FIG 3.

Lysates of rVVs encoding individual IAV proteins stimulated the same immunodominance hierarchy that was stimulated by purified IAV antigens. (A) P815 cells were infected with rVVs encoding each of the 11 IAV proteins at an MOI of 10 for 1 h at 37°C in PBS containing 0.1% BSA (Sigma). Following overnight incubation, rVV HA-infected P815 cells were stained with mouse MAb TW2.3, specific for rVV RNA-binding protein (15), and H28-E23, specific for HA (16) (kind gifts from Jonathan Yewdell and Jack Bennink, National Institutes of Health, Bethesda, MD), and representative FACS plots are shown. Uninfected P815 cells were used as a control (dotted lines). (B) Autologous BLCLs pulsed with individual recombinant IAV proteins, HA, NA, M1, and NP, at different concentration were used to compare their activities to activate IAV-specific CD4⁺ T cell responses in the multispecificity T cell line with that of the lysates from P815 cells infected by rVVs encoding corresponding individual IAV proteins.