Non radioactive HPRT assay
Including active human HPRT enzyme
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.
#REF | SIZE | PRICE | |
---|---|---|---|
#K0709-01-2 | 24 analyses (8 samples in triplicate) with HPRT enzyme | 330.00 € | Inquiry |
Updated on October 2nd, 2024.
Kit is provided in stable lyophilized form and shipped without dry ice
Download HPRT assay Protocol (User manual)
Download Material Safety Data Sheet (MSDS)
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 d eficiencies
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 hours.
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 procedure.
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.
Scientific Articles citing PRECICE® HPRT Assay kit from NOVOCIB:
-
Ablation of Atp5if1 impairs metabolic reprogramming and proliferation of T lymphocytes and compromises mouse survival
I. Romero-Carraminana, S. Dominguez-Zorita, P.B. Esparza-Molto, J. M. Cuezva (2024) iScience 27, 109863 -
Electron transport chain inhibition increases cellular dependence on purine transport and salvage
Z. Wu, D. Bezwada, F. Cai, J. Garcia-Bermudez, M. Ni, R.J. DeBerardinis, Cell Metab. 2024 Jun 7:S1550-4131(24)00190-6 -
Therapeutic gene correction for Lesch-Nyhan syndrome using CRISPR-mediated base and prime editing (2023)
G. Jang , H. Rim Shin, H.-S. Do , J. Kweon , S. Hwang , S. Kim, S. Hee Heo, Y. Kim, B. Lee Molecular Therapy - Nucleic Acids ; 31: 586-595. -
Rescuing compounds for Lesch-Nyhan disease identified using stem cell-based phenotypic screening (2020)
V. Ruillier, J. Tournois, C. Boissart, M. Lasbareilles, G. Mahé, L. Chatrousse, M. Cailleret, M. Peschanski, A. Benchoua JCI Insight 5(4): e132094. -
Hypoxanthine-guanine phosphoribosyltransferase and inosine 5'-monophosphate dehydrogenase activities in three mammalian species: aquatic (Mirounga angustirostris), semi-aquatic (Lontra longicaudis annectens) and terrestrial (2015)
M. Barjau Pérez-Milicua, T. Zenteno-Savín, D.E. Crocker, J.P. Gallo-Reynoso Front Physiol.; 6: 212. -
Prolonged fasting increases purine recycling in post-weaned northern elephant seals
José Guadalupe Soñanez-Organis, José Pablo Vázquez-Medina, Tania Zenteno-Savín, Andres Aguilar, Daniel E. Crocker, Rudy M. Ortiz J Exp Biol (2012) 215 (9): 1448-1455.