[Histonet] RE: Fading of Hematoxylin

Gayle Callis gcallis <@t> montana.edu
Mon Jul 12 10:01:46 CDT 2004

Barry gave good advice, look for a permanent mounting media that contains
antioxidative or a good reducing agent.  Many mounting medias are
available.  Also when doing your hematoxylin staining, be sure you
incorporate good water rinses between steps to have the correct pH for
staining. 1 minute of running tap water rinse between each step should do
the job nicely.   A 70% alcohol rinse before eosin (or use the same
concentration of alcohol of the eosin solution) removes any cations that
can interfere with the eosin staining.  

Gayle Callis
Research Histopathology Supervisor
Veterinary Molecular Biology 
Montana State University - Bozeman
PO Box 173610
Bozeman MT 59717-3610
406 994-6367 (lab with voice mail)
406 994-4303 (FAX)

At 08:26 AM 7/11/2004 -0500, you wrote:
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>Most permanent mounting media tend to cause some fading of hematoxylin
probably due to its continued oxidation. This is however, generally not as
rapid as the time period that you indicate. 
>We have seen some problems in the past when batches of clearing agent
(xylene, which is really a mixture of xylenes) had some contaminants and
this problem was eliminated using better quality xylene.
>One medium that retained hematoxylin was Euparal vert from George Gurr Co.
This contained a copper salt that in effect produced a copper hematoxylin,
much more resistant to fading. I wonder whether converting the hematoxylin
to a copper hematoxylin or an iron hematoxylin might solve this problem  or
perhaps incorporating a reducing agent in the mountant. Eosin fading is
another problem. If you have old faded slides can look at them using
fluorescence microscopy for the eosin.
>Would be interesting to have a Histonet discussion of this.
>-----Original Message----- 
>From: histonet-bounces <@t> lists.utsouthwestern.edu on behalf of Sulekha Ravi 
>Sent: Sun 7/11/2004 6:39 AM 
>To: histonet <@t> lists.utsouthwestern.edu 
>Subject: [Histonet] Fading of Hematoxylin
>	Hi
>	Please help me in overcoming this problem. Hematoxylin in my H&E stained
slides are fading in 1 month. I am using 'Gurr' make Hematoxylin powder to
make Harrys' Hematoxylin. Fresh stain is prepared every 3 months. Kindly
advise what to do.
>	Sulekha
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