Re: [Histonet] Grocott's Methenamine Silver Stain

From:Kathleen Roberts

Mmmm, good point.  I made up the borax fresh, but the source bottle I 
used is quite old, I think.  Another addition to the "dump" list for haz 
waste pickup!


Amy Porter wrote:

> I would be suspicious about the borax.  We do decalcified mouse 
> turbinates with this method for fungal sinusitis.  Any time I have no 
> or weak reaction i make up fresh borax.  Since you are making 
> everything up fresh, ?????.  If your control hasn't been decaled then 
> it seems like that should be working if all your solutions are fresh. 
> I use a water-bath and prewarm my working silver before i add my 
> slides.  Hope you figure out what is wrong, very frustrating.
> Kathleen Roberts  wrote:
>     Hey, all-
>     I've been trying to stain some FFPE slides of decalcified rat sinuses
>     for fungi using the AFIP protocol for Grocott's stain, and despite
>     using
>     fresh reagents, the positive control and unknowns do not stain at
>     all,
>     except for some random deposition of silver that does not even
>     begin to
>     look like fungi. The positive control is loaded with Candida, and can
>     be seen with H&E easily.
>     So far, we don't see any fungi in the rat sinuses and lungs (they
>     were
>     sneezing, and the bedding came back positive for Aspergillus) with
>     H&E
>     staining, which is why we decided to try the Grocott's-one way or
>     another we should be able to visualize it in the tissues.
>     Any ideas why the Candida-loaded positive control would not stain
>     with
>     Grocott's? Right now I am thinking that perhaps they didn't sit long
>     enough in the 4% chromi c acid, even though the protocol says to
>     soak the
>     slides for an hour to oxidize them. Either that or a longer soak
>     in the
>     methenamine-silver solution at 60 degrees C (also for one hour).
>     Thanks in advance for your help-
>     Kathleen Roberts
>     Principal Lab Technician
>     Neurotoxicology Labs
>     Dept of Pharmacology & Toxicology
>     Ernest Mario School of Pharmacy
>     Rutgers University
>     41 B Gordon Rd
>     Piscataway, NJ 08854
>     _______________________________________________
>     Histonet mailing list
> Amy S.Porter, HT(ASCP)
> Michigan State University
> Department of Physiology
> Division of Human Pathology
> College of Human Medicine
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