In this study, immunotoxins containing monoclonal anti-human T-cell leukemia antibodies are shown to be capable of specific killing of human T-leukemia cells in vitro. These immunotoxins were prepared by conjugating ricin A chain (RIA) with our recently generated murine monoclonal antibodies, SN1 and SN2, the latter of which was obtained from a hybridoma clone N6/D11 described previously by Negoro and Seon (Cancer Res., 42: 4259-4262, 1982), directed to two unique human T-cell leukemia antigens. We have shown previously that these monoclonal antibodies do not react with non-T-leukemia cells nor with various normal cells including normal T-cells, thymocytes, and bone marrow cells (Proc. Natl. Acad. Sci. U. S. A., 80: 845, 1983; Cancer Res., 42: 4259, 1982). Control conjugate was also prepared by conjugating RIA with a murine monoclonal immunoglobulin G (IgG), the isotype of which is the same as that of SN1 and SN2, i.e., IgG1-kappa. In initial experiments, the cytotoxic activity of an SN1 IgG:RIA conjugate preparation and the control IgG conjugate preparation against leukemic T-cell lines and normal B-cell lines was tested by two different test procedures, i.e., by measuring direct killing of the cells and by measuring inhibitory activity against protein synthesis in the cells. In each test, the SN1 conjugate showed specific cytotoxic activity against T-leukemia cells, whereas the control conjugate was not cytotoxic against either T-leukemia cells or normal B-cells. Nearly complete killing of T-leukemia cells and inhibition of protein synthesis in T-leukemia cells were observed at the concentrations of 10(-8) to 10(-7) M of the SN1 IgG:RIA conjugate. In subsequent experiments, another preparation of SN1 IgG:RIA conjugate and an SN2 IgG:RIA conjugate preparation were tested individually and together for their inhibitory activity against protein synthesis in T-leukemia cells and control cells. With T-leukemia cells, specific inhibition was observed for both SN1 IgG:RIA and SN2 IgG:RIA. The combined use of these conjugates did not display a synergistic effect. Nevertheless, the combined use of different immunotoxins directed to different antigen molecules will be important in clinical use, since uncultured tumor cells derived freshly from patients, in general, display heterogeneity with respect to the expression of tumor-associated antigens. These immunotoxins may be useful for the in vitro eradication of tumor cells in the bone marrow taken from patients with T-cell leukemia.(ABSTRACT TRUNCATED AT 400 WORDS)