Another IMAC question

James Fethiere james at mpimf-heidelberg.mpg.de
Fri Aug 21 08:51:50 EST 1998


I am using a 1.6ml POROS Ni column to purify a protein eluted from a
FLAG-M1 column with 2mM EDTA.  After a certain time I start to loose
binding to the Ni column.

I would like to know if it's possible that after approximately 1.5L of
these 2mM EDTA preps, there is enough stripping of the Ni from the
column to explain what I see.  I should mention that in the previous
step of purification, EDTA is necessary to chelate Ca (3mM) that is
required for binding of the protein to the FLAG M1 column.  Everything
is done in 50mM Tris pH 8.

 
THank you

James



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