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CAS:62669-70-9
Storage:Powder:2-8℃，2 years;Insolvent(Mother Liquid):-20℃，6 months;-80℃，1 year
Purity:≥98%
Appearance:Red Solid
Rhodamine 123 is a cell staining agent that can stain the mitochondria of living cells. Rhodamine 123 can penetrate the cell membrane and accumulate in the mitochondria of living cells and emit yellow-green fluorescence. Rhodamine 123 is widely used to detect mitochondrial membrane potential and is also commonly used to detect apoptosis. Since there is a correlation between the amount of ATP in cells and the fluorescence intensity of Rhodamine 123, this fluorescent dye is used to detect ATP in cells. Rhodamine 123 has a maximum excitation wavelength of 507nm and a maximum emission wavelength of 525nm. Under fluorescence microscope, the fluorescence was yellowish-green.
How to use (for reference only)
Working fluid preparation
Dilute the storage solution directly with buffer or preheated culture solution to the desired working solution concentration (1-20 μM) and mix thoroughly. The specific working fluid concentration should be adjusted by the user according to their own experimental system.
[Note] : For cell experiments, the overall dilution ratio should be controlled well, and the concentration of DMSO in the culture medium should not exceed 0.1% to avoid the influence of DMSO on cells.
Fluorescence microscopy
1) Prepare the cells with a slide. The number of cells should be 5×104 ~ 5×105 /mL.
2) Incubate the cells on a slide and wash the cells with PBS or HBSS.
3) Rhodamine 123 working solution was added to the slide and incubated at 37℃ for 30 min to 1 hour.
4) Rhodamine 123 solution was removed and cells were washed with culture solution (if cells were fixed after washing, 10% formalin buffer was added and incubated for 15-20 min, then washed with PBS).
5) Observe the cells with fluorescence microscope.
Flow cytometry analysis
1) Cells of logarithmic growth stage were taken, inoculated into pore plates, and cultured overnight.
2) For drug stimulation, add drugs of interest to the cells for intervention, continue to culture for a certain time, collect the cells, and wash the PBS twice.
3) Adding Rhodamine 123 working solution to resuspension cells and incubating at 37℃ for 15 min or longer without light.
[Note] : Due to different cell types and experimental systems, the concentration of Rhodamine 123 working solution and incubation time can be adjusted according to pre-experiments or references.
4) Detected by flow cytometry.
Solarbio experiment Case (for reference only)
1. A549 cells of logarithmic growth phase were placed on a six-well plate (1×106+2mlDMEM medium) and cultured in a 37℃ incubator for 4-24h to be attached to the wall for subsequent experimental operations.
2. Discard the culture medium and wash twice with PBS.
3. 1 ~ 20μM Rh123 working liquid was prepared by diluting Rh123 mother liquid with medium (without serum). The specific working concentration depends on the cell type and cell concentration.
4. Add Rh123 working liquid to six-well plate and incubate at 37℃ for 30 minutes.
5. Remove the Rh123 working liquid and wash the cells three times with PBS to remove the background color.
6. Fluorescence microscope observation.
