CMV-specific CTLs have an effector memory phenotype and a Tc1 cytokine secretion profile. (A) CMV-specific CTLs have an effector memory phenotype. CTLs were characterized by staining for (left to right) CD8, CD62L, CD45RA, CCR7, and CXCR4 (all FITC-labeled antibodies, x-axis), against the P495 tetramer (PE-labeled, y-axis). Top to bottom: CTLs from peripheral blood prior to any stimulation (PB), CTLs activated on ER^- PBMCs, BLCLs, or AAPC^A2pp65 (AAPC). The immunophenotyping of CTLs from peripheral blood before any stimulation or after in vitro expansion using autologous presenting cells or AAPCs (either AAPC^A2P495 or AAPC^A2pp65) was identical and consistent with an effector memory phenotype for the 3 donors. (B) The P495-specific CTLs have a Tc1 cytokine secretion profile. Intracellular cytokine staining was performed after a 10 to 14-day coculture with AAPC^A2pp65. At the end of the coculture, 48% of the cells stain positive for P495 tetramer (left panel, y-axis). After a brief restimulation on a monolayer of AAPCs in the presence of brefeldin A, a comparable percentage of CD8-positive cells stained strongly positive for IFN-γ and TNF-α. Similar results were obtained with 3 APCs and for the 3 donors.