The CD94/NKG2A heterodimer is an inhibitory receptor expressed on a subset of mouse NK cells. CD94/NKG2A recognizes the non-classical MHC class I (class Ib) molecule Qa-1^b and inhibits NK cytotoxicity. Qa-1^b presents a peptide derived from the leader sequence of classical MHC class I molecules. Here, we examined the role of CD94/NKG2A in T cell-mediated cytotoxicity. Soluble tetrameric Qa-1^b bound to almost all CD8⁺, but not CD4⁺, T cells. This binding seems to be mediated by CD8, because COS cells transfected with CD8 also bound Qa-1^b tetramer. Therefore, the expression of CD94/NKG2 in T cells was further examined by single-cell RT-PCR. Most murine CD8⁺ T cells constitutively expressed CD94 and NKG2A transcripts, whereas they were not detected in CD4⁺ T cells. Co-expression of Qa-1^b and D^k on target cells significantly inhibited cytotoxicity of D^k-specific cytotoxic T lymphocytes generated by mixed lymphocyte reaction, indicating that Qa-1^b on antigen-presenting cells interacts with CD94/NKG2A on CD8 T cells and regulates classical MHC class I-restricted cytotoxic T cells. These results suggest a significant role of CD94/NKG2A as an inhibitory receptor on CD8⁺ T cells.