From: Phillips, Jim (JPhillip@med.miami.edu)
Date: Thu Dec 11 2003 - 10:08:21 EST
I agree that the concentration of PI would make a difference in the peaks moving.. but usually it would still be a single peak.. My experience with this situation is that the machines' flow cell has a partial block..not a total blockage but enough to throw off the stream and get double peaks.. try using a syringe with a little bit of diluted soap and pump it back and forth. Sometimes it works and sometimes it doesn't. Jim Phillips university of miami school of medicine -----Original Message----- From: Michael G. Ormerod [mailto:Michael_Ormerod@compuserve.com] Sent: Wednesday, November 26, 2003 3:23 PM To: cyto-inbox Subject: Re: Cell cycle conundrum If you are staining at different cell concentrations, depending on the PI concentration, the cells may be taking up different amounts of dye. I would suggest that, after recording the two cell cycles, you mix the two sets of cells in roughly equal numbers. Give the cells an hour or so for the PI to equilibriate and record the DNA histogram. If you still have two G1 and G2 peaks, it appears that either the DNA content (and hence the ploidy) has changed or the chromatin structure has altered to allow the cells to take up more dye. To me, the latter explanantion is less likely. Regards, Michael G. Ormerod, 34 Wray Park Road, Reigate, RH2 0DE, UK +44 (0)1737 241726 mobile: 07802 293242 ----- Original Message ----- From: "Corry, David" <David.Corry@uwe.ac.uk> To: cyto-inbox Sent: Wednesday, November 26, 2003 10:26 AM Subject: Cell cycle conundrum > > Hi Flowers > > I've recieved the following question from a collaborator and it's got me > beat... Can anyone else shed any light on the matter? > > Cheers > > Dave > > We are novices at cell cycle analysis but have begun some expts (on a newly > acquired machine) to study the response to particular transcription factors in > beta cells. We are using an inducible expression system to regulate the > transcript and propidium iodide staining to monitor the cell cycle. Control > cells have a nice profile corresponding to G1 and G2 peaks etc. The induced > cells have a very similar pattern but, in these, the peaks are shifted to the > left (with no change in their profile). This was unexpected and we don't know > how to interpret it......... I am not sure whether you have become skilled in > cell cycle analysis but, if so (or even if not) - do you have any ideas what > this might mean???? The peaks are nice and sharp under both conditions, they > just appear in a different place on the axis. Have you ever encountered this > type of shift? > ---------------------------------------- > David Corry > Centre for Research in Biomedicine > Faculty of Applied Sciences > University of the West of England > 0117 328 3397 > Email: David.Corry@uwe.ac.uk >
This archive was generated by hypermail 2.1.6 : Thu Jan 01 2004 - 17:44:05 EST
