WO2023281674A1

WO2023281674A1 - Film-type surface-stress sensor and method for manufacturing film-type surface-stress sensor

Info

Publication number: WO2023281674A1
Application number: PCT/JP2021/025694
Authority: WO
Inventors: 賢司宮崎; 久萩原
Original assignee: 日本電気株式会社
Priority date: 2021-07-07
Filing date: 2021-07-07
Publication date: 2023-01-12
Also published as: JPWO2023281674A1

Abstract

Provided is a film-type surface-stress sensor having insulation between circuitry.　A film-type surface-stress sensor 1 according to this invention comprises a film 13 and a sensor substrate 10. The film 13 deforms in response to surface stress. The sensor substrate 10 comprises a support region 15 and a circuit 12A. The support region 15 supports the film 13 and comprises a piezoresistive element 14. The piezoresistive element 14 detects the deformation of the film 13. The circuit 12A is connected to the piezoresistive element 14. The circuit 12A includes metal that can be made to form an oxide film through oxidation treatment. The circuit 12A has an oxide film layer 122 on the surface thereof.

Description

MEMBRANE-TYPE SURFACE STRESS SENSOR AND METHOD FOR MANUFACTURING MEMBRANE-TYPE SURFACE STRESS SENSOR

The present invention relates to a membrane-type surface stress sensor and a method for manufacturing the membrane-type surface stress sensor.

Target detection is important in a wide variety of fields such as food and medicine, and various methods have been proposed. In recent years, a membrane-type surface stress sensor has attracted attention (see Patent Document 1). The film-type surface stress sensor can analyze the presence and amount of the target by, for example, binding a target to a film such as a silicon film, deforming the film, and measuring variations in electrical resistance caused by the deformation. .

WO2011/148774

Most of the biological samples of blood are liquid. A membrane-type surface stress sensor (hereinafter also referred to as "MSS") has a circuit (wiring) formed on a sensor substrate. may occur. Therefore, the present inventor tried to insulate the circuits by coating the circuits of the MSS with a resin. However, if the resin adheres to the piezoresistive element that detects the distortion of the film when the circuit is coated with the resin, the resulting MSS sensor cannot be used because there is a problem in detecting the distortion by the piezoresistive element. However, when the resin is used for insulation, there arises a problem that the rate of breakage during manufacturing is high.

Therefore, the first object of the present invention is to provide an MSS insulated between circuits.

In addition, the present inventors invented MSS in which a binding substance such as an aptamer capable of binding to a target is immobilized on the surface of the MSS membrane. However, the inventors have found a problem that the immobilization efficiency of the binding substance on the membrane of MSS is low.

Therefore, the second object of the present invention is to provide an MSS capable of improving the binding substance immobilization efficiency.

In order to achieve the first object, a first film-type surface stress sensor (hereinafter also referred to as "first sensor") of the present invention comprises a film and a sensor substrate,
The membrane is a membrane that deforms in response to surface stress,
the sensor substrate comprises a support area and circuitry;
the support region supports the membrane and comprises a piezoresistive element;
The piezoresistive element is an element that detects deformation of the film,
The circuit is connected to the piezoresistive element,
The circuit includes a metal capable of forming an oxide film by oxidation treatment,
The circuit has an oxide film layer on its surface.

A method for manufacturing a film-type surface stress sensor according to the present invention (hereinafter also referred to as a "first manufacturing method") is a film-type stress sensor comprising a film and a circuit, wherein the circuit includes a metal capable of forming an oxide film by oxidation treatment. and forming an oxide film layer on the surface of the circuit by oxidizing the circuit.

In order to achieve the second object, a second film-type surface stress sensor (hereinafter also referred to as "second sensor") of the present invention comprises a film and a sensor substrate,
The membrane is a membrane that deforms in response to surface stress,
the sensor substrate comprises a support area;
the support region supports the membrane and comprises a piezoresistive element;
The piezoresistive element is an element that detects deformation of the film,
The film comprises an amorphous layer.

A method for manufacturing a film-type surface stress sensor of the present invention (hereinafter also referred to as a "second manufacturing method") is a film-type stress sensor comprising a film and a circuit, wherein the film is subjected to an amorphization treatment (amorphization process). quenching treatment) to form an amorphous layer on the surface of the film.

According to the method of manufacturing a semiconductor device of the present invention, in a semiconductor device comprising a film and a circuit, a circuit containing a metal capable of forming an oxide film by oxidation treatment is subjected to oxidation treatment to form an oxide film layer on the surface of the circuit. and
and forming an amorphous layer on the surface of the film by subjecting the film to amorphization treatment.

The target analysis method of the present invention includes an applying step of applying a voltage to a membrane surface stress sensor in a sample liquid;
an analysis step of analyzing the target in the sample liquid by measuring the stress change of the piezoresistive element in the film-type surface stress sensor;
The film-type surface stress sensor is the first film-type surface stress sensor of the present invention or the second film-type surface stress sensor of the present invention.

According to the first MSS of the present invention, it is possible to insulate between circuits. In addition, according to the second MSS of the present invention, it is possible to improve the immobilization efficiency of the binding substance on the MSS membrane.

FIG. 1 is a schematic diagram showing an example of the configuration of the MSS in Embodiment 1, (A) is a schematic plan view and a schematic partial enlarged view of the MSS, and (B) is II in (A). It is a schematic cross section seen from the direction. FIG. 2 is a schematic diagram showing an example of a method for producing MSS according to Embodiment 1. FIG. FIG. 3 is a schematic diagram showing another example of the method for producing MSS according to Embodiment 1. FIG. FIG. 4 is a schematic diagram showing an example of the configuration of the MSS in Embodiment 2, (A) is a schematic plan view of the MSS, and (B) is a schematic cross section viewed from the II-II direction in (A). It is a diagram. FIG. 5 is a schematic diagram showing an example of a method for producing MSS according to Embodiment 2. FIG. FIG. 6 is a schematic cross-sectional view showing another example of the silicon film of the MSS in Embodiment 2. FIG. FIG. 7 is a schematic diagram showing another example of the method for producing MSS according to Embodiment 2. FIG. FIG. 8 is a schematic diagram showing an example of the configuration of the MSS according to the third embodiment. 9 is a photograph showing a transmission electron microscope image of the silicon film of MSS of Embodiment 3 in Example 1. FIG.

As used herein, the term "membrane-type surface-stress sensor (MSS)" refers to a sensor in which a film that deforms in response to surface stress is supported by a support having a piezoresistive element. do. In the MSS, when the film receives stress, the film deforms (generates strain) due to the generation of strain or the like. Then, according to the amount of deformation of the film, stress is generated in the piezoresistive element of the support supporting the film, and the resistance value of the piezoresistive element changes in proportion to the stress. Therefore, according to the MSS sensor, the presence or absence of the target bound to the membrane can be qualitatively analyzed indirectly by applying a voltage to the MSS and measuring an electrical signal accompanying a change in resistance value. can. In addition, according to the MSS sensor, by applying a voltage to the MSS and measuring an electrical signal accompanying a change in resistance value, it is possible to quantitatively analyze the amount of the target that has contributed to the generation of stress in the film. . Further, in the present invention, for example, in such MSS, a binding substance that binds to a target is used, specifically, by immobilizing the binding substance on the membrane, the target is bound to the binding substance can be coupled to the MSS via Therefore, in the MSS of the present invention, when the binding substance is immobilized on the film, the amount of the target that has contributed to the generation of stress on the film due to the binding of the target to the binding substance can be qualitatively or quantitatively determined. can also be analyzed.

In the present invention, the "target" is not particularly limited and can be set arbitrarily. Said target may be, for example, a substance that can come into contact with said membrane or said binding substance in a liquid, ie in liquid phase. The target has, for example, one or more regions to which the binding substance binds, ie epitopes of the binding substance.

In the present invention, the targets include, for example, microorganisms including bacteria such as anthrax, Escherichia coli, salmonella, and tubercle bacillus; viruses such as SARS-CoV-2 and influenza virus; allergens; Examples of the allergen include grains such as wheat; eggs; meat; fish; shellfish; vegetables; The type of the target is not particularly limited, and examples thereof include high-molecular compounds such as proteins, sugar chains, nucleic acids, and polymers; low-molecular-weight compounds; and the like.

In the present invention, the "binding substance" may be any substance capable of binding to the target, that is, a binding substance. Examples of the binding substance include nucleic acid molecules and proteins, and specific examples thereof include antibodies, aptamers, and the like. Where the target is a receptor or its ligand, the binding agent may be the ligand or receptor, respectively. When using a receptor for a ligand as the binding substance, the receptor may be a fusion protein with the Fc region of an immunoglobulin, that is, a receptor-Fc fusion protein, preferably with the Fc region of an IgG protein. fusion protein, ie receptor-IgG Fc. The Fc fusion protein can be prepared, for example, by linking the C-terminal amino acid of the receptor directly or via a linker to the N-terminal amino acid of the CL region or CH1 region of an immunoglobulin.

In the present invention, "antibody" can also be referred to as a soluble immunoglobulin that has binding properties to a target. Types of the antibody include, for example, IgA, IgD, IgE, IgG, or IgM. IgA includes, for example, IgA1 or IgA2. IgG includes, for example, IgG1, IgG2, IgG3, or IgG4. The antibody may be an antigen-binding fragment thereof, that is, a partial peptide of the antibody that has binding properties to the target. Said antigen-binding fragment is for example a polypeptide comprising a part of said antibody, more particularly the binding or variable region of said antibody. Said antigen-binding fragments are, for example, Fab, Fab', F(ab')2, Fv fragments, rIgG (half IgG) fragments, single chain antibodies (scFv), dual variable domain antibodies (DVD-Ig™ ), diabodies, triabodies, tetrabodies, tandabs, flexibodies that are combinations of scFvs and diabodies, tandem scFvs (e.g., BiTE ( Micromet), DART® (MacroGenics), Fcab™ or ^mAb2 ™ (F-star), Fc engineering antibody (Xencor) or DuoBody® (Genmab company), etc. As the antibody, a known antibody or an antigen-binding fragment thereof having a binding property to a target may be used, or a new antibody or an antigen-binding fragment thereof obtained by immunizing an animal or the like with a target may be used. good. Moreover, the antibody may be a monoclonal antibody or a polyclonal antibody. The antibody may be a blood-derived fraction such as serum or plasma that contains antibodies capable of binding to the target.

In the present invention, an "aptamer" is a nucleic acid molecule that has binding properties to a target. The aptamer can also be referred to as, for example, a nucleic acid molecule that specifically binds to a target. The constituent units of the aptamer are, for example, nucleotide residues and non-nucleotide residues. Said nucleotide residues include, for example, deoxyribonucleotide residues and ribonucleotide residues, and said nucleotide residues may, for example, be modified or unmodified. Examples of the aptamer include DNA aptamers composed of deoxyribonucleotide residues, RNA aptamers composed of ribonucleotide residues, aptamers containing both, aptamers containing modified nucleotide residues, and the like. The length of the aptamer is not particularly limited, and is, for example, 10-200 bases. As the aptamer for the target, for example, an existing aptamer may be used, or an aptamer newly obtained using, for example, the SELEX method or the like may be used depending on the target.

In the present invention, "bind" or "capable of binding" may mean that the binding substance of interest actually binds to the binding entity (target) bound to the binding substance, It may mean that they are combined in a simulation using a molecular docking method or the like, but the former is preferable. The binding between the binding substance and the binding target can be detected, for example, using a protein-protein interaction analysis method, for example, antibody-antigen reaction such as co-immunoprecipitation, pull-down assay, ELISA method, and flow cytometry. It can be detected using the method used. As a specific example, the binding between the binding substance and the binding target is detected by, for example, contacting a cell expressing the binding target with a labeled binding substance and then detecting the label in the cell. can.

