Improving oxygen delivery to implantable sensors

Abstract

Embodiments of the present invention provide multilayer analyte sensors having elements and / or structures for improving oxygen delivery to a sensor enzyme in a manner that enhances sensor functionality, and methods for making and using such sensors. Typical embodiments of the present invention include glucose sensors for diabetes management.

Description

Technical Field

[0001] The present invention relates to methods, materials and components that can be used in analyte sensor systems, such as glucose sensors for diabetes management. Background Art

[0002] Sensors are used to monitor a variety of compounds in various environments, including analytes in the body. Quantitative determination of analytes in the human body is of great importance in the diagnosis and maintenance of many pathological conditions. Exemplary analytes that are commonly monitored in a large number of individuals include glucose, lactate, cholesterol, and bilirubin. Determination of glucose concentration in body fluids is particularly important for diabetic individuals who must frequently check the glucose levels in their body fluids to regulate the glucose intake in their diet. The results of such tests are crucial for determining how much (if any) insulin and / or other drugs need to be used.

[0003] Analyte sensors typically include components that convert the interaction with the analyte into a detectable signal, which can be associated with the concentration of the analyte. For example, some glucose sensors use amperometry to monitor glucose in vivo. This amperometric glucose sensor typically includes an electrode coated with glucose oxidase, which is an enzyme that catalyzes the reaction between glucose and oxygen to produce gluconic acid and hydrogen peroxide (H2O2). The H2O2 formed in the reaction changes the electrode current to form a detectable and measurable signal. Based on this signal, the glucose concentration in an individual can be measured.

[0004] A typical electrochemical glucose sensor works based on the following chemical reaction:

[0005]

[0006] H2O2→O2+2H + +2e - Reaction 2

[0007] Glucose oxidase is used to catalyze the reaction between glucose and oxygen to produce gluconic acid and hydrogen peroxide, as shown in reaction equation 1. H2O2 reacts electrochemically, as shown in reaction equation 2, and the current is measured by a potentiostat. The stoichiometry of the reaction poses a challenge to the development of in vivo sensors. In particular, to obtain optimal glucose oxidase-based sensor performance, the sensor signal output should be determined solely by the target analyte (glucose) and not by any co-substrate (O2) or kinetic control parameters such as diffusion. If oxygen and glucose are present in equimolar concentrations, then H2O2 is stoichiometrically related to the amount of glucose that reacts with glucose oxidase; and the associated current that produces the sensor signal is proportional to the amount of glucose that reacts with the enzyme. However, if there is not enough oxygen for all the glucose to react with the enzyme, then the current will be proportional to the oxygen concentration, not the glucose concentration. Therefore, in order for a glucose sensor to provide a signal that depends only on the glucose concentration, glucose must be the limiting reagent, that is, the O2 concentration must be in excess for all potential glucose concentrations. However, one problem with using such glucose sensors in vivo is that the oxygen concentration at the site where the sensor is implanted is low relative to glucose, a phenomenon that can compromise the accuracy of the glucose sensor readings (and thus, this phenomenon is referred to as the "oxygen starvation problem").

[0008] In view of problems such as those described above (eg, oxygen deficiency problems), there is a need in the art for electrochemical sensors whose structures and materials are selected to avoid oxygen deficiency problems and facilitate sensor function. Embodiments of the invention disclosed herein meet this need and other needs. Summary of the invention

[0009] Embodiments of the invention disclosed herein provide layered electrochemical glucose sensor designs, which include sensor materials, elements and architectures that contribute to sensor function (such as sensor enzyme stoichiometry and / or sensor real-time operation). As noted above in the discussion of the oxygen deficiency problem, sensor enzyme stoichiometry is a key parameter for optimal operation of implantable sensors that use glucose oxidase to measure glucose concentration. Sensor break-in time (or start-up time) is another key parameter for optimal operation of these types of implantable sensors. Fast sensor break-in time is very desirable because it allows the sensor to quickly provide relevant data for diagnosis and treatment of the disease. As discussed in detail below, we have designed many sensor elements, materials and architectures that improve the oxygen delivery and break-in time of the sensor to the oxidase disposed therein. For example, embodiments of the present invention include sensors designed to include "micropores" or "through holes" and structures that create paths / channels for faster diffusion of oxygen to the sensor electrodes and enhanced break-in time and oxygen delivery to the enzyme layer. Embodiments of the present invention include sensors that include high oxygen permeability and oxygen conductive materials (e.g., materials disposed on the exterior of the sensor and / or materials disposed within the micropores or through-holes of the sensor) that can further enhance oxygen delivery to the sensor enzyme layer. Embodiments of the present invention include sensors that include an oxygen generating electrode that can generate oxygen continuously or on demand when the oxygen concentration is below a set level.

[0010] The invention disclosed herein has many embodiments. Embodiments of the invention include, for example, an electrochemical analyte sensor comprising a substrate, a working electrode disposed on the substrate, and a multilayer analyte sensor stack disposed on the working electrode. In such embodiments, the multilayer analyte sensor stack includes at least one analyte sensing layer disposed directly on the working electrode, wherein the analyte sensing layer detectably changes the current at the working electrode in the presence of an analyte; and an analyte modulating layer disposed on the analyte sensing layer, wherein the analyte modulating layer promotes diffusion of O2 from an external environment to the analyte sensing layer. In such embodiments, the electrochemical analyte sensor is designed to include one or more channels disposed in the electrochemical analyte sensor so as to operably couple the analyte sensing layer to an external environment, such that the plurality of channels promote diffusion of O2 from the external environment to the analyte sensing layer.

[0011] Embodiments of the present invention include electrochemical analyte sensors having materials and component groups that facilitate the delivery of O2 to an oxidase (e.g., glucose oxidase) disposed within the sensor. For example, as shown in the relevant figures, one or more channels designed to form a conduit between an O2 source and an oxidase are disposed in the electrochemical analyte sensor in various positions and configurations. Typically, in such embodiments, at least one channel forms an O2 conduit from the external environment on the first side of the sensor through the substrate and the electrode and to the analyte sensing layer. In some embodiments of the present invention, the electrochemical analyte sensor also includes an oxygen diffusion composition disposed within a plurality of channels, wherein the oxygen diffusion composition is selected to promote O2 diffusion therethrough. In addition, some embodiments of the electrochemical analyte sensor disclosed herein also include one or more silicon bridges having a first portion operably contacted with air in an in vitro environment and a second portion operably contacted with at least one channel, wherein the silicon bridge promotes O2 diffusion from air to the oxidase in the analyte sensing layer. In certain embodiments of the present invention, the electrochemical analyte sensor includes an oxygen generating electrode, wherein the oxygen generating electrode is disposed in the sensor so that the oxygen generated by the electrode is operably contacted with the analyte sensing layer. In some embodiments of the present invention, the oxygen generating electrode is also used as a counter electrode in an electrochemical analyte sensor. Optionally, the oxygen generating electrode is in operative contact with one or more channels and / or bridges that operatively couple the analyte sensing layer to the oxygen generating electrode.

[0012] Other embodiments of the present invention include methods of making the electrochemical analyte sensors disclosed herein. Generally, such methods include: providing a substrate; forming a conductive layer on the substrate, wherein the conductive layer includes a working electrode; forming an analyte sensing layer on the conductive layer, wherein the analyte sensing layer includes a composition capable of changing the current at the working electrode in the conductive layer in the presence of an analyte; forming an analyte regulating layer on the analyte sensing layer; and forming one or more channels within the electrochemical analyte sensor in the region of the electrochemical analyte sensor so as to operably couple the analyte sensing layer to an external environment, wherein a plurality of channels promote diffusion of O2 from the external environment to the analyte sensing layer, thereby making an electrochemical analyte sensor. In certain embodiments of the present invention, additional elements (such as a silicon bridge and / or an oxygen generating electrode) are provided in the sensor to improve oxygen delivery to an enzyme (such as glucose oxidase) provided in the analyte sensing layer.

[0013] Other embodiments of the present invention are methods of sensing an analyte in a mammal. Generally, these methods include implanting an electrochemical analyte sensor disclosed herein into a mammal; sensing a change in current at a working electrode in the presence of an analyte; and then correlating the change in current with the presence of the analyte, thereby sensing the analyte.

[0014] For those skilled in the art, other objects, features and advantages of the present invention will become apparent from the following detailed description. However, it is to be understood that, although some embodiments of the present invention are indicated, detailed description and specific examples are provided by way of illustration and not limitation. Without departing from the spirit of the present invention, many changes and modifications within the scope of the present invention may be made, and the present invention includes all such modifications. BRIEF DESCRIPTION OF THE DRAWINGS

[0015] Figure 1 Bottom view schematics of cross sections of the following sensors are provided: a conventional / nominal sensor (left and upper right); a sensor having a channel (e.g., formed by laser ablation) disposed in an electrochemical analyte sensor so as to operably couple the analyte sensing layer to an external environment (middle right); and a sensor having a channel in which an oxygen diffusion component is disposed (lower right).

[0016] Figure 2A-2B A schematic diagram is provided showing a conventional (prior art) sensor design comprising an amperometric analyte sensor formed from a plurality of planar layered elements including an albumin layer and an adhesion promoter layer ( Figure 2A ); and a schematic diagram of the difference between a conventional multilayer sensor stack and a sensor stack with a high-density amine layer ( Figure 2B ).

[0017] Figure 3 Perspective views are provided illustrating a subcutaneous sensor insertion kit, a telemetric characteristic monitor transmitter device, and a data receiving device embodying features of the present invention.

