WO2017169717A1 - Inspection device, inspection apparatus, and inspection method - Google Patents

Inspection device, inspection apparatus, and inspection method Download PDF

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Publication number
WO2017169717A1
WO2017169717A1 PCT/JP2017/010100 JP2017010100W WO2017169717A1 WO 2017169717 A1 WO2017169717 A1 WO 2017169717A1 JP 2017010100 W JP2017010100 W JP 2017010100W WO 2017169717 A1 WO2017169717 A1 WO 2017169717A1
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Prior art keywords
inspection device
substance
inspection
pores
solution
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PCT/JP2017/010100
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French (fr)
Japanese (ja)
Inventor
弘隆 渡野
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富士フイルム株式会社
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Publication of WO2017169717A1 publication Critical patent/WO2017169717A1/en

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    • GPHYSICS
    • G01MEASURING; TESTING
    • G01NINVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N21/00Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
    • G01N21/62Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
    • G01N21/63Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
    • G01N21/64Fluorescence; Phosphorescence
    • GPHYSICS
    • G01MEASURING; TESTING
    • G01NINVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N21/00Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
    • G01N21/75Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated
    • G01N21/77Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated by observing the effect on a chemical indicator
    • G01N21/78Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated by observing the effect on a chemical indicator producing a change of colour
    • GPHYSICS
    • G01MEASURING; TESTING
    • G01NINVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/53Immunoassay; Biospecific binding assay; Materials therefor
    • GPHYSICS
    • G01MEASURING; TESTING
    • G01NINVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/53Immunoassay; Biospecific binding assay; Materials therefor
    • G01N33/543Immunoassay; Biospecific binding assay; Materials therefor with an insoluble carrier for immobilising immunochemicals
    • G01N33/551Immunoassay; Biospecific binding assay; Materials therefor with an insoluble carrier for immobilising immunochemicals the carrier being inorganic
    • G01N33/553Metal or metal coated

Definitions

  • the present invention relates to a test device for detecting an antigen, antibody, deoxyribonucleic acid, or the like, which is a test substance, a test apparatus equipped with the test device, and a test method.
  • the binding phenomenon between a test substance and a capture substance is optically determined.
  • a method of detecting is known. In this method, a test substance is bound to a capture substance fixed at a predetermined position and emits fluorescence upon receiving excitation light, or a label that catalyzes a substrate reaction to generate color, fluorescence, or chemiluminescence. Is applied to the test substance, and light generated due to the label is detected.
  • a method of detecting a fluorescence generated from a fluorescent label by attaching a fluorescent label to a binding substance such as an antibody that specifically binds to the test substance, a binding of an antibody that specifically binds to the test substance
  • a binding substance such as an antibody that specifically binds to the test substance
  • a binding of an antibody that specifically binds to the test substance There are known methods for detecting color development, fluorescence, chemiluminescence, etc. generated from a chromogenic substrate, fluorescent substrate, or chemiluminescent substrate that reacts with this enzyme as a catalyst. Identification becomes possible.
  • a multi-item simultaneous detection chip in which biologically related molecules are regularly arranged on a two-dimensional substrate has been conventionally used.
  • a device made of a porous substrate in which a large number of through-holes (pores) are arranged on a support has been studied.
  • a porous silicon (Si) base material As a porous substrate, a porous silicon (Si) base material is known, but silicon has low reflectivity, and the light generated in the pores is reflected multiple times in the pores. Since the signal intensity is greatly attenuated, there is a problem that the light extraction efficiency of the optical signal generated in the pore is very low.
  • Patent Document 1 discloses a porous substrate having a large number of through holes in an aluminum (Al) base material. Since aluminum has a higher reflectance than silicon, it is considered that the light extraction efficiency can be improved as compared with the case where a silicon substrate is used. However, it cannot be said that the light extraction efficiency can be sufficiently improved because the light exit openings of the individual pores are small. On the other hand, if the pore diameter is increased in order to widen the light extraction opening, the pore density is lowered, and the amount of trapped substance that can be fixed as a whole is reduced.
  • Patent Document 2 as a means for solving the above problem, porous silicon is thermally oxidized to be locally transparent silicon oxide (SiO 2 ), and the oxidized region is surrounded by a frame (wall) made of silicon.
  • a configuration has been proposed in which the light output aperture is substantially widened to improve the light extraction efficiency.
  • JP 2004-93152 A Japanese Patent No. 4125244
  • Patent Document 2 Although the light extraction efficiency can be improved as the optical aperture ratio increases, the reflectance of silicon used as a frame wall is low, so that light incident on the silicon wall is also attenuated. Therefore, the extraction efficiency of light incident on the frame wall is not improved.
  • it is considered necessary to increase the thickness of the chip, expand the detection area, or increase the density of the pores.
  • the aspect ratio with the opening diameter As the aspect ratio of the pores increases, the aspect ratio of the region surrounded by the silicon frame also increases accordingly, so the light exit aperture of the region surrounded by the silicon frame becomes substantially narrow, The influence of optical signal attenuation due to multiple reflection is increased.
  • the inspection device of the present invention is an inspection device having a plurality of pores penetrating from one surface of the plate-like substrate to the other surface, A region including at least a part of the pores, wherein at least a part of the wall member constituting the inner wall surface of the at least a part of the pores is made of alumina, and when viewed from the one side, A frame wall made of aluminum surrounding the region,
  • a capture substance that specifically binds to a specific substance is fixed to an inner wall surface of the at least some pores.
  • the wall member has a fine hole extending in parallel with the pore and having a diameter smaller than the pore.
  • the wall member is preferably made of anodized alumina.
  • the frame wall is preferably provided in contact with the wall member.
  • the other surface is preferably made of aluminum.
  • the capture substance is preferably an antigen, an antibody, or deoxyribonucleic acid (DNA).
  • the inspection device of the present invention comprises a plurality of the above-mentioned regions surrounded by a frame wall made of aluminum, and the same type of capture material is bound as a capture material to the inner wall surface of the pore in one of the plurality of regions. It is preferable that the plurality of regions have regions in which the bound capture substances are different from each other.
  • the equivalent circle diameter of the open region on one surface of the pore is preferably 1 ⁇ m to 100 ⁇ m.
  • the thickness of the plate-like substrate is preferably 100 ⁇ m to 2000 ⁇ m.
  • the inspection apparatus of the present invention includes the inspection device of the present invention, A solution supply unit for supplying an inspection solution into the pores of the inspection device;
  • the inspection apparatus includes a photodetector that is disposed on one surface or the other surface of the inspection device and detects fluorescence emitted from the inspection device.
  • a specimen liquid containing a specific substance is supplied to the at least some of the pores of the test device of the present invention to bind the specific substance to the capture substance
  • a labeling substance that specifically binds to a specific substance is bound to the specific substance
  • the inspection method detects light emitted from the inspection device in a state where the inspection solution is supplied to the pores and the inspection solution is retained in the pores.
  • a substance containing an enzyme label is used as the label
  • the light emitted from the inspection device is as follows: You may detect the light which a substrate in a reaction liquid produces when catalyzed by an enzyme label
  • the substrate include a chromogenic substrate, a fluorescent substrate, and a chemiluminescent substrate, and these substrates are appropriately selected depending on the type of enzyme label.
  • the light emitted from the inspection device differs depending on the substrate, and the detected light is light absorption (coloration), fluorescence, or chemiluminescence.
  • a substance containing a fluorescent label is used as a labeling substance, Irradiate the test device with excitation light that excites the fluorescent label, As the light emitted from the inspection device, fluorescence generated from the labeling substance by irradiation with excitation light may be detected.
  • the inspection device of the present invention is an inspection device having a plurality of pores penetrating from one surface of the plate-like substrate to the other surface, and is an area including at least some of the pores, Since at least a part of the wall member constituting the inner wall surface of some of the pores has a region made of alumina and a frame wall made of aluminum surrounding the region when viewed from the one surface, The extraction efficiency of the light generated in the pores to at least one surface side of the inspection device can be improved.
  • FIG. 1 is a perspective view of an inspection device according to a first embodiment of the present invention
  • FIG. 2 is a plan view of the inspection device of FIG. 1
  • FIG. 3 is a sectional view taken along line III-III of the inspection device of FIG. 4 is a bottom view of the inspection device of FIG.
  • the inspection device 1 of the present embodiment is a wall made of alumina that constitutes an inner wall surface 14a of a pore in a plate-like substrate 10 having a plurality of pores 14 penetrating from one surface 11 to the other surface 12. It has the area
  • FIG. In the present embodiment a total of nine pores 14 in the vertical and horizontal directions are used as one region 20, and a frame wall 18 is provided so as to surround each region 20.
  • a trapping substance 30 that specifically binds to a specific substance is fixed to the inner wall surface 14 a of the pore 14.
  • the plan view shown in FIG. 2 is a view of the inspection device 1 as viewed from one surface 11, and the bottom view of FIG. 4 is a view of the inspection device 1 as viewed from the other surface 12.
  • the external shape of the inspection device 1 is configured by the external shape of the plate-like base material 10
  • one surface 11 and the other surface 12 of the plate-like base material 10 are hereinafter referred to as one of the inspection devices 1.
  • the surface and the other surface are hereinafter referred to as the surface and the other surface.
  • the wall member constituting the inner wall surface 14a of the four regions 20 is made of alumina except for a part on the other surface 12 side.
  • a frame wall 18 made of aluminum is provided surrounding the region 20 and in contact with the wall member 16 made of alumina.
  • the lower surface member 19 which comprises the other surface 12 of the plate-shaped base material 10 consists of aluminum.
  • the lower surface member 19 is continuous with the frame wall 18 and is integrally formed.
  • the plate-like substrate 10 is composed of aluminum and alumina over the entire region.
  • the lower surface member 19 made of aluminum, a component of the light generated in the pores 14 that is emitted to the other surface 12 side and incident on the lower surface member 19 is reflected to the one surface 11 side. Therefore, the light extraction efficiency to the one surface 11 side can be improved.
  • the inspection device may not have the lower surface member 19.
  • the same result can be obtained even if the light detection is performed from either one of the surfaces 11 side and the other surface 12 side.
  • the wall member constituting the inner wall surface 14a of the region 20 including at least a part of the pores 14 is made of alumina. That is, in the region 20 surrounded by the frame wall 18, the wall member may include a portion formed of alumina and a portion formed of another material (for example, aluminum).
  • the frame wall 18 is formed so as to surround the outer periphery of the region 20, but is partially cut off if the length of the unconnected portion is about 20% of the length of the outer periphery of the region 20. It may be.
  • the plate-like base material 10 is preferably one in which at least a part of an aluminum base material is oxidized to form a wall member 16 made of alumina. According to such a configuration, an inexpensive aluminum base material can be used, and since it can be easily manufactured, it is preferable because it can be manufactured at low cost.
  • the plate-like substrate 10 may include a portion made of a different material other than aluminum and alumina.
  • the thickness of the plate-like substrate 10 is not particularly limited, but is preferably about 100 ⁇ m to 2000 ⁇ m.
  • the pores 14 provided in the plate-like substrate 10 are preferably arranged in an aligned manner as shown in the present embodiment, but may be arranged randomly.
  • the region 20 surrounded by the frame wall 18 (hereinafter referred to as “inspection region 20”) may include only one pore 14, but a plurality of pores 14 are included. It is preferable to include.
  • nine pores of 3 rows ⁇ 3 columns are provided in one inspection region 20 for easy visual recognition, but, for example, 100 10 rows ⁇ 10 columns in one inspection region.
  • the inspection area 20 provided in one inspection device 1 may be singular or plural. Although four inspection areas 20 are shown in the present embodiment, for example, ten or more inspection areas can be provided.
  • the opening and cross-sectional shape of the pore 14 are square in this embodiment, but are not limited to a rectangle such as a square or a rectangle, but may be a circle, an ellipse, a triangle, or a polygon that is a pentagon or more. Note that the corners of the polygon may be rounded due to manufacturing reasons.
  • the pores 14 are generally columnar and the cross-sectional shape does not change, but the cross-sectional shape may partially change or the cross-sectional size may change.
  • the equivalent circle diameter of the opening in at least one surface 12 of the pore 14 is preferably about 1 ⁇ m to 100 ⁇ m. More preferably, it is 3 ⁇ m to 50 ⁇ m, and particularly preferably 5 ⁇ m to 30 ⁇ m.
  • the equivalent circle diameter refers to the diameter of a circle having an area equivalent to the area of the opening region.
