KR20210062982A - Bio sensor - Google Patents

Abstract

A biosensor according to embodiments of the present invention includes a substrate, a working electrode including a reaction electrode layer disposed on the substrate and an enzyme reaction layer disposed on the reaction electrode layer, and a reference electrode spaced apart from the working electrode. The surface roughness (Ra) of the reaction electrode layer is 0.2 to 1.5 μm. By increasing the measurement range of the biosensor, it is possible to accurately measure the concentration of a small amount of a target substance.

Description

Biosensor {BIO SENSOR}

The present invention relates to a biosensor. More particularly, it relates to a biosensor including an enzyme reaction layer.

As the life expectancy of humans increases, the health care industry is expanding rapidly. In particular, there is an increasing demand for a portable and small biosensor that can conveniently measure various biosignals anywhere.

Conventional biosensors use enzymes that react with chemical species contained in body fluids (sweat, tears, blood, etc.). When the enzyme reacts with the species to generate an electric current, it is measured to determine the concentration of the species.

Of these, sweat, tears, and urine contain smaller amounts of some detectable substances than blood. When the concentration of the sensing target material is less than a specific level, the measurement current value competes with noise and measurement error may increase. In addition, when the concentration is extremely small, detection may not be possible.

For example, Korean Patent Application Laid-Open No. 2001-0110272 discloses a biosensor in which Prussian blue is introduced into an electrode, but additional research is needed to improve the sensitivity of the biosensor.

Republic of Korea Patent Publication No. 2001-0110272

One object of the present invention is to provide a biosensor with improved sensing performance.

1. Substrate; a working electrode including a reaction electrode layer disposed on the substrate and an enzyme reaction layer disposed on the reaction electrode layer; and a reference electrode spaced apart from the working electrode on the substrate, wherein the surface roughness (Ra) of the reaction electrode layer is 0.2 to 1.5 μm.

2. The method of 1 above, wherein the working electrode further includes a first wiring electrode disposed between the substrate and the reaction electrode layer, wherein the first wiring electrode is made of gold (Au), silver (Ag), or copper (Cu). , platinum (Pt), titanium (Ti), nickel (Ni), tin (Sn), molybdenum (Mo), cobalt (Co) and silver-palladium- containing at least one selected from the group consisting of copper alloy (APC) , biosensors.

3. The biosensor of 1 above, wherein the reaction electrode layer includes an electron transport material.

4. The method of 3 above, wherein the electron transport material is Prussian blue, potassium ferricyanide, potassium ferrocyanide, hexaamineruthenium (III) chloride (hexaammineruthenium (III)) chloride), ferrocene (ferrocene), ferrocene derivatives, quinones (quinones), comprising at least one selected from the group consisting of quinone derivatives and hydroquinone (hydroquinone), a biosensor.

5. The biosensor according to the above 1, wherein the reaction electrode layer includes a carbon-based electrode protective material.

6. The biosensor of 5 above, wherein the carbon-based electrode protection material includes a carbon paste.

7. The above 1, wherein the enzyme reaction layer comprises glucose oxidase, cholesterol oxidase, lactate oxidase, ascorbic acid oxidase, alcohol oxidase, glucose dehydrogenase, glutamic acid dehydrogenase, lactate dehydrogenase and alcohol dehydrogenase. A biosensor comprising at least one of.

8. The biosensor of 1 above, wherein the working electrode further comprises an ion filter layer disposed on the enzyme reaction layer.

9. The biosensor according to the above 1, wherein the surface roughness is 0.3 to 1 μm.

10. The biosensor of 1 above, further comprising a first wiring electrode disposed between the reaction electrode layer and the substrate.

11. The biosensor according to 1 above, further comprising a second wiring electrode disposed between the substrate and the reference electrode.

A biosensor according to exemplary embodiments of the present invention includes a reactive electrode layer having a surface roughness Ra of a specific range formed thereon. As the surface area of the reaction electrode layer increases, the contact area (reaction area) between the reaction electrode layer and the ion reaction layer may increase. Accordingly, the sensitivity and range of the biosensor may be increased, and the concentration of a small sample may be accurately measured.

