A tracheal cell culture system which can be used for detection of hazardous environmental agents is described. The culture system makes use of primary tracheal cells that are isolated from rats by protease digestion of the tracheal epithelium. The epithelial cells are plated on a film of collagen or onto a layer of gamma-ray- or mitomicin-C-inactivated mouse 3T3 cells. One day after the rat tracheal cells are plated they are exposed to toxicants, and the effect on colony formation is tabulated after 1 week. By altering the culture conditions it has become possible to construct an assay for carcinogen-induced transformation of rat tracheal cells, which takes advantage of the observation that normal rat tracheal cells have a finite lifetime in culture. Rat tracheal cell transformants are visualized as discrete colonies of proliferating epithelial cells that arise and survive after the normal population has died out. Their number is proportional to concentration of carcinogen to which the cells were initially exposed. Agents that have been shown to induce tracheal cell transformation in culture include polycyclic hydrocarbons, metal salts, nitroso compounds, and cigarette smoke condensate.