KR100998648B1

KR100998648B1 - biosensor

Info

Publication number: KR100998648B1
Application number: KR20080039332A
Authority: KR
Inventors: 이금필
Original assignee: 이금필
Priority date: 2008-04-28
Filing date: 2008-04-28
Publication date: 2010-12-07
Also published as: WO2009133983A1; KR20090113550A

Abstract

The present invention relates to a biosensor in which a conductive material is formed on a bonding layer for bonding an upper substrate or a lower substrate provided with a working electrode and used as an auxiliary electrode or a sample recognition electrode.

At least one working electrode formed on at least one of an upper substrate and an insulating substrate, and configured to measure a signal formed by the analyte; At least one conductive bonding layer coupling the two substrates and having electrical conductivity; The combined two substrates and the conductive bonding layer form a reaction space of at least one analyte, and one side of the conductive bonding layer exposed to the reaction space assists in measuring a signal formed by the analyte. Characterized in that formed into an electrode.

Biosensor, Conductive Bonding Layer

Description

Biosensor

The present invention relates to a biosensor, and more particularly, to a biosensor used as an auxiliary electrode or a sample recognition electrode by forming a conductive material on a bonding layer for bonding an upper substrate or a lower substrate having a working electrode.

Background arts related to biosensors include: The following numbers are all US registered patent numbers.

5,437,999; After the electrode is patterned on the insulator substrate using a photolithography method with a precise area of a working eletrode or a counter electrode, the substrate on which the electrodes are formed is bonded to each other using an intermediate bonding layer. To form an electrode. On one side of the intermediate bonding layer is formed an empty space for injection of the measurement sample, the empty space is formed with a reagent reacting with the analyte, the air discharge port for the air discharge on the substrate on which the working electrode or auxiliary electrode is formed Formed.

5,582,697; A biosensor for quantifying an enzyme substrate present in a sample by an electrochemical method, a working electrode, an auxiliary electrode, and a third electrode on an insulating substrate, and a reaction layer including an oxidoreductase Biosensor formed on the working electrode and the auxiliary electrode. The third electrode is used as an electrode for sensing the sample injection, and is located at least far from the working electrode and the auxiliary electrode from the sample inlet.

6,071,391 and 6,156,173; The upper substrate and the lower substrate are joined by an intermediate bonding layer to form a space for sample injection, and a working electrode or auxiliary electrode is formed on the two substrates to face each other, wherein the two substrates have air The discharge port is not formed, and the substrate provided with the working electrode and the auxiliary electrode at the portion injecting the sample has a sharp tip. In addition, in order to connect the electrode formed on the upper substrate to the conductive wire formed on the lower substrate, a hole is formed in a portion of the bonding layer to connect the conductive material.

6,576,101; In the biosensor measuring the sample electrochemically using a sample of 1uL or less, a method of manufacturing the electrode in the form of a coating like an enzyme participating in the reaction on the working electrode by binding a redox material that can be oxidized in the air to the polymer In addition, the oxidation / reduction material bound to the polymer serves as an electron transfer material (mediator) when reacted with a working electrode or an enzyme, and has a property of not diffusing into a sample solution.

6,618,934; In a method of manufacturing a plurality of electrochemical sensors, a plurality of working electrodes and auxiliary electrodes are formed on the substrate in the first electrode region and the second electrode region, a spacer layer is formed in one of the electrode regions, and then the sample After removing a portion of the spacer to form a reaction chamber, and folding the substrate to form a layer of the first electrode zone and the second electrode zone to produce an electrochemical sensor, each sensor is separated into It consists of at least one working electrode, auxiliary electrode and sample reaction chamber.

6,863,800; In a biosensor configuration in which an analyte is positioned between an electrode formed on an electrode substrate and an electrode formed on a substrate to cover the electrode, the working electrode includes an ink composed of a reagent, a conductive material, and an electron mediator reacting with the analyte. The reference electrode is formed by coating a material reacting with an analyte and an electron transporting material on a conductive material, and the auxiliary electrode is positioned to face an electrode substrate made of a conductive material and to cover each other.

6,885,196; In the biosensor structure in which the first insulating substrate on which the working electrode is formed and the second insulating substrate on which the auxiliary electrode is formed face each other, a sample inflow space is formed between the two substrates, and the working electrode and the auxiliary electrode are formed in the sample inflow space. The reaction layer including the electrode and the oxidoredutase is exposed, and the distance between the working electrode and the auxiliary electrode is 150 micrometers or less, and the area of the auxiliary electrode exposed to the sample inflow space is larger than that of the exposed working electrode. It is a small biosensor.

6,942,769B2; In the biosensor assembled by combining an electrode, an insulating layer, an adhesive layer, a layer having a porosity of 10 to 40%, an adhesive layer, and a cover plate in this order after forming an electrode on the substrate for electrode formation, the insulating film Holes are formed in the cover plate and the porous layer controls the flow and volume of the analyte by holes formed in the insulating film and the cover plate.