[0018] Figure 4 Schematic diagram of a potentiostat that can be used to measure current in an embodiment of the present invention is shown. Figure 4 As shown, the potentiostat 300 may include an operational amplifier 310 connected in a circuit to have two inputs: Vset and Vmeasure. As shown, Vmeasure is a measurement of the voltage between the reference electrode and the working electrode. On the other hand, Vset is the optimal desired voltage across the working electrode and the reference electrode. The current between the counter electrode and the reference electrode is measured, thereby creating a current measurement result (isig) output from the potentiostat.

[0019] Figures 5A-5D A bottom view schematic diagram of a sensor embodiment having different element populations disclosed herein is shown. Figure 5AA bottom view schematic diagram showing a rear view (left), a top view (middle), and a side view (left) of a sensor embodiment having an air channel / conduit formed by a silicon bridge on the back side of a sensor substrate / flexure and having a first portion operably contacting air in an external extracorporeal environment and a second portion operably contacting at least one channel such that the silicon bridge facilitates diffusion of O2 from the air to an analyte sensing layer. Figure 5B A bottom view schematically illustrates a rear view (left), a top view (middle), and a side view (left) of a sensor embodiment having rear and front air channels / conduits formed by a silicon bridge and having a first portion operably contacting air in an external extracorporeal environment and a second portion operably contacting at least one channel such that the silicon bridge facilitates diffusion of O2 from the air to an analyte sensing layer. Figure 5C A bottom view schematically illustrates a rear view (left), a top view (middle), and a side view (left) of a sensor embodiment having rear and front air channels / conduits formed by a silicon bridge and having a first portion operably contacting air in an external extracorporeal environment and a second portion operably contacting at least one channel such that the silicon bridge facilitates diffusion of O2 from the air to an analyte sensing layer. Figure 5D A bottom view schematically illustrates a rear view (left), a top view (middle), and a side view (left) of a sensor embodiment having rear and front air channels / conduits formed by a silicon bridge and having a first portion operably contacting air in an external extracorporeal environment and a second portion operably contacting at least one channel such that the silicon bridge facilitates diffusion of O2 from the air to an analyte sensing layer. Figure 5E Bottom view schematics showing a back view (left), top view (center), and side view (left) of a sensor embodiment having back and front side air channels / conduits formed by silicon bridges to form air channels from the silicone composition on the back side of the sensor flexure to the analyte sensing layer (in this embodiment, the analyte sensing layer includes glucose oxidase (GOx)).

[0020] Figure 6A-6B Data graphs comparing sensor signals observed in sensors designed to include an oxygen generating electrode to sensor signals observed in sensors that do not include an oxygen generating electrode are provided. Fig. 6A Shown are the graphical sensor signal (in the form of current) obtained from a sensor designed to include an oxygen generating electrode, the sensor signal observed in a sensor that does not include an oxygen generating electrode ("control"), and the signal from an oxygen meter. Figure 6BShown is the break-in time (in hours) for sensors designed to include micropores / channels that operably couple the analyte sensing layer to the external environment (better break-in time, comparable O2 response, higher isig, and lower impedance were observed compared to control sensors without such micropores / channels).

[0021] Figure 7 A schematic diagram of a bottom view of a sensor embodiment with an electrode dedicated to oxygen generation is shown. In this embodiment, a first potentiostat is used to measure glucose in a glucose measurement sensor, which includes a working electrode (WE), a reference electrode (RE), and a counter electrode (CE); and a second potentiostat is used to generate oxygen in a counter electrode sensor including a working electrode (WE), a reference electrode (RE), and a counter electrode (CE). DETAILED DESCRIPTION

[0022] Unless otherwise defined, all technical terms, symbols and other scientific terms or technical terms used herein are intended to have the meanings commonly understood by those skilled in the art to which the present invention belongs. In some cases, for clarity and / or for ease of reference, the terms with commonly understood meanings may be defined herein, and the inclusion of these definitions herein should not necessarily be interpreted as representing that there are substantial differences with the meanings commonly understood in the art. Many techniques and procedures described or referenced herein are well understood and commonly used by those skilled in the art using conventional methods. As required, unless otherwise noted, procedures involving the use of commercially available kits and reagents are generally performed according to the schemes and / or parameters defined by the manufacturer. Many terms are defined below.

[0023] All numbers described in the specification and related claims refer to values ​​that can be numerically characterized by values ​​other than integers (e.g., the diameter of a disk), and should be understood to be modified by the term "about". In the case of providing a range of values, it should be understood that each intermediate value between the upper and lower limits of the range, unless otherwise explicitly stated, to one tenth of the lower limit unit, and any other specified values ​​or intermediate values ​​in the range are included in the present invention. The upper and lower limits of these smaller ranges can be independently included in smaller ranges and are also included in the present invention, subject to any explicitly excluded limits in the stated ranges. When the stated range includes one or two limits, the range excluding any one or both of those included limits is also included in the present invention. In addition, all publications mentioned herein are incorporated herein by reference, thereby disclosing and describing methods and / or materials related to the cited publications. The publications cited herein are cited for their disclosures before the filing date of this application. Nothing herein shall be interpreted as admitting that the inventor has no right to advance the publication date by virtue of an earlier priority date or a priority invention date. In addition, the actual publication date may be different from the date shown and needs to be verified separately.

[0024] As used herein, the term "analyte" is a broad term and is used in its ordinary sense, including but not limited to referring to a substance or chemical component in a fluid such as a biological fluid (e.g., blood, interstitial fluid, cerebrospinal fluid, lymph, or urine) that can be analyzed. The analyte may include naturally occurring substances, artificial substances, metabolites, and / or reaction products. In a common embodiment, the analyte is glucose. However, embodiments of the present invention can be used with sensors designed to detect a variety of other analytes. Illustrative analytes include but are not limited to lactate and naturally occurring salts, sugars, proteins, fats, vitamins, and hormones in the body (e.g., in blood or interstitial fluid). The analyte may be naturally present in a biological fluid or be endogenous; for example, metabolites, hormones, antigens, antibodies, etc.

[0025] As discussed in detail below, embodiments of the present invention relate to the use of electrochemical analyte sensors that measure the concentration of a target analyte or an analyte in an indicating fluid or the concentration of a substance present. In some embodiments, the sensor is a continuous device, such as a subcutaneous, percutaneous, or intravascular device. In some embodiments, the device can analyze multiple intermittent blood samples. The sensor embodiments disclosed herein can use any known method, including invasive, minimally invasive, and non-invasive sensing techniques, to provide an output signal indicating the concentration of the target analyte. The product is then measured using an electrochemical method, and the output of the electrode system is therefore used as a measure of the analyte.

[0026] Embodiments of the invention disclosed herein provide sensors of the type used, for example, for subcutaneous or transcutaneous monitoring of blood glucose levels in diabetic patients. Various implantable electrochemical biosensors have been developed to treat diabetes and other life-threatening diseases. Many existing sensor designs use some form of immobilized enzyme to achieve their biospecificity. The embodiments of the invention described herein may be applied and implemented in a variety of known electrochemical sensors, including, for example, U.S. Patent Application No. 20050115832, U.S. Patent Nos. 6,001,067, 6,702,857, 6,212,416, 6,119,028, 6,400,974, 6,595,919, 6,141,573, 6,122,536, 6,512,939, 5,605,152, 4,431,004, 4,703,756, 6,514,718, 5,985,129, 5,390,691, 5,391,250, 5,482,473, 5,299,571, 5,568,806, 5,494,562, 6,120,676, 6,542,765, 7,033,336 and PCT International Publication No. WO 01 / 58348, WO 04 / 021877, WO 03 / 034902, WO 03 / 035117, WO 03 / 035891, WO 03 / 023388, WO 03 / 022128, WO 03 / 022352, WO 03 / 023708, WO 03 / 036255, WO 03 / 036310, WO 08 / 042,625 and WO 03 / 074107, and European Patent Application No. EP 1153571, the contents of each of which are incorporated herein by reference.

[0027] Illustrative embodiments of the invention and related features

[0028] Embodiments of the present invention disclosed herein provide sensors, which are designed to include a multilayer analyte sensor stack formed by selected materials and related elements, which provides enhanced functions and / or material properties for the sensor. The present disclosure also provides a method for manufacturing and using such a sensor. As discussed in detail below, a typical embodiment of the present invention relates to the use of a sensor, which measures the concentration of a target aqueous analyte (e.g., glucose) or indicates the concentration of an analyte concentration or a substance present in the body. In certain embodiments, the sensor is a subcutaneous, intramuscular, intraperitoneal, intravascular or transcutaneous device. Typically, the sensor can be used for continuous analyte monitoring. The sensor embodiments disclosed herein can use any known method, including invasive, minimally invasive and non-invasive sensing techniques, to provide an output signal indicating the concentration of a target analyte.

[0029] In typical embodiments of the present invention, the components of the multilayer analyte sensor element are formed by selected layers / materials and / or are operably coupled to additional elements and / or are arranged in a stacked structure in a specific direction, which is intended to provide these sensors with improved oxygen delivery to the oxidase arranged in the sensor. The present disclosure also provides methods for making and using such sensors. As discussed in detail below, typical embodiments of the present invention relate to the use of sensors that measure the concentration of a target aqueous analyte or indicate the concentration of an analyte in the body or the concentration of a substance present (e.g., a glucose sensor for diabetes management).

[0030] The invention disclosed herein has many embodiments. Embodiments of the invention include, for example, an electrochemical analyte sensor comprising a substrate (e.g., a polyimide carrier, optionally as part of a flexible circuit assembly), a working electrode disposed on the substrate, and a multilayer analyte sensor stack disposed on the working electrode. In such embodiments, the multilayer analyte sensor stack includes at least one analyte sensing layer disposed directly on the working electrode, wherein the analyte sensing layer detectably changes the current at the working electrode in the presence of an analyte; and an analyte modulating layer disposed on the analyte sensing layer, wherein the analyte modulating layer promotes diffusion of O2 from an external environment (such as interstitial fluid or blood) to the analyte sensing layer (and optionally limits the diffusion of glucose). In such embodiments, the electrochemical analyte sensor is designed to include one or more channels disposed in the electrochemical analyte sensor so as to operably couple the analyte sensing layer to O2 in the external environment, wherein multiple channels promote diffusion of O2 from the external environment to the analyte sensing layer.

