p-nitropenyl phosphate (PNPP) substrate tabs and Buffer for ELISA
pNPP is a soluble substrate used in ELISA and for determination of alkaline and acid phosphatase in physiological fluids. For determination of acid phosphatase low pH buffer is required, not supplied.
10X buffer (25 mls) (For alkaline phosphatase).
Dissolve each tablet in 5-10 ml of 1X buffer.
ELISA PROCEDURE: Dissolve each tablet in 5-10 mls of 1X buffer. (dilute 10X buffer in distilled or deionized water). Add 5o microliters to each ELISA well and react for 30-60 mins. The reaction can be stopped with 50 microliters of 3 N NaOH and the yellow soluble color is read in spectrophotometer at 405 nm (ELISA reader)
Reacts with alkaline phosphatase ( buffer supplied) and acid phosphatase (buffer pH about 5 not supplied).
p-nitrophenol (MW 139.1) is the hydrolysis product of PNPP and can be used as a std. to determine enzyme activity.
It can also be used to make standard curve for quantitation of enzyme.