JP3979574B2 - Array electrode for biological sample and production method thereof - Google Patents

Description

[0001]
BACKGROUND OF THE INVENTION
The present invention relates to an array electrode (multi-point electrode) for a biological sample and a manufacturing method thereof.
[0002]
[Prior art]
Among the measurements on biological samples, electrical measurement and electrochemical measurement using electrodes have been widely used as highly sensitive measurement methods with excellent time resolution. For example, a patch clamp method in which measurement is performed by inserting an electrode into a glass capillary that is thinly drawn into a needle shape is widely used as a method for monitoring the electrical activity of a single cell or a cell in a living tissue.
[0003]
In addition, an on-line sensor that extracts a solution in the vicinity of a cell from an acicular capillary and analyzes it electrochemically is expected to be increasingly expected as a highly sensitive analytical method with excellent spatial resolution (reference: Niwa). O, Torimitsu K, Morita M, Osborne P, Yamamoto K. Concentration of extracellular L-glutamate released from cultured nerve cells measured with a small-volume online sensor. Anal. Chem., 1996; 68: 1865-1870).
[0004]
On the other hand, the extracellular action potential is a technique that can measure the cell activity using an electrode placed outside the cell without damaging the sample (non-invasion). We have succeeded in measuring the electrical activity of multiple neurons simultaneously at multiple points using multiple electrodes arranged in a flat plate (Reference: Jimbo Y, Tateno T, Robinson). HPC. Simultaneous induction of pathway-specific potentiation and depression in networks in cortical neurons. Biophys. J., 1999; 76: 670-678). In this method, the extracellular action potential of cultured neurons can be measured simultaneously at multiple points using an array-type electrode fabricated by photolithography, so that the activity of connected cells and tissues can be measured with high temporal resolution. It is very effective.
[0005]
In addition, the array type plate electrode can be provided with selectivity by, for example, coating an enzyme thereon, and can also measure a specific chemical substance electrochemically. Very suitable for distribution measurement (reference: Kasai N, Jimbo Y, Niwa O, Matsue T, and Torimitsu K, Real-time multisite observation of glutamate release in rat hippocampal slices, Neuroscience Letters, 2001, 304, 112− 116).
[0006]
Furthermore, these electrodes are effective not only for measuring a biological sample, but also for applying a local stimulus to the sample, regardless of the needle type or the flat type. In this case, it is possible to apply electrical stimulation directly with the electrode, and an electric field is formed in the capillary using the electrode inserted into the capillary, and the stimulation solution previously placed in the capillary is placed in the biological sample. Alternatively, local stimulation by drug administration can be applied by injecting (injecting) the vicinity.
[0007]
[Problems to be solved by the invention]
The brain (aggregation of a large number of nerve cells) has a three-dimensional structure, and it is difficult to perform measurement for a large number of cells in the state of the three-dimensional structure. Therefore, conventionally, when measuring brain neurons and the like, the brain is dissected, the neurons are dissociated, the cells are held in a petri dish by culture, and measurement is performed using a planar electrode. Specifically, measurement is performed using an electrode having a large number of electrodes (for example, an 8 × 8 array of planar electrodes) on the bottom of the petri dish, which is useful when measuring separated cells or the like. Met.
[0008]
However, in recent years, with the worldwide interest in elucidating biological functions including the brain, a biological sample having a three-dimensional structure such as the brain or part of the brain has been decomposed into individual cells. Therefore, there is an increasing need for measurement while maintaining the shape.
[0009]
For example, an attempt is made to measure a biological sample having a three-dimensional structure by using a slice obtained by slicing a brain (usually 200 to 500 μm in thickness) and preparing a sample that retains a certain three-dimensional structure. However, since the above-described planar electrode can only measure cells on the surface of the section, a sufficient measurement signal cannot be obtained. The main reason for this is thought to be that the cells on the surface of the slice are greatly damaged when the slice sample is prepared, and there is a need for a measuring means that targets inner cells that are not damaged more than the cells on the surface. It has been.
[0010]
Furthermore, in order to elucidate the function of a living body having a three-dimensional spread, it is essential to obtain three-dimensional living body information, and therefore measurement in three dimensions is important. These are expected to become increasingly necessary with the advancement of the nanotechnology field that enables finer measurement in the future.
[0011]
In addition to elucidating biological functions, attempts have been made to apply electrical stimuli to arbitrary microscopic areas as a treatment for various diseases or as a substitute for sensory organs, and the position in three dimensions can be grasped with higher accuracy. However, there is a need for a method of applying a stimulus while doing so.
