Re: NBT/BCIP
From: | "t.hacker@har.mrc.ac.uk" <T.Hacker@har.mrc.ac.uk> |
Date sent: Thu, 12 Jul 2001 16:43:20 +0100
From: Maura Farquharson <M.Farquharson@udcf.gla.ac.uk>
Subject: NBT/BCIP
To: histonet@pathology.swmed.edu
> Hi folks,
> Several enzyme histochemical methods e.g. for dehydrogenases use NBT as the
> chromogen.At the end of the development of the formazan reaction
> product,several methods recommend fixing for 10-15 mins in 10% formaldehyde
> then dehydating through alcohols,clearing in xylene and mounting.I find
> this loses some of the reacion product and the results are weaker than if I
> aqueous mount.
Allan, I have done a lot of work in the past demonstrating
dehydrogenases (DT diaphorase actually) using NBT as the
chromagen.After the post fix as you mention (and counterstaining),
I would wash and drain the slides then give a quick (5-10 second)
rinse in acetone to remove any soluble (pink) formazan followed by
a rapid (final) dehydration in alcohol, clear and mount.The blue to
blue/black staining was always clear and strong.
Terry.
Terry Hacker,
Medical Research Council,
Harwell,
Didcot
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