Sequential intranasal or intramuscular vaccination with chimeric hemagglutinins induces stalk reactive antibodies.

<p>ELISA was used to determine the HA stalk-specific seroreactivity induced by the vaccination strategy for cohort I (cH6/1, cH5/1 and H1 protein) to cH9/1 hemagglutinin in a 7+1 reassortant PR8 virus. Total stalk-directed IgG titers were assayed for sera after the third immunization for individual IN (A) or IM (D) vaccinees. *p<0.01. Significant differences in stalk-specific antibody titers were detected at the indicated points in the groups receiving adjuvant compared to the unadjuvanted control group (A & D). The unadjuvanted IM vaccinees had significantly higher titers compared to naïve mice (D). The HA stalk seroreactivity induced by each immunization demonstrates the effect of the protein prime-boost-boost strategy adjuvanted with IVT SeV DI, poly(I:C), Addavax or protein administered alone for the IN (B) and IM (E) routes of administration. Stalk-specific IgG antibodies were subtyped by ELISA for the IN (C) and IM (F) groups. (F) The IgG1 signal in the Addavax group was saturated at OD₄₀₅=4. Error bars represent the standard error of the mean for each of five individual mice (A, B, D, & E) or the standard deviation for replicates of pooled serum samples (C & F). (C & F) Statistical analyses have not been included because replicates of pooled serum samples were assayed rather than biological replicates. The analyses for IN and IM immunizations in this figure were conducted independently and are not directly comparable.