Single Domain Antibodies: Methods and Protocols

Part I: Overview of Single Domain Antibodies

1. From Whole Monoclonal Antibodies to Single Domain Antibodies: Think Small

Jean-Luc Teillaud

2. Introduction to Heavy Chain Antibodies and Derived Nanobodies

Ccile Vincke and Serge Muyldermans

3. Overview and Discovery of IgNARs and Generation of VNARs

Stewart D. Nuttall

Part II: Single Domain Antibody Library Construction

4. Creation of the Large and Highly Functional Synthetic Repertoire of Human VH and Vκ
Domain Antibodies

Olga Ignatovich, Laurent Jespers, Ian M, Tomlinson, and Ruud M.T. de Wildt

5. Preparation of a Nave Library of Camelid Single Domain Antibodies

Aurelien Olichon and Ario de Marco

Part III: Selection of Single Domain Antibodies

6. Selection by Phage Display of Single Domain Antibodies Specific to Antigens in Their Native Conformation

Peter Verheesen and Toon Laeremans

7. Semi-Automated Panning of Naive Camelidae Libraries and Selection of Single-Domain Antibodies Against Peptide Antigens

Jyothi Kumaran, Roger MacKenzie, and Mehdi Arbabi-Ghahroudi

8. Pichia Surface Display: A Tool for Screening Single Domain Antibodies

Kristof De Schutter and Nico Callewaert

9. Bacterial Two Hybrid: A Versatile One-Step Intracellular Selection Method

Mireille Pellis, Serge Muyldermans, and Ccile Vincke

10. Intracellular Antibody Capture (IAC) Methods for Single Domain Antibodies

Tomoyuki Tanaka and Terence H. Rabbitts

11. Selection of Functional Single Domain Antibody Fragments for Interfering with Protein-Protein Interactions Inside Cells: A "One Plasmid" Mammalian Two-Hybrid System

Tomoyuki Tanaka and Terence H. Rabbitts

12. Cell-Free Selection of Domain Antibodies by In Vitro Compartmentalization

Armin Sepp and Andrew Griffiths

13. Selection of VHHs Under Application Conditions

Edward Dolk, Theo Verrips, and Hans de Haard

14. Isolation and Characterization of Clostridium difficile Toxin-Specific Single-Domain Antibodies

Greg Hussack, Mehdi Arbabi-Ghahroudi, C. Roger MacKenzie, and Jamshid Tanha

15. Selection of VHH Antibody Fragments that Recognize Different Aβ
Depositions Using Complex Immune Libraries

Rinse Klooster, Kim S. Rutger, and Sylvre van der Maarel

Part IV: Expression of Single Domain Antibodies and Derivatives

16. Expression of Single-Domain Antibodies in Bacterial Systems

Toya Nath Baral and Mehdi Arbabi-Ghahroudi

17. Expression of VHHs in S. cerevisiae

Andrea Gorlani, Hans de Haard, and Theo Verrips

18. Stable Expression of Chimeric Heavy Chain Antibodies in CHO Cells

Vishal Agrawal, Igor Slivac, Sylvie Perret, Louis Bisson, Gilles St-Laurent, Yanal Murad, Jianbing Zhang, and Yves Durocher

19. Production of Camel-Like Antibodies in Plants

Sylvie De Buck, Vikram Virdi, Thomas De Meyer, Kirsten De Wilde, Robin Piron, Jonah Nolf, Els Van Lerberge, Annelies De Paepe, and Ann Depicker

Part V: Improvement and Applications of Single Domain Antibodies

20. Selecting and Purifying Autonomous Human Variable Heavy (VH) Domains

Raffi Tonikian and Sachdev S. Sidhu

21. Solubility and Stability Engineering of Human VH Domains

Dae Young Kim, Wen Ding, and Jamshid Tanha

22. Improvement of Proteolytic Stability through In Silico Engineering

Lucy Rutten, Hans de Haard, and Theo Verrips

23. Selection of Human VH Single Domains with Improved Biophysical Properties by Phage Display

Kip Dudgeon, Romain Rouet, Kristoffer Famm, and Daniel Christ