Abstract
Recombinant Nanobodies, derived from camelid heavy-chain antibodies, are a unique form of monoclonal antibodies composed of a single antigen-binding domain. Nanobodies are the smallest intact antigen-binding entities that can be derived from an in vivo affinity-matured antibody. They are small (15 kDa, 2.2 nm diameter, 4 nm height), soluble, stable, robust, and easy to produce in various host cells. Nanobodies bind their cognate antigens with nanomolar affinities and often interact with epitopes that are less accessible to/or less immunogenic for classical antibodies. Nanobodies are easily tailored for various applications. We have developed a robust technology to identify antigen-specific Nanobodies. Here, we describe, in detail, the protocols for the generation of Nanobody phage display libraries and for the isolation and characterization of antigen-specific Nanobodies from such libraries.
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References
Alvarez-Reuda N, Behar G, Ferré V, Pugnière M, Roquet F, Gastinel L, Jacquot C, Aubry J, Baty D, Barbet J, Birklè S (2007) Generation of llama single-domain antibodies against methotrexate, a prototypical hapten. Mol Immunol 44:1680–1690
Ghahroudi MA, Desmyter A, Wyns L, Hamers R, Muyldermans S (1997) Selection and identification of single domain antibody fragments from camel heavy-chain antibodies. FEBS Lett 414:521–526
Hamers-Casterman C, Atarhouch T, Muyldermans S, Robinson G, Hamers C, Bajyana Songa E, Bendahman N, Hamers R (1993) Naturally occurring antibodies devoid of light chains. Nature 363:446–448
Koch-Nolte F, Reyelt J, Schöβow B, Schwarz N, Scheuplein F, Rothenburg S, Haag F, Alzogaray V, Cauerhff A, Goldbaum FA (2007) Single domain antibodies from llama effectively and specifically block T cell ecto-ADP-ribosyltransferase ART2.2 in vivo. FASEB J 21:3490–3498
Ladenson RC, Crimmins DL, Landt Y, Ladenson JH (2006) Isolation and characterization of a thermally stable recombinant anti-caffeine heavy-chain antibody fragment. Anal Chem 78:4501–4508
Lauwereys M, Ghahroudi MA, Desmyter A, Kinne J, Hölzer W, De Genst E, Wyns L, Muyldermans S (1998) Potent enzyme inhibitors derived from dromedary heavy-chain antibodies. EMBO J 17:3512–3520
Maass DR, Sepulveda J, Pernthaner A, Shoemaker CB (2007) Alpaca (Lama pacos) as a convenient source of recombinant camelid heavy chain antibodies (VHHs). J Immunol Methods 324:13–25
Nguyen VK, Muyldermans S, Hamers R (1998) The specific variable domain of camel heavy-chain antibodies is encoded in the germline. J Mol Biol 257:413–418
Nguyen VK, Hamers R, Wyns L, Muyldermans S (2000) Camel heavy-chain antibodies: diverse germline VHH and specific mechanisms enlarge the antigen-binding repertoire. EMBO J 19:921–931
Nguyen VK, Desmyter A, Muyldermans S (2001) Functional Heavy-Chain antibodies in camelidae. Adv Immunol 79:261–296
Sambrook J, Fritsch EF, Maniatis T (1989) Molecular Cloning: A Laboratory Manual, 2nd edn. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY
Van der Linden R, de Geus B, Stok W, Bos W, van Wassenaar D, Verrips T, Frenken L (2000) Induction of immune responses and molecular cloning of the heavy chain antibody repertoire of Lama glama. J Immunol Meth 240:185–195
Vu KB, Ghahroudi MA, Wyns L, Muyldermans S (1997) Comparison of llama VH sequences from conventional and heavy chain antibodies. Mol Immunol 34:1121–1131
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Ghassabeh, G.H., Saerens, D., Muyldermans, S. (2010). Isolation of Antigen-Specific Nanobodies. In: Kontermann, R., Dübel, S. (eds) Antibody Engineering. Springer Protocols Handbooks. Springer, Berlin, Heidelberg. https://doi.org/10.1007/978-3-642-01147-4_20
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DOI: https://doi.org/10.1007/978-3-642-01147-4_20
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