JC-10 Mitochondrial Membrane Potential Assay Kit (Microplate) (ab112134)
Key features and details
- Detection method: Fluorescent
- Platform: Microplate reader
- Assay time: 1 hr
- Sample type: Adherent cells, Suspension cells
Overview
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Product name
JC-10 Mitochondrial Membrane Potential Assay Kit (Microplate)
See all Mitochondrial Membrane Potential kits -
Detection method
Fluorescent -
Sample type
Adherent cells, Suspension cells -
Assay time
1h 00m -
Product overview
JC-10 Mitochondrial Membrane Potential Assay Kit (Microplate) ab112134 enables researchers to analyze a JC-10 assay with a microplate reader. The JC-10 assay provides the most robust assay method for monitoring mitochondria membrane potential changes.
This mitochondrial membrane potential assay protocol is based on the detection of the mitochondrial membrane potential changes in cells by the cationic, lipophilic JC-10 dye. In normal cells, JC-10 concentrates in the mitochondrial matrix where it forms red fluorescent aggregates. However, in apoptotic and necrotic cells, JC-10 diffuses out of mitochondria. It changes to monomeric form and stains cells in green fluorescence.
Although JC-1 is widely used in many labs, its poor water solubility causes great inconvenience. Even at 1 µM concentration, JC-1 tends to precipitate in aqueous buffer. JC-10 is developed to be a superior alternative to JC-1 when high dye concentration is desired. Compared to JC-1, JC-10 has much better water solubility. JC-10 is capable of selectively entering mitochondria, and reversibly changes its color from green to orange as membrane potentials increase. This property is due to the reversible formation of JC-10 aggregates upon membrane polarization that causes shifts in emitted light from 520 nm (i.e. emission of JC-10 monomeric form) to 570 nm (i.e. emission of J-aggregate form). When excited at 490 nm, the color of JC-10 changes reversibly from green to greenish orange as the mitochondrial membrane becomes more polarized.
In normal cells, JC-10 concentrates in the mitochondrial matrix where it forms red fluorescent aggregates. However, in apoptotic and necrotic cells, JC-10 exists in monomeric form and stains cells green. The green emission can be analyzed in fluorescence channel 1 (FL1) and greenish orange emission in channel 2 (FL2). Besides its use in fluorescence microplate platform, it can also be used in fluorescence imaging and flow cytometry.
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Notes
A microplate reader with bottom-reading mode is essential to perform this assay.
If you would like to use JC-10 on a flow cytometer, we recommend JC-10 Mitochondrial Membrane Potential Assay Kit (Flow Cytometry) (ab112133).
Related assays
Review the cell health assay guide to learn about kits to perform a cell viability assay, cytotoxicity assay and cell proliferation assay.
Review the metabolism assay guide to learn about assays for metabolites, metabolic enzymes, mitochondrial function, and oxidative stress, and also about how to assay metabolic function in live cells using your plate reader.
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Platform
Microplate reader
Properties
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Storage instructions
Store at -20°C. Please refer to protocols. -
Components 5 x 96 tests 100X JC-10 in DMSO 1 x 250µl Assay Buffer A 1 x 25ml Assay Buffer B 1 x 25ml -
Research areas
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Relevance
Mitochondrial Membrane Potential is an important parameter of mitochondrial function used as an indicator of cell death. The collapse of the mitochondrial Membrane potential coincides with the opening of the mitochondrial permeability transition pores, leading to the release of cytochrome c into the cytosol, which in turn triggers other downstream events in the apoptotic cascade. -
Alternative names
- mitochondrial membrane potential
Images
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Mitochondrial membrane potential was measured using ab112134 Kysenius K et al., PLoS One, 9(9). Fig2f. doi: 10.1371/journal.pone.0107129 Reproduced under the Creative Commons license http://creativecommons.org/licenses/by/4.0/
CGN were cultured on 96-well white-walled clear-bottom plates in phenol-red free Neurobasal until 7 DIV. Thirty minutes before the end of the treatment, 50 μl of JC-10 dye-loading solution was added to each well and incubated for 30 minutes before measuring fluorescence intensities (Ex/Em = 485/515 nm and Ex/Em = 540/590 nm). The change of mitochondrial membrane potential was measured as the ratio between aggregate (Em = 515 nm) and monomeric forms (Em = 590 nm) of JC-10. Increasing ratios indicate mitochondrial membrane depolarization.
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Functional Studies - JC-10 Mitochondrial Membrane Potential Assay Kit (Microplate) (ab112134)JC-10 Mitochondrial Membrane Potential Assay Kit (Microplate) (ab112134). Camptothecin-induced mitochondria membrane potential changes with JC-10 and JC-1 in Jurkat cells. JC-1 (blue bar) and JC-10 (red bar) dye loading solutions were added to Jurket cells untreated (0 µM) or treated with camptothecin (10 µM for 4 hours) and incubated for 30 minutes. The fluorescent intensities for both J-aggregates and monomeric forms of JC-1 and JC-10 were measured at Ex/Em = 490/525 nm and 490/590 nm with a microplate reader.
