Fig. 3. Memory T cell recall responses against P. yoelii candidate antigens, P36 and P52.

Splenocytes and liver-mononuclear cells from immunized mice groups depicted in panel a of Fig. 2 were stimulated overnight in vitro with the P36 or P52 proteins and were tracked through flowcytometry to identify the activation of CD4⁺ and CD8⁺ memory T cells using the IFN-γ marker. Memory populations (Tcm and Tem/e) among CD4⁺ and CD8⁺ T cells of spleen (a and b) and liver (c and d) were tracked for IFN-γ production following P36 or P52 protein stimulation. A fluorescence minus one (FMO) control was used to gate on IFN-γ expressing CD8⁺ and CD4⁺ T cells (S Fig. 2, panels c and d). To quantify T cell activation following antigen stimulation, protein antigen was not added to unstimulated wells (UN). Frequency of T cell activation following either P36 or P52 protein stimulation was compared using the graphs on right side or at bottom of each panel using the marker IFN-γ. N = 10 mice per group. Data is from two or three independent experiments. Data are the mean ± SEM. Data were analyzed by the Mann-Whitney test. P < 0.05 is considered significant. *P < 0.05, **P < 0.01, ***P < 0.001 ****P < 0.0001.