A Competitive Protein-Binding Assay for 25-Hydroxyvitamin D

1976 ◽  
Vol 51 (6) ◽  
pp. 605-607 ◽  
Author(s):  
J. M. Pettifor ◽  
F. P. Ross ◽  
J. Wang
Keyword(s):  
Protein Binding ◽  
Binding Assay ◽  
Specific Binding ◽  
Normal Children ◽  
Bovine Albumin ◽  
Hydroxyvitamin D ◽  
Protein Binding Assay ◽  
Competitive Protein Binding Assay ◽  
25 Hydroxyvitamin D ◽  
Competitive Protein Binding

1. We describe a modified competitive protein-binding assay for 25-hydroxyvitamin D, which does not require the prior separation of vitamin D from 25-hydroxyvitamin D. 2. Bovine albumin was used in the buffer, at a concentration of 1 mg/ml, as a protein stabilizer and lipid solubilizer. Bovine albumin from different manufacturers produced very different non-specific binding of 25-[3H]hydroxycholecalciferol (ranging from 3·3% to 58·9%). 3. The mean concentration of 25-hydroxyvitamin D in sera from normal children in Johannesburg was 76·75 ± 24·75 μnol/ml (30·7 ± 9·9 ng/ml).

scholarly journals A Simple Micromethod for 25-Hydroxyvitamin D Estimation

1979 ◽  
Vol 16 (1-6) ◽  
pp. 106-110 ◽  
Author(s):  
Angela Fairney ◽  
C. Turner ◽  
S. Hanson ◽  
Mary Zambon
Keyword(s):  
Binding Assay ◽  
Clinical Biochemistry ◽  
Simple Method ◽  
Hydroxyvitamin D ◽  
Protein Binding Assay ◽  
Competitive Protein Binding Assay ◽  
25 Hydroxyvitamin D ◽  
Competitive Protein Binding ◽  
Working Day ◽  
Clinical Biochemistry Laboratory

A quick and simple method for estimating 25-hydroxyvitamin D is described. It involves dichloromethane extraction followed by competitive protein-binding assay without chromatography. This assay can be performed on 100 μl of serum and enables 50 samples to be estimated in one working day. The simplicity, speed, and sample size of this method make it very suitable for use in a routine clinical biochemistry laboratory. It is particularly useful when calcium disorders secondary to abnormalities in vitamin D intake and metabolism are suspected.

A specific competitive protein binding assay for serum 24,25-dihydroxyvitamin d in normal children and patients with nephrotic syndrome

1980 ◽  
Vol 106 (2) ◽  
pp. 145-154 ◽  
Author(s):  
Tsunesuke Shimotsuji ◽  
Teisuke Hiejima ◽  
Yoshiki Seino ◽  
Kanji Yamaoka ◽  
Tsuneyasu Ishii ◽  
...  
Keyword(s):  
Nephrotic Syndrome ◽  
Protein Binding ◽  
Binding Assay ◽  
Normal Children ◽  
Dihydroxyvitamin D ◽  
Protein Binding Assay ◽  
Competitive Protein Binding Assay ◽  
Competitive Protein Binding

scholarly journals HPLC Method for 25-Hydroxyvitamin D Measurement: Comparison with Contemporary Assays

2006 ◽  
Vol 52 (6) ◽  
pp. 1120-1126 ◽  
Author(s):  
Gary L Lensmeyer ◽  
Donald A Wiebe ◽  
Neil Binkley ◽  
Marc K Drezner
Keyword(s):  
Binding Assay ◽  
Hplc Method ◽  
Hydroxyvitamin D ◽  
Protein Binding Assay ◽  
Liquid Chromatography Tandem Mass ◽  
Competitive Protein Binding Assay ◽  
25 Hydroxyvitamin D ◽  
Competitive Protein Binding ◽  
Functional Indicator ◽  
Comparison Data

Abstract Background: The concentration of 25-hydroxyvitamin D [25(OH)D] in serum has been designated the functional indicator of vitamin D (VitD) nutritional status. Unfortunately, variability among 25(OH)D assays limits clinician ability to monitor VitD status, supplementation, and toxicity. Methods: We developed an HPLC method that selectively measures 25-hydroxyvitamin D2 [25(OH)D2] and D3 [25(OH)D3] and compared this assay with a liquid chromatography–tandem mass spectrometry (LC-MS/MS) method, a competitive protein-binding assay (CPBA) on the Nichols Advantage™ platform, and an RIA from Diasorin. Results: For the new HPLC assay, between-run CVs were 2.6%–4.9% for 25(OH)D3 and 3.2%–13% for 25(OH)D2; recoveries were 95%–102%; and the assay was linear from 5 μg/L to at least 200 μg/L. Comparison data were as follows: for HPLC vs LC-MS/MS, y = 1.01x − 4.82 μg/L (Sy|x = 4.93 μg/L; r = 0.996) for 25(OH)D3, and y = 0.902x − 0.566 μg/L (Sy|x = 2.56 μg/L; r = 0.9965 for 25(OH)D2; for HPLC vs Diasorin RIA, y = 0.709x − 5.86 μg/L (Sy|x = 7.35 μg/L; r = 0.7509); and for HPLC vs Nichols Advantage CPBA, y = 1.00x − 3.60 μg/L (Sy|x = 32.7 μg/L; r = 0.6823). Conclusions: The new HPLC method is reliable, robust, and has advantages compared with the Nichols Advantage CPBA and the Diasorin RIA. The Nichols Advantage CPBA overestimated or underestimated 25(OH)D concentrations predicated on the prevailing metabolite present in patients’ sera.

Interlaboratory comparison of 25-hydroxyvitamin D determination.

1984 ◽  
Vol 30 (7) ◽  
pp. 1199-1204 ◽  
Author(s):  
E Mayer ◽  
H Schmidt-Gayk
Keyword(s):  
Interlaboratory Comparison ◽  
Binding Assay ◽  
Reference Method ◽  
Serum Samples ◽  
Hydroxyvitamin D ◽  
Protein Binding Assay ◽  
Competitive Protein Binding Assay ◽  
25 Hydroxyvitamin D ◽  
Competitive Protein Binding

Abstract This interlaboratory study on determination of 25-hydroxyvitamin D (25-OH-D) in serum involved 15 laboratories in eight European countries. All could distinguish between normal (50 +/- 31 nmol/L, mean +/- SD) and grossly increased concentrations, but for eight laboratories the results for serum samples with low and normal 25-OH-D content overlapped. In general, values were well reproducible, but interlaboratory variation in 25-OH-D measurement was large, 24,25(OH)2D3 interfering in most of the assays. We present evidence in favor of chromatography before assay, as opposed to nonchromatographic methods. Liquid chromatography with ultraviolet detection for quantifying 25-OH-D2 and 25-OH-D3 appears to be an appropriate reference method, whereas competitive protein binding assay is the method of choice for routine determinations. Control sera with subnormal, normal, and above-normal concentrations of 25-OH-D3 are needed for use in standardization of 25-OH-D assays.

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