EmTh-1 cells impair tumor-induced iTreg generation. CD4⁺CD25⁻CD62L⁺ naive T cells were isolated from mouse spleens using magnetic activated cell sorting. The cells were activated using T-cell expander beads (cell:bead ratio 1:1), resuspended in the culture medium from different tumor cell lines (12B1, B16, 4T1), and treated with emTh-1 supernatant. (A) FoxP3 expression was determined by flow cytometry. (B) Pooled data from 3 independent experiments are depicted. Student t tests were used to analyze the diagrams. *P < .001, a significant difference compared with the corresponding group without emTh-1 supernatant.