WO2017169714A1 - Inspection device, inspection apparatus and inspection method - Google Patents

Inspection device, inspection apparatus and inspection method Download PDF

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Publication number
WO2017169714A1
WO2017169714A1 PCT/JP2017/010091 JP2017010091W WO2017169714A1 WO 2017169714 A1 WO2017169714 A1 WO 2017169714A1 JP 2017010091 W JP2017010091 W JP 2017010091W WO 2017169714 A1 WO2017169714 A1 WO 2017169714A1
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Prior art keywords
pores
inspection device
inspection
substance
substrate
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PCT/JP2017/010091
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French (fr)
Japanese (ja)
Inventor
孝明 伊藤
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富士フイルム株式会社
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Publication of WO2017169714A1 publication Critical patent/WO2017169714A1/en

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    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
    • G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
    • G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
    • G01N21/64—Fluorescence; Phosphorescence
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
    • G01N21/75—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated
    • G01N21/77—Systems 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
    • G01N21/78—Systems 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
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
    • G01N33/543—Immunoassay; Biospecific binding assay; Materials therefor with an insoluble carrier for immobilising immunochemicals

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 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.
  • the outermost surface material (first layer) of the pore wall is made of a material having a refractive index larger than the refractive index of the lower layer (second layer).
  • a laminated structure has been proposed. With this laminated structure, the light reflectance on the inner wall surface can be improved, and as a result, the light detection efficiency can be improved.
  • Patent Document 1 it is possible to improve the light extraction efficiency accompanying an increase in the light aperture ratio, but the light incident on the silicon wall is also attenuated because the reflectance of silicon used as the frame wall is low. Therefore, the extraction efficiency of light incident on the frame wall is not improved.
  • 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.
  • Patent Document 2 discloses a specific device manufacturing method in which holes are formed by electrochemical etching in a support constituting the second layer, and the first layer is formed by CVD (Chemical Vapor Deposition). Has been.
  • CVD Chemical Vapor Deposition
  • material restrictions are large, the second layer is substantially limited to a silicon-based material, and the first layer is limited to a material that can be formed by CVD film formation. May be high.
  • 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 cavity formed around the region extending in the depth direction of the pores, not penetrating at least the other surface, and including at least some of the plurality of pores when viewed from one surface.
  • Have The portion of the plate-like base material constituting the region including at least a part of the pores has light transmittance,
  • This is a test device in which a capture substance that specifically binds to a specific substance is fixed to the inner wall surface of at least some of the pores.
  • the plate-like substrate is preferably made of SiO 2 or Al 2 O 3 (alumina).
  • a plurality of pores are arranged in a plurality of regions partitioned by the cavity, and one of the plurality of regions It is preferable that the same kind of trapping substance as the trapping substance is bonded to the inner wall surface of each of the pores, and the plurality of regions have areas where the trapping substances are different from each other.
  • the hollow portion may be formed so as to extend from one surface of the plate-like substrate to the front of the other surface, and may be non-penetrating on the other surface.
  • the inspection device of the present invention is a member whose material is different from that of the plate-like substrate in the opening of the other surface side of the through hole in which the hollow portion is provided so as to penetrate from one surface of the plate-like substrate to the other surface. May be blocked by the other surface and non-penetrated on the other surface.
  • the capture substance is preferably an antigen, an antibody, or deoxyribonucleic acid (DNA).
  • 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 light 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 labeling substance, a reaction solution containing a substrate that is catalyzed by the enzyme label and reacts as the test solution, and the light emitted from the inspection device
  • the light produced when the substrate in the reaction solution is catalyzed by the enzyme label may be detected.
  • 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.
  • An inspection device of the present invention is an inspection device having a plurality of pores penetrating from one surface of a plate-like substrate to the other surface, extending in the depth direction of the pores, and non-existing on at least the other surface. It is a penetration and has a cavity formed around a region including at least some of the plurality of pores when viewed from one side, and constitutes a region including at least some of the pores Since the plate-like base material portion is light transmissive, it is possible to improve the extraction efficiency of light generated in the pores to at least one surface side of the inspection device. In addition, since the base material can be made of a single material, it can be manufactured at low cost.
  • FIG. 1 is a perspective view of an inspection device according to a first embodiment of the present invention. It is a top view of the inspection device shown in FIG. It is sectional drawing of the test
  • 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 extends in the depth direction of the pores 14 on the plate-like substrate 10 having a plurality of pores 14 penetrating from one surface 11 to the other surface 12, and at least the other surface. 12 has a cavity 16 formed around a region including at least some of the plurality of pores 14 when viewed from one surface 11.
