The Relationship of Binding Affinity to b4 Allotype in Rabbit Anti-Dinitrophenyl Antibody

Abstract

Abstract The b4:b- ratio has been studied in anti-DNP antibody populations of different affinities, obtained from b4b4 rabbits immunized with DNP185-KLH. The average intrinsic association constant (K0) as well as the b4 percentage were determined in fractions of antibodies obtained by the subsequent addition to serum of limiting amounts of DNP45-BGG. In two reported experiments the sera of two lots of animals bled at several intervals of time after the initial injection of immunogen were pooled, in order to have a wide range of combining affinities. In a third experiment, the whole antibody of single bleedings, taken at the same intervals of time from individual animals, was isolated by means of a DNP-lysine-Sepharose adsorbent. In one of these rabbits the antibody was fractionated by the addition of antigen to each bleeding, in order to study higher affinity populations. An exponential relationship between antibody binding affinity and b4 light chain content was found in both types of experiments. Results reported in this paper show: a) an absolute increase in b4 (or κ) light chains in the pool of rabbit antibody populations during the maturation of the immune response and, conversely, a decrease in b- (or λ) chains, b) the coexistence, in the serum of immune animals, of several quite discrete populations of b+ molecules of different binding properties, the relative proportions of which vary with the time of immunization, c) the probable existence of a lower level of average binding affinity in b- than in b+ molecules.