Figure 6. HAC1 activation of multifunctional T cells. Human PBMCs isolated 21 d post vaccination were stimulated in vitro with HAC1 or the vaccine antigen or left non-stimulated (Neg) overnight. Cells were then fixed and analyzed for the presence of multifunctional T cells producing one or more of the cytokines TNFα, IL-2 and IFNγ by multi-parametric flow cytometry. (A) Total frequency of CD4⁺ cells producing each of the cytokines investigated. (B) Frequency of CD4⁺ cells producing only one, any two or all three of the cytokines investigated. *, ** and *** indicate significant difference with p < 0.05, p < 0.005 and p < 0.0005, respectively. Bars represent the mean ± standard error of the mean (SEM). n = 19.