Electrochemical electrode assembly

Abstract

An electrochemical electrode assembly having micro / nano-scale characteristics, in particular at least one micro / nano-scale working electrode "active" surface, for selective oxidation / reduction of a target analyte or reaction products thereof. The working electrode includes a surface area that is minimal relative to the surface area size of the surrounding insulating material surface such that a target analyte in a biological fluid (e.g., blood, interstitial fluid, sweat, saliva, etc.) needs to diffuse across at least one region of the insulating surface toward the working electrode surface.

Description

Technical Field

[0001] This concept relates to an electrochemical electrode assembly, and particularly to a microelectrode suitable for use as part of an electrochemical sensor, including but not limited to. Background Art

[0002] Electrochemical sensors in implantable or skin-contact devices have received much attention in the field of electroanalysis because they can quantitatively and qualitatively characterize target analytes in complex biological systems. According to existing manufacturing technologies, such micron-scale sensors can achieve high-time-resolution and time-sensitivity analysis.

[0003] The first electrochemical sensor for oxygen was reported in the 1960s. Specifically, Clark and Lyons proposed the concept of glucose sensing (in vitro) in 1962 (based on an oxygen sensor), and the first commercial in vitro electrochemical glucose sensor was manufactured by YSI in 1975 (based on a hydrogen peroxide sensor). The first commercial in vivo glucose sensor came out in 1999. Recently, due to the development of micron- and nanoscale manufacturing technologies, micro / nanoelectrode array sensors have become an attractive type of biofluid sensor, which provides multiplexing capabilities and robustness for bioanalysis in various different environments such as cells, tissues, and organs, and can be used as implantable and non-invasive wearable devices.

[0004] As invasive or wearable intelligent devices, micro- or nanoelectrode array sensors for intelligent sensing generally include electrodes with micron- and nanoscale dimensions. For different measurement conditions, various types of micro / nanoelectrode arrays have been developed. Generally speaking, such electrode arrays are prepared by layer deposition or layer growth, in which layers of metals, carbon, ceramics, etc. are deposited on template substrates such as silicon, glass, polymers, ceramics, etc. Lithography, screen printing, film formation, laser ablation, and 3D printing technologies have been used to fabricate complex electrode micro-patterns. An example of an electrode assembly is described in WO2010 / 061229A1, where the stacked structure includes an insulating cover layer disposed on a conductive layer, which is designed to provide an exposed electrical contact area at an etched void extending through the cover layer.

[0005] However, existing microelectrochemical electrode assemblies and sensors have drawbacks for various reasons. Typically, existing devices cannot achieve separation between individual electrode elements, thus failing to fully exploit the advantages of micro / nano-scale electrodes. Existing structures and sensors involve the use of thin-film technologies that require precise and complex surface patterning to expose areas of the conductive layer. Additionally, existing devices cannot adequately control the diffusion of target analytes (within a carrier biological fluid) towards the conductive regions, resulting in excessive oxidation or reduction of the analytes, a situation that can only be mitigated by an undesirable long pause when applying a potential (to allow fresh analytes to continue diffusing to the active conductive surface) or by adding a restricting membrane (which reduces both the signal and the time response). Accordingly, there is a need for an electrochemical electrode and sensor arrangement that addresses the above problems. SUMMARY OF THE INVENTION

[0006] The goal of this concept is to provide an electrode assembly that can be manufactured conveniently and efficiently, while providing a device suitable for use as an implantable or wearable (skin-contact) electrochemical sensor that results in enhanced sensitivity, reliability, response speed, and reproducibility.

[0007] Another goal is to provide an electrode assembly and an electrochemical electrode-based sensor that provide significantly enhanced analyte diffusion control (non-planar diffusion control) in use. Another specific goal is to provide an assembly that achieves a diffusion-controlled current independent of time during operation (typically when polarized to a potential sufficient to oxidize or reduce the target analyte). Another specific goal is to provide an electrode assembly that controls the diffusion of target analytes in a biological fluid towards the working electrode surface such that the diffusion rate of the target analytes is greater than the oxidation / reduction rate at the electrode surface. Another specific goal is to avoid a significant volume depletion problem of the target analyte near / within the region of the working electrode surface, which would otherwise manifest as a significant reduction or zero Faradaic current in the working electrode region (in response to excessive or complete oxidation / reduction of the target analyte or reaction products of the target analyte with the conductive surface coating material).

[0008] The target analytes referred to in this specification include substances of interest and / or substances produced as a result of the presence of the target analyte. Such substances can be produced by enzyme reactions as described herein.

[0009] A further specific goal is to provide an electrochemical sensor, particularly an implantable or wearable device capable of detecting multiple target analytes in series or in parallel.

[0010] This objective is achieved by this electrochemical electrode assembly with micro / nano-scale characteristics, in particular with a micro / nano-scale working electrode "active" surface for selectively detecting the target analyte. The surface area of this working electrode is minimized with respect to the surface area of the surrounding insulating material surface. According to this concept, the conductive surface is constrained and surrounded by at least one insulating material surface, such that the target analyte in a biological fluid (such as blood, sweat, saliva, etc.) needs to diffuse across at least one region of the insulating surface towards the working electrode surface. By configuring the relative size, geometry, and / or relative spatial orientation of the conductive surface with respect to the surrounding insulating surface, the diffusion of the target analyte in the biological fluid is mainly hemispherical diffusion and exceeds the consumption (i.e., reaction, oxidation, or reduction) rate of the target analyte on the conductive surface.

[0011] This concept is configured to respond to the appearance / presence of the target analyte on the active electrode surface, thereby effectively determining the concentration or rate of change of the concentration of the target analyte in a fluid sample (such as a biological fluid). According to a specific embodiment, the active electrode surface includes an analyte-responsive layer, which has a material configured to bind to, and undergo chemical and / or physical interactions with, the target analyte. In particular, the analyte-responsive layer can contain an enzyme material for interacting or reacting chemically with the target analyte and generating an electrochemically active product resulting from the enzymatic reaction with the target analyte. The target analyte can include an electrochemically active product resulting from an enzymatic reaction with an initial substance. That is to say, the target analyte can be a chemical product resulting from an enzymatic reaction with an initial target substance. For example, when this concept is configured to identify glucose level / concentration, the target analyte can include the product resulting from the enzymatic reaction with glucose, i.e., hydrogen peroxide. This concept can be configured to measure the current between the working electrode and the auxiliary electrode resulting from the direct oxidation or reduction of the target analyte. Or, alternatively or additionally, this concept can be configured to measure the current between the working electrode and the auxiliary electrode resulting from the oxidation or reduction of the product of the electrochemical reaction of the target analyte with the analyte-responsive layer. For example, when the target analyte is glucose, the substances undergoing oxidation and reduction on the active electrode surface can include hydrogen peroxide (i.e., the electrochemically active reaction product of the enzyme analyte-responsive layer material coated and / or located near the active electrode surface). The oxidation / reduction of the target analyte mentioned herein includes directly oxidizing / reducing the analyte or indirectly oxidizing / reducing the analyte through the oxidation / reduction of its reaction product.

[0012] Therefore, this electrochemical method for interrogating at least one target analyte in a biological fluid includes applying an oxidation or reduction potential to the working electrode to oxidize or reduce the target analyte or its reaction product, while measuring the Faraday current on the working electrode. Optionally, the step of applying the oxidation or reduction potential includes performing cyclic voltammetry.

