Analyte sensing microneedle electrode

Abstract

The present invention relates to a microneedle electrode for detecting at least one analyte, such as glucose, ketones and / or lactate. The present invention also relates to a wearable device comprising one or more microneedle electrodes, as well as a process for detecting at least one analyte using the microneedle electrode and / or the wearable device of the present invention.

Description

ANALYTE SENSING MICRONEEDLE ELECTRODEINTRODUCTION

[0001] The present invention relates to a microneedle electrode for detecting at least one analyte, such as glucose, ketones and / or lactate. The present invention also relates to a wearable device comprising one or more microneedle electrodes, as well as a process for detecting at least one analyte using the microneedle electrode and / or the wearable device of the present invention.BACKGROUND OF THE INVENTION

[0002] Detecting and monitoring analyte concentration in the human and animal body is well documented as an important metric in a number of performance and disease related fields. For example, monitoring the concentration of analytes such as glucose and lactate, two key performance metrics, can allow a subject to predict exhaustion times, reduce the risk of injuries and maximise performance levels by tailoring a specific dietary plan based on their glucose and lactate levels. In an alternative field, monitoring the concentration of analytes such as glucose is of critical importance in the management of diseases such as diabetes mellitus and hypoglycaemia, both of which are the result of an imbalance in glucose levels. The imbalance in glucose levels typically arises from a lack of insulin, a hormone that regulates the level of glucose and is produced by the pancreas. A common practice in monitoring glucose concentration levels has been to employ devices such as blood glucose meters and continuous glucose monitors. Blood glucose meters are biosensors which are equipped with a needle, a lancet or the like and a measuring device in order to determine the glucose concentration in blood. This is typically done by acquiring a small drop of blood by pricking the skin of a subject. The blood sample is subsequently placed onto a disposable test strip or similar measuring device in order to determine the glucose concentration in the blood of the subject. A continuous glucose monitor differs in a number of areas but most notably this technology measures glucose concentration in interstitial fluid, which is a thin layer of fluid surrounding cells in the body, as opposed to blood, as well as monitoring the glucose concentration continuously, rather than a one-off measurement.

[0003] In spite of the advances in the technology of blood glucose meters and continuous glucose monitors, there are a number of drawbacks associated with this technology. Firstly, the invasive nature of blood glucose meters means that the subject has a regular level of pain and discomfort, as well as risking infection when pricking the skin to obtain a blood sample. A continuous glucose monitoring device is somewhat less invasive, but still requires the insertion of small sensor wire under the skin of the subject and some devices also still require a finger prick sample, typically twice a day, in order to maintain the calibration of the device. Thus,these continuous glucose monitoring devices are still considered to be invasive techniques, which often leads to discomfort and lack of subject compliance. Secondly, continuous glucose monitors, when inserted, typically have poor in-vivo mechanical stability. This issue is particularly noticeable in performance studies during kinematic movement of a subject as the in-vivo mechanical instability can lead to inaccuracies in detection and measurement readings. This limits the effectiveness of such devices for optimum performance analysis of glucose concentration. Thirdly, detection devices inserted into a subject, such as continuous glucose monitors, are susceptible towards interfering substances in the blood and interstitial fluid, such as sweat and ascorbic acid. This poor biocompatibility with the surrounding environment and susceptibility to substance interference typically leads to inaccurate measurements and accelerated sensory drift with time.

[0004] In general, certain prior art devices suffer from drawbacks including their high price, their reliability (i.e., high error rate), and their invasiveness (which may lead to inflammation for the subject). Moreover, single sensor continuous glucose monitors carry the risk that the single sensor may break off and become embedded in the skin. This, of course, carries serious risks for the subject and prevents normal functioning of the device.

[0005] In view of the above, there is a need for an analyte monitoring device which is minimally invasive, mechanically stable and biocompatible in a range of different physiological environments.

[0006] The present invention was devised with the foregoing in mind.SUMMARY OF THE INVENTION

[0007] According to a first aspect of the present invention there is provided a microneedle electrode for detecting at least one analyte, wherein at least a portion of the surface of the microneedle electrode is coated with a composite material,wherein the composite material comprises:a semi-permeable outer layer; anda sensing layer positioned underneath the semi-permeable outer layer that is capable of generating an electrochemical response in the presence of the analyte(s).

[0008] According to a second aspect of the present invention there is provided a wearable device for detecting at least one analyte, wherein the wearable device comprises one or more microneedle electrodes according to the first aspect of the present invention.

[0009] According to a third aspect of the present invention there is provided a method of detecting at least one analyte, wherein the method comprises contacting a microneedle electrode according to the first aspect of the present invention with the analyte(s).

[0010] According to a fourth aspect of the present invention there is provided a method of detecting at least one analyte, wherein the method comprises contacting a wearable deviceaccording to the second aspect of the present invention with the analyte(s).DETAILED DESCRIPTION OF THE INVENTIONDefinitions

[0011] Unless otherwise stated, the following terms used in the specification and claims have the following meaning set out below:

[0012] The term "(m-nC)" or "(m-nC) group" used alone or as a prefix, refers to any group having m to n carbon atoms.

[0013] The term “alkyl” as used herein refers to straight or branched chain alkyl moieties, typically having 1, 2, 3, 4, 5 or 6 carbon atoms. This term includes reference to groups such as methyl, ethyl, propyl (n-propyl or isopropyl), butyl (n-butyl, sec-butyl or tert-butyl), pentyl (including neopentyl), hexyl and the like. In particular, an alkyl may have 1, 2, 3 or 4 carbon atoms.

[0014] The term “alkoxy” as used herein refers to -O-alkyl, wherein alkyl is a straight or branched chain and comprises 1, 2, 3, 4, 5 or 6 carbon atoms. In one class of embodiments, alkoxy has 1, 2, 3 or 4 carbon atoms. This term includes reference to groups such as methoxy, ethoxy, propoxy, isopropoxy, butoxy, tert-butoxy, pentoxy, hexoxy and the like.

[0015] The term “alkenyl” as used herein refers to straight or branched chain alkenyl moieties, typically having 2, 3, 4, 5 or 6 carbon atoms. The term includes reference to alkenyl moieties containing 1, 2 or 3 carbon-carbon double bonds (C=C). This term includes reference to groups such as ethenyl (vinyl), propenyl (allyl), butenyl, pentenyl and hexenyl, as well as both the c / s and trans isomers thereof.

[0016] The term “alkynyl” as used herein refers to straight or branched chain alkynyl moieties, typically having 2, 3, 4, 5 or 6 carbon atoms. The term includes reference to alkynyl moieties containing 1, 2 or 3 carbon-carbon triple bonds (C=C). This term includes reference to groups such as ethynyl, propynyl, butynyl, pentynyl and hexynyl.

[0017] The term “carbocyclyl”, “carbocyclic” or “carbocycle” means a non-aromatic saturated or partially saturated monocyclic, or a fused, bridged, or spiro bicyclic carbocyclic ring system(s). Monocyclic carbocyclic rings contain from about 3 to 13 (suitably from 3 to 7) ring atoms. Bicyclic carbocycles contain from 7 to 17 carbon atoms in the rings, suitably 7 to 12 carbon atoms, in the rings. Bicyclic carbocyclic rings may be fused, spiro, or bridged ring systems.

[0018] The term “aryl” or “aromatic” as used herein means an aromatic ring system comprising 6, 7, 8, 9 or 10 ring carbon atoms. Aryl is often phenyl but may be a polycyclic ring system, having two or more rings, at least one of which is aromatic. This term includes reference to groups such as phenyl, naphthyl and the like.

[0019] The term “heterocyclyl”, “heterocyclic” or “heterocycle” means a non-aromaticsaturated or partially saturated monocyclic, fused, bridged, or spiro bicyclic heterocyclic ring system(s). Monocyclic heterocyclic rings contain from about 3 to 12 (suitably from 3 to 7) ring atoms, with from 1 to 5 (suitably 1, 2 or 3) heteroatoms selected from nitrogen, oxygen or sulfur in the ring. Bicyclic heterocycles contain from 7 to 17 member atoms, suitably 7 to 12 member atoms, in the ring. Bicyclic heterocyclic(s) rings may be fused, spiro, or bridged ring systems. Examples of heterocyclic groups include cyclic ethers such as oxiranyl, oxetanyl, tetrahydrofuranyl, dioxanyl, and substituted cyclic ethers. Heterocycles containing nitrogen include, for example, azetidinyl, pyrrolidinyl, piperidinyl, piperazinyl, tetrahydrotriazinyl, tetrahydropyrazolyl, and the like. Typical sulfur containing heterocycles include tetrahydrothienyl, dihydro-1, 3-dithiol, tetrahydro-2H-thiopyran, and hexahydrothiepine. Other heterocycles include dihydro oxathiolyl, tetrahydro oxazolyl, tetrahydro-oxadiazolyl, tetrahydrodioxazolyl, tetrahydro oxathiazolyl, hexahydrotriazinyl, tetrahydro oxazinyl, morpholinyl, thiomorpholinyl, tetrahydropyrimidinyl, dioxolinyl, octahydrobenzofuranyl, octahydrobenzimidazolyl, and octahydrobenzothiazolyl. For heterocycles containing sulfur, the oxidized sulfur heterocycles containing SO or SO2 groups are also included. Examples include the sulfoxide and sulfone forms of tetrahydrothienyl and thiomorpholinyl such as tetrahydrothiene 1,1 -dioxide and thiomorpholinyl 1,1 -dioxide. Heterocycles may comprise 1 or 2 oxo (=0) or thioxo (=S) substituents. A suitable value for a heterocyclyl group which bears 1 or 2 oxo (=0) or thioxo (=S) substituents is, for example, 2 oxopyrrolidinyl, 2 thioxopyrrolidinyl, 2 oxoimidazolidinyl, 2 thioxoimidazolidinyl, 2 oxopiperidinyl, 2,5 dioxopyrrolidinyl, 2,5 dioxoimidazolidinyl or 2,6 dioxopiperidinyl. Particular heterocyclyl groups are saturated monocyclic 3 to 7 membered heterocyclyls containing 1, 2 or 3 heteroatoms selected from nitrogen, oxygen or sulfur, for example azetidinyl, tetrahydrofuranyl, tetrahydropyranyl, pyrrolidinyl, morpholinyl, tetrahydrothienyl, tetrahydrothienyl 1,1-dioxide, thiomorpholinyl, thiomorpholinyl 1,1-dioxide, piperidinyl, homopiperidinyl, piperazinyl or homopiperazinyl. As the skilled person would appreciate, any heterocycle may be linked to another group via any suitable atom, such as via a carbon or nitrogen atom. However, reference herein to piperidino or morpholino refers to a piperidin-1-yl or morpholin-4-yl ring that is linked via the ring nitrogen.

[0020] By “bridged ring systems” is meant ring systems in which two rings share more than two atoms, see for example Advanced Organic Chemistry, by Jerry March, 4th Edition, Wiley Interscience, pages 131-133, 1992. Examples of bridged heterocyclyl ring systems include, aza-bicyclo[2.2.1]heptane, 2-oxa-5-azabicyclo[2.2.1]heptane, aza-bicyclo[2.2.2]octane, aza-bicyclo[3.2.1]octane and quinuclidine.

[0021] By “spiro bi-cyclic ring systems” we mean that the two ring systems share one common spiro carbon atom, i.e. the heterocyclic ring is linked to a further carbocyclic or heterocyclic ring through a single common spiro carbon atom. Examples of spiro ring systems include 6-azaspiro[3.4]octane, 2-oxa-6-azaspiro[3.4]octane, 2-azaspiro[3.3]heptanes, 2-oxa-6-azaspiro[3.3]heptanes, 7-oxa-2-azaspiro[3.5]nonane, 6-oxa-2-azaspiro[3.4]octane, 2-oxa-7-azaspiro[3.5]nonane and 2-oxa-6-azaspiro[3.5]nonane.

[0022] The term “heteroaryl” or “heteroaromatic” means an aromatic mono, bi, or polycyclic ring incorporating one or more (for example 1 4, particularly 1, 2 or 3) heteroatoms selected from nitrogen, oxygen or sulfur. The term heteroaryl includes both monovalent species and divalent species. Examples of heteroaryl groups are monocyclic and bicyclic groups containing from five to twelve ring members, and more usually from five to ten ring members. The heteroaryl group can be, for example, a 5- or 6-membered monocyclic ring or a 9- or 10-membered bicyclic ring, for example a bicyclic structure formed from fused five and six membered rings or two fused six membered rings. Each ring may contain up to about four heteroatoms typically selected from nitrogen, sulfur and oxygen. Typically the heteroaryl ring will contain up to 3 heteroatoms, more usually up to 2, for example a single heteroatom. In one embodiment, the heteroaryl ring contains at least one ring nitrogen atom. The nitrogen atoms in the heteroaryl rings can be basic, as in the case of an imidazole or pyridine, or essentially non-basic as in the case of an indole or pyrrole nitrogen. In general the number of basic nitrogen atoms present in the heteroaryl group, including any amino group substituents of the ring, will be less than five.

[0023] Examples of heteroaryl include furyl, pyrrolyl, thienyl, oxazolyl, isoxazolyl, imidazolyl, pyrazolyl, thiazolyl, isothiazolyl, oxadiazolyl, thiadiazolyl, triazolyl, tetrazolyl, pyridyl, pyridazinyl, pyrimidinyl, pyrazinyl, 1,3,5 triazenyl, benzofuranyl, indolyl, isoindolyl, benzothienyl, benzoxazolyl, benzimidazolyl, benzothiazolyl, benzothiazolyl, indazolyl, purinyl, benzofurazanyl, quinolyl, isoquinolyl, quinazolinyl, quinoxalinyl, cinnolinyl, pteridinyl, naphthyridinyl, carbazolyl, phenazinyl, benzisoquinolinyl, pyridopyrazinyl, thieno[2,3 b]furanyl, 2H furo[3,2 b] pyranyl, 5H pyrido[2,3 d] o oxazinyl, 1H pyrazolo[4,3 d] oxazolyl, 4H imidazo[4,5 d]thiazolyl, pyrazino[2,3 d]pyridazinyl, imidazo[2,1 b]thiazolyl, imidazo[1,2 b][1,2,4]triazinyl. “Heteroaryl” also covers partially aromatic bi- or polycyclic ring systems wherein at least one ring is an aromatic ring and one or more of the other ring(s) is a non-aromatic, saturated or partially saturated ring, provided at least one ring contains one or more heteroatoms selected from nitrogen, oxygen or sulfur. Examples of partially aromatic heteroaryl groups include for example, tetrahydroisoquinolinyl, tetrahydroquinolinyl, 2-oxo-1,2,3,4-tetrahydroquinolinyl, dihydrobenzthienyl, dihydrobenzfuranyl, 2,3-dihydro-benzo[1,4]dioxinyl, benzo[1,3]dioxolyl, 2,2-dioxo-1,3-dihydro-2-benzothienyl, 4,5,6,7-tetrahydrobenzofuranyl, indolinyl, 1,2, 3, 4 tetrahydro 1,8 naphthyridinyl, 1,2, 3, 4 tetrahydropyrido[2,3 b]pyrazinyl and 3,4 dihydro 2H pyrido[3,2 b][1,4]oxazinyl.

[0024] Examples of five membered heteroaryl groups include but are not limited to pyrrolyl, furanyl, thienyl, imidazolyl, furazanyl, oxazolyl, oxadiazolyl, oxatriazolyl, isoxazolyl, thiazolyl,isothiazolyl, pyrazolyl, triazolyl and tetrazolyl groups.

[0025] Examples of six membered heteroaryl groups include but are not limited to pyridyl, pyrazinyl, pyridazinyl, pyrimidinyl and triazinyl.

[0026] A bicyclic heteroaryl group may be, for example, a group selected from:a benzene ring fused to a 5- or 6-membered ring containing 1, 2 or 3 ring heteroatoms; a pyridine ring fused to a 5- or 6-membered ring containing 1, 2 or 3 ring heteroatoms; a pyrimidine ring fused to a 5- or 6-membered ring containing 1 or 2 ring heteroatoms; a pyrrole ring fused to a 5- or 6-membered ring containing 1, 2 or 3 ring heteroatoms;a pyrazole ring fused to a 5- or 6-membered ring containing 1 or 2 ring heteroatoms;a pyrazine ring fused to a 5- or 6-membered ring containing 1 or 2 ring heteroatoms;an imidazole ring fused to a 5- or 6-membered ring containing 1 or 2 ring heteroatoms; an oxazole ring fused to a 5- or 6-membered ring containing 1 or 2 ring heteroatoms; an isoxazole ring fused to a 5- or 6-membered ring containing 1 or 2 ring heteroatoms; a thiazole ring fused to a 5- or 6-membered ring containing 1 or 2 ring heteroatoms;an isothiazole ring fused to a 5- or 6-membered ring containing 1 or 2 ring heteroatoms; a thiophene ring fused to a 5- or 6-membered ring containing 1, 2 or 3 ring heteroatoms; a furan ring fused to a 5- or 6-membered ring containing 1, 2 or 3 ring heteroatoms;a cyclohexyl ring fused to a 5- or 6-membered heteroaromatic ring containing 1, 2 or 3 ring heteroatoms; anda cyclopentyl ring fused to a 5- or 6-membered heteroaromatic ring containing 1, 2 or 3 ring heteroatoms.