1 is a schematic plan view of a biosensor according to exemplary embodiments of the present invention.
2 and 3 are schematic cross-sectional views of biosensors according to exemplary embodiments of the present invention.

Embodiments of the present invention are disposed on a substrate and include a working electrode including a reaction electrode layer and an enzyme reaction layer, and a reference electrode spaced apart from the working electrode, wherein the surface roughness (Ra) of the reaction electrode layer is 0.2 to 1.5 μm. provides By increasing the measurement range of the biosensor, it is possible to accurately measure the concentration of a small amount of a target substance.

Hereinafter, embodiments of the present invention will be described in detail. However, this is merely exemplary and the present invention is not limited to the specific embodiments described by way of example.

1 is a schematic plan view of a biosensor according to exemplary embodiments of the present invention.

Referring to FIG. 1 , a biosensor 10 according to example embodiments includes a substrate 100 , a working electrode 200 , and a reference electrode 300 . The biosensor 10 may include an auxiliary sensor 130 and a wiring unit 140 . The wiring unit 140 may include a first wiring 142 , a second wiring 144 , and a third wiring 146 .

The substrate 100 may be provided as a substrate on which the working electrode 200 , the reference electrode 300 , and the like are disposed.

For example, the substrate 100 may be a film having flexible characteristics. For example, the film may include a polyester-based resin such as polyethylene terephthalate, polyethylene isophthalate, polyethylene naphthalate, and polybutylene terephthalate; Cellulose resins, such as a diacetyl cellulose and a triacetyl cellulose; polycarbonate-based resin; acrylic resins such as polymethyl (meth)acrylate and polyethyl (meth)acrylate; styrenic resins such as polystyrene and acrylonitrile-styrene copolymer; polyolefin-based resins such as polyethylene, polypropylene, polyolefin having a cyclo-based or norbornene structure, and an ethylene-propylene copolymer; vinyl chloride-based resin; amide resins such as nylon and aromatic polyamide; imide-based resin; polyether sulfone-based resin; sulfone-based resins; polyether ether ketone resin; sulfide polyphenylene-based resin; vinyl alcohol-based resin; vinylidene chloride-based resin; vinyl butyral-based resin; allylate-based resin; polyoxymethylene-based resins; It may include a film formed of a thermoplastic resin, such as an epoxy resin, or a combination thereof. In addition, it may include a film formed of a thermosetting resin or ultraviolet curable resin such as (meth)acrylic, urethane, acrylic urethane, epoxy, or silicone.

The thickness of the substrate 100 may be appropriately determined, but may be 1 to 500 μm in consideration of strength, handleability, workability, thin layer properties, and the like. 1-300 micrometers is preferable and 5-200 micrometers is more preferable.

For example, the substrate 100 may contain an additive. For example, the additive may include a UV absorber, an antioxidant, a lubricant, a plasticizer, a mold release agent, a color inhibitor, a flame retardant, a nucleating agent, an antistatic agent, a pigment, a colorant, and the like.

In some embodiments, the substrate 100 may include a functional layer on one or both surfaces of the film. The functional layer may include, for example, a hard coating layer, an antireflection layer, a gas barrier layer, and the like.

In some embodiments, the substrate 100 may be surface-treated. For example, the surface treatment may include a plasma treatment, a corona treatment, a dry treatment such as a primer treatment, and a chemical treatment such as an alkali treatment including a saponification treatment.

2 is a schematic cross-sectional view of a biosensor according to exemplary embodiments of the present invention.

Referring to FIG. 2 , a biosensor 10 according to example embodiments includes a substrate 100 , a working electrode 200 , and a reference electrode 300 . The working electrode 200 includes a reaction electrode layer 220 and an enzyme reaction layer 230 . The reference electrode 300 may include a reference electrode layer 320 . The working electrode 200 and the reference electrode 300 may include a first wire electrode 210 and a second wire electrode 310 , respectively.

