Hct116

The human colorectal cancer cell line HT29 was purchased from Cell Bank of Type Culture Collection of the Chinese Academy of Sciences (Shanghai Institute of Cell Biology, Chinese Academy of Sciences). The human colorectal cancer cell line HCT116, DLD1 and HCT8 were granted by Department of Immunology of the Fourth Military Medical University. HT29 and HCT116 cell lines were maintained in McCOY's 5A medium (MACGENE)supplemented with 10% fetal calf serum (Gibco), DLD1 and HCT8 cell lines were maintained in RPMI 1640 medium (HyClone) supplemented with 10% fetal calf serum(Gibco). These cell lines were incubated in a humidified atmosphere with 5% CO₂ at 37°C. To generate spheroid bodies, single cells were seeded on ultralow attachment plates (Corning) at a density of 1,000 cells/mL. Cells were maintained in DMEM/F12 (Gibco) supplemented with B-27 (Gibco), basic fibroblast growth factor (PeproTech), epidermal growth factor (PeproTech), 0.4% bovine serum albumin (Sigma), and 4 mg/mL Insulin (Sigma) and were incubated in a humidified atmosphere with 5% CO₂ at 37°C. The average number of spheroids (with a size>50μm) was calculated 5-7 days after seeding under microscope.

The human colorectal tumor cell lines HCT8, HT29, HCT116, HCT15, RKO, and CW2 were purchased from the Cell Culture Center of Peking Union Medical College (PUMC, Beijing, China). The human colorectal tumor cell lines HCT8/L with OXA resistance were purchased from ShangHai MEIXUAN Biological science and technology LTD (Shanghai, China). The human normal colon epithelial cell line NCM460 was purchased from Beijing Mintai Yuan Technology Co., Ltd. (Beijing, China). HCT8, HCT15, CW2, HCT8/L, and NCM460 cells were cultured in RPMI 1640 (MacGene, Beijing, China). HT29 was cultured in DMEM/F12 (1:1) (Gibco, CA, USA). HCT116 was cultured in DMEM (MacGene). RKO was cultured in MEM (Gibco). All cells were cultured in medium supplemented with 10% fetal bovine serum (Gibco, CA, USA), 100 mg/mL streptomycin, and 100 U/mL penicillin (MacGene) at 37 °C in 5% CO₂. All cell lines were maintained and used in ≤20 passages.