[Histonet] Grocott Stain - quick method <pkarlisch@psu.edu>

[Paul Bradbury]

Hi Patricia,

A modification that I developed a number of years ago permits the 
Grocott Silver method to be completed in less than 45 minutes. When I 
devised this method, I was teaching Histology in Vancouver and the 
allotted lab periods were only two hours in length, so a number of 
methods had to be "tweaked" to allow them to be completed by even the 
slowest students within that time period.

This method has actually been adopted by several clinical labs as it is 
so much quicker, but without sacrificing the specificity.

1.   Bring sections to water as usual.
2.   Oxidize in 10% chromic acid for 10 minutes (this step is the 
biggest time saver)
       (While this is happening, make up the Grocott silver working 
solution in a very clean glass Coplin jar))
3.   Wash thoroughly in running water
4.   Treat with 1% sodium metabisulphite for 1 minute
       (While this is happening, place the Coplin jar in a 56 degree 
water bath)
5.   Wash well in several changes of distilled water
6.   Place the section(s) in the warm silver solution. Use plastic 
forceps as metal ones will cause the silver to precipitate.
7.   Watch the sections carefully, after a couple of minutes they will 
begin to turn golden-brown.
8.   Place the sections in a Coplin jar of distilled water. Examine the 
control section microscopically to gauge the intensity of the silver 
impregnation. Ideally, fungi (or protozoa, Pneumocystis, etc) will 
appear black on a golden background.
If the fungi appear too pale, rinse the sections again in distilled 
water and place them back in the silver solution for a few more seconds. 
Re-examine the sections, etc.
9.   When the fungi appear correctly impregnated, wash thoroughly in 
distilled water.
10.   Treat with gold chloride (whatever concentration you have on hand)
11.   Wash.
12.   Treat with 1% sodium thiosulphate
13.    Wash
14.    Dehydrate, Clear and mount

Do not be tempted to pre-heat the silver solution to soon. If you do 
this, or if the Coplin jar is not scrupulously clean, the silver 
solution will precipitate to form a silver mirror on the glass.

Paul Bradbury
Kamloops, Canada

Patricia Karlisch wrote:
>  Histonetters,
>   Does anyone have a quick method to do the Grocott stain.  
>   Thank you,
>    Pat Karlisch pkarlisch <@t> psu.edu 
>  
> Pat Karlisch
> Supervisor, Histology, Pathology and Laboratory Medicine
> Penn State Milton S. Hershey Medical Center
> Mail Code H179
> Hershey, PA 17033
> Phone (717) 531-6072
> Fax: (717) 531- 7741
> email: pkarlisch <@t> psu.edu 
>  
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