobrinus is located on both sides of the crystalline style sac and the conducting chamger of the intestine. Standard histochemical techniques were employed for the detection of mucosubstances elaborated by the mid-gut gland. The results show that he gland secretes neutral mucins and sialomucins which act as lubricants and are involved in converying good particles during feeding.


Differentiation between stains of human blood and blood of monkey. The precipitin tube test fails to differentiate between blood stains of closely related animals such as man and monkey because of the close similarity between antigenic determinants of serum proteins of such closely related species. The quantitative precipitin technique cannot be applied to forensic work. This paper cites a case where definite opinion on the species of origin of some blood stains suspected to have come from a monkey was possible. In the method used, extracts of the questioned stain were tested with similar extracts of known human and known monkey blood stains against human globulin antiserum, by comparative double diffusion method in agar gel on microslides.


[Pathophysiological significance of endotoxins]. The biological activities of endotoxin are manifold. Besides the drop in platelets and the biphasic change in the number of leukocytes, severe disturbances in the capillary bed are among the first changes observed following the administration of endotoxin. It is well known that endotoxins cause the release of various vasoactive mediators. According to our results endotoxins enhance the activity of histamine and serotonin; these findings may contribute to a better understanding of the action of histamine and serotonin in the early post-endotoxin phase. The histamine-sensitizing effect of endotoxins is not prevented by antihistamines. Endotoxins and biogenic amines cause similar disturbances in the capillary bed. The changes that are observed in the content of the vessels, the vessel wall, and the perivascular region are the following: slowing down of the blood stream, degranulation of perivascular mast cells, granulocytosis, wall adhering granulocytes, plasma skimming, rouleaux-formation of erythrocytes, reduction in plasticity of many erythrocytes, acanthocytes, acanthocytosis, appearance of spherocytes and microcytes, and formation of massive aggregates of platelets and of microthrombi. Also, occasionally cell aggregates dissolve and as microemboli form new thrombi. Swelling of pericytes, endothelial, and periendothelial cells is observed, and dissociation and deformation of the endothelial cells occur. By means of contact of the vessel contents with the collagen, there is an additional activation of the coagulation system by factor XII possible. The changes of the epithelial lining and the wall adhering cells enhance the narrowing of the vessel lumen. Prestasis and occasionally stasis occur. One observes increased swelling of the endothelial and periendothelial cells, increased permeability of the vessel wall, passage of plasma and occasionally blood cells, especially erythrocytes, through the endothelium and massive microbleedings. While stasis is observed in the nutritive capillaries, in regions where arteriolar-venular shunts exist the flow continues. The systemic blood pressure may therefore remain unchanged during this phase, although the severe disturbances described occur in the capillary bed. Metabolic alterations, especially in the carbohydrate metabolism are mentioned.


[Gamma globulin therapy of acute viral hepatitis. Studies on the therapeutic effect of gamma globulin on the course and late prognosis of manifested acute viral hepatitis in man]. In a prospective study 148 consecutive patients with biopsyproved acute viral hepatitis were observed serially and followed for 5 years. They were divided into three groups on the basis of being treated with high or low doses of gamma globulin and compared with a control group, not treated. As the efficacy of gamma globulin for the prophylaxis or modification of infectious hepatitis has been well documented by many investigators during the past 25 years, we were interested in evaluating the therapeutic effect of gamma globulin on the course of viral hepatitis. The purpose of the study was to determine the comparative efficacy of various doses of gamma globulin in preventing complications and in influencing the severity and the length of time of acute viral hepatitis and in preventing the development of chronic hepatitis and cirrhosis. For controlling the clinical, biochemical and histopathologic course 12 functional parameters were repeatedly measured under stable clinical conditions and 3--12 liver biopsies were performed in an individual patient using the Menghini needle with an intercostal approach. During the 5-year trial an overall of 825 liver biopsis were performed with this 148 patients. We conclude from this study, that in about 80% of patients with acute viral hepatitis recovery is complete, but takes several month's. A protracted course of 4 month's duration until recovery was found in 45 patients (30,4%), persistent hepatitis with recovery after 1--4 years duration occurred in 37 patients (25%), global liver necrosis with hepatic coma in 3 (2,3%), chronic hepatitis in 22 (14,8%), 8 of them as chronic aggressive hepatitis and cirrhosis in 3 (2,3%). The study demonstrated no therapeutic efficacy of gamma globulin in modifying the course or preventing complications of both AuAg+ and AuAg-neg. acute viral hepatitis in man. There was no striking difference in the groups treated with various doses of gamma globulin compared with a control group.


[Cellular immunity in poxvirus infections. Demonstration, significance and cellular cross reactivity with brain matter]. Its proof, significance, and the cellular cross reactivity to basic encephalitogenic protein of vaccinia-stimulated T-cells. Vaccination against smallpox induces both, a cellular allergy and immunity, and the production of humoral antibodies. Of greater importance for the defense of the organism against infections by orthopox viruses is the cellular immunity. New methods for the proof of cellular immunity and an in vitro-technique for its examination by challange with variola vera virus are described. In close connection with induction of cellular allergy by vaccination an immunological cell-bound reactivity of isolated lymphocytes develops against the basic encephalitogenic protein. In this antigen-stimulated resp.-irritated stage isolated lymphocytes show an enhanced rate of chromosomal aberrations and a high rate of spontaneous transformations. Lymphocytes of patients with Multiple Sclerosis demonstrate a high stimulation rate (3H- resp. 14C-Thymidine-inlay) after challenge with antigen of vaccinia virus. The T cell-mediated cross reactivity between vaccinia and myelin extracts is discussed as an hypothetic pathogenic factor of CNS complications after primary smallpox vaccination.


