Immunofluorescent staining of septins in primary cilia

Primary cilia are cellular antennae that receive and transduce extracellular cues. These microtubule-rich structures are comprised of at least three distinct ciliary compartments: basal bodies, transition zone, and axoneme. Septins have been implicated in cilia function at the transition zone, but accumulating evidence suggests that they localize predominantly within the axoneme. Here, we describe three fixation conditions that preserve the substructure of primary cilia and demonstrate known ciliary proteins that localize to these distinct ciliary substructures. Finally, we show immunostaining and live microscopy methods to detect septins within the axoneme.