Enzyme-immobilized adhesive layers for analyte sensors.

Abstract

To provide devices for measuring a biological analyte in a host.SOLUTION: The devices comprise a sensor configured to generate a signal associated with a concentration of an analyte, and a sensing membrane located over the sensor. The sensing membrane comprises an enzyme layer, where the enzyme layer comprises an enzyme and a polymer comprising polyurethane and / or polyurea segments and one or more zwitterionic repeating units.SELECTED DRAWING: None

Description

[Technical Field]

[0001] INCORPORATION BY REFERENCE OF RELATED APPLICATIONS Any and all priority claims identified in the Application Data Sheet, or any corrections thereto, are incorporated herein by reference under 37 CFR 1.57. This application claims the benefit of U.S. Provisional Application No. 62 / 273,155, filed December 30, 2015, U.S. Provisional Application No. 62 / 273,142, filed December 30, 2015, and U.S. Provisional Application No. 62 / 273,219, filed December 30, 2015. Each of the foregoing applications is incorporated herein by reference in its entirety, and each is hereby expressly made a part of this specification.

[0002] The subject matter disclosed herein relates to devices for measuring biological analytes in a host, and components of such devices. [Background technology]

[0003] Electrochemical sensors are useful for determining the presence or concentration of biological analytes, such as blood glucose. Such sensors are effective, for example, in monitoring glucose in diabetic patients and lactate during critical care events. A variety of intravascular, transcutaneous, and implantable sensors have been developed to continuously detect and quantify blood analytes, such as blood glucose levels.

[0004] Such analyte sensors have a membrane layer or domain containing an enzyme responsible for the conversion of the analyte to a drug that can be registered as a measurable signal. For example, a glucose sensor contains an enzyme that converts glucose to hydroperoxide, which is further converted into a sensor signal. Therefore, the performance of an enzymatic glucose sensor, like other sensors that rely on enzymatic conversion, can be affected by the amount of active enzyme incorporated into the sensor's membrane layer.

[0005] It is often difficult to incorporate and maintain sufficient active enzymes in membranes to efficiently catalyze analyte reactions (e.g., glucose vs. hydrogen peroxide). Enzymes can leach from membranes in their hydrated state. Leached enzymes can also result in severe foreign body responses (FBRs). These events alter sensor sensitivity, degrade the resistive layer, and ultimately reduce sensor accuracy and lifetime. Furthermore, enzyme degradation can occur through many different mechanisms, leading to irreversible or reversible enzyme inactivation. Enzymes can be sensitive to environmental conditions, including temperature and pH changes, as well as exposure to reaction chemistries, including crosslinkers often used for enzyme immobilization, such as glutaraldehyde and carbodiimides, as well as byproducts from redox reactions, such as hydrogen peroxide and gluconic acid, and endogenous byproducts. Enzyme degradation severely limits the functional lifetime of analyte sensors in vivo, resulting in a gradual decrease in sensor sensitivity and premature sensor end-of-life.

[0006] Incorporation and immobilization of enzymes into various carriers or binders, including polymers, sol-gels, particles, and mixtures thereof, to create enzyme layers have been attempted to prevent enzyme leaching in analyte sensors. However, these layers suffer from the problem of swelling and decomposing in aqueous environments, resulting in poor adhesion to adjacent layers in the membrane system. As a result, these by-products can also leach from the membrane and contribute to FBR, affecting the sensitivity and accuracy of the sensor. Poor adhesion can further result in reduced mechanical stability and delamination of the membrane layer in vivo.

[0007] Therefore, modified enzyme layers are desired in which the enzyme is immobilized within the membrane through strong molecular-level interactions between the enzyme and the base polymer material. Such layers can reduce (or prevent) enzyme leaching, which reduces FBR and improves sensor lifetime, sensitivity, and accuracy. There is also a need for enzyme layers that are modified with physiochemical and catalytic performance stability in aqueous environments and have good adhesion to other layers in the sensor's membrane system. The compositions, methods, and devices disclosed herein address these and other needs. [Prior art documents] [Patent documents]

[0008] [Patent Document 1] Patent Publication No. US-2011-0027127-A1 Summary of the Invention [Means for solving the problem]

[0009] In accordance with the objectives of the disclosed materials and methods, as embodied and broadly described herein, the disclosed subject matter relates in one aspect to compounds, compositions, and methods of making and using the compounds and compositions, as well as devices containing the compounds and compositions. In a first aspect, a device for determining an analyte concentration (e.g., glucose) is provided, comprising a sensor configured to generate a signal associated with the analyte concentration and a sensing membrane positioned over the sensor. The sensing membrane includes an enzyme layer, the enzyme layer comprising an enzyme and a polymer including polyurethane and / or polyurea segments and one or more zwitterionic repeat units. The enzyme layer protects the enzyme and prevents it from leaching from the sensing membrane into a host without adversely affecting the enzyme's activity. The enzyme layer can be 0.01 μm to about 250 μm thick.

[0010] In a second aspect, a device for determining an analyte concentration (e.g., glucose) is provided, comprising a sensor configured to generate a signal associated with the analyte concentration and a sensing membrane positioned over the sensor. The sensing membrane includes an enzyme layer, the enzyme layer comprising an enzyme and a polymer including polyurethane and / or polyurea segments and one or more zwitterionic repeat units. The enzyme layer protects the enzyme and prevents it from being inactivated by dynamic changes in its environment caused by endogenous and exogenous compounds and other stressors, including temperature and pH. The enzyme layer can be 0.01 μm to about 250 μm thick. In further embodiments of the disclosed device, the enzyme can be selected from the group consisting of glucose oxidase, glucose dehydrogenase, galactose oxidase, cholesterol oxidase, amino acid oxidase, alcohol oxidase, lactate oxidase, and uricase. In certain embodiments, the enzyme is glucose oxidase.

[0011] In embodiments of the device of this aspect, one or more of the zwitterionic repeat units comprises a betaine compound or a derivative thereof. In embodiments of the device of this aspect, one or more of the zwitterionic repeat units comprises a betaine compound or a precursor thereof.

[0012] In embodiments of devices of this aspect, one or more zwitterionic repeat units comprise at least one moiety selected from the group consisting of carboxylbetaine, sulfobetaine, phosphobetaine, and derivatives thereof.

[0013] In an embodiment of the device of this aspect, one or more of the zwitterionic repeat units is derived from a monomer selected from the group consisting of: [ka] wherein Z is a branched or straight chain alkyl, heteroalkyl, cycloalkyl, cycloheteroalkyl, aryl, or heteroaryl; R 1is H, alkyl, heteroalkyl, cycloalkyl, heterocycloalkyl, aryl, or heteroaryl, and R 2 , R 3 , and R 4 is independently selected from alkyl, heteroalkyl, cycloalkyl, heterocycloalkyl, aryl, or heteroaryl; R 1 , R 2 , R 3 , R 4 and one or more of Z is substituted with a polymerizable group.

[0014] In an embodiment of the device of this aspect, one or more of the zwitterionic repeat units is derived from a monomer selected from the group consisting of: [ka] wherein Z is a branched or straight chain alkyl, heteroalkyl, cycloalkyl, cycloheteroalkyl, aryl, or heteroaryl; R 1 is H, alkyl, heteroalkyl, cycloalkyl, heterocycloalkyl, aryl, or heteroaryl, and R 2 and R 3 is independently selected from alkyl, heteroalkyl, cycloalkyl, heterocycloalkyl, aryl, or heteroaryl; R 1 , R 2 , R 3 and one or more of Z is substituted with a polymerizable group.

[0015] In an embodiment of the device of this aspect, one or more of the zwitterionic repeat units is derived from a monomer selected from the group consisting of: [ka] wherein Z is a branched or straight chain alkyl, heteroalkyl, cycloalkyl, cycloheteroalkyl, aryl, or heteroaryl; R 1is H, alkyl, heteroalkyl, cycloalkyl, heterocycloalkyl, aryl, or heteroaryl, and R 2 and R 3 is independently selected from alkyl, heteroalkyl, cycloalkyl, heterocycloalkyl, aryl, or heteroaryl; R 1 , R 2 , R 3 and one or more of Z is substituted with a polymerizable group.

[0016] In embodiments of the device of this aspect, the polymerizable group is selected from an alkene, alkyne, epoxide, lactone, amine, hydroxyl, isocyanate, carboxylic acid, anhydride, silane, halide, aldehyde, and carbodiimide.

[0017] In embodiments of devices according to this aspect, the one or more zwitterionic repeat units are at least about 1 wt %, based on the total weight of the polymer.

[0018] In embodiments of devices according to this aspect, the polyurethane and / or polyurea segments are from about 15% to about 75% by weight based on the total weight of the polymer.

[0019] In an embodiment of the device of this aspect, the polymer in the enzyme layer further comprises at least one segment selected from the group consisting of epoxides, polyolefins, polysiloxanes, polyamides, polystyrenes, polyacrylates, polyethers, polyesters, and polycarbonates.

[0020] In embodiments of the device of this aspect, the polymer in the enzyme layer further comprises polyethylene oxide segments, in some embodiments from about 5% to about 60% by weight based on the total weight of the enzyme layer polymer.

[0021] In an embodiment of the device of this aspect, the polymer in the enzyme layer has a molecular weight of about 10 kDa to about 500,000 kDa, a polydispersity index of about 1.4 to about 3.5, and / or a contact angle of about 10° to about 90°.

[0022] In a second aspect, a device is provided, wherein the enzyme layer further comprises a base polymer and an enzyme stabilizing and / or immobilizing polymer, wherein the enzyme stabilizing and / or immobilizing polymer comprises a polymer chain having both hydrophilic and hydrophobic regions and one or more zwitterionic repeat units, and the base polymer is selected from silicones, epoxides, polyolefins, polystyrenes, polyoxymethylenes, polysiloxanes, polyethers, polyacrylic acids, polymethacrylic acids, polyesters, polycarbonates, polyamides, poly(etherketones), poly(etherimides), polyurethanes, and polyurethaneureas.

[0023] In a third aspect, a device is provided, wherein the enzyme layer further comprises an enzyme stabilizing reagent. In certain examples, the enzyme stabilizing reagent may be selected from the group consisting of one or more zwitterions selected from the group consisting of cocamidopropyl betaine, oleamidopropyl betaine, octyl sulfobetaine, capryl sulfobetaine, lauryl sulfobetaine, myristyl sulfobetaine, palmityl sulfobetaine, stearyl sulfobetaine, betaine (trimethylglycine), octyl betaine, phosphatidylcholine, glycine betaine, poly(carboxybetaine), poly(sulfobetaine), and derivatives thereof.

[0024] In all of the devices disclosed herein, they can be configured for continuous measurement of analyte concentration.

[0025] Additional advantages will be set forth in part in the description that follows, and in part will be obvious from the description, or may be learned by practice of the following aspects. The advantages will be realized and attained by means of the elements and combinations particularly pointed out in the appended claims. It is to be understood that the foregoing general description and the following detailed description are exemplary and explanatory only and are not restrictive.

[0026] The accompanying drawings, which are incorporated in and constitute a part of this specification, illustrate several aspects described below. [Brief explanation of the drawings]

[0027] [Figure 1] FIG. 1 is a schematic diagram of a continuous analyte sensor system attached to a host and in communication with other devices. [Figure 2] 1A-1C are general cross-sectional views of a sensor showing various embodiments of a membrane system. [Figure 3A] FIG. 1 is a schematic side view illustrating an in vivo portion of a continuous analyte sensor in one embodiment. [Figure 3B] FIG. 1 is a perspective schematic diagram illustrating an in vivo portion of a continuous analyte sensor in one embodiment. [Figure 3C] FIG. 1 is a schematic side view illustrating an in vivo portion of a continuous analyte sensor in one embodiment. [Figure 3D] FIG. 1 is a cross-sectional / side schematic diagram showing the in vivo portion of a continuous analyte sensor in one embodiment. [Figure 4] 1 is a graph showing the % of active enzyme leached over time into water from a 200 μm thick film of a control polymer blend (P3) with a hydrophilic polymer additive that does not contain betaine, or from a polymer blend with a hydrophilic polymer additive that contains betaine in the polymer backbone disclosed herein. [Figure 5] 10 is a graph comparing various sensor metrics for a glucose sensor (P3) constructed with an enzyme layer formed with the same polymer binder as used in (P3) but with 30 wt % betaine-containing polymer as the enzyme immobilization polymer additive, and a glucose sensor (P3) constructed with an enzyme layer without the 30 wt % betaine-containing polymer. [Figure 6] 1 is a graph showing the normalized elution of total protein enzyme over time in water from 200 μm thick films of a control polymer without betaine, a control polymer with a small molecule betaine added to the formulation, or a polymer disclosed herein with betaine in the polymer backbone. [Figure 7]1 is a graph showing water absorption over time for enzyme layers prepared from WB-7 and a control polymer (P3) without betaine. [Figure 8] 1 is a graph showing the sensitivity of sensors with and without a betaine-containing polymer in the enzyme layer. The enzyme-coated sensors were treated at 70°C and 95% humidity. After this accelerated aging treatment, a resistive layer was added and the sensitivity was measured. The data show a certain decrease in sensitivity for the sensors without the betaine-containing polymer, which is the result of enzyme inactivation due to heat stress and / or high humidity. [Figure 9] 1 is a graph showing the accuracy of sensors with and without a betaine-containing polymer in the enzyme layer. The sensors were coated with an enzyme layer and treated at 70°C and 95% humidity. After this accelerated aging treatment, a resistive layer was added and sensor performance was measured in the form of sensor accuracy, expressed as the mean absolute relative difference (MARD), which is calculated from the average of the absolute relative differences between the calculated and measured values ​​from a least-squares linear fit. The average of |(Vcalculated - Vobserved) / Vobserved| for each step of glucose concentration. [Figure 10] 1 is a schematic illustrating one particular embodiment of an enzyme layer polymer. [Figure 11A] FIG. 1 is a schematic diagram of a portion of one embodiment of an interference domain comprising multiple polycation and polyanion layers. [Figure 11B] 11A illustrates one embodiment of a layer-by-layer method using layer-by-layer adsorption of polycations and polyanions to create the structure shown in FIG. 11A. [Figure 12] 1 is a graph showing results from an adhesive pull-out test. [Figure 13] 10 is a graph showing further results from an adhesive pull-out test. DETAILED DESCRIPTION OF THE INVENTION

[0028] The methods, compositions, and devices described herein may be understood more readily by reference to the following detailed description of certain aspects of the disclosed subject matter and the examples and figures included therein.

[0029] Before the methods, compositions, and devices are disclosed and described, it is to be understood that the following aspects are not limited to particular synthetic methods or to particular reagents, and as such may, of course, vary. It is also to be understood that the terminology used herein is for the purpose of describing particular aspects only and is not intended to be limiting.

[0030] Also, throughout this specification, various publications are referenced. The disclosures of these publications in their entireties are hereby incorporated by reference into this application in order to more fully describe the state of the art to which the disclosed matter pertains. The references disclosed are also individually and specifically incorporated by reference herein for the material contained therein that is discussed in the sentence in which the reference is relied upon.

[0031] definition In this specification and in the claims that follow, reference will be made to a number of terms that shall be defined to have the following meanings.

[0032] As used herein, the term "about" is intended to qualify the numerical value it modifies and indicates such value as being variable within a certain range of error. When a specific error, such as a standard deviation for the average value shown in a chart or table of data, is not recited, the term "about" should be understood to mean a range encompassing the recited value, taking into account significant digits, as well as a range encompassed by rounding up or down to that number.

[0033] As used herein, the term "analyte" is a broad term given its ordinary and customary meaning to those skilled in the art (and is not limited to any special or customized meaning) and refers to, but is not limited to, a substance or chemical constituent in a biological fluid (e.g., blood, interstitial fluid, cerebrospinal fluid, lymphatic fluid, urine, sweat, saliva, etc.) that may be analyzed. Analytes may include naturally occurring substances, man-made substances, metabolites, and / or reaction products. In some embodiments, the analyte for measurement by the sensing region, devices, and methods is glucose. However, other analytes are contemplated as well, including acarboxyprothrombin, acylcarnitines, adenine phosphoribosyltransferase, adenosine deaminase, albumin, α-fetoprotein, amino acid profile (arginine (Krebs cycle), 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, cholesterol, cholinesterase, conjugated 1-β-hydroxycholic acid, cortisol, creatine kinase, creatine kinase MM isoenzyme, cyclosporin A, d-penicilla amine, de-ethylchloroquine, dehydroepiandrosterone sulfate, DNA (acetylation polymorphism), alcohol dehydrogenase, α1-antitrypsin, cystic fibrosis, Duchenne / Becker muscular dystrophy, glucose-6-phosphate dehydrogenase, hemoglobin A, hemoglobin S, hemoglobin C, hemoglobin D, hemoglobin E, hemoglobin F, D Punjab, β-thalassemia, hepatitis B virus , HCMV, HIV-1, HTLV-1, Leber's hereditary optic neuropathy, MCAD, RNA, PKU, Plasmodium vivax, sex differentiation, 21-deoxycortisol), desbutylhalofantrine, dihydropteridine reductase, diphtheria / tetanus antitoxin, erythrocyte arginase, erythrocyte protoporphyrin, esterase D, fatty acids / acylglycines, free β-human chorionic gonadotropin, free erythrocyte porphyrin,Free thyroxine (FT4), free tri-iodothyronine (FT3), fumarylacetoacetase, galactose / gal-1-phosphate, galactose-1-phosphate uridyltransferase, gentamicin, glucose-6-phosphate dehydrogenase, glutathione, glutathione peroxidase, glycocholate, glycosylated hemoglobin, halofantrine, hemoglobin variants, hexosaminidase A, human erythrocyte carbonic anhydrase I, 17-α-hydroxyprogesterone, hypoxanthine phosphoribosyltransferase, immunoreactive trypsin, lactate , lead, lipoproteins ((a), B / A-1, β), lysozyme, mefloquine, netilmicin, phenobarbitone, phenytoin, phytanic acid / pristanic acid, progesterone, prolactin, prolidase, purine nucleoside phosphorylase, quinine, inverted tri-iodothyronine (rT3), selenium, serum pancreatic lipase, sisomicin, somatomedin C, specific antibodies (adenovirus, antinuclear antibody, anti-zeta antibody, arbovirus, Aujeszky's disease virus, dengue virus, guinea worm, Echinococcus granulosus, Entamoeba histolytica, enterovirus, Giardia lamblia (giardia duodenalisa), Helicobacter pylori, Hepatitis B virus, Herpes virus, HIV-1, IgE (atopic disease), Influenza virus, Leishmania donovani, Leptospirosis, Measles / Mumps / Rubella, Mycobacterium leprae, Mycoplasma pneumoniae, Myoglobin, Onchocerciasis volvulus, Parainfluenza virus, Plasmodium falciparum, Poliovirus, Pseudomonas aeruginosa, Respiratory syncytial virus, Rickettsia (scrub typhus), Schistosoma mansoni, Toxoplasma gondii, Treponema pallidum, Trypanosoma cruzi / Langer, Vesicular stomatitis virus virus), Wuchereria bancrofti, Yellow fever virus), specific antigens (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, leukocytes, and zinc protoporphyrin.The present invention is not limited to the above. Salts, sugars, proteins, fats, vitamins, and hormones naturally occurring in blood or interstitial fluids may also constitute analytes in certain embodiments. Analytes, such as metabolites, hormones, antigens, antibodies, etc., may be naturally present in biological fluids or may be endogenous. Alternatively, analytes, such as contrast agents for diagnostic imaging, radioisotopes, chemical agents, fluorocarbon-based artificial blood, or drugs or pharmaceutical compositions, may be introduced into the body or may be exogenous, including insulin, ethanol, cannabis (marijuana, tetrahydrocannabinol, hashish), inhalants (nitrous oxide, amyl nitrite, butyl nitrite, chlorohydrocarbons, hydrocarbons), cocaine (crack cocaine), stimulants (amphetamines, methamphetamines, Ritalin, Cylert, Preludin, Didrex, PreState, Voranil, Sandrex, Plegine), depressants (barbiturates, methaqualone, psychotropic drugs), and the like. These include, but are not limited to, stabilizers (e.g., Valium, Librium, Miltown, Serax, Equanil, Tranxene), hallucinogens (phencyclidine, lysergic acid, mescaline, peyote, psilocybin), narcotics (heroin, codeine, morphine, opium, meperidine, Percocet, Percodan, Tussionex, Fentanyl, Darvon, Talwin, Lomotil), designer drugs (fentanyl, meperidine, amphetamine, methamphetamine, and phencyclidine analogs, e.g., Ecstasy), anabolic steroids, and nicotine. Metabolites of drugs and pharmaceutical compositions may also be contemplated as analytes. Analytes such as neurochemicals and other chemicals produced in the body, such as ascorbic acid, uric acid, dopamine, noradrenaline, 3-methoxytyramine (3MT), 3,4-dihydroxyphenylacetic acid (DOPAC), homovanillic acid (HVA), 5-hydroxytryptamine (5HT), and 5-hydroxyindoleacetic acid (FHIAA), may also be analyzed.

[0034] As used herein, the term "baseline" is a broad term given its ordinary and customary meaning to those skilled in the art (and is not limited to any special or customized meaning), and refers to, but is not limited to, a component of an analyte sensor signal that is not related to analyte concentration. In one example of a glucose sensor, the baseline is substantially composed of signal contributions attributable to factors other than glucose (e.g., interfering species, unreacted associated hydrogen peroxide, or other electroactive species with an oxidation potential that overlaps with hydrogen peroxide). In some embodiments where calibration is defined by solving the equation y=mx+b, the value of b represents the baseline of the signal.

[0035] As used herein, the term "continuous (or continuous) analyte sensing" is a broad term given its ordinary and customary meaning to those skilled in the art (and is not limited to any special or customized meaning), and refers, without limitation, to a period during which analyte concentration monitoring is performed continuously, continuously, and / or intermittently (but regularly), for example, about every 5-10 minutes.

[0036] As used herein, the term "counts" is a broad term given its ordinary and customary meaning to those skilled in the art (and is not limited to any special or customized meaning), and refers to, but is not limited to, a unit of measurement for a digital signal. In one example, the raw data stream measured in counts is directly related to a voltage (e.g., converted by an A / D converter), which is directly related to the current from the working electrode. In another example, the counter electrode voltage measured in counts is directly related to a voltage.

[0037] As used herein, the terms "dipole" or "dipolar compound" are broad terms that are given their ordinary and customary meaning to those skilled in the art (and are not limited to any special or customized meaning), and refer to, but are not limited to, compounds in which a neutral molecule of the compound bears positive and negative charges at different locations within the molecule. The positive and negative charges within the molecule can be any non-zero charge up to and including a full unit charge.

[0038] As used herein, the term "distal" is a broad term given its ordinary and customary meaning to those skilled in the art (and is not limited to any special or customized meaning), and refers to, but is not limited to, the spatial relationship between various elements relative to a particular reference point. For example, some embodiments of a sensor include a membrane system having a biointerface domain and an enzyme domain. If the sensor is considered to be the reference point and the biointerface domain is positioned further from the sensor than the enzyme domain, then the biointerface domain is more distal to the sensor than the enzyme domain.

[0039] As used herein, the term "domain" is a broad term that is given its ordinary and customary meaning to those skilled in the art (and is not limited to any special or customized meaning), and refers to, but is not limited to, a region of a film that may be a uniform or non-uniform gradient (i.e., anisotropic) layer or may be provided as part of a film.

[0040] As used herein, the term "electrical potential" is a broad term given its ordinary and customary meaning to those skilled in the art (and is not limited to any special or customized meaning), and refers to, but is not limited to, the potential difference between two points in a circuit that is responsible for the flow of electric current.

[0041] As used herein, the terms "electrochemically reactive surface" or "electroactive surface" are broad terms that are given their ordinary and customary meaning to those skilled in the art (and are not limited to any special or customized meaning) and refer to, but are not limited to, the surface of an electrode on which an electrochemical reaction occurs. As an example, at the working electrode, H2O2 (hydrogen peroxide), produced by an enzyme-catalyzed reaction of the analyte being detected, reacts, thereby creating a measurable electrical circuit. For example, in the detection of glucose, glucose oxidase produces H2O2 as a by-product. H2O2 reacts with the surface of the working electrode to release two protons (2H + ), two electrons (2e - ), and one oxygen molecule (O), which generates a current that is detected. In the case of the counter electrode, a reducible species, e.g., O, is reduced at the electrode surface to balance the current generated by the working electrode.

[0042] As used herein, the term "host" is a broad term given its ordinary and customary meaning to those skilled in the art (and is not limited to any special or customized meaning), and refers to, but is not limited to, animals (e.g., humans) and plants. In some examples, hosts can include domestic animals (e.g., dogs, cats, etc.), livestock (e.g., cows, horses, pigs, sheep, goats, etc.), laboratory animals (e.g., mice, rabbits, rats, guinea pigs, etc.), and birds. In other examples, hosts can include mammals such as primates or humans.

[0043] As used herein, the terms "interferent" and "interfering species" are broad terms given their ordinary and customary meanings to those skilled in the art (and are not limited to any special or customized meanings), and refer to, but are not limited to, effects or species that interfere with the measurement of an analyte of interest in a sensor, producing a signal that does not accurately represent the analyte measurement. In an exemplary electrochemical sensor, an interfering species can include compounds that have an oxidation potential that overlaps with the oxidation potential of the analyte being measured.

[0044] As used herein, the terms "non-zwitterionic dipole" and "non-zwitterionic zwitterionic compound" are broad terms that are given their ordinary and customary meaning to those of skill in the art (and are not limited to any special or customized meaning), and refer to, but are not limited to, compounds in which a neutral molecule of the compound has positive and negative charges at different positions within the molecule. The positive and negative charges within the molecule may be any non-zero charge, although they are less than a full unit.

[0045] As used herein, the terms "operably connected," "operably connected," and "operably linked" are broad terms given their ordinary and customary meanings to those skilled in the art (and are not limited to any special or customized meanings), and refer, without limitation, to one or more components that are connected to another component(s) in a manner that allows for the transmission of a signal between the components. For example, one or more electrodes can be used to detect the amount of analyte in a sample and convert that information into a signal that can then be transmitted to a circuit. In this case, the electrodes are "operably linked" to the electronic circuit.

[0046] The term "optional" or "optionally" means that the subsequently described event or circumstance may or may not occur, and that the description includes instances where the event or circumstance occurs and instances where it does not occur.

[0047] As used herein, the term "polyampholyte polymer" is a broad term given its ordinary and customary meaning to those skilled in the art (and is not limited to any special or customized meaning), and refers to, but is not limited to, polymers containing both cationic and anionic groups. Such polymers can be prepared to have approximately equal numbers of positive and negative charges, and thus the surface of such polymers can be approximately net neutral in charge. Alternatively, such polymers can be prepared to have an excess of either positive or negative charges, and thus the surface of such polymers can be net positive or negative, respectively.

[0048] As used herein, the term "polyzwitterion" is a broad term, given its ordinary and customary meaning to those skilled in the art (and not limited to any special or customized meaning), and refers to, but is not limited to, polymers in which the repeat units of the polymer chain are zwitterionic moieties. Polyzwitterions are also known as polybetaines. Polyzwitterions are a type of polyampholyte polymer because they have both cationic and anionic groups. However, because both cationic and anionic groups are part of the same repeat unit, they are unique, meaning that polyzwitterions have the same number of cationic and anionic groups, while other polyampholyte polymers may have more of some ionic groups than others. Polyzwitterions also have cationic and anionic groups as part of the repeat unit. Polyampholyte polymers need not have cationic groups connected to anionic groups; they can be on different repeat units and thus distributed apart from each other at random intervals, or one ionic group may outnumber the others.

[0049] As used herein, the term "proximal" is a broad term given its ordinary and customary meaning to those skilled in the art (and is not limited to any special or customized meaning), and refers to, but is not limited to, the spatial relationship between various elements relative to a particular reference point. For example, some embodiments of a device include a membrane system having a biointerface layer and an enzyme layer. If the sensor is considered to be the reference point and the enzyme layer is positioned closer to the sensor than the biointerface layer, then the enzyme layer is more proximal to the sensor than the biointerface layer.

[0050] As used herein, the terms "raw data stream" and "data stream" are broad terms given their ordinary and customary meanings to those skilled in the art (and are not limited to any special or customized meanings) and refer to, but are not limited to, an analog or digital signal directly related to a measured glucose concentration from a glucose sensor. In one embodiment, the raw data stream is digital data in "counts" (e.g., voltage or amperage) converted from an analog signal by an A / D converter and represents a glucose concentration. These terms broadly encompass multiple time-spaced data points from a substantially continuous glucose sensor, including individual measurements taken at time intervals ranging from less than one second up to, for example, one, two, or five minutes or more.

[0051] As used herein, the terms "sensing membrane" and "membrane system" are broad terms given their ordinary and customary meanings to those skilled in the art (and are not limited to any special or customized meanings), and refer, without limitation, to a permeable or semi-permeable membrane composed of a few microns or more of material that may contain one or more domains or layers, that is permeable to oxygen, and that may or may not be permeable to an analyte of interest. In one example, the sensing membrane or membrane system may include immobilized glucose oxidase enzyme, which allows an electrochemical reaction to occur to measure the concentration of glucose.

[0052] As used herein, the terms "sensing area," "sensor," and "sensing mechanism" are broad terms that are given their ordinary and customary meanings to those skilled in the art (and are not limited to any special or customized meanings) and refer to, but are not limited to, an area or mechanism of a monitoring device that is involved in the detection of a particular analyte.

