title: A simple micro-extraction plate assay for automated LC-MS/MS analysis of human serum 25-hydroxyvitamin D levels.
abstract: This short application note describes a simple and automated assay for determination of 25-hydroxyvitamin D (25(OH)D) levels in very small volumes of human serum. It utilizes commercial 96-well micro-extraction plates with commercial 25(OH)D isotope calibration and quality control kits. Separation was achieved using a pentafluorophenyl liquid chromatography column followed by multiple reaction monitoring-based quantification on an electrospray triple quadrupole mass spectrometer. Emphasis was placed on providing a simple assay that can be rapidly established in non-specialized laboratories within days, without the need for laborious and time consuming sample preparation steps, advanced calibration or data acquisition routines. The analytical figures of merit obtained from this assay compared well to established assays. To demonstrate the applicability, the assay was applied to analysis of serum samples from patients with chronic liver diseases and compared to results from a routine clinical immunoassay.
authors.affiliation: Institute of Bioanalytical Chemistry, Department of Chemistry, Saarland University, Saarbrücken, Germany.
authors.forename: Timon
authors.initials: T
authors.lastname: Geib
authors.forename: Florian
authors.initials: F
authors.lastname: Meier
authors.forename: Pascal
authors.initials: P
authors.lastname: Schorr
authors.forename: Frank
authors.initials: F
authors.lastname: Lammert
authors.forename: Caroline S
authors.initials: CS
authors.lastname: Stokes
authors.forename: Dietrich A
authors.initials: DA
authors.lastname: Volmer
mesh_terms: D001331:Automation; D002138:Calibration; D005591:Chemical Fractionation / Q000295:instrumentation*; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D002853:Chromatography, Liquid / Q000295:instrumentation / Q000379:methods*; D002908:Chronic Disease; D004867:Equipment Design; D006801:Humans; D008107:Liver Diseases / Q000097:blood* / Q000188:drug therapy; D008163:Luminescent Measurements / Q000379:methods; D011786:Quality Control; D015203:Reproducibility of Results; D021241:Spectrometry, Mass, Electrospray Ionization / Q000295:instrumentation; D053719:Tandem Mass Spectrometry / Q000295:instrumentation / Q000379:methods*; D014807:Vitamin D / Q000031:analogs & derivatives* / Q000097:blood / Q000627:therapeutic use
chemical_list: D014807:Vitamin D; C104450:25-hydroxyvitamin D
keywords: 25-hydroxyvitamin D; LC-MS/MS; chronic liver diseases; electrospray; micro-extraction; sample preparation; serum; vitamin D