[0031] Embodiments of the present invention include electrochemical analyte sensors having a variety of materials and component groups. For example, as shown in the relevant figures (see, for example, Figure 1 and 5A-5D), one or more channels can be disposed in an electrochemical analyte sensor in a variety of positions and configurations. Typically, in such embodiments, at least one channel forms an O2 conduit from an external environment (e.g., interstitial fluid) on a first side of the sensor through the substrate and electrodes and to the analyte sensing layer. In some embodiments of the present invention, the electrochemical analyte sensor also includes an oxygen diffusion composition disposed within the plurality of channels, wherein the oxygen diffusion composition is selected to promote diffusion of O2 therethrough (and optionally limit diffusion of glucose). Sensors that typically include such channels exhibit improved properties, such as reduced break-in time. For example, if an analyte sensor exhibits a long break-in time (e.g., multiple hours), these types of such sensors may not be able to diagnose or treat medical emergencies within the break-in time. In this context, a working prototype of such a sensor was built in-house and tested in vitro and in vivo (in animals). Data from the prototype sensor indicated a significant improvement in the break-in time of the sensor (see, e.g., Figure 6B Data shown). Thus, sensors with the component population disclosed herein achieve better sensor break-in time and first day performance. In addition, hypoxia is considered a key factor in sensor sensitivity loss. Sensor embodiments disclosed herein improve the local oxygen concentration in the enzyme environment and in this way improve sensor sensitivity / lifetime. Thus, such sensors exhibit enhanced sensor life and minimized sensitivity loss.

[0032] In addition, some embodiments of the electrochemical analyte sensors disclosed herein further include one or more silicon bridges having a first portion in operable contact with air in an in vitro environment and a second portion in operable contact with at least one channel, wherein the silicon bridges are located in the sensor and are disposed in a structure designed to facilitate diffusion of O2 from the air to the analyte sensing layer (see, e.g. Figures 5A-5D ). Some embodiments disclosed herein further include one or more silicon bridges having a first portion in operable contact with air in an in vitro environment and a second portion in operable contact with at least one channel, wherein the second silicon bridge facilitates diffusion of O2 from the air to the analyte sensing layer.

[0033] The silicone bridge can be formed from a variety of materials known in the art (see, for example, AS Carpenter and DFT Wiss, Rubber Chemistry and Technology, 13, 326 (1940), Oxygen permeability of silicone hydrogel contact lens materials, Morgan, Philip & Efron, Nathan & D. Cameron, Ian & Brennan, Noel & Goodwin and Marie (2006) and Evolution of Silicone Hydrogel Lenses Contact Lens Spectrum, (June 2008). Silicone hydrogels (rather than pure silicone rubber) are generally the preferred material for forming the bridge element because such hydrogels can transport both oxygen and water. Many such silicone hydrogel formulations are used in the contact lens industry, and these formulations primarily modify the polysiloxane chains to introduce hydrophilic functional groups to assist in the transport of O2 and water. Such silicone hydrogel modifications include modification of polysiloxane chains with functional groups such as N-vinyl pyrrolidone, polyethylene glycol, N,N-dimethylacrylamide, and 2-hydroxyethyl methacrylate. The following table lists some illustrative materials that can be used to form the silicone bridges.

[0034]

[0035]

[0036] Without being limited by a particular theory or mechanism of action, in certain embodiments of the present invention, the materials used for the bridge are designed so that free volume or "pores" exist in the bridge / matrix material. In these materials, as the polymer chains move, the "pores" are formed and disappear in a thermal manner. Gases are soluble in such materials. When the material is exposed to the gas, a solution will appear on the surface and the dissolved gas molecules will diffuse into the interior. The diffusion of gas molecules in the material is the process of gas molecules migrating from the "pores" (free volume) to the "pores" (free volume). For example, the permeation of a gas through a bridge material / membrane may include a solution on one side, diffusing through the bridge material / membrane to the other side, and finally evaporating from the bridge material / membrane.

[0037] As shown herein (see, e.g. Figures 5A-5E), embodiments of the present invention may have a variety of element groups. For example, embodiments of the present invention include an electrochemical analyte sensor comprising a substrate, a working electrode disposed on the substrate, and a multilayer analyte sensor stack disposed on the working electrode. The multilayer analyte sensor stack may include an analyte sensing layer disposed directly on the working electrode, wherein the analyte sensing layer detectably changes the current at the working electrode in the presence of an analyte. The multilayer analyte sensor stack may also include an analyte modulation layer disposed above the analyte sensing layer, wherein the analyte modulation layer promotes diffusion of O2 from an external environment to the analyte sensing layer. In such sensor embodiments, one or more channels are disposed in the electrochemical analyte sensor architecture, which operably couples the analyte sensing layer to an external environment on opposite sides of the sensor (e.g., in a sensor comprising a first side and a second side), such that the plurality of channels promote diffusion of O2 from the external environment to the analyte sensing layer. Other embodiments of the present invention include an electrochemical analyte sensor comprising a substrate, a working electrode disposed on the substrate; and a multilayer analyte sensor stack disposed on the working electrode, wherein the analyte sensing layer is disposed directly on the working electrode, and the analyte sensing layer detectably changes the current at the working electrode in the presence of an analyte. The layered element in this embodiment also includes an oxygen transport layer disposed above the back side of the base layer, wherein the oxygen transport layer promotes diffusion of O2 from the external environment to the analyte sensing layer; and one or more channels disposed in the electrochemical analyte sensor architecture, which operably couples the analyte sensing layer to the external environment, wherein multiple channels promote diffusion of O2 from the external environment to the analyte sensing layer.

[0038] In order to reliably maintain the oxygen synergy required for the operation of an enzyme sensor based on an oxidase (e.g., glucose oxidase used in a glucose sensor), an oxygen generating electrode can be used in close association with the enzyme working electrode. This oxygen generating electrode can provide the oxygen required for sensor operation in the absence of endogenous oxygen (see, e.g., Fig. 6AData from working examples of the present invention are shown). In this context, in typical embodiments of the present invention, the electrochemical analyte sensor includes an oxygen generating electrode, wherein the oxygen generating electrode is arranged in the sensor so that the oxygen generated by the electrode is in operative contact with the analyte sensing layer. Typically, the oxygen generating electrode is in operative contact with one or more channels that operatively couple the analyte sensing layer to the oxygen generating electrode. In some embodiments of the present invention, the oxygen generating electrode is also used as a counter electrode in the electrochemical analyte sensor. Such an oxygen generating electrode can be designed to generate oxygen continuously, or to generate oxygen on demand when the oxygen concentration is below a set level. Optionally, the electrode can be made of a (precious) metal (Pt, Pd, Ru) electrode that can generate oxygen electrolytically or by some other oxidation or reduction process. In working embodiments of such sensors, it was observed that the sensor oxygen sensitivity was significantly improved when oxygen reached 1% and below, and it was observed that the sensor signal using the oxygen generating electrode changed less than the sensor signal without the oxygen generating electrode.

[0039] Other embodiments of the present invention include methods for making an electrochemical analyte sensor disclosed herein. Generally, such methods include: providing a base layer; forming a conductive layer on the base layer, wherein the conductive layer includes a working electrode; forming an analyte sensing layer on the conductive layer, wherein the analyte sensing layer includes a composition capable of changing the current at the working electrode in the conductive layer in the presence of an analyte; forming an analyte regulating layer on the analyte sensing layer; and forming one or more channels within the electrochemical analyte sensor in the region of the electrochemical analyte sensor so as to operably couple the analyte sensing layer to an external environment, wherein a plurality of channels promote diffusion of O2 from the external environment to the analyte sensing layer, thereby making an electrochemical analyte sensor. In this context, improved run-in (sensor start-up) time and O2 delivery to an enzyme (such as glucose oxidase) can be achieved by creating a path for the analyte to diffuse through an oxidase in the analyte sensing layer through a sensor base element (e.g., on the back of the sensor). A person skilled in the art can use methods such as drilling or laser processing (e.g., under ultraviolet light at 355nm) to create through-holes / channel holes on sensor elements such as working electrodes. Typically, the through-hole / channel micropore diameter is between ˜0.5 μm and ˜200 μm (eg, between ˜10 μm and ˜100 μm, ˜80 μm, etc.) to facilitate delivery of such O 2 to the sensor enzyme (eg, glucose oxidase).

[0040] In certain methods of making the sensors disclosed herein, additional elements (such as a silicon bridge and / or an oxygen generating electrode) are provided in the sensor to improve oxygen delivery to an enzyme (such as glucose oxidase) provided in the analyte sensing layer (see, e.g. Figures 5A-5D). In some embodiments of the present invention, such a silicon bridge has a first portion that is operably in contact with air in an in vitro environment and a second portion that is operably in contact with at least one channel, wherein such a silicon bridge promotes diffusion of O2 from the air to the analyte sensing layer. In some embodiments of the present invention, such a silicon bridge has a first portion that is operably in contact with O2 in an in vitro environment (e.g., interstitial fluid) and a second portion that is operably in contact with at least one channel, wherein such a silicon bridge promotes diffusion of O2 from the in vitro environment to the analyte sensing layer. Some embodiments of the present invention also include at least one additional element disclosed herein, such as: a layer comprising a high-density amine layer comprising a poly-L-lysine polymer having a molecular weight between 30KDa and 300KDa; a layer comprising albumin; a layer comprising a silicone adhesion promoter; or a layer comprising glutaraldehyde.

