JPH06296595A - Integrated composite electrode - Google Patents

Abstract

(57) [Abstract] [Purpose] To provide an integrated composite electrode which is capable of performing multipoint simultaneous stimulation and recording of nerve cells for a long period of time and has excellent responsiveness. [Structure] An ITO film is vapor-deposited on the entire surface of a hard glass insulating substrate 3, the center of each electrode 1 is located at each intersection on an 8 × 8 grid, and the distance between the centers of the nearest electrodes of each electrode is set. Are equal, and the ITO film is etched into a shape in which the lead wires 2 extend radially. Then, a negative photosensitive polyimide is spin-coated as the insulating layer 4, and an insulating layer pattern is formed by exposure so that a square hole 5 having a side of 50 μm is formed at the center of each electrode. Furthermore, in the exposed part of each electrode (that is, inside a square of 50 μm on a side),
Gold 6 is vapor-deposited to a film thickness of 500 angstrom. The contact point between the lead wire 2 and the external circuit in the vicinity of the end opposite to the electrode 1 is coated with gold 7 and nickel 8.

Description

Detailed Description of the Invention

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to an integrated composite electrode having multiple electrodes, which is used in the field of electrical measurement of biological activity, particularly neurophysiology for measuring electrical activity of nerve cells.

[0002]

2. Description of the Related Art In recent years, medical examination of nerve cells and possibility of application as an electric element have been actively conducted. When nerve cells are activated, action potentials are generated. The action potential is caused by a change in ion concentration inside and outside the cell membrane with a change in ion permeability of nerve cells. Nerve activity is being detected and studied by measuring potential changes associated with changes in ion concentration (that is, ion current) in the vicinity of nerve cells using electrodes.

Conventionally, to measure the electrical activity of nerve cells, a recording electrode made of a glass electrode or the like and a stimulation electrode made of a metal electrode or the like are inserted inside or between cells, and a stimulation current is applied from the stimulation electrode. The electrical activity of nerve cells when (or voltage) was applied was usually measured by a recording electrode.

In addition to this, for example, the cell body is pierced by a thin glass suction electrode, the inside of the cell body is circulated by the liquid in the glass suction electrode, and an electric signal is given from this glass suction electrode to electrically connect the cell membrane. There are many variations such as the so-called patch clamp method for observing the characteristics.

Further, an electrode having a diameter of 15 to 20 μm is formed of a conductive substance such as ITO (indium tin oxide) on an insulating base, and nerve cells are cultured on the electrode to puncture the electrode. The present inventors have separately proposed a method of applying an electrical stimulus to a cell and recording the electrical activity of a nerve cell without inserting the cell.

As an improved method, the diameter of the electrode is set to 2
When the thickness is 0 to 200 μm, the potential difference generated between the electrodes when a constant current stimulus is applied to the nerve cells is reduced, and as a result, the ITO is less likely to be destroyed, and it is possible to observe for a longer period of time. Proposed separately.

[0007]

In the above-mentioned conventional technique and its modified method, since an electrode such as a glass electrode which has a size considerably larger than that of a cell is used, a space limitation is mainly caused. Due to the limitation of operation accuracy, there is a problem that it is very difficult to perform multipoint simultaneous measurement in which two or more recording electrodes are inserted into one sample at a time and the electrical activity of nerve cells is recorded.

In order to study the function of the entire neural network, it is necessary to record the activity of many nerve cells at the same time. As the number of measurement points increases, the degree of difficulty increases, and the observation between multiple cells is increased. There was a problem that it was difficult to do.

Furthermore, since it is necessary to insert electrodes of glass, metal or the like into cells or between cells, there is a problem that damage to cells is large and it is difficult to perform measurement for a long time of several hours or more. It was

On the other hand, if a circular (or square) electrode having a diameter (or one side) of 15 to 20 μm is formed on the insulating substrate by using a conductive material such as ITO, signal transmission across multiple cells can be achieved. Observation becomes possible. However,
Since the electrode area is small as 177μm ² ~400μm ^2,
The electrode resistance at the interface of the culture solution is several MΩ, and the stimulus is usually given by a constant current. Therefore, if the electric resistance is large, an extremely large potential difference occurs between the electrodes. If it is given, the ITO is destroyed, which makes it difficult to observe for a long period of time.

