Mouse TNF alpha ELISA Kit (A311790) employs the sandwich enzyme immunoassay technique for the quantitative measurement of mouse TNF alpha in An antibody specific for TNF alpha has been pre-coated onto a 96-well microtiter plate. The standards and test samples are added into the wells and the TNF alpha present in each sample is bound to the wells by the immobilized antibody. Biotinylated Anti-TNF alpha Antibody, which also binds the TNF alpha present in each sample, and Streptavidin-HRP, which binds the Biotinylated Anti-TNF alpha Antibody, are added and the microtiter plate is incubated. Following incubation, unbound Biotinylated Anti-TNF alpha 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 TNF alpha 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 TNF alpha 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.