Hemagglutinin from multiple divergent influenza A and B viruses bind to a distinct branched, sialylated poly-LacNAc glycan by surface plasmon resonance.

Influenza viruses initiate infection via specific interactions of hemagglutinin (HA) with host cell surface sialic acid-containing glycans. Antigenic drift has resulted in HA amino acid sequence changes that affect binding properties for sialic acids. Further, viral propagation in eggs and cell culture for vaccine production can yield variants with mutations that affect the conformation and affinity of HA for sialic acids. Therefore, influenza vaccine researchers and manufacturers need robust analytical methods to assess directly the ability of vaccine candidates to bind to their specific sialic acid ligand. We developed a surface plasmon resonance method that uses an extended, biantennary glycan terminating with α-2,6 linked sialic acids to bind influenza HA and assess this interaction. Recombinant HA (rHA) from both influenza A and B viruses isolated from 1999 to 2017 strongly and specifically bind this sialic acid ligand, suggesting the binding ability of divergent HA for this ligand is resistant to antigenic drift. Importantly, the method can differentiate between wild type and mutant rHA for which binding to this sialylated glycan and red blood cells in hemagglutination assays is compromised. We believe this method can be a powerful tool to screen influenza A and B vaccine candidates and final vaccine preparations for their functional ability to bind sialic acids, which allows manufacturers to identify preparations in which mutations that affect sialic acid binding have arisen during propagation. Evaluation of vaccine rHA antigen integrity by confirmation of the receptor binding site functionality is a prudent cautionary step to assure the antigenic quality of seasonal influenza vaccines.