Co-Ip - bands in my pre-clearing controls - pre-clearing step - how to improve it? (Nov/05/2007 )
Hello hello.
I am currently immunoprecipitating my protein complex on Sepharose G.
The thing is, I get the appropiate band in the preclearing step as well.
Is my IP non-specific or does my protein have some affinity to protein G?
And what to do to ensure specificity?
The cell extract controls work just fine.
I do not have any other antibody for my protein suitable for IP and there's none available at the moment.
-Telomerase-
Your protein seems to have some affinity for protein G.
You could try to increase NaCl concentration up to 0.5M for incubation and washing steps.
-Missele-
QUOTE (Missele @ Nov 5 2007, 02:20 PM)
Your protein seems to have some affinity for protein G.
You could try to increase NaCl concentration up to 0.5M for incubation and washing steps.
You could try to increase NaCl concentration up to 0.5M for incubation and washing steps.
Hmmm, I am currently using 50mM KCl, maybe the stringency is too low indeed.
Thanks, I am going to try this one out. Maybe not 0.5M, nevertheless higher...
-Telomerase-