title: Single-domain antibodies and their utility.
abstract: Engineered monoclonal antibody fragments have gained market attention due to their versatility and tailor-made potential and are now considered to be an important part of future immunobiotherapeutics. Single-domain antibodies (sdAbs), also known as nanobodies, are derived from VHHs [variable domains (V) of heavy-chain-only antibodies (HCAb)] of camelid heavy-chain antibodies. These nature-made sdAbs are well suited for various applications due to their favorable characteristics such as small size, ease of genetic manipulation, high affinity and solubility, overall stability, resistance to harsh conditions (e.g., low pH, high temperature), and low immunogenicity. Most importantly, sdAbs have the feature of penetrating into cavities and recognizing hidden epitopes normally inaccessible to conventional antibodies, mainly due to their protruding CDR3/H3 loops. In this unit, we will present and discuss comprehensive and step-by-step protocols routinely practiced in our laboratory for isolating sdAbs from immunized llamas (or other members of the Camelidae family) against target antigens using phage-display technology. Expression, purification, and characterization of the isolated sdAbs will then be described, followed by presentation of several examples of applications of sdAbs previously characterized in our laboratory and elsewhere.
authors: Human Health Therapeutics, Life Sciences Division, National Research Council Canada, Ottawa, Ontario, Canada.
authors: Toya Nath
authors: TN
authors: Baral
authors: Human Health Therapeutics, Life Sciences Division, National Research Council Canada, Ottawa, Ontario, Canada.|University of Guelph, Guelph, Ontario, Canada.
authors: Roger
authors: R
authors: MacKenzie
authors: Human Health Therapeutics, Life Sciences Division, National Research Council Canada, Ottawa, Ontario, Canada.|University of Guelph, Guelph, Ontario, Canada.|Department of Biology, Carleton University, Ottawa, Ontario, Canada.
authors: Mehdi
authors: M
authors: Arbabi Ghahroudi
mesh_terms: D000818:Animals; D001419:Bacteria / Q000276:immunology; D015153:Blotting, Western; D002161:Camelids, New World / Q000276:immunology; D002850:Chromatography, Gel; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D018604:Epitope Mapping; D015723:Gene Library; D006801:Humans; D007114:Immunization; D061905:Single-Domain Antibodies / Q000276:immunology*; D020349:Surface Plasmon Resonance
chemical_list: D061905:Single-Domain Antibodies
keywords: VHH; antibody fragment libraries dimeric VHH; heavy-chain antibodies; multivalency; pentabodies; single-domain antibody