HIS re-purification

Stephan Wenkel xyz at xyz.com
Wed Jan 21 05:08:31 EST 2004


I have purified a HIS-tagged protein from E.coli. After dialysis against a
buffer containing no imidazole I would like to put it again on the Ni-column
to get rid of some minor contaminations. Unfortunately it looks like that
the protein binds very tight to the resin that I can not get it down
anymore. Why? I can only elute it by boiling @99 degrees in the presence of
0,5M imidazole plus gel loading buffer.

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