The binding substance is preferably an aptamer or an antibody.

The binding substance may be labeled. The label is not particularly limited, and examples thereof include fluorescent substances, dyes, isotopes, enzymes, and the like. Examples of the fluorescent substance include pyrene, TAMRA, fluorescein, Cy3 dye, Cy5 dye, FAM dye, rhodamine dye, Texas Red dye, and fluorophores such as JOE, MAX, HEX, and TYE. Examples include Alexa dyes such as Alexa488 and Alexa647. Examples of the enzyme include luciferase, alkaline phosphatase, peroxidase, β-galactosidase, glucuronidase and the like.

When the binding substance is a nucleic acid, the label is bound, for example, to at least one of the 5' end and the 3' end of the nucleic acid. On the other hand, if the first binding substance is a protein, the label and carrier are, for example, attached to the N-terminus, C-terminus or side chain of the protein.

The label is, for example, directly or indirectly bound to the binding substance. In said indirect attachment, said label is attached via a linker.

In the present invention, the "sample liquid" may be any liquid. When the specimen to be collected is liquid, it may be used as a liquid sample as it is, or may be a liquid sample prepared by diluting, suspending, dispersing, or the like with a liquid solvent. When the specimen to be collected is solid, it may be a liquid sample prepared by dissolving, suspending, dispersing, or the like in a liquid solvent. When the specimen to be collected is a gas, for example, it may be a liquid sample obtained by concentrating an aerosol in the gas, or a liquid sample prepared by dissolving, suspending, or dispersing in a liquid solvent. The type of the liquid solvent is not particularly limited. For example, it is a solvent that hardly affects the binding between the binding substance and the target, and specific examples thereof include water and buffer solutions. Examples of the specimen to be collected include foods, biological substances (e.g., biological samples such as blood, urine, saliva, and body fluids), soil, wastewater, rainwater, tap water, ponds, rivers, seawater, air, atmosphere, and the like. . The sample liquid may be, for example, a liquid containing targets, a liquid containing no targets, or a liquid whose presence or absence of targets is unknown.

Hereinafter, the manufacturing method of a semiconductor such as MSS and MSS of the present invention will be described in detail with reference to the drawings. However, the invention is not limited to the following description. In FIGS. 1 to 8 below, the same parts are denoted by the same reference numerals, and their description may be omitted. In addition, in the drawings, for convenience of explanation, the structure of each part may be simplified as appropriate, and the dimensional ratio of each part may be schematically shown unlike the actual one.

[Embodiment 1]
This embodiment is an example of the first MSS of the present invention and a method of manufacturing a semiconductor such as the MSS. FIG. 1 is a schematic diagram showing an MSS 1 of Embodiment 1. FIG. In FIG. 1, (A) is a schematic plan view of the MSS 1 and a schematic enlarged view of the periphery of the circuit 12A, and (B) is a schematic cross-sectional view seen from the II direction in (A). As shown in FIG. 1, the MSS 1 of Embodiment 1 comprises a sensor substrate 10, electrodes 11, aluminum wires 12A (circuits), MSS films 13 (films), piezoresistive elements 14, and support regions 15. FIG. The aluminum wire 12A is composed of a conductive layer 121 and an insulating layer 122 (oxide layer). The MSS film 13 is supported by the sensor substrate via a support region 15 having a piezoresistive element 14 formed thereon. Also, the MSS film 13 is connected to the aluminum wire 12A through the piezoresistive element 14, and the aluminum wire 12A is connected to the electrode 11 respectively. In Embodiment 1, the aluminum wire 12A constitutes a Wheatstone bridge circuit including four piezoresistive elements 14 . In addition, although the MSS 1 of Embodiment 1 includes the electrode 11, the electrode 11 has an arbitrary configuration and may or may not be present. If the MSS 1 does not have the electrode 11, the aluminum wire 12A is connected to the voltage applying device.

The sensor substrate 10 is a substrate on which various configurations of the MSS 1 such as the electrodes 11, aluminum wires 12A, and the MSS film 13 can be arranged. As the sensor substrate 10, a semiconductor substrate can be used, and as a specific example, a wafer made of silicon can be used. The sensor substrate 10 supports the MSS film 13 by means of support regions 15 . The sensor substrate 10 preferably partially supports the MSS film 13 , and specifically preferably partially supports the side surfaces of the MSS film 13 .

In the MSS 1 of Embodiment 1, the sensor substrate 10 supports the MSS film 13 via four supporting regions 15 (supporting portions), but the present invention is not limited to this, and the sensor substrate 10 may be any A number of support regions 15 may support the MSS membrane 13 . In addition, the sensor substrate 10 supports one MSS film with four support regions 15, but the present invention is not limited to this. Multiple support regions 15 may each support an MSS membrane 13 . In this case, the number of support regions 15 and the number of supported MSS films 13 on one sensor substrate 10 are not particularly limited, and each may be one or two or more.

The electrode 11 can be configured to apply a voltage to the MSS1 by being connected to a voltage applying device outside the MSS1. The electrode 11 thereby makes it possible to measure the change in stress on the piezoresistive element 14 as a voltage value. The material of the electrode 11 may be any conductive metal, and the description of the material of the circuit in the description of the aluminum wire 12A, which will be described later, can be used. The material of the electrodes 11 may be the same as or different from that of the circuit. In the MSS 1, the electrode 11 is composed of a conductive layer made of a conductive metal, but the present invention is not limited to this. may be formed.

The aluminum wire 12A electrically connects the electrode 11 and the piezoresistive element 14, and makes it possible to measure the change in stress on the piezoresistive element 14 as a voltage value when voltage is applied by the voltage application device outside the MSS1. As shown in FIG. 1B, the aluminum wire 12A has a conductive layer 121 laminated on the substrate 10 on the center side thereof, and the conductive layer 121 is covered with an insulating layer 122. As shown in FIG. That is, the conductive layer 121 is sealed between the substrate 10 and the insulating layer 122 . Therefore, the conductive layer 121 is insulated from the outside of the aluminum wire 12A. Therefore, since the insulation between the aluminum wires 12A is ensured in the MSS1, short-circuiting between the aluminum wires 12A can be suppressed even if the MSS1 is immersed in the sample liquid and a voltage is applied to the MSS1. In the aluminum wire 12A, the conductive layer 121 is made of aluminum that exhibits conductivity, and more specifically, is made of non-oxidized aluminum. On the other hand, in the aluminum wire 12A, the insulating layer 122 is made of aluminum exhibiting insulating properties, specifically, made of aluminum oxide. In addition, the insulation means insulation to the extent that the short circuit between the circuits is significantly suppressed when the MSS 1 is used under normal conditions, as compared with the MSS without the insulation layer 122 .

In the MSS 1 of Embodiment 1, the material of the circuit is aluminum, but in the present invention, the material of the circuit is not limited to this, and may be any metal capable of forming an oxide film layer by oxidation treatment. Examples of metals capable of forming the oxide film layer include valve metals, and specific examples include aluminum, tantalum, niobium, titanium, hafnium, zirconium, zinc, tungsten, bismuth, antimony, molybdenum, chromium, iron, or alloys thereof. In this way, by using a metal that forms an oxide film by oxidation treatment as the material of the circuit, the MSS 1 can ensure insulation between circuits without forming an insulating layer using resin.

In the aluminum wire 12A, the width and height of the conductive layer 121 are sufficient as long as they are thick enough to electrically connect the electrodes 11 and the piezoresistive elements 14 . The width of the conductive layer 121 is, for example, 1-100 μm, 10-20 μm. The height of the conductive layer 121 is, for example, 0.1 to 100 μm, 1 to 20 μm. In addition, the thickness of the insulating layer 122 is such that when the MSS 1 is used under normal conditions, compared to the MSS without the insulating layer 122, the insulating property is such that the short circuit between the aluminum wires 12A is significantly suppressed. Any thickness can be used. The thickness of the insulating layer 122 is, for example, 0.5 to 50 nm, 1 to 10 nm.

In MSS1, the aluminum wire 12A forms a Wheatstone bridge circuit with the piezoresistive element 14, but the circuit structure of the aluminum wire 12A is not limited to this, and the change in stress on the piezoresistive element 14 can be electrically measured. circuit structure.

The MSS film 13 deforms according to the surface stress, and the deformation applies stress to the piezoresistive element 14 . The material of the MSS film 13 is not particularly limited as long as it deforms in response to surface stress and applies stress to the piezoresistive element 14 due to the deformation. The MSS film 13 is, for example, a thin film, and its thickness and surface area are not particularly limited, and are similar to MSS films used in commercially available MSS, for example. Although the planar shape of the MSS film 13 is circular, the present invention is not limited to this, and may be any shape. The planar shape of the MSS film 13 is preferably a perfect circle because it can increase distortion due to surface stress. The material of the MSS film 13 is not particularly limited, and is silicon, for example. A silicon film can be used as the MSS film 13, and as a specific example, any silicon film can be used regardless of p-type or n-type polarity. For example, Si(100) can be used.

The piezoresistive element 14 is an element that detects deformation of the MSS film 13 . When stress is applied to MSS membrane 13 in MSS 1 , the stress change is concentrated in support region 15 . Therefore, in the MSS 1 , the piezoresistive element 14 is formed in the supporting region 15 at a location supporting the MSS film 13 . Moreover, the piezoresistive element 14 changes its resistance value according to the stress change. Therefore, in the MSS1, by applying a voltage through the electrode 11 and the aluminum wire 12A, an output voltage corresponding to the stress can be obtained due to the change in the resistance value. This allows the MSS 1 to analyze the presence or amount of the target in the sample liquid. The piezoresistive element 14 can be manufactured, for example, by doping an arbitrary region of the support region 15 made of a silicon film to make it p-type by doping an impurity. Therefore, the piezoresistive element 14 preferably uses p-type Si formed on a silicon film. Note that the piezoresistive element 14 is formed at the location where the support region 15 supports the MSS film 13, but may be formed in the vicinity of that location.

Next, the manufacturing method of MSS1 will be explained using FIG. FIG. 2 is a schematic cross-sectional view showing an example of a method for manufacturing the MSS1. In the manufacturing method of MSS1 of Embodiment 1, first, MSS is prepared. As MSS, commercially available MSS may be used, or self-prepared. When the MSS 1 is self-prepared, the MSS 1 forms the piezoresistive element 14 by, for example, connecting the MSS film 13 to the supporting region 15 of the sensor substrate 10 and then doping the supporting region 15 . Then, the manufacturing method of the first embodiment can form the electrodes 11 and the aluminum wires 12A, which are circuits, by wiring aluminum by sputtering, vapor deposition, or the like.

As shown in FIG. 2(A), the aluminum wire 12A formed on the MSS is made of conductive aluminum, that is, non-oxidized aluminum. Therefore, in the manufacturing method of the first embodiment, the insulating layer 122 is formed on the surface of the aluminum wire 12A. Specifically, in the manufacturing method of the first embodiment, the aluminum wire 12A is oxidized. In the manufacturing method of Embodiment 1, the oxidation treatment oxidizes the aluminum on the surface of the aluminum wire 12A, thereby forming an insulating oxide film as shown in FIG. is formed.

The oxidation treatment may be any treatment as long as it causes an oxidation reaction with a metal such as aluminum. be done.