[0013] According to a first aspect of the present concept, there is provided an electrochemical electrode assembly comprising: an electrically insulating material having at least one insulating surface; a conductive material having at least one conductive surface, the perimeter of which is constrained and surrounded by the insulating surface, wherein the surface area of the conductive surface or, when the assembly includes a plurality of conductive surfaces, the surface area of each conductive surface is less than 5% of the surface area of the insulating surface surrounding the perimeter of the conductive surface or the respective surface areas of the insulating surfaces surrounding the respective perimeters of each conductive surface, wherein the surface areas of the insulating surfaces are defined between the inner boundaries of the perimeters of the conductive surface or the respective inner boundaries of the perimeters of each conductive surface and the respective outer boundaries, the outer boundaries being at a distance of at least five times the diameter or minimum width from the respective inner boundaries, the diameter or minimum width being the diameter or minimum width of the conductive surface or, when the assembly includes a plurality of conductive surfaces, the respective conductive surfaces.

[0014] To control the depletion of the target analyte at the working electrode surface, the assembly is configured such that the active oxidation / reduction electrode surface forms a discontinuity within the insulating material surface, thereby representing a relatively “active” surface area that is greatly reduced within the surrounding insulating material surface. The present concept is further configured to include physical properties that facilitate the flow and diffusion of the target analyte to the active working electrode surface. That is, the present concept is configured to minimize or avoid positioning the active surface within hidden, restricted, or otherwise limiting partitions or regions within micro / nano-scale structures that would impede the free flow of the target analyte to the active surface. In a preferred embodiment, both the active surface and the surrounding insulating surface are substantially flat and have no grooves, channels, pits, peaks, or valleys in the nano / micron-scale structure. However, depending on certain manufacturing techniques, such a profile design may be necessary, and the active electrode surface may be provided on the sidewalls, edges, grooves, or valleys of a profiled / heterogeneous nano / micro-assembly structure, particularly a stacked structure in which a conductive layer is covered or constrained by one or more insulating layers, and portions of the stacked structure are ablated or cut (by creating channels or cavities within the structure) to provide regions of the exposed conductive layer surface that provide regions of the active oxidation / reduction surface. Such an arrangement can provide an active electrode surface within a cavity that forms part of a 3D array or structure. The present electrochemical electrode assembly can be fabricated according to various techniques, not limited to atomic layer deposition (ALD), chemical vapor deposition (CVD), plasma-enhanced chemical vapor deposition (PECVD), slot-die extrusion coating, screen printing, inkjet printing, spraying, 3D printing, and other existing microprocessor or chip manufacturing techniques in the field of silicon chip manufacturing.

[0015] Optionally, the surface area of an individual conductive surface or each conductive surface among a plurality of conductive surfaces is less than 300 μm 2 , 200 μm 2 , 150 μm 2 or 100 μm2 Optionally, the surface area of the conductive surface or each conductive surface among a plurality of conductive surfaces is less than 300 μm 2 , 200 μm 2 , 150 μm 2 , 100 μm 2 , 80 μm 2 , 60 μm 2 , 40 μm 2 , 20 μm 2 , 10 μm 2 , 5 μm 2 , 3 μm 2 or 1 μm 2 .

[0016] According to some embodiments of a working electrode with a minimized active surface area, the total conductive surface area of a plurality of active conductive surfaces can be less than 100 μm2. Optionally, according to some embodiments of the electrode configuration and / or the electrochemical sensor, the total surface area of the plurality of conductive surfaces can be greater than 100 μm2. However, in such a configuration, the surface area of each individual conductive surface can be less than 100 μm2.

[0017] Optionally, the surface area of the conductive surface or the total surface area of the plurality of conductive surfaces when the component includes a plurality of conductive surfaces is in the range of 0.001 to 500000 μm 2 , 0.001 to 250000 μm 2 , 0.001 to 100000 μm 2 , 0.001 to 10000 μm 2 , 0.01 to 8000 μm 2 , 0.001 to 6000 μm 2 , 0.001 to 4000 μm 2 , 0.001 to 2000 μm 2 , 0.001 to 1000 μm 2 .

[0018] Optionally, the component is configured to be used with a minimally invasive device such as an implantable or skin-contact device, etc., to serve as an electroanalysis part for quantitatively and qualitatively characterizing a target analyte in a complex biological system. Optionally, for such a configuration and use, when the component includes a plurality of conductive surfaces, the total surface area of the conductive surfaces is in the range of 0.001 to 1000 μm 2 , 0.001 to 800 μm 2 , 0.001 to 600 μm 2 , 10 to 600 μm 2 , 100 to 600 μm 2 , 100 to 400 μm 2, from 100 to 200 μm 2 , from 0.001 to 500 μm 2 , from 0.001 to 300 μm 2 , from 0.01 to 300 μm 2 , from 0.01 to 200 μm 2 , from 1 to 300 μm 2 , from 1 to 200 μm 2 , from 50 to 300 μm 2 , from 50 to 250 μm 2 , from 50 to 200 μm 2 , from 5 to 200 μm 2 or from 10 to 200 μm 2 within the range. This arrangement helps to control the diffusion of the target analyte towards the conductive surface.

[0019] Preferably, the surface area of a single conductive surface, or each conductive surface when the electrochemical electrode assembly includes a plurality of independent conductive surfaces, is at least one order of magnitude smaller than the surface area of the surrounding insulating surface that surrounds the single conductive surface or each independent conductive surface. In particular, the conductive surface or each conductive surface can be less than 300 μm 2 , 200 μm 2 , 150 μm 2 or 100 μm 2 , while the surrounding insulating material can be on the order of at least 1000 μm2. Optionally, the surface area of the respective insulating surface that surrounds the conductive surface or each individual conductive surface can be greater than 5000 μm 2 , 10000 μm 2 , 15000 μm 2 , 20000 μm 2 , 50000 μm 2 , 100000 μm 2 , 500000 μm 2 or 1 mm 2 .

[0020] Optionally, the width, diameter or minimum distance of a single conductive surface or each individual conductive surface may be in the range of 1 to 1000 nm, 1 to 800 nm, 1 to 600 nm, 1 to 400 nm, 1 to 800 nm, 1 to 600 nm, 1 to 400 nm, 1 to 200 nm, 1 to 100 nm, 10 to 100 nm. Optionally, the width, diameter or minimum distance of a single conductive surface or each individual conductive surface may be in the range of 10 to 1000 nm, 50 to 800 nm, 50 to 600 nm, 50 to 400 nm, 50 to 200 nm, 100 to 1000 nm, 100 to 800 nm, 100 to 600 nm, 100 to 400 nm or 100 to 200 nm. Optionally, the width, diameter or minimum distance of each individual conductive surface is less than 100, 80, 60, 40, 20 or 10 nm.

[0021] Optionally, the assembly may include 10 to 100000, 10 to 50000, 10 to 10000, 10 to 8000, 10 to 3000, 10 to 1000, 10 to 800, 10 to 600, 10 to 400, 10 to 200, 10 to 100, 10 to 80 or 10 to 60 active surfaces.

[0022] Optionally, the conductive surface and / or the insulating surface may be substantially flat, convex, concave, shaped to include a shape with peaks or valleys, or a combination thereof. Preferably, the conductive surface is located in a region that does not impede the diffusion of the target analyte towards the conductive surface.

[0023] Optionally, the conductive material is a layer disposed on a substrate, and the electrically insulating material is a layer disposed on the conductive material layer, wherein the insulating material layer is discontinuous to expose regions in the conductive material layer that provide at least one conductive surface. Such exposed regions may be provided by ablating or cutting a portion of the laminated structure.

[0024] Optionally, the assembly may include a single conductive surface. Alternatively, the assembly may include a plurality of conductive surfaces, wherein the perimeter of each conductive surface is separated from each other by a distance that is at least 5 times the diameter or minimum width of any one of the conductive surfaces. That is, the distance by which each conductive surface is separated from each other is greater than the width, diameter or minimum distance across the conductive surfaces. The distance across the conductive surfaces mentioned herein refers to a straight line extending from a first point at the perimeter of at least one conductive surface and a second point at the respective perimeter that bisects the conductive surface. Preferably, the distance is on the nanometer scale. Preferably, the corresponding distance between the inner boundary and the outer boundary of the insulating surface is on the micrometer scale.

