[Histonet] Immediate Early Gene expression in FISH

[yangpw <@t> umich.edu]

HI, 

I have been trying to do Fluorescent In situ for Immediate Early Gene (IEG such
as c-fos, arc, etc.). I use riboprobe and tried bio-UTP/dig-UTP, rabbit
anti-bio/dig, and then use fluorescein conjugated anti rabbit. I tried same
protocol on enk mRNA and it turned out just fine. But IEG signals were always
weak. Anything I can do to improve? Do I need to use any amplification kit like
TSA? Any suggestion is welcome. Also I tried Alexa 350 for blue fluorescence,
it seems photobleach very quick even with anti-fade media. Under microscope in
couple minutes, it is fading. Does anyone have any experience on this? Which
blue fluorescein is good? THanks a lot.

Pengwei Yang, MD
University of Michigan