When the oxidation treatment is performed by the acid treatment, in the manufacturing method of Embodiment 1, the acid can be applied to the surface of the aluminum wire 12A, and the aluminum wire 12A and the acid can be reacted for a predetermined time. The acid used for the acid treatment is an acid that can act as an oxidizing agent, and specific examples include nitric acid, sulfuric acid, phosphoric acid, oxalic acid, chromic acid, etc., preferably concentrated nitric acid, concentrated sulfuric acid, concentrated phosphoric acid, etc. acid, concentrated oxalic acid, concentrated chromic acid. The reaction time with the acid in the acid treatment can be set according to the type of the acid and the metal forming the circuit, and is, for example, 15 to 180 minutes, 60 to 90 minutes. The reaction temperature of the acid treatment is, for example, 0 to 50°C, preferably 30°C or less.

The plasma treatment can be performed by placing the MSS1 in a plasma generator and generating plasma while supplying oxygen to the plasma generator. The oxygen concentration in the gas supplied to the plasma generator may be, for example, the default setting of each device, eg, 80-100%, 85-95%, about 90%. The plasma treatment time can be set according to the type of metal forming the circuit and the thickness of the insulating layer 122 to be formed. One hour. After the oxygen plasma treatment, an insulating layer 122 containing aluminum oxide and aluminum hydroxide is formed on the surface of the aluminum wire 12A. Therefore, in the manufacturing method of Embodiment 1, after the oxygen plasma treatment, the formed aluminum hydroxide is preferably converted into aluminum oxide by a step including a heat treatment such as an annealing step to be described later. Thereby, the manufacturing method of Embodiment 1 can improve the insulating property of the formed insulating layer 122 .

Thus, the manufacturing method of Embodiment 1 can manufacture MSS1.

Next, an example of a method for analyzing targets in samples using MSS1 will be described. First, in the target analysis method of Embodiment 1, the sample liquid is brought into contact with the MSS1 (contact step). The conditions for immersing MSS1 in the sample liquid are not particularly limited, and examples thereof include a temperature of 20 to 35° C. and a temperature of 0.1 to 120 minutes and a temperature of 50 to 60° C. and 0.1 to 120 minutes. When the MSS1 has a plurality of MSS membranes, for example, the plurality of MSS membranes in the MSS1 may be immersed in the same sample liquid at the same time.

Next, in the analysis method of Embodiment 1, a voltage is applied to the MSS1 in the sample liquid (liquid phase) (application step). Conditions for applying the voltage are not particularly limited, and for example, conditions similar to those for commercially available MSS can be exemplified. The liquid phase may be, for example, the sample liquid in the contacting step, or may be another solvent. In the latter case, the MSS1 after the contacting step is removed from the sample solution, immersed in a new solvent, and voltage is applied. The solvent is not particularly limited, and examples thereof include PBS, buffer solutions such as Tris-HCl, water, and the like.

The timing of the application is the start of the contact process or after a predetermined time has elapsed since the start of the contact process.

Next, according to the analysis method of the first embodiment, the target in the sample liquid is analyzed by measuring the stress change of the piezoresistive element 14 in MSS1. The stress change can be measured, for example, by measuring electrical signals, and a commercially available measurement module (eg, MSS-8RM, NANOSENSOR) can be used. In the analysis method of Embodiment 1, for example, a reference measurement value (signal) may be obtained using a control sample solution that does not contain the target. In this case, in the analysis step, for example, the target may be analyzed using a reference measurement value and a measurement value (signal) of the sample liquid.

In the MSS 1 of Embodiment 1, an insulating layer 122 is formed on the surface of the aluminum wire 12A, which is the line. Therefore, in the MSS1, since the insulation between the aluminum wires 12A is ensured, it is possible to perform measurement in a liquid phase such as a sample liquid. Therefore, MSS1 can be suitably used for the analysis of liquid samples. In addition, in the manufacturing method of the first embodiment, by using a metal that forms an oxide film by oxidation treatment as the material of the circuit, an oxide film layer can be formed by oxidation treatment, and the oxide film layer is used as the insulating layer 122 . be able to. Therefore, the manufacturing method according to the first embodiment does not require coating of the lines with resin, and damage due to adhesion of the resin to the piezoresistive element 14 can be avoided. Therefore, according to the manufacturing method of Embodiment 1, the breakage rate during manufacturing can be reduced, so the yield in manufacturing the MSS1 can be improved.

In the manufacturing method of Embodiment 1, one type of oxidation treatment is performed, but the present invention is not limited to this, and multiple oxidation treatments may be performed. Moreover, the manufacturing method of Embodiment 1 may further include a step of annealing the formed oxide film layer. Another example of the manufacturing method of Embodiment 1 will be described with reference to FIG.

FIG. 3 is a schematic diagram showing another example of the method for manufacturing the MSS1 of Embodiment 1. FIG. In the manufacturing method of another example of the first embodiment, the conductive layer 121 of the aluminum wire 12A is subjected to two-stage oxidation treatment (FIGS. 3B and 3C), and then heat treatment to remove the oxide formed. Annealing the film layer forms an insulating layer 122 having a higher insulating property (FIG. 3A).

First, in the manufacturing method of another example of the first embodiment, the aluminum wire 12A formed on the MSS is oxidized by acid treatment. Thereby, as shown in FIG. 3B, in the manufacturing method of another example of the first embodiment, an insulating layer 122 which is an oxide film layer is formed on the surface of the conductive layer 121 .

Next, in another example of the manufacturing method of Embodiment 1, the MSS after the acid treatment is subjected to plasma treatment in an oxygen atmosphere. As a result, as shown in FIG. 3C, in the manufacturing method of the other example of the first embodiment, the oxide film layer formed on the surface of the aluminum wire 12A is grown to increase the thickness of the insulating layer 122. As shown in FIG. Further, in the oxygen plasma treatment, aluminum hydroxide is formed in the insulating layer 122 in addition to aluminum oxide.

Then, in the manufacturing method of another example of Embodiment 1, heat treatment is performed on the MSS after the oxygen plasma treatment. As a result, as shown in FIG. 3D, in the manufacturing method of another example of the first embodiment, the insulating layer 122 formed on the surface of the aluminum wire 12A is annealed, and the aluminum hydroxide in the insulating layer 122 is removed. can be converted to aluminum oxide to improve insulation. The lower limit of the temperature in the heat treatment is preferably 100° C. or higher, 200° C. or higher, or 300° C. or higher, for example. The upper limit of the temperature in the heat treatment is, for example, 400° C. or less. Further, the temperature in the heat treatment is, for example, 100 to 400°C, 200 to 400°C, 300 to 400°C, and about 350°C. The heat treatment time is, for example, 60 to 180 minutes.

Thereby, the manufacturing method of another example of Embodiment 1 can manufacture MSS1. According to another example of the manufacturing method of Embodiment 1, the thickness of the insulating layer 122 in the MSS1 can be increased. Therefore, according to the manufacturing method of another example of the first embodiment, the MSS1 with higher insulation can be manufactured.

In the MSS 1 of this embodiment, the binding substance is not immobilized on the MSS membrane 13 . However, the present invention is not limited to this, and the binding substance may be immobilized on MSS1. In this case, the binding substance may be immobilized on one surface of the MSS membrane 13, or may be immobilized on both surfaces. When the binding substance is immobilized on both sides of the MSS membrane 13, the binding substance on one surface and the binding substance on the other surface are preferably the same binding substance that binds to the same target, for example. Hereinafter, a method for immobilizing a binding substance on the MSS membrane 13 will be described by taking as an example a case where an aptamer is used as the binding substance, but the present invention is not limited to this, and a protein such as an antibody may be immobilized. good too. Further, in the following description, the surface of the MSS film 13 may be, for example, one surface or both surfaces.

The method for immobilizing the aptamer on the MSS membrane 13 is not particularly limited, and the aptamer may be immobilized directly or indirectly on the MSS membrane 13. In the former case, for example, the MSS membrane 13 and the aptamer can be immobilized by covalent bonding or the like by chemically treating the aptamer. The direct fixing method includes, for example, a method using photolithography, and for specific examples, reference can be made to US Pat. No. 5,424,186. Another direct immobilization method is, for example, a method of synthesizing the aptamer on the MSS membrane 13 . This method includes, for example, a so-called spot method, and for specific examples, reference can be made to US Pat. No. 5,807,522. In the latter case, for example, the aptamer can be immobilized on the MSS membrane 13 via a linker. The type of linker is not limited at all, and examples thereof include a combination of biotin or a biotin derivative (hereinafter referred to as biotins) and avidin or an avidin derivative (hereinafter referred to as avidins). Examples of the biotin derivative include biocytin, and examples of the avidin derivative include streptavidin. The length of the linker is, for example, the length of the shortest molecular chain (main chain long). The main chain length of the linker is 1 to 20, and is preferably 1 to 15, 1 to 13, 3 to 13, 5 to 13, 1 to 11, 3 to 11 because it can improve the sensitivity of the MSS membrane 13. , 1-10, 3-10, 1-8, 3-8, 1-5, 1-3, 1 or 2. Examples of immobilization methods are shown below, but the present invention is not limited to these.

As a first example, the biotins are bound to one of the MSS membrane 13 and the aptamer, and the avidins are bound to the other. Then, the aptamer can be indirectly immobilized on the MSS membrane 13 by binding the biotins and the avidins.

In the first example, the aptamer was indirectly immobilized on the MSS membrane by utilizing the specific binding between avidins and biotins, that is, the affinity of biotins to avidins. Not limited to this, affinity tags other than avidins-biotins may be used. Examples of the affinity tag include His tag (His×6 tag)-nickel ion, glutathione-S-transferase-glutathione, maltose binding protein-maltose, epitope tag (myc tag, FLAG tag, HA (hemagglutinin) tag)- Antibodies or antigen-binding fragments can be used. The point that other affinity tags may be used is the same in the second to fourth examples described later.

As a second example, the aptamer may be immobilized on the MSS membrane 13 via an intervening membrane, for example. Forming the intervening membrane on the MSS membrane 13, binding the biotins to one of the intervening membrane and the aptamer in the same manner as in the first example, and binding the avidins to the other, By binding the biotins and the avidins, the aptamer can be immobilized on the MSS through the intervening membrane. The intervening film is, for example, a film of metal such as gold, and can be formed by vapor-depositing the metal on the MSS film 13 . The thickness of the intervening film is not particularly limited, and is, for example, 10 to 100 nm. The intervening film may be, for example, one layer or two or more layers. When the surface of the intervening film is made of gold, the intervening film is, for example, two-layered, and a metal film (adhesive film) for adhesion to the MSS film 13 is provided in order to improve the adhesiveness of the gold film. It is preferable to form the gold film through the metal. Examples of the metal of the adhesive film include titanium and chromium. The thickness of the adhesive film is, for example, 0.1 to 10 nm, and the thickness of the gold film is, for example, 0.1 to 100 nm. When the biotins are bound to the intervening membrane, for example, self-assembled monolayers (SAMs) of thiolalkanes are formed on the surface of the intervening membrane using thiolalkanes to which the biotins are bound. The aptamer formed and bound with the biotins may be brought into contact, and the aptamer may be immobilized by binding the biotins and the avidins.

As a third example, there is a method of binding an amino group to the MSS film 13 and further binding glutaraldehyde to bind the streptavidins. That is, the MSS film 13 is reacted with a silane coupling agent having an amino group to bond the amino group onto the MSS film 13 . The reaction can be carried out, for example, by coating the MSS film 13 with a solution containing a silane coupling agent having an amino group. Furthermore, for the MSS membrane 13, a cross-linking agent capable of binding the amino group and the main chain or side chain of the amino acid, or for the MSS membrane 13, a linker between the amino group and the main chain or side chain of the amino acid is reacted with a cross-linking agent such as glutaraldehyde capable of forming a , and one end of the cross-linking agent such as glutaraldehyde is bound to the amino group on the MSS film 13 . Specifically, after silane coupling, the surface of the MSS film is washed, and a solution containing a cross-linking agent is applied to the MSS film 13 to bond the amino groups with the cross-linking agent. The conditions for the cross-linking reaction can be appropriately determined according to, for example, the type of cross-linking agent. Next, the avidins are bound to the other end of the cross-linking agent such as glutaraldehyde. Specifically, after cross-linking, the surface of the MSS membrane is washed, and a solution containing avidins is applied to bind the other end of the cross-linking agent to the main chain or side chain of amino acids of avidins. Then, the biotin-bound aptamer is brought into contact with the MSS membrane 13 treated in this way, and the aptamer can be immobilized by binding the biotins and the avidins.

