The identity of cells that mediate positive selection of CD8+T cells was investigated in two T cell receptor (TCR) transgenic systems. Irradiated β2-microglobulin mutant mice or mice with mutations in both the Kband Dbgenes were repopulated with fetal liver cells from class I+TCR transgenic mice. In the case of the 2C TCR, mature transgene-expressing CD8+T cells appeared in the thymuses of the chimeras and in larger numbers in the peripheral lymphoid organs. These CD8+T cells were functional, exhibited a naive, resting phenotype, and were mostly thymus-dependent. Their development depended on donor cell class I expression. These results establish that thymic hematopoietic cells can direct positive selection of CD8+T cells expressing a conventional TCR. In contrast, no significant development of HY (male antigen)–TCR+CD8+T cells was observed in class I+into class I-deficient chimeras. These data suggest that successful positive selection directed by hematopoietic cells depends on specific properties of the TCR or its thymic ligands. The possibility that hematopoietic cell-induced, positive selection occurs only with TCRs that exhibit relatively high avidity interactions with selecting ligands in the thymus is discussed.