Recipients that received transplants of marrow and CD4⁺CD25⁺ T cells demonstrate proliferative responses to polyclonal stimulators but do not proliferate or generate CTLs to donor or host antigens. Three to 4 months after BMT, chimeric BALB/c recipients of B6 BM and CD4⁺CD25⁺ T cells (n = 5-7), BALB/c recipients of B6 BM only who rejected the donor marrow (n = 3-4), and B6 mice that did not receive transplants (n = 4-6) were killed, and spleen and lymph node cells were isolated to assess immune responses. Polyclonal proliferation was measured in response to anti-CD3 mAb (▪) and LPS (▦) (A). Unstimulated cells are represented by □. Generation of proliferative (B) and CTL (C) responses was measured against donor (B6, left) and host (BALB/c, right) background stimulators. In B, light gray bars indicate C57BL/6; dark gray bars, BALB/c. In C, • indicates BM only; ♦, BM + CD4⁺CD25⁺; ▪, nontransplanted B6. Each experiment was performed 2 to 3 times. Error bars are mean cpm and SEM of triplicate wells (A,B).