in situ hybridization

Gabriele Kerber kerber at alf.biochem.mpg.de
Thu Mar 6 04:45:19 EST 1997


William wrote:
> 
> Hi, I'm doing in situ hybridization of cartilage. The problem is that
> the cyro sections do not stay on the slide. I've used all kinds of
> slides such poly lysine, superfrost, probe on, you name it. A collegue
> told me to incubate the sections with the slides for 2 hrs at 60C. I've
> done that and indeed the sections stay on, but I'm afraid there might be
> degradation of the rna. Does anybody else have a better way.
> Thanks
> 
> William


Hi William,

I´m also making in situ hybridizations, but with nervous tissue.
I´m using slides, coated with 3aminomethyltriethoxysilane and don´t 
have any trouble with that.
Once I had also the problem, that the cryo-sections did not stay on the slides.
But there was the SDS in one washing step the problem. Now I use SDS only in the
hybridization mix and my sections stay where they should.

I take normal slides (no superfrost...), bake them for 4 h at 200 °C, wash them 5 min
in aceton, dry them 5 min at room temperatur, then 5 min aceton (200 ml) + 5 ml silane,
wash 2 times in DEPC-water and dry them at 37° - 50°C over night. 

Good luck
Gabriele


***********************************
Gabriele Kerber
Max Planck Institut fuer Psychatrie
Abteilung Neuromorphologie
Am Klopferspitz 18a
82152 Martinsried
Germany
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