7,022,218; In the biosensor for the analysis of the trace amount, the working electrode having a plurality of branches and the auxiliary electrode having a plurality of branches are alternately arranged alternately on the first insulating substrate, and another auxiliary electrode branch is formed on the second insulating substrate. The two substrates are assembled to face each other, and a working electrode and an auxiliary electrode are alternately arranged with a reaction reagent composed of a oxidoreductase in a sample inflow space formed between the two substrates.

7,050,843; An insulating first substrate having a conductive surface and a second substrate having a conductive surface are bonded to each other by an intermediate layer, and the capillary channel formed by the two substrates and the intermediate layer has a biosensor in which a reagent reacting with a sample is dried. And forming an insulating pattern line having a “V” shape on the conductive surface to adjust the flow of the sample, and dividing the conductive surface into two sections.

2006/0008581 A1; In the method of manufacturing an electrochemical sensor, having a working electrode on the first insulator and forming a plurality of insulating layers on the working electrode, the hole is formed through the working electrode to form a container, the cross section of the cut working electrode The container is exposed to the wall of the container, and additionally, a reference electrode is formed on the uppermost layer of the plurality of insulators, and the bottom plate is additionally bonded.

The biosensor measuring the analytical material electrochemically described in the prior art, basically consists of an upper insulator substrate, a lower insulator substrate and an intermediate layer that combines the two substrates, the sample injection or sample by the two substrates and the intermediate layer Comprising a space for the reaction, the two substrates are provided with a working electrode and an auxiliary electrode for measuring the electrochemical oxidation or reduction.

In particular, in order to measure the electrochemical oxidation / reduction current, the area where the working electrode and the auxiliary electrode formed on the substrate are in contact with the sample is important, and in general, the electrochemical signal increases in proportion to the electrode area. In order to obtain a larger electrochemical signal using the same volume of the sample, the patents include a working electrode on one of the two substrates, an auxiliary electrode on the other substrate, and then use the intermediate layer to connect the two electrodes to each other. Form facing each other. The arrangement of the electrodes has an advantage of maximizing the electrode area more efficiently than forming the two electrodes on the same substrate.

However, since the conductive material used in most of the electrodes is opaque, when the electrode is formed on both substrates, there is a disadvantage that it is difficult to visually check when the sample is injected.

To solve this problem, a method of forming an electrode other than the working electrode and the auxiliary electrode to confirm the sample injection is used, but the area of the working electrode or the auxiliary electrode is reduced or measured due to the formation of a third electrode in a limited area. There is a disadvantage that the amount of sample for the increase.

In order to solve this problem, an object of the present invention is to provide a biosensor to form a conductive material in the bonding layer for coupling the upper substrate or lower substrate provided with a working electrode to use as an auxiliary electrode or a sample recognition electrode. .

Biosensor according to the present invention for solving the above problems,

In the biosensor for measuring the analyte in the sample,

At least one working electrode formed on at least one of an upper substrate and an insulating substrate, and configured to measure a signal formed by the analyte; At least one conductive bonding layer combining the two substrates and having electrical conductivity; The combined two substrates and the conductive bonding layer form a reaction space of at least one analyte, and one side of the conductive bonding layer exposed to the reaction space assists in measuring a signal formed by the analyte. Characterized in that formed into an electrode.

In addition, the working electrode is characterized in that it comprises at least any one of a conductive polymer, gold, palladium, graphite, carbon, ITO particles, metal particles, carbon nanotubes (carbon nanotube, CNT).

In addition, irregularities or bends are formed on one side of the conductive bonding layer to increase the contact area with the sample.

In addition, the conductive bonding layer is formed by printing or coating a conductive adhesive material on both sides of the insulating layer.

In addition, the conductive bonding layer is characterized in that the conductive adhesive material is formed on one surface of the insulating layer, the non-conductive adhesive material is formed on the other surface of the insulating layer.

In addition, the conductive bonding layer is characterized in that formed only with a conductive adhesive material.

In addition, the conductive bonding layer is characterized in that it comprises at least one of a conductive polymer, gold, silver, silver chloride, palladium, graphite, carbon, ITO particles, metal particles, carbon nanotubes (carbon nanotube, CNT).

In addition, the coupling between the upper substrate and the insulating substrate, and the conductive bonding layer is characterized in that the coupling using pressure, heat, or light.

In addition, the reaction space is characterized in that a reaction layer comprising at least one enzyme reacting with the analyte, and the electron transfer material capable of oxidation or reduction by reacting with the enzyme. In this case, the reaction layer is preferably formed under the reaction space and the conductive bonding layer. In this case, the reaction layer preferably contains a water-soluble polymer. In this case, the enzyme contained in the reaction layer is preferably at least one enzyme of oxidoreductase, transfer enzyme, hydrolase, degrading enzyme, isomerase, synthetase. In this case, the reaction layer preferably further includes a buffer material of pH 2 to pH 9.