[0041] Method embodiments of the present invention also include placing an oxygen diffusion composition in one or more channels, wherein the oxygen diffusion composition is selected to promote diffusion of O2 therethrough. Certain embodiments of the present invention include providing a silicon bridge in an electrochemical analyte sensor such that a first portion of the silicon bridge is in operably contact with air in an in vitro environment, and a second portion of the silicon bridge is in operably contact with at least one channel, such that the first silicon bridge promotes diffusion of O2 from air to the analyte sensing layer. Optionally, the silicon bridge is provided in the electrochemical analyte sensor such that a portion is in contact with interstitial fluid when the electrochemical analyte sensor is implanted in the body. In addition, embodiments of the present invention may include providing an oxygen generating electrode in the electrochemical analyte sensor, wherein the oxygen generating electrode is provided in the electrochemical analyte sensor such that oxygen generated by the electrode is in operably contact with the analyte sensing layer. Typically, the oxygen generating electrode is provided in the electrochemical analyte sensor so as to be in operably contact with one or more channels, which operably couple the analyte sensing layer to the oxygen generating electrode. Optionally, the oxygen generating electrode is provided in the electrochemical analyte sensor so as to be used as a counter electrode in the electrochemical analyte sensor.

[0042] Other embodiments of the present invention are methods of sensing an analyte in a mammal. Generally, these methods include implanting an electrochemical analyte sensor disclosed herein into a mammal; sensing a change in current at a working electrode in the presence of an analyte; and then correlating the change in current with the presence of the analyte, thereby sensing the analyte.

[0043] In typical embodiments of the invention, the electrochemical sensor is operably coupled to a sensor input capable of receiving a signal from the electrochemical sensor; and a processor coupled to the sensor input, wherein the processor is capable of characterizing one or more signals received from the electrochemical sensor. In certain embodiments of the invention, the electrical conduit of the electrode is coupled to a potentiostat (see, e.g., Figure 4). Optionally, a pulse voltage is used to obtain a signal from the electrode. In a typical embodiment of the invention, the processor is capable of comparing a first signal received from the working electrode in response to a first working potential with a second signal received from the working electrode in response to a second working potential. Optionally, the electrode is coupled to a processor adapted to convert data obtained by observing fluctuations in the current from a first format to a second format. Such embodiments include, for example, a processor designed to convert a sensor current input signal (e.g., ISIG measured in nA) to a blood glucose concentration.

[0044] In many embodiments of the present invention, the sensor includes a biocompatible region suitable for implantation in vivo. In some embodiments, the sensor includes a discrete probe that pierces the in vivo environment. In an embodiment of the present invention, the biocompatible region may include a polymer that contacts the in vivo tissue. Optionally, the polymer is a hydrophilic polymer (e.g., a polymer that absorbs water). In this way, the sensor used in the system of the present invention can be used to sense a variety of analytes in different aqueous environments. In some embodiments of the present invention, an electrode is coupled to a puncture member (e.g., a needle) suitable for implantation in vivo. Although the sensor embodiment of the present invention may include one or two puncture members, optionally, this type of sensor device may include 3 or 4 or 5 or more puncture members, which are coupled to and extend from a substrate element, and are operably coupled to 3 or 4 or 5 or more electrochemical sensors (e.g., microneedle arrays, embodiments of which are disclosed in, for example, U.S. Patents 7,291,497 and 7,027,478, and U.S. Patent Application No. 20080015494, the contents of which are incorporated herein by reference).

[0045] In some embodiments of the invention, the device includes one or more working electrodes, counter electrodes, and reference electrodes, optionally, they are aggregated together in a unit consisting essentially of one working electrode, one counter electrode, and one reference electrode; and the aggregated unit is longitudinally distributed on the base layer in a repeating unit pattern. In some sensor embodiments, the distributed electrodes are organized / set within a flexible circuit assembly (i.e., a circuit assembly that utilizes flexible rather than rigid materials). Such flexible circuit assembly embodiments provide an interconnect assembly of elements (e.g., electrodes, electrical conduits, contact pads, etc.) that is configured to contribute to wearer comfort (e.g., by reducing pad stiffness and wearer discomfort).

[0046] As described above, the sensor electrodes of the present invention are coated with a variety of materials having properties that, for example, facilitate analyte sensing. In a typical embodiment of the present invention, the analyte sensing layer is directly disposed on the working electrode and includes a reagent selected for its ability to detectably change the current at the working electrode in the presence of an analyte. In the working embodiments of the present invention disclosed herein, the reagent is glucose oxidase, a protein that undergoes a chemical reaction in the presence of glucose, the chemical reaction resulting in a change in the current at the working electrode. These working embodiments also include an analyte regulating layer disposed above the analyte sensing layer, wherein the analyte regulating layer regulates the diffusion of glucose when glucose migrates from the in vivo environment to the analyte sensing layer. In certain embodiments of the present invention, the analyte regulating layer comprises a hydrophilic comb copolymer having a central chain and a plurality of side chains coupled to the central chain, wherein at least one side chain comprises a siloxane moiety. In certain embodiments of the present invention, the analyte regulating layer comprises a blended mixture of a linear polyurethane / polyurea polymer and a branched acrylate polymer; and the linear polyurethane / polyurea polymer and the branched acrylate polymer are blended in a ratio of 1:1 to 1:20 (e.g., 1:2) weight %. Typically, the analyte modulating layer composition includes a first polymer formed from a mixture comprising a diisocyanate; at least one hydrophilic diol or hydrophilic diamine; and a siloxane; the first polymer is blended with a second polymer formed from a mixture comprising: 2-(dimethylamino)ethyl methacrylate; methyl methacrylate; monomethacryloxypropyl polydimethylsiloxane; poly(ethylene oxide) methyl ether methacrylate; and 2-hydroxyethyl methacrylate. As disclosed herein, additional material layers may be included in such a device. For example, in some embodiments of the present invention, the device includes a high-density amine layer disposed between and in direct contact with the analyte sensing layer and the analyte modulating layer to exhibit a number of beneficial properties, including the ability to provide a smoother surface structure and further promote adhesion between the analyte sensing layer and the analyte modulating layer. Without being bound by a particular scientific theory or mechanism of action, it is believed that the adhesion between the layers is promoted by: a smoother layer contact architecture, and van der Waals interactions between the HDA polymers in the HDA layer and compounds present in the analyte sensing layer disposed on a first side of the HDA layer, and van der Waals interactions between the HDA polymers and compounds present in the analyte modulating layer disposed on a second side of the HDA layer (i.e., giving the HDA layer a "sandwich" configuration).

[0047] Figure 2AOne prior art conventional sensor embodiment shown in is an amperometric sensor 100 having multiple layered elements, which includes a base layer 102, a conductive layer 104 (e.g., a conductive layer including multiple conductive members) disposed on and / or coupled to the base layer 102. The following comments relate to this conventional sensor, which are described to help understand the differences between such conventional sensors and the present invention disclosed herein. Generally, the conductive layer 104 includes one or more electrodes. An analyte sensing layer 110 (typically including an enzyme, such as glucose oxidase) is disposed on one or more exposed electrodes of the conductive layer 104. A protein layer 116 is disposed on the analyte sensing layer 110. An analyte regulating layer 112 is disposed above the analyte sensing layer 110 to regulate the proximity of an analyte (e.g., glucose) to the analyte sensing layer 110. As Figure 2A As shown, an adhesion promoter layer 114 is disposed between layers such as the analyte modulating layer 112 and the analyte sensing layer 110 to facilitate their contact and / or adhesion. This embodiment also includes a covering layer 106, such as a polymer coating that can be disposed on a portion of the sensor 100. The hole 108 can be formed in one or more layers of this type of sensor. Amperometric glucose sensors with this type of design are disclosed, for example, in U.S. Patent Application Publication Nos. 20070227907, 20100025238, 20110319734, and 20110152654, the contents of each of which are incorporated herein by reference. Figure 2B A comparison between these conventional multilayer sensor stacks including HDA layers is shown.

[0048] Embodiments of the present invention also provide articles and test kits for observing analyte concentrations. In illustrative embodiments, the test kit includes a sensor, and the sensor includes a multilayer sensor stack as described herein. In a typical embodiment, the sensor is arranged in a test kit in a sealed sterile dry package. Optionally, the test kit includes an insertion device that helps the sensor to be inserted. The test kit and / or sensor kit typically include a container, a label and an analyte sensor as described above. Suitable containers include, for example, easy-to-open packaging, bottles, vials, syringes and test tubes made of materials such as metal foil. The container can be formed of a variety of materials, such as metal (e.g., foil) paper products, glass or plastics. The label on or associated with the container indicates that the sensor is used to analyze the selected analyte. The test kit and / or sensor kit may include other materials desired from a commercial and user perspective, including buffer, diluent, filter, needle, syringe and a package insert with instructions for use.

[0049] Specific aspects of embodiments of the invention are discussed in detail in the following sections.

