NBT/BCIP in situ detection problem

Peter Ashby p.r.ashby at dundee.MAPS.ac.uk
Sat Oct 21 06:14:40 EST 2000

In article <39F073EE.DC083AFE at neurochem.u-strasbg.fr>, Paul Klosen 
<klosen at neurochem.u-strasbg.fr> wrote:

> The problem is not with the in situ. We perform ISH on sections. When we
> follow the reaction development under a scope, it develops normally.
> Just, when we later on look at the slides on the microscope, we have a
> granular deposit instead of the more diffuse typical deposit. This is
> very annoying, because it seems to be located above the section, and
> does not outline the cells. We have the problem with several probes and
> only with alk phos detection. With TSA and perox detection the result is
> the same as usual, but we need the alk phos for our double in situs.

Your description confirms that it is most likely a problem with your 
mounting medium as outlined by someone else. I have seen this effect 
with a number of different enzyme stain products including LacZ from 
transgenic embryos. Have a look at your mounting medium as that is the 
most likely culprit unless you have suddenly increased the magnification 
you are using to view your sections.


Peter Ashby
Wellcome Trust Biocentre
University of Dundee
Dundee, Scotland
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