Mcf 10a

All cell lines and the materials used for cell culture were ordered from Procell (Wuhan, China, https://www.procell.com.cn/) unless specified otherwise. Human breast epithelial cell MCF-10A (CL-0525) and BC cell lines MCF-7 (CL-0149), MDA-MB-231 (CL-0150), MDA-MB-436 (CL-0383), and SK-BR-3 (CL-0211) were ordered and cultured as needed.
MCF-10A cells were maintained in Dulbecco’s modified Eagle’s medium/F12 medium (PM150312) blended with 5% horse serum (164215), 20 ng/mL of epidermal growth factor (P00033, Solarbio Lifesciences, China), 0.5 μg/mL of hydrocortisone (G8450, Solarbio Lifesciences, China), 10 μg/mL of insulin (I8830, Solarbio Lifesciences, China), 1% non-essential amino acid (PB180424), and 1% penicillin−streptomycin (PB180120). MCF-7 cell was cultured in minimum essential medium (PM150140) with 0.01 mg/mL of insulin. Leibovitz’s L-15 medium (PM151010) was used to ensure the growth of MDA-MB-231 and MDA-MB-436 cells. SK-BR-3 cell was grown in McCoy’s 5A medium (PM150710). All media for BC cells were supplemented with 10% fetal bovine serum (164210) and 1% penicillin−streptomycin.
All cells were finally incubated in the Sanyo MCO-18AIC(UV) CO₂ incubator (SA-MC018, Marshall Scientific, LLC., Hampton, NH, USA) at 37°C and 5% CO₂. As TMTC3 was lowly expressed in MCF-7 and MDA-MB-231 cells, these two cells were used for subsequent studies.

Human breast cancer cell lines MCF-7, BT-474 and BT-549 and normal human breast epithelial cell line MCF-10A were obtained from American Type Cultural Collection (ATCC). MCF-10A were maintained in DMEM/F12 (Solarbio Science & Technology Co., Ltd, Beijing, China) supplemented with 10% fetal bovine serum (FBS) and 1% penicillin and streptomycin solution plus 3.5 μg/mL human insulin, 20 ng/mL epidermal growth factor and 0.5 μg/mL hydrocortisone. MCF-7, BT-474 and BT-549 were maintained in RPMI 1640 (Solarbio Science & Technology Co., Ltd, Beijing, China) supplemented with 10% fetal bovine serum (FBS) and 1% penicillin and streptomycin solution. These cells were cultured in an incubator with a 5% CO2 humidified atmosphere at 37°C. All reagents were commercially obtained from the Procell Life Science&Technology Co., Ltd (China)