Characterization of the reactivity pattern of murine monoclonal antibodiesagainst wild-type hepatitis B surface antigen to G145R and other naturallyoccurring "a" loop escape mutations

The hepatitis B surface antigen (HBsAg) "a" domain harbors major B-cell epi topes. Viruses with mutations in this region emerge after vaccination or du ring hepatitis B immune globulin (HBIg) prophylaxis. A strain with G145R re placement has been almost invariably isolated as a major escape mutant. We investigated mutant antigen-antibody interactions with direct binding assay s. G145R and 16 other naturally occurring recombinant HBsAg mutants were ex pressed in mammalian Cos-1 cells. The reactivity of a panel of 28 murine an ti-hepatitis B surface antigen (anti-HBs) monoclonal antibodies to mutant a ntigens was measured with enzyme immunoassay and expressed as percentage co mpared with the wild-type (wt) HBsAg signal for each antibody. All point-mu tated proteins displayed diffuse intracellular immunofluorescent labeling c orresponding to a secretory pathway. Monoclonal antibodies (mAbs) were clas sified according to different binding patterns. The effect of mutations on antibody binding differs depending on the amino acid involved and on the lo cation within the "a" loop. As expected, most antibodies had absent or negl igible binding (<40%), notably with residue 145 replacements. However, we i dentified antibodies that reacted with conformational epitopes but neverthe less had adequate reactivity (>40%) with all mutant antigens, including G14 5R. The effect of G145R was more pronounced than that of G145A. A subgroup of antibodies had substantially increased recognition (>120%) of antigens w ith mutations in the first loop. We demonstrated that antibodies can be sel ected or combined that react with all mutants investigated, including G145R . These data offer perspectives for improving anti-HBs-based protection aga inst hepatitis B.