cFos IHC in frozen brain

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Foteini Delis
Foteini Delis's picture
cFos IHC in frozen brain

Hi,
I have a bunch of frozen mouse brains stored at -80C that will be sectioned at 15um and thaw mounted on slides for receptor autoradiography.

Does anyone know if I will be able to get a good immunohistochemical reaction for cFos if I post-fix the sections?
Does anyone know which is the best post-fix solution for cFos? 4% PFA? Methanol? Something else?
And which is the most used and reliable antibody?

Thx!
Foteini

Fraser Moss
Fraser Moss's picture
Generally the practice is to

Generally the practice is to perfuse the animal under deep anesthesia with 1 l of heparinized saline followed by 1.5 l of a solution of 4% paraformaldehyde, 15% saturated picric acid and 0.5% of glutaraldehyde in phosphate buffer (PB, 0.1 M, pH 7.4). This is followed by perfusion with 500 ml of the same solution with 10% sucrose .

You'd then remove the brain and place in postfixative solution for 24 h (solution consisting of of 2% paraformaldehyde, 15% saturated picric acid and 10% of sucrose in PB). You'd then place the tissue for 3 days in a solution of sucrose (25%) and sodium azide (0.1%)

I'd be worried about the state of the tissue if your brains were not fixed before you froze them.

A rabbit polyclonal Fos antiserum Fos Ab5; is available from Oncogene Research Products/Calbiochem, La Jolla, CA, USA)