The sensory epithelia of the mammalian inner ear consist of a highly precise pattern of sensory hair cells and supporting cells. The mechanisms regulating this patterning are only beginning to be determined. The present study describes a method for culturing dissociated embryonic inner ear cells and the resulting patterning that occurs in these cultures. The results indicate that developing inner ear cells aggregate into precise patterns on a two-dimensional substrate, suggesting that intrinsic patterning mechanisms remain active in vitro. Using antibodies and scanning electron microscopy to detect hair cells and nonsensory cells, it was determined that only a subset of aggregates contained sensory hair cells. The hair cells were organized into specific patterns and surrounded by supporting cells, similar to the in vivo pattern. Additionally, hair cells increased their immunoreactivity and number of stereocilia over time, suggesting that hair cells continue to mature in vitro. Thus, the study reveals that the cells of the developing inner ear provide the necessary signals that direct sensory hair cells and supporting cells to reassociate into very precise patterns in vitro and that these patterns are reminiscent of the patterning that occurs in vivo.