RE: [Histonet] quantitative iron testing/processor solutions

From:"Connie McManus"

Donna Willis has the right idea... send fresh tissues for any quant anal
that needs to be done.  

My question is.. why do  you color your ETOHs?  I don't do this at all
and have no problem seeing small bxs or scrapings. I'm not criticizing,
just curious. 

Below my comments, Vinnie speaks of the "bigger problem"  in how to
determine contaminants in dye.  We always think in terms of chemical
analysis ... gas chromatography, ICP, etc.  These are very good and
sensitive procedures, but has anyone considered using EM to do this?  I
wish I had paid better attention to the things my husband has told my
about this so I could talk with more intelligence about it.  Elemental
analysis can be done utilizing EM (whether TEM or SEM, I forget). Bill
has done this numerous times in the past and with the more advanced
technology in detectors, I believe the sensitivity and range of elements
is as good as the chemistry methods currently available.

Just another Monday morning thought...

Connie McManus
Utah Veterinary Diagnostics Laboratory
Utah State University
Logan, UT
Phone:  435/797-1891
fax: 435/797-2805

-----Original Message-----
[] On Behalf Of Vinnie
Della Speranza
Sent: Friday, June 18, 2004 12:19 PM
Subject: Re: [Histonet] quantitative iron testing/processor solutions

I've spoken with our Clinical Chemistry director about this issue. She
informed me that the assay for iron is extremely sensitive and difficult
to perform while avoiding contamination of any sort. Indeed she was not
at all surprised that is problem occurred. She further mentioned that
even if we had available a detailed analysis of the dye listing all
impurities, any trace iron, even in the range of 0.0001 % would be
sufficient to affect the results.

the bigger problem is how to determine whether any dye source will
contain iron impurities. I don't think the issue is specific to
methylene blue and probably could result from any dye. 
since there is no iron in the molecular structure of methylene blue (or
eosin for that matter) the contaminant likely arises in the impurities
that accompanying the dyestuff in powdered form. any solution of the dye
would also contain those same impurities. if you dye is labeled to be
80% pure, this essentially means that 20% of the stuff in the bottle is
something else that we usually are not interested in in terms of the
staining we desire.

I don't know if there is any simple method we might employ to assay for
or remove the contaminants accompanying the dye in the bottle. I suspect
not, but I would be grateful to any chemists on the list for their

I suspect that there will be no simple answer to Dana's question however
it is a really good one for those of us who sometimes utilize paraffin
embedded tissues for this iron assay.



Vinnie Della Speranza
Manager for Anatomic Pathology Services
Medical University of South Carolina
165 Ashley Avenue  Suite 309
Charleston, SC 29425
Ph: 843-792-6353
fax: 843-792-8974

>>> "Hoops, Maureen"  06/18/04 10:06AM >>>

Our lab recently had a case sent out for quantitative iron analysis that
yielded abnormal results.  The pathologists requested we submit a sample
the methylene blue dye we had been using to color our processing
Based upon the sample assays, the methylene blue was the cause of the
elevated metal levels. This raises several questions --
Do other laboratories utilize methylene blue coloring in their processor
If so, have you ever experienced a similar situation with heavy metal
Does anyone use Fast Green (Light Green) for a coloring agent in the
alcohols of their processor?  If so, does it interfere with testing?
Does anyone know of an agent we can use to color our alcohols (tissues)
does not hinder quantitative iron and/or heavy metal analysis?
Thanks for your help

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