title: Four years of LC-MS/MS method for quantification of 25-hydroxyvitamin D (D2+D3) for clinical practice.
abstract: Many methods for routine total plasma/serum 25-hydroxyvitamin D (25OHD) measurements are available from automated immunoassays to the most specific LC-MS/MS techniques. These last ones are nowadays numerous, still perfectible but more powerful than immunoassays in specific illnesses. We presented a robust method for simple quantification of 25-hydroxyvitamin D(2) (25OHD(2)) and 25-hydroxyvitamin D(3) (25OHD(3)) in human plasma by LC-APCI-MS/MS. This method is reliable and easy to perform for clinical measurements as we report a 4-year of clinical laboratory use. A brief off-line sample pretreatment (protein precipitation with addition of the internal standard) was realized then the supernatant was loaded into 96-well plates and analyzed by an online SPE-LC/MS/MS method on an APCI mode. 25OHD(2) and 25OHD(3) were both measured. The chromatographic system was thought and optimized for providing a dedicated line for this measurement on a shared instrument. The linearity was tested up to 380 nmol/L for both 25OHD(2) and 25OHD(3). The limit of quantification (LOQ) was 7 and 8 nmol/L for 25OHD3 and 25OHD(2), respectively. Routine imprecision and bias were found in agreement with recommended limits for routine testing, CV≤10% and bias≤5%. Since July 2010, our participation to DEQAS was successfully validated. This simple robust online SPE-LC/MS/MS method is suitable for routine measurement of 25OHD(2) and 25OHD(3) in human adult plasma. The assay operates for 4 years and has performed more than 40,000 patient samples on a shared instrument.
authors: Unit of Hormone and Nutrition, Department of Biochemistry, Toxicology and Pharmacology, University Hospital Center, Grenoble, France.
authors: Cécile
authors: C
authors: Meunier
authors: Unit of Hormone and Nutrition, Department of Biochemistry, Toxicology and Pharmacology, University Hospital Center, Grenoble, France.
authors: Julien
authors: J
authors: Montérémal
authors: Unit of Hormone and Nutrition, Department of Biochemistry, Toxicology and Pharmacology, University Hospital Center, Grenoble, France.
authors: Patrice
authors: P
authors: Faure
authors: Unit of Hormone and Nutrition, Department of Biochemistry, Toxicology and Pharmacology, University Hospital Center, Grenoble, France. Electronic address: VDucros@chu-grenoble.fr.
authors: Véronique
authors: V
authors: Ducros
mesh_terms: D000328:Adult; D002762:Cholecalciferol / Q000097:blood*; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D003955:Diagnostic Tests, Routine / Q000379:methods*; D004872:Ergocalciferols / Q000097:blood*; D005260:Female; D006801:Humans; D008297:Male; D015203:Reproducibility of Results; D053719:Tandem Mass Spectrometry / Q000379:methods*; D055815:Young Adult
chemical_list: D004872:Ergocalciferols; D002762:Cholecalciferol
keywords: Clinical practice; High throughput; Mass spectrometry; Vitamin D