[0050] Typical components, configurations, and analyte sensor embodiments of the present invention

[0051] A. Typical Elements Present in Embodiments of the Invention

[0052] Figure 2A A cross section of a conventional sensor embodiment 100 is shown. Components of the sensor are typically characterized herein as layers of the multilayer electrochemical sensor stack because, for example, it can be easily characterized as Figure 2A The conventional sensor structure shown in Figure 2B The distinction of the present invention disclosed herein is shown in Figure 5 (i.e., a sensor including an HDA layer 500). The skilled person will understand that in certain embodiments of the present invention, the sensor components are combined such that the multiple components form one or more heterogeneous layers. In this context, the skilled person will understand that although certain layers / components of conventional sensor embodiments may be used in the HDA sensors disclosed herein, the arrangement and composition of the layered components are very different in the HDA sensor embodiments of the present invention. The skilled person will understand that in certain embodiments, the present invention includes elements / layers present in conventional sensors, excludes other elements, and / or includes new material layers / elements. For example, Figure 2A Certain elements disclosed in the invention may also be present in the invention disclosed herein (e.g., a base layer, an analyte sensing layer, an analyte modulating layer, etc.), and Figure 2B As shown, other elements are not present (e.g., a separate HSA protein layer, a layer comprising a silicone adhesion promoter, etc.). Similarly, embodiments of the present invention include layers / elements having materials arranged in a unique configuration that are not present in conventional sensors (e.g., a high density amine (HDA) polymer layer 500).

[0053] Figure 2A The conventional embodiment shown includes a substrate 102 that supports the sensor 100. The substrate 102 can be made of a material such as a metal and / or a ceramic and / or a polymer substrate, which can be self-supporting or further supported by another material known in the art. An embodiment may include a conductive layer 104 disposed on and / or combined with the substrate 102. Typically, the conductive layer 104 includes one or more conductive elements used as electrodes. The operating sensor 100 typically includes a plurality of electrodes, such as a working electrode, a counter electrode, and a reference electrode. Other embodiments may also include a plurality of working electrodes and / or counter electrodes and / or reference electrodes and / or one or more electrodes that perform multiple functions, such as an electrode that is used as both a reference electrode and a counter electrode.

[0054] As discussed in detail below, many known techniques and materials can be used to produce the base layer 102 and / or the conductive layer 104. In certain embodiments of the present invention, the circuit of the sensor is defined by etching the conductive layer 104 into a desired conductive path pattern. A typical circuit for the sensor 100 includes two or more adjacent conductive paths having an area at the proximal end to form a contact pad and an area at the distal end to form a sensor electrode. An electrically insulating cover layer 106 such as a polymer coating can be provided on a portion of the sensor 100. Acceptable polymer coatings used as an insulating protective cover layer 106 can include, but are not limited to, non-toxic biocompatible polymers such as silicone compounds, polyimides, biocompatible solder masks, epoxy acrylate copolymers, and the like. In the sensor of the present invention, one or more exposed areas or holes 108 can be formed through the cover layer 106 to open the conductive layer 104 to the external environment and, for example, allow an analyte such as glucose to penetrate the layer of the sensor and be sensed by the sensing element. The hole 108 can be formed by a variety of techniques, including laser ablation, masking with a mask, chemical grinding or etching, or photolithographic development, and the like. In certain embodiments of the present invention, during fabrication, a second photoresist may also be applied to the protective layer 106 to define areas of the protective layer to be removed to form one or more holes 108. The exposed electrodes and / or contact pads may also undergo secondary processing (e.g., through the holes 108), such as additional electroplating processes, to prepare the surface and / or enhance the conductive areas.

[0055] exist Figure 2A In the conventional sensor configuration shown in , the analyte sensing layer 110 is disposed on one or more exposed electrodes of the conductive layer 104. Typically, the analyte sensing layer 110 is an enzyme layer. Most typically, the analyte sensing layer 110 includes an enzyme capable of producing and / or utilizing oxygen and / or hydrogen peroxide, such as glucose oxidase. Optionally, the enzyme in the analyte sensing layer is combined with a second carrier protein such as human serum albumin, bovine serum albumin, etc. In an illustrative embodiment, an oxidoreductase such as glucose oxidase in the analyte sensing layer 110 reacts with glucose to produce hydrogen peroxide, which then regulates the current at the electrode. Since this regulation of the current depends on the concentration of hydrogen peroxide, and the concentration of hydrogen peroxide is related to the concentration of glucose, the concentration of glucose can be determined by monitoring this regulation of the current. In a specific embodiment of the present invention, hydrogen peroxide is oxidized on a working electrode (also referred to herein as an anode working electrode) as an anode, and the resulting current is proportional to the concentration of hydrogen peroxide. This modulation of the current caused by changes in hydrogen peroxide concentration can be monitored by any of a variety of sensor detector devices, such as a Universal Sensor amperometric biosensor detector or one of a variety of other similar devices known in the art, such as the glucose monitoring device produced by Medtronic Diabetes.

[0056] In an embodiment of the present invention, the analyte sensing layer 110 can be applied over a portion of the conductive layer or over the entire area of ​​the conductive layer. Typically, the analyte sensing layer 110 is disposed on a working electrode, which can be a positive electrode or a negative electrode. Optionally, the analyte sensing layer 110 is also disposed on a counter electrode and / or a reference electrode. Methods for producing a thin analyte sensing layer 110 include brushing the layer onto a substrate (e.g., a reactive surface of a platinum black electrode), as well as spin coating processes, dip coating and drying processes, low shear spray processes, inkjet printing processes, screen processes, and the like.

[0057] In this context, a variety of spin coating materials and methods are known in the art (see, for example, Sahu et al., Indian J. Phys. 83(4)493-502 (2009), and U.S. Patent Publications 20020127878, 20020127878, 20090285982, and 20140272704). In certain embodiments of the present invention, a material comprising a high density amine layer of a polymer having multiple repeating amine groups (e.g., a poly-L-lysine polymer) is mixed with another material such as a solvent or other agent that adjusts the viscosity of the solution in order to optimize spin coating uniformity. In this context, in order to prepare a HDA layer for spin coating, a viscosity modifier and / or one or two or more solvents can be mixed together. For example, when two solvents are used, a major component of something that evaporates relatively quickly and a minor component of something that evaporates relatively slowly can be used. By using this combination, it is often possible to optimize various aspects of the process because during the spin coating process, the primary component evaporates quickly to provide good coverage and uniformly thick films, while the remaining secondary component still leaves enough plasticity to organize the molecules before the film is completely dry.

[0058] The analyte modulating membrane layer 112 may include a glucose limiting membrane that modulates the amount of glucose that contacts an enzyme such as glucose oxidase present in the analyte sensing layer. Such glucose limiting membranes may be made of a variety of materials known to be suitable for such purposes, for example, silicone compounds such as polydimethylsiloxane, polyurethane, polyurea acetate cellulose, perfluorosulfonic acid, polyester sulfonic acid (e.g., Kodak AQ), hydrogels, or any other suitable hydrophilic membrane known to those skilled in the art.

[0059] In a typical embodiment of the present invention, Figure 2BAs shown, a material layer including a high-density amine composition layer 500 is disposed between the analyte modulating layer 112 and the analyte sensing layer 110 to facilitate their contact and / or adhesion. In a typical embodiment of the present invention, the high-density amine layer 500 includes a first side in direct contact with the analyte sensing layer and a second side in direct contact with the analyte modulating layer, and serves as an adhesive layer for bonding the analyte sensing layer to the analyte modulating layer. Optionally, the analyte sensing layer includes a glucose oxidase disposed in the layer so that the analyte sensor senses glucose; and the high-density amine layer 500 also has the function of reducing the sensor signal changes caused by fluctuations in oxygen (O2) levels. In certain embodiments of the present invention, the poly-L-lysine in the high-density amine layer 500 has a molecular weight between 30KDa and 300KDa, for example, a molecular weight between 150KDa and 300KDa. Typically, the amount of poly-L-lysine in the high-density amine layer 500 is 0.1 wt% to 0.5 wt%. Optionally, the high-density amine layer 500 is 0.1 to 0.4 microns thick.

[0060] Typical Analyte Sensor Components of the Invention

[0061] The following disclosure provides examples of typical elements / ingredients used in sensor embodiments of the present invention. Although these elements can be described as discrete units (e.g., layers), it is understood by those skilled in the art that the sensor can be designed to include elements having some or all of the material properties and / or combinations of functions of the elements / ingredients discussed below (e.g., the element is used as both a supporting substrate component and / or a conductive component and / or a matrix for an analyte sensing component, and further used as an electrode in the sensor). It is understood by those skilled in the art that selected elements from these thin film analyte sensors can be suitable for many sensor systems, such as those described below.

[0062] Base ingredients

[0063] The sensors of the present invention typically include a substrate component (see, e.g. Figure 2A 102 in the device). The term "substrate component" is used herein according to a term recognized in the art and refers to a component in the device that generally provides a supporting matrix for multiple components stacked on top of each other and including a functional sensor. In one form, the substrate component includes a thin film sheet of insulating (e.g., electrically insulating and / or impermeable) material. This substrate component can be made of a variety of materials with desired qualities (such as dielectric properties, impermeability, and air tightness). Some materials include metals and / or ceramics and / or polymer substrates, etc. In some sensor embodiments, the distributed electrodes are organized / arranged within a flexible circuit assembly (i.e., a circuit assembly that utilizes flexible rather than rigid materials).

[0064] Conductive components

[0065] The electrochemical sensor of the present invention typically includes a conductive component disposed on a substrate component, the conductive component including at least one electrode for contacting an analyte to be analyzed or its byproducts (e.g., oxygen and / or hydrogen peroxide) (see, e.g., Figure 2A 104 in the analyte sensing component 110). The term "conductive component" is used herein according to the recognized terminology in the art. An illustrative example thereof is a conductive component forming a working electrode that can measure an increase or decrease in current in response to exposure to a stimulus (such as a change in concentration of an analyte or its byproducts) compared to a reference electrode that does not experience a change in analyte concentration, the analyte being a co-reactant (such as oxygen) used when the analyte interacts with a composition (such as the enzyme glucose oxidase) present in the analyte sensing component 110, or the analyte is a reaction product of this interaction (such as hydrogen peroxide). Illustrative examples of such elements include electrodes that are capable of generating a variable detectable signal in the presence of a variable concentration of a molecule such as hydrogen peroxide or oxygen.

