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Background. The clinical relevance of circulating 1,25-dihydroxyvitamin D (1,25(OH)2D) is probably underappreciated, but variations in the measurement of this difficult analyte between different methods limit comparison of results. Methods. In 129 clinical samples, we compared a new automated assay with a commercially available liquid chromatography tandem mass spectrometry (LC-MS/MS) kit. Results. Median (interquartile range) 1,25(OH)2D concentrations with the automated assay and the LC-MS/MS method were 26.6 pg/mL (18.5–39.0 pg/mL) and 23.6 pg/mL (16.1–31.3 pg/mL), respectively ( P = 0.001 ). Using the method-specific cut-offs for deficient 1,25(OH)2D levels (<20 pg/mL for the automated assay and <17 pg/mL for the LC-MS/MS method), the percentage of patients classified as 1,25(OH)2D deficient was 28.7% and 27.1%, respectively. However, concordance between the two methods for deficient levels was only 62% and the concordance correlation coefficient was poor (0.534). The regression equation resulted in an intercept of −1.99 (95% CI: −7.33–1.31) and a slope of 1.27 (95% CI: 1.04–1.52) for the automated assay. The mean bias with respect to the mean of the two methods was −3.8 (1.96 SD: −28.3–20.8) pg/mL for the LC-MS/MS method minus the automated assay. Conclusions. The two methods show only modest correlation and further standardization is required to improve reliability and comparability of 1,25(OH)2D test procedures.

Measurement of Circulating 1,25-Dihydroxyvitamin D: Comparison of an Automated Method with a Liquid Chromatography Tandem Mass Spectrometry Method