Which trypsin EDTA to use? - (Jun/21/2010 )

Hi everyone,

I've been culturing some human granulosa cells for a while now and I normally use 0.05% trypsin 0.02% EDTA in hanks balanced salt solution with phenol red to trypsinise them. Last time I trypsinised them I had run out of this trypsin so I used 10x 0.5%trypsin 0.2% EDTA diluted to 1X with PBS instead thinking it would have the same effect. Only a few of the cells would detatch and they ended up clumping together so I had to throw all of the cells away! Does anyone know why the cells reacted differently to this trypsin when the concentrations were the same?

Thanks

-punkyneen-

punkyneen on Jun 21 2010, 04:41 AM said:

Last time I trypsinised them I had run out of this trypsin so I used 10x 0.5%trypsin 0.2% EDTA diluted to 1X with PBS instead thinking it would have the same effect. Only a few of the cells would detatch and they ended up clumping together...
Thanks

was the source of the pbs clean? i.e. no lps?

-telamon-

Does your PBS have magnesium and calcium? Mg and Ca promote cell adhesion.

-navjay-

In addition to navjay's comment; EDTA captures Mg and Ca, so the result is probably that your cells detach from the plate but clump together.

-SusieQ-