Q: Is Azide Inhibition of HRP reversible? (I hope not)
Frank O. Fackelmayer
fof1 at chclu.chemie.uni-konstanz.de
Wed Dec 10 04:12:27 EST 1997
In article <66jq41$m2l at mserv1.dl.ac.uk>, wgschech at med.uni-tuebingen.de wrote:
> Hi all,
> I noticed that sodium azide inhibits ECL detection of proteins in
> western blots. I'm too lazy to strip my blot (officially, I'm
> afraid that stripping will worse my detection level, of course...)
> My primary antibody is stored in a buffer containig some amount of
> sodium azide for preservation, so I wonder if incubating my
> already probed blot with this AB will abolish the peroxidase
> conjugated to the secondary AB of my first probe.
> Any ideas, comments, flames, suggestions are appreciated!
> This message is RNAse free - please don't touch!
> Wolfgang Schechinger
> University of Tuebingen
> email: wgschech at med.uni-tuebingen.de
> !Junk mail is *not* appreciated!
> SPAMs will be answered automatically by sending my 128MByte virtual
> This is *NO* joke.
> usual disclaimers apply
Sodium azide is known to kill peroxidase activity completely, as you
noticed. I don´t know if this can be used instead of stripping, but I
won´t try because there is always some unreacted primary antibody from the
first detection that will then bind secondary ab in your second detection.
So, you´ll get NEW signals at the old site. It way work if the primary abs
are from different species (e.g. mouse and rabbit), and you have secondary
abs that do not cross-react.
Hope this helps
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