A silane coupling agent is represented by, for example, Y—Si(CH ₃ ) _3-n (OR) _n . When the silane coupling agent is a silane coupling agent having an amino group, examples of n, R, and Y are given below. The "n" is 2 or 3. Examples of R include alkyl groups such as methyl group and ethyl group; acyl groups such as acetyl group and propyl group; and the like. Y is a reactive functional group terminating with an amino group.

Silane coupling agents having an amino group include, for example, N-(2-aminoethyl)-3-aminopropylmethyldimethoxysilane (eg, KBM-602 (manufactured by Shin-Etsu Silicone Co., Ltd.)), N-(2-aminoethyl )-3-aminopropyltrimethoxysilane (e.g., KBM-603 (manufactured by Shin-Etsu Silicone Co., Ltd.)), 3-aminopropyltrimethoxysilane (e.g., KBM-903 (manufactured by Shin-Etsu Silicone Co., Ltd.)), 3-aminopropyltriethoxy Silane (e.g., KBE-903 (manufactured by Shin-Etsu Silicone Co., Ltd.)), 3-(2-aminoethylamino)propyltrimethoxysilane (e.g., GENIOSIL (registered trademark) GF 91 (manufactured by Asahi Kasei Wacker Silicone Co., Ltd.)), 3-( 2-aminoethylamino)propylmethyldimethoxysilane (eg, GENIOSIL (registered trademark) GF 95 (manufactured by Asahi Kasei Wacker Silicone Co., Ltd.)) and the like.

The cross-linking agent can be appropriately determined according to the functional group of the main chain or side chain of the amino acid that binds to the linker. Examples of the functional group include an amino group (--NH ₂ ), a thiol group (--SH), a carboxyl group (--COOH) and the like. Said amino groups are present, for example, at the N-terminus of proteins or peptides or at the side chains of lysines. The thiol group is, for example, a side chain of cysteine. Said carboxyl groups are present, for example, at the C-terminus of proteins or peptides or at the side chains of aspartic acid or glutamic acid.

When the amino group of the main chain or side chain of the amino acid is used, examples of the cross-linking agent include a cross-linking agent having aldehyde groups at both ends such as glutaraldehyde; bis(sulfosuccinimidyl) suberate (BS3); Disuccinimidyl glutarate (DSG), Disuccinimidyl suberate (DSS), Dithiobis(succinimidylpropionate), Dithiobis(sulfosuccinimidylpropionate) (DSP), Dithiobis(succinimidylpropionate) (DTSP) , dithiobis(sulfosuccinimidyl propionate) (DTSSP), disuccinimidyl tartrate (DST), ethylene glycol bis(succinimidyl succinate) (ESG), ethylene glycol bis(sulfosuccinimidyl succinate) ) (Sulfo-ESG), N-hydroxysuccinimide active esters (N-hydroxysuccinimide reactive groups) such as PEGylated bis(sulfosuccinimidyl) (BS(PEG)5, BS(PEG)9, etc.) at both ends cross-linking agents having imide ester reactive groups at both ends, such as dimethyl adipimidate (DMA), dimethyl pimelimidate (DMP), and dimethyl pimelimidate (DMS);

When the thiol group of the side chain of the amino acid is used, examples of the cross-linking agent include N-(6-maleimidocaproyloxy) succinimide (EMCS), N-(6-maleimidocaproyloxy) sulfosuccinimide (Sulfo -EMCS), N-(8-maleimidocaryloxy) succinimide (HMCS), N-(8-maleimidocaryloxy) sulfosuccinimide (Suflo-HMCS), N-α-maleimidoacet-oxysuccinimide ester (AMAS), N-β -maleimidopropyl-oxysuccinimide ester (BMPS), N-γ-maleimidobutyryl-oxysuccinimide ester (GMBS), N-γ-maleimidobutyryl-oxysulfosuccinimide ester (Sulfo-GMBS), m-maleimidobenzoyl-N-hydroxysuccinimide ester (MBS), m-maleimidobenzoyl -N-hydroxysulfosuccinimide ester (Sulfo-MBS), succinimidyl 4-(N-maleimidomethyl) cyclohexane-1-carboxylate (SMCC), sulfosuccinimidyl 4-(N-maleimidomethyl) cyclohexane-1-carboxylate (Sulfo-SMCC), succinimidyl 4- (p-maleimidophenyl) butyrate (SMPB), sulfosuccinimidyl 4-(N-maleimidophenyl) butyrate (Sulfo-SMPB), Succinimidyl 6-((beta-maleimidopropionamido) hexanoate) (SMPH), succinimidyl 4-(N-maleimidomethyl)cyclohexane- 1-carboxy-(6-amidocaproate) (LC-SMCC), N-κ-maleimidoundecanoyl-oxysulfosuccinimide ester (Sulfo-KMUS) and other maleimide groups and N-hydroxysuccinimide active esters at both ends of the molecule N-hydroxysuccinimide such as succinimidyl iodoacetate (SIA), succinimidyl 3-(bromoacetamido) propionate (SBAP), succinimidyl (4-iodoacetyl) aminobenzoate (SIAB), sulfosuccinimidyl (4-iodoacetyl) aminobenzoate (Sulfo-SIAB) Crosslinkers with active esters and haloacetyl-reactive groups on both ends; (3(2-pyridyldithio) propionamido) hexanoate (Sulfo-LC-SPDP), 4-succinimidyloxycarbonyl-alpha-methyl-α(2-pyridyldithio) toluene (SMPT), 2-Pyridyldithiol-tetraoxatetradecane-N-hydoxysuccinimide (PEG4-SPDP) ), 2-Pyridyldithiol-tetraoxaoctatriacontane-N-hydoxysuccinimide (PEG12-SPDP) or other N-hydroxysuccinimide active ester and a cross-linking agent having pyridyldithiol reactive groups at both ends;

When using the carboxyl group of the main chain or side chain of the amino acid, examples of the cross-linking agent include dicyclohexylcarbodiimide (DCC), 1-Ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC), N-hydroxysuccinimide (NHS ), N-hydroxysulfosuccinimide (Sulfo-NHS), acetic anhydride, and the like. In addition, DCC, EDC, NHS, Sulfo-NHS, and acetic anhydride, for example, directly bond the amino group and the carboxyl group, do not remain between the carboxyl group and the amino group, the linker region derived from the cross-linking agent (group) does not occur.

The cross-linking agent is preferably a cross-linking agent that does not substantially cause self-condensation because the length of the linker can be made substantially constant or constant. The above-mentioned "constant linker length" means, for example, that in the linkers of a plurality of aptamers, the linkers of each aptamer have substantially the same or the same length. The length of the linker can be achieved, for example, by making the structures of the linkers the same. In the third example, the sensitivity of MSS can be improved by using such a cross-linking agent. The improvement in sensitivity is presumed to be due to the following reasons. In addition, the present invention is not limited to the following estimation. When a target binds to an aptamer, steric hindrance caused by the target occurs around the aptamer bound to the target. If the aptamers are immobilized at different distances from the MSS membrane 13, the targets are more likely to come into contact with aptamers that are distal to the MSS membrane 13. FIG. Therefore, it is presumed that the target preferentially binds to the aptamer on the distal end side from the MSS membrane 13 . In this case, even if steric hindrance due to the target occurs around the target-bound aptamer, other aptamers are present on the MSS membrane 13 side compared to the target-bound aptamer. Because of its large number, it is less susceptible to steric hindrance caused by the target. Therefore, even if the target binds to the aptamer, the surrounding aptamers are less likely to move due to steric hindrance. Therefore, when the aptamers are immobilized at different distances with respect to the MSS membrane 13, distortion of the MSS membrane may occur on the MSS membrane 13 due to the displacement of the surrounding aptamers. relatively low. On the other hand, when the aptamers are immobilized on the MSS membrane 13 at substantially the same distance, when the aptamers bind to the targets, the surrounding aptamers are affected by steric hindrance caused by the targets. Therefore, there is a high possibility that the positions of the surrounding aptamers will move, and the possibility that the MSS membrane will be distorted due to the movement of the positions of the surrounding aptamers is also relatively high. That is, when the aptamers are immobilized on the MSS membrane 13 at approximately the same distance, the binding of one aptamer to the target on the MSS membrane 13 also causes the position of the surrounding aptamers to move. The distortion of the MSS film 13 is amplified. Therefore, when the aptamer is immobilized on the MSS membrane 13 at substantially the same distance, that is, when the linker length is substantially constant, the sensitivity of the MSS membrane 13 is presumed to be improved. be.

Specific examples of the cross-linking agent that does not substantially cause self-condensation include, for example, a cross-linking agent having N-hydroxysuccinimide active esters at both ends, a cross-linking agent having imide ester reactive groups at both ends, and a maleimide group and N-hydroxysuccinimide. a cross-linking agent having an active ester at both ends of the molecule, a cross-linking agent having an N-hydroxysuccinimide active ester and a haloacetyl-reactive group at both ends, a cross-linking agent having an N-hydroxysuccinimide active ester and a pyridyldithiol-reactive group at both ends, DCC, EDC, NHS, Sulfo-NHS, acetic anhydride and the like.

The linker is represented, for example, by the following formula (1). In the following formula (1), M ₁ represents an atom bonded to the silane coupling agent on the MSS film, L ₁ represents a region (group) derived from the silane coupling agent, and L ₂ represents a cross-linking Represents the agent _- derived region (group), L2 is optional, and _M2 represents the atom that binds to the crosslinker or NH in the affinity tag. NH represents an amine derived from an amino group of a silane coupling agent having an amino group.
M ₁ -L ₁ -NH-L ₂ -M ₂ (1)

L ₁ is, for example, (M ₁ )—Si(CH ₃ ) _2-m (OR ₄ ) _m —R ₁ —(NH) or (M ₁ )—Si(CH ₃ ) _2-m (OR ₄ ) _m —R ₂ —NH—R ₃ —(NH). R ₁ is a straight or branched alkyl group having 1 to 5 carbon atoms. R ₂ and R ₃ are, for example, each independently a linear or branched alkyl group having 1 to 5 carbon atoms, and may be the same or different. Examples of the alkyl group include methyl group, ethyl group, propyl group, butyl group and pentyl group. _R4 is, for example, a hydrogen atom or a bond. m is 1 or 2;

When 3-aminopropyltriethoxysilane is used as the silane coupling agent, L ₁ is represented by, for example, (M ₁ )-Si(OR ₄ ) ₂ --(CH ₂ ) ₃ --(NH). . _R4 is, for example, a hydrogen atom or a bond. Further, when glutaraldehyde is used as the cross-linking agent, L ₂ is, for example, (NH)=CH--C ₃ H ₆ --CH=(CH(CHO)--C ₂ H ₄ --CH) _n =CH( CHO)—C ₂ H ₄ —C=(M ₂ ).

The length of the linker is, for example, the shortest molecular chain length (main chain length) between the functional group on the MSS membrane (for example, the oxygen atom of the silanol group on the silicon membrane) and the affinity tag such as avidin. can be represented. The main chain length of the linker is 1 to 20, and since it can improve the sensitivity of MSS, it is preferably 1 to 15, 1 to 13, 3 to 13, 5 to 13, 1 to 11, 3 to 11, 1 ~10, 3-10, 1-8, 3-8, 1-5, 1-3, 1 or 2.

