Affimers as an alternative to antibodies for protein biomarker enrichment
Publication year
2020Source
Protein Expression and Purification, 174, (2020), article 105677ISSN
Publication type
Article / Letter to editor

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Organization
Laboratory Medicine
Internal Medicine
Paediatrics
Analytical Chemistry
Journal title
Protein Expression and Purification
Volume
vol. 174
Subject
Radboudumc 0: Other Research RIMLS: Radboud Institute for Molecular Life Sciences; Radboudumc 6: Metabolic Disorders RIMLS: Radboud Institute for Molecular Life SciencesAbstract
INTRODUCTION: Assessing the specificity of protein binders is an essential first step in protein biomarker assay development. Affimers are novel protein binders and can potentially replace antibodies in multiple protein capture-based assays. Affimers are selected for their high specificity against the target protein and have benefits over antibodies like batch-to-batch reproducibility and are stable across a wide range of chemical conditions. Here we mimicked a typical initial screening of affimers and commercially available monoclonal antibodies against two non-related proteins, IL-37b and proinsulin, to assess the potential of affimers as alternative to antibodies. METHODS: Binding specificity of anti-IL-37b and anti-proinsulin affimers and antibodies was investigated via magnetic bead-based capture of their recombinant protein targets in human plasma. Captured proteins were analyzed using SDS-PAGE, Coomassie blue staining, Western blotting and LC-MS/MS-based proteomics. RESULTS: All affimers and antibodies were able to bind their target protein in human plasma. Gel and LC-MS/MS analysis showed that both affimer and antibody-based captures resulted in co-purified background proteins. However, affimer-based captures showed the highest relative enrichment of IL-37b and proinsulin. CONCLUSIONS: For both proteins tested, affimers show higher specificity in purifying their target proteins from human plasma compared to monoclonal antibodies. These results indicate that affimers are promising antibody-replacement tools for protein biomarker assay development.
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- Academic publications [204024]
- Electronic publications [102375]
- Faculty of Medical Sciences [80459]
- Faculty of Science [32153]
- Open Access publications [71013]
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