Product nameComplex IV Human Enzyme Activity Dipstick Assay Kit
See all Complex IV kits
Sample typeCell culture extracts, Tissue
Assay typeSandwich (quantitative)
Species reactivityReacts with: Cow, Human
Does not react with: Mouse, Rat
Contains 30 or 90 dipsticks and necessary components to quantify the activity of the cytochrome c oxidase enzyme complex from human and bovine samples. The kit includes sufficient materials to generate a standard curve and evaluate several unknown samples.
The isolation of mitochondria is not necessary for the performance of this assay. In this kit the specificity of anti-COX monoclonal antibodies (mAbs) is combined with traditional methods for determining COX enzyme activity by histochemical methods and in-gel activity assays. First, the COX enzyme complex is immnocaptured (i.e immunoprecipated in active form) on the dipstick. Second, the dipstick is immersed in COX activity buffer containing reduced cytochrome c and di-amino benzidinetetrachloride (DAB), which serves as the reporter of COX activity. Immunocaptured COX oxidizes cytochrome c, which then oxidizes DAB to form a red-colored precipitate at the COX antibody line on the dipstick. In addition to being quick, the reaction is cyanide-sensitive. The signal intensity of this precipitate corresponds to the level of COX activity in the sample. The signal intensity is best measured by a dipstick reader or may be analyzed by another imaging system.
Store dipsticks at room temperature in their provided container and out of direct sunlight. High humidity conditions should be avoided.
Store Buffer A, B, and C at 4°C or at -20°C for long term storage.
Store Tubes 1 and 2 at -80°C; they can also be aliquoted upon receipt to prevent freeze/thaw cycles.
Tube 3 can be stored at room temperature.
Range of complex IV / cytochrome c oxidase assay kits
Biochemical assay - ab239711
Immunocapture with biochemical assay and ELISA - ab109910 (human)
ELISA - ab179880 (human)
Tested applicationsSuitable for: Functional Studiesmore details
Storage instructionsPlease refer to protocols.
Components 90 tests 30 tests 96-well microplate 6 units 2 units Buffer B (10X Blocking solution) 6 x 1ml 2 x 1ml Dipsticks 3 x 30 units 1 x 30 units Extraction Buffer (ab260490) 3 x 15ml 1 x 15ml Tube 1 (DAB 100X stock) 3 x 100µl 1 x 100µl Tube 2 (reduced cytochrome c 20X stock) 3 x 500µl 1 x 500µl Tube 3 (Buffer for activity assay) 3 x 10ml 1 x 10ml Wash buffer 3 x 1.5ml 1 x 1.5ml
- Cytochrome c oxidase
- Cytochrome oxidase
Our Abpromise guarantee covers the use of ab109876 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.|
Abcam's enzyme activity assays apply a novel approach, whereby target enzymes are first immunocaptured from tissue or cell samples before subsequent functional analysis. Dipstick ELISA Kits extend this concept by utilizing the well-established lateral flow concept, wherein capture antibodies are striped onto nitrocellulose membrane and a wicking pad draws the sample through the antibody bands. 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.
Figure 1. An example using ab109876 to measure Complex IV activity in fibroblast protein extracts. Developed dipsticks from a 1:2 dilution series using a positive control sample and the associated standard curve. Starting material was 100 µg of fibroblast protein extract.
Figure 2. An example using ab109876 to measure Complex IV activity in fibroblast protein extracts. Based on the standard curve, 50 µg of protein extract were loaded onto a dipstick for each sample. The figure shows four developed dipsticks, a control sample (1) and four unknowns (2-6). The analysis of the signal intensity and interpolation from the standard curve showed that the unknown samples have between 15-61% of normal Complex IV activity levels.
ab109876 has been referenced in 10 publications.
- Li H et al. Mitochondrial dysfunction and mitophagy defect triggered by heterozygous GBA mutations. Autophagy 15:113-130 (2019). PubMed: 30160596
- Rodríguez-García ME et al. An innovative strategy to clone positive modifier genes of defects caused by mtDNA mutations: MRPS18C as suppressor gene of m.3946G>A mutation in MT-ND1 gene. Hum Genet 136:885-896 (2017). PubMed: 28526948
- Tang G et al. Mitochondrial abnormalities in temporal lobe of autistic brain. Neurobiol Dis 54:349-61 (2013). PubMed: 23333625
- Calvo SE et al. Molecular diagnosis of infantile mitochondrial disease with targeted next-generation sequencing. Sci Transl Med 4:118ra10 (2012). PubMed: 22277967
- Fassone E et al. FOXRED1, encoding an FAD-dependent oxidoreductase complex-I-specific molecular chaperone, is mutated in infantile-onset mitochondrial encephalopathy. Hum Mol Genet 19:4837-47 (2010). PubMed: 20858599
- Calvo SE et al. High-throughput, pooled sequencing identifies mutations in NUBPL and FOXRED1 in human complex I deficiency. Nat Genet 42:851-8 (2010). PubMed: 20818383
- Marusich MF et al. Novel antibody-based strategies for the rapid diagnosis of mitochondrial disease and dysfunction. Int J Biochem Cell Biol 41:2081-8 (2009). PubMed: 19460456
- Baughman JM et al. A computational screen for regulators of oxidative phosphorylation implicates SLIRP in mitochondrial RNA homeostasis. PLoS Genet 5:e1000590 (2009). PubMed: 19680543
- Willis JH et al. Isolated deficiencies of OXPHOS complexes I and IV are identified accurately and quickly by simple enzyme activity immunocapture assays. Biochim Biophys Acta 1787:533-8 (2009). PubMed: 19041632
- Sugiana C et al. Mutation of C20orf7 disrupts complex I assembly and causes lethal neonatal mitochondrial disease. Am J Hum Genet 83:468-78 (2008). PubMed: 18940309