Enhanced electrochemical activity of redox-labels in multi-layered protein films on indium tin oxide nanoparticle-based electrode

Abstract

a b s t r a c tFacile electrical communication between redox-active labeling molecules and electrode is essential in the electrochemical detection of bio-affinity reactions. In this report, nanometer-sized indium tin oxide (ITO) particles were employed in the fabrication of porous thick film electrodes to enhance the otherwise impeded electrochemical activity of redox labels in multi-layered protein films, and to enable quantitative detection of avidin/biotin binding interaction. To carry out the affinity reaction, avidin immobilized on an ITO electrode was reacted with mouse IgG labeled with both biotin and ruthenium Tris-(2,2 -bipyridine) (Ru-bipy). The binding reaction between avidin and biotin was detected by the catalytic voltammetry of Ru-bipy in an oxalate-containing electrolyte. On sputtered ITO thin film electrode, although a single layer of Ru-bipy labeled avidin exhibited substantial anodic current, attaching the label to the outer IgG layer of the avidin/biotin-IgG binding pair resulted in almost complete loss of the signal. However, electrochemical current was recovered on ITO film electrodes prepared from nanometer-sized particles. The surface of the nanoparticle structured electrode was found by scanning electron microscopy to be very porous, and had twice as much surface binding capacity for avidin as the sputtered electrode. The results were rationalized by the assumption of different packing density of avidin inner layer on the two surfaces, and consequently different electron transfer distance between the electrode and Ru-bipy on the IgG outer layer. A linear relationship between electrochemical current and IgG concentration was obtained in the range of 40-4000 nmol L −1 on the nanoparticle-based electrode. The approach can be employed in the electrochemical detection of immunoassays using non-enzymatic redox labels.

Highly ordered transparent mesoporous TiO2 thin films: an attractive matrix for efficient immobilization and spectroelectrochemical characterization of cytochrome c

“…These efforts have been motivated by the potential for developing unique spectroelectrochemical strategies for the characterization of redox proteins, 1,2 as well as by the possibilities of developing new electrochemical biosensors. 3,4 These novel protein host matrixes combine the attractive properties of transparency, high surface area, electrical semiconductivity, biocompatibility, ease of fabrication and high chemical, mechanical and thermal stability, with the possibility of direct electron transfer between the protein redox sites and the host mesoporous metal oxide.…”

“…Attention to date has been largely focused on the electrochemical investigations of heme proteins immobilized within semiconductive mesoporous metal oxide films (M x O y , with M = Ti, Sn, Zn) produced from randomly sintered nanosized particles deposited on an electrode surface (film thickness of a few micrometers). [1][2][3][4][5][6] The structure and porosity of these films is however relatively ill-defined since they are constituted by irregular aggregates of interconnected metal oxide nanoparticles (possibly with a binder) and, in the case of electrochemical applications, such heterogeneity can significantly affect and complicate mass transport within the film. 7,8 Moreover, despite a noteworthy increase of the electrical conductivity at potentials above the conduction band edge, the electron transport through such metal oxide nanoparticulate films was shown significantly to be altered by poor interparticle electron transfer, 9 leading thus to an electrical conductivity much lower than at compact polycrystalline or monocrystalline materials.…”

High catalytic activity of indium tin oxide nanoparticle modified electrode towards electro-oxidation of ascorbic acid

“…ITO nanoparticle-modified electrodes were fabricated on sputtered ITO glass from a commercially available colloidal solution of dispersed nanoparticles by following the procedure described previously [24,25]. As shown in the scanning electron micrographs (Fig.…”

Development of redox-labeled electrochemical immunoassay for polycyclic aromatic hydrocarbons with controlled surface modification and catalytic voltammetric detection

“…Obviously, the detection limit obtained in our work is not sufficient for most practical applications. To improve the sensitivity, study is currently underway which employs nano-porous ITO film electrodes to enhance electron transfer kinetics of the redox label (Yang and Guo, 2009). It is hoped that the new electrochemical assay method can be used in the simultaneous detection of multiple analytes on antibody biochips.…”