Two continuous cell lines derived from the neoplastic urothelium had been maintained in culture for more than two years. The first cell line derived from the urothelium of a fusion papillocarcinoma on the left lateral wall of the bladder was designated as TBC-1 and grown in vitro for more than 150 generations. The second cell line derived from the urothelium of a papillocarcinoma in the left renal pelvis was designated as TPC-1 and grown in vitro for more than 100 generations. Characterization studies made on both cell lines showed that the cells had a rapid doubling time, exhibited multilayering and produced tumors in sc of BALB/c. Tumor nodules that produced sc of BALB/c kept similar cellular and pathological features to those of the primary biopsy specimens under light and electron microscopes. TPC-1 cell line exhibited a three-dimensional structure of transitional epithelium on the nylon-mesh disk which was coated with a layer of rat tail collagen. Both TBC-1 and TPC-1 cell lines formed colonies in soft agar. Their forming rates were 35% and 28%, respectively. The chromosome number of TBC-1 cells ranged from 17 to 84, with a modal number of 54; whereas that of TPC-1 cells ranged from 28 to 139, with a modal number of 49. The TBC-1 cells showed mutant p53 and ras p21 protein expression and expressed weakly ABH blood group isoantigens. Analysis of lactic dehydrogenase (LDH) isozymes showed the highest levels of LDH isozyme 4 sonicated cell lysates of TBC-1 and TPC-1 respectively.