P.C. <PC at no.email.sorry> wrote:
> I find it very strange that lysozyme would bind at all to Q sepharose
> (not that I tried it myself). Can it be that the real peak actually did
> not elute from SP yet?
I redid the experiment. From Q-sepharose 2-3 overlapping peaks in
gradient. From SP one peak in gradient. SDS-PAGE shows identical bands
in all relevant fractions, corresponding to lysozyme size.
> Are your columns fresh and clean? If not, wash them in 8M GuHCl.
> Or NaOH as APB recommends.
They were freshly packed, hence the lysozyme test.
> And finally, are you sure that the vial that you used for chromatography
> was actually chicken egg lysozyme and not chicken egg albumin? (please
> do not take offence, it is just another thing to check).
No offence taken, strange things happen. I checked the vial, it is
lysozyme (hen egg white, 3 x crystallized).
I only wanted to use the Q-sepharose column and it worked in my
isolation so I leave the mysterious binding to SP in the folder
"mysteries". But if someone has any explanation to offer I would
be grateful.
--
Kaj
