Principle of Assay
Universal Free Thyroxine ELISA Kit (A335451) is a Competitive ELISA for the quantitative measurement of universal Free Thyroxine in serum, plasma, cell culture supernatant, cell or tissue lysate, and other liquid samples. An antibody specific for Free Thyroxine has been pre-coated onto a 96-well microtiter plate. During the incubation, Free Thyroxine in the sample or standard competes with a fixed amount of Biotinylated-Free Thyroxine. Following incubation, Biotinylated-Free Thyroxine and free Free Thyroxine in the sample or standard are washed from the plate. HRP-Streptavidin (SABC) is then added to each well and incubated. Unbound SABC is washed away with wash buffer. TMB substrate solution is added and after incubation the enzyme-substrate reaction is terminated by the addition of sulfuric acid stop solution. The colour change is measured spectrophotometrically at a wavelength of 450nm and the concentration of Free Thyroxine in the samples is then determined by comparing the O.D. 450nm of the samples to the standard curve. O.D. 450nm is inversely proportional to the concentration of Free Thyroxine in the sample.