In addition, the thickness of the conductive bonding layer is characterized in that 1μm to 1000μm.

In addition, the sheet resistance of the conductive bonding layer is characterized in that 10Ω to 500kΩ.

In addition, it is formed on the working electrode, it characterized in that it further comprises a porous layer bonded by the conductive bonding layer. At this time, the porous layer is preferably made of microporous of less than 10 ㎛ capable of removing red blood cells in the sample. In this case, the porous layer preferably includes an enzyme that reacts with the analyte in the sample.

In addition, the upper substrate, the insulating substrate, and the conductive bonding layer formed in the reaction space formed by the conductive coupling layer, characterized in that it further comprises a sample recognition electrode for detecting a sample introduced into the reaction space; do. In this case, the sample recognition electrode is formed spaced apart from the sample injection unit for the sample injection, it is preferable to be formed at a position farther than the working electrode.

The apparatus may further include an air outlet connected to the upper substrate, the insulating substrate, and the reaction space formed by the conductive bonding layer.

According to the biosensor according to the present invention as described above, since the conductive bonding layer is used as an auxiliary electrode or a sample recognition electrode, there is no need to separately provide the electrodes on the substrate, thereby maximizing the area of the working electrode.

Hereinafter, embodiments of the present invention will be described in detail with reference to the accompanying drawings. First, it should be noted that the same components or parts in the drawings represent the same reference numerals as much as possible. In describing the present invention, detailed descriptions of related well-known functions or configurations are omitted in order not to obscure the gist of the present invention.

1 is a process diagram illustrating a process of manufacturing a biosensor according to a first embodiment of the present invention, FIG. 2 is a cross-sectional view of a biosensor according to a first embodiment of the present invention, and FIG. 3 is a first embodiment of the present invention. 4 is a cross-sectional view illustrating that a reaction layer is formed in a reaction space in a biosensor according to an example, and FIG. 4 illustrates that a reaction layer is formed below a portion of a reaction space and a conductive coupling layer in a biosensor according to a first embodiment of the present invention. 5 is a cross-sectional view showing a conductive bonding layer of the biosensor according to the present invention, FIG. 6 is a biosensor including a sample recognition electrode for detecting a sample introduced into the reaction space in the biosensor according to the present invention. 7 is a process diagram illustrating a process for manufacturing a biosensor formed on a working electrode in the biosensor according to the present invention and including a porous layer bonded by a conductive bonding layer. 8 is a process diagram illustrating a process of manufacturing a biosensor having a sample injection unit and a sample reaction space on both sides of the biosensor in the biosensor according to the present invention, and FIG. Process diagram showing a process of manufacturing a biosensor having a plurality of reaction spaces according to a second embodiment, Figure 10 is a process diagram showing a process of manufacturing a face-type biosensor according to a third embodiment of the present invention 11 is a cross-sectional view of a facing biosensor according to a third exemplary embodiment of the present invention, and FIG. 12 is a process diagram illustrating a process of manufacturing a biosensor having a plurality of reaction spaces according to a fourth exemplary embodiment of the present invention. 13 is a cross-sectional view of a biosensor having a plurality of reaction spaces according to a fourth embodiment of the present invention, and FIG. 14 is a diagram illustrating a biosensor according to the present invention mounted on a rotatable rotor.

1 is a process diagram illustrating a process of manufacturing a biosensor according to a first embodiment of the present invention.

Referring to FIG. 1A, first, the conductive layers 20 and 30 are formed on the surface of the insulating substrate 10. The conductive layer may be formed by various methods such as printing, coating, and deposition.

Next, an insulating layer 40 is formed on the conductive layers 20 and 30 and the insulating substrate 10 in a pattern as shown in FIG. 1 (b) and a conductive bonding layer 50A to be formed in a later process. After covering a part of the conductive layer 20 in contact, the working electrode 30A and the measuring device connection part 30B of the working electrode are formed, and for the electrical connection between the conductive coupling layer 50B and the conductive layer 20. The connecting portion 20A and the measuring instrument connecting portion 20B are formed.

Then, the conductive bonding material is formed in a pattern as shown in FIG. 1C to form the conductive bonding layers 50A and 50B. FIG. 1 (c) illustrates a case in which the conductive bonding layers 50A and 50B and the exposed portion 20C, which is part of the conductive material in contact with the sample, are used together as an auxiliary electrode, and in FIG. The case where the layers 50A and 50B are used as auxiliary electrodes in the electrochemical sensor is shown.

The conductive bonding layer is a layer made of a material having electrical conductivity and adhesion, which will be described later with reference to FIG. 5.

