title: Two direct (nonchromatographic) assays for 25-hydroxyvitamin D.
abstract: We describe direct, nonchromatographic assays for 25-hydroxyvitamin D3 in which we use either rat vitamin D-binding protein or rabbit antibodies to 25-hydroxyvitamin D3-3-hemisuccinate-bovine serum albumin as binding protein. Nonspecific interferences in serum could be eliminated by an appropriate extraction method or in obtaining data for the calibration curve, by using vitamin D-free human serum. The latter is prepared by affinity chromatography with use of immobilized antibodies against human vitamin D-binding protein. Values obtained by the direct assays and those obtained by two different chromatographic methods correlated well (r greater than 0.95). The direct competitive protein-binding assay overestimated the true 25-hydroxyvitamin D3 concentration by about 20%, but this percentage was constant from 5 to 600 micrograms/L. Overestimation by the direct radioimmunoassay was less than 10%. These two direct assays for 25-hydroxyvitamin D3 allow reliable, rapid, and simple screening for vitamin D deficiency or excess.
mesh_terms: D000085:Acetates; D000818:Animals; D001667:Binding, Competitive; D002112:Calcifediol / Q000097:blood*; D002845:Chromatography / Q000379:methods; D003510:Cyclohexanes; D000431:Ethanol; D006801:Humans; D007118:Immunoassay / Q000379:methods; D011485:Protein Binding; D011863:Radioimmunoassay; D051381:Rats; D011919:Rats, Inbred Strains; D013056:Spectrophotometry, Ultraviolet; D014809:Vitamin D-Binding Protein
chemical_list: D000085:Acetates; D003510:Cyclohexanes; D014809:Vitamin D-Binding Protein; D000431:Ethanol; C007650:ethyl acetate; D002112:Calcifediol