Single-use disposable oxygen sensor

Abstract

The electrochemical oxygen sensor includes a sensing surface having a working electrode and a reference electrode, a hydrophilic layer formed from an oxygen diffusion limiting layer emulsion and overlying the working electrode, and a hydrophobic membrane formed from a hydrophobic solution disposed above the hydrophilic layer. The hydrophilic layer includes an epoxy network and a hydrophilic polymer. The hydrophobic layer includes an acetate copolymer and a crosslinker that reacts with a liquid epoxy resin in the hydrophilic layer to form an epoxy network, and the hydrophobic member is water vapor and oxygen permeable.

Description

Detailed Description of the Invention

[0001] [Background of the invention] 1. FIELD OF THEINVENTION

[0002] The present invention relates generally to planar electrochemical sensors with membrane coatings used to perform chemical analyses, and more particularly to disposable oxygen sensors.

[0003] 2. Description of the Prior Art

[0004] One of the biggest challenges in single-use sensors is the fabrication of electrochemical planar oxygen sensors. Single-use sensor substrates consist of a metal-coated plastic substrate covered with an insulating layer. The electrode dimensions are determined by the opening dimensions in this layer. The dimensions of the oxygen working electrode in this format are usually larger than typical electrochemical oxygen sensors, and upon contact with the sample, they produce high currents that cause sample depletion and produce erratic values. The dimensions of the working electrode are proportional to the amount of current produced, which is based on the Cotterell equation. The Cotterell equation describes the response of current in time as a function of a step in potential. The Cotterell equation is:

number

[0005] Although it is technically possible to fabricate "small electrodes," they are not cost-effective for use as single-use disposable sensors.

[0006] Several different technologies for single-use oxygen sensors have been reported. One approach comes from i-Stat Systems by Abbott Diagnostics. The oxygen sensor uses microfabrication techniques to fabricate small electrodes in a silicon nitride substrate. A multi-step process involves masking, etching, and lifting to fabricate a small-sized oxygen working electrode similar in size to a typical oxygen working electrode. Silicon nitride-based substrates are expensive and require complex processes to fabricate the oxygen sensor described above.

[0007] Another approach comes from Epocal, Inc., Ottawa, Ontario, Canada (a subsidiary of Siemens Healthineers), which introduces a heterogeneous membrane. This membrane consists of a mixture of oil and water fractions. This mixture is emulsified in a cross-linkable polyvinyl alcohol aqueous medium with a salt and redox couple and a cross-linkable hydrophobic polymer to facilitate a manufacturing process using either dosing or printing. The next step requires settling of the deposition layer, degassing, and a final UV cure to immobilize all fractions. This procedure is a complex and time-dependent process due to the phase separation nature of the heterogeneous membrane, which can lead to sensor-to-sensor variability.

[0008] Another related approach discloses a method for determining oxygen concentration in a biological sample. In U.S. Patent No. 7,648,624 (Cai et al., 2010), an oxygen sensor is disclosed that includes a working electrode, a reference electrode, and a reagent matrix disposed on at least the working electrode, the reagent matrix containing a reduced form of a redox mediator, an oxidase, and a peroxidase to obtain an initial oxygen measurement, and the oxygen sensor has a known correlation between oxygen and a predefined analyte, the correlation being expressed as a function of the oxygen sensor's response to both oxygen and the predefined analyte. The process of manufacturing the oxygen sensor is burdensome and time-consuming, which correlates to the reagent and mixing time to provide a solution that can be used to dispense onto the working electrode to form the reagent matrix.

[0009] [Summary of the invention] Disposable arterial blood gas (ABG) sensors have many advantages over conventional blood gas analyzers, such as no maintenance required, ready availability, ease of use, reduced contamination, cost-effectiveness, rapid analysis, and simplicity.

[0010] Because disposable oxygen sensors tend to use relatively large electrode surfaces that produce relatively large currents compared to oxygen electrodes found in conventional blood gas analyzers, an alternative method to produce sufficient "small currents" on a large electrode is to develop a diffusion layer that limits oxygen diffusion from the sample to the electrode. The requirements for the diffusion limiting layer are a fixed path length, electrical conductivity, and linear oxygen permeability when hydrated.

[0011] In the present invention, a multi-layered reagent matrix is ​​formed over a working electrode to fabricate an oxygen sensor. The reagent matrix includes a hydrophilic polymer layer and a hydrophobic polymer layer. The hydrophilic polymer layer is formed by a combination of a liquid epoxy resin and a hydrophilic polymer emulsion, and the combination is used as an oxygen diffusion limiting layer. A hydrophobic polymer layer containing an epoxy hardener is used for a cover membrane and to solidify the liquid epoxy in the hydrophilic polymer layer.

[0012] Liquid epoxy resins, including bisphenol A diglycidyl ether, i.e., epichlorohydrin and bisphenol A, can be made into water-based epoxy emulsions by mixing them with hydrophilic polymers such as PVA, PVP, etc. Epoxy is the material that limits oxygen diffusion, while the water-soluble hydrophilic polymer allows oxygen diffusion and can make electrical connections in the mechanism of electrochemical sensors. This epoxy / water-based hydrophilic polymer mixture results in a homogeneous emulsion that is easy to dispense and creates a uniform film. However, this layer can be washed away once in direct contact with an aqueous solution. The hydrophobic cover membrane prevents this layer from being washed away and also allows the diffusion of water vapor and oxygen across the membrane. Unfortunately, if the liquid epoxy is left uncured, the diffused water vapor will begin to dissolve (i.e., hydrate) the hydrophilic moieties in the layer, eventually increasing the path length (i.e., layer thickness), thereby degrading the sensor performance.

