Specificity and effector function ofCD4⁺CD8⁺ cells in healthy blood donors. (A-C) PBMCs of healthy blood donors (A) or HLA-A2-positive healthy blood donors (B-C) were ex vivo stimulated with the indicated virus-infected cell lysates (A) or HLA-A2-restricted peptides (B) or proteins (C) and analyzed by intracellular cytokine staining or cytokine secretion assay. The percentage of cytokine-producing cells is indicated in each quadrant. Dot plots are representative for 10 healthy blood donors tested. DP indicates CD4⁺CD8⁺ T cells. (D) Cumulative frequency of all CMV, EBV, IV, and VV peptide-specific, cytokine-producing CD4⁺, CD8⁺, and CD4⁺CD8⁺ cells in the PBMC population of each healthy blood donor (HD). Results are representative for 10 healthy donors. Similar results were obtained with virus-infected cell lysates and protein stimulation. (E) Mean granzyme A and perforin expression in the indicated cell populations of 5 healthy blood donors. Error bars indicate SD.