Induction of cell differentiation by twenty-one derivatives of vitamin D₃ and their binding affinity for the receptor of 1α, 25-dihydroxyvitamin D₃ an active vitamin D₃ metabolite, were examind using HL-60 human promyelocytic leukemia cells and chick intestinal cytosol, respectively. 1α, 24(R)-dihydroxy-26-chlorovitamin D₃ is found to be more active than 1α, 24(R)-dihydroxyvitamin D₃ and 1α, 25-dihydroxyvitamin D₃ in the two abilities to induce the differeniation and to bind to the receptor. the results suggest that a hydroxy group at C-1 position of vitamin D₃ and a hydroxy or oxo group at C-25 or C-24 position are essential for both abilities. In diastereoisomers of 1α, 25-dihydroxyvitamin D₃-26, 23-lactone, synthesized (23S, 25S)1α, 25-dihydroxyvitamin D₃-26, 23-lactone was more active in both abilities than the natural metabolite, (23S, 25R)1α, 25-dihydroxy-vitamin D₃-26, 23-latone, which shows anti-vitamin D action.