From: Albert Tai (acktai@exelixis.com)
Date: Tue May 20 2003 - 17:03:10 EST
Hi Roger, Did you fix MCF-7 prior to the PI staining ? My experience was that fixing the cells with cold ethanol by adding dropwise to cells while vortexing tends to help prevent cell aggregation. After fixation, I usually wash the cells twice with PBS before staining with PI/RNase mixture. Hope this help. Albert Tai Exelixis Inc. SSF, Calif ----- Original Message ----- From: "Roger Smith" <rosmith@cvm.tamu.edu> To: cyto-inbox Sent: Monday, May 19, 2003 2:37 PM Subject: sticky problem with MCF-7 cells > Greetings, > > I work with several students from a lab that uses MCF-7 cells for many > of their studies. My involvement is cell cycle analysis with PI. The > problem is that there are always significant numbers of aggregates, > some of which clog the instrument even after filtering. They lift cells > off the plates with trypsin (probably with EDTA, but the student today > was not sure) and syringe them with 25 ga needles. Still, there are > aggregates. people in the same lab working with other cell lines have > no problem obtaining single cell suspensions. They tell me that MCF-7's > are very difficult to disaggregate. Any suggestions? > > thanks, > Roger > > ---------- > Roger Smith III > Dept. Veterinary Pathobiology > Texas Veterinary Medical Center > Texas A&M University > College Station, TX 77843-4467 > voice: 979-845-5167 > fax: 979-862-2344 > e-mail: rosmith@cvm.tamu.edu >
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