Figure 3.

Annexin V is an easy non-toxic, and robust reporter for use in high-contrast cell imagers. (a and b) MEFs were treated with either vehicle, CHX (50 μg/ml) alone, or CHX (50 μg/ml) and ABT-737 (1 μM), co-incubated with both Annexin V-488 (1 μg/ml) and YOYO3 (200 nM), and scanned at 2-h intervals up to 24 h for green or red fluorescence, respectively. (c) Graph depict the difference of values collected at each time point from cells in (a and b). (d) Data from (a and b) represented as bar graphs for comparison between Annexin V- and YOYO3-labelling at indicated time points. (e) Captured images from IncuCyte Zoom using a × 10 objective for CHX-treated cells from (a and b); 50 μm scale bars. Data shown in (a—e) are from a single experiment and representative of at least three experiments. (f) MEFs treated as in (a) but analysed by flow cytometry. (g) MEFs treated with CHX (50 μg/ml) and ABT-737 (1 μM) were co-incubated with Annexin-594 (1 μg/ml) and CellEvent Caspase-3/-7 Green Reporter (labelled as ‘DEVD’, 2 μM), and scanned every 2 h for 24 h. (h) MEFs treated as in (g) were incubated with either Annexin-488 (1 μg/ml), YOYO3 (200 nM) or DEVD reporter (2 μM), and scanned repeatedly every 2 h for 48 h to assess photostability