Next, as shown in FIG. 1 (d), the insulating substrate 10 and the upper substrate 80 are bonded by using the adhesive force of the conductive bonding layer. 1A through 1D, the insulating substrate and the upper substrate are joined in a pattern as shown in FIG. 1D, so that the sample inlet 90 and the measuring analyte react for the measurement sample injection. A space 70 and an air discharge unit 60 for discharging air in the sample reaction space 70 by the sample injection through the sample injection port 90 are formed. In this case, the upper substrate, the insulating substrate, and the coupling between the conductive bonding layer are preferably bonded using pressure, heat, or light. In addition, since the conductive bonding layers 50A and 50B are made of a conductive material, the conductive bonding layers 50A and 50B electrically connect to the conductive layer 20 through the connection part 20A.

2 is a cross-sectional view of a biosensor according to a first embodiment of the present invention.

Referring to FIG. 2, the conductive bonding layers 50C and 50D exposed in the sample reaction space 70 are in contact with an analytical sample, and the sample from the electrochemical sensor to the auxiliary electrode or the reference electrode or the sample reaction space 70. It is used as a sample recognition electrode to detect inflow. The use of the conductive bonding layer 50A, 50B, 50C, 50D as such an electrode has a minimum space occupancy within the limited sample reaction space 70, thus providing a relatively maximum working electrode area of 30A. It has a merit to be maintained. In this case, the working electrode includes at least one of a conductive polymer, gold, palladium, graphite, carbon, ITO particles, metal particles and carbon nanotubes (CNTs).

3 is a cross-sectional view showing a reaction layer is formed in the reaction space in the biosensor according to the first embodiment of the present invention, Figure 4 is a portion of the reaction space and the conductive bonding layer in the biosensor according to the first embodiment of the present invention It is sectional drawing which shows that the reaction layer was formed in the lower part.

3 and 4, a reaction layer 500 for measuring analyte of a sample is formed in the sample reaction space 70 of the biosensor 100, and the reaction layer 500 is a working electrode ( 30A), and portions 500A and 500B of the reaction layer 500 may be formed under the conductive bonding layers 50A and 50B. The reaction layer 500 reacts with an analyte to provide an analytical signal measurable at the working electrode 30A, and is composed of various kinds of enzymes according to the analyte, and forms the reaction layer 500 together with a polymer.

In particular, as shown in FIG. 4, when portions 500A and 500B of the reaction layer are formed under the conductive bonding layers 50A and 50B, the reaction layer is dissolved or absorbs wet or sample by inflow of a sample. It is connected to the conductive bonding layer in contact, and the contact area between the sample and the conductive bonding layer can be relatively widened through the reaction layer. As the polymer of the reaction layers 500, 500A, and 500B, a water-soluble polymer or a water-insoluble polymer may be used. Preferably, the water-soluble polymer is suitable.

At this time, the enzyme provided in the reaction layer 500 is oxidoreductase (oxidoreductase), transferase (transferase), hydrolase, hydrolase (lyase), isomerase (isomerase), At least one enzyme may be selected and used among synthetase, and a buffer material between pH 2 and pH 9 may be further used to maintain activity of the selected enzyme. In addition, the reaction layer 500 includes an electron transport material (mediator) capable of oxidation-reduction by reacting with the enzyme, and the electron transport material is oxidized or reduced when a constant voltage is applied from a working electrode, thereby providing an oxidation current or Generate a reducing current.

The description of the reaction layer is equally applicable to the second to fourth embodiments described below as well as the first embodiment of the present invention.

5 is a cross-sectional view showing a conductive bonding layer of the biosensor according to the present invention.

Referring to FIG. 5, the conductive bonding layer 50 combines an insulating substrate and an upper substrate to form a sample injection unit and a sample reaction space. In addition, the conductive bonding layer exposed to the reaction space may include an auxiliary electrode and a sample in a biosensor. Used as a recognition electrode. In the biosensor according to the present invention, the conductive bonding layer 50 may be composed of a single conductive layer 400 (see FIG. 5 (a)), and both electrical conductivity and adhesiveness for bonding an insulating substrate and an upper substrate. Have

In addition, the conductive bonding layer 50 may be in the form of a multilayer. For example, the conductive layer 400 is provided on both surfaces of the insulating layer 450 (see FIG. 5 (b)), or the conductive layer 400 is formed on one surface of the insulating layer 450, and the non-conductive adhesive layer is formed on the other surface of the insulating layer 450. 5 (see FIG. 5C), both layers 400 and 500 have adhesiveness to an upper substrate or an insulating substrate. In addition, the insulating layer 450 formed between the non-conductive adhesive layer 500 and the conductive layer 400 may be removed and used (see FIG. 5 (d)).

In addition, irregularities or bends may be formed on one side of the conductive bonding layer to increase the contact area with the sample.