  • the test substance (target molecule) to be tested in this testing device 1 is mainly a biological molecule, such as proteins such as antigens and antibodies, saccharides, peptides, DNA, ribonucleic acid (RNA), peptides Examples thereof include nucleic acids (peptide nucleic acid: PNA).
  • the capture substance 30 that is fixed to the inner wall surface 14a of the pore 14 and specifically binds to a specific substance is a substance that specifically binds to these test substances.
  • the test device 1 is suitable for an allergy test provided with an allergen that is a kind of antigen as a capture substance.
  • the present inspection device 1 has a plurality of inspection regions 20 partitioned by a frame wall 18.
  • a trapping substance of the same type is bound (fixed) to the inner wall of the pore 14 in one inspection region 20.
  • Different capture substances can be fixed between the examination regions 20. Accordingly, it is possible to simultaneously inspect a plurality of test substances with one inspection device 1.
  • the plurality of inspection regions there may be two or more regions to which the same type of capture substance is bound.
  • inspection regions 20 in which different capture substances are fixed may be alternately arranged, and inspection regions in which the same type of capture material is periodically fixed may be disposed. According to such a configuration, it is possible to suppress variation and obtain a highly reliable test result.
  • a plurality of allergen test regions 20 are provided by fixing the same type of allergen as a capture substance in one or a plurality of test regions 20 in one test device. It is preferable that the response to can be simultaneously examined.
  • FIG. 6 is an enlarged perspective view showing a part of the inspection device 1.
  • the wall member 16 made of alumina preferably has fine holes 17 extending in parallel with the pores 14 and having a diameter smaller than the pores 14.
  • Such fine holes 17 can be formed by anodizing aluminum to transform it into alumina.
  • porous alumina having a large number of aligned fine holes 17 can be easily obtained.
  • the diameter of the fine hole formed by anodic oxidation is about several nm to several tens of nm, and preferably 5 nm to 20 nm.
  • the fine hole 17 is preferably not a through-hole penetrating from one surface 11 to the other surface 12 but a non-through hole with at least the other surface 12 closed.
  • a non-through hole can be formed by providing the lower surface member 19 described above.
  • a barrier layer (dense alumina layer) formed between the base material base and the anodizing treatment may be provided.
  • the one surface 11 side may remain open, it is also preferable that a sealing process for closing the opening of the one surface 11 is performed.
  • the fine hole 17 is made into the closed cavity part 17a.
  • the hollow portion 17a formed in this way can also be regarded as a kind of fine hole. According to such a configuration, the infiltration of the solution from any one of the one surface 11 and the other surface 12 can be suppressed, and the cavity can be used as an air layer, contributing to the improvement of light extraction efficiency described later. Becomes easy.
  • FIG. 7 is a schematic view showing a part of the inspection device 1 in an enlarged manner.
  • the present inspection device 1 binds a test substance to a capture substance (not shown in FIG. 7) fixed to the inner wall surface 14a of the pore 14, and specifically detects the test substance.
  • a binding substance labeled with a binding enzyme By binding a binding substance labeled with a binding enzyme and detecting a light signal generated by a luminescence reaction in which this enzyme acts as a catalyst, the presence or absence of the test substance or the amount of the test substance is detected. It is used in an inspection method for detecting the presence or absence of a test substance and the amount of a test substance.
  • the detection of the optical signal from the present inspection device 1 is performed in a state where the pore 14 is filled with an inspection solution 61 such as a buffer solution.
  • the wall member 16 constituting the inner wall surface 14a of the pore 14 in the inspection region 20 is made of alumina and has light transmittance.
  • a part of light generated in one pore 14 is repeatedly reflected in one pore and emitted from the surface.
  • the wall member 16 has optical transparency, a part of the light generated in one pore 14 can be guided to the surface side through the adjacent pore. As a result, the aperture ratio of light emitted to the surface of the light generated in the pores 14 increases.
  • the frame wall 18 made of aluminum is provided so as to surround the inspection region 20, a part of the light generated in the pores 14 in the inspection region 20 is reflected by the interface C 0 with the frame wall 18 and is a plate. The light is guided to the surface side of the substrate. Since aluminum has a higher reflectance than silicon and is less attenuated by reflection, more light components can be extracted to the device surface.
  • the lower surface member 19 constituting the other surface 12 is made of aluminum as in the inspection device 1 of the present embodiment, the light that is generated in the pore 14 and directly or indirectly incident on the lower surface member 19. Can be reflected on the one surface 11 side. In this case, the light emitted to the one surface 11 side can be further increased, and the extraction efficiency can be further improved.
  • FIG. 8 is an enlarged view of a region VIII indicated by a broken line in FIG.
  • the fine holes 17 provided in the wall member 16 are used so that the solution 61 does not enter, and the fine holes 17 constitute an air layer.
  • the wall member 16 has a multi-layered structure of the wall member layer 16a and the air layer (the minute hole 17) microscopically as shown in FIG.
  • the wall member layer 16a has a refractive index of about 1.6 to 1.9, and the inspection solution 61 has a refractive index of about 1.3 to 1.6, whereas the refractive index of the air layer is about 1. .0. That is, since the refractive index difference between the wall member 16 and the air layer is very large, at least a part of the light incident on the interface between the wall member layer 16a and the air layer is reflected at the interface. Since the wall member layer 16a has a larger refractive index than the air layer, the light incident on the interface C 1 at an incident angle theta 1 larger than the critical angle from the wall member 16 side to the air layer is totally reflected, a plate-shaped base It will go to the surface side of the material 10.
  • the light generated from the pores 14 in the region 20 can be reflected by the frame wall 18 made of aluminum having high reflectivity and guided to the surface side. Light attenuation can be suppressed and light extraction efficiency can be improved. Furthermore, when the micro hole 17 is provided in the wall member 16, light is condensed on the surface side of the device by causing reflection at a plurality of interfaces by the wall member 16, the air layer, and the multi-layered structure. Therefore, the light extraction efficiency can be greatly improved as compared with the case where a frame made of silicon disclosed in Patent Document 2 is provided.
  • FIG. 9 is a diagram illustrating a manufacturing process of the inspection device 1.
  • an aluminum substrate 100 is prepared (A1).
  • a mask 102 is formed on a portion to be the frame wall 18 made of aluminum on one surface and the entire surface of the other surface (A2).
  • the mask can be formed by a known lithography method or screen printing. For example, a method described in JP 2014-198353 A can be employed.
  • the aluminum base material 100 provided with the mask 102 at the part where the frame wall is to be formed is anodized.
  • known anodic oxidation methods described in JP-A-2015-132837 and JP-A-2014-198453 can be employed.
  • the portion of the aluminum substrate 100 that is not provided with the mask 102 is anodized to form an anodized alumina portion 104 (A3).
  • a mask is removed (A4).
  • the pores 14 are formed in the anodized alumina portion 104 (A5).
  • the pores 14 can be formed by a mechanical method using a drill or the like.
  • a mask may be provided in a portion other than the portion where the pore 14 is formed in the anodized alumina portion 104, and the pore 14 may be formed by chemical etching.
  • the method for producing the plate-like substrate 10 is not limited to the above procedure, and anodization is performed by providing the aluminum substrate with pores 14 first, and then providing a mask at the frame wall forming portion. It may be a procedure.
  • the anodized alumina portion 104 formed in the above procedure A3 is porous alumina and has a large number of fine holes. By performing a wide process on the desired fine holes, the diameter is larger than that of the fine holes. Large pores 14 can also be formed.
  • a known method can be used as the power wide processing, and for example, a method described in Japanese Patent Application Laid-Open No. 2014-198453 can be employed.
  • the other surface 12 is composed of a lower surface member 19 made of aluminum, and the fine hole 17 does not penetrate to the other surface 12 side.
  • An enlarged view of a region X indicated by a broken line of the plate-like substrate 10 shown in A5 of FIG. 9 is shown in A5 of FIG.
  • the barrier layer 101 and the aluminum substrate 100 that is the substrate substrate remain between the micro holes 17 and the other surface 12.
  • the other surface 12 side can be a closed hole.
  • the lower surface member 19 is comprised by the base-material base
  • the fine hole 17 can be formed as a non-through hole closed on the other surface 12 side without penetrating on the other surface 12 side.
  • the sealing treatment include high-temperature pressurized steam treatment and boiling treatment in boiling water.
  • the alumina on the surface becomes a hydrate, and the opening is substantially closed by the alumina hydrate layer 16b.
  • FIG. 10A5 the opening is shown as being completely blocked, but it is sufficient that the opening is blocked to the extent that the solution can be prevented from entering, and the opening is formed by the alumina hydrate layer 16b. It does not have to be completely blocked.
  • the capture substance which is a specific binding substance that specifically binds to the test substance is bound (immobilized) to the inner wall surface 14a of the pore 14 of the plate-like substrate 10 manufactured as described above.
  • the inspection device 1 can be manufactured.
  • a known method can be applied without any particular limitation.
  • the method disclosed in the above-described Patent Document 2 can be used.
  • FIG. 11 is a diagram schematically showing the configuration of the inspection apparatus 50 according to an embodiment of the present invention.
  • the inspection apparatus 50 according to the present embodiment is disposed on the inspection device 1, the solution supply unit 60 that supplies the inspection solution into the pores 14 of the inspection device 1, and the one surface 11 side of the inspection device 1.
  • a photodetector 70 a photodetector 70.
  • the light detector 70 detects light emitted from the inspection device 1.
  • FIG. 12 is a diagram illustrating a schematic configuration of the solution supply unit 60.
  • the solution supply unit 60 stores the inspection solution 61 installed on the other surface 12 side of the inspection device 1, and the inspection installed in the upper part of the storage unit 62 and stored in the storage unit 62.
  • the pipette part 64 for sucking the solution 61 for use, and the pressure of the depressurizing space part 66 and the depressurizing space part 66 arranged on the one surface 11 side of the inspection device 1 installed on the pipette part 64 are reduced.
  • a pump 68 for pressurization is provided.
  • the reduced pressure space 66 is decompressed by the pump 68, whereby the inspection solution 61 passes through the pipette part 64 and the pore 14 of the inspection device 1. Supplied in. Note that the inspection solution 61 is supplied to the pores 14 of the inspection device 1 and the inspection solution 61 is discharged from the pores 14 by depressurization and pressurization of the reduced pressure space 66 by the pump 68. it can.
  • the test solution 61 is a solution supplied into the pores 14 at the time of light detection, but the solution supply unit 60 includes a sample solution, a label solution, a cleaning solution, and the like to be supplied into the pores 14 in the test process. It is also used for supply. That is, the solution supply unit 60 supplies a necessary solution to the pores 14 for each inspection process.
  • the fine holes 17 (not shown in FIG. 12) provided in the wall member 16 are non-open on the other surface 12 side, so that the supplied solution is fine holes from the other surface 12 side. 17 does not enter.
  • the solution supply unit 60 supplies the solution so that the solution does not enter the fine hole 17 from the one surface 11 where the fine hole 17 is opened. Control. If the above-described sealing treatment is applied to the opening on the one surface 11 side of the fine hole 17, the solution does not enter the fine hole 17 from the one surface 11. Is preferable.
  • the light emitted from the test device and detected by the photodetector is, for example, fluorescence generated by excitation of a label attached to the test substance, or binding of an antibody that specifically binds to the test substance.
  • the substance is given the same label as above, and light is emitted from the label, or an enzyme is labeled on a binding substance such as an antibody that specifically binds to the test substance, and light emission by the reaction using this enzyme as a catalyst (in the following) , Or “chemiluminescence”)).
  • chemiluminescence if the test substance generates autofluorescence, no label is necessary, and autofluorescence may be detected.
  • the optical signal includes absorbance (colorimetric) in addition to fluorescence and chemiluminescence light.
  • FIG. 13 is a diagram schematically showing the inspection process.
  • a capturing substance 30 such as an allergen is fixed to the inner wall surface 14a of the pore 14 of the inspection device 1 (S1).
  • a specimen liquid containing a test substance (for example, a specific IgE antibody that specifically binds to the allergen) is supplied to the pores 14 of the test device 1, and the test substance 32 is bound to the capture substance 30 (S2). ).
  • the above-described solution supply unit 60 is used for supplying the sample liquid.
  • the sample liquid is efficiently brought into contact with the capture substance 30 in the pores 14 by repeatedly pumping and discharging the sample liquid by the pipette unit 64 while the sample liquid is stored in the storage unit 62. Can do.
