Re: In situ with AP and Fast Red TR/Naphthol
I use BioGenex's New Fuchsin (www.biogenex.com) which is slightly soluble in
xylene but not soluble in the non-aliphatic hydrocarbons, I use Richard-Allan's
Clear-Rite 3 for clearing instead of xylene when I use this chromogen and
coverslip with Permount.
The New Fuchsin gives a nice bright red end product with Alk. Phos. and is
permanent. It is also available from Dako and others, I just happen to like
the BioGenex kit.
Laboratory of Molecular Carcinogenesis & Mutagenesis
University of North Carolina at Chapel Hill
Lisa C Ranford Cartwright wrote:
> Hello histo folk
> We are performing in situ PCR on tissue sections (fixed, embedded,
> sectioned, etc). The PCR results in DIG incorporation into the DNA, which
> we then detect with an anti-DIG antibody conjugated to alkaline
> phosphatase. This part works fine (using a fluorescent detection system).
> We now wish to use a colorimetric substrate rather than a fluorescent one,
> and FAST Red TR/Naphthol had been suggested. This is supposed to give a
> bright red stain. However we also wish to counterstain with Giemsa, and I
> understand the Fast Red is soluble in organics. The final methanol
> concentration in the Giemsa stain will of course be low (5% stain) - is
> this likely to dissolve the fast red deposits?
> I've never used FAst Red TR before. Has anyone tried Fast Red TR and Giemsa
> together? Any thoughts on whether it will work? Any alternatives (note we
> don't want to use NBT/BCIP substrate as our tissue already as some blackish
> material in it which we think will be confused with the signal).
> Thanks for any suggestions, comments and assistance.
> Lisa Ranford-Cartwright,Ph.D.
> Division of Infection & Immunity
> Institute of Biomedical & Life Sciences
> Joseph Black Building
> University of Glasgow
> Joseph Black Building, Glasgow G12 8QQ
> Scotland, UK
> Tel: 00 44 141 330 2639
> Fax: 00 44 141 330 4600
> Email: L.C.Ranford-Cartwright@bio.gla.ac.uk
> Lab webpages : http://www.gla.ac.uk/ibls/II/lrc/
<< Previous Message | Next Message >>