[Serum glycoside concentration and digoxin intoxication]. Serum digoxin levels, estimated by radioimmuno assay, technique, and the pattern of cardiac arrhythmias due to digoxin intoxication seem to be correlated in cases with high serum digoxin levels. The prognostic value of  those correlations is demonstrated and discussed. Serial observations of the course of cardiac arrhythmias under the suspicion of digoxin intoxication together with blood level measurements allow to introduce more objective criteria in clinical management of poor risk patients receiving digoxin.


Experimental investigations in post-mortem protein degradation. Samples of thoracic fluid were obtained at regular intervals from the putrefying bodies of dead dogs. For quantitative estimation of the proteins two-dimensional immunoelectrophoresis was employed and for the amino acids and amines two-dimensional thin-layer chromatography. The results lead to speculation that the initially lagging proteolytic changes are due to catalysis of autoenzymes, whereas bacteriological processes are responsible for a distinct acceleration from the 20th day after death. At this time the amino-acid concentrations showed a marked increase, which is followed by a maximum of bacterial growth, that ultimately leads to a sudden regression of amino acids. Simultaneously amines appear. The drying up of the thoracic cavity after 35 to 45 days marks the change from liquefaction to the terminal decay.


Banding of human chromosomes with basic fuchsin. In this paper a technique is described for the banding of human metaphase chromosomes with basic fuchsin. The main characteristics of the G-banding pattern obtained with this cationic triphenylmethane dye are: the secondary constriction regions of chromosomes Nos. 1 and 16 are strongly stained, especially in the latter one; the heterochromatic area of chromosome No. 9, usually negative with most other G-banding techniques, is clearly visible as an intensely stained band adjacent to the centromere; the chromosomal outline is often very distinct, facilitating the study of the telomeres; a number of chromosomal regions with bright Q fluorescence such as the polymorphic regions of the chromosomes Nos. 3, 4, and Y also stain strongly with basic fuchsin. The basic fuchsin technique combines therefore properties of G-, C-, and Q-banding methods and seems very suitable for use in e.g., family and linkage studies. Several triphenylmethanes closely related to basic fuchsin produce similar banding patterns. The band-producing ability is, however, diminished in those dyes which contain methylated amino groups. If the methyl groups are attached to the carbon atoms at the 3-positions in the phenyl rings, band formation seems unaffected. The way in which basic fuchsin and chromatin may interact as well as the possible mechanism(s) of band formation with this dye are discussed.


Blocking the encephalitogenic activity of the C-terminal part of the basic myelin protein in rabbits by treatment with serum. Peptides HNB-92-169 and 116-169, tyr, derivatives of the C-terminal part of bovine myelin basic protein, sensitize rabbits for experimental allergic encephalomyelitis (EAE). Dissolving these peptides in heat-inactivated normal guinea-pig, human, or monkey serum immediately before emulsification with Freund's complete adjuvant (FCA) abrogated their disease-inducing activity. Dissolving them in homologous or autologous serum only slightly affected the encephalitogenicity.


Xenopus laevis 19S immunoglobulin. Ultrastructure and J chain isolation. Electron microscopy examination of the 19S immunoglobulin of Xenopus laevis revealed a hexameric structure with a central core. The molecules measured 360-430 A across the span of the arms and the average diameter of the central region was 140 A. A polypeptide, homologous to human J chain, was isolated by chromatography on DEAE-cellulose from the reduced and alkylated X. laevis hexameric macroglobulin. This polypeptide had a fast mobility in alkaline-urea gel electrophoresis, with distinct antigenicity, as compared to heavy and light chains. It shared common antigenic determinants with human J chain.


Chemically induced temperature-sensitive mutants of dengue virus type 2: comparison of temperature sensitivity in vitro with infectivity suckling mice, hamsters, and rhesus monkeys. A series of temperature-sensitive (ts) mutants of dengue virus type 2 (DEN-2, TH-36 isolate) were induced by treatment with 5-azacytidine. These mutants and parental viruses were compared for the ts trait and/or attenuation in four systems: primary hamster kidney cells, suckling mice, golden Syrian hamsters, and rhesus monkeys. Seven clones judged to possess the ts trait in virto demonstrated a variety of patterns in vivo. On initial isolation, five of seven ts mutants exhibited reduced mouse lethality. The remaining two mutants possessed parental levels of mouse lethality. In hamsters, neither ts mutant nor parental viruses replicated very well, and then only when inoculated intracerebrally. Studies in rhesus monkeys indicated that all seven ts clones and parental viruses were capable of inducing abtibody responses; however, ts-1 and ts-2 failed to produce detectable viremia. After challenge with parental virus, all vaccinated monkeys demonstrated rapid secondary-type antibody response. Reversion from ts to ts(+) was confirmed to ts-1 in mice and ts-3 in monkeys, and was strongly suspected in several other instances.


Cell-mediated cytotoxicity against murine cells infected with 6/94 virus, a parainfluenza type 1 isolate from multiple sclerosis brain tissue. Maximum cytotoxicity to parainfluenza type 1 viral determinants occurs on day 5 post-immunization in mice with and without preexisting anti-6/94 virus antibody. Animals with preexisting anti-6/94 serum antibody, however, exhibited a markedly higher cytotoxic response. Syngeneic, allogeneic, and xenogeneic virus-infected cells induce virus-specific cytotoxicity, but only infected cells histocompatible with the effector cells can serve as susceptible target cells. The effector cell is a T cell, and hence cytotoxicity is thymus dependent. Coincident with the maximum cytotoxic activity on day 5 is the occurrence of pathological lesions, the sites of which are dependent on the route of inoculation.


