How terminally differentiating cells are selectively expelled from the basal layer of epidermis has been a source of interest and speculation for many years. The problem can now be studied in culture, using involucrin synthesis as an early marker of terminal differentiation in human keratinocytes. When keratinocytes are forced to grow as a monolayer by reducing the calcium ion concentration of the culture medium, they still begin to synthesize involucrin. Raising the level of calcium ions induces stratification, and cells that are synthesizing involucrin are selectively expelled from the basal layer. I have found that during calcium-induced stratification no new proteins or glycoproteins are synthesized, and the rate of cell division does not change. Movement of involucrin-positive cells out of the basal layer was found to be unaffected by cycloheximide, tunicamycin, or cytosine arabinoside. These results suggest that keratinocytes growing as a monolayer already have the necessary properties to determine their position when stratification is induced. Addition of calcium simply allows formation of desmosomes and other intimate cell contacts required for stratification. The properties of involucrin-positive cells that determine their suprabasal position include a reduced affinity for the culture substrate and preferential adhesion to other cells at the same stage of terminal differentiation. The molecular basis of these adhesive changes is discussed.