JP4197085B2 - Biosensor - Google Patents

Description

[0001]
BACKGROUND OF THE INVENTION
The present invention relates to a biosensor for quantifying a substrate contained in a sample solution.
[0002]
[Prior art]
A biosensor is a sensor that utilizes the molecular recognition ability of biological materials such as microorganisms, enzymes, and antibodies, and applies biological materials as molecular identification elements. That is, the immobilized biological material utilizes a reaction that occurs when a target substrate is recognized, oxygen consumption due to respiration of microorganisms, an enzymatic reaction, luminescence, and the like.
[0003]
Among biosensors, enzyme sensors have been put to practical use. For example, enzyme sensors for glucose, lactose, urea, and amino acids are used in medical measurement and the food industry. The enzyme sensor reduces the electron acceptor by electrons generated by the reaction between the substrate and the enzyme contained in the sample liquid that is the specimen, and the measurement device electrochemically measures the reduction amount of the electron acceptor, Perform quantitative analysis of specimens.
[0004]
Hereinafter, a conventional biosensor will be described with reference to the drawings.
FIG. 3 is a diagram showing perspective views of a conventional biosensor in the order of creation steps, and FIG. 4 is a diagram showing a sample supply path of the conventional biosensor. Reference numeral 101 denotes an insulating substrate made of polyethylene terephthalate or the like. Reference numeral 102 denotes an electrically conductive layer made of carbon, a metal material, or the like, which is formed on the entire surface of the substrate 101. Reference numerals 103 a, 103 b, 103 c, and 103 d are slits formed in the electrically conductive layer 102. Reference numerals 105, 106, and 107 denote electrodes formed by dividing the electrically conductive layer 102 by slits 103a, 103b, 103c, and 103d, which are a measurement electrode, a counter electrode, and a detection electrode. Reference numeral 108 denotes a spacer that covers the measurement electrode 105, the counter electrode 106, and the detection electrode 107. Reference numeral 109 denotes a rectangular notch that is provided in the center of the front edge of the spacer 108 and forms a specimen supply path. Reference numeral 110 denotes an inlet of the sample supply path. Reference numeral 111 denotes a reagent layer formed by applying a reagent containing an enzyme to the measurement electrode 105, the counter electrode 106, and the detection electrode 107 by dropping. A cover 112 covers the spacer 108. Reference numeral 113 denotes an air hole provided at the center of the cover 112.
[0005]
As shown in FIG. 3A, an electrically conductive layer 102 is formed on the entire surface of the substrate 101 by a screen printing method or the like. Next, as shown in FIG. 3B, slits 103a, 103b, 103c, and 103d are formed in the electrically conductive layer 102 using a laser, and the electrically conductive layer is formed on the measurement electrode 105, the counter electrode 106, and the detection electrode 107. 102 is divided. Next, as shown in FIG. 3C, in the case of a blood glucose sensor, a reagent composed of glucose oxidase, which is an enzyme, and potassium ferricyanide, etc., is dropped as an electron acceptor onto the measurement electrode 105, the counter electrode 106, and the detection electrode 107. To form the reagent layer 111. Next, a spacer 108 having a notch 109 for forming a specimen supply path is placed on the measurement electrode 105, the counter electrode 106, and the detection electrode 107. The sample supply path is in a state as shown in FIG. Further, a cover 112 is installed thereon. Here, one end of the notch 109 of the spacer 108 communicates with an air hole 113 provided in the cover 112.
[0006]
According to this configuration, when a sample liquid, which is a specimen such as blood, is supplied to the inlet 110 of the specimen supply path, a certain amount of specimen is sucked into the specimen supply path by capillary action through the air holes 113, and the counter electrode 106 and the measurement are performed. It reaches on the electrode 105 and the detection electrode 107. The reagent layer 111 formed on the electrode is dissolved by blood, for example, an oxidation-reduction reaction occurs between the reagent and the specimen, and an electrical change occurs between the measurement electrode 105 and the counter electrode 106. At the same time, if the sample is correctly filled in the sample supply path, an electrical change occurs between the measurement electrode 105 and the detection electrode 107. When this is sensed and a voltage is applied to the measurement electrode 105 and the counter electrode 106, for example, a blood glucose level sensor generates a current proportional to the glucose concentration, and the blood glucose level can be measured from the value.
