Cleaning up primary antibodies??
ajardi at po-box.mcgill.ca
Wed Dec 13 09:07:14 EST 2000
you can apply you crude antisera to a protein A column and the IgG Abs will
bind. The IgG can then be eluted with either a 50 mM glycine solution pH 3.0
of a 50 mM ethanolamine solution pH 11.0. In any case the column effluent
contain the IgG fraction should be collected into a tube containing enough 1 M
Tris pH 7.2 to quickly neutralize the pH of the effluent. You will then have
to test the acid (glycine) and base (ethanolamine) eluted IgG by ELISA or
Western blot to ensure that the purified IgG Abs are still active.
Hope this helps
Institute of Parasitology
Neal Robert Melvin wrote:
> Is it possible to 'clean up' a primary antisera by incubating with protein
> A, and then somehow gently eluting the IgGs from the protein A to be able
> to use?? What is the best way to go about this??
> Any help would be appreciated!
> Neal Melvin
> University of Calgary - Faculty of Medicine
> Department of Neuroscience - Neuroscience Research Group (NRG)
> Health Sciences Centre
> 3330 Hospital Drive, NW
> Calgary, Alberta, Canada
> T2N 4N1
> e-mail: nrmelvin at ucalgary.ca
> Phone: (403) 220-4490 (lab)
> (403) 220-7035 (office)
> (403) 283-8731 (fax)
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