1,25-Dihydroxyvitamin D Testing in Clinical Practice: A Literature Review on Diagnostic Challenges Arising From Analytical Variability

As our understanding of vitamin D and its metabolites has improved, different assays have been developed for measuring their serum concentrations. The predominant circulating metabolite, 25-hydroxyvitamin D (25(OH)D), has been extensively studied and is now widely used for assessing vitamin D status. Of current interest is the biologically active metabolite, 1,25-dihydroxyvitamin D (1,25(OH)2D), which is involved in numerous physiological processes related to bone metabolism and various non-calcaemic functions. Quantification of 1,25(OH)2D serves not only as a diagnostic tool for numerous diseases, but also as a way to ensure optimal levels in the circulation to maintain physiological functioning. Measuring 1,25(OH)2D, however, proves challenging as its concentration in the blood is very low, and the molecule itself is highly lipophilic. A variety of analytical assays have been developed for measuring 1,25(OH)2D, each differing in its analytical principles. These methods exhibit variations in their performances, showcasing both strengths and limitations in their underlying analytical principles; some of these assays continue to be used today, while others have become obsolete. Comparability of these methods to this day is challenging due to the absence of a standardisation scheme for 1,25(OH)2D.