HPRT Assay Kit

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 by tracking the production of IMP, which is oxidized by recombinant IMP dehydrogenase. This process leads to the formation of NADH2, continuously monitored spectrophotometrically at 340 nm for real-time measurements.

HPRT enzyme Molecular Structure — PRECICE® HPRT Assay Kit

#K0709-01-2

#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 Detection Range

Detect HPRT activity from 6.75 nmol/hour/ml to 340 nmol/hour/ml, enabling the characterization of complete and partial deficiencies.

Accurate

Measurement imprecisions: 1.5% (within-run), 5% (between-day), and 6.5% (total).

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.

HPRT Activity

Cells	PRECICE® HPRT Assay Kit	Published Data	References
RBC specimen 1 (n=4)	76.94±2.5 nmol/h/mg of Hgb	80–120 nmol/h/mg of hemoglobin	4
RBC specimen 2 (n=12)	78.03±5.52 nmol/h/mg of Hgb
RBC specimen 3 (n=4)	88.7±3.41 nmol/h/mg of Hgb
PBMC specimen 1 (n=4)	159.91±3.60 nmol/h/mg of protein 19.7±0.6 nmol/h/10⁶ PBMC	343±18 nmol/h/mg of protein 10.2–18.0 nmol/h/10⁶ PBMC	4
PBMC specimen 2 (n=4)	152.30±4.94 nmol/h/mg of protein 21.3±0.42 nmol/h/10⁶ PBMC	343±18 nmol/h/mg of protein 10.2–18.0 nmol/h/10⁶ PBMC	5
Human Dermal Fibroblasts (Invitrogen)	70.91±2.67 nmol/h/mg of protein	81–127 nmol/h/mg of protein	6
W20-17 cells (ATCC)	91.95±4.49 nmol/h/mg of protein	81–127 nmol/h/mg of protein	6

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.