[0053] As used herein, the term "sensitivity" is a broad term given its ordinary and customary meaning to those skilled in the art (and is not limited to any special or customized meaning), and refers, without limitation, to the amount of signal (e.g., in the form of current and / or voltage) generated by a given amount (unit) of measured analyte. For example, in one embodiment, the sensor has a sensitivity (or slope) of about 1 to about 100 picoamps of current per 1 mg / dL of glucose analyte.

[0054] As used herein, the terms "zwitterion" and "zwitterionic compound" are broad terms that are given their ordinary and customary meaning to those of skill in the art (and are not limited to any special or customized meaning), and refer to, but are not limited to, compounds in which a neutral molecule of the compound bears a unit positive charge and a unit negative charge at different positions within the molecule. Such compounds are a type of zwitterionic compound and are sometimes referred to as "inner salts."

[0055] As used herein, the terms "zwitterion precursor" or "zwitterionic compound precursor" are broad terms, given their ordinary and customary meanings to those skilled in the art (and are not limited to any special or customized meanings), and refer to, but are not limited to, any compound that is not itself a zwitterion but can become a final or transitional zwitterion through chemical reaction. In some embodiments described herein, the device includes a zwitterion precursor that can be converted to a zwitterion prior to in vivo implantation of the device. Alternatively, in some embodiments described herein, the device includes a zwitterion precursor that can be converted to a zwitterion through some chemical reaction that occurs after in vivo implantation of the device. Such reactions are known to those skilled in the art and include ring-opening reactions, addition reactions such as Michael addition, and other reactions. This method is particularly useful when polymerization of betaine-containing monomers to achieve desired physical properties, such as molecular weight and mechanical strength, is difficult due to technical challenges, such as the solubility of the betaine monomer. Post-polymerization modification or conversion of betaine precursors can be a practical method for achieving desired polymer structures and compositions. Examples of such precursors include tertiary amines, quaternary amines, pyridine, and others detailed herein.

[0056] As used herein, the terms "zwitterionic derivative" or "zwitterionic compound derivative" are broad terms that are given their ordinary and customary meaning to those skilled in the art (and are not limited to any special or customized meaning) and refer to, but are not limited to, any compound that is not itself a zwitterion, but rather is the product of a chemical reaction in which a zwitterion is converted to a non-zwitterion. Such reactions can be reversible, so that under certain conditions, a zwitterionic derivative can act as a zwitterionic precursor. For example, a hydrolyzable betaine ester formed from a zwitterionic betaine is a cationic zwitterionic derivative that can undergo hydrolysis back to the zwitterionic betaine under appropriate conditions.

[0057] As used herein, the following abbreviations apply: Eq and Eqs (equivalents), mEq (milliequivalents), M (mole), mM (millimolar), μM (micromolar), N (normal), mol (mole), mmol (millimolar), μmol (micromolar), nmol (nanomole), g (gram), mg (milligram), μg (microgram), Kg (kilogram), L (liter), mL (milliliter), dL (deciliter), μL (microliter), cm (centimeter), mm (millimeter), μm (micrometer), nm (nanometer), h and hr (hours), min (minute), and sec (second), °C (degrees Celsius).

[0058] Sensor System FIG. 1 is a schematic diagram of a continuous analyte sensor system 100 attached to a host and in communication with a number of other exemplary devices 110-113. The transdermal analyte sensor system is shown comprising an on-skin sensor assembly 600 secured to the host's skin via a disposable housing (not shown). The system includes a transdermal analyte sensor 200 and an electronics unit (interchangeably referred to as "sensor electronics" or "transmitter") 500 for wirelessly transmitting analyte information to a receiver. During use, a sensing portion of the sensor 200 resides beneath the host's skin, and a contact portion of the sensor 200 is operably connected (e.g., electrically connected) to the electronics unit 500. The electronics unit 500 is engaged with a housing attached to an adhesive patch secured to the host's skin.

[0059] The on-skin sensor assembly 600 may be attached to the host using an applicator adapted to provide convenient and safe application. Such an applicator may also be used to insert the sensor 200 through the skin of the host. Once the sensor 200 is inserted, the applicator is detached from the sensor assembly.

[0060] In general, continuous analyte sensor system 100 includes any sensor configuration that provides an output signal indicative of an analyte concentration. The output signal (e.g., including sensor data such as a raw data stream, filtered data, smoothed data, and / or otherwise transformed sensor data) is transmitted to a receiver, which may be, for example, a smartphone, a smartwatch, a dedicated device, or the like. In one embodiment, analyte sensor system 100 includes a transcutaneous glucose sensor such as described in U.S. Patent Publication No. US-2011-0027127-A1 (the contents of which are incorporated herein by reference in their entirety). In some embodiments, sensor system 100 includes a continuous glucose sensor, such as a transcutaneous sensor such as described in U.S. Patent No. 6,565,509 to Say et al. In another embodiment, sensor system 100 includes a continuous glucose sensor, such as a subcutaneous sensor such as described with reference to U.S. Patent No. 6,579,690 to Bonnecaze et al. and U.S. Patent No. 6,484,046 to Say et al. In another embodiment, sensor system 100 includes a continuous glucose sensor, such as a subcutaneous sensor as described with reference to U.S. Pat. No. 6,512,939 to Colvin et al. In another embodiment, sensor system 100 includes a continuous glucose sensor, such as an intravascular sensor as described with reference to U.S. Pat. No. 6,477,395 to Schulman et al. In another embodiment, sensor system 100 includes a continuous glucose sensor, such as an intravascular sensor as described with reference to U.S. Pat. No. 6,424,847 to Mastrototaro et al. Other signal processing techniques and glucose monitoring system embodiments suitable for use with the embodiments described herein are described in U.S. Patent Publication Nos. US-2005-0203360-A1 and US-2009-0192745-A1, the contents of which are incorporated herein by reference in their entireties. The sensor extends through a housing that maintains the sensor on the skin and is provided in an electronics unit that provides electrical connection between the sensor and the sensor electronics.

[0061] In one embodiment, the sensor is formed from or in the form of a wire. For example, the sensor may include an elongated conductor, such as a bare, elongated conductive core (e.g., a metal wire) or an elongated conductive core coated with one, two, three, four, five, or more layers of material, each of which may or may not be conductive. The elongated sensor may be long and thin, yet flexible and strong. For example, in some embodiments, the smallest dimension of the elongated conductor is less than about 0.1 inches, less than about 0.075 inches, less than about 0.05 inches, less than about 0.025 inches, less than about 0.01 inches, less than about 0.004 inches, or less than about 0.002 inches. The sensor may have a circular cross-section. In some embodiments, the cross-section of the elongated conductor may be oval, rectangular, triangular, polygonal, star-shaped, C-shaped, T-shaped, X-shaped, Y-shaped, irregular, etc. In one embodiment, a conductive wire electrode is used as the core. One or two additional conductive layers may be added to such clad electrodes (e.g., with an intervening insulating layer to provide electrical isolation). The conductive layers may be composed of any suitable material. In certain embodiments, it may be desirable to use a conductive layer that includes conductive particles (i.e., particles of a conductive material) in a polymer or other binder.

[0062] In certain embodiments, the materials used to form the elongated conductors (e.g., stainless steel, titanium, tantalum, platinum, platinum-iridium, iridium, certain polymers, and / or the like) can be strong and rigid, thereby preventing damage. For example, in some embodiments, the ultimate tensile strength of the elongated conductors is between about 80 kPsi and about 500 kPsi. In other examples, in some embodiments, the Young's modulus of the elongated conductors is between about 160 GPa and about 220 GPa. In yet other examples, in some embodiments, the yield strength of the elongated conductors is between about 60 kPsi and about 2200 kPsi. In some embodiments, the small diameter of the sensor provides (e.g., imparts, enables) flexibility to these materials, and thus to the sensor as a whole. Thus, the sensor can withstand repeated forces exerted on it by the surrounding tissue.

[0063] In addition to providing structural support, resilience, and flexibility, in some embodiments, the core (or components thereof) provides electrical conduction for an electrical signal from the working electrode to the sensor electronics (not shown). In some embodiments, the core comprises a conductive material such as stainless steel, titanium, tantalum, a conductive polymer, and / or the like. However, in other embodiments, the core is formed from a non-conductive material such as a non-conductive polymer. In still other embodiments, the core comprises multiple layers of material. For example, in one embodiment, the core comprises an inner core and an outer core. In further embodiments, the inner core is formed from a first conductive material and the outer core is formed from a second conductive material. For example, in some embodiments, the first conductive material is stainless steel, titanium, tantalum, a conductive polymer, an alloy, and / or the like, and the second conductive material is a conductive material selected to provide electrical conduction between the core and the first layer and / or to adhere the first layer to the core (e.g., when the first layer is formed from a material that does not adhere well to the core material). In another embodiment, the core is formed of a non-conductive material (e.g., a non-conductive metal and / or a non-conductive polymer) and the first layer is a conductive material such as stainless steel, titanium, tantalum, a conductive polymer, and / or the like. The core and the first layer may be a single (or the same) material, e.g., platinum. Those skilled in the art will appreciate that additional configurations are possible.

[0064] In the illustrated embodiment, electronics unit 500 is removably attachable to sensor 200. Electronics unit 500 includes electronics circuitry associated with measuring and processing continuous analyte sensor data and is configured to execute algorithms associated with processing and calibrating sensor data. For example, electronics unit 500 can provide various aspects of the functionality of the sensor electronics module described in U.S. Patent Publication No. US-2009-0240120-A1 and U.S. Patent Application No. 13 / 247,856, filed September 28, 2011, entitled "ADVANCED CONTINUOUS ANALYTE MONITORING SYSTEM," the contents of which are incorporated herein by reference in their entireties. Electronics unit 500 can include hardware, firmware, and / or software that enable measurement of analyte levels via a glucose sensor, such as analyte sensor 200. For example, the electronics unit 500 may include a potentiostat, a power supply for providing power to the sensor 200, other components useful for signal processing and data storage, and preferably a telemetry module for one-way or two-way data communication between the electronics unit 500 and one or more receivers, repeaters, and / or display devices, such as devices 110-113. The electronics may be affixed to a printed circuit board (PCB) or the like and may take a variety of forms. For example, the electronics may take the form of an integrated circuit (IC), such as an application specific integrated circuit (ASIC), a microcontroller, and / or a processor. The electronics unit 500 may include sensor electronics configured to process the sensor information, e.g., store the data, analyze the data stream, calibrate the analyte sensor data, estimate analyte values, compare the estimated analyte values ​​to time-corresponding measured analyte values, and analyze the various estimated analyte values.Examples of systems and methods for processing sensor analyte data are described herein and in U.S. Pat. No. 7,310,544, U.S. Pat. No. 6,931,327, U.S. Patent Publication No. 2005-0043598-A1, U.S. Patent Publication No. 2007-0032706-A1, U.S. Patent Publication No. 2007-0016381-A1, U.S. Patent Publication No. 2008-0033254-A1, U.S. Patent Publication No. 2005-0016381-A1, U.S. Patent Publication No. 2008 ... Nos. 2005-0203360-A1, 2005-0154271-A1, 2005-0192557-A1, 2006-0222566-A1, 2007-0203966-A1, and 2007-0208245-A1, the contents of which are incorporated herein by reference in their entireties.

[0065] One or more transponders, receivers, and / or display devices, such as a key fob transponder 110, a medical device receiver 111 (e.g., an insulin delivery device and / or a dedicated glucose sensor receiver), a smartphone 112, a portable computer 113, etc., are operably connected to the electronics unit 500, which is also referred to herein as a transmitter and / or sensor electronics body, to receive data from the electronics unit 500 and, in some embodiments, transmit data to the electronics unit 500. For example, sensor data may be transmitted from the sensor electronics unit 500 to one or more of the key fob transponder 110, the medical device transponder 111, the smartphone 112, the portable computer 113, etc. In one embodiment, the display device includes an input module having a quartz crystal operably connected to an RF transceiver (not shown), which together function to transmit, receive, and synchronize the data stream from the electronics unit 500. However, the input module may be configured in any manner capable of receiving data from the electronics unit 500. Once received, the input module transmits the data stream to a processor, which processes the data stream, as described in more detail below. The processor is a central control unit that performs processes such as storing data, analyzing the data stream, calibrating analyte sensor data, estimating analyte values, comparing estimated analyte values ​​with time-corresponding measured analytes, analyzing variations in estimated analyte values, downloading data, and controlling the user interface by providing analyte values, prompts, messages, warnings, alarms, etc. The processor includes hardware that performs the processes described herein, such as read-only memory (ROM) providing permanent or semi-permanent storage of data, storage of data such as sensor ID (sensor identifier), receiver ID (receiver identifier), and programming for processing the data stream (e.g., programming for performing estimation and other algorithms described elsewhere herein), and random access memory (RAM) storing cache memory for the system and aiding in data processing.The output module, which may be integrated with and / or operatively connected to the processor, includes programming (and any processing that occurs within the processor) for generating output based on sensor data received from the electronics unit.

[0066] In some embodiments, the analyte value is displayed on a display device. In some embodiments, prompts or messages can be displayed on the display device to convey information to the user, such as reference outliers, a request for a reference analyte value, a treatment recommendation, deviation of the measured analyte value from the estimated analyte value, etc. Additionally, prompts can be displayed to guide the user through calibration or troubleshooting the calibration.

[0067] Additionally, data output from the output module can provide wired or wireless one-way or two-way communication between the receiver and an external device. The external device can be any device that interfaces or communicates with the receiver. In some embodiments, the external device is a computer, and the receiver can download current or historical data for retrospective analysis, for example, by a physician. In some embodiments, the external device is a modem, and the receiver can send alerts, warnings, emergency messages, etc. to a physician or other party, such as a family member, over a communication line. In some embodiments, the external device is an insulin pen, and the receiver can communicate treatment recommendations, such as insulin amount and time, to the insulin pen. In some embodiments, the external device is an insulin pump, and the receiver can communicate treatment recommendations, such as insulin amount and time, to the insulin pump. The external device can include other technology or medical devices, such as, for example, a pacemaker, an implantable analyte sensor patch, other infusion devices, telemetry devices, etc. The receiver may communicate with the external device and / or any number of additional devices via any suitable communication protocol, including radio frequency, Bluetooth®, Universal Serial Bus (USB), Wireless Local Area Network (WLAN) communication standards (including IEEE 802.11, 802.15, 802.20, 802.22, and other 802 communication protocols), ZigBee, wireless (e.g., cellular) telecommunications, paging network communications, magnetic induction, satellite data communications, GPRS, ANT, and / or proprietary communication protocols.

[0068] The implementations described herein generally discuss sensors configured with one or more sensor wires. However, it will be understood that sensors are not limited to such wire shapes or linear arrangements. Rather, sensors may be implemented as planar sensors, volume sensors, point sensors, or in other shapes that will be understood in light of this description.

[0069] Membrane System The membrane systems disclosed herein are suitable for use with implantable devices in contact with biological fluids. For example, the membrane systems can be utilized with implantable devices such as devices for monitoring and determining analyte levels in biological fluids, e.g., devices for monitoring glucose levels in individuals with diabetes. In some embodiments, the analyte measuring device is a continuous device. The analyte measuring device provides a raw signal using any suitable sensing element, including, but not limited to, those involving enzymatic, chemical, physical, electrochemical, spectrophotometric, polarimetric, calorimetric, radiometric, immunochemical, or similar elements.

[0070] Although some of the following description, including the described membrane systems and methods for their use, is directed to glucose measurement devices, these membrane systems are not limited to use in devices that measure or monitor glucose. These membrane systems are suitable for use in any of a variety of devices, including, for example, devices that detect and quantify other analytes present in biological fluids (e.g., cholesterol, amino acids, alcohols, galactose, and lactate), cell transplantation devices (see, e.g., U.S. Pat. Nos. 6,015,572, 5,964,745, and 6,083,523), drug delivery devices (see, e.g., U.S. Pat. Nos. 5,458,631, 5,820,589, and 5,972,369), and the like.

[0071] In one embodiment, the analyte measuring device is an implantable glucose sensor as described with reference to U.S. Patent No. 6,001,067 and U.S. Patent Publication No. US-2005-0027463-A1, which are incorporated by reference in their entireties. In another embodiment, the analyte measuring device is a glucose sensor as described with reference to U.S. Patent Publication No. US-2006-0020187-A1, which are incorporated by reference in their entireties. In yet other embodiments, the sensor is configured to be implanted in a host vessel or extracorporeally, as described in U.S. Patent Publication Nos. US-2007-0027385-A1, US-2008-0119703-A1, US-2008-0108942-A1, and US-2007-0197890-A1, which are incorporated by reference in their entireties. In some embodiments, the sensor is configured as a dual-electrode sensor, such as those described in U.S. Patent Publication Nos. US-2005-0143635-A1, US-2007-0027385-A1, US-2007-0213611-A1, and US-2008-0083617-A1 (incorporated herein by reference in their entireties). In one alternative embodiment, the continuous glucose sensor includes a sensor such as that described in U.S. Patent No. 6,565,509 to Say et al. In another alternative embodiment, the continuous glucose sensor includes a subcutaneous sensor such as that described with reference to U.S. Patent No. 6,579,690 to Bonnecaze et al. or U.S. Patent No. 6,484,046 to Say et al. In another alternative embodiment, the continuous glucose sensor includes a refillable subcutaneous sensor such as that described with reference to U.S. Patent No. 6,512,939 to Colvin et al. In yet another alternative embodiment, the continuous glucose sensor comprises an intravascular sensor, for example, as described with reference to U.S. Patent No. 6,477,395 to Schulman et al. In another alternative embodiment, the continuous glucose sensor comprises an intravascular sensor, for example, as described with reference to U.S. Patent No. 6,424,847 to Mastrototaro et al.In some embodiments, the electrode system may be any of a variety of known in vivo analyte sensors or monitors, such as U.S. Pat. No. 7,157,528 to Ward, U.S. Pat. No. 6,212,416 to Ward et al., U.S. Pat. No. 6,119,028 to Schulman et al., U.S. Pat. No. 6,400,974 to Lesho, U.S. Pat. No. 6,595,919 to Berner et al., U.S. Pat. No. 6,141,573 to Kurnik et al., U.S. Pat. No. 6,122,536 to Sun et al., European Patent Publication No. EP 1153571 to Varall et al., U.S. Pat. No. 6,122,536 to Co. No. 6,512,939 to Ivin et al., U.S. Patent No. 5,605,152 to Slate et al., U.S. Patent No. 4,431,004 to Bessman et al., U.S. Patent No. 4,703,756 to Gough et al., U.S. Patent No. 6,514,718 to Heller et al., U.S. Patent No. 5,985,129 to Gough et al., PCT International Publication No. WO 4 / 021877 to Caduff, U.S. Patent No. 5,494,562 to Maley et al., U.S. Patent No. 6,120,676 to Heller et al., and U.S. Patent No. 6,542,765 to Guy et al. In general, the disclosed embodiments are applicable to a variety of continuous analyte measurement device configurations.

[0072] In some embodiments, long-term sensors (e.g., fully implantable or intravascular) are configured and positioned to function for a period (e.g., a sensor session) of about 30 days or less to about a year or more. In some embodiments, short-term sensors (e.g., transcutaneous or intravascular) are configured and positioned to function for a period of about several hours to about 30 days, including a period (e.g., a sensor session) of about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, or 29 days. As used herein, the term "sensor session" is a broad term and refers to, but is not limited to, the period during which a sensor is applied to a host (e.g., implanted) or used to obtain sensor values. For example, in some embodiments, a sensor session extends from the time of sensor implantation (e.g., including inserting the sensor into subcutaneous tissue and placing the sensor in fluid communication with the host's circulatory system) to the time the sensor is removed.

[0073] Generally, membrane systems include multiple domains, such as an electrode domain, an interference domain, an enzyme domain, a resistance domain, and a biointerface domain. Membrane systems can be deposited onto exposed electroactive surfaces using known thin-film techniques (e.g., vapor deposition, spraying, electrodeposition, dipping, brush coating, film coating, droplet coating, etc.). Following membrane material deposition, additional steps, such as drying, baking, and curing (e.g., UV curing, thermal curing, moisture curing, radiation curing, etc.), can be applied to enhance certain properties, such as mechanical properties, signal stability, and selectivity. In a typical process, upon deposition of the interference layer membrane, the enzyme layer has a "dry film" thickness of about 0.05 μm to about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, or 16 μm. "Dry film" thickness refers to the thickness of a cured film cast from a coating formulation using standard coating techniques.

[0074] In certain embodiments, the enzyme layer is formed from an enzyme layer polymer and an active enzyme, where the enzyme layer polymer comprises a polyurethane and / or polyurea segment and one or more zwitterionic repeat units. In some embodiments, the enzyme layer coating is formed from a polyurethaneurea having carboxyl betaine groups incorporated into the polymer and nonionic hydrophilic polyethylene oxide segments. The polyurethaneurea polymer is dissolved in an organic or non-organic solvent system according to a predetermined coating formulation, crosslinked with an isocyanate crosslinker, and cured at a moderate temperature of about 50°C. The solvent system can be a single solvent or a mixture of solvents that aid in the dissolution or dispersion of the polymer. The solvent can be the one selected as the polymerization medium or added after polymerization is complete. The solvent is preferably selected to have a lower boiling point to facilitate drying, a low ability to denature the enzyme, and low toxicity for implant applications. Examples of these solvents include water, lipid-soluble ketones, esters, ethers, alcohols, carbohydrates, etc. Depending on the final thickness of the enzyme layer and the solution viscosity (related to the percent of polymer solids), the coating can be applied in a single step or multiple repeated steps of a selected process, such as dipping, to build up the desired thickness. In yet another embodiment, the enzyme layer polymer is formed of a polyurethaneurea having carboxylic acid and carboxyl betaine groups incorporated into the polymer, as well as nonionic hydrophilic polyethylene oxide segments; this polyurethaneurea polymer is dissolved in an organic or non-organic solvent system in the coating formulation, crosslinked with a carbodiimide (e.g., 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC)), and cured at moderate temperatures of about 50°C. Other crosslinkers, such as multifunctional aziridines, can be used as well.

[0075] In another embodiment, the enzyme layer is formed of a polyurethaneurea with sulfobetaine groups incorporated into the polymer and nonionic hydrophilic polyethylene oxide segments. This polyurethaneurea polymer is dissolved in an organic or non-organic solvent system according to the coating formulation, crosslinked with an isocyanate crosslinker, and cured at a moderate temperature of approximately 50°C. The solvent system can be a single solvent or a mixture of solvents that aid in dissolving or dispersing the polymer. The solvent can be selected as the polymerization medium or added after polymerization is complete. The solvent is preferably selected to have a lower boiling point to facilitate drying and low toxicity for implant applications. Examples of these solvents include lipid-soluble ketones, esters, ethers, alcohols, carbohydrates, etc. Depending on the final thickness of the enzyme layer and the solution viscosity (related to the percent polymer solids), the coating can be applied in a single step or multiple repeated steps of a selected process, such as dipping, to build up the desired thickness. In yet another embodiment, the enzyme layer polymer is formed of a polyurethaneurea having unsaturated hydrocarbon and sulfobetaine groups incorporated into the polymer, and non-ionic hydrophilic polyethylene oxide segments, which polyurethaneurea polymer is dissolved in an organic or non-organic solvent system in a coating formulation, crosslinked by heat and irradiation, including UV, LED light, e-beam, etc., in the presence of an initiator, and cured at a moderate temperature of about 50° C. Examples of unsaturated hydrocarbons include allyl groups, vinyl groups, acrylates, methacrylates, alkenes, alkynes, etc.

[0076] 3A-3C illustrate an embodiment of an in vivo portion of a continuous analyte sensor 400, including an elongated conductor 402. The elongated conductor 402 includes a core 410 (see FIG. 3B) and a first layer 412 at least partially surrounding the core. The first layer includes a working electrode (e.g., located within a window 406) and a membrane 408 located on the working electrode. In some embodiments, the core and first layer can be a single material (e.g., platinum). In some embodiments, the elongated conductor is a composite of at least two materials, such as two conductive materials, or at least one conductive material and at least one non-conductive material. In some embodiments, the elongated conductor includes multiple layers. In certain embodiments, there are at least two concentric or annular layers, such as a core formed of a first material and a first layer formed of a second material. However, in some embodiments, additional layers may be included. In some embodiments, the layers are coaxial.

[0077] The elongated conductors can be long and thin, yet flexible and strong. For example, in some embodiments, the smallest dimension of the elongated conductor is less than about 0.1 inches, 0.075 inches, 0.05 inches, 0.025 inches, 0.01 inches, 0.004 inches, or 0.002 inches. While the elongated conductors are shown in FIGS. 3A-3C as having a circular cross-section, in other embodiments, the cross-section of the elongated conductors can be oval, rectangular, triangular, polygonal, star-shaped, C-shaped, T-shaped, X-shaped, Y-shaped, irregular, etc. In one embodiment, a conductive wire electrode is used as the core. Two additional conductive layers may be added to such a clad electrode (e.g., with an intervening insulating layer to provide electrical isolation). The conductive layers may be composed of any suitable material. In certain embodiments, it may be desirable to use a conductive layer comprising conductive particles (i.e., particles of a conductive material) in a polymer or other binder.

[0078] The materials used to form the elongated conductors (e.g., stainless steel, titanium, tantalum, platinum, platinum-iridium, iridium, certain polymers, and / or the like) can be strong and rigid, preventing breakage. In some embodiments, the small diameter of the sensor provides flexibility to these materials, and thus to the sensor as a whole. Thus, the sensor can withstand repeated forces exerted on it by the surrounding tissue.

[0079] In addition to providing structural support, resilience, and flexibility, in some embodiments, the core 410, or components thereof, provides electrical conduction for an electrical signal from the working electrode to the sensor electronics (not shown). In some embodiments, the core 410 comprises a conductive material such as stainless steel, titanium, tantalum, a conductive polymer, and / or the like. However, in other embodiments, the core is formed from a non-conductive material, such as a non-conductive polymer. In still other embodiments, the core comprises multiple layers of material. For example, in one embodiment, the core comprises an inner core and an outer core. In further embodiments, the inner core is formed from a first conductive material and the outer core is formed from a second conductive material. For example, in some embodiments, the first conductive material is stainless steel, titanium, tantalum, a conductive polymer, an alloy, and / or the like, and the second conductive material is a conductive material selected to provide electrical conduction between the core and the first layer and / or to adhere the first layer to the core (i.e., if the first layer is formed from a material that does not adhere well to the core material). In another embodiment, the core is formed of a non-conductive material (e.g., a non-conductive metal and / or a non-conductive polymer, etc.) and the first layer is formed of a conductive material, such as stainless steel, titanium, tantalum, a conductive polymer, and / or the like. The core and the first layer may be a single (or the same) material, e.g., platinum. Those skilled in the art will appreciate that additional configurations are possible.

[0080] 3A-3C, the first layer 412 may be formed of an electrically conductive material, and the working electrode may be an exposed portion of the surface of the first layer 412. Thus, the first layer 412 may be formed of a material configured to provide a suitable electroactive surface for the working electrode, such as, but not limited to, platinum, platinum-iridium, gold, palladium, iridium, graphite, carbon, a conductive polymer, an alloy, and / or the like.

[0081] As shown in FIGS. 3B and 3C , the second layer 404 surrounds at least a portion of the first layer 412, thereby defining the boundary of the working electrode. In some embodiments, the second layer 404 functions as an insulator and is formed of an insulating material, such as polyimide, polyurethane, parylene, or any other known insulating material. For example, in one embodiment, the second layer is disposed on the first layer and configured such that the working electrode is exposed through a window 406. In some embodiments, an elongated conductor is provided that includes a core, a first layer, and a second layer. A portion of the second layer can be removed to form the window 406, thereby exposing the electroactive surface of the working electrode (i.e., the exposed surface of the first layer 412). In some embodiments, a portion of the second layer and (optionally) the third layer can be removed to form the window 406, thus exposing the working electrode. Removal of coating material from one or more layers of the elongated conductor (e.g., to expose the electroactive surface of the working electrode) can be accomplished by hand, excimer laser treatment, chemical etching, laser ablation, grit blasting, or the like.

[0082] The sensor may further include a third layer 414 comprising a conductive material. For example, the third layer 414 may include a reference electrode, which may be formed of a silver-containing material, and is applied over the second layer 404 (i.e., the insulator).

[0083] The elongated conductor 402 may further include one or more intermediate layers (not shown) positioned between the core 410 and the first layer 412. For example, the intermediate layer may be one or more of an insulator, a conductor, a polymer, and / or an adhesive.

[0084] It is contemplated that the ratio of the thickness of the silver / silver chloride layer to the thickness of the insulator (e.g., polyurethane or polyimide) layer can be controlled to allow for certain errors (i.e., errors associated with the etching process) due to defects in the etching process (e.g., defects resulting from the etching process cutting deeper than intended, thereby unintentionally exposing the electroactive surface). This ratio can vary depending on the type of etching process used, whether it is laser ablation, grit blasting, chemical etching, or some other etching method. In one embodiment in which laser ablation is used to remove the silver / silver chloride layer and the polyurethane layer, the ratio of the thickness of the silver / silver chloride layer to the thickness of the polyurethane layer can be from about 1:5 to about 1:1, or from about 1:3 to about 1:2.

[0085] In some embodiments, the core 410 comprises a non-conductive polymer and the first layer 412 comprises a conductive material. Such a sensor configuration advantageously provides reduced material costs in that typically expensive materials are replaced with less expensive materials. For example, the core 410 may be formed of a non-conductive polymer, such as a nylon or polyester filament, string, or cord, which may be coated and / or plated with a conductive material, such as platinum, platinum-iridium, gold, palladium, iridium, graphite, carbon, conductive polymers, and alloys, or combinations thereof.

