I've been told that imidazole might cleave your protein under certain
conditions; therefore I don't like it. Try pH elution, followed by rapid
neutralization after elution. Should work fine
hope this helps
Arnoud
--
Dr. A.H.M. van Vliet
Vrije Universiteit
Amsterdam
The Netherlands
Sigrid Van Boxstael <svboxsta at vub.ac.be> wrote in message
news:38DA31B9.6ABBFDAC at vub.ac.be...
> Hi,
>> I'm purifying my protein with the His-TAG-method.
> My protein, the ATCase of pyrococcus abyssi consists of two different
> subunits. The His-Tag is attached to one of them.
> After elution with Imidazol (100 mM Imidazol), I concentrated my protein
> and put it on gel.
> On SDS gel it looks very nice and pure, on a native gel on the other
> hand, there is a smear in which I can distinguish one band.
> The native gel is a 6 % Tris-glycine gel.
> Is it possible that the high imidazol concentration has an influence on
> the migration in my native gel.
> Is it possible that because of the His-Tag the protein forms higher
> molecular weight aggregates, dimers?, Trimers?
> Can I remove the Imidazol just by dialyzing?
>>> Thanks in advance
>> Sigrid
>> Sigrid Van Boxstael
> VUB MICROBIOLOGIE
> E. Gryzonlaan 1
> 1070 Brussels
> Belgiuml
>>