  • a labeled solution containing a labeled substance 35 in which a label F is added to a substance 33 (for example, a secondary antibody) that specifically binds to the test substance 32 is supplied to the pores 14.
  • a substance 33 for example, a secondary antibody
  • the labeling substance 35 is bound to the test substance 32 (S3).
  • the label solution is supplied to the pores 14 by the solution supply unit 60 in the same manner as the sample solution.
  • the labeling substance 35 is bound to the test substance 32 after the test substance 32 is bound to the capture substance 30. It is also possible to combine the test substance 32 and the labeling substance 35 by mixing and supply the mixed liquid into the pores 14. In this case, by supplying the mixed liquid to the pores 14, the test substance 32 to which the labeling substance 35 is bound can be bound to the capture substance 30 fixed in the pores 14.
  • the cleaning solution is supplied to the pores 14 using the solution supply unit 60 in the same manner as the supply of the sample solution, and nonspecific adsorption is performed in the pores 14.
  • the test substance 32 and the labeling substance 35 are removed.
  • the cleaning liquid is discharged, and the optical signal from the luminescence reaction that reacts with the label F as a catalyst is detected by the photodetector 70 with the inspection solution such as the buffer solution filled in the pores 14 ( S4).
  • the wall member 16 is transparent and has a frame wall 18 around it. Since the air layer 17 constitutes an air layer, light detection can be performed efficiently.
  • the buffer solution is supplied into the pore 14 as a test solution, and the fluorescence measurement is performed with the pore 14 filled with the buffer solution. .
  • the inspection device is irradiated with light having a wavelength that excites the fluorescent label as excitation light, and fluorescence from the label excited by the excitation light is detected.
  • the photodetector 70 used for detecting the fluorescent label is provided with an excitation light irradiation unit.
  • the label F is an enzyme or the like that acts as a catalyst for a chemical reaction in which luminol is oxidized by hydrogen peroxide to generate luminescence
  • the chemical by contacting with the enzyme that acts as a catalyst after the washing treatment.
  • a reaction solution containing a substance that emits light by the reaction is supplied into the pores 14 as a test solution.
  • the photodetector 70 the reaction solution is supplied to the pores 14 so that the chemiluminescent label attached to the test substance 32 captured by the capture material 30 in the pores 14 is a substance in the reaction solution.
  • Luminescence generated by reaction with is detected. In this case, luminescence detection is performed in a state where the pores 14 are filled with the reaction solution.
  • enzyme labels that generate chemiluminescence include enzymes that react with chemiluminescent substrates such as luminol, lophine, lucigenin and oxalate.
  • HRP horseradish peroxidase
  • a reaction liquid luminol reaction liquid
  • HRP functions as a catalyst an enzyme for converting HRP to alkaline phosphatase. It is preferable to use a reaction solution containing a dioxetane chemiluminescent substrate.
  • the luminol reaction solution contains at least a luminol substrate and a hydrogen peroxide solution.
  • the enzyme label catalyzes the oxidation of luminol in the presence of hydrogen peroxide.
  • the reaction solution preferably contains a sensitizer that sensitizes chemiluminescence.
  • chemiluminescent substrate described above but also a reaction solution containing a luminescent substrate or a fluorescent substrate may be used to detect a color (absorption) reaction or fluorescence. it can.
  • the inspection device of the present invention by using the inspection device of the present invention, the light generated in the pores is efficiently emitted from the surface of the inspection device, and the light extraction efficiency is high. Is possible.

Abstract

[Problem] To provide: an inspection device that enables light extraction efficiency to be improved and can be produced at low cost; an inspection apparatus equipped with the inspection device; and an inspection method. [Solution] An inspection device (1) provided with a plurality of holes (14) that penetrate from one surface (11) to the other surface (12) of a sheet-like substrate (10), said inspection device (1) comprising: regions (20), each of which contains at least some of the holes (14), has at least a portion of wall members (16), which constitute the inner wall surfaces (14a) of the at least some of the holes (14), comprising alumina; and frame walls (18) comprising aluminum that surround the regions (20) when seen from the one surface (11); wherein a trapping substance that specifically binds with a specific substance is fixed on the inner wall surfaces (14a) of the at least some of the holes (14).

Description

検査デバイス、検査装置および検査方法Inspection device, inspection apparatus, and inspection method
 本発明は、被検物質である抗原、抗体もしくはデオキシリボ核酸などを検出するための検査デバイス、その検査デバイスを備えた検査装置および検査方法に関する。 The present invention relates to a test device for detecting an antigen, antibody, deoxyribonucleic acid, or the like, which is a test substance, a test apparatus equipped with the test device, and a test method.
 生化学的な反応、例えば、酵素反応、核酸ハイブリダイゼーション、抗原-抗体反応などの特異的結合反応を検査する方法の一つとして、被検物質と捕捉物質との間の結合現象を光学的に検出する方法が知られている。この方法は、所定位置に固定されている捕捉物質に被検物質を結合させ、励起光を受けて蛍光を発する標識、あるいは基質の反応を触媒して発色、蛍光もしくは化学発光を生じさせる標識などをその被検物質に付与し、かかる標識に起因して生じる光を検出するものである。より具体的には、被検物質に特異的に結合する抗体などの結合物質に蛍光標識を付与し、蛍光標識から生じる蛍光を検出する方法、被検物質に特異的に結合する抗体などの結合物質に酵素を標識し、この酵素を触媒として反応する発色基質、蛍光基質、あるいは化学発光基質から生じる発色や蛍光、化学発光を検出する方法等が知られており、これらにより、被検物質の特定が可能となる。 As one of the methods for examining biochemical reactions, for example, specific binding reactions such as enzyme reactions, nucleic acid hybridizations, and antigen-antibody reactions, the binding phenomenon between a test substance and a capture substance is optically determined. A method of detecting is known. In this method, a test substance is bound to a capture substance fixed at a predetermined position and emits fluorescence upon receiving excitation light, or a label that catalyzes a substrate reaction to generate color, fluorescence, or chemiluminescence. Is applied to the test substance, and light generated due to the label is detected. More specifically, a method of detecting a fluorescence generated from a fluorescent label by attaching a fluorescent label to a binding substance such as an antibody that specifically binds to the test substance, a binding of an antibody that specifically binds to the test substance There are known methods for detecting color development, fluorescence, chemiluminescence, etc. generated from a chromogenic substrate, fluorescent substrate, or chemiluminescent substrate that reacts with this enzyme as a catalyst. Identification becomes possible.
 このような検査に用いられるバイオチップとしては、二次元基板上に生体関連分子が規則的に配列された多項目同時検出のためのチップが従来用いられている。近年では、支持体に多数の貫通孔(細孔)が整列配置されてなる多孔性基板からなるデバイスの検討が進められている。細孔内に捕捉物質を固定化し、被検物質を含有する検体液を多孔性基板の裏面側から表面へと貫通孔を介してポンプで汲み上げ、循環させることにより、検体液が捕捉物質に効率的に接触されて、被検物質を捕捉物質に結合させることができ、測定時間の大幅な短縮化を図ることができる。 As a biochip used for such an inspection, a multi-item simultaneous detection chip in which biologically related molecules are regularly arranged on a two-dimensional substrate has been conventionally used. In recent years, a device made of a porous substrate in which a large number of through-holes (pores) are arranged on a support has been studied. By immobilizing the capture substance in the pores, pumping the sample liquid containing the test substance from the back side to the surface of the porous substrate through the through-hole and circulating it, the sample liquid becomes efficient as the capture substance The test substance can be bonded to the capture substance and the measurement time can be greatly shortened.
 多孔性を有する基板としては、多孔性のシリコン(Si)基材が知られているが、シリコンは反射率が低く、細孔内で生じた光は細孔内で多重反射するうちに、その信号強度が大幅に減衰してしまうため、細孔内で生じた光信号の光取り出し効率が非常に低いという問題がある。 As a porous substrate, a porous silicon (Si) base material is known, but silicon has low reflectivity, and the light generated in the pores is reflected multiple times in the pores. Since the signal intensity is greatly attenuated, there is a problem that the light extraction efficiency of the optical signal generated in the pore is very low.
 特許文献1には、アルミニウム(Al)基材に多数の貫通孔を有してなる多孔性基板が開示されている。アルミニウムはシリコンと比較して反射率が高いため、シリコン基材を用いた場合と比較すると光取り出し効率を向上できると考えられる。しかしながら、個々の細孔の光出射開口は小さいために、光取り出し効率を十分に向上できるとは言えない。一方で、光取り出し開口を広がるために細孔径を大きくすると、細孔密度が低下し、全体として固定できる捕捉物質量が減少してしまう。 Patent Document 1 discloses a porous substrate having a large number of through holes in an aluminum (Al) base material. Since aluminum has a higher reflectance than silicon, it is considered that the light extraction efficiency can be improved as compared with the case where a silicon substrate is used. However, it cannot be said that the light extraction efficiency can be sufficiently improved because the light exit openings of the individual pores are small. On the other hand, if the pore diameter is increased in order to widen the light extraction opening, the pore density is lowered, and the amount of trapped substance that can be fixed as a whole is reduced.
 特許文献2では、上記問題を解決する手段として、多孔性シリコンを熱酸化処理して局部的に透明な酸化シリコン(SiO)化し、その酸化された領域をシリコンからなる枠(壁)によって取り囲むことにより、光の出射開口を実質的に広げて光取り出し効率を向上させた構成が提案されている。 In Patent Document 2, as a means for solving the above problem, porous silicon is thermally oxidized to be locally transparent silicon oxide (SiO 2 ), and the oxidized region is surrounded by a frame (wall) made of silicon. Thus, a configuration has been proposed in which the light output aperture is substantially widened to improve the light extraction efficiency.
特開2004-93152号公報JP 2004-93152 A 特許第4125244号公報Japanese Patent No. 4125244
 しかしながら、特許文献2では、光開口率の増加に伴う光取り出し効率の向上は可能であるが、枠壁として用いられているシリコンの反射率が低いため、やはりシリコンの壁に入射する光は減衰されてしまい枠壁に入射する光の取り出し効率は向上しない。特に、チップの高性能化を図るためには、チップを厚くして、検出領域を広げたり、細孔を高密度化したりする必要が生じると考えられ、細孔の長さ(基板厚み)と開口径とのアスペクト比を大きくする必要性が生じる。細孔のアスペクト比が大きくなると、シリコン枠で囲まれた領域のアスペクト比も追従して大きくなるために、シリコン枠で囲まれた領域の光出射開口が実質的に狭くなり、シリコン枠での多重反射による光信号減衰の影響が大きくなる。 However, in Patent Document 2, although the light extraction efficiency can be improved as the optical aperture ratio increases, the reflectance of silicon used as a frame wall is low, so that light incident on the silicon wall is also attenuated. Therefore, the extraction efficiency of light incident on the frame wall is not improved. In particular, in order to improve the performance of the chip, it is considered necessary to increase the thickness of the chip, expand the detection area, or increase the density of the pores. There is a need to increase the aspect ratio with the opening diameter. As the aspect ratio of the pores increases, the aspect ratio of the region surrounded by the silicon frame also increases accordingly, so the light exit aperture of the region surrounded by the silicon frame becomes substantially narrow, The influence of optical signal attenuation due to multiple reflection is increased.
 本発明は、上記事情に鑑み、従来のデバイスよりも光取り出し効率を向上させることができる検査デバイスを提供することを目的とする。また、本発明は高い光取り出し効率を実現した検査デバイスを備えた検査装置および検査方法を提供することを目的とする。 In view of the above circumstances, an object of the present invention is to provide an inspection device capable of improving the light extraction efficiency as compared with a conventional device. Another object of the present invention is to provide an inspection apparatus and an inspection method provided with an inspection device that realizes high light extraction efficiency.
 本発明の検査デバイスは、板状基材の一方の面から他方の面に貫通する複数の細孔を備えた検査デバイスであって、
 少なくとも一部の細孔を含む領域であって、その少なくとも一部の細孔の内壁面を構成する壁部材の少なくとも一部がアルミナからなる領域と、上記一方の面から見た場合に、その領域を囲むアルミニウムからなる枠壁とを有し、
 前記少なくとも一部の細孔の内壁面に特定の物質と特異的に結合する捕捉物質が固定されている検査デバイスである。
The inspection device of the present invention is an inspection device having a plurality of pores penetrating from one surface of the plate-like substrate to the other surface,
A region including at least a part of the pores, wherein at least a part of the wall member constituting the inner wall surface of the at least a part of the pores is made of alumina, and when viewed from the one side, A frame wall made of aluminum surrounding the region,
In the inspection device, a capture substance that specifically binds to a specific substance is fixed to an inner wall surface of the at least some pores.
