Heart-cut nano-LC-CZE-MS for the characterization of proteins on the intact level
- Multidimensional separation techniques play an increasingly important role in separation science, especially for the analysis of complex samples such as proteins. The combination of reversed-phase liquid chromatography in the nanoscale and CZE is especially beneficial due to their nearly orthogonal separation mechanism and well-suited geometries/dimensions. Here, a heart-cut nano-LC-CZE-MS setup was developed utilizing for the first time a mechanical 4-port valve as LC-CE interface. A model protein mixture containing four different protein species was first separated by nano LC followed by a heart-cut transfer of individual LC peaks and subsequent CZE-MS analysis. In the CZE dimension, various glycoforms of one protein species were separated. Improved separation capabilities were achieved compared to the 1D methods, which was exemplarily shown for ribonuclease B and its different glycosylated forms. LODs in the lower μg/mL range were determined, which are considerably lower compared to traditional CZE-MS. In addition, this study represents the first application of an LC-CE-MS system for intact protein analysis. The nano-LC-CZE-MS system is expected to be applicable to various other analytical challenges.
Author: | Kevin Jooß, Nico Scholz, Jens Meixner, Christian Neusüß |
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Institutional Author: | Kevin Jooß |
Institutional Author: | Nico Scholz |
Institutional Author: | Jens Meixner |
Institutional Author: | Christian Neusüß, |
DOI: | https://doi.org/10.1002/elps.201800411 |
ISSN: | 0173-0835 |
eISSN: | 1522-2683 |
Pubmed Id: | https://pubmed.ncbi.nlm.nih.gov/30575976 |
Source Title (English): | Electrophoresis |
Document Type: | Article |
Language: | English |
Year of Completion: | 2019 |
Release Date: | 2021/05/11 |
Tag: | Chromatography, Reverse-Phase/instrumentation/methods; Electrophoresis, Capillary/instrumentation/methods; Glycosylation; Hydrogen-Ion Concentration; Limit of Detection; Ribonucleases/analysis; Tandem Mass Spectrometry/instrumentation/methods |
Volume: | 40 |
First Page: | 1061 |
Last Page: | 1065 |
Faculty: | Chemie |
Relevance: | peer reviewed |