[Histonet] cresyl violet staining on unfixed frozen brain sec tions

[Frank, Matthew]

Sounds like you may be over-differentiating the sections.  I would try going
straight from the staining solution to a water rinse and dehydrate quickly
95% and 100% ethanol.  The staining procedure is simple but you can over
differentiate.

In "Troubleshooting Histology Stains" by Richard Horobin & John Bancroft
1998 it list differentiation as the cause of weak staining.


-----Original Message-----
From: Hector Gonzalez Pardo [mailto:hgpardo <@t> uniovi.es]
Sent: Friday, February 11, 2005 11:41 AM
To: histonet <@t> lists.utsouthwestern.edu
Subject: [Histonet] cresyl violet staining on unfixed frozen brain
sections


Hi everyone!
I am trying to perform Nissl staining using cresyl violet acetate on 30
micrometer-tick cryostat sections from rat brain (I usually don't need
gelatinized slides). After re-hydriting the sections in alcohols (70%, 80%,
96% and 100%), I immerse the sections for 10-15 min in a solution of cresyl
violet acetate (Sigma, 0,5 % at pH 4). I differentiate the sections using
70% ethanol with a few drops of glacial acetic acid (for abut 15-30 secs.),
but when I try to deydrate the sections using 80%, 96% and 100 % ethanol, I
always lose the staing. Finally, I get a blueish pale-looking Nissl staining
on my sections, and I cannot see anything. Does anyone know how to solve it?
I've tried to first de-fat the sections using ethanol for a few minutes but
I did'nt work. Thank you!
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