I used to have this problem, but started sonicating my samples just before
reading them (after staining with PI), as well as the sonication step
before fixation. The sonication before reading completely cleaned up the
problem. Good luck.
Todd Hryciw ph:(306)-966-4309
fax:(306)-966-4311
Department of Microbiology
University of Saskatchewan
107 Wiggins Road
Saskatoon SK
Canada
S7N 5E5
On 28 Apr 1998, Robert J.D. Reid wrote:
> Dear Yeast group,
>> I am having trouble with flow cytometry after alpha factor
> synchronization. A significant number of the G1 cells after alpha arrest
> have gone through cytokinesis but remain stuck to their previous bud. I
> sonicated these before flow cytometry, but this did not disrupt the
> pairs. My flow cytometry profile shows two sets of G1 and G2/M peaks
> through the first and second cell cycles. I can gate out the second set
> of peaks, but it is a significant fraction of the population and it
> varies somewhat by sample. Any ideas about breaking up these pairs? I
> am going to try a light zymolyase treatment and perhaps more sonication,
> but if anyone has any ideas or more specific protocols I would appreciate
> the advice.
>> Thanks,
>> Bob.
>
