Dataset Information

Development and analytical validation of a novel bioavailable 25-hydroxyvitamin D assay.

ABSTRACT:

Background

Bioavailable 25-hydroxyvitamin D (25OHD) may be a better indicator of vitamin D sufficiency than total 25OHD. This report describes a novel assay for measuring serum bioavailable 25OHD.

Methods

We developed an assay for 25OHD % bioavailability based on competitive binding of 25OHD tracer between vitamin D-binding protein (DBP)-coated affinity chromatography beads and serum DBP. Bioavailable 25OHD, total 25OHD, albumin, and DBP protein concentrations were measured in 89 samples from hospitalized patients and 42 healthy controls to determine how the DBP binding assay responds to differences in concentrations of DBP and compares to calculated bioavailable 25OHD values.

Results

DBP binding assay showed a linear relationship between DBP-bound 25OHD tracer recovered from bead supernatant and DBP calibrator concentrations (y = 0.0017x +0.731, R2 = 0.9961, p<0.001). Inversion of this relationship allowed interpolation of DBP binding equivalents based upon 25OHD tracer recovered. The relationship between DBP binding equivalents and % bioavailability fits a non-linear curve, allowing calculation of % bioavailable 25OHD from DBP binding equivalents (y = 10.625x-0.817, R2 = 0.9961, p<0.001). In hospitalized patient samples, there were linear relationships between DBP protein concentrations and DBP binding equivalents (y = 0.7905x + 59.82, R2 = 0.8597, p<0.001), between measured vs. calculated % bioavailability (y = 0.9528 + 0.0357, R2 = 0.7200, p<0.001), and between absolute concentrations of measured vs. calculated bioavailable 25OHD (y = 1.2403 + 0.1221, R2 = 0.8913, p<0.001).

Conclusions

The DBP-binding assay for bioavailable 25OHD shows expected changes in 25OHD % bioavailability in response to changes in DBP concentrations and concordance with calculated bioavailable 25OHD concentrations.

SUBMITTER: Berg AH

PROVIDER: S-EPMC8270209 | biostudies-literature |

REPOSITORIES: biostudies-literature

ACCESS DATA

Similar Datasets

Project description:Background and aimsFree and bioavailable 25-hydroxyvitamin D (25OHD) are emerging measurements of vitamin D status. It remains unclear whether circulating free or bioavailable 25OHD are relevant to hepatocellular carcinoma (HCC) prognosis. Our aim was to test the hypothesis that bioavailable 25OHD may be a better serum biomarker of vitamin D status than total 25OHD on the association with HCC survival.Approach and resultsWe included 1,031 newly diagnosed, previously untreated patients with HCC from the Guangdong Liver Cancer Cohort enrolled between September 2013 and April 2017. Serum total 25OHD levels were measured using an electrochemiluminescence immunoassay. Serum-free 25OHD levels were measured using a two-step enzyme-linked immunosorbent assay. Bioavailable 25OHD levels were calculated from measured free 25OHD and albumin using a previously validated equation. Primary outcomes were liver cancer-specific (LCSS) and overall survival (OS). Cox proportional hazards models were performed to calculate the multivariable hazard ratios (HRs) and 95% confidence intervals (CIs). During a median follow-up of 726 days, 430 patients had deceased, including 393 deaths from HCC. In multivariable analyses, higher bioavailable 25OHD levels were significantly associated with better survival, independent of nonclinical and clinical prognostic factors including serum C-reactive protein, Barcelona Clinic Liver Cancer stage, and cancer treatment. The multivariable-adjusted HRs in the highest versus lowest quartile of bioavailable 25OHD levels were 0.69 (95% CI: 0.51, 0.93; P for trend = 0.014) for LCSS and 0.71 (95% CI: 0.53, 0.94; P for trend = 0.013) for OS. In contrast, neither total nor free 25OHD levels were associated with LCSS or OS.ConclusionsHigher bioavailable, rather than total, 25OHD levels were independently associated with improved survival in a population-based HCC cohort, suggesting a potential utility of bioavailable 25OHD in HCC prognosis.

Project description:ObjectivesVitamin D is essential for bone health. Not only total but also free 25-hydroxyvitamin D (25OHD) may contribute to bone mass. We sought to determine which vitamin D measure best reflected clinical and bone parameters in healthy children.MethodsA cross-sectional study including 146 healthy children (71 boys, 9.5 ± 1.9 years) conducted at a tertiary medical center. We used a multiplex liquid chromatography-tandem mass spectrometry-based assay to simultaneously measure vitamin D metabolites. The bioavailable and free 25OHD (25OHDBioA and 25OHDFree) levels were calculated using the genotype-specific or genotype-constant affinity coefficients of vitamin D-binding proteins (yielding spe-25OHDBioA, spe-25OHDFree and con-25OHDBioA, con-25OHDFree respectively). The 25OHDFree level was directly measured (m-25OHDFree). Bone mineral content (BMC) and bone mineral density (BMD) were assessed via dual-energy X-ray absorptiometry.ResultsThe total 25OHD (25OHDTotal), the two forms of 25OHDBioA, the three forms of 25OHDFree, and 24,25-dihydroxyvitamin D3 levels correlated with parathyroid hormone level (all p < 0.01). Serum 25OHDTotal and m-25OHDFree levels were influenced by age, pubertal status, season, body mass index (BMI), daylight hours, and vitamin D intake (all p < 0.05). The con-25OHDBioA and con-25OHDFree levels better reflected pubertal status and daylight hours than did the spe-25OHDBioA and spe-25OHDFree levels (both p < 0.01). The association between the 25OHDTotal level and bone parameters varied according to the BMI (interaction p < 0.05). In 109 normal-weight children, the con-25OHDBioA and con-25OHDFree levels correlated with total body BMC and BMD (both p < 0.05), whereas the 25OHDTotal and 24,25-dihydroxyvitamin D3 levels were associated with total body BMC (both p < 0.05). No such association was found in overweight or obese children.ConclusionsIn healthy children, total, bioavailable, and free 25OHD levels comparably reflected lifestyle factors. In normal-weight children, the con-25OHDBioA and con-25OHDFree, but not m-25OHDFree levels, reflected bone mass, as did the 25OHDTotal level.