Description
Principle of the assay: NFKB P65 ELISA kit applies the competitive enzyme immunoassay technique utilizing a monoclonal anti-NFKB P65 antibody and an NFKB P65-HRP conjugate. The assay sample and buffer are incubated together with NFKB P65-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 NFKB P65 concentration since NFKB P65 from samples and NFKB P65-HRP conjugate compete for the anti-NFKB P65 antibody binding site. Since the number of sites is limited, as more sites are occupied by NFKB P65 from the sample, fewer sites are left to bind NFKB P65-HRP conjugate. A standard curve is plotted relating the intensity of the color (O.D.) to the concentration of standards. The NFKB P65 concentration in each sample is interpolated from this standard curve