[0012]
However, with the conventional electrode shapes, it is very difficult to perform three-dimensional measurement and stimulus application that can obtain information inside the tissue, and even if possible, it is necessary to establish a complicated system. It was.
[0013]
Therefore, three-dimensional measurement / stimulation application is in an indispensable stage using a three-dimensionally arranged array electrode as a plurality of arbitrary electrodes.
[0014]
In view of the above problems, the present invention provides an array electrode for a biological sample that enables three-dimensional measurement and stimulus application, and a method for producing the same.
[0016]
[Means for Solving the Problems]
  An array electrode for a biological sample according to the present invention that solves the above problems isA plurality of electrodes, and an electrode fixing jig for three-dimensionally arranging the tips of the electrodes,
  The electrodeIs ledIt consists of an electric wire and an insulating material covering the conductive wire.The
  The electrode fixing jig includes an electrode alignment jig provided with a plurality of electrode alignment / fixing holes for three-dimensionally arranging the tip of the electrode, and a body portion for fixing the other end of the electrode. And withIt is characterized by that.
[0017]
The array electrode for a biological sample according to the present invention that solves the above-described problems is characterized in that the conductive wire is a metal wire and the insulating material is a glass tube.
[0018]
  The array electrode for a biological sample according to the present invention for solving the above-described problems is obtained by, GuidanceEnclose the wire and the conductive wireRukiCapilaryWhat consists ofIt is characterized by that.
[0019]
  The array electrode for a biological sample according to the present invention for solving the above-mentioned problems is characterized in that the conductive wire is a silver / silver chloride wire,CapillaryIs a glass tube.
[0029]
  A method for producing an array electrode for a biological sample according to the present invention that solves the above problems,A method for producing an array electrode for a biological sample in which a plurality of electrodes are arranged in three dimensions,
  The electrode, GuidanceAn electric wire and an insulating material covering the conductive wire;And
  Using an electrode fixing jig provided with an electrode alignment jig provided with a plurality of electrode alignment / fixing holes for fixing the electrode tips, the tips of the electrodes are arranged in three dimensions.It is characterized by doing.
[0030]
The manufacturing method of the array electrode for biological samples which solves the said subject based on this invention uses the said electrically conductive wire as a metal wire, and uses the said insulating material as a glass tube, It is characterized by the above-mentioned.
[0031]
  A method for producing an array electrode for a biological sample according to the present invention that solves the above-described problems,, GuidanceEnclose the wire and the conductive wireRukiCapilaryWhat consists ofIt is characterized by.
[0032]
  The method for producing an array electrode for a biological sample according to the present invention for solving the above-mentioned problems is characterized in that the conductive wire is a silver / silver chloride wire,CapillaryIs a glass tube.
[0041]
DETAILED DESCRIPTION OF THE INVENTION
In the present invention, by arranging a plurality of electrodes or capillaries three-dimensionally (three-dimensionally), a sample having a three-dimensional structure such as a biological sample can be measured in a three-dimensional region and applied with a stimulus to the three-dimensional region. It becomes possible. Moreover, it is also possible to arrange an electrode at an arbitrary position by changing the three-dimensional shape of the electrode or the like.
[0042]
That is, the present invention is characterized in that a three-dimensional array electrode in which electrodes and the like are arranged three-dimensionally is produced and applied to a biological sample so that measurement / stimulation can be applied.
[0043]
  The simplest method for producing the three-dimensional array electrode as described above is considered as follows.
  (I)Needle-shaped electrodes are arranged in three dimensions.
  (II)The capillaries are arranged in three dimensions.
[0044]
  Further, as another manufacturing method, the following can be considered.
  (III)Conductive materials and semiconductor materials containing metals, metal oxides, carbon, and organic substances are processed into three-dimensional electrodes, and unnecessary portions are covered with an insulating material such as a resist.
  In this method, not only a method of laminating a conductive material / semiconductor material by elution, etc., but also using a currently available fine processing technique, it is conceivable to produce a plurality of minute and various convex portions as electrodes. . Specific microfabrication techniques include an etching method using photolithography, a lift-off method, a plating method, and the like.
[0045]
  Furthermore, as another manufacturing method, the following can be considered.
  (IV)A convex portion is formed in advance using an insulating material, and the convex portion is covered with a conductive material / semiconductor material containing a metal, a metal oxide, or an organic substance to form an electrode.
