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High-Density Antigen Microarrays for the Assessment of Antibody Selectivity and Off-Target Binding

  • Ronald SjöbergEmail author
  • Eni Andersson
  • Cecilia Hellström
  • Cecilia Mattsson
  • Jochen M. Schwenk
  • Peter Nilsson
  • Burcu Ayoglu
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1785)

Abstract

With the increasing availability of collections of antibodies, their evaluation in terms of binding selectivity becomes an important but challenging task. Planar antigen microarrays are very suitable tools to address this task and provide a powerful proteomics platform for the characterization of the binding selectivity of antibodies toward thousands of antigens in parallel. In this chapter, we describe our in-house developed procedures for the generation of high-density planar antigen microarrays with over 21,000 features. We also provide the details of the assay protocol, which we routinely use for the assessment of binding selectivity of the polyclonal antibodies generated within the Human Protein Atlas.

Key words

Affinity proteomics Protein microarrays Antigen microarrays Antibody selectivity 

Notes

Acknowledgments

We thank the entire staff of the Human Protein Atlas for producing all the antigens and antibodies utilized in this study. This work was supported by the ProNova VINN Excellence Centre for Protein Technology (VINNOVA, Swedish Governmental Agency for Innovation Systems) and by grants from the Knut and Alice Wallenberg Foundation, SciLifeLab Stockholm and the KTH Center for Applied Proteomics funded by the Erling-Persson Family Foundation. The authors declare no conflict of interest.

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Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2018

Authors and Affiliations

  • Ronald Sjöberg
    • 1
    Email author
  • Eni Andersson
    • 1
  • Cecilia Hellström
    • 1
  • Cecilia Mattsson
    • 1
  • Jochen M. Schwenk
    • 2
  • Peter Nilsson
    • 2
  • Burcu Ayoglu
    • 2
  1. 1.SciLifeLab, Division of Affinity ProteomicsKTH–Royal Institute of TechnologyStockholmSweden
  2. 2.Affinity Proteomics, SciLifeLab, School of Engineering Sciences in Chemistry, Biotechnology and HealthKTH–Royal Institute of TechnologyStockholmSweden

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