Key features and details
- Assay type: Enzyme activity
- Detection method: Colorimetric
- Platform: Microplate reader
- Assay time: 30 min
- Sample type: Adherent cells, Cell culture extracts, Cell Lysate, Suspension cells, Tissue Extracts, Tissue Homogenate, Tissue Lysate
Product nameNQO1 Activity Assay Kit
Intra-assay Sample n Mean SD CV% Overall 3 2.183% Inter-assay Sample n Mean SD CV% Overall 9 6.035%
Sample typeCell culture extracts, Adherent cells, Suspension cells, Tissue Extracts, Cell Lysate, Tissue Homogenate, Tissue Lysate
Assay typeEnzyme activity
Assay time0h 30m
Species reactivityReacts with: Mouse, Rat, Cow, Human
NQO1 Activity Assay kit (ab184867) is designed for the sensitive and accurate measurement of NQO1 (NAD(P)H dehydrogenase [quinone] 1) activity.
The enzyme activity is determined by following the reduction of Menadione with cofactor NADH and the simultaneous reduction of WST1 which leads to increased absorbance at 440 nm. Dicoumarol is used as an inhibitor of NQO1 as part of the assay.
NQO1 assay protocol summary:
- add samples to wells
- add reaction buffer + inhibitor and reaction buffer to separate sample wells
- analyze with a microplate reader for 5 min
NQO1 serves as a quinone reductase in connection with conjugation reactions of hydroquinones involved in detoxification pathways, NQO1 protects against quinone-induced damage by competing with potentially toxic one-electron pathways. It also functions in biosynthetic processes such as the vitamin K-dependent gamma-carboxylation of glutamate residues in prothrombin synthesis.
Storage instructionsStore at -20°C. Please refer to protocols.
Components 1 x 96 tests 1000X Cofactor (lyophilized) 1 vial 1000X Inhibitor (lyophilized) 1 vial 100X Dye (lyophilized) 1 vial 20X Basic Buffer 1 x 3ml 2X Extraction Buffer 1 x 15ml 5000X Menadione 1 x 100µl 500X NADH (lyophilized) 1 vial 96-well Microplate 2 units
- Bovine Heart Mitochondria (ab110338)
- Rat liver tissue lysate - mitochondrial extract (ab110346)
- Rat heart tissue lysate - mitochondrial extract (ab110347)
- Rat brain tissue lysate - mitochondrial extract (ab110348)
- Mouse liver tissue lysate - mitochondrial extract (ab110349)
- Mouse heart tissue lysate - mitochondrial extract (ab110350)
- Mouse brain tissue lysate - mitochondrial extract (ab110351)
An example is shown below where the rate/slope is calculated between these time points.
This raw data is expressed as rate (mOD/min) per microgram of cell extract added per well as shown above. The molar extinction coefficient factor for the WST1 dye is 25.9/mM/well.
ab184867 has been referenced in 6 publications.
- Guo G et al. NQO1 is a determinant for cellular sensitivity to anti-tumor agent Napabucasin. Am J Cancer Res 10:1442-1454 (2020). PubMed: 32509390
- Tsao YC et al. Discovery of Isoplumbagin as a Novel NQO1 Substrate and Anti-Cancer Quinone. Int J Mol Sci 21:N/A (2020). PubMed: 32575541
- Ling Y et al. Development of novel amino-quinoline-5,8-dione derivatives as NAD(P)H:quinone oxidoreductase 1 (NQO1) inhibitors with potent antiproliferative activities. Eur J Med Chem 154:199-209 (2018). PubMed: 29803003
- Liu X et al. The novel triterpenoid RTA 408 protects human retinal pigment epithelial cells against H2O2-induced cell injury via NF-E2-related factor 2 (Nrf2) activation. Redox Biol 8:98-109 (2016). PubMed: 26773873
- Madajewski B et al. Depleting Tumor-NQO1 Potentiates Anoikis and Inhibits Growth of NSCLC. Mol Cancer Res 14:14-25 (2016). PubMed: 26553038
- Tumer TB et al. Direct and Indirect Antioxidant Activity of Polyphenol- and Isothiocyanate-Enriched Fractions from Moringa oleifera. J Agric Food Chem 63:1505-13 (2015). PubMed: 25605589