Flow cytometry

An Annexin V-Fluorescein Isothiocyanate (FITC)/Propidium Iodide (PI) Staining kit (BD Biosciences, Franklin Lakes, NJ, USA) was used to detect apoptotic cells. Cells were harvested 48 h following transfection and resuspended in 1X binding buffer at a concentration of 1×10⁶ cells/ml. Subsequently, cells were stained with 5 µl Annexin V-FITC and 5 µl PI for 15 min at room temperature in the dark. Cells were acquired using a BD Accurri^™ C6 flow cytometer (BD Biosciences) and data was analyzed using FlowJo software version 7.6.2 (Tree Star, Inc., Ashland, OR USA).