Quick navigation:        Home   |    Site Map   ||    References   |    Biography   ||    Copyright   |    Other copyright   |    Contact us   |   
Protein structure
 

Re: [ccp4bb] rescuing crashing-out protein eluted from Nickel column
 

Basic tutorials: 		 
 

CCP4bb navigation

CCP4bb <-- 2008 <-- February 2008 <-- 14 February 2008
Previous message:
Subject: rescuing crashing-out protein eluted from Nickel column
From: Jacob Wong jacob {- dot -} j {- dot -} wong {- at -} GMAIL {- dot -} COM
Date: 2008-02-14 		Next message:
Subject: Re: rescuing crashing-out protein eluted from Nickel column
From: Jacob Wong jacob {- dot -} j {- dot -} wong {- at -} GMAIL {- dot -} COM
Date: 2008-02-14


Subject: Re: rescuing crashing-out protein eluted from Nickel column
From: Chun Luo chun {- at -} ACCELAGEN {- dot -} COM
Date: 2008-02-14
JJ



Dilute your fractions for this prep.



Do gradient elution in the future. Again dilute the fractions immediately.



Figure out a better buffer will be helpful.



Cheers,



Chun Luo, Ph.D.
The Protein Expert
Accelagen, Inc.
11585 Sorrento Valley Road, Suite 107
San Diego, CA 92121
TeL: 858-350-8085 ext 111
Fax: 858-350-8001
chun@accelagen.com
www.accelagen.com





_____

From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of Jacob
Wong
Sent: Thursday, February 14, 2008 4:26 PM
To: CCP4BB@JISCMAIL.AC.UK
Subject: [ccp4bb] rescuing crashing-out protein eluted from Nickel column



Dear all,



I just ran into this problem and would like to see if I could get some
helpful tips before my protein completely crashes out.



I have a protein as 6His fusion and it remained bound to the Ni resin with
40 mM Imidazole wash (added to 1XPBS) but then was eluted off with 200 mM
(added to 1XPBS). The protein seemed to be highly concentrated in the
elution and began to get cloudy right away, with more and more precipitation
produced over a matter of minutes. I felt so helpless, didn't know what to
do, and then decided to add 5% of glycerol into one of the fractions but
that made it even more cloudy (ohh no...).



While the protein is dying in the tube, do you have some quick remedy for
me? Thanks very much, -J.J.
ProteinCrystallography.org: Copyright 2006-2007 by Quid United Ltd