For quantitative determination of phosphatase activity.

• The detection limit is generally 3 ng phosphatase or below
• No wash or reagent transfer steps are involved
• All reagents are compatible with high-throughput liquid handling instruments

p-Nitrophenyl phosphate (pNPP) is a chromogenic substrate for most phosphatases such as alkaline phosphatases, acid phosphatases, protein tyrosine phosphatases and serine/threonine phosphatases. The reaction yields para-nitrophenol, which becomes an intense yellow soluble product under alkaline conditions and can be conveniently measured at 405 nm on a spectrophotometer. This homogeneous "mix-and-measure" assay involves simply adding a single reagent to the phosphatase and measuring the product formation on an absorbance reader. The assay can be conveniently performed in cuvettes, tubes or multi-well plates at either room temperature or 37°C.