  • a total of nine pores 14 in the vertical and horizontal directions are used as one region 20, and the cavity 16 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.
  • the inspection device 1 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.
  • a cavity is formed around a region including at least a part of the pores. That is, in the inspection device, it is sufficient that a pore region surrounded by the cavity is formed at least in part. Further, in the region surrounded by the cavity, the plate-like base material constituting the pores and the cavity, that is, the pores and the wall constituting the cavity have light transmittance.
  • the entire plate-like substrate may be made of a light transmissive material, or only the region surrounded by the cavity may be made of a light transmissive material.
  • having light transmittance means that the transmittance of detection light described later is 50% or more. Specifically, the average transmittance in the visible light wavelength range of 400 to 700 nm is 50% or more. If there is, it will be regarded as having light transparency.
  • the plate-like base material is preferably made of SiO 2 or Al 2 O 3 . If the whole plate-shaped base material is comprised from the same material, preparation will be easy and it is especially preferable, but the plate-shaped base material may be provided with the part which consists of a different material. For example, only the region surrounded by the cavity is made of SiO 2 and the other part is made of Si, or only the region surrounded by the cavity is made of Al 2 O 3 and the other part is made of Al. A substrate or the like may be used.
  • the cavity 16 is a space that is open on one surface 11 side and closed on the other surface 12 side.
  • the hollow portion 16 is formed to extend from one surface 11 of the plate-like base material 10 to the front of the other surface 12, and an opening connected to the hollow portion 16 is provided on the other surface 12 of the plate-like base material 10. (See FIG. 4).
  • “formed to extend to the front” means that it extends toward the other surface 12 but does not reach the other surface 12.
  • the length of the cavity 16 extending from the one surface 11 to the other surface 12 side, that is, the depth along the depth direction of the pores 14 is at least half the thickness of the plate-like substrate 10. Is more preferable, and more preferably 3/4 or more.
  • 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.
  • in the region 20 surrounded by the cavity 16 hereinafter referred to as “inspection region 20”
  • 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).
  • acquisition substance 30 specifically couple
  • the test device 1 is suitable for an allergy test provided with an allergen that is a kind of antigen as a capture substance.
  • a plurality of pores 14 are arranged in a plurality of inspection regions 20 partitioned by a cavity 16 as shown in a view of the inspection device 1 viewed from one surface 11.
  • a trapping substance of the same type is bound (fixed) to the pores 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. Note that there may be two or more regions containing the same type of capture substance between a plurality of inspection regions.
  • the same kind of allergen that is a capture substance is fixed to one or a plurality of examination areas 20, and a plurality of allergen examination areas are provided in one examination device. Thereby, the reaction with respect to a several allergen can be test
  • the cavity 16 is provided over the entire circumference of the inspection region 20, but the cavity 16 does not have to be provided over the entire circumference.
  • the total length S 1 of the portions facing the inspection region 20 of the opening of the cavity 16 disposed around the inspection region 20 is the inspection region 20.
  • the “cavity formed around the region” means a cavity arranged so as to satisfy S 1 ⁇ S 2 .
  • S 2 is , is preferably 1/4 or more the length S 3 of the pores outer peripheral edge, 1/2 or more preferably of S 3, and particularly preferably S 3 or more.
  • one side of the plurality of pores 14 arranged on the outermost periphery among the plurality of pores 14 in the inspection region 20 is a 0 , and the cavity portion 16 of the pore 14 arranged on the outermost periphery.
  • the total length S 2 12a 0 of the portions facing the surface.
  • a pore outer peripheral edge 22 shown by a broken line in the drawing
  • the length S 3 of the outer circumferential edge 22 of the pore is 4a.
  • A>a> 3a 0 S 1 ⁇ S 2 is satisfied.
  • FIG. 5 is a plan view of the inspection device 2 of the second embodiment in which the shape of the cavity is different from that of the inspection device 1 of the first embodiment.
  • the inspection region 20 including the plurality of pores 14 is configured by a plurality (here, four) of isolated rectangular cavities 17.
  • the inspection region 20 including the plurality of pores 14 is the same as that shown in FIG. 2, and the cavity 17 of the plurality of pores 14 arranged on the outermost periphery among the plurality of pores 14 in the inspection region 20.
  • the length of the portion of the opening of one cavity portion 17 facing the inspection region 20 is the length B of the long side of the rectangular cavity portion 17 and faces the inspection region 20 for the four cavity portions 17.
