2.12. Filipin staining

A total of 5 × 10⁴ cells/well were seeded onto a sterile slide (Solarbio) placed in a 12-well plate and incubated at 37 °C overnight. After being fixed by 4% PFA, the cells were stained for 2 h with filipin (50 μg/mL, Sigma). Forty microliters/slide of prolonged gold anti-fade reagent (Invitrogen) was used to mount the cells. A DAPI filter was required to view the filipin staining. A fluorescence microscope (Zeiss) was used to scan and record the fluorescence. The experiments were detected in triplicate.