Thanks for the pat on the back. Actually, the ion-exchange/RPC combination
is useful, but in the opposite order. Take a look at Link et al., Nature
Biotechnol. 17 (1999) 676 [John Yates' group, U. of Wash.]. They're
performing proteome analysis by trapping lots of proteins in a
cation-exchange cartridge or capillary segment. The SCX material is pulsed
with 10 mM salt, stepping off a collection of the least basic proteins.
These are trapped downstream on a RPC capillary segment, which is eluted with
the usual ACN gradient into MS/MS for characterization. After
reequilibration of the RPC material, a pulse of saltier buffer is used to
step a more basic collection of proteins off the SCX segment, etc. This is
continued for a total of 12 pulses off the SCX material. Each pulsed
collection is resolved at the RPC step and characterized by MS/MS. I have a
chromatogram that John Yates gave me of the ribosomal proteins of C. elegans;
it looks like a collection of 12 peak clusters along the X-axis. If it's OK
with him, I'll send it as an attached file to anyone who requests it. In
short, the IEX/RPC combination is a quick way to characterize large
protein/peptide collections online.
Andy Alpert
PolyLC Inc.
tel: (410) 992-5400 e-mail: PolyLC@aol.com
*************************************************************************
<< Subj: IE HPLC
Date: 12/03/1999 2:33:57 PM Eastern Standard Time
From: mcmillen@morel.uoregon.edu (Deb McMillen)
Sender: abrf-request@aecom.yu.edu (Association of Biomolecular Resource
Facilities)
To: abrf@aecom.yu.edu (Recipients of ABRF List)
Thanks to Jenny Shipway, Andy Alpert and Amos Heckendorf for a great
discussion of ion exchange chromatography. I appreciate these
mini-symposia. And, Jenny, someday, that reverse phase column in line
before the ion exchange column might be a useful tool--so don't dismiss
the idea!
Deb McMillen
Institute of Molecular Biology
University of Oregon
Eugene OR 97403