Figure 2.

Schematic of strategy used to identify recurring immunoglobulin classes that recognize conserved hemagglutinin (HA) stem epitopes. Two weeks after an H5N1 vaccine boost, peripheral blood IgG⁺ memory B cells capable of binding HA from different influenza subtypes were detected using an HA probe mutated to prevent nonspecific binding to sialic acid. B cells able to bind multiple group 1 subtypes (H5 and H1) or both group 1 and group 2 subtypes (H5 and H3) were single-cell sorted, and immunoglobulin heavy and light chains for each cell were polymerase chain reaction (PCR) amplified and sequenced. Representative heavy and light chain pairs were cloned, expressed as monoclonal antibodies and tested for binding and neutralization. The pie charts show the prominent IGHV genes expressed by B cells within each category with the number of sequences analyzed indicated in the center of each chart. The predominant IGHV gene used by group 1 HA-binding B cells was IGHV1-69. Few group 1/2 HA-binding B cells expressed IGHV1-69. Instead, three classes of cross-group HA stem-binding and neutralizing B cells encoded by IGHV1-18 and IGHV6-1 were identified in multiple donors after H5N1 vaccination as indicated. IGHV genes, including IGHV3-30 and others not referenced here, are colored in grey. More details can be found in Joyce et al. (2016).