- Harvest 2x10⁵ cells per staining and wash cells twice in cold phosphate- buffered-saline (PBS).
- Resuspend cells in 100µl binding buffer (10mM HEPES, pH 7.4, 140mM sodium chloride, 2.5mM calcium dichloride), add 5µl annexin V-Alexa Fluor^® 488 to the cells and mix gently.
  For detection of necrosis add additionally appropriate amounts of propidium iodide (final concentration 1µg/ml or 1.5µM).
- Incubate for 15 min. at room temperature in the dark.
- Add 400µl binding buffer and analyze the staining in flow cytometry as soon as possible (within an hour).

Swiss-Prot link P48036: Annexin V (mouse)
AfCS Signalling Gateway link A000281: Annexin V (mouse)