Re: Mouse brain cryostat sections

<< Previous Message | Next Message >>
From:"kunihiro uryu" <> (by way of histonet)
Content-Type:text/plain; charset="us-ascii"

I would think that this problem is mainly due to the slow freezing, although
I am bit curious that if 5 hours fixation is long enough for fixing brain.

You seem to take to long time to freeze the brain in not too cold
temperature.  I always freeze brain in dry ice powder, which you prepare by
smashing dry ice.  (Some people use Isopantain precooled by dry ice.  This
is nice method, too.  However dry ice power works fine and easier.)  You
embedded the brain in the powder for 20 min at least.  Then if you wish you
can embed the brain in OCT compound.  Or if you wish to embed brains in OCT,
you can prepare samll container with alminum foil and put brain and OCT.
Freeze whole thing in the Isopantain pre-cooled with dry ice.

However, I do not embed brains in the OCT, becuase OCT can be used only to
glue the brain to a specimen holder in the cryostat when you preapre
cryosectioning.  In this way you can see the brain clearly and change
angles, when needed.  Also based on my experience, the optimal temperature
for cutting fix brains is lower than fresh frozen brains.  This temperature
makes OCT hard.  I found out brain without OCT surrounded is easier to cut
than brain embedded in OCT.

I prefer not to use cryoprotectant for freezing brain.  Sugar makes cutting
brain sections in the cryostat more difficult.  If you use sugar, You tend
to have more curly sections than brains without sugar treatment.  DMSO may
be good for preservation, but it also effect on membrane structure.  You
need to aware of that.  As long as you freeze brain in a rapid manner in low
temperture as cold as dry ice is (about -80C degree) you do not need any
cryoprotectant.  I like this simple easy preparation.

Hiro (kunihiro Uryu)
CNDR, U of Penn
Get Your Private, Free Email at

<< Previous Message | Next Message >>