CN1902478A - Method of reducing the effect of direct interference current in an electrochemical test strip - Google Patents

Abstract

The present invention describes a method for reducing the influence of interfering compounds when using an electrochemical sensor (62) to measure an analyte. The method of the present invention is particularly applicable to electrochemical sensors, wherein the sensor (62) includes a substrate (50), first and second working electrodes (10, 12) and a reference electrode (14), and the first and second Both working electrodes or only the second working electrode include regions free of reagent (22). In the present invention, an algorithm is described that mathematically corrects for interferent effects using the test strip embodiments of the present invention.

Description

Method for reducing the effect of direct interfering currents in electrochemical test strips

Field of Invention

The present invention relates generally to methods of reducing the effect of interfering compounds on measurements made by an analyte measurement system, and more particularly, to methods of reducing the effect of directly interfering currents in glucose monitoring systems using electrochemical test strips that The test strip has electrodes with uncoated areas.

Background of the Invention

In many cases, electrochemical measurement systems can have increased oxidation currents due to the oxidation of interfering compounds commonly found in physiological fluids, such as paracetamol, ascorbic acid, bilirubin, dopamine, gentisic acid, glutathione peptides, levodopa, methyldopa, tolazamide, tolbutamide, and uric acid. Thus, by reducing or eliminating the portion of the oxidation current generated by interfering compounds, the accuracy of the glucose meter can be improved. Ideally, there should be no oxidation current due to any interfering compounds, such that the overall oxidation current depends only on the glucose concentration.

Therefore, it is desirable to improve the accuracy of electrochemical sensors in the presence of possible interfering compounds such as ascorbate, urate and paracetamol which are commonly found in physiological fluids. For such electrochemical sensors, examples of analytes may include glucose, lactate, and fructosamine. While glucose is the primary analyte discussed, it will be apparent to those skilled in the art that the present invention is applicable to other analytes as well.

Oxidation currents can be generated through several pathways. In particular, the desired oxidation current results from the interaction of the redox mediator with the analyte of interest (e.g., glucose), whereas the undesired oxidation current typically results from oxidation at the electrode surface and through interaction with the redox mediator. Interfering compounds that are oxidized by the interaction are produced. For example, certain interfering compounds such as paracetamol are oxidized on the electrode surface. Other interfering compounds, such as ascorbic acid, are oxidized through chemical reactions with redox mediators. In a glucose measurement system, oxidation of interfering compounds causes the measured oxidation current to depend on both the concentration of glucose and the concentration of any interfering compounds. Therefore, when the interfering compound oxidizes with the same efficiency as glucose, and the concentration of the interfering compound is high relative to the glucose concentration, the measurement of the glucose concentration can be improved by reducing or eliminating the contribution of the interfering compound to the total oxidation current.

A known strategy to reduce the effect of interfering compounds is to cover the working electrode with a negatively charged thin film. As an example, sulfonated fluoropolymers such as NAFION ^™ can be used to repel all negatively charged chemicals. In general, most interfering compounds such as ascorbate and urate are negatively charged, therefore, the negatively charged film prevents negatively charged interfering compounds from reaching and being oxidized on the electrode surface. However, this technique is not always successful since certain interfering compounds such as paracetamol are not negatively charged and thus can pass through negatively charged membranes. The technique also does not reduce oxidation currents due to the interaction of interfering compounds with certain redox mediators. The use of a negatively charged film on the working electrode also prevents certain commonly used redox mediators such as ferricyanide from exchanging electrons with the electrode through the negatively charged film.

Another strategy that can be used to reduce the effect of interfering compounds is to use a size selective membrane on top of the working electrode. As an example, a 100 Dalton size exclusion membrane, such as a cellulose acetate membrane, can be placed over the working electrode to exclude all chemicals with a molecular weight greater than 100 Daltons. Most interfering compounds have a molecular weight greater than 100 Daltons and are therefore excluded from being oxidized at the electrode. However, such selective membranes generally complicate the manufacture of test strips and increase measurement time since oxidized glucose must diffuse through the selective membrane to reach the electrodes.

Another strategy that can be used to reduce the effect of interfering compounds is to use a redox mediator with a low redox potential, such as a redox mediator with a redox potential of about -300 mV to +100 mV (when measured with respect to a saturated calomel electrode) . Because the redox mediator has a low redox potential, the voltage applied to the working electrode can also be lower, which reduces the rate at which interfering compounds are oxidized by the working electrode. Examples of redox mediators having a lower redox potential include osmium bipyridyl complexes, ferrocene derivatives, and quinone derivatives. The disadvantage of this strategy is that redox mediators with lower redox potentials are often difficult to synthesize, are less stable, and have low aqueous solubility.

Another strategy that can be used to reduce the effect of interfering compounds is to use pseudo-electrodes coated with redox mediators. In some cases, it is also possible to use inert proteins or inactivated oxidoreductases as pseudo-electrodes. The purpose of the dummy electrode is to oxidize interfering compounds and/or redox mediators reduced by interfering compounds on the electrode surface. In this strategy, the current measured at the dummy electrode is subtracted from the total oxidation current measured at the working electrode to remove interfering effects. The disadvantage of this strategy is that it requires the test strip to include additional electrodes and additional electrical connections (ie dummy electrodes) that cannot be used to measure glucose. Including dummy electrodes is an inefficient use of electrodes in a glucose measurement system.

Invention overview

The present invention relates to methods of reducing the effect of interferents when using electrochemical sensors to detect analytes. An electrochemical sensor useful in the methods of the invention includes a substrate, at least first and second working electrodes and a reference electrode. Place the reagent layer on the electrodes such that the reagent layer completely covers the entire area of the first working electrode and only partially covers the second working electrode. In the method of the invention, the oxidation current generated on the part of the second working electrode not covered by the reagent layer is used to correct for the influence of interfering substances on the glucose measurement.

