Supplementary Figure 2: The nucleic acid-binding dye TO-PRO-3 stains viable, early apoptotic and necrotic cells differentially.
From: Monitoring the progression of cell death and the disassembly of dying cells by flow cytometry
a, Histogram showing TO-PRO-3 uptake by viable, A5- early apoptotic, A5+ early apoptotic and necrotic cells. b, Uptake of TO-PRO-3 by cells at different stages of cell death. Data is presented as median fluorescence intensity (MFI) relative to viable cells (n = 3, experimental replicates). Human Jurkat T cells were induced to undergo apoptosis by anti-Fas treatment (31.25 ng ml−1, 4 h). Samples were stained with A5-FITC (1 in 200 dilution), 7-AAD (1 μg ml−1) and TO-PRO-3 (0.5 μM) in 1x A5 Binding Buffer. 30,000 events (cells and cell fragments) were recorded by the FACSCanto II flow cytometer. Resultant data were analysed by the FlowJo 8.8.6 software using the electronic gating strategy as described in Figure 3. Alternative STAGE 1 gating step was used to identify necrotic cells based on the 7-AAD parameter. Error bars represent standard error of the mean. Data are representative of at least two independent experiments.