In the third example, avidin-biotin binding is used, but the third example is not limited to this, and a linker is directly bound to the hydroxyl group or phosphate group of the aptamer. good too. In this case, the aptamer can be immobilized on the MSS membrane 13 by amidating the 3' terminal phosphate group and reacting it with the linker.

As a fourth example, a methacryl group (--C(=O)--C(CH ₃ )=CH ₂ ) is bound to the MSS membrane 13, and an amino acid or derivative thereof (hereinafter referred to as "amino acid derivative"). and binding the streptavidins via. That is, the MSS film 13 is reacted with a silane coupling agent having a methacrylic group to bond an amino group onto the MSS film 13 . The reaction can be carried out, for example, by coating the MSS film 13 with a solution containing a silane coupling agent having a methacrylic group. Furthermore, after reacting an amino acid derivative such as N-acetylcysteine with the MSS film 13, a linker is formed between the main chain or side chain of the amino acid derivative and the main chain or side chain of the amino acid of the avidins. A possible cross-linking agent is reacted to bind one end of the cross-linking agent to the amino acid derivative on the MSS membrane 13 . Specifically, the surface of the MSS membrane after treatment with the amino acid derivative is washed, and a solution containing a cross-linking agent is applied to the MSS membrane 13 to bind the amino acid derivative and the cross-linking agent. The conditions for the cross-linking reaction can be appropriately determined according to, for example, the type of cross-linking agent. Next, the avidins are bound to the other end of the cross-linking agent. Specifically, after cross-linking, the surface of the MSS membrane is washed, and a solution containing avidins is applied to bind the other end of the cross-linking agent to the main chain or side chain of amino acids of avidins. Then, the biotin-bound aptamer is brought into contact with the MSS membrane 13 treated in this way, and the aptamer can be immobilized by binding the biotins and the avidins.

As mentioned above, the silane coupling agent is represented by, for example, Y—Si(CH ₃ ) _3-n (OR) _n . When the silane coupling agent is a silane coupling agent having a methacryl group, examples of n, R, and Y are given below. The "n" is 2 or 3. Examples of R include alkyl groups such as methyl group and ethyl group; acyl groups such as acetyl group and propyl group; and the like. Y is a reactive functional group terminated with a methacryl group.

The silane coupling agent having a methacrylic group includes, for example, 3-(methacryloyloxy)propylmethyldimethoxysilane (eg, KBM-502 (manufactured by Shin-Etsu Silicone Co., Ltd.)), 3-(methacryloyloxy)propyltrimethoxysilane (eg, KBM-503 (manufactured by Shin-Etsu Silicone Co., Ltd.), GENIOSIL (registered trademark) GF31 (manufactured by Asahi Kasei Wacker Silicone Co., Ltd.)), 3-(methacryloyloxy) propylmethyldimethoxysilane (e.g., KBE-502 (manufactured by Shin-Etsu Silicone Co., Ltd.)), ( 3-methacryloyloxypropyl)triethoxysilane (for example, KBE-503 (manufactured by Shin-Etsu Silicone Co., Ltd.)) and the like.

The amino acid or amino acid derivative has, for example, a functional group capable of reacting with a methacryl group and a carboxyl group. Examples of the functional group capable of reacting with the methacrylic group include a thiol group (--SH). Examples of the amino acid or amino acid derivative having a thiol group include cysteine; cysteine having an amino group modified such as N-acetylcysteine; and the like.

The cross-linking agent can be appropriately determined according to, for example, the functional group of the amino acid derivative subjected to cross-linking and the functional group of the amino acid of the avidins to be subjected to cross-linking. As a specific example, when the two functional groups are amino groups, the cross-linking agent can refer to the description of the cross-linking agent in the case of using the amino group of the main chain or side chain of the amino acid in the third example. Further, when one of the two functional groups is an amino group and the other is a thiol group, the cross-linking agent is the side chain thiol group of the amino acid in the third example. can be used. Furthermore, when one of the two functional groups is an amino group and the other is a carboxyl group, the cross-linking agent can be used for cross-linking when utilizing the carboxyl group of the main chain or side chain of the amino acid in the third example. The description of the agent can be used.

The cross-linking agent is preferably a cross-linking agent that does not substantially cause self-condensation because the length of the linker can be made constant. In the fourth example, by using such a cross-linking agent, the sensitivity of MSS can be improved by a mechanism similar to the mechanism described in the third example. Specific examples of cross-linking agents that do not substantially cause self-condensation include cross-linking agents having N-hydroxysuccinimide active esters at both ends, cross-linking agents having imide ester reactive groups at both ends, and maleimide groups and N-hydroxysuccinimide active groups. cross-linking agent having an ester at both ends of the molecule, cross-linking agent having an N-hydroxysuccinimide active ester and a haloacetyl reactive group at both ends, cross-linking agent having an N-hydroxysuccinimide active ester and a pyridyldithiol reactive group at both ends, DCC , EDC, NHS, Sulfo-NHS, acetic anhydride and the like.

The linker is represented, for example, by the following formula (2). In the following formula (2), M ₁ represents an atom bonded to the silane coupling agent on the MSS film, L ₁ represents a region (group) derived from the silane coupling agent, and A is an amino acid derivative. , L2 represents a crosslinker _- derived region ₍ group), L2 may or may not be present, and _M2 represents an atom bonded to the crosslinker or NH in the affinity tag.
M ₁ -L ₁ -AL ₂ -M ₂ (2)

L ₁ is represented, for example, by (M ₁ )—Si(CH ₃ ) _2-m (OR ₄ ) _m —R ₅ —C(═O)—CH ₁ (CH ₃ ) _2-1 —(A) be. _R4 is, for example, a hydrogen atom or a bond. R ₅ is a linear or branched alkyl group having 1 to 5 carbon atoms. Examples of the alkyl group include methyl group, ethyl group, propyl group, butyl group and pentyl group. m is 1 or 2; l is 0 or 1;

When 3-(methacryloyloxy)propyltrimethoxysilane is used as the silane coupling agent, L ₁ is, for example, (M ₁ )—Si(OR ₄ ) ₂ —(CH ₂ ) ₃ —OC( ═O)—C(CH ₃ ) ₂ —(A). _R4 is, for example, a hydrogen atom or a bond. Further, when N-acetylcysteine is used as the amino acid derivative, A is, for example, (L ₁ )-S-CH ₂ -CH(-NHCOCH ₃ )-C(=O)-(M ₂ ). be done. When acetic anhydride is used as the cross _- linking agent, L2 is absent, for example.

The length of the linker can be represented, for example, by the shortest molecular chain length (main chain length) between the functional group on the MSS membrane (eg, silanol group on the silicon membrane) and the affinity tag such as avidin. can. The main chain length of the linker is 1 to 20, and since it can improve the sensitivity of MSS, it is preferably 1 to 15, 1 to 13, 1 to 11, 1 to 10, 1 to 8, 1 to 5, 1 ~3, 1 or 2.

In the fourth example, avidin-biotin binding is used, but the fourth example is not limited to this, and a linker is directly bound to the hydroxyl group or phosphate group of the aptamer. good too. In this case, the aptamer can be immobilized on the MSS membrane 13 by amidating the 3' terminal phosphate group and reacting it with the linker.

The site for immobilizing the aptamer on the MSS membrane 13 is not particularly limited, and examples include the 3' end or 5' end.

[Embodiment 2]
This embodiment is an example of the second MSS of the present invention and a method of manufacturing a semiconductor such as the MSS. FIG. 4 is a schematic diagram showing the MSS2 of the second embodiment. In FIG. 4, (A) is a schematic plan view of the MSS2, and (B) is a schematic cross-sectional view of (A) viewed from the II-II direction. As shown in FIG. 4, the MSS 2 of Embodiment 2 comprises a sensor substrate 10, electrodes 11, aluminum wires 121 (circuits), MSS films 13A (films), piezoresistive elements 14, and support regions 15. FIG. The aluminum wire 12 is composed of a conductive layer 121 . The MSS film 13A is composed of a support layer 131 and an amorphous layer 132. As shown in FIG. The MSS film 13A is supported by the sensor substrate via a support region 15 having a piezoresistive element 14 formed thereon. Also, the MSS film 13A is connected to the aluminum wire 121 through the piezoresistive element 14, and the aluminum wire 121 is connected to the electrode 11 respectively. In Embodiment 2, the aluminum wire 121 constitutes a Wheatstone bridge circuit including four piezoresistive elements 14 . Except for this point, the MSS2 of the second embodiment has the same configuration as the MSS1 of the first embodiment, and the description thereof can be used. Although the MSS 2 of Embodiment 2 includes the electrode 11, the electrode 11 has any configuration and may or may not be present. If the MSS 2 does not have electrodes 11, an aluminum wire 121 is connected to the voltage application device.

As shown in FIG. 4B, the MSS film 13A is composed of a support layer 131 and an amorphous layer 132, and the amorphous layer 132 is laminated on the support layer 131. When the MSS film 13A is a silicon film, the support layer 131 is made of crystalline silicon, for example. On the other hand, the amorphous layer 132 is composed of amorphous silicon (eg, amorphized silicon). In the MSS film 13A, one surface of the film is composed of amorphous molecules, so that the contact area with other substances and the number of atoms that can be used for bonding are reduced compared to the case where the film is composed only of crystalline molecules. can be relatively increased. Therefore, by including the amorphous layer 132, the MSS film 13A can immobilize a larger amount of the binding substance on the surface of the MSS film 13A than the MSS film 13 does.

When the MSS film 13A is a silicon film, the amorphous layer 132 may contain other molecules in addition to amorphous silicon. Examples of the other molecules include silicon carbide (silicon carbide), oxides of silicon carbide, and the like.

Next, the manufacturing method of MSS2 will be explained using FIG. FIG. 5 is a schematic cross-sectional view showing an example of a method for manufacturing the MSS2. In the manufacturing method of MSS2 of Embodiment 2, first, MSS is prepared. As MSS, commercially available MSS may be used, or self-prepared.

As shown in FIG. 5(A), the MSS film 13 of the MSS is composed of a support layer 131 composed of crystalline silicon. Therefore, in the manufacturing method of Embodiment 2, the amorphous layer 132 is formed on the surface of the support layer 131 . Specifically, in the manufacturing method of the second embodiment, the support layer 131 is subjected to an amorphization treatment (amorphization treatment). In the manufacturing method of Embodiment 2, the crystalline silicon on the surface of the support layer 131 is made amorphous by the amorphization treatment, and as shown in FIG. Therefore, an amorphous layer 132 made of amorphous silicon is formed.

The amorphization treatment may be any treatment as long as it causes a change in crystal structure. Growth treatment (for example, plasma CVD (Chemical Vapor Deposition), reactive ion etching (RIE), etc.) and the like can be mentioned.

Thus, the manufacturing method of Embodiment 2 can manufacture MSS2.

In the MSS2 of Embodiment 2, an amorphous layer 132 is formed on the surface of the MSS film 13A. Therefore, in MSS2, in the MSS film, the surface area of the MSS film that can be bound to the binding substance is increased, and the contact area with other substances and the number of atoms that can be used for binding are increased as compared with MSS1. Relatively increasing. Therefore, in the MSS2 of Embodiment 2, the immobilization efficiency of the binding substance can be improved.

In the MSS 2 of Embodiment 2, the entire surface of one surface of the MSS film 13A is the amorphous layer 132, but the present invention is not limited to this, and part of one surface may be the amorphous layer 132. . Further, in the MSS2 of Embodiment 2, part or the entire surface of both surfaces of the MSS film 13A may be the amorphous layer 132. FIG. As a result, it is possible to prevent damage to the piezoresistive element 14 and the like during modification processing of the MSS film in the manufacturing method of the MSS2, which will be described later.