 本発明の検査デバイスは、壁部材が、細孔と平行に伸び、かつ、細孔より小さい直径の微細穴を有するものであることが好ましい。 In the inspection device of the present invention, it is preferable that the wall member has a fine hole extending in parallel with the pore and having a diameter smaller than the pore.
 本発明の検査デバイスは、壁部材が、陽極酸化アルミナからなることが好ましい。 In the inspection device of the present invention, the wall member is preferably made of anodized alumina.
 本発明の検査デバイスは、枠壁が壁部材と接して設けられていることが好ましい。 In the inspection device of the present invention, the frame wall is preferably provided in contact with the wall member.
 本発明の検査デバイスは、他方の面がアルミニウムからなることが好ましい。 In the inspection device of the present invention, the other surface is preferably made of aluminum.
 本発明の検査デバイスにおいては、捕捉物質が、抗原、抗体またはデオキシリボ核酸(deoxyribonucleic acid:DNA)であることが好ましい。 In the test device of the present invention, the capture substance is preferably an antigen, an antibody, or deoxyribonucleic acid (DNA).
 本発明の検査デバイスは、アルミニウムからなる枠壁により囲まれた上記領域を複数備え、その複数の領域の1つの領域中の細孔の内壁面には捕捉物質として同一種の捕捉物質が結合されており、その複数の領域は、結合されている捕捉物質が互いに異なる領域を有することが好ましい。 The inspection device of the present invention comprises a plurality of the above-mentioned regions surrounded by a frame wall made of aluminum, and the same type of capture material is bound as a capture material to the inner wall surface of the pore in one of the plurality of regions. It is preferable that the plurality of regions have regions in which the bound capture substances are different from each other.
 本発明の検査デバイスは、細孔の一方の面における開口領域の円相当直径が1μm~100μmであることが好ましい。 In the inspection device of the present invention, the equivalent circle diameter of the open region on one surface of the pore is preferably 1 μm to 100 μm.
 本発明の検査デバイスは、板状基材の厚さが100μm~2000μmであることが好ましい。 In the inspection device of the present invention, the thickness of the plate-like substrate is preferably 100 μm to 2000 μm.
 本発明の検査装置は、本発明の検査デバイスと、
 検査デバイスの細孔中に検査用溶液を供給する溶液供給部と、
 検査デバイスの一方の面または他方の面の側に配置され、検査デバイスから出射される蛍光を検出する光検出器とを備えた検査装置である。
The inspection apparatus of the present invention includes the inspection device of the present invention,
A solution supply unit for supplying an inspection solution into the pores of the inspection device;
The inspection apparatus includes a photodetector that is disposed on one surface or the other surface of the inspection device and detects fluorescence emitted from the inspection device.
 本発明の検査方法は、本発明の検査デバイスの上記少なくとも一部の細孔に、特定の物質を含有する検体液を供給して、特定の物質を捕捉物質に結合させ、
 特定の物質と特異的に結合する標識物質を特定の物質に結合させ、
 細孔に検査用溶液を供給して細孔に検査用溶液を留めた状態で、検査デバイスから出射される光を検出する検査方法である。
In the test method of the present invention, a specimen liquid containing a specific substance is supplied to the at least some of the pores of the test device of the present invention to bind the specific substance to the capture substance
A labeling substance that specifically binds to a specific substance is bound to the specific substance,
The inspection method detects light emitted from the inspection device in a state where the inspection solution is supplied to the pores and the inspection solution is retained in the pores.
 本発明の検査方法においては、上記標識として酵素標識を含む物質を用い、上記検査用溶液として酵素標識により触媒されて反応する基質を含む反応液を用い、上記検査デバイスから出射される光として、反応液中の基質が酵素標識により触媒されて生じる光を検出してもよい。
 なお、基質としては、発色基質、蛍光基質および化学発光基質などが挙げられ、これらの基質は酵素標識の種類に応じて適宜選択される。また、この基質に応じて、検査デバイスから出射される光は異なり、検出される光は、吸光(呈色)、蛍光または化学発光である。
In the inspection method of the present invention, a substance containing an enzyme label is used as the label, a reaction solution containing a substrate that is catalyzed by the enzyme label and reacts as the inspection solution, and the light emitted from the inspection device is as follows: You may detect the light which a substrate in a reaction liquid produces when catalyzed by an enzyme label | marker.
Examples of the substrate include a chromogenic substrate, a fluorescent substrate, and a chemiluminescent substrate, and these substrates are appropriately selected depending on the type of enzyme label. Moreover, the light emitted from the inspection device differs depending on the substrate, and the detected light is light absorption (coloration), fluorescence, or chemiluminescence.
 本発明の検査方法においては、標識物質として蛍光標識を含む物質を用い、
 蛍光標識を励起させる励起光を検査デバイスに照射し、
 上記検査デバイスから出射される光として、励起光の照射により標識物質から生じる蛍光を検出してもよい。
In the inspection method of the present invention, a substance containing a fluorescent label is used as a labeling substance,
Irradiate the test device with excitation light that excites the fluorescent label,
As the light emitted from the inspection device, fluorescence generated from the labeling substance by irradiation with excitation light may be detected.
 本発明の検査デバイスは、板状基材の一方の面から他方の面に貫通する複数の細孔を備えた検査デバイスであって、少なくとも一部の細孔を含む領域であって、その少なくとも一部の細孔の内壁面を構成する壁部材の少なくとも一部がアルミナからなる領域と、上記一方の面から見た場合に、その領域を囲むアルミニウムからなる枠壁を有しているので、細孔内において生じる光の検査デバイスの少なくとも一方の面側への取り出し効率を向上させることができる。 The inspection device of the present invention is an inspection device having a plurality of pores penetrating from one surface of the plate-like substrate to the other surface, and is an area including at least some of the pores, Since at least a part of the wall member constituting the inner wall surface of some of the pores has a region made of alumina and a frame wall made of aluminum surrounding the region when viewed from the one surface, The extraction efficiency of the light generated in the pores to at least one surface side of the inspection device can be improved.
本発明の実施形態にかかる検査デバイスの斜視図である。It is a perspective view of the inspection device concerning the embodiment of the present invention. 図1に示す検査デバイスの平面図である。It is a top view of the inspection device shown in FIG. 図1に示す検査デバイスの断面図である。It is sectional drawing of the test | inspection device shown in FIG. 図1に示す検査デバイスの下面図である。It is a bottom view of the inspection device shown in FIG. 検査デバイスの設計変更例を示す断面図である。It is sectional drawing which shows the example of a design change of a test | inspection device. 検査デバイスの一部拡大斜視図である。It is a partial expansion perspective view of an inspection device. 本発明の検査デバイスの効果を説明するための模式図である(その1)。It is a schematic diagram for demonstrating the effect of the test | inspection device of this invention (the 1). 本発明の検査デバイスの効果を説明するための模式図である(その2)。It is a schematic diagram for demonstrating the effect of the test | inspection device of this invention (the 2). 本発明の検査デバイスの作製工程を示す図である(その1)。It is a figure which shows the manufacturing process of the test | inspection device of this invention (the 1). 本発明の検査デバイスの作製工程を示す図である(その2)。It is a figure which shows the manufacturing process of the test | inspection device of this invention (the 2). 本発明の検査デバイスを備えた一実施形態の検査装置の概略構成を示す図である。It is a figure which shows schematic structure of the test | inspection apparatus of one Embodiment provided with the test | inspection device of this invention. 検査装置における溶液供給部を説明するための模式図である。It is a schematic diagram for demonstrating the solution supply part in a test | inspection apparatus. 本発明の検査デバイスを用いた検査工程を示す模式図である。It is a schematic diagram which shows the test | inspection process using the test | inspection device of this invention.
 以下、図面を参照して本発明の実施形態を詳細に説明する。なお、本明細書において「~」とは、その前後に記載される数値を下限値および上限値として含む意味で使用される。 Hereinafter, embodiments of the present invention will be described in detail with reference to the drawings. In the present specification, “to” is used to mean that the numerical values described before and after it are included as a lower limit value and an upper limit value.
 図1は本発明の第1の実施形態にかかる検査デバイスの斜視図であり、図2は図1の検査デバイスの平面図、図3は図2の検査デバイスのIII-III線断面図、図4は図1の検査デバイスの下面図である。 1 is a perspective view of an inspection device according to a first embodiment of the present invention, FIG. 2 is a plan view of the inspection device of FIG. 1, and FIG. 3 is a sectional view taken along line III-III of the inspection device of FIG. 4 is a bottom view of the inspection device of FIG.
 本実施形態の検査デバイス1は、一方の面11から他方の面12に貫通する複数の細孔14を備えた板状基材10に、細孔の内壁面14aを構成する、アルミナからなる壁部材16を有する領域20と、一方の面11から見た場合に、その領域20を囲むアルミニウムからなる枠壁18とを有する。本実施形態においては、縦横に3本ずつの計9本の細孔14を1つの領域20として、各領域20を囲むように枠壁18が設けられている。そして、細孔14の内壁面14aには特定の物質と特異的に結合する捕捉物質30が固定されている。 The inspection device 1 of the present embodiment is a wall made of alumina that constitutes an inner wall surface 14a of a pore in a plate-like substrate 10 having a plurality of pores 14 penetrating from one surface 11 to the other surface 12. It has the area | region 20 which has the member 16, and the frame wall 18 which consists of aluminum surrounding the area | region 20 when it sees from the one surface 11. FIG. In the present embodiment, a total of nine pores 14 in the vertical and horizontal directions are used as one region 20, and a frame wall 18 is provided so as to surround each region 20. A trapping substance 30 that specifically binds to a specific substance is fixed to the inner wall surface 14 a of the pore 14.
 図2に示す平面図は、検査デバイス1を一方の面11から見た図であり、図4に示す下面図は、検査デバイス1を他方の面12から見た図である。なお、検査デバイス1の外形は板状基材10の外形により構成されるものであるため、以下において、板状基材10の一方の面11および他方の面12を、検査デバイス1の一方の面および他方の面と称する場合がある。 The plan view shown in FIG. 2 is a view of the inspection device 1 as viewed from one surface 11, and the bottom view of FIG. 4 is a view of the inspection device 1 as viewed from the other surface 12. In addition, since the external shape of the inspection device 1 is configured by the external shape of the plate-like base material 10, one surface 11 and the other surface 12 of the plate-like base material 10 are hereinafter referred to as one of the inspection devices 1. Sometimes referred to as the surface and the other surface.
 本実施形態の検査デバイス1においては、図3に示すように4つの領域20の内壁面14aを構成する壁部材のうち他方の面12側の一部を除きアルミナから構成されている。そして、この領域20を取り囲みアルミナからなる壁部材16に接してアルミニウムからなる枠壁18が設けられている。また、板状基材10の他方の面12を構成する下面部材19はアルミニウムからなる。この下面部材19は枠壁18と連続し一体的に構成されている。このように、本実施形態の検査デバイス1は板状基材10が全ての領域に亘って、アルミニウムとアルミナとから構成されている。 In the inspection device 1 of the present embodiment, as shown in FIG. 3, the wall member constituting the inner wall surface 14a of the four regions 20 is made of alumina except for a part on the other surface 12 side. A frame wall 18 made of aluminum is provided surrounding the region 20 and in contact with the wall member 16 made of alumina. Moreover, the lower surface member 19 which comprises the other surface 12 of the plate-shaped base material 10 consists of aluminum. The lower surface member 19 is continuous with the frame wall 18 and is integrally formed. As described above, in the inspection device 1 of the present embodiment, the plate-like substrate 10 is composed of aluminum and alumina over the entire region.
 なお、アルミニウムからなる下面部材19を備えたことにより、細孔14中で発生した光のうち、他方の面12側に放射され、下面部材19に入射する成分が一方の面11側に反射されるので、一方の面11側への光取り出し効率を向上させることができる。 In addition, by providing the lower surface member 19 made of aluminum, a component of the light generated in the pores 14 that is emitted to the other surface 12 side and incident on the lower surface member 19 is reflected to the one surface 11 side. Therefore, the light extraction efficiency to the one surface 11 side can be improved.
