Cell Cycle Group home pageCell Cycle Group lab membersCell Cycle Group research projectsCell Cycle Group publicationsCell Cycle Group laboratory protocolsLinks to journals, databases and other web sites related to the Cell Cycle GroupCell Cycle Group social events and humourPhD, postdoctoral and other research opportunities in the Cell Cycle GroupContact the Cell Cycle Group MRC Clinical Sciences Centre Cell Cycle Group, Luis Aragón

Yeast protocols, molecular biology protocols, media and general protocols used by the Cell Cycle Group
     Yeast Protocols | Molecular Biology Protocols | Media | General Protocols

Propidium Iodide Staining for FACS Analysis

  1. Add 400 µl (300 µl) culture to 1.2 ml (0.9 ml) 100% ethanol in Eppendorf (volume not critical, therefore depends how much culture can be spared).
  2. 1 h-2+ weeks at 4° C or –20° C.
  3. Spin down cells.
  4. Prepare n x 1 ml SSC (n = number of samples).
  5. From that, prepare n x 0.25 ml SSC + 0.25 µl RNaseA.
  6. Resuspend cells in 250 µl SSC RNase by vortexing.
  7. Incubate at 50° C overnight (or at least 1 h if in a rush — but overnight gives better results).
  8. Prepare n x 50 µl SSC + 1.5 µl proteinase K.
  9. Add 50 µl SSC protK to samples.
  10. Incubate for exactly 1 h at 50° C.
  11. Prepare n x 0.5 ml SSC + propidium iodide (final concencentration is 3 µg/ml).
  12. Add 0.5 ml SSC propidium iodide to samples.
  13. Incubate at room tempearture for 1+ h then store at 4° C until use. (Overnight or over weekend gives best results but can store for at least a month and still get results.)