Identification of two novel anti-HCV E2 412-423 epitope antibodies by screening a Chinese-specific phage library

The hepatitis C virus (HCV) E2 412-423 linear epitope has been found to be highly conserved across multiple HCV genotypes. The antibodies against this epitope have broadly neutralizing activity. Considering the poor immunogenicity of the epitope in humans and significant diversity in the global distribution of HCV genotypes, the aim of this study was to construct an anti-HCV phage library by using a series of optimal strategies to screen novel broadly neutralizing antibodies from Chinese donors. mRNA was isolated from peripheral blood samples of 39 patients who were anti-HCV positive. A phage library was constructed by inserting a single-chain variable fragment (scFv) gene repertoire into the T7Select10-3b vector. A synthetic peptide representing the HCV E2 N-terminal 412-423 region was used as "bait" for bio-panning. The binding affinities of phage clones to the synthetic peptide were evaluated through peptide-ELISA. Two scFv clones (R3-19 and R4-85) showing the strongest binding affinities were selected. The complementarity-determining regions (CDRs) of these clones were aligned with those of other previously reported broadly neutralizing anti-HCV antibodies, and multiple conserved amino acid sites were found. The optimized procedures ensured that two novel scFv antibodies were isolated from a constructed phage library and showed specific binding to the poorly immunogenic HCV E2 412-423 linear epitope. Keywords: phage antibody library; hepatitis C virus; broadly neutralizing antibody; synthetic peptide.