Sensitive and convenient quantitation of antibody binding to cellular antigens using glutaraldehyde preserved cells.
As a preliminary step in the identification and isolation of antibodies to human cancers, we have developed a sensitive and convenient assay for antibody binding to cellular antigens. The basis for the method is antibody binding to glutaraldehyde-fixed cells (AbGfC) and quantitation with radioiodinated staphylococcal protein A (SpA). Glutaraldehyde fixation of intact cells, which does not appear to effect the ability to form antigen-antibody complexes, provides a convenient and standard supply of target cells which may be stored at 4 degrees C and used in the assay over a period of several months. The amount of antibody specifically bound to the cells is quantitated by the addition of 125I-labeled SpA. The sensitivity of the method was compared with two complement-dependent cytotoxicity methods (trypan blue exclusion and 51Cr release assays) and tested with two antisera to human lung cancer and one antiserum to a membrane antigen of a murine lymphoma. These comparisons indicated much greater sensitivity when compared with the trypan blue exclusion assay and equivalent sensitivity with greater dose response characteristics when compared with the 51Cr release assay.
D000447:Aldehydes; D000818:Animals; D000912:Antibodies, Neoplasm / Q000302:isolation & purification*; D000937:Antigen-Antibody Reactions; D000951:Antigens, Neoplasm / Q000276:immunology; D000954:Antigens, Surface / Q000276:immunology; D003601:Cytotoxicity Tests, Immunologic; D004306:Dose-Response Relationship, Immunologic; D005976:Glutaral; D006801:Humans; D011817:Rabbits; D013205:Staphylococcal Protein A
D000447:Aldehydes; D000912:Antibodies, Neoplasm; D000951:Antigens, Neoplasm; D000954:Antigens, Surface; D013205:Staphylococcal Protein A; D005976:Glutaral