  • the opening shape of the pores 14 and the opening shape of the hollow portion 17 are not limited to a rectangle.
  • FIG. 6 shows a case where the pore 114 and the cavity 116 both have a circular opening, and the region 120 including the plurality of pores 114 is surrounded by the plurality of cavities 116.
  • S 1 , S 2 and S 3 are defined as follows.
  • the outer peripheral edge 122 of the pores is represented by a straight line in contact with the pores 114 arranged on the outermost periphery among the plurality of pores 114 and a line passing through the edges of some of the pores 114. .
  • the length of the portion of the pore 114 facing the cavity 116 disposed on the outer periphery (outermost periphery) 122 of the pore is the length of the arc facing the cavity 116 in the circumference of the opening of each pore 114. In this example, it is a semicircle or 3/4 circular arc portion indicated by a thick line in FIG.
  • the lengths of the portions of the openings of the plurality of cavities 116 arranged so as to surround the region 120 facing the inspection region 120 are the lengths of the arcs facing the inspection region 120 in the circumference of the openings of the cavities 116, respectively. In this example, it is a semicircle or a quarter of a circular arc indicated by a thick line in FIG.
  • the pores in the inspection region can be defined in the same manner as in FIG.
  • FIG. 7 is a plan view of the inspection device 3 according to the third embodiment
  • FIG. 8 is a cross-sectional view taken along line VIII-VIII of the inspection device 3 of FIG.
  • the pores 14 and the cavities 18 are alternately arranged vertically and horizontally.
  • one pore 14 is arranged in an inspection region 28 surrounded by four cavities 18.
  • the pore 14 and the cavity 18 have the same opening shape.
  • the pore 14 is a through hole, and the cavity 18 is provided as a blind hole that does not penetrate the other surface 12 of the plate-like substrate 10.
  • FIG. 9 shows a design change example.
  • the cavity 118 is formed on the plate-like substrate 10 after the through-hole 40 penetrating from one surface 11 to the other surface 12 is provided, and then on the other surface 12 side.
  • the opening may be formed so as to be closed with a closing member 42 made of a material different from that of the plate-like substrate 10. That is, in FIG. 9, the cavity 118 is constituted by the through hole 40 and the closing member 42 provided in the plate-like base material 10.
  • the cavity 118 is made of a material whose opening on the one surface 11 side of the plate-like substrate 10 is different from that of the plate-like substrate 10.
  • the second closing member 44 may be closed. That is, the cavity 118 may be a closed space.
  • the inside of the cavity may be a vacuum, or a gas such as air having a refractive index of approximately 1.0 may be enclosed.
  • the material constituting the closing member 42 and the second closing member 44 constituting the hollow portion 118 is not particularly limited, but it can be configured as a closing member by being hardened after being dropped by an ink jet device, such as a metal or a resin material. Material is preferred.
  • the inspection device including one pore in the inspection region surrounded by the cavity has been described.
  • a plurality of pores are included in the inspection region surrounded by the cavity. It is more preferable to have.
  • FIG. 11 is an enlarged schematic view of a part of the inspection device 1.
  • a test substance is bound to a capture substance (not shown in FIG. 11) fixed to the inner wall surface 14a of the pore 14, and the test substance is further labeled with an enzyme. It is used in a test method for detecting the presence or absence of a test substance by detecting the chemiluminescence using a labeling substance as a catalyst by binding the substance or detecting the presence and quantity of the 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. In addition, it uses so that the solution 61 may not enter into the cavity part 16, and the cavity part 16 comprises an air layer.
  • the base material forming the pores 14 in the inspection region 20 has light transmittance, the surface of the light generated in the pores 14 is compared with the pores 14 formed of the base material impermeable to silicon.
  • the exit aperture ratio increases.
  • the refractive index of the light-transmitting plate-like substrate 10 and the refractive index of the test solution are equivalent in the range 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 plate-like substrate 10 and the air layer is very large, a large reflection occurs at the interface.
  • the plate-like substrate 10 since the plate-like substrate 10 has a higher refractive index than the air layer, light incident on the interface C 1 at an incident angle ⁇ 1 greater than the critical angle from the plate-like substrate 10 side toward the air layer is totally reflected. Then, it goes to the surface side of the plate-like substrate 10. Even when the incident light is incident on the interface C 1 from the plate-like substrate 10 side at an incident angle ⁇ 2 smaller than the critical angle, a part of the incident light is specularly reflected at the interface C 1 and It will go to the surface side.
