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® HPRT Assay Kit

Ref. #K0709-01-2
Size Price*
24 analyses (8 samples in triplicate) with HPRT enzyme** € 330.00

* Pricing updated August 28th, 2015. ** As positive control.
Kit is provided in stable lyophilized form and shipped without dry ice

To buy HPRT Assay Kit click here or ask for Quotation


Download HPRT assay Protocol  
(User Manual)

Non radioactive HPRT assay

PRECICE® HPRT Assay kit provides the first non radioactive protocol for measurement of HPRT activity in a convenient 96-well plate format.

Hypoxanthine-guanine phosphoribosyltransferase (HPRT, EC 2.4.2.8), a key enzyme of the purine salvage pathway, is encoded by highly variable HPRT1 gene. More than 300 mutations in HPRT1 gene associated with genetic disorders in humans have been described leading to partial or complete deficiency of HPRT enzyme. Complete HPRT deficiency is a rare human genetic disorder, also known as Lesch-Nyhan syndrom. In some cases, gout, more frequent purine disorder, can also be secondary to HPRT deficiency (Ceballos-Picot I. et al 2010). High variability of HPRT1 gene complicates genetic testing. Currently, HPRT enzyme functions are tested in red blood cell lysates in a chromatographic assay using radiolabeled 14C-hypoxanthine (Cartier P. et al, 1968).

Principle
HPRT activity is measured as a rate of production of IMP, which is oxidized by recombinant IMP dehydrogenase with concomitant formation of NADH2. The formation of NADH2 is continuously monitored spectrophotometrically at 340nm.

Wide linearity range

    The detection of as low as 6.75 nmol/hour/ml up to 340 nmol/hour/ml of HPRT activity enables the characterization of complete and partial deficiencies

Accurate

    Within-run,  between-day and total measurement imprecisions of assay run in are 1.5%, 5% and 6.5%

Fast

    Simultaneous analysis of up to 15 samples in triplicate in 2 h

Convenient

    No sample preparation nor prior inactivation of cellular 5’-nucleotidase required. Cell lysates are directly used for continuous monitoring of HPRT activity

Validated

    Specific activities of HPRT in lysates of erythrocytes, PBMC and cultured cells measured by PRECICE® HPRT Assay kit are comparable to those obtained previously by radiochemical procedureHPRT activity assay

 



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Calibration curve of IMPDH-based PRECICE® HPRT Assay
The rate of the increase in absorbance at 340nm per hour as a function of the concentration of human recombinant HPRT enzyme (NovoCIB, ref. E-Nov-9). The changes in absorbance corresponding to 3 different control hemolysates diluted in complete reaction mixture to final hemoglobin concentration 1mg/ml (n=4) are shown by filled squares, filled triangles and filled circles. The insert shows a linear correlation observed in whole range of 21ng/ml up to 1.5µg/ml of recombinant HPRT; the units are the same.
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References
1. J.G. Soñanez-Organis, J.P. Vázquez-Medina, T. Zenteno-Savín, A. Aguilar, D.E. Crocker, R.M. Ortiz (2012) Prolonged fasting increases purine recycling in post-weaned northern elephant seals. J Exp Biol. 215:1448-55.
2. Z.M. Zaïr, G.E. Johnson The Applicable Use of the HPRT Gene Mutation Assay as a Practical Tool in Mutagenesis and DNA Repair Studies (2014) in "Genotoxicity and DNA Repair : A Practical Approach": 185-197
3. M. Barjau Pérez-Milicua M, T. Zenteno-Savín, D.E. Crocker , J.P. Gallo-Reynoso (2015) Hypoxanthine-guanine phosphoribosyltransferase and inosine 5'-monophosphate dehydrogenase activities in three mammalian species: aquatic (Mirounga angustirostris), semi-aquatic (Lontra longicaudis annectens) and terrestrial (Sus scrofa). Front Physiol. 6:212
4. R.I. López-Cruz , D.E. Crocker, Gaxiola-R. Robles, J.A. Bernal, R.A. Real-Valle, O. Lugo-Lugo, T. Zenteno-Savín (2016). Plasma Hypoxanthine-Guanine Phosphoribosyl Transferase Activity in Bottlenose Dolphins Contributes to Avoiding Accumulation of Non-recyclable Purines. Front Physiol. 2016 Jun 8;7:213.


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Related Links
HGPRT enzymeBacterial Recombinant IMPDH
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