[0027] Particular examples of bicyclic heteroaryl groups containing a six membered ring fused to a five membered ring include but are not limited to benzfuranyl, benzthiophenyl, benzimidazolyl, benzoxazolyl, benzisoxazolyl, benzthiazolyl, benzisothiazolyl, isobenzofuranyl, indolyl, isoindolyl, indolizinyl, indolinyl, isoindolinyl, purinyl (e.g., adeninyl, guaninyl), indazolyl, benzodioxolyl and pyrazolopyridinyl groups.

[0028] Particular examples of bicyclic heteroaryl groups containing two fused six membered rings include but are not limited to quinolinyl, isoquinolinyl, chromanyl, thiochromanyl, chromenyl, isochromenyl, chromanyl, isochromanyl, benzodioxanyl, quinolizinyl, benzoxazinyl, benzodiazinyl, pyridopyridinyl, quinoxalinyl, quinazolinyl, cinnolinyl, phthalazinyl, naphthyridinyl and pteridinyl groups.

[0029] The term “substituted” as used herein in reference to a moiety means that one or more of the hydrogen atoms in said moiety are replaced independently of each other by the corresponding number of the described substituents. Preferably, “substituted” as used herein in reference to a moiety means that 1, 2 or 3, of the hydrogen atoms in said moiety are replaced independently of each other by the corresponding number of the described substituents. Even more preferred, “substituted” as used herein in reference to a moietymeans that 1 or 2, of the hydrogen atoms in said moiety are replaced independently of each other by the corresponding number of the described substituents. The term “optionally substituted” as used herein means substituted or unsubstituted.

[0030] It will, of course, be understood that substituents are only at positions where they are chemically possible, the person skilled in the art being able to decide (either experimentally or theoretically) without inappropriate effort whether a particular substitution is possible.

[0031] In this specification, the terms “comprise” and “contain” and variations of them mean “including but not limited to”, and they are not intended to (and do not) exclude other moieties, additives, components, integers or steps. Throughout the description and claims of this specification, the singular encompasses the plural unless the context otherwise requires. In particular, where the indefinite article is used, the specification is to be understood as contemplating plurality as well as singularity, unless the context requires otherwise.

[0032] Features, integers, characteristics, compounds, chemical moieties or groups described in conjunction with a particular aspect, embodiment or example of the invention are to be understood to be applicable to any other aspect, embodiment or example described herein unless incompatible therewith. All of the features disclosed in this specification (including any accompanying claims, abstract and drawings), and / or all of the steps of any method or process so disclosed, may be combined in any combination, except combinations where at least some of such features and / or steps are mutually exclusive. The invention is not restricted to the details of any foregoing embodiments. The invention extends to any novel one, or any novel combination, of the features disclosed in this specification (including any accompanying claims and drawings), or to any novel one, or any novel combination, of the steps of any method or process so disclosed.Microneedle electrode

[0033] As described herein, the present invention provides a microneedle electrode for detecting at least one analyte, wherein at least a portion of the surface of the microneedle electrode is coated with a composite material,wherein the composite material comprises:a semi-permeable outer layer; anda sensing layer positioned underneath the semi-permeable outer layer that is capable of generating an electrochemical response in the presence of the analyte(s).

[0034] Through extensive investigations, the inventors have found that the microneedle electrodes of the present invention are particularly well suited for analyte detection, as well as determination of analyte concentration. The microneedle electrodes can be used to monitor analytes (e.g., glucose, ketones and lactate) in performance analysis, as well as diseases such as diabetes mellitus and hypoglycaemia. In particular, the inventors have found that themicroneedle electrodes are themselves minimally invasive when applied to a subject and can be used in minimally invasive methods for monitoring glucose concentration. Suitably, the subject is a human. The microneedle electrodes have also been found to exhibit mechanical stability during kinematic movement of a subject as well as biocompatibility in a range of physiological environments such as interstitial fluid in layers of the skin. The inventors have also demonstrated that the microneedle electrodes display anti-interferent properties to potential interferents such as ascorbic acid and uric acid, as well as a high enzyme stability, thereby circumventing a number of the drawbacks associated with known blood glucose meters and continuous glucose monitors.

[0035] The microneedle electrode may comprise a substrate, an optional non-conductive passivation layer, and a conductive electrode layer. Suitably, the microneedle electrode comprises a substrate, a non-conductive passivation layer, and a conductive electrode layer. The substrate may comprise (or is formed from) metals (e.g., titanium, tungsten, silver, gold, copper, aluminium, steel (e.g., stainless steel), iron, platinum, tantalum, Nitinol, Elgiloy or alloys thereof), carbon (e.g., graphite, graphene, carbon nanotubes, pyrolytic carbon or carbon fibre), plastics, liquid crystal polymers, silicon or composites (e.g. glass-filled, mineral-filled, or carbon-filled composites). The non-conductive passivation layer is suitably present when the substrate comprises (or is formed from) a conductive material (e.g., steel). The non-conductive passivation layer may comprise (or is formed from) insulating polymers (e.g., parylene (e.g., parylene-C, parylene-F or parylene-N), polyimide, polyvinylchloride, benzocylcobutene or acrylate) or metal oxides (e.g., titanium dioxide, silicon dioxide or aluminium oxide). The conductive electrode layer may comprise (or is formed from) carbon (e.g., graphite, graphene, carbon nanotubes, pyrolytic carbon or carbon fibre), platinum, gold, or rhodium, palladium, silver, silver-silver chloride, steel (e.g. stainless steel), copper, nickel, zinc, or aluminium, tantalum or chromium. Suitably, the substrate and the conductive electrode layer comprise (or are formed from) different materials from one another. More suitably, the microneedle electrode comprises a substrate, a non-conductive passivation layer and a conductive electrode layer, wherein the substrate comprises (or is formed from) steel, the non-conductive passivation layer comprises (or is formed from) parylene (e.g., parylene-C, parylene-F or parylene-N), polyimide, polyvinylchloride, benzocylcobutene, acrylate, titanium dioxide, silicon dioxide or aluminium oxide, and the conductive electrode layer comprises (or is formed from) carbon (e.g., graphite, graphene, carbon nanotubes, pyrolytic carbon or carbon fibre), platinum, gold or rhodium. Most suitably, the microneedle electrode comprises a substrate, a non-conductive passivation layer and a conductive electrode layer, wherein the substrate comprises (or is formed from) stainless steel, the non-conductive passivation layer comprises (or is formed from) parylene-C, and the conductive electrode layer comprises (or is formed from) carbon ink. The configuration of the substrate, non-conductivepassivation layer and conductive electrode layer, in its simplest form, is illustrated below (i.e., the non-conductive passivation layer overlays the substrate and the conductive electrode layer overlays the non-conductive passivation layer):

[0036] The microneedle electrode may comprise a substrate, a non-conductive passivation layer and a conductive electrode layer, wherein:the non-conductive passivation layer surrounds the entirety of the substrate (i.e., the substrate is encapsulated by the non-conductive passivation layer); andthe conductive electrode layer overlays at least a portion of the non-conductive passivation layer. For example, the configuration of the microneedle electrode may be illustrated as follows:

[0037] The term analyte is known in the art as an analysable substance or chemical constituent in a biological fluid, such as blood, interstitial fluid, cerebral spinal fluid, lymph fluid, urine, sweat and saliva. Suitably, the analyte is present in blood, interstitial fluid, cerebral spinal fluid, lymph fluid, urine, sweat and / or saliva. More suitably, the analyte is present in interstitial fluid. The interstitial fluid may be in the dermis of the skin. The dermis of the skin is the layer of skin which lies beneath the epidermis and above the subcutaneous tissue layer. The analyte may alternatively be present in other layers of the skin, such as the stratum corneum, the epidermis and the subcutaneous tissue of the skin.

[0038] Interstitial fluid is the body fluid found in the spaces around cells and blood vesselsand typically comprises, inter alia, a number of substances such as sugars, salts, fatty acids, amino acids and hormones. Therefore, the analyte may be a sugar, a salt, a fatty acid, an amino acid or a hormone. Suitably, the analyte is a sugar. More suitably, the analyte is selected from the group consisting of glucose, ketones (e.g., p-hydroxybutyrate), histamine, alcohols, cholesterol, vitamins, iodine, potassium, sodium, magnesium, calcium, zinc, copper, iron, chloride, phosphate, ammonium, lithium, bicarbonate, C-reactive protein (CRP), thrombin, therapeutic drugs (e.g. aminoglycosides, doxorubicin, tetracyclines), hormones (e.g., testosterone, cortisol, oestrogen, serotonin and progesterone), pH (e.g., hydrogen ion), interleukin-6 (IL-6), creatinine, acarboxyprothrombin, acylcarnitine, adenine phosphoribosyl transferase, adenosine deaminase, albumin, a-fetoprotein, amino acid profiles (e.g., arginine, histidine / urocanic acid, homocysteine, phenylalanine / tyrosine, tryptophan), andrenostenedione, antipyrine, arabinitol enantiomers, arginase, benzoylecgonine (cocaine), biotinidase, biopterin, c-reactive protein, carnitine, carnosinase, CD4, ceruloplasmin, chenodeoxycholic acid, chloroquine, cholinesterase, conjugated 1- hydroxy-cholic acid, Cortisol, cyclosporin A, d-penicillamine, de-ethylchloroquine, dehydroepiandrosterone sulphate, DNA (e.g., acetylator polymorphism, alcohol dehydrogenase, alpha 1 -antitrypsin, cystic fibrosis, Duchenne / Becker muscular dystrophy, glucose-6-phosphate dehydrogenase, haemoglobin A, haemoglobin S, haemoglobin C, haemoglobin D, haemoglobin E, haemoglobin F, D-Punjab, p- thalassemia, hepatitis B virus, HCMV, HIV-1, HTLV-1, Leber hereditary optic neuropathy, MCAD, RNA, PKU, Plasmodium vivax, sexual differentiation, 21-deoxycortisol), desbutylhalofantrine, dihydropteridine reductase, diptheria / tetanus antitoxin, erythrocyte arginase, erythrocyte protoporphyrin, esterase D, fatty acids / acylglycines, free -human chorionic gonadotropin, free erythrocyte porphyrin, free thyroxine (FT4), free triiodothyronine (FT3), fumarylacetoacetase, galactose / gal-1 -phosphate, galactose-1-phosphate uridyltransferase, gentamicin, glucose-6-phosphate dehydrogenase, glycocholic acid, glycosylated haemoglobin, halofantrine, haemoglobin variants, hexosaminidase A, human erythrocyte carbonic anhydrase I, 17-a-hydroxyprogesterone, hypoxanthine phosphoribosyl transferase, immunoreactive trypsin, lactate, lead, lipoproteins ((a), B / A-1, P), lysozyme, mefloquine, netilmicin, phenobarbitone, phenyloin, phytanic / pristanic acid, progesterone, prolactin, prolidase, purine nucleoside phosphorylase, quinine, reverse triiodothyronine (rT3), selenium, serum pancreatic lipase, sissomicin, somatomedin C, specific antibodies (e.g., adenovirus, anti-nuclear antibody, anti-zeta antibody), arbovirus, Aujeszky's disease virus, dengue virus, Dracunculus medinensis, Echinococcus granulosus, Entamoeba histolytica, enterovirus, Giardia duodenalisa, Helicobacter pylori, hepatitis B virus, herpes virus, HIV-1, IgE (atopic disease), influenza virus, Leishmania donovani, leptospira, measles / mumps / rubella, Mycobacterium leprae, Mycoplasma pneumoniae, Myoglobin, Onchocerca volvulus, parainfluenza virus, Plasmodium falciparum, poliovirus, Pseudomonasaeruginosa, respiratory syncytial virus, rickettsia (scrub typhus), Schistosoma mansoni, Toxoplasma gondii, Trepenoma pallidium, Trypanosoma cruzi / rangeli, vesicular stomatis virus, Wuchereria bancrofti, yellow fever virus, specific antigens (e.g., hepatitis B virus, HIV-1), succinylacetone, sulfadoxine, theophylline, thyrotropin (TSH), thyroxine (T4), thyroxine-binding globulin, trace elements, transferrin, UDP-galactose-4-epimerase, urea, uroporphyrinogen I synthase, vitamin A, white blood cells, zinc protoporphyrin, salts, sugar, protein, fat, vitamins, and hormones naturally occurring in blood or interstitial fluids, a metabolic product, a hormone, an antigen, an antibody, a radioisotope, a chemical agent, a fluorocarbon-based synthetic blood, or a drug or pharmaceutical composition, including: insulin; ethanol; cannabis (marijuana, tetrahydrocannabinol, hashish); inhalants (e.g., nitrous oxide, amyl nitrite, butyl nitrite, chlorohydrocarbons, hydrocarbons); cocaine (crack cocaine); stimulants (e.g., amphetamines, methamphetamines, Ritalin, Cylert, Preludin, Didrex, PreState, Voranil, Sandrex, Plegine); depressants (e.g., barbituates, methaqualone, tranquilizers such as Valium, Librium, Miltown, Serax, Equanil, Tranxene); hallucinogens (e.g., phencyclidine, lysergic acid, mescaline, peyote, psilocybin); narcotics (e.g., heroin, codeine, morphine, opium, meperidine, Percocet, Percodan, Tussionex, Fentanyl, Darvon, Talwin, Lomotil); designer drugs (e.g., analogs of fentanyl, meperidine, amphetamines, methamphetamines, and phencyclidine, for example, Ecstasy); anabolic steroids; and nicotine, and metabolic products of drugs and pharmaceutical compositions such as neurochemicals and other chemicals generated within the body such as, for example, noradrenaline, 3-methoxytyramine (3MT), 3,4-dihydroxyphenylacetic acid (DOPAC), homovanillic acid (HVA), 5-hydroxytryptamine (5HT), and 5- hydroxyindoleacetic acid (FHIAA). Even more suitably, the analyte is selected from the group consisting of glucose, ketones (e.g., p- hydroxy butyrate), lactate, histamine, urea, creatinine, alcohols, cholesterol, vitamins, hormones (e.g., testosterone, cortisol, serotonin, oestrogen and progesterone), pH (e.g., hydrogen ion), iodine, potassium, sodium, magnesium, calcium, zinc, copper, iron, chloride, phosphate, ammonium, lithium, bicarbonate, thrombin, IL-6 and therapeutic drugs (e.g. aminoglycosides, doxorubicin, tetracyclines). Yet even more suitably, the analyte is selected from the group consisting of glucose, ketones (e.g., p-hydroxybutyrate), lactate, histamine, urea, alcohols, cholesterol, vitamins, iodine, potassium, sodium, hormones (e.g., testosterone, cortisol, oestrogen and progesterone) and IL-6. Yet even more suitably, the analyte is selected from the group consisting of glucose, ketones and lactate. In some embodiments, the analyte is lactate. In other embodiments, the analyte is glucose.

[0001] The microneedle electrode is suitable for detecting at least one analyte. Suitably, the microneedle electrode detects one to ten analytes. More suitably, the microneedle electrode detects one to five analytes. Even more suitably, the microneedle electrode detects one to three analytes. Most suitably, the microneedle electrode detects one or two analytes. Inembodiments wherein the microneedle electrode detects more than one analyte, the analytes will be different. Each analyte that can be detected is as defined herein.

[0039] In some embodiments, the microneedle electrode detects one to five analytes, wherein each analyte is independently selected from the group consisting of glucose, ketones (e.g., p-hydroxybutyrate), lactate, histamine, urea, creatinine, alcohols, cholesterol, vitamins, hormones (e.g., testosterone, cortisol, serotonin, oestrogen and progesterone), pH (e.g., hydrogen ion), iodine, potassium, sodium, magnesium, calcium, zinc, copper, iron, chloride, phosphate, ammonium, lithium, bicarbonate, thrombin, IL-6 and therapeutic drugs (e.g. aminoglycosides, doxorubicin, tetracyclines). In some embodiments, the microneedle electrode detects one or two analytes, wherein each analyte is independently selected from glucose, ketones or lactate.

[0040] The microneedle electrode may comprise an insulation layer. The insulation layer is particularly useful in blocking interferents such as sweat on the skin, thereby increasing the sensing accuracy of the microneedle electrodes. Suitably, the insulation layer is a dielectric layer. The dielectric layer may comprise (or is formed from) a dielectric ink. Suitably, the ink solid content (i.e., the proportion of ink that is not solvent) of the dielectric layer is greater than 70%. More suitably, the ink solid content of the dielectric layer is greater than 80%. Even more suitably, the ink solid content of the dielectric layer is greater than 90%. Yet even more suitably, the ink solid content of the dielectric layer is greater than 95%. Yet still even more suitably, the ink solid content of the dielectric layer greater than 99%. Most suitably, the ink solid content of the dielectric layer is 100%. The dielectric layer may be a uniform layer having a thickness of 2 pm to 10 pm, suitably 3 pm to 5 pm. Suitably, the insulation layer coats a portion of the surface of the microneedle electrode that is not is coated with the composite material as defined herein.

[0041] The composite material comprises a semi-permeable outer layer and a sensing layer positioned underneath the semi-permeable outer layer that is capable of generating an electrochemical response in the presence of the analyte(s). The configuration is such that the sensing layer is present between the microneedle electrode and the semi-permeable outer layer. This configuration is illustrated below, in its simplest form:

[0042] Through detailed analysis, the present inventors have found that the composite material, based on its composition and design, optimises the analyte sensing capabilities of the microneedle electrode of the present invention. Not only this, it is thought that the composite material provides increased mechanical and physiological stability to themicroneedle electrode, as well as increased sensitivity to analytes, offering improved analyte concentration measurements. The improvements in mechanical and physiological stability are in part due to the semi-permeable outer layer, which suitably forms the outermost layer of the composite material of the present invention.

