[Histonet] Processing Problem
Rene J Buesa
rjbuesa <@t> yahoo.com
Tue Sep 17 10:47:21 CDT 2013
A sample 2mmx2mmx1mm is quite small and there should not be any problems with either fixation of processing but there is where the problem my reside.
1- fixing in 10% formalin: is it neutral buffered? For such a small piece 24 hours will be enough but to be absolutely sure try leaving the pieces for 48 hours.
2- sectioning should not be a problem but I advise that you check the times in the reagents. Such small pieces should be less time in the alcohols. Since it is pig skin try using more time in the "ante-medium". What are you using, xylene? And give extra time in the infiltration steps (melted paraffin wax).
With these precautions you should have no problems sectioning the samples.
From: Sandra Cheasty <cheastys <@t> svm.vetmed.wisc.edu>
To: "histonet <@t> lists.utsouthwestern.edu" <histonet <@t> lists.utsouthwestern.edu>
Cc: Sandra Cheasty <cheastys <@t> svm.vetmed.wisc.edu>
Sent: Tuesday, September 17, 2013 11:26 AM
Subject: [Histonet] Processing Problem
I need help with two issues. I have a researcher who is sending us porcine skin, about 2mm x 2mm x 1mm tall.
Issue 1: There is a lesion in the center, and although he wants the skin sectioned through the lesion eventually, he says if he bisects the chunk of skin before fixation, the lesion becomes distorted. So, he is fixing them in the 2x2x1 chunks, and the 10% formalin (and subsequent processing reagents) are not penetrating. Does anyone know of either a more penetrating fixative or a less distorting one so we can bisect the skin before fixation?
Issue 2: Even on smaller sections that fixed and processed well, we are having issues with the porcine skin sections staying on the slides. We use Superfrost Plus slides, drain them, air dry them overnight, and then they go on the "Extra Oven" program on the stainer. (25 minutes in the oven.) Any suggestions on other slides, drying techniques to try?
My background is that of a certified Histologist for 30 years, with experience in many labs in various parts of the country. This research project has me stymied!
Thanks for your help!
School of Veterinary Medicine
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