CD8 T_reg that undergo conversion after BMT exert contact-dependent, antigen-specific suppression and are more potent than CD4 T_reg. B6D2F1 recipients were lethally irradiated and transplanted with BM and T cells from B6.FoxP3-GFP donors 24 hours later. At day 7 after transplantation, LN and spleens were removed, and CD4⁺Foxp3⁺, CD8⁺Foxp3⁺, and CD8⁺Foxp3⁻ cells sort purified. These cells were cultured with the following responding cells: (A) CFSE-labeled CD45.1⁺CD3⁺ T cells, CD45.2⁺ DCs, and 1 μg/mL CD3 with CFSE dilution quantified in CD45.1⁺ cells 3 days later. IL-10 and IFN-γ in the tissue culture supernatant were assessed. (B) Sorted CFSE-labeled CD45.1⁺CD4 or CD45.1⁺CD8 T cells and CD45.2⁺ recipient (DBA/2) or third party (C3H/Hej) DCs with CFSE dilution analyzed in CD45.1⁺ CD8 or CD4 responders. (C) CFSE-labeled CD45.1⁺CD3⁺ T cells, CD45.2⁺ DCs, and 1 μg/mL CD3 with CFSE dilution quantified in CD45.1⁺ cells 3 days later. (D) Sorted Cell Trace-labeled responder CD4⁺ T cells and DCs from B6.WT, B6.β₂m^−/−, or B6.bcl-2 Tg mice with or without CD8⁺FoxP3⁺ T_reg as shown and violet dilution quantified 72 to 96 hours later. (E) Sorted CFSE-labeled CD45.1⁺CD4⁺ T cells and DBA/2 DC in transwell plates with CFSE dilution quantified in CD45.1⁺ cells 3 days later. (F) FoxP3⁺ populations were sort purified and cultured with CD45.1⁺CD4⁺ T cells and B6.MHC class II-GFPxDBA2.F1 DC. Costimulatory molecules were analyzed on GFP⁺ DCs after 72 hours of culture with histograms shown (open represents isotype; and tinted, costimulatory marker). For all experiments, representative plots are shown from 2 duplicate experiments.