TcR gamma/delta+ lymphocytes represent a small subset homogeneously composed of cytolytic T cells displaying unique motility and homing properties. Since the lytic machinery of these cells can be efficiently triggered by monoclonal antibodies (MAbs) directed to the TcR gamma/delta, such MAbs were used for the construction of bispecific MAbs in conjunction with an MAb specific for ovarian carcinoma cells. Hybrid hybridomas were obtained by fusing the Mov19 MAb (IgG2a)-producing hybridoma with either GI (IgG2a) or A13 (IgG1) hybridomas, secreting MAbs specific for 2 peripheral blood subsets of TcR gamma/delta+ lymphocytes. Hybrid hybridomas producing bispecific MAbs were screened according to their ability to induce ovarian carcinoma (IGROVI) target cell lysis by GI+ or A13+ T cell clones, respectively. The GI-derived GM33.9 bispecific MAb induced selective lysis of Mov19+ ovarian carcinoma target cells only by GI+ clones, whereas the A13-derived AM18.4 MAb was effective only in combination with A13+ clones. Neither the anti-TcR gamma/delta nor the Mov19 parental MAbs (used alone or in combination) induced target-cell lysis. The hybrid nature (IgG1/IgG2a) of the AM18.4 bispecific MAb was indicated by 2-color immunofluorescence experiments. Thus, both ovarian carcinoma and A13+ effector cells were double stained by AM18.4 bispecific MAb followed by PE-conjugated anti-IgG1 and FITC-conjugated anti-IgG2a second reagents. Polyclonal TcR gamma/delta+ cells were obtained by direct stimulation of peripheral blood mononuclear cells with Sepharose bead-conjugated anti-TcR gamma/delta MAbs and IL-2. The lines so obtained contained more than 90% of TcR gamma/delta+ cells after 4 weeks of culture, with an increase in TcR gamma/delta+ cell numbers ranging from 200 to 1,000-fold. These TcR gamma/delta+ cell lines efficiently lysed ovarian carcinoma target cells in the presence of bispecific MAb and may therefore represent a suitable source of effector cells for induction of ovarian carcinoma cell lysis.