Western blot analysis of proteins related to apoptosis. MCF-7 cells at 80% confluence were treated with 30 μM chalcone-imidazolone conjugates for 24 h. 50 μg protein from total cell lysate was run on 10-15% SDS-PAGE and protein expression was detected by Western blotting. The expression of pro-apoptotic protein (Bax), active caspase-9 and the protein that undergo cleavage during apoptosis (Rb), the retinoblastoma protein was determined by using anti-Bax, anti-active caspase-9 and anti Rb-cleavage specific antibodies. β- actin was used as loading control. Here C represents control untreated cells.