I need help removing IgG from protein homogenates!

Martin Offterdinger a8803349 at unet.univie.ac.at
Fri Jan 10 02:27:08 EST 1997


On Thu, 09 Jan 1997 18:39:18 -0500, Jil Tardiff <jtardiff at westnet.com>
wrote:

>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
Hello
What about removing IgG with a Protein G column- this will bind
Immunoglobulins -the column can be recycled by acid elution with
Glycin /Hcl pH=2.7-this is a common method to purify Antibodies and
should also work for your purposes.
Martin



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