Key features and details
- Assay type: Quantitative
- Detection method: Colorimetric
- Platform: Microplate reader
- Assay time: 5 hr
- Sample type: Inhibitor compounds
Product nameMitoTox™ Complex I OXPHOS Activity Assay Kit
See all Complex I kits
Sample typeInhibitor compounds
Assay time5h 00m
MitoTox™ Complex I OXPHOS Activity Assay (ab109903) is designed for testing the direct inhibitory effect of compounds on Complex I activity in only 5 hours. Complex I extracted from the provided bovine heart mitochondria (a rich source of Complex I) is immunocaptured by specific antibodies on the plate. Complex I activity can be observed as decrease in absorbance at OD 340 nm. The intra-assay and inter-assay variation of this assay are both < 10%.
Inhibitory effects of compounds on Complex I activity can be tested in two different ways: 1. Screening format, where up to 23 compounds can be tested at a single concentration in triplicate; 2. Dose response (IC50) format, where two compounds known to affect Complex I activity can be tested at 11 different data points in triplicate.
Testing for mitochondrial function has become a key aspect of drug discovery. Mitochondria can be affected by drug treatment, resulting into cardio- and hepatotoxic side effects that can lead to drug withdrawal from the market. Therefore, there is increasing emphasis on testing the impact on mitochondria early on in the drug development process to reduce failure rates during preclinical and clinical phases.
Store Phospholipids, Bovine heart mitochondria, Ubiquinone and Activity Buffer at -80°C. Store all other components at 4°C.
Review the mitochondrial assay guide, or the full metabolism assay guide to learn about more assays for metabolites, metabolic enzymes, mitochondrial function, and oxidative stress, and also how to assay metabolic function in live cells using your plate reader.
Storage instructionsPlease refer to protocols.
Components 96 tests 12-channel reagent reservoirs 2 units 1X Mito Buffer 1 x 5ml 20X Wash Buffer 1 x 5ml Bovine heart mitochondria 1 x 360µl Complex I Activity Buffer 1 x 24ml Detergent 1 x 100µl Phospholipids 1 x 6ml Pre-coated 96-well microplate 1 unit Ubiquinone 1 1 x 60µl
- NADH dehydrogenase
Typical dose response curve for rotenone. Assay was performed following the Dose Response Assay Procedure using rotenone, a well known Complex I inhibitor. Rotenone was prepared in DMSO to generate a 10 mM stock. Starting with a 50 µM final concentration in well, 1:10 serial dilutions of rotenone were generated.
ab109903 has been referenced in 10 publications.
- Park SY & Koh HC FUNDC1 regulates receptor-mediated mitophagy independently of the PINK1/Parkin-dependent pathway in rotenone-treated SH-SY5Y cells. Food Chem Toxicol 137:111163 (2020). PubMed: 32001317
- Wang Y et al. Metformin Improves Mitochondrial Respiratory Activity through Activation of AMPK. Cell Rep 29:1511-1523.e5 (2019). PubMed: 31693892
- Thomas LW et al. CHCHD4 confers metabolic vulnerabilities to tumour cells through its control of the mitochondrial respiratory chain. Cancer Metab 7:2 (2019). PubMed: 30886710
- Baccelli I et al. Mubritinib Targets the Electron Transport Chain Complex I and Reveals the Landscape of OXPHOS Dependency in Acute Myeloid Leukemia. Cancer Cell 36:84-99.e8 (2019). PubMed: 31287994
- Cameron AR et al. Metformin selectively targets redox control of complex I energy transduction. Redox Biol 14:187-197 (2018). PubMed: 28942196
- Choi SE et al. NF-?B/p53-activated inflammatory response involves in diquat-induced mitochondrial dysfunction and apoptosis. Environ Toxicol N/A:N/A (2018). PubMed: 29484840
- Kim EM et al. Pro-apoptotic Bax promotes mesenchymal-epithelial transition by binding to respiratory complex-I and antagonizing the malignant actions of pro-survival Bcl-2 proteins. Cancer Lett 424:127-135 (2018). PubMed: 29596889
- Zhang C et al. Sorafenib targets the mitochondrial electron transport chain complexes and ATP synthase to activate the PINK1-Parkin pathway and modulate cellular drug response. J Biol Chem 292:15105-15120 (2017). PubMed: 28673964
- Kalghatgi S et al. Bactericidal antibiotics induce mitochondrial dysfunction and oxidative damage in Mammalian cells. Sci Transl Med 5:192ra85 (2013). PubMed: 23825301
- Lai K et al. Integrated Compound Profiling Screens Identify the Mitochondrial Electron Transport Chain as the Molecular Target of the Natural Products Manassantin, Sesquicillin, and Arctigenin. ACS Chem Biol N/A:N/A (2012). PubMed: 23138533