[0025] Optionally, each conductive surface is separated from each other by a distance in the range of 10 to 100,000 times, 10 to 10,000 times, 10 to 1,000 times, or 10 to 100 times the width, diameter, or minimum distance across the respective conductive surfaces.

[0026] Preferably, when the assembly includes multiple conductive surfaces, each conductive surface is separated from another conductive surface by a distance of at least five times the diameter or minimum width of the respective conductive surface. Optionally, the distance can be at least 10, 50, 100, or 1,000 times the diameter or minimum width of the respective conductive surface. This arrangement ensures that the conductive surfaces do not interfere with each other and, in particular, can absorb the target analyte from the electrolyte medium in a controlled manner, thereby providing controlled diffusion, particularly oxidation / reduction characteristics, and an electrode assembly with a time-independent response.

[0027] Optionally, the assembly can include an analyte-responsive layer disposed on or at the conductive surface. Optionally, the analyte-responsive layer includes any of the following: a material configured to bind the target analyte; an enzyme material that interacts or undergoes a chemical reaction with the target analyte. Optionally, the analyte-responsive layer includes at least one of an aptamer, an antibody, a molecularly imprinted polymer, or a combination thereof. Optionally, the assembly can include multiple analyte-responsive layers, each analyte-responsive layer including a different material to bind a different respective target analyte.

[0028] Optionally, the assembly can include multiple conductive surfaces, each conductive surface including a different material to attract, capture, adsorb, undergo a chemical reaction with, and / or bind a different respective target analyte. Thus, a single sensor can be configured to sense multiple different target analytes in parallel (either directly or indirectly through oxidation or reduction) during one or more data collection cycles.

[0029] According to another aspect of the present concept, there is provided an electrochemical sensor including: at least one working electrode including the electrode assembly described and claimed herein; and at least one auxiliary electrode. It should be understood that in use, the working electrode is electrically coupled to another working electrode through a sample fluid (biofluid) in which the target analyte is present. That is, the working electrode and the auxiliary electrode are in an electrically isolated state when not in use (e.g., in an "open-air" environment). The electrical coupling between the working electrode and the auxiliary electrode is only completed through the sample fluid (biofluid) to form a complete circuit.

[0030] Optionally, the sensor can include an additional reference electrode (relative to the working and / or auxiliary electrodes) such that the sensor includes two, three, or more electrodes.

[0031] Optionally, the sensor may further comprise: a potentiostat electrically connected to the electrodes for applying a potential to at least the working electrode and / or measuring the potential between the working electrode and one of the auxiliary electrodes; and a control program for receiving charge, current, and / or charge or current data generated by the oxidation or reduction of the analyte or the reaction product of the target analyte at the working electrode, and / or measuring the potential on the electrodes. Optionally, the sensor may further comprise an electrochemical potential module for generating a potential and applying it to the working electrode. The module may be implemented as software and / or firmware, stored on and running on a suitable operating system / platform (e.g., potentiostat device), which is familiar to those skilled in the art. Optionally, the electrochemical potential module is configured to apply a pulsed potential to the working electrode.

[0032] According to another aspect of the present concept, there is provided a sensor as described and claimed herein. According to another aspect of the present concept, there is provided a wearable device attachable to the body of a human or animal, comprising a sensor as described and claimed herein. According to another aspect of the present concept, there is provided a method of manufacturing an electrode assembly as described and claimed herein. According to another aspect of the present concept, there is provided a method of manufacturing an electrochemical sensor as described and claimed herein.

[0033] According to another aspect of the present concept, there is provided an electrochemical method for interrogating at least one target analyte in a biological fluid, comprising: providing an electrochemical sensor as described and claimed herein; contacting the sensor with a body containing a biological fluid comprising at least one target analyte; applying an oxidation or reduction potential to the working electrode to oxidize or reduce the target analyte or the reaction product of the target analyte; and measuring the current between the working electrode and the auxiliary electrode generated by the oxidation or reduction of the target analyte or the reaction product of the target analyte.

[0034] Optionally, the step of applying the oxidation or reduction potential includes performing at least one electroanalytical technique, such as voltammetry (e.g., cyclic voltammetry), potentiometry, or amperometry. Optionally, the electroanalytical technique includes electrochemical impedance spectroscopy (EIS). As understood, many electroanalytical techniques are compatible with the present concept. Optionally, the method includes applying a potential for a predetermined period of time. Optionally, the method includes applying a pulsed potential to the working electrode, the pulsed potential having a period of a first potential and a period of a second potential that is zero or lower than the first potential.

[0035] According to another aspect of the present concept, there is provided an electrochemical method for interrogating at least one target analyte in a biological fluid, comprising: providing an electrochemical sensor having an electrode assembly, the electrode assembly comprising: an electrically insulating material having at least one insulating surface; a conductive material having at least one conductive surface, the conductive surface being constrained and surrounded by the insulating surface; bringing the sensor into contact with a body comprising the biological fluid containing at least one target analyte; applying an oxidation or reduction potential at a working electrode to oxidize or reduce the target analyte or a reaction product of the target analyte; and measuring a current between the working electrode and a counter electrode resulting from the oxidation or reduction of the target analyte or a reaction product of the target analyte; wherein the surface area of the conductive surface relative to the surface area of the surrounding insulating surface or, when the assembly comprises a plurality of conductive surfaces, the surface area of each conductive surface is configured to: control the diffusion rate of the target analyte to the conductive surface within the biological fluid such that the volumetric consumption rate of the target analyte at the conductive surface is less than the diffusion rate of the same volume of the target analyte in the biological fluid to the conductive surface, thereby maintaining supply at the conductive surface by diffusion of the target analyte.

[0036] This electrochemical electrode structure comprises at least one active electrode surface that is small enough and optionally forms a discontinuity within a relatively large surface area of the insulating material such that the transport of the electroactive species (target analyte) proceeds under non-linear diffusion control. Preferably, the inner boundary perimeter of the surrounding insulating surface is at least 10 times smaller than the outer boundary perimeter of the insulating surface. In practice, the outer boundary determines the minimum distance to any nearest or adjacent conductive surface or edge within the electrode assembly. Preferably, the conductive surface comprises a width, diameter or other cross-surface distance of less than 10 μm, more preferably less than 100 nm in at least one dimension (two-dimensional or three-dimensional). For example, for a conductive surface with an area of a and a diameter or minimum width of x, the perimeter of the conductive surface must be at least 5x separated in all directions and in the main plane of the insulating surface and comprise a minimum area of 25a. The relative dimensions of the conductive surface (oxidation / reduction) to the surrounding insulating material surface are used to provide the required diffusion control for the target analyte. The present concept provides a sensor arrangement that responds under spherical or hemispherical diffusion control, thereby providing an electrochemical sensing arrangement and method that is independent of time and biological fluid volume.

[0037] According to another aspect of the present concept, there is provided an electrochemical electrode assembly, comprising: an electrically insulating material having at least one insulating surface; a conductive material having at least one conductive surface constrained and surrounded by the insulating surface; wherein the surface area of the conductive surface is less than 5% of the surface area of the insulating surface, the surface area of the insulating surface being defined between an inner boundary and an outer boundary of the perimeter of the conductive surface, the distance between the outer boundary and the inner boundary being at least five times the diameter or minimum width of the conductive surface.

[0038] According to another aspect of the present concept, there is provided an electrochemical electrode assembly comprising: an electrically insulating material having an insulating surface; a conductive material having a conductive surface, the perimeter of which is constrained and surrounded by the insulating surface; wherein the surface area of the conductive surface is less than 5% of the surface area of the insulating surface surrounding the perimeter of the conductive surface, and wherein the insulating surface is defined between an inner boundary and an outer boundary at the perimeter of the conductive surface, the distance between the outer boundary and the inner boundary being at least five times the diameter or width of the conductive surface.

