Ni-agarose elution

Charles A Miller oravaxcm at world.std.com
Sun Sep 21 23:42:00 EST 1997


If you don't want to lose your nickel resin and don't want
to use Imidazole (or don't have any...or its too expensive for
your budget), then why not just drop the pH to 5.6 or 4.56?

This should elute your protein really well, while leaving your nickel 
resin intact. Then you could just readjust the pH of your protein
or dialyze it into another buffer. 

One question, if you elute with EDTA (which will strip the
nickel and protein complex off your column) will you still have
nickel complexed with protein? Well, I guess not, it'll just chelate
the nickel...which could get pretty expensive.

Chuck
oravaxcm at world.std.com








brett (brett at BORCIM.WUSTL.EDU) wrote:
: >Hi
: >I am on my way to purify a 6xHis tagged protein.  Does anybody know if I 
: >can elute the protein from Ni-agarose by just adding SDS-PAGE loading 
: >buffer, instead of using imidazole buffer?
: >
: >nam
: >
: >-- 
: >***************************************************
: >Tsunehisa Namba
: >Department of Anesthesia, Kyoto University Hospital
: >Sakyo-ku, Kyoto 606-01, Japan
: >Fax:+81-75-752-3259
: >E-mail: drnamba at kuhp.kyoto-u.ac.jp


: I take it you're not interested in saving the resin.  Why not make your sample
: buffer 100mM EDTA? That seems to work for me.
: Brett Lindenbach
: brett at borcim.wustl.edu

: Dept Molecular Microbiology
: Washington University School of Medicine
: Box 8230
: 660 S. Euclid Ave
: St Louis, MO 63110




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