Antibody subpopulation in antihorse cytochrome c serum

Abstract

A specific antibody subpopulation(s) in antihorse cytochrome c serum was detected for peptide fragment 81–104 of cyanogen bromide (CNBr) cleaved horse cytochrome c (HCytc). This antiserum was made in the rabbit against polymeric horse cytochrome c. The presence of the peptide-specific antibody subpopulation(s) was demonstrated utilizing HCytc, CNBr-peptide 81–104 and isolated chymotrypsindigested HCytc fragments 60–67, 83–97 and 98–104 to compete with radio-labeled peptide 81–104 and antiHCytc serum in a competitive radioimmunoassay (RIA). This antibody subpopulation(s) in anti-HCytc serum was demonstrated to be specific for peptide 81–104. At the 50% inhibition level in competitive RIA, 100- and 1000-fold molar excesses of HCytc and its peptide 1–65, respectively, were required to affect an equivalent binding to that of the HCytc peptide 81–104. Competitive RIAs have been performed utilizing three different kinds of antigen to compete with HCytc peptide 81–104 and antihorse cytochrome c sera. These three kinds of antigens are: (1) endopeptidase digests of HCytc, (2) cytochrome c peptides 81–104 of several species and (3) several isolated chymotryptic peptide fragments of HCytc. The results have indicated that this peptide-specific antibody subpopulations(s) in antiHCytc serum is similar to antibodies made against peptide 81-104-BSA. Regions of antigenicity have been identified at positions 92, 100, 103 and 104 with both antisera.