Nanobiosensing with Arrays and Ensembles of Nanoelectrodes

Abstract

Since the first reports dating back to the mid-1990s, ensembles and arrays of nanoelectrodes (NEEs and NEAs, respectively) have gained an important role as advanced electroanalytical tools thank to their unique characteristics which include, among others, dramatically improved signal/noise ratios, enhanced mass transport and suitability for extreme miniaturization. From the year 2000 onward, these properties have been exploited to develop electrochemical biosensors in which the surfaces of NEEs/NEAs have been functionalized with biorecognition layers using immobilization modes able to take the maximum advantage from the special morphology and composite nature of their surface. This paper presents an updated overview of this field. It consists of two parts. In the first, we discuss nanofabrication methods and the principles of functioning of NEEs/NEAs, focusing, in particular, on those features which are important for the development of highly sensitive and miniaturized biosensors. In the second part, we review literature references dealing the bioanalytical and biosensing applications of sensors based on biofunctionalized arrays/ensembles of nanoelectrodes, focusing our attention on the most recent advances, published in the last five years. The goal of this review is both to furnish fundamental knowledge to researchers starting their activity in this field and provide critical information on recent achievements which can stimulate new ideas for future developments to experienced scientists.

Keywords: array; bioelectroanalysis; biosensor; ensemble; nanoelectrode; voltammetry.

Figure 1

Schematic diagram of a nanoelectrode ensemble in a template membrane: (a) overall view; (b) cross section (redrawn and reprinted with permission from [4]).

Figure 2

Scheme illustrating three template deposition methods used which differ for the way used to contact the PC membrane (8) to the flat disk Cu electrode (7). From left to right: (a) adhesion by the pressure furnished by the melamine foam (5); (b) adhesion by using a Nafion interlayer (9) as polyelectrolytic glue; (c) as (b), but using a PC membrane pre-sputtered with the gold interlayer (10). Other components: (1) Cu counter electrode; (2) Pt counter electrode; (3) working electrode; (4) Ag/AgCl KCl sat reference electrode; (6) 0.4 M CuSO₄, 0.01 M H₂SO₄ solution (reprinted with permission from [29]).

Figure 3

Electrochemical reduction current as a function of time for the potentiostatic platin of Ni and Co in the pores of PC membrane with 80 nm of nominal diameter (reprinted with permission from [24]).

Figure 4

Schematic representation of a NEE, prepared by using a track-etched polycarbonate membrane as template: (a) track-etched golden membrane; (b) copper adhesive tape with conductive glue to connect to instrumentation; (c) aluminium adhesive foil with nonconductive glue; (d) thermoadhesive insulating tape (e.g., Monokote by Topflite). Note: the dimensions of the pores (nanofibres) are only indicative and not to scale (reprinted with permission from [52]).

Figure 5

Scheme illustrating the preparation procedure used to obtain Au nanowires decorated with hierarchically branched ZnO. (a) scheme and SEM image of the template membrane; (b) scheme and SEM image of an ensemble of gold nanowires obtained by electroless deposition; (c) scheme and SEM image of ZnO nanostructures grown electrochemically on the Au nanowires (reprinted with permission from [55]).

Figure 6

(a) Simulated concentration profiles and relevant voltammetric patterns, for microelectrode arrays representing the five main categories of possible diffusion modes (from I to V). In the scale bar, the red and blue colour represents the bulk concentration and zero concentration, respectively. The second scale bar represents a relative concentration scale for the contour lines. (b) Zone diagram of cyclic voltammetric behaviour at electrode arrays: d is the centre-to-centre distance of individual electrodes in the array (measured in units of a), V is the dimensionless scan rate, and θ is the fraction of electrochemically active area in the array (reprinted with permission from [64]).

Figure 7

SEM images of NEAs with holes 500 nm in diameter with gold electrochemically deposited inside for 0 s (a); 10 s (b); 20 s (c); and 30 s (d). Estimated recession depths: (a) 450 nm; (b) 300 nm; (c) 150 nm; (d) 0 nm (reprinted with permission from [68]).

Figure 8

CVs recorded at a gold nanoelectrodes array in 10⁻⁴ M ferrocene methanol and 0.5 M NaNO₃. Scan rates: 5 (full line), 10 (dashed line), 20 (dotted line), and 50 mV·s⁻¹ (dash–dot line). Geometrical characteristics: nanodisk radius = 75 nm, distance centre-to-centre = 3 μm, number of nanoelectrodes in the array = 1.1 × 10⁴ (reprinted with permission from [68]).

Figure 9

CVs recorded at a NEA (full line, see Figure 8) and at a NEE (dashed line, nanoelectrode radius 15 nm, A_geom = 0.07 cm², A_act= 4.5 × 10⁻³ cm²) made of Au-nanodisks in polycarbonate, plotted using current densities calculated respect to the geometric area (a) and active area (b); scan rate 10 mV·s⁻¹, in 10⁻⁴ M ferrocenemethanol. For further details, see the original article (reprinted with permission from [68]).

Figure 10

ECL images of a BDD-NEA obtained in phosphate buffer (pH 7.4) containing 1 mM Ru(bpy)₃²⁺ and increasing concentrations of TPrA (indicated in top-left corner of each box). Images were recorded in the dark with a ×50 objective when applying a constant potential of 1.2 V vs. Ag/AgCl/KCl. All images were coded according to the same false colour scale (right) (reprinted with permission from [88]).

Figure 11

Different geometries for template nanoelectrode ensembles: (a) ensemble of nanodisks; (b) ensemble of partially naked nanowires; (c) ensemble of completely naked.

Figure 12

Sketch of the effect of different electron transfer kinetics on electrochemical responses at three-dimensional nanoelectrode ensemble (3D-NEE) (reprinted with permission from [101]).

Figure 13

Schematic representation of the analytical protocol used to detect immunoglobulin IgY, extracted from tempera paints. MB and LB are the oxidized and reduced forms of the redox mediator methylene blue and HRP is horseradish peroxidase, used as enzymatic label for the anti-IgY antibody (a) capture of immunoglobulin IgY on the polycarbonate of the NEE; (b) molecular recognition by anti-IgY labeled with HRP; (c) generation of the electrocatalytic cycle by adding the enzyme substrate H₂O₂ and the mediator MB. (reprinted with permission from [109]).

Figure 14

Electrical nanowell biosensor design, assembly and operation. Sensor device and operation (a) optical micrograph showing the gold microelectrodes, PDMS encapsulant and nanoporous alumina membrane (b) modified Randles equivalent circuit for label-free non-faradaic impedance spectroscopy and (c) schematic representation of binding events at the electrical double layer. PDMS: Polydimethyl siloxane; PSA: Prostate-specific antigen (reprinted with permission from [125]).