FIG 8.

T-cell responses to CO92 challenge during long-term heterologous prime-boost vaccination study. A cohort (n = 5 per group) of immunized and naive control mice as described for Fig. 5 was challenged on day 105 of the study (84 days after 2nd vaccination) with 100 LD₅₀ of CO92. Spleens were harvested on day 3 p.i., and the isolated splenocytes were then stimulated with PMA, ionomycin, and brefeldin A. Cells were stained with T-cell surface markers CD3, CD4, and CD8, followed by intracellular IFN-γ and IL-17A staining. Percentages of CD4⁺ IFN-γ⁺ (A), CD8⁺ IFN-γ⁺ (B), and CD4⁺ IL-17⁺ cells (C) are shown. Cells were then analyzed by flow cytometry. Statistical significance was determined using one-way ANOVA with Tukey’s post hoc test as well as Student's t test. Asterisks above columns represented comparison to the control group, while asterisks with comparison bars denoted significance between other indicated groups. *, P < 0.05; **, P < 0.01; ***, P < 0.001; ****, P < 0.0001. Two biological replicates were performed and data plotted. In vitro studies had 3 replicates.