Accuracy of 6 Routine 25-Hydroxyvitamin D assays; Influence of Vitamin D Binding Protein Concentration

A.C. Heijboer, M.A. Blankenstein, I.P. Kema, and M.M. Buijs

March 2012

www.clinchem.org/content/article/58/3/543.full

© Copyright 2012 by the American Association for Clinical Chemistry

© Copyright 2009 by the American Association for Clinical Chemistry

Introduction IIntroduction I

Vitamin D

High percentage of the population is vitamin D deficientVitamin D has skeletal and extraskeletal effects

Laboratory vitamin D requests on the increaseTherefore automation is desirable

© Copyright 2009 by the American Association for Clinical Chemistry

Introduction IIIntroduction II

Beware: vitamin D is a difficult analyte! Editorial of Carter, Clin Chem March 2012

Analysis of vitamin D Prerequisite: release vitamin D from binding protein (DBP) Traditional methods: RIA and HPLC preceded by extraction Automated methods: unclear whether displacement of

vitamin D from DBP is sufficient

© Copyright 2009 by the American Association for Clinical Chemistry

Question IQuestion I

What problem can arise if the displacement of vitamin D from its binding protein (DBP) is not sufficient?

© Copyright 2009 by the American Association for Clinical Chemistry

AimsAims

To test the accuracy of the currently available 25(OH)D assays

To assess the sensitivity of the various assays to differences in circulating DBP concentrations

© Copyright 2009 by the American Association for Clinical Chemistry

Material & MethodsMaterial & MethodsPlasma and serum samples

51 healthy individuals, 52 pregnant women, 50 hemodialysis patients, 50 intensive care (IC) patients

Methods vitamin D ID-XLC-MS/MS (accuracy established by measuring standard & control with reference method) 6 routine assays

• Architect i2000SR (Abbott Diagnostics) • Centaur XP (Siemens Diagnostics) • iSYS (IDS) • Liaison (Diasorin) • Elecsys (Roche Diagnostics)• RIA preceded by extraction (Diasorin)

Method DBPELISA (R&D systems)

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Question IIQuestion II

As compared with healthy controls, do you expect DBP to be increased or decreased?

In pregnant women In hemodialysis patients In intensive care patients

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Figure. 1. 25(OH)D concentrations [mean (SD)] measured by different methods in healthy individuals (n=51), pregnant women (n=52), hemodialysis patients (n=50), and IC patients (n=50). 25(OH)D was not measured in plasma from IC patients using the iSYS. To convert 25(OH)D concentrations to ng/mL, multiply by 0.4.

Main ResultsMain Results

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Figure 2. DBP concentrations in the circulation of healthy individuals, pregnant women, dialysis patients, and IC patients. *P <0.001.

Main ResultsMain Results

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Main ResultsMain Results

Table 1. Correlation coefficient, slope, and intercept according to Passing–Bablok regression for each 25(OH)D assay compared with ID-XLC-MS/MS in each clinical subject group.

a To convert 25(OH)D concentrations to ng/mL, multiply by 0.4b Significantly different from 1.00 (slope) and 0.00 (intercept) (P< 0.05)c ND, not determined

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Figure 3. Correlation (R) between the difference of 25(OH)D results between 6 routine 25(OH)D assays and the ID-XLC-MS/MS, and DBP concentration. For the Architect (P< 0.005), Centaur, iSYS, and Liaison (P< 0.0001), the correlation was statistically significant. To convert 25(OH)D concentrations to ng/mL, multiply by 0.4.

Main ResultsMain Results

© Copyright 2009 by the American Association for Clinical Chemistry

Question IIIQuestion III

Did you anticipate an inverse relationship between DBP concentrations and deviations of measured 25(OH)D concentrations from ID-XLC-MS/MS results?

Based on the answer to Question II, what is your explanation for this finding?

© Copyright 2009 by the American Association for Clinical Chemistry

ConclusionsConclusions Some of the assays used to measure 25(OH)D are not well standardized and report significantly different results from measurements performed with ID-XLC-MS/MS

The deviations which are sometimes serious are different in various patient groups, and are dependent on the concentration of DBP and other still-unknown interfering factors

Laboratory specialists, physicians, researchers, reviewers and authorities who provide advice on cutoffs for sufficiency and supplementation of 25(OH)D should carefully consider the 25(OH)D assay used in studies, before any conclusions are drawn or decisions made

© Copyright 2009 by the American Association for Clinical Chemistry

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