A preparation of isolated intestinal cells has been developed which permits the incubation of these cells in culture for 90 min. with preservation of viability and responsiveness to vitamin D analogues added in vitro. Characterization of phosphate transport suggested the presence of two transport processes which differed in their affinity for phosphate, calcium stimulation, and pH dependence. Both processes were facilitated by sodium transport. This cell preparation promises to be a valuable experimental model for the elucidation of the action of vitamin D on phosphate transport.