Description
Principle of the Assay: This assay employs the quantitative sandwich enzyme immunoassay technique. Antibody specific for NID has been pre-coated onto a microplate. Standards and samples are pipetted into the wells and any NID present is bound by the immobilized antibody. After removing any unbound substances, a biotin-conjugated antibody specific for NID is The microtiter plate provided in this kit has been pre-coated with an antibody specific to NP-Y. Standards or samples are then added to the appropriate microtiter plate wells with a biotin-conjugated antibody preparation specific for NP-Y and Avidin conjugated to Horseradish Peroxidase (HRP) is added to each microplate well and incubated. Then a TMB (3,3',5,5' tetramethyl-benzidine) substrate solution is added to each well. Only those wells that contain NP-Y, biotin-conjugated antibody and enzyme-conjugated Avidin will exhibit a change in color. The enzyme-substrate reaction is terminated by the addition of a sulphuric acid solution and the color change is measured spectrophotometrically at a wavelength of 450 nm +/- 2 nm. The concentration of NP-Y in the samples is then determined by comparing the O.D. of the samples to the standard curve.added to the wells. After washing, avidin conjugated Horseradish Peroxidase (HRP) is added to the wells. Following a wash to remove any unbound avidin-enzyme reagent, a substrate solution is added to the wells and color develops in proportion to the amount of NID bound in the initial step. The color development is stopped and the intensity of the color is measured