Re: Phase of Cell Cycle at Time of Death

Dear Ann, 
If there are many dividing cells in a cell population that is stained with a nuclear
viability dye, like PI or 7-AAD, a sign of good staining that we usually see is that
dead, stained cells form two peaks with the second peak being of double intensity of the
first one. The first peak is made of cells in G0/G1 and the second one of G2/M, i.e. 4N
cells. In other words, the dead cells produce the same PI staining picture as cells
permeabilised for cells cycle analysis. Because it is usually on a log scale the peaks
are close to each other and the S phase area is not clear, but this look may give some
idea of how many cells are cycling and in what phase they are.
If the phases need to be recognised more precisely PI fluorescence can be displayed as
linear and the voltage adjusted so as the dead cells are seen similarly to the usual view
of cells permeabilised for cell cycle analysis.
I think this is the simplest approach and the above is OK with cells that died recently,
but if, like in your case, they died some time ago a potential complication may be that
they may have lost some DNA by the time of staining.
Regards.
Vladislav Rozenkov, MD, PhD
-----Original Message-----
From: Byrne, Ann <Ann.Byrne@genzyme.com>
To: cyto-inbox
Sent: Mon, 10 Apr 2006 16:34:25 -0400
Subject: Phase of Cell Cycle at Time of Death


Hello,
I stained some cells with PKH67.  The cells were treated with a test compound and then
incubated for three days.  Samples taken on each day were also stained with propidium
iodide.  The PKH fluorescence on the PI positive cells gives some information about when
the cells died.  Is it possible to get further information about what phase (of the cell
cycle) each cell was in when it died?	
Regards,
Ann
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