Re: Eosin bleeding out

Hi Michael,
If all your reagents are Shandon, I would suggest you call their technical
service department for their help.

Rande Kline
EM Science

Michael Becker  on 08/08/2002 07:50:41 PM

To:    HistoNet Server 
Subject:    Re: Eosin bleeding out


Long time since I posted.  Please if anyone out there in HistoLand can tell
me what causes and how can I get rid of Eosin counterstain bleeding out of
frozen tissue sections after they are coverslipped and are drying.  Once in
awhile this is a problem.  I have not changed any solutions or technique or
mounting media.  I use the following protocol:

1.  Rapid Fixx-10 to 15 secs
2.  Tap water rinse x2
3. Gill 3 Hemotoxylin 35-60 seconds depending on age of stain
4. Tap water rinse at the sink until water is clear
5.  Lithium Carbonate Bluing-5 to 7 dips
6. 200 proof Ethanol-10 dips
7. 1% Alcoholic Eosin-2 quick dips
8. 200 proof Ethanol-4 containers-10 dips each
9. 200 proof Ethanol-1 container-1 minute
10.Xylene Substitute-Histosolve for 30 secs to a minute or longer.

Coverslip with Histosolve and a Xylene Substitute mounting media-Histomount

I get all my reagents except Ethanol from Shandon and have been very happy
with staining quality.

Any ideas!  Thanks in advance to all who may respond.

Hope summer is treating you all well.  Have fun.  Be safe.

Sue Becker HTL (ASCP)
c/o Dr. Michael J. Mulvaney
MOHS Micrographic Surgery
Albany, NY 12203

"No one knows noses like MOHS!"

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