Thermo Scientific Pierce PNPP Substrates are powders, tablets, solutions and complete kits of p-nitrophenyl phosphate, a colorimetric, soluble substrate of alkaline phosphatase for use in ELISA applications.
PNPP (p-Nitrophenyl Phosphate, Disodium Salt) is a widely used substrate for detecting alkaline phosphatase in ELISA applications. When alkaline phosphatase and PNPP are reacted, a yellow water-soluble reaction product is formed. This reaction product absorbs light at 405nm. The substrate is offered in several formats.
- One-component alkaline phosphatase (AP) substrate
- Absorbance maximum of 405nm
- Develops a yellow hue in solution upon reaction with AP
- Safer to use — contains a non-mercury, non-azide preservative
PNPP is available either as a crystalline powder or 5mg tablets. Also available is the Phosphatase Substrate Kit, which contains PNPP tablets and a Diethanolamine Buffer to yield more than one liter of substrate. The Diethanolamine Substrate Buffer (Part No. 34064) is also provided individually as a 5X concentrate. The Pierce formulation has an optimal and consistent pH and it is stable even at a 1X concentration.
1-Step PNPP is also available. Until now, the method for preparing PNPP solutions required dissolving the powder or tablets in buffer and then diluting the solution to the desired concentration. Substrate instability made it necessary to prepare it on a daily basis. Pierce 1-Step PNPP circumvents this time-consuming and variable introducing step by providing a single component PNPP substrate. This substrate is stable for 12 months at 2°C-8°C and can be stopped with conventional methods.
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Assay development and ELISA technical guide
ELISA substrate selection guide