david.kirke at virgin.net
Mon Aug 31 09:54:59 EST 1998
If you know the molecular weight of your protein, you could look at it on a
zymogram gel. This is a non-denaturing gel containing caesin or gelatine.
After running, the gel is incubated for a few hours in a suitable buffer to
allow any proteolysis to occur. The gel is then stained with coomassie
blue. The caesin or gelatine background will be stained blue and any
proteases present will appear as clear bands.
These gels are usually quite sensitive and you should be able to see whether
the protease band corresponds to the molecular weight of your protein. You
will have to run prestained molecular weight markers which will show up on
the blue background.
Alan Tunnicliffe wrote in message <35EAA742.167E at imb-jena.de>...
>We currently have a protein solution which is 99% (plus) pure,
>however we have discovered protease activity within our sample.
>This protease activity can be prevented by addition of EDTA
>and using several protease inhibitors.
>The question I have, is how can I discover if the protein we are working
>has intrinsic protease activity (it is not supposed to) or if the less
>1% impurity is infact a protease ?
>We have tried silver stained gels
>and Native gels to see if we can detect the contaminant.
>Any advice or help would be greatly appreciated
>atunn at imb-jena.de
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