Figure 3

TCR induced CD25⁺Foxp3⁺ T cells were immunosuppressive to CD4⁺CD25^- T cell proliferation in vitro. A. CD4⁺CD25^- T cells were cultured with anti-CD3 and anti-CD28 for 5 days. The converted CD4⁺CD25⁺Foxp3⁺ T cells were purified and washed extensively. The converted Tregs were then used at varying concentrations in a co-culture suppression assay along with CD4⁺CD25^- (5 × 10⁴) T cells pre-labeled with CFSE as responders and autologous monocytes (2 × 10⁵) as accessory cells. Anti-CD3 antibody was added into the start of the co-culture suppression assay (0.5 μg/ml). CFSE dilution of responder cells was measured after 72 hrs using flow cytometry. B. IFN-γ production of responder cells in the co-culture assay as detected by flow cytometry after 72 hrs. iTreg: induced Foxp3⁺CD4⁺CD25⁺ T cells. The experiment was repeated for three times with similar results.