In the manufacturing method of Embodiment 2, the amorphization treatment was performed as the modification treatment of the MSS film, but the present invention is not limited to this, and the amorphization treatment is combined with other modification treatments. good too. Another example of the method for manufacturing MSS will be described with reference to FIGS.

Another example of the MSS 2 of Embodiment 2 includes an MSS film 13B instead of the MSS film 13A. As shown in FIG. 6, the MSS film 13B is composed of a supporting layer 131 and an amorphous layer 132. As shown in FIG. The amorphous layer 132 is composed of a silicon carbide layer 133 and a silicon carbide oxide layer 135 . In the MSS 13B, the support layer 131, the silicon carbide layer 133, and the silicon carbide oxide layer 135 are arranged (stacked) in this order. The silicon carbide layer 133 is a layer containing amorphous silicon and silicon carbide. The silicon carbide oxide layer 135 is a layer containing amorphous silicon and silicon carbide oxide. In the MSS film 13B, since the silicon carbide oxide layer 135 on the surface contains silicon carbide oxide, the types of functional groups increase compared to crystalline silicon and amorphous silicon. It is excellent in reactivity with the above-mentioned cross-linking agents such as Specifically, since the MSS film 13B contains carbon atoms in its surface layer, carbon atoms can be selected as atoms to be bonded in addition to silicon atoms. Therefore, according to MSS2, binding molecules such as nucleic acid molecules or proteins can be introduced more easily. Therefore, according to MSS2, the binding substance can be more efficiently immobilized using the cross-linking agent.

FIG. 7 is a schematic diagram showing another example of a method for producing MSS2. In another example of the method of manufacturing the MSS2, the MSS film 131, which is a crystalline silicon film, is subjected to the amorphization treatment to form an amorphous layer 132 (FIG. 7B). Next, carbon is introduced into the amorphous layer 132 to form a silicon carbide layer 133 containing silicon carbide (FIG. 7(C)), after which part of it is converted into an oxide layer 135 of silicon carbide. (Fig. 7(D)).

First, in the method of manufacturing the MSS 2 of another example, the crystalline silicon film 131, which is the MSS film, is subjected to an amorphization treatment by ion beam treatment. Thereby, an amorphous layer 132 is formed on the surface of the support layer 131 as shown in FIG. 7B.

Next, in another example of the method for manufacturing MSS2, carbon is introduced into the MSS film after the amorphization treatment. Thereby, as shown in FIG. 7C, a silicon carbide layer 133 and a carbon layer 134 are formed.

The method for introducing the carbon can be implemented by, for example, a vapor deposition method, a CVD method, a coating method, or the like.

In another example of the method for manufacturing MSS2, after forming the silicon carbide layer 133 and the carbon layer 134, heat treatment may be performed in order to promote the reaction between the introduced carbon and the silicon in the amorphous silicon. The lower limit of the temperature in the heat treatment is preferably 100° C. or higher, 200° C. or higher, or 300° C. or higher, for example. The upper limit of the temperature in the heat treatment is, for example, 400° C. or less. Further, the temperature in the heat treatment is, for example, 100 to 400°C, 200 to 400°C, 300 to 400°C, and about 350°C. The heat treatment time is, for example, 60 to 180 minutes.

Next, in the method of manufacturing the MSS2 of another example, the carbon layer 134 is removed from the MSS film after the introduction of carbon by plasma treatment in an oxygen atmosphere. Thereby, as shown in FIG. 7D, a silicon carbide layer 133 and a silicon carbide oxide layer 135 are formed.

For the oxygen plasma treatment, for example, the above explanation of the oxygen plasma treatment can be used.

The removal of the carbon layer 134 may be performed by a method other than the oxygen plasma treatment, and specific examples thereof include sputtering treatment in an inert gas atmosphere such as helium, neon, argon, and dry etching treatment. .

As a result, the manufacturing method of Embodiment 2 can manufacture another example of MSS2.

[Embodiment 3]
This embodiment is an example of the first and second MSS of the present invention. FIG. 8 is a schematic diagram showing the MSS3 of the third embodiment. As shown in FIG. 8, the MSS 3 of Embodiment 3 comprises a sensor substrate 10, electrodes 11, aluminum wires 12A (circuits), MSS films 13B (films), piezoresistive elements 14, and support regions 15. FIG. Aluminum wire 12A is composed of conductive layer 121 and insulating layer 122 . The MSS film 13B is composed of a support layer 131 and an amorphous layer 132, and the amorphous layer 132 is composed of a silicon carbide layer 133 and a silicon carbide oxide layer 135. FIG. The MSS film 13B is supported by the sensor substrate via a support region 15 having a piezoresistive element 14 formed thereon. Also, the MSS film 13B is connected to the aluminum wire 12A through the piezoresistive element 14, and the aluminum wire 12A is connected to the electrode 11, respectively. In Embodiment 3, the aluminum wire 12A constitutes a Wheatstone bridge circuit including four piezoresistive elements 14. As shown in FIG. Except for this point, the MSS3 of Embodiment 3 has the same configuration as the MSS1 of Embodiment 1, and the description thereof can be used. Although the MSS 3 of Embodiment 3 includes the electrode 11, the electrode 11 has any configuration and may or may not be present. If the MSS 2 does not have the electrode 11, the aluminum wire 12A is connected to the voltage applying device.

In the MSS3 of Embodiment 3, an insulating layer 122 is formed on the surface of the aluminum wire 12A, which is the line. Therefore, in the MSS3, since the insulation between the aluminum wires 12A is ensured, it is possible to perform measurement in a liquid phase such as a sample liquid. Therefore, MSS3 can be suitably used for the analysis of liquid samples. In the MSS 3 of Embodiment 3, an amorphous layer 132 is formed on the surface of the MSS film 13B, and the amorphous layer 132 is composed of a silicon carbide layer 133 and a silicon carbide oxide layer 135 . Further, in the MSS film 13B, an oxide layer 135 of silicon carbide is arranged (stacked) on the surface. In the MSS film 13B, since the silicon carbide oxide layer 135 on the surface contains silicon carbide oxide, the types of functional groups increase compared to crystalline silicon and amorphous silicon. It is excellent in reactivity with the above-mentioned cross-linking agents such as Therefore, according to MSS3, the binding substance can be more efficiently immobilized using the cross-linking agent.

Next, an embodiment of the present invention will be described. However, the present invention is not limited by the following examples. Commercially available reagents were used according to their protocol unless otherwise indicated.

[Example 1]
MSS3 of Embodiment 3 was manufactured, and it was confirmed that a silicon carbide layer and a silicon carbide oxide layer were formed on the MSS film.

(1) Configuration of MSS Using a commercially available MSS (SD-MSS-1K2G, NANOSENSOR), MSS3 of Embodiment 3 was created. The commercially available MSS has the same configuration as MSS1 in FIG.

(2) Formation of Insulating Layer An insulating layer 122 was formed by oxidizing the aluminum wire 121 of the MSS. Specifically, nitric acid (1.38 g/ml or more) was applied onto the aluminum wire 121 of the MSS. The amount of nitric acid applied was 50 μl per MSS. After the coating, the aluminum wire 121 was oxidized by reacting at room temperature (about 25° C., hereinafter the same) for 60 minutes. After the oxidation treatment, the MSS was rinsed under pure water at room temperature for 30 minutes to remove nitric acid.

Next, the MSS was subjected to oxygen plasma treatment. The oxygen plasma treatment was performed in a plasma generator under an oxygen atmosphere for 16 hours. The oxygen concentration was the default setting (100%) of the plasma generator. The treated MSS was heat-treated at 350° C. for 3 hours to anneal the formed oxide film and convert the aluminum hydroxide formed by the oxygen plasma treatment into aluminum oxide. As a result, the aluminum wire 12 of the MSS was converted into an aluminum wire 12A composed of a conductive layer 121 and an insulating layer 122. FIG.

(3) Formation of Silicon Carbide Layer and Silicon Carbide Oxide Layer Regarding the MSS of Example 1 (2), crystalline silicon is converted to amorphous silicon (amorphized silicon) to form an amorphous layer 132. bottom.

Next, carbon was introduced into the amorphous layer 132 . After the deposition, the MSS was heat treated to react the introduced carbon with the silicon in the amorphous layer 132 and convert it to silicon carbide. Then, the carbon layer 134 was removed from the obtained MSS, and the surface of the silicon carbide layer 133 was converted to an oxide layer 135 of silicon carbide.

(4) Structure of MSS film The MSS film 13B was recovered from the MSS3 obtained in Example 1(3), and its structure in the stacking direction was confirmed with a transmission electron microscope. The results are shown in FIG.

FIG. 9 is a photograph showing an electron microscope image. As shown in FIG. 9, the MSS film 13B consists of a support layer 131 made of crystalline silicon, a silicon carbide layer 133 made of amorphous silicon and silicon carbide, and a silicon carbide layer 133 made of amorphous silicon and silicon carbide. It was confirmed that an oxide layer 135 of silicon carbide composed of an oxide of silicon carbide was laminated. It was also found that when the MSS film was processed under the conditions described above, the thickness of the silicon carbide layer 133 was about 70 nm, and the thickness of the silicon carbide oxide layer 135 was about 5 nm. Amorphous silicon also has an improved surface area compared to crystalline silicon. Therefore, it can be said that MSS3 can more efficiently immobilize binding substances such as nucleic acid molecules and antibodies on the surface. Regarding the MSS3, when the region including the MSS film 13B was immersed in an aqueous solution and a voltage was applied, an electric signal could be measured. For this reason, it was confirmed that the aluminum wire 12A was able to secure insulation between circuits, ie, between the aluminum wires 12A, by forming the insulating layer 122 .

Although the present invention has been described with reference to the embodiments and examples, the present invention is not limited to the above embodiments and examples. Various changes that can be understood by those skilled in the art can be made to the configuration and details of the present invention within the scope of the present invention.

