A calcium binding protein has been demonstrated in human duodenal mucosa which we believe to be human vitamin D dependent CaBP. Sephadex gel filtration demonstrated that the duodenal mucosa of both the human and pig contained a calcium binding protein with a similar molecular weight to the reported vitamin D dependent rat CaBP (M.W. 12 000–13 000) but dissimilar to chick CaBP (M.W. 24 000–28 000). In addition the rat, pig, and human mucosa contained a high molecular weight calcium binding protein which is probably not vitamin D dependent.A relatively simple procedure, utilizing the tagging of CaBP by 47Ca, has been developed to partially purify normal pig CaBP by Sephadex gel filtration. Further fractionation of the 12 000–13 000 M.W. area, using disc electrophoretic procedures, separated two calcium binding proteins which had similar electrophoretic mobilities and calcium binding formation constants (3.5 and 5.5 × 106, respectively), indicating that both are forms of CaBP but that either one or both have been altered during the procedures. The electrophoretic mobilities of both of these proteins are relatively low in the presence of calcium ions but much greater when calcium ions are removed by chelation with EDTA. This finding should facilitate both the identification and purification of mammalian CaBP.