Dataset Information

Development of a Chemiluminescence Immunoassay for Quantification of 25-Hydroxyvitamin D in Human Serum.

ABSTRACT: In this study, a chemiluminescence immunoassay (CLIA) for human serum 25-hydroxyvitamin D (25(OH)D) was established by a competition model. In serum, more than 99% of total circulating 25(OH)D binds to protein and less than 1% of 25(OH)D is in free form (Jassil et al., 2017). Before measuring concentration of 25(OH)D in serum, a releasing procedure should be conducted. A new reagent is used to release binding 25(OH)D to free form. Streptavidin (SA) was labeled to magnetic beads by a 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide/N-hydroxysuccinimide (EDC/NHS) method. Biotinylated VD was used as a competitor of 25(OH)D in samples. Anti-VD antibody (aby) was labeled to horseradish peroxidase (HRP) by EDC to react with 25(OH)D and biotinylated-VD molecules. The pretreated samples or standards were added into the reaction tube with biotin-VD and anti-VD aby-HRP, free 25(OH)D in the sample competes with biotinylated VD for binding to anti-VD aby-HRP, an SA-labeled magnetic particle is added to isolate the signal-generating complex, and the signal is inversely proportional to the 25(OH)D concentration in the sample. The method established shows good thermostability and performance. The limitation of detection (LoD) is 1.43?ng/mL. The intra-assay coefficient of variation (CV) is 3.66%-6.56%, the interassay CV is 4.19%-7.01%, and the recovery rate is 93.22%-107.99%. Cross-reactivity (CR) was remarkably low with vitamin D2, vitamin D3, 1, 25-dihydroxyvitamin D3, and 1, 25-dihydroxyvitamin D2. At the same time, the cross-reaction values with 25-hydroxyvitamin D2 and 25-hydroxyvitamin D3 were 97% and 100%, respectively. The developed method shows good correlation with the total VD product from Roche and DiaSorin. 1096 clinical patient samples were measured with developed reagent kit in this study. 7 types of disease were involved, and the concentration of 25(OH)D is less than 30?ng/mL in 94.98% of patients.

SUBMITTER: Han S

PROVIDER: S-EPMC7416287 | biostudies-literature | 2020

REPOSITORIES: biostudies-literature

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Development of a Chemiluminescence Immunoassay for Quantification of 25-Hydroxyvitamin D in Human Serum.

Han Shuang S Qiu Wuxian W Zhang Junlan J Bai Zhonghu Z Tong Xiao X

Journal of analytical methods in chemistry 20200801

In this study, a chemiluminescence immunoassay (CLIA) for human serum 25-hydroxyvitamin D (25(OH)D) was established by a competition model. In serum, more than 99% of total circulating 25(OH)D binds to protein and less than 1% of 25(OH)D is in free form (Jassil et al., 2017). Before measuring concentration of 25(OH)D in serum, a releasing procedure should be conducted. A new reagent is used to release binding 25(OH)D to free form. Streptavidin (SA) was labeled to magnetic beads by a 1-ethyl-3-(3 ...[more]

PMID: 32802551

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Dataset Information

Development of a Chemiluminescence Immunoassay for Quantification of 25-Hydroxyvitamin D in Human Serum.

Publications

Development of a Chemiluminescence Immunoassay for Quantification of 25-Hydroxyvitamin D in Human Serum.

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