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DIG labeled probes for ISH

Mario Nenno [Bio.] nenno at rhrk.uni-kl.de
Thu Oct 24 06:34:09 EST 1996


I don't have a reference at the moment. But I use DIG-labeled oligos
for ISH. The problem with small probes or low-copy sequences is, that 
you have to use a good amplification system.

In our hands a good starting point is the following:
- Anti-DIG antibody from mouse  
- Anti-mouse-DIG antibody
- Anti-DIG-antibody conjugated with either FITC or AP or similar

Boehringer Mannheim calls this combination 'DIG amplification system'.

Good luck,

>In article <3264B583.44D8 at alf.biochem.mpg.de>, Gabriele Kerber
><kerber at alf.biochem.mpg.de> wrote:

># Ms. K.K. Martin wrote:
># > 
># > I have used both 35S and DIG for ISHH.  I far prefer DIG and will never,
># > ever use 35S again.  DIG is easier, safer, faster (results in hours rather
># > than days or weeks you get good spatial localisation and, best
># > of all, you avoid all that nasty paperwork associated with keeping track
># > of your radioactive waste.  Boehringer Mannheim have very helpful manuals
># > for using DIG in ISHH.  Good luck.
># DIG is wonderful for IHS with riboprobes. But if you want to use 
># oligonucleotides, you will probably have a lot of problems with 
># DIG. With oligos it4s better to use 35S or even 32P.
># Gabriele

>Is there a published side by side comparison of 35-S vs. DIG in the
>literature?  I have the impression, from ISHers I've talked to, that 35-S
>ISH is considerably more sensitive than DIG.  However, I have no reference
>for this claim.

>Gene S. Huh, Ph.D.
>Department of Molecular and Cell Biology
>Life Sciences Addition, Room 221
>University of California at Berkeley
>Berkeley, California   94720-3200

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