The electrode area is 300 μm ^{2 to} 4000.
When it is set to 0 μm ² , the electrode resistance at the interface of the culture solution becomes small, so that the potential difference generated between the electrodes becomes relatively small. No destruction of ITO was observed microscopically even when a stimulating current was applied for a long period of time. However, when a stimulating current was applied from a certain electrode and the potential change due to stimulation was recorded at another electrode, a large change was observed in the recorded waveform before and after long-term stimulation. That is, after long-term stimulation, the effect of the stimulation current application on the recording waveform (ie, artifact) was larger than that before long-term stimulation. It is considered that the cause of the waveform change is that the electrode surface is polarized. In the worst case, the electrical activity of nerve cells was hidden by artifacts and became unmeasurable. In addition, there is a problem that it is difficult to compare the nerve activity intensities before and after the long-term stimulation even when the artifact is not so large.

The present invention solves the above-mentioned conventional problems,
Simultaneous multi-point simultaneous stimulation and measurement of nerve cells, etc., enabling signal transmission observation between multiple cells for several hours or more, suppressing the occurrence of artifacts due to the application of stimulation current, and recording potential waveforms before and after long-term stimulation It is an object of the present invention to provide an integrated composite electrode that enables the comparison of

[0013]

In order to solve the above-mentioned problems, an integrated composite electrode of the present invention comprises a plurality of electrodes having the same closest distance between electrodes on an insulating substrate, and leads from the electrodes. A wiring portion in which wires are arranged substantially radially and an insulating layer for covering the lead wires are provided, and an electrode area is 3 ×.
The range is 10 ² μm ² or more and 4 × 10 ⁴ μm ² or less,
The surface resistance of the electrode portion is 10 Ω / cm ² or less.

In the integrated composite electrode of the present invention,
The distance between the nearest electrodes is preferably 10 μm or more and 1000 μm or less. In the integrated composite electrode of the present invention, the insulating layer that covers the lead wire has a hole on each electrode, and substantially the entire surface of the insulating substrate except for the vicinity of the contact point of the lead wire with the external circuit. It is preferable that the insulating layer is provided on the.

Further, in the integrated composite electrode of the present invention, it is preferable that a plurality of electrode central portions are located at respective intersections on an 8 × 8 grid.

[0016]

The integrated composite electrode of the present invention comprises a plurality of electrodes having the same distance between the closest electrodes on the insulating base, and a wiring portion in which lead wires are arranged in a substantially radial manner from the electrodes, Since an insulating layer covering the lead wire is provided, the electrode area is in the range of 3 × 10 ² μm ² or more and 4 × 10 ⁴ μm ² or less, and the surface resistance of the electrode portion is 10 Ω / cm ² or less, When giving a signal to a nerve cell cultured on the integrated composite electrode of the invention and measuring the transmission of the signal between cells at the same time,
By adjusting the distance between the closest electrodes to the length of the nerve cell to be measured (that is, the cell body, dendrites, and axons), and arranging these electrodes at equal intervals, one cell body The probability that the cell body placed on the upper side and via the cell projection extending from this cell body is located on the adjacent electrode is high. Therefore, signal transmission between adjacent cell bodies can be detected.

Moreover, since the lead wires extended from the electrodes are arranged in a substantially radial pattern, the capacitance component between the lead wires is smaller than that in the case where the lead wires are arranged in parallel, and a pulse which is an electric signal is obtained. Since the collapse of the signal waveform can be reduced and the time constant of the circuit is reduced, the responsiveness to a fast pulse signal is improved and the followability to a fast component of nerve cell activity is improved.