<Appendix>
Some or all of the above-described embodiments and examples can be described as in the following appendices, but are not limited to the following.
<First Membrane Surface Stress Sensor>
(Appendix 1)
comprising a membrane and a sensor substrate;
The membrane is a membrane that deforms in response to surface stress,
the sensor substrate comprises a support area and circuitry;
the support region supports the membrane and comprises a piezoresistive element;
The piezoresistive element is an element that detects deformation of the film,
The circuit is connected to the piezoresistive element,
The circuit includes a metal capable of forming an oxide film by oxidation treatment,
The circuit has an oxide film layer on the surface,
Membrane type surface stress sensor.
(Appendix 2)
2. The sensor of clause 1, wherein the membrane comprises an amorphous layer.
(Appendix 3)
3. The sensor of Clause 2, wherein the amorphous layer comprises amorphous silicon or amorphized silicon.
(Appendix 4)
4. The sensor of Clause 2 or 3, wherein the amorphous layer comprises silicon carbide.
(Appendix 5)
The amorphous layer is
a silicon carbide layer;
an oxide layer of silicon carbide;
the silicon carbide layer includes amorphous silicon or amorphized silicon and silicon carbide;
the oxide layer comprises amorphous silicon or amorphized silicon and silicon carbide and/or oxidized silicon carbide;
5. The sensor according to any one of appendices 2 to 4, wherein the silicon carbide layer and the oxide layer are laminated in this order.
(Appendix 6)
the membrane comprises a support layer;
the support layer comprises a crystalline component of the amorphous layer;
6. The sensor of any one of Clauses 2-5, wherein the amorphous layer is laminated to the support layer.
(Appendix 7)
the support layer comprises crystalline silicon;
wherein the amorphous layer comprises amorphous silicon or amorphized silicon;
A sensor according to Appendix 6.
(Appendix 8)
The amorphous layer is
a silicon carbide layer;
an oxide layer of silicon carbide;
the silicon carbide layer includes amorphous silicon or amorphized silicon and silicon carbide;
the oxide layer comprises amorphous silicon or amorphized silicon and silicon carbide and/or oxidized silicon carbide;
8. The sensor of Claim 7, wherein the support layer, the silicon carbide layer and the oxide layer are stacked in this order.
(Appendix 9)
9. The sensor of any one of the clauses 1-8, wherein the membrane is a silicon membrane.
(Appendix 10)
10. The sensor of any preceding clause, wherein the support region partially supports the membrane.
(Appendix 11)
The support region includes a plurality of piezoresistive elements,
11. The sensor according to any one of appendices 1 to 10, wherein the circuit constitutes a Wheatstone bridge circuit including the plurality of piezoresistive elements.
(Appendix 12)
the sensor substrate having a plurality of support areas;
12. The sensor of any one of Clauses 1 to 11, wherein the plurality of support regions each support the membrane.
(Appendix 13)
13. The sensor of any one of Clauses 1-12, wherein the metal is a valve metal.
(Appendix 14)
14. A sensor according to any preceding clause, wherein the metal is aluminium, tantalum, niobium, titanium, hafnium, zirconium, zinc, tungsten, bismuth, antimony, molybdenum, chromium, iron, or alloys thereof.
(Appendix 15)
15. The sensor of any one of Clauses 1-14, wherein the metal is aluminum.
(Appendix 16)
the membrane comprises a binding substance that binds to a target;
16. The sensor of any one of Clauses 1 to 15, wherein the membrane deforms upon binding of a target to the binding substance.
(Appendix 17)
17. The sensor of clause 16, wherein the binding substance is a nucleic acid molecule or a protein.
(Appendix 18)
18. The sensor of clause 16 or 17, wherein the binding substance is immobilized on one surface of the membrane.
(Appendix 19)
18. The sensor of clause 16 or 17, wherein the binding substance is immobilized on both sides of the membrane.
(Appendix 20)
20. The sensor according to any one of Appendices 16 to 19, wherein the binding substance is immobilized on the membrane via a conjugate of avidin or an avidin derivative and biotin or a biotin derivative.
(Appendix 21)
The membrane comprises a metal film on the surface,
21. The sensor according to any one of appendices 16 to 20, wherein the binding substance is immobilized on the surface of the film via the metal film.
(Appendix 22)
22. The sensor according to any one of appendices 16 to 21, wherein the binding substance is immobilized on the membrane surface via a linker.
(Appendix 23)
23. The sensor of clause 22, wherein the length of the linker is substantially constant.
(Appendix 24)
24. The sensor of clause 22 or 23, wherein the linker comprises a silane coupling agent.
(Appendix 25)
25. A sensor according to any one of the clauses 1 to 24, wherein the circuit is not covered with resin.
(Appendix 26)
26. A sensor according to any of clauses 1 to 25 for use in analyzing liquid samples.
<Second Membrane Surface Stress Sensor>
(Appendix 27)
comprising a membrane and a sensor substrate;
The membrane is a membrane that deforms in response to surface stress,
the sensor substrate comprises a support area;
the support region supports the membrane and comprises a piezoresistive element;
The piezoresistive element is an element that detects deformation of the film,
the film comprises an amorphous layer;
Membrane type surface stress sensor.
(Appendix 28)
28. The sensor of Clause 27, wherein the amorphous layer comprises amorphous silicon or amorphized silicon.
(Appendix 29)
29. The sensor of clause 27 or 28, wherein the amorphous layer comprises silicon carbide.
(Appendix 30)
The amorphous layer is
a silicon carbide layer;
an oxide layer of silicon carbide;
the silicon carbide layer includes amorphous silicon or amorphized silicon and silicon carbide;
the oxide layer comprises amorphous silicon or amorphized silicon and silicon carbide and/or oxidized silicon carbide;
30. The sensor according to any one of appendices 27 to 29, wherein the silicon carbide layer and the oxide layer are laminated in this order.
(Appendix 31)
the membrane comprises a support layer;
the support layer comprises a crystalline component of the amorphous layer;
31. The sensor of any of Clauses 27-30, wherein the amorphous layer is laminated to the support layer.
(Appendix 32)
the support layer comprises crystalline silicon;
wherein the amorphous layer comprises amorphous silicon or amorphized silicon;
32. The sensor of clause 31.
(Appendix 33)
The amorphous layer is
a silicon carbide layer;
an oxide layer of silicon carbide;
the silicon carbide layer includes amorphous silicon or amorphized silicon and silicon carbide;
the oxide layer comprises amorphous silicon or amorphized silicon and silicon carbide and/or oxidized silicon carbide;
33. The sensor of clause 32, wherein the support layer, the silicon carbide layer and the oxide layer are stacked in this order.
(Appendix 34)
34. The sensor of any of clauses 27-33, wherein the membrane is a silicon membrane.
(Appendix 35)
35. The sensor of any of clauses 27-34, wherein the support region partially supports the membrane.
(Appendix 36)
The sensor substrate has a circuit,
The support region includes a plurality of piezoresistive elements,
36. The sensor according to any one of appendices 27 to 35, wherein said circuit constitutes a Wheatstone bridge circuit including said plurality of piezoresistive elements.
(Appendix 37)
the sensor substrate having a plurality of support areas;
37. The sensor of any of Clauses 27-36, wherein each of the plurality of support regions supports the membrane.
(Appendix 38)
the membrane comprises a binding substance that binds to a target;
38. The sensor of any of clauses 27-37, wherein the membrane is deformed upon binding of a target to the binding substance.
(Appendix 39)
39. The sensor of clause 38, wherein said binding agent is a nucleic acid molecule or a protein.
(Appendix 40)
40. The sensor of clause 38 or 39, wherein the binding substance is immobilized on one surface of the membrane.
(Appendix 41)
40. The sensor of clause 38 or 39, wherein the binding substance is immobilized on both sides of the membrane.
(Appendix 42)
42. The sensor according to any one of Appendices 38 to 41, wherein the binding substance is immobilized on the membrane via a conjugate of avidin or an avidin derivative and biotin or a biotin derivative.
(Appendix 43)
The membrane comprises a metal film on the surface,
43. The sensor according to any one of Appendices 38 to 42, wherein the binding substance is immobilized on the surface of the membrane via the metal membrane.
(Appendix 44)
44. The sensor according to any one of Appendices 38 to 43, wherein the binding substance is immobilized on the membrane surface via a linker.
(Appendix 45)
Clause 45. The sensor of Clause 44, wherein the linker length is substantially constant.
(Appendix 46)
46. The sensor of clause 44 or 45, wherein the linker comprises a silane coupling agent.
(Appendix 47)
47. A sensor according to any of clauses 27 to 46 for use in analyzing liquid samples.
<Manufacturing Method of First Membrane Surface Stress Sensor>
(Appendix 48)
A film-type stress sensor comprising a film and a circuit, the film-type surface comprising a step of subjecting a circuit containing a metal capable of forming an oxide film by oxidation treatment to an oxidation treatment to form an oxide film layer on the circuit surface. A method of manufacturing a stress sensor.
(Appendix 49)
49. The manufacturing method according to appendix 48, wherein the oxidation treatment is performed by acid treatment and/or plasma treatment in an oxygen atmosphere.
(Appendix 50)
49. The manufacturing method according to appendix 48 or 49, comprising the step of subjecting the circuit to acid treatment to form an oxide film layer on the surface of the circuit.
(Appendix 51)
51. The production method according to appendix 49 or 50, wherein the acid treatment is nitric acid treatment, sulfuric acid treatment, phosphoric acid treatment, oxalic acid treatment, and/or chromic acid treatment.
(Appendix 52)
52. The manufacturing method according to any one of appendices 48 to 51, wherein a heat treatment is performed after forming the oxide film.
(Appendix 53)
53. The manufacturing method according to Appendix 52, wherein the heat treatment is performed at 100° C. or higher.
(Appendix 54)
54. The method according to any one of appendices 48 to 53, including a step of subjecting the film of the film-type surface stress sensor to an amorphization treatment (amorphization treatment) to form an amorphous layer on the film surface. Production method.
(Appendix 55)
55. The manufacturing method according to appendix 54, comprising the step of introducing carbon into the amorphous layer.
(Appendix 56)
56. The manufacturing method according to appendix 55, comprising removing carbon from the film by plasma treatment and/or sputtering treatment in an oxygen atmosphere.
(Appendix 57)
57. The manufacturing method according to any one of appendices 48 to 56, wherein the film is a silicon film.
(Appendix 58)
58. The manufacturing method according to any one of appendices 48 to 57, wherein the metal is a valve metal.
(Appendix 59)
59. The manufacturing method according to any one of Appendices 48 to 58, wherein the metal is aluminum, tantalum, niobium, titanium, hafnium, zirconium, zinc, tungsten, bismuth, antimony, molybdenum, chromium, iron, or alloys thereof.
(Appendix 60)
60. The manufacturing method according to any one of appendices 48 to 59, wherein the metal is aluminum.
<Method for Manufacturing Second Membrane Surface Stress Sensor>
(Appendix 61)
1. A method of manufacturing a film-type surface stress sensor comprising a film and a circuit, comprising the step of subjecting said film to amorphization treatment to form an amorphous layer on said film surface.
(Appendix 62)
62. The manufacturing method according to appendix 61, wherein the amorphization treatment is performed by an amorphization treatment.
(Appendix 63)
63. The manufacturing method according to appendix 61 or 62, including the step of introducing carbon into the amorphous layer.
(Appendix 64)
64. The manufacturing method according to appendix 63, comprising removing carbon from the film by plasma treatment and/or sputtering treatment in an oxygen atmosphere.
(Appendix 65)
65. The manufacturing method according to any one of appendices 61 to 64, wherein the film is a silicon film.
<Method for manufacturing a semiconductor device>
(Appendix 66)
In a semiconductor device comprising a film and a circuit, a step of oxidizing a circuit containing a metal capable of forming an oxide film by oxidation treatment to form an oxide film layer on the surface of the circuit;
A method of manufacturing a semiconductor device, comprising: subjecting the film to an amorphization treatment to form an amorphous layer on the surface of the film.
(Appendix 67)
67. The manufacturing method according to appendix 66, wherein the oxidation treatment is performed by acid treatment and/or plasma treatment in an oxygen atmosphere.
(Appendix 68)
a step of acid-treating the circuit to form an oxide film layer on the surface of the circuit;
68. The manufacturing method according to appendix 66 or 67, comprising performing a plasma treatment in an oxygen atmosphere on the circuit.
(Appendix 69)
69. The production method according to appendix 67 or 68, wherein the acid treatment is nitric acid treatment, sulfuric acid treatment, nitric acid treatment, sulfuric acid treatment, phosphoric acid treatment, oxalic acid treatment, and/or chromic acid treatment.
(Appendix 70)
69. The manufacturing method according to any one of appendices 66 to 69, wherein a heat treatment is performed after forming the oxide film.
(Appendix 71)
71. The manufacturing method according to appendix 70, wherein the heat treatment is performed at 100° C. or higher.
(Appendix 72)
72. The manufacturing method according to any one of appendices 66 to 71, wherein the amorphization treatment is performed by ion beam treatment.
(Appendix 73)
73. The manufacturing method according to any one of Appendixes 66 to 72, comprising introducing carbon into the amorphous layer.
(Appendix 74)
74. The manufacturing method according to appendix 73, comprising removing carbon from the film by plasma treatment and/or sputtering treatment in an oxygen atmosphere.
(Appendix 75)
75. The manufacturing method according to any one of appendices 66 to 74, wherein the film is a silicon film.
(Appendix 76)
76. The manufacturing method according to any one of appendices 66 to 75, wherein the metal is a valve metal.
(Appendix 77)
77. The manufacturing method according to any of appendices 66 to 76, wherein the metal is aluminum, tantalum, niobium, titanium, hafnium, zirconium, zinc, tungsten, bismuth, antimony, molybdenum, chromium, iron, or alloys thereof.
(Appendix 78)
78. The manufacturing method according to any one of appendices 66 to 77, wherein the metal is aluminum.
<Target analysis method>
(Appendix 79)
an applying step of applying a voltage to the membrane surface stress sensor in the sample liquid;
an analysis step of analyzing the target in the sample liquid by measuring the stress change of the piezoresistive element in the film-type surface stress sensor;
48. A method of analyzing a target, wherein the film-type surface stress sensor is the film-type surface stress sensor according to any one of Appendices 1 to 47.