Redox reaction of the sensing target material may occur in the working electrode 200 . The working electrode 200 may detect an electrical signal generated by the reaction between the enzyme of the enzyme reaction layer 230 and the sensing target material in the sample.

The sample may be a body fluid such as sweat, tears, urine, or the like of a human body and blood. When the sample is sweat, tears, or urine, it may contain a smaller amount of a sensing target material (eg, glucose) than blood. According to exemplary embodiments of the present invention, it is possible to accurately measure the concentration of a small amount of a sensing target material by increasing the sensitivity of the biosensor.

The first wiring electrode 210 may be disposed on the substrate 100 . For example, the first wiring electrode 210 may contact the substrate 100 . The first wiring electrode 210 may serve as a path through which electrons or holes generated in an oxidation-reduction reaction of a sensing target material are transmitted. The first wire electrode 210 may be connected to the driving integrated circuit chip through the first wire 142 , and may be provided integrally with the first wire 142 .

The first wiring electrode 210 may further include a metal layer and a metal protective layer disposed on an upper surface of the metal layer. For example, the metal protective layer may cover the metal layer. The metal protective layer has electrical conductivity and may prevent oxidation-reduction of the metal layer due to the oxidation-reduction reaction of the working electrode 200 .

In example embodiments, the metal layer may include at least one of Au, Ag, Cu, Pt, Ti, Ni, Sn, Mo, Co, Pd, and alloys thereof. For example, an APC alloy (Ag-Pd-Cu alloy) may be used. The Au, Ag, APC alloy (Ag-Pd-Cu alloy), and Pt may improve electrical conductivity and reduce resistance of the first wiring electrode 210 . Accordingly, the detection performance of the biosensor 10 can be improved.

In example embodiments, the metal protective layer may include indium tin oxide (ITO) or indium zinc oxide (IZO). The ITO and IZO are chemically stable while having electrical conductivity, so that the metal layer can be effectively protected from oxidation-reduction reactions.

For example, the metal protective layer may prevent the metal layer from being in direct contact with the atmosphere to prevent oxidation of a metal component constituting the metal layer. Accordingly, the reliability of the electrical signal sensed by the metal layer may be improved.

The reaction electrode layer 220 may be disposed on the substrate 100 . In some embodiments, the reaction electrode layer 220 may be disposed on the first wire electrode 210 . The reaction electrode layer 220 may cover an upper surface of the first wire electrode 210 . In this case, the reaction electrode layer 220 may protect the first wiring electrode 210 from the oxidation-reduction reaction of the biosensor 10 .

In example embodiments, the reactive electrode layer 220 may include an electron transport material and a carbon-based electrode protection material. For example, the electron transport material may be dispersed in the carbon-based electrode protective material to form the reactive electrode layer 220 .

The electron transport material may improve electron and hole transport speed and stability of the reactive electrode layer 220 .

The electron transport material may include, for example, a material that is oxidized or reduced by receiving electrons/holes generated in an oxidation-reduction reaction of the sensing target material in the enzyme reaction layer 230 . Through the oxidation or reduction, electrons/holes may be transferred to the reaction electrode layer 220 .

In exemplary embodiments, it may include a cyanide complex of iron (Fe). The cyanide complex of iron can be easily oxidized or reduced. Therefore, electrons and holes can be transported easily and quickly.

For example, the sensing target material may react with the enzyme of the enzyme reaction layer 230 _{to form hydrogen peroxide (H 2} O ₂ ), and the biosensor 10 may measure the current generated while decomposing the hydrogen peroxide. .

Hydrogen peroxide may have a high redox potential. In the comparative example, when the cyanide complex of iron is not used, hydrogen peroxide must be decomposed at a high potential. In this case, substances other than hydrogen peroxide may be decomposed together to generate an electric current. Accordingly, the selectivity of the biosensor 10 may be reduced.

