imidazol eluent alternative in his-tagged proteins

David Tucker David.Tucker at REMOVEME.hsn.csiro.au
Thu Nov 4 07:40:41 EST 1999


>A second possible way to elute your protein is to chelate the nickel
>with EDTA. Try elution with 100mM EDTA. The column (which is usually
>slightly blue) will turn white as the nickel elutes together with your
>protein. Elution with EDTA also works very well, but has the drawback
>that the eluted protein will have to be dialysed to get rid of nickel
>and EDTA.
>

Using EDTA to strip the column worked quite well in lab where I did my honours 
degree. We simply collected the blue stuff as it came of the column (we used a 
Cu2+ charged HiTrap (Pharmacia) column), and then stuck it onto a little PD10 
gel filtration column (again, Pharmacia). Purifications were therefore quick 
and very very easy...even for a bumbling honours student!

Whether the EDTA is detrimental to your protein/haeme complex, I guess you 
would know more about that than me!

Best of luck

David 




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