A rapid, automated colorimetric assay for measuring antibody binding to cell surface antigens.
An automated, colorimetric procedure is described for detecting antibodies specific for cell surface antigens. The procedure entails (a) coating the wells of 96-well microplates with either protein A or anti-immunoglobulin antibodies and (b) preincubating either the microplate or target cells with the test antibody. Target cells which react with the test antibody bind to the wells of the microplate and bound cells are quantitated by staining with the dye Rose Bengal. A microplate spectrophotometer is used to measure absorbance in each well of the plate, providing a rapid, automated measure of antibody titre. The assay is simple to perform, uses readily available reagents and gives comparable sensitivity to rosetting assays. With these features, and the capacity for handling large numbers of trays quickly, this method has obvious advantages in screening for antibody activity in culture supernatants of hybridoma clones.
D000818:Animals; D000911:Antibodies, Monoclonal / Q000032:analysis* / Q000276:immunology; D000918:Antibody Specificity; D000954:Antigens, Surface / Q000276:immunology*; D001322:Autoanalysis; D001666:Binding Sites, Antibody; D003601:Cytotoxicity Tests, Immunologic; D007163:Immunosorbent Techniques; D008214:Lymphocytes / Q000276:immunology / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D011863:Radioimmunoassay; D012397:Rosette Formation
D000911:Antibodies, Monoclonal; D000954:Antigens, Surface