An ultrastructural comparison of the tracheal epithelium of Golden Syrian hamsters revealed that some variability between in vivo and organ culture specimens occurs although ultrastructural integrity is maintained to a remarkable degree. The most striking difference between in vivo and in vitro specimens proved to be a proliferation of tight junction type sealing strands in the membranes of cultured tracheal epithelial cells. The strands run randomly and in whorled patterns over the membrane, possibly sealing areas of damaged and deteriorated membranes in vitro. Other ultrastructural features of the membranes appeared similar in vivo and in vitro. The cellular organelles of organ culture cells maintained morphological identity with their in vivo counterparts and cellular ultrastructure appeared to remain generally intact up to 48 hr.