Use of the hemadsorption phenomenon for determining virus and neutralizing antibody titers of rabies. Chicken embryo cells infected with the HEP Flury strain of rabies virus adapted to tissue culture produced a hemadsorption (HAD) phenomenon by using goose erthyrocytes. The optimal conditions for HAD included the incubation of cell cultures at 37C for 3 days after virus inoculation, the use of a 0.4% suspension of goose erythrocytes in phosphate buffer adjusted at pH 6.2, and adsorption of erythrocytes at 4C. This phenomenon was inhibited with anti-rabies serum. Virus titer obtained with the HAD technique was almost the same as with the fluorescent antibody technique or the intracerebral inoculation of suckling mice. Results of the neutralization test by using the HAD technique could be easily determined 3 days after inoculation of chicken embryo cells with the mixture of 100 mean tissue culture infective doses of virus and diluted serum. The neutralizing antibody titers coincided with those obtained in mice.


DNA synthesis in vitro by cells from mice immunized with picryl chloride: effect of injection of immune cells. Mice were immunized with picryl chloride and the regional nodes taken at various times afterwards. These cells spontanesouly synthesized DNA in vitro as measured by thymidine incorporation over an 18-hour period and the peak incorporation occurred when the cells were takin on day 3. When the mice were injected with cells taken 5 days after immunization with picryl chloride and then immunized, there was a depression of the spontaneous DNA synthesis in vitro. This was absent on day 2, most marked on day 3 and still present on day 4. Cells from donors immunized with 4-ethoxymethylene-2-phenyloxazolone had a smaller but definite effect. Attempts to reproduce the phenomenon by in vitro mixtures of cells taken at various times after immunization in vivo were unsuccessful.


Specific acquired immune unresponsiveness to contact allergens with cyclophosphamide in the mouse. Treatment with cyclophosphamide (Cy) can modulate the acquisition of allergic contact dermatitis in the mouse. We compared the effect of a single dose of Cy given at different times before or after allergen. Cy one or more days prior to allergen intensified, and Cy several days after allergen inhibited the acquisition of sensitivity. Mice whose immunological response to an allergen had been suppressed by Cy were specifically immunologically tolerant to that allergen, but not to an unrelated allergen. This tolerance probably represents a combination of clone deletion and inhibition; almost certainly it does not depend on the generation of enhancing ('blocking') antibody.


Chemical modification of crude timothy grass pollen extract. I. Antigenicity and immunogenicity changes following amino group modification. Glutaraldehyde modification was found to reduce the allergenic potency of crude timothy pollen extracts yet even highly substituted materials retained the capability of inducing the formation of allergen-specific antibody in animals. Experiments showed that these antibodies were capable of blocking skin test reactions to native timothy allergens in pollen-sensitive human volunteers. The value of glutaraldehyde-modified allergen extracts for use in desensitization vaccine therapy is discussed.


Inhibition by nicotinamide of antigen-induced histamine release from mouse peritoneal mast cells. The in vitro antigen-induced histamine release from mouse peritoneal mast cells actively sensitized with IgE antibodies was inhibited by nicotinamide. The drug was either given in vivo to the sensitized mice (once daily 100 mg/kg) for 7 days before an in vitro experiment or incubated in vitro (in concentrations 1-40 mM) with sensitized mast cells before an antigen challenge. The possible action of nicotinamide on the mechanisms involved in the regulation of antigen-induced histamine release from mast cells is discussed.


Carcinoembryonic antigen-like substances of human bile. Isolation and partial characterization. Three species of carcinoembryonic antigen (CEA)-like macromolecules, called the biliary glycoprotein I, II and III (BGP I, II and III) were identified in human bile. BGP I was found in normal gall-bladder bile and hepatic bile but not in bile subjected to inflammation ("white bile"). It was immunologically related to CEA and NGP (the "normal CEA-like glycoprotein". Synonyms: NCA, CCEA-2, etc.). BGP I differed immunologically from CEA in that it lacked the tumor-associated determinants of CEA. It was different from NGP (and CEA) in that it contained BGP I specific determinants as revealed by anti-BGP I antibodies. In bile from gallbladders with obstructed outlet and subjected to cholecystitis, a non-malignant inflammatory process, BGP I was replaced by BGP II and BGP III. Immunologically and physicochemically, BGP II and BGP III appeared to be closely similar to NGP and CEA, respectively.


Ability of delayed-type hypersensitivity reactions to distinguish tumor-associated antigens and histocompatibility antigens in soluble extracts from murine fibrosarcomas. The footpad swelling (FPS) test for delayed-type hypersensitivity in the mouse was evaluated for its ability to measure both tumor-associated antigens (TAA) and histocompatibility (H) antigens solubilized from methylcholanthrene (MCA) induced fibrosarcomas of C57B1/6 (B6) mice. Tests for TAA were performed in mice immune to syngeneic tumors while H-antigens were assayed in mice immunized with skin allografts. FPS was most intense in B6 mice challenged with TAA from the immunizing B6 tumor, but also occurred in response to cross-reactive TAA solubilized from another B6 fibrosarcoma. Tests for tumor-associated H-antigens in allograft immune mice were strongly positive in response to donor/recipient H-antigen differences and proved sensitive to shared third-party H-antigen differences. Comparison of soluble antigens from the same tumor maintained in vitro and in vivo revealed that, while both TAA and H-antigens could be detected in preparations from the in vivo tumor line, only TAA and not H-antigens could be detected by RPS in extracts prepared from the in vitro tumor line. These experiments have demonstrated that the mouse FPS test can distinguish both TAA and H-antigen specificities persent in the same complex mixture of tumor-cell antigens.


Human pregnancy-specific beta1-globulin and its relation to chorioepithelioma. Human serum pregnancy-specific beta1-globulin (beta1-GP) was localized in paraffin sections of placenta and chorioepithelioma of the uterus by indirect immunofluorescence. The structures containing beta1-GP but not human serum albumin were regarded as specifically associated with beta1-GP metabolism. Beta1-GP was found in trophoblastic cells of chorion and Langhans cells of chorioepithelioma. Using the immunoautoradiographic method (I131), we only found beta1-GP in sera of patients with trophoblastic tumours: in 74.3% of 35 patients with chorioepithelioma and in 80% of 25 patients with hydatidiform mole and destructive hydatidform mole. The diagnostic and prognostic significance of the immunochemical test for beta1-GP in trophoblastic tumours is discussed.