[0007]
[Problems to be solved by the invention]
However, in conventional biosensors, even if a reagent is applied dropwise to the electrode to form a reagent layer, the reagent is uniform on the electrode due to the difference in the surface condition of the electrode and how the reagent spreads depending on the reagent solution composition. The amount of reagent on the electrode varies. That is, even if the same amount of reagent is applied by dropping, the spread of the reagent varies, so that the position and area of the reagent layer also vary. Therefore, there was a problem that the performance of the biosensor deteriorated.
[0008]
The present invention has been made in view of the above problems, and an object thereof is to provide a biosensor in which a reagent layer is uniformly arranged on an electrode regardless of the reagent solution composition and the performance is uniform.
[0009]
[Means for Solving the Problems]
In order to solve the above-described object, the biosensor according to claim 1 is a biosensor for quantifying a substrate contained in a sample solution, and includes an insulator substrate and an entire surface or a part of the insulator substrate. A plurality of electrodes formed by providing first slits in the electrically conductive layer formed thereon, a reagent layer disposed on the plurality of electrodes and formed by dropping a reagent, and the electrodes and reagents disposed on the layer comprises a spacer having a cutout portion of the sample solution to form a sample supply path for supplying to said electrodes, placed in front SL on the spacer and a cover having an air hole communicating with the specimen supply path A second slit that partially surrounds the position where the reagent is applied is provided around the position where the reagent is dropped on the electrode.
[0010]
The biosensor according to claim 2 is the biosensor according to claim 1, wherein the electrically conductive layer is formed on the insulating substrate by a sputtering method.
[0011]
The biosensor according to claim 3 is the biosensor according to claim 1 or 2, wherein the first slit and the second slit are formed by a laser. And
[0012]
The biosensor according to claim 4 is the biosensor according to any one of claims 1 to 3, wherein the second slit has an arc shape.
[0013]
DETAILED DESCRIPTION OF THE INVENTION
Embodiment 1 FIG.
The biosensor according to the first embodiment will be described with reference to the drawings.
FIG. 1 is a diagram showing perspective views of a biosensor according to the first embodiment in the order of creation steps, and FIG. 2 is a diagram showing a specimen supply path of the biosensor according to the first embodiment. Reference numeral 1 denotes an insulating substrate made of polyethylene terephthalate or the like. Reference numeral 2 denotes an electrically conductive layer formed on the entire surface of the substrate 1 and made of an electrically conductive material such as a noble metal such as gold or palladium or carbon. Reference numerals 3 a, 3 b, 3 c and 3 d are first slits provided in the electrically conductive layer 2. Reference numerals 5, 6, and 7 denote electrodes formed by dividing the electrically conductive layer 2 by the first slits 3a, 3b, 3c, and 3d. The measurement electrode, the counter electrode, and the specimen are surely placed inside the specimen supply path. It is a detection electrode which is an electrode for confirming whether it was attracted | sucked to. Reference numerals 4a and 4b denote second slits for regulating the position and area where the reagent on the electrode is applied. A spacer 8 covers the measurement electrode 5, the counter electrode 6, and the detection electrode 7. Reference numeral 9 denotes a rectangular notch that forms a sample supply path provided in the center of the front edge of the spacer 8. Reference numeral 10 denotes an inlet of the sample supply path. Reference numeral 11 denotes a reagent layer formed by applying a reagent containing an enzyme to the measurement electrode 5, the counter electrode 6, and the detection electrode 7 by dropping. A cover 12 covers the spacer 8. Reference numeral 13 denotes an air hole provided in the central portion of the cover 12.
[0014]
As shown in FIG. 1A, an electrically conductive layer 2 of a noble metal thin film such as gold or palladium is formed by sputtering, which is a method for forming a thin film on the entire surface of the substrate 1. The electrically conductive layer 2 may be formed not only on the entire surface of the substrate 1 but only on a portion necessary for forming the electrode.
[0015]
Next, as shown in FIG. 1B, the first slits 3a, 3b, 3c, and 3d are formed in the electrically conductive layer 2 using a laser, and the electrically conductive layer 2 is measured with the measuring electrode 5 and the counter electrode 6. And divided into detection electrodes 7. In addition, arc-shaped second slits 4 a and 4 b are formed in the electrically conductive layer 2 around the position where the reagent is dropped by using a laser so as to surround the position where the reagent is applied.
