Fig. 1.
The chimeric H5/1 construct with consensus H5 globular head and consensus H1 stem (cHA) and broadly cross-protective, stem-specific antibodies elicited by vaccination with cHAmg immunogens. (A) The constructs of swap H1/5 (H1 globular head and H1+H5[HA2] stem), swap H5/1 (H5 globular head and H5+H1[HA2] stem), and chimeric H5/1 (cHA: H5 globular head and H1 stem). (B) Neutralization activity against H1N1 California/07/2009 and H5N1 Vietnam/1194/2004 viruses. Titers of <400 (low) are shown in green, of 400 to 2,000 (medium) in yellow or orange, and >2,000 (high) in red. (C) The number of granzyme B (GrzB)-producing CD8+ T cells in splenocytes stimulated with HA (black bar) or PBS control (white bar) for 2 d in mice vaccinated with PBS (control), HA+Alu, or HA+C34 was evaluated by flow cytometric analysis. (D–I) The antibody titers from the mice vaccinated with cHAfg (blue) and cHAmg (red) adjuvanted with Al(OH)3 vs. cHAfg (green) and cHAmg (purple) adjuvanted with C34 were measured on day 42 by ELISA with the A/California/07/2009 H1N1 HA protein (D), A/Brisbane/59/2007 H1N1 HA protein (E), A/Brisbane/10/2007 H3N2 HA protein (F), A/Vietnam/1194/2004 H5N1 HA protein (G), A/Shanghai/2/2013 H7N9 HA protein (H), and the A/Brisbane/59/2007 (Bris/07) stem HA (no. 4900) protein (I) as the coating antigen. The endpoint antibody titer was defined as the last dilution of antisera to produce an absorbance 2.5 times higher than the optical absorbance produced by the negative control (preimmune serum). Data were examined by using Student’s t test and two-way ANOVA from Prism; differences were considered statistically significant at *P < 0.05; **P < 0.01. Data represent the mean ± SEM.