Oral and epidermal rat keratinocytes when cultured on a matrix of type I collagen fibrils at the interface between the gaseous and liquid phases of a culture form a highly ordered stratified squamous epithelium. Autoradiographic studies of cells labeled by tritiated thymidine indicate that the keratinocytes are capable of autoregulating cell division. Early confluent cultures exhibit 51 % of basal cells labeled, a percentage that decreases to 18% when a fully differentiated stratified squamous epithelium is formed. Such a decrease in labeling occurs in cultures where the mitotically active basal cells have unimpeded access to culture medium supplied from below and when no cell type other than the keratinocyte is present in the culture. Additionally, the transit of keratinocytes from the basal cell layer through other viable cell strata to the layer of terminally differentiated cells can be followed by tracking cells labeled with tritiated thymidine. In cultures of oral keratinocytes, cells move from the basal cell layer to the cornified layer at a maximum rate of 7 days, while virtually all labeled cells (91 %) are localized in the terminally differentiated cell layer 14 days following labeling. Keratinocyte cultures grown in culture at an air-liquid interface exhibit tissue organization that closely resembles the native, parent tissue. Such cultures can be useful in studying the effects of pharmacologic mediators of cell division and cell transit.