The Heterogeneity of Combining Sites of Anti-Benzylpenicilloyl Antibodies Obtained from Individual Rabbits: Changes in Combining Sites of γG and γM Antibodies During the Immune Response

Abstract

Summary: Nine rabbits were immunized with benzylpenicilloyl-human γG conjugate (BPO-HGG). Radioiodinated γG and γM were prepared from the anti-BPO-HGG sera of individual rabbits collected 10 days and 10 weeks after the first immunization. These antibodies were charged on a specific immunoadsorbent and eluted step by step with three haptens corresponding to constituents of the BPO molecule. Three antibody fractions were separated. The antibody fractions were adapted to 6-acetylaminopenicilloic acid (nuclear portion), phenylacetylglycine (acyl side chain portion) and benzylpenicilloyl-ε-aminocaproate (whole BPO molecule), respectively. The binding properties of these antibodies for BPO antigens were examined by equilibrium dialysis. The binding affinity of antibodies directed to the whole BPO molecule was stronger than that of antibodies directed to smaller parts of the BPO molecule. By PCA reactions the responses of antibodies complementary to the smaller parts of the BPO molecule were shown to be weaker than those of antibodies complementary to the whole BPO molecule. The combining sites of γG and γM antibodies changed with time after the immune response. At a late stage in the immune response a significant increase was observed in antibodies adapted to the whole BPO molecule. These changes of combining sites were more characteristic in γG than in γM. However, at an early stage of immune response no significant differences were observed between γG and γM. It is concluded that changes in the binding affinity and heterogeneity of antibodies during the immune response are caused by changes in the combining sites of the antibodies.