To begin, incubate ferroptosis-induced medulloblastoma cells with body PC-11 and remove the staining solution under a laminar hood. After washing the cells with two milliliters of PBS, incubate the cells with 150 microliters of a cell dissociation reagent. To mechanically harvest the cells, add 250 microliters of flow cytometry buffer, and tap the plate gently.
Then, transfer the cells into a labeled flow cytometry tube with a filter cap. Next, turn on the FACS machine. Create a new experiment and choose the FITC filter and 488 nanometer laser.
In View Data, select the cell size greater than 12 micrometers and gate the dot plot. Then add a new plot histogram with FITC-A on the axis and logarithmic scale. Then, vortex the sample taken in the tube and load it into the FACS machine.
Finally, record 10, 000 forward scatter singlet events. To stain the dead cells, add propidium iodide solution just before the FACS analysis. Flow cytometry analysis of lipid hydroperoxidation, verified ferroptosis-induction in medulloblastoma cells treated with the erastin.
