Biomedical subjects
Publications and source records attributed to D Bout.
A prototype of automatized enzyme-linked immunosorbent assay (ELISA) in tubes is described, using a commercially available basic material, easily modified. Nine hundred samples could be completely studied in a day by only one person. The different steps of the automatized ELISA were systematically studied to obtain the best performance. Its application is described in toxoplasmosis serodiagnosis.
A complex antigenic preparation obtained from Listeria monocytogenes serovariant 4b by freeze-pressing, centrifugation, and gel filtration treatment was studied by crossed immunoelectrophoresis, with the aim of preparing an antigenic fraction that could be used to investigate the serological response to listeric infection. Of 17 immunoprecipitates revealed in the soluble extract, one of three major antigens (designated antigen 2) was shown to be a strong antigen in humans or rabbits infected with L. monocytogenes serovariant 4b. A monospecific antiantigen 2 serum was obtained and used to prepare a serologically homogeneous antigen by immunoadsorption. Antigen 2, most probably located on the bacterial surface, is common to all serovariants of L. monocytogenes and to Listeria grayi and is not shared by the main bacterial species known to have common antigens with L. monocytogenes.
Microdouble diffusion technique using whole hydatid antigen and monospecific antiserum against Echinococcus genus-specific antigen 5 was applied to the diagnosis of human hydatid disease. The use of this simple and economical method may be extended to the specific diagnosis of parasitic diseases.
Using Schistoma mansoni schistosomula collected in vitro and sensitized with 42-day-infected rat serum, normal rat peritoneal cells enriched in eosinophils were proven to subsequently adhere to and kill the schistosomula within 24 to 48 h. The cell-dependent, heat-stable antibody in infected rat serum reached a peak between 30 and 42 days after infection. Inhibition experiments with aggregated immunoglobulins indicated the role of IgG2a antibody in the adherence of eosinophils to sensitized schistosomula. The immune absorption technique showed that IgG2a antibody was involved in the mechanism of cytotoxicity of effector to target cells, whereas the role of IgE antibody could be excluded. Ultrastructural studies revealed the constant presence of eosinophils and mast cells in contact with schistosomula. The use of purified cell populations showed that the cytotoxic effect of the original cell population was significantly decreased after depletion of mast cells and partially restored after addition of mast cells. These observations, together with those concerning the role of IgE immune complexes in macrophage cytotoxicity, suggest the possible participation of anaphylactic antibodies in immunity to schistosomes in the rat.
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Inhibition enzyme immunoassay was applied to human apolipoprotein B (apo-B) from plasma. The technical conditions of the assay were determined. The detection limits of the assay were 200 ng to 10 microgram/ml. Correlation coefficients obtained between enzymoassay and rocket immunoelectrophoresis on one hand and radial immunodiffusion on the other were respectively 0.84 and 0.80. The inhibition enzymoassay provides a specific and highly sensitive method for the quantitation of apo-B.
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We used enzyme immunoassay to measure apolipoprotein B concentration in human plasma. Pure lipoprotein B was isolated from serum samples of fasting normolipidemic subjects by sequential preparative ultracentrifugation and coated to a polystyrene tube surface by adsorption. Human serum samples and rabbit antiserum to human apolipoprotein B were incubated with the solid-phase lipoprotein B. Soluble antigen competed with solid-phase antigen for binding to antibodies. After washing, peroxidase-labeled sheep antibodies against rabbit immunoglobulins were added, and after further washing the bound label was assayed. This provided a direct measurement of the soluble antigen. The best technical conditions for the assay were determined. The minimum detectable concentration was 1 microgram per assay. The enzyme immunoassay yielded values that compare favorably with those obtained by radial immunodiffusion (r = 0.84) and by rocket immunoelectrophoresis (r = 0.80). The assay offers several advantages over existing techniques: sensitivity, specificity, simplicity, ane non-use of radioisotopes.
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Schistosomicide drugs are used as ligands to isolate the target antigens of schistosome by affinity chromatography. Anti-target antigens immune sera produced in rabbit permit their localization on the parasite using immunofluorescence. Target antigens of some drugs determine a relatively high immunoprotection in rat or mouse schistosomiasis infection.
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Immunoglobulins, anti-Schistosoma mansoni antibodies, complement components and schistosome antigens were investigated in milk from mothers infected with S. mansoni. No significant differences of immunoglobulins or complement component levels were observed between infected and control mothers. Anti-S. mansoni antibodies were detected in the milk of 8 out of 25 infected mothers. A significant relationship was observed between serum and "4", were demonstrated in milk from infected patients by the double diffusion micromethod. The function of these immunologically active substances transmitted by milk from mother to child is discussed.
A review of some applications of the Enzyme-Linked Immunosorbent Assay (ELISA) for the serodiagnosis of parasitic infections is presented. Examples are given of applications with trichinellosis, hydatidosis, schistosomiasis, toxoplasmosis, african trypanosomiasis and amoebiasis. ELISA is believed to be a valuable addition to existing serological tools, especially due to its sensitivity and ease of performance.
A new purified antigen (E2) of Paracoccidioides brasiliensis mycelial growth phase was isolated by immunoadsorption from a crude metabolic soluble extract of the fungus. The antiserum prepared in a rabbit by inoculation of E2 antigen developed only one immunodiffusion line with the crude metabolic extract. Findings on immunological analysis showed that E2 antigen is the antigenic component of immunoelectrophoretic band E. The isolated antigens did not possess detectable alkaline phosphatase activity. It reacted in immunodiffusion tests with all the sera (14/14) from P. brasiliensis infected patients containing precipitating antibodies.
Circulating immune complexes (CIC) were investigated by the [125I]Clq binding test, the complement fixation test (CFT) and optical density measurement after redissolving 3% polyethylene glycol precipitates of serum from patients infected by Schistosoma mansoni. A highly significant correlation was obtained among these three techniques. More than 60% of the patients demonstrated significantly higher values than control individuals. The level of CIC was found to be higher in the mild than in the hepatosplenic form of the disease. Parasite antigen, IgG, IgM and IgE were characterized in these CIC. In experimental schistosomiasis in mice, maximum levels of CIC, evaluated by the CFT, were observed between the 40th and the 70th day of infection.
Circulating immune complexes (IC) were detected and quantified in serum of patients with rheumatoid arthritis (RA) by three methods: radiolabeled Clq binding test, anticomplementary activity and optical density of 3% PEG precipitated sera. Rheumatoid factor (RF) is characterized in PEG precipitates by the Waaler-Rose reaction. The relationship between IC, complement and RF levels are analyzed.
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