A critical number of CD8⁺ central memory T cells are needed to resist tolerance. Naive B6 mice (CD45.2) received T cells from sensitized congenic B6 mice (CD45.1). (a) Titration of memory cell numbers. Naive recipients received varying doses of T cells (calculated number of memory cells transferred in brackets) from sensitized hosts (2 × 10⁷ [400,000], filled squares; 10⁷ [200,000], filled diamonds; 5 × 10⁶ [100,000], filled triangles; 2 × 10⁶ [40,000], open diamonds; 10⁶ [20,000], open triangles; 2 × 10⁷ naive cells [none], open squares). (b) CD8⁺ memory T cells represent a potent barrier to tolerance induction. Purified CD4⁺, CD8⁺, or whole T cells were isolated from sensitized mice and transferred to naive congenic mice. Mice receiving either naive selected cells (data not shown) or CD4⁺ T cells (filled diamonds) from sensitized donors developed high-level donor cell chimerism and accepted donor-type skin grafts indefinitely. The transfer of CD8⁺ T cells (filled triangles) from sensitized animals prevented tolerance induction in a similar fashion to transferred whole T cells (filled squares) (MST, 21 days). (c) CD8⁺ central memory T cells promote rejection more efficiently than “effector” memory T cells. Equal numbers of CD8⁺ (open squares), CD8⁺ CD62L^hi (filled squares), or CD8⁺ CD62L^lo (open triangles) antigen-specific memory T cells were transferred to naive congenic hosts, which were then subjected to the tolerance protocol (control naive CD8⁺ T cells, filled triangles). Both unseparated CD8⁺ memory T cells and central CD8⁺ memory T cells efficiently rejected allogeneic skin grafts in less than 3 weeks (MST, 21 and 18 days, respectively). In contrast, effector memory T cells of the traditional CD62L^lo memory phenotype were less effective at promoting rejection (MST, more than 70 days; P < 0.01).