Naive CD4 T cells acquire the potential to produce IFN-γ and IL-4 by culture in the presence of their cognate Ag, APC, and appropriate cytokines. In this study, we show that commitment to IFN-γ production on the part of rigorously purified naive CD4 T cells can occur without cell division. Indeed, even entry into S phase is not essential. Moreover, both CD4 and CD4/CD8 thymocytes from TCR-transgenic mice (5CC7 mice) on a Rag2−/− background can acquire IFN-γ-producing capacity when stimulated by peptide, APC, and IL-12. These cells can do so without dividing and some acquire IFN-γ-producing activity without entry into S phase. Not only is cell division not required for acquisition of cytokine-producing potential, cell populations that have undergone the same numbers of divisions can have quite different proportions of IFN-γ-or IL-4-producing cells, depending on the duration of priming or, in the case of IL-4, on the concentration of peptide. Thus, cell division is not a clock for the expression of these cytokines. Factors associated with priming conditions including strength of stimulation, duration of priming, and number of divisions each play a role.