Antibody Engineering Methods and Protocols

This detailed new edition provides complete and easy access to a variety of antibody engineering techniques.  The volume explores topics such as the generation of native, synthetic, or immune antibody libraries, the selection of lead candidates via the different powerful and innovative display technologies, Fc engineering, as well as their production, characterization, and optimization of antibodies. Written for the highly successful Methods in Molecular Biology series, 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. 

Part I: In Silico Methods

 

1. Antibody Design and Humanization via In Silico Modeling

            Vinodh B. Kurella and Reddy Gali

 

2. Antibody Affinity Maturation by Computational Design

            Daisuke Kuroda and Kouhei Tsumoto

 

3. Use of IMGT® Databases and Tools for Antibody Engineering and Humanization

            Marie-Paule Lefranc, François Ehrenmann§, Sofia Kossida, Véronique Giudicelli, and Patrice Duroux

 

Part II: Generation of Diversity

 

4. Construction of Human Naïve Antibody Gene Libraries

            Michela Pasello, Alessandra Mallano, Michela Flego, Silvia Zamboni, Anna Maria Giudice, and Katia Scotlandi

 

5. Construction of Synthetic Antibody Libraries

            Déborah Caucheteur, Gautier Robin, Vincent Parez, and Pierre Martineau

 

6. Construction of Histidine-Enriched Shark IgNAR Variable Domain Antibody Libraries for the Isolation of pH-Sensitive vNAR Fragments

            Doreen Könning, Steffen Hinz, Julius Grzeschik, Christian Schröter, Simon Krah, Stefan Zielonka, and Harald Kolmar

 

7. Display Technologies for Generation of Ig Single Variable Domains

            Vladimir Bobkov, Bas van der Woning, and Hans de Haard

 

8. A Streamlined Approach for the Construction of Large Yeast Surface Display Fab Antibody Libraries

            Simon Krah, Julius Grzeschik, Simon Rosowski, Ramona Gaa, Iris Willenbuecher, Deniz Demir, Lars Toleikis, Harald Kolmar, Stefan Becker, and Stefan Zielonka

 

Part III: Selections of Lead Candidate

 

9. Phage Display and Selections on Purified Antigens

            Magali Colazet and Patrick Chames

 

10. Selection of Antibodies to Transiently Expressed Membrane Proteins Using Phage Display

            Martina L. Jones, Stephen M. Mahler, and Sumukh Kumble

 

11. Selection of Antibody Fragments against Structured DNA by Phage Display

            Mahdi Zeraati, Marcel E. Dinger, and Daniel Christ

 

12. Selection of Antibody Fragments by Yeast Display

            Nathalie Scholler

 

13. Rapid Selection of High Affinity Antibody scFv Fragments Using Ribosome Display

            Birgit Dreier and Andreas Plückthun

 

14. In Vitro Selection of Single Domain Antibody (VHH) Using cDNA Display

            Naoto Nemoto, Shigefumi Kumachi, and Hidenao Arai

 

15. Sequencing and Affinity Determination of Antigen-Specific B Lymphocytes from Peripheral Blood

            Peter Schofield, Rodrigo Vazquez-Lombardi, Mahmoud Abdelatti, Damien Nevoltris, Christopher C. Goodnow, Daniel Christ, and Joanne H. Reed

 

Part IV: Production of Recombinant Antibodies

 

16. Expression of IgG Monoclonals with Engineered Immune Effector Functions

            Rodrigo Vazquez-Lombardi, Damien Nevoltris, Romain Rouet, and Daniel Christ

 

17. An IRES-Mediated Tricistronic Vector for Efficient Generation of Stable, High Level Monoclonal Antibody Producing CHO DG44 Cell Lines

            Jessna H.M. Yeo, Mariati, and Yuansheng Yang

 

18. Production, Purification, and Characterization of Antibody-TNF Superfamily Ligand Fusion Proteins

            Martin Siegemund, Nadine Beha, and Dafne Müller

 

Part V: Fc Engineering

 

19. Chemoenzymatic Defucosylation of Therapeutic Antibodies for Enhanced Effector Functions Using Bacterial a-Fucosidases

            Chao Li, Tiezheng Li, and Lai-Xi Wang

 

20. Fc Glyco- and Fc Protein-Engineering: Design of Antibody Variants with Improved ADCC and CDC Activity

            Christian Kellner, Stefanie Derer, Katja Klausz, Sophia Rosskopf, Tim Wirt, Thies Rösner, Anna Otte, Elisa Cappuzzello, and Matthias Peipp

 

21. Fc Engineering: Tailored Synthetic Human IgG1-Fc Repertoire for High Affinity Interaction with FcRn at pH 6.0

            Abhishek Saxena, Bingxin Bai, Shin-Chen Hou, Lianlian Jiang, Tianlei Ying, Shane Miersch, Sachdev S. Sidhu, and Donghui Wu

 

Part VI: Characterization, Optimization, and Innovative Formats

 

22. Measuring Antibody-Antigen Binding Kinetics Using Surface Plasmon Resonance

            Stephen Hearty, Paul Leonard, Hui Ma, and Richard O’Kennedy

 

23. Parallel Evolution of Antibody Affinity and Thermal Stability for Optimal Biotherapeutic Development

            Edward Franklin, Orla Cunningham, and Brian Fennell

 

24. The Use of Somatic Hypermutation for the Affinity Maturation of Therapeutic Antibodies

            Peter M. Bowers, William J. Boyle, and Robert Damoiseaux

 

25. Selection and Use of Intracellular Antibodies

            Sandrine Moutel, Clément Nizak, and Franck Perez

 

26. Site-Specific Radioactive Labeling of Nanobodies

            Maxine Crauwels, Sam Massa, Charlotte Martin, Cecilia Betti, Steven Ballet, Nick Devoogdt, Catarina Xavier, and Serge Muyldermans