[Histonet] Thionin staining method

[Geoff McAuliffe]

Hi Caroline:

    John Kiernan is might be the best person to ask about this but for 
now.........

Lack of de-fatting should not be the cause of your problems, at least 
not in my experience.
There was considerable de-fatting in the graded ethanols, there is 
experimental evidence for this.
Did you put some acetic acid in your thionin? If the stain is not acidic 
it will wash right out.
If all else fails, try an oxidation with 0.25% potassium permanganate 
for a few minutes followed by bleaching the
ugly brown color with 2% oxalic acid, then wash and restain with thionin.

Geoff


Runyan, Caroline (NIH/NIMH) wrote:

>I am trying to re-stain brain sections with thionin (they had been stained
>four years ago initially but were originally very light and have now faded
>beyond recognition).  I popped off the coverslips, and then left the
>sections to sit in xylenes for about five days, at which point the dpx used
>for coverslipping appeared to have been removed from the tissue.  Then, I
>re-hydrated and incubated in thionin for fifteen minutes-the thionin did not
>stick to the tissue at all.  As soon as I drained the slides in water, all
>color was removed.  I then went back and looked at the protocol that was
>apparently used to stain the tissue originally, and it looks as though the
>tissue was not de-fatted prior to thionin.  The slides went from 70% EtOH
>(no water or 50%) through to 100%, into thionin, and then straight to 95%,
>100%, xylene.  
> 
>Is tissue's current lack of thionin reactivity due to the fact that the
>original stain had been done so long ago, or because of the procedure
>originally used?  Thanks.
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Geoff McAuliffe, Ph.D.
Neuroscience and Cell Biology
Robert Wood Johnson Medical School
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