Deb McMillen
Institute of Molecular Biology
University of Oregon
Not affiliated with the Nest Group--but always glad to listen when Amos
Heckendorff gives advice about chromatography!
On Thu, 15 Jan 1998, Brett Phinney wrote:
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> I have a pretty basic chromatography question that I hope someone can help me with. I am trying to separate 2 integral membrane proteins. The problem is that these two proteins have almost the same MW (approx. 56 KD each) so SDS-PAGE does not work very well. I have an HPLC and have been trying to separate these proteins using a RP-C18 column, but this also has not worked so well, I think that the proteins may be too hydrophobic and thus are sticking very tightly to the column and not coming off. I wanted to try either ion-exchange or maybe a less hydrophobic column like a C4 or CN. The proteins have pI's of 6 and 9 so I was thinking Ion exchange with a non-ionic detergent such as octly glucoside may be the way to go.
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> What should I try? If ion-exchange is the most appropriate, what would be a good column to try. Considering the cost I don't really feel like spending $700-1000 dollars on a column that will not separate my proteins, although I may not have a choice...
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> Thanks a lot in advance for the help
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> Brett Phinney
> North Carolina State University
> Phone (919) 515-5786
> Fax (919) 515-2047
> phinney@bchserver.bch.ncsu.edu
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