Splenocytes of nu/nu mice treated with serum thymic factor (FTS) for 3 or more days followed by stimulation with either phorbol myristate acetate (PMA) or concanavalin A (Con A) produced interleukin 2 (IL 2) as determined in two indicator systems, namely, support of growth of IL 2-dependent T cells and promotion of Con A-initiated mitogenesis of thymocytes. However, neither mitogens nor FTS alone could induce nude mice cells to product IL 2. Supernatants derived from the tumor cell line WEHI-3 (WEHI-3 conditioned media) induced and supported continuous growth in culture of Thy-1.2+, Lyt-1+2- lymphocytes from athymic nude mice capable of producing IL 2 after their stimulation by either PMA or Con A. The growth of these IL 2-producer cell lines strickly depends on the presence of WEHI-3 conditioned media, as in the absence of it they die 24-48 h later. In addition, WEHI-3 conditioned media have been supporting the growth of IL 2-producer cell lines derived from nude mice for 3 1/2 months. The helper factor contained in WEHI-3 conditioned media responsible for the above biological activity has an apparent mol. wt. of approximately 40 000 as determined by Sephadex G-100 chromatography and lacks IL 1 and IL 2 activities, but efficiently supports the growth of IL 2-producer cells derived from nude mice and the peripheral blood of normal human volunteers. These results indicate that the helper factor in WEHI-3 conditioned media which enables the generation and continuous proliferation in culture of IL 2-producer cells in nude mice is distinct from interleukin 1, IL 2 and FTS (mol. wt. 864). Finally, the possibly functional relationship of FTS and the helper factor produced by WEHI-3 cells is discussed.