  • the light transmitted through the interface C 1 is incident on the plate-like base material 10 constituting the cavity 16 from the air layer, and at least a part of the light is regularly reflected at the interface C 2 , and the surface of the plate-like base material 10 is again formed. Head to the side.
  • a function of condensing light generated in the pores 14 toward the surface of the plate-like substrate 10 is provided.
  • the amount of light emitted from the surface of the inspection device 1 is increased, and the light extraction efficiency is improved.
  • the surface of the base material the surface of the inspection device.
  • the light amount can be reflected at the interface between the plate-like base material and the air layer with almost no attenuation, and the light can be condensed on the surface side of the device.
  • the light extraction efficiency can be greatly improved as compared with the case where a frame made of silicon is provided.
  • a plate-like substrate made of SiO 2 having a plurality of pores and cavities can be produced as follows. First, a plate-like substrate made of Si is prepared, and pores and cavities are formed from one surface side by wet etching or dry etching. For example, a microporous Si manufacturing process known in MEMS (Micro Electro Mechanical System) technology can be used. In the case where the cavity and the pore as shown in FIG. 2 or FIG. 5 are manufactured, when the pore is formed by etching, a mask is provided outside the pore formation region and the cavity is formed by etching. In this case, the mask may be formed sequentially with a mask other than the cavity forming region. The cavity can be formed by etching from one side and stopping the etching before the other side.
  • MEMS Micro Electro Mechanical System
  • the pore and the cavity may be etched at the same time. Since the etching speed of the cavity portion having a small diameter is slower than the etching speed of the pore portion, the pore portion penetrates the other surface, but the cavity portion can be left unpenetrated.
  • a large cavity can be formed by etching in the lateral direction so as to connect the holes.
  • the whole base material can be made transparent by changing the base material made of Si having pores and cavities formed as described above to SiO 2 by oxidation treatment.
  • oxidation treatment for example, thermal oxidation treatment is performed at a temperature of 1100 ° C. for a predetermined time.
  • the thermal oxidation treatment time may be appropriately adjusted according to the size and shape of the plate-like substrate and the desired oxidation treatment region.
  • the cavity 118 is constituted by the through hole 40 and the closing member 42 as shown in FIG. 9, first, the through hole 40 constituting a part of the cavity is formed by etching simultaneously with the pore 14. . Thereafter, the Si base material is changed to SiO 2 by the above thermal oxidation treatment, and then the opening on the other surface side of the through hole 40 constituting a part of the cavity is closed with a closing member 42 made of a material different from the base material. . As a result, the cavity 118 with the other surface closed can be formed.
  • the blocking member 42 only needs to be able to block the opening on the other surface side of the through hole 40, and may be formed so as to partially enter the through hole 40.
  • the closing member 42 can be formed, for example, by dripping a metal or a resin material so as to close the opening on the other surface side of the through hole using an ink jet device, and solidifying or curing.
  • a plate-like substrate made of Al 2 O 3 having a plurality of pores and cavities can be produced as follows.
  • a base material made of Al By preparing a base material made of Al and subjecting this base material to known anodization treatment and power wide treatment described in, for example, JP-A-2015-132737, JP-A-2014-198453, etc., A plate-like substrate made of porous alumina can be produced.
  • an Al base material is anodized to form a porous alumina coating.
  • a pore having a desired diameter is formed by performing a pore wide process to widen the diameter of the micropore.
  • the pores used as the cavity may be provided with a blocking member that covers the opening on the other surface side as in the case of the device made of SiO 2. .
  • the trapping substance which is a specific binding substance that specifically binds to the test substance, is fixed to the inner wall surface of the pores of the plate-like substrate having a plurality of pores and cavities produced as described above.
  • an inspection device can be manufactured.
  • a method for fixing the trapping substance to the inner wall surface of the pore a known method can be applied without particular limitation. For example, the method disclosed in Patent Document 1 or Patent Document 2 described above can be used.
  • FIG. 12 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 and is disposed on one surface 11 side of the inspection device 1 here, but is disposed on the other surface 12 side. It may be.
  • the inspection apparatus 50 is not limited to the inspection device 1 and may include any of the inspection devices described above.
  • FIG. 13 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 cavity 16 is not open on the other surface 12 side, so that the supplied solution does not enter the cavity 16 from the other surface 12 side.
  • the solution supply unit 60 supplies the solution so that the solution does not enter the cavity from one surface 11 where the cavity 16 is opened. Control. As shown in FIG.
  • the solution supply unit 60 was provided with a lid portion 67 that covers the opening of the cavity portion 16 in a part of the depressurizing space portion 66, and overflowed from the opening of the pores 14 on one surface 11.
