Solid-phase Competitive ELISA designed for the quantitative determination of Porcine citrullinated histone H3 (cit H3).
cit H3 ELISA kit applies the competitive enzyme immunoassay technique utilizing an
anti-cit H3 antibody and an cit H3-HRP conjugate. The assay sample and buffer are
incubated together with cit H3-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 cit H3 concentration since cit H3 from samples and cit H3-HRP conjugate compete for the anti-cit H3 antibody binding site. Since the
number of sites is limited, as more sites are occupied by cit H3 from the sample,
fewer sites are left to bind cit H3-HRP conjugate. A standard curve is plotted relating the intensity of the color (O.D.) to the concentration of standards. The cit H3
concentration in each sample is interpolated from this standard curve.