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FDA-approved vitamin D immunoassays may be inaccurate

NEW YORK (Reuters Health) – An evaluation of two FDA-approved automated immunoassays for 25 hydroxyvitamin D found high variability in the tests’ results. Neither Abbott Laboratories’ Architect nor Siemens’ Centaur2 met even a minimum quality standard for analytical bias, researchers say.

“The two tests tend to underestimate vitamin D levels, so they have the potential to overcall vitamin D deficiency, said Dr. Earle Holmes of Loyola University Medical Center in Maywood, Illinois, who led the study, in an interview.

Consequently, “You might over-diagnose vitamin D insufficiency and prescribe supplements because of an inaccurate test result,” Dr. Holmes said.

He presented the research Monday at the Endocrine Society’s annual meeting in Houston, Texas.

Dr. Holmes and his team tested 163 randomly selected blood samples with both immunoassays, then sent the samples to another laboratory for a comparison test by liquid chromatography with tandem mass spectrometry (LCMS).

While LCMS found a vitamin D deficiency (defined as less than 20 ng/mL) in 33 samples, Abbott’s test found 45 with deficiencies and the Siemens’ assay found 71.

The tests generally skewed toward over-reporting deficiencies, but there were differences in how each brand reported the different types of vitamin D.

Ninety-eight samples had hydroxyvitamin D2 alone, and 65 had hydroxyvitamin D2 as well as D3.

Abbott’s test tended to under-report the presence of vitamin D2 and Siemens’ test tended to over-report it. Siemens’ test also over-reported vitamin D3 at high levels, Dr. Holmes says.

The findings may support a growing body of evidence about the inaccuracies in these kinds of tests. A study published in March in the journal Clinical Chemistry found variability between five immunoassays, two of which were included in the tests by Dr. Holmes’ team.

“Variability between assays for 25OHD levels is not a new problem, but has been recognized for several years,” says Dr. Daniel Bikle, a professor of medicine at the University of California, San Francisco, who has led research on vitamin D but was not involved in this study.

“All assays have limitations, but those dependent on antibodies appear to be more problematic than those dependent on physical-chemical characteristics such as LCMS or HPLC (high-performance liquid chromatography). It is a major concern to those of us making decisions regarding who should receive vitamin D supplementation and judging the effectiveness of such supplementation that the assays remain so variable,” Dr. Bikle says.

There is an effort underway to address the problem. The National Institutes of Health, the Centers for Disease Control and Prevention, and the National Institute for Standards and Technology launched the Vitamin D Standardization Program in November 2010, to standardize the detection and treatment of vitamin D deficiency.

In the meantime, Dr. Holmes doesn’t think any other immunoassays are more reliable than the two tested here. “I would be reluctant to use another immunoassay again because I think that in general, as a class, the immunoassays have issues like the ones we reported for these two,’ he said. “They have a lot of variability and trouble calculating vitamin D accurately and may be calibrated differently in some cases. There are new candidates coming online all the time. But the important thing is, they have to be tested rigorously.”

Don Braakman, a spokesperson for Abbott, told Reuters Health by email that in other studies, including an Abbott-sponsored study presented at the same meeting by Dr. Brian Keevil of the University of Manchester, UK, et al, “the ARCHITECT vitamin D test showed excellent precision and reproducibility of results from lab to lab…The Holmes study used the LC Mass Spec method, which is a very hands-on, labor intensive, manual procedure for which there currently is no standard protocol or calibration method. As a result, this leads to variability of results from lab to lab. While LC Mass Spec may be the appropriate method for Dr. Holmes’ lab, based on his patient population, it may not be the test of choice for a lab with a typical patient population, as seen worldwide.”

Dr. Beth Schodin, Abbott’s Global Scientific Affairs Manager, told Reuters Health that immunoassays are easily automated, provide a quick turnaround, and at low cost compared to chromatography.

As for the under-reporting of vitamin D2 by the Architect assay in Dr. Holmes’ research, she noted that vitamin D2 reflects the form provided by very large supplemental doses available only by prescription. People’s vitamin D levels from sun exposure and from most over-the-counter supplements are reflected by vitamin D3 measurements.

Even from one LCMS machine to another, she says, results can differ because the systems are calibrated to different standards.

“In an ideal world,” every assay would give standardized results, Dr. Schodin said. But she believes the Architect assay provides clinically useful answers to “Is there a severe deficiency?” and “Is there a directional change with supplementation?”

Dr. Holmes advises clinicians to contact the labs they work with to discuss vitamin D testing.

“There’s a pretty substantial and growing literature out there on these immunoassays and what’s being evaluated,” he said. “They can find data similar to ours that has been published. We’re at the beginning of a groundswell of enlightenment that these assays aren’t very good. I know there’s a lot of upcoming meetings where researchers will recapitulate data like ours.”

A representative of Siemens did not respond to a request for comment.