[0013] Liquid epoxies can be cured by reacting with a hardener (e.g., an amine) that turns the water-miscible epoxy into a water-insoluble rigid polymer network. However, epoxy-amines exhibit a very fast curing reaction upon contact with liquid epoxy. Once the epoxy-amine is mixed with the liquid epoxy, the reaction begins immediately. As a result, it is very difficult to add the hardener to the epoxy / hydrophilic polymer emulsion prior to the dosing process.

[0014] The present invention avoids this problem by incorporating an epoxy-amine hardener into the hydrophobic cover membrane solution. The hydrophobic cover membrane solution also contains an acetate copolymer along with the amine hardener. The epoxy-amine hardening reaction begins immediately after dispensing the cover membrane solution on top of the epoxy / hydrophilic polymer layer. As a result of dispensing the hydrophobic cover membrane solution containing the epoxy-amine hardener, the liquid epoxy embedded in the hydrophilic layer emulsion becomes a rigid epoxy network that contains the hydrophilic portion of the layer. The rigid epoxy network formed in the diffusion layer that is coupled with the hydrophobic cover membrane formed above the diffusion layer prevents any change in the path length (i.e., thickness) of the multi-layered reagent matrix upon hydration.

[0015] It is an object of the present invention to provide an electrochemical oxygen sensor that accurately measures dissolved oxygen in small fluid samples. It is another object of the present invention to provide an amperometric oxygen sensor that is disposable and measures oxygen with high accuracy and precision.

[0016] The present invention achieves these and other objects by providing, in one embodiment, an electrochemical oxygen sensor comprising: a sensing surface having a working electrode and a reference electrode; a hydrophilic layer formed from an oxygen diffusion-limiting layer emulsion, the hydrophilic layer overlying the working electrode, the diffusion-limiting layer containing an epoxy network and a hydrophilic polymer; and a hydrophobic membrane formed from a hydrophobic solution disposed over the hydrophilic layer, the hydrophobic solution containing an acetate copolymer and a crosslinker that reacts with a liquid epoxy resin in the hydrophilic layer to form an epoxy network; film has a hydrophobic membrane, which is water vapor permeable and oxygen permeable.

[0017] In another embodiment of the invention, the hydrophilic polymer is polyvinyl alcohol.

[0018] In another embodiment of the invention, the acetate copolymer of the hydrophobic membrane is an ethylene vinyl acetate copolymer.

[0019] In one embodiment, the epoxy network is formed by reaction between the liquid epoxy in the hydrophilic layer emulsion and the crosslinker from the hydrophobic layer.

[0020] In one embodiment, the electrochemical oxygen sensor is a single-use oxygen sensor.

[0021] In one embodiment, the acetate copolymer is ethylene vinyl acetate, with fifty percent (50%) of the ethylene vinyl acetate polymerized into the ethylene vinyl polymer backbone.

[0022] In another embodiment, a method of forming an electrochemical oxygen sensing device is disclosed that includes providing a sensor body having a base layer with at least two independent electrically conductive pathways and an insulating and reagent holding layer disposed on the base layer, the insulating and reagent holding layer having at least two reagent holding openings, one of the at least two reagent holding openings exposing a portion of one of the at least two independent electrically conductive pathways and another of the at least two reagent holding openings exposing a portion of another of the at least two independent conductive pathways, disposing an oxygen diffusion limiting emulsion including a liquid epoxy and a hydrophilic polymer in one of the at least two reagent holding openings, drying the oxygen diffusion limiting emulsion to form a hydrophilic layer, disposing a cover membrane solution including an acetate copolymer and an epoxy hardener over the hydrophilic layer, and drying the cover membrane solution to form a hydrophobic layer.

[0023] In one embodiment of this method, the oxygen diffusion limited emulsion comprises adding together a plurality of components comprising a predefined amount of a liquid epoxy resin, a predefined amount of polyvinyl alcohol, a predefined amount of a surfactant, and a predefined amount of distilled water, and mixing the plurality of components to form the emulsion.

[0024] In another embodiment of this method, the method includes measuring 1.6 grams of liquid epoxy resin, measuring 1.4 grams of 10% polyvinyl alcohol, and measuring a volume of 1 milliliter of distilled water.

[0025] In one embodiment, the method further comprises measuring a predefined amount of an antifoam agent and adding the antifoam agent to the hydrophilic emulsion.

[0026] In one embodiment, the method includes forming a cover film solution, where forming the cover film solution includes adding together a plurality of cover film components including 50 wt % acetate copolymer, 3 wt % epoxy hardener, and 1 wt % pentaerythritol tetrakis 3-mercaptopropionate, and mixing the plurality of cover film components in a predefined amount of THF / cyclohexanone to form the cover film solution.

[0027] In one embodiment, the method includes selecting 2% 2,4,6-tris(dimethylaminomethyl)phenol as the epoxy hardener.

[0028] In one embodiment, the method further includes forming a reference electrode in another of the at least two reagent holding openings.

[0029] In another embodiment, a reagent matrix for making a working electrode into an oxygen sensor is disclosed, the reagent matrix comprising: a hydrophilic layer formed from an oxygen diffusion limiting layer emulsion, the hydrophilic layer overlying the working electrode, the diffusion limiting layer containing an epoxy network and a hydrophilic polymer; and a hydrophobic membrane formed from a hydrophobic solution disposed above the hydrophilic layer, the hydrophobic solution containing an acetate copolymer and a crosslinker that reacts with the liquid epoxy resin in the hydrophilic layer to form an epoxy network; film The membrane comprises a hydrophobic membrane, which is water vapor permeable and oxygen permeable.

