Dear Ian,
I'm not convinced one way or the other about the impact of different
clearing reagents.  I have used several different clearing agents for
various projects over the years but the only times I have had real
trouble was when our satellite lab used "Histolene" (a limonene based
product) and when "Mr Nobody" tampers with the tissue processors.  On
one ocassion Mr Nobody forgot to change the processors' reagents for a
fortnight which led to the same appalling IHC which resulted on tissue
from the satellite lab.  The latter event has led me to wonder if the
problem in the satellite lab was that the solutions were not changed
frequently enough rather than histolene being the lousy option for
clearing which I had first thought.  I suspect that people use the
processing schedules which were designed for xylene and undenatured
alcohol but have neglected to make the appropriate alterations in
processing times and frequency of reagent changes to reflect the use of
different clearing reagents and quality of alcohol.  

In short, I believe that the processing schedule plays a far more
critical part in IHC than we acknowledge.  When Mr Nobody has changed
our processing schedule or the grade or reagents I observe the results
in the IHC long before any changes are noticed in the morphology, H&E
staining or routine special stains.  I wish I had more time to explore
the topic further.

Penny Marr


> -----Original Message-----
> From:	Ian Montgomery [SMTP:ian.montgomery@bio.gla.ac.uk]
> Sent:	Thursday, 8 July 1999 3:26
> To:	HistoNet@Pathology.swmed.edu
> Subject:	Clearing agents.
> 
> 	Clearing agents for tissue processing in immunohistochemistry.
> 	Xylene is ok but what about the others, chloroform, benzene,
> toluene, amyl acetate etc. I'm a wee bit suspicious about loss of
> antigenicity with chloroform but nothing I'd like to stake the
> contents of
> my wifes wallet on.
> 	Haven't found a lot in the literature regarding use, any
> thoughts.
> Ian.
> 
> 
> 
> Dr. Ian Montgomery,
> West Medical Building,
> University of Glasgow,
> Glasgow,
> G12 8QQ,
> Scotland.
> Tel: 0141 339 8855 Extn. 6602.
> Fax: 0141 330 4100.
> e-mail: ian.montgomery@bio.gla.ac.uk
> 
> 
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