[0046]
For example, the protrusions can be sharply processed only by applying a potential to each protrusion (electrolytic etching or the like). By sharpening the tip of the convex portion, the tip of the electrode can reach the inside of the biological sample, and can be embedded in the living body if the entire size is reduced. It is also possible to cover the fabricated array electrode with an insulating film and form an electric field on the required electrode, thereby removing the insulating film only at the tip of the electrode and disposing the electrode at an arbitrary position.
[0047]
The three-dimensional array electrode produced by the above method enables electrical measurement and electrochemical measurement at different positions by independently connecting each electrode, and further, electrical stimulation is locally applied through each electrode. By applying to, electrical stimulation can be applied.
[0048]
Hereinafter, the present invention will be described more specifically with reference to examples and drawings. In addition, this invention is not limited only to a following example.
[0049]
Example 1
FIG. 1 is a schematic view of an array electrode for a biological sample showing an example of an embodiment according to the present invention. Part of it has been enlarged to show the detailed structure.
[0050]
  The array electrode for a biological sample of this embodiment is a three-dimensional (three-dimensional) arrangement using a plurality of needle-like electrodes used in electrical measurement of a biological sample. This was mentioned earlier(I)This is based on the method for producing an array electrode for a biological sample.
[0051]
As shown in FIG. 1, the biological sample array electrode 1 has a plurality of needle-like electrodes 2, and the tips of the electrodes 2 are arranged in a three-dimensional manner by an electrode fixing jig 3. It is fixed to. A lead terminal 4 for connection to a measuring instrument or the like is connected to the other end of the electrode 2 and is fixed to the main body portion of the electrode fixing jig 3.
[0052]
The electrode fixing jig 3 includes an electrode alignment jig 5 for arranging the tip of the electrode 2 three-dimensionally. An electrode alignment / fixing hole 5a (hole) provided in the electrode alignment jig 5 is provided. The electrode 2 is fixed using a diameter of 0.17 millimeters and the distance between the hole centers is 0.3 millimeters, and electrical measurement of an arbitrary position of the biological sample 7 on the substrate 6 can be performed (enlargement of FIG. 1). (See Fig. A. Enlarged view A shows a top view and a side view of the electrode alignment jig 5). The electrode 2 has a thin tungsten wire 8 serving as a conductive wire and a glass tube 9 serving as an insulating material, and the glass tube 9 covers the tungsten wire 8 except for its tip portion, and has a needle-like shape. It is an electrode (see enlarged view B in FIG. 1).
[0053]
If the electrode alignment jig 5 is used, it is possible to simultaneously measure electrical signals at a plurality of positions separated by 0.3 mm. Further, if the distance between the centers of the electrode alignment / fixing holes 5a is further reduced, measurement with higher spatial resolution is possible. Further, by using the needle-like electrodes 2 having different lengths, the electrodes 2 can be arranged at arbitrary height positions, and a three-dimensional biological sample can be electrically measured.
[0054]
In this embodiment, tungsten is used as the material of the conductive wire of the electrode. However, other metals, metal oxides, semiconductor materials, carbon, etc. can be used. Furthermore, although the electrode is insulated and sealed using a glass tube as an insulating material, other organic materials such as resist and semiconductor insulating materials such as silicon nitride can be used.
[0055]
(Example 2)
FIG. 2 is a schematic view of an array electrode for a biological sample showing another example of the embodiment according to the present invention. Part of it has been enlarged to show the detailed structure.
[0056]
  The array electrode for a biological sample of this embodiment is a three-dimensionally (three-dimensionally) arrangement using a plurality of capillaries for performing electrical measurement in or near a minute region in the biological sample. This was mentioned earlier(II)This is based on the method for producing an array electrode for a biological sample.
[0057]
As shown in FIG. 2, the biological sample array electrode 11 has a plurality of capillaries / electrodes 12, and the tips of the capillaries / electrodes 12 are arranged in three dimensions by capillaries / electrode fixing jigs 13. It is fixed to become. A lead terminal 14 for connecting to a measuring instrument or the like is connected to the other end of the capillary / electrode 12 and is fixed to the main body portion of the capillary / electrode fixing jig 13.
