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Early experiments of this investigator used PI for cell cycle analysis, with
little success. Since he has switched to DAPI, we get very nice cell cycle
distributions for his control (no hydroxyurea treatment) cells (nuclei).
Also, it seems that he has achieved the desired block with hydroxyurea.
The problem is that there is a shift to the left (less fluorescence) of the
DNA distribution on the hydroxyurea-treated nuclei relative to the control
nuclei; i.e., the Go/G1 peaks do not occur at the same channel numbers. The
shift is approximately 30 channels on a linear scale. This makes it
difficult to analyze his data, since the Go/G1 peak is hard to distinguish
in cell-cycle blocked nuclei. Though he does not run any internal control
(such as CRBC), it is apparent to me that the changes are not due to machine
variation since I can run the samples multiple times with the same results.
Does anyone have any idea what may be causing this problem? Could the
hydroxyurea be effecting the DAPI staining?
Thanks for your help.
Rick Meister
* * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * *
* Richard K. Meister Email: Meister.1@osu.edu *
* The Ohio State University Voice: (614) 292-9716 *
* Dept. of Veterinary Biosciences FAX: (614) 292-6473 *
* Cytometry Instrumentation Lab *
* 1925 Coffey Road *
* Columbus, OH 43210 U.S.A. *
* * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * *
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