[0043] Suitably, the semi-permeable outer layer, as highlighted above, is the outermost layer of the composite material, thereby coming into contact with substances in the immediate environment surrounding the microneedle electrodes. The properties of the semi-permeable outer layer are therefore important in terms of how the microneedle electrode detects the analyte and its suitability in different physiological environments within a subject. Suitably, the semi-permeable outer layer has greater permeability to some substances, but less permeability to other substances, thereby controlling the amount of certain substances reaching the sensing layer positioned underneath the semi-permeable outer layer.

[0044] The semi-permeable outer layer may be permeable to the analyte(s) and optionally oxygen, the analyte(s) being as defined herein. Permeable, in the context of the semi-permeable outer layer, suitably means that greater than 0% of the substance (e.g., the analyte(s) and optionally oxygen) permeates through the semi-permeable outer layer. More suitably, permeable, in the context of the semi-permeable outer layer, means that approximately 30%, 25%, 20%, 15%, 10%, 5%, 3%, 2% or 1% (but greater than 0%) of the substance (e.g., the analyte(s) and optionally oxygen) permeates through the semi-permeable outer layer.

[0045] To avoid high current density (-200 pA / cm2) and minimise saturation of the microneedle electrode, the analyte diffusivity of the semi-permeable outer layer is suitably controlled. The analyte diffusivity of the semi-permeable outer layer is measured using a vertical diffusion (Franz) cell to determine the change in concentration gradient with time, the calculation applied is then derived from Fick's 1st Law of diffusion, rearranged to find "D". Further information on calculating the analyte diffusivity can be found in D N H Pg Sulaiman et al 2020 IOP Conf. Ser.: Mater. Sci. Eng. 991 012103. It will be understood that analyte diffusivity is applicable to the total concentration of analyte(s) to be measured (e.g., glucose and lactate when both are to be detected) Suitably, the analyte diffusivity of the semi-permeable outer layer is 5x10-10cm2 / s to 1x10-7cm2 / s. More suitably, the analyte diffusivity of the semi-permeable outer layer is 5x10-9cm2 / s to 1x10-8cm2 / s. Even more suitably, the analyte diffusivity of the semi-permeable outer layer is 5x10-9cm2 / s to 2x10-8cm2 / s. Most suitably, the analyte diffusivity of the semi-permeable outer layer is 8x10-9cm2 / s. The inventors have found that by controlling the analyte diffusivity of the semi-permeable outer layer, the linear range of detection of the microneedle electrodes can be extended up to -25 mM.

[0046] In some embodiments, the analyte diffusivity of the semi-permeable outer layer is 5x1 O'10cm2 / s to 1x10-7cm2 / s and the analyte is selected from glucose, ketones and / or lactate.Suitably, the analyte diffusivity of the semi-permeable outer layer is 5x10-9cm2 / s to 2x10-8cm2 / s.

[0047] The semi-permeable outer layer may have a reduced permeability - relative to the analyte(s) and optionally oxygen - to interferents. Interferents can lead to inaccuracies in detecting and measuring the analyte(s) (e.g., interferents). Suitably, the semi-permeable outer layer has greater permeability to the analyte(s) and optionally oxygen relative to interferents. Accordingly, if the semi-permeable outer layer permeates 30% of the analyte(s) and optionally oxygen, the semi-permeable outer layer will permeate <30% of the interferents. The nature of the interferent will be dependent on the analyte(s) to be detected (i.e., the analyte(s) to be detected cannot be the same as the interferents). For example, it may be that when glucose is the analyte to be detected, ascorbic acid may be an interferent. It may also be that when potassium is the analyte to be detected, sodium may be an interferent. Suitably, the interferent is acetaminophen (i.e., paracetamol), ascorbic acid, bilirubin, cholesterol, sodium, creatine, dopamine, ephedrine, ibuprofen, L-dopa, methyldopa, salicylate, tetracycline, tolazamide, tolbutamide, triglycerides, uric acid, L-glutathione and / or L-cystine, with the proviso that the interferent is different from the analyte(s) to be detected. More suitably, the interferent is ascorbic acid, uric acid, acetaminophen, L-glutathione and / or L-cystine. Even more suitably, the interferent is ascorbic acid, uric acid and / or acetaminophen.

[0048] The semi-permeable outer layer may be a unform layer covering all or part of the sensing layer. Suitably, the semi-permeable outer layer is a uniform layer covering all of the sensing layer. The semi-permeable outer layer may have a thickness of 0.01 pm to 80 pm. Suitably, the semi-permeable outer layer has a thickness of 0.1 pm to 70 pm. More suitably, the semi-permeable outer layer has a thickness of 1 pm to 60 pm. Even more suitably, the semi-permeable outer layer has a thickness of 5 pm to 50 pm. Yet even more suitably, the semi-permeable outer layer has a thickness of 7 pm to 40 pm. Yet even more suitably, the semi-permeable outer layer has a thickness of 10 pm to 37 pm. Most suitably, the semi-permeable outer layer has a thickness of 10 pm to 30 pm.

[0049] Suitably, the semi-permeable outer layer comprises (or is formed from) a first polymeric material. The first polymeric material may comprise a hydrophobic portion and / or a hydrophilic portion. The hydrophobic portion may have log Kow (also known as pKow) values of more than 0. The hydrophilic portion may have log ow values of less than 0. Log ow values are common in the art to determine hydrophobicity / hydrophilicity on the basis of the partition of a species between octanol and water. The presence of a hydrophobic portion and a hydrophilic portion prevents I minimises non-specific adsorption (e.g., of a protein to the microneedle electrode surface), a common problem associated with biosensor technology. Non-specific adsorption may decrease sensitivity and specificity of the microneedle electrode.

[0050] The first polymeric material may be selected from the group consisting of zwitterionicpolymers (e.g., betaines), polyacrylic acid, polyethylene glycol-NH2, polyethylene glycol, polyethylenimine, Parylene C, epoxy-polyurethane, Nation, poly(vinylpyridine), copolymers of vinylpyridine with styrene and / or butylmethacrylate, polyvinylimidazole, polyallylamine, polyaniline, polyvinylpyrrolidone, polystyrene, poly(styrene-ethylene-butylene), poly(styrene-butadiene-styrene), poly(styrene-isobutylene-styrene), urethanes, polyurethanes, copolymers of (1 -13C)alkyl chains, (2-13C) alkenyl chains, (2-13C) alkynyl chains, carbocycles and / or aryls, polysiloxanes and / or polyethers (linked by urethane, carbonate and / or urea linkages). Suitably, the first polymeric material is a zwitterionic polymer (e.g., betaines, carboxyl betaines, sulfo betaines or phosphor betaines), Parylene C and / or Nation. Suitably, the first polymeric material is a betaine (e.g., cocam idopropyl betaine, oleamidopropyl betaine, octyl sulfobetaine, caprylyl sulfobetaine, lauryl sulfobetaine, myristyl sulfobetaine, palmityl sulfobetaine, stearyl sulfobetaine, betaine (trimethylglycine), octyl betaine, phosphatidylcholine, glycine betaine, poly(carboxybetaine) (pCB), and poly(sulfobetaine) (pSB)).

[0051] The first polymeric material may be optionally substituted with one or more groups selected from (1-20C)alkyl, (2-20C)alkenyl, (2-20C)alkynyl, (1-20C)alkoxy (any of which may be optionally substituted), optionally substituted zwitterionic polymers, optionally substituted carbocycles, optionally substituted aryls, amino, hydroxy, carboxy, sulphonate and phosphate.

[0052] The first polymeric material may be a co-polymer (e.g., alternating, random or block co-polymer). Suitably, the first polymeric material is a random co-polymer. The co-polymers may be formed by any suitable monomers, such as zwitterionic, hydrophobic and hydrophilic monomers.

[0053] Suitably, the first polymeric material is a co-polymer (e.g., alternating, random or block co-polymer) comprising monomers A, B, C and D, wherein A, B, C and D are each independently selected from a monomer comprising a zwitterionic portion, a monomer comprising a pyridine portion, a monomer comprising a phenyl portion and a monomer comprising a hydrophilic portion. In some embodiments, the first polymeric material is a random co-polymer.

[0054] Suitably, the first polymeric material is a co-polymer (e.g., alternating, random or block co-polymer) comprising monomers A, B, C and D:-Aa_Bb-Cc- Dd_wherein:A, B, C and D are each independently selected from a monomer comprising a zwitterionic portion, a monomer comprising a pyridine portion, a monomer comprising a phenyl portion and a monomer comprising a hydrophilic portion;a is 0 to 25 mol%;b is 50 to 90 mol%;c is 0.1 to 10 mol%; andd is 0 to 25 mol%,wherein the sum total of a, b, c and d is 100 mol%.

[0055] Most suitably, the first polymeric material is a random co-polymer. A random copolymer comprising monomers A, B, C and D, means that monomers A, B, C and D are in a random order. For example, the first polymeric material may be a random co-polymer comprising monomers A, B, C and D in any one or more of the following configurations: -Aa-Bb-Cc-Dd-, -Aa-Bb-Dd-Cc", -Aa-Cc-Bb-Dd-, -Aa-Cc-Dd-B -, -Aa-Dd-B -Cc", -Aa-Dd-Cc-B -, -B -Aa-Cc" Dd_, -B -Aa-Dd-Cc", -B -Cc-Aa-Dd", -B -Cc-Dd-Aa-, -B -Dd-Aa-Cc", -B -Dd-Cc-Aa-, -Cc-Aa-B -Dd-, -Cc-Aa-Dd-B ", -Cc-B -Aa-Dd-, -Cc-B -Dd-Aa-, -Cc-Dd-Aa-B -, -Cc-Dd-B -Aa-, -Dd-Aa-B -Cc", -Dd-Aa- Cc-Bb-, -Dd-Bb-Aa-Cc-, -Dd-Bb-Cc-Aa-, -Dd-Cc-Aa-Bb- and / or -Dd-Cc-Bb-Aa-. It will be understood that the either side of the aforementioned configurations indicates connection (e.g., a bond) to another monomer A, B, C or D.

[0056] All of A, B, C and D may be identical. Alternatively, three of A, B, C and D may be identical. Further alternatively, two of A, B, C and D may be identical. Suitably, each of A, B, C and D are different.

[0057] A may be a monomer comprising a zwitterionic portion, a monomer comprising a pyridine portion, a monomer comprising a phenyl portion or a monomer comprising a hydrophilic portion. Suitably, A is a monomer comprising a zwitterionic portion.

[0058] B may be a monomer comprising a zwitterionic portion, a monomer comprising a pyridine portion, a monomer comprising a phenyl portion or a monomer comprising a hydrophilic portion. Suitably, B is monomer comprising a pyridine portion.

[0059] C may be a monomer comprising a zwitterionic portion, a monomer comprising a pyridine portion, a monomer comprising a phenyl portion or a monomer comprising a hydrophilic portion. Suitably C is a monomer comprising a hydrophilic portion.

[0060] D may be a monomer comprising a zwitterionic portion, a monomer comprising a pyridine portion, a monomer comprising a phenyl portion or a monomer comprising a hydrophilic portion. Suitably, D is monomer comprising a phenyl portion.

[0061] The following definitions for the monomer comprising a zwitterionic portion, the monomer comprising a pyridine portion, the monomer comprising a phenyl portion, and the monomer comprising a hydrophilic portion are in relation to the first polymeric material.

[0062] Each monomer comprising a zwitterionic portion may comprise a functional group comprising N+and a functional group comprising O'. Suitably, each monomer comprising a zwitterionic portion comprises a pyridinium positively charged functional group. Suitably, each monomer comprising a zwitterionic portion independently comprises a CO2' negatively charged functional group or an SOs' negatively charged functional group. In some embodiments, each monomer comprising a zwitterionic portion comprises a pyridiniumpositively charged functional group and one of a CC>2' negatively charged functional group or an SOs' negatively charged functional group.

[0063] Suitably, each monomer comprising a zwitterionic portion is independently selected from:

[0064] More suitably, each monomer comprising a zwitterionic portion is independently selected from:

[0065] More suitably, each monomer comprising a zwitterionic portion is independently selected from:

[0066] Most suitably, each monomer comprising a zwitterionic portion is:

[0067] Each monomer comprising a pyridine portion may be:

[0068] Each monomer comprising a hydrophilic portion may comprise a polyethylene glycol (PEG) chain. The PEG chain of the hydrophilic portion (i.e., not the whole monomercomprising a hydrophilic portion) may have a molecular weight of 100 to 10000 Da. Suitably, the PEG chain of the hydrophilic portion has a molecular weight of 200 to 5000 Da. More suitably, the PEG chain of the hydrophilic portion has a molecular weight of 300 to 2000 Da. Even more suitably, the PEG chain of the hydrophilic portion has a molecular weight of 350 to 1000 Da. Yet even more suitably, the PEG chain of the hydrophilic portion has a molecular weight of 400 to 700 Da. Most suitably, the PEG chain of the hydrophilic portion has a molecular weight of 500 to 600 Da.

[0069] Each monomer comprising a hydrophilic portion may comprise a PEG chain and a pyridinium moiety.

[0070] Suitably, each monomer comprising a hydrophilic portion is independently selected from:

[0071] Most suitably, each monomer comprising a hydrophilic portion is:

[0072] Each monomer comprising a phenyl portion may be a styrene monomer:

[0073] Suitably, the first polymeric material is a random co-polymer comprising the following monomers:wherein a, b, c and d are as defined herein.

[0074] More suitably, the first polymeric material comprises:wherein a, b, c and d are as defined herein.

[0075] As described hereinbefore, a is 0 to 25 mol%, b is 50 to 90 mol%, c is 0.1 to 10 mol%, and d is 0 to 25 mol%, wherein the sum total of a, b, c and d is 100 mol%. It will be understood that a, b, c and d refer to the proportion of each monomer in the first polymeric material.

[0076] Suitably, a is 2 to 20 mol%. More suitably, a is 5 to 15 mol%. Even more suitably, a is 8 to 12 mol%. More suitably a is 9 to 11 mol%. More suitably a is 9.5 to 10.5 mol%. Most suitably, a is 10 mol%. In some embodiments, a is 9.5 to 10 mol %.

[0077] Suitably, b is 60 to 90 mol%. More suitably, b is 70 to 85 mol%. Even more suitably, bis 75 to 80 mol%. More suitably, b is 78 to 80 mol%. More suitably, b is 78 to 79.8 mol%. Most suitably, b is 78.25 to 79.8 mol%.

[0078] Suitably, c is 0.1 to 5 mol%. More suitably, c is 0.1 to 4 mol%. Even more suitably, c is 0.2 to 3 mol%. More suitably, c is 0.2 to 2 mol%. More suitably, c is 0.3 to 1 mol%. Most suitably, c is 0.5 to 0.75 mol%.

[0079] Suitably, d is 2 to 20 mol%. More suitably, d is 5 to 15 mol%. Even more suitably, d is 8 to 12 mol%. More suitably d is 9 to 11 mol%. More suitably d is 9.5 to 10.5 mol%. Most suitably, d is 10 mol%. In some embodiments, d is 10 to 10.5 mol %.

[0080] Suitably, a is 2 to 20 mol%, b is 60 to 90 mol%, c is 0.1 to 5 mol% and d is 2 to 20 mol%, wherein the sum total of a, b, c and d is 100 mol%. More suitably, a is 5 to 15 mol%, b is 70 to 85 mol%, c is 0.1 to 4 mol% and d is 5 to 15 mol%, wherein the sum total of a, b, c and d is 100 mol%. Even more suitably, a is 8 to 12 mol%, b is 75 to 80 mol%, c is 0.2 to 3 mol% and d is 8 to 12 mol%, wherein the sum total of a, b, c and d is 100 mol%. Most suitably, a is 10 mol%, b is 78 to 79.8 mol%, c is 0.2 to 2 mol% and d is 10 mol%. In some embodiments, a is 9 to 11 mol%, b is 78 to 80 mol%, c is 0.2 to 2 mol% and d is 9 to 11 mol%, wherein the sum total of a, b, c and d is 100 mol%. In some embodiments, a is 9.5 to 10.5 mol%, b is 78 to 79.8 mol%, c is 0.3 to 1 mol% and d is 9.5 to 10.5 mol%, wherein the sum total of a, b, c and d is 100 mol%. In some embodiments, a is 9.5 to 10 mol%, b is 78.25 to 79.8 mol%, c is 0.5 to 0.75 mol% and d is 10 to 10.5 mol%, wherein the sum total of a, b, c and d is 100 mol%.

[0081] It may be that the first polymeric material is crosslinked with one or more crosslinkers. While a plurality of crosslinkers may be used, it is preferred that one crosslinker is used. Suitably, the crosslinker is selected from the group consisting of isocyanate, carbodiimide, glutaraldehyde, aziridine, silane, or other aldehydes, epoxy, di epoxy, acrylates, free-radical based agents, such as ethylene glycol diglycidyl ether (EGDE), poly(ethylene glycol) diglycidyl ether (PEGDE), or dicumyl peroxide (DCP), glycerol triglycidyl ether, Tris(4-hydroxyphenyl)methane triglycidyl ether, trimethylolpropane triglycidyl ether, trimethylolethane triglycidyl ether, N, N-diglycidyl-4-glycidyloxyaniline, neopentyl glycol diglycidyl ether and 1,4-butanediol diglycidyl ether. More suitably, the crosslinker is selected from the group consisting of ethylene glycol diglycidyl ether (EGDE), poly(ethylene glycol) diglycidyl ether (PEGDE), dicumyl peroxide (DCP), glycerol triglycidyl ether, Tris(4-hydroxyphenyl)methane triglycidyl ether, trimethylolpropane triglycidyl ether, trimethylolethane triglycidyl ether, N, N-diglycidyl-4-glycidyloxyaniline, neopentyl glycol diglycidyl ether and 1,4-butanediol diglycidyl ether. Most suitably, the crosslinker is glycerol triglycidyl ether.