[0030] In one embodiment, the epoxy network is formed by reaction between the liquid epoxy in the hydrophilic layer emulsion and the crosslinker from the hydrophobic layer. [Brief description of the drawings]

[0031] [Brief description of the drawings] [Figure 1] FIG. 1 is a perspective view of one embodiment of the present invention showing an oxygen sensor. [Diagram 2] FIG. 2 is an exploded view of the embodiment of FIG. 1 showing the two component layers of the oxygen sensor. [Diagram 3] FIG. 2 is a top view of the base layer of the oxygen sensor. [Figure 4] FIG. 2 is a top view of a reagent holding layer. [Diagram 5] FIG. 5 is an enlarged cross-sectional view of the oxygen sensor taken along line 5-5 of FIG. [Figure 6] FIG. 2 is an enlarged view of the multi-layer reagent matrix of the working electrode showing the hydrophilic and hydrophobic layers. [Figure 7] FIG. 2 is an exemplary top view of an oxygen sensor connected to a flow cell. [Figure 8] 1 is a graphical representation showing the correlation between the readings of an oxygen sensor of the present invention and readings from a blood gas analyzer using a tonometer measured blood sample. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0032] [Detailed Description of the Invention] 1-8 illustrate some embodiments of the present invention. In one embodiment, an oxygen sensor 10 of the present invention is made using a two-layer structure (see FIGS. 1-4). The two-layer structure has a laminate 12 including an electrode end 14, an electrical contact end 16, at least a counter electrode 17, a working electrode 18, and a reference electrode 19 at the electrode end 14, and electrical contact pads 16a, 16b, and 16c at the electrical contact end 16. The laminate 12 also includes a base layer 20 and an insulating and electrode boundary definition layer 30. All layers of the laminate 12 are made of a dielectric material, preferably a plastic. Examples of preferred dielectric materials are polyvinyl chloride, polycarbonate, polysulfone, nylon, polyurethane, nitrocellulose, cellulose propionate, cellulose acetate, cellulose acetate butyrate, polyester, polyimide, polypropylene, polyethylene, polystyrene, and the like.

[0033] The base layer 20 has an electrically conductive layer 21 having at least three electrically conductive pathways 22, 24, and 26 defined thereon. The electrically conductive pathways 22, 24, and 26 may be formed by scribing or scoring the electrically conductive layer 21 or by silk screening the electrically conductive pathways 22, 24, and 26 onto the base layer 20. Scribing or scoring the electrically conductive layer 21 may be performed by mechanically scribing the electrically conductive layer 21 to produce non-electrically conductive score lines 28 sufficient to produce at least three independent conductive pathways 22, 24, and 26. A preferred scribing or scoring method of the present invention is by using a CO2 laser, a YAG laser, or an excimer laser. The electrically conductive layer 21 may be made of any electrically conductive material, such as, for example, copper, gold, tin oxide / gold, palladium, other precious metals or their oxides, or a carbon film composition. The electrically conductive material used in this embodiment is palladium. Acceptable thicknesses for base layer 20 are in the range of 0.002 inch (0.05 mm) to 0.010 inch (0.25 mm). One such usable material for base layer 20 is a 0.005 inch (0.125 mm) palladium polyester film (Stock Number: Melinex 329) sold by Marian, Inc. of Indianapolis, Indiana.

[0034] Insulation and electrode demarcation layer 30 has at least three openings 32, 34, and 36. Opening 32 exposes a portion of conductive pathway 22, opening 34 exposes a portion of conductive pathway 24 to create a reagent holding well, and opening 36 exposes a portion of conductive pathway 26. In this embodiment, insulation and electrode demarcation layer 30 is a medical grade single-sided adhesive tape / film available from Transcendia, Inc. of Franklin Park, Illinois. Acceptable thicknesses of this tape for use in the present invention are in the range of about 0.001 inch (0.025 mm) to about 0.005 inch (0.13 mm). One such tape / film, stock number PE31280 (about 0.002 inch (0.045 mm)), is used because of its ease of handling and good performance in terms of its ability to hold sufficient amounts of chemical reagents and promote capillary action through the fluid sample flow path of the sensor. It should be understood that the use of tape is not required: insulation and electrode demarcation layer 30 may be made from a plastic sheet and coated with a pressure sensitive adhesive, a photopolymer, ultrasonically bonded to base layer 20, silk screened onto base layer 20, or 3D printed onto base layer 20 to achieve the same results as the use of the polyester tape mentioned.

[0035] Three openings 32, 34 and 36 define electrode areas C, W and R, respectively, forming a counter electrode C, a working electrode W and a reference electrode R. Generally, the working electrode W is loaded with a hydrophilic polymer layer deposited directly on the portion of the conductive layer 21 exposed in the electrode area W and a hydrophobic polymer layer on top of the hydrophilic polymer layer, the hydrophobic polymer layer forming a cover membrane.

[0036] The counter, working, and reference electrodes are each in electrical contact with a separate conductive pathway 22, 24, and 26, respectively, that terminates on the end of the laminate 12 opposite the electrode end 14 and is exposed for making electrical connection to a reading device.

