Development and Certification of a Standard Reference Material for Vitamin D Metabolites in Human Serum

Abstract

The National Institute of Standards and Technology (NIST), in collaboration with the National Institutes of Health’s Office of Dietary Supplements (NIH-ODS), has developed a Standard Reference Material (SRM) for the determination of 25-hydroxyvitamin D [25(OH)D] in serum. SRM 972 Vitamin D in Human Serum consists of four serum pools with different levels of vitamin D metabolites and has certified and reference values for 25(OH)D2, 25(OH)D3, and 3-epi-25(OH)D3. Value assignment of this SRM was accomplished using a combination of three isotope-dilution mass spectrometry approaches, with measurements performed at NIST and at the Centers for Disease Control and Prevention (CDC). Chromatographic resolution of the 3-epimer of 25(OH)D3 proved to be essential for accurate determination of the metabolites.

Automated Competitive Protein‐Binding Assay for Total 25‐OH Vitamin D, Multicenter Evaluation and Practical Performance

“…The HPLC system was calibrated with the Chromsystems Serum Calibration Standard D3/D3 nr 38033. To summarize, all LC-MS/MS and HPLC methods used were calibrated against either NIST SRM 2972 or commercially available calibrators (Chromsystems, Munich, Germany), traceable to NIST SRM 2972 (5,21). Calibration of the immunoassays Liaison 25 OH Vitamin D total (DiaSorin, Inc. Stillwater MN) and IDS-iSYS (Immunodiagnostic System, Boldon, UK) is traceable to UV spectrophotometry (26) and the assays were performed according to the manufacturers' instructions.…”

“…An association of vitamin D status with cardiovascular diseases, cancer, and autoimmunity has been described (1)(2)(3)(4)(5)(6).…”

“…These differences cause method-specific effects and trouble comparison results between laboratories (2,5,7,14,15). Moreover, reports on the high variability and different standardization of assays raised questions about assay reliability and possible impact on medical decisions (16)(17)(18)(19).…”

“…Moreover, reports on the high variability and different standardization of assays raised questions about assay reliability and possible impact on medical decisions (16)(17)(18)(19). Recently introduced standards SRM 972 (vitamin D in human serum) and SRM 2972 (ethanolic solutions of either 25-OHD2 or 25-OHD3) from the National Institute of Standards and Technology (NIST) for use in quality control or as primary assay calibrator (5,14,(17)(18)(19) should improve reliability and traceability. This article describes the evaluation of an automated competitive protein-binding assay in a multicenter study with special focus on traceability and performance in routine laboratory testing.…”

What is novel about certified reference materials?

“…For example, Phinney et al [6] do not describe a new CRM but instead satisfy the novelty requirement by offering a significantly Bimproved^material. In 2012 Phinney et al [7] reported the first serum-based CRM with values assigned for three vitamin D metabolites, i.e., 25-hydroxyvitamin D 2 2 , and high levels of 3-epi-25(OH)D 3 , which must be chromatographically separated from 25(OH)D 3 for accurate measurement of total 25(OH)D. To achieve target levels, the normal level was diluted with horse serum, which contains no 25(OH)D, to provide a low level, and the remaining two levels were fortified with either 25(OH)D 2 or 3-epi-25(OH)D 3 . Even though SRM 972 was widely used, a major criticism was that the use of modified serum and/or the presence of non-endogenous metabolites in three of the four levels produced inaccurate results using some assays, particularly immunoassays.…”