In order to explore the influence of purified donor regulatory T cells (Treg) infused after allogeneic bone marrow transplantation (allo-BMT) on GVHD and GVL effect in mice, an EL4 leukemia allo-BMT model of BALB/c-->C57BL/6 mice was established. The CD4(+)CD25(+)T cells were purified by positive selection using MACS. The recipients were injected with CD4(+)CD25(+)T cells or CD4(+)CD25(-)T cells within 4 hours respectively along with allo-BMT. Survival time, clinical GVHD score or histopathological features were observed after allo-BMT. The results showed that the mean survival time in leukemia control group was (17.9 +/- 0.7) days and all mice died of leukemia. The mean survival times in transplantation control group and CD4(+)CD25(-)T group were (23.2 +/- 1.6) and (22.3 +/- 1.9) days. Histopathological analysis in several target organs (skin, liver and small intestine) confirmed the presence of severe GVHD. The mean survival time in Treg group was (47.3 +/- 6.5) days. 70% of recipients were alive until day 60 without features of GVHD or tumor progression. Clinical GVHD scores in Treg group significantly decreased as compared to transplantation control group and CD4(+)CD25(-)T group (p < 0.05). It is concluded that allo-BMT combined with injection of donor CD4(+)CD25(+)Treg cells can efficiently prevent recipients from lethal GVHD with preserving GVL effect during allo-BMT.