[0066] In addition to the working electrode, the analyte sensor of the present invention typically also includes a reference electrode or a combined reference and counter electrode (also referred to as a quasi-reference electrode or counter / reference electrode). If the sensor does not have a counter / reference electrode, it may include a separate counter electrode, which may be made of the same or different material as the working electrode. A typical sensor of the present invention has one or more working electrodes and one or more counter, reference and / or counter / reference electrodes. An embodiment of the sensor of the present invention has two, three or four or more working electrodes. These working electrodes in the sensor may be integrally connected or they may also remain separate. Alternatively, the electrode may be arranged on a single surface or one side of the sensor structure. Alternatively, the electrode may be arranged on multiple surfaces or sides of the sensor structure (and, for example, may be connected to the surface on which the electrode is arranged by passing through a through hole of the sensor material). In certain embodiments of the present invention, the reactive surface of the electrode has different relative areas / sizes, such as a 1X reference electrode, a 2.6X working electrode, and a 3.6X counter electrode.

[0067] Analyte sensing components

[0068] The electrochemical sensor of the present invention includes an analyte sensing component disposed on an electrode of the sensor (see, e.g. Figure 2A110 in). The term "analyte sensing component" is used herein according to professional terms recognized in the art, and refers to a component containing a material capable of identifying an analyte or reacting with an analyte, the presence of which will be detected by an analyte sensor device. Typically, this material in the analyte sensing component usually produces a detectable signal via an electrode of a conductive component after interacting with the analyte to be sensed. In this regard, the electrodes of the analyte sensing component and the conductive component work in a combined manner to produce an electrical signal read by a device associated with the analyte sensor. Typically, the analyte sensing component includes an oxidoreductase (e.g., glucose oxidase) that can react with a molecule and / or cause the molecule to produce a concentration change, which can be measured by measuring the current change at the electrode of the conductive component (e.g., oxygen and / or hydrogen peroxide). The enzyme capable of producing a molecule (e.g., hydrogen peroxide) can be placed on an electrode according to many methods known in the art. The analyte sensing component can coat all or part of each electrode of the sensor. In this context, the analyte sensing component can coat the electrode to an equivalent degree. Alternatively, the analyte sensing component may coat different electrodes to different extents, such as the working electrode having a coated surface that is greater than the coated surface of the counter and / or reference electrodes.

[0069] Some sensor embodiments of this element of the invention utilize an enzyme (e.g., glucose oxidase) that has optionally been combined with a second protein (e.g., albumin) in a fixed ratio (e.g., a ratio typically optimized for the stability properties of glucose oxidase) and then applied to an electrode surface to form a thin enzyme component. In a typical embodiment, the analyte sensing component includes a GOx and HSA mixture. In a typical embodiment of an analyte sensing component with GOx, GOx reacts with glucose present in the sensing environment (e.g., the body of a mammal) and produces hydrogen peroxide.

[0070] As mentioned above, the enzyme and the second protein (e.g., albumin) can be treated to form a cross-linked matrix (e.g., by adding a cross-linking agent to the protein mixture). As known in the art, the cross-linking conditions can be controlled to adjust factors such as the retained biological activity of the enzyme, its mechanical and / or operational stability. Illustrative cross-linking procedures are described in U.S. Patent Application Serial No. 10 / 335,506 and PCT Publication WO 03 / 035891, which are incorporated herein by reference. For example, an amine cross-linking agent (such as, but not limited to, glutaraldehyde) can be added to the protein mixture (but in certain embodiments of the present invention disclosed herein, glutaraldehyde is excluded because adding a cross-linking agent to the protein mixture will produce a less active protein paste).

[0071] An alternative embodiment of the analyte sensing component is not formed using glutaraldehyde, but is formed to include a polypeptide that is encapsulated and / or cross-linked, such as glucose oxidase cross-linked with a polyvinyl alcohol (PVA, see, e.g., CAS No. 9002-89-5) polymer. As is known in the art, polyvinyl alcohol reacts with aldehydes to form water-insoluble polyacetals. In a pure PVA medium at a pH of about 5.0, the polymer reacts with the dialdehyde to form an acetal cross-linked structure. In certain embodiments of the present invention, this cross-linking reaction can be carried out using a chemical vapor deposition (CVD) process. Due to the acidity of the PVA polymer solution, the cross-linking reaction in the CVD system is both simple and conventional. In addition, acidic conditions can be created by introducing compounds such as acetic acid into the glutaraldehyde solution, so the CVD system can provide acidic steam conditions. In addition, the pH of the polymer medium can be adjusted by adding acidic compounds such as citric acid, polymer additives such as polylysine, HBr, etc.

[0072] Embodiments of the analyte sensing composition include compositions having properties that make them particularly suitable for use in mobile glucose sensors of the type worn by diabetic individuals. Such embodiments of the present invention include PVA-SbQ compositions for use in layered analyte sensor structures, which contain 1 mol% to 12.5 mol% SbQ. In certain embodiments of the present invention suitable for or used in glucose sensors, the ingredients in the layer are selected so that the molecular weight of the polyvinyl alcohol is between 30 kilodaltons and 150 kilodaltons, and the amount of SbQ present in the polyvinyl alcohol is between 1 mol% and 4 mol%. In some embodiments of the present invention, the analyte sensing layer is formed to contain from 5% to 12% PVA by weight. In some embodiments of the present invention, the analyte sensing layer is formed to contain glucose oxidase in an amount of 10 kU / ml to 20 kU / ml.

[0073] Embodiments of analyte sensing components include analyte sensing layers, which are selected based on their ability to provide desired properties for implantable sensors. In certain embodiments of the present invention, the amount or ratio of PVA in the composition is used to adjust the water adsorption of the composition, the crosslinking density of the composition, etc. The effects of such preparations on phenomena such as H2O adsorption, sensor isig drift, and in vivo startup curves can be easily evaluated. Sufficient H2O adsorption can help maintain normal chemical and electrochemical reactions within amperometric analyte sensors. Therefore, it is desirable to form such sensors from compositions with appropriate hydrophilic chemistry. In this context, the PVA-GOx compositions disclosed herein can be used to produce electrolyte hydrogels that can be used for internal coatings / membrane layers, and can also be coated on top of an analyte regulating layer (e.g., a glucose limiting membrane or "GLM") to improve the biocompatibility and hydrophilicity of the GLM layer.

[0074] As described above, in some embodiments of the present invention, the analyte sensing component includes a reagent (e.g., glucose oxidase) capable of generating a signal (e.g., a change in oxygen and / or hydrogen peroxide concentration), which can be sensed by a conductive element (e.g., an electrode sensing a change in oxygen and / or hydrogen peroxide concentration). However, other useful analyte sensing components can be formed by any composition capable of generating a detectable signal that can be sensed by a conductive element after interacting with a target analyte to be detected for its presence. In some embodiments, the composition includes an enzyme that modulates the concentration of hydrogen peroxide when reacting with the analyte to be sensed. Alternatively, the composition includes an enzyme that modulates the concentration of oxygen when reacting with the analyte to be sensed. In this context, a variety of enzymes that are used or generate hydrogen peroxide and / or oxygen in reactions with physiological analytes are known in the art, and these enzymes can be easily incorporated into the analyte sensing component composition. Various other enzymes known in the art can generate and / or utilize compounds, and the modulation of the compound can be detected by electrodes of a conductive element (e.g., incorporated into the sensor design described herein). Such enzymes include, for example, those described specifically in Table 1, pages 15-29 and / or Table 18, pages 111-112 of Protein Immobilization: Fundamentals and Applications (Bioprocess Technology, Vol 14), by Richard F. Taylor (ed.), Publisher: Marcel Dekker; (January 7, 1991), the entire contents of which are incorporated herein by reference.

[0075] Protein composition

[0076] The electrochemical sensor of the present invention optionally includes a protein component (see, for example, element 116 in FIG. 2 ) disposed between the analyte sensing component and the analyte regulating component. The term "protein component" is used herein according to the specialized term recognized in the art, and refers to a component containing a carrier protein, etc., which is selected to be compatible with the analyte sensing component and / or the analyte regulating component. In a typical embodiment, the protein component includes albumin, such as human serum albumin. The HSA concentration may vary between about 0.5%-30% (w / v). Typically, the HSA concentration is about 1-10% w / v, and most typically about 5% w / v. In alternative embodiments of the present invention, collagen or BSA or other structural proteins used in these contexts may be used to replace or supplement HSA. According to a protocol recognized in the art, this component is typically cross-linked on the analyte sensing component.

[0077] Adhesion promoting ingredients

[0078] The electrochemical sensor of the present invention may include one or more adhesion promoting (AP) components (see, for example, element 114 in FIG. 2 ). The term "adhesion promoting component" is used herein according to a specialized term recognized in the art, and refers to a component that includes a material selected because it can promote adhesion between adjacent components in the sensor. The adhesion promoting component is usually placed between the analyte sensing component and the analyte regulating component. The adhesion promoting component is usually placed between the optional protein component and the analyte regulating component. The adhesion promoter component can be made of any of a variety of materials known in the art to facilitate bonding between such components, and can be applied by any of a variety of methods known in the art. Typically, the adhesion promoter component includes a silane compound, such as γ-aminopropyltrimethoxysilane.

