Genomic in situ hyb?

Mikael Niku mikael.niku at helsinki.fi
Tue Nov 17 02:07:50 EST 1998


I'm using in situ hybridization with DIG-labelled oligonucleotide probes to
genomic DNA in tissue sections.

The method works mostly very nicely. However, I'm having problems with
coverage with some tissues and some fixations. That is, I don't get
signal from as many cells as I should. I have tried varying proteinase
treatments and other tricks I know but with limited success.

The problem is especially severe with cross-linking fixatives like
paraformaldehyde,
which works very well for detecting mRNA. I guess this might be because the
chromosomal
proteins are fixed so that they prevent probe from accessing the target
sequence. Paraformaldehyde would be nice to use because it preserves tissue
morphology very well.

Any suggestions are welcome, as well as good protocols for genomic in situ
hyb.
I do have the B-M guide already.







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