  • the configuration may be such that the solution does not enter the cavity 16.
  • the light emitted from the inspection device and detected by the photodetector is, for example, fluorescence generated when the label attached to the test substance is excited, or luminescence generated by the label acting as a substrate of the reaction solution It is an optical signal resulting from a label, such as luminescence by reaction (hereinafter referred to as “chemiluminescence”).
  • chemiluminescence an optical signal resulting from a label, such as luminescence by reaction
  • the optical signal includes absorbance (colorimetric) in addition to fluorescence and chemiluminescence light.
  • FIG. 14 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.
  • the sample liquid and the labeling solution are mixed in advance outside the test apparatus 50.
  • the test substance 32 and the labeling substance 35 may be combined to supply the mixed liquid into 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 in which the label F acts as a catalyst is detected by the photodetector 70 in a state where the inspection solution such as the buffer solution is filled in the pores 14 ( S4).
  • the inspection solution such as the buffer solution is filled in the pores 14 ( S4).
  • 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 by supplying this reaction solution to the pores 14, the enzyme label imparted to the test substance 32 captured by the capture material 30 in the pores 14 becomes the substance in the reaction solution.
  • Luminescence generated by catalyzing a chemical reaction 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 can improve light extraction efficiency and that can be manufactured at low cost, in addition to an inspection apparatus equipped with the inspection device, and an inspection method. [Solution] An inspection device (1) comprising a plurality of pores (14) that pass through a plate-shaped substrate (10) from a first surface (11) to the other surface (12), and a cavity (16) that extends in the depth direction of the pores (14), that does not penetrate at least the second surface and is formed so as to surround an area (20) that includes at least some of the plurality of pores (14) when viewed from the first surface (11), wherein part of the plate-shaped substrate (10) that constitutes a region (20) including at least some of the pores (16) is light-transmissive, and a scavenger substance (30) that specifically bonds with a specific substance is immobilized on the inner wall surface (14a) of at least some of the pores (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（ＳｉＯ２）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 1, 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.
　特許文献２では、上記問題を解決する手段として、細孔の壁の最表面材料（第１層）の屈折率をその下層（第２層）の屈折率よりも大きな屈折率を有する材料から構成する積層構造が提案されている。この積層構造により、内壁面での光反射率を向上させることができ、結果として光検出効率を向上させることが可能となる。 In Patent Document 2, as a means for solving the above problem, the outermost surface material (first layer) of the pore wall is made of a material having a refractive index larger than the refractive index of the lower layer (second layer). A laminated structure has been proposed. With this laminated structure, the light reflectance on the inner wall surface can be improved, and as a result, the light detection efficiency can be improved.
特許第４１２５２４４号公報Japanese Patent No. 4125244 特表２００５－５２７８１２号公報JP 2005-527812 Gazette
　しかしながら、特許文献１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 1, it is possible to improve the light extraction efficiency accompanying an increase in the light aperture ratio, but the light incident on the silicon wall is also attenuated because the reflectance of silicon used as the frame wall is low. 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.
　また、特許文献２では、具体的なデバイスの作製方法として、第２層を構成する支持体に電気化学エッチングで孔を形成し、第１層をＣＶＤ（Chemical Vapor Deposition）で形成する方法が開示されている。この作製方法を用いるには材料の制約が大きく、第２層は実質的にシリコン系の材料、第１層はＣＶＤ成膜により形成可能な材料に限定されると同時に、作製工程数が多くコスト高となる恐れがある。 Patent Document 2 discloses a specific device manufacturing method in which holes are formed by electrochemical etching in a support constituting the second layer, and the first layer is formed by CVD (Chemical Vapor Deposition). Has been. In order to use this manufacturing method, material restrictions are large, the second layer is substantially limited to a silicon-based material, and the first layer is limited to a material that can be formed by CVD film formation. May be high.
　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、かつ、低コストで作製可能な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 that can improve the light extraction efficiency as compared with a conventional device and can be manufactured at a low cost. 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、
　細孔の深さ方向に延び、少なくとも他方の面に非貫通であり、一方の面から見た場合に複数の細孔の少なくとも一部の細孔を含む領域の周囲に形成された空洞部を有し、
　少なくとも一部の細孔を含む領域を構成する板状基材の部分が光透過性を有し、
　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 cavity formed around the region extending in the depth direction of the pores, not penetrating at least the other surface, and including at least some of the plurality of pores when viewed from one surface. Have
The portion of the plate-like base material constituting the region including at least a part of the pores has light transmittance,
This is a test device in which a capture substance that specifically binds to a specific substance is fixed to the inner wall surface of at least some of the pores.