AKR leukemia-specific surface antigens acquired by malignant transformation: their common and individual specificities. Antisera against five newly established AKR spontaneous leukemias (KSL) prepared by immunization of KSL cells in (C57B1/6 X C3Hf/He)F1 mice were preabsorbed with non-leukemic AKR lymphoid cells to remove antibodies against virus-associated surface antigens and alloantigens. In absorption tests the antisera showed no cross-reaction by immunofluorescence microscopy with thymocyte, fetal, male-specific H-Y, E and X.1 antigens; with known Gross murine leukemia virus-associated antigens; or with cell surface antigens on Friend, Moloney and Rauscher virus-induced tumors. It was thus shown that the antigens detected were leukemia-specific, were acquired by malignant transformation, and consisted of two types: (1) a common antigen in all the KSL, and (2) individual antigens found in four of the KSL which showed distinct patterns of partial cross-reactivity with the other KSL.


Immunogenic and immunosuppressive components in soluble preparations from Moloney murine leukemia virus-induced tumor cells. Soluble extracts from Moloney virus-induced leukemia cells were shown to contain immunogenic and immunosuppressive components. Immunogenic components were divided into two classes based on their ability to :(1) combine with humoral antibody to inhibit cytotoxic activity; or (2) stimulate blastogenesis in normal syngeneic lymphocytes. Immunosuppressive components were demonstrated by the inhibition of DNA synthesis in normal syngeneic lymphocytes and suppression of a mixed lymphocyte reaction. Stimulating components in many soluble preparations were completely masked by immunosuppressive components. However, all three components could be resolved by Sephadex G200 chromatography.


Collagenase inhibitors: rationale for their use in treating corneal ulceration. Tissue collagenases have been implicated in corneal ulceration in human corneal disease and in ulceration of the rabbit cornea that has served as a model system. Such enzymes from the rabbit and human cornea are inhibited by metal-binding agents of the EDTA type, by thiols, and by the human serum antiprotease alpha2-macroglobulin. Determination of the relative efficacies of collagenase inhibitors indicates that EDTA and Ca-EDTA are about one hundred times more effective on a molar basis than L-cysteine and its derivatives, N-acetyl-L-cysteine and D-penicillamine. The alpha2-macroglobulin on a molar basis, is superior as an inhibitor to the metal-binding agents and thiols. Although Ca may be a necessary cofactor of the corneal collagenases, such a requirement has not been established unequivocally. Inhibition and isotope studies do indicate a requirement for Zn. Thiols are thought to inhibit corneal collagenases by binding to or removing an intrinsic metal cofactor (Zn), and/or possibly by reducing one or more disulfide bonds. Inhibition by both EDTA-type agents and thiols is largely reversible by dialysis. The human alpha2-macroglobulin appears to inhibit corneal colleagenases irreversibly by forming tight complexes with them. Ca-EDTA, cysteine, and acetylcysteine, given as eyedrops, are able to prevent or retard ulceration in the alkali-burned rabbit cornea. They appear to have some efficacy in the prevention of corneal ulceration in humans. EDTA-type compounds are quite stable under routine storage, while acetylcysteine is more stable than cysteine. EDTA is quite toxic and should not be used as eye medication. Ca-EDTA has a low toxicity, and cysteine and acetylcysteine have even lower toxicity. It is not yet certain which inhibitor has the most favorable therapeutic index for clinical use, or is the optimal mode of drug delivery known. However, the collagenase inhibitors seem to have therapeutic promise in the prevention of corneal ulceration.


Human prostatic adenoma and carcinoma. Transplantation of cultured cells and primary tissue fragments in "nude" mice. Tissues from human prostatic carcinomas and adenomas as well as cells from permanent cell lines originating from such tumors were transplanted into the subcutaneous space of "nude" mice. A total of 208 such transplantations were carried out. For transplantation of primary tumor tissue several techniques were used. Transplantation of single pieces of about 50 mg of tissue from carcinomas and adenomas provided the best results. With this technique carcinoma tissue was recovered from five of 10 tumors transplanted into 46 "nude" mice. Carcinoma was found in seven of 25 "nude" mice that were castrated and treated with testosterone but only once in 21 transplantations into castrates without testosterone substitution. This may suggest that heterotransplants of human prostatic carcinoma tissue can be established more easily in testosteronized animals. In the case of adenoma transplants, the histological picture was strikingly similar to the original tumors. Secretory activity was found in some acini, squamous metaplasia in others. Heterotransplantation of cells from permanent cell lines resulted in fast growing carcinomas without similarity to the origianl tumors. Dependence on hormones for growth of EB 33 and MA 160 cells in "nude" mice was not shown.


Reverse transciptase activity associated with R-type virus-like particles of SV40-transformed hamster cells (TSV5 clone 2). A line of hamster cells transformed by SV40 (TSV5 clone 2) contains different DNA polymerase activities. One of them has the same template specificity as viral reverse transcriptases. Partial isolation of R-type virus-like particles from TSV5 clone 2 shows that a reverse transcriptase activity is associated with these RNA viruses. This reverse transcriptase was partially purified and its biochemical properties are described.


Mycoplasma in tissue culture: overview of detection methods. Mycoplasma contamination of tissue cultures is too costly, scientifically damaging and inappropriate, and perhaps, even medically hazardous to be ignored. Prevention by continuous surveillance demands adequate mycoplasma testing procedures. Despite a proliferation of indirect methods, the present consensus indicates that careful and extensive microbiological testing is still the cornerstone of any mycoplasma detection program. The finding of 'non-cultivable' or possible tissue-adapted micro-organisms plus the variability in microbiological expertise of many laboratories, makes the selection of at least one additional method imperative. At the present, though no one test will prove 100% satisfactory and selection should partly depend on available expertise, the method based on the ratio of the specific activities of incorporated radio-labelled uridine to uracil into RNA appears to be an acceptable choice due to its simplicity and general validity.