[0016]
In addition, a printing plate or a masking plate in which a pattern necessary for forming the electrically conductive layer 2 having the first slits 3a, 3b, 3c, and 3d and the second slits 4a and 4b was previously arranged was used. The electrodes and the first slits 3a, 3b, 3c, 3d and the second slits 4a, 4b may be formed on the substrate 1 by a screen printing method or a sputtering method.
[0017]
In addition, as a method of providing the first slits 3a, 3b, 3c, and 3d and the second slits 4a and 4b in the electrically conductive layer 2, a part of the electrically conductive layer 2 is formed by a jig having a sharp tip. You may sharpen.
[0018]
Next, as shown in FIG. 1 (c), in the case of a blood glucose sensor, a reagent composed of glucose oxidase which is an enzyme and potassium ferricyanide or the like as an electron acceptor is dropped onto the measurement electrode 5, the counter electrode 6 and the detection electrode 7. Apply by. Since the part to which the reagent is applied is a position sandwiched between the second slits 4a and 4b, the second slits 4a and 4b can be used as marks for applying the reagent. In addition, since the applied reagent is liquid, it spreads in a circular shape to the outside centering on the portion applied by dripping. However, the second slits 4a and 4b act as a breakwater and position of the reagent layer 11 And the area is restricted, and the second slits 4a and 4b do not spread. Therefore, the reagent layer 11 is formed at a predetermined position with a predetermined area.
[0019]
Next, a spacer 8 having a notch 9 for forming a specimen supply path is installed on the electrodes of the measurement electrode 5, the counter electrode 6 and the detection electrode 7. The sample supply path is in a state as shown in FIG.
[0020]
Next, the cover 12 is installed on the spacer 8. Here, one end of the notch 9 of the spacer 8 communicates with an air hole 13 provided in the cover 12.
In addition, after forming the spacer 8 on the electrode of the measurement electrode 5, the counter electrode 6, and the detection electrode 7, a reagent is dripped at the part exposed from the notch part 9 of the measurement electrode 5, the counter electrode 6, and the detection electrode 7. Thus, the reagent layer 11 may be formed.
[0021]
According to this configuration, when blood is supplied to the inlet 10 of the specimen supply path as a specimen liquid as a specimen, a certain amount of specimen is aspirated into the specimen supply path by capillary action through the air holes 13, and the counter electrode 6, the measurement electrode 5. It reaches on the detection electrode 7. The reagent layer 11 formed on the electrode is dissolved in blood as a sample, and an oxidation-reduction reaction occurs between the reagent and a specific component in the sample. Here, if the sample is correctly filled in the sample supply path, an electrical change occurs between the counter electrode 6 and the detection electrode 7. Thereby, it is confirmed that the specimen is sucked up to the detection electrode 7. In addition, since an electrical change also occurs between the measurement electrode 5 and the detection electrode 7, it may be confirmed that the specimen is sucked up to the detection electrode 7. After the specimen is aspirated to the detection electrode 7, the reaction between the specimen and the reagent is promoted for a certain period of time, and then a constant voltage is applied to the measurement electrode 5 and the counter electrode 6 or both the counter electrode 6 and the detection electrode 7. To do. Since it is a blood glucose level sensor, a current proportional to the glucose concentration is generated, and the blood glucose level can be measured from that value.
[0022]
Although the blood glucose level sensor has been described in the first embodiment, it can be used as a biosensor other than the blood glucose level sensor by changing the components of the reagent layer 11 and the specimen.
In the first embodiment, a biosensor having three electrodes has been described. However, the number of electrodes may be other than that.
[0023]
In the first embodiment, the second slits 4a and 4b are arc-shaped. However, the position and area of the reagent layer can be restricted, and the shape is limited to this shape as long as the accuracy of the electrode is not reduced. Is not to be done. For example, a straight line or a key shape may be used.
[0024]
As described above, the biosensor according to the first embodiment is a biosensor for quantifying a substrate contained in a sample solution, and includes an insulator substrate and the whole or a part of the insulator substrate. A plurality of electrodes created by providing a first slit in an electrically conductive layer formed by a sputtering method, and an arc-shaped first electrode for regulating the reagent application position and area provided in the electrically conductive layer. Two slits, a spacer disposed on the electrode and having a notch for forming a sample supply path for supplying a sample solution to the measurement electrode, and an enzyme provided on the electrode in the sample supply path And a second slit arranged on the spacer, the cover having an air hole leading to the sample supply path, and the spread of the applied reagent being restricted by the second slit. Therefore, when a reagent is applied on an electrode for forming a reagent layer, the reagent spreads uniformly, a reagent layer having no variation in position and area is formed, and accurate measurement without variation can be performed when measuring a specimen. It has the effect.
