Characteristics of CD4^highCD25⁺ alloantigen-specific Treg. Freshly purified naive CD4⁺CD25⁻ T cells were cocultured with CD40-activated allogeneic B cells for the indicated time. The expression of cell surface markers (A) and intracellular cytokines (B) was determined and analyzed by FACS as described in “Flow cytometric analysis.” The percentage of positive cells for each cell surface marker or intracellular cytokine within the CD4^highCD25⁺ and CD4^mediumCD25⁻ subsets are indicated. The results shown are representative of 4 independent experiments.