[0039] According to another aspect of the present concept, there is provided an electrochemical electrode assembly comprising: an electrically insulating material having an insulating surface; a conductive material having a conductive surface, the perimeter of which is constrained and surrounded by the insulating surface; wherein the surface area of the conductive surface is one order of magnitude smaller than the surface area of the insulating surface surrounding the perimeter of the conductive surface.

[0040] Optionally, the surface area of the insulating surface is defined between an inner boundary and an outer boundary at the perimeter of the conductive surface, the distance between the outer boundary and the inner boundary being several times the diameter or width of the conductive surface. Optionally, the distance is at least 5, 10, 50, 100, 500, 1000, 10000 or 100000 times the diameter or width of the conductive surface. Optionally, the distance is at least 10 to 100000 times, 10 to 10000 times, 10 to 1000 times or 10 to 100 times the diameter or width of the conductive surface.

[0041] Preferably, the width of the conductive surface is the minimum width. The minimum width refers to the shortest distance of the conductive surface when the surface width or thickness is variable / inconsistent.

[0042] According to another aspect of the present concept, there is provided an electrochemical electrode assembly comprising: an electrically insulating material having at least one insulating surface; a conductive material having a plurality of conductive surfaces, each conductive surface having a perimeter constrained and surrounded by the insulating surface; wherein the surface area of each conductive surface is less than 5% of the respective surface area of the insulating surface surrounding the perimeter of each conductive surface, and wherein the respective surface areas of the insulating surfaces are defined between an inner boundary and respective outer boundaries at the perimeters of the respective conductive surfaces, the distance between the outer boundary and the respective inner boundary being at least five times the diameter or minimum width of the respective conductive surface. BRIEF DESCRIPTION OF THE DRAWINGS

[0043] The specific embodiments of the present invention will now be described by way of example only and with reference to the accompanying drawings, wherein:

[0044] Figure 1A is a plan view of a first stage of a manufacturing process of an electrode suitable as an electrochemical sensor according to a specific embodiment;

[0045] Figure 1B is Figure 1APlan view of the second stage manufacturing process of the electrode;

[0046] Figure 1C is Figure 1A and 1B Enlarged plan view of the distal end of the electrode;

[0047] Figure 2A is Figures 1A to 1C Plan view of the third stage manufacturing process of the electrode;

[0048] Figure 2B is Figures 1A to 2A Plan view of the final stage manufacturing process of the electrode;

[0049] Figure 2C is Figure 2B Enlarged side view of the distal end of the electrode;

[0050] Figure 2D is Figure 2C Enlarged side view of the distal end of the electrode;

[0051] Figure 3 is the cyclic voltammogram of the electrode using a silver chloride reference electrode and a Pt wire in 1 mmol dm -3 PBS of FCA at a scan rate of 50 mV s -1 of, using Figure 2B and C;

[0052] Figure 4 is Figure 2B and 2C of the electrode in 1 mmol dm -3 PBS of FCA after polarization to 0.35 V;

[0053] Figure 5 is Figure 2B and 2C of the electrode in 1 mmol dm -3 PBS of FCA with 100 ms 0.35 V pulses repeated 5 times, with a 10 s rest at open circuit potential between each repetition;

[0054] Figure 6 is the current-time plot of the nanoribbon array electrode in 1 mmol dm -3 PBS of FCA with 100 ms 0.35 V pulses repeated 5 times, with a 10 s rest at open circuit potential between each repetition;

[0055] Figure 7 is a 100 μm diameter platinum disk in 1 mmol dm -3Current vs. time plot of 5 repetitions of 0.35 V pulses at 100 ms in PBS of FCA, resting at open circuit potential for 10 s between each repetition;

[0056] Figure 8 For the widest coherence window Figure 2B and 2C of the electrodes in 1 mmol dm -3 Response plots of the electrodes in PBS of FCA and PBS, 0.35 V pulses, 0.1 ms sampling interval within a 20 ms window, and OCP of 1 s between repetitions;

[0057] Figure 9 For including the Figure 2B and 2C with the earliest data points of FCA repetitions of the electrodes in 1 mmol dm -3 Response plots of the electrodes in PBS of FCA and PBS, 0.35 V pulses, 0.1 ms sampling interval within a 20 ms window, and OCP of 1 s between repetitions;

[0058] Figure 10A Is Figure 2B and 2C of the electrodes in 1 mmol dm -3 Plot of 10 repetitions of 5 ms 0.35 V pulses in PBS of FCA with a 50 ms rest at open circuit potential between each repetition;

[0059] Figure 10B Is of the nanobelt array electrodes using a commercial ASIC (ADuCM355) in 1 mmol dm -3 Plot of 10 repetitions of 5 ms 0.35 V pulses in PBS of FCA with a 50 ms rest at open circuit potential between each repetition;

[0060] Figure 11 Is of a 100 μm diameter platinum disk using a commercial ASIC (ADuCM355) in 1 mmol dm -3 Plot of 10 repetitions of 5 ms 0.35 V pulses in PBS of FCA with a 50 ms rest at open circuit potential between each repetition;

[0061] Figure 12A Is Figure 2B and 2C Side view of another embodiment of the electrodes, Figure 2B and 2C of the electrodes having an active electrode surface formed as a protrusion or bump protruding from the substrate;

[0062] Figure 12B Is Figure 2B and 2C Side view of another embodiment of the electrodes,Figure 2B and 2C The electrodes of 2C include active electrode surfaces formed as concave or serrated / recessed cavities on the substrate surface;

[0063] Figure 13 is a plan view of a dual - electrode system that includes a pair of electrodes disposed on the top and bottom surfaces of an intermediate substrate according to the embodiment of Figure 2B and 2C ;

[0064] Figure 14 is a side view of the distal ends of the electrodes of Figure 13 disposed on the top and bottom surfaces of the substrate;

[0065] Figure 15A is an initial - stage manufacturing process of another embodiment of the electrodes of Figure 2B and 2C ;

[0066] Figure 15B is Figure 15A the final - stage manufacturing process of the electrodes of Figure 15A The electrodes of Figure 15A include a triple - conductive - layer arrangement, where each layer includes a plurality of active electrode surfaces according to another embodiment;

[0067] Figure 16A is Figure 2B and 2C a plan view of another embodiment of the electrodes of

[0068] Figure 16B where the assembly includes a plurality of laterally - defined individually - addressable electrode regions; Figure 2B and 2C ;

[0069] Figure 17A is the first - stage manufacturing process of another embodiment of an electrode assembly;

[0070] Figure 17B is Figure 17A the second - stage manufacturing process of the electrodes of

[0071] Figure 17C is Figure 17A the third - stage manufacturing process of the electrodes of

[0072] Figure 17D is Figure 17A the fourth - stage manufacturing process of the electrodes of

[0073] Figure 18A is Figure 17D another plan view of the electrodes of

[0074] Figure 18B is Figure 18A a side sectional view along the CC direction;

[0075] Figure 19 is a plan view of another embodiment of an electrode assembly that includes a plurality of active electrode surfaces defined at open grooves, holes, or open cavities of an electrode assembly having a grid or mesh conductive layer structure;

[0076] Figure 20A is a first stage manufacturing process for another embodiment of an electrode assembly having a grid or mesh conductive layer structure;

[0077] Figure 20B is a second stage manufacturing process for another embodiment of an electrode assembly having a grid or mesh conductive layer structure;

[0078] Figure 20C is a third stage manufacturing process for another embodiment of an electrode assembly having a grid or mesh conductive layer structure;

[0079] Figure 20D is a fourth stage manufacturing process for another embodiment of an electrode assembly having a grid or mesh conductive layer structure;

[0080] Figure 20E is a fifth stage manufacturing process for another embodiment of an electrode assembly having a grid or mesh conductive layer structure;