[0058]
The capillary / electrode fixing jig 13 is the same as that shown in FIG. 1, and includes a capillary alignment jig 15 for arranging the tip of the capillary / electrode 12 in three dimensions. The capillary / electrode 12 is fixed using a capillary alignment / fixing hole 15a provided in the capillary alignment jig 15 (a hole diameter of 0.17 mm, and the distance between the hole centers is 0.3 mm), and the substrate 16 is fixed. Electrical measurement is performed at an arbitrary position of the biological sample 17 (see an enlarged view C in FIG. 2. The enlarged view C shows a top view and a side view of the capillary alignment jig 15). The capillary / electrode 12 has a thin silver / silver chloride wire 18 serving as a conductive wire and a glass tube 19 serving as an insulating material. The glass tube 19 has an opening at the tip thereof, The silver chloride wire 18 is completely encapsulated therein (see enlarged view D in FIG. 2). The silver / silver chloride wire 18 is composed of an appropriate composition ratio of silver and silver chloride. Further, since the glass tube 12 of the capillary / electrode 12 has a dropper-like shape, an arbitrary solution 20 can be held therein.
[0059]
The capillary alignment jig 15 is the same as the electrode alignment jig 5 of the biological sample array electrode 1 shown in FIG. 1, and when this is used, electrical connection at a plurality of positions separated by 0.3 mm is performed. Can be measured simultaneously. Further, if the distance between the centers of the capillary alignment / fixing holes 15a is further reduced, measurement with higher spatial resolution is possible. Furthermore, by using capillaries / electrodes 12 having different lengths, the capillaries / electrodes 12 can be arranged at arbitrary height positions, and three-dimensional electrical measurement of a biological sample can be performed.
[0060]
Further, by applying a potential pulse to the silver / silver chloride wire 18, it is possible to inject (inject) the solution 20 inside a specific capillary / electrode 12, thereby applying a local drug stimulus. Is possible. Further, the capillary / electrode 12 fixed at an arbitrary position enables electrochemical measurement by sampling the sample solution from any of a plurality of three-dimensional microregions in or near the biological sample.
[0061]
Here, silver / silver chloride is used as the material for the conductive wire of the electrode for electrical measurement, but other metals, metal oxides, semiconductor materials, carbon, and the like can also be used.
[0062]
  (referenceExample 3)
  FIG.Reference exampleIt is the schematic explaining the array electrode for biological samples, and its production method.
[0063]
  BookreferenceThe example array electrode for a biological sample is produced by three-dimensionally processing conductive wires on an insulating substrate and arranging them in three dimensions. This was mentioned earlier(III)This is based on the method for producing an array electrode for a biological sample. Hereinafter, this method will be described (refer to FIG. 3. Note that the following (1) to (6) correspond to steps (1) to (6) in FIG. 3).
[0064]
(1) A transparent acrylic substrate 22 to be an insulating substrate is prepared.
(2) The pad portion 23 is formed on the acrylic substrate 22 by photolithography.
(3) A plurality of fine tungsten wires 24 to be conductive wires are arranged and fixed to each other with an adhesive 25. The plurality of fine tungsten wires 24 are covered with insulation one by one.
(4) The coating of the tip end portion of the tungsten fine wire 24 is peeled off and bent vertically to form a three-dimensional electrode portion 26, and the other end side of the tungsten fine wire 24 is bent into an arbitrary shape. The lead part 27 is formed so as to overlap the position. At this time, it is possible to arrange the electrode portion 26 at a three-dimensional free position by changing the length of the tip portion of the tungsten thin wire 24 to be a three-dimensional electrode, the bending angle, or the like.
(5) The tungsten fine wire 24 is fixed so that the end portion of the lead wire 27 is aligned with the pad portion 23 on the acrylic substrate 22, and the lead portion 27 is fixed to the pad portion 23 using the conductive paste 28.
(6) As a final treatment, the portions other than the electrode portion 26 and the pad portion 23 are covered with an insulating coating material 29.
[0065]
The biological sample array electrode 21 has a plurality of electrodes arranged at a desired three-dimensional position by the above production procedure.
[0066]
  BookreferenceIn the example, a thin tungsten wire is used as the material of the conductive wire of the electrode, but other metals, metal oxides, semiconductor materials, carbon, etc. can also be used. In addition, a three-dimensional electrode is formed using a tungsten thin wire with insulation coating, and a final insulation coating is performed in the above production procedure (6), but a tungsten thin wire without insulation coating is used. Is also possible.
[0067]
  As a measuring method using the biological sample array electrode 21, the biological sample array 21 is arranged with the electrode part 26 facing upward in the center of the petri dish, and the pad part 23 is connected to a measuring instrument or the like. A slice sample such as brain is placed on top and measured. Since the electrode part 26 has a three-dimensional distribution, it is possible to measure the depth direction of the section sample, and it is possible to reliably measure the undamaged nerve cells and to increase the nerve cell activity as compared with the conventional method. A signal can be obtained. This is the otherreferenceThe same applies to the examples.
