John Philo wrote:
[snip]
>> It is possible to use ion exchange with (non-ionic) urea, although I
> don't think I have ever heard of anyone doing it with urea
> concentrations that high.
I did it as a PhD-student. I was working with large peptides, almost
protein-sized.
Worked fine; I used a FPLC and different ion-exchange columns. And,
before somebody starts complaining about the havoc the urea will do on
the pumps - every two to three months I had to replace the seals, that
was all.
[snip]
>> Also, one problem with ion exchange for proteins which tend to aggregate
> is that typically your protein all sticks near the top of the column and
> thus is very highly concentrated and prone to aggregate. Thus you may
> want to consider chromatography under conditions where the impurities
> will bind but your protein will flow through.
He could also adjust the pH to a value, where the binding is not as
strong; e.g., with an anion-exchange column he could use pH 7 or lower,
instead of pH 8. Another possibility would be to use weaker ion-exchange
materials, e.g., DEAE.
[snip]
