Data Availability StatementGlycan microarray data as well as the session file for the Pleased analysis are available within the NCFG site (https://ncfg. of chicken and guinea pig erythrocytes to gain insights into reduced agglutination properties THIQ displayed by drifted strains and show that both chicken and guinea pig erythrocytes contain complex sialylated N-glycans but that they differ with respect to the extent of branching, core fucosylation, and the abundance of poly-N-acetyllactosamine (PL) [-3Gal1-4GlcNAc1-]n structures. We also examined binding of the H3N2 viruses using three different glycan microarrays: the synthetic Consortium for Functional Glycomics array; the defined N-glycan array designed to reveal contributions to binding based on sialic acid linkage type, branched structures, and core modifications; and the human lung shotgun glycan microarray. The results demonstrate that H3N2 viruses have progressively lost their capacity to bind nearly all canonical sialylated receptors other than a selection of biantennary structures and PL structures with or without sialic acid. Significantly, all viruses displayed robust binding to nonsialylated high-mannose phosphorylated glycans, even as the recognition of sialylated structures is decreased through antigenic drift. IMPORTANCE Influenza subtype H3N2 viruses have circulated in humans THIQ for over 50?years, continuing to cause annual epidemics. Such viruses have undergone antigenic drift in response to immune pressure, reducing the protective effects of preexisting immunity to previously circulating H3N2 strains. The changes in hemagglutinin (HA) affiliated with drift have implications for the receptor binding properties of these viruses, affecting virus replication in the culture systems commonly used to generate and amplify vaccine strains. Therefore, the antigenic properties of the vaccines may not directly reflect those of the circulating strains from which they were derived, compromising vaccine efficacy. In order to reproducibly provide effective vaccines, it will be critical to understand THIQ the interrelationships between binding, antigenicity, and replication properties in different growth substrates. value and are indicated at the top of the spectrum. Blue box, value. Some glycans appear in THIQ two or all of the top three fractions. See Fig. 1 legend for cartoon key. Binding of H3N2 from the 2017-to-2018 season. We also examined three strains obtained from patients in 2017, during one of the worst influenza epidemics in several years. Sequence analysis revealed that each virus is of subtype H3N2 (Table 3). These infections were briefly amplified in MDCK-SIAT1 cells and useful for receptor binding research for the glycan microarrays then. These infections also didn’t agglutinate poultry RBCs and exhibited minimal agglutination of guinea pig RBCs (Desk 3). Binding towards the N-glycan array was negligible (Fig. 7), following a trend of the earlier H3N2 drift viruses. Interestingly, the profiles of these strains were highly restricted on the CFG arrays (Fig. 7) with a clear distinction between high levels of binding to a few determinants and moderate/low binding to the other available glycans. Similarly to the drift strains, these viruses were able to bind long-chain PL structures on the CFG array that did not terminate in sialic acid. Clinical isolates L and N exhibited high levels of binding to both 2,6-Sia-terminating and 2,3-Sia-terminating structures. Only the 2 2,6 linkage was found on the sialylated glycans in the top binders F2rl1 for medical isolate M. Nearly all glycan determinants identified by THIQ this stress took the proper execution of nonsialylated long-chain PL terminating in the galactose residue or for 2?h and were resuspended in 2?ml of PBS buffer.