I need help removing IgG from protein homogenates!

Jil Tardiff jtardiff at westnet.com
Thu Jan 9 18:39:18 EST 1997


Hi folks, 

I could really use a hand with a confounding problem.

I have some transgenic mice and I am trying to study the transgenic
protein (simple). The problem is that our animal facility has become
infected with *something* of late and the end result is that my mice are
really making boatloads of IgG. Thus, whenever I perform a protein
determination my results are skewed because of the IgG load. What's
more, when I run an SDS-PAGE the Tg proteins are not running correctly.
The subsequent Western (since we're using goat anti-mouse HRP with ECL
to detect) has a whopping IgG H+L signal (detectable without primary of
course) that is making a real shambles of my attempts to characterize
the mutant protein. The primary Ab is hybridoma supt of c-myc (9E10).

What I would *like* to do is figure out a way to *remove* the IgG from
the initial protein homogenate. The protein is isolated from whole mouse
hearts in a low-conc KCl-KPO4 buffer. And yes, I try mightily to wash
the blood out before homogenization.

Any and all help, suggestions, advice would be greatly appreciated!

Thanks,

 Jil Tardiff
 Albert Einstein College of Medicine and
 the Columbia-Presbyterian Medical Center

jtardiff at westnet.com



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