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MitoTox™ Complex IV OXPHOS Activity Assay Kit (ab109906) is designed for testing the direct inhibitory effect of compounds on Complex IV activity in only 5 hours. Complex IV extracted from the provided bovine heart mitochondria (a rich source of Complex IV) is immunocaptured by specific antibodies on the plate. Complex IV activity can be observed as decrease in absorbance at OD 550 nm. The intra-assay and inter-assay variation of this assay are both <10%.
Inhibitory effects of compounds on Complex IV 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 IV 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 Bovine Heart Mitochondria and Reagent C at -80°C. Store all other components at 4°C.
|12-channel reagent reservoirs||2 units|
|1X Blocking Buffer (Solution 1)||1 x 70ml|
|1X Complex IV Mito Buffer (Solution 2)||1 x 110ml|
|Bovine Heart Mitochondria||1 x 90µl|
|Detergent||1 x 20µl|
|Fat free powdered milk (Blocking powder)||1 x 1.8g|
|Pre-coated 96-well microplate||1 unit|
|Reagent C (Cytochrome c)||2 x 1ml|
Our Abpromise guarantee covers the use of ab109906 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
|Functional Studies||Use at an assay dependent dilution.|
Typical dose response curve for KCN. Assay was performed following the Dose Response Assay Procedure using KCN, a well known Complex IV inhibitor. KCN was prepared in 0.1 M NaOH to generate a 100 mM stock. Starting with a 100 µM final concentration in well, 1:10 serial dilutions of KCN were generated.