Identification of a novel linear B-cell epitope in the N-terminus of varicella-zoster virus glycoprotein B.

Varicella-zoster virus (VZV), a highly contagious human alphaherpesvirus, imposes a substantial economic burden on global healthcare systems. Glycoprotein B (gB), a central fusogen mediating viral entry and cell-cell fusion, represents a promising target for developing VZV vaccines and diagnostic reagents. This study aimed to systematically characterize the antigenicity of VZV gB and map novel linear epitopes to facilitate the development of targeted diagnostics. Methodologically, recombinant glycoprotein B was successfully expressed in a eukaryotic expression system and subsequently purified using nickel-affinity chromatography, and its antigenicity was confirmed with VZV-positive convalescent human serum. Six monoclonal antibodies (mAbs 3B9, 4D2, 5E2, 6G7, 8C9 and 10F11) specifically targeting gB were generated via hybridoma technology. Subsequently, three rounds of progressive truncation using a peptide-overlapping approach and alanine scanning mutagenesis, were employed to precisely identify the antibody binding sites. The results demonstrated that several generated mAbs, particularly the primary detection candidate mAb 6G7, exhibited high titers (up to 1:1024,000) and strong binding affinity (Ka ≈ 7.89 × 10⁸). Crucially, we identified a novel linear B-cell epitope, 97LGDGDEIR104, located within the flexible N-terminal region of gB, which was critical for antibody binding. This epitope lies proximal to previously reported functional motifs involved in membrane fusion. In conclusion, our findings reveal a new antigenic site on VZV gB, providing a robust experimental foundation for the development of epitope-specific serological assays and vaccine strategies.