Karen:
I don't have any references to the use of cold
fixatives for this purpose, but I can't believe that
perfusion with cold fixative would work that well. 
The vasculature will contract upon the initial
exposure to the cold unless a relaxant has been used,
and even then I'm not sure that this will obviate that
problem.  I have always used fixative that is at least
at room temp, and in some instances have run the
perfusion tubing through a warm water
bath--particularly since we were looking at the blood
brain barrier.

Roger Moretz
Dept of Toxicology
Boehringer Ingelheim Pharmaceuticals, Inc.


--- Karen Larison <larisonk@uoneuro.uoregon.edu>
wrote:
> Histonetters,
> 
> One of the labs here is doing in situ hybridzation
> on developing rat brains.  They 
> have been using ice-cold perfusate on these rats as
> they believe that this prevents 
> endogenous RNAses from acting.  I have a hard time
> believing this, particularly in 
> light of the evidence that paraformaldehyde fixation
> effectively neutralizes 
> endogenous RNAses (J Neurosci Meth 85 (1998) pp
> 129-139).  If this is true, why would 
> you want to slow the fixation rate by perfusing with
> ice-cold perfusates?
> 
> If anyone knows of any references that would clarify
> this issue or if anyone has had 
> some practical experience on perfusion and in situ
> techniques, I'd appreciate hearing 
> from you.
> 
> Thanks for your kind advice.
> 
> Karen in Oregon
> 
> 
> 

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