title: Oriented conjugation of single-domain antibodies and quantum dots.
abstract: Nanoparticle-based biodetection routinely employs monoclonal antibodies (mAbs) for targeting. However, the large size of mAbs limits the number of ligands per nanoparticle and severely restricts the bioavailability and distribution of these probes in a sample. Furthermore, conventional conjugation techniques provide nanoprobes with irregular orientation of mAbs on the nanoparticle surface and often provoke mAb unfolding. Here, we describe a protocol for engineering a new generation of ultrasmall diagnostic nanoprobes through oriented conjugation of semiconductor quantum dots (QDs) with 13 kDa single-domain antibodies (sdAbs) derived from llama immunoglobulin G (IgG). The sdAbs are conjugated with QDs in a highly oriented manner via an additional cysteine residue specifically integrated into the sdAb C-terminus. The resultant nanoprobes are <12 nm in diameter, ten times smaller in volume compared to the known alternatives. They have been proved highly efficient in flow cytometry and immunuhistochemical diagnostics. This approach can be easily extended to other semiconductor and plasmonic nanoparticles.
authors.affiliation: Laboratory of Nano-Bioengineering, National Research Nuclear University MEPhI "Moscow Engineering Physics Institute", 31 Kashirskoe sh., 115409, Moscow, Russian Federation.
authors.forename: Kristina
authors.initials: K
authors.lastname: Brazhnik
authors.forename: Igor
authors.initials: I
authors.lastname: Nabiev
authors.forename: Alyona
authors.initials: A
authors.lastname: Sukhanova
mesh_terms: D000818:Animals; D002161:Camelids, New World; D005434:Flow Cytometry; D006801:Humans; D006878:Hydroxides / Q000737:chemistry; D018796:Immunoconjugates / Q000737:chemistry*; D007074:Immunoglobulin G / Q000737:chemistry; D007150:Immunohistochemistry; D008958:Models, Molecular; D008968:Molecular Conformation; D011092:Polyethylene Glycols / Q000737:chemistry; D011786:Quality Control; D045663:Quantum Dots / Q000737:chemistry*; D061905:Single-Domain Antibodies / Q000737:chemistry*; D012995:Solubility; D014867:Water / Q000737:chemistry
chemical_list: D006878:Hydroxides; D018796:Immunoconjugates; D007074:Immunoglobulin G; D061905:Single-Domain Antibodies; D014867:Water; D011092:Polyethylene Glycols; C031356:hydroxide ion