[0086] As shown in Figures 3C and 3D, the sensor may also include a membrane 408, such as those discussed elsewhere herein, for example, with reference to Figures 2A-2C. The membrane 408 may include an enzyme layer (not shown), as described elsewhere herein. For example, the enzyme layer may include a catalyst or enzyme configured to react with the analyte. For example, the enzyme layer may be an immobilized enzyme layer including glucose oxidase. In other embodiments, the enzyme layer may be impregnated with other oxidases, including, for example, galactose oxidase, cholesterol oxidase, amino acid oxidase, alcohol oxidase, lactate oxidase, or uricase.

[0087] FIG. 3B is a schematic diagram illustrating an embodiment of an elongated conductor 402 or elongated body formed from at least two layers of materials and / or conductive materials, as described in more detail elsewhere herein. The term "electrode" may be used herein to refer to an elongated conductor that includes an electroactive surface that detects an analyte. In some embodiments, the elongated conductor provides an electrical connection between the electroactive surface (i.e., working electrode) and the sensor electronics (not shown). In certain embodiments, each electrode (i.e., the elongated conductor on which the electroactive surface is located) is formed from a thin wire having a diameter of about 0.001 inches or less to about 0.01 inches or more. Each electrode may be formed, for example, from plated insulators, plated wire, or bulk conductive material. For example, in some embodiments, the wire and / or elongated conductor used to form the working electrode is about 0.002, 0.003, 0.004, 0.005, 0.006, 0.007, 0.008, 0.009, 0.01, 0.015, 0.02, 0.025, 0.03, 0.035, 0.04, or 0.045 inches in diameter.

[0088] Additionally, the first layer may include an electroactive surface (i.e., the portion exposed through window 406). The exposed electroactive surface may be a working electrode. For example, if the sensor is an enzymatic electrochemical analyte sensor, the analyte reacts enzymatically with an enzyme within a membrane covering at least a portion of the electroactive surface. This reaction produces electrons (e - For example, in the detection of glucose, where glucose oxidase produces hydrogen peroxide as a by-product, hydrogen peroxide can be released by reacting with two protons (2H + ), two electrons (2e - ), and reacts with the surface of the working electrode producing one oxygen molecule (O2), which generates a current that is detected.

[0089] As described above with reference to FIG. 3A and shown in FIG. 3C , the insulator 404 is disposed on at least a portion of the elongated conductor 402. In some embodiments, the sensor is configured and arranged such that the elongated body includes a core 410 and a first layer 412, with a portion of the first layer 412 exposed through a window 406 in the insulator 404. In other embodiments, the sensor is configured and arranged such that the elongated body 402 includes a core 410 embedded in the insulator 404, with a portion of the core 410 exposed through a window 406 in the insulator 404. For example, the insulating material may be applied to the elongated body 402 (e.g., by screen printing, inkjet printing, and / or block printing) in a configuration designed to leave at least a portion of the surface of the first layer 412 (or the surface of the core 410) exposed. For example, the insulating material may be printed in a pattern that does not cover a portion of the elongated body 402. Alternatively, a portion of the elongated body 402 may be masked prior to application of the insulating material. Removal of the mask after application of the insulating material may expose a portion of elongate body 402 .

[0090] In some embodiments, the insulating material 404 comprises a polymer, such as a non-conductive (i.e., dielectric) polymer. The insulating material can be deposited on the elongate body 402 and / or core 410 using dip coating, spray coating, vapor deposition, printing, and / or other thin and / or thick film coating or deposition techniques. For example, in some embodiments, the insulating material is applied as a layer less than about 5 μm thick, or from about 5, about 10, or about 15 μm to about 20, about 25, about 30, or about 35 μm or thicker. The insulator can be applied as a single layer of material or as two or more layers composed of the same or different materials, as described elsewhere herein. Alternatively, the conductive core does not require a coating of insulator. In some embodiments, the insulating material defines the electroactive surface of the analyte sensor (i.e., working electrode). For example, the surface of the conductive core (e.g., a portion of the first layer 412) can remain exposed during insulator application, or, as described above, a portion of the applied insulator can be removed to expose a portion of the surface of the conductive core.

[0091] In some embodiments in which the sensor has an insulator disposed on an insulated elongate body or conductive structure, a portion of the insulating material can be peeled off or otherwise removed, for example, by hand, excimer laser treatment, chemical etching, laser ablation, grit blasting (e.g., with sodium bicarbonate or other suitable grit), etc., to expose the electroactive surface(s). In one exemplary embodiment, grit blasting is implemented to expose the electroactive surface(s), for example, by utilizing a grit material that is hard enough to cut through polymeric materials but soft enough to minimize or avoid damage to the underlying metal electrodes (e.g., platinum electrodes). Various "grit" materials (e.g., sand, talc, walnut shells, crushed plastic, sea salt, etc.) can be used, and in some embodiments, sodium bicarbonate is an advantageous grit material because it is hard enough to cut through, for example, a parylene coating without damaging, for example, the underlying platinum conductor. Additional benefits of sodium bicarbonate blasting include its abrasive action on metal as it peels off the polymer layer, thereby eliminating a cleaning step that may otherwise be necessary. Alternatively, portions of the electrodes or other electrical conductors may be masked prior to depositing the insulator to maintain exposed electroactive surface area.

[0092] The electroactive surface of the working electrode may be exposed by forming a window 406 in the insulator 404. The electroactive window 406 of the working electrode may be configured to measure the concentration of an analyte.

[0093] In some embodiments, a silver wire is formed on and / or fabricated within the sensor and later chlorinated to form a silver / silver chloride reference electrode. Advantageously, the chlorination of the silver wire described herein allows for the fabrication of a reference electrode with good in vivo performance. By controlling the quantity and amount of silver chlorination to form silver / silver chloride, improved intermittency, stability, and extended life of the reference electrode can be obtained in some embodiments. Furthermore, the use of silver chloride as described above allows for relatively inexpensive and simple fabrication of the reference electrode.

[0094] 3B and 3C, the reference electrode 414 may include a silver-containing material (e.g., silver / silver chloride) applied over at least a portion of the insulating material 404, as discussed in more detail elsewhere herein. For example, the silver-containing material may be applied using thin-film and / or thick-film techniques, such as, but not limited to, dipping, spraying, printing, electrodeposition, vapor deposition, spin coating, and sputter deposition, as described elsewhere herein. For example, a silver- or silver chloride-containing paint (or similar formulation) may be applied to a reel of insulated conductive core. Alternatively, a reel of insulated elongated body (or core) may be cut into single-unit pieces (i.e., "singulated") and a silver-containing ink may be pad-printed thereon. In yet other embodiments, the silver-containing material is applied as a silver foil. For example, an adhesive may be applied to the insulated elongated body, and then a silver foil may be wrapped around it. Alternatively, the sensor is wrapped with Ag / AgCl particles, whereby a sufficient amount of silver is affixed to and / or embedded in and / or otherwise adhered to the adhesive so that the particles function as a reference electrode. In some embodiments, the reference electrode of the sensor comprises a sufficient amount of silver chloride for the sensor to measure and / or detect an analyte for at least 3 days.

[0095] 2A is a cross-sectional view through the sensor showing one embodiment of membrane system 32. In this particular embodiment, the membrane system includes an electrode layer 42, an enzyme layer 44, a diffusion-resistant layer 46, and a biointerface layer 48, all of which are positioned around the working electrode of sensor 38, all of which are described in further detail elsewhere herein. In some embodiments, an integrated diffusion-resistant domain and biointerface layer may be included in the membrane system (e.g., the functionality of both layers is incorporated into one domain). In some embodiments, the sensor is configured for short-term implantation (e.g., about 1-30 days). However, membrane system 32 may be modified for use in other devices, for example, by including only one or more of the domains or additional domains.

[0096] 2B is a cross-sectional view through one embodiment of the sensor showing another embodiment of membrane system 32. In this particular embodiment, the membrane system includes an interference-reducing or blocking layer 43, an enzyme layer 44, a diffusion-resistant layer 46, and a biointerface layer 48 positioned around the working electrode of sensor 38, all of which are described in more detail elsewhere herein.

[0097] 2C is a cross-sectional view through one embodiment of the sensor showing yet another embodiment of the membrane system 32. In this particular embodiment, the membrane system includes an interference-reducing or blocking layer 43, an enzyme layer 44, and an integrated diffusion resistance / biointerface layer 47 positioned around the working electrode of the sensor, all of which are described in more detail elsewhere herein.

[0098] In some embodiments, the membrane system may include a biointerface layer 48 that includes a surface-modified biointerface polymer, which is described in more detail elsewhere herein. However, the sensing membrane 32 of some embodiments may include multiple domains or layers, including, for example, an electrode domain (e.g., as shown in FIG. 2A), an interference reduction or blocking domain (e.g., as shown in FIGS. 2B and 2C), or a cell disruption domain (not shown), which is described in more detail elsewhere herein and in U.S. Patent Publication No. US-2006-0036145-A1, which is incorporated herein by reference in its entirety.

[0099] For example, it is understood that sensing membranes modified for other sensors may include fewer or additional layers. For example, in some embodiments, a membrane system may include one electrode layer, one enzyme layer, and two biointerface layers, while in other embodiments, a membrane system may include one electrode layer, two enzyme layers, and one biointerface layer. In some embodiments, the biointerface layer may be configured to function as a diffusion resistance domain and control the flux of an analyte (e.g., glucose) to the underlying membrane layer.

[0100] In some embodiments, one or more domains of the sensing membrane may be formed from materials such as silicone, polytetrafluoroethylene, polyethylene-co-tetrafluoroethylene, polyolefins, polyesters, polycarbonates, biostable polytetrafluoroethylene, homopolymers, copolymers, terpolymers of polyurethane, polypropylene (PP), polyvinyl chloride (PVC), polyvinylidene fluoride (PVDF), polybutylene terephthalate (PBT), polymethyl methacrylate (PMMA), polyether ether ketone (PEEK), polyurethane, polyurethane urea, cellulose polymers, poly(ethylene oxide), poly(propylene oxide) and copolymers and blends thereof, polysulfone and its block copolymers (including, for example, diblock, triblock, alternating, random, and graft copolymers).

[0101] In some embodiments, the sensing membrane may be deposited on the electroactive surface of the electrode material using known thin or thick film techniques (e.g., spraying, electrodeposition, dipping, etc.) It should be understood that the sensing membrane located on the working electrode need not have the same structure as the sensing membrane located on the reference electrode; for example, an enzyme domain deposited on the working electrode does not necessarily need to be deposited on the reference or counter electrode.

[0102] Although the exemplary embodiment shown in Figures 2A-2C involves a circumferentially extending membrane system, the membranes described herein may be applied to any planar or non-planar surface.

[0103] Sensor Electronics Generally, an analyte sensor system has associated electronics, also referred to as a "computer system," which may include hardware, firmware, or software that enables measurement and processing of data related to analyte levels in a host. In an exemplary embodiment of an electrochemical sensor, the electronics include a potentiostat, a power supply for providing power to the sensor, and other components useful for signal processing. In additional embodiments, some or all of the electronics may communicate with the sensor or other pieces of electronics via wires or wirelessly. For example, a potentiostat disposed on the device may be hardwired to the rest of the electronics (e.g., processor, recorder, transmitter, receiver, etc.) at the bedside. In another example, one piece of electronics is wirelessly connected to another piece of electronics (e.g., receiver), for example, via infrared (IR) or radio frequency (RF). It is contemplated that other embodiments of the electronic device may be useful for providing sensor data outputs such as those described in U.S. Patent Publication No. US-2005-0192557-A1, U.S. Patent Publication No. US-2005-0245795-A1, U.S. Patent Publication No. US-2005-0245795-A1, U.S. Patent Publication No. US-2005-0245795-A1, U.S. Patent Publication No. US-2008-0119703-A1, and U.S. Patent Publication No. US-2008-0108942-A1 (each of which is incorporated by reference in its entirety herein).

[0104] In a preferred embodiment, a potentiostat is operably connected to the electrode(s) (e.g., as described elsewhere herein) and deflects the sensor to allow measurement of a current signal indicative of the analyte concentration (also referred to as the analog portion) in the host. In some embodiments, the potentiostat includes a resistor that converts the current to a voltage. In some alternative embodiments, a current-to-frequency converter configured to continuously integrate the measured current, for example, using a charge counting device, is provided. In some embodiments, the electronics includes an A / D converter that digitizes the analog signal into a digital signal, also referred to as "counts," for processing. Thus, the resulting raw data stream in counts (also referred to as raw sensor data) is directly related to the current measured by the potentiostat.

[0105] Generally, the electronics include a processor module, which includes a central control unit that controls the processing of the sensor system. In some embodiments, the processor module includes a microprocessor, although computer systems other than microprocessors can be used to process the data described herein, e.g., an ASIC can be used for some or all of the sensor's central processing. The processor typically provides semi-permanent storage of data, e.g., storage of data such as sensor identifiers (IDs), and programming for processing data streams (e.g., programming for data smoothing or signal artifact replacement, as described in U.S. Patent Publication No. US-2005-0043598-A1). Additionally, the processor can be used to temporarily store system cache memory, e.g., recent sensor data. In some embodiments, the processor module includes memory storage components, e.g., ROM, RAM, dynamic RAM, static RAM, non-static RAM, EEPROM, rewritable ROM, flash memory, etc.

[0106] In some embodiments, the processor module includes a digital filter, such as an infinite impulse response (IIR) or finite impulse response (FIR) filter, configured to smooth the raw data stream. Generally, the digital filter is programmed to filter data sampled at predetermined time intervals (also referred to as the sampling rate). In some embodiments in which the potentiostat is configured to measure the analyte at discrete time intervals, these time intervals determine the sampling rate of the digital filter. In some alternative embodiments in which the potentiostat is configured to measure the analyte continuously, for example, using a current-to-frequency converter as described above, the processor module can be programmed to request digital values ​​from the A / D converter at predetermined time intervals, also referred to as the acquisition time. In these alternative embodiments, the values ​​obtained by the processor are advantageously averaged over the acquisition time due to the continuity of the current measurements. Therefore, the acquisition time determines the sampling rate of the digital filter.

[0107] In some embodiments, the processor module is configured to construct data packets for transmission to an external source, e.g., RF transmission to a receiver. Generally, the data packets include a preamble unique identifier (e.g., a sensor ID code) that identifies the electronics unit, the receiver, or both, data (e.g., raw data, filtered data, or an integrated value), or multiple bits that may include error detection or correction. Preferably, the data (transmit) packets have a length of about 8 bits to about 128 bits, preferably about 48 bits, although larger or smaller packets may be desirable in certain embodiments. The processor module may be configured to transmit any combination of raw data or filtered data. In one exemplary embodiment, the transmit packet includes a fixed preamble unique ID of the electronics unit, a single 5-minute average (e.g., integrated) sensor data value, and a cyclic redundancy code (CRC).

[0108] In some embodiments, the processor further performs processes such as storing data, analyzing the data stream, calibrating analyte sensor data, estimating analyte values, comparing estimated analyte values ​​with time-corresponding measured analytes, analyzing variations in estimated analyte values, downloading data, and controlling a user interface by providing analyte values, prompts, messages, warnings, alarms, etc. In such cases, the processor includes hardware to perform the processes described herein, e.g., flash memory provides permanent or semi-permanent storage of data, storage of data such as sensor ID, receiver ID, and programming for processing the data stream (e.g., programming for performing the estimation and other algorithms described elsewhere herein), and random access memory (RAM) stores cache memory for the system and aids in data processing. Alternatively, certain portions of the data processing (e.g., described with reference to a processor elsewhere herein) can be accomplished in a separate (e.g., remote) processor, which may be configured for wired or wireless connection thereto.

[0109] In some embodiments, an output module integrated with or operably connected to the processor includes programming for generating an output based on the data stream received from the sensor system and the processing thereof occurring within the processor, hi some embodiments, the output is generated via a user interface.

[0110] Interfering substances Interferents are molecules or other species that can cause a sensor to produce a false-positive or false-negative analyte signal (e.g., a non-analyte-related signal). Some interferents are reduced or oxidized at the electrochemically reactive surface of the sensor, while others interfere with the ability of the enzyme used (e.g., glucose oxidase) to react with the analyte being measured. Still other interferents react with the enzyme (e.g., glucose oxidase) to produce electrochemically active by-products. Interferents can exaggerate or mask the response signal, leading to false or misleading results. For example, a false-positive signal can make a host's analyte concentration (e.g., glucose concentration) appear higher than the true analyte concentration. False-positive signals can pose a clinically significant problem in some conventional sensors. For example, in a severe hypoglycemic situation where a host has ingested an interfering substance (e.g., acetaminophen), the resulting artificially high glucose signal can lead the host to believe they are euglycemic or hyperglycemic. In response, the host may make an inappropriate therapeutic decision by injecting too much insulin or by doing nothing when the appropriate course of action would be to start a meal. In turn, this inappropriate action or inaction can lead to a hypoglycemic episode that is dangerous to the host. Accordingly, certain embodiments contemplated herein include membrane systems that substantially reduce or eliminate the effect of interferents on analyte measurements. These membrane systems may include one or more domains that can block or substantially reduce the flow of interferents onto the electroactive surface of the electrode, which may reduce noise and improve sensor accuracy, as described in more detail in U.S. Patent Publication No. US-2009-0247856-A1.

[0111] Drift As used herein, the term "drift" is a broad term, given its ordinary and customary meaning to those skilled in the art (and is not limited to any special or customized meaning), and refers to, but is not limited to, a change in the sensitivity of a sensor over time. Drift can be driven by changes in the permeability of the sensor membrane system, which can be particularly evident in embodiments using polyurethane diffusion-resistant domains. Without being bound by theory, it is believed that permeability changes in such systems result from rearrangements of the diffusion-resistant domain polyurethane polymer chains that bring more hydrophilic components to the surface or otherwise allow better access to the hydrophilic polymer components during hydration of the membrane system. Thus, increasing the hydration rate or increasing the wettability of the membrane system reduces system drift.

[0112] Due to electrostatically induced hydration, polymeric and crosslinked coatings of zwitterionic compounds have near-instantaneous wettability. As discussed in more detail below, including one or more zwitterionic compounds, their precursors, or derivatives (hydrolyzable cationic esters) in the outermost domain of a membrane system, or applying a coating of such compounds to the surface of a membrane system, results in reduced sensor drift.

[0113] membrane manufacturing The polymers of preferred embodiments can be processed by solution-based techniques such as spraying, dipping, casting, electrospinning, vapor deposition, spin-coating, and coating. Water-based polymer emulsions can be made to form films by methods similar to those used for solvent-based materials. In both cases, evaporation of the volatile liquid (e.g., organic solvent or water) leaves a film of polymer. Crosslinking of the deposited film or layer can be achieved by many methods, including the use of multifunctional reactive components. Liquid systems can be cured by heat, moisture, high-energy radiation, ultraviolet light, or by completing a reaction to produce the final polymer in the mold or on the substrate to be coated.

[0114] In some embodiments, the wettability of the membrane (and thereby the degree of sensor drift exhibited by the sensor) can be tuned and / or controlled by creating covalent crosslinks between surface-active group-containing polymers, functional group-containing polymers, polymers with zwitterionic groups (or precursors or derivatives thereof), and combinations thereof. Crosslinking can have a substantial effect on film structure, which in turn can affect the wettability of the film surface. Crosslinking can also affect film tensile strength, mechanical strength, water absorption rate, and other properties.

[0115] The crosslinked polymers may have different crosslink densities. In certain embodiments, crosslinking is used to promote crosslinking between layers. In other embodiments, heat is used to form crosslinks instead of (or in addition to) the crosslinking techniques described above. For example, in some embodiments, imide and amide bonds may form between two polymers as a result of elevated temperatures. In some embodiments, photocrosslinking is performed to form covalent bonds between the polycation layer(s) and the polyanion layer(s). One major advantage over photocrosslinking is that it offers the possibility of patterning. In certain embodiments, patterning using photocrosslinking is performed to modify the film structure and thus tune the wettability of the membrane.

[0116] Polymers having domains or segments functionalized to allow crosslinking can be prepared by methods known in the art. For example, polyurethaneurea polymers having aromatic or aliphatic segments with electrophilic functional groups (e.g., carbonyl, aldehyde, anhydride, ester, amide, isocyano, epoxy, allyl, or halo groups) can be crosslinked with crosslinkers having multiple nucleophilic groups (e.g., hydroxyl, amine, urea, urethane, or thio groups). In a further embodiment, polyurethaneurea polymers having aromatic or aliphatic segments with nucleophilic functional groups can be crosslinked with crosslinkers having multiple electrophilic groups. Furthermore, polyurethaneurea polymers having hydrophilic segments with nucleophilic or electrophilic functional groups can be crosslinked with crosslinkers having multiple electrophilic or nucleophilic groups. The unsaturated functional groups on the polyurethaneurea can also be used for crosslinking by reacting with a multivalent free radical agent. Non-limiting examples of suitable crosslinkers include isocyanates, carbodiimides, glutaraldehyde, aziridines, silanes or other aldehydes, epoxies, acrylates, free radical agents, ethylene glycol diglycidyl ether (EGDE), poly(ethylene glycol) diglycidyl ether (PEGDE), or dicumyl peroxide (DCP). In one embodiment, about 0.1% to about 15% w / w of crosslinker is added based on the total dry weight of the crosslinker and polymer when blending the components (in one example, about 1% to about 10%). During the curing process, substantially all of the crosslinker is believed to react, leaving substantially no detectable unreacted crosslinker in the final film.

[0117] The polymers disclosed herein can be formulated into a mixture that can be drawn into a film or applied to a surface using any method known in the art (e.g., spraying, painting, dip coating, vapor deposition, molding, 3D printing, lithographic techniques (e.g., photolithography), micro- and nano-pipetting printing techniques, silkscreen printing, etc.). The mixture can then be cured at elevated temperatures (e.g., 50-150°C). Other suitable curing methods can include, for example, ultraviolet light or gamma irradiation.

[0118] Biointerface Domain The biointerface layer is a domain or layer of an implantable device configured to interface with (i.e., contact) biological fluids when implanted in or connected to a host (e.g., via an intravascular access device providing intracorporeal access to blood vessels). When present on an analyte sensor, e.g., a continuous analyte sensor implanted in a host, the biointerface layer can increase sensor lifetime and reduce sensor inaccuracy by reducing biomaterial-associated inflammatory responses. The antifouling properties of the biointerface layer can inhibit the accumulation of cells, proteins, and other biological species on the sensor. In some embodiments, the biointerface domain may be formed from the biointerface domain described in U.S. Provisional Application No. 62 / 273,142, filed December 20, 2015, which is incorporated herein by reference in its entirety.

[0119] The biointerface layers disclosed herein are mechanically robust, preventing damage during implantation and resistant to degradation during sensor implantation. Furthermore, the disclosed biointerface layers do not affect the sensor's response time or the properties of the diffusion-resistant layer. The disclosed biointerface layers can also be hydrophilic, capable of high water absorption, fast water absorption, and rapid stabilization, thereby not adversely affecting sensor start-up. The disclosed biointerface layers are also permeable to analytes (e.g., glucose) but prevent protein adsorption.

[0120] Some embodiments described herein may include a membrane that includes a biointerface layer 48 (see Figures 2A-2C).

[0121] Additionally, the disclosed biointerface layers can host pharmaceutical or bioactive agents that, upon release from the biointerface layer into local tissue, can effectively reduce or delay inflammation. The anti-inflammatory agents can be steroidal or non-steroidal drugs and can be scavengers of reactive oxygen species (ROS). Suitable anti-inflammatory agents include, but are not limited to, nonsteroidal anti-inflammatory drugs (NSAIDS), such as acetomethaphen, aminosalicylic acid, aspirin, celecoxib, choline magnesium trisalicylate, diclofenac potassium, diclofenac sodium, diflunisal, etodolac, fenoprofen, flurbiprofen, ibuprofen, indomethacin, interleukin (IL)-10, IL-6 muteins, anti-IL-6 iNOS inhibitors (e.g., L-NAME or L-NMDA), interferons, ketoprofen, ketorolac, leflunomide, melenamic acid, mycophenolic acid, mizoribine, nabumetone, naproxen, naproxen sodium, oxaprozin, Piroxicam, rofecoxib, salsalate, sulindac, and trimethine; and corticosteroids such as cortisone, hydrocortisone, methylprednisolone, prednisone, prednisolone, betamethasone, beclomethasone dipropionate, budesonide, dexamethasone sodium phosphate, flunisolide, fluticasone propionate, paclitaxel, tacrolimus, tranilast, triamcinolone acetonide, betamethasone, fluocinolone, betamethasone dipropionate, betamethasone valerate, desonide, desoximetasone, fluocinolone, triamcinolone, triamcinolone acetonide, clobetasol propionate, and dexamethasone.

[0122] In some embodiments, the biointerface layer may include a polymer described as a bioprotective layer in U.S. Patent Publication No. 2014-0094671, which is incorporated by reference herein for its teachings regarding at least bioprotective layers in sensor membranes.

[0123] In other embodiments, the biointerface layer can include a biointerface polymer. The biointerface polymer is a polyzwitterion. A polyzwitterion is a polymer in which the repeat units of the polymer chain are constituent ionic moieties. As such, these polymers have equal numbers of cationic and anionic groups, with each zwitterionic repeat unit having both positive and negative charges, and therefore often having an overall zero charge over a wide pH range.

[0124] Polyzwitterions are distinguished from other polyampholytes in that they contain anionic and cationic groups, but the ionic groups are not correlated with each other as part of the same repeating unit. Thus, the anionic and cationic groups may be randomly spaced apart from each other, or there may be more of one ionic group than the other. Thus, polyampholytes typically have a net charge, except perhaps over a somewhat narrow pH range.

[0125] The disclosed polyzwitterions can have a variety of repeating units, shown below as i-vii), where n is an integer between 2 and 1000. [ka]

[0126] In structures i-iv), the zwitterionic units are attached to the backbone (~~~~), with the charges on side groups pendant to the chain. In structures v)-vii), the zwitterionic units are such that one or both charges are on the chain itself.

[0127] Examples of suitable zwitterionic monomers that can be used to generate polyzwitterions of any of structures i)-vii) include: Included are ammoniophosphates (phosphobetaines or lecithin analogs), ammoniophosphonates (phosphonobetaines), or ammoniophosphinates (phosphinobetaines), each having the structure: [ka] wherein Z is a branched or straight chain alkyl, heteroalkyl, cycloalkyl, cycloheteroalkyl, aryl, or heteroaryl; R 1 is H, alkyl, heteroalkyl, cycloalkyl, heterocycloalkyl, aryl, or heteroaryl, and R 2 , R 3 , and R 4 is independently selected from alkyl, heteroalkyl, cycloalkyl, heterocycloalkyl, aryl, or heteroaryl; R 1 , R 2 , R 3 , R 4 and one or more of Z is substituted with a polymerizable group.

[0128] "Polymerization group" refers to a functional group that allows a monomer to polymerize with itself to form a homopolymer or with a different monomer to form a copolymer. Depending on the type of polymerization method used, the polymerization group may be selected from alkenes, alkynes, epoxides, lactones, amines, hydroxyls, isocyanates, carboxylic acids, anhydrides, silanes, halides, aldehydes, and carbodiimides. In step-growth polymerizations, matching pairs of functional groups may be selected to facilitate polymerization, for example, to polymerize dihydroxyl groups bearing zwitterionic monomers, and comonomers containing diisocyanate, epoxide, or dicarboxylic acid groups may be selected to provide polymers formed with urethane, ether, and ester linkages.

[0129] Further examples of suitable zwitterionic monomers that can be used to generate polyzwitterions of any of structures i)-vii) include ammoniosulfonates (sulfobetaines), ammoniosulfates, each having the following structure: [ka] wherein Z is a branched or straight chain alkyl, heteroalkyl, cycloalkyl, cycloheteroalkyl, aryl, or heteroaryl; R 1 is H, alkyl, heteroalkyl, cycloalkyl, heterocycloalkyl, aryl, or heteroaryl, and R 2 and R 3 is independently selected from alkyl, heteroalkyl, cycloalkyl, heterocycloalkyl, aryl, or heteroaryl; R 1 , R 2 , R 3 and one or more of Z is substituted with a polymerizable group; Ammoniocarboxylate has the following structure: [ka] wherein Z is a branched or straight chain alkyl, heteroalkyl, cycloalkyl, cycloheteroalkyl, aryl, or heteroaryl; R 1 is H, alkyl, heteroalkyl, cycloalkyl, heterocycloalkyl, aryl, or heteroaryl, and R 2 and R 3 is independently selected from alkyl, heteroalkyl, cycloalkyl, heterocycloalkyl, aryl, or heteroaryl; R 1 , R 2 , R 3 and one or more of Z is substituted with a polymerizable group.

[0130] In each of these monomers, Z can have a length of 1 to 12 atoms, e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 atoms, and any of these values ​​can form the upper or lower endpoint of a range.

[0131] These monomers can be prepared by methods known to those skilled in the art, for example, as detailed in Laschewsky, "Structures and synthesis of zwitterionic polymers," Polymers 6:1544-1601, 2014. In certain examples, the disclosed polyzwitterions can have repeating zwitterionic units derived from any of the zwitterionic monomers disclosed above.

[0132] The biointerface polymer may also contain polyurethane and / or polyurea segments. For example, the biointerface polymer may include polyurethane copolymers, such as polyether-urethane-urea, polycarbonate-urethane, polyether-urethane, silicone-polyether-urethane, silicone-polycarbonate-urethane, polyester-urethane, polyurethane-urea, and the like. These polyurethane and / or polyurea segments are referred to herein as "hard segments" because they contain urea and / or urethane linkages formed from polyisocyanates and short-chain polyols or polyamines, moieties that are rich in hydrogen bonds. These segments may also be relatively hydrophobic.

