JP2001296267A - Forming method of membrane electrode, and biosensor equipped with same - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide a highly sensitive membrane electrode and a high- performance, inexpensive biosensor. SOLUTION: This highly sensitive membrane electrode which does not require pretreatment, such as surface grinding treatment or the like and having satisfactory wettability is manufactured, by forming a membrane electrode layer comprising a conductive material after forming a rough surface by colliding stimulated gas onto the substrate surface under a vacuum atmosphere. This high- performance and inexpensive biosensor is manufactured by developing a reagent layer on the membrane electrode.

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a biosensor for measuring a substance to be measured in a liquid sample, and a method for forming a thin film electrode suitable for the biosensor.

[0002]

2. Description of the Related Art As a biosensor for measuring a specific substance to be measured in a liquid sample, for example, Japanese Unexamined Patent Publication No. Hei.
As disclosed in Japanese Patent Publication No. 95, there is a method in which a blood glucose level or the like is obtained by measuring a current value obtained by a reaction between glucose in blood and a reagent such as glucose oxidase carried in a sensor.

FIG. 7 is a sectional view showing a schematic configuration of an electrode portion of such a conventional sensor for measuring blood glucose level. On a carbon electrode 2 formed by applying and drying a carbon paste by screen printing or the like on an insulating substrate 1 such as polyethylene terephthalate, a reaction reagent layer 3 containing an enzyme, an electron carrier and the like is carried. ing.

[0004]

In such a sensor, in order to improve the sensitivity of the sensor, it is necessary to increase the wettability between the reaction reagent layer and the carbon electrode and to improve the adhesion between them.

Conventionally, the surface of the electrode is polished or heat-treated after the formation of the carbon electrode, and the number of steps is large, so that the cost is increased and the accuracy of the sensor is also varied due to the variation of the polishing process. There was a problem.

[0006] The carbon paste used for screen printing is generally a composite material composed of a resin binder, graphite, carbon black, an organic solvent, etc., and depends on each raw material lot, production conditions at the time of kneading the paste, and the like. Strict control was required to mass-produce a stable sensor in which the characteristics of the paste tended to fluctuate.

[0007]

In order to solve the above-mentioned problems, the present invention provides a method for forming a roughened surface by impinging an excited gas on a substrate surface in a vacuum atmosphere and then forming the surface of the substrate on a conductive material. When a biosensor is obtained by providing a reagent layer on this electrode, a pretreatment such as a surface polishing treatment is not required as in the related art, so that the manufacturing cost can be reduced. Further, a highly sensitive biosensor having good surface wettability is provided.

[0008]

BEST MODE FOR CARRYING OUT THE INVENTION The invention according to claim 1 of the present invention is characterized in that, after the step of roughening the surface of the substrate by colliding the excited gas with the surface of the substrate in a vacuum atmosphere, Forming a thin film electrode layer made of a conductive material on the roughened surface of the substrate, without the need for pretreatment such as surface polishing treatment, and It is possible to obtain a thin film electrode having high adhesion between the substrate and the electrode layer, and when a reagent is formed on the electrode, the biosensor has good wettability with the reagent and good adhesion between the reagent and the electrode layer. Can be configured.

According to a second aspect of the present invention, the step of roughening the surface of the substrate includes the step of placing the substrate in a vacuum chamber, the step of evacuating the vacuum chamber, and the step of filling gas. And a step of exciting and ionizing the gas to collide with the substrate, wherein the method of claim 1 is further embodied. is there.

According to a third aspect of the present invention, there is provided the method of forming a thin film electrode according to the first aspect, wherein the substrate is made of a resin material. The surface treatment becomes easier.

According to a fourth aspect of the present invention, in the step of evacuating the vacuum chamber, the degree of vacuum is 1 × 10 ^{-1 to} 3 × 10 ^-3.
A method of forming a thin-film electrode according to claim 2, wherein the method is within the range of Pascal, and is a more specific example of the method of claim 2.

According to a fifth aspect of the present invention, there is provided the method for forming a thin film electrode according to the second aspect, wherein the gas is an inert gas, wherein the surface of the thin film electrode is not denatured. Can be roughened.

According to a sixth aspect of the present invention, the inert gas is a rare gas such as argon, neon, helium, krypton, or xenon, or nitrogen. The method is more specific about the type of the inert gas.

