Fontana Questions

From:Miriam Schroeder <>

Hi all.  I left a lengthy post awhile back and didn't
get any replies.  I'm splitting the two topics into
two posts and hope to get some replies this time:

Does anybody have any special hints about the
Fontana Stain (whether these cells do turn out to be
Fontana-positive, or not).  My protocol has a Gold
Chloride step, but the one in the text "Theory &
Practic of Histological Techniques" edited by Bancroft
& Stevens DOESN'T have a gold chloride step.  Can
someone tell me what the Gold Chloride is for / what
it is doing.  Also, none of my texts seem to go into
the chemical mechanisms behind this stain, so I'm also
wondering what the sodium thiosulfate step does. 
(Since I don't know anything about the chemistry
behind this stain, I really have no clue which steps I
might want to "play" with the times on to try & get
the stain to work.  Maybe some of the steps are
mordanting steps or others are differentiations?) 
Finally, my protocol said to make the ammoniacal
silver sol'n in the following manner:  To the silver
nitrate sol'n add ammonium hydroxide drop by drop
until the solution clears and no precipitate remains. 
The add the reserved Silver Nitrate Sol'n, drop by
drop, until the solution turns slightly cloudy. 
"Slightly cloudy" sounds like a pretty subjective
concept to me.  Does anyone have any hints on what is
the right "amount" of "cloudiness"?  (Back to the
chemistry questions):  If this sol'n was too cloudy or
not cloudy enough, could that have affected the stain?

By the way, my protocol was:
Ammoniacal Silver Solution 1 hr at 60C
Wash - Distilled water
0.1% Gold Chloride 1 minute
Wash - Distilled water
5% Sodium Thiosulfate 2 minutes
Wash - distilled water
Nuclear Fast Red 5 minutes
Dehydrate & Clear

ALSO - has anyone ever tried using hematoxylin as a
counterstain for the Fontana?  We don't like Nuclear
Fast Red very much.

OK, thanks for your help!

Sincerely, Miriam Schroeder
Research Associate, Berlex Biosciences

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