Key features and details
- Assay type: Enzyme activity (quantitative)
- Detection method: Colorimetric
- Platform: Microplate reader
- Assay time: 6 hr
- Sample type: Cell culture extracts, Tissue
Product nameComplex IV Rodent Enzyme Activity Microplate Assay Kit
See all Complex IV kits
Sample typeCell culture extracts, Tissue
Assay typeEnzyme activity (quantitative)
Assay time6h 00m
Species reactivityReacts with: Mouse, Rat
Complex IV Rodent Enzyme Activity Microplate Assay Kit (ab109911) is used to determine the activity of cytochrome c oxidase in a mouse sample with speed and simplicity. The COX enzyme is immunocaptured within the wells of the microplate and activity is determined colorimetrically by following the oxidation of reduced cytochrome c by the absorbance change at 550 nm. Included in this kit for performance of the activity assay are buffer, detergent, substrate, and 96-well microplate with monoclonal antibody pre-bound to the wells of the plate, allowing for a stream-lined assay.
Complex IV assay protocol summary:
- add samples to wells to capture enzyme and incubate for 3 hrs
- wash wells
- add reaction mix
- analyze with microplate reader in kinetic mode for 120 min
Buffer, detergent, and microplate should be stored at 4°C, Reagent C (reduced cytochrome c) should be stored at -80°C.
Range of complex IV / cytochrome c oxidase assay kits
Biochemical assay - ab239711
Immunocapture with biochemical assay and ELISA - ab109910 (human)
ELISA - ab179880 (human)
Other related products
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 96-well microplate 1 unit Tube 1 (Buffer) 1 x 10ml Detergent 1 x 1ml Reagent C (Reduced Cytochrome c) 1 x 1ml
- Cytochrome c oxidase
- Cytochrome oxidase
- 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)
Figure 1. To determine the activity in the sample, calculate the slope by using microplate software or by manual calculations using one of the two methods shown. Compare the sample rate with the rate of the control (normal) sample and with the rate of the null (background) to get the relative Complex IV activity. (A)The rate is determined by calculating the gradient of the initial slope over the linear region. (B) The rate is determined by calculating the slope between two points within the linear region.
Abcam's enzyme activity assays apply a novel approach, whereby target enzymes are first immunocaptured from tissue or cell samples before subsequent functional analysis. All of our ELISA kits utilize highly validated monoclonal antibodies and proprietary buffers, which are able to capture even very large enzyme complexes in their fully-intact, functionally-active states. Capture antibodies are pre-coated in the wells of premium Nunc MaxiSorp™ modular microplates, which can be broken into 8-well strips. After the target has been immobilized in the well, substrate is added, and enzyme activity is analyzed by measuring the change in absorbance of either the substrate or the product of the reaction (depending upon which enzyme is being analyzed). By analyzing the enzyme's activity in an isolated context, outside of the cell and free from any other variables, an accurate measurement of the enzyme's functional state can be understood.
Mitchondiral fractions from the heart tissue of Rats infected by S. pneumoniae, or given PBS sham control, were subjected to measurements of complex I-V activities. Complex I was measured with ab109721 (top left), Complex II + III were measured using ab109905 (top right), Complex IV was measured using ab109911 (bottom left) and Complex V was measured using ab109714 (bottom right).
Freshly isolated mitochondrial pellets were resuspened in PBS supplemented with 10% detergent provided in the kits. Protein concentrations of these mitochondrial lysates were estimated and 25 μg (for complex I, IV and V) or 100 μg (for complex II+III) mitochondrial protein was used per reaction. Enzyme activities were measured spectrophotometricly in triplicate and expressed as changes of absorbance per minute per mg protein.
ab109911 has been referenced in 54 publications.
- Black M et al. FOXM1 nuclear transcription factor translocates into mitochondria and inhibits oxidative phosphorylation. Mol Biol Cell N/A:mbcE19070413 (2020). PubMed: 32348194
- Santos MCFD et al. Irp2 regulates insulin production through iron-mediated Cdkal1-catalyzed tRNA modification. Nat Commun 11:296 (2020). PubMed: 31941883
- Lee DH et al. Withania somnifera Extract Enhances Energy Expenditure via Improving Mitochondrial Function in Adipose Tissue and Skeletal Muscle. Nutrients 12:N/A (2020). PubMed: 32046183
- Zhang R et al. Caloric Restriction Induces MicroRNAs to Improve Mitochondrial Proteostasis. iScience 17:155-166 (2019). PubMed: 31279933
- Caldwell CC et al. Treadmill exercise rescues mitochondrial function and motor behavior in the CAG140 knock-in mouse model of Huntington's disease. Chem Biol Interact 315:108907 (2019). PubMed: 31778667