[0133] In addition to the polyurethane and / or polyurea hard segments, the disclosed biointerface polymers may also contain soft segments that have relatively poor hydrogen bonding. The soft segments are typically composed of polyols such as polycarbonates, polyesters, polyethers, polyarylenes, and polyalkylenes. The soft segments can be either hydrophobic or hydrophilic.

[0134] Biointerface polymers useful in certain embodiments may comprise linear or branched polymers on the backbone structure of the polymer. Thus, either the hard or soft segments may comprise a branched or linear backbone.

[0135] The zwitterionic monomers, as described herein, can be part of either the hard or soft segments, or both.

[0136] In some embodiments, the hard segment portion of the biointerface polymer may comprise about 5% to about 50% by weight of the polymer, sometimes about 15% to 20% by weight, and sometimes about 25% to 40% by weight. The hard segment may have a molecular weight of about 160 daltons to about 10,000 daltons, and sometimes about 200 daltons to about 2,000 daltons. In some embodiments, the molecular weight of the soft segment may be about 200 daltons to about 10,000,000 daltons, sometimes about 500 daltons to about 5,000 daltons, and sometimes about 500 daltons to about 2,000 daltons.

[0137] As noted, the hard segments can be polyurethanes or polyureas. Polyurethanes are polymers produced by the condensation reaction of diisocyanates and difunctional hydroxyl-containing materials. Polyureas are polymers produced by the condensation reaction of diisocyanates and difunctional amine-containing materials. Preferred diisocyanates include aliphatic diisocyanates containing from about 4 to about 9 methylene units. Cycloaliphatic diisocyanates containing diisocyanates can also be useful in preparing the polymer and copolymer components of the membranes of preferred embodiments.

[0138] The soft segments used in preparing the biointerface polymer may be polyfunctional aliphatic polyols, polyfunctional aliphatic or aromatic amines, etc., which may be useful in effecting permeability of an analyte (e.g., glucose) therethrough, and may include, for example, polyoxazoline, poly(ethylene glycol) (PEG), polyacrylamide, polyimine, polypropylene oxide (PPO), PEG-co-PPO diol, silicone-co-PEG diol, silicone-co-PPO diol, polyethyl acrylate (PEA), polyvinylpyrrolidone (PVP), and modifications thereof (e.g., PVP vinyl acetate), where PEG and modifications thereof may be preferred due to their hydrophilicity.

[0139] In some embodiments, the soft segment portion of the biointerface polymer may comprise about 5% to about 50% by weight, sometimes about 15% to 20% by weight, and other times about 25% to 40% by weight of the polymer. The soft segment may have a molecular weight of about 160 daltons to about 10,000 daltons, and sometimes about 200 daltons to about 2,000 daltons. In some embodiments, the molecular weight of the soft segment may be about 200 daltons to about 10,000,000 daltons, sometimes about 500 daltons to about 5,000 daltons, and sometimes about 500 daltons to about 2,000 daltons.

[0140] In some embodiments, biointerface polymers comprising hard and soft segments and zwitterionic repeat units can have a molecular weight of about 10 kDa to about 500,000 kDa, e.g., about 10 kDa to about 100,000 kDa, about 1000 kDa to about 500,000 kDa, about 10,000 kDa to about 100,000 kDa, and about 100,000 kDa to about 500,000 kDa.

[0141] The hard and soft segments may each be selected for their properties, including but not limited to, tensile strength, flex life, modulus, etc. For example, polyurethanes are relatively strong and offer many reaction pathways, properties that may be advantageous as bulk properties for the membrane domain of a continuous sensor.

[0142] As noted above, biointerface polymers contain one or more zwitterionic repeat units; therefore, these groups are "internal" with respect to the polymer backbone. Such "internal" repeat units are distinguished from materials found at the ends of polymer chains, since such moieties are only attached to the polymer chain at one position. The disclosed biointerface polymers, in some embodiments, may have one or more zwitterionic groups at the ends of the polymer chain, but such groups are not the only zwitterionic groups in the chain; there is at least one internal zwitterionic group in the backbone.

[0143] In some preferred embodiments, zwitterionic moieties are selected for desirable properties, such as, for example, transient noise blocking ability, (reduced) intermittency, ability to repel charged species, cationic or ionic blocking, surface wettability, antifouling properties, etc. In some embodiments, the zwitterions or zwitterionic precursors are present as zwitterionic groups while the device is in vivo. As such, these groups present a mixed charge area on the device surface to the surrounding environment, thereby increasing surface hydration of the device and potentially reducing nonspecific protein adsorption and cell adhesion.

[0144] In some embodiments, the biointerface polymer comprises at least about 1%, about 2%, about 3%, about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, about 10%, about 11%, about 12%, about 13%, about 14%, about 15%, about 16%, about 17%, about 18%, about 19%, about 20%, about 21%, about 22%, about 23%, about 24%, about 25%, about 26%, about 27%, about 28%, about 30%, about 31%, about 32%, about 33%, about 34%, about 35%, about 36%, about 37%, about 38%, about 39%, about 40%, about 41%, about 42%, about 43%, about 44%, about 45%, about 46%, about 47%, about 48%, about 49%, about 50%, about 51%, about 52%, about 53%, about 54%, about 55%, about 56%, about 57%, about 58%, about 59%, about 60%, about 61%, about 62%, about 63%, about 64%, about 65%, about 66%, about 67%, about 68%, about 69%, about 70%, about 71%, about 72%, about 73%, about 74%, about 75%, about 76%, about 77%, about 78%, about 79%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92% by weight of the polymer. %, about 28%, about 29%, about 30% to about 31%, about 32%, about 33%, about 34%, about 35%, about 36%, about 37%, about 38%, about 39%, about 40%, about 41%, about 42%, about 43%, about 44%, about 45%, about 46%, about 47%, about 48%, about 49%, about 50%, about 51%, about 52%, about 53%, about 54%, or about 55% by weight of zwitterionic repeat units.

[0145] The zwitterionic repeat units can be betaines, precursors or derivatives thereof (e.g., alkylbetaines or aminobetaines), such as carboxyl, sulfo, or phosphobetaine compounds. These segments or moieties can be incorporated into the biointerface polymer in the hard segments, soft segments, or both, for example, up to about 55% by weight of the biointerface polymer.

[0146] In some embodiments, two or more different zwitterionic or zwitterionic precursor segments or moieties are used, while in other embodiments, a single zwitterionic or zwitterionic precursor segment or moiety may be used in the biointerface polymer.

[0147] Some examples of biointerface polymers are illustrated schematically in Figure 10. Generally, biointerface polymers include one or more hard segments and one or more soft segments. The hard segments can be aliphatic or aromatic monomers. The soft segments can be hydrophilic or hydrophobic oligomers, such as polyalkylene glycols, polycarbonates, polyesters, polyethers, polyvinyl alcohols, polyvinylpyrrolidones, and polyoxazolines. Zwitterionic groups (e.g., betaines) can be part of the soft segments, the hard segments, or both. As shown in Figure 10, various hard and soft segments can be present, allowing the properties of the biointerface polymer to be tailored by using different segments, different segment lengths, functionalization on certain segments, crosslinking certain segments, etc. In some embodiments, biocompatible segmented block polyurethane copolymers containing hard and soft segments can be used in the biointerface layer.

[0148] Incorporation of these zwitterionic repeat units into polymers can be achieved by using zwitterionic monomers bearing diols or diamines (e.g., at the Z position), or R 1 ~R 4 R 1 ~R 4 Attachment to a diol or diamine in can be achieved by reacting the corresponding precursor with a halo-substituted diamine or diol. Examples of such monomers are shown below: [ka] wherein W, Y, and Z are independently branched or straight chain alkyl, heteroalkyl, cycloalkyl, cycloheteroalkyl, aryl, or heteroaryl, any of which may be optionally substituted with O, OH, halogen, amino, or alkoxyl; R 1is H, alkyl, heteroalkyl, cycloalkyl, cycloheteroalkyl, aryl, or heteroaryl, and R 2 , R 3 , and R 4 is independently selected from alkyl, heteroalkyl, cycloalkyl, cycloheteroalkyl, aryl, or heteroaryl. In certain embodiments, W is C1-C4 alkyl. In certain embodiments, Y is C1-C4 alkyl. In other embodiments, Z is C1-C4 alkyl.

[0149] These compounds can be reacted with diisocyanates to form polyurethanes or polyureas. Alternatively, the carboxylate, sulfonate, phosphinate, or phosphonate moieties can be protected, and then the protecting groups can be removed after polymerization. In another alternative, the amines can be tertiary amines, which are then quaternized by alkylation after polymerization.

[0150] Another method involves the radical polymerization of zwitterionic monomers having unsaturated moieties substituted at the Z position in the monomers shown above. In another example, zwitterionic monomers in which the unsaturated moiety is attached to an ammonium group can be used in the radical polymerization. Examples of such monomers are shown below: [ka] wherein X is O, NH, or NR 4 Y and Z are independently branched or straight chain alkyl, heteroalkyl, cycloalkyl, cycloheteroalkyl, aryl, or heteroaryl, any of which may be optionally substituted with OH, halogen, or alkoxyl; R 1 is H, alkyl, heteroalkyl, cycloalkyl, cycloheteroalkyl, aryl, or heteroaryl, and R 3 and R 5 is independently selected from heteroalkyl, cycloalkyl, cycloheteroalkyl, aryl, or heteroaryl.5 is H or CH3. In other embodiments, X is O. In still other embodiments, X is NH or NCH3. In certain embodiments, Y is C1-C4 alkyl. In other embodiments, Z is C1-C4 alkyl.

[0151] Additional examples of suitable zwitterionic monomers include N-(2-methacryloyloxy)ethyl-N,N-dimethylammoniopropanesulfonate, N-(3-methacryloylimino)propyl-N,N-dimethylammoniopropanesulfonate, 2-(methacryloyloxy)ethylphosphatidylcholine, and 3-(2′-vinyl-pyridino)propanesulfonate.

[0152] In other embodiments, the biointerface polymer is crosslinked. For example, polyurethaneurea polymers having aromatic or aliphatic segments with electrophilic functional groups (e.g., carbonyl, aldehyde, anhydride, ester, amide, isocyanate, epoxy, allyl, or halo groups) can be crosslinked with crosslinkers having multiple nucleophilic groups (e.g., hydroxyl, amine, urea, urethane, or thio groups). In further embodiments, polyurethaneurea polymers having aromatic or aliphatic segments with nucleophilic functional groups can be crosslinked with crosslinkers having multiple electrophilic groups. Still further, polyurethaneurea polymers having hydrophilic segments with nucleophilic or electrophilic functional groups can be crosslinked with crosslinkers having multiple electrophilic or nucleophilic groups. The unsaturated functional groups on the polyurethaneurea can also be used for crosslinking by reacting with multivalent free radical agents.

[0153] Non-limiting examples of suitable crosslinkers include isocyanates, carbodiimides, glutaraldehyde or other aldehydes, aziridines, silanes, epoxies, acrylates, free radical agents, ethylene glycol diglycidyl ether (EGDE), poly(ethylene glycol) diglycidyl ether (PEGDE), or dicumyl peroxide (DCP). In one embodiment, about 0.1% to about 15% w / w of crosslinker is added based on the total dry weight of the crosslinker and polymer when blending the components (in one example, about 1% to about 10%). During the curing process, substantially all of the crosslinker is believed to react, leaving substantially no detectable unreacted crosslinker in the final layer.

[0154] Additionally, the disclosed biointerface layers can have zwitterions entrapped or embedded within the polymer network through non-covalent interactions. Thus, in further embodiments, the disclosed biointerface layers can include a biointerface polymer and an additional betaine blended therewith. For example, the biointerface polymer can be blended with 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). It will be understood that many more zwitterionic compounds or precursors or derivatives thereof may be applicable, and that this list of exemplary betaines is not intended to limit the scope of the embodiments.

[0155] The biointerface layer may further include a domain comprising a surface-modifying polymer attached to a base polymer, where the surface-modifying polymer comprises a polymer chain having both hydrophilic and hydrophobic regions, and where one or more zwitterionic compounds are covalently attached to an interior region of the polymer, and the base polymer may be selected from silicone, epoxy, polyolefin, polystyrene, polyoxymethylene, polysiloxane, polyether, polyacrylic, polymethacrylic, polyester, polycarbonate, polyamide, poly(etherketone), poly(etherimide), polyurethane, and polyurethaneurea.

[0156] In some embodiments, the biointerface layer may comprise a combination of one or more biointerface polymer(s), e.g., polyurethane or polyurethaneurea, and one or more hydrophilic polymers, e.g., PVA, PEG, polyacrylamide, polyacetate, polyzwitterion, PEO, PEA, PVP, and variations thereof (e.g., PVP vinyl acetate), e.g., as a physical blend or admixture, wherein each polymer maintains its unique chemical properties.

[0157] In some embodiments, the biointerface layer 48 is positioned most distal to the sensing region, such that its outermost domain contacts biological fluids when inserted in vivo. In some embodiments, the biointerface layer may be composed of a material that is resistant to cell attachment, impermeable to cells, and biostable. Without being bound by theory, if the biointerface domain 48 is resistant to cell attachment (e.g., attachment by inflammatory cells such as macrophages, and thus maintained at a sufficient distance from other domains, e.g., enzyme domains), hypochlorite and other oxidizing species are short-lived species in vivo, and biodegradation generally does not occur. Furthermore, preferred materials for forming the biointerface domain 48 are referred to as biodurable because they can resist the effects of these oxidizing species. In some embodiments, the biointerface domain controls the flux of oxygen and other analytes (e.g., glucose) to the underlying enzyme domain (e.g., the functionality of a diffusion resistance domain is incorporated into the biointerface domain, thereby eliminating the need for a separate diffusion resistance domain).

[0158] In some embodiments, the one or more zwitterionic compounds or precursors thereof applied to the surface of the membrane system are hydrolyzable cationic esters of zwitterionic compounds. In these embodiments, the hydrolyzable cationic esters provide the added benefit that hydrolysis of the cationic esters to non-fouling zwitterionic groups can kill microorganisms (such as bacteria) or condense DNA. Furthermore, the mixed-charge nature of the resulting zwitterionic groups results in the inhibition of nonspecific protein adsorption on the sensor surface. In these embodiments, cationic betaine esters, such as cationic pCB esters, are preferred.

[0159] In certain embodiments, the biointerface polymer may contain reactive groups that may be used for further functionalization. For example, unsaturated functional groups such as alkynes can be used to attach various moieties attached to dipolar groups such as azides to form covalent bonds. Such Huisgen cycloaddition chemistry is often referred to as click chemistry. Thus, in certain embodiments herein, the biointerface layer may contain alkyne functional groups pendant to the polymer backbone. Antiseptics, cytokines, anti-inflammatory agents, steroids, and other bioactive agents disclosed herein, such as proteins, attached to dipolar groups such as azides, can be conveniently attached to the polymer, resulting in triazole groups. Thus, biointerface layers, sensors including such layers containing alkynes, triazoles, or both, are disclosed herein. These reactive groups may be present in the zwitterionic repeat unit (e.g., as substituents on Z or Y).

[0160] Incorporating zwitterionic or zwitterionic precursor segments or moieties into the backbone of a polymer can be difficult due to solubility issues associated with the zwitterionic or zwitterionic precursor monomer. Such groups are typically soluble only in highly polar solvents, such as methanol and water, which are unfavorable for the synthesis of some biointerface polymers (e.g., polyurethanes). Therefore, the available functional groups that can be chemically incorporated into the backbone of biointerface polymers via solution-based polycondensation synthesis are limited. As an alternative method for incorporating zwitterionic or zwitterionic precursor segments or moieties into the backbone of a base polymer, precursors or derivatives of the zwitterionic or zwitterionic precursor can be used. For example, zwitterionic precursors and / or zwitterionic derivatives with more desirable solubility characteristics in low-polarity organic solvents can be used as monomers. Biointerface polymers (e.g., polyurethane-ureas) can be synthesized via polycondensation reactions to form well-defined polymers with high molecular weights and low polydispersity indices. These polymers can then be converted to zwitterionic group containing polymers via chemical reactions (hydrolysis, deprotection, thermally induced rearrangement, and UV-induced degradation) or biologically induced reactions after in vivo implantation of the device.

[0161] In certain embodiments, the thickness of the biointerface domain can be about 0.1, about 0.5, about 1, about 2, about 4, about 6, or about 8 μm or less to about 10, about 15, about 20, about 30, about 40, about 50, about 75, about 100, about 125, about 150, about 175, about 200, or about 250 μm or more. In some of these embodiments, the thickness of the biointerface domain can sometimes be about 1 to about 5 μm, and sometimes about 2 to about 7 μm. In other embodiments, the biointerface domain can be about 20 or about 25 μm to about 50, about 55, or about 60 μm thick. In some embodiments, the glucose sensor can be configured for transdermal or short-term subcutaneous implantation and can have a thickness of about 0.5 μm to about 8 μm, and sometimes about 4 μm to about 6 μm. In one glucose sensor configured for fluid communication with a host's circulatory system, the thickness can be about 1.5 μm to about 25 μm, and sometimes about 3 to about 15 μm. In some embodiments, the biointerface layer or any other layer of the electrode can have a consistent thickness, although it is contemplated that in other embodiments, the thickness can vary. For example, in some embodiments, the thickness of the biointerface layer can vary along the longitudinal axis of the electrode end.

[0162] The biointerface layer can be hydrophilic as measured by contact angle, for example, the biointerface layer can have a contact angle of about 20° to about 90°, about 60° to about 90°, about 70° to about 90°, about 80° to about 90°, about 60° to about 80°, at least about 50°, at least about 60°, or at least about 70°.

[0163] The biointerface layer may also have a low polydispersity index. For example, the polymer may have a polydispersity index of about 1.4 to about 3.5, about 1.75 to about 2.25, about 1.75 to about 2.5, or about 2.

[0164] The biointerface layer also cannot substantially affect the T95 response time of the sensor. For example, a sensor having a biointerface layer disclosed herein may have a T95 response time that is the same as or within 5% of the T95 response time of an otherwise identical sensor without the biointerface layer.

[0165] Diffusion Resistance Domain In some embodiments, a diffusion resistance domain 46, also referred to as a diffusion resistance layer, can be used and is positioned more proximal to the implantable device relative to the biointerface layer. In some embodiments, the functionality of the diffusion resistance domain can be incorporated into a biointerface layer comprising a polyzwitterionic biointerface polymer. Therefore, it should be noted that the description herein regarding the diffusion resistance domain can also apply to the biointerface layer. The diffusion resistance domain functions to control the flux of oxygen and other analytes (e.g., glucose) to the underlying enzyme domain. As described in more detail elsewhere herein, there is a molar excess of glucose relative to the amount of oxygen in blood, i.e., there are typically more than 100 glucose molecules for every free oxygen molecule in the extracellular fluid (see Updike et al., Diabetes Care 5:207-21 (1982)). However, immobilized enzyme-based sensors that use oxygen as a cofactor are supplied with oxygen in a non-rate-limiting excess so that they respond linearly to changes in glucose concentration but are unresponsive to changes in oxygen partial pressure. More specifically, when the glucose monitoring response is oxygen-limited, linearity is not achieved at glucose concentrations above a minimum. Without a semipermeable membrane located over the enzyme domain to control the flux of glucose and oxygen, a linear response to glucose levels can be obtained at only up to about 40 mg / dL. However, in a clinical setting, a linear response to glucose levels of at least up to about 500 mg / dL is desirable. In some embodiments, the diffusion resistance domain can be formed with the diffusion resistance domain described in U.S. Provisional Application No. 62 / 273,219, filed December 20, 2015, which is incorporated herein by reference in its entirety.

[0166] The diffusion resistance domain 46 comprises a semipermeable membrane that controls the flux of oxygen and glucose to the underlying enzyme domain 42, preferably providing a non-rate-limiting excess of oxygen. As a result, the upper linearity limit of glucose measurement is extended much higher than would be achieved without the diffusion resistance domain. In some embodiments, the diffusion resistance domain exhibits an oxygen-to-glucose permeability ratio of approximately 200:1, while in other embodiments, the oxygen-to-glucose permeability ratio may be approximately 100:1, 125:1, 130:1, 135:1, 150:1, 175:1, 225:1, 250:1, 275:1, 300:1, or 500:1. As a result of the high oxygen-to-glucose permeability ratio, one-dimensional reactant diffusion can provide sufficient excess oxygen at all reasonable glucose and oxygen concentrations found in the subcutaneous matrix (see Rhodes et al., Anal. Chem., 66:1520-1529 (1994)). In some embodiments, a lower oxygen-to-glucose ratio may be sufficient to provide excess oxygen by using a highly oxygen-soluble domain (e.g., a silicone material) to enhance oxygen supply / transport to the enzyme membrane or electroactive surface. By enhancing oxygen supply through the use of a silicone composition, for example, glucose concentration is less of a limiting factor. In other words, if more oxygen is supplied to the enzyme or electroactive surface, then more glucose can also be supplied to the enzyme without resulting in an oxygen rate-limiting excess.

[0167] In some embodiments, the diffusion resistance domain is formed from a base polymer synthesized to include a polyurethane membrane having both hydrophilic and hydrophobic regions to control the diffusion of glucose and oxygen to the analyte sensor. A suitable hydrophobic polymer component can be polyurethane or polyetherurethaneurea. Polyurethanes are polymers produced by the condensation reaction of diisocyanates and difunctional hydroxyl-containing materials. Polyureas are polymers produced by the condensation reaction of diisocyanates and difunctional amine-containing materials. Preferred diisocyanates include aliphatic diisocyanates containing about 4 to about 8 methylene units. Cycloaliphatic diisocyanates may also be useful in preparing the polymer and copolymer components of the membranes of preferred embodiments. The material forming the basis of the hydrophobic matrix of the diffusion resistance domain can be any of those known in the art to be suitable for use as a membrane in a sensor device, having sufficient permeability to allow relevant compounds to pass therethrough, for example, to allow oxygen molecules to pass from the sample under test through the membrane to reach an active enzyme or electrochemical electrode. Examples of materials that can be used to make non-polyurethane type membranes include vinyl polymers, polyethers, polyesters, polyamides, inorganic polymers (such as polysiloxanes and polycarbosiloxanes), natural polymers (such as cellulose and protein-based materials), and copolymers, mixtures, or combinations thereof.

[0168] In one embodiment of the polyurethane-based resistance domain, the hydrophilic soft segment polymer component can be polyethylene oxide. For example, one useful hydrophilic copolymer component is a polyurethane polymer containing about 20% hydrophilic polyethylene oxide. The polyethylene oxide portion of the copolymer is thermodynamically driven to separate from the hydrophobic portion and hydrophobic polymer component of the copolymer. The 20% polyethylene oxide-based soft segment portion of the copolymer used to form the final blend affects the water absorption and subsequent glucose permeability of the membrane.

[0169] Alternatively, in some embodiments, the diffusion-resistant domain may comprise a combination of a base polymer (e.g., polyurethane) and one or more hydrophilic polymers (e.g., PVA, PEG, polyacrylamide, acetate, PEO, PEA, PVP, and copolymers, blends, and / or modifications thereof). Any of a variety of polymer combinations may be used to provide the desired blend of glucose, oxygen, and interference permeability properties. For example, in some embodiments, the diffusion-resistant domain may be formed from a blend of a silicone polycarbonate-urethane base polymer and a PVP hydrophilic polymer, while in other embodiments, a blend of polyurethane, or another base polymer, and one or more hydrophilic polymers may be used instead. In some embodiments involving the use of PVP, the PVP portion of the polymer blend may comprise about 5% to about 50% by weight of the polymer blend, sometimes about 15% to about 20% by weight, and other times about 25% to about 40% by weight. It is contemplated that PVP of various molecular weights may be used. For example, in some embodiments, the molecular weight of the PVP used can be from about 25,000 daltons to about 5,000,000 daltons, sometimes from about 50,000 daltons to about 2,000,000 daltons, and other times from about 6,000,000 daltons to about 10,000,000 daltons.

[0170] In some embodiments, the diffusion resistance domain 46 may be formed as an integral structure with the biointerface domain 48, i.e., the inherent properties of the diffusion resistance domain 46 are incorporated into the biointerface domain 48, such that the biointerface domain 48 functions as the diffusion resistance domain 46.

[0171] In certain embodiments, the thickness of the diffusion resistance domain can be about 0.05 μm or less to about 200 μm or more. In some of these embodiments, the thickness of the diffusion resistance domain can be about 0.05, about 0.1, about 0.15, about 0.2, about 0.25, about 0.3, about 0.35, about 0.4, about 0.45, about 0.5, about 1, about 1.5, about 2, about 2.5, about 3, about 3.5, about 4, about 6, or about 8 μm to about 9, about 10, about 11, about 12, about 13, about 14, about 15, about 16, about 17, about 18, about 19, about 19.5, about 20, about 30, about 40, about 50, about 60, about 70, about 75, about 80, about 85, about 90, about 95, or about 100 μm. In some embodiments, the thickness of the diffusion resistance domain is from about 2, about 2.5, or about 3 μm to about 3.5, about 4, about 4.5, or about 5 μm for transcutaneously implanted sensors, or from about 20 or about 25 μm to about 40 or about 50 μm for fully implanted sensors.

[0172] Enzyme domain The enzyme layer, also referred to as the enzyme domain, is a domain or layer of an implantable device configured to immobilize an active enzyme that reacts with an analyte when implanted in or connected to a host (e.g., via an intravascular access device that provides extracorporeal access to blood vessels). In one embodiment, the enzyme domain comprises glucose oxidase. In other embodiments, the enzyme domain can be impregnated with other oxidases, such as galactose oxidase, cholesterol oxidase, amino acid oxidase, alcohol oxidase, lactate oxidase, or uricase. For example, for an enzyme-based electrochemical glucose sensor to perform well, the sensor's response should not be limited by either enzyme activity or cofactor concentration. In other embodiments, the enzyme may be a dehydrogenase, such as glucose dehydrogenase.

[0173] The enzyme layer disclosed herein is mechanically robust, prevents physiochemical degradation upon implantation, and can withstand adhesive degradation during sensor implantation. Furthermore, the disclosed enzyme layer does not affect the response time of sensors that are permeable to analytes and does not alter glucose rate-limiting control by resistive layers. The disclosed enzyme layer can have hydrophilic properties, with water absorption exceeding 10% by dry weight, and rapid water absorption and rapid stabilization, so that sensor start-up is not negatively affected.

[0174] The enzyme layer disclosed herein includes an enzyme layer polymer. The enzyme layer polymer is a polyzwitterion. A polyzwitterion is a polymer in which the repeating units of the polymer chain are zwitterionic moieties. As such, these polymers have equal numbers of cationic and anionic groups, with each zwitterionic repeating unit having both positive and negative charges, and therefore often have an overall zero charge over a wide pH range. Without being bound by theory, it is believed that the zwitterionic groups in the enzyme layer polymer provide charge centers for strong charge-charge interactions with the ionic groups in the enzyme, which may aid in immobilizing the enzyme in the enzyme layer and reducing leakage of the enzyme from the enzyme layer (and sometimes into the host). Furthermore, the zwitterionic groups are highly hydrophilic, retaining water and helping to prevent enzyme denaturation.

[0175] The enzyme layer polymers are polyzwitterions, which are distinguished from other polyampholytes in that polyampholytes contain anionic and cationic groups, but the ionic groups are not correlated with each other as part of the same repeating unit. Thus, the anionic and cationic groups may be randomly spaced apart from each other, or there may be more of one ionic group than the other. Thus, polyampholytes typically have a net charge, except perhaps over a somewhat narrow pH range.

[0176] The disclosed polyzwitterions can have a variety of repeating units, shown below as i-vii), where n is an integer between 2 and 1000. [ka]

[0177] In structures i through iv), the zwitterionic units are attached to the backbone (~~~~), with the charges on side groups pendant to the chain. In structures v) through vii), the zwitterionic units are such that one or both charges are on the chain itself.

[0178] Examples of suitable zwitterionic monomers that can be used to generate polyzwitterions of any of structures i)-vii) include: Included are ammoniophosphates (phosphobetaines or lecithin analogs), ammoniophosphonates (phosphonobetaines), or ammoniophosphinates (phosphinobetaines), each having the structure: [ka] wherein Z is a branched or straight chain alkyl, heteroalkyl, cycloalkyl, cycloheteroalkyl, aryl, or heteroaryl; R 1 is H, alkyl, heteroalkyl, cycloalkyl, heterocycloalkyl, aryl, or heteroaryl, and R 2 , R 3 , and R 4 is independently selected from alkyl, heteroalkyl, cycloalkyl, heterocycloalkyl, aryl, or heteroaryl; R 1 , R 2 , R 3 , R 4 and one or more of Z is substituted with a polymerizable group. Further examples of suitable zwitterionic monomers that can be used to generate polyzwitterions of any of structures i)-vii) include ammoniosulfonates (sulfobetaines), ammoniosulfates, each having the following structure: [ka] wherein Z is a branched or straight chain alkyl, heteroalkyl, cycloalkyl, cycloheteroalkyl, aryl, or heteroaryl; R 1 is H, alkyl, heteroalkyl, cycloalkyl, heterocycloalkyl, aryl, or heteroaryl, and R 2 and R 3 is independently selected from alkyl, heteroalkyl, cycloalkyl, heterocycloalkyl, aryl, or heteroaryl; R 1 , R 2 , R 3 and one or more of Z is substituted with a polymerizable group; Ammoniocarboxylate has the following structure: [ka] wherein Z is a branched or straight chain alkyl, heteroalkyl, cycloalkyl, cycloheteroalkyl, aryl, or heteroaryl; R 1 is H, alkyl, heteroalkyl, cycloalkyl, heterocycloalkyl, aryl, or heteroaryl, and R 2 and R 3 is independently selected from alkyl, heteroalkyl, cycloalkyl, heterocycloalkyl, aryl, or heteroaryl; R 1 , R 2 , R 3 and one or more of Z is substituted with a polymerizable group.

