Detection of Ferroptosis

The relative level of intracellular iron was detected by an Iron Assay Kit (Abcam, USA) following manufacturer’s protocol. The lipid ROS was detected by using C11-BODIPY (Invitrogen) staining. In brief, cells were planted on 12-well plates and incubated overnight. After indicated incubation, cells were collected and resuspended in 200 μL PBS, stained with C11-BODIPY (5 μM) at 37°C for 30 minutes. Samples were examined by flow cytometer (BD, USA). The level of malondialdehyde (MDA) was assessed by using a lipid peroxidation (MDA) assay kit (Sigma). Cells were lysed by MDA lysis buffer on ice, followed by centrifugation at 13,000 × g for 5 minutes. The absorbance values were detected at 532 nm by a spectrophotometer (BD Biosciences).