The present invention also includes a method of reducing interference in an electrochemical sensor comprising the steps of: measuring a first oxidation current at a first working electrode, wherein the first working electrode is covered by a reagent layer; measuring at a second working electrode a second oxidation current at the electrode where the reagent layer only partially covers the second working electrode; and calculating a corrected oxidation current representing the concentration of a preselected analyte (eg, glucose). In this calculation, the ratio of covered to uncovered area of the second working electrode was used to remove the effect of interferents on the oxidation current. More specifically, the correction current value can be calculated using the following formula,

$\mathrm{<span\; class="notranslate">\; G</span>}<span\; class="notranslate">\; =</span>{\mathrm{<span\; class="notranslate">\; we</span>}}_{\mathrm{<span\; class="notranslate">\; 1</span>}}<span\; class="notranslate">\; -</span><span\; class="notranslate">\; \{</span><span\; class="notranslate">\; (</span>\frac{{\mathrm{<span\; class="notranslate">\; A</span>}}_{\mathrm{<span\; class="notranslate">\; cov</span>}}}{{\mathrm{<span\; class="notranslate">\; A</span>}}_{\mathrm{<span\; class="notranslate">\; unc</span>}}}<span\; class="notranslate">\; )</span>\mathrm{<span\; class="notranslate">\; x</span>}<span\; class="notranslate">\; (</span>{\mathrm{<span\; class="notranslate">\; we</span>}}_{\mathrm{<span\; class="notranslate">\; 2</span>}}<span\; class="notranslate">\; -</span>{\mathrm{<span\; class="notranslate">\; we</span>}}_{\mathrm{<span\; class="notranslate">\; 1</span>}}<span\; class="notranslate">\; )</span><span\; class="notranslate">\; \}</span>$

where G is the corrected current density, VE ₁ is the uncorrected current density at the first working electrode, WE ₂ is the uncorrected current density at the second working electrode, and A _cov is the coating on the second working electrode area, A _unc is the uncoated area of the second working electrode.

In one embodiment of an electrochemical test strip useful in the present invention, the electrochemical glucose test strip includes first and second working electrodes, wherein the first working electrode is completely covered by a reagent layer and the second working electrode Only partially covered by the reagent layer. Thus, the second working electrode has a reagent coated area and an uncoated area. The reagent layer may include, for example, a redox enzyme such as glucose oxidase and a redox mediator such as ferricyanide. The first working electrode will have the superposition of two oxidation current sources, one from glucose and the other from the interferent. Similarly, the second working electrode will have a superposition of three current sources from the glucose, the interferent on the reagent coated part, and the interferent on the uncoated part. Since there is no reagent in this region, the uncoated portion of the second working electrode oxidizes only the interferents, but not the glucose. The oxidation current measured on the uncoated portion of the second working electrode can be used to estimate the total interferer oxidation current, and to calculate a corrected oxidation current that removes the effect of the interferer.

In another test strip embodiment useful in the methods of the present invention, an electrochemical glucose test strip includes first and second working electrodes, wherein the first and second working electrodes are only partially covered by a reagent layer. Thus, in this embodiment, both the first and second working electrodes have reagent-coated and uncoated portions. The first uncovered area of the first working electrode is different from the second uncovered area of the second working electrode. The oxidation current measured on the uncoated portion of the first and second working electrodes was used to estimate the interferent oxidation current in the uncoated portion, and to calculate the corrected glucose current.

The present invention also includes a method of reducing interference in an electrochemical sensor comprising the steps of: measuring a first oxidation current at a first working electrode, wherein the first working electrode is partially covered by a reagent layer; measuring at a second a second oxidation current on the working electrode, where the reagent layer only partially covers the second working electrode; and calculating a corrected oxidation current representing the concentration of a preselected analyte (eg, glucose). In this calculation, the ratio of covered to uncovered areas of the first and second working electrodes was used to remove the effect of interferents on the oxidation current. More specifically, the correction current value can be calculated using the following formula,

$\mathrm{<span\; class="notranslate">\; G</span>}<span\; class="notranslate">\; =</span>{\mathrm{<span\; class="notranslate">\; we</span>}}_{\mathrm{<span\; class="notranslate">\; 1</span>}}<span\; class="notranslate">\; -</span><span\; class="notranslate">\; \{</span><span\; class="notranslate">\; (</span>\frac{{\mathrm{<span\; class="notranslate">\; f</span>}}_{\mathrm{<span\; class="notranslate">\; 1</span>}}<span\; class="notranslate">\; +</span>{\mathrm{<span\; class="notranslate">\; f</span>}}_{\mathrm{<span\; class="notranslate">\; 2</span>}}}{{\mathrm{<span\; class="notranslate">\; f</span>}}_{\mathrm{<span\; class="notranslate">\; 2</span>}}<span\; class="notranslate">\; -</span>\mathrm{<span\; class="notranslate">\; 1</span>}}<span\; class="notranslate">\; )</span><span\; class="notranslate">\; \&times;</span><span\; class="notranslate">\; (</span>{\mathrm{<span\; class="notranslate">\; we</span>}}_{\mathrm{<span\; class="notranslate">\; 2</span>}}<span\; class="notranslate">\; -</span>{\mathrm{<span\; class="notranslate">\; we</span>}}_{\mathrm{<span\; class="notranslate">\; 1</span>}}<span\; class="notranslate">\; )</span><span\; class="notranslate">\; \}</span>$

where f ₁ is equal to A _cov1 /A _unc1 ; f ₂ is equal to A _cov2 /A _unc2 ; A _unc1 is the uncoated area of the first working electrode; A _unc2 is the uncoated area of the second working electrode; A _cov1 is Coated area of the first working electrode; A _cov2 is the coated area of the second working electrode; G is the corrected current value; WE ₁ is the uncorrected current density on the first working electrode, and WE ₂ is the uncorrected current density at the second working electrode.

Brief description of attached drawings

A better understanding of the features and advantages of the present invention may be obtained from the following detailed description of exemplary embodiments, using the principles of the invention and the accompanying drawings:

Figure 1 is an exploded perspective view of a test strip according to one embodiment of the present invention.

2 is a simplified plan view of a distal portion of a test strip according to the embodiment of the invention shown in FIG. 1 and including a conductive layer and an insulating layer.

3 is a simplified plan view of the distal portion of the test strip according to the embodiment of the invention shown in FIG. 1 showing the location of the reagent layer and the insulating and conductive layers.

Figure 4 is an exploded perspective view of a test strip according to another embodiment of the present invention.

5 is a simplified plan view of a distal portion of a test strip according to the embodiment of the invention shown in FIG. 4 and including a conductive layer and an insulating layer.

6 is a simplified plan view of the distal portion of the test strip according to the embodiment of the invention shown in FIG. 4 showing the reagent layer and the insulating and conductive layers.

7 is a simplified plan view of the distal portion of the test strip according to the embodiment of the invention shown in FIG. 4 showing the reagent layer and the conductive layer.

8 is a simplified plan view of the distal portion of a test strip showing a reagent layer with a conductive layer to help reduce IR drop effects according to another embodiment of the present invention.

9 is a simplified plan view of the distal portion of a test strip according to another embodiment of the present invention showing the reagent layer and the conductive and insulating layers such that there are two working electrodes with uncoated portions.

