Eve plus automated cell counter

BJ-5ta and KerCT, hTERT-immortalized cell lines used in this research were acquired from the ATCC collection. Cells were cultured as described previously [47 (link)]. Briefly, cells were cultured under standard aseptic conditions (37 °C, 95% humidity, 5% CO₂) using KGM-Gold medium with supplements and penicillin/streptomycin for KerCT cells and a 1:4 mixture of Medium 199 and DMEM with the addition of 10% FBS, penicillin/streptomycin, and 0.01 mg/mL hygromycin B for BJ-5ta cells. After retrieving the cells from cryostorage, the cells were passaged at least twice before experiments. EVE Plus Automated Cell Counter (NanoEntek, Seoul, Korea) was used for cell counting.

KerCT and BJ-5ta acquired from the ATCC collection were used in the research. Both cell lines are hTERT-immortalized. The BJ-5ta were cultured in a 4:1 mixture of DMEM and Medium 199 with the addition of 10% FBS, 50 U/mL of penicillin, 50 µg/mL of streptomycin, and 0.01 mg/mL hygromycin B. The KerCT were cultured in a KGM-Gold medium with supplements and 50 U/mL of penicillin, and 50 µg/mL of streptomycin. The cells were cultured under standard aseptic conditions (37 °C, 95% humidity, 5% CO₂). The cells used for the experiments (following between 3 and 18 passages) were grown in continuous cultures and were passaged after reaching 80–90% confluence with TrypLE. After thawing from the cell bank, the cells were passaged at least twice before experiments. After trypsinization, the cells were centrifuged at 400× g for 5 min. Finally, the cell pellets were resuspended in 1 mL of fresh media and counted with EVE Plus Automated Cell Counter (NanoEntek, Seoul, Korea).