We have previously described a limiting dilution assay (LDA) for estimating the frequency, in the spleens of unprimed mice, of alloantigen-responsive cells that, together with their immediate clonal progeny, can produce interleukin 2 (IL 2) in short-term culture. In this paper, we provide further evidence that the limiting cell in these cultures is, in fact, the immediate precursor of the IL 2-producing cell rather than merely a participant in a multicellular cascade that ultimately leads to production of this lymphokine. We also demonstrate the usefulness of the LDA method for estimating the frequency of the limiting cell for production of IL 3. By analyzing the supernatants of these short-term microcultures for both IL 2 and IL 3, we show that essentially all wells that produce IL 3 also produce IL 2. Furthermore, the amount of IL 2 produced in any individual well correlates strongly with the IL 3 production in the same well. Our data suggest that both these lymphokines are produced by a single class of T cell, and further that the regulatory events that control IL 2 generation in allostimulated helper cells act to control IL 3 levels in parallel.

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