[Histonet] Mammary Tissue Processing & Sectioning Troubles

[Joseph Saby]

Andrea-
I believe your tissue was not fully fixed.  Many labs think they know more than they do, so itis always a good idea to pin down exactly what they did to fix the tissue.  I am guessing they left thick 
blobs of tissue in an inadequate volume of formalin and expected that to do the trick.
I would deparaffinize and rehydrate the tissues, then place them back in an appropriate volume of formalin 
and fix them until you know they will be fixed.  If the sections are thick, cut them thinner for this fixation.  After 
reprossessing, these tissues should section much better.
Good luck!
Joe Saby, retired

 
      From: Andrea Calhoun via Histonet <histonet at lists.utsouthwestern.edu>
 To: "histonet at lists.utsouthwestern.edu" <histonet at lists.utsouthwestern.edu> 
 Sent: Wednesday, September 27, 2017 3:49 PM
 Subject: [Histonet] Mammary Tissue Processing & Sectioning Troubles
   
Hi All!

I'm working with primate mammary tissue that was given to us from a  pathology group off campus. They say the tissue was fixed in NBF for at least two days, and sent to us in 70% EtOH, where it has sat for a couple months.  We are treating these samples as we would with human tissue.  After grossing, processing (14hr schedule), and sectioning (4-5um, high-profile blade)), I find the tissue is difficult/ near impossible to section.  Even after leaving the blocks on ice for 30+min, the tissue continuously mushes against the blade.  If I do get it to cut, multiple knife marks develop quickly and I find I am going through blades like crazy.  I am staying superficial incase the fix didn't penetrate  deep into the tissue.    So far I've only processed a few samples from a larger group to resolve these issues before processing the remainder.

Does anyone have any tips for processing or sectioning?  Besides grossing the tissue into smaller pieces, would it help to re-fix them overnight or an additional day? How would you tell if the issue is from fixation or insufficient clearing/infiltration of paraffin?

Any information would be appreciated!

Andrea Calhoun B.S., CEMT
Research Assistant 2, Schedin Lab
Dept. of Cell, Developmental, & Cancer Biology
Oregon Health & Sciences University
calhouan at ohsu.edu , RJH-5350

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