Epitope Mapping Protocols

This third edition volume expands on the previous editions with more detailed research on the characterization of antibody antigen interactions between different users with different requirements. The chapters in this book are divided into four parts: Part One looks at the entire native antigen and covers traditional structural biology techniques such as nuclear magnetic resonance and x-ray crystallography. Part Two talks about protein fragments derived from antigens, and discusses binding regions within antigen sequence using bacterial surface display and ELISA, for example. Part Three describes the use of surface plasmon resonance spectroscopy and biolayer interferometry, and Part Four highlights methods used to identify new antigens and assess antibody cross-reactivity. Written in the highly successful Methods in Molecular Biology series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls.

Thorough and cutting-edge, Epitope Mapping Protocols, Third Edition is a valuable resource for anyone interested in furthering their research in this expanding field.

Preface...

Table of Contents...

Contributing Authors...

1. An Introduction to Epitope Mapping

Johan Nilvebrant and Johan Rockberg

Part I Whole Antigen Methods

2. Epitope Mapping of Antibody-Antigen Interactions with X-Ray Crystallography

Moeko Toride and Cory L. Brooks

3. Mapping Antibody Epitopes by Solution NMR Spectroscopy: Practical Considerations

Luca Simonelli, Mattia Pedotti, Marco Bardelli, Simon Jurt, Oliver Zerbe, and Luca Varani

4. Label-Free Cell-Based Assay for Characterization of Biomolecules and Receptors

Diluka Peiris and Lars M. Jørgensen 

5. Probing Antibody Binding Sites on G Protein-Coupled Receptors Using Genetically-Encoded Photo-Activatable Cross-Linkers

Thomas Huber and Thomas P. Sakmar

6. Rapid Mapping of Protein Binding Sites and Conformational Epitopes by Coupling Yeast Surface Display to Chemical Labeling and Deep Sequencing

Tariq Ahmad Najar, Shruti Khare, and Raghavan Varadarajan

7. Epitope Mapping Using Yeast Display and Next Generation Sequencing

Thomas Van Blarcom, Andrea Rossi, Davide Foletti, Purnima Sundar, Steven Pitts, Zea Melton, Dilduz Telman, Lora Zhao, Wai Ling Cheung, Jan Berka, Wenwu Zhai, Pavel Strop, Jaume Pons, Arvind Rajpal, and Javier Chaparro-Riggers

Part II Protein Domain and Fragment-Based Methods

8. B-Cell Epitope Mapping Using a Library of Overlapping Synthetic Peptides in an Enzyme-Linked Immunosorbent Assay

Thiru Vanniasinkam, Mary D. Barton, Tongted Phumoonna Das, and Michael W. Heuzenroeder 

9. Array-In-Well Epitope Mapping of Phage-Displayed Antibodies

Urpo Lamminmäki, Gaurav Batra, and Petri Saviranta

10. Epitope Mapping of Antibodies Using Bacterial Cell Surface Display of Gene Fragment Libraries

Anna-Luisa Volk, Francis Jingxin Hu, and Johan Rockberg

11. Discontinuous Epitope Mapping of Antibodies Using Bacterial Cell Surface Display of Folded Domains

Anna-Luisa Volk and Johan Rockberg

Part III Real-Time Based Methods

12. Kinetic Analysis and Epitope Binning Using Surface Plasmon Resonance

Johan Nilvebrant

13. Epitope Binning of Human Monoclonal Antibodies in Classical Sandwich and In-Tandem Orientation Using the Octet System Based on Biolayer Interferometry

Lisette Bronswijk-Deddens

Part IV Antibody Specificity and T-Cell Prediction

14. Antibody Specificity Profiling Using Protein Microarrays

Pedro Ramos-López, José Irizarry, Ignacio Pino, and Seth Blackshaw

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

Ronald Sjöberg, Eni Andersson, Cecilia Hellström, Cecilia Mattsson, Jochen M. Schwenk, Peter Nilsson, and Burcu Ayoglu

16. Multiplexed Antigen Bead Arrays for Assessment of Antibody Selectivity and Epitope Mapping

Burcu Ayoglu, Peter Nilsson, and Jochen M. Schwenk

John T. Ballew, Jack R. Reifert, and Patrick S. Daugherty

18. Bioinformatics Tools for the Prediction of T Cell Epitopes

Massimo Andreatta and Morten Nielsen