Heart-cut nano-LC-CZE-MS for the characterization of proteins on the intact level
| Autor: | Christian Neusüß, Nico Scholz, Jens Meixner, Kevin Jooß |
|---|---|
| Rok vydání: | 2018 |
| Předmět: |
Detection limit
Chromatography Reverse-Phase Chromatography Glycosylation Chemistry 010401 analytical chemistry Clinical Biochemistry Protein species Ribonuclease B Model protein Intact protein Electrophoresis Capillary 02 engineering and technology Hydrogen-Ion Concentration 021001 nanoscience & nanotechnology 01 natural sciences Biochemistry Mechanical valve 0104 chemical sciences Analytical Chemistry Characterization (materials science) Ribonucleases Limit of Detection Tandem Mass Spectrometry 0210 nano-technology |
| Zdroj: | Electrophoresis. 40(7) |
| ISSN: | 1522-2683 |
| Popis: | 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. |
| Databáze: | OpenAIRE |
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