Biomedical subjects
Publications and source records attributed to F Klein.
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We isolated and purified the main lipids of the rat sciatic nerve. After methanolysis, fatty acids were isolated and purified by thin layer chromatography, and studied by gas chromatography. C 16 and C 18 fatty acids are the most abundant. Among the long-chain fatty acids, arachidonic acid (20:4) is present in some lipids; highly desaturated fatty acids in C 22 and C 24 are also present. In general, the fatty acids are highly saturated; cholesterol esters and ethanolamine phosphoglyceride fatty acids are highly unsaturated.
Infectious Epstein-Barr virus (EBV) was produced from suspension cultures of P3HR-1 cells. A protocol for the scaled-up production and concentration of the virus was developed. Virus from the culture fluid was concentrated by continuous-flow pelletization or continuous flow with banding in sucrose. EBV prepared by pelletization yielded 1.7 X 10(4) infectious units/ml (100 X concentration) and less than 3.4 X 10(7) EVB particles/ml (1,000 X concentration), whereas EBV prepared by banding yielded 4.6 X 10(3) infectious units/ml (100 X concentration) aand 1.3 X 10(8) EBV particles 1 ml (1,000 X concentration). The majority of the virus particles observed were "empty" membrane-associated particles. A statistical test of the correlation between infectivity and virus particle count was made.
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Two enzymatic methods for the determination of cholesterol in serum have been modified for automatic use. These methods utilize the colour reactions according to Kageyama or Trinder to measure the hydrogen peroxide resulting from the enzymatic reaction. The procedures have been compared with the classical methods according to Abell and to Huang. Details regarding the procedures are given. Comparisons made during routine analysis of a large number of different sera have been expressed in the form of regression equations. The standard errors of the methods are given along with the standard deviations for differences between methods. Some conversions of values from one method into another are also given. It appears that the automated Kageyama method can be used for large scale practice. The manual Kageyama method is not very satisfactory because of its large experimental error. A disadvantage of the Kageyama methods is the necessity of having blank corrections for haemolytic sera. The Trinder method avoids the problem and has some additional advantages.
In two patients with a persisting, high antistreptolysin titre the antistreptolysin activity in both cases resided exclusively in a monoclonal IgG component in the serum. This component had all the characteristics of a true antibody. A history in both patients of arthritis with or without angina suggested that the monoclonal antibodies were reactive in origin, although definite proof was lacking. In one case there was a suggestion of incipient myeloma. Whenever an extremely high antistreptolysin titre persists after antibiotic treatment the possibility of paraproteinaemia should be considered.
In the course of our study on the lipids of the rat sciatic nerve, the analysis of the neutral lipids allowed us to detect and characterize cholesteryl esters present at a relatively high level (5%). Among the phospholipids, ethanolamine phosphoglyceride is the most abundant fraction and contains neraly all the plasmalogens (20% of total lipid phosphorus). The glycolipids consist of five different fractions; the cerebrosides with hydroxy fatty acids account for 38% of total glycolipids. Monogalactosyl diglyceride represents 7% of total glycolipids.
Three subgroups of human IgM can be distinguished on the basis of differences of slopes of the D2 versus absolute concentration plot in the radial immunodiffusion technique (Klein et al., 1973). Individual IgM fractions, whether mono- or polyclonal, always belong to only one of these groups. The differences between the subgroups disappear after reduction of the IgM to 7S subunits. These findings provide an explanation for the large discrepancies between absolute IgM determinations in different laboratories. It follows that most of the readings of individual IgM values in the Mancini technique must be wrong by any standard, including WHO reference preparations. The insertion of a simple reduction step in the assay abolishes the differences in quantitative reactivity between IgM subgroups as well as between natural 7S and 19S IgM. This allows an absolute determination of both forms together. The values thus obtained differ considerably from the estimates given by Humphrey and Batty (1974). It also appears that the International Units of the WHO do not represent the same quantity of IgM in different reference sera. The modified method allows the determination of total monoclonal as well as polyclonal human IgM by weight.
Procedures were developed to produce infectious Epstein-Barr virus in large glass fermentors with limited mechanical agitation by using an intermittent culture collection system. The system appears quite suitable for use in fermentation vessels of 50 liters or larger.
We present a pattern of fractionation of total lipids, leading to the isolation of individual glycolipids, MGDG contains as sugars, a high quantity of galactose and a low level of glucose. The study of the fatty acids distribution shows: a) a high level of saturated short chains fatty acids, b) a high level of palmitic acid (C 16:0, 68% of the total).
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(1) The phagocytosis of human IgG, IgM, and C3 by granulocytes from various rheumatoid and nonrheumatoid sera and synovial fluids (SF) was investigated by direct examination of the patient's leucocytes and indirect testing by incubation of normal donor leucocytes with various sera and SF. (2) In rheumatoid arthritis (RA) phagocytosis of IgM, IgG, and C3 was common from sera and SF. There was a strong correlation of IgM and C3 phagocytosis with the occurence of rheumatoid factor. The phagocytosed IgM is probably rheumatoid factor. In SF both the direct and indirect test method yielded equally positive results; in serum the direct test was negative throughout. (3) In systemic lupus erythematosus there was phagocytosis of IgG, IgM and C3 from serum (indirect test), IgM not being correlated with the latex-fixation test and probably of antinuclear antibody nature. Phagocytosis decreased after treatment of the disease. Sera from many other rheumatic disease frequently gave weak IgG phagocytosis, but rarely did IgM or C3. (4) IgG, and sometimes C3, was frequently taken up from IgG myeloma sera (indirect test). IgM and IgG were taken up from Waldenström's macroglobulinaemia sera, independent of IgM concentration. It is possible that an aggregation tendancy of particular paraproteins determines Ig uptake from these sera. (5) IgG was taken up from half of the studied sera of infectious diseases in the indirect test, including two cases with Hodgkin's disease as well. Three sera from patients with untreated trypanosomiasis were positive for IgG as well as for IgM. (6) Normal healthy control sera remained negative, even after prolonged preservation or frequent freezing and thawing: only among very old sera were a few positive observations recorded. Immunoglobulin phagocytosis appears to be a common phenomenon in a number of conditions. It seems probable that soluble immune complexes, or in other cases nonimmune aggregates, may cause phagocytosis.
JLS-V9 mouse bone marrow cells were readily adapted to suspension culture, chronically infected with Rauscher leukemia virus (RLV), and subsequently grown in 7.5- and 14-liter New Brunswick fermentors. The suspension-type cell system can be modified to produce virus with clearly defined properties, such as high ribonucleic acid-dependent deoxyribonucleic acid polymerase (RDDP) activity, high particle count, and high infectious particle count. Biological and biophysical properties of suspension-produced RLV were not affected by concentration and purification employing continuous-flow and rate-zonal centrifugation procedures. The RDDP assay was standardized and showed a linear incorporation of (3)H-thymidine 5'-monophosphate ((3)H-TMP) up to 30 min. Further characterization indicated that a high percentage of (3)H-TMP incorporation was due to RDDP.
An in vitro system for production, purification, and concentration of mouse mammary tumor virus is described. Monolayer cultures of C(3)H mouse mammary tumor cells propagated at 34 C in roller bottles in the presence of dexamethasone, a glucocorticoid hormone, release B-type particles which possess ribonucleic acid and a ribonucleic acid-dependent deoxyribonucleic acid polymerase. One thousandfold concentration by ultracentrifugation with subsequent gradient fractionation yielded > 7 x 10(10) particles per ml in the 1.16- to 1.18-g/ml region. Mouse mammary tumor virus produced in this system was free of detectable C-type virus.
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