10 is a simplified plan view of the distal portion of a test strip according to another embodiment of the present invention showing the reagent layer and the conductive and insulating layers such that there are two working electrodes with uncoated portions.

Figure 11 shows the current at the first working electrode of a test strip designed in accordance with the present invention, measured with blood samples of 70 mg/dL glucose spiked with various levels of uric acid.

Figure 12 shows the current at the first working electrode of a test strip designed according to the present invention, measured with blood samples of 240 mg/dL glucose spiked with various levels of uric acid.

Figure 13 is an exploded perspective view of a test strip with an integral blade.

14 is a simplified diagrammatic view showing a meter connected to a test strip having a first contact, a second contact, and a reference contact disposed on a substrate.

                    Invention Details

The present invention includes test strips and methods for improving the selectivity of electrochemical glucose measurement systems.

Figure 1 is an exploded perspective view of a test strip according to a first embodiment of the present invention. In the embodiment of the invention shown in FIG. 1, an electrochemical test strip 62 for measuring glucose concentration in a bodily fluid such as blood or interstitial fluid includes a first working electrode 10 and a second working electrode 12, wherein the first working electrode The electrode 10 is completely covered by the reagent layer 22 , while the second working electrode 12 is only partially covered by the reagent layer 22 . Thus, the second working electrode has a reagent-coated portion and an uncoated portion. Reagent layer 22 may include, for example, a redox enzyme such as glucose oxidase and a redox mediator such as ferricyanide. Since ferricyanide has a redox potential of about 400 mV at a carbon electrode (when measured against a saturated calomel electrode), introduction into body fluids such as blood can produce significant interference oxidation by redox mediators and/or the working electrode, This results in a significant undesired oxidation current. Therefore, the oxidation current measured at the first working electrode 10 will be the superposition of the oxidation current sources: the first desired oxidation current due to the oxidation of glucose, and the second undesired oxidation current by the interfering species. current. The oxidation current measured on the second working electrode 12 will also be a superposition of the oxidation current sources: the first desired oxidation current due to the oxidation of glucose, on the covered part of the second working electrode 12 by the interfering A second undesired oxidation current is generated, and a third oxidation current is generated by the interferent on the uncovered portion of the second working electrode 12 . The uncoated portion of the second working electrode 12 will only oxidize the interferents, not the glucose, since there is no reagent on the uncoated portion of the second working electrode 12 . Since the oxidation current measured on the uncoated portion of the second working electrode 12 is independent of glucose, and the uncoated areas of the two working electrodes 12 are known, the uncoated area of the second working electrode 12 can be calculated. Disturbance oxidation current of coated sections. Thus, using the calculated interferent oxidation current for the uncoated portion of the second working electrode 12 and knowing the area of the first working electrode 10 and the area of the coated portion of the second working electrode 12, the elimination of Glucose current corrected for the effect of oxidized interfering compounds on the electrodes.

Figure 1 is an exploded perspective view of a test strip 62 according to a first embodiment of the present invention. A test strip 62 as shown in FIG. 1 can be produced by a series of 6 sequential printing steps which place 6 layers of material on the substrate 50 . These 6 layers can be deposited on the substrate 50 by eg screen printing. In one embodiment of the invention, the six layers may include conductive layer 64 , insulating layer 16 , reagent layer 22 , adhesive layer 66 , hydrophilic layer 68 and top layer 40 . The conductive layer 64 may further include a first working electrode 10, a second working electrode 12, a reference electrode 14, a first contact point 11, a second contact point 13, a reference contact point 15 and a test strip detection plate 17 . The insulating layer 16 may further include a cutout 18 . Adhesive layer 66 may further include first adhesive pad 24 , second adhesive pad 26 and third adhesive pad 28 . The hydrophilic layer 68 may further include a first hydrophilic film 32 and a second hydrophilic film 34 . The top layer 40 may further include a transparent portion 36 and an opaque portion 38 . As shown in FIG. 1 , the test strip 62 has a first side 54 and a second side 56 , an electrode distal side 58 and an electrode proximal side 60 . The following sections describe the various layers of test strip 62 in more detail.

In one embodiment of the invention, substrate 50 is an electrically insulating material such as plastic, glass, ceramic, or the like. In a preferred embodiment of the invention, substrate 50 may be a plastic such as nylon, polycarbonate, polyimide, polyvinyl chloride, polyethylene, polypropylene, PETG or polyester. More specifically, the polyester may be, for example, Melinex(R) ST328 produced by DuPont Teijin Films. Substrate 50 may also include an acrylic coating applied to one or both sides to improve ink adhesion.

The first layer deposited on the substrate 50 is the conductive layer 64 which includes the first working electrode 10 , the second working electrode 12 , the reference electrode 14 and the test strip detection plate 17 . According to the present invention, a screen with a latex pattern can be used to deposit a material such as conductive carbon ink in the defined geometry shown in FIG. 1 . The reference electrode 14 can also be a counter electrode, a reference electrode/counter electrode or a quasi-reference electrode. Conductive layer 64 may be deposited on substrate 50 by screen printing, rotogravure printing, sputtering, evaporation, electroless plating, inkjet, sublimation, chemical vapor deposition, or the like. Suitable materials that may be used for the conductive layer 64 are Au, Pd, Ir, Pt, Rh, stainless steel, doped tin oxide, carbon, and the like. In one embodiment of the present invention, the carbon ink layer may have a height of 1-100 microns, more specifically 5-25 microns, even more specifically about 13 microns. The height of the conductive layer can vary depending on the desired resistance of the conductive layer and the conductivity of the material used to print the conductive layer.

The first contact point 11, the second contact point 13 and the reference contact point 15 can be used for electrical connection with the measuring instrument. This places the meter in electrical communication with the first working electrode 10 , the second working electrode 12 and the reference electrode 14 via the first contact point 11 , the second contact point 13 and the reference contact point 15 respectively.

