[Histonet] better perfusion and inclusion---thanks to y'all

[Charles Scouten]

It was my error in specifying 5% sucrose, but if you had done the Perfusion One procedure with 10% sucrose prewash (isotonic, or slightly hypertonic), and 300 mm Hg, you would have seen no shrinkage, and negligible ventricular distortion. I have done it.

Please do not hesitate to try again.  You have used enough pressure to fully wash out the red blood cells, so you have seen the clean tissue and thorough perfusion.  You can also get the unshrunk and undistorted effects.  The results you did get are fully predictable given the hypotonic prewash.  


Cordially,
Charles W.  Scouten, Ph.D. 
myNeuroLab.com 
5918 Evergreen Blvd. 
St. Louis, MO 63134 
Ph: 314 522 0300  
FAX  314 522 0377 
cwscouten <@t> myneurolab.com 
http://www.myneurolab.com 


-----Original Message-----
From: histonet-bounces <@t> lists.utsouthwestern.edu [mailto:histonet-bounces <@t> lists.utsouthwestern.edu] On Behalf Of Nancy.Walker <@t> sanofi-synthelabo.com
Sent: Thursday, June 10, 2004 10:52 AM
To: histonet <@t> lists.utsouthwestern.edu
Subject: RE: [Histonet] better perfusion and inclusion---thanks to y'all





More precisions:
The catheter diameter varies with animal size. For a 35 g mouse a n° 4 cathater works well (inside diameter = 0.76 mm outside = 1.22 mm).

We used 5% sucrose (saccharose was a little french slipping in) in water , as you had suggested. Unfortunately I didn't check out if this was in fact isotonic,  so that is a mistake! We perfused 1 minute at 300mm Hg. The histology looked bad, that is,  the ventricles were dilated.

Another source advised us to use Mannitol at 25% for 20 " at 110 mm Hg.
This too created ventricle dilation.

For each trial 3 mice brains were processed, coronally sliced at the hippocampal level and stained with H/E.

yours truely,
Nancy




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