[0037] The dimensions of the reagent holding openings are preferably as small as possible to allow the oxygen sensor to have a fluid sample flow path as short as possible while still retaining enough chemical reagent to function properly. The shape of the reagent holding openings in this embodiment is round and has a diameter of about 0.03 inches (0.75 mm). The two reagent holding openings 32 and 34 are aligned with each other and spaced about 0.0256 inches (0.65 mm) apart from each other. These circular reagent holding openings are for illustrative purposes only. It should be understood that the shape of the reagent holding openings is not critical and the dimensions of the openings are more dictated by the technical feasibility of dispensing the reagent matrix mixture into the openings and other manufacturing constraints.

[0038] The positional arrangement of the counter, working and reference electrodes is not critical to obtain usable results from the oxygen sensor. The possible electrode arrangements when the oxygen sensor is coupled to a flow cell may be CWR, WCR or any arrangement of the three electrodes, and the arrangements listed for the electrode arrangements appear based on the direction of sample flow across the counter electrode C, the working electrode W and the reference electrode R. The preferred position has been found to be CWR. That is, as the fluid sample enters the flow cell 70, the fluid sample first covers the counter electrode C, then the working electrode W and then the reference electrode R.

[0039] Preferably, the reference electrode 19 (electrode well 36) is loaded with an Ag / AgCl layer (e.g., by applying an Ag / AgCl ink, or by (a) sputter coating a layer of Ag and then chlorinating the Ag, or (b) sputter coating a layer of Ag / AgCl), or other reference electrode material that does not require a redox mediator to function properly. It should be noted that the positional arrangement of the working, reference, and counter electrodes within the flow path is not critical to obtaining usable results from the sensor.

[0040] 3 and 4, a top view of the base layer 20 and the insulating and reagent holding layer 30 are illustrated. As illustrated in FIG. 3, the symmetry of the conductive pathways is such that one longitudinal end of the base layer 20 may be designated as either an electrode end 14 or an electrical contact end 16, depending on the orientation of the insulating and reagent holding layer 30 relative to the base layer 20 and the assembly process. In this embodiment, the base layer 20 has scribe marks in the conductive layer 21 that demarcate three separate conductive pathways. It should be understood that the base layer may have more than two conductive pathways and additional conductive pathways may be designated for similar or other analyte sensor reagents, making this oxygen sensor a multi-analyte sensor.

[0041] FIG. 4 is a top view of the insulating and reagent holding layer 30. The insulating and reagent holding layer 30 has three or more openings spaced apart from one another such that each opening coincides with one of the conductive paths defined on the base layer 20. It should be understood that the electrically conductive paths disclosed herein may be made of any non-corroding metal. Carbon deposits, such as carbon paste or carbon ink, may also be used as the electrically conductive paths, all of which are well known to those skilled in the art. It should also be understood that the counter electrode C is used to pass current from the working electrode W to the counter electrode C. A two-electrode system (i.e., a working electrode and a reference electrode) may be used, but the use of a relatively large working electrode has disadvantages in this case compared to conventional oxygen sensors. The challenge is that the reference electrode in an amperometric system is required to maintain a high charge density. In a two-electrode system, when there is a high current flow from the working electrode to the reference electrode, the Ag-AgCl reference electrode may deteriorate due to the changing current density. This in turn causes a shift in the bias potential window and a decrease in the accuracy of the determined oxygen concentration level.

[0042] Turning now to Figure 5, there is illustrated an enlarged cross-sectional view of oxygen sensor 10 taken along line 5-5 of Figure 1. It should be understood that the relative dimensions of layers 20 and 30 and electrode wells 32, 34, and 36, and the thicknesses of the working and reference electrode reagent matrices, are not drawn to scale, but are merely for the purpose of illustrating the various components of oxygen sensor 10. As can be seen in Figure 5, base layer 20 has an electrically conductive layer 21 disposed thereon, with scoring lines 28 creating non-electrically conductive breaks in conductive layer 21. Insulating and reagent-retaining layer 30 has reagent-retaining opening 34 containing working electrode oxygen diffusion limiting multilayer reagent matrix 34a, and reagent-retaining opening 36 containing reference electrode reagent matrix 60.

[0043] FIG. 6 shows a multi-layer reagent matrix 3 4 FIG. 1 is an enlarged view of multi-layer reagent matrix 3. 4 a includes a hydrophilic polymer layer 50 and a hydrophobic polymer layer 40. The hydrophilic layer 50 includes an epoxy network 52 and a hydrophilic portion 54. The hydrophobic layer 40, as the name suggests, is not water soluble but is water vapor and oxygen permeable.

[0044] The polymer used as the hydrophilic moiety in the hydrophilic layer should be sufficiently water-soluble and also be capable of stabilizing all other chemicals in the reagent to the conductive surface layer in the electrode area. Suitable polymers include, but are not limited to, low and high molecular weight polyethylene oxide (PEO), polyethylene glycol, polyvinylpyrrolidone (PVP), starch, methyl cellulose, hydroxypropyl cellulose, polyvinyl alcohol (PVA), carboxymethyl cellulose (CMC), and polyamino acids. The hydrophilic moiety can be a single polymer or a combination of polymers, preferably in a concentration range of about 0.02% (w / w) to about 7.0% (w / w). The preferred hydrophilic moiety in the hydrophilic layer of the present invention is PVA. PVA is available from Scientific Polymer Products, New York, USA.

[0045] The hydrophilic layer also contains an epoxy network that is not water soluble, however, the epoxy network is made from a hydrophilic emulsion that contains a water soluble liquid epoxy resin.