 しかしながら、図5に設計変更例の検査デバイスの断面図を示すように、検査デバイスとしては、下面部材19を備えていない構成であってもよい。下面部材19を備えない場合には、光検出を一方の面11側、他方の面12側のいずれの方向から行っても同様の結果を得ることができる。 However, as shown in the cross-sectional view of the inspection device of the design change example in FIG. 5, the inspection device may not have the lower surface member 19. When the lower surface member 19 is not provided, the same result can be obtained even if the light detection is performed from either one of the surfaces 11 side and the other surface 12 side.
 なお、本発明の検査デバイスとしては、少なくとも一部の細孔14を含む領域20の内壁面14aを構成する壁部材の少なくとも一部がアルミナにより構成されていればよい。すなわち、枠壁18で囲まれた領域20において、壁部材はアルミナで形成されている部分と他の材料(例えばアルミニウム)で形成された部分が混在していてもよい。また、本実施形態においては、枠壁18は領域20の外周を囲むように形成されているが、繋がっていない部分の長さが領域20外周の長さの20%程度であれば一部分断されていてもよい。 In the inspection device of the present invention, it is only necessary that at least a part of the wall member constituting the inner wall surface 14a of the region 20 including at least a part of the pores 14 is made of alumina. That is, in the region 20 surrounded by the frame wall 18, the wall member may include a portion formed of alumina and a portion formed of another material (for example, aluminum). In this embodiment, the frame wall 18 is formed so as to surround the outer periphery of the region 20, but is partially cut off if the length of the unconnected portion is about 20% of the length of the outer periphery of the region 20. It may be.
 板状基材10はアルミニウム基材の少なくとも一部が酸化処理されてアルミナからなる壁部材16が形成されてなるものが好ましい。係る構成によれば、安価なアルミニウム基材を用いることができ、かつ作製が容易であるため、低コストで作製することができるため好ましい。しかしながら、板状基材10は、アルミニウムおよびアルミナ以外の異なる材料からなる部分を備えていてもよい。 The plate-like base material 10 is preferably one in which at least a part of an aluminum base material is oxidized to form a wall member 16 made of alumina. According to such a configuration, an inexpensive aluminum base material can be used, and since it can be easily manufactured, it is preferable because it can be manufactured at low cost. However, the plate-like substrate 10 may include a portion made of a different material other than aluminum and alumina.
 板状基材10の厚さには、特に制限はないが、100μm~2000μm程度が好ましい。
 板状基材10に設けられる細孔14は、本実施形態に示すように、整列配置されていることが好ましいが、ランダムに配置されていてもよい。また、枠壁18で囲まれた領域20(以下において、「検査領域20」と称する。)内には、細孔14が1本のみ含むものであってもよいが、複数の細孔14を含むことが好ましい。本実施形態においては、視認容易のために3行×3列の9本の細孔を1つの検査領域20に備えるものとしたが、例えば、1つの検査領域に10行×10列の100本の細孔を含むなど、1つの検査領域中の細孔の数に制限はない。また、1つの検査デバイス1中に備えられる検査領域20も単数であっても、複数であってもよい。本実施形態においては4つの検査領域20を示しているが、例えば、10以上の検査領域を備えることもできる。
The thickness of the plate-like substrate 10 is not particularly limited, but is preferably about 100 μm to 2000 μm.
The pores 14 provided in the plate-like substrate 10 are preferably arranged in an aligned manner as shown in the present embodiment, but may be arranged randomly. In addition, the region 20 surrounded by the frame wall 18 (hereinafter referred to as “inspection region 20”) may include only one pore 14, but a plurality of pores 14 are included. It is preferable to include. In the present embodiment, nine pores of 3 rows × 3 columns are provided in one inspection region 20 for easy visual recognition, but, for example, 100 10 rows × 10 columns in one inspection region. There is no limit to the number of pores in one inspection region, such as a plurality of pores. Further, the inspection area 20 provided in one inspection device 1 may be singular or plural. Although four inspection areas 20 are shown in the present embodiment, for example, ten or more inspection areas can be provided.
 細孔14の開口および断面形状は本実形態においては正方形状であるが、正方形、長方形などの矩形に限らず、円形、楕円形、三角形あるいは五角形以上の多角形であってもよい。なお、作製上の都合により多角形状の角部には丸みがつく場合がある。細孔14は、柱状で断面形状が変化しないものが一般的であるが、一部断面形状が変化したり、断面の大きさが変化したりしても構わない。 The opening and cross-sectional shape of the pore 14 are square in this embodiment, but are not limited to a rectangle such as a square or a rectangle, but may be a circle, an ellipse, a triangle, or a polygon that is a pentagon or more. Note that the corners of the polygon may be rounded due to manufacturing reasons. The pores 14 are generally columnar and the cross-sectional shape does not change, but the cross-sectional shape may partially change or the cross-sectional size may change.
 なお、細孔14の少なくとも一方の面12における開口の円相当直径が1μm~100μm程度であることが好ましい。より好ましくは3μm~50μmであり、特に好ましくは5μm~30μmである。なお、円相当直径とは、開口領域の面積と同等の面積を有する円の直径をいう。 It should be noted that the equivalent circle diameter of the opening in at least one surface 12 of the pore 14 is preferably about 1 μm to 100 μm. More preferably, it is 3 μm to 50 μm, and particularly preferably 5 μm to 30 μm. The equivalent circle diameter refers to the diameter of a circle having an area equivalent to the area of the opening region.
 本検査デバイス1において検査対象とされる被検物質(標的分子)は、主として、生体由来分子であり、抗原および抗体などのタンパク質、糖類、ペプチド、DNA、リボ核酸(ribonucleic acid:RNA)、ペプチド核酸(peptide nucleic acid:PNA)などである。そして、細孔14の内壁面14aに固定されている、特定の物質と特異的に結合する捕捉物質30としては、これらの被検物質と特異的に結合する物質である。特に、本検査デバイス1は、抗原の一種であるアレルゲンを捕捉物質として備えたアレルギー検査に好適である。 The test substance (target molecule) to be tested in this testing device 1 is mainly a biological molecule, such as proteins such as antigens and antibodies, saccharides, peptides, DNA, ribonucleic acid (RNA), peptides Examples thereof include nucleic acids (peptide nucleic acid: PNA). The capture substance 30 that is fixed to the inner wall surface 14a of the pore 14 and specifically binds to a specific substance is a substance that specifically binds to these test substances. In particular, the test device 1 is suitable for an allergy test provided with an allergen that is a kind of antigen as a capture substance.
 図2に示すように、本検査デバイス1は枠壁18より区画された複数の検査領域20を有している。1つの検査領域20中の細孔14の内壁には同一種の捕捉物質が結合(固定)されている。検査領域20間では、異なる捕捉物質を固定させておくことができる。これにより、1つの検査デバイス1で、複数の被検物質についての検査を同時に行うことが可能となる。なお、複数の検査領域には、同一種の捕捉物質が結合されている領域が2以上あってもよい。また、1つのデバイスにおいて、互いに異なる捕捉物質が固定されている検査領域20が交互に配置されて周期的に同一種の捕捉物質が固定された検査領域が配置されていてもよい。係る構成によれば、ばらつきを抑制し、信頼性の高い検査結果を得ることが可能となる。 As shown in FIG. 2, the present inspection device 1 has a plurality of inspection regions 20 partitioned by a frame wall 18. A trapping substance of the same type is bound (fixed) to the inner wall of the pore 14 in one inspection region 20. Different capture substances can be fixed between the examination regions 20. Accordingly, it is possible to simultaneously inspect a plurality of test substances with one inspection device 1. In the plurality of inspection regions, there may be two or more regions to which the same type of capture substance is bound. Further, in one device, inspection regions 20 in which different capture substances are fixed may be alternately arranged, and inspection regions in which the same type of capture material is periodically fixed may be disposed. According to such a configuration, it is possible to suppress variation and obtain a highly reliable test result.
 具体的には、1つの検査デバイス中において、1つもしくは複数の検査領域20に捕捉物質である同一種のアレルゲンを固定することとして、複数種のアレルゲン検査領域20を備えることにより、複数のアレルゲンに対する反応を同時に検査することができ、好ましい。 Specifically, a plurality of allergen test regions 20 are provided by fixing the same type of allergen as a capture substance in one or a plurality of test regions 20 in one test device. It is preferable that the response to can be simultaneously examined.
 図6は、検査デバイス1の一部を拡大して示した斜視図である。本実施形態の検査デバイス1のように、アルミナからなる壁部材16には、細孔14と平行に伸び、かつ、細孔14より小さい直径の微細穴17を有することが好ましい。このような微細穴17は、アルミニウムを陽極酸化してアルミナに変質させることにより形成できる。アルミニウム基材を陽極酸化することにより、整列した微細穴17を多数備えたポーラスアルミナを容易に得ることができる。陽極酸化により形成される微細穴の直径は数nm~数10nm程度であり、5nm~20nmであることが好ましい。 FIG. 6 is an enlarged perspective view showing a part of the inspection device 1. Like the inspection device 1 of the present embodiment, the wall member 16 made of alumina preferably has fine holes 17 extending in parallel with the pores 14 and having a diameter smaller than the pores 14. Such fine holes 17 can be formed by anodizing aluminum to transform it into alumina. By anodizing the aluminum substrate, porous alumina having a large number of aligned fine holes 17 can be easily obtained. The diameter of the fine hole formed by anodic oxidation is about several nm to several tens of nm, and preferably 5 nm to 20 nm.
 なお、微細穴17は、一方の面11から他方の面12に貫通する貫通孔ではなく、少なくとも他方の面12側が閉じた非貫通孔であることが好ましい。例えば、上述の下面部材19を備えることにより非貫通孔とすることができる。また、下面部材19を備えない場合には、陽極酸化処理の際に基材素地との間に形成されるバリア層(緻密なアルミナ層)を備えたものとすればよい。なお、一方の面11側は開口したままでもよいが、一方の面11の開口を塞ぐ封孔処理が施されていることも好ましい。これにより、微細穴17は、閉じた空洞部17aとされる。本発明においては、このようにして形成された空洞部17aも微細穴の1種と看做すことが可能である。係る構成によれば、一方の面11および他方の面12のいずれからの溶液の浸入も抑制することができ、空洞部を空気層として利用することができ、後述の光取り出し効率向上への寄与が容易となる。 The fine hole 17 is preferably not a through-hole penetrating from one surface 11 to the other surface 12 but a non-through hole with at least the other surface 12 closed. For example, a non-through hole can be formed by providing the lower surface member 19 described above. Further, when the lower surface member 19 is not provided, a barrier layer (dense alumina layer) formed between the base material base and the anodizing treatment may be provided. In addition, although the one surface 11 side may remain open, it is also preferable that a sealing process for closing the opening of the one surface 11 is performed. Thereby, the fine hole 17 is made into the closed cavity part 17a. In the present invention, the hollow portion 17a formed in this way can also be regarded as a kind of fine hole. According to such a configuration, the infiltration of the solution from any one of the one surface 11 and the other surface 12 can be suppressed, and the cavity can be used as an air layer, contributing to the improvement of light extraction efficiency described later. Becomes easy.
 次に、上記各実施形態の検査デバイス1の効果について図7を参照して説明する。
 図7は検査デバイス1の一部を拡大して示した模式図である。本検査デバイス1は、細孔14の内壁面14aに固定された捕捉物質(図7中では省略している。)に、一例として、被検物質を結合させ、この被検物質に特異的に結合する酵素で標識した結合物質を結合させ、この酵素が触媒として作用する発光反応により生じる光信号を検出することにより、被検物質の有無、または被検物質の量を検出する、あるいは、被検物質の有無及び被検物質の量を検出する検査方法に用いられる。本検査デバイス1からの光信号検出は、細孔14中にはバッファ液等の検査用溶液61が充填された状態で実施される。
Next, the effect of the inspection device 1 of each of the above embodiments will be described with reference to FIG.
FIG. 7 is a schematic view showing a part of the inspection device 1 in an enlarged manner. As an example, the present inspection device 1 binds a test substance to a capture substance (not shown in FIG. 7) fixed to the inner wall surface 14a of the pore 14, and specifically detects the test substance. By binding a binding substance labeled with a binding enzyme and detecting a light signal generated by a luminescence reaction in which this enzyme acts as a catalyst, the presence or absence of the test substance or the amount of the test substance is detected. It is used in an inspection method for detecting the presence or absence of a test substance and the amount of a test substance. The detection of the optical signal from the present inspection device 1 is performed in a state where the pore 14 is filled with an inspection solution 61 such as a buffer solution.
