Theoretical considerations of the ability of monoclonal antibodies to detect antigenic differences between closely related variants, with particular reference to heterospecific reactions

Abstract

In theory monoclonal antibodies can be used to analyse antigenic determinants in great detail by correlating differences in antibody affinity for variant antigens with their amino acid differences. In particular, heteroclitic antibodies should be detected, which would normally be masked in a polyclonal antiserum. Recognition of such antibodies may be important for our understanding of the scope of antibody repertoires particularly when the immunogen is closely related to a component of the immunised animal. In practice the immunoassays commonly used to measure affinity differences between different antigens fall short of these capabilities. Mathematical studies were carried out to identify factors controlling the sensitivity of 4 types of assay to differences in affinities for different antigens. The most important factors controlling assay sensitivity were found to be the ratio of antibody affinity ( K) to epitope density in direct binding assays, the ratio of K to antibody concentration in liquid phase competition assays, and the ratio of solid phase to liquid phase values of K for solid-phase competition assays. It is predicted that a combination of solid-phase competition assay with high epitope density and direct binding assay with low epitope density would result in optimal detection of (1) heteroclitic antibodies and (2) small differences in antibody affinity for cross-reactive antigens.