Figure EV1. miPEP31 does not affect the inhibitory function of T_reg cells.

• A, B
  WT mice were treated with synthetic scPEP or miPEP31 at day 8 postimmunization every 2 days for three injections. FoxP3^EGFP+ T_reg cells were sorted from splenocytes derived from mice treated with synthetic scPEP or miPEP31. The CellTrace™ Violet (CTV)‐labeled memory T_conv cells (1 × 10⁵) were cultured in 96‐well plates for 72 h together with a decreasing ratio of sorted T_reg cells in the presence of anti‐CD3 plus γ‐irradiated antigen‐presenting cells (1 × 10⁵). The suppressive function of T_reg cells was determined by the proliferation of activated responder T cells (T_resp) on the basis of CTV dilution. Data in (B) are presented as mean ± SEM of four biological replicates, ns, not significant (1:2 or 1:4 or 1:8 or 1:16 miPEP31 vs. scPEP), nonparametric one‐way ANOVA, Kruskal–Wallis test.
• C–G
  Flow cytometric analysis of ICOS (C), LAG3 (D), GITR (E), and CTLA‐4 (F) expression in T_reg‐cell induction in the presence of 10 μM synthetic scPEP or miPEP31. Data in (G) are presented as mean ± SEM of six biological replicates. ns, not significant (scPEP vs. miPEP31), unpaired two‐tailed Student’s t‐test.