　板状基材はＳｉＯ２またはＡｌ２Ｏ３（アルミナ）からなることが好ましい。 The plate-like substrate is preferably made of SiO 2 or Al 2 O 3 (alumina).
　本発明の検査デバイスは、板状基材の一方の面からみた場合に、複数の細孔が、空洞部により区画された複数の領域に配置されており、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、複数の領域は、その捕捉物質が互いに異なる領域を有することが好ましい。 In the inspection device of the present invention, when viewed from one surface of the plate-like substrate, a plurality of pores are arranged in a plurality of regions partitioned by the cavity, and one of the plurality of regions It is preferable that the same kind of trapping substance as the trapping substance is bonded to the inner wall surface of each of the pores, and the plurality of regions have areas where the trapping substances are different from each other.
　本発明の検査デバイスは、空洞部が、板状基材の一方の面から他方の面の手前まで延びて形成されてなり、他方の面に非貫通とされていてもよい。 In the inspection device of the present invention, the hollow portion may be formed so as to extend from one surface of the plate-like substrate to the front of the other surface, and may be non-penetrating on the other surface.
　本発明の検査デバイスは、空洞部が、板状基材の一方の面から他方の面に貫通して設けられた貫通孔の他方の面側の開口が板状基材とは異なる材質の部材により塞がれてなり、他方の面に非貫通とされていてもよい。 The inspection device of the present invention is a member whose material is different from that of the plate-like substrate in the opening of the other surface side of the through hole in which the hollow portion is provided so as to penetrate from one surface of the plate-like substrate to the other surface. May be blocked by the other surface and non-penetrated on the other surface.
　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、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 light 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、上記標識物質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 labeling substance, a reaction solution containing a substrate that is catalyzed by the enzyme label and reacts as the test solution, and the light emitted from the inspection device Alternatively, the light produced when the substrate in the reaction solution is catalyzed by the enzyme label may be detected.
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、細孔の深さ方向に延び、少なくとも他方の面に非貫通であり、一方の面から見た場合に複数の細孔の少なくとも一部の細孔を含む領域の周囲に形成された空洞部を有し、少なくとも一部の細孔を含む領域を構成する板状基材の部分が光透過性を有す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。また、基材を単一の材料から構成することも可能であるため、低コストで作製可能である。 An inspection device of the present invention is an inspection device having a plurality of pores penetrating from one surface of a plate-like substrate to the other surface, extending in the depth direction of the pores, and non-existing on at least the other surface. It is a penetration and has a cavity formed around a region including at least some of the plurality of pores when viewed from one side, and constitutes a region including at least some of the pores Since the plate-like base material portion is light transmissive, it is possible to improve the extraction efficiency of light generated in the pores to at least one surface side of the inspection device. In addition, since the base material can be made of a single material, it can be manufactured at low cost.
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る。1 is a perspective view of an inspection device according to a first 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. 本発明の第２の実施形態にかかる検査デバイスの平面図である。It is a top view of the inspection device concerning a 2nd embodiment of the present invention. 細孔の周囲に配置された空洞部を説明するための図である。It is a figure for demonstrating the cavity part arrange | positioned around the pore. 本発明の第３の実施形態にかかる検査デバイスの平面図である。It is a top view of the inspection device concerning a 3rd embodiment of the present invention. 本発明の第３の実施形態にかかる検査デバイスの断面図である。It is sectional drawing of the test | inspection device concerning the 3rd Embodiment of this invention. 設計変更例の検査デバイスの断面図である。It is sectional drawing of the test | inspection device of the example of a design change. さらなる設計変更例の検査デバイスの断面図である。It is sectional drawing of the test | inspection device of the further example of a design change. 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。It is a schematic diagram for demonstrating the effect of the test | inspection device of this invention. 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１４の深さ方向に延び、少なくとも他方の面１２に非貫通であり、一方の面１１から見た場合に複数の細孔１４の少なくとも一部の細孔を含む領域の周囲に形成された空洞部１６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を囲むように空洞部１６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。なお、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 inspection device 1 of the present embodiment extends in the depth direction of the pores 14 on the plate-like substrate 10 having a plurality of pores 14 penetrating from one surface 11 to the other surface 12, and at least the other surface. 12 has a cavity 16 formed around a region including at least some of the plurality of pores 14 when viewed from one surface 11. In the present embodiment, a total of nine pores 14 in the vertical and horizontal directions are used as one region 20, and the cavity 16 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. 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.
