CD4(+)CD25(+) T regulatory (Treg) cells are critical in inducing and maintaining immunological self-tolerance as well as transplant tolerance. The effect of low doses of whole-body irradiation (WBI) on CD4(+)CD25(+)Foxp3(+) Treg cells has not been determined. The proportion, phenotypes and function of CD4(+)CD25(+) Treg cells were investigated 0.5, 5 and 15 days after euthymic, thymectomized or allogeneic bone marrow transplanted C57BL/6 mice received 2-Gy γ-rays of WBI. The 2-Gy WBI significantly enhanced the ratios of CD4(+)CD25(+) Treg cells and CD4(+)CD25(+)Foxp3(+) Treg cells to CD4(+) T cells in peripheral blood, lymph nodes, spleens and thymi of mice. The CD4(+)CD25(+) Treg cells of the WBI-treated mice showed immunosuppressive activities on the immune response of CD4(+)CD25(-) T effector cells to alloantigens or mitogens as efficiently as the control mice. Furthermore, 2-Gy γ-ray WBI significantly increased the percentage of CD4(+)CD25(+)Foxp3(+) Treg cells in the periphery of either thymectomized mice or allogeneic bone marrow transplanted mice. The in vitro assay showed that ionizing irradiation induced less cell death in CD4(+)CD25(+)Foxp3(+) Treg cells than in CD4(+)CD25(-) T cells. Thus, a low dose of WBI could significantly enhance the level of functional CD4(+)CD25(+)Foxp3(+) Treg cells in the periphery of naive or immunized mice. The enhanced proportion of CD4(+)CD25(+)Foxp3(+) Treg cells in the periphery by a low dose of WBI may make hosts more susceptible to immune tolerance induction.