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Attach cells fails to change their morphology after Trypsinization

Attach cells fails to change their morphology after Trypsinization - Stem Cell Forum

Attach cells fails to change their morphology after Trypsinization - Stem Cells and Stem Cell Culture. Embryonic Cell Forum and Mouse Transgenic Forum. Blastocyst injection, ES Cell Selection.


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  #1  
Old 04-02-2010, 11:17 AM
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Default Attach cells fails to change their morphology after Trypsinization



In Trpsinization of Mesenchymal stem cells i am using 0.05% Trypsin with 0.53mM EDTA but i noticed that they take very long for deatachment 20 to 25 min and after trypsinization most of the cells fails to reattach to the palstic surface or if they attach they didnt change thier morphology they remains round so next time i diluted the Trypsin and minimiza the duration of incubation but i got the same results.I use Ca & Mg free PBS for washing of cells and after trypsinization i add complete medium with 10% FBS to stop the action of trypsin.
i fail to sort out the problem .Can anyone give me some suggestion.
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Old 04-06-2010, 10:03 PM
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Default Re: Attach cells fails to change their morphology after Trypsinization

MSCs should detach very quickly. Your cells are probably dying because you're using too low a concentration of trypsin so they're taking way too long to detach.

I use 0.25% trypsin/1mM EDTA for 2-5 minutes, and I tap the underside of the plate and rock it back and forth to help them detach.

Hope that helps.
-Bonnie
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Old 04-07-2010, 11:22 AM
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Default Re: Attach cells fails to change their morphology after Trypsinization

Thanks for your suggestion.
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