A Direct Assay for Measuring Free 25-Hydroxyvitamin D

2017 ◽  
Vol 100 (5) ◽  
pp. 1318-1322 ◽  
Author(s):  
Nicolas Heureux ◽  
Ernst Lindhout ◽  
Leon Swinkels
Keyword(s):  
Liver Failure ◽  
Respiratory Diseases ◽  
Microtiter Plate ◽  
Cross Reactivity ◽  
Vitamin D Binding Protein ◽  
Calculation Methods ◽  
Hydroxyvitamin D ◽  
Pancreatic Cancers ◽  
Direct Elisa ◽  
25 Hydroxyvitamin D

Abstract Recent studies suggest that the concentration andgenotype of vitamin D binding protein (VDBP) are important factors that determine the bioavailability of25-hydroxyvitamin D [25(OH)D] in blood. Accumulatingdata indicate that, e.g., in pregnant women, hemodialysis patients, chronic kidney disease, liver failure, and bladder and pancreatic cancers, the measurement of free 25(OH)D in serum provides more relevant diagnostic information than measurement of total 25(OH)D. The aim of this study was to develop and validate an ELISA for direct measurement of free 25(OH)D in serum. A simple and direct ELISA was developed, based on a two-step immunoassay procedure performed ina microtiter plate. The assay has been characterizedin terms of precision (4–10% CV, according toconcentration), sensitivity (limits of blank = 0.5–1.0 pg/mL and LODs = 1.3–1.8 pg/mL), accuracy (correlation to dialysis, ELISA = 0.99xdialysis-0.5 pg/mL, r2 = 0.74), cross-reactivity of the antibody for the D2 form (77%),and addition of both VDBP and albumin (35–38%recovery upon addition of VDBP, 53–58% upon addition of albumin). The assay has already been usedin multiple studies, including its comparison with calculation methods and in studies of patients with liver failure, different ethnic groups, supplemented mice, respiratory diseases, and obesity. The free 25(OH)D ELISA can be used in studies as a valuable toolto establish the clinical relevance of free 25(OH)D.

Noncompetitive Immunoassay Detection System for Haptens on the Basis of Antimetatype Antibodies

2015 ◽  
Vol 61 (4) ◽  
pp. 627-635 ◽  
Author(s):  
Kazuya Omi ◽  
Tsuyoshi Ando ◽  
Takuya Sakyu ◽  
Takashi Shirakawa ◽  
Yoshiaki Uchida ◽  
...  
Keyword(s):  
Small Molecules ◽  
Limit Of Detection ◽  
Ex Vivo ◽  
Detection System ◽  
Sandwich Elisa ◽  
Microtiter Plate ◽  
Cross Reactivity ◽  
Serum Samples ◽  
Hydroxyvitamin D ◽  
25 Hydroxyvitamin D

Abstract BACKGROUND Small molecules classified as haptens are generally measured by competitive immunoassay, which is theoretically inferior to noncompetitive sandwich immunoassay in terms of sensitivity and specificity. We created a method for developing sandwich immunoassays to measure haptens on the basis of antimetatype antibodies. METHODS We generated antimetatype monoclonal antibodies against a hapten–antibody immunocomplex using an ex vivo antibody development system, the Autonomously Diversifying Library (ADLib) system. We selected 2 haptens, estradiol (E2) and 25-hydroxyvitamin D [25(OH)D], as analytes. Sandwich immunoassays for these 2 haptens were developed by use of a 96-well microtiter plate and a fully automated chemiluminescence analyzer, and the performances of these immunoassays were investigated. RESULTS The developed assays exhibited sensitivity high enough to detect target haptens in serum samples. The limit of detection of the ELISA for E2 was 3.13 pg/mL, and that of the fully automated chemiluminescent enzyme immunoassay (CLEIA) system was 2.1 ng/mL for 25(OH)D. The cross-reactivity with immunoreactive derivatives was effectively improved compared with the competitive assay. The CVs for the sandwich ELISA for E2 were 4.2%–12.6% (intraassay) and 6.2%–21.8% (total imprecision). The CVs for the sandwich CLEIA for 25(OH)D were 1.0%–2.3% (intraassay) and 1.9%–3.5% (total imprecision). In particular, the sandwich CLEIA for 25(OH)D showed correlations of r = 0.99 with both LC-MS/MS and a commercially available 125I RIA. CONCLUSIONS Our method represents a potentially simple and practical approach for routine assays of haptens, including vitamins, hormones, drugs, and toxins.

scholarly journals Circulating vitamin D binding protein, total, free and bioavailable 25-hydroxyvitamin D and risk of colorectal cancer

