Award
Dr Larissa Balakireva, CEO & Founder of NovoCIB, was awarded with the Trophy of
"Femmes en Or 2011, Femme de l'Innovation"
in September 2011
.

PRECICE® Phosphatase Assay Kit

Ref. #K0211-01
Size Price*
1 plate (96 assays) € 410.00

* Pricing updated April 16th, 2014.

To buy Phosphatase Assay Kit click here or ask for Quotation
Kit is provided in stable lyophilized form and shipped without dry ice

Download our User Manual In vitro phosphatase assay

Continuous Phosphatase Assay

PRECICE® Phosphatase Assay Kit developed by NovoCIB is a continuous spectrophotometric enzyme assay to monitor activity of any enzymes liberating inorganic phosphate (Pi) during enzymatic reactions. The kit can be used for:

  • Phosphatase activity assay
  • Alkaline phosphatase activity
  • Protein phosphatases activity assay
  • ATPase assay
  • GTPases assay
  • Helicases assay.

Principle
The release of inorganic phosphate is measured through coupling to PNP-XDH enzymatic system. In the presence of inorganic phosphate, PNP (purine nucleoside phosphorylase) catalyzes inosine phosphorolysis leading to the formation of hypoxanthine and P-ribose. XDH enzyme catalyzes irreversible oxidation of hypoxanthine to uric acid with simultaneous reduction of NAD to NADH2, measurable by absorbance at 340nm. The formation of NADH2 is continuously monitored spectrophotometrically at 340nm. The assay is linear in 5-300µM range.


Key Features:

Continuous

    • Activity is measured through continuous montoring of absorbance at 340nm

Convenient

    • Tolerant to contamination of ware and reagents by inorganic phosphate
    • Homogenous
    • Non radioactive
    • Wide pH range
    • Compatible with reducing agents

Quantitative

    • Wide linearity range (5-300µM)

High-Throughput Analysis

    • The assay is performed in standard multi-well microplates
    • Readout is peformed with plate reader fitted with 340nm filter

Specific

    • Only inorganic phosphate is quantified
 

V mouse over to enlarge
Calibration curves of PRECICE® Phosphate Release Assay done in phosphate-free solutions (left) and in solutions "contaminated" by 100µM inorganic phosphate (right).
The presence of contaminating phosphates does not affect the linearity of the assay.

References
M.R. Webb MR, J.L. Hunter JL. (1992): Interaction of GTPase-activating protein with p21ras, measured using a continuous assay for inorganic phosphate release. Biochem J. 287 ( Pt 2):555-9.
T. Nishino et al. (2005): Mechanism of the conversion of xanthine dehydrogenase to xanthine oxidase: identification of the two cysteine disulfide bonds and crystal structure of a non-convertible rat liver xanthine dehydrogenase mutant. J Biol Chem. 280(26):24888-94.
T. Nishino, T. Nishino. (1997): The conversion from the dehydrogenase type to the oxidase type of rat liver xanthine dehydrogenase by modification of cysteine residues with fluorodinitrobenzene. J Biol Chem. 272(47):29859-64.
Harrison R. (2002): Structure and function of xanthine oxidoreductase: where are we now? Free Radic Biol Med. 33(6):774-97. C. Enroth et al (2000) Crystal structures of bovine milk xanthine dehydrogenase and xanthine oxidase: Structure-based mechanism of conversion . PNAS 97(20): 10723–10728.

Download our User Manual 


Related Links
• Xanthine dehydrogenase
• PNP enzyme		 • Phosphate Assay Kit
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