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Alkaline Phosphatase (AP) is purified from bovine intestinal mucosa by chromatography and filtration. AP is a zinc metaloenzyme for catalyzing the hydrolysis of an orthophosporic monoester to yield an alcohol and an orthophosphate. AP can be coupled to antibodies resulting in conjugates, used in ELISA, membrane & IHC applications, biosensors etc.
In ELISA, the main colorimetric substrate for AP is p-nitrophenyl phosphate (pNPP) which yields a yellow reaction product. Absorbance can be quantified by reading the OD at 405 nm. The kinetic reaction is terminated by the addition of a stop-solution.
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