RE: Rat Tissue Processing
I use a Leica TP1050. The processing schedule is:
formalin: 15 min. (station 1)
70% ETOH to 3 Abs ETOH changes: 40 min each (stations 2-6)
abs ETOH/histoclear: 40 min (station 7)
histoclear, 3 changes, 40 min each (stations 8-10)
3 paraffin changes, 1 hour each.
For small tissues (like mouse livers) I have a short schedule that has 10
minutes in each station and 30 minutes for each paraffin bath. I don't know
or how big or thick the brains you're working with are, but if they're thin
and small (i.e. < 3mm thick X < 4mm/side), I would use a short processing
time. Use fresh reagents and be sure they are WELL FIXED. We use 10% BNF for
all our stuff.
To keep brain from heat damage, I have set my processor paraffin baths at 58C
rather than 60. This seems to help. the person who set up the processor had
it set at 60 C and there were a lot of dry, brittle livers, brains, etc.
Since changing to this temp, there has been less of this.
Last of all, what temperature is your waterbath? If it's too warm, the brain
sections will blow apart on the water bath. I set my waterbath to 36-38C, I
add approx. 100 mL 95% ETOH to the water (1800 mL of DI water). I pick up the
sections, wrinkles included *g*, place them falt on a slide warmer (set to
40C) for a few seconds (usually about 30 sec) until the wrinkles are gone.
Voila, they're ready for staining.
Hope this helps.
Veterinary Diagnostics Laboratory
Utah State University
>===== Original Message From C H <email@example.com> =====
>I wonder if any of you have a protocol that you wouldn't mind sharing for
>auto-processing rat CNS tissue for paraffin embedding. I am a novice at
>working with paraffin,using a Leica TP1050 and so far I have not found the
>right combination of steps to prevent the tissue from overly expanding and
>separating from the paraffin in the water bath.
>Any assistance would be greatly appreciated.
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