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Fixing Cells with GFP Expression

Fixing Cells with GFP Expression - Protein Science

Fixing Cells with GFP Expression - Post Questions and Discuss Protein Structure and Function, Protein Localization etc.. Please do not post Proteomic questions here, there is a Proteomic Forum below.


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Old 01-29-2009, 05:31 AM
Pipette Filler
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Question Fixing Cells with GFP Expression



Hey guys how to fix cells to retain GFP expression for later analysis?
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Old 01-29-2009, 05:32 AM
Pipette Filler
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Default Re: Fixing Cells with GFP Expression

Hello,
we routinely fix cells with methanol at -20 degrees for 15 minutes. This always gives a good GFP expression. Usually I don't even use an antibody however, a post-doc in my lab is amazed because in his previous lab the methanol killed the GFP.

I'm not sure if it's the quality (we use 99.9% pure) of methanol, the time allowed to fix or the GFP itself (there are now many different mutant forms). I also maintain GFP when fixing with 3.7% paraformaldehyde (PFA) for 20 mins at room temp. Really, the only time I've had trouble maintaining GFP is when trying to co-stain for BrdU but I've got that one worked out now as well.
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