According to exemplary embodiments, the redox potential of hydrogen peroxide may be reduced by using a cyanide complex of iron. In this case, it is possible to selectively decompose hydrogen peroxide at a lower potential. Accordingly, the selectivity of the biosensor 10 can be improved.

In example embodiments, the electron transport material is Prussian blue (Fe ₄ [Fe(CN) ₆ ] ₃ ), potassium ferricyanide (Potassium ferricyanide, K ₃ [Fe(CN) ₆ ]) , potassium ferrocyanide (Potassium ferrocyanide, K ₄ [Fe(CN)] ₆ ), hexaamineruthenium (III) chloride (hexaammineruthenium (III) chloride), ferrocene, ferrocene derivatives, quinones, quinone derivatives , and may include hydroquinone and the like. For example, Prussian blue is a blue pigment and may have high oxidation properties. When Prussian blue as an electron transport material is disposed on the reactive electrode layer 220 , the electrical sensitivity of the working electrode 200 may be improved.

The carbon-based electrode protection material may have electrical conductivity. Accordingly, the carbon-based electrode protective material may transport electrons and/or holes generated in the enzyme reaction layer 230 . In example embodiments, the carbon-based electrode protection material may include a carbon paste.

The electron transport material may be included in an amount of 0.01 to 5 wt% based on the total weight of the reaction electrode layer 220 . In the above-described content range, the electron transport material may be uniformly dispersed, and electrons and holes may be effectively transported. Preferably, the electron transport material may be included in an amount of 0.1 to 3 wt% based on the total weight of the reactive electrode layer 220 .

The surface roughness (Ra, center line average) of the reaction electrode layer 220 may be 0.2 to 1.5 μm. The surface roughness may increase the surface area of the reaction electrode layer 220 . In this case, an area in which electrons/holes can move between the reaction electrode layer 220 and the enzyme reaction layer 230 (hereinafter, referred to as a reaction area) may increase. Accordingly, the sensitivity and the sensitivity range of the biosensor 10 may be increased.

When the surface roughness is less than 0.2 μm, the effect of increasing the sensitivity and the sensitivity range may be insignificant. When the surface roughness exceeds 1.5 μm, the dispersion of the measured current value may increase. Preferably, the surface roughness may be 0.3 to 1㎛. More preferably, the surface roughness may be 0.3 to 0.8 μm.

The enzyme reaction layer 230 may be disposed on the reaction electrode layer 220 . For example, the upper surface of the reaction electrode layer 220 may be covered. In the enzyme reaction layer 230 , an oxidation-reduction reaction of the sensing target material may occur.

In exemplary embodiments, the enzyme reaction layer 230 may include an oxidizing enzyme or a dehydrogenase. The oxidative enzyme and the dehydrogenase may be selected according to the type of the substance to be tested.

In exemplary embodiments, the oxidizing enzyme is glucose oxidase, cholesterol oxidase, lactate oxidase, ascorbic acid oxidase, or alcohol oxidase. (alcohol oxidase) may include at least one of.

In exemplary embodiments, the dehydrogenase may include at least one of glucose dehydrogenase, glutamate dehydrogenase, lactate dehydronase, or alcohol dehydrogenase. can

In this case, the biosensor 10 may measure the concentration of glucose, cholesterol, lactate, ascorbic acid, alcohol, glutamic acid, and the like.

For example, when a sample including a detection target material is injected into the biosensor 10 , the detection target material included in the sample reacts with an oxidase or dehydrogenase to generate byproducts such as _{hydrogen peroxide (H 2} O _{2 )} can be In this case, the electron transport material may reduce the by-product and may itself be oxidized. The oxidized electron transport material can be reduced again by obtaining electrons from the electrode surface to which a predetermined voltage is applied.

The concentration of the sensing target material in the sample may be proportional to the amount of current generated during the oxidation of the electron transport material. Accordingly, the concentration of the sensing target material may be measured by measuring the amount of current.

