About Glucose-6-Phosphate Dehydrogenase:
GLUCOSE-6-PHOSPHATE DEHYDROGENASE (G6PDH) is a cytosolic enzyme in the pentose phosphate pathway which supplies reducing energy to cells by maintaining the level of the co-enzyme nicotinamide adenine dinucleotide phosphate (NADPH). G6PDH reduces nicotinamide adenine dinucleotide phosphate (NADP) to NADPH while oxidizing glucose-6-phosphate (G6P). Humans with a genetic deficiency of G6PDH are predisposed to non-immune hemolytic anemia. Recently, studies have found G6PDH plays a critical role in survival, proliferation, and metastasis of cancer cells. Therefore, inhibitors of the enzyme are attractive candidates for new cancer therapeutics.
How our G6PDH Assay Kits Work:
Assay Genie's non-radioactive, colorimetric G6PDH assay is based on the reduction of the tetrazolium salt MTT in a NADPH-coupled enzymatic reaction to a reduced form of MTT which exhibits an absorption maximum at 565 nm. The increase in absorbance at 565 nm is proportional to the enzyme activity. The percent inhibition of a test compound can be determined by comparing the activity of G6PDH treated with a test compound to the activity of untreated G6PDH.
- Safe. Non-radioactive assay.
- Rapid and reliable. Can be completed in 30 minutes and no 37°C heater is needed.
|Application:||HTS for inhibitor screening and evaluation of glucose-6-phosphate dehydrogenase inhibitors.|
|Detection limit:||The following protocol is optimized for L. Mesenteroides G6PDH. Dilute purified G6PDH to 0.0003 U/µL using dH2O. If another species is being analyzed, we recommend that you experimentally determine the Km and then adjust the volume of substrate in the Working reagent so that the final concentration of the substrate in the 100 µL reaction is near the Km.|
|Shelf Life:||6 months|
Materials required, but not provided:
Purified G6PDH (e.g. Calzyme Cat# 078A0020) and if desired a control Zn2+ inhibitor (e.g. Zn(NO3)2 · 6H2O, Sigma Cat# 228737). Pipetting devices and accessories (e.g. multi-channel pipettor), clear flat bottom 96-well plates (e.g. VWR cat# 82050-760), and plate reader.