[0081] Figure 20F is a final stage manufacturing process for another embodiment of an electrode assembly having a grid or mesh conductive layer structure;

[0082] Figure 21A is another embodiment of an electrode assembly configured with relatively large active electrode surfaces formed within a conductive layer grid arrangement;

[0083] Figure 21B is another embodiment of an electrode assembly configured with relatively large active electrode surfaces formed within a conductive layer grid arrangement;

[0084] Figure 21C is another embodiment of an electrode assembly configured with relatively large active electrode surfaces formed within a conductive layer grid arrangement;

[0085] Figure 22A is a plan view of a hybrid electrode assembly including a nanoribbon array according to another embodiment;

[0086] Figure 22B is a plan view of a hybrid electrode assembly including a nanoribbon array according to another embodiment;

[0087] Figure 23FIG. 0 is a plan view of another embodiment of an electrode assembly that includes a plurality of active electrode surfaces, each disposed on the sidewalls of respective channels or grooves formed within a multi-layer structure;

[0088] Figure 24 FIG. 4 is another embodiment of an electrode assembly that includes a plurality of cavities or holes formed within a multi-layer structure to expose a plurality of active electrode surfaces at open cavities, holes, and / or open grooves. DETAILED DESCRIPTION

[0089] Referring Figures 1A to 1C to FIG. 11, electrode assembly 10 includes a substrate 18 formed of an electrically insulating material. Figures 1A to 2C All dimensions shown in FIG. 13 are in μm. Conductive layer 11 is formed as an elongated strip disposed on surface 19 (FIG. 2) of substrate 18, having a proximal end connected to conductive tab 12 and a distal end 13 that includes a plurality of laterally extending conductive fingers 14. Each conductive finger 14 includes a respective tip 15. According to a particular embodiment, conductive layer 11 has a thickness of approximately 0.05 μm and is disposed on substrate 18, which has a thickness of approximately 250 μm. The approximate width across the electrode at conductive fingers 14 (between respective tips 15) is approximately 600 μm. The width of conductive elongated strip 11 is approximately 60 μm, and the approximate total length of the elongated electrode (including strip 11 and fingers 14) is approximately 40,000 μm. Figure 1A FIG. 15 represents the manufacturing process of a first stage, involving depositing conductive strip 11 and fingers 14 on substrate 18. Figure 1B FIG. 17 represents the manufacturing process of a second stage, in which a layer of insulating material 16 is deposited on top of substrate 18 and conductive layer 11. Insulating layer 16 has a width of approximately 600 μm and a length that extends approximately the full length of conductive strip 11, also approximately 40,000 μm.

[0090] Figure 2A FIG. 21 represents the manufacturing process of a third stage, in which a selected area of the Figure 1B assembly is identified for ablation using a laser cutter or similar technique. After ablation cutting, the completed electrode assembly is as shown in Figure 2B FIG. 25, where the Figure 2A structure is cut / ablated at ablation line 17 (shown schematically). Thus, referring to Figure 2CSide view, where the active electrode surface 15 is provided at each respective end of each conductive finger 14, exposed to the respective side wall surfaces of the stacked electrode assembly including the substrate 18, the conductive layer 11, and the insulating material 16. Importantly, each electrode active surface 15 is spatially separated from each other to provide discrete conductive surface areas for the oxidation / reduction of target analytes contained in biological fluids such as sweat, saliva, blood, etc. According to the present embodiment described herein, the insulating layer 16 is a material configured and selected to be suitable for the end use, which includes biocompatibility when deployed as a minimally invasive device. Such a layered material provides compatibility, adhesion, etc. for any additional functions required for chemical interactions, i.e., binding, reacting, absorbing the target analyte at the region of each active surface 15, etc. for oxidation / reduction, reaction, or interaction with the surface, thereby resulting in a potential change.

[0091] According to the present embodiment of the electrode assembly described herein, an active electrode coating can be provided on each exposed active electrode surface 15. According to the existing arrangement, such a coating is configured to chemically interact with the target analyte so as to bind, react, absorb, etc. the target analyte, so that the analyte remains on the active surface 15 for oxidation and / or reduction, or to produce a change on the surface, thereby resulting in a potential change. In some cases, the desired properties of the active electrode coating can be incorporated into the insulating layer 16, thus eliminating one process step. Such active electrode coatings are well known to those skilled in the field of sensor manufacturing, especially in the field of biosensor manufacturing. Additional coatings can be implemented to beneficially modify the performance of the entire sensor, including but not limited to the purpose of excluding potential interferents. This increases the possibility of using this structure for potential measurement, increases the possibility that the insulating layer 16 is passivated, and also increases the possibility of including an "exclusion membrane" to exclude interferents.

[0092] All embodiments described herein are applicable to implantable or wearable electrochemical sensors as minimally invasive diagnostic sensors.

[0093] According to Figure 2B and 2D For the specific electrode assembly shown, the material of the substrate 18 includes polyethylene terephthalate (PET) to provide a first insulating layer. The conductive layer 11 optionally includes platinum deposited using appropriate techniques. Figure 2B and 2D The final electrode structure of can be achieved by, for example, laser ablation or lithography to create a conductive pattern for the oxidation / reduction or surface interaction of the target analyte, or other substances produced due to the presence of the target analyte. The conductive pattern has discrete spatially separated regions of the active electrode surface. The other substances are, for example, reaction products of the analyte with the active coating, and the active coating is, for example, an enzyme-active coating that reacts with the analyte.

[0094] according to Figure 2B and 2D The electrode assembly includes fingers 14 sized and spaced to ensure that the electrode structure is electrically addressable, and includes 12 active conductive areas 15 that are appropriately sized (each active area is 30 μm long and 50 nm wide) to ensure high diffusion characteristics (50 nm wide), while being sufficiently separated from adjacent active areas 15 (90 μm) to minimize any interaction between adjacent active areas 15. At the proximal end, second and third contact pads 12 enable the incorporation of (optional) auxiliary electrodes (not shown) as required. The size of the pads 12 can be varied to accommodate the selected electrical connection method (pad size, spacing, etc.). These auxiliary electrodes can be incorporated on the back side of the structure that requires a through hole for connection, or on the same side as the contact pads 12 adjacent to the working electrode structure.

[0095] Reference Figure 2D , the insulating surface surrounding each active electrode surface 15 is formed by a layer of substrate 18 and insulating material 16. The substrate 18 and insulating layer 16 together provide a surrounding insulating surface that limits and surrounds each active electrode surface 15. As shown, the relative size of each active electrode surface 15 is significantly smaller than the surface area surrounding the insulating material (16, 18). In particular, the surface area of ​​each active electrode surface 15 can be approximately 1.2 μm 2 , or less than 1μm 2 , 0.5μm 2 or 0.1μm 2 .

[0096] Each active electrode surface 15 is bounded and surrounded by an area 24 of insulating material. Each area 24 comprises a surface area on the order of square micrometers relative to the area of ​​each active electrode surface 15 being on the order of square nanometers. In particular, the area 24 of insulating material is defined between an inner boundary 21 and a respective outer boundary 23, the inner boundary 21 corresponding to the perimeter 20 of the respective active electrode surface 15. The outer boundaries 23 are separated by a distance d, which is five times the distance e, which is the diameter or minimum width of the respective active electrode surface 15. This arrangement facilitates the control of the diffusion of analytes (target substances) to the active conductive surface 15, in particular avoiding undesirably high analyte diffusion rates, which in turn provides controlled oxidation / reduction of the target substance.

