RhoNox-1,IC-0121510

RhoNox-1 is a fluorescent probe for the specific detection of divalent iron ions, and when RhoNox-1 reacts with Fe2+. RhoNox-1 can generate an irreversible orange (red) fluorescent product (Ex/Em£º540/575 nm) . FeRhoNox-1 can enter the cell well, suitable for the detection of Fe2+ in living cells, and tends to be localized in the Golgi apparatus[1].

Usage
1. Preparation of RhoNox-1 working fluid
1.1 Preparation of storage solution
Prepare 50 ¦Ì g of RhoNox-1 with 110 ¦Ì L anhydrous DMSO to obtain a 1 mM stock solution.
1.2 Preparation of working fluid
Dilute the storage solution with preheated serum-free cell culture medium or PBS to prepare a 1-10 ¦Ì M RhoNox-1 working solution.
Note: Please adjust the concentration of RhoNox-1 working solution according to the actual situation, and use and prepare it now.
2. Cell staining (suspended cells)
2.1 Suspension cells: Collect cells by centrifugation and wash twice with PBS for 5 minutes each time. Cell density is 1 ¡Á 106/mL
2.2 Add 1 mL of working solution and incubate at room temperature for 5-30 minutes.
2.3 400 g, centrifuge for 3-4 minutes, discard the supernatant.
2.4 Wash the cells twice with PBS, each time for 5 minutes.
After resuspending cells in 1 mL serum-free medium or PBS, observe them using a fluorescence microscope or flow cytometer.
3. Cell staining (adherent cells)
3.1 Cultivate adherent cells on sterile coverslips.
3.2 Remove the cover glass from the culture medium and aspirate excess culture medium.
3.3 Add 100 ¦Ì L of dye working solution, gently shake to completely cover the cells, and incubate at room temperature for 5-30 minutes.
3.4 Remove the dye working solution, wash 2-3 times with culture medium for 5 minutes each time, and observe using a fluorescence microscope or flow cytometer.

[1]. Mukaide T, et al. Histological detection of catalytic ferrous iron with the selective turn-on fluorescent probe RhoNox-1 in a Fenton reaction-based rat renal carcinogenesis model. Free Radic Res. 2014 Sep;48(9):990-5.
[2]. Jamnongkan W, et al. Upregulation of transferrin receptor-1 induces cholangiocarcinoma progression via induction of labile iron pool. Tumour Biol. 2017 Jul;39(7):1010428317717655.  
[3]. Ito F, et al. Contrasting intra- and extracellular distribution of catalytic ferrous iron in ovalbumin-induced peritonitis. Biochem Biophys Res Commun. 2016 Aug 5;476(4):600-606.