 検査領域20の細孔14の内壁面14aを構成する壁部材16はアルミナからなり光透過性を有する。従来の不透過なシリコン基材により構成された細孔では、1つの細孔14内で発生した光の一部は1つの細孔内で反射を繰り返して表面から出射される。これに対し、壁部材16が光透過性を有するので、1つの細孔14内で発生した光の一部は隣接する細孔を介して表面側へと導光されうる。その結果として、細孔14内で発生した光の表面に出射する光開口率が増加する。また、検査領域20を囲むようにアルミニウムからなる枠壁18が備えられているため、検査領域20の細孔14において生じた光の一部は枠壁18との界面Cで反射されて板状基材の表面側へと導光される。アルミニウムはシリコンと比較して反射率が高く、反射による減衰も小さいため、より多くの光成分をデバイス表面へと取り出すことが可能となる。 The wall member 16 constituting the inner wall surface 14a of the pore 14 in the inspection region 20 is made of alumina and has light transmittance. In a conventional pore formed of an impermeable silicon substrate, a part of light generated in one pore 14 is repeatedly reflected in one pore and emitted from the surface. On the other hand, since the wall member 16 has optical transparency, a part of the light generated in one pore 14 can be guided to the surface side through the adjacent pore. As a result, the aperture ratio of light emitted to the surface of the light generated in the pores 14 increases. Further, since the frame wall 18 made of aluminum is provided so as to surround the inspection region 20, a part of the light generated in the pores 14 in the inspection region 20 is reflected by the interface C 0 with the frame wall 18 and is a plate. The light is guided to the surface side of the substrate. Since aluminum has a higher reflectance than silicon and is less attenuated by reflection, more light components can be extracted to the device surface.
 さらに、本実施形態の検査デバイス1のように、他方の面12を構成する下面部材19がアルミニウムから形成されていれば、細孔14において生じ、直接もしくは間接的に下面部材19に入射する光を、一方の面11側に反射させることができる。この場合、一方の面11側に出射する光をより増加させることができ、取り出し効率をさらに向上させることができる。 Further, if the lower surface member 19 constituting the other surface 12 is made of aluminum as in the inspection device 1 of the present embodiment, the light that is generated in the pore 14 and directly or indirectly incident on the lower surface member 19. Can be reflected on the one surface 11 side. In this case, the light emitted to the one surface 11 side can be further increased, and the extraction efficiency can be further improved.
 また、壁部材16に微細穴17が設けられている場合の効果について説明する。図8は、図7中の破線で示す領域VIIIを拡大して示す図である。図8に示すように、壁部材16に設けられた微細穴17には溶液61が入り込まないようにして用いられ、微細穴17は空気層を構成する。これにより、壁部材16は、微視的には図8に示すように、壁部材層16aと空気層(微細穴17)との多重積層構造を有している。 Further, the effect when the fine hole 17 is provided in the wall member 16 will be described. FIG. 8 is an enlarged view of a region VIII indicated by a broken line in FIG. As shown in FIG. 8, the fine holes 17 provided in the wall member 16 are used so that the solution 61 does not enter, and the fine holes 17 constitute an air layer. Thereby, the wall member 16 has a multi-layered structure of the wall member layer 16a and the air layer (the minute hole 17) microscopically as shown in FIG.
 壁部材層16aの屈折率は1.6~1.9程度、検査用溶液61の屈折率は1.3~1.6程度の範囲であるのに対して、空気層の屈折率は略1.0である。すなわち、壁部材16と空気層との屈折率差は非常に大きいため、壁部材層16aと空気層との界面に入射した光は界面において少なくとも一部が反射される。壁部材層16aは空気層よりも屈折率が大きいため、壁部材16側から空気層に向かって臨界角以上の入射角θで界面Cに入射する光は全反射して、板状基材10の表面側に向かうこととなる。また、壁部材層16a側から界面Cに臨界角より小さい入射角θで入射した場合であっても、入射光の一部は界面Cで正反射して板状基材10の表面側に向かう。さらに、界面Cを透過した光は空気層から次の壁部材16に入射し、この空気層と壁部材16との界面Cで少なくとも一部が正反射されて、やはり板状基材10の表面側に向かう。このように、壁部材層16aと微細穴17による空気層とが交互に複数配置されていることから細孔14内で発生した光は空気層と壁部材層16aとの複数の界面で反射されて板状基材10の表面側へと向けて導光される。なお、図8においては一部の界面における反射のみを表しているが、実際には、光線が入射する(図8において光線が横切る)すべての界面において反射が生じる。 The wall member layer 16a has a refractive index of about 1.6 to 1.9, and the inspection solution 61 has a refractive index of about 1.3 to 1.6, whereas the refractive index of the air layer is about 1. .0. That is, since the refractive index difference between the wall member 16 and the air layer is very large, at least a part of the light incident on the interface between the wall member layer 16a and the air layer is reflected at the interface. Since the wall member layer 16a has a larger refractive index than the air layer, the light incident on the interface C 1 at an incident angle theta 1 larger than the critical angle from the wall member 16 side to the air layer is totally reflected, a plate-shaped base It will go to the surface side of the material 10. Further, even when the incident light is incident on the interface C 1 from the wall member layer 16 a side at an incident angle θ 2 smaller than the critical angle, a part of the incident light is regularly reflected by the interface C 1 and the surface of the plate-like substrate 10. Head to the side. Further, the light transmitted through the interface C 1 is incident on the next wall member 16 from the air layer, and at least a part of the light is regularly reflected at the interface C 2 between the air layer and the wall member 16. Head toward the surface side of the. As described above, since the plurality of wall layers 16a and the air layers formed by the fine holes 17 are alternately arranged, the light generated in the pores 14 is reflected at the plurality of interfaces between the air layer and the wall member layer 16a. Then, the light is guided toward the surface side of the plate-like substrate 10. Although FIG. 8 shows only reflection at some interfaces, in reality, reflection occurs at all interfaces where light rays enter (light rays cross in FIG. 8).
 このように、細孔14を構成する壁部材16に微細穴17による空気層を設けることにより、細孔14中で発生した光を板状基材10の表面に向けて集光させる機能が付与される。結果として、検査デバイス1の表面から出射する光量を増加させ、光取り出し効率の向上効果を奏する。なおここで、板状基材10の一方の面11、他方の面12を区別しない場合には、基材の表面(検査デバイスの表面)と称している。 As described above, by providing an air layer by the fine holes 17 on the wall member 16 constituting the pores 14, a function of condensing the light generated in the pores 14 toward the surface of the plate-like substrate 10 is provided. Is done. As a result, the amount of light emitted from the surface of the inspection device 1 is increased, and the light extraction efficiency is improved. Here, when one surface 11 and the other surface 12 of the plate-like base material 10 are not distinguished, they are referred to as the surface of the base material (the surface of the inspection device).
 以上の通り、本検査デバイス1では、領域20内において細孔14から生じた光を反射率の高いアルミニウムからなる枠壁18により反射して表面側に導光させることができるので、多重反射による光の減衰を抑制し、光取り出し効率を向上させることができる。さらに、壁部材16中に微細穴17を備えている場合には、壁部材16と空気層と多重積層構造により複数の界面で反射を生じさせて、デバイスの表面側に光を集光させることができるので、特許文献2に開示されているシリコンからなる枠を備えた場合と比較して大幅に光取り出し効率を向上させることができる。 As described above, in the present inspection device 1, the light generated from the pores 14 in the region 20 can be reflected by the frame wall 18 made of aluminum having high reflectivity and guided to the surface side. Light attenuation can be suppressed and light extraction efficiency can be improved. Furthermore, when the micro hole 17 is provided in the wall member 16, light is condensed on the surface side of the device by causing reflection at a plurality of interfaces by the wall member 16, the air layer, and the multi-layered structure. Therefore, the light extraction efficiency can be greatly improved as compared with the case where a frame made of silicon disclosed in Patent Document 2 is provided.
 次に、本検査デバイス1の作製方法について説明する。図9は、本検査デバイス1の作製工程を示す図である。 Next, a manufacturing method of the inspection device 1 will be described. FIG. 9 is a diagram illustrating a manufacturing process of the inspection device 1.
 まず、アルミニウム基材100を用意する(A1)。検査デバイス1において一方の面のアルミニウムからなる枠壁18となるべき部分および他方の面の全面にマスク102を形成する(A2)。マスクは、公知のリソグラフィー法やスクリーン印刷により形成することができる。例えば、特開2014-198353号公報に記載の方法を採用することができる。 First, an aluminum substrate 100 is prepared (A1). In the inspection device 1, a mask 102 is formed on a portion to be the frame wall 18 made of aluminum on one surface and the entire surface of the other surface (A2). The mask can be formed by a known lithography method or screen printing. For example, a method described in JP 2014-198353 A can be employed.
 その後、枠壁形成予定の部分にマスク102を備えたアルミニウム基材100を陽極酸化する。例えば、特開2015-132837号公報、特開2014-198453号公報等に記載の公知の陽極酸化方法を採用することができる。これによりアルミニウム基材100のマスク102を備えていない部分が陽極酸化されて陽極酸化アルミナ部104が形成される(A3)。 Thereafter, the aluminum base material 100 provided with the mask 102 at the part where the frame wall is to be formed is anodized. For example, known anodic oxidation methods described in JP-A-2015-132837 and JP-A-2014-198453 can be employed. As a result, the portion of the aluminum substrate 100 that is not provided with the mask 102 is anodized to form an anodized alumina portion 104 (A3).
 そして、陽極酸化により凸になった部分をフライス加工等の機械的手法で除去し、略平坦な面を形成すると共に、マスクを除去する(A4)。この陽極酸化アルミナ部104に細孔14を形成する(A5)。細孔14はドリル等を用いた機械的手法に形成することができる。あるいは、陽極酸化アルミナ部104の、細孔14を形成する部分以外にマスクを設け、化学エッチングにより細孔14を形成してもよい。
 以上の工程により、細孔14の内壁を構成するアルミナ壁部材16を有する領域20がアルミニウム枠壁18により囲まれた構成の板状基材10を作製することができる。
And the part which became convex by anodization is removed by mechanical methods, such as a milling process, while forming a substantially flat surface, a mask is removed (A4). The pores 14 are formed in the anodized alumina portion 104 (A5). The pores 14 can be formed by a mechanical method using a drill or the like. Alternatively, a mask may be provided in a portion other than the portion where the pore 14 is formed in the anodized alumina portion 104, and the pore 14 may be formed by chemical etching.
Through the above steps, the plate-like substrate 10 having a configuration in which the region 20 having the alumina wall member 16 constituting the inner wall of the pore 14 is surrounded by the aluminum frame wall 18 can be produced.
 なお、上記板状基材10の作製方法は、上記手順に限るものではなく、アルミニウム基材に、先に細孔14を設け、その後、枠壁形成部分にマスクを設けて、陽極酸化を行う手順でもよい。 The method for producing the plate-like substrate 10 is not limited to the above procedure, and anodization is performed by providing the aluminum substrate with pores 14 first, and then providing a mask at the frame wall forming portion. It may be a procedure.
 また、上記手順A3において形成される陽極酸化アルミナ部104はポーラスアルミナであり、多数の微細穴を有するので、そのうちの所望の微細穴に対してポワワイド処理を施すことにより、微細穴よりも径の大きな細孔14を形成することも可能である。ポワワイド処理としては、公知の手法を用いることができ、例えば、特開2014-198453号公報に記載されている方法を採用することできる。 In addition, the anodized alumina portion 104 formed in the above procedure A3 is porous alumina and has a large number of fine holes. By performing a wide process on the desired fine holes, the diameter is larger than that of the fine holes. Large pores 14 can also be formed. A known method can be used as the power wide processing, and for example, a method described in Japanese Patent Application Laid-Open No. 2014-198453 can be employed.
 なお、図3に示す検査デバイス1は、他方の面12がアルミニウムからなる下面部材19から構成されており、微細穴17は他方の面12側に貫通していない。図9のA5に示す板状基材10の破線で示す領域Xの拡大図を図10のA5に示す。図10に示すように、アルミニウム基材100を一方の面11側から陽極酸化すると、微細穴17と他方の面12との間にはバリア層101および基材素地であるアルミニウム基材100が残る。アルミニウム基材100を剥離することなく用いることにより、他方の面12側は閉じた穴とすることができる。また、このとき、基材素地部分によって下面部材19が構成される。 In the inspection device 1 shown in FIG. 3, the other surface 12 is composed of a lower surface member 19 made of aluminum, and the fine hole 17 does not penetrate to the other surface 12 side. An enlarged view of a region X indicated by a broken line of the plate-like substrate 10 shown in A5 of FIG. 9 is shown in A5 of FIG. As shown in FIG. 10, when the aluminum substrate 100 is anodized from the one surface 11 side, the barrier layer 101 and the aluminum substrate 100 that is the substrate substrate remain between the micro holes 17 and the other surface 12. . By using the aluminum substrate 100 without peeling, the other surface 12 side can be a closed hole. Moreover, the lower surface member 19 is comprised by the base-material base | substrate part at this time.
