2.12 Iron content analysis
The intracellular iron level was measured by a calcein-AM kit. After cell plating and drug treatment for 24 h, HK-2 cells were treated with AM (2 μM) in the dark at 37 ℃ for 30 minutes. Next, the cells were washed and analyzed with a fluorescence microscope. To perform fluorescence imaging of Fe2+ in living cells, HK-2 cells were inoculated into confocal dishes and cultured overnight at 37 ℃ in a 5% CO2 incubator. FerroOrange working solution (1 μM), purchased from Dojindo (Kumamoto, Kyushu, Japan), was added to the cells, which were cultured in a 5% CO2 incubator at 37 ℃ and then observed by a fluorescence confocal microscope. In vivo, an iron assay kit was used to detect the iron ion content in mouse serum, which was determined by detecting the absorbance at 405 nm.