CM-H2DCFDA (General Oxidative Stress Indicator)
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CM-H2DCFDA (General Oxidative Stress Indicator)
Citations & References (115)
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CM-H2DCFDA (General Oxidative Stress Indicator)

CM-H2DCFDA is a chloromethyl derivative of H2DCFDA, useful as an indicator for reactive oxygen species (ROS) in cells. This indicatorRead more
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Catalog NumberQuantity
C682720 x 50 μg
Catalog number C6827
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4,875.00
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Quantity:
20 x 50 μg
Price (CNY)
4,875.00
Online Exclusive
Ends: 31-Dec-2025
6,564.00
Save 1,689.00 (26%)
Each
Add to cart

CM-H2DCFDA is a chloromethyl derivative of H2DCFDA, useful as an indicator for reactive oxygen species (ROS) in cells. This indicator exhibits much better retention in live cells than H2DCFDA. CM-H2DCFDA passively diffuses into cells, where its acetate groups are cleaved by intracellular esterases and its thiol-reactive chloromethyl group reacts with intracellular glutathione and other thiols. Subsequent oxidation yields a fluorescent adduct that is trapped inside the cell, thus facilitating long-term studies.

ROS Indicator Specifications:
• Excitation: ∼492–495 nm
• Emission: 517–527 nm
• Product is air sensitive and should be stored under dry argon or nitrogen
• Product may be dissolved in DMSO, DMF, or ethanol for use
• Indicator is cell permeant (cell loading protocols are available in the literature)
• Fluorescence can be monitored using a flow cytometer, fluorometer, microplate reader, or fluorescence microscope, using excitation sources and filters appropriate for fluorescein

Find more ROS indicators
We offer an assortment of Molecular Probes™ products for the generation of reactive oxygen species (ROS), including singlet oxygen, superoxide, hydroxyl radical and various peroxide and hydroperoxides, as well as for their fluorometric detection in solution. Review Generating and Detecting Reactive Oxygen Species—Section 18.2 in the Molecular Probes™ Handbook for more information on these products.

Dissolve the dye using high-quality, anhydrous dimethylsulfoxide (DMSO), dimethylformamide (DMF), or 100% ethanol to prepare stock concentration up to 5 mM. Working solutions should be freshly prepared. This probe oxidizes readily in solution thus presence of moisture will facilitate the decomposition of the dye.

For Research Use Only. Not for use in diagnostic procedures.
Specifications
Quantity20 x 50 μg
Product TypeROS Indicator
Unit SizeEach

Frequently asked questions (FAQs)

I want to assay cells for reactive oxygen species using carboxy-H2DCFDA, but I want to do so with a plate reader instead of microscope. Will it work?

It has been done. The problem is that plate readers are less sensitive than microscopes, with far less signal-to-background difference. It is worth trying, but first optimize concentrations and loading times with control cells, use a plate with little to no autofluorescence, and possibly optimize the gain setting in order to get the best signal possible. But don't expect the same sensitivity, even with optimization.

Find additional tips, troubleshooting help, and resources within our Cell Analysis Support Center.

I have GFP-transfected cells and need to label for reactive oxygen species. Can I use H2DCFDA?

This is not recommended as the two dyes overlap in the emission wavelength. There are other ROS reagents available in different wavelengths, such as CellROX Deep Red, which emits in the far-red range (665 nm), or dihydroethidium, which is emits in the visible red range (620 nm).

Find additional tips, troubleshooting help, and resources within our Cell Analysis Support Center.

I labeled my cell with CM-H2DCFDA for reactive oxygen detection, but upon illuminating the cell there is a significant increase in fluorescence in the control cells. Why?

If the cell is overloaded with dye, the high intracellular concentration of the dye may lead to dye-dye quenching. Upon illumination, photobleaching will occur, which will reduce the dye-dye quenching and actually increase the fluorescence (for a while, but then it will start decreasing). To solve the problem, reduce the concentration and incubation time, and try a range of incubation times and concentrations.

Find additional tips, troubleshooting help, and resources within our Cell Analysis Support Center.

I need a formaldehyde-fixable reactive oxygen species detection assay. Is H2 DCFDA fixable?

