[Histonet] rescue tissues

From:"Osborn, Sharon"

1.  Wash the tissues as is in cassettes with the paraffin under cool
running water for 5-10 min to remove the buffer.
    Using paper toweling, pat off the excess water. 
2.  Allow to air dry or, place into oven to evaporate the water and to
melt off excess paraffin.
3.  Place cassettes with melted outside paraffin into xylene or xylene
substitute for 30-45 min X3 changes.
4.  Place into 100% alcohol x2 changes for 30 min each.  Continue until
you have the 70% alcohol then you can properly process on the correct
program for the machine.
   Essentially, you are running the tissues backwards down to the 70%
alcohol then processing properly.  I assume the tissues were already
fixed a correct amount of time.  If not, then you need to start in the
10% NBF on the processor.

Sharon Osborn
Palo Alto, CA

Date: Tue, 21 Nov 2006 09:41:10 -0700
From: "Tamara A Howard" 
Subject: [Histonet] oops - samples directly to paraffin
To: histonet@lists.utsouthwestern.edu
Content-Type: text/plain; charset="ISO-8859-1"; format="flowed"

Help! I need the collective wisdom of the HistoNet this 
morning. Some students (who should know better) loaded 
their samples on to the processor yesterday afternoon but 
didn't have it set up correctly; the basket went directly 
in to one of the paraffin beakers and they didn't notice 
the error. Their PI went a few hours later to check up on 
the run and caught it; she put the cassettes directly in a 
beaker of buffer. Now we are wondering about the best way 
to rescue the samples: Take them to xylene-substitute to 
deparaffinize *OR* warm them up to melt the paraffin and 
fish out the pieces? Or some other brilliant solution? The 
tissue pieces are too small to chip out of the solid 
paraffin without risking a lot of damage. These were mouse 
bits destined for IHC - are they totally screwed?

Advice, experience, horror stories all welcome.



Tamara Howard
Cell Biology & Physiology
Albuquerque, NM
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