Re: GFP/fixation/paraffin sections

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From:Jamie Erickson <>,
Date:Wed, 28 Apr 1999 17:03:26 -0400

Hi All,
          Here is what I known about the GFP that our molecular group used in our GFP experiments, the GFP is from Clonetech (pEGFP-n1 MCS cat # 6085-1) and I use for the VIP processing I use Ethanol and Xylene in the VIP. 
         The first GFP experiment we did that worked with paraffin processing was one  to evaluate GFP as an alternate method of in-vivo detection of transfection. We used B-gal in one mouse and GFP in another to see if we could get comparable expression. 
We only looked at day 4 but previous results with B-gall showed maximum liver expression at around day 7. It was hard to compare B-gal and GFP samples, GFP seemed to be specific where the 
B-gal livers were stained very blue and it was hard to tell specifically how many cells or what cells were staining. We really should do a longer time course to find out maximal expression.
I have not tried Immuno for GFP on paraffin samples for GFP due to the success of the paraffin processing
               We also did frozen sections that were fixed for 24 hours in 10% Neutral buffered Formalin and that improved the morphology greatly but this was still not as good as paraffin.  The Journal that this image is cover of is the  Journal of Toxicologic Pathology   Vol 27, Number 1, Jan-Feb 1999. It is just the cover no article or method of GFP staining.



>>> Gayle Callis <> 04/28 1:24 PM >>>
I had (much to my happiness!) two replies from people who had used
GFP with SUCCESS in formalin fixed tissues/paraffin processing.  

Could it be that the GFP they are using is a stable form (heat stabile,
resistant to solvents?).  If so I am hoping they share the specifics
of what was done, exact GFP used (there are chimeras out there!) and
some of the processing times/solvents/paraffin.  

At least we had one thing in common, formalin fixation of tissues, time
of fixation may be a big factor also.

ladies, are you willing to share these goodies, it would be a big help.

Gayle Callis


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