In addition, the thickness of the conductive bonding layer is preferably 1μm to 1000μm. Because, when the thickness of the conductive bonding layer is more than 1000μm, the area of the auxiliary electrode in contact with the sample is increased to obtain a stable electrical signal, but the volume of the formed reaction space is required a large amount of samples, 1μm or less Due to the low thickness of the bonding layer, it is difficult to bond due to low adhesion between the insulating substrate 10 and the upper substrate 80, and the volume of the reaction space formed is not constant because of the error of the thickness, and the area of the auxiliary electrode in contact with the sample. This is because the role of the auxiliary electrode is reduced.

In addition, the sheet resistance of the conductive bonding layer is preferably 10Ω to 500kΩ. If the surface resistance of the conductive bonding layer is less than 10Ω, the ratio of the conductive material to the conductive bonding layer should be increased. As a result, the content of the adhesive material decreases, so that the adhesive strength decreases, and if the resistance is more than 500 kΩ, the electrical resistance is high and stable. This is because signal measurement is difficult.

Adhesiveness of the insulating substrate or the upper substrate of the conductive bonding layer 50 is made of a method such as heat, pressure, ultraviolet irradiation to the conductive bonding layer, conductive polymer, gold, silver, silver chloride, to the conductive layer 400, Palladium, graphite, carbon, ITO particles, metal particles, carbon nanotubes (carbon nanotube, CNT) can be produced by mixing a conductive material and an adhesive material, and other commercially available materials can also be used.

The description of the conductive bonding layer may be equally applied to the second to fourth embodiments described below as well as the first embodiment.

6 is a process diagram illustrating a process of manufacturing a biosensor including a sample recognition electrode for sensing a sample introduced into the reaction space in the biosensor according to the present invention.

First, the conductive material 30 for use as the working electrode in the electrochemical sensor on the surface of the insulating substrate 10, is electrically connected to the conductive bonding layer to be formed in a later process so that the conductive bonding layers 50A and 50B are auxiliary electrodes. A sample recognition electrode that is electrically connected to the conductive material 20 to be used as a conductive bonding layer 50E to be formed in a later process to detect the sample introduced into the sample reaction space 70 by the conductive bonding layer 50E. The conductive material 110 for use as is formed in a pattern as shown in Figure 6 (a).

Next, as shown in FIG. 6B, an insulating layer 40 is formed on a part of the conductive materials 20, 30, and 110 to electrically connect the conductive bonding layers 50A and 50B used as auxiliary electrodes. A connecting portion 20A for connection, a conductive bonding layer 50E used as a sample recognition electrode, a connecting portion 110A for electrical connection, and an automatic electrode 30A are formed.

Then, the conductive bonding layers 50A, 50B, and 50E are formed in a pattern as shown in FIG. 6 (c). The conductive bonding layers 50A and 50B are used as auxiliary electrodes, and the conductive bonding layers 50E are used as sample recognition electrodes for sample injection recognition.

Then, as shown in Figure 6 (d), by coupling the insulating substrate 10 and the upper substrate 80 using the conductive bonding layer (50A, 50B, 50E), the sample reaction for the reaction of the sample Space 70, the air discharge portion 60 for discharging air in the space according to the sample inlet into the reaction space, the sample injection portion 90 for the sample injection, each of the conductive material (20, 30, 110) To form the respective connection lines (20B, 30B, 110B) for electrical connection with the measuring device.

In addition, a material reacting with the analyte to measure the analyte of the sample may be configured in the sample reaction space 70, for example, the working electrode 30A exposed to the sample reaction space 70 or the exposed material. Reagents that react with the analyte of the sample may be formed on the upper substrate 80.

The sample is injected through the sample injecting unit 90 to be introduced into the sample reaction space 70, and the air inside the reaction space is discharged to the outside through the air discharge unit 60. The sample can be detected electrochemically whether the sample is completely injected by the conductive bonding layer 50E for sample recognition formed in a part of the sample reaction space 70, and after the sample is detected, reacted in the reaction space for a predetermined time The sample is measured by an electrochemical method using the working electrode 30A and the conductive bonding layers 50A and 50B for the auxiliary electrode.

When the electrode for sample recognition is formed on the insulating substrate 10 or the upper substrate 80, an extra area is used, and as a result, the volume of the sample reaction space 70 is increased, so that a relatively large amount of sample analyte is measured. A positive sample is required.

In order to solve this problem and form a conductive bonding layer 50E for sample recognition in order to measure analytes using a relatively small amount of sample, an electrode for sample recognition of the insulating substrate 10 or the upper substrate 80 may be separately No need to use

In addition, the description of the sample recognition electrode can be equally applied to the second to fourth embodiments described below as well as the first embodiment of the present invention.

