Methodological issues in assessing plasma 25-hydroxyvitamin D concentration in newborn infants

Sina Gallo; Kathryn Comeau; Sherry Agellon; Catherine Vanstone; Atul Sharma; Glenville Jones; Mary L'abbé; Ali Khamessan; Hope Weiler; Celia Rodd

doi:10.1016/j.bone.2014.01.012

Methodological issues in assessing plasma 25-hydroxyvitamin D concentration in newborn infants

Bone. 2014 Apr:61:186-90. doi: 10.1016/j.bone.2014.01.012. Epub 2014 Jan 25.

Authors

Affiliations

¹ Department of Nutrition and Food Studies, George Mason University, Fairfax, VA, USA.
² School of Dietetics and Human Nutrition, McGill University, Montréal, Québec, Canada.
³ Montréal Children's Hospital, McGill University Health Centre, Montréal, Québec, Canada.
⁴ Department of Biomedical and Molecular Sciences, Department of Medicine, Queen's University, Kingston, Ontario, Canada.
⁵ Department of Nutritional Sciences, University of Toronto, Toronto, Ontario, Canada.
⁶ Euro-pharm International Canada Inc., Montréal, Québec, Canada.
⁷ School of Dietetics and Human Nutrition, McGill University, Montréal, Québec, Canada; Montréal Children's Hospital, McGill University Health Centre, Montréal, Québec, Canada. Electronic address: crodd@hsc.mb.ca.

PMID: 24473373
DOI: 10.1016/j.bone.2014.01.012

Abstract

Background: Although no gold standard exists, liquid chromatography tandem mass spectrometry (LC-MS/MS) is a precise and accurate method for the analysis of plasma 25-hydroxyvitamin D (25(OH)D). Immunoassays are more readily available and require small volume sampling, ideal for infant testing. The objective was to compare two commercially available immunoassays for measuring circulating 25(OH)D concentration in infant plasma against LC-MS/MS.

Methods: Capillary blood samples from 103 infants were analyzed for plasma 25(OH)D using an enzyme immunoassay (EIA, Octeia, IDS Ltd.) and radioimmunoassay (RIA, DiaSorin). Plasma 25(OH)D(3), C-3 epimer of 25(OH)D(3) (3-epi-25(OH)D(3)) and 24,25-dihydroxyvitamin D (24,25(OH)(2)D(3)) were measured on the same samples using LC-MS/MS. To establish whether plasma 24,25(OH)(2)D(3) or 3-epi-25(OH)D(3) interferes with these immunoassay results, the zero 25(OH)D calibrator from each assay kit was spiked with increasing amounts of 24,25(OH)(2)D(3) or 3-epi-25(OH)D(3).

Results: Classifying infants below the common vitamin D status targets of 50 nmol/L and 75 nmol/L respectively, 58% and 99% fell below using the RIA, 19% and 56% with the EIA and 31% and 76% with LC-MS/MS. Compared to LC-MS/MS, both immunoassays showed poor Bland-Altman limits of agreement for 25(OH)D concentrations (RIA: limits of agreement -27 to +13%; EIA: -12 to +41%), and mountain plots (folded cumulative distribution) depicted significant skew and bias. Spiked 24,25(OH)2D3 concentrations, but not 3-epi-25(OH)D3, appeared as >100% of known values on the EIA but not on the RIA thus, suggesting that the EIA may cross-react with 24,25(OH)(2)D(3) to a greater extent than 3-epi-25(OH)D(3).

Conclusion: Two common immunoassays resulted in very different classifications of vitamin D status possibly related to the interference of other vitamin D metabolites. Based on these data, LC-MS/MS assessment of vitamin D status is recommended in young infants (4-6 weeks of age).

Keywords: 25-Hydroxyvitamin D; Enzyme immunoassay; Gold standard; Infants; Liquid chromatography tandem mass spectrometry; Radioimmunoassay.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Chromatography, Liquid / methods*
Female
Hematologic Tests / methods*
Humans
Immunoassay / methods*
Infant
Infant, Newborn
Male
Tandem Mass Spectrometry / methods*
Vitamin D / analogs & derivatives*
Vitamin D / blood

Substances

Vitamin D
25-hydroxyvitamin D

Grants and funding

Canadian Institutes of Health Research/Canada