TCR repertoire analysis and functional analyses of the different types of HA-2–TCR-modified CMV-specific T clones. (A) The endogenous TCR repertoire was analyzed by RT-PCR and sequencing. To measure the expression level of HA-2 and CMV-TCR on the HA-2–TCR–transferred CMV-specific T-cell clones, the T cells were labeled with either PE-conjugated HA-2^A2 tetramers or CMV^A2 tetramers for 2 hours at 4°C. Four different types of HA-2–TCR–modified CMV^A2-specific T clones were identified. (B) Representative T-cell clones of these 4 different types of HA-2–TCR–modified CMV^A2-specific T cells, control-transduced CMV^A2-specific T cells, and the original HA-2–specific T-cell clone HA2.27 were tested at an E/T ratio of 10:1 against HLA-A2⁺ HA-2⁻ EBV-LCLs (EBV-Z), HLA-A2⁺ HA-2⁺ EBV-LCLs (EBV-RZ), EBV-Z transduced with the CMV pp65 gene (EBV-Z pp65), HLA-A2⁺ HA-2⁻ CML cells (CML-Z), and HLA-A2⁺ HA-2⁺ CML cells (CML-T) in an 18-hour cytotoxicity assay. Error bars indicate SD of triplicate culture.