Re: [Histonet] Rodent Processing Schedule?
No, whole rat brains are larger and would require a longer processing
We have processed whole mouse brains using automated VIP (Sakura Finetek)
with NO TEMPERATURE added to dehydration stations nor clearing as this will
add to tissue drying.
Make sure your mouse brains are totally fixed, perfusion is ideal, followed
by immersion overnight.
We process 70, 80, 95 X 3, 100 X 3, Xylene X 1, Clearite 3 X 1 (or you can
use 2 changes of Clearite 3), and 4 changes of paraffin 30 min X 2 changes,
1 hour x 2 changes. All stations are at 1 hour with exception of
paraffins, just to shorten total time in that for 3 hours. Use vacuum and
pressure for all stations, and do not exceed 60C with paraffin
infiltration, avoiding excess heat helps. What you want to avoid is
removing the bound water on proteins (this leads to hard, dry
tissues) only free water in tissues spaces. Some labs like to have first
dehydration stations at lower concentrations of alcohol, 50, 70, 80, 95 X
2, 100 X 2 or 3, clearing, etc. Xylene tends to harden tissue, but
Clearite 3 (Richard Allan) and Propar (ANATECH) are more forgiving, single
aliphatic hydrocarbon xylene substitutes. These are only guidelines as
your conditions may be slightly different.
Recommended is do some processing runs on normal brain just to see what is
optimal for your fixation and processor conditions. We recently had to
customize processing schedules after some test runs for both
Periodiate/lysine/paraformaldehyde perfused hamster brain (coronal
sections) and tongue, with a separate schedule for mouse coronal brain
At 07:48 AM 7/30/2004, you wrote:
>Does anyone have a protocol for processing whole, formalin fixed mouse
>brains? The mouse brains need to be processed whole.
>I am concerned that over-processing, using a schedule meant for human
>brain tissue, will dry the tissue out excessively and make the serial
>sectioning a nightmare.
>Also, would the same schedule apply to whole rat brains, or should the
>times be increased a bit?
>Harvard Brain Tissue Resource Center
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