We have investigated the conditions for the production of a T cell growth factor (TCGF) capable of maintaining the long-term growth of normal human T cells and present in the supernatants of human-stimulated peripheral blood leukocytes. T cell lectins induced TCGF release with maximal production by using PHA (1%) and lower activity with Con A (60 µg/ml). TCGF activity was also detected in the supernatants of MLC reactions, but in a smaller amount than after lectin stimulation. Furthermore, the cells activated by the MHC differences to produce TCGF, seemed to be included into those stimulated by PHA, since supernatants from a MLC reaction carried out in the presence of PHA did not show higher TCGF activity than did a pool of supernatants from the same donors individually stimulated with PHA. TCGF release was possible in serum-free media with inhibition of its production at serum concentrations higher than 2%. The kinetics of TCGF release were highly dependent on the cellular concentration of the cultures with a peak of activity that occurred early (18 hr) in high density cultures and later (60 hr) in low density cultures. Afterward, there was a progressive fall in activity that did not seem to be due to the release of suppressive factors. We also observed significant individual differences in the production of TCGF that were closely related to the individual variability in PHA-induced DNA synthesis.