[0179] In each of these monomers, Z can have a length of 1 to 12 atoms, e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 atoms, and any of these values ​​can form the upper or lower endpoint of a range.

[0180] These monomers can be prepared by methods known to those skilled in the art, for example, as detailed in Laschewsky, "Structures and synthesis of zwitterionic polymers," Polymers 6:1544-1601, 2014. In certain examples, the disclosed polyzwitterions can have repeating zwitterionic units derived from any of the zwitterionic monomers disclosed above.

[0181] The enzyme layer polymer may also contain polyurethane and / or polyurea segments. For example, the enzyme layer polymer may include polyurethane copolymers such as polyether-urethane-urea, polycarbonate-urethane, polyether-urethane, silicone-polyether-urethane, silicone-polycarbonate-urethane, polyester-urethane, polyurethane-urea, etc. These polyurethane and / or polyurea segments are referred to herein as "hard segments" because they contain urea and / or urethane linkages formed from polyisocyanates and short-chain polyols or polyamines, moieties that are rich in hydrogen bonds. These segments may also be relatively hydrophobic.

[0182] In addition to the polyurethane and / or polyurea hard segments, the disclosed enzyme layer polymers may also contain soft segments that have relatively poor hydrogen bonding. The soft segments are typically composed of polyols such as polycarbonates, polyesters, polyethers, polyarylenes, and polyalkylenes. The soft segments can be either hydrophobic or hydrophilic.

[0183] Enzyme layer polymers useful in certain embodiments may comprise linear or branched polymers on the backbone structure of the polymer. Thus, either the hard or soft segments may comprise a branched or linear backbone.

[0184] The zwitterionic monomers, as described herein, can be part of either the hard or soft segments, or both.

[0185] In some embodiments, the hard segment portion of the enzyme layer polymer may comprise about 5% to about 50% by weight of the polymer, sometimes about 15% to 20% by weight, and sometimes about 25% to 40% by weight. The hard segment may have a molecular weight of about 160 daltons to about 10,000 daltons, and sometimes about 200 daltons to about 2,000 daltons. In some embodiments, the molecular weight of the soft segment may be about 200 daltons to about 10,000,000 daltons, sometimes about 500 daltons to about 5,000 daltons, and sometimes about 500 daltons to about 2,000 daltons.

[0186] As noted, the hard segments can be polyurethanes or polyureas. Polyurethanes are polymers produced by the condensation reaction of diisocyanates and difunctional hydroxyl-containing materials. Polyureas are polymers produced by the condensation reaction of diisocyanates and difunctional amine-containing materials. Preferred diisocyanates include aliphatic diisocyanates containing from about 4 to about 9 methylene units. Cycloaliphatic diisocyanates containing diisocyanates can also be useful in preparing the polymer and copolymer components of the membranes of preferred embodiments.

[0187] The soft segments used in preparing the enzyme layer polymer may be polyfunctional aliphatic polyols, polyfunctional aliphatic or aromatic amines, etc., which may be useful in effecting permeability of the analyte (e.g., glucose) therethrough, and may include, for example, polyoxazoline, poly(ethylene glycol) (PEG), polyacrylamide, polyimine, polypropylene oxide (PPO), PEG-co-PPO diol, silicone-co-PEG diol, silicone-co-PPO diol, polyethyl acrylate (PEA), polyvinylpyrrolidone (PVP), and modifications thereof (e.g., PVP vinyl acetate), where PEG and modifications thereof may be preferred due to their hydrophilicity.

[0188] In some embodiments, the soft segment portion of the enzyme layer polymer may comprise about 5% to about 70% by weight of the polymer, sometimes about 15% to 20% by weight, and sometimes about 25% to 40% by weight. The soft segment may have a molecular weight of about 160 daltons to about 10,000 daltons, and sometimes about 200 daltons to about 2,000 daltons. In some embodiments, the molecular weight of the soft segment may be about 200 daltons to about 10,000,000 daltons, sometimes about 500 daltons to about 5,000 daltons, and sometimes about 500 daltons to about 2,000 daltons.

[0189] In some embodiments, the enzyme layer polymer comprising hard and soft segments and zwitterionic repeat units can have a molecular weight of about 10 kDa to about 500,000 kDa, e.g., about 10 kDa to about 100,000 kDa, about 1000 kDa to about 500,000 kDa, about 10,000 kDa to about 100,000 kDa, and about 100,000 kDa to about 500,000 kDa.

[0190] The hard and soft segments may each be selected for their properties, such as, but not limited to, tensile strength, flex life, modulus, etc. For example, polyurethanes are relatively strong and offer many reaction pathways, properties that may be advantageous for the membrane domain of a continuous sensor.

[0191] As noted above, the enzyme layer polymers contain one or more zwitterionic repeat units, and therefore these groups are "internal" with respect to the polymer backbone. Such "internal" repeat units are distinguished from materials found at the ends of polymer chains, since such moieties are only attached to the polymer chain at one position. While the disclosed enzyme layer polymers may, in some embodiments, have zwitterionic groups at the ends of the polymer chains, such groups are not the only zwitterionic groups in the chain, and there is at least one internal zwitterionic group in the backbone.

[0192] In some embodiments, the enzyme layer polymer comprises at least about 1%, about 2%, about 3%, about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, about 10%, about 11%, about 12%, about 13%, about 14%, about 15%, about 16%, about 17%, about 18%, about 19%, about 20%, about 21%, about 22%, about 23%, about 24%, about 25%, about 26%, about 27%, about 28%, about 30%, about 31%, about 32%, about 33%, about 34%, about 35%, about 36%, about 37%, about 38%, about 39%, about 40%, about 41%, about 42%, about 43%, about 44%, about 45%, about 46%, about 47%, about 48%, about 49%, about 50%, about 51%, about 52%, about 53%, about 54%, about 55%, about 56%, about 57%, about 58%, about 59%, about 60%, about 61%, about 62%, about 63%, about 64%, about 65%, about 66%, about 67%, about 68%, about 69%, about 70%, about 71%, about 72%, about 73%, about 74%, about 75%, about 76%, about 77%, about 78%, about 79%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92% by weight of the polymer. The enzyme layer polymer may comprise about 28%, about 29%, about 30% to about 31%, about 32%, about 33%, about 34%, about 35%, about 36%, about 37%, about 38%, about 39%, about 40%, about 41%, about 42%, about 43%, about 44%, about 45%, about 46%, about 47%, about 48%, about 49%, about 50%, about 51%, about 52%, about 53%, about 54%, or about 55% by weight of zwitterionic repeat units. In a preferred embodiment, the enzyme layer polymer comprises at least about 20% zwitterionic repeat units by weight of the polymer.

[0193] The zwitterionic repeat units can be betaines, precursors or derivatives thereof (e.g., alkylbetaines or aminobetaines), such as carboxyl, sulfo, or phosphobetaine compounds. These segments or moieties can be incorporated into the enzyme layer polymer, whether in the hard segments, soft segments, or both, for example, at up to about 55% by weight of the enzyme layer polymer.

[0194] In some embodiments, two or more different zwitterionic or zwitterionic precursor segments or moieties are used, while in other embodiments, a single zwitterionic or zwitterionic precursor segment or moiety may be used in the enzyme layer polymer.

[0195] Some examples of enzyme layer polymers are shown schematically in Figure 10. Generally, enzyme layer polymers include one or more hard segments and one or more soft segments. The hard segments can be aliphatic or aromatic monomers. The soft segments can be hydrophilic or hydrophobic oligomers, such as polyalkylene glycols, polycarbonates, polyesters, polyethers, polyvinyl alcohols, polyvinylpyrrolidones, and polyoxazolines. Zwitterionic groups (e.g., betaines) can be part of the soft segments, the hard segments, or both. As shown in Figure 10, a variety of hard and soft segments can be present, allowing for tailoring the properties of the enzyme layer polymer by using different segments, different segment lengths, functionalization on certain segments, crosslinking certain segments, etc. In some embodiments, biocompatible segmented block polyurethane copolymers containing hard and soft segments can be used in the enzyme layer.

[0196] Incorporation of these zwitterionic repeat units into polymers can be achieved by using zwitterionic monomers bearing diols or diamines (e.g., at the Z position), or R 1 ~R 4 R 1 ~R 4 Attachment to a diol or diamine in can be achieved by reacting the corresponding precursor with a halo-substituted diamine or diol. Examples of such monomers are shown below: [ka] wherein W, Y, and Z are independently branched or straight chain alkyl, heteroalkyl, cycloalkyl, cycloheteroalkyl, aryl, or heteroaryl, any of which may be optionally substituted with O, OH, halogen, amino, or alkoxyl; R 1 is H, alkyl, heteroalkyl, cycloalkyl, cycloheteroalkyl, aryl, or heteroaryl, and R 2 , R 3 , and R 4 is independently selected from alkyl, heteroalkyl, cycloalkyl, cycloheteroalkyl, aryl, or heteroaryl. In certain embodiments, W is C1-C4 alkyl. In certain embodiments, Y is C1-C4 alkyl. In other embodiments, Z is C1-C4 alkyl.

[0197] These compounds can be reacted with diisocyanates to form polyurethanes or polyureas. Alternatively, the carboxylate, sulfonate, phosphinate, or phosphonate moieties can be protected, and then the protecting groups can be removed after polymerization. In another alternative, the amines can be tertiary amines, which are then quaternized by alkylation after polymerization.

[0198] Another method involves the radical polymerization of zwitterionic monomers having unsaturated moieties substituted at the Z position in the monomers shown above. In another example, zwitterionic monomers in which the unsaturated moiety is attached to an ammonium group can be used in the radical polymerization. Examples of such monomers are shown below: [ka] wherein X is O, NH, or NR 4 Y and Z are independently branched or straight chain alkyl, heteroalkyl, cycloalkyl, cycloheteroalkyl, aryl, or heteroaryl, any of which may be optionally substituted with OH, halogen, or alkoxyl; R 1is H, alkyl, heteroalkyl, cycloalkyl, cycloheteroalkyl, aryl, or heteroaryl, and R 3 and R 5 is independently selected from heteroalkyl, cycloalkyl, cycloheteroalkyl, aryl, or heteroaryl. 5 is H or CH3. In other embodiments, X is O. In still other embodiments, X is NH or NCH3. In certain embodiments, Y is C1-C4 alkyl. In other embodiments, Z is C1-C4 alkyl.

[0199] Additional examples of suitable zwitterionic monomers include N-(2-methacryloyloxy)ethyl-N,N-dimethylammoniopropanesulfonate, N-(3-methacryloylimino)propyl-N,N-dimethylammoniopropanesulfonate, 2-(methacryloyloxy)ethylphosphatidylcholine, and 3-(2′-vinyl-pyridino)propanesulfonate.

[0200] In other embodiments, the enzyme layer polymer can be crosslinked. For example, polyurethaneurea polymers having aromatic or aliphatic segments with electrophilic functional groups (e.g., carbonyl, aldehyde, anhydride, ester, amide, isocyanate, epoxy, allyl, or halo groups) can be crosslinked with crosslinkers having multiple nucleophilic groups (e.g., hydroxyl, amine, urea, urethane, or thio groups). In further embodiments, polyurethaneurea polymers having aromatic or aliphatic segments with nucleophilic functional groups can be crosslinked with crosslinkers having multiple electrophilic groups. Still further, polyurethaneurea polymers having hydrophilic segments with nucleophilic or electrophilic functional groups can be crosslinked with crosslinkers having multiple electrophilic or nucleophilic groups. The unsaturated functional groups on the polyurethaneurea can also be used for crosslinking by reacting with multivalent free radical agents.

[0201] Non-limiting examples of suitable crosslinkers include isocyanates, carbodiimides, glutaraldehyde or other aldehydes, aziridines, silanes, epoxies, acrylates, free radical agents, ethylene glycol diglycidyl ether (EGDE), poly(ethylene glycol) diglycidyl ether (PEGDE), or dicumyl peroxide (DCP). In one embodiment, about 0.1% to about 15% w / w of crosslinker is added based on the total dry weight of the crosslinker and polymer when blending the components (in one example, about 1% to about 10%). During the curing process, substantially all of the crosslinker is believed to react, leaving substantially no detectable unreacted crosslinker in the final layer.

[0202] Additionally, the disclosed enzyme layers may have zwitterions entrapped or embedded within the polymer network through non-covalent interactions. Thus, in further embodiments, the disclosed enzyme layers may include an enzyme layer polymer and an additional betaine blended therewith. For example, the enzyme layer polymer may be blended with 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). It is understood that many more zwitterionic compounds or precursors or derivatives thereof may be applicable, and this list of exemplary betaines is not intended to limit the scope of the embodiments.

[0203] In certain embodiments, the enzyme layer can include a polyzwitterionic enzyme layer polymer and an enzyme. In other embodiments, the enzyme layer can include a polyzwitterionic enzyme layer polymer and an enzyme blended with a base polymer. Suitable base polymers can include, but are not limited to, silicone, epoxy, polyolefin, polystyrene, polyoxymethylene, polysiloxane, polyether, polyacrylic, polymethacrylic, polyester, polycarbonate, polyamide, poly(etherketone), poly(etherimide), polyurethane, and polyurethaneurea, where polyurethane and polyurethaneurea can include polyurethane copolymers such as polyether-urethane-urea, polycarbonate-urethane, polyether-urethane, silicone-polyether-urethane, silicone-polycarbonate-urethane, polyester-urethane, etc. In some embodiments, the base polymer can be selected for its bulk properties, such as, but not limited to, tensile strength, flex life, and modulus. For example, polyurethane is known to be relatively strong and provide many reaction pathways, which can be advantageous as a bulk property for the membrane domain of a continuous sensor.

[0204] In some embodiments, a base polymer may be used that includes a biocompatible segmented block polyurethane copolymer containing hard and soft segments. In some embodiments, the hard segment of the copolymer may have a molecular weight of about 160 daltons to about 10,000 daltons, and sometimes about 200 daltons to about 2,000 daltons. In some embodiments, the molecular weight of the soft segment may be about 200 daltons to about 10,000,000 daltons, and sometimes about 500 daltons to about 5,000,000 daltons, and sometimes about 500,000 daltons to about 2,000,000 daltons. It is contemplated that the polyisocyanate used to prepare the hard segment of the copolymer may be an aromatic or aliphatic diisocyanate. The soft segments used in preparing the polyurethanes may be polyfunctional aliphatic polyols, polyfunctional aliphatic or aromatic amines, etc., which may be useful in effecting permeability of an analyte (e.g., glucose) therethrough, and may include, for example, polyvinyl acetate (PVA), poly(ethylene glycol) (PEG), polyacrylamide acetate, polyethylene oxide (PEO), polyethyl acrylate (PEA), polyvinylpyrrolidone (PVP), poly(2-oxazoline (POX), and variants thereof (e.g., PVP vinyl acetate), where PVP, POX, and variants thereof may be preferred in some embodiments due to their hydrolytic stability.

[0205] In some embodiments, the one or more zwitterionic compounds or precursors thereof applied to the surface of the membrane system are hydrolyzable cationic esters of zwitterionic compounds. In these embodiments, the hydrolyzable cationic esters provide the added benefit that hydrolysis of the cationic esters to non-fouling zwitterionic groups can kill microorganisms (such as bacteria) or condense DNA. Furthermore, the mixed-charge nature of the resulting zwitterionic groups results in the inhibition of nonspecific protein adsorption on the sensor surface. In these embodiments, cationic betaine esters, such as cationic pCB esters, are preferred.

[0206] Incorporating zwitterion or zwitterion precursor segments or moieties into the polymer backbone can be difficult due to solubility issues associated with the zwitterion or zwitterion precursor monomer. Such groups are typically soluble only in highly polar solvents, such as methanol and water, which are unfavorable for the synthesis of some enzyme layer polymers (e.g., polyurethanes). Therefore, the available functional groups that can be chemically incorporated into the backbone of enzyme layer polymers via solution-based polycondensation synthesis are limited. As an alternative to incorporating zwitterion or zwitterion precursor segments or moieties into the backbone of a base polymer, precursors or derivatives of the zwitterion or zwitterion precursor can be used. For example, zwitterion precursors and / or zwitterion derivatives with more desirable solubility characteristics in low-polarity organic solvents can be used as monomers. Enzyme layer polymers (e.g., polyurethaneureas) can be synthesized via polycondensation reactions to form well-defined polymers with high molecular weights and low polydispersity indices. These polymers can then be converted to zwitterionic group containing polymers via chemical reactions (hydrolysis, deprotection, thermally induced rearrangement, and UV-induced degradation) or biologically induced reactions after in vivo implantation of the device.

[0207] In some embodiments, the enzyme contained in the enzyme layer is susceptible to heat- or pH-induced degradation. In some related embodiments, the enzyme layer may also include one or more enzyme stabilizers. Such agents improve the enzyme's ability to withstand heat- or pH-induced denaturation. Thus, the inclusion of enzyme stabilizers facilitates device fabrication by enabling the use of manufacturing processes that would otherwise reduce enzyme activity. The inclusion of these agents has the added benefit of extending the usable shelf life of the sensor. Any material that improves the thermal and / or pH stability of the enzyme without affecting the oxygen permeability of the analyte or the enzyme layer until the enzyme layer is no longer suitable for use in the sensor may be used as an enzyme stabilizer. In some embodiments, the enzyme stabilizer may be bipolar. Without being bound by theory, it is believed that bipolar enzyme stabilizers stabilize the enzyme by orienting themselves around the enzyme in a manner that provides a localized charge environment that stabilizes the enzyme's tertiary structure.

[0208] Zwitterionic or non-zwitterionic zwitterionic enzyme stabilizers can be neutral molecules with positive and negative charges at different positions. In some embodiments, the positive and negative charges are full unit charges (i.e., the molecule is zwitterionic). In other embodiments, the positive and negative charges are less than full unit charges (i.e., the molecule is zwitterionic, but non-zwitterionic).

[0209] In some embodiments, the zwitterionic enzyme stabilizer can be a betaine, such as glycine betaine, poly(carboxybetaine) (pCB), or poly(sulfobetaine) (pSB), or some other zwitterion, such as cocamidopropyl betaine, oleamidopropyl betaine, octyl sulfobetaine, caprylyl sulfobetaine, lauryl sulfobetaine, myristyl sulfobetaine, palmityl sulfobetaine, stearyl sulfobetaine, betaine (trimethylglycine), octyl betaine, phosphatidylcholine, ectoine, or hydroxyectoine. In a preferred embodiment, the zwitterionic enzyme stabilizer is glycine betaine. In some embodiments, the non-zwitterionic enzyme stabilizing reagent can be an amine oxide.

[0210] When used, enzyme stabilizing reagents may be present at up to about 0.1, about 0.2, about 0.5, about 1, about 2, or about 5% by weight of the enzyme layer. It will be understood that many more zwitterionic groups, or precursors or derivatives thereof, may be applicable, and this exemplary list of betaines is not intended to limit the scope of the embodiments. In some embodiments, hydrolyzable cationic esters of zwitterionic groups (discussed elsewhere) may be used at similar concentrations for incorporation into the enzyme layer.

[0211] In embodiments in which the enzyme layer includes an enzyme stabilizing reagent, the amount of enzyme stabilizing reagent present in the enzyme domain is sufficient to provide improved thermal and / or pH stability of the enzyme, but does not interfere with the permeability characteristics of the enzyme layer, thereby allowing the sensor to retain high glucose sensitivity. While the identity and amount of the enzyme stabilizing reagent used in the enzyme layer may vary based on the particular enzyme used in the sensor, the amount of enzyme stabilizing reagent is generally less than about 50% by weight, e.g., less than about 25% by weight, e.g., less than about 10% by weight, of the amount of enzyme. In a preferred embodiment, the enzyme is glucose oxidase and the enzyme stabilizing reagent is a betaine, such as glycine betaine.

[0212] In some embodiments, the enzyme and enzyme layer polymer, as well as any enzyme stabilizing agents, can be impregnated or otherwise immobilized in the biointerface layer or diffusion-resistant domain, such that a separate enzyme layer is not required (e.g., where a unitary domain is provided that includes the functionality of the biointerface layer, diffusion-resistant domain, interference domain, and enzyme layer). In some embodiments, the enzyme layer is formed from an aqueous dispersion of polyurethane, e.g., a colloidal polyurethane polymer that includes the enzyme and enzyme stabilizing reagent. Again, in some embodiments, the polymer system of the enzyme layer may not be crosslinked, although in other embodiments, it is contemplated that crosslinking may be used and may be achieved by any of a variety of methods, for example, by adding a crosslinking agent.

[0213] In some other embodiments, the blend of polymers containing two or more surface-active groups contains one negatively charged surface-active group and one positively charged surface-active group. In some embodiments, the number of negatively and positively charged surface-active groups is such that the enzyme domain formed from the blend carries approximately a net neutral charge. In other embodiments, the number of positively and negatively charged surface-active groups may not be equal, and there may be more of either the positively or negatively charged surface-active groups.

[0214] In some embodiments, membranes are disclosed that include an enzyme layer 44 (see Figures 2A-2C). The enzyme layers disclosed herein can include an enzyme layer polymer and an enzyme, or in alternative embodiments, the enzyme layer can include an enzyme layer polymer and one or more other polymers that form a polymer blend, and an enzyme.

[0215] In some embodiments, an enzyme layer 44 can be used and is located closer to the electrochemical reaction surface than the diffusion resistance domain 46. The enzyme layer includes an enzyme configured to react with the analyte. In one embodiment, the membrane includes an immobilized enzyme layer 44 including glucose oxidase. In other embodiments, the enzyme layer 44 can be impregnated with other oxidases, such as galactose oxidase, cholesterol oxidase, amino acid oxidase, alcohol oxidase, lactate oxidase, or uricase. For example, for an enzyme-based electrochemical glucose sensor to perform well, the sensor's response should not be limited by enzyme activity or cofactor concentration.

[0216] In some embodiments, the enzyme can be impregnated or otherwise immobilized in the biointerface or diffusion-resistant domain, thereby eliminating the need for a separate enzyme domain 44 (e.g., where a unitary domain is provided that includes the functionality of the biointerface domain, the diffusion-resistant domain, and the enzyme domain). In some embodiments, the enzyme domain 44 is formed from a polyurethane, e.g., an aqueous dispersion of a colloidal polyurethane polymer containing the enzyme.

[0217] In certain embodiments, the thickness of the enzyme layer can be about 0.01, about 0.05, about 0.1, about 0.5, about 1, about 2, about 4, about 6, or about 8 μm to about 10, about 15, about 20, about 30, about 40, about 50, about 75, about 100, about 125, about 150, about 175, about 200, or about 250 μm. In some of these embodiments, the thickness of the enzyme layer can sometimes be about 1 to about 5 μm, and sometimes about 2 to about 7 μm. In other embodiments, the enzyme layer can be about 20 or about 25 μm to about 50, about 55, or about 60 μm thick. In some embodiments, the glucose sensor can be configured for transdermal or short-term subcutaneous implantation and can have a thickness of about 0.5 μm to about 8 μm, and sometimes about 4 μm to about 6 μm. In one glucose sensor configured for fluid communication with a host's circulatory system, the thickness can be from about 1.5 μm to about 25 μm, and sometimes from about 3 to about 15 μm. In some embodiments, the enzyme layer or any other layer of the electrode can have a consistent thickness, but in other embodiments, it is contemplated that the thickness can vary. For example, in some embodiments, the thickness of the enzyme layer can vary along the longitudinal axis of the electrode end.

[0218] In another aspect, the enzyme layer may have a biomimetic adhesive polymer as an additive blended into the enzyme layer to enhance adhesion of the enzyme layer to the diffusion-resistant and interference domains and reduce delamination. A suitable biomimetic adhesive polymer that may be used in this embodiment is a 3,4-dihydroxy-L-phenylalanine-containing polymer. 3,4-dihydroxy-L-phenylalanine (DOPA), an active ingredient in marine mussel proteins, can be converted into a polymerizable monomer and polymerized to form linear, non-degradable homopolymers or copolymers.

[0219] Interference Domain In some embodiments, it is contemplated that an interference domain 43, also referred to as an interference layer, may be provided in addition to (or instead of) the biointerface layer, for example, in the sensor configuration shown in FIG. 2B . The interference domain 43 may substantially reduce the permeation of one or more interferents to the electrochemical reaction surface. The interference domain 43 may be configured to be much less permeable to one or more interferents than the measured species. In some embodiments, a separate interference domain is not present, where interference blocking may be provided by the biointerface layer (e.g., via a surface-active group-containing polymer in the biointerface layer). In other embodiments, the membrane includes both an interference domain and a biointerface layer, and both domains are configured to reduce the permeation of one or more interferents. In further embodiments, the interference domain and the biointerface layer are each configured to reduce the permeation of different interfering species. For example, the interference domain may have greater specificity than the biointerface layer for reducing the permeation of one interfering species, while the biointerface layer may have greater specificity than the interference domain for reducing the permeation of another interfering species. In some embodiments, both the interference domain and the biointerface layer are configured to target a specific interfering species for permeation reduction.

[0220] In certain embodiments, the implantable sensor employs a membrane system comprising a resistance domain, an enzyme domain, and an interference domain. The interference domain can be proximal to the sensor, and the resistance domain can be distal to the sensor, with the enzyme domain between them. The interference domain can consist of a single layer or multiple layers of the same material. However, in some embodiments, the interference domain comprises layers of two or more different species in an alternating configuration. For example, a layer of a first species can be represented by X, a layer of a second species can be represented by Y, and a layer of a third species can be represented by Z. An interference domain comprising alternating layers can have the following exemplary configurations: [ka]

[0221] The above configurations, which are merely exemplary, illustrate various embodiments. In certain embodiments, the first and last layers are the same (e.g., X and X), while in other embodiments, the first and last layers are different (e.g., X and Y). A domain can include one or more layers and can be monolithic (i.e., a single layer is deposited, e.g., X) or composite (e.g., a first layer of material is deposited, followed by second, third, etc. layers of the same material deposited over the first, e.g., XXX). The pattern of alternating layers can be regular (e.g., XYXYXYXYXY) or irregular (e.g., ZYXZXYZYZ).

[0222] In some embodiments, the alternating layers comprise polyanion and polycation layers. The following are exemplary interference domain configurations, where the polyanion layers (unitary, multiple, and / or consecutive with the same or different polyanions) are represented by A, and the polycation layers (unitary, multiple, and / or consecutive with the same or different polyanions) are represented by C: [ka] [ka]

[0223] Other configurations (e.g., those including additional layers and / or additional materials) are also contemplated for some embodiments. In some embodiments, each A layer is a monolithic or composite layer of the same polyanion, and each C layer is a monolithic or composite layer of the same polycation. Both of the outermost layers of an interference domain can be polycationic layers, with the polyanion layer being present only as an internal layer. Any suitable number of alternate layers, e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, or more bilayers (defined as a polycationic layer adjacent to a polyanion layer), can be used in the interference domain. In some embodiments, a final polycationic layer is added to result in an interference domain having a polycationic layer as its outermost layer. In other embodiments, a final anionic layer is added to result in an interference domain having a polyanionic layer as its outermost layer.

[0224] Polyanions and polycations belong to a class of polymers commonly referred to as polyelectrolyte-polymers, in which at least some of the repeating units (or monomers) contain one or more ionic moieties. Polyelectrolytes that carry both cationic and anionic moieties are commonly referred to as polyampholytes. Certain polyelectrolytes form self-assembled monolayers, in which one end of the molecule exhibits a specific, reversible affinity for a substrate, allowing the deposition of an organized, close-packed monolayer of polyelectrolytes.

[0225] The polycation can be any biocompatible polycationic polymer. In some embodiments, the polycation is a biocompatible, water-soluble polycationic polymer. In certain embodiments, water solubility can be enhanced by grafting the polycationic polymer with a water-soluble polynonionic material, such as polyethylene glycol. Representative polycationic materials can include, for example, natural and unnatural polyamino acids, positively charged polysaccharides, and positively charged synthetic polymers that have a net positive charge at neutral pH. Additional examples of suitable polycationic materials include polyamines having amine groups either in the polymer backbone or in the polymer side chains, such as poly-L-lysine and other positively charged polyamino acids of natural or synthetic amino acids or mixtures of amino acids, including poly(D-lysine), poly(ornithine), poly(arginine), and poly(histidine)), as well as non-peptide polyamines, such as poly(aminostyrene), poly(aminoacrylate), poly(N-methylaminoacrylate), poly(N-ethylaminoacrylate), poly(N,N-dimethylaminoacrylate), poly(N,N-diethylaminoacrylate), poly(diallyldimethylammonium chloride), poly(aminomethacrylate), poly(N-methylamino-methacrylate), poly(N,N-dimethylamino-methacrylate), poly(N,N-diethylaminoacrylate), poly(diallyldimethylammonium chloride), poly(aminomethacrylate), poly(N-methylamino-methacrylate), poly(N,N-dimethylamino-methacrylate), poly(N,N-diethylamino-meth ...diethylamino-methacrylate), poly(N,N-diethylamino-methacrylate), poly(diallyldimethylammonium chloride), poly( Poly(N,N-dimethylaminomethacrylate), poly(N,N-diethylaminomethacrylate), poly(ethyleneimine), polymers of quaternary amines such as poly(N,N,N-trimethylaminoacrylate chloride), poly(methylacrylamidopropyltrimethylammonium chloride), and natural or synthetic polysaccharides such as chitosan, poly(allylamine hydrochloride), poly(diallyldimethylammonium chloride), poly(vinylbenzyltrimethylamine), polyaniline or polyaniline sulfonate (p-doped), polypyrrole (p-doped), polyallylamine gluconolactone, and poly(pyridinium acetylene).