[0082] The crosslinker(s) may be present in an amount of 0.01 to 25 wt% of the total weight of the semi-permeable outer layer. Suitably, the crosslinker(s) is I are present in amount of 1 to 20 wt% of the total weight of the semi-permeable outer layer. More suitably, thecrosslinker(s) is I are present in amount of 2 to 15 wt% of the total weight of the semi-permeable outer layer. Even more suitably, the crosslinker(s) is I are present in amount of 3 to 10 wt% of the total weight of the semi-permeable outer layer. Most suitably, the crosslinker(s) is I are present in an amount of 6 wt% of the total weight of the semi-permeable outer layer.

[0083] In an embodiment, the crosslinker(s) is I are present in amount of 1 to 20 wt% of the total weight of the semi-permeable outer layer, and the crosslinker(s) is I are selected from the group consisting of ethylene glycol diglycidyl ether (EGDE), poly(ethylene glycol) diglycidyl ether (PEGDE), dicumyl peroxide (DCP), glycerol triglycidyl ether, Tris(4-hydroxyphenyl)methane triglycidyl ether, trimethylolpropane triglycidyl ether, trimethylolethane triglycidyl ether, N, N-diglycidyl-4-glycidyloxyaniline, neopentyl glycol diglycidyl ether and 1,4-butanediol diglycidyl ether. In an embodiment, the crosslinker is present in amount of 3 to 10 wt% of the total weight of the semi-permeable outer layer, and the crosslinker is glycerol triglycidyl ether.

[0084] When a crosslinker is present, the remaining weight of the semi-permeable outer layer may be made up by the first polymeric material. Accordingly, the first polymeric material may be present in an amount of 75 to 99.99 wt% of the total weight of the semi-permeable outer layer. Suitably, the first polymeric material is present in amount of 80 to 99 wt% of the total weight of the semi-permeable outer layer. More suitably, the first polymeric material is present in amount of 85 to 98 wt% of the total weight of the semi-permeable outer layer. Even more suitably, the first polymeric material is present in amount of 90 to 97 wt% of the total weight of the semi-permeable outer layer. Most suitably, the first polymeric material is present in an amount of 94 wt% of the total weight of the semi-permeable outer layer.

[0085] Suitably, the semi-permeable outer layer comprises 0.01 to 25 wt% of the crosslinker(s) and 75 to 99.99 wt% of the first polymeric material, wherein the sum total of the crosslinker(s) and the first polymeric material is 100 wt%.

[0086] Suitably, the semi-permeable outer layer comprises I consists of I consists essentially of 1 to 20 wt% of the crosslinker(s) and 80 to 99 wt% of the first polymeric material, wherein the sum total of the crosslinker(s) and the first polymeric material is 100 wt%.

[0087] Suitably, the semi-permeable outer layer comprises I consists of I consists essentially of 2 to 15 wt% of the crosslinker(s) and 85 to 98 wt% of the first polymeric material, wherein the sum total of the crosslinker(s) and the first polymeric material is 100 wt%.

[0088] Suitably, the semi-permeable outer layer comprises I consists of I consists essentially of 3 to 10 wt% of the crosslinker(s) and 90 to 97 wt% of the first polymeric material, wherein the sum total of the crosslinker(s) and the first polymeric material is 100 wt%.

[0089] Suitably, the semi-permeable outer layer comprises I consists of I consists essentially of 6 wt% of the crosslinker(s) and 94 wt% of the first polymeric material.

[0090] The semi-permeable outer layer may optionally comprise an outermost biocompatible layer. It will be understood that the term “outermost” means that this layer is located on the surface of the semi-permeable outer layer which is exposed to the external environment. This external environment could be, for example, interstitial fluid in the dermis of the skin. The outermost biocompatible layer is particularly useful in adapting the microneedle electrodes such that they exhibit the appropriate host response, or are more compatible in the environment in which they are located. The outermost biocompatible layer can also improve resistance to insertion-related impact forces, which would typically not be a consideration for previous technologies relying on the use of hypodermic needles for insertion. The outermost biocompatible layer may be a second polymeric material, wherein the second polymeric material has a definition as described above in relation to the first polymeric material. In some embodiments, the outermost biocompatible layer is a secondary polymeric material identical to the first polymeric material. In some embodiments, the outermost biocompatible layer is a secondary polymeric material different to the first polymeric material.

[0091] The outermost biocompatible layer may have a viscoelasticity of > 1x104Pa. It will be understood that the viscoelasticity is determined when the outermost biocompatible layer is fully hydrated by immersion in water for at least 24 hours. The viscoelasticity can be measured according to ISO 6721-1 by characterising the relationship between the storage and loss moduli using a rheometer.

[0092] The sensing layer is positioned underneath the semi-permeable outer layer and is capable of generating an electrochemical response in the presence of the analyte(s). The sensing layer allows the microneedle electrode to detect and measure the concentration of the analyte that has permeated through the semi-permeable outer layer by producing a detectable electrochemical response (also referred to as a current signal). The current signal is proportional to the concentration of the analyte and is relayed to the microneedle electrode. By measuring the current signal, the microneedle electrode can therefore measure the concentration of the analyte.

[0093] The sensing layer may have a mean sensing area of 10,000 to 250,000 pm2. The mean sensing area will be understood as the average area of the sensing layer which can generate an electrochemical response in the presence of the analyte(s). Suitably, the sensing layer has a mean sensing area of 15,000 to 185,000 pm2. More suitably, the sensing layer has a mean sensing area of 20,000 to 150,000 pm2. More suitably, the sensing layer has a mean sensing area of 30,000 to 125,000 pm2. More suitably, the sensing layer has a mean sensing area of 40,000 to 110,000 pm2. Most suitably, the sensing layer has a mean sensing area of 50,000 to 100,000 pm2.

[0094] The sensing layer may comprise: i) an enzyme or an ionophore; and ii) a third polymeric material. The enzyme or ionophore may be immobilized within the sensing layer(e.g., the enzyme or ionophore is located within the sensing layer with restricted movement). For example, the enzyme or ionophore may be covalently bound to the third polymeric material of the sensing layer. Suitably, the enzyme or ionophore (e.g., via functional amine groups) is covalently bound via a di-epoxy crosslinker to the functional groups of the third polymeric material of the sensing layer (e.g., amine or hydroxyl groups).

[0095] In some embodiments, the sensing layer comprises: i) an enzyme; and ii) a third polymeric material.

[0096] The enzyme may be selected from the group consisting of glucose oxidase, lactate oxidase, glucose dehydrogenase, diamine oxidase, p-hydroxybutyrate dehydrogenase, alcohol oxidase, cholesterol oxidase, choline oxidase, alcohol dehydrogenase, lactate dehydrogenase, glutamate dehydrogenase, glutamate oxidase, glycerophosphate oxidase, glycerol dehydrogenase, leucine dehydrogenase, p-hydroxybenzoate hydroxylase, pyruvate oxidase, sarcosine oxidase, uricase, catechol oxidase, laccase, tyrosinase, and xanthine oxidase. Suitably, the enzyme is selected from the group consisting of glucose oxidase, lactate oxidase, glucose dehydrogenase, p-hydroxybutyrate dehydrogenase and lactate dehydrogenase. More suitably, the enzyme is selected from the group consisting of glucose oxidase, lactate oxidase, p-hydroxybutyrate dehydrogenase and lactate dehydrogenase. Even more suitably, the enzyme is glucose oxidase.

[0097] The enzyme may be present in an amount of 10 wt% to 80 wt% of the total weight of the sensing layer. Suitably, the enzyme is present in an amount of 20 wt% to 70 wt% of the total weight of the sensing layer. More suitably, the enzyme is present in an amount of 30 wt% to 60 wt% of the total weight of the sensing layer. Even more suitably, the enzyme is present in an amount of 33 wt% to 58 wt% of the total weight of the sensing layer. Yet even more suitably, the enzyme is present in an amount of 35 wt% to 55 wt% of the total weight of the sensing layer. Most suitably, the enzyme is present in an amount of 40 wt% to 50 wt% of the total weight of the sensing layer. In some embodiments, the enzyme is present in an amount of 15 wt% to 25 wt% of the total weight of the sensing layer. In some embodiments, the enzyme is present in an amount of 30 wt% to 40 wt% of the total weight of the sensing layer. In some embodiments, the enzyme is present in an amount of 50 wt% to 60 wt% of the total weight of the sensing layer. The remaining weight of the sensing layer may be made up of the third polymeric material.

[0098] In an embodiment, the enzyme is present in an amount of 30 wt% to 60 wt% of the total weight of the sensing layer, and the enzyme is selected from the group consisting of glucose oxidase, lactate oxidase, glucose dehydrogenase, diamine oxidase, p-hydroxy butyrate dehydrogenase and alcohol oxidase. In an embodiment, the enzyme is present in an amount of 20 wt% to 70 wt% of the total weight of the sensing layer, and the enzyme is selected from the group consisting of glucose oxidase, lactate oxidase, glucosedehydrogenase, p-hydroxybutyrate dehydrogenase and lactate dehydrogenase. In an embodiment, the enzyme is present in an amount of 35 wt% to 55 wt% of the total weight of the sensing layer, and the enzyme is glucose oxidase. In an embodiment, the enzyme is present in an amount of 40 wt% to 50 wt% of the total weight of the sensing layer, and the enzyme is glucose oxidase. In an embodiment, the enzyme is present in an amount of 33 wt% to 58 wt% of the total weight of the sensing layer, and the enzyme is glucose oxidase, lactate oxidase, lactate dehydrogenase or p-hydroxybutyrate dehydrogenase. In an embodiment, the enzyme is present in an amount of 33 wt% to 58 wt% of the total weight of the sensing layer, and the enzyme is lactate oxidase or lactate dehydrogenase. In an embodiment, the enzyme is present in an amount of 15 wt% to 25 wt% of the total weight of the sensing layer, and the enzyme is p-hydroxybutyrate dehydrogenase.

[0099] The sensing layer may comprise 0.01 pg to 1 pg loading of the enzyme per microneedle electrode. Suitably, the sensing layer comprises 0.05 pg to 0.5 pg loading of the enzyme per microneedle electrode. More suitably, the sensing layer comprises 0.075 pg to 0.2 pg loading of the enzyme per microneedle electrode. Most suitably, the sensing layer comprises 0.09 pg to 0.12 pg loading of the enzyme per microneedle electrode.

[0100] In some embodiments, the sensing layer comprises: i) an ionophore; and ii) a third polymeric material.

[0101] Ionophores are chemical species that reversibly bind and transport ions (e.g., sodium, potassium, magnesium, calcium, hydrogen (e.g., for pH monitoring), bicarbonate, zinc, chloride, ammonium, lithium, copper, iron or phosphate), for example, across a membrane. These exemplary ions are analytes are described herein.

[0102] The ionophore may be present in an amount of 0.5 wt% to 20 wt% of the total weight of the sensing layer. Suitably, the ionophore is present in an amount of 0.75 wt% to 15 wt% of the total weight of the sensing layer. More suitably, the ionophore is present in an amount of 1 wt% to 10wt% of the total weight of the sensing layer. Most suitably, the ionophore is present in an amount of 2 wt% to 5 wt% of the total weight of the sensing layer. The remaining weight of the sensing layer may be made up of the third polymeric material.

[0103] The third polymeric material may be selected from the group consisting of zwitterionic polymers (e.g., betaines), polyacrylic acid, polyethylene glycol-NH2, polyethylene glycol, polyethylenimine, Parylene C, epoxy-polyurethane, Nation, poly(vinylpyridine), polyvinylchloride, copolymers of vinylpyridine with styrene and / or butylmethacrylate, polyvinylimidazole, polyallylamine, polyaniline, polyvinylpyrrolidone, polystyrene, poly(styrene-ethylene-butylene), poly(styrene-butadiene-styrene), poly(styrene-isobutylene-styrene), urethanes, polyurethanes, co-polymers of (1-13C)alkyl chains, (2-13C) alkenyl chains, (2-13C) alkynyl chains, carbocycles and / or aryls, polysiloxanes and / or polyethers (linked by urethane, carbonate and / or urea linkages). Suitably, the third polymeric material isa zwitterionic polymer, Parylene C, Nation, poly(vinylpyridine) and / or copolymers of vinylpyridine with styrene and / or butylmethacrylate. More suitably, the third polymeric material is a zwitterionic polymer (e.g., betaines, carboxyl betaines, sulfo betaines or phosphor betaines), poly(vinylpyridine), Parylene C and / or Nation. Suitably, the third polymeric material is a betaine (e.g., cocamidopropyl betaine, oleamidopropyl betaine, octyl sulfobetaine, caprylyl sulfobetaine, lauryl sulfobetaine, myristyl sulfobetaine, palmityl sulfobetaine, stearyl sulfobetaine, betaine (trimethylglycine), octyl betaine, phosphatidylcholine, glycine betaine, poly(carboxybetaine) (pCB), and poly(sulfobetaine) (pSB)).

[0104] The third polymeric material may be optionally substituted with one or more groups selected from (1-20C)alkyl, (2-20C)alkenyl, (2-20C)alkynyl, (1-20C)alkoxy (any of which may be optionally substituted), optionally substituted zwitterionic polymers, optionally substituted carbocycles, optionally substituted aryls, amino, hydroxy, carboxy, sulphonate and phosphate.

[0105] In some embodiments, the third polymeric material comprises a monomer comprising a redox mediator, wherein the monomer comprising a redox mediator is as defined herein.

[0106] The third polymeric material may be a co-polymer (e.g., alternating, random or block co-polymer). Suitably, the third polymeric material is a random co-polymer. The copolymers may be formed by any suitable monomers, such as zwitterionic, hydrophobic and hydrophilic monomers.

[0107] Suitably, the third polymeric material is a co-polymer (e.g., alternating, random or block co-polymer) comprising monomers X, Y and Z, wherein X, Y and Z are each independently selected from a monomer comprising a zwitterionic portion, a monomer comprising a pyridine portion and a monomer comprising a redox mediator. In some embodiments, the third polymeric material is a random co-polymer.

[0108] Suitably, the third polymeric material is a co-polymer (e.g., alternating, random or block co-polymer) comprising monomers X, Y and Z:-Xx1 -Yy1-Zz1"wherein:X, Y and Z are each independently selected from a monomer comprising a redox mediator, a monomer comprising a pyridine portion and a monomer comprising a zwitterionic portion; x1 is 5 to 25 mol%;y1 is 70 to 95 mol%; andz1 is 0 to 25 mol%,wherein the sum total of x1, y1 and z1 is 100 mol%.

[0109] Most suitably, the third polymeric material is a random co-polymer. A random co-polymer comprising monomers X, Y and Z, means that monomers X, Y and Z are in arandom order. For example, the third polymeric material may be a random co-polymer comprising monomers X, Y and Z in any one or more of the following configurations: -Xxi-Yyi-Zzi-, -Xxi-Zzi-Yyi -, “Yyi “Xxi -Zzi -, -Yyi -Zzi -Xxi -Zzi “Xxi- Yyi - and / or -Zzi-Yyi-Xxi-. It will be understood that the either side of the aforementioned configurations indicates connection (e.g., a bond) to another monomer X, Y or Z.

[0110] All of X, Y and Z may be identical. Alternatively, two of X, Y and Z may be identical. Suitably, each of X, Y and Z are different.

[0111] X may be a monomer comprising a zwitterionic portion, a monomer comprising a pyridine portion or a monomer comprising a redox mediator. Suitably, X is a monomer comprising a redox mediator.

[0112] Y may be a monomer comprising a zwitterionic portion, a monomer comprising a pyridine portion or a monomer comprising a redox mediator. Suitably, Y is monomer comprising a pyridine portion.

[0113] Z may be a monomer comprising a zwitterionic portion, a monomer comprising a pyridine portion or a monomer comprising a redox mediator. Suitably Z is a monomer comprising a zwitterionic portion.

[0114] The following definitions for the monomer comprising a zwitterionic portion, the monomer comprising a pyridine portion, and the monomer comprising a redox mediator are in relation to the third polymeric material.

[0115] Each monomer comprising a zwitterionic portion may comprise a functional group comprising N+and a functional group comprising O'. Suitably, each monomer comprising a zwitterionic portion comprises a pyridinium positively charged functional group. Suitably, each monomer comprising a zwitterionic portion comprises a CO2' negatively charged functional group. In some embodiments, each monomer comprising a zwitterionic portion comprises a pyridinium positively charged functional group and a CC>2' negatively charged functional group.