Carboxylic ester hydrolases in the thyroid gland of the guinea-pig. A light microscopic study. The location of cholinesterase and non-specific esterase in the thyroid gland of the guniea-pig was studied with the light microscope. It was found that the idoxyl method for non-specific esterase activity under special conditions is superior to the cholinesterase method in a number of respects for the demonstration of the intra-, inter- and parafollicular cells. When using the indoxyl method the incubation period can be reduced from 2.5-3 hr to 40-50 min. Further, the reaction can be followed during the incubation. False localization of the reaction products is avoided,and nerves and erythrocytes are not stained. By varying the fixation time and the time of storage in gum arabic-sucrose, it was found that the miscellaneous activity of non-specific esterase in APUD cells (C-cells) and follicle cells may be due to both factors. In fresh tissue the activity of the enzyme was equal in follicle and C cells. Special cyst-like structures containing an esterase which is NaF-resistanct when alpha-naphthyl acetate is employed as a substrate and which gives a strong reaction at low pH values when 5-bromo indoxyl acetate is the substrate, are described, and their nature and possible origin are discussed.


The effects of ductal ligation on the parenchyma of salivary glands of cat studied by enzyme histochemical methods. Submandibular, sublingual and parotid glands of cat have been studied following periods of ductal ligation ranging from 1 day to 1 year. An increased prominence of granules of acid phosphatase, beta-glucuronidase and E600-resistant esterase reaction products was sometimes seen in acinar cells and atropic striated ductal cells, and probably represents increased lysosomal enzymic activity, which may be of importance in the adapation of the parenchyma to the altered environment. Other reaction products often appeared to be at normal levels in parenchymal structures that were not very atrophic, and at reduced levels in those that were very atrophic, suggesting a reduction of functional activity in these structures.


Properties and kinetics of development of Rous sarcoma virus-infected cells evidenced by methylene blue staining. Three different kinds of areas of infected cells corresponding to different focus formation stages can be evidenced by methylene blue (MB) staining in cultures of chick embryo (CE) fibroblasts infected at low multiplicity with the temperature-sensitive (ts) mutant of Rous sarcoma virus (RSV), FU19, which transforms these fibroblasts at 37 degrees but not at 41 degrees. These are: (a) areas of MB-stainable cells with transformed phenotype (STP areas=foci); (b) areas of MB-stainable cells with normal phenotype (SNP areas), and (c) areas of MB-unstainable cells with normal phenotype (USNP areas). DNA and RNA synthesis and virus production were followed in these various stages at 37 degrees and at 41 degrees. The results show that when cultures are shifted from 37 degrees to 41 degrees, virus production in the SNP and USNP areas which arise by phenotypic reversion of STP foci remains comparable to that of the latter foci. On the contrary, DNA and RNA synthesis are markedly reduced in SNP and USNP areas, DNA synthesis falling down to the level of uninfected cells, and RNA synthesis remaining somewhat higher. The kinetics of development of SNP and STP areas in cultures infected with FU19 and with the parental virus SR4 were also compared. The results confirm that SNP areas are precursors of STP areas but that this passage occurs at a slower rate in cultures infected with FU19.


Acid mucopolysaccharides in hair papillae of the stump-tailed macaque (Macaca speciosa). Acid mucopolysaccharides in dermal papillae of hair follicles from both bald and on-bald regions of the scalp of stump-tailed macaques were studies histochemically. Alcian Blue, Azure A and Periodic acid Schiff methods were used for staining mucopolysaccharides, and Bromphenol Blue for staining basic proteins. In an attempt to identify various polyanions, staining was carried out with Alcian Blue containing different concentrations of electrolytes. Methylation, saponification, mild acid hydrolysis and digestion with streptomyces or testicular hyaluronidase, chondroitinase ABC, or sialidase, were also used. The results indicate that chondroitin sulphate B is present in the papillae of terminal hair follicles in early and intermediate anagen, and degraded chondroitin sulphates are present in the papillae of vellus and terminal hair follicles in late anagen.


Developmental changes of carbonic anhydrase in the retina of the mouse: a histochemical study. In the optic cup of early mouse embryos, carbonic anhydrase activity is seen only in the pigment epithelial cells. During the late prenatal period the enzyme appears in the Müller cells, first in the perikarya. During the postnatal development the enzyme activity appears in the radial processes of the Müller cells starting at the vitreal border along with the separation of the outer retinal layers. This is followed by increased activity in the plexiform layers, presumably in the lateral processes. Carbonic anhydrase localization in pigment epithelial and Müller cells attains the adult pattern at about 16 days corresponding with the initiation of retinal function.


A comparison of two methods of preparing cells or nuclei for high-resolution microspectrophotometry. Suspensions of isolated mouse thymus cells were subjected to two preparative methods: either they were dropped through several mm of 9:1 v/v ethanol-acetic acid fixative, allowed to stand for 1hr and then processed for staining; or they were fixed, passed through a graded ethanol series to 70% ethanol, centrifuged on to slides in a modified Shandon cytocentrifuge and then carried wet into the staining procedure. All preparations were stained by the Feulgen reaction and evaluated by high-resolution microspectrophotometry. While the two preparative procedures yielded similar results, there appeared to be less variability in the data obtained from the centrifuged cell populations.


An improved method for staining cells of the endocrine polypeptide (APUD) series by masked metachromasia: application of the principle of 'fixation by excluded volume'. Masked metachromasia is demonstrated by staining with a metachromatic basic dye, after acid hydrolysis of suitably fixed tissue. We report that the addition of 20% Carbowax 20M (an inert polyer, mol. wt. about 20,000) to the hydrolysis mixture improved the reaction. The improved method gives increased metachromasia , greater tolerance to variations in hydrolysis conditions, and demonstrates a greater proportion of cells--presumably due to a lower threshold of sensitivity. Lower molecular weight polymers (Carbowax 1000, Carbowax 6000) are less effective.


