The Free 25OH Vitamin D ELISA is a competitive ELISA assay, based on patented monoclonal antibodies, that allows for the direct measurement of the free fraction of 25OH Vitamin D.
The assay is calibrated against a symmetric dialysis method. It shows excellent sensitivity, precision and performance characteristics, as performed according to the CLSI guidelines.
The assay is unique and exclusively available at DiaSource Immunoassays.
The Free 25(OH)Vitamin D ELISA is based on a two-step immunoassay procedure performed in a microtiterplate .
During the first incubation step free 25(OH)Vitamin D [25(OH)Vitamin D2 and D3] is bound to the anti-Vitamin D antibody coated on the wall of the microtiterplate. The in vivo equilibrium between free and bound 25(OH)Vitamin D is minimally disturbed. After washing, a fixed amount of biotinylated 25(OH)Vitamin D is added to each well. The non-bound biotinylated 25(OH)Vitamin D is removed by washing and a streptavidin peroxidase conjugate is added. In a next step TMB chromogenic substrate is added. Finally the reaction is stopped by addition of Stop reagent and the absorbance [A450nm] is measured using a plate spectrophotometer.
The concentration of free Vitamin D in the sample is inversely proportional to the absorbance in each sample well.
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