[Histonet] Water stable counterstain?

I.B. beldorth.msu+hist <@t> gmail.com
Fri Sep 29 18:22:50 CDT 2006

Hi All,

I just sent a long message about ISH, but I have another question perhaps
someone can answer.  I am using Nuclear Fast Red (Vector Labs) to counter
stain tissue stained with NBT/BCIP (also Vector Labs), and, in order to
reduce the number of steps in the protocol, I would like to use an aqueous
based mounting media (at least with the aqueous media I have tried).
However, Nuclear Fast Red washes out of the tissue when mounted aqueously,
making contrast under the microscope terrible.  Can anyone

1) recommend a counterstain, preferably something red, that will be stable
in aqueous mounting media, i.e. not wash out?
2) recommend a water-stable non-red counterstain that has excellent contrast
to NBT/BCIP ( i.e., is there a different color counterstain that is
compatible with NBT AND aqueous mounting media?)
3) recommend a completely different substrate/counterstain combination that
works with DIG/Alkaline Phosphatase and that can be mounted aqueously?  (
i.e., if the NBT/BCIP just plain sucks, and I am getting the impression that
it does, is there anything out there I can use with Alkaline Phosphatase so
I don't have to completely rework my probes?)


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