Annexin V-FITC Apoptosis Assay

The assay was carried out using Annexin V FITC Kit (BD Pharmigen, Franklin Lakes, NJ, USA) according to manufacturer’s protocol to evaluate the apoptosis induction of rAF-IL12 towards cancer cells. Briefly, the seeded cells (2 x 10⁵ cells/well) were infected for 72 h before the cell pellets were harvested and resuspended in the 1X Binding buffer prior to staining with FITC Annexin V and propidium iodide (PI) dyes as described by Najmuddin et al. [19 (link)]. The cell suspension was then allowed to stand in the dark at room temperature for 15 min before being analysed by NovoCyte Flow Cytometer (ACEA Biosciences Inc., San Diego, CA, USA) using NovoExpress^® version 1.2.4 software (ACEA Biosciences Inc., San Diego, CA, USA).