Mammary epithelial cell growth media

MDA-MB-231 cells (ATCC HTB-26, Manassas, VA, USA) were cultured in DMEM (Cellgro, Manassas, VA, USA) with 10% fetal bovine serum and 1% penicillin–streptomycin. MCF-10A cells (ATCC CRL-10317) were cultured in mammary epithelial cell growth media (Lonza, Walkersville, MD, USA) with 1% penicillin–streptomycin. Cells were authenticated by STR profiling (ATCC). CT20p (Ac-VTIFVAGVLTASLTIWKKMG-NH2) and RHO-tagged CT20p were commercially synthesized (Biopeptide Co., Inc, San Diego, CA, USA) at >98% purity. Human tissues were transported in RPMI on ice. Upon receipt, tissues were washed thoroughly in PBS containing 1% penicillin–streptomycin and 5 μg/ml Fungizone (Life Technologies, Carlsbad, CA, USA). Tissues were minced and digested in 0.1% collagenase I (Gibco, Carlsbad, CA, USA) at 37 °C for 2 h. Differential centrifugation as described by Speirs et al.^{34 (link)} was used to obtain an epithelial-enriched cell fraction. Cells were maintained in mammary epithelial cell growth media (Lonza) at 37 °C and 5% CO₂. Human tissues were deidentified and the protocol for use was approved by the Institutional Review Boards at the University of Central Florida and Florida Hospital.