[Histonet] question of the day - embedding

[Hofecker, Jennifer L]

I was starting to feel all alone in the world, Rene'!
First, let me say that animal tissue always seems a littler "drier" to me and I typically do work with human (neuro) tissue these days. I've never had a problem leaving cassettes in molten wax, as long as the processing was adequate and the temperature is correct. I have also worked without submerging in liquid paraffin and there are a few problems I've encountered. We use a lot of Histogel in our lab. If you embed those blocks without submerging them in wax they Histogel will likely pop right out of the paraffin when you attempt to section it. Another similar problem occurred when tissues had "cooled" a little in the holding chamber and were not re-warmed adequately before the block was embedded. The tissue was white and surrounded by a "circle" and when you tried to cut it, the "circle" and the tissue came out too. Of course, those of us who have embedded without submersion (we call that "dry" embedding here) know to be sure the tissue has the appropriate appearance in the molten paraffin (in the mold.)
In my opinion, either way is acceptable but like everything else we do, it requires training and education for us to know potential pitfalls.
Have a great rest of the week!

Jennifer L. Hofecker HT(ASCP)
Vanderbilt University Medical Center
Division of Neuropathology
Nashville, TN 
ph 615.343.0083
fax 615.343.7089
-----Original Message-----
From: Rene J Buesa [mailto:rjbuesa <@t> yahoo.com] 
Sent: Wednesday, February 18, 2009 8:40 AM
To: histonet <@t> lists.utsouthwestern.edu; Tracy Bergeron
Subject: [Histonet] question of the day - embedding

Tracy:
Let me try to dispel some misconceptions:
1- the tissues do not cook if are left in melted paraffin as long as the paraffin is in just its melting point.
2- the tissue are already infiltrated with the paraffin, so there is no additional infiltration to occur
3- the tissues are already dehydrated when they get to the paraffin so they will not "dry out" (they are already dried)
4- the only way there could be some difficulty sectioning later is if the tissues are left a very long time in melted paraffin, like over the weekend.
Otherwise there is no real adverse effect caused by leaving the tissues in melted paraffin during the short time that it takes to embed them, in the same way that if the tissue processor ends the cycle at a given moment and the embedding starts a few hours after that.
I personally consider more problematic leaving the tissues outside the melted paraffin in a warm empty embedding center because there will always be a film of semisolid paraffin surrounding the tissue that will have to melted when the block is casted, and that is what can cause problems.
I know it will very difficult for you to change what it seems you have been doing for years, but I would advise you to fill the holding tank of the embedding center with melted paraffin and place there the tissues until the blocks are done.
René J.

--- On Tue, 2/17/09, Tracy Bergeron <tracy.bergeron <@t> biogenidec.com> wrote:

From: Tracy Bergeron <tracy.bergeron <@t> biogenidec.com>
Subject: [Histonet] question of the day - embedding
To: histonet <@t> lists.utsouthwestern.edu
Date: Tuesday, February 17, 2009, 4:14 PM

Hi all question/dilemma of the day.

        I have been of the view that the longer tissue sat in melted 
paraffin the harder it got, especially animal tissue.  So with that said, 
for the past nearly 10 years I have not used melted paraffin in the 
holding chamber of the embedding center.  I just keep the chamber warm, 
and work that way.  Thus keeping the tissue from continuing to cook and 
harden in the wax.

        Everyone else I am currently working with has never seen the 
method I use, and firmly believe that this causes harm to the tissues if 
they are not in paraffin.

        Thoughts ideas etc.  I am dying to know if I am the only one that 
worries about length of time that animal tissue sits in paraffin.

Thanks.

Sincerely,
Tracy E. Bergeron, B.S., HT, HTL (ASCP)
Associate Scientist III, Pathology
Comparative Pathology Laboratory
Biogen Idec
14 Cambridge Center
Cambridge, MA 02142
Direct:  617-914-1115
Fax:  617-679-3208
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