Principle of Assay
Total Cholesterol ELISA Kit (A326911) employs the competitive enzyme immunoassay technique for the quantitative measurement of Total Cholesterol in serum, plasma, cell culture supernatant, cell or tissue lysate, and other liquid samples. The 96-well microtiter plate has been pre-coated with Total Cholesterol antigen. During the incubation, Total Cholesterol present in the samples or standards competes with the fixed amount of immobilized Total Cholesterol for binding sites on the Biotinylated Anti-Total Cholesterol Antibody. The more Total Cholesterol present in a sample or standard, the less Biotinylated Anti-Total Cholesterol Antibody that binds to the plate. Following incubation, unbound Biotinylated Anti-Total Cholesterol 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 inversely proportional to the amount of Total Cholesterol present in each sample or standard. The concentration of Total Cholesterol can then be calculated by reading the O.D. absorbance at 450nm in a microplate reader and referring to the standard curve.