The second layer deposited on substrate 50 is insulating layer 16 . As shown in FIG. 1 , insulating layer 16 is deposited on at least a portion of conductive layer 64 . FIG. 2 is a simplified plan view of the distal portion of test strip 62 highlighting the positions of first working electrode 10 , second working electrode 12 , and reference electrode 14 relative to insulating layer 16 . The insulating layer 16 also includes a cutout 18 which may have a T-shaped structure as shown in FIGS. 1 and 2 . The cutout 18 exposes a portion of the first working electrode 10, the second working electrode 12 and the reference electrode 14 that can be wetted by the liquid. The incision 18 further includes a distal incision width W1, a proximal incision width W2, a distal incision length L4, and a proximal incision length L5. As shown in FIG. 2 , the distal incision width W1 corresponds to the width of the first working electrode 10 and the reference electrode 14 . The length corresponding to the distal incision length L4 is greater than the sum of the lengths of the first working electrode 10 and the reference electrode 14 . Proximal incision width W2 and proximal incision length L5 form a rectangular cut that exposes the width and length of second working electrode 12 . According to the present invention, the distal incision width W1, the proximal incision width W2, the distal incision length L4 and the proximal incision length L5 may have dimensions of about 0.7, 1.9, 3.2 and 0.43 mm, respectively. In one embodiment of the invention, the first working electrode 10, the reference electrode 14 and the second working electrode 12 have lengths L1, L2 and L3, respectively, which may be about 0.8, 1.6 and 0.4 mm, respectively. The electrode spacing S1 is the distance between the first working electrode 10 and the reference electrode 14; and the distance between the reference electrode 14 and the second working electrode 12, which may be about 0.4 mm.

The third layer deposited on substrate 50 is reagent layer 22 . As shown in FIG. 1 , reagent layer 22 is disposed on conductive layer 64 and at least a portion of insulating layer 16 . 3 is a simplified plan view of the distal portion of a test strip 62 according to a first embodiment of the present invention, which highlights the relationship of the reagent layer 22 to the first working electrode 10, the second working electrode 12, the reference The position of electrode 14 and insulating layer 16. Reagent layer 22 may be rectangular as shown in Figures 1 and 3, having a reagent width W3 and a reagent length L6. In one embodiment of the invention, the reagent width W3 may be about 1.3 mm and the reagent length L6 may be about 4.7 mm. In another embodiment of the present invention, the reagent layer 22 has a width W3 and a length L6 sufficiently large such that the reagent layer 22 completely covers the first working electrode 10 and the reference electrode 14 . However, the reagent layer 22 has a width W3 and a length L6 of appropriate size such that the second working electrode is not completely covered by the reagent layer 22 . In such an arrangement, as shown in FIG. 3, the second working electrode 12 has a coated portion 12c and an uncoated portion 12u. The uncoated portion 12u may be in the shape of two rectangles, wherein the uncoated portion 12u has a wing width W4 and a length corresponding to the second working electrode length L3. As a non-limiting example, wing width W4 may be about 0.3 mm. In one embodiment of the invention, the reagent layer 22 may include an oxidoreductase such as glucose oxidase or PQQ-glucose dehydrogenase (where PQQ is an acronym for pyrrolo-quinoline-quinone), and a redox mediator bodies such as ferricyanide.

The fourth layer deposited on the substrate 50 is an adhesive layer 66 that includes the first adhesive pad 24 , the second adhesive pad 26 and the third adhesive pad 28 . A first adhesive pad 24 and a second adhesive pad 26 form the walls of the sample receiving chamber. In one embodiment of the invention, the first adhesive pad 24 and the second adhesive pad 26 may be disposed on the substrate 50 such that neither adhesive pad contacts the reagent layer 22 . In another embodiment of the present invention where reducing the volume of the test strip is not required, the first adhesive pad 24 and/or the second adhesive pad 26 may be disposed on the substrate 50 such that it overlaps the reagent layer 22 . In one embodiment of the invention, the adhesive layer 66 has a height of about 70-110 microns. Adhesive layer 66 may comprise a double sided pressure sensitive adhesive, UV curable adhesive, heat activated adhesive, heat curable adhesive or other adhesives known to those skilled in the art. As a non-limiting example, adhesive layer 66 may be formed by screen printing a pressure sensitive adhesive such as a water-based acrylic copolymer pressure sensitive adhesive available from Tape Specialties LTD in Tring , Herts, United Kingdom (part #A6435).

The fifth layer deposited on substrate 50 is hydrophilic layer 68 , which includes first hydrophilic film 32 and second hydrophilic film 34 as shown in FIG. 1 . The hydrophilic layer 68 forms the "roof" of the sample receiving chamber. The "side walls" and "floor" of the sample receiving chamber are formed by a portion of the adhesive layer 66 and the substrate 50, respectively. As a non-limiting example, hydrophilic layer 68 may be an optionally clear polyester with a hydrophilic anti-fog coating, such as those available from 3M. The hydrophilic nature of the coating is used in the design of the test strip 62 as it facilitates the filling of liquid into the sample receiving chamber.

The sixth layer deposited on substrate 50 is top layer 40 , which includes transparent portion 36 and opaque portion 38 as shown in FIG. 1 . According to the invention, the top layer 40 comprises polyester coated on one side with a pressure sensitive adhesive. The top layer 40 has an opaque portion 38 that helps the user see high contrast when blood is beneath the transparent portion 36 . This enables the user to visually verify that the sample receiving chamber is sufficiently filled. After the test strip 62 has been fully laminated, it is cut along the cut line A-A', in the process creating the sample inlet 52 as shown in Figure 3 .

The first test strip embodiment shown in FIGS. 1-3 can have a possible disadvantage because the reagent layer 22 can dissolve in the liquid sample and move a portion of the dissolved reagent layer to the uncoated portion of the second working electrode 12. Part 12u on. If this occurs, the uncoated portion 12u will also measure an oxidation current which is also proportional to the glucose concentration. This will reduce the ability to use mathematical algorithms to remove the effects of oxidation of interferents. In another embodiment of the invention, the reagent layer 22 should be designed to dissolve in such a way that it does not migrate onto the uncoated portion 12u. For example, reagent layer 22 may be chemically bonded to first working electrode 10, second working electrode 12, and reference electrode 14, or may have a thickener that minimizes migration of dissolved reagent layer 22.

Another embodiment of the present invention is shown in FIG. 4. The embodiment shown in FIG. 4 reduces the migration of dissolved reagents to the uncoated portion of the second working electrode, and in some cases, reduces this migration. to a minimum. In this embodiment, the second working electrode 102 has a C-shaped geometry, as shown in FIG. According to the present invention, as shown in FIG. 6, the reagent layer 110 is only disposed on a portion of the second working electrode 102, so as to eliminate the uncoated portion 102u and the coated portion 102c. Uncoated portion 102u is adjacent to sample inlet 52 . The coated portion 102c is adjacent to the first working electrode 100 . When liquid is applied to the sample inlet 52 of the assembled test strip 162, the liquid will flow from the sample inlet 52 onto the coated portion 102c until all electrodes are covered with liquid. By arranging the location of the uncoated portion 102u upstream of the liquid flow, this almost completely prevents the reagent layer 110 from dissolving and migrating onto the uncoated portion 102u. This enables the mathematical algorithm to precisely remove the influence of interfering substances on the measured oxidation current.

