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-   -   Protein G on the western? (http://www.molecularstation.com/forum/immunoprecipitation-forum/48143-protein-g-western.html)

Enzy70 01-14-2009 07:16 PM

Protein G on the western?
 
Hi!
Can anybody tell me if it's possible for the protein G used for the IP to show up as a ~23KDa band?

The reason why I'm asking this is that I've been using the TrueBlot secondary ab which, according to the manufacturer, doesn't recognize the reduced forms of heavy and light chain of the ab but only the intact one.
If what they show is true, I must assume that the band that I identify at ~23KDa after IP, is something else and not the light chain of the ab... could it be the protein G used for the IP? Is it even possible?

Thanks

danfive 01-14-2009 11:25 PM

Re: Protein G on the western?
 
Protein G should not wash off.
23 KDa band may be a residual protein, and you may be able to get rid of it with extraneous washes.
My off the cuff suggestion, is for you to try robust washes.
For example if you did 4 washes try 12, if you did 8 try 20-24.

More info, if you want more specific suggestions.


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