[Cell-biology] Re: Cell pellets
(by allison from nospam.com)
Tue Feb 13 11:39:23 EST 2007
Kevin Carbajal wrote:
> It seems silly, but I'm having a horrible time pelleting and
> recovering my mouse embryonic stem cells during a wash-instensive
> protocol that labels them for FACS (TUNEL staining and surface marker
> labeling). I have changed obvious procedures such as centrifugation
> speeds and solution replacing techniques.... but I keep losing too
> many cells during the many washes I have to perform. The pellets seem
> loosely attached to my tubes... I keep ending up with either full,
> half, or no pellets in my tubes.
> Any ideas on how to make sure I don't lose so many cells... or at
> least how to lose them equally in every tube? Thanks!
How are you removing the supernatant after spinning? Can you see your
cell pellet clearly?
I use vacuum and a pasteur pipette with a yellow Eppendorf tip on the
end. The vacuum creates enough suction to 'pick up' the tip from a
racked box. Because the hole in the tip is very fine I find it easy to
aspirate off the supernatant down close to the cell pellet. And it is
easier to change the tip between samples than to change pasteur pipettes.
I also use polystyrene tubes instead of polypropylene - the clear walls
let me see the pellet better.
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