Succinate dehydrogenase activity in the developing molar of the Mongolian gerbil Meriones unguiculatus. The distribution of succinate dehydrogenase in the odontogenic tissues in the Mongolian gerbil, ranging in age from 18 days prenatal to 8 days postnatal, was studied. Activity levels were designated negative, trace, slight, moderate, strong and very, dependent upon visual observations. The results indicated increased levels of activity as the tissue layers differentiated from the lamina stage through early apposition. The cell layer displaying the most succinate dehydrogenase activity was the ameloblastic layer while the stratum intermedium layer was the next most active cell layer. Succinate dehydrogenase activity appeared to be related to the degree of differentiation and functional competency of the odontogenic tissues in the Mongolian gerbil.


Glycogen, its chemistry and morphological appearance in the electron microscope. III. Identification of the tissue ligands involved in the glycogen contrast staining reaction with the osmium (VI)--iron(II) complex. By application of appropriate blocking reactions (acetylation, de-amination, methylation and NaHSO3-treatment) it is demonstrated that the tissue ligands involved in the selective glycogen contrast staining reaction with the OsVI. FeII complex (known to be present in the combination K2OsO4K4Fe(CN)6) are the glycogen C2-C3 di-hydroxyl groups. Deliberate conversion of the diols into di-aldehydes and (di-)carboxyl groups by the application of specific oxidative agents followed by application of the OsVI. FeII-complex results morphologically in identical selective contrast staining of glycogen. By applying appropriate blocking reactions to such pre-oxidized aldehyde fixed glycogen, evidence is accumulated that K2OSO4 and K3Fe(CN)6 are unable to oxidize diols, whereas OSO4 and H2O2 are able to convert diols into carboxyl groups. From these results it is concluded that in the combination K2OSO4K4Fe(CN)6 the OsVI.FeII complex reacts with unchanged diols in the glycogen, whereas the OSO4 in the combination OSO4K3Fe(CN)6 can potentially create carbocyl groups in the aldehyde-fixed glycogen. The addition of urea to the two glycogen contrasting combinations (K2OSO4K4Fe(CN)6 or OSO4K3Fe(CN)6), also emphasizes that, although morphologically both combinations produce identical contrast stained glycogen, chemically the contrast staining is apparently obtained in a different way, as urea prevented the contrast formation in the glycogen by the combination K2OsO4Fe(CN)L, but not by the combination OSO4K3F e(CN)6.


An ultrastructural application of silver methenamine to the study of mucin changes in the colonic mucosa adjacent to and remote from carcinoma. A periodic acid-chromic acid-silver methenamine techniqe for visualizing glycoproteins at the electron microscope level was applied to colonic mucosa taken from areas adjacent to and remote from carcinoma. Normal control mucosa was obtained by biopsy of patients with no known gastrointestinal disease. Non-oxidized control sections were run in parallel. Quantitative and qualitative differences in glycoproteins were detected in the mucosa adjacent to carcinoma ('transitional' mucosa, as we call it) as compared with the normal. Furthermore, the vesicles in both the 'intermediate' and absorptive cells elaborate a glycoprotein product and it seems that a direct relationship exists between the increased vesiculation and the markedly developed 'fuzzy coat' in the 'transitional' mucosa. It is suggested that these findings may represent one of the features of an early stage of carcinogenesis. Histochemical and ultrastructural techniques of the kind used in this study may thus be of value in identifying or predicting malignancy in the colonic epithelium.


Movement of horseradish peroxidase in rabbit submandibular glands after ductal injection. Horseradish peroxidase (HRP) has been used as a tracer to study movements of solutions injected retrogradely via the duct of submandibular glands in rabbits. 0.1 ml of solution was injected either manually or by a constant hydrostatic pressure, and the subsequent distribution of HRP in the gland and duct at different times after injection has been examined histochemically at light and electron microscopical levels. Shortly after the injections, strong interstitial staining for peroxidase resulted from passage between acinar cells. Some sites of cellular uptake were observed and staining occurred in some ductal cells even when the duct had been cut at the hilum to minimize pressure effects. It is not known whether this diffuse uptake represents a physiological or pathological phenomenon. Some interstitial activity still remained 24 hr after injection but had disappeared by 48 hr. Inflammatory cells first appeared in the gland about 4hr after the injection and slowly increased up to about 24 hr after injection. The results indiate that the HRP reaches the interstices of the gland principally by penetration between acinar cells, and that the junctional complexes between striated  duct cells appear to be more resistant to disruption by luminal pressures.


Electron microscopic autoradiography of 35SO4-labelled material closely associated with collagen fibrils in mammalian synovium and ear cartilage. Electron microscopic autoradiography of connective tissue obtained from mice and rabbits previously injected with 35SO4 indicated that sulphated proteoglycans are localized on collagen fibrils. Ruthenium Red-positive transverse belts surrounding fibrils near the a-bands were heavily labelled, but fine lateral filaments of Ruthenium Red-positive material were not. These filaments, which interconnect collagen fibrils in a variety of connective tissues may represent linear aggregations of hyaluronic acid, glycoproteins and non-sulphated or long-lived sulphated proteoglycans.


Physiologic/clinical comparisons of a sustained-release decongestant combination, its components and placebo in patients with allergic rhinitis. Fifteen subjects with chronic allergic rhinitis had measurements of nasal airways flow/resistance (Rn) made before, and for 12 hours after, single doses of a sustained release decongestant combination, some of its components given alone or together, and placebo, in a randomized double-blind trial. The magnitude of improvement in Rn was statistically greater for the the four active preparations than for placebo over the first 8 hours; the effects of phenylpropanolamine/chlorpheniramine were still present after 10 hours, while at the end of 12 hours only the full triple-drug capsule had significant activity. Patient-estimates of symptomatic improvement generally mirrored these physiologic changes although statistical differences among the active capsules were not delineated. The data confirm the ability of electronic posterior rhinometry to discriminate between the effects of active medications and placebo at the 95 per cent confidence level or better and suggest that observed decreases in elevated Rn mean reflected helpful clinical activity as well as increased nasal patency.


