Kainos

The FGF-23 ELISA Kit is a two-site enzyme-linked immunosorbent assay kit for the measurement of FGF-23 in serum. Two specific murine monoclonal antibodies bind to full-length FGF-23. One antibody is immobilized onto the microtiter plate well for capture. The other antibody is conjugated to HRP(horseradish peroxidase) for detection.

In first reaction, a sample containing FGF-23 is incubated with the immobilized antibody in a microtiter well. FGF-23 in the sample is captured with the antibody. At the end of this reaction, the well is washed to remove unbound FGF-23 and other components.

In second reaction, this immobilized FGF-23 is incubated with HRP labeled antibody to form a “sandwich”complex :

Anti FGF-23 antibody - N-terminal　FGF-23　C-terminal - HRP labeled anti FGF-23 antibody

At the end of this reaction, the well is washed to remove unbound components.

In enzyme reaction, the sandwich complex immobilized on the well is incubated with a substrate solution and then measured by a spectrophotometric microtiter plate reader. The enzymatic activity of the complex bound to the well is directly proportional to the amount of FGF-23 in the sample.

A standard curve is generated by plotting the absorbance versus the each concentration of FGF-23 standard. The concentration of FGF-23 in the sample is determined from this curve.

This kit captures ADAMTS13-cleaved products using a sandwich method with the anti-GST mouse monoclonal antibody and the peroxidase-conjugated mouse monoclonal anti-N10 antibody. First, the anti-GST mouse monoclonal antibody immobilized on to the microplate reacts with the VWF73 substrate（GST-VWF73-His）. The sample is then applied to the microplate, on which ADAMTS13 cleaves the VWF73 substrate. By applying the mouse monoclonal anti-N10 antibody conjugated with horseradish peroxidase（HRP conjugated antibody）, the cleavage product is sandwiched between the two antibodies. Because the amount of the cleavage products depends on the ADAMTS13 activity, the amount of the enzyme-labeled antibodies that bind to the cleavage products also reflect the level of ADAMTS13 activity. Therefore, the plasma ADAMTS13 activity can be determined by colorimetrically measuring the amount of detached oxidized （colored）TMBZ using urea hydrogen peroxide（H2O2）and 3,3’,5,5’-Tetramethyl- benzidine（TMBZ）as the substrates.