His-tag purification

Artem Evdokimov AEVDOKIMOZ at cinci.rr.com
Thu Oct 31 22:14:07 EST 2002

Ni-NTA purification will inevitably give you some background. In order to
*minimize* background (note the emphasis on minimize) you can optimize your
pH and salt concentration (experimentally, of course) - the result would be
individual for a specific protein. You also should try binding the protein
to the beads in ~20 mM imidazole - this often reduces the background
dramatically. Also, you should try to maximize the protein/resin ratio - it
will help compete out junk.

In the end, if everything else fails you can follow your first step of
purification with some sort of secondary step.


More information about the Proteins mailing list