Impact of Lectin biotinylation for surface plasmon resonance and enzyme-linked Lectin assays for protein glycosylation.

Autor: Serafin B; Department of Chemical Engineering, Polytechnique Montreal, Montreal, QC, Canada., Kamen A; Department of Bioengineering, McGill University, Montreal, QC, Canada., De Crescenzo G; Department of Chemical Engineering, Polytechnique Montreal, Montreal, QC, Canada. Electronic address: gregory.decrescenzo@polymtl.ca., Henry O; Department of Chemical Engineering, Polytechnique Montreal, Montreal, QC, Canada. Electronic address: olivier.henry@polymtl.ca.
Jazyk: angličtina
Zdroj: Analytical biochemistry [Anal Biochem] 2025 Jan; Vol. 696, pp. 115693. Date of Electronic Publication: 2024 Oct 19.
DOI: 10.1016/j.ab.2024.115693
Abstrakt: Lectins are widely employed for the assessment of protein glycosylation as their carbohydrate binding specificities have been well characterized. In glycosylation assays, lectins are often conjugated with biotin tags, which interact with streptavidin to functionalize biosensing surfaces or recruit signal generating molecules, depending on the assay configuration. We here demonstrate that a high degree of biotin conjugation can limit total capture to streptavidin functionalized SPR surfaces due to multipoint binding, and can additionally bias the reported kinetic evaluations when measuring the interaction between lectins and glycoproteins by SPR. For microplate assays using different configurations, high biotinylation ratios can effectively amplify the signal obtained when using Streptavidin conjugates for detection, in some cases significantly lowering the limit of detection. The cumulative results express the importance of customizing the ligand biotinylation ratios for different assay configurations, as commercially obtained pre-biotinylated lectins are not necessarily optimized for different assay configurations.
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.
(Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)
Databáze: MEDLINE
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