Further, by adjusting the electrode area within the range of 3 × 10 ² μm ² or more and 4 × 10 ⁴ μm ² or less, the cells are electrically stimulated for a long time of several hours or more, and the electrical activity of the cells is increased. Can be measured.

Further, the surface resistance of the electrode portion is 10 Ω / cm ²
Because of the following, when a stimulation current is applied to a nerve cell for a long time with one electrode and the electrical activity (potential change) of the nerve cell corresponding to the stimulation current is recorded with another electrode, the polarization of the stimulation electrode surface is Since it does not easily occur, the influence (that is, artifact) of the stimulation current on the electrogram waveform is reduced. In particular,
Since the artifact is small and the morphology does not change even after the stimulation current is applied for a long period, it is possible to compare the electrical activity of nerve cells before and after the long-term stimulation.

In the integrated composite electrode of the present invention, the distance between the closest electrodes is 10 μm or more and 1000 μm.
By the following preferred embodiment, the neurite length of the nerve cell is generally within this range, so that the cell body is located on the electrode and is highly likely to be bound via the neurite. The distance between the electrodes is convenient for the measurement of nerve cells.

In the integrated composite electrode of the present invention, the insulating layer covering the lead wire has holes on each electrode, and the insulating layer is provided except in the vicinity of the contact portion of the lead wire with the external circuit. By adopting a preferable mode in which the insulating layer is provided on almost the entire surface of the substrate, as compared with the case where the insulating layer is selectively provided only on the lead wire, an insulating material made of a photosensitive resin is used to cover almost the entire surface. Applying this resin and removing the insulating layer on each electrode by photo-etching method, you can easily form the required insulating layer by photo-etching such as making holes so that the electrodes are exposed, thus facilitating production. This is preferable because it is possible to reduce the probability of insulation failure.

Furthermore, in the integrated composite electrode of the present invention, a plurality of electrode central portions are located at respective intersections on an 8 × 8 grid, so that the lead wires are substantially omitted from the electrode of the present invention. It is preferable because the maximum number of electrodes that can be arranged radially can be obtained.

[0023]

EXAMPLES As the insulating base material used in the present invention, a transparent base is preferable because it needs to be observed under a microscope after cell culture. Glass such as quartz glass, lead glass, borosilicate glass, or inorganic such as quartz. Substances, or organic substances having transparency such as polymethylmethacrylate or its copolymer, polystyrene, polyvinyl chloride, polyester, polypropylene, urea resin, melamine resin, etc. When added, inorganic substances are preferable.

Examples of the electrode material used in the present invention include indium tin oxide (ITO), tin oxide, Cr, Au,
Cu, Ni, Al, etc. can be used. In particular, when ITO or tin oxide is used, the electrode becomes slightly yellowish and transparent, and the visibility of nerve cells under a microscope is good, which is advantageous in experimental operation, but ITO is especially a good conductor. It is desirable because it is sex.

The same material can be applied to the lead wire material,
Again, ITO is preferable for the same reason as the electrode material. Although not particularly limited, the thickness of these electrodes and lead wires is usually about 500 to 5000 angstroms, and usually, these materials are vapor-deposited on an insulating substrate and a desired pattern is formed by etching using a photoresist. Can be formed into

The insulating layer material for insulating the lead wire used in the present invention is, for example, polyimide (P
Examples of the transparent resin include transparent resin such as I) resin, epoxy resin, acrylate resin, polyester resin, and polyamide resin.

An insulating layer is formed by applying these resins on a lead wire by a usual method. It is preferable that the insulating layer material is a photosensitive resin such as a photopolymerizable resin because it is possible to form a pattern such as forming a hole in the insulating layer portion on the electrode to expose the electrode as described above.

