Re: Mystery:Disappearing melanin!

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From:"R.Wadley" <> (by way of histonet)
To:histonet <>
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	Dear Greg,

	Sounds like the granules are not fixed, ie not fixed into place in the
cells & not cohesive within themselves.  Have you tried drying the slides
after staining?  My favorite xylene replacement fluid was not miscible with
my xylene mountant I got excellent results be drying the slides (on a hot
plate) then coverslipping.
	In your case maybe using glycerine as the mountant (whether or not
you dry
the slide first) will fix your problem.  When I was Curator of the Rodda
Pathology Museum, University of Tasmania, the manual recommended gelatin
solutions, glycerine and liquid paraffin as a mounting mediums for
melanomas because it prevented the melanin (& other colouring agents) from
leaking out of the specimen & discolouring the mounting fluid.  A little
clear nail polish (or even your usual mounting medium) around the edge of
the coverslip prevents the mountant from drying out.

	Hope this helps.


	Rob W.

At 08:24 AM 2/19/99 -0400, you wrote:
>One of our graduate students is working on lobster hemolymph (blood).
>She is trying to use cytochemistry to demonstrate phenyloxidase
>activity in lobster hemocytes.
>She is obtaining nice brown-black staining of intracellular
>granules, BUT, the staining disappears when the mounting media is
>placed on the cells prior to coverslipping.
>The staining survives hematoxylin counterstaining and subsequent tap
>water wash, survives the alcohol dehydration, and xylene clearing,
>but disappears when mounting media added. She tried both solvent
>based and aqueous mounting medias. ANY IDEAS???
R. Wadley, B.App.Sc, M.L.S
Laboratory Manager
Cellular Analysis Facility
School of Microbiology & Immunology
UNSW, New South Wales, Australia, 2052
Ph (BH) 	+61 (2) 9385 3517
Ph (AH)	+61 (2) 9555 1239
Fax 	+61 (2) 9385 1591
	(Under development)

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