[0079] High density amine ingredients

[0080] The electrochemical sensor of the present invention comprises one or more high-density amine component layers (see, e.g. Figure 2B Element 500 in the sensor), which provides many beneficial functions for the sensor. Such a layer can optimize the sensor function, for example, by acting as an adhesion promoting component of a layer adjacent to the HDA layer, by reducing the fluctuations that may occur in a sensor based on glucose oxidase in the presence of oxygen concentration fluctuations, by improving the sensor initialization configuration, etc. The term "adhesion promoting component" is used herein according to the special words recognized in the art, and refers to such a component, which contains a material selected because it can promote adhesion between adjacent components in the sensor. Generally, the high-density amine adhesion promoting component is arranged between the analyte sensing component and the analyte regulating component, and is in direct contact with the analyte sensing component and the analyte regulating component. In a typical embodiment, the high-density amine layer 500 includes poly-L-lysine with a molecular weight between 30KDa and 300KDa (e.g., between 150KDa and 300KDa). In such a high-density amine layer 500, the concentration of poly-L-lysine is generally 0.1 wt% to 0.5 wt%, and the thickness of the high-density amine layer 500 is 0.1 to 0.4 microns. In an embodiment, the analyte sensing layer includes glucose oxidase so that the analyte sensor senses glucose, and the high-density amine layer 500 functions to reduce sensor signal variations due to fluctuations in oxygen (O 2 ) levels.

[0081] Analyte Modulating Components

[0082] The electrochemical sensors of the present invention include an analyte modulating component disposed on the sensor (see, e.g. Figure 2A112 in). The term "analyte modulating component" is used herein according to professional terms recognized in the art, and refers to a component that typically forms a film on the sensor, which is used to regulate the diffusion of one or more analytes, such as glucose, through the component. In certain embodiments of the present invention, the analyte modulating component is an analyte limiting membrane, the function of which is to prevent or limit the diffusion of one or more analytes (such as glucose) through the component. In other embodiments of the present invention, the analyte modulating component is used to promote the diffusion of one or more analytes through the component. Alternatively, such an analyte modulating component can be formed to prevent or limit the diffusion of one type of molecule through the component (such as glucose), while allowing or even promoting the diffusion of other types of molecules through the component (such as O2).

[0083] Regarding glucose sensors, in known enzyme electrodes, glucose and oxygen from blood and some interferents such as ascorbic acid and uric acid diffuse through the primary membrane of the sensor. When glucose, oxygen and interferents reach the analyte detection component, enzymes (such as glucose oxidase) catalyze the conversion of glucose into hydrogen peroxide and gluconolactone. Hydrogen peroxide can diffuse back through the analyte regulating component, or it can diffuse to the electrode, where hydrogen peroxide can react to form oxygen and protons to produce an electric current proportional to the glucose concentration. The analyte regulating sensor membrane assembly is used for several functions, including selectively allowing glucose access to pass therethrough (see, e.g., U.S. Patent Application No. 2011-0152654).

[0084] Covering ingredients

[0085] The electrochemical sensor of the present invention may include one or more covering components (see, for example, element 106 in FIG. 2 ) that are generally electrically insulating protective components. Typically, such covering components may be in the form of a coating, sheath or tube and are disposed on at least a portion of the analyte modulating component. Typically, such features include a polymer comprising a surface having a feature group disclosed herein, which is used to modulate an immune response. Acceptable polymer coatings used as insulating covering components may include, but are not limited to, non-toxic biocompatible polymers such as silicone compounds, polyimides, biocompatible solder masks, epoxy acrylate copolymers, and the like. In addition, these coatings may be photoimageable to facilitate photolithographic formation of holes through the conductive component. Typical covering components include spinning on silicone. As known in the art, this component may be a commercially available RTV (room temperature vulcanized) silicone composition. In this context, a typical chemical is polydimethylsiloxane (based on acetoxy).

[0086] C. Typical Analyte Sensor System Embodiments of the Present Invention

[0087] Embodiments of sensor elements and sensors can be operably coupled to a variety of other system elements (e.g., structural elements such as puncture members, insertion kits, etc., and electronic components such as processors, monitors, drug infusion pumps, etc.) typically used with analyte sensors, such as to make them suitable for use in various environments (e.g., implanted in mammals). One embodiment of the present invention includes a method for monitoring a user's physiological characteristics using an embodiment of the present invention, the embodiment including an input element, which can receive a signal based on a physiological characteristic value sensed by the user from a sensor, and a processor for analyzing the received signal. In a typical embodiment of the present invention, the processor determines the dynamic behavior of the physiological characteristic value and provides an observable indicator based on the dynamic behavior of the physiological characteristic value so determined. In some embodiments, the physiological characteristic value is a measure of the user's blood glucose concentration. In other embodiments, the method for analyzing the received signal and determining the dynamic behavior includes repeatedly measuring the physiological characteristic value to obtain a series of physiological characteristic values ​​to, for example, incorporate comparative redundancy into the sensor device in a manner designed to provide confirmation information about sensor function, analyte concentration measurement results, the presence of interference, etc.

[0088] Figure 4 Schematic diagram of a potentiostat that can be used to measure current in an embodiment of the present invention is shown. Figure 4 As shown, the potentiostat 300 may include an operational amplifier 310 connected in a circuit to have two inputs: Vset and Vmeasure. As shown, Vmeasure is a measurement of the voltage between the reference electrode and the working electrode. On the other hand, Vset is the optimal desired voltage across the working electrode and the reference electrode. The current between the counter electrode and the reference electrode is measured, thereby creating a current measurement result (isig) output from the potentiostat.

[0089] Embodiments of the present invention include a device for processing display data from measurements of sensed physiological characteristics (e.g., blood glucose concentration) in a manner and format that is customized to allow the user of the device to easily monitor and (when necessary) modulate the physiological state of the characteristic (e.g., modulate blood glucose concentration by insulin administration). An illustrative embodiment of the present invention is a device comprising: a sensor input capable of receiving a signal from a sensor based on a sensed physiological characteristic value of a user; a memory for storing a plurality of measurements of the sensed physiological characteristic value of the user of the received signal from the sensor; and a display for presenting a text and / or graphical representation (e.g., text, line graph, etc., bar graph, etc., grid pattern, etc., or a combination thereof) of a plurality of measurements of the sensed physiological characteristic value. Typically, the graphical representation displays real-time measurements of the sensed physiological characteristic value. Such devices can be used in various situations, such as in combination with other medical devices. In some embodiments of the present invention, the device is used in combination with at least one other medical device (e.g., a glucose sensor).

[0090] The illustrative system embodiment is composed of a glucose sensor, a transmitter and a pump receiver and a glucose meter. In this system, a radio signal from a transmitter can be sent to a pump receiver every 5 minutes to provide a real-time sensor glucose (SG) value. The value / graph is displayed on the monitor of the pump receiver so that the user can monitor blood sugar by themselves and use their own insulin pump to deliver insulin. Typically, the embodiments of the device disclosed herein communicate with a second medical device via a wired or wireless connection. Wireless communication can include, for example, the reception of an emitted radiation signal that occurs when a signal is transmitted by RF telemetry, infrared transmission, optical transmission, sound waves, and ultrasonic transmission. Optionally, the device is an integral part of a drug infusion pump (e.g., an insulin pump). Typically, in such a device, a physiological characteristic value includes multiple measurements of blood sugar.

[0091] Figure 3 A perspective view of a broad embodiment of a subcutaneous sensor insertion system and a block diagram of a sensor electronics device according to an illustrative embodiment of the present invention is provided. Additional elements commonly used with such sensor system embodiments are disclosed, for example, in U.S. Patent Application No. 20070163894, the contents of which are incorporated by reference. Figure 3A perspective view of a telemetric characteristic monitoring system 1 is provided, which includes a subcutaneous sensor kit 10 that is provided for subcutaneous placement of an active portion of a flexible sensor 12 or the like at a selected location on a user's body. The subcutaneous or percutaneous portion of the sensor kit 10 includes a hollow slotted insertion needle 14 having a sharp tip 44 and a cannula 16. Inside the cannula 16 is a sensing portion 18 of the sensor 12 to expose one or more sensor electrodes 20 to the user's bodily fluids through a window 22 formed in the cannula 16. The sensing portion 18 is joined to a connecting portion 24 that terminates in a conductive contact pad or the like, which is also exposed through one of the insulating layers. The connecting portion 24 and the contact pads are typically adapted for direct wired electrical connection to a suitable monitor 200 that is coupled to a display 214 to monitor a condition of the user in response to signals originating from the sensor electrodes 20. The connecting portion 24 can be conveniently electrically connected to the monitor 200 or the characteristic monitor transmitter 100 via a connector block 28 (or the like).

[0092] like Figure 3 As shown, according to an embodiment of the present invention, the subcutaneous sensor kit 10 can be configured or formed to work with a wired or wireless feature monitoring system. The proximal portion of the sensor 12 is mounted in a mounting base 30 suitable for placement on the user's skin. The mounting base 30 can be a pad having a lower surface, the lower surface of which is coated with a suitable pressure-sensitive adhesive layer 32, wherein a peeling paper tape 34 is generally provided to cover and protect the adhesive layer 32 until the sensor kit 10 is ready for use. The mounting base 30 includes an upper layer 36 and a lower layer 38, wherein the connecting portion 24 of the flexible sensor 12 is sandwiched between the layers 36 and 38. The connecting portion 24 has a front portion that is joined to the active sensing portion 18 of the sensor 12, and the front portion is folded at an angle to extend downward through a hole 40 formed in the lower base layer 38. Optionally, the adhesive layer 32 (or another portion of the device that contacts the tissue in the body) includes an anti-inflammatory agent to reduce the inflammatory response and / or an antibacterial agent to reduce the chance of infection. The insertion needle 14 is adapted to be received in a sliding fit through a needle port 42 formed in the upper substrate 36 and through a lower hole 40 in the lower substrate 38. After insertion, the insertion needle 14 is withdrawn to leave the cannula 16 with the sensing portion 18 and the sensor electrode 20 at the selected insertion location. In this embodiment, the telemetry characteristic monitor transmitter 100 is coupled to the sensor kit 10 via a wire 102 through a connector 104, which is electrically coupled to the connector block 28 of the connector portion 24 of the sensor kit 10.

