Tumour-infiltrating lymphocytes (TIL) are often observed in human tumours and their presence has been correlated with a better prognosis. It has been suggested that TIL are enriched for tumour-specific cytotoxic cells, and TIL activated and expanded in vitro by interleukin-2 (IL-2) are currently used in the therapy of human cancer. We have studied the T cell repertoire in IL-2-expanded TIL cells from patients with ovarian and renal carcinoma using T-cell-receptor-V-beta-specific monoclonal antibodies and a polymerase-chain-reaction-based Southern blot technique for analysis of J-beta usage. In TIL lines derived from three of nine patients with ovarian carcinomas and from two of eight patients with renal carcinomas, selective usage of the V-beta 6 or V-beta 5 T-cell receptor gene products was found. The majority of the cells were CD4+, with up to 40% of the T cells utilizing the same V-beta gene. T-cell lines derived from peripheral blood lymphocytes from patients or healthy donors contained normal levels of V-beta subsets. Only moderate levels of V-beta 6+ T cells were detected from freshly isolated TIL and the increase of this subpopulation appeared as a result of in vitro culture. The level of clonal restriction, as measured by the usage of J-beta gene segments within the V-beta 5 or V-beta 6 families, was analysed using a recently developed technique based on the polymerase chain reaction. Evidence for restricted J-beta usage was detected only in TIL expanded in vitro, while this was not the case in freshly isolated tumour-derived lymphocytes or T cell lines obtained from peripheral blood lymphocytes. The presence of a population with biased T cell receptor expression in cells derived from tumour tissue could be explained by their activation in vivo as a result of contact with tumour antigens and should be taken into consideration when discussing the therapeutic efficiency of IL-2-expanded TIL.