title: Electrochemical biosensors featuring oriented antibody immobilization via electrografted and self-assembled hydrazide chemistry.
abstract: Appropriate site-directed chemistry is essential to maximize the performance of immunosensors. We present two new functionalization strategies that preserve proper folding and binding potential of antibodies by forcing their oriented immobilization. Both strategies are based on the formation of hydrazone bonds between aldehyde groups on the Fc moieties of periodate-oxidized antibodies and hydrazide groups on functionalized gold electrodes. Those hydrazide groups are introduced by electrografting of diazonium salts or by self assembly of mono- and dithiolated hydrazide linkers, resulting in films with tailored functional groups and, thus, antibody distribution and spacing. Their barrier properties and permeability toward electroactive species are evaluated. To demonstrate the potential of these new functionalization strategies, detection of bacteriophage MS2 is performed through either a direct assay using electrochemical impedance spectroscopy (EIS) or through a sandwich assay using differential pulse voltammetry (DPV). Diazonium and monothiolated self-assembled monolayer-modified electrodes enable the detection of less than 1 plaque forming unit (pfu)/mL in a direct EIS assay. However, nonspecific adsorption renders measurements in river water samples difficult. In contrast, sandwich-assays on electrodes with electrografted diazonium salts and monothiolated self-assembled monolayers do not show significant matrix effects using river water samples, but the limits of detection are 10(8) times higher than those of the direct assay. Best results are achieved for immunosensors based on mixed monolayers of hydrazide and hydroxyl diothiolated linkers (15 pfu/mL). These new functionalization techniques are facile to implement. They afford the possibility to tune the surface composition and tailor the electrochemical properties of electrochemical sensors. These advantages should translate into broad interest in this type of surface chemistry for biosensor development.
authors.affiliation: Mawson Institute, University of South Australia , Mawson Lakes, South Australia 5001, Australia.
authors.forename: Beatriz
authors.initials: B
authors.lastname: Prieto-Simón
authors.forename: Christopher
authors.initials: C
authors.lastname: Saint
authors.forename: Nicolas H
authors.initials: NH
authors.lastname: Voelcker
mesh_terms: D055910:Antibodies, Immobilized / Q000737:chemistry* / Q000276:immunology; D015374:Biosensing Techniques / Q000379:methods*; D003979:Diazonium Compounds / Q000737:chemistry; D004563:Electrochemistry; D006046:Gold / Q000737:chemistry; D006834:Hydrazines / Q000737:chemistry; D007118:Immunoassay; D017909:Levivirus / Q000302:isolation & purification
chemical_list: D055910:Antibodies, Immobilized; D003979:Diazonium Compounds; D006834:Hydrazines; C029424:hydrazine; D006046:Gold