Murine TNBC and ER+ Cell Culture Protocol

The 4T1 and TS/A spontaneous murine cells are TNBC and ER+ phenotypic lines, respectively, originated in BALB/C mice [37 (link),38 (link),39 (link),40 (link)]. The 4T1 (ATCC) cells were cultured in RPMI 1640 medium supplemented with 10% FBS, 100 units of penicillin, and 100 μg/mL of streptomycin. TS/A cells (Sigma-Aldrich Inc., St. Louis, MO, USA) were grown in Dulbecco’s modified MEM (GIBCO, Life Technologies, Monza, MB, Italy) supplemented with 2 mM of glutamine, 100 U/mL of penicillin, 100 μg/mL of streptomycin, and 10% heat-inactivated fetal calf serum (GIBCO, Life Technologies, Monza, MB, Italy) at 37 °C in humidified 5% of CO₂ atmosphere. Culture medium was changed every 2–3 days, and cells were passaged with 0.25% trypsin/EDTA. Cells were routinely tested for Mycoplasma using a MycoAlert mycoplasma detection kit (BioWhittaker-Lonza, Euroclone S.p.a., Milan, Italy).