In some embodiments, the oxidizing enzyme or dehydrogenase may be immobilized through a binder. The binder may include a binder commonly used in the art, for example, Nafion, a derivative thereof, or chitosan may be included.

3 is a schematic cross-sectional view of a biosensor according to exemplary embodiments of the present invention.

Referring to FIG. 3 , in the biosensor 20 according to example embodiments, the working electrode 200 may further include an ion filter layer 240 .

In exemplary embodiments, the ion filter layer 240 may be disposed on the enzyme reaction layer 230 . For example, the ion filter layer 240 may cover the upper surface of the enzyme reaction layer 230 .

The ion filter layer 240 may protect the enzyme reaction layer 230 from chemical substances other than the test target material. In addition, it is possible to prevent the oxidizing enzyme or dehydrogenase of the enzyme reaction layer 230 from being exposed to the external environment.

The ion filter layer 240 may pass only the sensing target material. Accordingly, it is possible to prevent the enzyme reaction layer 230 from being denatured or damaged by a material other than the component to be measured.

As the ion filter layer 240 , an ion exchange membrane commonly used in the art may be used as long as the sensing target material passes through, and a fluorine-based polymer compound may be used. For example, the ion filter layer 240 may include Nafion, a Nafion derivative, polyaniline, or the like, but is not limited thereto.

In example embodiments, the reference electrode 300 may be disposed on the substrate 100 to be spaced apart from the working electrode 200 . The reference electrode 300 and the working electrode 200 may be electrically disconnected. The reference electrode 300 may be disposed on the same surface of the substrate 100 on which the working electrode 200 is disposed, or may face the working electrode 200 with the substrate 100 interposed therebetween.

The reference electrode 300 may provide a reference value for a current value or a potential value measured by the working electrode 200 during measurement. By using the potential value of the reference electrode 300 as a reference value, the oxidation-reduction reaction of the sensing target material occurring in the working electrode 200 may be specified.

In addition, by comparing the reference value of the current value with the current value measured by the working electrode 200, it is possible to calculate the amount of current changed purely by the measurement target component (eg, the sensing target material), and from the current amount The concentration of the component to be measured can be derived.

The reference electrode 300 may include a second wire electrode 310 and a reference electrode layer 320 . The reference electrode layer 320 may be formed on the second wire electrode 310 .

The second wiring electrode 310 may be formed of the same material as the first wiring electrode 210 of the working electrode 200 .

The reference electrode layer 320 may include an Ag/AgCl electrode. The Ag/AgCl electrode layer may be formed from Ag/AgCl paste. In some embodiments, when the second wire 144 is connected to the side of the reference electrode layer 320 , the second wire electrode 310 may be omitted.

In some embodiments, a protective layer may be disposed on the upper surface of the reference electrode 300 . The protective layer may protect the reference electrode 300 from external impact and environment. The protective layer may be formed in the same manner as the ion filter layer 240 of the working electrode 200 .

In example embodiments, the working electrode 200 may be formed by sequentially stacking the reaction electrode layer 220 and the enzyme reaction layer 230 on the substrate 100 . The reference electrode 300 may be formed to be spaced apart from the working electrode 200 by a predetermined distance.

In some embodiments, the first wiring electrode 210 may be formed between the substrate 100 and the reaction electrode layer 220 , and the second wiring electrode 310 may be formed between the substrate 100 and the reference electrode layer 320 . ) can be formed.

The first wiring electrode 210 forms a metal film including at least one of Au, Ag, Cu, Pt, Ti, Ni, Sn, Mo, Co, Pd, and alloys thereof on the substrate 100 and then patterning it. (patterning) can be formed. During the patterning, the first wiring electrode 210 and the first wiring 142 connected thereto may be formed together. The reference electrode 300 may be formed in the same way.

For the patterning, a patterning method commonly used in the art may be used. For example, photolithography may be used.

