4.10. Analysis of Cellular Apoptosis by Hoechst 33342 and SYTOX™ Green Double Staining

Apoptosis and necrosis were investigated in MCF-7 cells by double staining assay. Cells were seeded in a 96-well plate and transfected with siRNA as described in the transfection section. After 48 h of treatment, cells were then stained with a mixture solution containing 5 µg/mL of Hoechst 33342 and 5 µg/mL of SYTOX™ Green (Invitrogen, Carlsbad, CA, USA) for 15 min. Within 1 h after staining, a picture of the cells was taken under an inverted fluorescence microscope (Eclipse TE 2000-U; Model: T-DH Nikon^®, Tokyo, Japan). The apoptosis cells (bright blue fluorescence) and the death cells (green fluorescence) were quantified and presented as percentages from three different microscopic fields.