Effect of molar ratio of fluorescent dye to IgG on the detection of lymphocyte surface immunoglobulins by immunofluorescence.
Labeled antibodies with different F/P molar ratios of FITC to protein (F/P molar ratio) were used for the detection of surface immunoglobulin (S-Ig) of human and mouse lymphocytes by membrane immunofluorescence, and the following results were obtained. 1. The percentage of S-Ig bearing cells increased markedly when labeled anti-human H- or L-chains antibodies were used with higher F/P molar ratios. The investigation of frozen kidney sections of mice injected with human immunoglobulin revealed that such an increase of the positive ratio in S-Ig was caused by increased non-specific adsorption of the fraction of labeled antibody with a high F/P molar ratio. 2. This non-specific adsorption phenomenon was observed at various intensities in materials from different species; materials from mcie showed less non-specific adsorption than those from humans. 3. It was possible to exclude reactivity with an Fc receptor using the top one third of the supernatant of labeled antibody centrifuged at 150,000 for 30 min.
D000818:Animals; D000888:Antibodies, Anti-Idiotypic; D005455:Fluorescent Antibody Technique; D005456:Fluorescent Dyes; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007074:Immunoglobulin G; D007143:Immunoglobulin Heavy Chains / Q000276:immunology; D007147:Immunoglobulin Light Chains / Q000276:immunology; D008214:Lymphocytes / Q000276:immunology*; D051379:Mice; D011947:Receptors, Antigen, B-Cell / Q000032:analysis*; D014357:Trypsin / Q000494:pharmacology
D000888:Antibodies, Anti-Idiotypic; D005456:Fluorescent Dyes; D007074:Immunoglobulin G; D007143:Immunoglobulin Heavy Chains; D007147:Immunoglobulin Light Chains; D011947:Receptors, Antigen, B-Cell; D014357:Trypsin