FerroOrange Fluorescent Method Ferrous Lon Detection Kit

 

Product Name	Cat.No.	Spec.
FerroOrange Fluorescent Method Ferrous lon Detection Kit	G1727-100T	100 T

 

 

FerroOrange, also known as RhoNox-4, specifically detects intracellular free ferrous (II) ions (Fe²⁺). The basic principle of the kit is that after FerroOrange reacts with ferrous ions, an orange fluorescence product is irreversibly generated, and its fluorescence intensity is proportional to the content of ferrous ions in the cell. Neither iron (III) ions (Fe³⁺) nor divalent metal ions other than iron increase their fluorescence intensity. FerroOrange is highly permeable to the cell membrane and can remain in the cell after rapid reaction, so it is suitable for live cell imaging.

Based on the above detection principles, this kit takes FerroOrange as the main body, and provides special supporting buffer and positive inducer. Fluorescence microscope, enzyme labeling instrument and other corresponding instruments can be used to detect ferrous ions in living cells, which is convenient to use and sensitive to detect.

 

 

Ship with dry ice; Store at -80℃ away from light for 6 months.

 

 

Component Number	Component	G1724-100T
G1727-1	FerroOrange fluorescent probe	20 μL
G1727-2	Ferrous ion detection buffer	20 mL
G1727-3	Ferrous ion positive inducer	2×1 mL
Manual		1 pc

Note: The above kit reaction times correspond to the 96-well plate assay system.

 

 

1. Pre-experiment preparation: Cells are inoculated in 96-well plates and pre-treated accordingly to the experimental purpose;

2. Configuration of FerroOrange working solution: Prepare FerroOrange working solution according to the need, the configuration system can be referred to the following table, note that different cell characteristics are different, according to the actual situation of the experiment can be added or reduced the concentration of the probe used, the initial recommended dilution ratio of 1:500; Before preparation, vortex and centrifuge the FerroOrange fluorescent probe briefly to ensure that all reagents are immersed in the bottom of the tube;

 

	Experimental group	Control group
FerroOrange fluorescent probe	0.2 μL	0.2 μL
Ferrous ion detection buffer	100 μL	90 μL
Ferrous ion positive inducer		10 μL

 

3. Staining markers (with adherent cells as an example;Suspension cell treatment requires centrifugation):

3.1. Remove the original cell culture medium and wash the cells 1-2 times with PBS or other buffers;;

3.2. Remove the washing buffer, add 100μL Fluo-4 AM working solution to each well, and incubate in a cell incubator at 37℃ for 30min away from light;

3.3. Remove the FerroOrange working solution at the end of the incubation and wash the cells 1-2 times using PBS or other buffers;

3.4. Add 100 μL PBS or other buffer solution to cover adherent cell, or resuspend cell pellet, for the next fluorescence detection;

4. Fluorescence detection: Use fluorescence microscope, enzyme marker and other instruments to carry out its detection. FerroOrange is orange fluorescent with EX/EM ≈ 543/580 nm. The fluorescence intensity of positive control group treated with positive inducer should be weaker than that of experimental group.

 

 

1. Due to different intercellular properties and different pre-treatment methods, the ratio of probe use and incubation time range described in the steps are for reference only and can be adjusted according to specific circumstances.

2. Fluorescent dyes have quenching problems, please pay attention to avoid the light operation to slow down the fluorescence quenching.

3. FerroOrange is very sensitive to humidity, so please pack it properly according to the experimental arrangement when using it for the first time.

4. FerroOrange can only detect free bivalent iron and cannot detect bound iron.

5. The increase of fluorescence intensity after the reaction of FerroOrange with Fe²⁺ is irreversible and cannot be used for real-time dynamic monitoring of ferrous ion levels.

6. Based on the working principle of this probe, it is not possible to treat the cells such as fixation or perforation.

7. Some precipitation after thawing is a normal phenomenon, and it is fully shaken before use to ensure that it is completely dissolved.

8. For your health and safety, please wear lab coat and gloves during operation.

 

For Research Use Only!