NACB - Scientific Shorts
NACB - Scientific Shorts (formerly NACB Blog)
By Ravinder Singh, PhD
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In the US, millions of 25-OH-D tests have been performed in last few years. Testing has increased at the rate of 80% to 90% per year and has overwhelmed the clinical laboratories. Vitamin D deficiency is an important concern, but assays for serum 25-hydroxy vitamin D (25-OH-D), the accepted marker for vitamin D nutritional status, are not standardized. This raises questions about the adequacy of the methods used to make these measurements. Recognition that vitamin D deficiency may be more prevalent in most patient populations than earlier assumed has resulted in an unexpected and marked increase in the volume of testing for 25-OH-D in clinical laboratories.

 

Various methods are available for measuring circulating concentrations of 25-OH-D. Current methods include HPLC, RIA with low throughput to high throughput, automated chemiluminescence immunoassays, and liquid chromatography-tandem mass spectrometry (LC-MS/MS). This variability in methods has already aroused controversy.

 

Institute of Medicine Report

In the latest Institute of Medicine report, experts concluded that <12 ng/ml is indication of deficiency and 20 ng/ml is the optimum level for most North Americans. This report didn’t find any negative correlation with the PTH which was earlier assumed to be the basis of determining optimum Vitamin D for sufficiency.

 

In contrast, labs have been raising the cut-off for deficiency levels every year and some labs may have flagged results between 20-30 ng/ml to be low. How quickly will/should labs change their reference ranges? Should labs revise/recall their reports since lab results may have triggered treatment in thousands of patients?

 

When should the test be utilized?

Should clinical labs even offer or charge for 25-OH-vitamin D for screening vitamin D deficiency when there is no consensus on the optimum value or reference ranges?

 

There is probably no need to have this test done or ordered in healthy individuals. However, vitamin D should always be ordered and interpreted in the context of disease (osteomalacia, secondary hyperparathyroidism) and interpreted in relation to markers like PTH and renal function. Vitamin D is part of the endocrine system and levels of active vitamin D (1,25 dihydroxy vitamin D) are influenced by the circulating calcium levels. Calcium intake influences PTH which further regulates the conversion of vitamin D to 1,25 dihydroxy vitamin D.

 

What is the value to patient care?

Is it cost effective to screen populations for determining vitamin D deficiency when the treatment could be a change in life style (reasonable sun exposure) or inexpensive Vitamin D supplementation? Despite the performance of millions of tests in the last five years, I am not sure if we have added any value to the outcomes for patient care in any disease.

 

What about test performance?

What should the coefficient of variation (CV) of the 25-OH-vitamin D test be at LOQ? Calcium, like most automated assays performed in various labs, has CVs down in the 1-2% range. Other testing related to calcium metabolism, like PTH, have precision performance that is better than 25-OH-Vitamin D tests (10-20%) as observed in proficiency testing data.

 

Are we measuring, and should we be reporting both the D3 and D2 forms?

Like Vitamin D3 (Cholecaliferol), Vitamin D2 (Ergocalciferol) is metabolized to 25-OH-vitamin D2. Assays should quantify these two forms equally, but do we know if this is true for various assays used clinically? Immunoassays may not measure D2 at all, or may measure it at 100%. The HPLC and LC-MS/MS assays, by design, separate and quantify the two vitamin forms, yet 25-OH-Vitanmin D is always reported as total by adding the two together, rather than reporting them individually.

 

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Posted by AACC\bgarman
On 6/20/2011

The role and status of Vitamin D nutritionally and in skeletal disorders is profound and undisputed. The science of measurement for steroids is quite a challenge. Dr Singh has given a brilliant account of the various issues that require consideration in testing of Vitamin D. The IOM report has been a source of caution and balance on the status of Vitamin D-and laboratory scientists, pathologists, and clinicians must understand that the IOM recommendation is from a population model perspective and not a medical model. Furthermore, the metrology of Vitamin D testing is just beginning to mature with the availability of standards for calibration from the NIST. With higher order reference materials having traceable and commutable characteristics and with reference methods for testing Vitamin D there will be better harmonization of results between testing sites serving interests of all stakeholders. With the cost and expertise to operate the LC-MS/MS becoming easier and affordable, moreover, Immunoassays are also becoming more focused on standardization, the diagnostic testing of Vitamin D will become more valuable. The final Arbiter in this interesting debate will review to distinguish: which laboratorians can maintain higher testing performance for Vitamin D by participating in an External Proficiency & Accuracy Testing surveys and using standardized testing methods and which clinical research and epidemiologist group can establish a causative relation of Vitamin D to non-skeletal disorders through credible RCT data. Shoukat Khan Pathology Quality Specialist Riyadh Military Hospital

Posted by AACC\bgarman
On 6/16/2011

The IOM report also makes it clear that the only unequivocal health benefits of vitamin D supplementation concern bone health. The therapeutic value of vitamin D in cancer prevention, improvement in immune function, etc., has not been unquestionably proven. We must remember that association studies do not provide proof of causality. Note: For the blog readers, always check the 25-OHD units that are used when you are reading the literature or when you are comparing patient results and reference intervals. Ravinder - Should patients on very high dose vitamin D (e.g., >1000- 2000 units /day) be monitored for their 25-OHD levels? Should clinicians test for hypercalcemia, hyperphosphatemia and hypercalciuria in patients on very high dose vitamin D?

About the Author
Ravinder Singh, PhD
Ravinder Singh, PhD