Method Development for Multiplexing CGE-LIF Based Glycoanalysis

Motivation

In cooperation with the glyXera GmbH in Magdeburg/Germany, a method for HT/HP/HR glycoanalysis, based on multiplexed capillary gel electrophoresis with laser induced fluorescence detection (xCGE-LIF) utilizing a DNA-sequencer, is developed at the MPI. The application of this technique to glycoanalysis using instruments with up to 96 capillaries in parallel, results in massive reduction of the effective separation time per sample combined with an impressive sensitivity achieved due to LIF detection and therefore, shows high potential for HT/HP/HR glycoprofiling of glycoconjugates.

Workflow of high-throughput glycosylation pattern analysis of glycoproteins utilizing a multiplexing capillary-DNA-sequencer.

© MPI Magdeburg (BPE)

Workflow of high-throughput glycosylation pattern analysis of glycoproteins utilizing a multiplexing capillary-DNA-sequencer.

© MPI Magdeburg (BPE)

Aim of the project

The aim of this project is to further investigate and to improve this innovative approach for use in various fields of application. Therefore, the sample preparation method and workflow are further optimized with respect to performance and feasibility regarding HT, and the separation parameters (e.g.: gel matrix composition, voltage or capillary length) are evaluated regarding HP and HR, to find the optimum for an automated separation for each of a set of different types of samples.

Publications

Hennig, R.; Rapp, E.; Kottler, R.; Cajic, S.; Reichl, U.: N -Glycosylation Fingerprinting of Viral Glycoproteins by xCGE-LIF. In: Carbohydrate-Based Vaccines: Methods and Protocols, pp. 123 - 143. Springer, New York (2015)

MPG.PuRe

DOI

Borowiak, M.; Hennig, R.; Kottler, R.; Reichl, U.; Rapp, E.: Tailored methods for CGE-LIF based multiplexed high-throughput glycoanalysis with respect to sample characteristics. In Glycoconjugate Journal, 28 (5 ), p. 332. 21st International Symposium on Glycoconjugates - Glyco 21, Wien, Östereich, August 21, 2011 - August 26, 2011. (2011)

MPG.PuRe

DOI

publisher-version

Schwarzer, J.; Rapp, E.; Reichl, U.: N-Glycan Analysis by CGE-LIF – Profiling Influenza A Virus Hemagglutinin N-Glycosylation during Vaccine Production. Electrophoresis 29 (20), pp. 4203 - 4214 (2008)

MPG.PuRe

DOI

Patents

[1] E Rapp, J Schwarzer, U Reichl, C Bohne: Method for automated high throughput identification of carbohydrates and carbohydrate mixture composition patterns as well as systems therefore. European patent: EP2533039 (A1) (2008).

[2] E Rapp, J Schwarzer, U Reichl, C Bohne: Method for automated high throughput identification of carbohydrates and carbohydrate mixture composition patterns as well as systems therefore. US patent: US2013075258 (A1) (2009).