 なお、陽極酸化した部分をアルミニウム基材100から剥離することにより、図5に示した、他方の面12側に下面部材19を備えない検査デバイスを作製することができる。この場合、バリア層101を残すことにより、微細穴17は他方の面12側に貫通することなく、他方の面12側で閉じた非貫通孔とすることができる。 It should be noted that an inspection device that does not include the lower surface member 19 on the side of the other surface 12 shown in FIG. In this case, by leaving the barrier layer 101, the fine hole 17 can be formed as a non-through hole closed on the other surface 12 side without penetrating on the other surface 12 side.
 さらに、図10のA5に示すような微細穴17に対して封孔処理を行い、一方の面11における開口を塞ぐことが好ましい(A6)。この封孔処理としては、高温加圧水蒸気処理や沸騰水中での沸騰処理などが挙げられる。この処理により、表面のアルミナが水和物となり、アルミナの水和物層16bにより開口が略塞がれた状態となる。なお、図10のA5においては、完全に開口が塞がれた状態として示しているが、溶液の浸入を抑制できる程度に塞がれていればよく、アルミナの水和物層16bにより開口が完全に塞がれていなくても構わない。 Furthermore, it is preferable to perform a sealing process on the fine holes 17 as shown in A5 of FIG. 10 to close the opening on one surface 11 (A6). Examples of the sealing treatment include high-temperature pressurized steam treatment and boiling treatment in boiling water. By this treatment, the alumina on the surface becomes a hydrate, and the opening is substantially closed by the alumina hydrate layer 16b. In FIG. 10A5, the opening is shown as being completely blocked, but it is sufficient that the opening is blocked to the extent that the solution can be prevented from entering, and the opening is formed by the alumina hydrate layer 16b. It does not have to be completely blocked.
 なお、以上のようにして作製された板状基材10の細孔14の内壁面14aに被検物質と特異的に結合する特異的結合物質である捕捉物質を結合させる(固定化する)ことにより、検査デバイス1を作製することができる。
 細孔14の内壁面14aへの捕捉物質の固定方法は公知の方法を特に制限なく適用することができる。例えば、既述の特許文献2等に開示されている方法を用いることができる。
In addition, the capture substance which is a specific binding substance that specifically binds to the test substance is bound (immobilized) to the inner wall surface 14a of the pore 14 of the plate-like substrate 10 manufactured as described above. Thus, the inspection device 1 can be manufactured.
As a method for fixing the trapping substance to the inner wall surface 14a of the pore 14, a known method can be applied without any particular limitation. For example, the method disclosed in the above-described Patent Document 2 can be used.
 次に、本発明の検査デバイスを備えた検査装置および本発明の検査デバイスを用いた検査方法を説明する。 Next, an inspection apparatus provided with the inspection device of the present invention and an inspection method using the inspection device of the present invention will be described.
 図11は、本発明の一実施形態の検査装置50の構成を模式的に示す図である。本実施形態の検査装置50は、上記検査デバイス1と、この検査デバイス1の細孔14中に検査用溶液を供給する溶液供給部60と、検査デバイス1の一方の面11側に配置された光検出器70とを備えている。光検出器70は、検査デバイス1から出射される光を検出するものである。 FIG. 11 is a diagram schematically showing the configuration of the inspection apparatus 50 according to an embodiment of the present invention. The inspection apparatus 50 according to the present embodiment is disposed on the inspection device 1, the solution supply unit 60 that supplies the inspection solution into the pores 14 of the inspection device 1, and the one surface 11 side of the inspection device 1. And a photodetector 70. The light detector 70 detects light emitted from the inspection device 1.
 図12は溶液供給部60の概略構成を示す図である。
 溶液供給部60は、検査デバイス1の他方の面12側に設置される検査用溶液61を貯留する貯留部62と、貯留部62の上部に設置されて、貯留部62に貯留されている検査用溶液61を吸引するピペット部64と、このピペット部64上に設置される検査デバイス1の一方の面11側に配置される減加圧空間部66および減加圧空間部66の圧力を減圧もしくは加圧するためのポンプ68を備えている。
FIG. 12 is a diagram illustrating a schematic configuration of the solution supply unit 60.
The solution supply unit 60 stores the inspection solution 61 installed on the other surface 12 side of the inspection device 1, and the inspection installed in the upper part of the storage unit 62 and stored in the storage unit 62. The pipette part 64 for sucking the solution 61 for use, and the pressure of the depressurizing space part 66 and the depressurizing space part 66 arranged on the one surface 11 side of the inspection device 1 installed on the pipette part 64 are reduced. Alternatively, a pump 68 for pressurization is provided.
 ピペット部64の先端が検査用溶液61中に浸漬された状態で、ポンプ68により減加圧空間部66を減圧することにより検査用溶液61がピペット部64を介して検査デバイス1の細孔14内に供給される。なお、ポンプ68による減加圧空間部66の減圧および加圧により、検査デバイス1の細孔14への検査用溶液61の供給および、細孔14からの検査用溶液61の排出を行うことができる。 In a state where the tip of the pipette part 64 is immersed in the inspection solution 61, the reduced pressure space 66 is decompressed by the pump 68, whereby the inspection solution 61 passes through the pipette part 64 and the pore 14 of the inspection device 1. Supplied in. Note that the inspection solution 61 is supplied to the pores 14 of the inspection device 1 and the inspection solution 61 is discharged from the pores 14 by depressurization and pressurization of the reduced pressure space 66 by the pump 68. it can.
 ここで、検査用溶液61は、光検出時に細孔14中に供給される溶液であるが、溶液供給部60は、検査工程において細孔14中に供給すべき検体液、標識溶液、洗浄液等の供給にも用いられる。すなわち、溶液供給部60は検査の工程毎に必要な溶液を細孔14に供給するものである。なお、壁部材16に備えられている微細穴17(図12においては図示していない。)は、他方の面12側が非開口であるため、供給される溶液は他方の面12側から微細穴17には浸入しない。そして、既述の空気層を備えた効果を得るために、微細穴17が開口されている一方の面11からも溶液が微細穴17中に浸入しないように溶液供給部60は溶液の供給を制御する。なお、微細穴17の一方の面11側の開口に対して既述の封孔処理が施されていれば、一方の面11からも溶液は微細穴17には入り込まないため、溶液の供給制御が容易になり好ましい。 Here, the test solution 61 is a solution supplied into the pores 14 at the time of light detection, but the solution supply unit 60 includes a sample solution, a label solution, a cleaning solution, and the like to be supplied into the pores 14 in the test process. It is also used for supply. That is, the solution supply unit 60 supplies a necessary solution to the pores 14 for each inspection process. Note that the fine holes 17 (not shown in FIG. 12) provided in the wall member 16 are non-open on the other surface 12 side, so that the supplied solution is fine holes from the other surface 12 side. 17 does not enter. In order to obtain the effect provided with the air layer described above, the solution supply unit 60 supplies the solution so that the solution does not enter the fine hole 17 from the one surface 11 where the fine hole 17 is opened. Control. If the above-described sealing treatment is applied to the opening on the one surface 11 side of the fine hole 17, the solution does not enter the fine hole 17 from the one surface 11. Is preferable.
 以下、上記検査装置50を用いた本発明の検査方法について説明する。
 本発明において検査デバイスから出射され、光検出器により検出される光は、例えば、被検物質に付与された標識が励起されて生じる蛍光、もしくは被検物質に特異的に結合する抗体などの結合物質に、上記と同様の標識を付与し、標識から生じる光、あるいは被検物質に特異的に結合する抗体などの結合物質に酵素を標識し、この酵素を触媒とする反応による発光(以下において、「化学発光」という。)など、標識に起因する光信号である。なお、被検物質が自家蛍光を生じるものであれば、標識は不要であり、自家蛍光を検出すればよい。また、ここで光信号とは、蛍光、化学発光光のほか、吸光度(比色)を含むものとする。
Hereinafter, the inspection method of the present invention using the inspection apparatus 50 will be described.
In the present invention, the light emitted from the test device and detected by the photodetector is, for example, fluorescence generated by excitation of a label attached to the test substance, or binding of an antibody that specifically binds to the test substance. The substance is given the same label as above, and light is emitted from the label, or an enzyme is labeled on a binding substance such as an antibody that specifically binds to the test substance, and light emission by the reaction using this enzyme as a catalyst (in the following) , Or “chemiluminescence”)). In addition, if the test substance generates autofluorescence, no label is necessary, and autofluorescence may be detected. Here, the optical signal includes absorbance (colorimetric) in addition to fluorescence and chemiluminescence light.
 図13は検査工程を模式的に示す図である。
 検査デバイス1の細孔14の内壁面14aにはアレルゲンなどの捕捉物質30が固定されている(S1)。この検査デバイス1の細孔14に被検物質(例えば、上記アレルゲンと特異的に結合する特異的IgE抗体)を含む検体液を供給して、被検物質32を捕捉物質30に結合させる(S2)。検体液の供給には、上述の溶液供給部60を用いる。貯留部62に検体液を貯留させた状態で、ポンプを動作させてピペット部64による検体液の吸引および排出を繰り返すことにより、検体液を細孔14内の捕捉物質30に効率よく接触させることができる。
FIG. 13 is a diagram schematically showing the inspection process.
A capturing substance 30 such as an allergen is fixed to the inner wall surface 14a of the pore 14 of the inspection device 1 (S1). A specimen liquid containing a test substance (for example, a specific IgE antibody that specifically binds to the allergen) is supplied to the pores 14 of the test device 1, and the test substance 32 is bound to the capture substance 30 (S2). ). The above-described solution supply unit 60 is used for supplying the sample liquid. The sample liquid is efficiently brought into contact with the capture substance 30 in the pores 14 by repeatedly pumping and discharging the sample liquid by the pipette unit 64 while the sample liquid is stored in the storage unit 62. Can do.
 次に、検体液を排出した後、被検物質32と特異的に結合する物質33(例えば、二次抗体)に標識Fが付与されてなる標識物質35を含む標識溶液を細孔14に供給して、被検物質32に標識物質35を結合させる(S3)。標識溶液の細孔14へ供給は上記検体液の供給と同様に溶液供給部60により行う。 Next, after discharging the sample liquid, a labeled solution containing a labeled substance 35 in which a label F is added to a substance 33 (for example, a secondary antibody) that specifically binds to the test substance 32 is supplied to the pores 14. Then, the labeling substance 35 is bound to the test substance 32 (S3). The label solution is supplied to the pores 14 by the solution supply unit 60 in the same manner as the sample solution.
 なお、上記測定方法においては、被検物質32の捕捉物質30への結合の後に、標識物質35を被検物質32に結合させているが、検査装置50外において、あらかじめ検体液と標識溶液を混合して被検物質32と標識物質35を結合させ、その混合液を細孔14内に供給するようにしてもよい。この場合、混合液の細孔14への供給により、細孔14内に固定された捕捉物質30に、標識物質35が結合された被検物質32を結合させることができる。 In the measurement method described above, the labeling substance 35 is bound to the test substance 32 after the test substance 32 is bound to the capture substance 30. It is also possible to combine the test substance 32 and the labeling substance 35 by mixing and supply the mixed liquid into the pores 14. In this case, by supplying the mixed liquid to the pores 14, the test substance 32 to which the labeling substance 35 is bound can be bound to the capture substance 30 fixed in the pores 14.
 そして、標識溶液もしくは混合液を排出した後、検体液の供給と同様の方法で、溶液供給部60を用いて細孔14に洗浄液を供給して、細孔14中に非特異吸着している被検物質32や標識物質35を除去する。 Then, after discharging the labeling solution or the mixed solution, the cleaning solution is supplied to the pores 14 using the solution supply unit 60 in the same manner as the supply of the sample solution, and nonspecific adsorption is performed in the pores 14. The test substance 32 and the labeling substance 35 are removed.
