APF 

APF is a cell-permeable fluorogenic probe to measure hydroxyly radical of ROS. APF is nonfluorescent and produces bright green fluorescence upon reaction with hydroxyl radical. APF can be used to detect intracellular reactive oxygen species (ROS) (Ex/Em=490/525 nm)^[1].

Guide (The following is our recommended protocol. This protocol is only a guide and should be modified according to your specific needs).
1.1 Preparation of stock solution
Use anhydrous DMSO to prepare a 10-20 mM APF stock solution.
Note: It is recommended that the APF stock solution be stored at -20 °C or -80 °C in the dark after aliquoting.
1.2 Preparation of working solution
Use preheated serum-free cell culture medium or PBS to dilute the stock solution to prepare a 1-10 μM APF working solution.
Note: Please adjust the concentration of APF working solution according to actual conditions and prepare it before use.
2. Cell staining (suspended cells)
2.1 Collect cells by centrifugation and wash twice with PBS for 5 minutes each time. The cell density is 1×10⁶/mL.
2.2 Add 1 mL of dye working solution and incubate at 37 °C for 20-60 minutes.
2.3 Centrifuge at 400 g for 3-4 minutes and discard the supernatant.
2.4 Add PBS to 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 the adherent cells on a sterile coverslip.
3.2 Remove the coverslip from the culture medium and remove the excess culture medium.
3.3 Add 100 μL of dye working solution, gently shake to completely cover the cells, and incubate at 37 °C for 20-60 minutes.
3.4 Aspirate the dye working solution, wash 2-3 times with culture medium, 5 minutes each time, and observe using a fluorescence microscope or flow cytometer.
Note: If flow cytometry is required, the cells need to be digested with trypsin and resuspended before staining.

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

423.42

C₂₆H₁₇NO₅

359010-70-1

Solid

Yellow to orange

O=C1OC2(C3=CC=C(OC4=CC=C(N)C=C4)C=C3OC5=CC(O)=CC=C25)C6=CC=CC=C16

Room temperature in continental US; may vary elsewhere.

-20°C, protect from light

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

* Please refer to the solubility information to select the appropriate solvent. Once prepared, please aliquot and store the solution to prevent product inactivation from repeated freeze-thaw cycles.
Storage method and period of stock solution: -80°C, 6 months; -20°C, 1 month (protect from light). When stored at -80°C, please use it within 6 months. When stored at -20°C, please use it within 1 month.

• [1]. Setsukinai K, et al. Development of novel fluorescence probes that can reliably detect reactive oxygen species and distinguish specific species. J Biol Chem. 2003;278(5):3170-3175.  [Content Brief]