[0025]
【The invention's effect】
As described above, the biosensor according to claim 1 of the present invention is a biosensor for quantifying a substrate contained in a sample solution, and includes an insulator substrate and the entire surface of the insulator substrate. Alternatively, a plurality of electrodes formed by providing a first slit in an electrically conductive layer formed on a part, and a notch that is disposed on the electrode and forms a sample supply path for supplying a sample solution to the electrode a spacer having a section, the disposed on the electrode in the sample supply path, the cover having a reagent layer formed by reagents are dripped, disposed on the spacer, the air hole communicating with the specimen supply path And a second slit is provided around the position where the reagent on the electrode is dropped, so as to partially surround the position where the reagent is applied. Do not drop the reagent on the electrode. When the coating is applied, the reagent spreads uniformly, and a reagent layer with a predetermined area is formed at a predetermined position, so that a uniform reagent layer with no variation in position and area is formed, and accurate measurement without variation can be performed. It has the effect.
[0026]
Further, according to the biosensor according to claim 2 of the present invention, in the biosensor according to claim 1, the electrically conductive layer is formed on the insulating substrate by a sputtering method. Therefore, a highly accurate thin film can be formed, a highly accurate electrode can be created, and the measurement accuracy can be improved.
[0027]
According to the biosensor as set forth in claim 3 of the present invention, in the biosensor as set forth in claim 1 or 2, the first slit and the second slit are formed by a laser. Therefore, the area of each electrode can be defined with high accuracy. In addition, the distance between the electrodes can be made very short to reduce the size of the specimen supply path. In addition, there is an effect that the position and area of the reagent layer can be defined with high accuracy.
[0028]
Moreover, according to the biosensor according to claim 4 of the present invention, in the biosensor according to any one of claims 1 to 3, the second slit has an arc shape. Since the spread of the reagent is regulated by a slit equal to the shape of the spread of the reagent, there is an effect that the area and position of the reagent layer can be regulated more accurately.
[Brief description of the drawings]
FIG. 1 is a diagram illustrating perspective views of a biosensor according to a first embodiment in the order of creation steps.
FIG. 2 is a diagram showing a specimen supply path of the biosensor according to the first embodiment.
FIG. 3 is a diagram showing a perspective view of a conventional biosensor in the order of creation steps.
FIG. 4 is a diagram showing a sample supply path of a conventional biosensor.
[Explanation of symbols]
DESCRIPTION OF SYMBOLS 1 Board | substrate 2 Electrically conductive layer 3a 1st slit 3b 1st slit 3c 1st slit 3d 1st slit 4a 2nd slit 4b 2nd slit 5 Measuring electrode 6 Counter electrode 7 Detection electrode 8 Spacer 9 Notch Part 10 Entrance of specimen supply path 11 Reagent layer 12 Cover 13 Air hole 101 Substrate 102 Electrically conductive layer 103a Slit 103b Slit 103c Slit 103d Slit 105 Measuring electrode 106 Counter electrode 107 Detection electrode 108 Spacer 109 Notch 110 Entrance of specimen supply path 111 Reagent layer 112 Cover 113 Air hole

Claims (4)

A biosensor for quantifying a substrate contained in a sample solution,
An insulator substrate;
A plurality of electrodes formed by providing a first slit in an electrically conductive layer formed on the whole surface or a part of the insulator substrate;
A reagent layer disposed on the plurality of electrodes and formed by dropping a reagent; and
A spacer disposed on the electrode and reagent layer and having a notch for forming a sample supply path for supplying a sample solution to the electrode ;
Disposed before SL on the spacer, and a cover having an air hole communicating with the specimen supply path,
Around the position where the reagent is dripped on the electrode, a second slit is provided to partially surround the position where the reagent is applied,
A biosensor characterized by that. The biosensor according to claim 1, wherein
The electrically conductive layer is formed on the insulating substrate by a sputtering method.
A biosensor characterized by that. The biosensor according to claim 1 or 2,
The first slit and the second slit are formed by a laser.
A biosensor characterized by that. The biosensor according to any one of claims 1 to 3,
The second slit has an arc shape.
A biosensor characterized by that.

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