H2DCFDA and similar derivatives are not fixable. The same goes for dihydroethidium and dihydrorhodamine. However, CellROX Deep Red and CellROX Green are retained for a limited time upon fixation with formaldehyde. CellROX Green may be retained upon subsequent Triton X-100 permeabilization. Avoid the use of any acetone or alcohol-based fixatives or fixatives that include alcohol, such as formalin.

Find additional tips, troubleshooting help, and resources within our Cell Analysis Support Center.

Why don't I see a significant change in signal for my live-cell fluorescent indicator dye?

Regardless of the type of live-cell indicator dye (e.g., calcium indicators, pH indicator, metal ion indicators), make sure there is no serum during the loading step, which can prematurely cleave dyes with AM esters and bind dyes non-specifically. Always optimize the dye concentration and staining time with a positive control before you run your test samples, to give the best signal-to-background. Always run a positive control with a buffer containing free ions of known concentration and an ionophore to open pores to those ions (for instance, for calcium indicators like Fluo-4 AM, this would include a buffer with added calcium combined with calcimycin, or for pH indicators, buffers of different pHs combined with nigericin). Reactive oxygen indicators, such as CellROX Green or H2DCFDA would require a cellular reactive oxygen species (ROS) stimulant as a positive control, such as menadione. Finally, make sure your imaging system has a sensitive detector. Plate readers, for instance, have much lower detector efficiency over background, compared to microscopy or flow cytometry.

Find additional tips, troubleshooting help, and resources within our Cell Analysis Support Center.

Citations & References (115)

Citations & References
Abstract
Authors:
Journal:
PubMed ID:18258751
Large-scale chemical dissection of mitochondrial function.
Authors:Wagner BK,Kitami T,Gilbert TJ,Peck D,Ramanathan A,Schreiber SL,Golub TR,Mootha VK
Journal:Nature biotechnology
PubMed ID:18297058
Mitochondrial oxidative phosphorylation (OXPHOS) is central to physiology and disease pathogenesis. To systematically investigate its activity and regulation, we performed a wide range of assays of OXPHOS physiology and nuclear and mitochondrial gene expression across 2490 chemical perturbations in muscle cells. Through mining of the resulting compendium, we discovered that: ... More
Reactive oxygen species production via NADPH oxidase mediates TGF-beta-induced cytoskeletal alterations in endothelial cells.
Authors:Hu T, Ramachandrarao SP, Siva S, Valancius C, Zhu Y, Mahadev K, Toh I, Goldstein BJ, Woolkalis M, Sharma K
Journal:Am J Physiol Renal Physiol
PubMed ID:16159901
'Cytoskeletal alterations in endothelial cells have been linked to nitric oxide generation and cell-cell interactions. Transforming growth factor (TGF)-beta has been described to affect cytoskeletal rearrangement in numerous cell types; however, the underlying pathway is unclear. In the present study, we found that human umbilical vein endothelial cells (HUVEC) have ... More
The yeast prion Ure2p native-like assemblies are toxic to mammalian cells regardless of their aggregation state.
Authors:Pieri L, Bucciantini M, Nosi D, Formigli L, Savistchenko J, Melki R, Stefani M
Journal:J Biol Chem
PubMed ID:16571726
'The yeast prion Ure2p assembles in vitro into oligomers and fibrils retaining the alpha-helix content and binding properties of the soluble protein. Here we show that the different forms of Ure2p native-like assemblies (dimers, oligomers, and fibrils) are similarly toxic to murine H-END cells when added to the culture medium. ... More
Role of extracellular signal-regulated protein kinase in neuronal cell death induced by glutathione depletion in neuron/glia mesencephalic cultures.
Authors:de Bernardo S, Canals S, Casarejos MJ, Solano RM, Menendez J, Mena MA
Journal:J Neurochem
PubMed ID:15485497
'To date, glutathione (GSH) depletion is the earliest biochemical alteration shown in brains of Parkinson''s disease patients, but the role of GSH in dopamine cell survival is debated. In this study we show that GSH depletion, produced with GSH synthesis inhibitor, L-buthionine-(S,R)-sulfoximine (BSO), induces selectively neuronal cell death in neuron/glia, ... More
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