[0116] Suitably, each monomer comprising a zwitterionic portion is independently selected from:

[0117] More suitably, each monomer comprising a zwitterionic portion is independently selected from:

[0118] Even more suitably, each monomer comprising a zwitterionic portion is independently selected from:

[0119] Most suitably, each monomer comprising a zwitterionic portion is:

[0120] Each monomer comprising a pyridine portion may be:

[0121] Each monomer comprising a redox mediator may comprise a pyridinium moiety or an imidazolium moiety. The pyridinium moiety or imidazolium moiety may be bound directly to the redox mediator, or via a linking group. The linking group may be a (1-20C)alkyl chain optionally comprising an amide group within the (1-20C)alkyl chain.

[0122] Suitably, each monomer comprising a redox mediator is independently selected from:wherein RM is a redox mediator as defined herein.

[0123] Most suitably, each monomer comprising a redox mediator is:wherein RM is a redox mediator as defined herein.

[0124] Redox mediators are electron transfer agents suitable for carrying electrons between an analyte, an analyte-reduced enzyme or an analyte-oxidized enzyme and the surface of the microneedle electrode (e.g., the conductive electrode layer), either directly, or via one or more additional electron transfer agents. Notwithstanding this, redox mediators also have a redox potential which is designed for the electrooxidation of an analyte.

[0125] The redox mediator may be a transition metal complex comprising iron, ferrocene, cobalt, ruthenium, osmium, manganese, copper and / or vanadium. Most suitably, the redox mediator is a transition metal complex comprising osmium.

[0126] Suitably, the redox mediator comprises at least one bidentate ligand. More suitably, the redox mediator comprises at least one bidentate ligand comprising a heterocycle or a heteroaryl ring. Even more suitably, the redox mediator comprises at least one bidentate ligand comprising a N-containing heterocycle or a N-containing heteroaryl. Yet still more suitably, the redox mediator comprises at least one bidentate ligand comprising a 5- or 6-membered N-containing heterocycle or a 5- or 6-membered N-containing heteroaryl. Yet still even more suitably, the redox mediator comprises at least one bidentate ligand comprising an imidazole ring and / or a pyridine ring. Yet still even more suitably, the redox mediator comprises at least one bidentate ligand comprising a bis-imidazole moiety, at least one bidentate ligand comprising a bipyridyl moiety and / or at least one bidentate ligand comprising a pyridyl-imidazole moiety. Most suitably, the redox mediator comprises two bidentate ligands comprising a bis-imidazole moiety and one bidentate ligand comprising a pyridyl-imidazolemoiety. The bidentate ligands may each independently be optionally substituted with one or more (1 -6C)alkyl groups and / or (1-6C)alkoxy groups.

[0127] Examples of suitable bidentate ligands include 2,2'-bisimidazole, 2,2'-bisoxazole, 2,2'-bisthiazole, 2,2'-bipyridine (bipy), 2-(2-pyridyl) imidazole, and derivatives thereof, all of which may be optionally substituted as described above. Suitably, the redox mediator comprises at least one bidentate ligand selected from the group consisting of 2,2'-bisimidazole, 2,2'-bisoxazole, 2,2'-bisthiazole, 2,2'-bipyridine (bipy), 2-(2-pyridyl) imidazole, all of which may be optionally substituted with (1 -3C)alkyl, preferably methyl. The redox mediator is particularly well suited to accept electrons from, or transfer electrons to the enzyme or at least one analyte at a high rate and also exchange electrons rapidly with the surface of the microneedle electrode (e.g., the conductive electrode layer).

[0128] Suitably, each monomer comprising a redox mediator is independently selected from:wherein M is selected from the group consisting of iron, cobalt, ruthenium, osmium, manganese, copper and vanadium;bipy is bi-pyridine; andeach X2 is independently selected from H, (1-6C) alkyl and (1-6C) alkoxy.

[0129] Most suitably, M is osmium

[0130] Suitably, each X2 is independently selected from H, (1-3C) alkyl and (1-3C) alkoxy. More suitably, each X2 is independently selected from H, methyl or methoxy. Most suitably, each X2 is methyl.

[0131] More suitably, each monomer comprising a redox mediator is independently selected from:wherein M is selected from the group consisting of iron, cobalt, ruthenium, osmium, manganese, copper and vanadium; andbipy is bi-pyridine.

[0132] Most suitably, M is osmium.

[0133] More suitably, each monomer comprising a redox mediator is independently selected from:wherein M is selected from the group consisting of iron, cobalt, ruthenium, osmium, manganese, copper and vanadium; andeach X2 is independently selected from H, (1-6C) alkyl and (1-6C) alkoxy.

[0134] Most suitably, M is osmium.

[0135] Suitably, each X2 is independently selected from H, (1-3C) alkyl and (1-3C) alkoxy. More suitably, each X2 is independently selected from H, methyl or methoxy. Most suitably, each X2 is methyl.

[0136] Suitably, the third polymeric material is a random co-polymer comprising the following monomers:wherein x1, y1 and z1 are as defined herein.

[0137] More suitably, the third polymeric material comprises:wherein x1, y1 and z1 are as defined herein.

[0138] Suitably, the third polymeric material is a random co-polymer comprising the following monomers:wherein x1, y1 and z1 are as defined herein.

[0139] More suitably, the third polymeric material comprises:wherein x1, y1 and z1 are as defined herein.

[0140] As described hereinbefore x1 is 5 to 25 mol%, y1 is 70 to 95 mol%; and z1 is 0 to 25 mol%, wherein the sum total of x1, y1 and z1 is 100 mol%. It will be understood that x1, y1 and z1 refer to the proportion of each monomer in the third polymeric material.

[0141] Suitably, x1 is 7 to 20 mol%. More suitably, x1 is 8 to 15 mol%. More suitably, x1 is 10 to 13 mol%. More suitably, x1 is 10 to 12.5 mol%. Most suitably, x1 is 10.2 to 12.3mol%.

[0142] In some embodiments, x1 is 8 to 18 mol%. More suitably, x1 is 10 to 18 mol%. More suitably, x1 is 15 to 17 mol%.

[0143] Suitably, y1 is 75 to 90 mol%. More suitably, y1 is 77 to 90 mol%. More suitably, y1 is 80 to 90 mol%. More suitably, y1 is 84 to 87 mol%. Most suitably, y1 is 85 to 86 mol%.

[0144] Suitably, z1 is 0 to 15 mol%. More suitably, z1 is 0 to 10 mol%. More suitably, z1 is 0 to 5 mol%. More suitably, z1 is 1.5 to 4.5 mol%. More suitably, z1 is 2.1 to 4.3 mol%.

[0145] Suitably, x1 is 7 to 20 mol%, y1 is 75 to 90 mol% and z1 is 0 to 15 mol%, wherein the sum total of x1, y1 and z1 is 100 mol%. More suitably, x1 is 8 to 15 mol%, y1 is 77 to 90 mol% and z1 is 0 to 10 mol%, wherein the sum total of x1, y1 and z1 is 100 mol%. More suitably, x1 is 10 to 13 mol%, y1 is 80 to 90 mol% and z1 is 0 to 5 mol%, wherein the sum total of x1, y1 and z1 is 100 mol%. More suitably, x1 is 10 to 12.5 mol%, y1 is 84 to 87 mol% and z1 is 1.5 to 4.5 mol%, wherein the sum total of x1, y1 and z1 is 100 mol%. Most suitably, x1 is 10.2 to 12.3 mol%, y1 is 85 to 86 mol% and z1 is 2.1 to 4.3 mol%, wherein the sum total of x1, y1 and z1 is 100 mol%.

[0146] In some embodiments, x1 is 8 to 18 mol%, y1 is 75 to 90 mol% and z1 is 0 to 15 mol%, wherein the sum total of x1, y1 and z1 is 100 mol%. More suitably, x1 is 10 to 18 mol%, y1 is 77 to 90 mol% and z1 is 0 to 10 mol%, wherein the sum total of x1, y1 and z1 is 100 mol%. More suitably, x1 is 15 to 17 mol%, y1 is 80 to 90 mol% and z1 is 0 to 5 mol%, wherein the sum total of x1, y1 and z1 is 100 mol%.

[0147] The third polymeric material may be present in an amount of 0.1 wt% to 99.5 wt% of the total weight of the sensing layer.

[0148] In some embodiments, the third polymeric material is present in an amount of 1 wt% to 75 wt% of the total weight of the sensing layer. Suitably, the third polymeric material is present in an amount of 10 wt% to 65 wt% of the total weight of the sensing layer. More suitably, the third polymeric material is present in an amount of 15 wt% to 60 wt% of the total weight of the sensing layer. More suitably, the third polymeric material is present in an amount of 25 wt% to 55 wt% of the total weight of the sensing layer. Most suitably, the third polymeric material is present in an amount of 30 wt% to 40 wt% of the total weight of the sensing layer. In some embodiments, the third polymeric material is present in an amount of 15 wt% to 40 wt% of the total weight of the sensing layer. In some embodiments, the third polymeric material is present in an amount of 15 wt% to 25 wt% of the total weight of the sensing layer. In some embodiments, the third polymeric material is present in an amount of 25 wt% to 35 wt% of the total weight of the sensing layer.

[0149] In some alternative embodiments, the third polymeric material is present in an amount of 50 wt% to 99.5 wt% of the total weight of the sensing layer. Suitably, the third polymeric material is present in an amount of 70 wt% to 99.25 wt% of the total weight of thesensing layer. More suitably, the third polymeric material is present in an amount of 80 wt% to 99 wt% of the total weight of the sensing layer. Most suitably, the third polymeric material is present in an amount of 90 wt% to 98 wt% of the total weight of the sensing layer.

[0150] It may be that the third polymeric material is crosslinked with one or more crosslinkers. While a plurality of crosslinkers may be used, it is preferred that one crosslinker is used. Suitably, the crosslinker is selected from the group consisting of isocyanate, carbodiimide, glutaraldehyde, aziridine, silane, or other aldehydes, epoxy, di epoxy, acrylates, free-radical based agents, such as ethylene glycol diglycidyl ether (EGDE), poly(ethylene glycol) diglycidyl ether (PEGDE), dicumyl peroxide (DCP), glycerol triglycidyl ether, Tris(4-hydroxyphenyl)methane triglycidyl ether, trimethylolpropane triglycidyl ether, trimethylolethane triglycidyl ether, N, N-diglycidyl-4-glycidyloxyaniline, neopentyl glycol diglycidyl ether and 1,4-butanediol diglycidyl ether. More suitably, the crosslinker is selected from the group consisting of ethylene glycol diglycidyl ether (EGDE), poly(ethylene glycol) diglycidyl ether (PEGDE), dicumyl peroxide (DCP), glycerol triglycidyl ether, Tris(4-hydroxyphenyl)methane triglycidyl ether, trimethylolpropane triglycidyl ether, trimethylolethane triglycidyl ether, N, N-diglycidyl-4-glycidyloxyaniline, neopentyl glycol diglycidyl ether and 1,4-butanediol diglycidyl ether. Most suitably, the crosslinker is poly(ethylene glycol) diglycidyl ether (PEGDE).

[0151] The crosslinker(s) may be present in an amount of 4 to 50 wt% of the total weight of the sensing layer. Suitably, the crosslinker(s) is I are present in amount of 6 to 45 wt% of the total weight of the sensing layer. Suitably, the crosslinker(s) is I are present in amount of 10 to 40 wt% of the total weight of the sensing layer. More suitably, the crosslinker(s) is I are present in amount of 15 to 35 wt% of the total weight of the sensing layer. Most suitably, the crosslinker(s) is I are present in amount of 15 to 20 wt% of the total weight of the sensing layer. In some embodiments, the crosslinker(s) is / are present in amount of 6 to 18 wt% of the total weight of the sensing layer. In some embodiments, the crosslinker(s) is I are present in amount of 5 to 10 wt% of the total weight of the sensing layer. In some embodiments, the crosslinker(s) is I are present in amount of 10 to 15 wt% of the total weight of the sensing layer. In some embodiments, the crosslinker(s) is I are present in amount of 7 to 12 wt% of the total weight of the sensing layer.

[0152] In an embodiment, the crosslinker(s) is I are present in amount of 10 to 40 wt% of the total weight of the sensing layer, and the crosslinker(s) is I are selected from the group consisting of ethylene glycol diglycidyl ether (EGDE), poly(ethylene glycol) diglycidyl ether (PEGDE), dicumyl peroxide (DCP), glycerol triglycidyl ether, Tris(4-hydroxyphenyl)methane triglycidyl ether, trimethylolpropane triglycidyl ether, trimethylolethane triglycidyl ether, N, N-diglycidyl-4-glycidyloxyaniline, neopentyl glycol diglycidyl ether and 1,4-butanediol diglycidyl ether. In an embodiment, the crosslinker is present in amount of 15 to 20 wt% of the totalweight of the sensing layer, and the crosslinker is poly(ethylene glycol) diglycidyl ether (PEGDE). In an embodiment, the crosslinker is present in amount of 6 to 18 wt% of the total weight of the sensing layer, and the crosslinker is poly(ethylene glycol) diglycidyl ether (PEGDE).

[0153] The sensing layer may comprise one or more additives, such as albumin (e.g., ovalbumin and / or human serum albumin). The one or more additives may be present in an amount of 10 to 50 wt% of the total weight of the sensing layer. Suitably, the one or more additives is present in an amount of 20 to 40 wt% of the total weight of the sensing layer. More suitably, the one or more additives is present in an amount of 30 to 40 wt% of the total weight of the sensing layer.

[0154] Suitably, the sensing layer comprises I consists of I consists essentially of 10 wt% to 80 wt% of the enzyme, 1 wt% to 75 wt% of the third polymeric material, and 4 wt% to 50 wt% of the crosslinker(s), wherein the sum total of the enzyme, the third polymeric material and the crosslinker(s) is 100 wt%.

[0155] Suitably, the sensing layer comprises I consists of I consists essentially of 30 wt% to 60 wt% of the enzyme, 10 wt% to 65 wt% of the third polymeric material, and 10 wt% to 40 wt% of the crosslinker(s), wherein the sum total of the enzyme, the third polymeric material and the crosslinker(s) is 100 wt%.

[0156] Suitably, the sensing layer comprises I consists of I consists essentially of 35 wt% to 55 wt% of the enzyme, 25 wt% to 55 wt% of the third polymeric material, and 15 wt% to 35 wt% of the crosslinker(s), wherein the sum total of the enzyme, the third polymeric material and the crosslinker(s) is 100 wt%.

[0157] Suitably, the sensing layer comprises I consists of I consists essentially of 40 wt% to 50 wt of the enzyme, 30 wt% to 40 wt% of the third polymeric material, and 15 wt% to 20 wt% of the crosslinker(s), wherein the sum total of the enzyme, the third polymeric material and the crosslinker(s) is 100 wt%.

[0158] Suitably, the sensing layer comprises I consists of I consists essentially of 20 wt% to 70 wt of the enzyme, 15 wt% to 40 wt% of the third polymeric material, and 6 wt% to 18 wt% of the crosslinker(s). It may be that the sum total of the enzyme, the third polymeric material and the crosslinker(s) is 100 wt%. It may alternatively be that the sensing layer also comprises one or more additives, as described herein. In this embodiment, the enzyme may be selected from the group consisting of glucose oxidase, lactate oxidase, glucose dehydrogenase, p-hydroxybutyrate dehydrogenase and lactate dehydrogenase.

[0159] Suitably, the sensing layer comprises I consists of I consists essentially of 40 wt% to 50 wt of the enzyme, 30 wt% to 40 wt% of the third polymeric material, and 15 wt% to 20 wt% of the crosslinker(s). It may be that the sum total of the enzyme, the third polymeric material and the crosslinker(s) is 100 wt%. It may alternatively be that the sensing layer alsocomprises one or more additives, as described herein. In this embodiment, the enzyme may be glucose oxidase.

[0160] Suitably, the sensing layer comprises I consists of I consists essentially of 30 wt% to 40 wt of the enzyme, 15 wt% to 25 wt% of the third polymeric material, and 5 wt% to 10 wt% of the crosslinker(s). It may be that the sensing layer also comprises one or more additives, as described herein. In this embodiment, the sensing layer may also comprise 10 wt% to 50 wt%, suitably 20 wt% to 40 wt%, albumin (e.g., ovalbumin and / or human serum albumin). In this embodiment, the enzyme may be lactate oxidase.

[0161] Suitably, the sensing layer comprises I consists of I consists essentially of 50 wt% to 60 wt of the enzyme, 25 wt% to 35 wt% of the third polymeric material, and 10 wt% to 15 wt% of the crosslinker(s). It may be that the sum total of the enzyme, the third polymeric material and the crosslinker(s) is 100 wt%. It may alternatively be that the sensing layer also comprises one or more additives, as described herein. In this embodiment, the enzyme may be lactate dehydrogenase.

[0162] Suitably, the sensing layer comprises I consists of I consists essentially of 15 wt% to 25 wt of the enzyme, 15 wt% to 25 wt% of the third polymeric material, and 7 wt% to 12 wt% of the crosslinker(s). It may be that the sensing layer also comprises one or more additives, as described herein. In this embodiment, the sensing layer may also comprise 10 wt% to 50 wt%, suitably 20 wt% to 40 wt%, albumin (e.g., ovalbumin and / or human serum albumin). In this embodiment, the enzyme may be p-hydroxybutyrate dehydrogenase.

[0163] Suitably, the sensing layer comprises 0.5 wt% to 20 wt% of the ionophore and 50 wt% to 99.5 wt% of the third polymeric material, wherein the sum total of the ionophore and the third polymeric material does not exceed 100 wt%. In this embodiment, the sensing layer may suitably comprise 80 wt% to 99.5 wt% of the third polymeric material.

