Description
Principle of the Assay: AKA ELISA kit applies the competitive enzyme immunoassay technique utilizing keratin antigen and an AKA-HRP conjugate. The assay sample and buffer are incubated together with AKA-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 AKA concentration since AKA from samples and AKA-HRP conjugate compete for the keratin antigen binding site. Since the number of sites is limited, as more sites are occupied by AKA from the sample, fewer sites are left to bind AKA-HRP conjugate. A standard curve is plotted relating the intensity of the color (O.D.) to the concentration of standards. The AKA concentration in each sample is interpolated from this standard curve