Organotypic culture is an invaluable technique that allows researchers with the tool to analyze a tissue development in an isolated and well-defined environment. This technique also permits one to study the roles of different signaling systems/signaling molecules and to take advantage of the modern real-time imaging techniques, including confocal microscopy. With great success, our lab has used organotypic culture of the urogenital tract (UGT) to study growth and extension of the mesonephric (Wolffian) duct and its cloaca connection, ureter maturation, and bladder urothelium development (Batourina et al. Nat Genet 32:109, 2002; Batourina et al. Nat Genet 37:1082, 2005; Mendelsohn Organogenesis 5:306, 2009).