[Histonet] RE: embedding multiple brains

From:"Pixley, Sarah \(pixleysk\)"

Dear Zsuzsa:

We cryosection brains also and this way might work for your needs. We
cut the unfrozen brains (after sucrose) down into sections that include
the area we are interested. We make the most posterior slice as close as
possible to a true vertical plane, so that when we set the brains down
on that, they will be flat and we will be cutting as close as possible
to true coronal sections. Then we lay the brain down flat in a plastic
weigh boat (Fisher Sci. has medium and large ones that might hold
several brains at once). Then we fill the weigh boat with OCT compound
and mark the outside edges of the weigh boat with the specimen number,
mark out the dimensions of the brain and put D and V for dorsal/ventral.
Then we sit that weigh boat flat onto a flat piece of dry ice. It takes
about 15 mins for the whole thing to freeze. As it freezes, it moves
down into the dry ice block. When it is done, it is transferred to the
cryostat and we let it equilibrate for at least 15 mins. Sometimes, we
have to wait, so we put the frozen block in the weigh boat into a Ziploc
bag to keep humidity constant and put in the freezer. After freezing, we
also mark the OCT with important numbers and dimensions. When ready to
cut, we pop it out of the weigh boat and use a razor blade to cut down
the outside OCT to a manageable size rectangular block. We use a small
amount of Oct to attach the block to the cryostat chuck and start
sectioning. If the top remained flat during freezing, you can use it to
get the specimen cutting angle correct and you should be set to go once
you get to the tissue. Sometimes we have to fine tune once getting to
the tissue.

I could envision that you could put several mouse brains in one of the
medium or large size weigh boats, or even make home-made boats out of
aluminum foil. However, you will be limited by the size of the cryostat
chuck and by how well you are able to cut such a large block.

Good luck, hope this helps.
Sarah Pixley

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