Novocyte 1000

2 × 10⁵ cells/well were seeded in a six-well plate and incubated overnight. The next day, various concentrations of PeME were treated for 24 h, and then cells were harvested with trypsin-EDTA (Welgene) and transferred into 1.5 ml microtubes (1 × 10⁴ cell/tube). JC-1 dyes were diluted to a final concentration of 2.5 µg/ml. The cells were incubated for 15 min at 21‒23°C in the dark. After incubation, cells were washed with PBS, and resuspended in 500 µl PBS. Flow cytometry analysis was performed immediately. The solutions were divided by NovoCyte 1000 and visualized by NovoExpress (ACEA biosciences).

Each cell was seeded into six-well plates at 2 × 10⁵ cells/well and incubated overnight. PeME was treated into each well and incubated for 24 h. Then, the cells were washed twice and resuspended in 1 ml of 1x Annexin V binding buffer at a concentration of 1 × 10⁶ cell/ml. Following this, 100 µl of the resuspended solution (1 × 10⁵ cells) was transferred into a 1.5 ml microtube and 5 µl of FITC-Annexin V and 5 µl of PI were added. After incubation for 20 min at 21‒23°C in the dark, 400 µl of 1x Annexin V binding buffer was added to each sample. Then, cells were analyzed by flow cytometry using NovoCyte 1000 and visualized by NovoExpress (ACEA Biosciences, San Diego, CA, USA).