[Histonet] IHC on 3 year fixed tissue?

[Kevin Egan]

Hello Histonetters,

I've been asked to perform anti-GFP IHC on some mouse bones which have 
been sitting in NBF for 3-4 years. My Abcam rabbit polyclonal gets great 
results in GFP mouse control tissue, but I can't seem to get any results 
out of these bones. I've tried citrate buffer HIER using autoclave, 
hotplate, and 65 degree water batch over night, but my signal is 
non-existent (or really the noise is so huge you can't distinguish).Do 
the great minds of histonet have any ideas on how to achieve good 
staining? Is enzymatic epitope retrieval worth  a try? Does anyone have 
a time machine I can use to stop my predecessors from just sticking the 
bones in NBF and forgetting about them for years?

This listserv has helped me countless times, I am truly appreciative of 
the knowledge and experience available on Histonet.

Thanks,
Kevin