: Thursday, June 20, 2013 4:34 PM
To: Zhang, Zhen
Cc: CCP4BB@JISCMAIL.AC.UK
Subject: Re: [ccp4bb] Puzzling observation about size exclusion chromatography
Hi Zhen -
Superdex is known to have some ion-exchange characteristics, so that it
can weakly interact with some proteins. This is why
@JISCMAIL.AC.UK
Subject: Re: [ccp4bb] Puzzling observation about size exclusion chromatography
Dear Zhen,
I should also point out that the statement Matt made (Superdex is known to
have some ion-exchange characteristics, so that it can weakly interact with
some proteins.) is not completely correct
Dear all,
I just observed a puzzling phenomenon when purifying a refolded protein with
size exclusion chromatography. The protein was solubilized by 8M Urea and
refolded by dialysis against 500mM Arginine in PBS. The protein is 40KDal and
is expected to be a trimer. The puzzling part is the
If your protein elutes very late, that means it's binding to the column matrix
(so all estimates of size go into the trash). Check to see that the ionic
strength of buffer is reasonable (equivalent to, say, 150 mM NaCl). If so, then
the only solution is to go to a different matrix type.
Pat
On
@JISCMAIL.AC.UK
Subject: Re: [ccp4bb] Puzzling observation about size exclusion chromatography
If your protein elutes very late, that means it's binding to the column matrix
(so all estimates of size go into the trash). Check to see that the ionic
strength of buffer is reasonable (equivalent to, say, 150 mM
To: CCP4BB@JISCMAIL.AC.UK
Subject: Re: [ccp4bb] Puzzling observation about size exclusion chromatography
Isn't 18 mLs into a Superdex 200 10/300 column run out near where the 670kD
marker is, just after the void at ~15 mLs? Zhen, did you mean ~500kD rather
than 5kD?.
Kushol
Kushol Gupta, Ph.D
: [ccp4bb] Puzzling observation about size exclusion
chromatography
Hi Kushol,
No. The void for the column is 8ml and the whole volume of the column is
24ml. You must be talking about a different column.
Zhen
-Original Message-
From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK
Hi Zhen -
Superdex is known to have some ion-exchange characteristics, so that it
can weakly interact with some proteins. This is why the manufacturer
recommends including a certain amount of salt in the running buffer. I
have had the same experience with a few proteins, including one that
: [ccp4bb] Puzzling observation about size exclusion chromatography
Hi Zhen -
Superdex is known to have some ion-exchange characteristics, so that it
can weakly interact with some proteins. This is why the manufacturer
recommends including a certain amount of salt in the running buffer. I
have
