Principle of Assay
Human HE4 ELISA Kit (A330017) employs the sandwich enzyme immunoassay technique for the quantitative measurement of human HE4 in serum, plasma, cell culture supernates, cell lysates, tissue homogenates and other biological fluids. An antibody specific for HE4 has been pre-coated onto a 96-well microtiter plate. The standards and test samples are added into the wells and the HE4 present in each sample is bound to the wells by the immobilized antibody. Following incubation, the wells are washed and then incubated with Biotinylated Anti-HE4 Antibody, which binds the captured HE4 present in each well. Following incubation, unbound biotinylated detection antibody is removed by washing, and an HRP-Streptavidin conjugate is added to the wells and the microtiter plate is incubated. Following incubation and washing, TMB substrate solution is then used to visualize the HRP enzymatic reaction by catalysis to produce a blue-coloured product that changes to yellow after addition of acidic stop solution. The density of yellow is proportional to the amount of HE4 captured in each well. The concentration of HE4 can then be calculated by reading the O.D. absorbance at 450nm in a microplate reader and referring to the standard curve.