According to a seventh aspect of the present invention, the step of forming a thin-film electrode layer made of a conductive material on a substrate includes the steps of placing the substrate in a vacuum chamber and evacuating the vacuum chamber. And a step of filling with a gas, and exciting and ionizing the gas to collide with a conductive substance, thereby knocking out atoms of the conductive substance and forming a film on the substrate. The method for forming a thin film electrode according to any one of claims 1 to 6, wherein a pretreatment such as a surface polishing treatment is not required, and a thin film electrode having high adhesion to a substrate can be obtained. When a reagent is formed on the electrode, a biosensor having good wettability with the reagent and good adhesion between the reagent and the electrode layer can be formed.

According to an eighth aspect of the present invention, the step of forming a thin film electrode layer made of a conductive material on a substrate includes the step of placing the substrate in a vacuum chamber and the step of evacuating the vacuum chamber. 7. A method for forming a thin-film electrode according to claim 1, further comprising: heating and evaporating a conductive substance to form a film on the substrate. A pretreatment such as a polishing treatment is not required, and a thin film electrode having high adhesion to a substrate can be obtained. When a reagent is formed on the electrode, a biosensor having good wettability with the reagent and good adhesion between the reagent and the electrode layer can be formed.

According to a ninth aspect of the present invention, in the step of evacuating the vacuum chamber, the degree of vacuum is 1 × 10 ^{-1 to} 3 × 10 ^-3.
A method of forming a thin-film electrode according to claim 7 or 8, wherein the method is pascal, and the method of claims 7 and 8 is further embodied.

According to a tenth aspect of the present invention, the gas is an inert gas.
9. The method for forming a thin-film electrode according to claim 7, wherein
Is embodied.

According to the eleventh aspect of the present invention, the inert gas is a rare gas such as argon, neon, helium, krypton, or xenon, or nitrogen. A method, wherein the method comprises:
This is a concrete example of the contents of 0.

According to a twelfth aspect of the present invention, there is provided a method of making a substrate surface rough by bombarding a gas excited in a vacuum atmosphere with a thin film electrode layer made of a conductive material on the substrate. 12. The method of forming a thin-film electrode according to claim 1, wherein the step of forming is performed continuously in the same space, wherein two steps are performed simultaneously in the same space. As a result, the number of manufacturing steps can be reduced and the production tact time can be improved.

According to a thirteenth aspect of the present invention, there is provided the method of forming a thin film electrode according to the first aspect, wherein the conductive substance is a noble metal or carbon. By using it as an electrode material, mass production of a stable electrode that is hardly influenced by manufacturing conditions and has a small difference between material lots is possible.

According to a fourteenth aspect of the present invention, there is provided the method for forming a thin film electrode according to the first aspect, wherein the thickness of the thin film electrode is in a range of 5 to 100 nm. By reducing the thickness, it is possible to improve the production tact time and reduce the cost of the material to reduce the cost of the electrode.

According to a fifteenth aspect of the present invention, there is provided a biosensor including a thin-film electrode formed by the method according to any one of the first to fourteenth aspects, wherein a reagent layer is formed on the thin-film electrode. This is a biosensor characterized by the fact that the thin-film electrodes reflect the unevenness of the surface of the substrate that has been processed into a rough surface, thereby increasing the wettability and adhesion between the electrodes and the reagent, and providing a high-performance biosensor. Can be realized. In forming the electrodes of this biosensor, the functions and effects of the above-described claims are obtained.

The invention according to claim 16 of the present invention is the biosensor according to claim 15, wherein the reagent layer contains an enzyme.

The invention according to claim 17 of the present invention is the biosensor according to claim 16, wherein the reagent layer contains an electron carrier.

The invention according to claim 18 of the present invention is the biosensor according to claim 17, wherein the reagent layer contains a water-soluble polymer.

(Embodiment 1) An embodiment of the present invention will be described below with reference to FIGS.

