MitoSOX Red

1. Preparation of MitoSOX Red working solution

1.1 Preparing storage solution

Prepare a 5 mM stock solution by diluting 50 μg of MitoSOX Red with 13 μL of anhydrous DMSO.

Note: MitoSOX Red storage solution is recommended to be aliquoted and stored at -20℃ or -80℃ in the dark.

1.2 Preparation of working fluid

Dilute the stock solution with pre-warmed serum-free cell culture medium or PBS to prepare a 1-5 μM MitoSOX Red working solution.

Note: Please adjust the concentration of MitoSOX Red working fluid according to the actual situation, and prepare it when used.

2. Cell staining (suspension cells)

2.1 Collect cells by centrifugation, add PBS and wash twice, 5 minutes each time. The cell density was 1×10⁶/mL.

2.2 Add 1 mL MitoSOX Red working solution and incubate at room temperature for 5-30 minutes.

2.3 Centrifuge at 400 g for 3-4 minutes and discard the supernatant.

2.4 Add PBS and wash the cells twice, 5 minutes each time.

2.5 Resuspend the cells in 1 mL of serum-free medium or PBS and observe using a fluorescence microscope or flow cytometer.

3. Cell staining (adherent cells)

3.1 Culture adherent cells on sterile coverslips.

3.2 Remove the coverslip from the culture medium and aspirate excess culture medium.

3.3 Add 100 μL dye working solution, shake gently to completely cover the cells, and incubate for 5-30 minutes.

3.4 Aspirate the dye working solution, wash with culture medium 2-3 times, 5 minutes each time, and observe using a fluorescence microscope or flow cytometer. (Ex/Em=396/610nm), If MitoSOX Red dye excites at 396 nm rather than 510 nm, it may be a more selective marker of mitochondrial superoxide.

Storage Conditions

Store in dark at -20 ℃ for one year

Precautions

1. Please adjust the MitoSOX Red working solution concentration and incubation time according to the actual situation.

2. This product is only for scientific research by professionals and may not be used for clinical diagnosis or treatment, and may not be used in food or medicine.

3. For your safety and health, please wear a lab coat and disposable gloves.