[0068]
  (referenceExample 4)
  FIG.Another reference exampleIt is the schematic explaining the array electrode for biological samples, and its production method.
[0069]
  BookreferenceThe example array electrode for a biological sample is a three-dimensional body formed by molding a non-conductive material on an insulating substrate, and the three-dimensional body is covered with a conductive material to form a three-dimensionally arranged electrode. Is. This was mentioned earlier(IV)This is based on the method for producing an array electrode for a biological sample. Hereinafter, this method will be described (see FIG. 4. Note that the following (1) to (7) correspond to the steps (1) to (7) in FIG.
[0070]
(1) A transparent acrylic substrate 32 to be an insulating substrate is prepared.
(3) A non-conductive resin 33 (for example, polydimethylsiloxane) that can be molded is thickly coated on the entire surface of the acrylic substrate 32, and an aluminum-made metal mask 34 for three-dimensional processing is pressed thereon. The three-dimensional body 35 having an arbitrary shape is formed.
(4) The acrylic substrate 32 is masked using a metal mask 36 made of aluminum.
(6) Platinum is deposited by sputtering only on necessary portions of the electrode portion 37, the lead portion 38, and the pad portion 39 on the acrylic substrate 32 by masking. At this time, the electrode part 37 is formed by covering the three-dimensional body 35 with platinum as a conductive material.
(7) As a final treatment, unnecessary portions such as the lead portion 38 other than the electrode portion 37 and the pad portion 39 are coated with a resist 40 and insulated so as not to hinder measurement or stimulus application.
[0071]
Further, as another manufacturing procedure, the following procedure is also possible.
(1) A transparent acrylic substrate 32 to be an insulating substrate is prepared.
(2) The lead portion 38 and the pad portion 39 are prepared in advance on the acrylic substrate 32 by photolithography.
(3) A non-conductive resin 33 (for example, polydimethylsiloxane) that can be molded is thickly coated on the entire surface of the acrylic substrate 32, and an aluminum-made metal mask 34 for three-dimensional processing is pressed thereon. The three-dimensional body 35 having an arbitrary shape is formed.
(5) The acrylic substrate 32 is masked using an aluminum metal mask 36b.
(6) By masking, platinum is deposited only on the electrode portion 37 on the acrylic substrate 32 by sputtering. At this time, the electrode part 37 is formed by covering the three-dimensional body 35 with platinum as a conductive material.
(7) As a final treatment, unnecessary portions such as the lead portion 38 other than the electrode portion 37 and the pad portion 39 are coated with a resist 40 and insulated so as not to hinder measurement or stimulus application.
[0072]
  The biological sample array electrode 31 has a plurality of electrodes arranged at a desired three-dimensional position by the above-described production procedure. BookreferenceIn the example, a three-dimensional electrode having an arbitrary shape can be formed by changing the shape of the metal mask 34, and the processing can be simplified by using a resin as a constituent of the three-dimensional body.
[0073]
  (referenceExample 5)
  FIG.Another reference exampleIt is the schematic explaining the array electrode for biological samples, and its production method.
[0074]
  BookreferenceThe example array electrode for a biological sample is a three-dimensional body formed by molding a non-conductive material on an insulating substrate, and the three-dimensional body is covered with a conductive material to form a three-dimensionally arranged electrode. Is. This was mentioned earlier(IV)This is based on the method for producing an array electrode for a biological sample. Hereinafter, this method will be described (refer to FIG. 5. Note that the following (1) to (7) correspond to steps (1) to (7) in FIG. 5).
[0075]
(1) A transparent acrylic substrate 42 to be an insulating substrate is prepared.
(3) The entire surface of the acrylic substrate 42 is coated with a non-conductive photoreactive resin 43 (for example, a photoresist) thickly, and a portion to be an electrode is patterned by photolithography (shown in FIG. 5 (3)). White part).
(4) Unnecessary portions (unpatterned portions) of the photoreactive resin 43 are removed by etching, and the three-dimensional body 44 constituting the electrode portion is formed.
(6) Masking is performed using a metal mask similar to the metal mask 36a shown in FIG. 4 and patterning is performed by sputtering platinum to form the electrode portion 45, the lead portion 46, and the pad portion 47 on the acrylic substrate 42. Conductivity is given only to necessary parts. At this time, the electrode portion 45 is formed by covering the three-dimensional body 44 with platinum as a conductive material.