[0164] Suitably, the sensing layer comprises 0.75 wt% to 15wt% of the ionophore and 70 wt% to 99.25 wt% of the third polymeric material, wherein the sum total of the ionophore and the third polymeric material does not exceed 100 wt%. In this embodiment, the sensing layer may suitably comprise 85 wt% to 99.25 wt% of the third polymeric material.

[0165] Suitably, the sensing layer comprises 1 wt% to 10 wt% of the ionophore and 80 wt% to 99 wt% of the third polymeric material, wherein the sum total of the ionophore and the third polymeric material does not exceed 100 wt%. In this embodiment, the sensing layer may suitably comprise 90 wt% to 99 wt% of the third polymeric material.

[0166] Suitably, the sensing layer comprises 2 wt% to 5 wt% of the ionophore and 90 wt% to 98 wt% of the third polymeric material, wherein the sum total of the ionophore and the third polymeric material does not exceed 100 wt%. In this embodiment, the sensing layer may suitably comprise 95 wt% to 98 wt% of the third polymeric material.

[0167] The composite material may optionally comprise an inner layer positionedbetween the sensing layer and the microneedle electrode. The inner layer can reduce noise and vibration, thereby providing support to the sensing layer. The inner layer may also improve the anti-interference properties (to supplement the anti-interference properties of the semi-permeable outer layer). The inner layer may comprise (or is formed from) a fourth polymeric material, wherein the fourth polymeric material has a definition as described above in relation to the first polymeric material, with the proviso that the fourth polymeric material is different from the first, second and third polymeric materials. Suitably, the fourth polymeric material is selected from the group consisting of zwitterionic polymers (e.g., betaines), polyacrylic acid, polyethylene glycol-NH2, polyethylene glycol, polyethylenimine, Parylene C, epoxypolyurethane, Nation, poly(vinylpyridine), copolymers of vinylpyridine with styrene and / or butylmethacrylate, polyvinylimidazole, polyallylamine, polyaniline, polyvinylpyrrolidone, polystyrene, poly(styrene-ethylene-butylene), poly(styrene-butadiene-styrene), poly(styrene-isobutylene-styrene), urethanes, polyurethanes, co-polymers of (1-13C)alkyl chains, (2-13C) alkenyl chains, (2-13C) alkynyl chains, carbocycles and / or aryls, polysiloxanes and / or polyethers (linked by urethane, carbonate and / or urea linkages). More suitably, the fourth polymeric material is a zwitterionic polymer (e.g., betaines, carboxyl betaines, sulfo betaines or phosphor betaines), Parylene C and / or Nation. Most suitably, the fourth polymeric material is Nation.

[0168] The inner layer may be permeable to hydrogen peroxide and / or electrons. Permeable, in the context of the inner layer, suitably means that greater than 0% of the substance (e.g., the hydrogen peroxide and / or electrons) permeates through the inner layer. More suitably, permeable, in the context of the inner layer, means that greater than or equal to 70%, 75%, 80%, 85%, 90%, 95%, 97% or 99% of the substance permeates through the inner layer.

[0169] The inner layer may have a reduced permeability - relative to the hydrogen peroxide and / or electrons - to interferents. Interferents can lead to inaccuracies in detecting and measuring the analyte(s) (e.g., interferents). Suitably, the inner layer has greater permeability to the hydrogen peroxide and / or electrons relative to interferents. Accordingly, if the inner layer permeates 70% of hydrogen peroxide and / or electrons, the inner layer will permeate <70% of the interferents. Suitably, the interferent is acetaminophen (i.e., paracetamol), ascorbic acid, bilirubin, cholesterol, sodium, creatine, dopamine, ephedrine, ibuprofen, L-dopa, methyldopa, salicylate, tetracycline, tolazamide, tolbutamide, triglycerides, uric acid, L-glutathione and / or L-cystine. More suitably, the interferent is ascorbic acid, uric acid, acetaminophen, L-glutathione and / or L-cystine.

[0170] The inner layer may be a uniform layer. Suitably, the inner layer has a thickness of 0.01 pm to 50 pm. More suitably, the inner layer has a thickness of 0.1 pm to 40 pm. Even more suitably, the inner layer has a thickness of 1 pm to 30 pm. Yet even more suitably, theinner layer has a thickness of 1.5 m to 20 pm. Most suitably, the inner layer has a thickness of 2 pm to 10 pm.

[0171] The inner layer may have a water contact angle of 40-140° More suitably, the inner layer has a water contact angle of 50-130° Even more suitably, the inner layer has a water contact angle of 60-120°. Yet even more suitably, the inner layer has a water contact angle of 70-115°. Most suitably, the inner layer has a water contact angle of 80-110°.

[0172] In some embodiments, the microneedle electrode comprises:a semi-permeable outer layer, wherein the semi-permeable outer layer comprises (or is formed from) a first polymeric material, wherein the first polymeric material is a random copolymer comprising the following monomers:wherein a is 9.5 to 10 mol%, b is 78.25 to 79.8 mol%, c is 0.5 to 0.75 mol% and d is 10 to 10.5 mol%, wherein the sum total of a, b, c and d is 100 mol%; anda sensing layer, wherein the sensing layer comprises: i) an enzyme; and ii) a third polymeric material, whereinthe enzyme is selected from the group consisting of glucose oxidase, lactate oxidase, glucose dehydrogenase, p-hydroxybutyrate dehydrogenase and lactate dehydrogenase; and the third polymeric material is a random co-polymer comprising the following monomers:wherein, x1 is 10.2 to 12.3 mol%, y1 is 85 to 86 mol% and z1 is 2.1 to 4.3 mol%, wherein the sum total of x1, y1 and z1 is 100 mol%.

[0173] In some embodiments, the microneedle electrode comprises:a semi-permeable outer layer, wherein the semi-permeable outer layer comprises (or is formed from) a first polymeric material, wherein the first polymeric material comprises:wherein a is 9.5 to 10 mol%, b is 78.25 to 79.8 mol%, c is 0.5 to 0.75 mol% and d is 10 to 10.5 mol%, wherein the sum total of a, b, c and d is 100 mol%; anda sensing layer, wherein the sensing layer comprises: i) an enzyme; and ii) a third polymeric material, whereinthe enzyme is selected from the group consisting of glucose oxidase, lactate oxidase, -hydroxy butyrate dehydrogenase and lactate dehydrogenase; andthe third polymeric material comprises:wherein, x1 is 10.2 to 12.3 mol%, y1 is 85 to 86 mol% and z1 is 2.1 to 4.3 mol%, wherein the sum total of x1, y1 and z1 is 100 mol%.

[0174] In these embodiments, the first polymeric material may be crosslinked with glycerol trigylcidyl ether. In these embodiments, the third polymeric material may be crosslinked with poly(ethylene glycol) diglycidyl ether (PEGDE).Configuration of the microneedle electrode

[0175] As described hereinbefore, at least a portion of the surface of the microneedle electrode is coated with a composite material, wherein the composite material comprises: a semi-permeable outer layer; and a sensing layer positioned underneath the semi-permeable outer layer that is capable of generating an electrochemical response in the presence of the analyte(s).

[0176] While it may be that all of the microneedle electrode is coated with the composite material, it is preferable that one or more portions of the microneedle electrode are coated with the composite material. For example, it may be that less than 100% of the surface area of the microneedle electrode is coated with the composite material. Suitably, less than 75% of the surface area of the microneedle electrode is coated with the composite material. More suitably, less than 50% of the surface area of the microneedle electrode is coated with the composite material. Even more suitably, less than 25% of the surface area of themicroneedle electrode is coated with the composite material.

[0177] In some embodiments, more than one portion of the microneedle electrode is coated with the composite material. Suitably, more than two (e.g., three) portions of the microneedle electrode are coated with the composite material. Most suitably, three portions of the microneedle electrode are coated with the composite material. Portions of the microneedle electrode which are not coated with the composite material, may be coated with the semi-permeable outer layer and optionally the insulation layer (i.e., the sensing layer is absent).

[0178] For illustrative purposes, figure 3 highlights one embodiment of the invention wherein three portions of the microneedle electrode are coated with the composite material. In this embodiment, the remainder of the microneedle electrode (i.e., portions of the microneedle electrode which are not coated with the composite material) are coated with the semi-permeable outer layer and optionally the insulation layer.

[0179] Each microneedle electrode comprises a microneedle body and an insertion tip at a distal end thereof. Suitably, at least a portion of the surface of the microneedle body is coated with a composite material as defined herein. Suitably, at least a portion of the surface of the insertion tip is coated with a composite material as defined herein.

[0180] The form of the insertion tip of each microneedle electrode facilitates improved insertion of the respective microneedle electrode in an application surface of a subject. Figure 4 shows a microneedle electrode of the present invention. The microneedle electrode 108 comprises a substrate 1302 upon which a conductive electrode layer 1304 is provided. An insulation layer 1306 covers a portion of the conductive electrode layer 1304 whilst leaving another portion of the conductive electrode layer 1304 exposed. On the exposed portion of the conductive electrode layer 1304, several separate sensing layers 1308 are provided. Figure 5 shows an alternative electrode of the present invention, wherein the several separate sensing layers are replaced with a single sensing layer 1308.

[0181] The microneedle electrode may be a substantially flat sheet as is evident from the side view of the microneedle electrode in Figure 11. Suitably, the microneedle electrode or each microneedle electrode is substantially flat. Consequently, the microneedle electrode or each microneedle 108 has a first planar surface 714 and a second planar surface 716 opposing the first planar surface 714. The distance between the first planar surface 714 and the second planar surface 716 defines the thickness T1 of the respective microneedle 108. Suitably, the microneedle electrode or each microneedle electrode has an identical thickness. The thickness of the microneedle electrode or each microneedle electrode (i.e., T1) may be 0.025 mm to 0.4 mm. Suitably, the thickness of the microneedle electrode or each microneedle electrode is 0.05 mm to 0.2 mm. More suitably, the thickness of the microneedle electrode or each microneedle electrode is 0.075 mm and 0.1 mm.Wearable device

[0182] In accordance with a second aspect, the present invention provides a wearable device for detecting at least one analyte, wherein the wearable device comprises one or more microneedle electrodes according to the first aspect of the present invention.

[0183] The analyte(s) are as defined anywhere herein. Suitably, the analyte is selected from the group consisting of glucose, ketones (e.g., p-hydroxybutyrate), lactate, histamine, urea, alcohols, cholesterol, vitamins, iodine, potassium, sodium, hormones (e.g., testosterone, cortisol, oestrogen and progesterone) and IL-6. Most suitably, the analyte is selected from the group consisting of glucose, ketones and lactate.

[0184] The wearable device may comprise two or more microneedle electrodes, wherein the microneedle electrodes are each independently as defined herein. Suitably, the wearable device comprises three or more microneedle electrodes, wherein the microneedle electrodes are each independently as defined herein. More suitably, the wearable device comprises five or more microneedle electrodes, wherein the microneedle electrodes are each independently as defined herein. Most suitably, the wearable device comprises ten or more microneedle electrodes, wherein the microneedle electrodes are each independently as defined herein.

[0185] When the wearable device comprises two or more microneedle electrodes, each microneedle electrode may be identical or different. Suitably, each microneedle electrode is identical. Each microneedle electrode is independently as defined herein.

[0186] When the wearable device comprises two or more microneedle electrodes, the wearable device may be considered as a microneedle array.

[0187] The wearable device may be applied to an application surface of a subject. Suitably the subject is a human. When the subject is a human, the application surface may be skin on the arm, leg or abdomen.

[0188] The wearable device may comprise adhesive means. The adhesive means can ensure that the wearable device remains adhered (i.e., affixed) to an application surface of a subject. Suitably, when the wearable device is affixed to an application surface of a subject, the microneedle electrode(s) are maintained in a configuration in which the microneedle electrode(s) penetrate and are continually urged through the application surface of a subject. Maintaining this configuration means that the microneedle electrode(s) are in continuous contact with the analyte(s) (e.g., the interstitial fluid in the dermis of the skin). This ensures that the sensing accuracy of the microneedle electrode remains high.

[0189] The wearable device may further comprise one or more microneedle electrodes not according to the first aspect of the present invention. The microneedle electrodes not according to the first aspect of the present invention may be considered to be reference microneedle electrodes, counter microneedle electrodes, pseudo-referencemicroneedle electrodes or sentinel microneedle electrodes. The microneedle electrodes not according to the first aspect of the present invention may comprise a semi-permeable outer layer, but do not comprise a sensing layer, as is the case for microneedle electrodes according to the first aspect of the present invention.

[0190] The microneedle electrodes may be arranged into distinct conductive zones, wherein in each conductive zone, microneedle electrodes are electrically connected to one another, and microneedle electrodes of one conductive zone are not directly electrically connected to microneedle electrodes of another conductive zone. Any of the conductive zones may be one of a working microneedle electrode zone, a reference microneedle electrode zone, a counter microneedle electrode zone, a pseudo-reference microneedle electrode zone, or a sentinel microneedle electrode zone.

[0191] Suitably, the wearable device comprises at least one working microneedle electrode zone and one of a pseudo-reference microneedle electrode zone or a reference microneedle electrode zone. In embodiments in which a reference microneedle electrode zone is provided, at least one counter microneedle electrode zone may also be provided.

[0192] Suitably, more than one working microneedle electrode zone is provided, wherein the microneedle electrodes of each working microneedle electrode zone are arranged to detect the presence of the analyte(s), and wherein the working microneedle electrode zones are arranged to detect a common analyte or distinct analytes. Suitably, at least one of the conductive zones is a reference microneedle electrode zone, and other present conductive zones are arranged equidistant from the reference microneedle electrode zone relative to one another.

[0193] Two conductive zones may be provided. The two conductive zones may, for example, comprise one working microneedle electrode zone and one pseudo-reference microneedle electrode zone.

[0194] Three conductive zones may be provided. The three conductive zones may, for example, comprise one working microneedle electrode zone, one reference microneedle electrode zone, and one counter microneedle electrode zone. Alternatively, the three conductive zones may, for example, comprise two working microneedle electrode zones and one pseudo-reference microneedle electrode zone.

[0195] Four conductive zones may be provided. The four conductive zones may, for example, comprise one working microneedle electrode zone for a first analyte (e.g., glucose), one working microneedle electrode zone for a second analyte that is different to the first analyte (e.g., lactate), one reference microneedle electrode zone, and one counter microneedle electrode zone. Alternatively, the four conductive zones may, for example, comprise two working microneedle electrode zones for a common analyte (e.g., glucose), one reference microneedle electrode zone, and one counter microneedle electrode zone.Alternatively, the four conductive zones may, for example, comprise three working microneedle electrode zones (e.g., for a common analyte, such as glucose) and one pseudoreference microneedle electrode zone. Alternatively, the four conductive zones may, for example, comprise one working microneedle electrode zone for an analyte (e.g., glucose), one sentinel microneedle electrode zone, one reference microneedle electrode zone, and one counter microneedle electrode zone.

[0196] Five or more conductive zones may be provided. The five or more conductive microneedle electrode zones may, for example, include at least one working microneedle electrode zone, and at least one reference microneedle electrode zone or pseudo-reference microneedle electrode zone.Methods of detecting at least one analyte

[0197] In accordance with a third aspect, the present invention provides a method of detecting at least one analyte, wherein the method comprises a step of contacting a microneedle electrode according to the first aspect of the present invention with at least one analyte.

[0198] In accordance with a fourth aspect, the present invention provides a method of detecting at least one analyte, wherein the method comprises a step of contacting a microneedle electrode of a wearable device according to the second aspect of the present invention with at least one analyte.

[0199] The analyte(s) are as defined anywhere herein. Suitably, the analyte is selected from the group consisting of glucose, ketones (e.g., p-hydroxybutyrate), lactate, histamine, urea, alcohols, cholesterol, vitamins, iodine, potassium, sodium, hormones (e.g., testosterone, cortisol, oestrogen and progesterone) and IL-6. Most suitably, the analyte is selected from the group consisting of glucose, ketones and lactate.

[0200] The semi permeable outer layer may be permeable to the analyte(s) and optionally oxygen. Accordingly, the methods may comprise a second step wherein the analyte(s) and optionally oxygen permeate the semi-permeable outer layer.

[0201] Once permeated through the semi-permeable outer layer, the analyte(s) and optionally oxygen reach the sensing layer. The sensing layer is positioned underneath the semi-permeable outer layer and is capable of generating an electrochemical response in the presence of the analyte(s). Accordingly, the methods may comprise a third step wherein the sensing layer generates an electrochemical response in the presence of the analyte(s) permeated through the semi-permeable outer layer.

[0202] In an embodiment, the methods comprise:contacting a microneedle electrode according to the first aspect of the present invention I a microneedle electrode of a wearable device according to the second aspect of the presentinvention with at least one analyte;permeating the analyte(s) and optionally oxygen through the semi-permeable outer layer; and generating an electrochemical response in the presence of the analyte(s) permeated through the semi-permeable outer layer.

[0203] The step of generating an electrochemical response in the presence of the analyte(s) permeated through the semi-permeable outer layer may have more than one mechanism, and this can depend on the nature of the sensing layer.

[0204] Suitably, the sensing layer comprises: i) an enzyme or an ionophore; and ii) a third polymeric material. In embodiments wherein the sensing layer comprises an enzyme, the enzyme can react with the analyte(s) and oxygen permeated through the semi-permeable outer layer, thereby generating an oxidised form of the analyte(s) and a reduced species or an electron.

