AbstractT lymphocytes derived from peripheral blood of a patient with adenosine deaminase (ADA) deficiency were expandedin vitro. The human ADA (hADA) gene was introduced into these replicating ADA−T cells with the use of an amphotropic recombinant retrovirus carrying the hADA gene. Subsequently, infected T cells were selected on the basis of their ADA expression, by exposure to a combination of the toxic agent xylofuranosyl‐adenine and the specific ADA inhibitor 2′‐deoxycoformycin. CD4+and CD8+T cells could be infected and selected with equal efficiencies. The genetically modified T cells were shown to contain intact copies of the provirus and to express normal levels of hADA. As expected, uninfected T cells from the ADA‐deficient patient displayed an increased sensitivity to 2′‐deoxyadenosine. Following genetic modification, however, this sensitivity was restored to normal levels in both CD4+and CD8+T cells. The introduction of the hADA gene into the genome of thein vitroexpanded T cells did not alter their phenotype, proliferative capacity or cytotoxic potential. These characteristics were identical to those of T cells derived from healthy individuals. These findings are of critical importance for the clinical application of hADA gene‐tran