When the first wiring electrode 210 further includes a metal protective layer, the metal layer is first patterned and then the metal protective layer is formed, or ITO (Indium Tin Oxide) or IZO (Indium Zinc Oxide) conductivity is performed on the metal layer. After forming the oxide layer, the metal layer and the conductive oxide layer may be patterned together to form a metal layer and a metal protective layer together.

The reaction electrode layer 220 may be formed by applying a composition obtained by mixing the electron transport material and the carbon-based electrode protection material to the first wiring electrode 210 and then drying the composition.

For the application, a coating method commonly used in the art may be used, for example, various printing methods may be used.

In some embodiments, the mixed composition may be screen-printed using a screen of 200 to 500 mesh. In this case, a roughness of Ra 0.2 to 1.5 μm may be formed on the surface of the reaction electrode layer 220 .

In some embodiments, after applying and drying the mixed composition on the first wiring electrode 210 , the exposed surface may be surface-treated. The surface treatment may include plasma treatment, corona treatment, ultraviolet cleaning treatment, primer treatment, saponification treatment, and the like. Preferably, a surface roughness of 0.2 to 1.5 μm Ra can be effectively formed through plasma treatment, corona treatment or ultraviolet cleaning treatment.

The enzyme reaction layer 230 may be formed, for example, by applying a composition obtained by mixing an oxidizing enzyme or a dehydrogenase with a binder on the reaction electrode layer 220 and then drying.

In exemplary embodiments, the ion filter layer 240 may be formed on the upper surface of the enzyme reaction layer 230 . The ion filter layer 240 may be formed by drying a material having an ion filter function, such as Nafion, after coating.

In some embodiments, the wiring unit 140 may be connected to each of the working electrode 200 and the reference electrode 300 . The first wiring 142 connected to the working electrode 200 and the second wiring 144 connected to the reference electrode 300 may be electrically spaced apart from each other. The wiring unit 140 may be connected to a driving integrated circuit (IC) chip.

For example, the wiring unit 140 may be formed of the same material as the reactive electrode layer 220 of the working electrode 200 , and may be formed of the same material as the reference electrode 300 .

In some embodiments, the first wiring 142 and the second wiring 144 may be integrally formed with the working electrode 200 and the reference electrode 300 , respectively. A carbon paste film and/or a metal film is formed on the substrate 100 and the reactive electrode layer 220 and the reference electrode 300 are respectively formed by patterning the first wiring 142 and the second wiring 144 through screen printing. can be formed integrally with

Electrical signals measured from the working electrode 200 and the reference electrode 300 may be transmitted to the driving IC chip through the first wiring 142 and the second wiring 144 , and the driving IC chip determines the concentration of the component to be measured. can be calculated.

In some embodiments, the biosensor 10 may further include an auxiliary sensor 130 . The auxiliary sensor 130 may include a thermal sensor, a pH sensor, and/or a humidity sensor. For example, the auxiliary sensor 130 may measure temperature, pH, and/or humidity to correct a measurement error of the biosensor. The auxiliary sensor 130 may be connected to the driving IC chip by a third wire 146 .

Hereinafter, preferred embodiments are presented to help the understanding of the present invention, but these examples are only illustrative of the present invention and do not limit the appended claims, and are within the scope and spirit of the present invention. It is obvious to those skilled in the art that various changes and modifications are possible, and it is natural that such variations and modifications fall within the scope of the appended claims.

Examples and Comparative Examples

A 180-micrometer-thick PET substrate was prepared as an insulating substrate.

A first wiring electrode was formed by sequentially stacking an APC layer with a thickness of about 2000 Å and IZO with a thickness of about 500 Å on a substrate. A carbon paste electrode (including 3 wt% of Prussian blue, about 10 μm thick) was formed on the first wiring electrode.

At this time, the surface roughness of the carbon paste electrode was adjusted as shown in Table 1 below by adjusting the mesh of the screen.

A second wiring electrode was formed to be spaced apart from the first wiring electrode using the same material as the first wiring electrode. A reference electrode was manufactured by forming an Ag/AgCl electrode on the second wiring electrode.