[0097] For clarity, Figures 1A to 1C The auxiliary electrode is omitted. In this embodiment, the auxiliary electrode is made by using a commercial silver / silver chloride screen printing ink on the back side of the substrate 18 and patterning it. Alternative auxiliary electrodes and materials for making them can be defined and are known to those skilled in the art. The structure is then screen printed using a commercially available dielectric ink to produce aFigure 1B The dielectric layer 16 shown, which covers the entire structure except for the electrical contact pads 12. The thickness of the dielectric layer is approximately 30 μm (which is sufficient to ensure that the active regions exposed in the last step (described in detail below) are fully encapsulated in the electrochemically passivating material to achieve optimal diffusion (sometimes described as hemispherical). Alternatively, a second screen printing step can be added to minimize any areas of exposed conductor (commonly referred to as pinholes). The back side is also screen printed with dielectric ink up to 700 μm from the distal end of the structure, exposing the silver / silver chloride layer at the far end (not shown for clarity). In Figure 2A the scribed ablation line 17 of the structure 10 is shown, which cuts through the dielectric / electrical conductor / PET substrate at the distal end, exposing six electrical conductor end faces 15, each 30 μm long and 50 nm wide on each side. As Figure 2B and Figure 2C shown, each surface 15 of the divided device is 90 μm apart from the adjacent surface (120 μm center to center). The total working electrode area in this embodiment is 18 square μm, compared to a typical structure in this scenario of approximately 8000 square μm (i.e., the surface area is reduced by approximately 400 times). This is beneficial for reducing the inherent capacitance of the electrode. The scribing process is carried out using a femtosecond infrared laser, and the width of the final structure is approximately 400 μm. It should be noted that the preliminary estimation of the device's performance is independent of its width, and the performance depends on the number, size, spacing, and environment of the active regions (surfaces 15). This is not the case when considering traditional electrode structures. This is important because the size of the overall structure has a significant impact on the foreign body reaction (FBR), which is in situ initiated when this example is used in applications such as minimally invasive implants (e.g., when applied to applications for continuous glucose monitoring). The larger the implant, the faster the FBR, and thus the shorter the lifespan of the implant. Therefore, being able to freely adopt the optimal width without compromising performance is an important feature.

[0098] The electrode embodiments in Figure 2B and Figure 2C are tested with ferrocene carboxylic acid (FCA) and hydrogen peroxide (HP). These two analytes are selected as surrogates for first-generation and second-generation biosensors. The first-generation biosensor utilizes an enzyme that produces HP, which is subsequently measured at the indicator electrode, and the second-generation biosensor utilizes an artificial mediator (FCA). The first-generation biosensor requires a catalytically active platinum electrode surface for HP electrochemistry. Unless otherwise stated, the measurements are carried out at room temperature in phosphate buffer saline (PBS). Regarding the potential range for cyclic voltammetry of silver / silver chloride, it is 0–0.45 V for FCA and -0.2 to 0.75 V for HP (scan rate of 50 μmVs -1) Chronoamperometric measurements were carried out under conditions of 0.35 V for FCA and 0.6 V for HP. For comparison, measurements were made using a 100 μm platinum disk electrode as a model of the current implantable device and a conventional 50 nm platinum nanobelt array electrode (30 μm holes on 60 μm pitch) as an example of the current teachings in terms of nanobelt electrode structures.

[0099] From Figure 3 it can be seen that when interrogated using cyclic voltammetry in the presence of 1 mmol dm -3 FCA, Figure 2B and 2C the electrodes in [[ ]] exhibit typical nanobelt capabilities as there are no signs of mass transport limitations (the oxidation wave remains constant above 0.35 V rather than reaching a maximum). Similar waveforms were observed when interrogating the conventional nanobelt array.

[0100] When interrogating this example using a potential step, a step to 0.35 V with respect to the silver / silver chloride electrode, it can be seen that there is a deviation from the behavior of the conventional nanobelt array or a platinum electrode area of 10 mm 2 at the macroscopic scale ([[]] Figure 4 [[]]). In the case of the nanobelt array and the macroscopic electrode, the continuous decrease in current within the 60 s measurement window indicates that these systems are limited by mass transport and cannot replenish the FCA consumed at the electrode surface. In the case of this example, it can be seen that when the current decrease appears to stop, a steady state is reached within approximately 6 s. This time-independent response is characteristic of the establishment of optimal (hemispherical) diffusion. At this point, the mass transport rate of the analyte to the electrode surface is sufficient to replenish the FCA consumed at the electrode. This situation is typically only observable when the flow in the electrode surface area is very carefully managed, whereas in this case the solution is stationary.

[0101] In PBS with 1 mmol dm -3 FCA and PBS (“blank”), this example was challenged with 100 ms 0.35 V pulses with a 10 s rest (electrode held at open circuit potential) between pulses. The behavior was compared to that of a 100 μm Pt disk electrode under the same conditions. The results are shown in [[ ]] Figures 5 to 7 [[]]. It can be seen that in the case of this example, the 5 repeated pulses are randomly ordered, while the response of the 100 μm Pt disk is ordered, decreasing from the initial response with 5 repeated pulses. This indicates that even with a short consumption period (100 ms) and a long recovery time (10 s), the 100 μm disk consumes more FCA than can be replenished by mass transport under stationary conditions, but this is not the case for this example. For applications where it is not possible or not desirable to manage the solution flow, this provides improved measurement accuracy. ​​​​

[0102] This example was challenged within a 20 ms sampling window (with 1 s of OCP between repeats) at the shortest time interval available on the instrument used (0.1 ms) to demonstrate that the response of this example remains predominantly a Faradaic response (i.e., dominated by redox processes rather than charging phenomena). Five pulse repeats were measured. At these high sampling rates using a potentiostat, there are instrument artifacts that manifest as complex waveforms. When the waveforms of each response are synchronized, a clear Faradaic response can be seen ( Figure 8 and 9 ). With a sampling interval of 0.1 ms, it can be seen that for 4 repeats of the FCA starting from the first data point (t = '0'), there is no sign of charging, whether at the initial point t = '0' or the first point t = 0.1 ms ( Figure 9 ). Thus, it is clear that the example exhibits a very low non-Faradaic (capacitive charging) component and may be polled at kilohertz frequencies. Although t = '0' will occur at some point in time after polarization of the example, the period will be less than 0.1 ms. Therefore, it is obvious that the example may be able to be interrogated at higher frequencies / shorter sampling intervals with appropriate hardware.

[0103] In addition, this example was challenged using a commercial application-specific integrated circuit (ASIC) designed for mobile / wearable scenarios (using a PalmSens Sensit BT potentiostat utilizing the Analogue Devices ADuCM355 ASIC). The shortest sampling interval allowed by the software, 1 ms, was used for the minimum number of samples (5). It should be noted that the native capability of the ADuCM355 is 400 thousand samples per second. In PBS with 1 mmol dm -3 of FCA and in PBS only ("blank"), 5 ms 0.35 V pulses were used with a 50 ms rest (electrode held at open circuit potential) within the software limitations. The responses under these conditions are as shown in Figure 10A , 10B and 11. Results are given for the electrodes in Figure 2B and 2C , a 100 um diameter disk electrode, and a nanoribbon array.

[0104] This electrode assembly can be implemented in different forms to adapt to the application, the analyte of interest, and its diffusion characteristics in the matrix under consideration. The sensitivity of the device will be linearly proportional to the length. Thus, if the fingers 14 are defined to be 1440 μm in length (i.e., 12 fingers 14 on each side instead of 6, with the same arm width and spacing), the signal current will double. If the number of fingers 14 on the same 720-μm length is doubled, a similar increase in the signal current will be observed. Therefore, the structure can vary from a single monolithic finger 14 (720 μm long in this exemplary embodiment) to a highly segmented finger structure (e.g., 72 fingers 5 μm wide separated by 5 μm), and then to a sparsely distributed finger structure (such as the current embodiment). A suitable structure can be fabricated (designed) according to the application requirements under consideration to provide optimal performance. In addition, without compromising performance, this device can be made significantly narrower (400 μm in this embodiment). The limitation of the width of this device depends on the length and resistivity of the conductor tracks 11 ( Figures 1A to 2C ), as well as the manufacturing tolerances of the manufacturing methods and tools used. In the case of this embodiment using the manufacturing tools employed, a 60-μm-wide embodiment would be possible. Further practical considerations can be whether the desired mechanical properties are maintained when the width of the embodiment is reduced. However, it should be noted that the electrochemical performance will not be weakened.

