Candidate Reference Measurement Procedures for Serum 25-Hydroxyvitamin D^sub 3^ and 25-Hydroxyvitamin D^sub 2^ by Using Isotope-Dilution Liquid Chromatography-Tandem Mass Spectrometry

25-hydroxyvitamin D [25(OH)D] assays are characterized by poor between-assay comparability. This result emphasizes the need for reference measurement procedures (RMPs) to establish calibration traceability and assist in method validation. We aimed at developing candidate RMPs on the basis of isotope...

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Bibliographic Details
Published in:Clinical chemistry (Baltimore, Md.) Md.), 2011-03, Vol.57 (3), p.441
Main Authors: Stepman, Hedwig C M, Vanderroost, An, Van Uytfanghe, Katleen, Thienpont, Linda M
Format: Article
Language:English
Subjects:
Calibration
Chromatography
Dietary supplements
Isotopes
Liquid chromatography
Mass spectrometry
Metabolites
Methods
Nutrition
Vitamin D
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Summary:25-hydroxyvitamin D [25(OH)D] assays are characterized by poor between-assay comparability. This result emphasizes the need for reference measurement procedures (RMPs) to establish calibration traceability and assist in method validation. We aimed at developing candidate RMPs on the basis of isotopedilution liquid chromatography-tandem mass spectrometry (ID-LC-MS/MS) for separate quantification of serum 25(OH)D^sub 2^ and 25(OH)D^sub 3^. Hexa-deuterated 25(OH)D^sub 3^/D^sub 2^ was added to serum. This mixture was extracted with n-hexane and fractionated on Sephadex LH-20 before 2-dimensional LC-MS/MS. In the first dimension, both procedures used a C4 column; however, in the second dimension, the 25(OH)D^sub 2^ procedure used a C18 and the 25(OH)D^sub 3^ procedure used a Zorbax SB-CN column. Calibration was traceable to the NIST Standard Reference Material (SRM) 2972. Validation comprised assessment of interference and limit of quantification/detection. Imprecision and trueness were validated by analysis of the SRM 972 against specifications (CV
ISSN:0009-9147
1530-8561