Targeted Metabolomics Services

Vitamin D

Updated 21/12/2022

What is measured?

Synonyms: 25-hydroxyvitamin-D3 (25OH-D3), 25-hydroxycholecalciferol, calcifediol, calcidiol; 25-hydroxyvitamin-D2 (25OH-D2), 25-hydroxyergocalciferol.
25-hydroxyvitamin-D3 and 25-hydroxyvitamin-D2 (1).
Method(s): LC-MS/MS.


Assessment of vitamin D status. 25OH-D3 and 25OH-D2 are the main circulating vitamin D forms. 25OH-D3 is most abundant and the important marker of vitamin D status, whereas 25OH-D2 is detected in a small portion of individual, is mainly derived from supplements, and may be inversely related to 25OH-D3 (2).

Specimen, collection and processing

Patient/subject: Vitamin D levels show a high level of seasonality due to the influence of sunlight.
Matrix: Serum or EDTA plasma.
Volume: Minimum volume is 50 µL, but 200 µL is optimal and allows reanalysis.
Preparation and stability: Samples should be put on ice, and protected from light.


Protected from light, frozen, on dry ice (for general instruction on transportation, click here)

Reported values, interpretation

Reported values (25OH-D3/25OH-D2): 10-280/0-60 nmol/L.
To convert from SI unit (nmol/L) to conventional unit (µg/L), multiply the SI value by the conversion factor = 0.4011. Intraclass correlation coefficient (ICC) (25OH-D3/25OH-D2): 0.71/na.


1. Midttun, Ø., McCann, A., Aarseth, O., Krokeide, M., Kvalheim, G., Meyer, K., and Ueland, P.M. (2016). Combined measurement of 6 fat-soluble vitamins and 26 water-soluble functional vitamin markers and amino acids in 50 μL of serum or plasma by high-throughput mass spectrometry. Anal Chem 88, 10427-436.
2. Yu, S., Zhang, R., Zhou, W., Cheng, X., Cheng, Q., Xia, L., Xie, S., Yin, Y., Sun, D., et al. (2018). Is it necessary for all samples to quantify 25OHD2 and 25OHD3 using LC-MS/MS in clinical practice? Clin Chem Lab Med 56, 273-77.

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