 最後に、洗浄液を排出して、細孔14内にバッファ溶液等の検査用溶液を充填させた状態で、標識Fを触媒として反応する発光反応からの光信号を光検出器70により検出する(S4)。なお、光検出器70による検査デバイス1からの出射光の検出時、壁部材16が透明であり、かつ周囲に枠壁18を備えた作用により、また、壁部材16に設けられている微細穴17が空気層を構成することにより、光取り出し効率よく光検出を行うことができる。 Finally, the cleaning liquid is discharged, and the optical signal from the luminescence reaction that reacts with the label F as a catalyst is detected by the photodetector 70 with the inspection solution such as the buffer solution filled in the pores 14 ( S4). Note that when the light emitted from the inspection device 1 is detected by the photodetector 70, the wall member 16 is transparent and has a frame wall 18 around it. Since the air layer 17 constitutes an air layer, light detection can be performed efficiently.
 標識Fが蛍光色素、量子ドットなどの蛍光標識である場合には、バッファ溶液を検査用溶液として細孔14内に供給し、細孔14内をバッファ溶液で満たした状態で、蛍光測定を行う。具体的には、蛍光標識を励起する波長の光を励起光として検査デバイスに照射し、その励起光により励起された標識からの蛍光を検出する。なお、蛍光標識の検出の際に用いられる光検出器70には、励起光照射部が備えられている。 When the label F is a fluorescent label such as a fluorescent dye or a quantum dot, the buffer solution is supplied into the pore 14 as a test solution, and the fluorescence measurement is performed with the pore 14 filled with the buffer solution. . Specifically, the inspection device is irradiated with light having a wavelength that excites the fluorescent label as excitation light, and fluorescence from the label excited by the excitation light is detected. The photodetector 70 used for detecting the fluorescent label is provided with an excitation light irradiation unit.
 一方、標識Fが、ルミノールが過酸化水素により酸化されて発光を生じる化学反応の触媒として作用する酵素等である場合には、上記洗浄処理の後、触媒として作用する酵素と接触させることによる化学反応で発光する物質を含む反応液を検査用溶液として細孔14中に供給する。そして、光検出器70では、細孔14にこの反応液を供給にすることにより、細孔14の捕捉物質30に捕捉された被検物質32に付与された化学発光標識が反応液中の物質と反応して生じる発光を検出する。この場合、反応液により細孔14が満たされた状態で発光検出を行う。 On the other hand, when the label F is an enzyme or the like that acts as a catalyst for a chemical reaction in which luminol is oxidized by hydrogen peroxide to generate luminescence, the chemical by contacting with the enzyme that acts as a catalyst after the washing treatment. A reaction solution containing a substance that emits light by the reaction is supplied into the pores 14 as a test solution. In the photodetector 70, the reaction solution is supplied to the pores 14 so that the chemiluminescent label attached to the test substance 32 captured by the capture material 30 in the pores 14 is a substance in the reaction solution. Luminescence generated by reaction with is detected. In this case, luminescence detection is performed in a state where the pores 14 are filled with the reaction solution.
 化学発光を生じる酵素標識としては、ルミノール、ロフィン、ルシゲニンおよびシュウ酸エステルなどの化学発光基質と反応する酵素などが挙げられる。HRP(西洋わさびペルオキシダーゼ)酵素が標識として用いられる場合には、HRPが触媒として機能するルミノール系の化学発光基質を含有する反応液(ルミノール反応液)を、ALP(アルカリホスファターゼ)酵素が用いられる場合にはジオキセタン系化学発光基質を含有する反応液を用いることが好ましい。 Examples of enzyme labels that generate chemiluminescence include enzymes that react with chemiluminescent substrates such as luminol, lophine, lucigenin and oxalate. When HRP (horseradish peroxidase) enzyme is used as a label, a reaction liquid (luminol reaction liquid) containing a luminol-based chemiluminescent substrate in which HRP functions as a catalyst is used, and an ALP (alkaline phosphatase) enzyme is used. It is preferable to use a reaction solution containing a dioxetane chemiluminescent substrate.
 ルミノール反応液には、少なくともルミノール基質と過酸化水素水が含まれる。酵素標識は、過酸化水素水存在下において、ルミノールの酸化を触媒するものである。反応液中には、化学発光を増感する増感剤を含むことが好ましい。 The luminol reaction solution contains at least a luminol substrate and a hydrogen peroxide solution. The enzyme label catalyzes the oxidation of luminol in the presence of hydrogen peroxide. The reaction solution preferably contains a sensitizer that sensitizes chemiluminescence.
 なお、酵素標識を用いた光検出においては、上記の化学発光基質のみならず、発光基質あるいは蛍光基質を含む反応液を用い、呈色(吸光)反応や、蛍光を検出するものとすることもできる。 In light detection using an enzyme label, not only the chemiluminescent substrate described above but also a reaction solution containing a luminescent substrate or a fluorescent substrate may be used to detect a color (absorption) reaction or fluorescence. it can.
 いずれの検出方法であっても、本発明の検査デバイスを用いることにより、細孔中で生じた光は検査デバイスの表面から効率よく出射され、光の取り出し効率が高いので、高精度な光検出が可能である。 Regardless of the detection method, by using the inspection device of the present invention, the light generated in the pores is efficiently emitted from the surface of the inspection device, and the light extraction efficiency is high. Is possible.
 1  検査デバイス
 10 板状基材
 11 板状基材(検査デバイス)の一方の面
 12 板状基材(検査デバイス)の他方の面
 14 細孔
 14a 細孔の内壁面
 16 壁部材
 16a 壁部材層
 16b 水和物層
 17 微細穴
 17a 閉じた空洞部
 18 枠壁
 19 下面部材
 20 検査領域
 30 捕捉物質
 32 被検物質
 33 被検物質に特異的に結合する物質
 35 標識物質
 50 検査装置
 60 溶液供給部
 61 検査用溶液
 62 貯留部
 64 ピペット部
 66 減加圧空間部
 68 ポンプ
 70 光検出器
 100 アルミニウム基材
 101 バリア層
 102 マスク
 104 陽極酸化アルミナ部
 F 標識
DESCRIPTION OF SYMBOLS 1 Inspection device 10 Plate-like base material 11 One side of plate-like base material (inspection device) 12 Other side of plate-like base material (inspection device) 14 Pore 14a Inner wall surface of pore 16 Wall member 16a Wall member layer 16b Hydrate layer 17 Fine hole 17a Closed cavity 18 Frame wall 19 Lower surface member 20 Inspection region 30 Captured substance 32 Test substance 33 Substance specifically binding to test substance 35 Labeling substance 50 Inspection apparatus 60 Solution supply part 61 Test Solution 62 Reservoir 64 Pipette 66 Reduced Pressurization Space 68 Pump 70 Photodetector 100 Aluminum Base 101 Barrier Layer 102 Mask 104 Anodized Alumina Part F Label

Claims (13)

  1.  板状基材の一方の面から他方の面に貫通する複数の細孔を備えた検査デバイスであって、
     少なくとも一部の細孔を含む領域であって、該少なくとも一部の細孔の内壁面を構成する壁部材の少なくとも一部がアルミナからなる領域と、前記一方の面から見た場合に、該領域を囲むアルミニウムからなる枠壁とを有し、
     前記少なくとも一部の細孔の内壁面に特定の物質と特異的に結合する捕捉物質が固定されている検査デバイス。
    An inspection device having a plurality of pores penetrating from one surface of a plate-like substrate to the other surface,
    A region including at least a part of the pores, and at least a part of the wall member constituting the inner wall surface of the at least a part of the pores when viewed from the one surface, A frame wall made of aluminum surrounding the region,
    An inspection device in which a capture substance that specifically binds to a specific substance is fixed to an inner wall surface of the at least some pores.
  2.  前記壁部材が、前記細孔と平行に伸び、かつ、該細孔より小さい直径の微細穴を有する請求項1記載の検査デバイス。 The inspection device according to claim 1, wherein the wall member has a fine hole extending in parallel with the pore and having a diameter smaller than the pore.
  3.  前記壁部材は、陽極酸化アルミナからなる請求項1または2記載の検査デバイス。 3. The inspection device according to claim 1, wherein the wall member is made of anodized alumina.
  4.  前記枠壁が前記壁部材と接して設けられている請求項1から3のいずれか1項に記載の検査デバイス。 The inspection device according to any one of claims 1 to 3, wherein the frame wall is provided in contact with the wall member.
  5.  前記他方の面がアルミニウムからなる請求項1から4のいずれか1項に記載の検査デバイス。 5. The inspection device according to claim 1, wherein the other surface is made of aluminum.
  6.  前記捕捉物質が、抗原、抗体またはデオキシリボ核酸である請求項1から5のいずれか1項に記載の検査デバイス。 6. The inspection device according to claim 1, wherein the capture substance is an antigen, an antibody, or deoxyribonucleic acid.
  7.  前記アルミニウムからなる枠壁により囲まれた前記領域を複数備え、
     該複数の領域の1つの領域中の細孔の内壁面には前記捕捉物質として同一種の捕捉物質が結合されており、
     前記複数の領域は、前記結合されている捕捉物質が互いに異なる領域を有する請求項1から6いずれか1項に記載の検査デバイス。
    A plurality of the regions surrounded by the frame wall made of aluminum,
    A trapping substance of the same type as the trapping substance is bound to the inner wall surface of the pore in one of the plurality of areas,
    The inspection device according to any one of claims 1 to 6, wherein the plurality of regions include regions in which the combined capture substances are different from each other.
  8.  前記細孔の前記一方の面における開口領域の円相当直径は1μm~100μmである請求項1から7のいずれか1項に記載の検査デバイス。 The inspection device according to any one of claims 1 to 7, wherein a circle-equivalent diameter of an opening region on the one surface of the pore is 1 µm to 100 µm.
  9.  前記板状基材の厚さは100μm~2000μmである請求項1から8のいずれか1項に記載の検査デバイス。 The inspection device according to any one of claims 1 to 8, wherein the thickness of the plate-like substrate is 100 µm to 2000 µm.
  10.  請求項1から9のいずれか1項に記載の検査デバイスと、
     前記検査デバイスの前記細孔中に検査用溶液を供給する溶液供給部と、
     前記検査デバイスの前記一方の面側に配置され、前記検査デバイスから出射される光を検出する光検出器とを備えた検査装置。
    The inspection device according to any one of claims 1 to 9,
    A solution supply unit for supplying a test solution into the pores of the test device;
    An inspection apparatus including a photodetector that is disposed on the one surface side of the inspection device and detects light emitted from the inspection device.
  11.  請求項1から9のいずれか1項に記載の検査デバイスの前記少なくとも一部の細孔に、前記特定の物質を含有する検体液を供給して、前記特定の物質を前記捕捉物質に結合させ、
     前記特定の物質と特異的に結合する標識物質を前記特定の物質に結合させ、
     前記細孔に検査用溶液を供給して該細孔に前記検査用溶液を留めた状態で、前記検査デバイスから出射される光を検出する検査方法。
    A specimen liquid containing the specific substance is supplied to the at least some of the pores of the inspection device according to any one of claims 1 to 9, and the specific substance is bound to the capture substance. ,
    Binding a labeling substance that specifically binds to the specific substance to the specific substance;
    An inspection method for detecting light emitted from the inspection device in a state where an inspection solution is supplied to the pores and the inspection solution is retained in the pores.
  12.  前記標識物質として酵素標識を含む物質を用い、
     前記検査用溶液として、前記酵素標識により触媒されて反応する基質を含む反応液を用い、
     前記出射される光として、前記反応液中の前記基質が前記酵素標識により触媒されて生じる光を検出する請求項11記載の検査方法。
    Using a substance containing an enzyme label as the labeling substance,
    As the test solution, a reaction solution containing a substrate that is catalyzed by the enzyme label and reacts,
    The inspection method according to claim 11, wherein as the emitted light, light generated when the substrate in the reaction solution is catalyzed by the enzyme label is detected.
  13.  前記標識物質として蛍光標識を含む物質を用い、
     該蛍光標識を励起させる励起光を前記検査デバイスに照射し、
     前記出射される光として、前記励起光の照射により前記標識物質から生じる蛍光を検出する請求項11記載の検査方法。
    Using a substance containing a fluorescent label as the labeling substance,
    Irradiating the inspection device with excitation light for exciting the fluorescent label;
    The inspection method according to claim 11, wherein fluorescence emitted from the labeling substance by irradiation with the excitation light is detected as the emitted light.
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