1,25 (OH)2 Vitamin D ELISA

The 1,25 (OH)2 Vitamin D ELISA Kit is intended for the quantitative determination of 1,25-dihydroxy vitamin D in serum and plasma.

For Research Use Only. Not for use in diagnostic procedures.

For Laboratory Professional Use Only.

Manufactured by Immundiagnostik AG.

Test Principle of the 1,25 (OH)2 Vitamin D ELISA

This assay utilizes a competitive enzyme immunoassay (EIA) technique with a selected monoclonal antibody recognizing 1,25-dihydroxy vitamin D. Standards, controls, and samples that are assayed for 1,25-dihydroxy vitamin D are incubated after the extraction step with the detection antibody.

The pre-incubated solution is then transferred to the microplate coated with 1,25-dihydroxy vitamin D. During this incubation step, 1,25-dihydroxy vitamin D in the sample and a fixed amount of 1,25-dihydroxy vitamin D bound to the microtiter well compete to bind the detection antibodies. Then a peroxidase-conjugated anti-mouse antibody is added into each microplate well, and a complex of 1,25-dihydroxy vitamin D – detection antibody – peroxidase conjugate is formed.

Tetramethylbenzidine (TMB) is used as a peroxidase substrate. Finally, an acidic stop solution is added to terminate the reaction, whereby the color changes from blue to yellow. The intensity of the yellow color is inversely proportional to the concentration of 1,25-dihydroxy vitamin D.

A dose-response curve of the absorbance unit (optical density, OD at 450 nm) vs. concentration is generated using the values obtained from the standard. The presence of 1,25-dihydroxy vitamin D in the samples is determined from this curve.