Re: [Histonet] quantitative iron testing/processor solutions

From:"Vinnie Della Speranza"

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 comments

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 of
the methylene blue dye we had been using to color our processing alcohols.
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) that
does not hinder quantitative iron and/or heavy metal analysis?
Thanks for your help

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