Preparation of anti-GM4 antiserum and its assay by a solid-phase radioimmunoassay.
Anti-GM4 antiserum was prepared from rabbits by immunization with pure human brain GM4 ganglioside in complete Freund's adjuvant and methylated bovine serum albumin. None of the immunized animals developed any clinically apparent neurological dysfunction. The antiserum titer and specificity were analyzed by double immunodiffusion and a novel solid-phase radioimmunoassay (RIA). In the latter procedure, microtiter plate wells were coated first with the glycolipid antigen, followed by sequential application of antiserum and [125I]-Staphylococcal Protein A. The absorbed radioactivity in the well was then counted. Employing the RIA procedure, anti-GM4 antibody achieved a titer of 1:1600. The antiserum also exhibited a high degree of specificity to GM4; cross-reactivity with glycolipids of similar structure was negligible. The production of highly specific antiserum to GM4 and the feasibility of detecting antibodies to glycolipid antigens by a convenient solid-phase RIA should be useful to further study the biological and immunological roles of GM4 and other glycolipids in the central nervous system.
D000818:Animals; D000917:Antibody Formation; D000918:Antibody Specificity; D001666:Binding Sites, Antibody; D001667:Binding, Competitive; D003429:Cross Reactions; D005260:Female; D005732:Gangliosides / Q000276:immunology* / Q000378:metabolism; D006017:Glycolipids / Q000378:metabolism; D007106:Immune Sera / Q000032:analysis; D011817:Rabbits; D011863:Radioimmunoassay
D005732:Gangliosides; D006017:Glycolipids; D007106:Immune Sera; C028253:ganglioside, GM4