['Genetically modified T-lymphocytes are potential therapeutic agents for the treatment of various disorders. Successful retroviral infection of primary murine T-lymphocytes is a prerequisite to the study of adoptive cell therapies in a small animal model. The definition of factors controlling retroviral infection of T-lymphocytes would also be useful to better understand retroviral diseases. However, retroviral-mediated gene transfer into murine primary T-cells has remained elusive.', 'In order to define the requirements for stable and efficient gene transfer in primary murine T-lymphocytes, we investigated factors capable of affecting retroviral infection. These include activation conditions (using mitogens or monoclonal antibodies), culture conditions (including media composition and cytokine addition), timing of viral exposure, retroviral receptor selection (ecotropic, amphotropic, or vesicular stomatitis virus G (VSV-G) glycoprotein receptor), and viral titer.', 'We show that efficient gene transfer can be achieved in murine T-lymphocytes, provided that a number of favorable conditions are met, in particular optimized T-cell activation conditions, optimal timing of infection, adequate interleukin-2 concentration and T-cell density, and a high viral titer. On a particulate basis, we find that ecotropic particles are more effective than amphotropic or VSV-G-pseudotyped particles, and recommend the use of a specifically selected retroviral packaging cell line. CD4+ T-cells are equally or more infectible than CD8+ lymphocytes, depending on the activation conditions. The Th1 and Th2 subsets are comparably susceptible to retroviral infection, in contrast to what has been reported in some instances of human T-cell infection by human immunodeficiency virus (HIV).'] Do activation conditions determine susceptibility of murine primary T-lymphocytes to retroviral infection?