| Abstract | BACKGROUND: Measurement of vitamin D molecules are important in the management of patients with bone disease. We developed a liquid chromatography-tandem mass spectrometry (LC-MS/MS) method to measure 25OHD(3) and 25OHD(2) in human serum and compared it to the traditionally used DiaSorin radioimmunoassay (RIA). METHODS: Serum samples (200 microl) were treated with acetonitrile and centrifuged to remove protein. An online solid-phase extraction procedure was used. Calibration solutions (5-100 ng/ml) of 25OHD(2) and 25OHD(3) were prepared using 4% albumin in phosphate-buffered saline. Chromatography: C18 column, isocratic ethanol/water (83/17, v/v). Mass spectrometry system: atmospheric pressure chemical ionization in the positive ion mode. Transitions: 25OHD(3), m/z 401.4-->383.4; 25OHD(2), m/z 413.4-->395.4; and the internal standard hexadeuterated-25OHD(3), m/z 407.7-->389.7. RESULTS: Detection limits were 0.49 ng/ml (25OHD(3)) and 1.86 ng/ml (25OHD(2)). Intra- and inter-assay coefficients of variation (CV) were <7% and <11%, respectively, for 25OHD(3) and <9% and <16%, respectively, for 25OHD(2). Recovery averaged (SD) 99% (2%) for 25OHD(3) and 95% (0.8%) for 25OHD(2). In a method comparison of 551 specimens from the National Health and Nutrition Examination Survey, the LC-MS/MS method gave values that were on average 13% higher (95%CI: 11-15%) than RIA results. CONCLUSIONS: This high throughput candidate reference method requires minimal sample preparation and is suitable for routine use for analysis of vitamin D status. |
| Authors | Huiping Chen, Leslie F McCoy, Rosemary L Schleicher, Christine M Pfeiffer
(Affiliation: Centers for Disease Control and Prevention, Atlanta, GA, USA.)
|
| Journal | Clinica chimica acta; international journal of clinical chemistry
(Clin Chim Acta)
Vol. 391
Issue 1-2
Pg. 6-12
(May 2008)
ISSN: 0009-8981 Netherlands |
| PMID | 18279671
(Publication Type: Comparative Study, Journal Article)
|
| Chemical References |
- Calcifediol
- 25-Hydroxyvitamin D 2
|
| Topics |
- 25-Hydroxyvitamin D 2
(blood)
- Calcifediol
(blood)
- Chromatography, Liquid
(methods)
- Humans
- Radioimmunoassay
(methods)
- Tandem Mass Spectrometry
(methods)
|
