Home Immunology & Inflammation ROS chemical H2DCFDA (DCFH-DA) For research use only.

H2DCFDA (DCFH-DA)

Synonyms: 2',7'-Dichlorodihydrofluorescein diacetate

H2DCFDA (DCFH-DA) is a cell-permeable probe used to detect intracellular reactive oxygen species (ROS) production. It is de-esterified intracellularly and turns to highly fluorescent 2′,7′-dichlorofluorescein upon oxidation. It is used in sensitive and rapid quantitation of oxygen-reactive species in response to oxidative metabolism; microplate assay for detecting oxidative products in phagocytic cells, and quantitative multiwell myeloid differentiation assay.(Ex/Em=488/525 nm)

H2DCFDA (DCFH-DA) Chemical Structure

H2DCFDA (DCFH-DA) Chemical Structure

CAS No. 4091-99-0

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H2DCFDA (DCFH-DA) Related Products

Biological Activity

Description H2DCFDA (DCFH-DA) is a cell-permeable probe used to detect intracellular reactive oxygen species (ROS) production. It is de-esterified intracellularly and turns to highly fluorescent 2′,7′-dichlorofluorescein upon oxidation. It is used in sensitive and rapid quantitation of oxygen-reactive species in response to oxidative metabolism; microplate assay for detecting oxidative products in phagocytic cells, and quantitative multiwell myeloid differentiation assay.(Ex/Em=488/525 nm)
Targets
ROS [1]
In vitro
In vitro

1. H2DCFDA is dissolved in DMSO to obtain a 10 mM stock solutions and further diluted before use.
2. Cells are incubated with 5 μM H2DCFDA solution in PBS in the dark for 30 min at 37°C, then harvested with 0.05% trypsin-EDTA solution, suspended in a fresh medium, and immediately analyzed with flow cytometer.
3. Along with the H2DCFDA probe, if indicated, use ROS-insensitive modification of the fluorescein dye DCFDA as a positive control. The staining procedure is the same as for the H2DCFDA。
Emission (Em)=517 Excitation (Ex)=492 517

Cell Research Cell lines hCECs
Concentrations 10 μM
Incubation Time 30 min
Method

Intracellular ROS levels in hCECs are measured by incubating the cells with the redox-sensitive fluorescent dye 2'7'-dichlorofluorescein diacetate (DCFH-DA) (10 μM) at 37℃ for 30 minutes in the dark. The cells are then detached from the culture wells using 0.25% trypsin-EDTA and washed twice using ice-cold PBS. Flow cytometry measurements are performed three times for each treatment. Mean fluorescence intensity is quantified using software. Cells that had been incubated without DCFH-DA are used as negative control. The distribution of fluorescent dichlorofluorescein (DCF) on the cell monolayer is visualized and photographed using a fluorescence microscope.

Chemical Information & Solubility

Molecular Weight 487.29 Formula

C24H16Cl2O7
CAS No. 4091-99-0 SDF --
Smiles CC(=O)OC1=CC2=C(C=C1Cl)C(C3=C(O2)C=C(OC(C)=O)C(=C3)Cl)C4=C(C=CC=C4)C(O)=O
Storage (From the date of receipt) 3 years -20°C(in the dark) powder

In vitro
Batch:

DMSO : 97 mg/mL ( (199.06 mM) Moisture-absorbing DMSO reduces solubility. Please use fresh DMSO.)

Ethanol : 7 mg/mL

Water : Insoluble


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In vivo
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Method for preparing in vivo formulation: Take μL DMSO master liquid, next add μL Corn oil, mix and clarify.

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Tech Support

Answers to questions you may have can be found in the inhibitor handling instructions. Topics include how to prepare stock solutions, how to store inhibitors, and issues that need special attention for cell-based assays and animal experiments.

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