H2DCFDA (DCFH-DA)

Catalog No.S9687 Batch:S968702

Technical Data

Formula

C₂₄H₁₆Cl₂O₇

Molecular Weight

487.29

CAS No.

4091-99-0

Solubility (25°C)*

In vitro

DMSO

97 mg/mL (199.06 mM)

Ethanol

15 mg/mL (30.78 mM)

Water

Insoluble

In vivo (Add solvents to the product individually and in order)

Clear solution	5%DMSO 1 40%PEG300 2 5%Tween80 3 50%ddH2O	4.85mg/ml	Taking the 1 mL working solution as an example, add 50 μL of 97 mg/ml clarified DMSO stock solution to 400 μL of PEG300, mix evenly to clarify it; add 50 μL of Tween80 to the above system, mix evenly to clarify; then continue to add 500 μL of ddH2O to adjust the volume to 1 mL. The mixed solution should be used immediately for optimal results.
Clear solution	5% DMSO 1 95% Corn oil	0.8mg/ml	Taking the 1 mL working solution as an example, add 50 μL of 16 mg/ml clear DMSO stock solution to 950 μL of corn oil and mix evenly. The mixed solution should be used immediately for optimal results.

* <1 mg/ml means slightly soluble or insoluble.
* Please note that Selleck tests the solubility of all compounds in-house, and the actual solubility may differ slightly from published values. This is normal and is due to slight batch-to-batch variations.
* Room temperature shipping (Stability testing shows this product can be shipped without any cooling measures.)

Preparing Stock Solutions

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

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

Protocol (from reference)

Cell Assay:^{^[2]}	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.

Cell Assay:^{^[2]}

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.

References

Selleck's H2DCFDA (DCFH-DA) has been cited by 10 publications

S-nitrosothiol homeostasis maintained by ADH5 facilitates STING-dependent host defense against pathogens [ Nat Commun, 2024, 15(1):1750]	PubMed: 38409248
Fusobacterium nucleatum-triggered neutrophil extracellular traps facilitate colorectal carcinoma progression [ J Exp Clin Cancer Res, 2023, 42(1):236]	PubMed: 37684625
Fusobacterium nucleatum-triggered neutrophil extracellular traps facilitate colorectal carcinoma progression [ J Exp Clin Cancer Res, 2023, 42(1):236]	PubMed: 37684625
Erianin inhibits the growth and metastasis through autophagy-dependent ferroptosis in KRASG13D colorectal cancer [ Free Radic Biol Med, 2023, 204:301-312]	PubMed: 37217090
FLRT2 suppresses bladder cancer progression through inducing ferroptosis [ J Cell Mol Med, 2023, 10.1111/jcmm.17855]	PubMed: 37480224
Reactive sulfur species disaggregate the SQSTM1/p62-based aggresome-like induced structures via the HSP70 induction and prevent parthanatos [ J Biol Chem, 2023, 299(6):104710]	PubMed: 37060999
[A Novel Chinese Medicine Formula Inhibits Non-small Cell Lung Cancer by Triggering Oxidative Stress Dependent on Pentose Phosphate Pathway] [ Zhongguo Fei Ai Za Zhi, 2023, 10.3779/j.issn.1009-3419.2023.101.27]	PubMed: 37985150
Elevated neutrophil extracellular traps by HBV-mediated S100A9-TLR4/RAGE-ROS cascade facilitate the growth and metastasis of hepatocellular carcinoma [ Cancer Commun (Lond), 2022, 10.1002/cac2.12388]	PubMed: 36346061
S100A8-Mediated NLRP3 Inflammasome-Dependent Pyroptosis in Macrophages Facilitates Liver Fibrosis Progression [ Cells, 2022, 11(22)3579]	PubMed: 36429008
The potential efficacy and mechanism of bendamustine in entra-nodal NK/T cell lymphoma [ Hematol Oncol, 2022, 10.1002/hon.3007]	PubMed: 35439335

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SHIPPING AND STORAGE
Selleck products are transported at room temperature. If you receive the product at room temperature, please rest assured, the Selleck Quality Inspection Department has conducted experiments to verify that the normal temperature placement of one month will not affect the biological activity of powder products. After collecting, please store the product according to the requirements described in the datasheet. Most Selleck products are stable under the recommended conditions.

NOT FOR HUMAN, VETERINARY DIAGNOSTIC OR THERAPEUTIC USE.