title: Orientation of llama antibodies strongly increases sensitivity of biosensors.
abstract: Sensitivity of biosensors depends on the orientation of bio-receptors on the sensor surface. The objective of this study was to organize bio-receptors on surfaces in a way that their analyte binding site is exposed to the analyte solution. VHH proteins recognizing foot-and-mouth disease virus (FMDV) were used for making biosensors, and azides were introduced in the VHH to function as bioorthogonal reactive groups. The importance of the orientation of bio-receptors was addressed by comparing sensors with randomly oriented VHH (with multiple exposed azide groups) to sensors with uniformly oriented VHH (with only a single azide group). A surface plasmon resonance (SPR) chip exposing cyclooctyne was reacted to azide functionalized VHH domains, using click chemistry. Comparison between randomly and uniformly oriented bio-receptors showed up to 800-fold increase in biosensor sensitivity. This technique may increase the containment of infectious diseases such as FMDV as its strongly enhanced sensitivity may facilitate early diagnostics.
authors.affiliation: Plant Research International, Wageningen, The Netherlands; Laboratory of Organic Chemistry, Wageningen University, Wageningen, The Netherlands.
authors.forename: Anke K
authors.initials: AK
authors.lastname: Trilling
authors.affiliation: Plant Research International, Wageningen, The Netherlands.
authors.forename: Thamara
authors.initials: T
authors.lastname: Hesselink
authors.affiliation: Plant Research International, Wageningen, The Netherlands.
authors.forename: Adèle
authors.initials: A
authors.lastname: van Houwelingen
authors.affiliation: Plant Research International, Wageningen, The Netherlands.
authors.forename: Jan H G
authors.initials: JH
authors.lastname: Cordewener
authors.affiliation: Plant Research International, Wageningen, The Netherlands.
authors.forename: Maarten A
authors.initials: MA
authors.lastname: Jongsma
authors.affiliation: Department of Bioorganic Chemistry, Institute for Molecules and Materials, Radboud University Nijmegen, Nijmegen, The Netherlands.
authors.forename: Sanne
authors.initials: S
authors.lastname: Schoffelen
authors.affiliation: Department of Bioorganic Chemistry, Institute for Molecules and Materials, Radboud University Nijmegen, Nijmegen, The Netherlands.
authors.forename: Jan C M
authors.initials: JC
authors.lastname: van Hest
authors.affiliation: Laboratory of Organic Chemistry, Wageningen University, Wageningen, The Netherlands; Department of Chemical and Materials Engineering, King Abdulaziz University, Jeddah, Saudi Arabia.
authors.forename: Han
authors.initials: H
authors.lastname: Zuilhof
authors.affiliation: Plant Research International, Wageningen, The Netherlands. Electronic address: jules.beekwilder@wur.nl.
authors.forename: Jules
authors.initials: J
authors.lastname: Beekwilder
mesh_terms: D000818:Animals; D000914:Antibodies, Viral / Q000276:immunology*; D015374:Biosensing Techniques / Q000295:instrumentation*; D002161:Camelids, New World / Q000276:immunology*; D004867:Equipment Design; D019544:Equipment Failure Analysis; D029544:Foot-and-Mouth Disease Virus / Q000276:immunology* / Q000302:isolation & purification*; D007118:Immunoassay / Q000295:instrumentation*; D011487:Protein Conformation; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D020349:Surface Plasmon Resonance / Q000295:instrumentation*
chemical_list: D000914:Antibodies, Viral
keywords: Foot-and-mouth disease virus; Llama heavy-chain antibodies; Orientation; Site-specific azide incorporation; Surface plasmon resonance