FIG. 2.

Frequencies of specific cytokine-secreting splenic CD8⁺ T cells increase after infectious challenge with T. cruzi in mice vaccinated with the heterologous prime-boost vaccination regimen. (A) C57BL/6 mice were immunized and challenged as depicted. Priming and boosting immunizations were performed as described in the Materials and Methods section. The mouse groups were the same as those described in the legend of Fig. 1. (B) Pools of splenic cells from 4 mice were restimulated in vitro in the absence (Med) or presence of the peptide VNHRFTLV (Pep), anti-CD28, BdGolgiPlug, and monensin. After 12 hr, the cells were stained with anti-CD8, anti-IFNγ, and anti-TNF. (C) The same as above expect that they represent only the cells stained with anti-CD8 and anti-IFNγ but not with anti-TNF. (D) C57BL/6 mice were immunized and challenged as depicted. (E) Splenic cells were restimulated in vitro in the presence of the peptide VNHRFTLV, anti-CD28, BdGolgiPlug, and monensin. After 12 hr, the cells were stained with anti-CD8, anti-IFNγ, and anti-TNF. The results are represented by medians (bars) from four individual mouse per group (dots). Two asterisks denote that the values of these groups were higher than those of the naïve control mice or all other groups (p<0.01), respectively. (F) Pie charts show the fraction of peptide-specific cells expressing the indicated molecules. The results are expressed as the mean values for four mice per group.