D8418 250ml

SCAP at a passage of 3–5 were used in this experiment. SCAP were isolated from a human tooth. We obtained Institutional Review Board approval at Seoul National University Hospital (Seoul, South Korea; IRB number CRI05004). Informed consent was obtained from all participants. All experiments were performed in accordance with the Declaration of Helsinki. The manuscript does not contain information or images to identify study participants. Therefore, patient consent for the publication of our experimental data does not seem necessary. Basal media is composed of Alpha-MEM, FBS, and an antibiotic (all of them from Welgene, Korea) at a ratio of 100:10:1. This was used as a complete media. The cells were cultured in an incubator (371 Steri Cycle CO₂ Incubator, Thermofisher, USA) at a temperature of 37 °C and 5% carbon dioxide. For cryopreservation, EDTA-trypsinized cells were suspended in freezing media of basal media, FBS, and dimethyl sulfoxide (DMSO) (D8418-250ML, Sigma, USA) in a ratio of 6:3:1. A mixture of cells and freezing media was placed in a freezing vial and placed in isopropanol at − 70 °C for 12 h. After 12 h, freezing vials were moved to the liquid nitrogen tank for more than 24 h. Frozen cells were used in experiments after thawing in a 37 °C-waterbath for 2 min.