FIG. 4 is an exploded perspective view of test strip 162 . Test strip 162 is fabricated in a similar manner to test strip 62 , but with changes in geometry or location to conductive layer 164 , insulating layer 106 , and reagent layer 110 . For the second embodiment of the present invention, the substrate 50, adhesive layer 66, hydrophilic layer 68 and top layer 40 are the same as the first test strip embodiment. The test strip 162 has a first side 54 and a second side 56 , an electrode distal side 58 and an electrode proximal side 60 . It should also be noted that the first and second test strip embodiments of the present invention may include components having similar structures, and these parts are designated by the same number and designation. If similar components of various test strip embodiments are structurally different, those components may have the same name but be given different part numbers. The following sections describe the various layers of test strip 162 in more detail.

For the test strip embodiment shown in FIG. 4, the first layer disposed on substrate 50 is conductive layer 164, which includes first working electrode 100, second working electrode 102, reference electrode 104, first A contact point 101, a second contact point 103, a reference contact point 105 and a test strip detection plate 17. According to the present invention, a screen with a latex pattern can be used to deposit a material such as conductive carbon ink in the defined geometry shown in FIG. 4 . The first contact point 101, the second contact point 103 and the reference contact point 105 can be used for electrical connection with the meter. This places the meter in electrical communication with the first working electrode 100, the second working electrode 102, and the reference electrode 104 via the first contact point 101, the second contact point 103, and the reference contact point 15, respectively.

In FIG. 4 , the second layer deposited on substrate 50 is insulating layer 106 . As shown in FIG. 4 , insulating layer 106 is deposited on at least a portion of conductive layer 164 . FIG. 5 is a simplified plan view of the distal portion of test strip 162 highlighting the positions of first working electrode 100 , second working electrode 102 , and reference electrode 104 relative to insulating layer 106 .

In FIG. 4 , the third layer deposited on substrate 50 is reagent layer 110 , which is disposed on at least a portion of conductive layer 164 and insulating layer 106 as shown in FIG. 6 . 6 is a simplified plan view of the distal portion of a test strip 162 according to a second embodiment of the present invention, which highlights the relationship of the reagent layer 110 to the first working electrode 100, the second working electrode 102, the reference The location of the electrode 104 and insulating layer 106. Reagent layer 110 may be rectangular having a reagent width W13 and a reagent length L16. In one embodiment of the invention, reagent width W13 may be about 1.3 mm and reagent length L16 may be about 3.2 mm. In a preferred embodiment of the present invention, the reagent layer 110 has a width W13 and a length L16 sufficiently large such that the reagent layer 110 completely covers the first working electrode 100, the coated portion 102c, and the reference electrode 104, but does not cover the remaining electrodes. Coated portion 102u.

Figure 7 is a simplified plan view of the distal portion of the test strip according to the embodiment of the invention shown in Figure 4, showing the reagent layer and the conductive layer. Unlike FIG. 6 , FIG. 7 does not show the insulating layer 106 . This helps to demonstrate the conductive relationship between the uncoated portion 102u and the coated portion 102c, which is hidden beneath the opaque symbols of the insulating layer 106. FIG.

For the test strip embodiment shown in FIG. 4 , the insulating layer 106 is used to define the width of the first working electrode 100 , the second working electrode 102 , and the reference electrode 104 . The insulating layer 106 also includes a cutout 108, which may have a T-shaped structure as shown in FIGS. 4-6. The cutout 108 exposes a portion of the first working electrode 100, the second working electrode 102, and the reference electrode 104 that can be wetted by the liquid. As shown in FIGS. 5 and 6 , the incision 108 further includes a distal incision width W11 , a proximal incision width W12 , a distal incision length L14 , and a proximal incision length L15 . The distal cutout width W11 corresponds to the width of the uncoated portion 102u. The distal cut length L14 is greater than the length of the uncoated portion 102u. Proximal cut width W12 and proximal cut length L15 form a rectangular cut that generally exposes the width and length of first working electrode 100, reference electrode 104, and coated portion 102c.

According to the present invention, distal incision width W11, proximal incision width W12, distal incision length L14, and proximal incision length L15 may have dimensions of about 1.1, 0.7, 2.5, and 2.6 mm, respectively.

In the embodiment of FIG. 4, uncoated portion 102u, reference electrode 104, first working electrode 100, and coated portion 102c have lengths L10, L12, L11, and L13, respectively, which may be about 0.7, 0.7, 0.4 and 0.4mm. The electrode spacing S11 is the distance between the uncoated portion 102u and the reference electrode 104, which may be about 0.2-0.75 mm, more preferably about 0.6-0.7 mm. The electrode spacing S10 is the distance between the reference electrode 104 and the first working electrode 100; and the distance between the coated portion 102c and the first working electrode 100, which may be about 0.2 mm. It should be noted that the electrode spacing S11 is greater than the electrode spacing S10 to reduce the possibility of reagent dissolution and migration onto the uncoated portion 102u. In addition, the electrode spacing S11 is larger than the electrode spacing S10 to reduce the possibility that the reagent layer 110 is disposed on the uncoated portion 102u due to a difference in printing process. The fourth through sixth layers are sequentially deposited on test strip 162 in the same manner as the first test strip embodiment. The relative positions and shapes of the adhesive layer 66, the hydrophilic layer 68 and the top layer 40 are shown in FIG.

In the embodiment of the invention shown in FIG. 8, the C-shape of the second working electrode 102 can be partially altered such that the sequence of liquid wetting the electrodes is the uncoated portion 102u, the first working electrode 100, the reference electrode 104 and coating portion 102c. In another form, the first working electrode 100 and the coated portion 102c are equidistant from the reference electrode 104, which is desirable from an IR drop perspective. In the second test strip embodiment (i.e., test strip 162) shown in FIG. The portion 102c is coated. For test strip 162 , the distance between coated portion 102 c and reference electrode 104 is greater than the distance between first working electrode 100 and reference electrode 104 .

Therefore, an algorithm can be used to calculate the corrected glucose current unaffected by interferents. After applying the sample to the test strip, apply a constant potential to the first working electrode and the second working electrode, and measure the current at both electrodes. On the first working electrode where the reagent covers the entire electrode area, the following formula can be used to describe the composition that contributes to the oxidation current,

WE ₁ =G+I _cov (Formula 1)

where _WE is the current density at the first working electrode, G is the current density due to glucose unaffected by the interferer, and I _cov is due to the interferer on the part of the working electrode covered by the reagent the current density.

