[Histonet] to cut or not to cut mouse brains

[Helen Fedor]

When the blocks are not croyprotected you can cut them at warmer temperatures. But when they are cryoprotected with the 30% sucrose, they are soft at -18, that is why you need the colder temperature. they will compress at the warmer temperatures and give you a lot of wrinkles.

Helen

-----Original Message-----
From: histonet-bounces <@t> lists.utsouthwestern.edu [mailto:histonet-bounces <@t> lists.utsouthwestern.edu] On Behalf Of Grantham, Andrea L - (algranth)
Sent: Thursday, November 17, 2011 2:57 PM
To: McLaughlin, Terry
Cc: histonet <@t> lists.utsouthwestern.edu
Subject: Re: [Histonet] to cut or not to cut mouse brains

I'm interested in this answer because this has happened to me - but not always.
I usually cut brains at warmer temps. like -17 or -18 as opposed to -25 and they are usually handled in the same manner before being brought to the lab frozen in OCT.

Andi



On Nov 17, 2011, at 12:24 PM, McLaughlin, Terry wrote:

> Hello All,
> I am having trouble cutting frozen mouse brains and was wondering if 
> someone can offer some help. The mouse was perfused in 4% PFA , the 
> brain removed and kept in PFA for 22 hrs. We placed it in 30% sucrose 
> for 20 hrs, until the brain sank in this solution. It was frozen back 
> by using a culture plate floating in liquid nitrogen with OCT .
> The problem I am having is many folds, and air bubbles under the tissue.
> What I have done so far:
> Cut at temp of -25, then tried - 22 and -17.
> Cut at 10um, then tried 16-25um.
> Tried making the knife colder by adding dry ice to it for a few seconds.
> Added dry ice to sample for a couple seconds.
> The sample cuts fine, it appears to happen when picking up on a slide. 
> I tried picking up by starting at one side or end and letting tissue 
> spread onto slide. Did not work.
> Also tried gently laying slide on top of sample and removing 
> gently-still folds and air bubbles. Any help would be greatly 
> appreciated as we have 6 more to do.
> 
> Signed,
> Help!
> Terry Kokas
> 
> 
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