Background With potent suppressive effect on responder T cells, CD 4 +CD 25 + regulatory T (Treg) cells have become the focus of attention only recently and they may play an important role in transplantation tolerance However, the mechanism of action is not clear This study was designed to assess the possibility of using CD 4 +CD 25 + Treg cells to induce transplantation tolerance and to investigate their mechanism of action KH*2/5DMethods CD 4 +CD 25 + Treg cells were isolated using magnetic cell separation techniques Mixed lymphocyte reactions were used to assess the ability of Treg cells to suppress effector T cells Before skin transplantation, various numbers of CD 4 +CD 25 +Treg cells, which have been induced using complex skin antigens from the donor, were injected into the host mice either intraperitoneally (0 5×10 5, 1×10 5, 2×10 5, 3×10 5, 4×10 5, or 5×10 5) or by injection through the tail vein (5×10 3, 1×10 4, 2×10 4, 5×10 4, 1×10 5, 2×10 5) Skin grafts from two different donor types were used to assess whether the induced Treg cells were antigen-specific The survival time of the allografts were observed Single photon emission computed tomography was also used to determine the distribution of Treg cells before and after transplantation Results Treg cells have suppressive effect on mixed lymphocyte reactions Grafts survived longer in mice receiving CD 4 +CD 25 + Treg cell injections than in control mice There was a significant difference between groups receiving intraperitoneal injection of either 2×10 5 or 3×10 5 CD 4 +CD 25 +Treg cells and the control group ( P <0 05, respectively) Better results were achieved when Treg cells were injected via the tail vein than when injected intraperitoneally The transplantation tolerance induced by CD 4 +CD 25 + Treg cells was donor-specific Analysis of the localization of Treg cells revealed that Treg cells mainly migrated from the liver to the allografts and the spleen KH*2/5DConclusions CD 4 +CD 25 +Treg cells can induce donor-specific transplantation tolerance Cell-to-cell contact may be the primary mechanism by which Treg cells act on effector T cells Background With potent suppressive effect on responder T cells, CD 4 + CD 25 + regulatory T (Treg) cells have become the focus of attention only recently and they may play an important role in implantation tolerance However, the mechanism of action is not clear This study was designed to assess the possibility of using CD 4 + CD 25 + Treg cells to induce transplantation tolerance and to investigate their mechanism of action KH * 2 / 5DMethods CD 4 + CD 25 + Treg cells were isolated using magnetic cell separation techniques Mixed lymphocyte reactions were used to assess the ability of Treg cells to suppress effector T cells before skin transplantation, various numbers of CD 4 + CD 25 + Treg cells, which have been induced using complex skin antigens from the donor, were injected into the host mice either intraperitoneally (0 5 × 10 5, 1 × 10 5, 2 × 10 5, 3 × 10 5, 4 × 10 5, or 5 × 10 5) or by injection through the tail vein (5 × 10 3, 1 × 10 5 4, 2 × 10 4, 5 × 10 4, 1 × 10 5, 2 × 10 5) Skin grafts from two different donor types were used to assess whether the induced Treg cells were antigen-specific The survival time of the allografts were observed Single photon emission computed tomography was also used to determine the distribution of Treg cells before and after transplantation Treg cells have suppressive effect on mixed lymphocyte reactions Grafts survived longer in mice receiving CD 4 + CD 25 + Treg cell injections than in control mice There was a significant difference between groups The results of intraperitoneal injection of either 2 × 10 5 or 3 × 10 5 CD 4 + CD 25 + Treg cells and the control group (P <0 05, respectively) Better results were achieved when Treg cells were injected via the tail vein than then injected intraperitoneally The transplantation tolerance induced by CD 4 + CD 25 + Treg cells was donor-specific Analysis of the localization of Treg cells that that Treg cells mainlymigrated from the liver to the allografts and the spleen KH * 2 / 5DConclusions CD 4 + CD 25 + Treg cells can induce donor-specific transplantation tolerance Cell-to-cell contact may be the primary mechanism by which Treg cells act on effector T cells