On the second working electrode partially covered by the reagent, the following formula can be used to describe the components contributing to the oxidation current,

WE ₂ ＝G+I _cov +I _unc (Formula 2)

where _WE2 is the current density at the second working electrode and I _unc is the current density due to interferents at the part of the working electrode not covered by the reagent. Additional embodiments of the invention can be obtained using different reagent coated areas on the first and second working electrodes, but the formula must take into account the different uncoated areas.

To reduce the influence of interferents, a formula was developed describing the relationship between the interference current on the coated portion of the second working electrode and the interference current on the uncoated portion of the second working electrode. The interferer oxidation current density measured on the coated portion was estimated to be about the same as the current density measured on the uncoated portion. This relationship is further described by the following formula,

$I_{\mathrm{cov}}=\frac{A_{\mathrm{cov}}}{A_{\mathrm{unc}}}\&times;I_{\mathrm{unc}}$ (Formula 3a)

where A _cov is the area of the second working electrode covered by the reagent, and A _unc is the area of the second working electrode not covered by the reagent.

It should be noted that uncoated portion 12u and coated portion 12c may have areas designated A _unc and A _cov , respectively. The uncoated portion 12u can oxidize interferents, but not glucose, since there is no reagent layer 22 coated thereon. In contrast, the coated portion 12c can oxidize glucose and interferents. Since it was found experimentally that the uncoated portion 12u oxidizes the interfering species in a manner proportional to the area of the coated portion 12c, the proportion of the interfering current to the total current measured at the second working electrode 12 can be predicted. This allows the total current measured at the second working electrode 12 to be corrected by subtracting the interfering current. In one embodiment of the invention, the ratio of _Aunc : _Acov may be from about 0.5:1 to 5:1, and is preferably about 3:1. A more detailed description of this mathematical algorithm for current correction will be described in the following sections.

In another embodiment of the invention, the interferer oxidation current density measured on the coated portion may be different from the current density measured on the uncoated portion. This may be due to more or less efficient interference oxidation on the coated part. In one instance, the presence of a redox mediator can enhance the oxidation of interferents relative to the uncoated portion. In another instance, the presence of a viscosity-increasing substance such as hydroxyethylcellulose may reduce interferer oxidation relative to the uncoated portion. Depending on the components included in the reagent layer to which the second working electrode portion is coated, the interferer oxidation current measured on the coated portion may be greater or smaller than the uncoated portion. This property can be modeled phenomenologically by rewriting Equation 3a as,

I _cov =f×I _unc (Formula 3b)

where f is a correction factor that incorporates the effect of the interference oxidation efficiency of the coated part on the uncoated part.

In one embodiment of the present invention, Equations 1, 2 and 3a can be manipulated to derive an equation that outputs a corrected glucose current density that is not affected by interferents. It should be noted that these three formulas (Equations 1, 2 and 3a) have a total of 3 unknowns which are G, I _cov and I _unc However, Equation 1 can be rearranged into the following form.

G＝WE ₁ -I _cov (Formula 4)

Next, I _cov from Equation 3a can be substituted into Equation 4 to obtain Equation 5.

$G={\mathrm{we}}_1-[\frac{A_{\mathrm{cov}}}{A_{\mathrm{unc}}}\&times;I_{\mathrm{unc}}]$ (Formula 5)

Next, Equation 1 and Equation 2 can be combined to obtain Equation 6.

I _unc =WE ₂ -WE ₁ (Equation 6)

Next, _Iunc from Equation 6 can be substituted into Equation 5, resulting in Equation 7a.

$G={\mathrm{we}}_1-\left\{\left(\frac{A_{\mathrm{cov}}}{A_{\mathrm{unc}}}\right)x({\mathrm{we}}_2-{\mathrm{we}}_1)\right\}$ (Formula 7a)

Equation 7a outputs the corrected glucose current density G, which eliminates the influence of interfering substances. This formula only needs the current density output from the first working electrode and the second working electrode, and the coating of the second working electrode. Ratio of cloth area to uncoated area. In one embodiment of the invention, the ratio A _cov /A _unc can be programmed into the glucose meter, for example into the read only memory of the meter. In another embodiment of the invention, the ratio A _cov /A _unc can be communicated to the gauge via a calibration code chip that can eliminate manufacturing variations in A _cov or A _unc .

In another embodiment of the invention, Equations 1, 2 and 3b may be used when the interferer oxidation current density of the coated portion is different from the interferer oxidation current density of the uncoated portion. In such a case, another correction formula 7b as shown below is derived.

G=WE ₁ -{f×(WEX-WE ₁ )} (Formula 7b)

In another embodiment of the present invention, the meter may use the corrected glucose current formula 7a or 7b only when a certain threshold is exceeded. For example, if WE ₂ is about 10% or more larger than WE ₁ , the meter will use Equation 7a or 7b to correct for output current. However, if WE ₂ is about 10% larger than WE ₁ or less, the meter will simply take the average current value between WE ₁ and WE ₂ to increase the accuracy and precision of the measurement. The strategy of using equation 7a or 7b mitigates the risk of overcorrection of the measured glucose current only in some cases where significant levels of interfering compounds are present in the sample. It should be noted that when WE ₂ is sufficiently larger than WE ₁ (eg, about 20% or more larger), this is an indication of having a very high concentration of interfering compounds. In such cases, it may be desirable to output an error message instead of a glucose value, since very high levels of interferents can cause a breakdown in the accuracy of Equation 7a or 7b.

In the embodiment of the invention shown in Figures 9 and 10, the first and second working electrodes are partially covered by the reagent layer in such a way that the uncoated portions of the first and second working electrodes are different of. This is in contrast to the first and second test strip embodiments described above in which the first working electrode is completely covered by the reagent layer.

9 is a simplified plan view of the distal portion of a test strip 2000 showing reagent layer 22 and conductive and insulating layers 2002, with two working electrodes having uncoated portions, according to another embodiment of the present invention. Test strip 2000 is prepared in a manner similar to test strip 62, but with geometric changes to cutout 18 shown in FIG. 1 . Test strip 2002 has the same substrate 50 , conductive layer 64 , reagent layer 22 , adhesive layer 66 , hydrophilic layer 68 and top layer 40 as test strip 62 . The test strip 2002 is modified to have a cutout 2004 having a dumbbell-like shape as shown in FIG. 9 . The altered shape of the cutout 2004 is such that the first working electrode 2008 includes a first coated portion 2008c and a first uncoated portion 2008u; the second working electrode 2006 includes a second coated portion 2006c and a second Uncoated portion 2006u. In order for the test strip 2000 to effectively reduce the influence of interferents, the first uncoated portion 2008u must have a different total area than the second uncoated portion 2006u.

