Immunological Activity of Cytochrome c III. ENHANCEMENT OF ANTIBODY DETECTION AND IMMUNE RESPONSE INITIATION BY CYTO- CHROME c POLYMERS

Antibodies which fail to precipitate with the monomeric homologous cytochromes c were elicited in rabbits by the conjugate of bovine y-globulin and cytochrome c (tuna and turkey proteins) or by free monomeric cytochrome c (kangaroo and, in one case, the horse protein). Such antibodies do form precipitates and fix complement when allowed to react with the polymerized homologous antigen. In contrast, antisera elicited by monomeric human, MQCQCQ mulaffa, or horse cytochrome c precipitate with the monomeric homologous antigen; these antisera also give much stronger reactions, both in precipitin and complement fixation tests, when reacted with polymeric preparations of the antigens than with monomers. In all cases but one, the reactions of the polymers were completely or nearly completely inhibited by an excess of the monomeric protein, and these inhibitions were highly efficient. Such differences in immunological behavior between polymeric and monomeric preparations of cytochrome c can be accounted for by the increase in antigenie valence on polymerization. As immunogens, polymeric preparations of the cytochromes c from the horse, tuna, M. mulaffa, kangaroo, and rabbit are much more effective in rabbits than the corresponding monomeric proteins, and except for the rabbit protein, yield antibodies in amounts comparable to those elicited by other effective soluble protein antigens. In the case of human cytochrome c, polymers and the monomer are good and equally effective immunogens. Polymers fix more complement and precipitate more antibody than monomeric preparations when reacted with antisera elicited by polymeric, as well as monomeric preparations. These antisera contain no antibodies specific to the polymerized forms of the cytochromes c. Possible reasons for the large enhancement of cytochrome c immunogenicity on polymerization are discussed. Of all cytochromes c examined, only the human protein is strongly immunogenic as the monomer.