Induction of FOXP3 in naive human CD4⁺ T cells was TGFβ-dependent. (A) Flow cytometric analyses of FOXP3 with 259D on postsorted and day 5-activated CD45RA⁺ cells treated without exogenous TGFβ1 with or without anti-TGFβ mAb or with TGFβ1. (B) FOXP3 expression on postsorted and day 5-activated CD45RA⁺ cells and CD25^hi nTregs treated with and without TGFβ1. The level of FOXP3 expression for each population is represented by the mean fluorescence intensity (MFI) gating on FOXP3⁺ cells. (C) FOXP3 expression on postsorted and day 5-activated CD45RA⁺ cells cultured in the presence of serum without exogenous TGFβ or in the absence of serum with exogenous TGFβ1 or without exogenous TGFβ1 with or without anti-TGFβ. Data above are representative of 3 independent experiments. The numbers in each quadrant represent the percentage of cells expressing the antigen.