β estradiol

The human parental MCF7, T47D and Tamoxifen-resistant (TamR) cell lines were derived from Osborne et al.^{54 (link)}. MCF7 cells were cultured in Dulbecco’s modified Eagle’s medium (DMEM) supplemented with 10% fetal bovine serum (FBS), 2 mM l-glutamine and 1% penicillin/streptomycin (pen/strep) until 90% confluent. For temporal estrogen responsiveness, MCF7 cells were hormone-starved for 72 h followed by the addition of 100 nM β-Estradiol (MP Biomedicals, Inc.) at 1, 4, 16 and 24 h. To hormone-starve MCF7 or T47D cells, these cells were grown to 80% confluency as described above. Once the desired confluency was reached, the cells were washed one time with phosphate-buffered saline and the media was replaced with phenol-red free DMEM supplemented with 5% charcoal-stripped FBS, 2 mM (l-glutamine) and 1% (pen/strep). For the 0 h time point, cells were immediately crosslinked following 72 h of hormone starvation.
TamR cells were cultured in phenol-red free DMEM supplemented with 10% charcoal-stripped FBS, 2 mM l-glutamine, 1% pen/strep, and 100 nM Tamoxifen (Sigma-Aldrich). Tamoxifen was replenished every 48 h and cells were crosslinked at 90% confluency.