Adherent cells
Chance, Tim (jtc3@CIDDAS1.EM.CDC.GOV)
Tue, 03 Dec 96 14:41:00 EST
You should be able to dissociate cells attached to glass or plastic with a
PBS solution plus 0.53mM EDTA. It is very important that your buffer does not
contain Ca++ or Mg++ or any of the other divalent cations because EDTA
chelates the ions. It also works better if you do not allow the cells to grow
to confluency because they come off in clumps because cell-cell attachment is
more complex than cell to plastic. Wash the monolayer free on media, serum,
etc with PBS+ EDTA then decant this solution and immediately add more
PBS+EDTA and incubate for approximately 15 min at 35c..you may need to "rock"
the flasks gently to help detach the cells...for the stubborn ones, a good
gentle "spanking" of the flask bottom should help finish them off. This
should give you cells with the surface membrane integrity retained. If you
have clumps to remove, Costar makes great cell strainer.. Hope this helps
John Tim Chance
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