[0105] In the said embodiment, the active region surface 15 is planar. According to another embodiment, such an active surface 15 can be concave or convex, as Figure 12A and 12B shown, being two-dimensional or three-dimensional.

[0106] In another alternative embodiment, the second electrode is defined on the back side of the insulating substrate 18 by depositing and patterning an additional conductor layer, followed by the insulating layer 16 as described above, as Figure 13 and 14 shown. In this embodiment, the "top surface" and "bottom surface" electrodes coincide. In another embodiment, the active regions 15 of the two electrodes are offset or staggered. In yet another embodiment, the active regions 15 and the spacing between the active regions 15 of the two electrodes are not the same.

[0107] In an alternative embodiment, by adding additional conductive and insulating layers and patterning processes, on a single substrate ( Figure 15A and 15B) generates two or more stacked independently addressable active regions 15 (collectively referred to as electrodes) on one side. In the illustrated embodiment, the active regions 15 of the two electrode structures are the same and coincide. In some embodiments, the active region structures may not be the same (e.g., if configured to oxidize / reduce different target analytes or their reaction products), and in other embodiments, the active region structures may not coincide.

[0108] Multiple independently addressable active regions 15 can also be defined laterally, and some examples are shown in FIG. 16. The substrate insulating substrate 18 is blocked by the conductive layer 11 and the dielectric insulating top layer 16. The size, spacing, and structure of the active regions 15 are the same, but in another embodiment, they can be different according to the sensing application. Any combination of one or all of the above methods can be employed.

[0109] In other embodiments of this assembly, the PET substrate 18 can be replaced with any suitable material having the required mechanical, biological, biocompatible, and electrical properties (such as polyester, polyalkane, etc.). Similarly, the Pt conductive layer 11 can be replaced with any suitable material having the required mechanical, biological, biocompatible, electrical, and electrochemical properties (such as platinum group metals, metals, organic conductors, etc.). In addition, the thickness of the conductive layer 11 can vary as needed (typically in the range of 0.1 - 100 nm). Furthermore, the insulating top layer 16 can be replaced with any suitable material having the required mechanical, biological, biocompatible, and electrical properties.

[0110] In some cases, it may be necessary to separate the process of creating the active regions 15 from the segmentation process. Figures 17A to 17D and Figure 18A and 18B Examples of this situation are shown. The segmentation can be performed anywhere outside or inside the ablation region 17, as long as the line is outside the inner line of the ablation.

[0111] In this example, the same basic electrode pattern is used (as shown in Figure 2B and 2C ), and it includes six biaxial fingers 14 that project laterally from the central conductor strip at the distal end. The patterning and layer structure are also the same as in the previous example. Specifically, the starting point of the fabrication is a PET substrate 18 with a 50 - nm Pt layer, in which a suitable pattern is ablated in the Pt layer 11. For this other example, the width of the fingers 14 is 30 μm, and the center - to - center spacing is 120 μm. Most of the pattern and the remaining Pt / PET regions are screen - printed with a suitable dielectric ink layer 16 to electrically isolate the Pt layer 11 from the small remaining regions that define the electrical contact pads 12, as shown in Figure 20B shown. As in Figure 2B and Figure 2CAs in the previous examples, the contact pad size can be optimized for the electrical connection system used. Figure 20C The marking for the ablation trace 17 is shown (if desired, there can be a single trace area), and Figure 20D The component after the ablation trace (through the dielectric layer 16, the conductor layer 11 and reaching the PET substrate 18, which acts as an active "etch stop" in this case) is completed. Ablation can be achieved using laser machining, die cutting or similar methods including methods such as reactive ion etching or chemical etching. Figure 20E A cross-section is shown, which in this example extends slightly into the insulating substrate 18. The device can be segmented into any size and shape suitable for the desired application. As in the first embodiment described, the materials and structures can vary.

[0112] These two methods can be combined to enhance the generated signal. Figure 19 An example is shown in which eight additional active surfaces 15 are defined in the surface to enhance the ten active surfaces 15 defined when the structure is segmented. The number, spacing and size of the active regions defined in the surface can be changed in the form of changing the pattern of the conductor layer and / or the holes themselves to optimize the performance of the structure in the selected application.

[0113] The benefits of the present invention have been demonstrated using an extreme embodiment, which is suitable for in vivo minimally invasive applications such as continuous glucose monitoring. The inventors have recognized that the size of the present electrode assembly is important as this determines the lifespan and performance of the device in use. In applications where this is not a critical issue but the management of sample flow is impossible / undesirable and / or the volume of the sample being addressed is unknown and / or small, the basic method can be extended to assemblies having active surfaces 15 on a larger surface area. An example of this method is shown in Figures 20A to 20F For some applications, it may be desirable to repeat the surface ablation pattern over a wider area. Examples of such methods are given below.

[0114] In this case, the manufacturing starting point is a PET substrate 18 with a 50 nm Pt layer 11 having a suitable pattern ablated into the Pt layer 11. The pattern and most of the remaining Pt / PET layers 11, 18 are screen printed with a suitable dielectric ink 16 to electrically isolate the Pt from the small remaining areas defining the electrical contact pads 12. Figure 20C The ablation trace 17 is shown, which alternatively can be a series of five separate traces 17. Figure 20D The component after the ablation trace 17 has been completed (through the stack and into or reaching the PET substrate 18) is shown. Figure 20E The marking line 17 for the cutting / laser machining 22 for device segmentation is shown, and in Figure 20FThe completed structure is shown. The materials and structure can vary as in the first embodiment described. In this example, active regions 15 are defined at the sidewalls of the layered material structure (formed by ablation cutting) during the separation process, in addition to those created during the surface ablation process. This is optional and, if not required for a given application, the separation can be carried out in such a way that no additional active regions 15 are created during the process.

[0115] The concept can include other electrode structures, such as having a nanoribbon array as shown in Figure 22A and Figure 22B . In the embodiment of Figure 22A , the nanoribbon array has been defined and constructed as an independent addressable electrode juxtaposed to the embodiments described previously. In this example, the contact pads 12 are on opposite sides of the structure, but they can alternatively be arranged on the same side of the structure to provide a convenient way to connect to a selected multi-channel electrical connector. In the embodiment of Figure 22B , the structure is addressed such that the present invention and the nanoribbon array electrodes are addressed as a single working / indicating electrode through a single connection pad.

[0116] The described embodiments can also be combined with one or more conventional larger-scale electrodes, for example, using a screen printing method if required.

[0117] In another embodiment, as shown in Figure 23 , a series of interdigitated fingers 14 are defined in the conductor layer 11. The active surfaces 15 defined by ablation / cutting are present on the sidewalls of the channels or grooves (formed in the laminated structure) that have been formed. In this example, the surfaces 15 on each opposite sidewall are not opposite each other (within each channel or groove), but are offset or staggered in the longitudinal direction of each channel or groove. This helps to ensure that each active surface 15 faces a passivated region in the structure (i.e., does not directly face another active surface 15), further reducing the ability of an active region 15 to strip a relative active region 15 immediately adjacent to an oxidizable / reducible analyte. In this example, no additional edges or active surfaces 15 are created during the separation (although these edges or active surfaces 15 can of course be created if additional fingers 14 are defined to extend to the perimeter of the construction).

[0118] If required, the surface patterning method can be restricted to discrete regions, and an example is shown in Figure 24 . The number of discrete ablation holes (open cavities, recesses or pits) 22 can vary as required, and the number, size and spacing of the active regions 15 defined within each ablation hole 22 can also vary to provide the required performance for the selected application.

