The tight junction defines epithelial organization. Structurally, the tight junction is comprised of transmembrane and membrane-associated proteins that are thought to assemble into stable complexes to determine function. In this study, we measure tight junction protein dynamics in live confluent Madin–Darby canine kidney monolayers using fluorescence recovery after photobleaching and related methods. Mathematical modeling shows that the majority of claudin-1 (76 ± 5%) is stably localized at the tight junction. In contrast, the majority of occludin (71 ± 3%) diffuses rapidly within the tight junction with a diffusion constant of 0.011 μm²s⁻¹. Zonula occludens-1 molecules are also highly dynamic in this region, but, rather than diffusing within the plane of the membrane, 69 ± 5% exchange between membrane and intracellular pools in an energy-dependent manner. These data demonstrate that the tight junction undergoes constant remodeling and suggest that this dynamic behavior may contribute to tight junction assembly and regulation.