Profiling, Relative Quantification, and Identification of Sialylated N-Linked Oligosaccharides by UPLC-FLR-ESI/MS After Derivatization with Fluorescent Anthranilamide. Butré, C. I., Largy, E., Cantais, F., & Delobel, A. Methods in Molecular Biology (Clifton, N.J.), 2271:237–247, 2021.
Profiling, Relative Quantification, and Identification of Sialylated N-Linked Oligosaccharides by UPLC-FLR-ESI/MS After Derivatization with Fluorescent Anthranilamide [link]Paper  doi  abstract   bibtex   
The presence of sialic acids is one characteristic of glycosylated therapeutic proteins. The presence of these charged monosaccharides is critical for the immunogenicity properties and structural properties of the proteins. Profiling of the N-glycans and their charge state is a requisite for complete protein characterization. Two analytical methods developed on released N-glycans are described in this chapter, allowing for the determination of the sialoglycosylation with different levels of details. In the first method (AEX-HILIC/FLR), N-glycans are separated based on their charge and the average charge state can be determined from the fluorescence profile. In the second method (AEX-RP-FLR-MS), N-glycans are also separated based on their charge and the sialylation level is determined based on the fluorescence signal. In addition, in this method, the N-glycans are also separated by type and identified with the hyphenated MS. For both methods, an optimized protocol with fast and high-throughput sample preparation and purification is presented.
@article{butre_profiling_2021,
	title = {Profiling, {Relative} {Quantification}, and {Identification} of {Sialylated} {N}-{Linked} {Oligosaccharides} by {UPLC}-{FLR}-{ESI}/{MS} {After} {Derivatization} with {Fluorescent} {Anthranilamide}},
	volume = {2271},
	copyright = {All rights reserved},
	issn = {1940-6029},
	url = {https://link.springer.com/protocol/10.1007/978-1-0716-1241-5_17},
	doi = {10.1007/978-1-0716-1241-5_17},
	abstract = {The presence of sialic acids is one characteristic of glycosylated therapeutic proteins. The presence of these charged monosaccharides is critical for the immunogenicity properties and structural properties of the proteins. Profiling of the N-glycans and their charge state is a requisite for complete protein characterization. Two analytical methods developed on released N-glycans are described in this chapter, allowing for the determination of the sialoglycosylation with different levels of details. In the first method (AEX-HILIC/FLR), N-glycans are separated based on their charge and the average charge state can be determined from the fluorescence profile. In the second method (AEX-RP-FLR-MS), N-glycans are also separated based on their charge and the sialylation level is determined based on the fluorescence signal. In addition, in this method, the N-glycans are also separated by type and identified with the hyphenated MS. For both methods, an optimized protocol with fast and high-throughput sample preparation and purification is presented.},
	language = {eng},
	journal = {Methods in Molecular Biology (Clifton, N.J.)},
	author = {Butré, Claire I. and Largy, Eric and Cantais, Fabrice and Delobel, Arnaud},
	year = {2021},
	pmid = {33908012},
	keywords = {AXH, AXR, Mass spectrometry, N-glycans, Sialic acids},
	pages = {237--247},
}
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