Claims

1. An electrochemical electrode assembly, comprising: an electrically insulating material having at least one insulating surface; a conductive material having at least one conductive surface, the conductive surface having a perimeter constrained and surrounded by the insulating surface, wherein the surface area of the conductive surface or, when the assembly includes a plurality of conductive surfaces, the surface area of each conductive surface is less than 5% of the surface area of the insulating surface surrounding the perimeter of the conductive surface or the respective surface areas of the insulating surfaces surrounding the respective perimeters of each conductive surface, wherein the respective surface areas of the insulating surfaces are defined between an inner boundary at the perimeter of the conductive surface or the respective inner boundaries at the perimeters of each conductive surface and respective outer boundaries, the outer boundaries being at a distance of at least five times the diameter or minimum width across the conductive surface or, when the assembly includes a plurality of conductive surfaces, across each conductive surface, the diameter or minimum width being the diameter or minimum width across the conductive surface or across each conductive surface when the assembly includes a plurality of conductive surfaces.

2. The component according to claim 1, wherein The surface area of each of the individual conductive surfaces or the multiple conductive surfaces is less than 300 μm 2 , 200 μm 2 , 150 μm 2 , 100 μm 2 , 80 μm 2 , 60 μm 2 , 40 μm 2 , 20 μm 2 , 10 μm 2 , 5 μm 2 , 3 μm 2 or 1 μm 2 .

3. The component according to claim 1, wherein, When the component includes a plurality of conductive surfaces, the total surface area of the conductive surfaces is in the range of 0.001 to 500000 μm 2 , 0.001 to 250000 μm 2 , 0.001 to 100000 μm 2 , 0.001 to 10000 μm 2 , 0.01 to 8000 μm 2 , 0.001 to 6000 μm 2 , 0.001 to 4000 μm 2 , 0.001 to 2000 μm 2 , 0.001 to 1000 μm 2 .

4. The component according to claim 1, wherein When the component includes a plurality of conductive surfaces, the total surface area of the conductive surfaces is in the range of 0.001 to 1000 μm 2 、0.001 to 800 μm 2 、0.001 to 600 μm 2 、10 to 600 μm 2 、100 to 600 μm 2 、100 to 400 μm 2 、100 to 200 μm 2 、0.001 to 500 μm 2 、0.001 to 300 μm 2 、0.01 to 300 μm 2 、0.01 to 200 μm 2 、1 to 300 μm 2 、1 to 200 μm 2 、50 to 300 μm 2 、50 to 250 μm 2 、50 to 200 μm 2 、5 to 200 μm 2 or 10 to 200 μm 2 in the range.

5. The component according to claim 1, wherein The width, diameter or minimum distance of the single or each individual conductive surface is in the range of 10 to 1000 nm, 50 to 800 nm, 50 to 600 nm, 50 to 400 nm, 50 to 200 nm, 100 to 1000 nm, 100 to 800 nm, 100 to 600 nm, 100 to 400 nm or 100 to 200 nm.

6. The assembly according to any one of the preceding claims, comprising 10 to 100000, 10 to 50000, 10 to 10000, 10 to 10000, 10 to 8000, 10 to 3000, 10 to 1000, 10 to 800, 10 to 600, 10 to 400, 10 to 200, 10 to 100, 10 to 80 or 10 to 60 conductive surfaces.

7. The component according to any one of the preceding claims, wherein, The conductive surface and / or the insulating surface is substantially flat, convex, concave, shaped to include any one or a combination of peaks or valleys.

8. The component according to any one of the preceding claims, wherein, The conductive material is a layer disposed on a substrate, and the electrically insulating material is a layer disposed on the conductive material layer, wherein the insulating material layer is discontinuous to expose regions of the conductive material layer providing the at least one conductive surface.

9. The component according to any one of the preceding claims, comprising a plurality of conductive surfaces, wherein, The perimeters of each of the conductive surfaces are separated from each other by at least five times the distance of the diameter or minimum width across any one of the conductive surfaces.

10. The assembly according to any one of the preceding claims, further comprising an analyte-responsive layer disposed on the conductive surface.

11. The component according to claim 10, wherein, The analyte-responsive layer comprises any one of the following: a material configured to bind the target analyte; an enzyme material that interacts or chemically reacts with the target analyte.

12. The assembly according to claim 10 or 11, comprising a plurality of analyte-responsive layers, each analyte-responsive layer comprising a different material to bind different respective target analytes.

13. The assembly according to claim 12, comprising a plurality of conductive surfaces, each conductive surface comprising one of the different materials.

14. An electrochemical sensor, comprising: At least one working electrode, the working electrode comprising an electrode assembly according to any one of claims 1 to 13; and at least one auxiliary electrode.

15. The sensor according to claim 14, further comprising: A potentiostat, the potentiostat being electrically connected to the working electrode and the auxiliary electrode to apply a potential to the working electrode and / or measure the potential between the working electrode and one of the auxiliary electrodes; and A control utility, the control utility being configured to receive charge, current, and / or charge or current data generated by oxidation or reduction of the analyte or a reaction product of the target analyte at the working electrode, and / or to measure the potential at the electrode.

16. The sensor according to claim 14 or 15, further comprising an electrochemical potential module configured to generate the potential and apply the potential to the working electrode.

17. The sensor according to claim 16, wherein, The electrochemical potential module is configured to apply a pulsed potential to the working electrode.

18. An implantable device comprising a sensor according to any one of claims 14 to 17.

19. A wearable device attachable to the body of a human or animal, comprising a sensor according to any one of claims 14 to 17.

20. A method of manufacturing an electrode assembly, the electrode assembly being an electrode assembly according to any one of claims 1 to 13.

21. A method of manufacturing an electrochemical sensor, the electrochemical sensor being an electrochemical sensor according to any one of claims 14 to 17.

22. An electrochemical method of interrogating at least one target analyte in a biological fluid, comprising: Providing an electrochemical sensor according to any one of claims 14 to 17; Contacting the sensor with a body containing a biological fluid comprising at least one target analyte; Applying an oxidation or reduction potential at the working electrode to oxidize or reduce the target analyte or a reaction product of the target analyte; and Measuring the current generated by oxidation or reduction of the target analyte or a reaction product of the target analyte between the working electrode and the auxiliary electrode.

23. The method according to claim 22, wherein, The step of applying an oxidation or reduction potential includes performing any one of voltammetry, potentiometry, or amperometry.

24. The method according to claim 22 or 23, comprising applying a potential for a predetermined period of time.

25. The method according to claim 22 or 23, comprising applying a pulsed potential at the working electrode having a period of a first potential and a period of a second potential that is zero or lower than the first potential.

26. An electrochemical method of interrogating at least one target analyte in a biological fluid, comprising: Providing an electrochemical sensor having an electrode assembly comprising: an electrically insulating material having at least one insulating surface; a conductive material having at least one conductive surface constrained and surrounded by the insulating surface; Contacting the sensor with a body containing a biological fluid comprising at least one target analyte; Applying an oxidation or reduction potential at the working electrode to oxidize or reduce the target analyte or a reaction product of the target analyte; and Measuring the current generated between the working electrode and the auxiliary electrode due to the oxidation or reduction of the target analyte or the reaction product of the target analyte wherein the surface area of the conductive surface relative to the surface area of the surrounding insulating surface or, when the assembly includes a plurality of conductive surfaces, the surface area of each conductive surface is configured to control the diffusion rate of the target analyte to the conductive surface in the biological fluid such that the volumetric consumption rate of the target analyte at the conductive surface is less than the diffusion rate of the same volume of the target analyte to the conductive surface in the biological fluid, thereby maintaining the supply by diffusion of the target analyte at the conductive surface.

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