Cytotoxic T lymphocytes (CTL) specific for melanoma antigens can exert useful therapeutic effects in melanoma patients, and are also the primary targets of promising new immunomodulatory agents, such as humanised monoclonal antibodies to CTLA4 and PD1/PDL1. It’s now well recognised that IL2-based cell culture methods for melanoma-specific CTL have deleterious effects on T cell function, impairing their subsequent survival and efficacy in adoptive T cell therapy. We have developed new culture techniques that dramatically improve the quality of human antigen-specific CTL, in particular by preserving their ability to home to lymph nodes, and their ability to take co-stimulatory signals, while reducing their vulnerability to activation-induced cell death. The substitution of other cytokines for IL2 in this culture system allows generation of resting CTL – raising questions about whether such quiescent cells can exert rapid effector function against melanoma. Using flow cytometry-directed CTL cloning, we generated melanoma-specific CTL clones with a very early-stage differentiation phenotype, similar to T memory stem cells. Although these memory cells become quiescent in our culture system, and downregulate their effector molecules, on exposure to melanoma cells they rapidly become effector cells without needing to divide. In particular, they kill melanoma cells with only slightly delayed kinetics compared with IL2-cultured cells, due to rapid upregulation of cytotoxic molecules. They also secrete a full complement of effector cytokines on exposure to melanoma cells alone. These data clarify the re-activation kinetics of human memory CTL on encountering melanoma cells, and also suggest new approaches to T cell therapy for melanoma.