Particularly, when the insulating layer material is PI and the cells to be cultured are nerve cells, it is desirable because it shows good growth. Further, among the PIs, negative photosensitive polyimide (NPI) is preferable because holes can be formed in an electrode shape by using a photoetching process after applying the negative photosensitive polyimide on substantially the same surface as in the pattern formation of the wiring portion.

The thickness of the insulating layer is not particularly limited as long as it can impart an insulating property, but is usually 0.1 to 10 μm, preferably 1 to 5 μm.

Using the integrated composite electrode of the present invention, cells were directly cultured and the electrical activity of the cells was measured and recorded. The size of the cell body or the length of cell projections such as dendrites and axons varies depending on the culture conditions or the type of cells, but the closest interelectrode distance of the integrated composite electrode is 10 to 1000 μm.
m is preferred. If the distance between the electrodes is less than 10 μm, the cell bodies are too close to each other, and thus the probability that the cell bodies are adjacent to each other via the cell projections is reduced, and the wiring of the lead wire is also difficult. On the other hand, if the thickness exceeds 1000 μm, the lead wire can be easily wired, but the cell protrusion rarely extends up to about 1000 μm, and thus the probability that the cell body is located on the electrode is reduced. Even under general conditions, the length of cell projections of cultured cells is about 200 to 300 μm on average in the case of mammalian central nerve cells, so the distance between electrodes is preferably about 200 to 300 μm.

The electrode area is required to have a certain size or more because it is necessary to reduce the resistance at the interface with the culture solution in order to avoid electrode destruction when applying electrical stimulation to cells for a long period of time. . However, when the electrode area increases and the resistance at the interface with the culture solution decreases,
The measured electrical activity of the cells is reduced and the S / N ratio is reduced. That is, assuming that the current value I is constant, I = V
Since / R, the smaller the resistance value R, the smaller the change in the measured potential V. That is, the electrical activity of the measured cell is reduced and the S / N ratio is reduced. Therefore, the electrode area needs to be carefully adjusted, and in the case of a circular electrode, the diameter is more than 20 μm and 200 μm or less, and particularly 100 μm to 200 μm is preferable.

The surface resistance of the electrode portion is 10 Ω / cm.
^A metal was coated on the upper surface of the ITO in order to make it ² or less. As the coating material, Ag, Al, Bi, Au, Cu, C
Although r, Pt, Co, etc. can be used, it is preferable to use Au, Pt in consideration of low toxicity to nerve cells. The thickness of the coat is not particularly limited,
It is about 500 Å, and usually, these materials can be formed into a desired pattern by vapor deposition on an insulating substrate and etching using a photoresist.

Further, according to the above-described preferred embodiment of the present invention, the pores in the insulating layer of the integrated composite electrode simultaneously provide electrical stimulation to the cell bodies cultured on the integrated composite electrode,
In order to detect electrical activity from adjacent cell bodies, the electrodes are formed to expose the electrodes and are located at the center of the electrodes.

Further, by arranging the lead wires extended from the electrodes in a substantially radial shape, a capacitance component between the lead wires is eliminated, noise is reduced, and measurement accuracy is improved. Further, when the electrode central portion of the integrated composite electrode of the present invention is located at each intersection on a concentric circle or a lattice of 8 × 8 or less,
It is desirable to provide electrodes at each intersection on the 8 × 8 grid from the viewpoint that the lead wires can be arranged in a radial pattern, and in particular, as many electrodes as possible are configured and multi-point simultaneous stimulation / recording is performed.

The integrated composite electrode of the present invention will be described in more detail with reference to specific examples below. Example 1 FIG. 1 is a plan view showing a pattern of a wiring part of an integrated composite electrode of the present example in which an electrode 1 and a lead wire 2 are formed on an insulating substrate 3 without an insulating layer. FIG. 2 is a partially cutaway view of a plan view of only the insulating layer formed on the member shown in FIG. FIG. 3 is a sectional view of a part of the integrated composite electrode of this embodiment. Hereinafter, description will be given with reference to these drawings.