7 is a process diagram illustrating a process of manufacturing a biosensor formed on a working electrode in the biosensor according to the present invention and including a porous layer bonded by a conductive bonding layer.

First, conductive materials 20 and 30 are formed on the insulating substrate 10 as shown in FIG. 7A.

Next, the insulating layer 40 is formed on a portion of the conductive material in a pattern as shown in FIG. 7 (b) to electrically connect the working electrode 30A and the conductive bonding layer 50 used as the auxiliary electrode. Connections 20A for forming the connection lines 20B and 30B for electrically connecting the conductive material to the electrochemical measuring device.

Next, as shown in FIG. 7C, the conductive bonding layer 50 is formed on the insulating layer 40. In this case, the conductive bonding layer 50 does not cover the working electrode (30A).

Then, as shown in Figure 7 (d), after bonding the porous layer 200 using the conductive bonding layer 50 on the working electrode (30A), the upper substrate is bonded to the biosensor 100 Manufacture. The porous layer 200 is composed of microporous, and serves to remove the microparticles present in the sample. At this time, the porous layer is preferably made of microporous of less than 10μm to remove the red blood cells in the sample. In this case, the porous layer preferably includes an enzyme that reacts with the analyte.

For example, when a whole blood sample is applied onto the porous layer 200, only serum components from which red blood cells are removed from the whole blood sample are passed through the porous layer 200 to the working electrode 30A, and some separated serum The sample is introduced into the conductive bonding layer 50 as the auxiliary electrode, and electrically connects the working electrode 30A and the auxiliary electrode conductive bonding layer 50.

8 is a process diagram illustrating a process of manufacturing a biosensor having a sample injection unit and a sample reaction space on both sides of the biosensor according to the present invention.

First, the conductive materials 20 and 30 are formed on the insulating substrate 10 in a pattern as shown in FIG. 8A.

Next, the insulating layer 40 is formed in a pattern as shown in FIG. 8 (b) to form the working electrode 30A and the auxiliary bonding layer 50A and 50B on the conductive material 20 and 30. ) 20A for electrical connection to the (), and connecting lines 20B and 30B for electrically connecting the measuring device and the respective conductive materials 20 and 30.

Next, as illustrated in FIG. 8C, conductive bonding layers 50A and 50B are formed on the insulating layer 40.

Then, by using the conductive bonding layer to combine the insulating substrate 10 and the upper substrate 80, the biosensor having a sample injection unit 90 and a sample reaction space 70 for sample injection on both sides ( 100) is prepared.

9 is a flowchart illustrating a process of manufacturing a biosensor having a plurality of reaction spaces according to a second embodiment of the present invention.

First, a plurality of conductive materials 20, 30, and 110 are formed on an insulating substrate in a pattern as shown in FIG. 9A.

Next, an insulating layer 40 is formed on the plurality of conductive materials in a pattern as shown in FIG. 9 (b) to provide a plurality of working electrodes 30A, a plurality of conductive bonding layers 50A, 50B, and 50E. A plurality of connecting portions 20A, 30A, 110A for electrical connection of the plurality of conductive materials and a plurality of connecting lines 20B, 30B, 110B for electrically connecting the plurality of conductive materials with the measuring device are formed.

Next, as shown in FIG. 9C, conductive bonding layers 50A, 50B, and 50E are formed on the insulating layer 40.

Then, the conductive bonding layer 50E for the sample recognition electrode for detecting the sample introduced into the sample reaction space 70 by combining the insulating substrate 10 and the upper substrate 80 using the conductive bonding layer. In addition, the conductive bonding layers 50A and 50B for the auxiliary electrode are manufactured, and a plurality of air discharge parts 60 are configured to discharge air in the reaction space 70 by the sample injection.

10 is a flowchart illustrating a process of manufacturing a face-to-face biosensor according to a third embodiment of the present invention.

First, conductive materials 20, 30, and 120 are formed on the insulating substrate 10 in a pattern as shown in FIG. 10 (a) on the insulating substrate, and an upper substrate in a pattern as shown in FIG. 10 (b). Conductive material 130 is formed on 80.

Next, an insulating layer 40 is formed on the insulating substrate 10 in a pattern as shown in FIG. 10 (c) to form a working electrode 30A, a conductive part of the conductive material 20, 30, 120. Connecting portions 20A and 120A for electrical connection between the coupling layers 50A and 50B and the conductive materials 20 and 120, and connecting lines for electrical connection between the conductive materials 20, 30 and 120 and the measuring device ( 20B, 30B, 120B). Conductive bonding layers 50A, 50B, and 50E are formed on the insulating layer 40 and the connecting portions 20A and 120A, and the conductive bonding layer 50E is not directly in contact with the analyte sample and is formed on the upper substrate 80. Is electrically connected to 130B.

