Application of lectin immobilized on polyHIPE monoliths for bioprocess monitoring of glycosylated proteins.

Autor: Stantič M; Faculty for Chemistry and Chemical Technology, University of Ljubljana, Večna pot 113, 1000 Ljubljana, Slovenia., Gunčar G; Faculty for Chemistry and Chemical Technology, University of Ljubljana, Večna pot 113, 1000 Ljubljana, Slovenia., Kuzman D; Technical development biosimilars, Global drug development, Novartis, Kolodvorska 27, 1234 Mengeš, Slovenia., Mravljak R; Faculty for Chemistry and Chemical Technology, University of Ljubljana, Večna pot 113, 1000 Ljubljana, Slovenia., Cvijić T; Technical development biosimilars, Global drug development, Novartis, Kolodvorska 27, 1234 Mengeš, Slovenia., Podgornik A; Faculty for Chemistry and Chemical Technology, University of Ljubljana, Večna pot 113, 1000 Ljubljana, Slovenia; COBIK, Tovarniška 26, 5270 Ajdovščina, Slovenia. Electronic address: ales.podgornik@fkkt.uni-lj.si.
Jazyk: angličtina
Zdroj: Journal of chromatography. B, Analytical technologies in the biomedical and life sciences [J Chromatogr B Analyt Technol Biomed Life Sci] 2021 Jun 01; Vol. 1174, pp. 122731. Date of Electronic Publication: 2021 Apr 25.
DOI: 10.1016/j.jchromb.2021.122731
Abstrakt: In-process monitoring of glycosylated protein concentration becomes very important with the introduction of perfusion bioprocesses. Affinity chromatography based on lectins allows selective monitoring when carbohydrates are accessible on the protein surface. In this work, we immobilized lectin on polyHIPE type of monoliths and implemented it for bioprocess monitoring. A spacer was introduced to lectin, which increased binding kinetics toward Fc-fusion protein, demonstrated by bio-layer interferometry. Furthermore, complete desorption using 0.25 M galactose was shown. Affinity column exhibited linearity in the range between 0.5 and 8 mg/ml and flow-unaffected binding for the flow-rates between 0.5 and 8 ml/min. Long-term stability over at least four months period was demonstrated. No unspecific binding of culture media components, including host cell proteins and DNA, was detected. Results obtained by affinity column matched concentration values obtained by a reference method.
(Copyright © 2021 Elsevier B.V. All rights reserved.)
Databáze: MEDLINE
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