Certain sublines of EL-4 thymoma cells can be stimulated to produce interleukin-2 (IL-2) by the mitogens phorbol-12-myristate-13-acetate and concanavalin A. EL-4 cells were cultured under conditions found to be most effective for stimulating the release of IL-2 into the culture medium. The resultant conditioned medium (CM) was used to maintain the IL-2-dependent T-cell line, CTLL-2. The response of the CTLL-2 cells to the CM was assessed by measuring [(3)H]thymidine incorporation. Since IL-2 is one of the major mediators in immune responses, it was felt that inhibition of the response of CTLL-2 cells might provide a means of screening for chemicals with the potential to affect immune responsiveness in vivo. Fourteen chemicals, including known immunoreactive compounds, were tested in the CTLL-2 responding system and ID(50) values determined. The basic cytotoxicities of the same chemicals were established by measuring growth inhibition in 3T3-L1 fibroblast-like cells. Significant differences between these two indices of toxicity for some chemicals suggested that the CTLL-2 responding system might be demonstrating effects other than basic cytotoxicity. As a result, this system is undergoing further examination to assess whether it could be developed as an assay for evaluating the potential immunomodulatory effects of chemicals.