FIG. 1 is a schematic diagram of a thin film electrode obtained by the present invention and a biosensor in which a reaction reagent layer is developed thereon. The most different point from the conventional structure of FIG.
The surface of an insulating resin substrate 1 such as polyethylene terephthalate or polycarbonate is subjected to a surface roughening treatment for improving the adhesion between the substrate 1 and the electrode layer 2 and between the electrode layer 2 and the reagent layer 3. It is. The material constituting the electrode layer 2 is a simple material made of noble metal or carbon,
Another difference is that the thickness of the electrode layer 2 is controlled to 5 to 100 nm.

Hereinafter, a specific method of the surface roughening treatment of the surface of the substrate 1 will be described. Materials suitable for the substrate 1 include polyethylene terephthalate, polycarbonate, polybutylene terephthalate, polyamide, polyvinyl chloride,
Examples thereof include polyvinylidene chloride, polyimide, and nylon. After placing the substrate 1 in a vacuum chamber, the substrate 1 is evacuated to a certain degree of vacuum (a range of 1 × 10 ^{−1 to} 3 × 10 ⁻³ Pascal is sufficient). Thereafter, the vacuum chamber is filled with an inert gas (the degree of vacuum after filling is in the range of about 0.1 to 10 Pascal), and when a high frequency voltage of about 0.01 to 5 KV is applied, the inert gas is excited. And ionized, the substrate 1
It is beaten to the surface. This ion has a high kinetic energy and is very short (about 0.1 to 10 seconds)
By applying the high frequency voltage, a sufficient surface roughening effect can be obtained. The same surface roughening effect can be obtained by applying a DC voltage or the like in addition to the application of the high-frequency voltage.

As the inert gas, nitrogen can be used in addition to the rare gases of argon, neon, helium, krypton, and xenon. Also, when an active (reactive) gas such as oxygen is used, the substrate surface can be roughened, but an oxide film is formed on the substrate surface, and the electrode characteristics and sensor response This is not desirable because it may adversely affect the characteristics.

Next, a method for forming a thin-film electrode layer made of a conductive substance on the surface of the substrate 1 subjected to the surface roughening treatment will be specifically described. As in the case of the surface roughening treatment of the surface of the substrate 1, the chamber 1 is evacuated to a certain degree of vacuum (the range may be 1 × 10 ^{-1 to} 3 × 10 ^-3 Pa). Thereafter, the vacuum chamber is filled with an inert gas (the degree of vacuum after the filling is in the range of about 0.1 to 10 Pascal), and a high-frequency voltage of about 0.01 to 5 KV is applied, whereby the inert gas is applied. Are excited and ionized. The ionized gaseous gas collides with a target plate made of a conductive material to strike out atoms of the conductive substance, and the atoms are formed on the substrate to form a thin film electrode layer. Further, it is also possible to form a thin film electrode layer by heating and evaporating a conductive substance after evacuation and forming a film of the vapor on the substrate. The above-mentioned typical method is sputtering vapor deposition, and a typical method described later is vacuum vapor deposition.

Here, examples of the material of the conductive material forming the target plate include noble metals such as palladium, platinum, gold and ruthenium, carbon and the like. This makes it possible to mass-produce a stable electrode that is hardly influenced by the difference between material lots.

The substrate surface roughening process and the thin film electrode forming process can be performed discontinuously in an independent space. However, as shown in FIG. 2, the substrate surface roughening process is performed. By continuously performing the process of forming the thin film electrode in the same space, the number of manufacturing steps can be reduced, the productivity can be improved by improving the production tact time, and the cost of the biosensor can be reduced accordingly.

As described above, when the two processes are continuously performed in the same space, it is difficult to perform vacuum deposition, and high-frequency sputtering deposition, bias sputtering deposition, asymmetric AC sputtering deposition, ion plating, and the like are effective. .

It is needless to say that the manufacturing cost can be reduced by reducing the thickness of the electrode layer as much as possible, but by reflecting the rough surface of the substrate directly as the rough surface of the electrode layer, The effect that the adhesiveness between the electrode layer 2 and the reaction reagent layer 3 composed of an enzyme, an electron carrier or the like is remarkably improved is also obtained.

Here, in order to reflect the rough surface of the substrate surface as the rough surface of the electrode layer, the thickness of the electrode layer must be 100 nm or less. To provide, the thickness of the electrode layer is 5-5.
Desirably, it is 0 nm.

