Fig. 4.

Transgenic parasites were able to induce granzyme B tumor-specific CD8⁺ T cells. gzmBCreERT2/ROSA26EYFP mice were vaccinated with two doses of PBS or the T. cruzi strain CL-14 expressing or not expressing NY-ESO-1. Treatment with tamoxifen was performed during 5 days as described in the methods section, and the splenocytes were collected 21 and 45 days after the boost vaccination. a The expression of EYFP, which is restricted to cells expressing granzyme B (gzmB), was used for gating. b The frequencies of EYFP⁺CD3⁺CD4⁺ and EYFP⁺CD3⁺CD8⁺ cells were determined. c The different subtypes of CD3⁺CD8⁺/gzmB⁺ lymphocytes were defined from the markers CD44, CD62L, and CD127, and the numbers of cells were plotted. d Representative pseudo-color plots demonstrates the gating of CD8⁺/gzmB⁺ cells (upper panels) collected 21 days after the boost and the assessment of the NY-ESO-1-tetramer⁺ (lower panels). The numbers represent the percentage observed, and the bar graph on the right summarizes the data obtained in three independent experiments. *P < 0.05, **P < 0.01, and ***P < 0.001. Within each time interval, the groups were compared with the control group PBS. Statistical analyses were performed using one-way ANOVA with Bonferroni posttest. Data are the mean ± s.e.m. of three to four independent experiments performed in triplicate