(7) As a final treatment, unnecessary portions such as the lead portion 46 other than the electrode portion 45 and the pad portion 47 are coated with a resist 48 and insulated so as not to hinder measurement or stimulus application.
[0076]
Further, as another manufacturing procedure, the following procedure is also possible.
(1) A transparent acrylic substrate 42 to be an insulating substrate is prepared.
(2) Leads 46 and pads 47 are prepared in advance on the acrylic substrate 42 by photolithography.
(3) The entire surface of the acrylic substrate 42 is coated with a non-conductive photoreactive resin 43 (for example, a photoresist or the like), and a portion to be an electrode is patterned by photolithography.
(5) The unnecessary portion of the photoreactive resin 43 is removed by etching, and the three-dimensional body 44 constituting the electrode portion is formed.
(6) Masking is performed using a metal mask similar to the metal mask 36b shown in FIG. 4, patterning is performed by sputtering platinum, and conductivity is imparted to a necessary portion to be the electrode portion 45 on the acrylic substrate 42. . At this time, the electrode portion 45 is formed by covering the three-dimensional body 44 with platinum as a conductive material.
(7) As a final treatment, unnecessary portions such as the lead portion 46 other than the electrode portion 45 and the pad portion 47 are coated with a resist 48 and insulated so as not to hinder measurement or stimulus application.
[0077]
The biological sample array electrode 41 has a plurality of electrodes arranged at a desired three-dimensional position by the above production procedure.
[0078]
In this manufacturing method, it is difficult to arbitrarily form the shape of the three-dimensional electrode, but it is possible to use a commonly used photoresist or the like, so that a convex electrode can be easily processed. Excellent in that you can.
[0079]
  (referenceExample 6)
  FIG.Another reference exampleIt is the schematic explaining the array electrode for biological samples, and its production method.
[0080]
  BookreferenceThe array electrode for a biological sample in the example is an electrode arranged three-dimensionally by molding a conductive material on an insulating substrate. This was mentioned earlier(III)This is based on the method for producing an array electrode for a biological sample. Hereinafter, this method will be described (refer to FIG. 6. Note that the following (1) to (8) correspond to steps (1) to (8) in FIG. 6).
[0081]
(1) A transparent acrylic substrate 52 to be an insulating substrate is prepared.
(3) Platinum 53 as a conductive material is thickly coated on the entire surface of the acrylic substrate 52 by sputtering, and a portion to be an electrode is patterned by photolithography (the white portion in FIG. 6 (3)).
(4) Unnecessary portions (unpatterned portions) of platinum 53 are removed by etching to form electrode portions 54 that are conductive three-dimensional bodies.
(6) Masking is performed using a metal mask similar to the metal mask 36a shown in FIG. 4, patterning is performed by sputtering platinum, and the conductive portions are formed on the necessary portions to be the lead portions 55 and the pad portions 56 on the substrate. give.
(7) Unnecessary portions such as the lead portion 55 other than the electrode portion 54 and the pad portion 56 are coated with a resist 57 and are covered with insulation so as not to hinder measurement and stimulus application.
(8) As a final treatment, the electrode portion 54 is formed into an arbitrary shape, for example, a needle shape, using etching, electrolytic etching, plating, or the like. However, this step is not always necessary.
[0082]
Further, as another manufacturing procedure, the following procedure is also possible.
(1) A transparent acrylic substrate 52 to be an insulating substrate is prepared.
(2) The lead 55 and the pad 56 are formed on the acrylic substrate 52 in advance by photolithography.
(3) The entire surface of the acrylic substrate 52 is coated with platinum 53, which is a conductive material, by sputtering, and a portion to be an electrode is patterned by photolithography.
(5) The unnecessary portion of the platinum 53 is removed by etching, and the electrode portion 54 that becomes a conductive three-dimensional body is formed.
(7) Unnecessary portions such as the lead portion 55 other than the electrode portion 54 and the pad portion 56 are coated with a resist 57 and are covered with insulation so as not to hinder measurement and stimulus application.
(8) As a final treatment, the electrode portion 54 is formed into an arbitrary shape, for example, a needle shape, using etching, electrolytic etching, plating, or the like. However, this step is not always necessary.
[0083]
The biological sample array electrode 51 has a plurality of electrodes arranged at a desired three-dimensional position by the above production procedure. Furthermore, in this manufacturing method, the shape of the three-dimensional electrode can be arbitrarily formed by electrolytic etching or the like.
