Thy-1+cell depletion with anti-Thy-1.2 mAb and complement markedly reduced the capacity of C57BL/6J, H-2bbone marrow to establish mixed lymphoid chimerism and induce tolerance to C57BL/6J skin grafts across an entire MHC disparity in BALB/c, H-2dhosts conditioned with sublethal, fractionated 7.5 Gy total-body irradiation. In this model tolerance can be transferred to secondary irradiated BALB/c hosts only by cells of C57BL/6J donor, not host, genotype isolated from the spleens of tolerant hosts. Thy-1+cell depletion abolished the capacity of C57BL/6J donor cells from tolerant BALB/c host spleens to transfer tolerance. The capacity of semiallogeneic BALB/c×C57BL/6J F1, H-2d/bdonor BM and spleen cells to induce chimerism and tolerance to C57BL/6J skin grafts in BALB/c parental hosts was also reduced by Thy-1+cell depletion. Thus the requirement for donor Thy-1+cells cannot be explained simply on the basis of alloaggression. It is unlikely that the requisite Thy-1+cells are nonspecific suppressor cells: Thy-1+cell depletion had no effect on the slight but significant prolongation of third-party C3H/HeJ, H-2kskin grafts in irradiated BALB/c hosts injected with allogeneic C57BL/6J or semiallogeneic BALB/c×C57BL/6J F1BM compared to irradiated controls injected with medium only. Furthermore, injections of semiallogeneic F1spleen cells had no significant effect on the survival of the third-party grafts, although these cells were fully capable of inducing tolerance, and their capacity to induce tolerance was significantly reduced by Thy-1+cell depletion. The requirement for a specific population of lymphoid cells, i.e. Thy-1+, remains unexplained but suggests that donor cells might play a role in the induction or maintenance of tolerance in this model other than merely providing a circulating source of donor antigens.