Culturing and Sorting CNE-2 Cell Line

The CNE-2 cell line was presented from Prof. Yunfei Xia at Sun Yat-sen University Cancer Center (Guangzhou, China)21 (link). CNE-2 cell was cultured in RPMI-1640 medium (Hyclone, Logan, UT, USA) with 10% fetal bovine serum (Hyclone), 100 units/ml penicillin and 100 mg/ml streptomycin (Hyclone). The Sorting of cell populations was referred to a previously reported method 22 (link). Sorted CNE-2s cell was cultured in DMEM/F12 (1:1) medium (Hyclone) with 20μg/L EGF (PeproTech, Rocky Hill, USA), 20μg/L bFGF (PeproTech) ,2% B27(no vitamin A, Gibco, Carlsbad, Calif, USA),100 units/ml penicillin and 100 mg/ml streptomycin (Hyclone). All the two cell lines were cultured in a 37℃ incubator (Sanyo, Japan) with 5% CO₂. Cells were digested by 0.25% trypsin and 0.02% EDTA solution (Sigma, St. Louis, MO, USA).