[0093] exist Figure 3In the illustrated embodiment, the telemetry characteristic monitor 100 includes a housing 106 that supports a printed circuit board 108, a battery 110, an antenna 112, and a wire 102 with a connector 104. In some embodiments, the housing 106 is formed of an upper housing 114 and a lower housing 116 that are ultrasonically welded to form a waterproof (or water-resistant) seal, thereby allowing cleaning by immersion (or wiping) with water, detergent, alcohol, etc. In some embodiments, the upper housing 114 and the lower housing 116 are made of medical grade plastic. However, in alternative embodiments, the upper housing 114 and the lower housing 116 can be connected together by other methods, such as snap fit, sealing rings, RTV (silicone sealant), and bonding together, etc., or formed of other materials, such as metals, composite materials, ceramics, etc. In other embodiments, a separate housing can be omitted and the components are simply encapsulated in an epoxy or other moldable material that is compatible with the electronics and reasonably moisture-proof. As shown, the lower housing 116 may have an underside surface coated with a suitable pressure sensitive adhesive layer 118, with a release paper strip 120 typically provided to cover and protect the adhesive layer 118 until the sensor suite telemetry signature monitor transmitter 100 is ready for use.

[0094] exist Figure 3 In the illustrative embodiment shown, the subcutaneous sensor kit 10 facilitates the precise placement of a type of flexible thin film electrochemical sensor 12 for monitoring specific blood parameters indicative of a user's condition. The sensor 12 monitors glucose levels in the body and can be used in conjunction with an external or implantable automatic or semi-automatic drug infusion pump such as described in U.S. Pat. Nos. 4,562,751, 4,678,408, 4,685,903, or 4,573,994 to control insulin delivery to a diabetic patient.

[0095] exist Figure 3 In the illustrative embodiment shown in , the sensor electrode 10 can be used for a variety of sensing applications and can be configured in a variety of ways. For example, the sensor electrode 10 can be used in physiological parameter sensing applications in which some types of biomolecules are used as catalysts. For example, the sensor electrode 10 can be used in a glucose and oxygen sensor having a glucose oxidase that catalyzes a reaction with the sensor electrode 20. The sensor electrode 10, along with the biomolecule or some other catalyst, can be placed in the human body in a vascular or non-vascular environment. For example, the sensor electrode 20 and the biomolecule can be placed in a vein and subjected to blood flow, or can be placed in the subcutaneous or peritoneal area of ​​the human body.

[0096] exist Figure 3In the illustrated embodiment of the invention, the monitor 200 of the sensor signal may also be referred to as the sensor electronics device 200. The monitor 200 may include a power supply, a sensor interface, processing electronics (i.e., a processor), and data formatting electronics. The monitor 200 may be coupled to the sensor kit 10 via a wire 102 through a connector that is electrically coupled to the connector block 28 of the connection portion 24. In an alternative embodiment, the wire may be omitted. In this embodiment of the invention, the monitor 200 may include a suitable connector for connecting directly to the connection portion 104 of the sensor kit 10. The sensor kit 10 may be modified so that the connector portion 104 is located in a different position, such as at the top of the sensor kit, to facilitate placement of the monitor 200 above the sensor kit.

[0097] Although the analyte sensors and sensor systems disclosed herein are typically designed to be implanted in a mammal, the invention disclosed herein is not limited to any particular environment, but can be used in a variety of situations, such as for analyzing most in vivo and in vitro fluid samples, including biological fluids such as interstitial fluid, whole blood, lymph, plasma, serum, saliva, urine, feces, sweat, mucus, tears, cerebrospinal fluid, nasal secretions, cervical or vaginal secretions, semen, pleural fluid, amniotic fluid, peritoneal fluid, middle ear fluid, joint fluid, gastric fluid, etc. In addition, solid or dried samples can be dissolved in an appropriate solvent to provide a liquid mixture suitable for analysis.

[0098] It should be understood that the present invention is not limited to the specific embodiments described, as they can certainly vary. It should also be understood that the special terms used herein are only for the purpose of describing specific embodiments, and are not intended to be restrictive, as the scope of the present invention is limited only by the appended claims. In the description of the preferred embodiments, reference is made to the accompanying drawings that form a part of the description, and in the accompanying drawings, specific embodiments that can practice the present invention are shown by way of illustration. It should be understood that other embodiments can be utilized without departing from the scope of the present invention, and structural changes can be made.

Claims

1. An electrochemical analyte sensor comprising: Grassroots; A working electrode disposed on the substrate; and A multi-layer analyte sensor stack disposed on the working electrode, the multi-layer analyte sensor stack comprising: (a) an analyte sensing layer disposed directly on the working electrode, wherein the analyte sensing layer detectably changes the current at the working electrode in the presence of an analyte; (b) an analyte modulating layer disposed on the analyte sensing layer, wherein the analyte modulating layer promotes diffusion of O2 from an external environment into the analyte sensing layer; one or more channels disposed in the electrochemical analyte sensor architecture, the channels operably coupling the analyte sensing layer to an external environment, wherein the plurality of channels facilitate diffusion of O2 from the external environment to the analyte sensing layer; and A first silicon bridge having a first portion operably in contact with air in an in vitro environment and a second portion operably in contact with at least one channel, wherein: the first silicon bridge facilitates diffusion of O2 from air to the analyte sensing layer, and the first silicon bridge includes a portion that contacts interstitial fluid when the electrochemical analyte sensor is disposed in vivo.

2. The electrochemical analyte sensor of claim 1, further comprising an oxygen diffusion composition disposed within the plurality of channels, wherein the oxygen diffusion composition is selected to facilitate diffusion of O2 therethrough.

3. The electrochemical analyte sensor of claim 2, wherein: forming the analyte modulating layer and the oxygen diffusion composition using the same composition; and / or At least one channel forms an O2 conduit from the external environment on a first side of the sensor, through the base layer and the electrodes, and to the analyte sensing layer.

4. The electrochemical analyte sensor of any one of claims 1 to 3, wherein the external environment comprises interstitial fluid.

5. The electrochemical analyte sensor of claim 1 , further comprising a second silicon bridge having a first portion operably in contact with air in an in vitro environment and a second portion operably in contact with at least one channel, wherein the second silicon bridge facilitates diffusion of O2 from the air to the analyte sensing layer.

6. The electrochemical analyte sensor of claim 1, further comprising an oxygen generating electrode, wherein the oxygen generating electrode is disposed in the sensor such that oxygen generated by the electrode is in operable contact with the analyte sensing layer.

7. The electrochemical analyte sensor of claim 6, wherein the oxygen generating electrode is in operative contact with one or more channels that operatively couple the analyte sensing layer to the oxygen generating electrode.

8. The electrochemical analyte sensor according to claim 6, wherein: The oxygen generating electrode also serves as a counter electrode in the electrochemical analyte sensor.

9. The electrochemical analyte sensor of claim 1, wherein the electrochemical analyte sensor further comprises at least one of the following: a layer comprising a high density amine layer comprising a poly-L-lysine polymer having a molecular weight between 30 KDa and 300 KDa; a layer containing albumin; a layer comprising a silicone adhesion promoter; or A layer comprising glutaraldehyde.

10. A method of manufacturing an electrochemical analyte sensor, comprising the steps of: Provide grass-roots; forming a conductive layer on the base layer, wherein the conductive layer comprises a working electrode; forming an analyte sensing layer on the conductive layer, wherein the analyte sensing layer comprises a composition that can change the current at the working electrode in the conductive layer in the presence of an analyte; forming an analyte modulating layer on the analyte sensing layer; and forming one or more channels within the electrochemical analyte sensor in a region of the electrochemical analyte sensor to operably couple the analyte sensing layer to an external environment, wherein the one or more channels facilitate diffusion of O2 from the external environment to the analyte sensing layer; as well as forming a first silicon bridge, the first silicon bridge being configured such that when the electrochemical analyte sensor is disposed in vivo, a first portion of the first silicon bridge is in operable contact with air in an in vitro environment, and a second portion of the first silicon bridge is in operable contact with at least one channel formed in the base layer, wherein: the first silicon bridge facilitates diffusion of O2 from air to the analyte sensing layer, and the first silicon bridge includes a portion that contacts interstitial fluid when the electrochemical analyte sensor is disposed in vivo; Thus, the electrochemical analyte sensor is manufactured.

11. The method of claim 10, further comprising placing an oxygen diffusion composition within the one or more channels, wherein the oxygen diffusion composition is selected to facilitate diffusion of O2 therethrough.

12. The method of claim 10, further comprising disposing an oxygen generating electrode in the electrochemical analyte sensor, wherein the oxygen generating electrode is disposed in the electrochemical analyte sensor such that oxygen generated by the electrode is in operable contact with the analyte sensing layer.

13. The method of claim 12, wherein the oxygen generating electrode is disposed in the electrochemical analyte sensor so as to be in operable contact with one or more channels that operably couple the analyte sensing layer to the oxygen generating electrode.

14. The method according to claim 12, wherein: The oxygen generating electrode is disposed in the electrochemical analyte sensor so as to function as a counter electrode in the electrochemical analyte sensor.

15. The method of claim 10, wherein forming the electrochemical analyte sensor to further include at least one of: a layer comprising a high density amine layer comprising a poly-L-lysine polymer having a molecular weight between 30 KDa and 300 KDa; a layer containing albumin; a layer comprising a silicone adhesion promoter; or A layer comprising glutaraldehyde.

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