Surface expression of pWT126-specific TCR on transduced primary human T lymphocytes. Human T lymphocytes retrovirally transduced with genes encoding the pWT126-specific TCR (Vα1.5/Vβ2.1) chains or mock transduced were triple stained with anti–human CD3-FITC or CD8-APC antibodies and anti–human Vβ2-PE antibody or HLA-A2/pWT126 tetramers. All panels display gated CD3⁺ T cells. Panel A shows staining of mock-transduced T cells and panel B the pWT126 TCR-transduced T cells 2 days after transduction. After 2 rounds of antigen-specific stimulation, the percentage of T cells binding Vβ2 and CD8 antibodies increased (C). After additional in vitro expansion, tetramer staining revealed that T cells specifically bound the HLA-A2/pWT126 tetramer (D) but not an A2/EBV (Epstein-Barr virus) control tetramer (E). Similarly, T cells of a patient with CML (F) and AML (G) showed a high percentage of Vβ2⁺/CD8⁺ T cells after pWT126 TCR transduction and 2 rounds of antigen-specific stimulation (control TCR-transduced T cells contained 2% and 3% Vβ2⁺/CD8⁺ T cells, respectively). Tetramer staining of the transduced T cells of the patient with CML (H) and AML (G) showed that they bound the pWT126 tetramer but not the EBV tetramer (J shows the CML patient cells; similar lack of staining was seen with AML patient cells). Numbers indicate the percentage of cells in the quadrants.