Next, an insulating layer 40 is formed on the upper substrate 80 in a pattern as shown in FIG. 10 (d) to connect the connection part 130B for electrical connection with the working electrode 130A and the conductive coupling layer 50E. ).

Next, conductive bonding layers 50A, 50B, and 50E are formed on the insulating substrate 10 in a pattern as shown in FIG. 10 (e).

Next, as shown in FIG. 10 (f), the insulating substrate 10 and the upper substrate 80 are combined by using the conductive bonding layer, and a sample injection unit 90 and a sample reaction space for sample injection. 70, the biosensor 100 including an air discharge unit 60 is manufactured.

FIG. 11 is a cross-sectional view of the large-area biosensor according to the third embodiment. Referring to this, the insulating substrate 10 and the upper substrate 80 are coupled to each other by the conductive bonding layers 50B and 50A. 70) and the conductive materials 130A and 30A exposed in the reaction space are used as working electrodes in the electrochemical sensor, and the portions 50C and 50D exposed in the direction of the reaction space among the conductive bonding layers 50A and 50B. Is used as an auxiliary electrode.

In the biosensor having such a structure, since a plurality of sample electrodes may be provided in a limited reaction space 70, an advantage of measuring a plurality of analytes using a small amount of sample and an exposed portion of the reaction space of the conductive bonding layer may be provided. Since 50C and 50D are used as auxiliary electrodes, it is not necessary to fabricate auxiliary electrodes corresponding to the respective working electrodes. In addition, a reactant may be formed in the working electrodes 30A and l30A in the reaction space 70 to generate an electrical detection signal by reacting with the sample analyte.

12 is a flowchart illustrating a process of manufacturing a biosensor having a plurality of reaction spaces according to a fourth embodiment of the present invention, and FIG. 13 is provided with a plurality of reaction spaces according to the fourth embodiment of the present invention. A cross section of the biosensor.

First, a plurality of conductive materials 20 and 30 are formed on the insulating substrate 10 in a pattern as shown in FIG. 12A.

Next, an insulating layer 40 is formed on the plurality of conductive materials 20 and 30 in a pattern as shown in FIG. 12 (b) to connect the plurality of working electrodes 30A and the conductive bonding layer 50. A plurality of connecting lines 20B and 30B are formed for the electrical connection between the 20A and the respective conductive materials 20 and 30 and the measuring device.

Next, as shown in FIG. 12 (c), a conductive bonding layer 50 is formed on the insulating layer 40.

Then, the plurality of sample reaction spaces 70 are formed by combining the insulating substrate 10 and the upper substrate 80 using the conductive bonding layer.

In the biosensor 100 manufactured as described above, the exposed portion 50F (see FIG. 13) of the plurality of conductive coupling layers 50 exposed to the plurality of sample reaction spaces 70 is used as an auxiliary electrode. Such biosensors have the advantage of being able to measure multiple samples at once.

14 is a view showing a biosensor according to the present invention mounted on a rotatable rotor.

Referring to FIG. 14, the biosensor according to the present invention includes an insulating substrate 10, an insulating layer 40, a conductive bonding layer 50, and an upper substrate 80 as described above, and is used as a working electrode. A conductive material may be formed on the insulating substrate, the upper substrate, or both substrates, and a plurality of analytes may be measured by one biosensor.

The biosensor forms an insulating layer 40 on the insulating substrate 10 or the upper substrate 80 to form a conductive material for a plurality of working electrodes and a plurality of conductive materials for electrical connection with the conductive coupling layer 50. After forming a plurality of connecting lines (20B, 30B) for electrical connection with the measuring device, a plurality of flow paths (310, 310A) through which the analytical sample or fluid is moved, reaction space 70 with a plurality of samples, a plurality of An air outlet 60 inside the sample reaction space is formed by using the conductive bonding layer 50.

In addition, the exposed portions of the flow paths 310 and 310A and the sample reaction space 70 of the conductive bonding layer 50 serve as an auxiliary electrode in an electrochemical biosensor for measuring analytes in a sample. , 31OA), the reaction space 70, and the air discharge unit 60 are formed by combining the insulating substrate 10 and the upper substrate 80 with the conductive bonding layer 50.

The biosensor 100 is implemented by the force of the rotation of the sample movement, mixing, reaction, pre-treatment, etc., the plurality of electrical connections with the measuring device is also made by the rotation of the sensor.

As described above with reference to the drawings illustrating a biosensor according to the present invention, the present invention is not limited by the embodiments and drawings disclosed herein, it is various within the technical scope of the present invention by those skilled in the art Of course, modifications can be made.

