4-Nitrophenyl phosphate (p-Nitrophenyl phosphate) disodium is a non-proteinaceous chromogenic substrate for alkaline and acid phosphatases used in ELISA and conventional spectrophotometric assays.

Guide (The following is our recommended protocol. This protocol is only a guide and should be modified according to your specific needs).
1. Equilibrate 4-Nitrophenyl phosphate to room temperature before opening.
2. Dissolve 10 mg of 4-Nitrophenyl phosphate in 10 mL of 1X Diethanolamine (HY-W035903) Substrate Buffer.
Note: Diethanolamine is effective at concentrations from 10 mM to 1 M.
3. Add 100 μL of the 4-Nitrophenyl phosphate solution to each microplate well.
4. Incubate plate at room temperature for 30 minutes, or until sufficient color develops.
5. Add 50 μL of 2 N NaOH to each well to stop the reaction.
6. Measure the absorbance at 405 nm. Alternatively, monitor reaction in a kinetic ELISA reader.

MedChemExpress (MCE) has not independently confirmed the accuracy of these methods. They are for reference only.

* Please refer to the solubility information to select the appropriate solvent. Once prepared, please aliquot and store the solution to prevent product inactivation from repeated freeze-thaw cycles.
Storage method and period of stock solution: -80°C, 6 months; -20°C, 1 month (sealed storage, away from moisture and light). When stored at -80°C, please use it within 6 months. When stored at -20°C, please use it within 1 month.

* Note: If you choose water as the stock solution, please dilute it to the working solution, then filter and sterilize it with a 0.22 μm filter before use.

• [1]. Crowther JR, et al. Evaluation of the use of chromogenic and fluorogenic substrates in solid-phase enzyme linked immunosorbent assays (ELISA). Biologicals. 1990 Oct;18(4):331-6.  [Content Brief]