　本発明の検査デバイスにおいては、少なくとも一部の細孔を含む領域の周囲に空洞部が形成されていればよい。すなわち、検査デバイスにおいては、少なくとも一部に、空洞部で囲まれた細孔領域が形成されていればよい。また、空洞部で囲まれた領域において、細孔および空洞部を構成する板状基材の部分、すなわち細孔および空洞部の壁面を構成する部分は光透過性を有するものとする。板状基材全体が光透過性材料により構成されていてもよいし、空洞部で囲まれた領域のみが光透過性材料で構成されているものであってもよい。ここで、光透過性を有するとは、後述の検出光の透過率が５０％以上であることをいうが、具体的には、可視光波長域４００～７００ｎｍの平均透過率が５０％以上であれば光透過性を有すると看做すこととする。 In the inspection device of the present invention, it is sufficient that a cavity is formed around a region including at least a part of the pores. That is, in the inspection device, it is sufficient that a pore region surrounded by the cavity is formed at least in part. Further, in the region surrounded by the cavity, the plate-like base material constituting the pores and the cavity, that is, the pores and the wall constituting the cavity have light transmittance. The entire plate-like substrate may be made of a light transmissive material, or only the region surrounded by the cavity may be made of a light transmissive material. Here, having light transmittance means that the transmittance of detection light described later is 50% or more. Specifically, the average transmittance in the visible light wavelength range of 400 to 700 nm is 50% or more. If there is, it will be regarded as having light transparency.
　板状基材は、具体的には、ＳｉＯ２もしくは、Ａｌ２Ｏ３からなるものが好ましい。板状基材の全体が同一の材料から構成されるものであれば作製が容易であり、特に好ましいが、板状基材は、異なる材料からなる部分を備えていてもよい。例えば、空洞部で囲まれた領域のみがＳｉＯ２からなり、その他の部分がＳｉからなる基材、あるいは空洞部で囲まれた領域のみがＡｌ２Ｏ３からなり、その他の部分がＡｌからなる基材などを用いてもよい。 Specifically, the plate-like base material is preferably made of SiO 2 or Al 2 O 3 . If the whole plate-shaped base material is comprised from the same material, preparation will be easy and it is especially preferable, but the plate-shaped base material may be provided with the part which consists of a different material. For example, only the region surrounded by the cavity is made of SiO 2 and the other part is made of Si, or only the region surrounded by the cavity is made of Al 2 O 3 and the other part is made of Al. A substrate or the like may be used.
　本実施形態の検査デバイス１において、空洞部１６は、一方の面１１側が開口され、他方の面１２側が閉塞した空間である。空洞部１６は、板状基材１０の一方の面１１から他方の面１２の手前まで延びて形成されており、板状基材１０の他方の面１２には空洞部１６に繋がる開口は設けられていない（図４参照）。ここで、「手前まで延びて形成される」とは他方の面１２に向かって延びているが、他方の面１２までは至っていないことを意味する。なお、空洞部１６の、一方の面１１から他方の面１２側に延びる長さ、すなわち、細孔１４の深さ方向に沿った深さは板状基材１０の厚みの半分以上であることが好ましく、さらには３／４以上であることがより好ましい。 In the inspection device 1 of the present embodiment, the cavity 16 is a space that is open on one surface 11 side and closed on the other surface 12 side. The hollow portion 16 is formed to extend from one surface 11 of the plate-like base material 10 to the front of the other surface 12, and an opening connected to the hollow portion 16 is provided on the other surface 12 of the plate-like base material 10. (See FIG. 4). Here, “formed to extend to the front” means that it extends toward the other surface 12 but does not reach the other surface 12. The length of the cavity 16 extending from the one surface 11 to the other surface 12 side, that is, the depth along the depth direction of the pores 14 is at least half the thickness of the plate-like substrate 10. Is more preferable, and more preferably 3/4 or more.