1 is a process chart showing a process of manufacturing a biosensor according to a first embodiment of the present invention;

2 is a cross-sectional view of a biosensor according to a first embodiment of the present invention;

3 is a cross-sectional view showing that the reaction layer is formed in the reaction space in the biosensor according to the first embodiment of the present invention;

4 is a cross-sectional view illustrating that a reaction layer is formed below a portion of a reaction space and a conductive coupling layer in a biosensor according to a first embodiment of the present invention;

5 is a cross-sectional view showing a conductive bonding layer of a biosensor according to the present invention;

6 is a process diagram illustrating a process of manufacturing a biosensor including a sample recognition electrode for sensing a sample introduced into the reaction space in the biosensor according to the present invention;

7 is a process diagram illustrating a process of manufacturing a biosensor formed on a working electrode in the biosensor according to the present invention and including a porous layer bonded by a conductive bonding layer;

8 is a process chart showing a process of manufacturing a biosensor having a sample injection unit and a sample reaction space on both sides of the biosensor in the biosensor according to the present invention;

9 is a process diagram illustrating a process of manufacturing a biosensor having a plurality of reaction spaces according to a second embodiment of the present invention;

10 is a process chart showing a process of manufacturing a face-to-face biosensor according to a third embodiment of the present invention;

11 is a cross-sectional view of a face type biosensor according to a third embodiment of the present invention;

12 is a flowchart illustrating a process of manufacturing a biosensor having a plurality of reaction spaces according to a fourth embodiment of the present invention;

13 is a cross-sectional view of a biosensor having a plurality of reaction spaces according to a fourth embodiment of the present invention;

Claims

In the biosensor for measuring the analyte in the sample,

At least one working electrode formed on at least one of an upper substrate and an insulating substrate, and configured to measure a signal formed by the analyte;

At least one conductive bonding layer coupling the two substrates and having electrical conductivity;

The combined two substrates and the conductive bonding layer form a reaction space of at least one analyte, and one side of the conductive bonding layer exposed to the reaction space assists in measuring a signal formed by the analyte. Including; an electrode;

And a reaction layer including at least one enzyme reacting with the analyte and an electron transfer material capable of being oxidized or reduced by reacting with the enzyme.

The method according to claim 1,

The working electrode is at least one of a conductive polymer, gold, palladium, graphite, carbon, ITO particles, metal particles, carbon nanotubes (carbon nanotube, CNT).

The method according to claim 1,

Biosensor, characterized in that the irregularities or bends formed on one side of the conductive bonding layer to increase the contact area with the sample.

The method according to claim 1,

The conductive bonding layer is a biosensor, characterized in that formed by printing or coating a conductive adhesive on both sides of the insulating layer.

The method according to claim 1,

The conductive coupling layer is a biosensor, characterized in that the conductive adhesive material is formed on one surface of the insulating layer, the non-conductive adhesive material is formed on the other surface of the insulating layer.

The method according to claim 1,

The conductive coupling layer is a biosensor, characterized in that formed only with a conductive adhesive material.

The method according to claim 1,

The conductive bonding layer is at least one of a conductive polymer, gold, silver, silver chloride, palladium, graphite, carbon, ITO particles, metal particles, carbon nanotubes (carbon nanotube, CNT).

The method according to claim 1,

Biosensor of the upper substrate, the insulating substrate, and the conductive coupling layer is coupled using pressure, heat, or light.

delete

The method according to claim 1,

The biosensor, characterized in that the reaction layer is formed below the reaction space and the conductive bonding layer.

The method according to claim 1,

The reaction layer is a biosensor comprising a water-soluble polymer.

The method according to claim 1,

The enzyme contained in the reaction layer is a biosensor, characterized in that at least any one of the enzyme oxidase, transfer enzyme, hydrolase, degrading enzyme, isomerase, synthetase.

The method according to claim 1,

The reaction layer is a biosensor, characterized in that further comprises a buffer material of pH2 to pH9.

The method according to claim 1,

The thickness of the conductive bonding layer is a biosensor, characterized in that 1μm to 1000μm.

The method according to claim 1,

The sheet resistance of the conductive bonding layer is a biosensor, characterized in that 10Ω to 500kΩ.

The method according to claim 1,

The biosensor is formed on the working electrode, further comprising a porous layer bonded by the conductive bonding layer.

18. The method of claim 16,

The porous layer is a biosensor, characterized in that made of microporous or less than 10 ㎛ capable of removing red blood cells in the sample.

18. The method of claim 16,

The porous layer is a biosensor comprising an enzyme reacting with an analyte in a sample.

The method according to claim 1,

The biosensor is formed of a conductive bonding layer in the reaction space formed by the upper substrate, the insulating substrate, and the conductive bonding layer, and further comprises a sample recognition electrode for sensing a sample introduced into the reaction space. .

The method of claim 19,

The sample recognition electrode is formed spaced apart from the sample injection unit for sample injection, characterized in that formed in a position farther than the working electrode.

The method according to claim 1,

And an air outlet connected to the reaction space formed by the upper substrate, the insulating substrate, and the conductive bonding layer.