First, the production of the composite electrode wiring portion will be described. The insulating substrate 3 of the integrated composite electrode is made of 50 × 50 × 1 mm hard glass (“IWAKI CODE 7740 GLASS”) as a transparent insulating material having high mechanical strength.
The same applies hereinafter) (manufactured by Iwaki Glass Co., Ltd.).

ITO is used as the material for the electrode 1 and the lead wire 2, and about 100 is formed on the entire surface of the insulating base 3 made of hard glass.
It was vapor-deposited to a thickness of 0 angstrom and then washed. Next, the center of each electrode 1 is located at each intersection (the position as shown by 5 in FIG. 2) on the 8 × 8 lattice, and the distance between the centers of the nearest electrodes of each electrode is equal, Moreover, exposure is performed using a photoresist so that the lead wire 2 has a pattern of the electrode 1 and the lead wire 2 having a radially extended shape, and pure water 5
IT in a mixed solution of 0, 50 hydrochloric acid and 1 nitric acid in a volume ratio
After etching O, the photoresist was removed. A wiring portion was formed in which the diameter of the electrode 1 was 60 μm, the width of the lead wire 2 was 30 μm, and the distance between the electrode centers was 300 μm.

Next, negative photosensitive polyimide (hereinafter abbreviated as NPI) is spin-coated as the insulating layer 4 so that the thickness after drying is 1 μm, and 1 is formed at the center of each electrode of the wiring portion as shown in FIG. The insulating layer pattern was formed by exposure so that square holes 5 having sides of 50 μm were formed. Furthermore, the exposed part of each electrode (that is, 50 μm on each side)
Gold 6 was vapor-deposited so that the film thickness would be 500 angstrom.

The contacts with the external circuit in the vicinity of the ends of the lead wire 2 in the direction opposite to the electrode 1 were coated with gold 7 and nickel 8 to improve the durability. In this embodiment, the electrodes 1 and the leads 2 are made of ITO, the insulating layer is made of NPI,
Although gold was used as the electrode surface coating material, it has already been described that the material used is not limited to these.

The process for constructing the integrated composite electrode of the present invention is not limited to the method of this embodiment. Example 2 Next, culture of nerve cells on the integrated composite electrode will be described.

On the integrated composite electrode constructed as in Example 1, rat cerebral visual cortex was cultured as nerve cells. Hereinafter, the culture method will be described in detail. (A) 16 to 18 days after the gestation, the brain of the SD rat fetus was extracted and ice-cooled Hanks balanced salt solution (hereinafter referred to as HBB).
Immerse in S). (B) A visual cortex is cut out from the brain in ice-cold HBBS and transferred into an Eagle minimum essential medium (hereinafter abbreviated as MEM) solution. (C) In the MEM solution, the visual cortex is cut as finely as possible to be 0.2 mm square at the maximum. (D) The finely cut visual cortex was placed in a centrifuge tube (test tube for centrifugation), washed three times with HBBS containing no calcium and magnesium (hereinafter abbreviated as CMF-HBBS), and then dispersed in an appropriate amount of the same solution. To do. (E) Trypsin CMF-in the centrifuge tube of (d) above.
HBBS solution (0.25 wt%) is added to double the total volume. While gently stirring, the enzyme reaction was carried out at 37 ° C. for 15 to 20 minutes while keeping a constant temperature. (F) DMEM / F-12 mixed medium prepared by mixing Dulbecco's modified Eagle medium (DMEM) containing 10 vol.% Fetal calf serum (FCS) and HamF-12 medium at a volume ratio of 1: 1, and the above (e) was used. Into the centrifuge tube, double the total volume. Using a Pasteur pipette with a small tip, use a Pasteur pipette and gently repeat pipetting (up to about 20 times) to loosen the cells. (G) 9806.65 m / sec ² (ie 1000
Centrifuge at g) for about 5 minutes. After the centrifugation, the supernatant was discarded and the precipitate was added to DMEM / F-1 containing 5 vol.% Of FCS.
2 Resuspend in mixed medium. (H) The above (G) and (H) are repeated two more times (three times in total). (I) The finally obtained precipitate is suspended in a DMEM / F-12 mixed medium containing 5 vol.% FCS, and the cell concentration in the suspension is measured using a red blood cell counter. The cell concentration is adjusted to 2-4 × 10 ⁶ cells / ml using the same medium. (V) 5 vol.% In advance in a cell culture well configured by adhering a plastic cylinder with a diameter of 25 mm and a height of 6 mm on the integrated composite electrode by aligning the center of the composite electrode with the center of the plastic cylinder. Add 500 μl of DMEM / F-12 mixed medium containing FCS, and add CO
₂ Warm in an incubator (air concentration 95 vol.%, CO ₂ concentration 5 vol.%, Relative humidity 97%, temperature 37 ° C.). (L) 100 μl of the cell concentration-adjusted suspension is gently added to the well of (nu) above, and the suspension is again placed in the CO ₂ incubator. (Wo) Three days after the operation in (l) above, half of the medium is replaced with a new one. Exchange medium is DM without FCS
EM / F-12 mixed medium is used. (W) Thereafter, the same medium exchange as above is performed every 4 to 5 days.

