Detection of 3H-labelled proteins

Jonathan B. Marder marder at agri.huji.ac.il
Thu Oct 9 04:47:04 EST 1997


REW wrote in message <61h2mb$lsa$1 at nntp.ucs.ubc.ca>...
>Hi,
>I'm trying to detect tritium-labelled proteins in dried SDS-PAGE gels
by
>autoradiography.   What's the best way to do this?  I've been told to
soak
>the gels in 1M salicylate before drying then to expose to film at -80C.


After fixing, wash the gel for 30 min. in water (running tap water is
good).
Then you can do the salicylate incubation.
BTW - be careful of sticky gels i.e. dry them properly!
Reference is Chamberlain, J.P. (1979) Analytical Biochem. 98, 132-135.

>Question 1: Is there a more sensitive method?
In my experience, the method is equal to PPO/DMSO for sensitivity and
resolution. This is only true for tritium. Salicylate gives fuzzy bands
with stronger
beta emitters (e.g. 35-S). One advantage of salicylate is that it can be
washed out
easily and the protein bands eluted.

>Question 2:  Is it really necessary to expose the film at -80C?
Yes! This is part of the fluorography process and needed with other
fluors too.


I hope that this is of help
Jonathan B. Marder   <MARDER at agri.huji.ac.il>
Department of Agricultural Botany, The Hebrew University of Jerusalem
Faculty of Agriculture, P.O.Box 12, Rehovot 76100, ISRAEL
Phone: +972 8 9481918   Fax:   +972 8 9467763
Web page:    http://www.agri.huji.ac.il/~marder





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