A neutral subtilopeptidase inhibitor from porcine serum some evidence for alpha2-macroglobulin. An inhibitor of neutral subtilopeptidase [EC 3.4.24.4] was purified from porcine serum by salting out with (NH4)2SO4, chromatography on anion exchange sephadex, gel filtration with Sepharose 6B, and isoelectric focusing. The preparation was homogeneous by electrophoretic and ultracentrifugal criteria, and was shown to be a glycoprotein with a molecular weight of 740,000. It inhibited the caseinolytic activities of thermolysin, subtilisin, trypsin [EC 3.4.21.4], and alpha-chymotrypsin [EC 3.4.21.1] as well as that of neutral subtilopeptidase by an equimolar binding to those proteolytic enzymes. SDS-polyacrylamide gel electrophoresis after reduction with beta-mercaptoethanol indicated that the inhibitor was made up of four subunit monomers having a molecular weight of 190,000. From comparisons of its physiocochemical and inhibitory properties with those of well-investigated plasma proteins, the inhibitor was identified as alpha2-macroglobulin. On treatment of the inhibitor with neutral subtilopeptidase, a protein with a molecular weight of 95,000 appeared after treatment with SDS and beta-mercaptoethanol, suggesting that a peptide bond susceptible to the enzyme exists near the mid-point of the subunit chains.


Synthesis and release of procollagenase by cultured fibroblasts. An inactive collagenase was harvested from both serum-free and serum-supplemented fibroblast monolayer cultures in periods of active collagen synthesis. The latent collagenase did not hydrolyze collagen and did not bind the potent collagenase inhibitor alpha2-macroglobulin. Activation with trypsin imparted to the enzyme the ability to hydrolyze collagen at neutral pH in a typical manner and to form an inhibited complex with alpha2-macroglobulin. The molecular weights, determined by calibrated gel filtration, were 78,000 and 60,000 for the latent and active enzymes, respectively. The data indicate that collagenase is released from the cells in inactive form, as a zymogen.


Long-term effects of chondrocyte death on rabbit articular cartilage in vivo. The effects of freezing localized areas of rabbit articular cartilage in vivo were studied at two to twelve months morphologically (gross and microscopic examination, including scanning electron microscopy), histochemically (toluidine blue and safranin O), and metabolically (35S uptake). Up to six months the frozen cartilage was intact but appeared to be dead, as shown by the absence of stainable chondrocytes, severely decreased acid mucopolysaccharide content, and absence of significant uptake of 35S. At twelve months fibrillation and softening were evident, clusters of new chondrocytes with surrounding acid mucopolysaccharides were visible microscopically, and scanning electron microscopy revealed an irregular pattern of collagen fibers which were larger than normal, of varying thickness, and broken in some areas. These changes resembled those seen in degenerative joint disease.


Coated-vesicle shells, particle/chain material, and tubulin in brain synaptosomes. An electron microscope and biochemical study. Coated vesicles (CVs), plain synaptic vesicles (PSVs), and nonvesicular flocculent material were isolated from synaptosomes and examined with goniometry and high-resolution electron microscopy after either negative staining or various biochemical procedures. The flocculent material (i.e. the presynaptic matrix material except CV shells) is largely composed of particulate or elongated (chainlike) structures; some of this material (here referred to as particle/chain material) is attached to PSVs. The results obtained were: (a) the proteinaceous properties of the CV coat (also referred to as CV shell) and the particle/chain material were demonstrated with chymotrypsin; (b) the CV shell, studied with various negative-staining techniques, differs from the particle/chain material since it has no 3-4-nm globular subunits and reacts differently to alkaline pH; (c) the particle/chain material consists of aggregates of 3-4-nm globular subunits, four of which yield 8-10-nm fine particles; and these particles can be further aggregated into chains 8-10 nm wide and up to 30-60 nm long showing a "hollow" core; (d) vinblastine sulfate induced ringlike or helical crystalloid precipitates closely resembling the vinblastine-induced microtubule crystals reported in the literature, but vinblastine had no effect on either the CV shell material or the particle/chain material.


Radioimmunoassay for serum thyroxine-binding globulin: results in normal subjects and in patients with hepatocellular carcinoma. Serum thyroxine-binding globulin (TBG) was measured by radioimmunoassay. The human TBG used in this study was purified by affinity, anion-exchange, and gel filtration chromatography. The serum TBG concentration in 98 euthyroid normals was 1.48 +/- 0.46 mg/100 ml (mean +/- SD), which is one-half that previously reported using a similar method. The level in females (1.66 +/- 0.56) was significantly higher than that in males (1.37 +/- 0.37). Comparison of the serum TBG level and the maximum binding capacity of serum TBG for thyroxine (T4) yielded a molar ratio of 1:1 for T4 and TBG. The mean serum TBG in 19 patients with hepatocellular carcinoma was 2.10 +/- 1.29 mg/100 ml; however, only 2 of these patients had serum TBG levels outside the normal range.


Immunoassay of human TSH using dried blood samples. A sensitive, semi-quantitative radioimmunoassay method to screen for elevations of TSH concentration in blood is described. The method requires two 0.32 cm dots of dried blood-impregnated filter paper (equivalent to 3 mul plasma) and a 3-day incubation. Separation of bound and free is obtained using polyethylene glycol. The method can recognize TSH concentrations as low as 22 muU/ml using a highly sensitive antiserum developed by one of us (AFP). TSH in dried cord and newborn blood from 4 infants with congenital hypothyroidism was clearly higher than in normal infants. The method is suitable for use in a newborn screening program to confirm the suspicion of primary hypothyroidism in specimens with low T4 concentrations.


