title: Single domain antibodies as a powerful tool for high quality surface plasmon resonance studies.
abstract: Single domain antibodies are recombinantly expressed functional antibodies devoid of light chains. These binding elements are derived from heavy chain antibodies found in camelids and offer several distinctive properties for applications in biotechnology such as small size, stability, solubility, and expression in high yields. In this study we demonstrated the potential of using single domain antibodies as capturing molecules in biosensing applications. Single domain antibodies raised against green fluorescent protein were anchored onto biosensor surfaces by using several immobilization strategies based on Ni2+:nitrilotriacetic acid-polyhistidine tag, antibody-antigen, biotin-streptavidin interactions and amine-coupling chemistry. The interaction with the specific target of the single domain antibodies was characterized by surface plasmon resonance. The immobilized single domain antibodies show high affinities for their antigens with KD = 3-6 nM and outperform other antibody partners as capturing molecules facilitating also the data analysis. Furthermore they offer high resistance and stability to a wide range of denaturing agents. These unique biophysical properties and the production of novel single domain antibodies against affinity tags make them particularly attractive for use in biosensing and diagnostic assays.
authors.affiliation: Department of Chemistry & Nano-Science Center, University of Copenhagen, Copenhagen, Denmark.
authors.forename: Eduardo Antonio
authors.initials: EA
authors.lastname: Della Pia
authors.affiliation: Department of Chemistry & Nano-Science Center, University of Copenhagen, Copenhagen, Denmark.
authors.forename: Karen L
authors.initials: KL
authors.lastname: Martinez
mesh_terms: D015374:Biosensing Techniques; D015870:Gene Expression; D017930:Genes, Reporter; D011485:Protein Binding; D054730:Protein Interaction Domains and Motifs; D055550:Protein Stability; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D061905:Single-Domain Antibodies / Q000737:chemistry* / Q000378:metabolism; D020349:Surface Plasmon Resonance / Q000379:methods*
chemical_list: D011993:Recombinant Fusion Proteins; D061905:Single-Domain Antibodies
references.citation: Appl Microbiol Biotechnol. 2011 Dec;92(6):1275-86
references.pmid: 22033566
references.citation: Expert Rev Mol Diagn. 2010 Sep;10(6):777-85
references.pmid: 20843201
references.citation: Nat Protoc. 2014 Mar;9(3):674-93
references.pmid: 24577359
references.citation: Structure. 2009 Feb 13;17(2):255-65
references.pmid: 19217396
references.citation: Nano Rev. 2012;3:
references.pmid: 22833780
references.citation: Transfus Apher Sci. 2012 Jun;46(3):343-6
references.pmid: 22475545
references.citation: ACS Nano. 2011 Jun 28;5(6):4319-28
references.pmid: 21609027
references.citation: Mol Cell Proteomics. 2008 Feb;7(2):282-9
references.pmid: 17951627
references.citation: Anal Chem. 2010 Sep 1;82(17):7202-7
references.pmid: 20687583
references.citation: Appl Environ Microbiol. 2005 Jan;71(1):442-50
references.pmid: 15640220
references.citation: Protein Sci. 2009 Mar;18(3):619-28
references.pmid: 19241371
references.citation: Biophys J. 1997 Nov;73(5):2782-90
references.pmid: 9370472
references.citation: Nat Methods. 2006 Apr;3(4):303-13
references.pmid: 16554836
references.citation: Plant J. 2009 Nov;60(4):744-54
references.pmid: 19686537
references.citation: Nature. 1993 Jun 3;363(6428):446-8
references.pmid: 8502296
references.citation: Biochim Biophys Acta. 1999 Apr 12;1431(1):37-46
references.pmid: 10209277
references.citation: Protein Sci. 2010 Dec;19(12):2389-401
references.pmid: 20945358
references.citation: ACS Nano. 2014 Feb 25;8(2):1844-53
references.pmid: 24476392
references.citation: Circ Res. 2012 Mar 30;110(7):927-37
references.pmid: 22461363
references.citation: Anal Chem. 2006 May 1;78(9):3072-9
references.pmid: 16642995
references.citation: PLoS One. 2012;7(5):e36967
references.pmid: 22606318
references.citation: Nat Methods. 2006 Nov;3(11):887-9
references.pmid: 17060912
references.citation: Gene. 1996;173(1 Spec No):33-8
references.pmid: 8707053
references.citation: Proteins. 2005 May 15;59(3):555-64
references.pmid: 15778955
references.citation: Anal Chem. 2005 Dec 1;77(23):7547-55
references.pmid: 16316161
references.citation: Vet Immunol Immunopathol. 2009 Mar 15;128(1-3):178-83
references.pmid: 19026455
references.citation: Annu Rev Biochem. 2013;82:775-97
references.pmid: 23495938
references.citation: J Am Chem Soc. 2006 Mar 29;128(12):3939-45
references.pmid: 16551101
references.citation: Protein Sci. 2002 Mar;11(3):500-15
references.pmid: 11847273
references.citation: Nat Struct Mol Biol. 2010 Jan;17(1):133-8
references.pmid: 20010839
references.citation: Anal Chem. 2012 Jun 19;84(12):5198-203
references.pmid: 22681495
references.citation: Biochemistry. 2005 Nov 8;44(44):14409-18
references.pmid: 16262241
references.citation: Biosens Bioelectron. 2005 Sep 15;21(3):483-90
references.pmid: 16076438