The University of Iowa Carver College of Medicine
DNA Staining with PI
The following protocol is for DNA staining with propidium iodide for cell cycle analysis (complex hypotonic solution).
- Stock Solution
- 20mg/ml propidium iodide (in 1X PBS) [store refrigerated, protected from light]
- 10X Lysis Buffer
- 10mM TRIS (1.211g/L in dd H2O) [MW=121.1g]
- 1mM EDTA (0.3722g/L in dd H2O) [EDTA Disodium Sulfate2H2O MW= 372.2g]
- 1% Triton X-100
- Store at room temperature
- Hypotonic Staining Solution (for 50ml)
- 50mg sodium citrate
- 5ml lysis buffer
- 45ml ddH2O
- Mix until sodium citrate dissolved
- 125µl stock propidium iodide solution
- Store refrigerated and protected from light [good for at least 2 weeks]
- Pellet 2 x 106 cells.
- Carefully aspirate supernatant leaving as little buffer as possible without aspirating cells.
- Add 1ml hypotonic propidium iodide staining solution.
- Store on ice for at least 15 minutes before running on flow cytometer.