Multimodal Cell Death Assay

Cell death was measured using a Cell Meter Apoptotic and Necrotic Detection kit (ATT Bioquest, Sunnyvale, CA, USA) as previously described.^{24 (link)} In brief, cells were incubated at 37 °C for 30 min with Apopxin Green for detection of phosphatidylserine on cell surface, PI or 7-ADD for labeling the nucleus of cells with membrane rupture, and CytoCalcein for labeling live cell cytoplasm. Cell death was then analyzed with an EVOS FL digital fluorescence microscope (AMG) or a FACSCalibur flow cytometer (BD Biosciences, San Jose, CA, USA). Cells with chromatin condensation were visualized by Hoechst 33342 (Invitrogen, Waltham, MA, USA) staining. Cell viability was also assessed using the Muse Count & Viability assay kit (Millipore, Billerica, MA, USA). In brief, cells were trypsinized, washed, and incubated with the Muse Count & Viability reagent, and cell viability was quantified on a Muse cell analyzer (Millipore).