Coupling imaged capillary isoelectric focusing with mass spectrometry using a nanoliter valve

Electrophoresis
Cristina Montealegre, Christian Neusüß

Abstract

Imaged capillary isoelectric focusing (iCIEF) is a powerful separation technique applied frequently for the analysis of biotherapeutics. However, direct mass spectrometric characterization is usually not possible. Here, focused peaks from an iCIEF system are transferred to a four-port nanoliter valve where peaks of interest are cut and transferred directly (flow injection) or via capillary zone electrophoresis to electrospray mass spectrometry. At first, flow injection coupling was tested with a mixture of peptides showing intraday precision (RSD) of 20.0 and 3.4% in area and 15.2 and 6.3% in intensity for angiotensin I and leucine-enkephalin, respectively. For the analysis of charge variants in an intact mAb, flow injection and CZE as second dimension were compared, demonstrating the usability of flow injection even for large proteins. However, improved spectra quality was achieved when CZE-MS was applied. In this way, accurate masses were obtained not only for the main isoform, but also for the main acidic and basic variants. These results demonstrate the power of iCIEF-CZE-MS for the analysis of biotherapeutics.

References

Mar 29, 2001·Journal of Chromatography. B, Biomedical Sciences and Applications·R J HarrisA B Chen
Oct 16, 2012·Journal of Separation Science·Oscar Salas-SolanoBrian Nunnally
Feb 1, 2015·Journal of Chromatography. B, Analytical Technologies in the Biomedical and Life Sciences·Bernd MoritzMark Lies
Aug 25, 2015·Electrophoresis·Jens HühnerChristian Neusüß
Apr 8, 2016·Analytical and Bioanalytical Chemistry·Jens Hühner, Christian Neusüß
Jun 7, 2016·Journal of Chromatography. B, Analytical Technologies in the Biomedical and Life Sciences·Rabah GahoualYannis-Nicolas François
Sep 30, 2017·Journal of Chromatography. B, Analytical Technologies in the Biomedical and Life Sciences·Alexandre GoyonAlain Beck

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Citations

Jul 20, 2018·Rapid Communications in Mass Spectrometry : RCM·Rabah GahoualYannis-Nicolas François
Dec 24, 2018·Electrophoresis·Kevin JooßChristian Neusüß
Oct 24, 2018·European Journal of Mass Spectrometry·Jérémie GiorgettiEmmanuelle Leize-Wagner
Oct 17, 2019·Analytical and Bioanalytical Chemistry·Jennifer RömerChristian Neusüß
Nov 2, 2019·Journal of Pharmaceutical and Biomedical Analysis·Julien CamperiNathalie Delaunay
May 28, 2019·Journal of Chromatography. B, Analytical Technologies in the Biomedical and Life Sciences·Antony LechnerYannis-Nicolas François
Apr 27, 2021·Frontiers in Chemistry·Tian Xu, Liangliang Sun
Nov 9, 2019·Analytical Chemistry·Courtney J KristoffLisa A Holland

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