Description
Principle of the assay: AFA ELISA kit applies the competitive enzyme immunoassay technique utilizing a monoclonal AFA antigen and an AFA-HRP conjugate. The assay sample and buffer are incubated together with AFA-HRP conjugate in pre-coated plate for one hour. After the incubation period, the wells are decanted and washed five times. The wells are then incubated with a substrate for HRP enzyme. The product of the enzyme-substrate reaction forms a blue colored complex. Finally, a stop solution is added to stop the reaction, which will then turn the solution yellow. The intensity of color is measured spectrophotometrically at 450nm in a microplate reader. The intensity of the color is inversely proportional to the AFA concentration since AFA from samples and AFA-HRP conjugate compete for the AFA antigen binding site. Since the number of sites is limited, as more sites are occupied by AFA from the sample, fewer sites are left to bind AFA-HRP conjugate. A standard curve is plotted relating the intensity of the color (O.D.) to the concentration of standards. The AFA concentration in each sample is interpolated from this standard curve