RE: Backing up a GMS for fungi

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From:"Reeder, Carole" <> (by way of histonet)
Date:Fri, 2 Jul 1999 01:15:09 -0400
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YES back it all the way up and start over! We do Chromic Acid for 10 minutes
then proceed with the rest of the stain. What temp are you using for the
silver sol step? We do ours in the microwave or water bath...we watch it and
gage it by its color (black). Also, you can use Periodic Acid in place of
Chromic Acid. Don't forget your borax. Good Luck give us a progress report.

-----Original Message-----
From: McCollough, Carol []
Sent: Tuesday, June 29, 1999 1:21 PM
To: 'histonet'
Subject: Backing up a GMS for fungi

Greetings Histonetters:

We seem to have an intermittent but chronic problem with our Gomori's
methenamine silver stain for fungi.  The organisms do not stain, although we
are using a known positive control.  We are using the method from Carson's
book, but using 5 percent chromic acid instead of 4 percent for the oxidizer
for 1 hour at room temperature.  I believe that the problem is due to
carelessness in formulating the stock methenamine silver, however in at
least one instance, the stain worked fine with one batch of stock and didn't
work about 2 weeks later with the same batch which had been stored at 4
degrees C .  The latest instance could have been an error in compounding the
working solution (left out the borax - maybe).

To make a long story shorter....... we cannot afford to do very many more
recuts on these tissues without losing them entirely.  Can this stain be
backed up?  To what step?  How?  My initial thoughts are to go back through
xylene, 100 percent alcohol and 95 percent alcohol (which will remove the
light green counterstain), distilled water, and reimpregnate.

Thanks for your kind answers to a simple question.

Regards -
Carol B. McCollough, HT(ASCP)
Diagnostics & Histology Laboratory Manager
Maryland Department of Natural Resources
Fisheries Service
Cooperative Oxford Laboratory
904 S. Morris Street
Oxford, MD 21654

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