Structural characterization of viral neutralizing monoclonal antibodies to hepatitis B surface antigen

In this study we describe the viral neutralizing activity of murine monoclonal antibodies (MAb) specific for hepatitis B surface antigen (HBsAg). This viral neutralizing activity was assessed in vitro by employing Hepatitis Delta Virus (HDV) and human hepatocytes as target cells. To further characterize these viral neutralizing antibodies we generated a panel of anti-idiotypic (anti-Id) reagents and serologically characterized these antibodies for epitope specificity, Id specificity, and Id heterogeneity. Direct binding and competitive inhibition solid phase enzyme immunoassay have demonstrated that two murine MAb specific for HBsAg (anti-HBs), designated A1.2 and A3.1, recognize similar or overlapping epitopes on HBsAg, while monoclonal anti-HBs, designated A2.1 recognizes a unique HBsAg epitope. Further, Id analysis using monoclonal and polyclonal anti-Id reagents have identified both a private and a cross-reactive Id, respectively, on the anti-HBs, A1.2 preparation. The source of the idiotypic cross-reactivity between A1.2 and A3.1 has been identified, using Western blot analysis, to conformational determinants expressed by the heavy (H) and light (L) chains of these monoclonal anti-HBs. Lastly, the intrastrain antibody repertoire induced following HBsAg immunization was found to be relatively restricted in heterogeneity by clonotype analysis using isoelectric focusing and affinity immunoblot analysis. Interspecies variability in the anti-HBs response was observed based on epitope recognition using purified anti-HBs from a variety of species.