Tumor dissociation

Kevin G Waddick waddi002 at tc.umn.edu
Tue Aug 25 13:10:23 EST 1998


   I would appreciate it if someone would provide me with the benefit of
their experience. When using adherent tumor cell lines, and wanting to get
them into suspension without using trypsin, a cell dissociation solution
from Gibco/BRL has worked well. Does anyone know the formula for this
solution (it is not listed by Gibco/BRL). Before this solution was substituted,
we used an EDTA/PIPES solution. Unfortunately, I never wrote down the
concentrations of these chemicals used in the mixture; does anyone have a
protocol for making and using this solution they would be willing to
share?
   My next question is about the best way to dissociate the cells from a
primary patient tumor for analysis and sorting by flow cytometry. In the
past, I used a tissue grinding pestle in a glass tube to break down organs
ex vivo and combined this with incubation in trypsin. Does anyone know of
a better way to do this such that degradation of cell-surface proteins by
trypsin can be avoided (or maybe an optimal concentration and incubation
time for trypsin that combines a good cell dissociation with minimal
protein digestion)? Do the solutions mentioned in the previous paragraph
work when used with solid tumors? Or are there better ways to do what I
want to do? Whatever seems to work well for you, could you outline for me
how it is done or provide me with a reference?
   I would be grateful for assistance in learning this without having to
put in a lot of time in the library.  

Kevin G. Waddick, Ph.D.
2520 Silver Lane NE, Apt. 201
St. Anthony, MN  55421




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