Principle of Assay
Bovine ACTH ELISA Kit (A7893) employs the competitive enzyme immunoassay technique for the quantitative measurement of bovine ACTH in serum, plasma, tissue homogenates, cell lysates, cell culture supernates or other biological fluids. The 96-well microtiter plate has been pre-coated with ACTH antigen. During the incubation, ACTH present in the samples or standards competes with the fixed amount of immobilized ACTH for binding sites on the Biotinylated Anti-ACTH Antibody. The more ACTH present in a sample or standard, the less Biotinylated Anti-ACTH Antibody that binds to the plate. Following incubation, unbound Biotinylated Anti-ACTH Antibody is removed by washing, and an HRP-Avidin 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 inversely proportional to the amount of ACTH present in each sample or standard. The concentration of ACTH can then be calculated by reading the O.D. absorbance at 450nm in a microplate reader and referring to the standard curve.
