AFFINITY AND HETEROGENEITY OF ANTIBODIES INDUCED BY EPSILON-2,4-DINITROPHENYLINSULIN

Bovine insulin, substituted exclusively on its sole lysine residue with the 2,4-dinitrophenyl group, acts as a potent immunogen in guinea pigs. Anti-2,4-dinitrophenyl antibodies have been isolated from sera at various times after immunization with ε-2,4-dinitrophenylinsulin and these have been compared with guinea pig antibodies made against heavily dinitrophenylated bovine γ-globulin. Despite the relative homogeneity of ε-2,4-dinitrophenylinsulin, the antibodies induced by this immunogen display heterogeneity of ligand binding constants and electrophoretic migration comparable with antibodies evoked by heavily dinitrophenylated bovine γ-giobulin. By analysis of the average association constants for binding a homologous series of 2,4-dinitrophenyl ligands including the immunogen, it appears that portions of the immunogen adjacent of the 2-4-dinitrophenyl group vary in their contributions to the total free energy of the ligandantibody reaction depending upon the time of antibody isolation during the immune response. Although the affinity for each ligand increases with time after immunization, the rate of increase seems to be greater for smaller 2,4-dinitrophenyl ligands. As a result, antibody molecules formed late in the immune response form nearly equally stable complexes with various members of a structurally homologous series of 2,4- dinitrophenyl compounds. This property of high-affinity molecules seems to correspond qualitatively to their broader range of cross-reactions with structurally similar ligands. © 1969, American Chemical Society. All rights reserved.