[Histonet] problems with Gomori's Trichrome on Frozen Muscles

[Due, Brice]

Hello Jennifer, I have noticed two things which cause red staining. 

a) incomplete rinse after hematoxylin. If the section is wrinkly or puckered and
traps the hematoxylin soln, those areas tend to stain red with the gomori.
Longer rinse after hematoxylin reduces this. This is probably not your trouble,
but you could try extending your rinse before the trichrome soln.

b) "freeze-dried-looking" regions of frozen sections. Sometimes two consecutive
sections will look different after they've dried on the slides. Thid can be
immediately obvious in the cryostat, but often shows up only after storage at
-80. The "good" section will look translucent, while the "bad" one will have
areas that look opaque, white and frosty -- as if it had freeze dried, or as if
it hadn't really stuck to the slide fully, or something. Those white areas tend
to show funky staining -- perhaps for the same reason as above: they may trap
solns more than usual. 

Are you picking up sections exactly the same way or handling them differently? I
know you said you are staining same day. I have cut and stained 30min later
without problems. 

c) another remote possibility is that the sections are getting exposed to some
solvent or formalin vapors at some point. Fixation and/or lipid extraction can
ruin gomori staining, although I have usually seen an absence of red rather than
excess. 

Curious to know what the problem is,
-brice
Neuropathology LAb
Brigham & Women's Hospital
Boston


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-----Original Message-----
From: histonet-bounces <@t> lists.utsouthwestern.edu
[mailto:histonet-bounces <@t> lists.utsouthwestern.edu]On Behalf Of Hofecker,
Jennifer L
Sent: Friday, August 12, 2005 3:04 PM
To: histonet
Subject: [Histonet] problems with Gomori's Trichrome on Frozen Muscles


Happy Friday everyone,
We are experiencing a strange phenomenon with our Gomori's.  All of a sudden the
staining is predominately red (instead of green).  Normal muscle control is
still green but the patient tissues are staining red.   We have tried several
different changes: new trichrome solution, recutting frozens, etc.  The only
thing that is reproducible is that sections which have been previously cut and
stored at -70 seem to stain fine.  If we cut a frozen then begin staining it
without time in the freezer, voila, the same muscle sections are now red instead
of green.  What could be causing this?  I even pulled out the same piece of
control tissue and cut fresh slides on it.  They were red also.  There was one
fresh slide that stained green in the whole "experiment" otherwise, anything
that doesn't spend time in -70 is turning red.  We are using the same procedure
that we have been using for quite a while, and the precut slides do stain, so I
don't think it's a procedural error.   The only change is that we have been
trying to stain the trichromes when the specimens are cut instead of storing in
the freezer to batch staining later.  Has anybody experienced this?  Any ideas
would be appreciated.
 
Thanks in advance,
Jennifer
 
 
 
 
Jennifer Hofecker, HT (ASCP)
Vanderbilt University Medical Center
Division of Neuropathology
(615) 343-0083
(615) 343-7089 fax
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