[0205] Depending on the nature of the sensing layer, which may comprise an enzyme and a third polymeric material, the step of generating an electrochemical response in the presence of the analyte(s) permeated through the semi-permeable outer layer may proceed via a first mechanism or a second mechanism, as described below:First mechanismthe sensing layer comprises an enzyme and a third polymeric material;the enzyme reacts with the analyte(s) and oxygen permeated through the semi-permeable outer layer thereby generating an oxidised form of the analyte(s) and a reduced species; the reduced species is transferred to the surface of the microneedle electrode (e.g., the conductive electrode layer), thereby generating a current signal;the microneedle electrode measures the current signal generated, which is proportional to the concentration of the analyte(s) before reaction with the enzyme.Second mechanismthe sensing layer comprises an enzyme and a third polymeric material, wherein the third polymeric material comprises a monomer comprising a redox mediator;the enzyme reacts with the analyte(s) permeated through the semi-permeable outer layer, thereby generating an oxidised form of the analyte(s) and an electron;the electron is transferred to the surface of the microneedle electrode (e.g., the conductive electrode layer) via the monomer comprising a redox mediator, thereby generating a current signal;the microneedle electrode measures the current signal generated, which is proportional to the concentration of the analyte(s) before reaction with the enzyme.

[0206] Accordingly, the present invention provides a method of detecting at least one analyte, wherein the method comprises:contacting a microneedle electrode according to the first aspect of the present invention or amicroneedle electrode of a wearable device according to the second aspect of the present invention with at least one analyte;permeating the analyte(s) and oxygen through the semi-permeable outer layer; and generating an electrochemical response in the presence of the analyte(s) permeated through the semi-permeable outer layer by:reacting an enzyme in the sensing layer with the analyte(s) and oxygen permeated through the semi-permeable outer layer thereby generating an oxidised form of the analyte(s) and a reduced species;transferring the reduced species to the surface of the microneedle electrode (e.g., the conductive electrode layer), thereby generating a current signal;measuring the current signal generated, which is proportional to the concentration of the analyte(s) before reaction with the enzyme.

[0207] Suitably, the microneedle electrode measures the current signal generated.

[0208] The present invention also provides a method of detecting at least one analyte, wherein the method comprises:contacting a microneedle electrode according to the first aspect of the present invention or a microneedle electrode of a wearable device according to the second aspect of the present invention with at least one analyte;permeating the analyte(s) and optionally oxygen through the semi-permeable outer layer; and generating an electrochemical response in the presence of the analyte(s) permeated through the semi-permeable outer layer by:reacting an enzyme in the sensing layer with the analyte(s) permeated through the semi-permeable outer layer thereby generating an oxidised form of the analyte(s) and an electron; transferring the electron to the surface of the microneedle electrode (e.g., the conductive electrode layer) via a monomer comprising a redox mediator in the sensing layer, thereby generating a current signal;measuring the current signal generated, which is proportional to the concentration of the analyte(s) before reaction with the enzyme.

[0209] Suitably, the microneedle electrode measures the current signal generated.

[0210] Depending on the nature of the sensing layer, which may comprise an ionophore and a third polymeric material, the step of generating an electrochemical response in the presence of the analyte(s) permeated through the semi-permeable outer layer may proceed via a third mechanism, as described below:Third mechanismthe sensing layer comprises an ionophore and a third polymeric material;the ionophore binds to the analyte(s) permeated through the semi-permeable outer layer, thereby generating a current signal;the microneedle electrode measures the current signal generated, which is proportional to the concentration of the analyte(s) before reaction with the enzyme.

[0211] In the third mechanism, the ionophore binds to the analyte(s) which shift the open circuit potential according to the Nerst equation. The open circuit potentiometry (i.e., voltage vs time) of the microneedle electrode of the present invention versus a reference electrode is measured overtime. By changing the analyte(s) concentration by known amounts, the measured shift in potential can be calibrated to the analyte(s) concentration.

[0212] The present invention therefore also provides a method of detecting at least one analyte, wherein the method comprises:contacting a microneedle electrode according to the first aspect of the present invention or a microneedle electrode of a wearable device according to the second aspect of the present invention with at least one analyte;permeating the analyte(s) and optionally oxygen through the semi-permeable outer layer; and generating an electrochemical response in the presence of the analyte(s) permeated through the semi-permeable outer layer by:reacting an ionophore in the sensing layer with the analyte(s) permeated through the semi-permeable outer layer thereby generating a current signal;measuring the current signal generated, which is proportional to the concentration of the analyte(s) before reaction with the ionophore.

[0213] Suitably, the microneedle electrode measures the current signal generated.Method of preparing a microneedle electrode

[0214] The microneedle electrode of the first aspect of the present invention may be prepared by any suitable method. For example, preparing the microneedle electrode may involve:Step 1 - Preparation of the sensing layer; andStep 2 - Preparation of the semi-permeable outer layer.Step 1 - Preparation of the sensing layer

[0215] The sensing layer may be prepared as follows:Preparation of a sensing layer solution;Applying the sensing layer solution to the microneedle electrode; andCuring the sensing layer solution applied to the microneedle electrode.

[0216] The sensing layer solution may comprise a third polymeric material, an optional crosslinker and an enzyme, as described herein, in a buffer. Example buffers include 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid (HEPES), Phosphate-buffered saline (PBS) and 2-(N-morpholino)ethanesulfonic acid (MES). The buffer may have a concentration of 1mM to 100 mM. Suitably, the buffer has a concentration of 5 mM to 20 mM. Most suitably, the buffer has a concentration of 10 mM. Suitably, the pH of the sensing layer solution is 5 to 9. Most suitably the pH of the sensing layer solution is 8.

[0217] The sensing layer solution may comprise 0.1 %w / v to 5 %w / v of the third polymeric material, 0 %w / v to 2 %w / v of the crosslinker and 0.5 %w / v to 10 %w / v of the enzyme. Suitably, the sensing layer solution comprises 0.5 %w / v to 2.5 %w / v of the third polymeric material, 0.25 %w / v to 1.5 %w / v of the crosslinker and 0.5 %w / v to 5 %w / v of the enzyme. More suitably, the sensing layer solution comprises 0.75 %w / v to 1.5 %w / v of the third polymeric material, 0.35 %w / v to 1.1 %w / v of the crosslinker and 0.75 %w / v to 2.5 %w / v of the enzyme.

[0218] The sensing layer solution may be applied to the microneedle electrode by immersing the microneedle electrode in the sensing layer solution (i.e., dip coating), drop casting, spray casting, aerosol jet printing, electrostatic spray coating, electro-depositing, electro-polymerising, chemical vapour deposition or low-volume dispensing of the sensing layer solution onto the microneedle electrode.

[0219] Curing the sensing layer solution may be conducted for 1 hour to 4 days. Suitably, curing the sensing layer solution is conducted for 2 hours to 3 days. More suitably, curing the sensing layer solution is conducted for 12 hours to 36 hours. Most suitably, curing the sensing solution is conducted for 24 hours.

[0220] Curing the sensing layer solution may be conducted at a temperature of 1 °C to 200 °C. More suitably, curing the sensing layer solution is conducted at a temperature of 10 °C to 150 °C. Even more suitably, curing the sensing layer solution is conducted at a temperature of 15 °C to 100 °C. Yet even more suitably, curing the sensing layer solution is conducted at a temperature of 20 °C to 50 °C. Most suitably, curing the sensing layer solution is conducted at a temperature of 25 °C.

[0221] Curing the sensing layer solution may be conducted at a relative humidity of 30% RH to 70% RH. Suitably, curing the sensing layer solution is conducted at a relative humidity of 40% RH to 60% RH. Most suitably, curing the sensing layer solution is conducted at a relative humidity of 55% RH.Step 2 Preparation of the semi-permeable outer layer

[0222] The semi-permeable outer layer may be prepared as follows:Preparing a semi-permeable outer layer solution;Applying the semi-permeable outer layer solution to the sensing layer and / or the microneedle electrode;Curing the semi-permeable outer layer solution applied to the sensing layer and / or the microneedle electrode.

[0223] The semi-permeable outer layer solution may comprise a first polymeric material and an optional crosslinker in a solvent. Example solvents include alcohols, such as ethanol and methanol, as well as dimethylformamide (DMF), dimethylsulfoxide (DMSO) and tetra hydrofuran (THF). The semi-permeable outer layer solution may comprise a buffer, such as HEPES, PBS or MES. Suitably, the pH of the semi-permeable outer layer solution is 6 to 10.

[0224] The semi-permeable outer layer solution may comprise 2 %w / v to 25 %w / v of the first polymeric material and 0.05 %w / v to 4 %w / v of the crosslinker. Suitably, the semi-permeable outer layer solution comprises 5 %w / v to 13 %w / v of the first polymeric material and 0.3 %w / v to 2 %w / v of the crosslinker. Most suitably, the semi-permeable outer layer solution comprises 8.5 %w / v to 11.5 %w / v of the first polymeric material and 0.5 %w / v to 0.8 %w / v of the crosslinker.

[0225] The semi-permeable outer layer solution may be applied to the sensing layer and / or the microneedle electrode by immersing the microneedle electrode in the semi-permeable outer layer solution (i.e., dip coating), drop casting, spray casting, aerosol jet printing, electrostatic spray coating, electro-depositing, electro-polymerising, chemical vapour deposition or low-volume dispensing of the semi-permeable outer layer solution onto the sensing layer and / or the microneedle electrode. Suitably, the semi-permeable outer layer solution is applied to the sensing layer and / or the microneedle electrode by dip coating.

[0226] Curing the semi-permeable outer layer solution applied to the sensing layer and / or the microneedle electrode may be conducted in two steps:In a first step, curing may be conducted for 1 hour to 4 days, suitably 2 hours to 3 days, more suitably 12 hours to 36 hours, most suitably 20 hours to 28 hours (e.g., 24 hours). In the first step, curing may be conducted at a temperature of 1 °C to 200 °C, suitably 10 °C to 150 °C, more suitably 15 °C to 100 °C, even more suitably 20 °C to 50 °C, most suitably 20 °C to 30 °C. In the first step, curing may be conducted at a relative humidity of 30% RH to 70% RH, suitably 40% RH to 60% RH, most suitably 50% RH to 60% RH.In a second step, curing may be conducted for 1 hour to 7 days, suitably 4 hours to 5 days, more suitably 1 day to 4 days, most suitably 2 days to 3 days. In the second step, curing may be conducted at a temperature of 1 °C to 200 °C, suitably 10 °C to 150 °C, more suitably 15 °C to 100 °C, even more suitably 30 °C to 60 °C, most suitably 40 °C to 50 °C. In the second step, curing may be conducted at a relative humidity of less than 10% RH.EXAMPLES

[0228] One or more examples of the invention will now be described, for the purpose of illustration only, with reference to the accompanying figures, in which:Fig. 1 shows an exemplary diagram of detecting at least one analyte using a microneedle electrode of the present invention proceeding via a first mechanism;Fig. 2 shows an exemplary diagram of detecting at least one analyte using a microneedle electrode of the present invention proceeding via a second mechanism;Fig. 3 shows an example microneedle electrode of the present invention;Fig. 4 shows an example microneedle electrode of the present invention;Fig. 5 shows an example microneedle electrode of the present invention;Fig. 6 shows the current response of a microneedle electrode of the present invention to step-wise additions of glucose under Test 2 in 0.1 M PBS at 35 °C (described herein). The subsequent concentration of glucose is added when a stable current has been held for at least 600 seconds;Fig. 7 shows calibration curves for 5 microneedle electrodes of the present invention (referred to as sensors 1-5, prepared according to Example 1A / Example 1B), all from the same manufacturing lot, indicating the stable current points relative to varying glucose concentration. The average gradient displays the sensitivity of the microneedle electrode towards changing glucose concentration (nA / mM);Fig. 8 shows the average sensitivity and standard deviation in sensitivity of consecutive manufacturing lots of microneedle electrodes of the present invention;Fig. 9 shows the average sensitivity and standard deviation across manufacturing lots of microneedle electrodes of the present invention. Each lot is labelled with the corresponding semi-permeable outer layer thicknesses (pm) of the microneedle electrode;Fig. 10 shows the average sensitivity of three wearable devices (i.e., microneedle arrays) of the present invention over the course of 14 days. Points are determined by a calibration curve according to Test 2 (described herein). Sensitivity loss is found to be on average 0.67% / day;Fig. 11 shows a side view of an example microneedle electrode of the present invention. Fig. 12 shows the current response of Example 2A to step-wise additions of lactate up to 20 mM;Fig. 13 shows the current response of Example 2B to step-wise additions of lactate up to 20 mM (N = 2 example datasets);Fig. 14 shows the current response of Example 3 to step-wise additions of - hydroxy butyrate up to 8 mM (N = 5 example datasets);Fig. 15 shows the current response of Example 4 (sensor 1) to step-wise additions of glucose;Fig. 16 shows the current response of Example 4 (sensor 2) to step-wise additions of glucose;Fig. 17 shows the current response of Example 4 (sensor 3) to step-wise additions of glucose;Fig. 18 shows the current response of Example 4 (sensor 4) to step-wise additions of glucose;Fig. 19 shows the current response of Example 4 (sensor 5) to step-wise additions of glucose;Fig. 20 shows the current response of Example 4 (sensor 6) to step-wise additions of glucose;Example 1ASensing layer: redox polymer (10 mg / ml), glucose oxidase enzyme (11.5 mg / ml, 278 U / mg) and poly(ethylene glycol) diglycidyl ether (4.6 mg / ml, Mw 500) in HEPES solution (10 mM, pH 8).Semi-permeable outer layer: outer layer polymer (110 mg / ml) and glycerol trigylcidyl ether (7.2 mg / ml) in 4:1 ethanol:HEPES solution (10 mM, pH 8).

[0229] The sensing layer solution was deposited onto a single microneedle electrode (i.e., the working microneedle electrode), and cured at 25 °C and 55% relative humidity (RH) for 24 hours. All of the microneedle electrodes (i.e., the working, reference and counter microneedle electrodes) were then dipped four times into the semi-permeable outer layer solution with a 7 minute wait in between coats. After dipping, the microneedle electrodes were stored at 25 °C and 55% RH for 24 hours. After this initial storage, the microneedle electrodes were cured at 48 °C and <10% RH for 60 hours.Example 1BSensing layer: redox polymer, glucose oxidase enzyme and poly(ethylene glycol) diglycidyl ether in buffered solution (pH 8).Semi-permeable outer layer: outer layer polymer and glycerol trigylcidyl ether in 5:1 ethanol: buffer solution (pH 8).

[0230] The sensing layer solution was deposited onto a single microneedle electrode (i.e., the working microneedle electrode), and cured at room temperature and humidity for about 1 day. All of the microneedle electrodes (i.e., the working, reference and counter microneedle electrodes) were then dipped four times into the semi-permeable outer layer solution with about a 10 minute wait in between coats. After dipping, the microneedle electrodes were stored in controlled temperature and humidity for 24 hours. After this initial storage, the microneedle electrodes were cured at 45 - 55 °C and low humidity for about 3days.

[0231] The sensors shown in figures 6 to 10 all comprise a microneedle electrode as described in Example 1A or Example 1B. In Examples 1A and 1B: the redox polymer is a random co-polymer comprising the following monomers:wherein, x1 is 10.2 to 12.3 mol%, y1 is 85 to 86 mol% and z1 is 2.1 to 4.3 mol%, wherein the sum total of x1, y1 and z1 is 100 mol%; andthe outer layer polymer is a random co-polymer comprising the following monomers:wherein a is 9.5 to 10 mol%, b is 78.25 to 79.8 mol%, c is 0.5 to 0.75 mol% and d is 10 to10.5 mol%, wherein the sum total of a, b, c and d is 100 mol%.The sensors shown in Figure 8 comprise a microneedle electrode as described in Example 1A or Example 1B (i.e., with the redox polymer and outer layer polymer described above) and the following:AnalysisTest 1: Sensitivity testing

[0232] The sensitivity testing involves a 3-part test:I. Open Circuit Potentiometry is employed to find any reference microneedle electrode drift in buffer (e.g., 10 mV) over a specified time interval, such as 1 hour;II. Multi-step amperometry is conducted at a constant glucose concentration, with determined steps (e.g., 10 mV) from the working microneedle electrode OCP up to 0.7 V, to determine the corresponding current increase (e.g., 1nA / 10mV), known as the “drift increase”. The bias voltage to be applied in the following chronoamperometry test is determined from the plateau of the amperometry;I I. Chronoamperometry with increasing injections of glucose under simulated physiological conditions in vitro to establish a linear model that describes the relationship between glucose and current using the collected set of stable current values and their corresponding glucose concentrations.Test 2: Calibration and Interference test

[0233] Chronoamperometry at a bias voltage value between -0.1 to 0.7 V under simulated physiological conditions in vitro. This involves the addition of the target analyte, such as glucose and potential interferents such as ascorbic acid into a stirred or flowing buffer. Linear and operational ranges, sensitivity, response time and %interference are quantified.

[0234] As shown in figure 6, the current increases to a stable value after each addition ofglucose. The proportional current change response after each addition of glucose is used to determine the sensitivity of the microneedle electrode. Figure 7 is the graphical representation of the increasing current response with glucose concentration of 5 individual microneedle electrodes (referred to as sensors 1-5, prepared according to Example 1A / Example 1B).