[0046] The polymer used in the hydrophobic layer is an ethylene vinyl acetate copolymer, which is 50 wt% vinyl acetate, available from Scientific Polymer Products, New York, USA.

[0047] A surfactant may be optionally included in the hydrophilic emulsion (used to fabricate the hydrophilic layer) to facilitate dispensing of the hydrophilic emulsion into the working electrode area W. The surfactant also aids in the rapid dissolution of the dry chemical reagent (i.e., PVA) as the fluid enters the sample chamber 17 of the oxygen sensor 10, 430. The amount and type of surfactant is selected to ensure the previously mentioned functions. The surfactant may be selected from, but is not limited to, a variety of anionic, cationic, nonionic, and zwitterionic detergents. Examples of acceptable surfactants are polyoxyethylene ether, Tween 20, sodium cholate hydrate, hexadecylpyridinium chloride monohydrate, and CHAPs. A preferred surfactant is polyoxyethylene ether. More preferably, it is t-octylphenoxypolyethoxyethanol, available from Sigma-Aldrich under the brand name TritonX-100. The concentration of the surfactant in the reagent matrix is ​​preferably from about 0.01% (w / w) to about 2%.

[0048] 7 is a top view representation of the oxygen sensor 10 connected to a flow cell 70. The flow cell 70 has a test chamber 74 in which the working electrode 18 and the reference electrode 19 are disposed. The test chamber 74 has a test chamber inlet 72 connected to a calibrant supply line 90 and a blood sample supply line 100. A predefined amount of calibrant is supplied to the test chamber 74 for a single point calibration. After calibration, the calibrant is moved out of the test chamber 74, followed by a predefined amount of blood sample. The blood sample is measured for the amount of oxygen (i.e., oxygen concentration) in the blood sample. It will be understood that the oxygen sensor 10 is electrically connected to appropriate electronic circuitry to perform chronoamperometric measurements.

[0049] [Preparation of Hydrophilic Layer Composition and Hydrophobic Layer Composition]

[0050] The preferred reagent layer composition for the hydrophilic emulsion used in fabricating the hydrophilic layer may be prepared in one step, but is preferably prepared in two steps.

[0051] Step 1: Add together 1.6 gm of liquid epoxy resin available under the trademark DER331 by The Dow Chemical Company (DER331), 1.4 gm of 10% polyvinyl alcohol (Mw. 130K), 1 ml of 1% Triton X-100 and distilled water, and 100 mg of antifoam agent.

[0052] Step 2: Mix all the ingredients in step 1 above using a homogenizer at 9,000 rpm for 1 minute. The prepared hydrophilic emulsion is stable for several days when stored at room temperature.

[0053] The reagent layer composition for the cover membrane solution used in fabricating the hydrophobic layer may also be prepared in one step, but is preferably prepared in two steps.

[0054] Step 1: Add 2 wt% of ethylene vinyl acetate copolymer, where the copolymer is ethylene vinyl acetate with 50% acetate copolymerized into the ethylene vinyl polymer backbone, 3% epoxy hardener (2% 2,4,6-tris(dimethylaminomethyl)phenol and 1% pentaerythritol tetrakis(3-mercaptopropionate)) in THF / cyclohexanone.

[0055] Step 2: The raw materials in step 1 are mixed together to form a cover membrane solution. The prepared cover membrane solution is stable for several weeks when stored at room temperature.

[0056] [Sensor structure]

[0057] Assembly of the various embodiments of the present invention is relatively simple: generally, the base layer and the insulating and reagent holding layer are laminated together, and then the appropriate reagent mixture is dispensed into each of the reagent holding openings.

[0058] More specifically, for the two-layer configuration shown in FIG. 1, a strip of palladium film is cut into the shape illustrated in FIG. 2 to form the base layer 20 of the sensor 10. The conductive palladium polyester film is scored, preferably using a laser, although mechanical scribing is optional. As illustrated in FIG. 2, the film is scored with a laser so that three electrode areas are formed at the sample fluid end 14 and three contact points 22, 24, and 26 are formed at the electrical contact end 16. The score lines are very thin, but sufficient to create three separate and distinct electrically conductive paths. A strip of single-sided adhesive tape is then cut to size and shape to form the insulating and electrode demarcation layer 30, such that the tape covers most of the conductive layer 21 of the base layer 20 except for exposing a small electrical contact area illustrated in FIG. 1 by reference numeral 16.

[0059] Prior to attaching the insulating and electrode boundary definition layer 30 to the base layer 20, at least three openings 32, 34, and 36 having substantially equal dimensions are punched by a laser or by mechanical means such as a die punch assembly to produce the electrode openings 32, 34, and 36 in the insulating and electrode boundary definition layer 30. The shape of the electrode openings may be any shape. In the illustrated embodiment, the openings are circular. The preferred hole size for the openings 32 and 34 has a typical diameter of about 0.030 inches (0.75 mm), but may be any size. As illustrated in FIG. 2, the electrode openings 32 and 34 (and optional electrode opening 36, if included) are aligned with one another and have a spacing between adjacent openings of about 0.020 inches (0.508 mm) to about 0.050 inches (1.27 mm). The circular openings are for illustrative purposes only. It should be understood that the shape and size of the openings, or the distance between the openings, is not critical. The circular openings do not need to be substantially equal in size so long as the surface area ratio remains substantially constant. Although the arrangement of electrodes can be any combination, a preferred arrangement of electrodes formed within openings 32, 34, and 36 is C (counter electrode), W (working electrode), and R (reference electrode) as positioned from test chamber inlet 72. An insulating and electrode boundary definition layer 30 is then attached to base layer 20 in a manner that defines electrode wells for fabricating counter electrode C, working electrode W, and reference electrode R.