[0036]

EXAMPLE 1 A high frequency voltage having a frequency of 13.56 MHz was applied at a power of 100 W for a certain period of time on an insulating substrate 1 made of polyethylene terephthalate to perform a surface roughening treatment. A noble metal thin film electrode in which palladium was formed to a thickness of about 10 nm on the surfaced substrate under the same conditions as described above was produced.

FIG. 4 shows that the application time of the high-frequency voltage is 0-60.
5 seconds shows the change in the wetting index (surface tension) of the substrate surface and the adhesion between the electrode layer and the substrate due to the surface roughening treatment for 5 seconds (0 second is a state where the surface roughening treatment is not performed). This indicates that the surface of the substrate is roughened by the above application, and that the surface wettability is improved and the adhesion between the electrode layer and the substrate is increased. Note that the present embodiment is a result at a high frequency voltage of 100 W, and the processing time can be further reduced by increasing the high frequency voltage.

The evaluation of the adhesion here is based on JIS 5600.
Performed in accordance with -5-10 (General Test Method for Paint: Mechanical Properties of Coating Film: Abrasion Resistance), and the adhesion value in the figure is until the time when the palladium thin film was worn away and the substrate surface was exposed. The number of stroke reciprocations indicates that the larger the value, the higher the adhesion.

FIG. 5 shows the relationship between the thickness of the palladium thin film and the wetting index (surface tension) of the electrode surface. Here, a substrate surface roughening treatment condition was used in which the high-frequency voltage was 100 W, the application time was 5 seconds, and the thickness of the palladium layer was arbitrarily adjusted in the range of 5 to 1000 nm. As is clear from FIG. 5, when the thickness of the palladium layer is in the range of 5 to 50 nm, the wetting index of 54 dyn / cm on the surface of the substrate subjected to the surface roughening treatment is maintained as it is.
A slight decrease in the wetting index of 52 dyn / cm by 00 nm is observed. If it exceeds 100 nm, the wetting index decreases to 48 dyn / cm, and thereafter becomes stable at that value. This indicates that up to a thickness of 100 nm, the rough surface of the substrate surface reflects the rough surface of the electrode surface.
It is shown that when the value exceeds the value, the wettability of the electrode material itself (palladium in this embodiment) is reflected.

Next, carboxymethylcellulose as a water-soluble polymer, glucose oxidase (GOD) as an enzyme, and potassium ferricyanide as an electron carrier were placed on a thin-film electrode with a 10 nm-thick palladium layer prepared under the above conditions. After the formation of the reaction reagent layer including the above, a biosensor for measuring a blood glucose level as shown in FIG.

FIG. 6 shows a blood glucose concentration of 40 to 600 mg.
This is a comparison of sensor sensitivity at / dl. The sensor sensitivity here means that after blood is sucked into the capillary,
After accelerating the reaction between the reaction reagent and glucose in the blood for about 25 seconds, a constant voltage was applied between the working electrode and the counter electrode terminal, and the current value was obtained 5 seconds after that. Since the conventional sensor and the sensor of the present embodiment have different electrode materials, the applied voltage was 0.5 V for the conventional carbon paste electrode and 0.2 V for the palladium thin film electrode of the present embodiment.

The number of measurements was n =
It was set to 10. As is clear from FIG. 6, the sensor sensitivity of this embodiment in which the electrode surface is not subjected to polishing treatment or heat treatment is subjected to polishing treatment or heat treatment conventionally required to increase the sensor sensitivity. It was confirmed that the sensor had the same or higher sensitivity.

Table 1 shows a comparison of the repetition accuracy (CV value) at the time of the ten measurements described above. In the conventional sensor, the deterioration of the CV value due to the variation in the polishing process and the like was remarkably recognized. In addition, it was confirmed that the sensor according to the present embodiment has excellent accuracy in which variations among individual sensors are reduced.

[0044]

[Table 1]

[0045]

As described above, according to the present invention, the substrate surface is roughened by bombarding a gas excited in a vacuum atmosphere, and then a thin film electrode layer made of a conductive material is formed. A high-performance and inexpensive biosensor can be formed by forming a thin film electrode having good surface wettability and high sensitivity which does not require pre-treatment such as surface polishing treatment, and developing a reagent layer on the thin film electrode. It is possible to do.