[0235] The relationship between glucose concentration and microneedle electrode current is described as a first order linear model. Defining glucose(t) as the glucose concentration in the solution, and current(t) as the electrical current measured by the sensor, the relationship is modelled as:current(t) = a ■ glucose(t) + bwhere a and b are defined as microneedle electrode sensitivity and baseline.

[0236] The microneedle electrode current response should be as linear as possible to ensure consistent glucose readings. From the estimated model, the linearity of the response can be characterized using the R-squared value of the estimation which is obtained as:where current indicates the average value of current.

[0237] Characterizing the microneedle electrode's response time to glucose is an important aspect of its performance. The time constant of the step response of an equivalent low-pass filter is determined. The semi-permeable outer layer, which acts as a physical membrane, slows the diffusion of glucose into the sensing layer, introducing a phase shift in the current signal. As such, the time it takes to reach 95% of the stable value (T95) is measured, from which the time constant as T95 divided by 3 is calculated.

[0238] The microneedle electrodes are found to have a linear response to glucose from 0 to 25 mM, R2= 0.99. The average sensitivity of this lot (i.e., the microneedle electrodes) is determined to 0.56 nA / mM, COV = 1.6%. This test demonstrates the ability of the microneedle electrodes to respond in a reliable and predictable manner over the clinically relevant range of glucose concentrations.Test 3: Stability test

[0239] Chronoamperometry at a bias voltage value between -0.1 to 0.7 V was held for fourteen days at 35 °C. The solution contains a known concentration of glucose in the physiological range (e.g., 5 - 25 mM) and may also contain protein (e.g., bovine serum albumin). The signal drift / decay was also quantified.

[0240] In a particular test, microneedle electrodes were polarized at an appropriate bias voltage in 25 mM glucose in PBS (pH 7.4) at 35 °C for 14 days. The sensitivity of the microneedle electrodes was periodically measured according to Test 2 in order to quantify theloss in sensitivity with time. As displayed in Figure 10, the average sensitivity loss over the course of 14 days was found to be 0.67% / day.Example 2

[0241] The sensors used in Example 2 (i.e., Examples 2A and 2B) are for detecting lactate. The sensors are manufactured and tested using similar procedures to Example 1A and Example 1B, but are modified for lactate detection.Example 2ASensing layer: redox polymer (10 mg / ml), lactate oxidase enzyme (20 mg / ml), ovalbumin (20 mg / ml) and poly(ethylene glycol) diglycidyl ether (4.6 mg / ml, Mw 400) in MES solution (10 mM, pH 5.5).Semi-permeable outer layer: outer layer polymer and glycerol trigylcidyl ether in ethanol: buffer solution (pH 8).Example 2BSensing layer: redox polymer (10 mg / ml), lactate dehydrogenase enzyme (20 mg / ml), and poly(ethylene glycol) diglycidyl ether (4.6 mg / ml, Mw 400) in MES solution (10 mM, pH 5.5). Semi-permeable outer layer: outer layer polymer and glycerol trigylcidyl ether in ethanol: buffer solution (pH 8).Example 3

[0242] The sensors used in Example 3 are for detecting ketones (e.g., p-hydroxy butyrate). The sensors are manufactured and tested using similar procedures to Example 1A and Example 1B, but are modified for ketone detection.Sensing layer: redox polymer, p-hydroxybutyrate dehydrogenase enzyme, diaphorase enzyme, Nicotinamide adenine dinucleotide coenzyme, human serum albumin and poly(ethylene glycol) diglycidylether (Mw 400) in MES buffer solution (10 mM, pH 5.5).Semi-permeable outer layer: outer layer polymer and glycerol trigylcidyl ether in ethanol: buffer solution (pH 8).Example 4

[0243] The sensors used in Example 4 are for detecting glucose. The sensors are manufactured and tested using identical procedures to Example 1A and Example 1B.

[0244] The data provided in figure 12 is generated using a microneedle electrode as described in Example 2A. The data provided in figure 13 is generated using a microneedle electrode as described in Example 2B. The data provided in figure 14 is generated using a microneedle electrode as described in Example 3. The data provided in figures 15 to 20 is generated using a microneedle electrode as described in Example 4. In Examples 2A, 2B, 3 and 4: the redox polymer is a random co-polymer comprising the following monomers:wherein, x1 is 10.2 to 12.3 mol%, y1 is 85 to 86 mol% and z1 is 2.1 to 4.3 mol%, wherein the sum total of x1, y1 and z1 is 100 mol%; andthe outer layer polymer is a random co-polymer comprising the following monomers:wherein a is 9.5 to 10 mol%, b is 78.25 to 79.8 mol%, c is 0.5 to 0.75 mol% and d is 10 to 10.5 mol%, wherein the sum total of a, b, c and d is 100 mol%.

[0245] While specific embodiments of the invention have been described herein for the purpose of reference and illustration, various modifications will be apparent to a person skilled in the art without departing from the scope of the invention as defined by the appended claims.

Claims

CLAIMS1. A microneedle electrode for detecting at least one analyte, wherein at least a portion of the surface of the microneedle electrode is coated with a composite material, wherein the composite material comprises:a semi-permeable outer layer; anda sensing layer positioned underneath the semi-permeable outer layer that is capable of generating an electrochemical response in the presence of the analyte(s).

2. The microneedle electrode of claim 1, wherein the microneedle electrode comprises a substrate, an optional non-conductive passivation layer, and a conductive electrode layer.

3. The microneedle electrode of claim 2, wherein the substrate comprises (or is formed from) metals (e.g., titanium, tungsten, silver, gold, copper, aluminium, steel (e.g., stainless steel), iron, platinum, tantalum, Nitinol, Elgiloy or alloys thereof), carbon (e.g., graphite, graphene, carbon nanotubes, pyrolytic carbon or carbon fibre), plastics, liquid crystal polymers, silicon or composites (e.g. glass-filled, mineral-filled, or carbon-filled composites).

4. The microneedle electrode of claim 2 or claim 3, wherein the non-conductive passivation layer comprises (or is formed from) insulating polymers (e.g., parylene, polyimide, polyvinylchloride, benzocylcobutene or acrylate) or metal oxides (e.g., titanium dioxide, silicon dioxide or aluminium oxide).

5. The microneedle electrode of any one of claims 2 to 4, wherein the conductive electrode layer comprises (or is formed from) carbon (e.g., graphite, graphene, carbon nanotubes, carbon ink, pyrolytic carbon or carbon fibre), platinum, gold, or rhodium, palladium, silver, silver-silver chloride, steel (e.g. stainless steel), copper, nickel, zinc, or aluminium, tantalum or chromium.

6. The microneedle electrode of any one of the preceding claims, wherein the analyte is present in blood, interstitial fluid, cerebral spinal fluid, lymph fluid, urine, sweat and / or saliva.

7. The microneedle electrode of any one of the preceding claims, wherein the analyte is selected from the group consisting of glucose, ketones (e.g., p-hydroxybutyrate), lactate, histamine, urea, creatinine, alcohols, cholesterol, vitamins, hormones (e.g., testosterone, cortisol, serotonin, oestrogen and progesterone), pH (e.g., hydrogen ion), iodine, potassium, sodium, magnesium, calcium, zinc, copper, iron, chloride, phosphate, ammonium, lithium, bicarbonate, thrombin, IL-6 and therapeutic drugs (e.g. aminoglycosides, doxorubicin,tetracyclines).

8. The microneedle electrode of any one of the preceding claims, wherein the analyte is selected from the group consisting of glucose, ketones and lactate.

9. The microneedle electrode of any one of the preceding claims, wherein the microneedle electrode detects one to ten analytes.

10. The microneedle electrode of any one of the preceding claims, wherein the semi-permeable outer layer is permeable to the analyte and / or oxygen.

11. The microneedle electrode of any one of the preceding claims, wherein the analyte diffusivity of the semi-permeable outer layer is 5x10-11cm2 / s to 1x10-7cm2 / s.

12. The microneedle electrode of any one of the preceding claims, wherein the semi-permeable outer layer has reduced permeability relative to the analyte for acetaminophen (i.e., paracetamol), ascorbic acid, bilirubin, cholesterol, creatine, dopamine, ephedrine, ibuprofen, L-dopa, methyldopa, salicylate, tetracycline, tolazamide, tolbutamide, triglycerides, uric acid, L-glutathione and / or L-cystine.

13. The microneedle electrode of any one of the preceding claims, wherein the semi-permeable outer layer comprises (or is formed from) a first polymeric material.

14. The microneedle electrode of claim 13, wherein the first polymeric material is a copolymer (e.g., alternating, random or block co-polymer) comprising monomers A, B, C and D, wherein A is a monomer comprising a zwitterionic portion; B is a monomer comprising a pyridine portion, C is a monomer comprising a hydrophilic portion; and D is a monomer comprising a phenyl portion.

15. The microneedle electrode of claim 13 or 14, wherein the first polymeric material is a co-polymer (e.g., alternating, random or block co-polymer) comprising monomers A, B, C and D:-Aa_Bb-Cc- Dd_wherein:A is a monomer comprising a zwitterionic portion; B is a monomer comprising a pyridine portion, C is a monomer comprising a hydrophilic portion; and D is a monomer comprising a phenyl portion;a is 0 to 25 mol%;b is 50 to 90 mol%;c is 0.1 to 10 mol%; andd is 0 to 25 mol%,wherein the sum total of a, b, c and d is 100 mol%.

16. The microneedle electrode of any one of claims 13, 14 or 15, wherein the first polymeric material is a random co-polymer comprising the following monomers:wherein a, b, c and d are as defined in claim 15.

17. The microneedle electrode of any one of claims 13 to 16, wherein the first polymeric material comprises:wherein a, b, c and d are as defined in claim 15.

18. The microneedle electrode of any one of claims 13 to 17, wherein the first polymeric material is crosslinked with one or more crosslinkers.

19. The microneedle electrode of claim 18, wherein the crosslinker is selected from the group consisting of isocyanate, carbodiimide, glutaraldehyde, aziridine, silane, or other aldehydes, epoxy, di epoxy, acrylates, free-radical based agents, such as ethylene glycol diglycidyl ether (EGDE), poly(ethylene glycol) diglycidyl ether (PEGDE), or dicumyl peroxide (DCP), glycerol triglycidyl ether, Tris(4-hydroxyphenyl)methane triglycidyl ether, trimethylolpropane triglycidyl ether, trimethylolethane triglycidyl ether, N, N-diglycidyl-4-glycidyloxyaniline, neopentyl glycol diglycidyl ether and 1,4-butanediol diglycidyl ether.

20. The microneedle electrode of any one of the preceding claims, wherein the sensing layer comprises: i) an enzyme or an ionophore; and ii) a third polymeric material.

21. The microneedle electrode of claim 20, wherein the enzyme is selected from the group consisting of glucose oxidase, lactate oxidase, glucose dehydrogenase, diamine oxidase, -hydroxy butyrate dehydrogenase, alcohol oxidase, cholesterol oxidase, choline oxidase, alcohol dehydrogenase, lactate dehydrogenase, glutamate dehydrogenase, glutamate oxidase, glycerophosphate oxidase, glycerol dehydrogenase, leucine dehydrogenase, p-hydroxybenzoate hydroxylase, pyruvate oxidase, sarcosine oxidase, uricase, and xanthine oxidase, preferably glucose oxidase, lactate oxidase, glucose dehydrogenase, diamineoxidase, p-hydroxybutyrate dehydrogenase, alcohol oxidase, cholesterol oxidase, lactate dehydrogenase, pyruvate oxidase, sarcosine oxidase, uricase, and xanthine oxidase.

22. The microneedle electrode of claim 20 or claim 21, wherein the third polymeric material is a co-polymer (e.g., alternating, random or block co-polymer) comprising monomers X, Y and Z, whereinX is a monomer comprising a redox mediator, Y is a monomer comprising a pyridine portion and Z is a monomer comprising a zwitterionic portion;23. The microneedle electrode of any one of claims 20 to 22, wherein the third polymeric material is a co-polymer (e.g., alternating, random or block co-polymer) comprising monomers X, Y and Z:-Xx1 -Yy1-Zz1"wherein:X is a monomer comprising a redox mediator, Y is a monomer comprising a pyridine portion and Z is a monomer comprising a zwitterionic portion;x1 is 5 to 25 mol%;y1 is 70 to 95 mol%; andz1 is 0 to 25 mol%,wherein the sum total of x1, y1 and z1 is 100 mol%.

24. The microneedle electrode of any of the claims 20 to 23, wherein the redox mediator is: (i) an organometallic complex bearing between 1 and 6 ligands;(ii) an organometallic complex bearing at least 1 ligand that forms a covalent linkage the polymer; or(iii) an organometallic complex where the metal is one of iron, cobalt, ruthenium, osmium, manganese, copper, ferrocene and vanadium.

25. The microneedle electrode of any one of claims 20 to 24, wherein the third polymeric material is a random co-polymer comprising the following monomers:wherein x1, y1 and z1 are as defined in claim 23.

26. The microneedle electrode of any one of claims 20 to 25, wherein the third polymeric material comprises:wherein x1, y1 and z1 are as defined in claim 23.

27. The microneedle electrode of any one of claims 20 to 26, wherein the third polymeric material is crosslinked with one or more crosslinkers.28 The microneedle electrode of claim 27, wherein the crosslinker is selected from the group consisting of ethylene glycol diglycidyl ether (EGDE), poly(ethylene glycol) diglycidyl ether (PEGDE), dicumyl peroxide (DCP), glycerol triglycidyl ether, Tris(4-hydroxyphenyl)methane triglycidyl ether, trimethylolpropane triglycidyl ether, trimethylolethane triglycidyl ether, N, N-diglycidyl-4-glycidyloxyaniline, neopentyl glycol diglycidyl ether and 1,4-butanediol diglycidyl ether.

29. The microneedle electrode of any one of the preceding claims, wherein the microneedle electrode is substantially flat.

30. The microneedle electrode of any one of the preceding claims, wherein the microneedle electrode has a thickness of 0.025 mm to 0.4 mm.

31. The microneedle electrode of any one of the preceding claims, whereinthe microneedle electrode comprise a substrate, an optional non-conductive passivation layer, and a conductive electrode layer, whereinthe substrate comprises metals (e.g., titanium, tungsten, silver, gold, copper, aluminium, steel (e.g., stainless steel), iron, platinum, tantalum, Nitinol, Elgiloy or alloys thereof), carbon (e.g., graphite, graphene, carbon nanotubes, pyrolytic carbon or carbon fibre), plastics, liquid crystal polymers, silicon or composites (e.g. glass-filled, mineral-filled, or carbon-filled composites;the non-conductive passivation layer comprises insulating polymers (e.g., parylene (e.g., parylene-C, parylene-F or parylene-N), polyimide, polyvinylchloride, benzocylcobutene or acrylate) or metal oxides (e.g., titanium dioxide, silicon dioxide or aluminium oxide); andthe conductive electrode layer comprises carbon (e.g., graphite, graphene, carbon nanotubes, carbon ink, pyrolytic carbon or carbon fibre), platinum, gold, or rhodium, palladium, silver, silver-silver chloride, steel (e.g. stainless steel), copper, nickel, zinc, or aluminium, tantalum or chromium;the semi-permeable outer layer comprises a first polymeric material, whereinthe first polymeric material is a co-polymer (e.g., alternating, random or block copolymer) comprising monomers A, B, C and D, wherein A, B, C and D are each independently selected from a monomer comprising a zwitterionic portion, a monomer comprising a pyridine portion, a monomer comprising a phenyl portion and a monomer comprising a hydrophilic portion;the sensing layer comprises: i) an enzyme or an ionophore; and ii) a third polymeric material,whereinthe third polymeric material is a co-polymer (e.g., alternating, random or block copolymer) comprising monomers X, Y and Z, wherein X, Y and Z are each independently selected from a monomer comprising a zwitterionic portion, a monomer comprising a pyridine portion and a monomer comprising a redox mediator; wherein the microneedle electrode is substantially flat; andwherein the thickness of the microneedle electrode is 0.025 mm to 0.4 mm.

32. The microneedle electrode of any one of the preceding claims, wherein:the microneedle electrode comprises a substrate, a non-conductive passivation layer and a conductive electrode layer, whereinthe substrate comprises stainless steel, and the conductive electrode layer comprises carbon ink;the semi-permeable outer layer comprises a first polymeric material, whereinthe first polymeric material is a random co-polymer comprising the following monomers:wherein a, b, c and d are as defined in claim 15;the sensing layer comprises: i) an enzyme; and ii) a third polymeric material, whereinthe enzyme is selected from the group consisting of glucose oxidase, lactate oxidase, glucose dehydrogenase, diamine oxidase, p-hydroxybutyrate dehydrogenase and alcohol oxidase;the third polymeric material is a random co-polymer comprising the following monomers:wherein x1, y1 and z1 are as defined herein in claim 23;wherein the microneedle electrode is substantially flat; andwherein the thickness of the microneedle electrode is 0.05 mm to 0.2 mm.

33. A wearable device for detecting at least one analyte, wherein the wearable device comprises one or more microneedle electrodes according to any one of the preceding claims.

34. The wearable device of claim 33, wherein the wearable device comprises two, three or more microneedle electrodes (e.g., the wearable device comprises a microneedle array).

35. A method of detecting at least one analyte, wherein the method comprises a step of contacting a microneedle electrode according to any one of claims 1 to 32, or a microneedle electrode of a wearable device according to claim 33 or claim 34, with at least one analyte.

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