[0060] A predefined amount of hydrophilic emulsion is dispensed into the working electrode well and allowed to dry. The hydrophilic emulsion may be air-dried for a few minutes at room temperature or dried for 30 seconds at 37° C. to form a hydrophilic layer. A shorter period of drying above room temperature allows for a more efficient manufacturing process. The hydrophilic emulsion and its composition are described above.

[0061] The cover membrane solution is then dispensed onto the hydrophilic layer, allowing the solution to completely cover the hydrophilic layer, and either air dried overnight at room temperature or dried at 37° C. for 30 seconds or more. During this process, the epoxy hardener in the cover membrane solution reacts with the liquid epoxy resin in the hydrophilic layer to form a rigid epoxy network containing the hydrophilic portion of the layer (i.e., polyvinyl alcohol), which maintains the predefined path length (i.e., thickness) of the hydrophilic layer when it is hydrated with the calibrant. As previously discussed, the hydrophobic layer allows the diffusion of water vapor and oxygen across the cover membrane, while the hydrophilic layer is an oxygen diffusion limiting layer containing a water-soluble hydrophilic polymer that allows oxygen dispersion and also makes an electrical connection to the working electrode.

[0062] The amount of time required to dry the reagents depends on the temperature at which the drying process is carried out.

[0063] [Oxygen sensor test]

[0064] The oxygen sensor 10 was connected to a flow cell as illustrated in Figure 7. When a fluid sample is applied to the oxygen sensor of the embodiment of the invention shown in Figure 1, the fluid sample enters the flow cell 70 and flows over electrodes C, W, and R where it remains for a predefined period of time. The length of time depends on whether the fluid is a calibrant or a blood sample.

[0065] The current response of the oxygen sensor 10 was measured by chronoamperometry using a Nova Biomedical blood gas analyzer, although a potentiostat may also be used. The oxygen sensor shown in FIG. 1 and constructed as described above was used to test the response of the oxygen sensor 10 of the present invention to the concentration of oxygen in a sample. A blood sample containing a predefined amount of oxygen is applied to the sample inlet 18 of the sensor strip and enters the sensor strip while a potential of −0.10 V to −0.70 volts (depending on the reference electrode used) is applied between the working and reference electrodes. In the particular example described here, the applied potential was −0.65 V versus the Ag-AgCl reference electrode. The output response current is proportional to the concentration in the blood sample. Using the Nova Biomedical blood gas analyzer, measurements were given as the partial pressure of oxygen in mmHg.

[0066] [Example 1]

[0067] [Different levels of pO 2 Reference pO 2 pO for sensor 2 [Illustration of

[0068] Different pO 2 Blood samples having a concentration of oxygen (pO) were tested with the oxygen sensor of the present invention using a pHox blood gas analyzer from Nova Biomedical. In the alternative, an electrochemical analyzer (Model 812 from CH Instruments, Austin, Texas, USA) may be used to measure the current response directly from the oxygen sensor strip 10. 2 ) was controlled using a Tonometer (Precision Gas Mixer, PGM-3, Medicor, Inc., Salt Lake City, UT, USA). Two milliliters of blood sample were placed in a temperature-controlled (37°C) cylindrical rotating cuvette and measured by the tonometer for 15 min.

[0069] A single-point calibration prior to measurement of a blood sample is required for hydration of the hydrophilic layer and to establish a reference point for the oxygen sensor 10. The single-point calibrator used had a pO 2 The single-point calibrant was a solution. This single-point calibrant was flowed through each oxygen sensor via the flow cell and allowed to reside within the flow cell for 90 seconds to hydrate the hydrophilic layer. At the end of the 90 second hydration period, a single-point calibration measurement is performed. Following calibration, a tonometer-measured blood sample was introduced into the oxygen sensor to replace the calibrant and a reading was taken 30 seconds after blood sample introduction. Concentration calculations were based on the single-point calibration and used the Cotterell equation. Performed in this manner, pO 2 The measurement response is from 50mmHg to 170mmHg pO at the working electrode. 2 The results were found to be linear over a range of 100-243 mmHg. Six blood samples were tested with tonometer-measured oxygen levels ranging from 49 mmHg to 243 mmHg. These tonometer-measured blood samples were also measured using a Nova Biomedical pHox blood gas analyzer to obtain reference readings for each blood sample for comparison with the readings obtained from the oxygen sensor of the present invention.

[0070] In this example, several biosensors using palladium substrates were fabricated to test the response of the oxygen sensor, and the results are shown in Table 1.

[0071] [Table 1]

[0072] Each blood concentration was tested at least ten times (i.e., ten disposable oxygen sensors 10 were used for each oxygen concentration level, and the average values ​​were calculated and displayed in Table 1). Standard deviation values ​​are also provided for each concentration level tested.

[0073] FIG. 8 shows the results of the various pO of the working electrode of the present invention (i.e., the hydrophilic layer / cover membrane layer electrode) of 49 mmHg, 82 mmHg, 103 mmHg, 122 mmHg, 167 mmHg, and 243 mmHg. 2 Measured pO versus level 2 These responses (i.e., current responses) are the first five pO 2 It is linear with respect to oxygen concentration over the entire range.

[0074] Advantages of the present invention over conventional blood gas analyzers include no maintenance, ready availability, ease of use, reduced contamination, cost effectiveness, rapid analysis, simplicity, etc.

