MitoSOX Red 

MitoSOX Red is a novel fluorescent probe that specifically targets mitochondria in living cells with cell membrane permeability. MitoSOX Red enters the mitochondria, it is oxidized by superoxide, but not by other ROS or RNS generating systems. MitoSOX Red then binds to intramitochondrial nucleic acids, producing strong red fluorescence. MitoSOX Red can be used as a fluorescent indicator to specifically detect superoxide. Additionally, Superoxide dismutase (SOD) prevents the oxidation of MitoSOX Red.
Excitation/emission wavelength: 396/610 nm.

General Protocol

1. Preparation of MitoSOX Red working solution
1.1 Preparation of the stock solution
Dissolve 50 μg MitoSOX Red in 13 μL DMSO to obtain 5 mM of stock solution.
Note: It is recommended to store the stock solution at -20°C and -80°C away from light and avoid repetitive freeze-thaw cycles.
1.2 Preparation of MitoSOX Red working solution
Dilute the stock solution in serum-free cell culture medium or PBS to obtain 1-10 μM of working solution.
Note: Please adjust the concentration of MitoSOX Red working solution according to the actual situation.
2. Cell staining
2.1 Suspension cells（6-well plate）
a. Centrifuge at 1000 g at 4°C for 3-5 minutes and then discard the supernatant. Wash twice with PBS, 5 minutes each time.The cell density is 1×10⁶/mL.
b. Add 1 mL of working solution, and then incubate at room temperature for 5-30 minutes.
c. Centrifuge at 400 g at 4°C for 3-4 minutes and then discard the supernatant.
d. Wash twice with PBS, 5 minutes each time.
e. Resuspend cells with serum-free cell culture medium or PBS. Observation by fluorescence microscopy or flow cytometry.
2.2 Adherent cells
a. Culture adherent cells on sterile coverslips.
b. Remove the coverslip from the medium and aspirate excess medium.
c. Add 100 μL of working solution, gently shake it to completely cover the cells,and then incubate at room temperature for 5-30 minutes.
d. Wash twice with medium, 5 minutes each time. Observation by fluorescence microscopy or flow cytometry.

Storage
-20°C, 1 year. Protect from light

Precautions
1. Please adjust the concentration of MitoSOX Red working solution according to the actual situation.
2. This product is for R&D use only, not for drug, household, or other uses.
3. For your safety and health, please wear a lab coat and disposable gloves to operate.

MCE has not independently confirmed the accuracy of these methods. They are for reference only.

759.70

Solid

C₄₃H₄₃IN₃P

1003197-00-9

580

510

NC1=CC(C(C2=CC=CC=C2)N(CCCCCC[P+](C3=CC=CC=C3)(C4=CC=CC=C4)C5=CC=CC=C5)C6=CC(N)=CC=C76)=C7C=C1.[I-]

Room temperature in continental US; may vary elsewhere.

-20°C, protect from light, stored under nitrogen

*In solvent : -80°C, 6 months; -20°C, 1 month (protect from light, stored under nitrogen)

• [1]. Price OT, et al. Quantitative fluorescence of 5-FU-treated fetal rat limbs using confocal laser scanning microscopy and Lysotracker Red. Cytometry A. 2003;53(1):9-21.  [Content Brief]

  [2]. Fengjiao Jin, et al. The PI3K/Akt/GSK-3β/ROS/eIF2B pathway promotes breast cancer growth and metastasis via suppression of NK cell cytotoxicity and tumor cell susceptibility. Cancer Biol Med. 2019 Feb;16(1):38-54.  [Content Brief]