[0084]
  (referenceExample 7)
  FIG.Another reference exampleIt is the schematic explaining the array electrode for biological samples, and its production method.
[0085]
  BookreferenceThe array electrode for a biological sample in the example is an electrode arranged three-dimensionally by molding a conductive material on an insulating substrate. This was mentioned earlier(III)This method is based on the lift-off method often used when a conductive material (metal) having low adhesion to the substrate is used for patterning. Hereinafter, this method will be described (see FIG. 7. Note that the following (1) to (7) correspond to steps (1) to (7) in FIG. 7).
[0086]
(1) A transparent acrylic substrate 62 to be an insulating substrate is prepared.
(2) The lead part 63 and the pad part 64 are prepared in advance on the acrylic substrate 62 by photolithography.
(3) Photoresist 65 serving as a photoreactive resin is coated on the entire surface of the acrylic substrate 62, and a portion serving as an electrode is patterned by photolithography (the white portion in FIG. 7 (3)). At this time, patterning is performed so that the photoresist 65 does not exist in the portion to be the electrode.
(4) The entire surface of the acrylic substrate 62 is coated with gold 66, which is a conductive material, by sputtering.
(5) By immersing in a lift-off solution, the patterned photoresist 65 is removed, and at the same time, the gold 66 coated thereon is also removed, and a three-dimensional electrode portion 67 is formed.
(6) Unnecessary parts such as the lead part 63 other than the electrode part 67 and the pad part 64 are coated with a resist 68 and insulated so as not to hinder measurement or stimulus application.
(7) As a final treatment, the electrode portion 67 is formed into an arbitrary shape, for example, a needle shape, using etching, electrolytic etching, plating, or the like. However, this step is not always necessary.
[0087]
The biological sample array electrode 61 has a plurality of electrodes arranged at a desired three-dimensional position by the above production procedure. Furthermore, this manufacturing method can also arbitrarily form a three-dimensional electrode shape by electrolytic etching or the like.
[0088]
  For referenceAs the substrate material in the examples, glass or the like can be used in addition to the acrylic resin, and if a transparent substrate such as glass or acrylic resin is used, it can be used while observing the biological sample with a microscope. In addition to tungsten, platinum, and gold, other materials such as metals, metal oxides, carbon, and conductive materials such as carbon, organic materials, and semiconductor materials can be used as electrode materials. It is also possible to combine these. Many other types of non-conductive resins can be used, and these can be combined if necessary.
[0089]
The array electrode having a three-dimensional electrode arrangement structure according to the present invention is extremely effective for measurement of biological signals and neurotransmitters for a living tissue having a three-dimensional structure. Furthermore, an enzyme (protein) is formed on the electrode surface. It is also possible to give selectivity by modifying a substance having specificity such as) and apply it to electrochemical measurement as a more sophisticated electrode. In this case, grasping the distribution of a specific substance and the change of the distribution to the stimulus leads to elucidation of the action and mechanism of the substance in the living body.
[0090]
  In addition to three-dimensional measurement, it is possible to give a stimulus to any three-dimensional position. In the case of electrical stimulation, sensory organ disorders such as vision and hearing, motor function disorders, It can be widely applied to rehabilitation.For example,Fabrication of minute three-dimensional array electrodes using microfabrication technologyifIn addition, it is possible to appropriately acquire information on necessary parts in a minute biological sample at intervals of several hundred μm, and to apply stimulation.
[0092]
【The invention's effect】
  Claim1To claims2According to the invention according toA plurality of electrodes, and an electrode fixing jig for three-dimensionally arranging the tips of the electrodes,The electrodeIs ledIt consists of an electric wire and an insulating material covering the conductive wire.The electrode fixing jig fixes an electrode alignment jig provided with a plurality of electrode alignment / fixing holes for arranging the tip of the electrode in three dimensions, and the other end of the electrode. With body partTherefore, electrical signals in a three-dimensional biological sample can be measured simultaneously in three dimensions with high sensitivity, high temporal resolution, and high spatial resolution, and stimulation can be applied locally in three dimensions. be able to.Furthermore, it is possible to provide selectivity by modifying a substance having specificity such as an enzyme on the electrode, so that it can be applied to electrochemical measurement as a highly functional electrode.
