NBT/BCIP in situ detection problem
pgrimm at ucsd.edu
Fri Oct 20 20:29:47 EST 2000
Is it possible that the mounting medium or technique is different and affecting
the NBT/BCIP deposits? I used glycerol gel to mount the cover slip when I was
using NBT/BCIP for in situ's. Sometimes a batch of glycerol gel would not work
as well as another.
Suggest that you try temporarily mounting one of your slides with water or
saline and a coverslip to see how the deposit looks under the microscope before
The other issue is whether you use polyvinyl alcohol in your NBT development. I
think PVA can give a more "granular" appearance.
Paul Klosen wrote:
> Recently we have run into a nasty problem with our NBT/BCIP in situ
> detections. The in situ part seems to work fine and the alk phos
> detections is sensitive, but when we observe our slides under the
> microscope, the precipitate is completely granular and seems to be
> located above the tissue. We have tried varying several parameter (pH,
> salt and MgCl2 conc., ..) and different lots and sources of both NBT and
> BCIP, but nothing seems to bring us back to our previous results, where
> we had a nice cell-filling precipitate, albeit a bit diffuse as expected
> with NBT/BCIP.
> Has anyone encountered a similar problem, and how did you get rid of it
> Paul Klosen, PhD
> CNRS UMR 7518 Neurobiologie des Fonctions Rythmiques et Saisonnieres
> Universite Louis Pasteur 12, rue de l'Universite F-67000 Strasbourg,
> Tel. 03.88.35.85.04 Fax. 03.88.24.04.61 klosen at neurochem.u-strasbg.fr
Paul C. Grimm
University of California at San Diego
Email pgrimm at ucsd.edu
Mail Stop 0831
9500 Gilman Drive,
La Jolla California
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