Abstract: In this paper, a chemiluminescent immunoassay (CLIA) kit for quantitative assay of 25-hydroxyvitamin D（25-OHD）in serum was developed and evaluated. The kit is established as follows: first, 25-hydroxyvitamin D monoclonal antibodies was coated with the 96 well microplate, then the serum sample and the release reagent were added into the microplate. After incubation, the 25-OHD of the sample was fully released and combined with the 25-OHD antibody coated on the microplate. HRP conjugate (mixture of biotinylated 25-OHD and streptavidin-labeled HRP ) compete with the 25-OHD released from sample to antibodies. After washing and adding the CLLIA substrate, the relative luminous intensity was read with chemiluminescent immunoassay analyzer. The minimum detection limit of the kit is 4.9 ng.mL-1, with the bias of less than 10％. The intra-assay and inter-assay coefficients of variation were less than 10%, and 15%, respectively. Its preserving duration could be as long as 12 months at 2-8℃. Two hundred clinical serum specimens were tested with this kit and a foreign made serum ferritin kit, the correlation coefficient was 0.9717. 25-hydroxy vitamin D (CLIA) quantification detection kit was successfully prepared, and all the technical parameters reached the industrial standards. The developed kit is sensitive, stable, fast and simple.

Key words: 25-hydroxyvitamin D, chemiluminescent immunoassay, kit