Human ACTH ELISA Kit (A311984) employs the sandwich enzyme immunoassay technique for the quantitative measurement of human ACTH in serum, plasma or other biological fluids. An antibody specific for ACTH has been pre-coated onto a 96-well microtiter plate. The standards and test samples are added into the wells and the ACTH present in each sample is bound to the wells by the immobilized antibody. Biotinylated Anti-ACTH Antibody, which also binds the ACTH present in each sample, and Streptavidin-HRP, which binds the Biotinylated Anti-ACTH Antibody, are added and the microtiter plate is incubated. Following incubation, unbound Biotinylated Anti-ACTH Antibody and unbound Streptavidin-HRP are removed by washing, and two substrate solutions are added to the wells. Color develops in proportion to the amount of ACTH captured in each well. The color development is stopped by addition of stop solution which changes the color from blue to yellow and the intensity of the color is then measured. The concentration of ACTH in the samples can then be calculated by reading the O.D. absorbance at 450nm in a microplate reader and referring to the standard curve.