Propidium Iodide Stain

From Genome@Yale

propidium iodide PI stock
RNase A

1. seed 75cm2 plates with 100,000 cells in each flask with 12mL
2. grow 96 hours
3. wash with PBS
4. trypsinize with 0.25X 1mL Trypsin/EDTA solution, 15 minutes at RT
5. quench trypsin with 9mL media + 10% FBS
6. add the detached cells to 15mL conical, spin 3000 rpm for 5 minutes
7. aspirate solution, add 10mL PBS
8. spin at 3000 rpm 5 minutes at 4°C in a centrifuge
9. resuspend cell pellet in 2mL PBS
10. add 8mL 100% EtOH drop-wise while vortexing the 15mL tube
11. incubate O/N at 4°C
12. spin down the cells 3000 rpm 5 minutes
13. resuspend cells in 1mL PI solution (30ug/mL PI, 40ug/mL RNase A in H2O)
14. analyze by flow cytometer