[0093]
  Claim3To claims4According to the invention concerning, GuidanceEnclose the wire and the conductive wireRukiCapilaryWhat consists ofAs a result, electrical signals in a three-dimensional biological sample can be measured in three dimensions at the same time with high sensitivity, high temporal resolution, and high spatial resolution, and stimulation can be applied. Since a drug or the like can be held inside, local drug stimulation can be applied, and electrochemical measurement can be performed by sampling a solution of a biological sample.
[0098]
  Claim5To claims6According to the invention according to
  A method for producing an array electrode for a biological sample in which a plurality of electrodes are arranged in three dimensions,The electrode, GuidanceAn electric wire and an insulating material covering the conductive wire;The electrode tips are arranged in a three-dimensional manner using an electrode fixing jig provided with an electrode alignment jig provided with a plurality of electrode alignment / fixing holes for fixing the electrode tips.Therefore, it is possible to produce an array electrode that can simultaneously measure electrical signals in a three-dimensional biological sample in a three-dimensional manner with high sensitivity, high temporal resolution, and high spatial resolution, and can apply a stimulus. .
[0099]
  Claim7To claims8According to the invention concerning, GuidanceEnclose the wire and the conductive wireRukiCapilaryWhat consists ofTherefore, electrical signals in a three-dimensional biological sample can be simultaneously measured in three dimensions with high sensitivity, high temporal resolution, and high spatial resolution, and can be stimulated. It is possible to make an array electrode capable of applying a typical drug stimulus and electrochemical measurement of a biological sample.
[Brief description of the drawings]
FIG. 1 is a schematic view of an array electrode for a biological sample showing an example of an embodiment according to the present invention.
FIG. 2 is a schematic view of an array electrode for a biological sample showing another example of the embodiment according to the present invention.
[Fig. 3]Reference exampleIt is the schematic explaining the array electrode for biological samples, and its production method.
[Fig. 4]Another reference exampleIt is the schematic explaining the array electrode for biological samples, and its production method.
[Figure 5]Another reference exampleIt is the schematic explaining the array electrode for biological samples, and its production method.
[Fig. 6]Another reference exampleIt is the schematic explaining the array electrode for biological samples, and its production method.
[Fig. 7]Another reference exampleIt is the schematic explaining the array electrode for biological samples, and its production method.
[Explanation of symbols]
  1 Array electrode for biological samples
  2 electrodes
  3 Electrode fixing jig
  4 Lead wire
  5 Electrode alignment jig
  5a Electrode alignment / fixing hole
  6 Substrate
  7 Biological samples
  8 Tungsten wire
  9 Glass tube

Claims (8)

A plurality of electrodes, and an electrode fixing jig for three-dimensionally arranging the tips of the electrodes,
The electrodes, Ri Do and an insulating material covering the conductive wire and the conductive wire,
The electrode fixing jig includes an electrode alignment jig provided with a plurality of electrode alignment / fixing holes for three-dimensionally arranging the tip of the electrode, and a body portion for fixing the other end of the electrode. biological sample for an array electrode, characterized in that it comprises and. The biological sample array electrode according to claim 1 ,
An array electrode for a biological sample, wherein the conductive wire is a metal wire and the insulating material is a glass tube. The biological sample array electrode according to claim 1,
The electrodes, the conductive lines and biological samples for an array electrode, characterized in that consisted of to Ruki Yapirari and enclosing the conductive line. The array electrode for biological samples according to claim 3 ,
An array electrode for a biological sample, wherein the conductive wire is a silver / silver chloride wire and the capillary is a glass tube. A method for producing an array electrode for a biological sample in which a plurality of electrodes are arranged in three dimensions ,
The electrode, consisted of an insulating material covering the conductive wire and the conductive wire,
Using an electrode fixing jig provided with an electrode alignment jig provided with a plurality of electrode alignment / fixing holes for fixing the electrode tips, the tips of the electrodes are arranged three-dimensionally. A method for producing an array electrode for a biological sample, characterized in that In the manufacturing method of the array electrode for biological samples of Claim 5 ,
A method for producing an array electrode for biological samples, wherein the conductive wire is a metal wire and the insulating material is a glass tube. In the manufacturing method of the array electrode for biological samples of Claim 5 ,
The electrode, a manufacturing method of a biological sample for the array electrodes, wherein the conductive lines and made of said conductive to enclosing the wire Ruki Yapirari. In the manufacturing method of the array electrode for biological samples of Claim 7 ,
A method for producing an array electrode for a biological sample, wherein the conductive wire is a silver / silver chloride wire and the capillary is a glass tube.

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