[0075] Although preferred embodiments of the invention have been described herein, the above description is merely illustrative. Further modifications of the invention disclosed herein will occur to those skilled in the art, and all such modifications are deemed to fall within the scope of the invention as defined by the appended claims.

Claims

1. a sensing surface having a working electrode and a reference electrode; a hydrophilic layer formed from an oxygen diffusion limiting layer emulsion, the hydrophilic layer overlying the working electrode, the hydrophilic layer comprising an epoxy network and a hydrophilic polymer; and a hydrophobic membrane formed from a hydrophobic solution disposed over the hydrophilic layer, the hydrophobic solution containing an acetate copolymer and a crosslinker that react to form the epoxy network, the hydrophobic membrane being water vapor and oxygen permeable; The electrochemical oxygen sensor comprises:

2. 2. The electrochemical oxygen sensor of claim 1, wherein the hydrophilic polymer is polyvinyl alcohol.

3. 2. The electrochemical oxygen sensor of claim 1, wherein said acetate copolymer of said hydrophobic membrane is an ethylene vinyl acetate copolymer.

4. 2. The electrochemical oxygen sensor of claim 1, wherein the epoxy network is formed by a reaction between a liquid epoxy in the hydrophilic layer and the cross-linker from the hydrophobic membrane.

5. 10. The electrochemical oxygen sensor of claim 1 which is a single-use oxygen sensor.

6. 4. The electrochemical oxygen sensor of claim 3, wherein said acetate copolymer is ethylene vinyl acetate, and 50 percent of said ethylene vinyl acetate is polymerized into an ethylene vinyl polymer backbone.

7. 1. A method of forming an electrochemical oxygen sensor, comprising: providing a sensor body having a base layer with at least two independent conductive pathways, and an insulating and reagent retaining layer disposed on the base layer, the insulating and reagent retaining layer having at least two reagent retaining openings, one of the at least two reagent retaining openings exposing a portion of one of the at least two independent conductive pathways, and another of the at least two reagent retaining openings exposing a portion of another of the at least two independent conductive pathways; disposing an oxygen diffusion limiting emulsion containing a liquid epoxy and a hydrophilic polymer in one of the at least two reagent holding openings; drying the oxygen diffusion limiting emulsion to form a hydrophilic layer; disposing a cover film solution containing an acetate copolymer and an epoxy hardener over the hydrophilic layer; and drying the cover membrane solution to form a hydrophobic layer; Including, the one of the at least two reagent-retaining openings containing the hydrophilic and hydrophobic layers forms a working electrode; method.

8. forming the oxygen diffusion limited emulsion; forming the oxygen diffusion limited emulsion, adding together a plurality of ingredients including a predefined amount of liquid epoxy resin, a predefined amount of polyvinyl alcohol, a predefined amount of a surfactant, and a predefined amount of distilled water; mixing the components to form an emulsion; The method of claim 7, comprising:

9. Measuring 1.6 grams of the liquid epoxy resin; Measure out 1.4 grams of 10% polyvinyl alcohol; and Measuring a volume of 1 milliliter of distilled water; The method of claim 8 , further comprising:

10. 10. The method of claim 8, further comprising adding a predefined amount of an antifoaming agent to the oxygen diffusion limited emulsion.

11. forming the cover membrane solution; forming the cover membrane solution, Adding together a plurality of cover film ingredients including 50 wt % acetate copolymer, 3 wt % epoxy hardener, and 1 wt % pentaerythritol tetrakis 3-mercaptopropionate; and mixing the cover film components in a predefined amount of THF / cyclohexanone to form the cover film solution; The method of claim 7, comprising:

12. The method of claim 11 further comprising selecting 2% 2,4,6-tris(dimethylaminomethyl)phenol as the epoxy hardener.

13. The method of claim 7 , further comprising forming a reference electrode in another of the at least two reagent holding openings.

14. A multi-layer reagent matrix for making a working electrode into an oxygen sensor, comprising: a hydrophilic layer formed from an oxygen diffusion limiting layer emulsion containing a liquid epoxy resin, the hydrophilic layer overlying the working electrode, the hydrophilic layer containing an epoxy network and a hydrophilic polymer; and a hydrophobic membrane formed from a cover membrane solution disposed over the hydrophilic layer, the cover membrane solution containing an acetate copolymer and a crosslinker that reacts with the liquid epoxy resin in the hydrophilic layer to form the epoxy network, the hydrophobic membrane being water vapor and oxygen permeable; The multi-layer reagent matrix comprises:

15. The reagent matrix of claim 14 , wherein the hydrophilic polymer is polyvinyl alcohol.

16. The reagent matrix of claim 14 , wherein the acetate copolymer of the hydrophobic membrane is an ethylene-vinyl acetate copolymer.

17. 1. A method of making a multi-layered reagent matrix that converts a working electrode into an oxygen sensor, comprising: forming an oxygen diffusion limited emulsion for deposition on a working electrode surface, the oxygen diffusion limited emulsion comprising: adding together a plurality of components comprising a predefined amount of a liquid epoxy resin, a predefined amount of polyvinyl alcohol, a predefined amount of a surfactant, and a predefined amount of distilled water, and mixing the plurality of components to form an emulsion; and forming a cover film solution for depositing on the oxygen diffusion limiting emulsion after drying the oxygen diffusion limiting emulsion, the cover film solution comprising: adding together a plurality of cover film components comprising a predefined amount of acetate copolymer, a predefined amount of epoxy hardener, and a predefined amount of pentaerythritol tetrakis 3-mercaptopropionate; and mixing the plurality of cover film components in a predefined amount of THF / cyclohexanone to form the cover film solution; Including, A method of making a multi-layer reagent matrix, wherein the oxygen diffusion limiting emulsion is used to fabricate a hydrophilic layer on the working electrode and the cover membrane solution is used to fabricate a hydrophobic layer on the hydrophilic layer.

