Semi-Automated Lectin Magnetic Bead Array (LeMBA) for Translational Serum Glycoprotein Biomarker Discovery and Validation.
Glycoproteomics
Glycosylation
Lectin
Liquid handler
Serum biomarkers
Journal
Methods in molecular biology (Clifton, N.J.)
ISSN: 1940-6029
Titre abrégé: Methods Mol Biol
Pays: United States
ID NLM: 9214969
Informations de publication
Date de publication:
2023
2023
Historique:
entrez:
13
2
2023
pubmed:
14
2
2023
medline:
16
2
2023
Statut:
ppublish
Résumé
Aberrant protein glycosylation is a characteristic of diverse diseases which has been explored as biomarkers. To support translational serum glycoprotein biomarker discovery and validation, we developed a semi-automated workflow using individual lectin-coupled magnetic beads to conduct lectin pulldowns in a high-throughput format. Lectins are naturally occurring glycoprotein binding proteins widely used in glycobiology. While lectin-affinity isolation has been coupled to mass spectrometry-based proteomics, the lectin magnetic bead array (LeMBA) platform allows technically robust screening and measurement of clinical cohorts. This chapter describes detailed lectin-magnetic bead coupling, serum denaturation, lectin magnetic bead pulldown, and on-bead trypsin digest. The resulting tryptic peptides are analyzed by untargeted or targeted liquid chromatography-mass spectrometry (LC-MS), for biomarker discovery, or qualification/validation, respectively. LeMBA-MS generates quantitative data for glycoforms based on lectin affinity of the glycoprotein coupled with MS measurement of one or more prototypic peptides and has successfully been used to discover and validate novel serum cancer glycoprotein biomarkers. This chapter includes detailed protocols for two different liquid handlers, along with recommendations on quality control measures for clinical biomarker studies.
Identifiants
pubmed: 36781799
doi: 10.1007/978-1-0716-2978-9_25
doi:
Substances chimiques
Lectins
0
Glycoproteins
0
Biomarkers, Tumor
0
Peptides
0
Types de publication
Journal Article
Langues
eng
Sous-ensembles de citation
IM
Pagination
395-411Informations de copyright
© 2023. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.
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