By these series of operations, the nerve cells of the visual cortex of rat cerebrum could be cultured on the integrated composite electrode. The cells grew well both on the insulating layer (NPI) and on the electrode on which platinum black was deposited. Therefore, if an electrode at an appropriate position is used as a stimulation electrode or recording electrode,
Simultaneous multipoint measurement of nerve cell electrical activity was possible.

In addition, before and after a constant current stimulation of 100 μA was applied at a frequency of 1 Hz for 1 week through the electrode at an appropriate position of the integrated composite electrode of one embodiment of the present invention, the nerve at the electrode at an appropriate position was applied. An example of recording the electrical response (potential change) of cells is shown in FIGS. 4 and 5. FIG. 4 shows recording of electrical response of nerve cells before stimulation, and FIG. 5 shows recording of electrical response of nerve cells after stimulation.

Furthermore, in FIGS. 6 and 7, using the integrated composite electrode in which the electrode surface is not coated with gold, the electrical response of nerve cells before and after long-term stimulation under the same conditions as described above was measured. An example of recording is shown. FIG. 6 shows recording of electrical response of nerve cells before stimulation, and FIG. 7 shows recording of electrical response of nerve cells after stimulation.

In FIG. 4 to FIG. 7, arrows indicate artifacts generated by applying stimulation current, and arrow heads indicate potential changes generated by electrical activity of nerve cells. As can be seen from FIG. 6, when the integrated composite electrode in which the electrode surface is not coated with gold is used, the occurrence of artifacts is large, whereas the integrated composite electrode of the embodiment of the present invention in FIG. 4 is used. If so, the occurrence of artifacts is suppressed.

Further, as can be seen from FIG. 7, when the integrated composite electrode in which the electrode surface is not coated with gold is used, the occurrence of artifacts is larger than before the stimulation, and the electrical activity of nerve cells is hidden by the artifacts and measured. It became impossible.
On the other hand, in the case of using the integrated composite electrode of the embodiment of the present invention shown in FIG. 5, as in the case shown in FIG. 4, the occurrence of artifacts is suppressed, and the electrical activity of nerve cells is sufficiently suppressed. I was able to record.

The method of culturing nerve cells is not limited to this embodiment, since there are many modified methods other than this embodiment.

[0048]

INDUSTRIAL APPLICABILITY As described above, the integrated composite electrode of the present invention is capable of culturing nerve cells, and is capable of simultaneously measuring multi-points of nerve cell electrical activity which has been impossible or extremely difficult in the past. A long-term observation of signal transmission for several hours or more can be realized, and an integrated composite electrode having excellent responsiveness can be provided.