Figure 10 is a simplified plan view of the distal portion of a test strip 5000 showing a reagent layer 820 and a conductive layer with two working electrodes having uncoated portions according to another embodiment of the present invention. Test strip 5000 was prepared in a manner similar to test strip 162, but with geometric changes to conductive layer 164 such that both first working electrode 4002 and second working electrode 4004 had a C-shape. Test strip 5000 has the same substrate 50 , insulating layer 106 , reagent layer 110 , adhesive layer 66 , hydrophilic layer 68 and top layer 40 as test strip 162 . This altered geometry is such that the first working electrode 4002 includes a first coated portion 4002c and a first uncoated portion 4002u; the second working electrode 4004 includes a second coated portion 4004c and a second uncoated portion 4002u. Coated portion 4004u. In order for the test strip 2000 to effectively reduce the influence of interferents, the first uncoated portion 4002u must have a different area than the second uncoated portion 4004u.

An advantage of test strips 2000 and 5000 is that they can be easily produced in terms of depositing the reagent layer to the desired location and any subsequently deposited layers. Furthermore, the first and second working electrodes will have to some extent identical chemical and electrochemical interactions with any interferents, thereby ensuring greater accuracy in the calibration method. Since both working electrodes have some level of uncoated area, the same but different degrees of reaction will occur on both electrodes. With a simple change to Equation 7a, the following Equation 7c can be used as the correction formula for glucose,

$G={\mathrm{we}}_1-\{\left(\frac{{\mathrm{<figure-callout\; id="1"\; label="f"\; filenames="A20048003953300241.png"\; state="\{\{state\}\}">f</figure-callout>}}_1+f_2}{f_2-1}\right)\&times;({\mathrm{we}}_2-{\mathrm{we}}_1)\}$ (Formula 7c)

where f ₁ =A _cov1 /A _unc1 , f ₂ =A _cov2 /A _unc2 , A _unc1 =uncoated area of the first working electrode, A _unc2 =uncoated area of the second working electrode, A _cov1 = The coated area of the first working electrode, and A _cov2 = the coated area of the second working electrode.

An advantage of the present invention is that the first and second working electrodes can be used to determine that the sample receiving chamber has been sufficiently filled with liquid. An advantage of the present invention is that the second working electrode not only corrects for interferent effects, but also enables the measurement of glucose. This allows for more accurate results as 2 glucose measurements can be averaged with one test strip.

Example 1

A test strip was prepared according to the first embodiment of the invention shown in Figures 1-3. These test strips were assayed in blood with varying concentrations of interfering substances. To measure these test strips, they were electrically connected to a potentiostat having means to apply 0.4 volts between the first working electrode and the reference electrode 810; and between the second working electrode and the reference electrode constant potential. Apply the blood sample to the sample inlet and allow the blood to soak into the sample receiving chamber and wet the first working electrode, the second working electrode and the reference electrode. The reagent layer becomes hydrated by blood, which then produces ferrocyanide, which can be directly proportional to the amount of glucose and/or the concentration of interferents present in the sample. Oxidation of ferrocyanide was measured as currents at the first working electrode and the second working electrode 5 seconds after the sample was applied to the test strip.

Figure 11 shows the current response at the first working electrode, measured with blood samples spiked with 70 mg/dL glucose at various levels of uric acid. The uncorrected current at the first working electrode (indicated by the squares) shows a current increase proportional to the concentration of uric acid. However, the corrected current (represented by triangles) processed by Equation 7a appears to be unaffected by increasing uric acid concentration.

Figure 12 shows the current response at the first working electrode, measured with blood samples spiked with 240 mg/dL glucose at various levels of uric acid. The purpose of the test strip for glucose determination at 240 mg/dL is to show that the correction algorithm of Equation 7a is also valid over a range of glucose concentrations. Similar to Figure 11, the uncorrected current at the first working electrode (indicated by the squares) exhibited a current increase proportional to the concentration of uric acid. However, the corrected current (indicated by triangles) appears to be unaffected by increasing uric acid concentration.

Example 2

In order to show that the method of correcting the interferor current is applicable to a variety of interferors, test strips constructed according to the embodiment of Figure 1 were also assayed with different concentration levels of paracetamol and gentisic acid in addition to uric acid. To quantify the magnitude of this effect, changes in glucose output greater than 10% (for glucose levels >70 mg/dL) or 7 mg/dL (for glucose levels <70 mg/dL) were defined as significant disturbances. Table 1 shows that the uncorrected current at the first working electrode exhibits significant interfering effects at lower interferent concentrations compared to the test strip determined by the corrected amperometric response using Equation 7a. This shows that using Equation 7a to correct the output current of the first working electrode is effective in correcting for interference. Table 1 shows that the current correction in Equation 7a is effective for paracetamol, gentisic acid, and uric acid interferences. Table 1 also shows the normal concentration ranges of the interferents found in blood. In addition, Table 1 also shows that the current correction in Equation 7a is valid at a glucose concentration level of 240 mg/dL.

FIG. 13 is an exploded perspective view of a test strip 800 that is designed to penetrate the user's skin layers so that physiological fluid is expressed and collected within the test strip 800 in a seamless manner. Test strip 800 includes substrate 50 , conductive layer 802 , insulating layer 804 , reagent layer 820 , adhesive layer 830 and top layer 824 . The test strip 800 also includes a distal end 5 and a proximal end 60 .

In test strip 800 , conductive layer 802 is the first layer disposed on substrate 50 . As shown in Figure 13, the conductive layer 802 includes a second working electrode 806, a first working electrode 808, a reference electrode 810, a second contact point 812, a first contact point 814, a reference contact point 816, a test Strip detection plate 17. The material of the conductive layer 802 and the method for printing the conductive layer 802 are the same as the test strip 62 and the test strip 800 .

Insulating layer 804 is a second layer disposed on substrate 50 . The insulating layer 16 includes a cutout 18 which may have a rectangular configuration. The cutout 18 exposes a portion of the second working electrode 806, the first working electrode 808, and the reference electrode 810 that are wettable by the liquid. The material used for insulating layer 804 and the method used to print insulating layer 804 are the same as test strip 62 and test strip 800 .