[0226] The polyanionic material can be any biocompatible polyanionic polymer, for example, any polymer having carboxylic acid groups attached as pendant groups. The polyionic layer can be hydrophilic (e.g., a material or portion thereof that will more readily associate with water than with lipids). In some embodiments, the polyanionic polymer is a biocompatible, water-soluble polyanionic polymer. Suitable materials include, but are not limited to, alginate, carrageenan, furcellaran, pectin, xanthan, hyaluronic acid, heparin, heparan sulfate, chondroitin sulfate, dermatan sulfate, dextran sulfate, polymethacrylic acid, polyacrylic acid, poly(vinyl sulfate), poly(thiophene-3-acetic acid), poly(4-styrenesulfonate), poly(styrenesulfonate), (poly[1-[4-(3-carboxy-4-hydroxy-phenylazo)benzenesulfonamido]-1,2-ethanediyl, sodium salt]), poly(4-[4-({4-[3-amino-2-(4-hydroxy-phenyl)propylcarbamoyl]-5-oxo-pentyl-}-methyl-amino)-phenylazo]-benzenesulfonic acid), oxidized cellulose, carboxymethylcellulose, and crossmarmellose, synthetic polymers, and copolymers containing pendant carboxyl groups, such as those containing maleic or fumaric acid in the backbone. Predominantly negatively charged polyamino acids are also suitable. Examples of these materials include polyaspartic acid, polyglutamic acid, and their copolymers with other natural and unnatural amino acids. Polyphenolic materials such as tannins and lignins can be used if they are sufficiently biocompatible.

[0227] The molecular weight of the polyionic material can be varied to alter coating characteristics such as coating thickness. As the molecular weight increases, the coating thickness generally increases. However, increasing the molecular weight can make handling more difficult. To achieve a balance between coating thickness, material handling, and other design considerations, the polyionic material can have a specific average molecular weight, Mn. In some embodiments, the average molecular weight of the polyionic material used is about 1,000, 10,000, or 20,000 to about 25,000, 50,000, 100,000, or 150,000 g / mol.

[0228] In some embodiments, interference domains can be prepared using a layer-by-layer method, in which a substrate (e.g., a sensor or a membrane layer on a sensor, such as a resistive layer or an enzyme layer) is first immersed in one polyelectrolyte bath and then in a bath of an oppositely charged polyelectrolyte. Optionally, the substrate can be immersed in a wash bath before or after immersion in the polyelectrolyte bath. During each immersion, a small amount of polyelectrolyte is adsorbed and the surface charge is reversed, thereby allowing for the gradual and controlled construction of an electrostatically crosslinked (or hydrogen-bonded) film of alternating polycation-polyanion layers. This method provides a technique for controlling functionality, as well as film thickness and functionality. For example, it can be used to deposit films as thin as a single monolayer or for thicker layers. Figure 11B shows one embodiment of the layer-by-layer method, which uses alternating adsorption of polycations and polyanions to create the structure shown in Figure 11A. 11B occurs through sequential exposure of substrate 938 to polycation and polyanion solutions, with each deposition step followed by washing to remove unadsorbed polymer. In a first step, polycation 942 is deposited onto substrate 938 (e.g., a wire or flat wafer substrate having an electroactive surface) to form polycation layer 942. As described in more detail elsewhere herein, layer deposition can be performed using any of a variety of techniques, such as dipping and / or spraying. In a second step, deposition of polycation layer 942 is followed by washing to remove unadsorbed polymer. Next, in a third step, polyanion 944 is deposited onto polycation layer 942. Then, in a fourth step, deposition of polyanion layer 944 is followed by washing to remove unadsorbed polymer. These steps can be repeated until the desired interference domain configuration and / or structure is achieved. In an alternative embodiment, instead of depositing a polycation layer onto substrate 938 as the first layer, a polyanion layer is deposited instead. A second layer formed of polycations is then deposited on top of the first, i.e., polyanion, layer, and this process continues until a particular desired interference domain configuration and / or structure is achieved.

[0229] In some embodiments, the method can also use other interactions, such as hydrogen or covalent bonds. Depending on the nature of the polyelectrolyte, polyelectrolyte cross-linking can occur, with a single polyelectrolyte chain adsorbing to two (or more) oppositely charged macroions, thereby establishing a molecular bridge. If only a monolayer of each polyelectrolyte is adsorbed in each deposition step, electrostatically cross-linked hydrogel-type materials can be built up on the surface a few microns at a time. If the substrate is not thoroughly washed between applications of the polyion film, thicker hydrogel-like structures can be deposited.

[0230] In some embodiments, the interference-blocking ability provided by the alternating polycation layer(s) and polyanion layer(s) can be tailored and / or controlled by creating covalent bonds between the polycation layer(s) and polyanion layer(s). Crosslinking can have a substantial effect on the mechanical properties and structure of the film, which in turn can affect the film's interference-blocking ability. Crosslinked polymers can have different crosslink densities. In certain embodiments, crosslinking is used to promote crosslinking between layers. In other embodiments, heat is used to form crosslinks instead of (or in addition to) the crosslinking techniques described above. For example, in some embodiments, imide and amide bonds can be formed between polycation and polyanion layers as a result of elevated temperatures. In some embodiments, photocrosslinking is performed to form covalent bonds between the polycation layer(s) and polyanion layer(s). One major advantage over photocrosslinking is that it offers the possibility of patterning. In certain embodiments, patterning using photocrosslinking is performed to modify the film structure and thus tailor the interference-blocking ability of the interference domains. Blocking ability may correspond to, but is not limited to, the ability to reduce the transport of certain interfering species or the selectivity of transport of a desired species (e.g., HO) over an interfering species. Post-deposition reactions, such as cross-linking and reduction of metal ions to form nanoparticles, provide additional methods for modifying film properties. In some embodiments, cross-linking can be performed between depositions of adjacent polycation or polyanion layers instead of (or in addition to) a post-deposition cross-linking process.

[0231] The overall thickness of an interference layer can affect its permeability to interferents. The overall thickness of the interference domain can be controlled by adjusting the number of layers and / or the degree of washing between layers. Layer deposition via spraying allows control of droplet size and density to provide a coating of a desired, selected thickness without necessarily requiring washing between layers. Furthermore, excess (unbound) material can be removed via other means, such as with an air jet. If residual polyelectrolytes from the previous layer are substantially removed before adding a subsequent layer, the thickness per layer is reduced. Thus, in one embodiment, a surface is first coated with polycations, then excess polycations are removed by washing the surface, followed by addition of polyanions, then the excess is removed, and the process is repeated as needed. In some embodiments, polycations or polyanions from different adjacent layers may be entangled. In further embodiments, they can be entangled across several layers.

[0232] In some embodiments, the level of ionization of the polyion can be controlled, for example, by controlling the pH of the immersion solution containing the polycation or polyanion. Varying the level of ionization of these polyions may alter and / or control the interference-blocking ability of a particular layer. For example, a first polycation layer with a higher level of ionization than a second polycation layer may interact better with a first interfering species and reduce its transport, while a second polycation layer may interact better with a second interfering species and reduce its transport. Varying the level of ionization of the polyion's charge groups can also affect the mechanical, structural, and certain other properties (e.g., diffusion properties) of the interference domain, which may affect its ability to reduce (or totally block) the transport of interfering species. For example, an alternating bilayer containing a polycation and a polyanion, both with high levels of ionization, may bind more tightly than a corresponding bilayer with low levels of ionization. Thus, structural differences between these two membranes, whether in the form of mechanical or other properties (e.g., domain thickness), can affect the performance of the interference domain.

[0233] In some embodiments, the linear charge density of a polyelectrolyte can be controlled, at least in part, by the average charge spacing along the polyion chain. The spacing between charged groups on the polycationic and / or polyanionic polymers that form interference domains can be controlled by polyelectrolyte polymer selection or polymer synthesis. The spacing of the charged groups can significantly affect the structural properties of the interference domain. For example, polyions with charged groups spaced closely together can result in small pores in the interference domain, thereby creating a structure that excludes medium- and large-sized interfering species but allows small pores to pass through them. Conversely, polyions with charged groups spaced a moderate distance from each other can result in medium-sized pores that exclude large-sized interfering species and allow medium- and small-sized interfering species to pass through them. In certain embodiments, the linear charge density of the polyanionic polymer is about 1-50 e / Å, sometimes about 10-25 e / Å, sometimes about 2-10 e / Å, and sometimes 2-3 e / Å, where e is the elementary charge of an electron / proton and Å is the distance in angstroms. In some embodiments, the linear charge density of the polyanionic polymer is about 1-50 e / Å, sometimes about 10-15 e / Å, sometimes about 2-10 e / Å, and sometimes 2-3 e / Å.

[0234] In some embodiments, the linear charge density of the polyanionic polymer is substantially similar to the linear charge density of the polycationic polymer. For example, in one embodiment, the polyanionic layer is formed of (i) poly(acrylic acid), which has an average linear charge density of about 2.5 e / Å, and (ii) poly(allylamine hydrochloride), which also has an average linear charge density of about 2.5 e / Å. In certain embodiments, the polycationic and polyanionic layers may have average linear charge densities that are substantially equal to one another, about 1-50 e / Å, sometimes about 2-25 e / Å, sometimes about 5-10 e / Å, other times about 10-15 e / Å, and other times about 15-25 e / Å.

[0235] By providing interference domains with different linear charge densities, interference domains can be formed that contain different polycation / polyanion bilayers specifically designed to exclude different interfering species based on certain characteristics (e.g., molecular diameter) of the targeted interfering species. For example, in one embodiment, the outermost bilayer of the interference domain is designed to have an intermediate average charge spacing, resulting in a bilayer that excludes only large molecular diameter species but allows the passage of medium and small molecular diameter species therethrough. Conversely, the innermost bilayer of the interference domain may be designed to have a low average charge spacing, resulting in a bilayer that excludes all molecules except those with very small molecular diameters, e.g., HO.

[0236] In some embodiments, the polycation layers may be formed of the same or substantially the same material (e.g., poly(allylamine hydrochloride) (PAH) in the case of a polycation, or poly(acrylic acid) (PAA) in the case of a polyanion), but have different levels of ionization. For example, in one embodiment, the interference domain comprises seven alternating polyelectrolyte layers, with the first, third, fifth, and seventh layers being polycation layers and the second, fourth, and sixth layers being polyanion layers, where the first and seventh layers form the outer layers of the interference domain. In one embodiment, each or some of the polycation layers may have different levels of ionization. For example, in one embodiment, the first, third, fifth, and seventh layers may each have different levels of ionization, with the first layer having the highest level of ionization and the seventh layer having the lowest level of ionization, or vice versa. In an alternative embodiment, some of the polycation layers may share substantially the same level of ionization. For example, in one embodiment, layers 1 and 7 may have substantially the same level of ionization, while layers 3 and 5 may have different levels of ionization. As described elsewhere herein, the ionization level of the polyions may be controlled by controlling the pH of the immersion solution containing the polycations or polyanions. By varying the ionization level of these polyions, the interference-blocking ability of a particular layer may be altered and / or controlled.

[0237] The design of interference domains with layers having levels of ionization can be applied to polyanion layers as well. For example, in one embodiment having seven alternating polyelectrolyte layers, the second, fourth, and sixth layers can each be polyanion layers, each with a different level of ionization, with the second layer having the highest level of ionization and the sixth layer having the lowest level, or vice versa. In alternative embodiments, some of the polyanion layers can share substantially the same level of ionization. For example, in one embodiment, the second and fourth layers can have substantially the same level of ionization, while the sixth layer can have a substantially different level of ionization from the others.

[0238] In certain embodiments, the particular polycation layer(s) and / or polyanion layer(s) selected to form the interference layer may depend, at least in part, on their ability to block, reduce, or prevent the passage of one or more interferents therethrough. For example, a polyanion layer may be selected for its ability to block, reduce, or prevent the passage of a first interferent, while a polycation layer may be selected for its ability to block, reduce, or prevent the passage of a second interferent. A layer may be designed to slow but not block the passage of interferents therethrough, or may be designed to substantially block (e.g., capture) the interferent therein. Additional polyion layers may still be included in interference domains that have specific selectivities for different interferents. Depending on the location of the interference domain in the membrane system relative to the electrode or electroactive surface of the sensor, the permeability of the layer to substances other than the interferents may be important. In sensor systems in which HO (hydrogen peroxide) is produced by an enzyme-catalyzed reaction of the analyte to be detected, the interference domain should be designed to allow HO to pass with minimal impedance when positioned between the electroactive surface and the enzyme layer. On the other hand, in a different membrane design, when the interference domain is positioned distal to the enzyme layer (with respect to the electroactive surface), in some embodiments, the interference domain can be designed to block HO not produced by the enzyme-catalyzed reaction from passing through it. Furthermore, with this particular membrane design, the interference domain can be configured to allow analyte and oxygen to pass with minimal impedance.

[0239] The application of layers in forming interference domains can be accomplished by various methods known in the art. One coating process embodiment simply involves dip-coating and dip-washing steps. Another coating process embodiment simply involves spray-coating and spray-washing steps. However, many alternative embodiments involve the use of various combinations of spray-coating, dip-coating, and / or washing steps. For example, one dip-coating method involves applying a coating of a first polyionic material to a substrate (e.g., a sensor or a membrane layer over a sensor, e.g., a resistive layer or an enzyme layer) by immersing the substrate in a first solution of the first polyionic material, washing the substrate by immersing the substrate in a wash solution, and optionally drying the substrate. This procedure is then repeated using a second polyionic material to form a polyionic bilayer, the second polyionic material having an opposite charge to that of the first polyionic material. This bilayer formation process can be repeated multiple times to generate interference domains. In some embodiments, the number of bilayers can be from 1 to about 16 bilayers, sometimes from 1 to about 10 bilayers, and sometimes from about 3 to about 7 bilayers. In certain embodiments, a final layer of oppositely charged polyionic material can be deposited, whereby the first and final layers have the same charge (both positive or both negative). The immersion time for each of the coating and washing steps can vary depending on many factors. For example, immersion of the substrate in the polyionic solution can occur for a period of about 1 to 30 minutes, or about 2 to 20 minutes, or about 1 to 5 minutes. Washing can be accomplished in a single step, although multiple washing steps can also be used. A series of about 2 to 5 washing steps can be used, with each immersion in the washing solution taking, for example, about 1 to about 3 minutes. In some embodiments, several polycation solutions and / or several polyanion solutions can be used.For example, in certain embodiments, a dip-coating sequence may involve applying a coating of a first polycationic material to a substrate to form a first layer, then applying a first anionic material to the first layer to form a second layer, then applying a second polycationic material to the second layer to form a third layer, then applying a second polyanionic material to form a fourth layer, and then applying either the first or second polycationic material to the fourth layer to form a fifth layer. In some of these embodiments, the dip-coating sequence may be interspersed with washing steps between coating steps. Any of a variety of permutations involving the steps and materials described may be used. In alternative embodiments, the materials used to form the polycationic and / or polyanionic layers may be substantially the same. However, individual polycationic layers may have different levels of ionization from one or more other polycationic layers in interference domains, and individual polyanionic layers may also have different levels of ionization from one or more other polyanionic layers. For example, in one embodiment, a dip-coating sequence involves the use of a first solution containing a polycationic material at a first pH, a second solution containing a polyanionic material at a second pH, a third solution containing the polycationic material at a third pH, a fourth solution containing the polyanionic material at a fourth pH, ​​and a fifth solution containing the polycationic material at a fifth pH. Even when the same polycationic material is used to form the first, third, and fifth layers, the ionization levels of the first, third, and fifth layers are different because the solutions used to form the first, third, and fifth layers have different pHs. Similarly, even when the same polyanionic material is used to form the second and fourth layers, the ionization levels of the second and fourth layers are different because the solutions used to form the second and fifth layers have different pHs. This difference in ionization levels can affect, among other things, the mechanical properties of the film, the structural properties of the film (e.g., porosity, roughness), the diffusion properties of the film, and the selectivity of a particular polyelectrolyte layer for one interfering species over another.All of these effects affect the ability of individual polyelectrolyte layers and interference domains to reduce the transport of various interfering species. In certain embodiments, at least two polycationic and / or two polyanionic layers of an interference domain are formed from the same polycationic / polyanionic material but through the use of solutions at different pHs. In some of these embodiments, the first polycationic layer has a higher selectivity for a particular interfering species over other interfering species, while the second polycationic layer has a higher selectivity for a different interfering species over other interfering species.

[0240] Alternatively, or in addition, spray coating techniques can be used. In one embodiment, the coating process generally involves applying a coating of a first polyionic material to a substrate by contacting the substrate with a first solution of the first polyionic material, washing the substrate by spraying a washing solution onto the substrate, and (optionally) drying the substrate. Similar to the dip-coating process, the spray-coating process can then be repeated with a second polyionic material, the second polyionic material having an opposite charge to that of the first polyionic material. Contact of the substrate with either the polyionic material or the washing solution can occur through various methods. For example, the substrate can be immersed in both solutions. One alternative is to apply the solutions in spray or mist form. Of course, various combinations are possible and within the scope of contemplated embodiments, such as immersing the substrate in the polyionic material and then spraying the washing solution. Spray coating application can be achieved through a number of methods known in the art. For example, a conventional spray coating arrangement may be used, i.e., the liquid material is sprayed by applying the fluid through a small diameter nozzle, which may or may not be at elevated or reduced pressure, directed towards a deposition target. Another spray coating technique involves the use of ultrasonic energy, whereby the liquid is atomized and thereby transformed into a spray by ultrasonic vibrations of a spray-forming tip.

[0241] Yet another technique involves electrostatic spray coating, in which an electric charge is transferred to the fluid or droplets, increasing the efficiency of the coating. Additional methods of atomizing liquids for spray coating require pure mechanical energy, for example, by contacting the liquid with a high-speed reciprocating member or a high-speed rotating disk. Yet another method of generating fine droplets for spray coating involves the use of piezoelectric elements to atomize the liquid. These techniques can be used with air assistance or with elevated solution pressure. Furthermore, a combination of two or more techniques may prove more useful for certain materials and conditions. A method of spray application involves dispensing a polyanion or polycation solution into an ultrasonic dispensing head using a metering pump. A polyion layer is sprayed, allowing the surface droplets to coalesce across the material surface. The resulting layer can then be allowed to interact for a period of time or immediately washed with water or saline solution (or other solutions without the polyanion or polycation).

[0242] In some embodiments, layers of an interference domain can include a polymer with a complex π system. A polymer with a complex π system can contain a delocalized electron system and can be electrically conductive. Layers of a polymer with a complex π system can interact with each other through intermolecular forces, such as electrostatic π-π interactions (i.e., π-stacking). The complex polymer can provide beneficial properties of the interference domain, such as increased domain rigidity, integrity, and / or reproducibility. In some embodiments, the polymer with a complex π system can be polyacetylene, polypyrrole, polythiophene, poly(p-phenylene), poly(p-phenylene vinylene), or poly(carbazole). An interference domain can include alternating layers of any of the above-mentioned complex polymers. In some embodiments, the number of layers of the complex polymer can be from 1 to about 20 layers, sometimes from about 3 to about 10 layers.

[0243] In some embodiments, the thickness of the interference domain can be from about 0.01 microns or less to about 20 microns or more. In some of these embodiments, the thickness of the interference domain can be from about 0.01, 0.05, 0.1, 0.15, 0.2, 0.25, 0.3, 0.35, 0.4, 0.45, 0.5, 1, 1.5, 2, 2.5, 3, or 3.5 microns to about 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 19.5 microns. In some of these embodiments, the thickness of the interference domain can be from about 0.2, 0.4, 0.5, or 0.6 microns to about 0.8, 0.9, 1, 1.5, 2, 3, or 4 microns.

[0244] Polyimine Film In some embodiments, certain polymer films can be used to form interference domains. For example, certain polyimides prepared from 2,2'-dimethyl-4,4'-diaminobiphenyl and the corresponding dianhydrides can be cast into films that can be used as hydrogen peroxide-selective membranes. See, for example, Ekinci et al., Turk. J. Chem. 30 (2006), 277-285. In one embodiment, the film is prepared using the following steps: First, n-methyl-2-pyrrolidone (NMP) is distilled over CaH2 under reduced pressure and stored over approximately 4 Å molecular sieves. Reagent-grade pyromellitic dianhydride (PMDA) is sublimed under reduced pressure at approximately 250°C and dried under vacuum at approximately 120°C before use. The diamine is purified via recrystallization from ethanol to obtain shiny crystals. Next, 2,20-dimethyl-4,40-diaminobiphenyl (approximately 1.06 g, approximately 5 mmol) is dissolved in NMP (approximately 15 mL) in a 50 mL Schlenk tube equipped with a nitrogen line, an overhead stirrer, a xylene-filled Dean-Stark trap, and a condenser. PMDA (approximately 1.09 g, approximately 5 mmol) is then added to the amine solution, followed by overnight stirring to obtain a viscous solution. After stirring for approximately 3 hours, the solution is heated to reflux at approximately 200 °C for approximately 15 hours. During the polymerization process, water produced from imidization is distilled from the reaction mixture along with approximately 1-2 mL of xylene. After cooling to ambient temperature, the solution is diluted with NMP and then slowly added to a vigorously stirred solution of 95% ethanol. The precipitated polymer is collected via filtration, washed with ethanol, and dried under reduced pressure at 150 °C. Prior to coating, the substrate (e.g., a Pt electrode) is cleaned and optionally polished to approximately 0.05 μm with an aqueous alumina slurry. Then, about 20 μL of a polymer solution prepared by dissolving about 70 mg of polyimide in about 2 mL of NMP is dropped onto the surface of the Pt electrode and dried at room temperature for about 3 days.

[0245] self-assembly techniques Using a self-assembly process, ultrathin multilayer films containing sequentially alternating anionic and cationic polyelectrolytes on a charged surface can be constructed. See, for example, Decher et al., Thin Solid Films, 210-211 (1992) 831-835. Ionic attraction between opposite charges is the driving force for multilayer assembly. Contrary to chemisorption techniques, which require approximately 100% reaction yield to maintain surface functional density in each layer, covalent bonds do not need to be formed in the self-assembly process. Furthermore, an advantage over classical Langmuir-Blodgett techniques is that solution processes are independent of substrate size and topology. Exemplary polyelectrolytes for use in such processes include, but are not limited to, polystyrene sulfonate sodium salt, polyvinyl sulfonate potassium salt, poly-4-vinylbenzyl-(N,N-diethyl-N-methyl-)-ammonium iodide, and poly(allylamine hydrochloride). Assembly of multilayer films can be carried out as follows: A solid substrate with a positively charged planar surface is immersed in a solution containing an anionic polyelectrolyte, and a monolayer of polyanions is adsorbed. Because adsorption is performed at a relatively high polyelectrolyte concentration, many ionic groups remain exposed at the interface with the solution, thus reversing the surface charge. After rinsing with pure water, the substrate is immersed in a solution containing a cationic polyelectrolyte. A monolayer is again adsorbed, but now the original surface charge is restored. By repeating both steps in a cyclical manner, alternating multilayer assemblies of both polymers are obtained. This multilayer formation process is based on the attraction of opposite charges and requires a minimum of two oppositely charged molecules. As a result, more than two molecules can be incorporated into the multilayer by simply immersing the substrate in as many polyelectrolyte solutions as desired, as long as the charges are reversed from layer to layer. Micro-oscillating multilayer assemblies can also be easily prepared. In this respect, this technique is more versatile than the Langmuir-Blodgett technique, which is limited to periodically alternating layer systems. Another advantage is that the immersion procedure does not impose major limitations on the size of the substrate or automation in a continuous process.

[0246] A specific example of the preparation of such a film follows. Polystyrene sulfonate (sodium salt, Mr = 100,000) and polyvinyl sulfonate (potassium salt, Mr = 245,000), and poly(allylamine hydrochloride), Mw = 50,000-65,000, are obtained from commercial sources and used without further purification. Poly-4-vinylbenzyl-(N,N-diethyl-N-methyl-)-ammonium iodide can be synthesized as described in Decher et al., Ber. Bunsenges. Phys. Chem., 95 (1992) 1430. Alternating multilayer assemblies of all materials can be characterized by UV / visible spectroscopy and small-angle X-ray scattering (SAXS) using techniques known in the art. Direct light microscopy and SAXS measurements can be performed using multilayer assemblies on suitable substrates. Multilayer films can be deposited, for example, on platinum or other metal electrodes, or suitable intervening layers can be deposited on top of the electrodes. For the adsorption of the first layer, an aqueous acidic solution of polystyrene sulfonate or polyvinyl sulfonate can be used. The substrate is then washed with water. After adsorption of the first layer, the substrate can be stored for several weeks without significant surface degradation. The cationic polyelectrolyte polyallylamine is then adsorbed from aqueous solution. In the case of non-quaternized polyallylamine, the polycation is adsorbed from an acidic solution. All subsequent layers of anionic polyelectrolyte (odd layer numbers) are adsorbed from aqueous solution. In the case of samples containing polyallylamine as a previously adsorbed layer, the polystyrene sulfonate layer can be adsorbed from an acidic solution. An adsorption time of approximately 20 minutes at ambient temperature can be used, although longer or shorter adsorption times may be acceptable in certain embodiments. A range of polymer concentrations (e.g., 20-30 mg / approximately 10 mL of water) can provide acceptable results.

[0247] Multilayer molecular films of polyelectrolyte:calixarene and polyelectrolyte:cyclodextrin hosts can be fabricated by alternating the adsorption of charged species from aqueous solution onto suitable substrates. See, for example, X. Yang, Sensors and Actuators B 45 (1997) 87-92. Using such a layer-by-layer molecular deposition approach, molecular recognition reagents can be incorporated into polymer films. This deposition process is highly reproducible, and the resulting films are uniform and stable. By substituting polyanions, highly negatively charged molecular species can be used in film fabrication. These molecular reagents can bind organic species and be deposited into thin films as functional components. This approach incorporates polymer and molecular elements into the film, thus resulting in a film with the physical properties of a polymer and the selectivity of a molecular film. Films can be prepared as follows: A substrate (e.g., a Pt electrode) can be first treated with aminopropyltrimethoxysilane in chloroform, followed by the deposition of PSS and then PDDA polyelectrolytes by immersion in aqueous solutions of the respective polyelectrolytes. This can be followed by alternating depositions of a negatively charged molecular host species (e.g., calix[6]arene or pt-butylcalix[4]arene) and PDDA until the desired number of bilayers is reached. Between each deposition, the substrate is thoroughly washed with deionized water. Polyelectrolyte and molecular-ion assembly can be monitored by UV-visible absorption spectroscopy, and mass loading can be measured with a surface acoustic wave (SAW) device.

[0248] In some embodiments, the interference domain is formed from one or more cellulose derivatives. Generally, the cellulose derivatives can include polymers such as cellulose acetate, cellulose acetate butyrate, 2-hydroxyethyl cellulose, cellulose acetate phthalate, cellulose acetate propionate, cellulose acetate trimellitate, or blends and combinations thereof.

[0249] In some alternative embodiments, other polymeric species that can be utilized as the base material for the interference domain include polyurethanes and / or polymers with controlled pore sizes. In one such alternative embodiment, the interference domain comprises a non-swelling, thin, hydrophobic membrane that restricts the diffusion of low molecular weight species. The interference domain is permeable to relatively low molecular weight substances, such as hydrogen peroxide, but restricts the passage of higher molecular weight substances, including glucose and ascorbic acid. Other systems and methods for reducing or eliminating interfering species that can be applied to the membrane system of the preferred embodiments are described in U.S. Pat. No. 7,074,307, U.S. Patent Publication No. US-2005-0176136-A1, U.S. Pat. No. 7,081,195, and U.S. Patent Publication No. US-2005-0143635-A1, each of which is incorporated herein by reference in its entirety.

[0250] In some embodiments, it is contemplated that the thickness of the interference domain can be from about 0.01 μm or less to about 20 μm or more. In some of these embodiments, the thickness of the interference domain can be from about 0.01, about 0.05, about 0.1, about 0.15, about 0.2, about 0.25, about 0.3, about 0.35, about 0.4, about 0.45, about 0.5, about 1, about 1.5, about 2, about 2.5, about 3, or about 3.5 μm to about 4, about 5, about 6, about 7, about 8, about 9, about 10, about 11, about 12, about 13, about 14, about 15, about 16, about 17, about 18, about 19, or about 19.5 μm. In some of these embodiments, the thickness of the interference domain can be from about 0.2, about 0.4, about 0.5, or about 0.6 microns to about 0.8, about 0.9, about 1, about 1.5, about 2, about 3, or about 4 microns.

