Intravesical immunotherapy with bacillus Calmette-Guérin (BCG) against recurrences of superficial bladder cancer and carcinoma in situ is a highly effective regimen in urology. Despite intensive efforts to clarify the immunological mechanisms of the most successful immunotherapy known today, the cellular mechanism of its antitumor activity remains unknown. In our approach to elucidate the way of action of intravesical BCG, we applied an in vitro adhesion assay to investigate the interaction of radiolabeled BCG with urothelial bladder-tumor cells. We demonstrated a BCG dose-dependent binding to bladder-tumor cell lines derived from tumors of different gradings. The binding of BCG is apparently specific, since competition experiments showed an inhibition by nonradioactive BCG but not by Escherichia coli. We also found that there was no difference between the binding of living or heat-killed mycobacteria. Control experiments showed only a low affinity of BCG for fibroblasts, smooth-muscle cells, and endothelial cells in comparison with the tumor cells. Furthermore, we investigated the role of fibronectin as an adhesion molecule that is also present in the bladder wall. We demonstrated that BCG was capable of binding to fibronectin-coated surfaces in a dose-dependent manner. However, competitive binding assays failed to reveal an inhibition of the binding of BCG to bladder-tumor cells by anti-fibronectin. Furthermore, binding was not influenced by soluble fibronectin. These data suggest that the in vitro attachment of BCG to bladder-tumor cells appears not to be mediated by fibronectin. In electron microscope studies an adhesion of BCG to bladder-tumor cells was observed after an incubation period of ony 30 min.(ABSTRACT TRUNCATED AT 250 WORDS)