2. Academic Validation
  3. Highly Specific Protein Identification by Immunoprecipitation-Mass Spectrometry Using Antifouling Microbeads

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

  • ACS Appl Mater Interfaces. 2022 May 10;14(20):23102-23116. doi: 10.1021/acsami.1c22734.
Esther van Andel 1 2,  Mark Roosjen 3,  Stef van der Zanden 2,  Stefanie C Lange 1,  Dolf Weijers 3,  Maarten M J Smulders 1,  Huub F J Savelkoul 2,  Han Zuilhof 1 4 5,  Edwin J Tijhaar 2
Affiliations

Affiliations

  • 1 Laboratory of Organic Chemistry, Wageningen University, Stippeneng 4, 6708 WE Wageningen, The Netherlands.
  • 2 Cell Biology and Immunology group, Wageningen University, De Elst 1, 6709 PG Wageningen, The Netherlands.
  • 3 Laboratory of Biochemistry, Wageningen University, Stippeneng 4, 6708 WE Wageningen, The Netherlands.
  • 4 School of Pharmaceutical Sciences and Technology, Tianjin University, 92 Weijin Road, Tianjin 300072, People's Republic of China.
  • 5 Department of Chemical and Materials Engineering, Faculty of Engineering, King Abdulaziz University, 21589 Jeddah, Saudi Arabia.
PMID: 35536557 DOI: 10.1021/acsami.1c22734
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.

Keywords

antibody functionalization; antifouling; click chemistry; immunoprecipitation; mass spectrometry; microbeads; proteomics; zwitterionic polymer brushes.

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