Normal rabbit intestinal cytosol as a source of binding protein for the 1,25-dihydroxyvitamin D3 assay

✍ Scribed by W.E. Duncan; T.C. Aw; P.G. Walsh; J.G. Haddad

• Publisher: Elsevier Science
• Year: 1983
• Tongue: English
• Weight: 460 KB
• Volume: 132
• Category: Article
• ISSN: 0003-2697
• DOI: 10.1016/0003-2697(83)90449-9

✦ Synopsis

Cytosol prepared from small intestine of vitamin D-sufficient rabbits contains a specific highaffinity binding protein for 1,25dihydroxyvitamin D, (1,2S(OH),Ds). This binding protein sediments at 3.0-3.5 S in sucrose. density gradients containing 0.3 M KCI. Scatchard analysis using intestinal cytosol demonstrated a Kd of 0.05 nM and a maximum binding capacity of 92 fmol/ mg cytosol protein for 1,25(OH)rD, at 4°C. Competitive binding studies with various metabolites of vitamin D showed a relative binding affinity of this protein for l,25(OH)zDr > 25-hydroxyvitamin Dr > vitamin Dg. With 200 Ng of rabbit intestinal cytosol protein, as little as 1.0-2.5 pg of 1,25(OH)zDs reproducibly displaced the tracer sterol from the binding protein. Analyses of human plasma 1,25(OH)2DS content yielded values consistent with published results. The vitamin D-replete rabbit provides a convenient, plentiful, and inexpensive source of binding protein for 1 ,25(OHhD3 assays.