Jump to content

  • Log in with Facebook Log in with Twitter Log in with Windows Live Log In with Google      Sign In   
  • Create Account

Submit your paper to J Biol Methods today!
Photo
- - - - -

Compatibility of Urea and Imidazole


Best Answer mdfenko, 25 August 2013 - 02:49 PM

there should be no problem mixing urea and imidazole.

 

keep in mind that imidazole is a buffer and addition of tris may not give you the pH you want (you may have to adjust).

Go to the full post


  • Please log in to reply
2 replies to this topic

#1 ddmf

ddmf

    member

  • Members
  • Pip
  • 2 posts
0
Neutral

Posted 25 August 2013 - 11:03 AM

I would like to make a buffer containing 8 M Urea, 250 mM Imidazole and 1 M NaCl; but I am not sure if Urea and Imidazole can be in the same buffer. Hence, I would greatly appreciate your feedback on this.

 

Background info:

I am purifying a protein using affinity chromatography (N-term Histag, cleavage with thrombin). And I would like to merge the last two steps of my current protocol, namely removal of cleaved Histag fragment and of thrombin. My idea is to pass my overnight cleaved sample through both nickel and benzamidine columns in series, hence leaving me with pure protein in the flow-through. After that I would like to remove the Histag fragment from the Nickel column and the thrombin from the bezamidine column. Thus, I thought about using a Tris buffer (pH 7.5 adjusted with HCl) with 8 M Urea, 250 mM Imidazole and 1 M NaCl; since it should remove everything that is bound by affinity and ionically. However I do not know the compatibility of Urea and Imidazole, indeed I never used Urea before.



#2 mdfenko

mdfenko

    an elder

  • Active Members
  • PipPipPipPipPipPipPipPipPipPip
  • 2,781 posts
132
Excellent

Posted 25 August 2013 - 02:49 PM   Best Answer

there should be no problem mixing urea and imidazole.

 

keep in mind that imidazole is a buffer and addition of tris may not give you the pH you want (you may have to adjust).


talent does what it can
genius does what it must
i do what i get paid to do

#3 ddmf

ddmf

    member

  • Members
  • Pip
  • 2 posts
0
Neutral

Posted 25 August 2013 - 03:09 PM

Thank you a lot for your comment. smile.png






Home - About - Terms of Service - Privacy - Contact Us

©1999-2013 Protocol Online, All rights reserved.