[Histonet] GFP IHC frozen samples

[anh2006 <@t> med.cornell.edu]

GFP is not destroyed in frozen sections but the tissue should be PFA fixed prior to sectioning, ideally. 

If you have already prepped your tissues and have no choice but to proceed with fresh frozen be aware that GFP is soluble and can leach out of fresh frozens. In addition, in fresh frozen preps it is deleterious to let them dry too much as GFP will quench when dried.

I suggest you fix your tissue immediately after sectioning and just put it on the scope (after coverslipping obviously) and see what you see, and go from there.

For anti-GFP use the rabbit anti-GFP from Molecular Probes/Invitrogen. It works well in frozen and paraffin, immunofluorescence and chromogenic.




-----Original Message-----
From: Garry Ashton <GAshton <@t> picr.man.ac.uk>

Date: Thu, 16 Jul 2009 12:01:32 
To: <histonet <@t> lists.utsouthwestern.edu>
Subject: [Histonet] GFP IHC frozen samples


Hi all,

 

I spent hours and looked at several antibodies on this subject.

 

Does anybody have a protocol that actually works for  GFP staining on
fresh frozen samples or is the signal simply destroyed?

Many thanks 
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