[0251] Generally, the membrane systems described herein can be formed or deposited on exposed electroactive surfaces (e.g., one or more of the working and reference electrodes) using known thin film techniques (e.g., casting, spray coating, spreading, electrodeposition, dip coating, etc.), although casting or other known application techniques can also be utilized. In some embodiments, the interference domain can be deposited by spraying or dip coating. In an exemplary embodiment, the interference domain is formed by dip coating the sensor into the interference domain solution using an insertion rate of about 0.5 inches / minute to about 60 inches / minute, sometimes about 1 inch / minute, a residence time of about 0.01 minutes to about 2 minutes, sometimes about 1 minute, and a withdrawal rate of about 0.5 inches / minute to about 60 inches / minute, sometimes about 1 inch / minute, and curing (drying) the domain for about 1 minute to about 14 hours, sometimes about 3 minutes to about 15 minutes (which can be accomplished at room temperature or under vacuum, e.g., about 20 to about 30 mmHg). In one exemplary embodiment including a cellulose acetate butyrate interference domain, a 3 minute cure (i.e., drying) time is used between each applied layer. In another exemplary embodiment using a cellulose acetate interference domain, a 15 minute cure time is used between each applied layer.

[0252] In some embodiments, the immersion process can be repeated at least once and up to 10 or more times. In other embodiments, only one immersion is preferred. The preferred number of repeated immersion processes can depend on the cellulose derivative(s) used, their concentration, conditions during deposition (e.g., immersion), and desired thickness (e.g., a thickness sufficient to provide functional blocking of a particular interferent), etc. In one embodiment, the interference domain is formed from three layers of cellulose acetate butyrate. In another embodiment, the interference domain is formed from ten layers of cellulose acetate. In yet another embodiment, the interference domain is formed from one layer of a blend of cellulose acetate and cellulose acetate butyrate. In alternative embodiments, the interference domain can be formed using any known method and combination of cellulose acetate and cellulose acetate butyrate, as will be understood by those skilled in the art.

[0253] Electrode Domain In some embodiments, such as the embodiment shown in FIG. 2A, an optional electrode domain 42, also referred to as an electrode layer, may be provided in addition to the biointerface domain and the enzyme domain, while in other embodiments the functionality of the electrode domain may be incorporated into an integral domain that includes the functionality of the biointerface domain, the diffusion resistance domain, the enzyme domain, and the electrode domain.

[0254] In some embodiments, the electrode domain is located proximal to the electrochemically reactive surface. To facilitate the electrochemical reaction, the electrode domain may include a semi-permeable coating that maintains hydrophilicity at the electrochemically reactive surface of the sensor interface. The electrode domain can enhance the stability of adjacent domains by protecting and supporting the materials that form the adjacent domains. The electrode domain can also help stabilize device operation by overcoming electrode starting and drift problems caused by improper electrolytes. The buffered electrolyte solution contained in the electrode domain can also protect against pH-mediated damage that can result from the formation of large pH gradients between the electrode and a substantially hydrophobic interference domain due to the electrochemical activity of the electrode.

[0255] In some embodiments, the electrode domains are between about 0.05 μm and about 100 μm, sometimes about 0.05, about 0.1, about 0.15, about 0.2, about 0.25, about 0.3, about 0.35, about 0.4, about 0.45, about 0.5, or about 1 μm to about 1.5, about 2, about 2.5, about 3, about 3.5, about 4, about 4.5, about 5, about 6, about 6.5, about 7, about 7.5, about 8, about 8.5, about The electrode domain may comprise a flexible, water-swellable, substantially solid, gel-like film (e.g., a hydrogel) having a "dry film" thickness of about 9, about 9.5, about 10, about 10.5, about 11, about 11.5, about 12, about 13, about 14, about 15, about 16, about 17, about 18, about 19, about 19.5, about 20, about 30, about 40, about 50, about 60, about 70, about 80, about 90, or about 100 μm. In some embodiments, the thickness of the electrode domain may be from about 2, about 2.5, or about 3 μm to about 3.5, about 4, about 4.5, or about 5 μm for transcutaneously implanted sensors, or from about 6, about 7, or about 8 μm to about 9, about 10, about 11, or about 12 μm for fully implanted sensors. As used herein, the term "dry film thickness" is a broad term given its ordinary and customary meaning to those skilled in the art (and is not limited to any special or customized meaning), and refers to, but is not limited to, the thickness of a cured film cast from a coating formulation on the surface of a membrane by standard coating techniques. The coating formulation may include a premix of a film-forming polymer and a crosslinker, and may be curable upon application of moderate heat.

[0256] In certain embodiments, the electrode domains may be formed from a curable mixture of a urethane polymer and a hydrophilic polymer. In some of these embodiments, the coating is formed from a polyurethane polymer having anionic carboxylate functional groups and nonionic hydrophilic polyether segments, which is crosslinked in the presence of polyvinylpyrrolidone and cured at a moderate temperature of about 50°C.

[0257] Aqueous dispersions of fully reacted colloidal polyurethane polymers with crosslinkable carboxyl functionality (e.g., BAYBOND™, Mobay Corporation) are particularly suitable for this purpose. These polymers are supplied in dispersion grades with a carboxylate-containing polycarbonate-polyurethane backbone identified as XW-121 and XW-123, and a carboxylate-containing polyester-polyurethane backbone identified as XW-110-2. In some embodiments, BAYBOND™ 123, an aqueous anionic dispersion of an aliphatic polycarbonate urethane polymer sold as a 35% by weight solution in water and the cosolvent N-methyl-2-pyrrolidone, may be used.

[0258] In some embodiments, the electrode domain is formed from a hydrophilic polymer that renders the electrode domain equal to or more hydrophilic than the overlying domain (e.g., interference domain, enzyme domain). Such hydrophilic polymers can include, for example, polyamides, polylactones, polyimides, polylactams, functionalized polyamides, functionalized polylactones, functionalized polyimides, functionalized polylactams, or combinations thereof.

[0259] In some embodiments, the electrode domain is formed primarily from a hydrophilic polymer, and in some of these embodiments, the electrode domain is formed substantially from PVP. PVP is a hydrophilic, water-soluble polymer commercially available from BASF Wyandotte and GAF ​​Corporation as the PVP K™ homopolymer series in a range of viscosity grades and average molecular weights ranging from about 18,000 to about 500,000. In certain embodiments, a PVP homopolymer having an average molecular weight of about 360,000, identified as PVP-K90 (BASF Wyandotte), can be used to form the electrode domain. Hydrophilic film-forming copolymers of N-vinylpyrrolidone, such as copolymers of N-vinylpyrrolidone and vinyl acetate, copolymers of N-vinylpyrrolidone, ethyl methacrylate, and methacrylic acid monomers, are also suitable.

[0260] In certain embodiments, the electrode domain is formed entirely from a hydrophilic polymer. Useful contemplated hydrophilic polymers include, but are not limited to, poly-N-vinylpyrrolidone, poly-N-vinyl-2-piperidone, poly-N-vinyl-2-caprolactam, poly-N-vinyl-3-methyl-2-caprolactam, poly-N-vinyl-3-methyl-2-piperidone, poly-N-vinyl-4-methyl-2-piperidone, poly-N-vinyl-4-methyl-2-caprolactam, poly-N-vinyl-3-ethyl-2-pyrrolidone, poly-N-vinyl-4,5-dimethyl-2-pyrrolidone, polyvinylimidazole, poly-N,N-dimethylacrylamide, polyvinyl alcohol, polyacrylic acid, polyethylene oxide, poly-2-ethyl-oxazoline, copolymers thereof, and mixtures thereof. Blends of two or more hydrophilic polymers may be suitable in some embodiments.

[0261] In certain embodiments, the hydrophilic polymer used may not be crosslinked, while in other embodiments, crosslinking may be used and may be achieved by any of a variety of methods, for example, by adding a crosslinking agent. In some embodiments, polyurethane polymers may be crosslinked in the presence of PVP by preparing a premix of the polymers and adding the crosslinking agent immediately prior to membrane formation. Suitable crosslinking agents contemplated include, but are not limited to, carbodiimides (e.g., 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride and UCARLNK™ XL-25 (Union Carbide)), epoxides, and melamine / formaldehyde resins. Alternatively, it is contemplated that crosslinking may be achieved by irradiation at a wavelength sufficient to promote crosslinking between hydrophilic polymer molecules, which is believed to create a more tortuous diffusion path through the domains.

[0262] The flexibility and rigidity of the coating can be varied as needed by varying the dry weight solids of the components in the coating formulation. As used herein, the term "dry weight solids" is a broad term, given its ordinary and customary meaning to those skilled in the art (and is not limited to any special or customized meaning), and refers to, but is not limited to, the dry weight percent based on the total coating composition after the crosslinker is included. In one embodiment, the coating formulation may contain about 6 to about 20% by dry weight, preferably about 8% by dry weight, PVP; about 3 to about 10% by dry weight, sometimes about 5% by dry weight, crosslinker; and about 70 to about 91% by weight, sometimes about 87% by weight, of a polyurethane polymer, such as a polycarbonate-polyolefin polymer. The reaction product of such a coating formulation is referred to herein as a water-swellable crosslinked matrix of polyurethane and PVP.

[0263] In some embodiments, beneath the electrode domain is an electrolyte phase, a free liquid phase comprising a solution containing at least one compound, typically a soluble chloride salt, that conducts current when hydrated. In one embodiment in which the membrane system is used with a glucose sensor as described herein, the electrolyte phase flows over the electrode and contacts the electrode domain. It is contemplated that certain embodiments can use any suitable electrolyte solution, including standard commercially available solutions. Generally, the electrolyte phase can have the same or a lower osmolality as the sample being analyzed. In a preferred embodiment, the electrolyte phase comprises normal saline.

[0264] Bioactive Agents It is contemplated that any of a variety of bioactive (therapeutic) agents can be used with the analyte sensor systems described herein, such as the analyte sensor system shown in FIG. 1 . In certain embodiments, the bioactive agent can be in the biointerface layer of the disclosed devices. In some embodiments, the bioactive agent is an anticoagulant. As used herein, the term “anticoagulant” is a broad term given its ordinary and customary meaning to those skilled in the art (and is not limited to any special or customized meaning) and refers to, but is not limited to, a substance that prevents clotting (e.g., minimizes, reduces, or stops blood embolism). In these embodiments, the anticoagulant included in the analyte sensor system can prevent clotting in or on the sensor.Anticoagulants suitable for incorporation into the sensor system include, but are not limited to, vitamin K antagonists (e.g., acenocoumarol, chlorindione, dicumarol, diphenadione, ethyl viscoum acetate, phenprocoumon, phenindione, thiochromarol, or warfarin), heparin-based anticoagulants (e.g., platelet aggregation inhibitors: antithrombin III, bemiparin, dalteparin, danaparoid, enoxaparin, heparin, nadroparin, parnaparin, reviparin, soldexide, tinzaparin), other platelet aggregation inhibitors (e.g., abciximab, acetylsalicylic acid (aspirin), aloxiprine, beraprost, diazol, carbasal). These include but are not limited to: calcium nitrite, chloricromene, clopidogrel, dipyridamole, epoprostenol, eptifibatide, indobufen, iloprost, picotamide, ticlopidine, tirofiban, treprostinil, triflusal), enzymes (e.g., alteplase, ancrod, anistreplase, brinase, drotrecogin alfa, fibrinolysin, protein C, reteplase, saruplase, streptokinase, tenecteplase, urokinase), direct thrombin inhibitors (e.g., argatroban, bivalirudin, desirudin, lepirudin, melagatran, ximelagatran), other antithrombotic agents (e.g., dabigatran, defibrotide, dermatan sulfate, fondaparinux, rivaroxaban), and the like.

[0265] In one embodiment, heparin is incorporated into the analyte sensor system, for example, by dipping or spraying. Without being bound by theory, heparin coated on a catheter or sensor can prevent blood from coagulating and embolizing on the analyte sensor system, thereby preventing thromboembolization (e.g., prevention of blood flow by clots or clotting) or subsequent complications. In some embodiments, heparin is admixed with one or more zwitterionic compounds or derivatives thereof, such as hydrolyzable cationic esters thereof (described above), prior to dipping or spraying, thus providing the sensor system with a mixed coating of heparin and one or more zwitterionic compounds or derivatives thereof.

[0266] In some embodiments, an antimicrobial agent is coated on the catheter (inner or outer diameter) or the sensor. In some embodiments, the antimicrobial agent may be incorporated into the analyte sensor system. Contemplated antimicrobial agents may include, but are not limited to, antibiotics, antiseptics, disinfectants, and synthetic moieties, as well as combinations thereof, and other agents that are soluble in organic solvents such as alcohols, ketones, ethers, aldehydes, acetonitrile, acetic acid, methylene chloride, and chloroform. The amount of each antimicrobial agent used to impregnate the medical device will vary to some extent, but will at least be a concentration effective to inhibit the growth of bacterial and fungal organisms, such as Staphylococcus aureus, Gram-positive bacteria, Gram-negative bacteria, and Candida.

[0267] In some embodiments, antibiotics can be incorporated into the analyte sensor system. Classes of antibiotics that can be used can include tetracyclines (e.g., minocycline), rifamycins (e.g., rifampin), macrolides (e.g., erythromycin), penicillins (e.g., nafeylline), cephalosporins (e.g., cefazolin), other β-lactam antibiotics (e.g., imipenem, aztreonam), aminoglycosides (e.g., gentamicin), chloramphenicol, sulfonamides (e.g., sulfamethoxazole), glycopeptides (e.g., vancomycin), quinolones (e.g., ciprofloxacin), fusidic acid, trimethoprim, metronidazole, clindamycin, mupirocin, polyenes (e.g., amphotericin B), azoles (e.g., fluconazole), and β-lactam inhibitors (e.g., sulbactam).

[0268] Examples of specific antibiotics that may be used include minocycline, rifampin, erythromycin, nafcillin, cefazolin, imipenem, aztreonam, gentamicin, sulfamethoxazole, vancomycin, ciprofloxacin, trimethoprim, metronidazole, clindamycin, teicoplanin, mupirocin, azithromycin, clarithromycin, ofloxacin, lomefloxacin, norfloxacin, nalidixic acid, sparfloxacin, pefloxacin, amifloxacin, enoxacin, fleroxacin, temafloxacin, tosufloxacin, clinafloxacin, sulbactam, clavulanic acid, amphotericin B, fluconazole, itraconazole, ketoconazole, and nystatin.

[0269] In some embodiments, antiseptics or disinfectants can be incorporated into the analyte sensor system. Examples of antiseptics and disinfectants include hexachlorophene, cationic bis-iguanides (e.g., chlorhexidine, cyclohexidine), iodines and iodophors (e.g., povidone-iodine), para-chloro-meta-xylenol, triclosan, furan medical preparations (e.g., nitrofurantoin, nitrofurazone), methenamine, aldehydes (glutaraldehyde, formaldehyde), and alcohols. Other examples of antiseptics and disinfectants will be readily apparent to those skilled in the art.

[0270] In some embodiments, anti-barrier cell agents can be incorporated into the analyte sensor system. Anti-barrier cell agents can include compounds that are effective against macrophages and foreign body giant cells (FBGCs). Anti-barrier cell agents are believed to prevent the closure of the barrier to lysate transport presented by macrophages and FBGCs at the device-tissue interface during FBC maturation. Anti-barrier cell agents can provide an anti-inflammatory or immunosuppressive mechanism that affects the wound healing process, for example, the healing of wounds caused by incisions through which implantable devices are inserted. Cyclosporine, which stimulates very high levels of angiogenesis around biomaterials, can be incorporated into the biointerface membrane of preferred embodiments (see, e.g., U.S. Pat. No. 5,569,462 to Martinson et al.). Alternatively, dexamethasone, which attenuates the intensity of FBC responses at the tissue-device interface, can be incorporated into the biointerface membrane of preferred embodiments. Alternatively, rapamycin, a potent and specific inhibitor of some macrophage inflammatory functions, can be incorporated into the biointerface membrane of preferred embodiments.

[0271] In some embodiments, an anti-inflammatory agent can be incorporated into the analyte sensor system to reduce acute or chronic inflammation adjacent to the implant or to reduce the formation of FBC capsules, e.g., to reduce the formation of a barrier cell layer. Suitable anti-inflammatory agents include, but are not limited to, nonsteroidal anti-inflammatory drugs (NSAIDS), such as acetomethaphine, aminosalicylic acid, aspirin, celecoxib, choline magnesium trisalicylate, diclofenac potassium, diclofenac sodium, diflunisal, etodolac, fenoprofen, flurbiprofen, ibuprofen, indomethacin, interleukin (IL)-10, IL-6 muteins, anti-IL-6 iNOS inhibitors (e.g., L-NAME or L-NMDA), interferons, ketoprofen, ketorolac, leflunomide, melenamic acid, mycophenolic acid, mizoribine, nabumetone, naproxen, naproxen sodium, oxaprozin, Piroxicam, rofecoxib, salsalate, sulindac, and trimethine; and corticosteroids such as cortisone, hydrocortisone, methylprednisolone, prednisone, prednisolone, betamethasone, beclomethasone dipropionate, budesonide, dexamethasone sodium phosphate, flunisolide, fluticasone propionate, paclitaxel, tacrolimus, tranilast, triamcinolone acetonide, betamethasone, fluocinolone, betamethasone dipropionate, betamethasone valerate, desonide, desoximetasone, fluocinolone, triamcinolone, triamcinolone acetonide, clobetasol propionate, and dexamethasone.

[0272] In some embodiments, immunosuppressants or immunomodulators may be incorporated into the analyte sensor system to directly interfere with some of the key mechanisms essential for the involvement of different cellular elements in the inflammatory response. Suitable immunosuppressants and immunomodulators include, but are not limited to, antiproliferative cell cycle inhibitors (e.g., paclitaxel, cytochalasin D, infiximab), taxol, actinomycin, mitomycin, thospromote VEGF, estradiol, NO donors, QP-2, tacrolimus, tranilast, actinomycin, everolimus, methotrexate, mycophenolic acid, angiopeptin, vincristine, mitomycin, statins, C These include MYC antisense, sirolimus (and analogs), RestenASE, 2-chloro-deoxyadenosine, PCNA ribozyme, batimustat, prolyl hydroxylase inhibitors, PPARγ ligands (e.g., troglitazone, rosiglitazone, pioglitazone), halofuginone, C-proteinase inhibitors, probucol, BCP671, EPC antibodies, catchin, glycating agents, endothelin inhibitors (e.g., ambrisentan, tesosentan, bosentan), statins (e.g., cerivastine), E. coli heat-labile enteroloxin, and advanced coatings.

[0273] In some embodiments, an anti-infective agent may be incorporated into the analyte sensor system. Generally, an anti-infective agent is a substance that can act against infection by inhibiting the spread of infectious agents or by immediately killing infectious agents, and may function, for example, to reduce an immune response without an inflammatory response at the implant site. Anti-infective agents include, but are not limited to, anthelmintics (e.g., mebendazole), antibiotics (e.g., aminoglycosides, gentamicin, neomycin, tobramycin), antifungal antibiotics (e.g., amphotericin b, fluconazole, glycerofulvin, itraconazole, ketoconazole, nystatin, micatin, tolnaftate), cephalosporins (e.g., cefaclor, cefazolin, cefotaxime, ceftazidime, ceftriaxone, cefuroxime, cephalexin), β-lactam antibiotics (e.g., cefotetan, meropenem), chloramphenicol, macrolides (e.g., azithromycin, clarithromycin, erythromycin), penicillins (e.g., penicillin G sodium salt, amoxicillin, ampicillin, dicloxacillin, nafcillin, pipera, cyclosporine, ticarcillin), tetracyclines (e.g., doxycycline, minocycline, tetracycline), bacitracin, clindamycin, colistin metasodium, polymyxin b sulfate, vancomycin, antivirals (e.g., acyclovir, amantadine, didanosine, efavirenz, foscarnet, ganciclovir, indinavir, lamivudine, nelfinavir, ritonavir, saquinavir, silver, stavudine, valacyclovir, valganciclovir, zidovudine), quinolones (e.g., ciprofloxacin, levofloxacin), sulfonamides (e.g., sulfadiazine, sulfisoxazole), sulfones (e.g., dapsone), furazolidone, metronidazole, pentamidine, sulfanilamidum crystallinum), gatifloxacin, and sulfamethoxazole / trimethoprim.

[0274] In some embodiments, an angiogenic agent can be incorporated into the analyte sensor system. An angiogenic agent can generally include substances with direct or indirect angiogenic properties. In some cases, an angiogenic agent can additionally affect the formation of barrier cells in vivo. Indirect angiogenesis means that angiogenesis can be mediated through inflammatory or immunostimulatory pathways. How agents that induce local angiogenesis inhibit barrier cell formation is not fully known, but without being bound by theory, it is believed that some barrier cell stiffening may result indirectly from the effects of the angiogenic agent.

[0275] Angiogenic agents may promote angiogenesis, accelerating wound healing around the membrane or providing a mechanism for minimizing ischemic periods by increasing angiogenesis near the tissue-device interface. Sphingosine-1-phosphate (S1P), a phospholipid with potent angiogenic activity, may be incorporated into the biointerface membrane. Monobutyrin, a vasodilatory and angiogenic lipid product of adipocytes, may also be incorporated into the biointerface membrane. In another embodiment, an antisense molecule (e.g., thrombospondin-2 antisense) capable of increasing angiogenesis is incorporated into the biointerface membrane.

[0276] Angiogenic agents may provide a mechanism for promoting inflammation, which is thought to accelerate angiogenesis and wound healing in vivo. In one embodiment, a xenogeneic carrier, such as bovine collagen, whose foreign nature elicits an immune response, stimulates angiogenesis and is incorporated into the biointerface membrane of some embodiments. In another embodiment, lipopolysaccharide, an immunostimulant, may be incorporated into the biointerface membrane. In another embodiment, proteins, such as bone morphogenetic proteins (BMPs), which regulate bone healing in tissue, may be incorporated into the biointerface membrane.

[0277] In some embodiments, angiogenic agents can be incorporated into the analyte sensor system. Angiogenic agents are substances that can stimulate angiogenesis, for example, accelerating and sustaining the development of a vascularized tissue bed at the tissue-device interface. Angiogenic agents include, but are not limited to, basic fibroblast growth factor (bFGF) (also known as heparin-binding growth factor-II and fibroblast growth factor II), acidic fibroblast growth factor (aFGF) (also known as heparin-binding growth factor-I and fibroblast growth factor-I), vascular endothelial growth factor (VEGF), platelet-derived endothelial growth factor BB (PDEGF-BB), angiopoietin-1, transforming growth factor beta (TGF-β), transforming growth factor alpha (TGFα), hepatocyte growth factor, and tumor necrosis factor-α. (TNFα), placental growth factor (PLGF), angiogenin, interleukin-8 (IL-8), hypoxia inducible factor-I (HIF-I), the angiotensin-converting enzyme (ACE) inhibitor quinaprilat, angiotropin, thrombospondin, the peptide KGHK, low oxygen tension, lactic acid, insulin, copper sulfate, estradiol, prostaglandins, cyclooxygenase inhibitors, endothelial cell binding agents (e.g., decorin or vimentin), glenipin, hydrogen peroxide, nicotine, and growth hormone.

[0278] In some embodiments, a pro-inflammatory agent may be incorporated into the analyte sensor system. Pro-inflammatory agents are generally substances that can stimulate an immune response in host tissue, accelerating or sustaining the formation of a mature vascularized tissue bed. For example, pro-inflammatory agents are generally irritants or other substances that induce chronic inflammation and chronic granular responses at wound sites. Without being bound by theory, it is believed that the formation of high tissue granularity induces blood vessels that provide an adequate or abundant supply of analytes to the device-tissue interface. Pro-inflammatory agents include, but are not limited to, xenogeneic carriers, lipopolysaccharides, S. aureus peptidoglycan, and proteins.

[0279] These bioactive agents can be used alone or in combination. The bioactive agent can be dispersed throughout the material of the sensor, for example, incorporated into at least a portion of the membrane system, or incorporated into the device (e.g., housing) and adapted to diffuse through the membrane.

[0280] There are various systems and methods by which a bioactive agent can be incorporated into a sensor membrane. In some embodiments, the bioactive agent can be incorporated during the manufacture of the membrane system. For example, the bioactive agent can be blended into the membrane system prior to curing or subsequent to membrane system manufacture, for example, by coating, imbibing, solvent casting, or absorbing the bioactive agent into the membrane system. In some embodiments, the bioactive agent is incorporated into the membrane system, while in other embodiments, the bioactive agent can be administered simultaneously with, before, or after in vivo device insertion, for example, by oral administration or locally by subcutaneous injection near the implantation site. A combination of a bioactive agent incorporated into the membrane system and local or systemic bioactive agent administration may be preferred in certain embodiments.

[0281] In general, bioactive agents can be incorporated into the membrane system or can be incorporated into the device and adapted to diffuse from it to modify the host's in vivo response to the membrane. In some embodiments, the bioactive agent can be incorporated into only a portion of the membrane system adjacent to the sensing region of the device, over the entire surface of the device except over the sensing region, or any combination thereof, which can serve to control different mechanisms or stages of the in vivo response (e.g., embolization). However, in some alternative embodiments, the bioactive agent can be incorporated into the device proximal to the membrane system, such that the bioactive agent diffuses through the membrane system into the host's circulatory system.

[0282] The bioactive agent comprises a carrier matrix, where the matrix comprises one or more of collagen, a particle matrix, an absorbent or non-absorbent matrix, a controlled-release matrix, or a gel. In some embodiments, the carrier matrix comprises a receptor, where the bioactive agent is encapsulated in microcapsules. The carrier matrix may comprise a system in which the bioactive agent is physically entrapped within a polymer network. In some embodiments, the bioactive agent is crosslinked with the membrane system, while in other embodiments, the bioactive agent is absorbed into the membrane system by, for example, adsorption, absorption, or wicking. The bioactive agent may be deposited in or on the membrane system by, for example, coating, filling, or solvent casting. In certain embodiments, the bioactive agent is incorporated into the membrane system using ionic and nonionic surfactants, detergents, micelles, emulsifiers, demulsifiers, stabilizers, aqueous and oil-based carriers, solvents, preservatives, antioxidants, or buffers.

[0283] In some embodiments, the surface of the membrane system comprises a tie layer found on the outermost surface of the sensor membrane to which the bioactive agent reversibly binds. In some embodiments, the tie layer comprises one or more zwitterionic compounds, or precursors or derivatives thereof, which are bound to surface-active groups of a polymer comprising the outermost domain of the membrane system. In some embodiments, the zwitterionic compound, or precursors or derivatives thereof, comprises one or more zwitterionic betaines, as described above. In some embodiments, the zwitterionic compound, or precursors or derivatives thereof, comprises a hydrolyzable cation ester of a zwitterionic compound, as described above. In a preferred embodiment, the tie layer comprises one or more hydrolyzable cation betaine esters, such as hydrolyzable cation pCB esters.

[0284] Bioactive agents can also be incorporated into polymers using techniques such as those described above, and the polymers can be used to form membrane systems, coatings on membrane systems, parts of membrane systems, or any part of sensor systems.

[0285] The membrane system can be manufactured using techniques known in the art. The bioactive agent can be absorbed into the membrane system, for example, by soaking the membrane system for a length of time (e.g., from about 1 hour or less to about 1 week, or more preferably from about 4, about 8, about 12, about 16, or about 20 hours to about 1, about 2, about 3, about 4, about 5, or about 7 days).

[0286] The bioactive agent can be blended into the uncured polymer prior to forming the membrane system, which is then cured, thereby crosslinking or encapsulating the bioactive agent within the polymer that forms the membrane system.

[0287] In yet another embodiment, microspheres are used to encapsulate bioactive agents. Microspheres can be formed of biodegradable polymers, most preferably synthetic or natural polymers, such as proteins and polysaccharides. As used herein, the term "polymer" refers to both synthetic polymers and proteins. U.S. Patent No. 6,281,015 discloses several systems and methods that can be used in conjunction with preferred embodiments. Generally, bioactive agents can be incorporated into (1) the polymer matrix that forms the microspheres, (2) microparticle(s) surrounded by the polymer that forms the microspheres, (3) a polymer core within a protein microsphere, (4) a polymer coating around a polymer microsphere, (5) mixed with microspheres that are aggregated into larger forms, or (6) a combination thereof. Bioactive agents can be incorporated as particles or by co-dissolving the agent with the polymer. Stabilizing agents can be incorporated by adding the stabilizer to the agent solution prior to the formation of the microspheres.

[0288] The bioactive agent may be incorporated into, coated on, or otherwise deposited within or on the membrane system. Some hydrogels suitable for use in preferred embodiments comprise a crosslinked, hydrophilic, three-dimensional polymer network that is highly permeable to the bioactive agent and can be triggered to release the bioactive agent upon stimulation.

[0289] The bioactive agent can be incorporated into the membrane system by solvent casting, where a solution containing the dissolved bioactive agent is deposited onto the surface of the membrane system, and the solvent is then removed to form a coating on the membrane surface.

[0290] The bioactive agent may be compounded into a plug of material that is placed within the device, as described in U.S. Patent Nos. 4,506,680 and 5,282,844. In some embodiments, it is preferable to place this plug beneath a membrane system; in this way, the bioactive agent is controlled by diffusion through the membrane, providing a mechanism for sustained release of the bioactive agent in the host.

[0291] Bioactive Agent Release Many variables can affect the pharmacokinetics of bioactive agent release. Bioactive agents of preferred embodiments may be optimized for short-term or long-term release. In some embodiments, bioactive agents of preferred embodiments are designed to aid or overcome factors associated with short-term effects of sensor insertion (e.g., acute inflammation or embolism). In some embodiments, bioactive agents of preferred embodiments are designed to aid or overcome factors associated with long-term effects, such as chronic inflammation or the buildup of fibrous tissue or plaque material. In some embodiments, bioactive agents of preferred embodiments combine short-term and long-term release to take advantage of the benefits of both.

[0292] As used herein, "controlled," "sustained," or "prolonged" release of a factor can be continuous or intermittent, linear or non-linear. This can be achieved using one or more polymer compositions, drug loading, selection of excipients or degradation-enhancing agents, or other modifications, administered alone, in combination, or sequentially to produce the desired effect.

