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Highly Specific Protein Identification by Immunoprecipitation-Mass Spectrometry Using Antifouling Microbeads.


ABSTRACT: A common method to study protein complexes is immunoprecipitation (IP), followed by mass spectrometry (thus labeled: IP-MS). IP-MS has been shown to be a powerful tool to identify protein-protein interactions. It is, however, often challenging to discriminate true protein interactors from contaminating ones. Here, we describe the preparation of antifouling azide-functionalized polymer-coated beads that can be equipped with an antibody of choice via click chemistry. We show the preparation of generic immunoprecipitation beads that target the green fluorescent protein (GFP) and show how they can be used in IP-MS experiments targeting two different GFP-fusion proteins. Our antifouling beads were able to efficiently identify relevant protein-protein interactions but with a strong reduction in unwanted nonspecific protein binding compared to commercial anti-GFP beads.

SUBMITTER: van Andel E 

PROVIDER: S-EPMC9136845 | biostudies-literature | 2022 May

REPOSITORIES: biostudies-literature

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Highly Specific Protein Identification by Immunoprecipitation-Mass Spectrometry Using Antifouling Microbeads.

van Andel Esther E   Roosjen Mark M   van der Zanden Stef S   Lange Stefanie C SC   Weijers Dolf D   Smulders Maarten M J MMJ   Savelkoul Huub F J HFJ   Zuilhof Han H   Tijhaar Edwin J EJ  

ACS applied materials & interfaces 20220510 20


A common method to study protein complexes is immunoprecipitation (IP), followed by mass spectrometry (thus labeled: IP-MS). IP-MS has been shown to be a powerful tool to identify protein-protein interactions. It is, however, often challenging to discriminate true protein interactors from contaminating ones. Here, we describe the preparation of antifouling azide-functionalized polymer-coated beads that can be equipped with an antibody of choice via click chemistry. We show the preparation of gen  ...[more]

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