I am not a big fan of sonication. Try changing your way of disrupting the
cells.
I have compared sonication vs mechanical stress on several unrelated proteins,
and for me a good old french press wins every time. If you want to get all
modern and fancy, a cell disruptor gives similar results.
Cheers,
Mads Gabrielsen
[Hide Quoted Text]
On Mar 2, 2008, at 11:47 PM, Tim Gruene wrote:
Hi all
sorry, for offtopic query...
I am trying to purify my protein by Ni-NTA affinity chromatography. After
sonication as i centrifuge bacterial lysate, soon after 10 min whole
lysates
get precipitated during loading on the column and some time it remain
soluble too. if i get purified through the column without precipitation,
it
gets precipitated during dialysis.
I have tried lot, by chnaging buffers, increasing salt or deacreasing salt
or no salt at are helpless.
I do purifiaction in cold room.
can any one suggest some solution?
Thanks in advance.
NSH
--
Dr. Mads Gabrielsen
GBRC, B217
Division of Biochemistry and Molecular Biology
IBLS
University of Glasgow Phone Office: 01413308119
G12 8QQ Phone Lab: 01413306449
UK E-mail: [log in to unmask]
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