[Brief description of the drawings]

FIG. 1 is a cross-sectional view illustrating a schematic configuration of a biosensor according to an embodiment of the present invention.

FIG. 2 is a schematic configuration diagram showing a step of forming a thin film electrode in the same embodiment.

FIG. 3 is an exploded perspective view showing a biosensor according to one embodiment of the present invention.

FIG. 4 is a graph showing the change in the wetting index of the substrate surface and the adhesion between the electrode layer and the substrate in one embodiment of the present invention.

FIG. 5 is a graph showing the change in the wetting index of the substrate surface and the adhesion between the electrode layer and the substrate in one embodiment of the present invention.

FIG. 6 is a graph comparing sensor whole blood sensitivity in one embodiment of the present invention.

FIG. 7 is a sectional view showing a schematic configuration of a conventional biosensor.

[Explanation of symbols]

 Reference Signs List 1 substrate 2 electrode layer 3 reaction reagent layer 4 vacuum tank 5 (substrate) sending roll 6 (substrate) winding roll 7 electrode for surface roughening treatment 8 cooling roller 9 cathode / target 10 gas inlet 11 measuring electrode 12 counter electrode 13 spacer 14 Cover 15 Measurement electrode terminal 16 Counter electrode terminal

Claims (18)

[Claims] In a vacuum atmosphere, after a step of roughening the surface of the substrate by colliding the excited gas with the surface of the substrate, a conductive material is applied on the roughened surface of the substrate. Forming a thin-film electrode layer. 2. The step of roughening the surface of the substrate includes the steps of: placing the substrate in a vacuum chamber; and evacuating the vacuum chamber.
The method for forming a thin-film electrode according to claim 1, further comprising: filling a gas; and exciting and ionizing the gas to collide with the substrate. 3. The method according to claim 1, wherein the substrate is made of a resin material. 4. The step of evacuating the vacuum chamber is performed at a degree of vacuum of 1.times.
3. The method for forming a thin film electrode according to claim 2, wherein the thickness is in a range of 10 ^{-1 to} 3 × 10 ^-3 Pascal. 5. The method according to claim 2, wherein the gas is an inert gas. 6. The method according to claim 5, wherein the inert gas is a rare gas of argon, neon, helium, krypton, or xenon or nitrogen. 7. A step of forming a thin-film electrode layer made of a conductive substance on a substrate, comprising the steps of: placing the substrate in a vacuum chamber; evacuating the vacuum chamber;
7. The method according to claim 1, further comprising the step of: exciting and ionizing the gas to collide with a conductive substance to strike out atoms of the conductive substance and form a film on the substrate. The method for forming a thin film electrode according to the above. 8. The step of forming a thin film electrode layer made of a conductive material on a substrate includes the steps of placing the substrate in a vacuum chamber, evacuating the vacuum chamber, and heating and evaporating the conductive substance. 7. A method for forming a thin film electrode according to claim 1, further comprising the step of: forming a film of the vapor on the substrate. 9. The process of evacuating the vacuum chamber is performed at a degree of vacuum of 1.times.
The method for forming a thin film electrode according to claim 7, wherein the pressure is 10 ^{−1 to} 3 × 10 ⁻³ Pascal. 10. The method for forming a thin film electrode according to claim 7, wherein the gas is an inert gas. 11. The method according to claim 10, wherein the inert gas is a rare gas such as argon, neon, helium, krypton, or xenon, or nitrogen. 12. A step of making a surface of a substrate rough by colliding a gas excited in a vacuum atmosphere and a step of forming a thin film electrode layer made of a conductive material on the substrate in the same space. The method for forming a thin-film electrode according to claim 1, wherein the method is performed continuously. 13. The method according to claim 1, wherein the conductive substance is a noble metal or carbon. 14. The method according to claim 1, wherein the thickness of the thin-film electrode is in the range of 5 to 100 nm. 15. A biosensor comprising a thin-film electrode formed by the method according to any one of claims 1 to 14,
A biosensor comprising a reagent layer formed on the thin film electrode. 16. The biosensor according to claim 15, wherein the reagent layer contains an enzyme. 17. The biosensor according to claim 16, wherein the reagent layer contains an electron carrier. 18. The biosensor according to claim 17, wherein the reagent layer contains a water-soluble polymer.