Reagent layer 820 is the third layer disposed on substrate 50 , first working electrode 808 and reference electrode 810 . The materials used for reagent layer 820 and the method used to print reagent layer 820 are the same as test strip 62 and test strip 800 .

Adhesive layer 830 is a fourth layer disposed on substrate 50 . The material used for adhesive layer 830 and the material used to print adhesive layer 830 are the same as test strip 62 and test strip 800 . Adhesive layer 830 functions to secure top layer 824 to test strip 800 . In one embodiment of the invention, the top layer 824 may be in the form of an integral blade as shown in FIG. 13 . In such an embodiment, the top layer 824 can include a blade 826 on the distal end 58 .

Blade 826, which may also be referred to as a penetrating member, may be adapted to pierce the user's skin and draw blood into test strip 800 such that second working electrode 806, first working electrode 808, and reference electrode 810 are moisten. Blade 826 includes a blade base 832 that terminates on distal portion 58 of the assembled test strip. Blade 826 may be made of insulating materials such as plastic, glass, and silicon, or conductive materials such as stainless steel and gold. Further descriptions of integrated medical devices using integrated blades can be found in International Application PCT/GB01/05634 and U.S. Patent Application 10/143,399. Additionally, the blade 826 may be manufactured, for example, by progressive die stamping techniques as disclosed in the aforementioned International Application PCT/GB01/05634 and U.S. Patent Application 10/143,399.

FIG. 14 is a simplified diagrammatic view showing the meter 900 connected to a test strip. In one embodiment of the invention, the following test strips may be suitable for use with meter 900: test strip 62, test strip 162, test strip 800, test strip 2000, test strip 3000 or test strip 5000. Meter 900 has at least 3 electrical contacts that form electrical connections to the second working electrode, the first working electrode, and the reference electrode. In particular, the second contact (13, 103 or 812) and the reference contact (15, 105 or 816) are connected to the first voltage source 910; the first contact (11, 101 or 814) and the reference The contact point (15, 105 or 816) is connected to a second voltage source 920.

When measuring, the first voltage source 910 applies the first potential E1 between the second working electrode and the reference electrode; the second voltage source 920 applies the first potential E1 between the first working electrode and the reference electrode. Two potentials E2. In one embodiment of the present invention, the first potential E1 and the second potential E2 may be the same, for example about +0.4V. In another embodiment of the invention, the first potential E1 and the second potential E2 may be different. Apply the blood sample so that the second working electrode, the first working electrode and the reference electrode are covered with blood. This allows the second working electrode and the first working electrode to measure a current proportional to glucose and/or a non-enzyme specific source. Five seconds after applying the sample, meter 900 measures the oxidation current of the second working electrode and the first working electrode.

Table 1. Summary of interferent effects using corrected and uncorrected output currents model interferer Glucose concentration (mg/dL) The effect is significant for the concentration of interferents Normal concentration range of interfering substances uncorrected paracetamol 70 11 1-2 uncorrected Gentisic acid 70 10 0.05-0.5 uncorrected uric acid 70 5 2.6-7.2 uncorrected paracetamol 240 16 1-2 uncorrected Gentisic acid 240 12 0.05-0.5 uncorrected uric acid 240 8 2.6-7.2 Correction paracetamol 70 120 1-2 Correction Gentisic acid 70 47 0.05-0.5 Correction uric acid 70 33 2.6-7.2 Correction paracetamol 240 59 1-2 Correction Gentisic acid 240 178 0.05-0.5 Correction uric acid 240 29 2.6-7.2

Claims (4)

1. A method of reducing interference in an electrochemical sensor, the method comprising: measuring a first current on a first working electrode covered by a reagent layer; measuring a second current on a second working electrode, wherein the reagent layer partially covers the second working electrode, the second working electrode having a covered area and an uncovered area; and A corrected current value representative of glucose concentration is calculated using the ratio of the covered area to the uncovered area of the second working electrode. 2. The method of claim 1, wherein said correction current value is calculated with the following formula: $\mathrm{<span\; class="notranslate">\; G</span>}<span\; class="notranslate">\; =</span>{\mathrm{<span\; class="notranslate">\; we</span>}}_{\mathrm{<span\; class="notranslate">\; 1</span>}}<span\; class="notranslate">\; -</span><span\; class="notranslate">\; \{</span><span\; class="notranslate">\; (</span>\frac{{\mathrm{<span\; class="notranslate">\; A</span>}}_{\mathrm{<span\; class="notranslate">\; cov</span>}}}{{\mathrm{<span\; class="notranslate">\; A</span>}}_{\mathrm{<span\; class="notranslate">\; unc</span>}}}<span\; class="notranslate">\; )</span>\mathrm{<span\; class="notranslate">\; x</span>}<span\; class="notranslate">\; (</span>{\mathrm{<span\; class="notranslate">\; we</span>}}_{\mathrm{<span\; class="notranslate">\; 2</span>}}<span\; class="notranslate">\; -</span>{\mathrm{<span\; class="notranslate">\; we</span>}}_{\mathrm{<span\; class="notranslate">\; 1</span>}}<span\; class="notranslate">\; )</span><span\; class="notranslate">\; \}</span>$ where G is the corrected current value, WE ₁ is the uncorrected current density at the first working electrode, WE ₂ is the uncorrected current density at the second working electrode, A _cov is the second is the coated area of the second working electrode, and A _unc is the uncoated area of the second working electrode. 3. A method of reducing interference in an electrochemical sensor, said method comprising: measuring a first current on a first working electrode, wherein the reagent layer partially covers the first working electrode, the first working electrode having a first covered area and a first uncovered area; measuring a second current on a second working electrode, wherein the reagent layer partially covers the second working electrode, the second working electrode having a second covered area and a second uncovered area; and A corrected current value representative of glucose concentration is calculated using the ratio of the covered area to the uncovered area of the first and second working electrodes. 4. The method of claim 3, wherein said correction current value is calculated with the following formula: $G={\mathrm{we}}_1-\{\left(\frac{f_1+f_2}{f_2-1}\right)\&times;({\mathrm{we}}_2-{\mathrm{we}}_1)\}$ (Formula 7c) in f ₁ =A _cov1 /A _unc1 ; f ₂ =A _cov2 /A _unc2 ; _Auncl is the uncoated area of the first working electrode; A _unc2 is the uncoated area of the second working electrode; A _cov1 is the coating area of the first working electrode; A _cov2 is the coating area of the second working electrode; G is the correction current value; _WE1 is the uncorrected current density on the first working electrode and _WE2 is the uncorrected current density on the second working electrode.

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