FIG 4.

Effector CD4 T cell responses to adjuvanted subunit vaccine in CCR2^−/−mice. Wild-type (WT) or CCR2^−/− mice were vaccinated as described in the legend to Fig. 2. To distinguish nonvascular cells from vascular cells in the lungs, mice were injected intravenously with fluorescence-labeled anti-CD45.2 antibodies, 3 min prior to euthanasia (CD45.2^+ve, vascular; CD45.2^−ve, nonvascular). On day 8 after vaccination, single-cell suspensions prepared from the lungs and bronchoalveolar lavage (BAL) fluid were stained with viability dye, followed by I-A^b/NP311 tetramers in combination with anti-CD4, anti-CD8, anti-CD44, anti-CD69, anti-KLRG1, anti-CD127, anti-OX40, and anti-CD62L antibodies (A and C to H). (A) FACS plots show percentages of tetramer-binding cells among CD4 T cells. (B) Percentages of vascular and nonvascular cells among NP311-specific CD4 T cells. Data are pooled from two independent experiments (A) or represent one of three independent experiments (B to H). Mann-Whitney U test; *, **, and *** indicate significance at P values of <0.05, <0.01, and <0.001, respectively.