Application Note A continuous spectrophotometric assay for rapid measurement of hypoxanthine-guanine phosphoribosyltransferase (HPRT) activity in cell lysates

Hypoxanthine-guanine phosphoribosyltransferase (HPRT) is a central enzyme in the purine salvage pathway. The sequence of HPRT1 gene is one of the best characterized because of its wide use as a biomarker of genotoxicity and the clinical significance of HPRT-deficiency for human health. The current methods of functional characterisation of HPRT are based on the end-point chromatographic quantification of IMP synthesized by HPRT. PRECICE HPRT Assay Kit is a spectrophotometric, enzymatic assay that allows a continuous monitoring of IMP formation in microplate format. In the presence of recombinant IMP-dehydrogenase, IMP is oxidized to XMP with the simultaneous formation of NADH2, measurable at 340nm. The specific activities of HPRT in lysates of erythrocytes, PBMC and cultured fibroblasts measured by this method are comparable to those previously determined by radiochemical procedures. PRECICE HPRT Assay Kit allows the detection of HPRT activity as low as 6.75nmol/hour/ml with a total imprecision of 6.5%.