Vitamin D deficiency is associated with poor bone health and other adverse health outcomes; however, the associations are greatly attenuated in black vs white individuals. One possible explanation for this attenuation is different concentrations of bioavailable vitamin D metabolites in plasma, which are estimated with equations that include the total concentration of vitamin D binding globulin (VDBG) and haplotype-specific dissociation constants. We developed a method to quantify VDBG with LC-MS/MS that could also identify the haplotypes/isoforms of VDBG present. We validated the method according to recent recommendations for publications of biomarker studies. We determined serum VDBG concentrations in samples from the Atherosclerosis Risk in Communities cohort and compared the results with a widely used monoclonal immunoassay.
With 10 μL of serum or plasma, the lower limit of quantification for the assay (<20% CV) was 71 μg/mL. The assay was linear from 62 to 434 μg/mL, with total imprecision of 7.3-9.0% CV at approximately 250 μg/mL. Significant hemolysis interfered with quantification. The identification of isoforms was 97% concordant with genotyping (κ coefficient). Method comparison with immunoassay revealed significant isoform-specific effects in the immunoassay. Mean concentrations (SD) of VDBG by mass spectrometry were similar in whites and blacks [262 (25) vs 266 (35) μg/mL, respectively; P = 0.43]. Validated mass spectrometric methods for the quantification of proteins in human samples can provide additional information beyond immunoassay. Counter to prior observations by immunoassay, VDBG concentrations did not vary by race.
- Henderson CM, Lutsey PL, Misialek JR, Laha TJ, Selvin E, Eckfeldt JH, Hoofnagle AN. Measurement by a Novel LC-MS/MS Methodology Reveals Similar Serum Concentrations of Vitamin D-Binding Protein in Blacks and Whites. Clin Chem. 2016 Jan;62(1):179-87. PMID: 26453697; PMCID: PMC4698095.
Read More: Clinical Chemistry
- Research Center: University of Washington