Further, since the distance between the closest electrodes is in the range of 10 μm or more and 1000 μm or less, it is possible to increase the possibility that each cell body is located on each electrode and binds via a neurite. It can be an integrated composite electrode that is convenient for measuring cells.

The insulating layer that covers the lead wire is an insulating layer that has holes on each electrode and is provided on almost the entire surface of the insulating substrate except in the vicinity of the contact point of the lead wire with the external circuit. By using an insulating material made of a photosensitive resin, the resin can be applied to almost the entire surface, and the necessary insulating layer pattern can be easily formed by photoetching.
The integrated composite electrode is easy to produce and has a low probability of insulation failure.

Further, since each electrode portion is coated with a substance having low surface resistance and low cytotoxicity, a stimulating current is applied using an appropriate electrode and a potential change is recorded using another appropriate electrode. In doing so, the polarization of the electrode is small even after applying a stimulus for a long period of time, and an integrated composite electrode capable of stable recording can be obtained.

Further, by arranging the center portions of the plurality of electrodes at respective intersections on the 8 × 8 grid, the maximum number of electrodes can be arranged so that the lead wires can be arranged substantially radially from the electrodes of the present invention. You can

[Brief description of drawings]

FIG. 1 is a plan view showing a pattern of an integrated composite electrode of the present invention in which an electrode and a lead wire are formed on an insulating substrate according to an embodiment of the present invention without an insulating layer.

FIG. 2 is a partially cutaway view of a plan view of only an insulating layer of an embodiment of an integrated composite electrode of the present invention.

FIG. 3 is a partial cross-sectional view of one embodiment of the integrated composite electrode of the present invention.

FIG. 4 shows an embodiment of an integrated composite electrode of the present invention,
FIG. 7 is a potential change waveform diagram recorded by using another appropriate electrode before applying a stimulating current for a long time using the appropriate electrode.

FIG. 5 shows an embodiment of an integrated composite electrode of the present invention,
FIG. 7 is a potential change waveform diagram recorded by using another appropriate electrode after applying a stimulating current for a long time using the appropriate electrode.

FIG. 6 is an example of an integrated composite electrode of the present invention, which is different from the integrated composite electrode only in that the surface of the electrode is not coated with gold, and a stimulating current is applied for a long time using a suitable electrode. FIG. 9 is a potential change waveform diagram recorded before using another appropriate electrode.

FIG. 7 is an example of an integrated composite electrode of the present invention, which is different from the integrated composite electrode of the present invention only in that the electrode surface is not coated with gold. FIG. 9 is a potential change waveform diagram recorded before using another appropriate electrode.

[Explanation of symbols]

 1 electrode 2 lead wire 3 insulating substrate 4 insulating layer 5 hole 6 gold 7 gold 8 nickel

─────────────────────────────────────────────────── ─── Continuation of the front page (51) Int.Cl. ⁵ Identification code Office reference number FI technical display location // H01B 5/14 Z 7638-4C A61B 5/04 300 J

Claims (4)

[Claims] 1. The distance between the electrodes closest to each other is matched on the insulating substrate.
Equivalent plural electrodes are provided, and lead wires are
Covers the wiring part arranged radially and the lead wire
An insulating layer is provided and the electrode area is 3 × 10²μm²that's all
4 x 10^Fourμm ²The following range, the surface resistance of the electrode part
Is 10 Ω / cm²An integrated composite electrode that is: 2. The distance between the closest electrodes is 10 μm or more 1
The integrated composite electrode according to claim 1, wherein the integrated composite electrode has a range of 000 μm or less. 3. The insulating layer for covering the lead wire is an insulating layer having holes on each electrode and provided on substantially the entire surface of the insulating substrate except in the vicinity of the contact point of the lead wire with an external circuit. The integrated composite electrode according to claim 1 or 2. 4. The integrated composite electrode according to claim 1, wherein a plurality of electrode central portions are located at respective intersections on an 8 × 8 grid.