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High-resolution separations of protein isoforms with liquid chromatography time-of-flight mass spectrometry using polymer monolithic capillary columns

Lookup NU author(s): David Blinco, Dr Glen Kemp, Dr Achim Treumann

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Abstract

The separation of intact proteins, including protein isoforms arising from various amino-acid modifications, employing a poly(styrene-co-divinylbenzene) monolithic capillary column in high-performance liquid chromatography coupled on-line to a time-of-flight mass spectrometer (MS) is described. Using a 250 mm × 0.2 mm monolithic capillary column high-sensitivity separations yielding peak capacities of >600 were achieved with a 2h linear gradient and formic acid added in the mobile phase as ion-pairing agent. The combination of high-resolution chromatography with high-accuracy MS allowed to distinguish protein isoforms that differ only in their oxidation and biotinylation state allowing the separation between structural isoforms. Finally, the potential to separate proteins isoforms due to glycosylation is discussed.

Publication metadata

Author(s): Eeltink S, Wouters B, Desmet G, Ursem M, Blinco D, Kemp GD, Treumann A

Publication type: Article

Publication status: Published

Journal: Journal of Chromatography A

Year: 2011

Volume: 1218

Issue: 32

Pages: 5504-5511

Print publication date: 21/06/2011

ISSN (print): 0021-9673

ISSN (electronic): 1873-3778

Publisher: Elsevier BV

URL: http://dx.doi.org/10.1016/j.chroma.2011.06.049

DOI: 10.1016/j.chroma.2011.06.049

PubMed id: 21741050

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