Development of a high-throughput assay to detect antibody inhibition of low pH induced conformational changes of influenza virus hemagglutinin
Fig 5
CCI showed virus neutralizing activity in TMN after removal of head-binding antibodies.
A. Correct folding of major epitopes for GH HA1 from A/Hong Kong/1/68 (H3N2) (Sino Biological, Inc. China) was confirmed by appropriate Ab binding profiles. GH HA1 from A/Hong Kong/1/68 (H3N2) and ectodomain H3 rHA from A/Aichi/1/68 (H3N2) were coated on a nickel-coated plate, an ELISA was performed by using rabbit antisera and a panel of conformation specific mAbs. B. Removal of HI antibodies by serum adsorption with GH HA1 rHAs. The convenient human serum pool (Pool) and #115, the highest positive sample in HCCIA, were adsorbed with GH HA1 from A/Hong Kong/1/68 (H3N2) or double adsorbed with GH HA1 proteins from A/Hong Kong/1/68 (H3N2) and A/Perth/16/2009 (H3N2). Hemagglutination inhibition assays were performed by using A/Aichi/2/68 and A/Perth/16/2009. CCI were consistently present after mock or serum adsorption with GH HA1 in HCCIA (C) and the proteinase susceptibility assay (D). E. CCI neutralized A/Aichi/2/68 (H3N2) and A/Perth/16/2009 after head-binding antibodies were removed by serum adsorption with GH HA1 rHAs. ND: not done.