2015 ◽  
Vol 5 (1) ◽  
Author(s):  
Hou-Qun Ying ◽  
Hui-Ling Sun ◽  
Bang-Shun He ◽  
Yu-Qin Pan ◽  
Feng Wang ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
Vitamin D ◽  
Binding Protein ◽  
Vitamin D Binding Protein ◽  
Hydroxyvitamin D ◽  
25 Hydroxyvitamin D

scholarly journals Circulating 25‐hydroxyvitamin D, vitamin D binding protein and risk of advanced and lethal prostate cancer

2018 ◽  
Vol 144 (10) ◽  
pp. 2401-2407 ◽  
Author(s):  
Chen Yuan ◽  
Irene M. Shui ◽  
Kathryn M. Wilson ◽  
Meir J. Stampfer ◽  
Lorelei A. Mucci ◽  
...  
Keyword(s):  
Prostate Cancer ◽  
Vitamin D ◽  
Binding Protein ◽  
Vitamin D Binding Protein ◽  
Hydroxyvitamin D ◽  
25 Hydroxyvitamin D ◽  
Lethal Prostate Cancer

scholarly journals Investigating the Role of Functional Polymorphism of Maternal and Neonatal Vitamin D Binding Protein in the Context of 25-Hydroxyvitamin D Cutoffs as Determinants of Maternal-Neonatal Vitamin D Status Profiles in a Sunny Mediterranean Region

Nutrients ◽  
2021 ◽  
Vol 13 (9) ◽  
pp. 3082
Author(s):  
Spyridon N. Karras ◽  
Erdinç Dursun ◽  
Merve Alaylıoğlu ◽  
Duygu Gezen-Ak ◽  
Cedric Annweiler ◽  
...  
Keyword(s):  
Vitamin D ◽  
Mediterranean Region ◽  
Binding Protein ◽  
Hypovitaminosis D ◽  
Vitamin D Status ◽  
Vitamin D Binding Protein ◽  
Hydroxyvitamin D ◽  
High Prevalence ◽  
25 Hydroxyvitamin D ◽  
Causative Effect

Recent results indicate that dysregulation of vitamin D-binding protein (VDBP) could be involved in the development of hypovitaminosis D, and it comprises a risk factor for adverse fetal, maternal and neonatal outcomes. Until recently, there was a paucity of results regarding the effect of maternal and neonatal VDBP polymorphisms on vitamin D status during pregnancy in the Mediterranean region, with a high prevalence of hypovitaminosis D. We aimed to evaluate the combined effect of maternal and neonatal VDBP polymorphisms and different maternal and neonatal 25-hydroxyvitamin D (25(OH)D) cut-offs on maternal and neonatal vitamin D profile. Blood samples were obtained from a cohort of 66 mother–child pairs at birth. Our results revealed that: (i) Maternal VDBP polymorphisms do not affect neonatal vitamin D status at birth, in any given internationally adopted maternal or neonatal cut-off for 25(OH)D concentrations; (ii) neonatal VDBP polymorphisms are not implicated in the regulation of neonatal vitamin D status at birth; (iii) comparing the distributions of maternal VDBP polymorphisms and maternal 25(OH)D concentrations, with cut-offs at birth, revealed that mothers with a CC genotype for rs2298850 and a CC genotype for rs4588 tended to demonstrate higher 25(OH)D (≥75 nmol/L) during delivery (p = 0.05 and p = 0.04, respectively), after adjustments for biofactors that affect vitamin D equilibrium, including UVB, BMI and weeks of gestation. In conclusion, this study from Southern Europe indicates that maternal and neonatal VDBP polymorphisms do not affect neonatal vitamin D status at birth, whereas mothers with CC genotype for rs2298850 and CC genotype for rs4588 demonstrate higher 25(OH)D concentrations. Future larger studies are required to establish a causative effect of these specific polymorphisms in the attainment of an adequate (≥75 nmol/L) maternal vitamin D status during pregnancy.

scholarly journals Vitamin D Binding Protein Affects the Correlation of 25(OH)D and Frailty in the Older Men

2014 ◽  
Vol 2014 ◽  
pp. 1-6 ◽  
Author(s):  
Yi Wang ◽  
Yan-Jiao Wang ◽  
Jun-Kun Zhan ◽  
Zhi-Yong Tang ◽  
Wu Huang ◽  
...  
Keyword(s):  
Vitamin D ◽  
Binding Protein ◽  
Linear Regression Analysis ◽  
Older Men ◽  
Joint Effect ◽  
Hunan Province ◽  
Multivariate Linear Regression Analysis ◽  
Vitamin D Binding Protein ◽  
Hydroxyvitamin D ◽  
25 Hydroxyvitamin D

