Measurement of 25-hydroxyvitamin D3 in serum.
We describe a method for measuring 25-hydroxyvitamin D3 in serum. Extraction with dichloromethane/methanol (2/1 by vol), followed by chromatography on a column of Sephadex LH-20, resulted in an overall analytical recovery of 82% +/- 3.5% (SD). Diluted normal rat serum was used as binding protein because it contains a transport protein that has both a high affinity (Ka = 2 X 10(10) liter/mol) and a high capacity (3 X 10(-6) mol/liter) for 25-hydroxyvitamin D3. There is no advantage in using more complex binding proteins derived either from rachitic animals or from cytosol preparations. Concentrations of 25-hydroxyvitamin D3 (13.4 +/- 4 mug/liter) in the serum of apparently normal Belgian subjects are lower than those reported for North Americans, but resemble those reported for the United Kingdom.
D000818:Animals; D001681:Biological Assay; D002352:Carrier Proteins / Q000097:blood; D002645:Chickens; D002852:Chromatography, Ion Exchange / Q000379:methods; D006801:Humans; D006863:Hydrogen-Ion Concentration; D006887:Hydroxycholecalciferols / Q000097:blood* / Q000302:isolation & purification; D007676:Kidney Failure, Chronic / Q000097:blood / Q000188:drug therapy; D011869:Radioligand Assay / Q000379:methods; D051381:Rats; D012279:Rickets / Q000378:metabolism
D002352:Carrier Proteins; D006887:Hydroxycholecalciferols