Antigen identification for cell-binding antibodies using ligand-directed crosslinking and biotin transfer

DOI	Trouver le DOI : https://doi.org/10.1007/978-1-4939-9597-4_10
Auteur	Rechercher : Tremblay, T.-L.¹; Rechercher : Hill, J. J.¹
Affiliation	Conseil national de recherches Canada. Thérapeutique en santé humaine
Format	Texte, Article
Sujet	receptor identification; crosslinking; proteomics; mass spectrometry; antibodies
Résumé	Panning approaches using antibody libraries often result in the isolation of antibodies that bind to cells through an unknown cellular receptor. Here, we describe a protocol that uses ligand-directed crosslinking with the aminooxy-sulfhydryl-biotin (ASB) trifunctional crosslinker followed by a proteomic analysis to identify the cellular receptors for orphan ligands. We describe the synthesis of the ASB crosslinker, labelling of the ligand with ASB, and cell binding of the labelled ligands. Next, biotin affinity purification and trypsin digestion of cell surface proteins that have been crosslinked by ASB are described. Lastly, several hints and tips to improve the proteomic analysis for these types of samples are provided.
Date de publication	2019-07-31
Maison d’édition	Springer
Dans	Immunoproteomics : 167–180.
Série	Methods in Molecular Biology 2024.
Langue	anglais
Publications évaluées par des pairs	Oui
Exporter la notice	Exporter en format RIS
Signaler une correction	Signaler une correction (s'ouvre dans un nouvel onglet)
Identificateur de l’enregistrement	312a76cc-7eb7-4a7e-953c-49f404b1847e
Enregistrement créé	2021-01-22
Enregistrement modifié	2021-01-22

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