Identification and characterization of monoclonal antibodies suitable for development of influenza A vaccine seed virus (VAC11P.1101)

Abstract

Abstract The seed viruses for influenza A vaccine production are high-yield (hy) reassortant viruses generated in embryonated chicken eggs by reassorting wild type (wt) viruses with a hy donor virus, A/Puerto Rico/8/1934 (PR8). Currently, reassortants with wt hemagglutinin (HA) and neuraminidase (NA) are selected for growth by using polyclonal antibodies (pAbs) against PR8. The efficiency and reproducibility of reassortment will be greatly enhanced with monoclonal antibodies (mAbs) which not only provide high specificity against PR8 but could also be prepared as defined reagents with guaranteed activity in unlimited quantities. In this study, from a panel of hybridoma clones prepared against PR8 surface glycoproteins, we found four HA specific mAbs with complete neutralization of PR8 virus in an in vitro plaque inhibition assay, which are designated as mAb-1B3, mAb-1H6, mAb-2A6, and mAb-2D11. All four mAbs have been shown highly specific in both in vitro and in ovo neutralization activity to PR8. Furthermore the epitopes of all four mAbs were mapped to the area around the receptor binding site (RBS) at the membrane-distal tip of HA. Their epitopes are composed of four regions of HA residues 158-172, 183-197, 208-222, and 253-267 (H1 numbering with signal sequence). The HA residues 158-172 and 253-267 are two common regions for all four mAbs, while residues 183-197 is specific for mAb-1H6 and mAb-2A6, and residues 208-222 is specific for mAb-2D11.