Vitamin D binding protein (DBP) may alter the biologic activity of 25-hydroxyvitamin D [25(OH)D]. The objective of our present study was to determine the joint effect of serum 25(OH)D and DBP on the risk of frailty. Five hundred sixteen male participants aged 70 years or older were recruited in Changsha city and its surrounding area in Hunan province of China. Frailty was defined as the presence of at least three of the five following criteria: weakness, low physical activity, slow walking speed, exhaustion, and weight loss. Multivariate linear regression analysis was performed to assess the relationship between 25(OH)D and DBP levels. Odds ratios (ORs) for frailty were evaluated across quartiles of 25(OH)D and DBP levels, adjusted age, education, and body mass index. The results showed that participants in the lowest quartile of 25(OH)D and the highest quartile of DBP levels, the lowest quartile of 25(OH)D and the lowest quartile of DBP levels, and those in the the lower quartile of 25(OH)D and lowest quartile of DBP levels had significantly higher OR of being frail compared with those in the highest quartile of 25(OH)D and lowest quartile of DBP, with OR of 3.18 (95% CI: 1.46–4.56,P<0.05), 2.63 (95% CI: 1.31–3.68,P<0.01), and 2.52 (95% CI: 1.22–3.52,P<0.05), respectively. The results indicate that the joint effect of serum 25(OH)D and DBP levels is associated with the risk of frailty, and serum DBP levels affects 25(OH)D-frailty relationship in the older men.

scholarly journals P431 Vitamin D binding protein in the limelight: IBD-related inflammation and circulating levels of vitamin D binding protein, total, free and bioavailable 25-hydroxyvitamin D

2020 ◽  
Vol 14 (Supplement_1) ◽  
pp. S393-S393
Author(s):  
A Aksan ◽  
K Böttger ◽  
N Hein ◽  
Y Caicedo-Zea ◽  
I Diehl ◽  
...  
Keyword(s):  
Vitamin D ◽  
Binding Protein ◽  
Transferrin Saturation ◽  
Simultaneous Detection ◽  
Full Blood Count ◽  
Vitamin D Binding Protein ◽  
Vitamin D Metabolism ◽  
Hydroxyvitamin D ◽  
Vitamin D Levels ◽  
25 Hydroxyvitamin D

Abstract Background Vitamin D deficiency occurs frequently in patients with Crohn’s disease (CD) and ulcerative colitis (UC). While recent cohort studies support an association of vitamin D with important clinical parameters and outcomes in IBD, the complex interplay of inflammation with vitamin D metabolism in IBD poses a viscious circle. We sought to further illucidate the relation between inflammation and different vitamin D parameters. To the best our knowledge, this was the first study to focus on the relationship between vitamin D binding protein (VDBP), circulating total, free, and bioavailable 25-hydroxyvitamin D (25(OH)D), and inflammation, in adult IBD patients. Methods This was a comparative, single-centred, cross-sectional study in patients with IBD aged 18–65 years. Full blood count, transferrin, albumin and hsCRP were determined by standard methods. The presence/absence of inflammation was assessed based on serum hsCRP levels (cutoff &lt;5mg/l). VDBP levels were determined by ELISA, and 25(OH)D by LCMS. Free and bioavailable vitamin D levels were calculated using the validated formula. IBM SPSS version 25.0 was used for statistical analysis. Results In total, 129 subjects with IBD (70 male/59 female; 82 CD/47 UC; mean age 41.7 ± 12.6 years) were enrolled. Of these, 38/129 had inflammation (19 m/19 f; 26 CD/12 UC; 39.6 ± 12.9 years) while 91/129 had no inflammation (40 m/51 f; 56 CD/35 UC; 42.5 ± 12.5 years). Subjects with disease activity had significantly higher leukocyte, erythrocyte sedimentation rate (ESR) and hsCRP, but lower transferrin, transferrin saturation (TSAT) and albumin levels than those without inflammation (p &lt; 0.05). Average serum levels of 25(OH)D (24.6[6.8–54.8] vs. 26.4[5.0–74.4]ng/ml), free 25(OH)D (5.9[1.3–13.3] vs. 1.0[1.0–21.4]ng/ml) and bioavailable 25(OH)D(2.3 [0.1–4.7] vs. 2.4[0.5–19.5]ng/ml) were similar in patients with vs. without inflammation (p &gt; 0.05). However, VDBP levels were significantly higher in inflammatory conditions (359.6[252.2–530.6] mg/l vs. 327.4[183.5–560.3]mg/l; p &lt; 0.05) and showed a positive correlation with CRP levels (0.293, p &lt; 0.001). Ratio of free/total 25(OH)D correlated negatively with CRP levels (−0.282, p = 0.002). Conclusion High levels of circulating VDBP were associated with inflammatory activity. Moreover, free/total 25(OH)D ratio was inversely associated with inflammation. Other vitamin D parameters including total, free and bioavailable 25(OH)D showed no association with inflammation. These findings suggest that VDBP may play a bigger role than thought as a modulator of vitamin D and inflammation, and that simultaneous detection and investigation of plasma VDBP may provide additional insights into this complex interaction.

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