The carbon paste electrode and the reference electrode were corona-treated (10 kV, 10 rpm).

A working electrode was prepared by forming an enzyme reaction layer in which glucose oxidase was fixed with chitosan on the corona-treated carbon paste electrode.

Experimental example

The concentration of the 0.1 mM glucose standard solution was measured through the biosensors of Examples and Comparative Examples.

Specifically, the working electrode and the reference electrode are connected to a current measuring device (CHI-630E, CH Instruments, INC), the standard solution is evenly dropped on the working electrode and the reference electrode, a voltage of -0.1 mV is applied, and 150 seconds The subsequent current value was measured.

In addition, in each of the Examples, the measurement was repeated 5 times to calculate the average current value and dispersion. The calculated values are shown in Table 1 below.

Surface roughness (㎛) Current value (nA) Spread (RSD, %) Example 1 0.2 82.9 5.6 Example 2 0.3 90.0 4.6 Example 3 0.36 90.4 4.3 Example 4 0.5 122 4.5 Example 5 0.55 131 4.7 Example 6 0.78 145.2 4.0 Example 7 0.8 146.3 3.9 Example 8 1.0 150.8 5.5 Example 9 1.5 152.4 5.8 Comparative Example 1 0.18 50.8 10.5 Comparative Example 2 1.6 90.9 23.2

Referring to Table 1, it was confirmed that the measured current value of the Examples was greatly improved and the dispersion was reduced compared to Comparative Examples in which the surface roughness of the reaction electrode layer was out of the range of 0.2 to 1.5 μm.

10: biosensor
100: substrate
200: working electrode 210: first wiring electrode
220: reaction electrode layer 230: enzyme reaction layer
240: ion filter layer
300: reference electrode 310: second wiring electrode
310: reference electrode layer

Claims (11)

Board;
a working electrode including a reaction electrode layer disposed on the substrate and an enzyme reaction layer disposed on the reaction electrode layer; and
and a reference electrode spaced apart from the working electrode on the substrate;
The surface roughness (Ra) of the reaction electrode layer is 0.2 to 1.5 ㎛, the biosensor.
The method according to claim 1, wherein the working electrode further comprises a first wiring electrode disposed between the substrate and the reaction electrode layer, the first wiring electrode is gold (Au), silver (Ag), copper (Cu), platinum (Pt), titanium (Ti), nickel (Ni), tin (Sn), molybdenum (Mo), cobalt (Co) and silver-palladium- containing at least one selected from the group consisting of copper alloy (APC), bio sensor.
The biosensor of claim 1 , wherein the reaction electrode layer comprises an electron transport material.
The method according to claim 3, wherein the electron transport material is Prussian blue (prussian blue), potassium ferricyanide (potassium ferricyanide), potassium ferrocyanide (potassium ferrocyanide), hexaamineruthenium (III) chloride (hexaammineruthenium (III) chloride) , A biosensor comprising at least one selected from the group consisting of ferrocene, ferrocene derivatives, quinones, quinone derivatives and hydroquinone.
The biosensor of claim 1, wherein the reaction electrode layer comprises a carbon-based electrode protective material.
The biosensor of claim 5 , wherein the carbon-based electrode protection material comprises a carbon paste.
The method according to claim 1, wherein the enzyme reaction layer comprises at least one of glucose oxidase, cholesterol oxidase, lactate oxidase, ascorbic acid oxidase, alcohol oxidase, glucose dehydrogenase, glutamic acid dehydrogenase, lactate dehydrogenase and alcohol dehydrogenase. A biosensor, including one.
The biosensor of claim 1, wherein the working electrode further comprises an ion filter layer disposed on the enzyme reaction layer.
The biosensor of claim 1, wherein the surface roughness is 0.3 to 1 μm.
The biosensor of claim 1, further comprising a first wiring electrode disposed between the reaction electrode layer and the substrate.
The biosensor of claim 1, further comprising a second wiring electrode disposed between the substrate and the reference electrode.

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