In vitro incubation of 24-epi-25-hydroxyvitamin D2 with chicken kidney homogenate produced several compounds, one of which had an affinity equal to that of 1,25-dihydroxyvitamin D2 for the chick intestinal receptor. The affinity of 24-epi-1,25-dihydroxyvitamin D2 for the same receptor was found to be half that of 1,25-dihydroxyvitamin D2. The unknown compound was produced only when homogenate was prepared from pooled kidneys taken from both vitamin D deficient and replete chickens. The compound has been tentatively identified as 1,25-dihydroxy-22-dehydro-26-homovitamin D3 by ultraviolet absorption spectrophotometry and mass spectrometry. Chemical synthesis of 1,25-dihydroxy-22-dehydro-26-homovitamin D3 provided additional evidence for the structure. Administration of this 26-homologue of 1,25-dihydroxyvitamin D3 at the dose level of 650 pmol/rat stimulated bone calcium mobilization in the hypocalcemic rat equal to that of 1,25-dihydroxyvitamin D3. Thus, this paper demonstrates unique methyl migration on the side chain of 24-epi-1,25-dihydroxyvitamin D3 to form a more biologically potent analogue.