[0293] Short-term release of a bioactive agent in preferred embodiments generally refers to release over a period of about a few minutes or hours to about 2, about 3, about 4, about 5, about 6, or about 7 days or more.

[0294] Bioactive agent loading The amount of bioactive agent loaded into the membrane system can depend on several factors. For example, the dosage and duration of bioactive agent administration can vary depending on the intended use of the membrane system, e.g., the intended duration of device use, patient-to-patient differences in effective doses of the bioactive agent, the location and method of loading the bioactive agent, and the release rate associated with the bioactive agent and, optionally, its carrier matrix. Therefore, one of ordinary skill in the art will appreciate the variability in bioactive agent loading levels for the reasons discussed above.

[0295] In some embodiments in which a bioactive agent is incorporated into a membrane system without a carrier matrix, the preferred level of bioactive agent loading into the membrane system can vary depending on the nature of the bioactive agent. The loading level of the bioactive agent is preferably high enough to observe a biological effect (e.g., embolic protection). Above this threshold, the bioactive agent can be loaded into the membrane system to assimilate up to 100% of the solid portion, coat all accessible surfaces of the membrane, or fill up to 100% of the accessible cavity space. Typically, the loading level (based on the weight of the bioactive agent(s), membrane system, and other materials present) is from about 1 ppm or less to about 1000 ppm or more, preferably from about 2, about 3, about 4, or about 5 ppm up to about 10, about 25, about 50, about 75, about 100, about 200, about 300, about 400, about 500, about 600, about 700, about 800, or about 900 ppm. In certain embodiments, loading levels can be from about 1% or less up to about 50% or more by weight, preferably from about 2, about 3, about 4, about 5, about 6, about 7, about 8, about 9, about 10, about 15, or about 20% by weight up to about 25, about 30, about 35, about 40, or about 45% by weight.

[0296] When a bioactive agent is incorporated into the membrane system with a carrier matrix, such as a gel, the gel concentration can be optimized, e.g., loaded with one or more test loads of bioactive agent. Generally, the gel contains about 0.1% or less to about 50% or more by weight of bioactive agent(s), preferably about 0.2, about 0.3, about 0.4, about 0.5, about 0.6, about 0.7, about 0.8, or about 0.9% by weight to about 6, about 7, about 8, about 9, about 10, about 15, about 20, about 25, about 30, about 35, about 40, or about 45% by weight or more of bioactive agent(s), more preferably about 1, about 2, or about 3% by weight to about 4 or about 5% by weight of bioactive agent(s). Non-bioactive substances can also be incorporated into the matrix.

[0297] Referring now to microencapsulated bioactive agents, release of the agent from these polymer systems generally occurs by two different mechanisms. The bioactive agent can be released by diffusion through aqueous fill channels created in the dosage form by dissolution of the agent, or by voids created by removal of the polymer solvent or pore-forming agent during the original microencapsulation. Alternatively, release can be enhanced due to degradation of the encapsulating polymer. Over time, the polymer erodes, creating more porosity and microstructure within the device. This creates additional pathways for release of the bioactive agent.

[0298] In some embodiments, the sensor is designed to be bioinert, for example, by the use of a bioinert material. The bioinert material does not substantially provoke any response from the host. As a result, cells may live adjacent to the material but do not form bonds with it. Bioinert materials include, but are not limited to, alumina, zirconia, titanium oxide, or other bioinert materials commonly used in the "catheter / catheterization" arts. Without being bound by theory, it is believed that the inclusion of a bioinert material within or on the sensor may reduce adhesion of blood cells or proteins to the sensor, embolization, or other host responses to the sensor. [Example]

[0299] Example 1: Synthesis of enzyme layer polymer and its characterization Betaine-containing polyurethaneurea polymers were synthesized via a two-step polycondensation reaction in organic solvent. In the first step, a homogeneous polyurethane prepolymer with isocyanate end groups on both chains was prepared. In the second step, small molecule diamine(s) were used as chain extender(s). These diamines reacted with the prepolymer in dilute organic solution to yield well-defined polyurethaneureas with linear structures and narrow molecular weight profiles.

[0300] As a representative example, a prepolymer was prepared by adding isophorone diisocyanate (IPDI), polyethylene oxide diol, polycarbonate diol, 2,2-bis(hydroxymethyl)propionic acid (Bis-MPA), and sulfobetaine prepolymer to a dry 200 mL reaction bottle equipped with a nitrogen inlet and mechanical stirrer at room temperature. The reaction mixture was heated to 65°C under nitrogen for 30 minutes with mechanical stirring (200 rpm) until all reactants were dissolved. 400 ppm of catalyst was added to the reaction mixture, which was then held at 65°C for 1 hour. The reaction temperature was increased to 85°C and held for 3 hours until no bubbles were observed in the reaction mixture. The reaction mixture was stirred at 100°C for an additional 2 hours to complete prepolymer formation. The viscous prepolymer was cooled to 50°C and dissolved in ethyl acetate to form a clear solution.

[0301] For the chain extension step, isophorone diamine was used as the chain extender and added to a dry 700 mL reaction bottle equipped with a mechanical stirrer and diluted with an ethyl acetate / isopropanol solvent mixture. The polyurethane prepolymer solution was added dropwise to the chain extender solution at room temperature with vigorous stirring. During the addition of the chain extender solution, a certain amount of solvent mixture (ethyl acetate / isopropanol) was added to the reaction mixture to maintain the reaction mixture at a suitable viscosity. After all the prepolymer solution had been added to the reaction mixture, the reaction mixture was continued stirring at room temperature for an additional 5 hours to complete the chain extension. The polymer thus formed was dried in an oven at 50 °C under a nitrogen flow to remove the solvent and then differentially dissolved or dispersed in a water-borne polyurethane dispersion along with the enzyme and, optionally, a crosslinker. The enzyme layer film was cast and dried at 50 °C for further characterization.

[0302] An active enzyme leaching assay, which measures enzyme activity, was used to determine the amount of active enzyme leaching from the film. The film was immersed in solution, and aliquots of the leachate were measured at specific time points for enzyme activity. Enzyme activity was determined by the rate of hydrogen peroxide produced in the presence of excess glucose. A reactive dye, coupled with peroxidase, was quantitatively converted by hydrogen peroxide to a colored compound, which was monitored spectrophotometrically. The rate of colorimetric change correlated with the activity of the sample, which reflects the active enzyme loading. Experiments were performed on 200 μm-thick films that were dried overnight in a convection oven at 50°C.

[0303] Figure 4 shows the percentage of enzyme leached, subtracted from the baseline, from a control film (P3) and a film prepared from the same polymer binder as used in P3 but containing 30 wt% betaine-containing polymer as the enzyme immobilization polymer additive. The results demonstrate effective GOX enzyme immobilization by the betaine-containing polymer.

[0304] The sensors were evaluated for cal-check metrics, including sensitivity, baseline signal, oxygen sensitivity, linearity, and acetaminophen blockade. Figure 5 shows certain sensor metrics (e.g., MARD and glucose gradient) for a sensor formed with the same polymer binder as used in P3 but with an enzyme layer containing 30 wt.% betaine-containing polymer as the enzyme immobilization polymer additive. Their performance under the various metrics is comparable. In this cal-check test, the following characteristics were measured: i. Glucose ramp (pA / mg / dL) - Ordinary least squares linear regression analysis of the electrical response of the sensor when placed in buffer solutions of increasing glucose concentrations. Also referred to as glucose sensitivity. ii. Baseline Equivalent (mg / dL) - mg / dL equivalent of non-glucose related signal iii. MARD (%) - Mean Absolute Relative Difference, a measure of change away from the ideal line iv. Hypoxic Response - defined as the percentage change in electrical response under reduced oxygen conditions (i.e., 0.25±0.05 mg O2 / L) compared to the signal obtained under ambient conditions. Also referred to as oxygen performance. v. Acetaminophen bias - mg / dL equivalent signal from a 2 mg / dL concentration of acetaminophen. Also called glucose equivalent. [Table 1]

[0305] Example 2: Synthesis of enzyme layer polymer and its characterization Betaine-containing polyurethaneurea polymers in aqueous solution were synthesized via a two-step polycondensation reaction in water. In the first step, a homogeneous polyurethane prepolymer with isocyanate end groups on both chains was prepared. In the second step, small molecule diamine(s) were used as chain extender(s). These diamines reacted with the prepolymer in dilute aqueous solution to yield a waterborne polyurethaneurea solution.

[0306] As a representative example, a prepolymer was prepared by adding polyether diol and 2,2-bis(hydroxymethyl)propionic acid to a dry 200 mL reaction bottle equipped with a nitrogen inlet and mechanical stirrer at room temperature. The reaction mixture was heated to 90°C under nitrogen for 30 minutes with mechanical stirring (200 rpm) until all reactants dissolved and a clear liquid was formed. The reaction mixture was cooled to 80°C, and carboxybetaine diol was added and stirred at 80°C for 1 hour. The reaction mixture was cooled to 65°C, and then isophorone diisocyanate (IPDI) was added. 400 ppm of catalyst was added to the reaction mixture, and the reaction mixture was held at 85°C under nitrogen with mechanical stirring for 4 hours. The reaction mixture was neutralized with trimethylamine and then added dropwise to water to form an aqueous prepolymer emulsion.

[0307] For the chain extension step, ethylenediamine was used as the chain extender and added to a 700 mL reaction bottle equipped with a mechanical stirrer and diluted with water. The polyurethane prepolymer aqueous emulsion was added dropwise to the chain extender solution at room temperature with vigorous stirring. After all the prepolymer solution had been added to the reaction mixture, the reaction was continued to stir at room temperature for an additional 5 hours to complete the chain extension.

[0308] In a different assay than that detailed in Example 1, total enzyme leaching from the enzyme layer film was determined using two separate tests. Figure 4 uses the bicinchoninic acid test, which determines total protein content by peptide bond reduction of copper II ions to copper I, with an associated color change of copper I complexing with bicinchoninic acid. This color change is measured via absorbance measurement at 562 nm using UV spectroscopy.

[0309] Total eluted protein was also measured by gel electrophoresis, followed by protein band quantification in Figure 6. Figure 6 shows a comparison of enzyme leaching from film samples prepared from a standard formulation (control, P3), a standard formulation with the addition of a small molecule betaine additive (Ralufon), and a film prepared from a water-borne polyurethane dispersion in which betaine was incorporated into the polymer as a building block, as disclosed in this example. Within 30 minutes at room temperature, the 200 μm-thick enzyme formulation control (P3) sample leached significantly more enzyme (approximately twice as much) than the water-borne polyurethane dispersion betaine film, indicating that the enzyme was immobilized within the film. Furthermore, the addition of an equal amount (3 wt%) of the small molecule sulfobetaine Ralufon to the P3 control formulation failed to improve enzyme retention. The test was continued for 24 hours, and the water-borne polyurethane dispersion film continued to effectively retain the enzyme.

[0310] Water adsorption of films prepared from the enzyme layer solution was performed in water at room temperature on 25-50 μm thick films. Figure 7 shows that both the betaine-containing polymer WB-7 and the standard (P3) enzyme layer films absorbed the majority of the water they took up within the first 5 minutes. However, the control film almost immediately began to leach large amounts of hydrophilic molecules, losing 10 wt% of water after 24 hours. Films prepared from the betaine-containing polyurethane dispersions with built-in betaine had a stable hydration state over time, were more hydrophilic, and absorbed more water than the control enzyme formulation film.

[0311] A half-sensor containing the enzyme layer (a sensor with all components except the resistive layer) was subjected to high heat and humidity treatment (70°C and 95% humidity). A standard resistive layer was then applied after treatment to avoid the effect of the treatment on the resistive layer, and the sensitivity was measured (Figure 8). The data show that higher sensitivity was maintained when a betaine-containing polymer was used in the enzyme layer.

[0312] The linearity was also determined and as shown in Figure 9, after humidity treatment the control (P3) enzyme sensor had poorer linearity compared to the sensor with the betaine-containing polymer in the backbone.

[0313] Sensors WB-9 and WB-14 containing an enzyme layer were prepared. A control sensor (P3) with a standard enzyme layer was also used. After immersion in heated interference solutions with and without glucose, the membrane-coated sensors were transferred from the sensor fixture to a clamp fixture with a rubber pad. A syringe needle was used to transfer the silicone tubing to the rear end of the membrane-coated sensor. The silicone tubing was placed onto the syringe needle, the sensor was inserted into the needle opening (to protect the membrane-coated area on the sensor), and then the silicone tubing was slid over the needle to reach the rear end of the sensor. The silicone tubing had a diameter small enough to be held firmly in place on the sensor. This was repeated for all sensors tested. The clamp with the sensor inserted into the silicone tubing was immersed in the same immersion buffer at the same temperature for 7 minutes. Using a sensor immersion tool, the control P3 and the carboxybetaine water-borne polymer sensors WB-9 and WB-14 were simultaneously pulled at a fixed withdrawal speed, forcing the tip and cutting area of ​​the sensor through the silicone tubing. This force and velocity motion is responsible for folding of the membrane coating on sensors that do not have good layer adhesion. After the pull-out test, the sensors were examined under an optical microscope to identify any obvious membrane delamination. The percentage of sensors that passed the adhesive pull-out test was determined by dividing the number of sensors that failed the test (exhibiting delamination) by the total number of sensors tested and multiplying by 100. The results are shown in Figures 12 and 13. In both cases, the waterborne polymers WB-9 and WB-14 performed better than the standard enzyme layer.

[0314] Suitable methods and devices for use in conjunction with aspects of the preferred embodiments are disclosed in U.S. Patent Nos. 4,757,022, 4,994,167, 6,001,067, 6,558,321, 6,702,857, 6,741,877, 6,862,465, 6,931,327, 7,074,307, 7,081,195, 7,108,778, 7,110,803, 7,134,999, 7,115,109, and 7,121,114. 36,689, U.S. Patent No. 7,192,450, U.S. Patent No. 7,226,978, U.S. Patent No. 7,276,029, U.S. Patent No. 7,310,544, U.S. Patent No. 7,364,592, U.S. Patent No. 7,366,556, U.S. Patent No. 7,379,765, U.S. Patent No. 7,424,318, U.S. Patent No. 7,460,898, U.S. Patent No. 7,467,003, U.S. Patent No. 7,471,972, U.S. Patent No. 7,494,465, U.S. Patent No. 7,497,827, U.S. Patent No. 7,519,408, U.S. Patent No. 7,583,990, U.S. Patent Nos. 7,591,801, 7,599,726, 7,613,491, 7,615,007, 7,632,228, 7,637,868, 7,640,048, 7,651,596, 7,654,956, 7,657,297, 7,711,402, 7,713,574, 7,715,893, 7,761,130, 7,771,352, 7,774,1 45, U.S. Patent No. 7,775,975, U.S. Patent No. 7,778,680, U.S. Patent No. 7,783,333, U.S. Patent No. 7,792,562, U.S. Patent No. 7,797,028, U.S. Patent No. 7,826,981, U.S. Patent No. 7,828,728, U.S. Patent No. 7,831,287, U.S. Patent No. 7,835,777, U.S. Patent No. 7,857,760, U.S. Patent No. 7,860,545, U.S. Patent No. 7,875,293, U.S. Patent No. 7,881,763, U.S. Patent No. 7,885,697, U.S. Patent No. 7,896,809, U.S. Patent No.899,511, U.S. Patent No. 7,901,354, U.S. Patent No. 7,905,833, U.S. Patent No. 7,914,450, U.S. Patent No. 7,917,186, U.S. Patent No. 7,920,906, U.S. Patent No. 7,925,321, U.S. Patent No. 7,927,274, U.S. Patent No. 7,933,639, U.S. Patent No. 7,935,057, U.S. Patent No. 7,946,984, U.S. Patent No. 7,949,381, U.S. Patent No. 7,955,261, U.S. Patent No. 7,959,569, U.S. Patent No. 7,970,448, U.S. Patent No. 7,974,672, U.S. Patent No. 7,976,492, U.S. Patent No. 7,979,104, U.S. Patent No. 7,986,986, U.S. Patent No. 7,998,071, U.S. Patent No. 8,000,901, U.S. Patent No. 8,005,524, U.S. Patent No. 8,005,525, U.S. Patent No. 8,010,174, U.S. Patent No. 8,027,708, U.S. Patent No. 8,050,731, U.S. Patent No. 8,052,601, U.S. Patent No. 8,053,018, U.S. Patent No. 8,060,173, U.S. Patent No. 8,064,977, U.S. Patent No. 8,070,079 3,519, U.S. Patent No. 8,073,520, U.S. Patent No. 8,118,877, U.S. Patent No. 8,128,562, U.S. Patent No. 8,133,178, U.S. Patent No. 8,150,488, U.S. Patent No. 8,155,723, U.S. Patent No. 8,160,669, U.S. Patent No. 8,160,671, U.S. Patent No. 8,167,801, U.S. Patent No. 8,170,803, U.S. Patent No. 8,195,265, U.S. Patent No. 8,206,297, U.S. Patent No. 8,216,139, U.S. Patent No. 8,229,534, U.S. Patent No. 8,229,535, U.S. Patent Nos. 8,229,536, 8,231,531, 8,233,958, 8,233,959, 8,249,684, 8,251,906, 8,255,030, 8,255,032, 8,255,033, 8,257,259, 8,260,393, 8,265,725, 8,275,437, 8,275,438, 8,277,713, 8,280,475, U.S. Patent No. 8,282,549, U.S. Patent No. 8,282,550, U.S. Patent No. 8,285,354, U.S. Patent No. 8,287,453, U.S. Patent No. 8,290,559, U.S. Patent No. 8,290,560, U.S. Patent No. 8,290,561, U.S. Patent No. 8,290,562, U.S. Patent No. 8,292,810, U.S. Patent No. 8,298,142, U.S. and U.S. Patent Nos. 8,311,749, 8,313,434, 8,321,149, 8,332,008, 8,346,338, 8,364,229, 8,369,919, 8,374,667, 8,386,004, and 8,394,021.

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[0316] Suitable methods and devices for use in conjunction with aspects of the preferred embodiments are disclosed in U.S. application Ser. No. 09 / 447,227, filed Nov. 22, 1999, entitled "DEVICE AND METHOD FOR DETERMINING ANALYTE LEVELS," and U.S. application Ser. No. 13 / 461,625, filed May 1, 2012, entitled "DUAL ELECTRODE SYSTEM FOR A CONTINUOUS ANALYTE SENSOR."

[0317] For ease of explanation and illustration, in some instances, the detailed description describes exemplary systems and methods with respect to a continuous glucose monitoring environment, but it should be understood that the scope of the present invention is not limited to that particular environment, and that those skilled in the art will appreciate that the systems and methods described herein may be embodied in a variety of forms. Accordingly, any structural and / or functional details disclosed herein should not be construed as limiting the systems and methods, but rather are provided as attributes of representative embodiments and / or arrangements that may be advantageous in other settings, to teach those skilled in the art one or more ways to implement the systems and methods.

[0318] For example, but not limited to, the described monitoring systems and methods may include sensors that measure the concentration of one or more analytes (e.g., glucose, lactate, potassium, pH, cholesterol, isoprene, and / or hemoglobin) and / or other blood or bodily fluid constituents in or associated with a host and / or another individual.

[0319] By way of example, and without limitation, embodiments of the monitoring systems and methods described herein may include fingerstick blood sampling, blood analyte test strips, non-invasive sensors, wearable monitors (e.g., smart bracelets, smart watches, smart rings, smart necklaces or pendants, workout monitors, fitness monitors, health and / or medical monitors, clip-on monitors, etc.), adhesive sensors, smart textile and / or garment-integrated sensors, shoe inserts and / or insoles containing sensors, transdermal (i.e., transcutaneous) sensors, and / or swallowed, inhaled, or implantable sensors.

[0320] In some embodiments, without limitation, the monitoring systems and methods may include, instead of or in addition to the sensors described herein, inertial measurement units including accelerometers, gyroscopes, magnetometers, and / or barometers; movement, altitude, position, and / or location sensors; biometric sensors; optical sensors including, for example, optical heart rate monitors, photoplethysmograms (PPG) / pulse oximeters, fluorescence monitors, and cameras; wearable electrodes; electrocardiogram (EKG or ECG), electroencephalography (EEG), and / or electromyography (EMG) sensors; chemical sensors; flexible sensors for measuring, for example, stretch, movement, pressure, weight, or impact; other sensors such as galvanometric sensors, capacitive sensors, electromagnetic field sensors, temperature / heat sensors, microphones, vibration sensors, ultrasonic sensors, piezoelectric / piezoresistive sensors, and / or transducers for measuring information about or relating to the host and / or another individual.

[0321] While the present disclosure has been illustrated and described in detail in the drawings and foregoing description, such illustration and description are considered illustrative or exemplary and not restrictive. The present disclosure is not limited to the disclosed embodiments. Variations to the disclosed embodiments can be understood and effected by those skilled in the art in practicing the claimed disclosure, from a study of the drawings, the disclosure, and the appended claims.

[0322] All references cited herein are incorporated herein by reference in their entirety. To the extent that publications and patents or patent applications incorporated by reference conflict with the disclosure contained herein, the present specification is intended to supersede and / or take precedence over any such conflicting material.

[0323] Unless otherwise defined, all terms (including technical and scientific terms) are to be construed as having their ordinary and customary meanings indicated to those skilled in the art, and are not to be limited to any special or customized meaning unless expressly defined as such herein. It should be noted that the use of a particular term when describing a particular feature or aspect of the present disclosure should not be construed as implying that the term is being redefined herein to be limited to include any particular characteristic of the feature or aspect of the present disclosure to which the term pertains. Particularly in the appended claims, terms and phrases used in this application, and variations thereof, should be construed as open-ended as opposed to limiting, unless expressly stated otherwise. As examples of the above, the term "including" should be interpreted to mean "including without limitation," "including but not limited to," etc.; the term "comprising," when used herein, is synonymous with "including," "containing," or "featuring," is inclusive or open-ended, and does not exclude additional, unrecited elements or method steps; the term "having" should be interpreted as "having at least," the term "including" should be interpreted as "including but not limited to," the term "embodiment" is used to provide illustrative examples of the items under discussion rather than an exhaustive or exclusive list of them, and does not refer to "known," "conventional," "ordinary," "ordinary," "ordinary" or "examples." Adjectives such as "standard," and terms of similar import should not be construed to limit the matter described to that available in a given period or at a given time, but rather should be construed to encompass known, conventional, or standard technology that may be available or known at any time now or in the future; and the use of terms such as "preferably," "preferred," "desired," or "desirable," and terms of similar import, should not be understood to imply that a particular feature is critical, essential, or even important to the structure or function of the invention, but rather should be intended merely to highlight alternative or additional features that may or may not be utilized in a particular embodiment of the invention.Similarly, a group of items joined by the conjunction "and" should not be construed as requiring the presence of every single one of the items in the group, but rather should be construed as "and / or" unless otherwise stated. Similarly, a group of items joined by the conjunction "or" should not be construed as requiring mutual exclusivity between the groups, but rather should be construed as "and / or" unless otherwise stated.

[0324] When a range of values ​​is provided, it is understood that the upper and lower limits, and each intervening value between the upper and lower limits of that range, are encompassed within an embodiment.

[0325] With respect to virtually any plural and / or singular term herein, those skilled in the art can convert from plural to singular and / or from singular to plural as appropriate to the context and / or application. Various singular / plural permutations may be expressly set forth herein for clarity. The indefinite articles "a" or "an" do not exclude plurals. A single processor or other unit may fulfill the functions of several items recited in the claims. The mere fact that certain criteria are recited in mutually distinct independent claims does not indicate that a combination of these criteria cannot be used to advantage. Reference signs in the claims should not be construed as limiting the scope.

[0326] When a specific number is intended in an introduced claim recitation, such intention will be clearly stated in the claim, and it will be further understood by those skilled in the art that, in the absence of such a statement, no such intention exists. For example, to aid in understanding, the following appended claims may include the use of the introductory phrases "at least one" and "one or more" to introduce claim recitations. However, the use of such phrases should not be construed as implying that introducing a claim recitation with the indefinite article "a" or "an" limits a particular claim that includes such an introduced claim recitation to embodiments containing only one such recitation (e.g., "a" and / or "an" should typically be construed to mean "at least one" or "one or more"). This also applies to the use of definite articles used to introduce claim recitations, even when the same claim includes the introductory phrases "one or more" or "at least one" and an indefinite article such as "a" or "an." Additionally, even when a specific number of recitations in an introduced claim is explicitly recited, those skilled in the art will recognize that such recitation should typically be interpreted to mean at least the recited number (e.g., a mere recitation of "two recitations" without other modifiers typically means at least two recitations, or more than two recitations). Furthermore, when a conventional expression similar to "at least one of A, B, or C, etc." is used, such a structure is generally intended in the sense that a person skilled in the art would understand the conventional expression (e.g., "a system having at least one of A, B, or C" includes, but is not limited to, systems having A alone, B alone, C alone, A and B, A and C, B and C, and / or A, B, and C, etc.). When a conventional expression similar to "at least one of A, B, or C, etc." is used, generally such a structure is intended in the sense that one of ordinary skill in the art would understand the conventional expression (e.g., "a system having at least one of A, B, or C" includes, but is not limited to, systems having A alone, B alone, C alone, A and B, A and C, B and C, and / or A, B, and C, etc.).It will be further understood by those skilled in the art that virtually any disjunctive word and / or phrase presenting two or more alternative terms, whether in the description, claims, or drawings, should be understood to contemplate the possibility of including one of the terms, either of the terms, or both terms. For example, the phrase "A or B" is understood to include the possibilities of "A" or "B" or "A and B."

[0327] It is to be understood that all numbers expressing quantities of ingredients, reaction conditions, and so forth used herein are generally modified by the term "about." Accordingly, unless indicated to the contrary, the numerical parameters set forth herein are approximations that may vary depending upon the desired properties sought to be obtained. At the very least, and not as an attempt to limit the application of the doctrine of equivalents to the scope of any claims in any application claiming priority to this application, each numerical parameter should be construed in light of the number of significant digits and ordinary rounding approaches.

[0328] Moreover, while the foregoing has been described in some detail by way of illustration and example for purposes of clarity and understanding, it will be apparent to those skilled in the art that certain changes and modifications may be practiced. Therefore, the descriptions and examples should not be construed as limiting the scope of the invention to the specific embodiments and examples described herein, but rather as embracing all modifications and alternatives that fall within the true scope and spirit of the invention.

Claims

1. 1. A device for measuring an analyte concentration, said device comprising: a sensor configured to generate a signal related to the concentration of an analyte; a sensing membrane overlying the sensor, the sensing membrane comprising an enzyme layer, the enzyme layer comprising an enzyme and a polymer capable of retaining at least 80% of an initial enzyme load when contacted with aqueous conditions for 1 hour at 37°C, the polymer comprising one or more zwitterionic repeat units, the one or more zwitterionic repeat units being formed from a zwitterionic monomer having a diol or a diamine.

2. 10. The device of claim 1, wherein the polymer further comprises one or more ionic or non-ionic emulsion stabilizers selected from carboxylic acid diols, polyethylene oxide diols, polyoxazolines.

3. 10. The device of claim 1, wherein the one or more zwitterionic repeat units are at least about 1 wt. %, based on the total weight of the polymer.

4. 10. The device of claim 1, wherein the polyurethane and / or polyurea segments, excluding zwitterionic repeat units, are 15% to 99% by weight based on the total weight of the polymer.

5. The device of claim 1 , wherein the polymer further comprises a polyethylene oxide segment.

6. 6. The device of claim 5, wherein the polyethylene oxide segments are 5% to 60% by weight based on the total weight of the enzyme layer polymer.

7. The device of claim 1, wherein the polymer has a molecular weight of from 10 kDa to 500,000 kDa.

8. The device of claim 1 , wherein the enzyme is glucose oxidase.

9. 10. The device of claim 1, wherein the enzyme is galactose oxidase, cholesterol oxidase, amino acid oxidase, alcohol oxidase, lactate oxidase, or uricase.

10. The device of claim 1 , wherein the polymer comprises polyurethane and / or polyurea segments.

11. 11. The device of claim 10, wherein the polymer is a polyurethane copolymer selected from polyether-urethane-urea, polycarbonate-urethane, polyether-urethane, silicone-polyether-urethane, silicone-polycarbonate-urethane, and polyester-urethane.

12. 10. The device of claim 1, wherein the polymer comprises a polymer selected from silicone, epoxy, polyolefin, polystyrene, polyoxymethylene, polysiloxane, polyether, polyacrylic acid, polymethacrylic acid, polyester, polycarbonate, polyamide, poly(ether ketone), poly(ether imide).

13. The device of claim 1 further comprising an enzyme stabilizing reagent.

14. 14. The device of claim 13, wherein the enzyme stabilizing reagent comprises one or more zwitterions selected from 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), and poly(sulfobetaine).

15. 14. The device of claim 13, wherein the enzyme stabilizing reagent comprises a biological agent selected from albumin, chitosan, and hyaluronate.

16. 10. The device of claim 1, wherein the polymer is crosslinked with a crosslinking agent, the crosslinking agent comprising a polymer or oligomer selected from a polyfunctional isocyanate, a polyfunctional aziridine, and a polyfunctional carbodiimide.

17. The device of claim 1, wherein the enzyme layer is between 0.01 μm and 250 μm thick.

18. The device of claim 1 , wherein the sensor comprises an electrode.

19. The device of claim 1 , wherein the device is configured for continuous measurement of an analyte concentration.

20. The device of claim 1 , wherein the analyte is glucose.

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