Assessing vitamin D status: Pitfalls for the unwary

Abstract

The use of vitamin D testing has grown rapidly in the recent times as a result of increased interest in the role of vitamin D in health. Although the generally accepted measure of vitamin D status is circulating 25(OH)D concentration, there is little consensus on which assay method should be used. Commonly used assays include competitive protein-binding assay, RIA, enzyme immunoassay, chemiluminescence immunoassays, HPLC, and LC-MS/MS, each with its own advantages and disadvantages. However, there is significant interassay and interlaboratory variability in measurements. Our simulation of the published data showed that using a deficiency cut-point of 50 nmol/L, 57% of samples assessed using a chemiluminescence immunoassay were classified as deficient compared with 41% of samples assessed using LC-MS/MS; a 20% misclassification rate. Similar rates of misclassification were seen at 75 nmol/L. This has implications for clinical practice and decision limits for vitamin D supplementation, suggesting that cut-points should be assay specific rather than universal and that greater harmonization between laboratories is required. Newer assays using alternative biological samples to determine the circulating 25(OH)D have been proposed and advances in the genetics of vitamin D and the role of vitamin D-binding protein may improve future assay accuracy.

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

“…The use of polyclonal antibodies could lead to lot-to-lot variability and the risk of a different epitope recognition pattern upon immunization of different animals (7,14,15). Roche Diagnostics has substantial experience in developing competitive assays based on binding proteins.…”

“…Hence, the calibration of the Elecsys assay is traceable to the NIST standards described above. This approach improves accuracy, reduces variability between the different methods, and contributes to global harmonization of 25-OHD measurements (14,15,(17)(18)(19).…”

“…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).…”

Interferences in Immunoassays

“…For example, cortisol assays can show significant cross-reactivity with fludrocortisone derivates and result in falsely elevated cortisol levels in patients using these drugs (Berthod et al, 1988). The problem of cross-reactivity in active vitamin D (1,25(OH) 2 D) determination due to possible positive interference of 25(OH) D is well known (Lai et al, 2010).…”

Vitamin D deficiency: is it real? Need to re-evaluate in context to the latest research findings

“…The unit of UV exposure is "1 standard vitamin D dose" (SDD), and is defined as the time required to obtain the recommended UV dose for adequate vitamin D synthesis. [4][5][6] The time duration for 1 SDD varies with latitudes and longitudes and with season of the year. Also important is the time of the day when exposure is taken; though there are a handful of recommendations, but almost all agrees that mid day or solar noon is the optimal time for exposure and adequate Vit D synthesis.…”