In allogeneic bone marrow transplantation (BMT) donor T cells are primarily responsible for antihost activity, resulting in graft-versus-host disease (GVHD), and for antileukemia activity, resulting in the graft-versus-leukemia (GVL) effect. The relative contributions of the Fas ligand (FasL) and perforin cytotoxic pathways in GVHD and GVL activity were studied by using FasL-defective or perforin-deficient donor T cells in murine parent → F1 models for allogeneic bone marrow transplantation. It was found that FasL-defective B6.gld donor T cells display diminished GVHD activity but have intact GVL activity. In contrast, perforin-deficient B6.pfp^−/− donor T cells have intact GVHD activity but display diminished GVL activity. Splenic T cells from recipients of B6.gld or B6.pfp^−/− T cells had identical proliferative and cytokine responses to host antigens; however, splenic T cells from recipients of B6.pfp^−/− T cells had no cytolytic activity against leukemia cells in a cytotoxicity assay. In experiments with selected CD4⁺ or CD8⁺ donor T cells, the FasL pathway was important for GVHD activity by both CD4⁺ and CD8⁺ T cells, whereas the perforin pathway was required for CD8-mediated GVL activity. These data demonstrate in a murine model for allogeneic bone marrow transplantation that donor T cells mediate GVHD activity primarily through the FasL effector pathway and GVL activity through the perforin pathway. This suggests that donor T cells make differential use of cytolytic pathways and that the specific blockade of one cytotoxic pathway may be used to prevent GVHD without interfering with GVL activity.