Abstract We evaluated the relative effects of donor CD19-targeting chimeric antigen receptors (CAR)+T cells on the elimination of CD19+B cells and endogenous TCR-mediated GVHD in mouse models of allo-HSCT. We generated a panel of retroviral vectors encoding mouse CD19-specific CARs: CD19-delta, a tail-less CAR lacking the CD3ζ signaling and CD19-CAR signaling through CD28 and CD3ζ. CD19-CAR+T cells mediated specific lysis of CD19+tumors in vitro, while CD19-delta+T cells did not. We next transferred the transduced donor T cells into lethally irradiated recipients of T cell-depleted allografts and CD19+lymphoma A20-TGL (B6→BALB/c+A20-TGL). CD19-CAR+T cells mediated clearance of A20 tumor leading to significantly improved tumor free survival (p<0.01), while mediating significantly less acute GVHD, resulting in improved survival (p<0.001). Donor CD19-delta+T cells caused lethal GVHD, indicating persistent endogenous TCR mediated alloreactivity in the absence of CAR signaling. Compared to controls, fewer alloactivated CD19-CAR+T cells were found in the lymphoid organs of allo-HSCT recipients. This was accompanied by a lower expression of IFN-γ and lower frequencies of phenotypically activated T cells (p<0.05). LPAM, associated with increased gut GVHD, was also significantly lower on CD19-CAR+T cells. CAR stimulation did not inhibit TCR-mediated killing. These results provide the first preclinical evidence suggesting that CAR+ donor T cells may be safely applied in an allogeneic context.