Preparation of monoclonal antibodies against poor immunogenic avian influenza virus proteins.

This study established a novel method of pre-screening peptides for monoclonal antibody (mAb) production. Whole virus particles were used as antigens to produce mAbs in the first stage. However, most mAbs obtained from this method were aimed toward hemagglutinin. For this reason, synthetic peptides were used as antigens for mAb production that aimed at the AIV proteins with low abundance or poor immunogenicity in the virus particle. The peptides that showed high immunogenicity were designed using bioinformatic tools for immunization. For high-throughput, a rabbit was used to screen the immunogenicities of the synthetic peptides. Those showed high immunity were used for mAb preparation in mice. Several new mAbs against PB2, PA, M1, M2, NS1 and NS2 proteins were successfully obtained in this study. Furthermore, the epitopes of M1 and NS1 mAbs were determined using competitive western blot assay and competitive ELISA. This study might simplify the mAb preparation and serves as the basis for developing mAb against poor immunogenic proteins.