Urothelium is the specialized endothelium that lines much of the mammalian urinary tract. This invention relates to the field of in vitro urothelial culture. Previous methods produce tissue lacking much of the functionality of native urothelium. In this invention, stratified, differentiated mammalian urothelium is produced by passaging urothelial cells through a nutrient medium containing serum, and redispersed, before going on in a like medium to form said urothelium.