Demonstration of the capsular antigens of bovine group B streptococci by the serum-soft agar method. The elaboration of type-specific capsular antigens by group B streptococci can be demonstrated by the serum-soft agar technique. Group B streptococci isolated from bovine mastitis, namely, strains 9F, 14Mi, 8Mo, 44B, and 4S, were shown to form diffuse and compact types of colony morphology in serum-soft agar. Immunochemical and chemical analyses of antigens isolated from diffuse and compact colonies of strain 9F indicated that the diffuse-type growth of this strain was due to the elaboration of a galactose-rich surface antigen, whereas the compact 9F strain was devoid of this antigen. Specific 9F antiserum was effective in converting the diffuse 9F colonies of the compact type, indicating the presence of capsular material. Preliminary evidence suggests that the serum-soft agar technique could also be used to determine the antigenic diversity of the surface antigens of group B streptococci, thus providing an effective means of typing those organisms.


Antigenic relationships among influenza virua A neuraminidase (N2) antigens by immunodiffusion and postinfection neutralization tests. The antigenic relationships among the neuraminidases of influenza A strains from 1957 to 1973 were examined by postinfection application of neuraminidase antisera. This procedure causes inhibition of virus spread and apparent neutralization. Neuraminidase (apparent) neutralization and neuraminidase inhibition tests with chicken antisera gave similar results. Neuraminidase inhibition tests were more discriminating than neuraminidase neutralization tests when rabbit and goat antisera were used. Antibody absorption studies revealed that the neuraminidase, like the hemagglutinin, may possess two kinds of antigenic determinants, which can give rise to "common," or "cross-reacting," and "specific" antibodies. "Specific" antibody appears to be more effective in the inhibition of enzyme activity than in the inhibition of virus spread.


Comparison of Chlamydia subgroup A detection from clinical specimens after 40 and 64 hours of incubation in 5-iodo-2-deoxyuridine-treated McCoy's cells. The time course of formation of inclusion bodies produced by Chlamydia in 5-iodo-2-deoxyuridine (IUdR)-treated McCoy's cells was studied with the use of a known isolate of Chlamydia trachomatis D/UW-184/Ur and 47 frozen clinical urethral specimens previously shown to be either positive or negative for chlamydial inclusions after 3 days of incubation. Subsequent examination of 369 clinical specimens from the genitourinary tract over a 6-month period revealed 47 (13%) Chlamydia-positive cultures, all of which demonstrated inclusion bodies by iodine staining at 40 and 64 h postinoculation. Another 146 similar detected by iodine staining from 22 (15%). This study indicates that, although Chlamydia subgroup A inclusions are larger at 64 h, they can be readily detected from clinical specimens in IUdR-treated McCoy's cells at 40 h postinfection.


Method for identifying Salmonella and Shigella directly from the primary isolation plate by coagglutination of protein A-containing staphylococci sensitized with specific antibody. A technique is described that allows presumptive identification of either Salmonella or Shigella organisms directly upon the original isolation plate, in this case, MacConkey agar. This was accomplished by applying a drop of specifically sensitized protein A-containing Staphylococcus aureus over a "suspected" colony or several colonies of organisms grown on MacConkey agar. The plate is tilted to and fro to allow mixing of the particles with specific antigen that ir readily solubilized from the colony and observing for agglutination of the sensitized particles by use of a dissecting microscope. The agglutination can frequently be seen within 15 s, increasing in intensity over a 2-min period. The polyvalent Salmonella antiserum was slower in developing strong agglutination (1.5 to 2 min) compared to particles sensitized with group-specific antisera (15 to 45 s). A high-titer antiserum was important for a test reagent to have the required sensitivity.


Nomemclature of the proteins of cow's milk: fourth revision. This report reviews the nomenclature of the milk proteins of cow's milk in light of more recent advances in our knowledge. With the establishment of the primary structures of a number of these proteins, we now have a definite identification of alphas1-, kappa-, beta-, and the gamma-caseins as well as beta-lactoglobulin and alpha-lactalbumin. On the basis of new information on their primary structures and relationship to beta-casein polymorphs, changes in nomenclature have been recommended for proteins of the gamma-casein fraction. Although the primary structure serves as the unambiguous definition of proteins for which it is known, a more practical identification is necessary. We recommend that their behavior in gel electrophoresis under suitable conditions be employed for this purpose for all of the "major" milk proteins of raw skim milk except the immunoglobulins where, because of their heterogeneity and molecular genetics, physical parameters are less useful and their identification must be based upon antigenic determinants and their homology with their human counterparts. More work is needed and, with the accumulation of more information, additional changes in nomenclature can be expected for such proteins as the minor components of alphas- and kappa-caseins, alpha-lactalbumin, and the proteose-peptone fraction as well as further confirmation of the presence of immunoglobulins IgE and additional IgG subclasses. Additional components and genetic variants also can be expected.


Development and evaluation of a self-instructional program in endodontic diagnosis and treatment planning. This study was undertaken to evaluate two methods of instruction, the traditional lecture and the slide-tape method, both used with learning material developed for a course in endodontic diagnosis and treatment planning. Student performance, the amount of time required to complete the work, and the attitude of the students toward the differing methods of instruction were studied. The student sample, consisting of 108 members of the freshman class at the University of Southern California, was divided into two groups; the lecture group received a traditional two-hour lecture, and the slide-tape group had access to two slide-tapes on the same material. Both groups received a pretest and a posttest, and the students evaluated the teaching material. Statistical analysis showed no significant differences in quality of performance, but the lecture group spent more time learning the material, and students in the slide-tape group of instruction had a more favorable opinion of both the instructor (the voice) and the course material.