18. The method of claim 17, wherein the predetermined amount of liquid epoxy resin is 1.6 grams, the predetermined amount of polyvinyl alcohol is 1.4 grams, and the predetermined amount of distilled water is 1 milliliter.

19. The method of claim 17, wherein the predetermined amount of the acetate copolymer is 50 wt %, the predetermined amount of the epoxy hardener is 3 wt %, and the predetermined amount of pentaerythritol tetrakis 3-mercaptopropionate is 1 wt %.

20. A multi-layer reagent matrix comprising: a hydrophilic layer formed at least in part as an epoxy network containing a hydrophilic polymer; a hydrophobic layer disposed above the hydrophilic layer and containing a crosslinking agent capable of reacting with the liquid epoxy resin of the hydrophilic layer in forming the epoxy network of the hydrophilic layer; The multi-layer reagent matrix comprises:

21. The multi-layer reagent matrix of claim 20, wherein the hydrophilic polymer comprises polyvinyl alcohol.

22. The multi-layer reagent matrix of claim 20, wherein the hydrophobic layer comprises an acetate copolymer.

23. The multi-layer reagent matrix of claim 22, wherein the acetate copolymer comprises ethylene vinyl acetate.

24. The multi-layer reagent matrix of claim 23, wherein 50% of the ethylene vinyl acetate is polymerized into the ethylene vinyl polymer backbone.

25. The multi-layer reagent matrix of claim 20, wherein the hydrophobic layer is water vapor permeable and oxygen permeable.

26. 1. A method of making a multi-layered reagent matrix, comprising: forming a hydrophilic emulsion by combining a predefined amount of a liquid epoxy resin, a predefined amount of a hydrophilic polymer, a predefined amount of a surfactant, and a predefined amount of distilled water; forming a membrane solution for depositing onto said hydrophilic emulsion by combining a predefined amount of acetate copolymer, a predefined amount of epoxy hardener, and a predefined amount of pentaerythritol tetrakis 3-mercaptopropionate. A method for making a multi-layered reagent matrix.

27. The method of claim 26, wherein the step of forming the hydrophilic emulsion further comprises combining a predetermined amount of an antifoaming agent with the liquid epoxy resin, hydrophilic polymer, surfactant, and distilled water.

28. The method of claim 26, wherein the predetermined amount of liquid epoxy resin is 1.6 grams, the predetermined amount of hydrophilic polymer is 1.4 grams, and the predetermined amount of distilled water is 1 milliliter.

29. The method of claim 26, wherein the predetermined amount of the acetate copolymer is 50 wt %, the predetermined amount of the epoxy hardener is 3 wt %, and the predetermined amount of pentaerythritol tetrakis 3-mercaptopropionate is 1 wt %.

30. The method of claim 26, wherein the hydrophilic polymer comprises polyvinyl alcohol.

31. The method of claim 26, further comprising drying the hydrophilic emulsion to form a hydrophilic layer.

32. The method of claim 31, further comprising depositing the membrane solution onto the hydrophilic layer.

33. The method of claim 32, further comprising drying the membrane solution to form a hydrophobic layer over the hydrophilic layer.

34. The method of claim 31, further comprising reacting the liquid epoxy resin with the epoxy hardener to form the hydrophilic layer at least in part as an epoxy network containing the hydrophilic polymer.

35. A method of forming an electrochemical oxygen sensor comprising the multi-layer reagent matrix of claim 26, comprising: providing a sensor body having a base layer with at least two independent conductive pathways and an insulating and reagent retaining layer disposed on the base layer, the insulating and reagent retaining layer having at least two reagent retaining openings, one of the at least two reagent retaining openings exposing a portion of one of the at least two independent conductive pathways and another of the at least two reagent retaining openings exposing a portion of another of the at least two independent conductive pathways; disposing the hydrophilic emulsion in one of the at least two reagent holding openings and drying the hydrophilic emulsion to form a hydrophilic layer; disposing the membrane solution over the hydrophilic layer and drying the membrane solution to form a hydrophobic layer; the one of the at least two reagent-retaining openings containing the hydrophilic layer and the hydrophobic layer forms a working electrode of the electrochemical oxygen sensor. method.

36. A sensing surface having a working electrode and a reference electrode; a hydrophilic layer overlying the working electrode, the hydrophilic layer being formed at least in part as an epoxy network containing a hydrophilic polymer; a hydrophobic layer disposed above the hydrophilic layer, the hydrophobic layer containing a crosslinking agent capable of reacting with a liquid epoxy resin of the hydrophilic layer in forming the epoxy network of the hydrophilic layer; The electrochemical oxygen sensor comprises:

37. The sensing surface has a base layer having at least two independent conductive paths and an insulating and reagent-retaining layer disposed on the base layer; the insulating and reagent retaining layer having at least two reagent retaining openings; one of the at least two reagent holding openings exposing a portion of one of the at least two independent conductive paths and another of the at least two reagent holding openings exposing a portion of another of the at least two independent conductive paths; 37. The electrochemical oxygen sensor of claim 36.

38. The electrochemical oxygen sensor of claim 36, wherein the hydrophilic polymer comprises polyvinyl alcohol.

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