According to the first MSS of the present invention, it is possible to insulate between circuits. In addition, according to the second MSS of the present invention, it is possible to improve the immobilization efficiency of the binding substance on the MSS membrane. Therefore, the present invention is useful in, for example, the analysis field of samples and the like, the medical field, and the like.

1, 2, 3 MSS
10 Sensor substrate 11

Electrodes

12, 12A Aluminum wire 121 Conductive layer 122 Insulating layer (oxide layer)
13, 13A MSS film 131 support layer 132 amorphous layer 14 piezoresistive element 15 support region

Claims

comprising a membrane and a sensor substrate;
The membrane is a membrane that deforms in response to surface stress,
the sensor substrate comprises a support area and circuitry;
the support region supports the membrane and comprises a piezoresistive element;
The piezoresistive element is an element that detects deformation of the film,
The circuit is connected to the piezoresistive element,
The circuit includes a metal capable of forming an oxide film by oxidation treatment,
The circuit has an oxide film layer on the surface,
Membrane type surface stress sensor.
2. The sensor of Claim 1, wherein the membrane comprises an amorphous layer.
3. The sensor of claim 2, wherein the amorphous layer comprises amorphous silicon or amorphized silicon.
4. The sensor of claim 2 or 3, wherein the amorphous layer comprises silicon carbide.
The amorphous layer is
a silicon carbide layer;
an oxide layer of silicon carbide;
the silicon carbide layer includes amorphous silicon or amorphized silicon and silicon carbide;
the oxide layer comprises amorphous silicon or amorphized silicon and silicon carbide and/or oxidized silicon carbide;
5. The sensor according to any one of claims 2 to 4, wherein said silicon carbide layer and said oxide layer are laminated in this order.
the membrane comprises a support layer;
the support layer comprises a crystalline component of the amorphous layer;
6. The sensor of any one of claims 2-5, wherein the amorphous layer is laminated to the support layer.
the support layer comprises crystalline silicon;
7. The sensor of claim 6, wherein the amorphous layer comprises amorphous silicon or amorphized silicon.
The amorphous layer is
a silicon carbide layer;
an oxide layer of silicon carbide;
the silicon carbide layer includes amorphous silicon or amorphized silicon and silicon carbide;
the oxide layer comprises amorphous silicon or amorphized silicon and silicon carbide and/or oxidized silicon carbide;
8. The sensor of claim 7, wherein the support layer, the silicon carbide layer and the oxide layer are laminated in this order.
9. A sensor according to any preceding claim, wherein the membrane is a silicon membrane.
10. The sensor of any one of claims 1-9, wherein the support area partially supports the membrane.
The support region includes a plurality of piezoresistive elements,
11. The sensor according to any one of claims 1 to 10, wherein said circuit constitutes a Wheatstone bridge circuit including said plurality of piezoresistive elements.
the sensor substrate having a plurality of support areas;
12. The sensor of any one of claims 1-11, wherein the plurality of support areas each support the membrane.
13. A sensor according to any preceding claim, wherein the metal is a valve metal.
14. Any one of claims 1 to 13, wherein the metal is aluminium, tantalum, niobium, titanium, hafnium, zirconium, zinc, tungsten, bismuth, antimony, molybdenum, chromium, iron, or alloys thereof. sensor.
15. A sensor according to any preceding claim, wherein the metal is aluminum.
the membrane comprises a binding substance that binds to a target;
16. The sensor of any one of claims 1-15, wherein the membrane is deformed upon binding of a target to the binding substance.
17. The sensor of claim 16, wherein said binding agent is a nucleic acid molecule or protein.
18. A sensor according to claim 16 or 17, wherein said binding substance is immobilized on one surface of said membrane.
18. A sensor according to claim 16 or 17, wherein said binding substance is immobilized on both sides of said membrane.
20. The sensor of any one of claims 1-19, wherein the circuitry is not coated with resin.
21. A sensor according to any preceding claim for use in analyzing liquid samples.
comprising a membrane and a sensor substrate;
The membrane is a membrane that deforms in response to surface stress,
the sensor substrate comprises a support area;
the support region supports the membrane and comprises a piezoresistive element;
The piezoresistive element is an element that detects deformation of the film,
the film comprises an amorphous layer;
Membrane type surface stress sensor.
23. The sensor of claim 22, wherein the amorphous layer comprises amorphous silicon or amorphized silicon.
24. A sensor according to claim 22 or 23, wherein the amorphous layer comprises silicon carbide.
The amorphous layer is
a silicon carbide layer;
an oxide layer of silicon carbide;
the silicon carbide layer includes amorphous silicon or amorphized silicon and silicon carbide;
the oxide layer comprises amorphous silicon or amorphized silicon and silicon carbide and/or oxidized silicon carbide;
25. The sensor of any one of claims 22-24, wherein the silicon carbide layer and the oxide layer are stacked in that order.
the membrane comprises a support layer;
the support layer comprises a crystalline component of the amorphous layer;
26. The sensor of any one of claims 22-25, wherein the amorphous layer is laminated to the support layer.
the support layer comprises crystalline silicon;
27. The sensor of Claim 26, wherein the amorphous layer comprises amorphous silicon or amorphized silicon.
The amorphous layer is
a silicon carbide layer;
an oxide layer of silicon carbide;
the silicon carbide layer includes amorphous silicon or amorphized silicon and silicon carbide;
the oxide layer comprises amorphous silicon or amorphized silicon and silicon carbide and/or oxidized silicon carbide;
28. The sensor of claim 27, wherein the support layer, the silicon carbide layer and the oxide layer are stacked in that order.
29. The sensor of any one of claims 22-28, wherein the membrane is a silicon membrane.
30. The sensor of any one of claims 22-29, wherein the support area partially supports the membrane.
The sensor substrate has a circuit,
The support region includes a plurality of piezoresistive elements,
31. A sensor according to any one of claims 22 to 30, wherein said circuit constitutes a Wheatstone bridge circuit comprising said plurality of piezoresistive elements.
the sensor substrate having a plurality of support areas;
32. The sensor of any one of claims 22-31, wherein said plurality of support areas each support said membrane.
the membrane comprises a binding substance that binds to a target;
33. The sensor of any one of claims 22-32, wherein the membrane deforms upon binding of a target to the binding substance.
34. The sensor of claim 33, wherein said binding agent is a nucleic acid molecule or protein.
35. A sensor according to claim 33 or 34, wherein said binding substance is immobilized on one surface of said membrane.
35. A sensor according to claim 33 or 34, wherein the binding substance is immobilized on both sides of the membrane.
37. A sensor according to any one of claims 22 to 36 for use in analyzing liquid samples.
A film-type stress sensor comprising a film and a circuit, the film-type surface comprising a step of subjecting a circuit containing a metal capable of forming an oxide film by oxidation treatment to an oxidation treatment to form an oxide film layer on the circuit surface. A method of manufacturing a stress sensor.
39. The manufacturing method according to claim 38, wherein said oxidation treatment is performed by acid treatment and/or plasma treatment in an oxygen atmosphere.
40. The manufacturing method according to claim 38 or 39, comprising the step of acid-treating the circuit to form an oxide film layer on the surface of the circuit.
41. The manufacturing method according to claim 39 or 40, wherein said acid treatment is nitric acid treatment, sulfuric acid treatment, phosphoric acid treatment, oxalic acid treatment and/or chromic acid treatment.
42. The manufacturing method according to any one of claims 38 to 41, wherein a heat treatment is performed after forming the oxide film.
The manufacturing method according to claim 42, wherein the heat treatment is performed at 100°C or higher.
44. The manufacturing method according to any one of claims 38 to 43, comprising the step of subjecting said film to amorphization treatment (amorphization treatment) to form an amorphous layer on said film surface.
45. The manufacturing method according to any one of claims 38 to 44, wherein said film is a silicon film.
46. The manufacturing method of any one of claims 38-45, wherein the metal is a valve metal.
47. Any one of claims 38-46, wherein the metal is aluminum, tantalum, niobium, titanium, hafnium, zirconium, zinc, tungsten, bismuth, antimony, molybdenum, chromium, iron, or alloys thereof. Production method.
48. The manufacturing method according to any one of claims 38 to 47, wherein said metal is aluminum.
1. A method of manufacturing a film-type surface stress sensor comprising a film and a circuit, comprising the step of subjecting said film to amorphization treatment to form an amorphous layer on said film surface.
50. The manufacturing method according to claim 49, wherein said amorphization treatment is performed by an amorphization treatment.
51. The manufacturing method according to claim 49 or 50, wherein said film is a silicon film.
In a semiconductor device comprising a film and a circuit, a step of oxidizing a circuit containing a metal capable of forming an oxide film by oxidation treatment to form an oxide film layer on the surface of the circuit;
A method of manufacturing a semiconductor device, comprising: subjecting the film to an amorphization treatment to form an amorphous layer on the surface of the film.
53. The manufacturing method according to claim 52, wherein said oxidation treatment is performed by acid treatment and/or plasma treatment in an oxygen atmosphere.
54. The manufacturing method according to claim 52 or 53, comprising the step of subjecting said circuit to acid treatment to form an oxide film layer on said circuit surface.
55. The manufacturing method according to claim 53 or 54, wherein said acid treatment is nitric acid treatment, sulfuric acid treatment, nitric acid treatment, sulfuric acid treatment, phosphoric acid treatment, oxalic acid treatment and/or chromic acid treatment.
56. The manufacturing method according to any one of claims 52 to 55, wherein a heat treatment is performed after forming the oxide film.
57. The manufacturing method according to claim 56, wherein the heat treatment is performed at 100[deg.]C or higher.
58. The manufacturing method according to any one of claims 52 to 57, wherein said amorphization treatment is performed by ion beam treatment.
59. The manufacturing method according to any one of claims 52 to 58, comprising introducing carbon into said amorphous layer.
60. The manufacturing method according to claim 59, comprising removing carbon from said film by plasma treatment and/or sputtering treatment in an oxygen atmosphere.
61. The manufacturing method according to any one of claims 52 to 60, wherein said film is a silicon film.
62. The manufacturing method of any one of claims 52-61, wherein the metal is a valve metal.
63. Any one of claims 52-62, wherein the metal is aluminum, tantalum, niobium, titanium, hafnium, zirconium, zinc, tungsten, bismuth, antimony, molybdenum, chromium, iron, or alloys thereof. Production method.
64. The manufacturing method of any one of claims 52-63, wherein the metal is aluminum.
an applying step of applying a voltage to the membrane surface stress sensor in the sample liquid;
an analysis step of analyzing the target in the sample liquid by measuring the stress change of the piezoresistive element in the film-type surface stress sensor;
A method for analyzing a target, wherein the membrane-type surface stress sensor is the membrane-type surface stress sensor according to any one of claims 1 to 37.