Dose-Response Curves for Cisplatin and p53

Dose-response curves were performed essentially as previously described [13 (link),14 (link)]. Briefly, EOC cells were seeded in 96-well culture plates and treated with increasing doses of cisplatin (CDDP) (TEVA Italia) for 72 h. For dose-response curves of p53-silenced MDAH, cells were seeded in 96-well culture plates and after 24 h transduced with sh-ctrl or sh-p53 lentiviral particles (Mission Sigma TRCN0000003756). Forty-eight hours after transduction, cells were treated with increasing doses of CDDP for 16 h and released with complete medium for 24 h. Cell viability was determined by MTS assay using the CellTiter 96 AQueous cell proliferation assay kit (Promega). 2′,7′-dichlorodihydrofluorescein diacetate (H2-DCF-DA) and cycloheximide (CHX) were acquired from Sigma. Double thymidine block was performed incubating cells with thymidine (2.5 mM) (Sigma).