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Development of a novel, label-free N-glycan method using charged aerosol detection.

Authors :
Knihtila R
Chemmalil L
Sawant P
Bhavsar S
Kuang J
Shao C
Atsma J
Li Z
Ding J
Source :
Journal of chromatography. B, Analytical technologies in the biomedical and life sciences [J Chromatogr B Analyt Technol Biomed Life Sci] 2022 Dec 01; Vol. 1212, pp. 123502. Date of Electronic Publication: 2022 Oct 17.
Publication Year :
2022

Abstract

Monoclonal antibodies (mAbs) are complex glycoproteins that are developed for treatment of various therapeutic indications such as cancer and autoimmune diseases. MAbs are glycosylated at conserved asparagine residues (N-X-S/T) of the Fc region at amino acid position 297 of the heavy chain. Glycans are important in governing the functions of efficacy and serum half-life of protein therapeutics and are part of the critical quality attribute panel for release testing. Traditionally, N-linked glycans are released from glycoproteins after denaturation and enzymatic digestion with PNGase F, followed by fluorescent labeling of the liberated glycans. The labeled glycans are then separated using hydrophilic liquid chromatography (HILIC) with fluorescence detection to generate chromatographic profile. Despite decades of use, this strenuous process remains unchanged, utilizing toxic reagents and extended sample preparation time. As an intervention, this report showcases a novel, label-free approach to detect and quantify N-glycans without using fluorescent labeling. Separation of glycans using mixed-mode PGC column along with detection of non-derivatized glycans using charged aerosol detector, the overall turnaround time can be greatly reduced with significant cost savings. The label-free method provides similar quantitative results as the conventional fluorescent labeled method, confirming the validity of the method for product release.<br />Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.<br /> (Copyright © 2022 Elsevier B.V. All rights reserved.)

Details

Language :
English
ISSN :
1873-376X
Volume :
1212
Database :
MEDLINE
Journal :
Journal of chromatography. B, Analytical technologies in the biomedical and life sciences
Publication Type :
Academic Journal
Accession number :
36279778
Full Text :
https://doi.org/10.1016/j.jchromb.2022.123502