　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、空洞部１６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本のみであってもよいが、複数であることが好ましい。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つの検査領域中に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, in the region 20 surrounded by the cavity 16 (hereinafter referred to as “inspection region 20”), there may be only one pore 14, but a plurality of pores are preferable. 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 single pore. 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、特定の物質である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). And as the capture | acquisition substance 30 specifically couple | bonded with the specific substance fixed to the inner wall face 14a of the pore 14, it is a substance couple | bonded specifically with these test substances which are specific 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、一方の面１１から検査デバイス１を見た図を示すように、複数の細孔１４が空洞部１６より区画された複数の検査領域２０に配置されている。１つの検査領域２０中の細孔１４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、複数の検査領域間では、同一種の捕捉物質を含む領域が２以上あってもよい。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た検査デバイスを用いれば、ばらつきを抑制した検査結果を得ることが可能となる。 2, a plurality of pores 14 are arranged in a plurality of inspection regions 20 partitioned by a cavity 16 as shown in a view of the inspection device 1 viewed from one surface 11. A trapping substance of the same type is bound (fixed) to the pores 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. Note that there may be two or more regions containing the same type of capture substance between a plurality of inspection regions. By using an inspection device in which inspection areas 20 in which different capture substances are fixed are alternately arranged and inspection areas in which the same kind of capture substances are periodically fixed are used, an inspection result with reduced variation is obtained. It becomes possible.
　例えば、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、１つの検査デバイスに複数種のアレルゲン検査領域を設ける。これにより、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る。 For example, the same kind of allergen that is a capture substance is fixed to one or a plurality of examination areas 20, and a plurality of allergen examination areas are provided in one examination device. Thereby, the reaction with respect to a several allergen can be test | inspected simultaneously.
　なお、本実施形態においては、検査領域２０の全周に亘って空洞部１６が設けられているが、空洞部１６は全周にわたって設けられている必要はない。板状基材１０の一方の面１１から見た場合において、検査領域２０の周囲に配置された空洞部１６の開口の検査領域２０に面する部分の長さの合計Ｓ１が、検査領域２０内に配置されている複数の細孔１４のうち、最外周に配置されている複数の細孔１４の空洞部１６に面する部分の長さの合計Ｓ２以上であればよい。なお、本発明においては、「領域の周囲に形成された空洞部」とは、Ｓ１≧Ｓ２を満たすように配置された空洞部を意味する。なお、検査領域２０内に配置されている複数の細孔１４のうち、最外周に配置されている複数の細孔１４を結ぶ細孔外周縁の長さをＳ３としたとき、Ｓ２は、細孔外周縁の長さＳ３の１／４以上であることが好ましく、Ｓ３の１／２以上がさらに好ましく、Ｓ３以上であることが特に好ましい。 In the present embodiment, the cavity 16 is provided over the entire circumference of the inspection region 20, but the cavity 16 does not have to be provided over the entire circumference. When viewed from one surface 11 of the plate-like substrate 10, the total length S 1 of the portions facing the inspection region 20 of the opening of the cavity 16 disposed around the inspection region 20 is the inspection region 20. among the plurality of pores 14 disposed within, or if the sum S 2 or more of the length portion facing the cavity 16 of the plurality of pores 14 disposed in the outermost periphery. In the present invention, the “cavity formed around the region” means a cavity arranged so as to satisfy S 1 ≧ S 2 . Of the plurality of pores 14 disposed in the examination region 20, when the length of the pore outer peripheral connecting a plurality of pores 14 disposed in the outermost periphery was S 3, S 2 is , is preferably 1/4 or more the length S 3 of the pores outer peripheral edge, 1/2 or more preferably of S 3, and particularly preferably S 3 or more.
　図２において、検査領域２０内の複数の細孔１４のうち最外周に配置されている複数の細孔１４の一辺はａ０であり、最外周に配置されている細孔１４の空洞部１６に面する部分の長さの合計Ｓ２＝１２ａ０である。また、検査領域２０内の複数の細孔１４のうち最外周に配置されている複数の細孔１４を結ぶ細孔外周縁２２（図中破線で示す。）は一辺の長さａの正方形であり、細孔外周縁２２の長さＳ３＝４ａである。ここでは、細孔１４の間隔は細孔１４の一辺と同等のａ０とする。したがって、Ｓ３＝２０ａ０であり、Ｓ２＞Ｓ３×１／２である。空洞部１６の開口の検査領域２０に面する部分の長さの合計Ｓ１は、一辺の長さがＡである正方形状の検査領域２０の外周２４の長さであり、Ｓ１＝４Ａである。ここではＡ＞ａ＞３ａ０であるのため、Ｓ１≧Ｓ２を満たす。 In FIG. 2, one side of the plurality of pores 14 arranged on the outermost periphery among the plurality of pores 14 in the inspection region 20 is a 0 , and the cavity portion 16 of the pore 14 arranged on the outermost periphery. The total length S 2 = 12a 0 of the portions facing the surface. Further, among the plurality of pores 14 in the inspection region 20, a pore outer peripheral edge 22 (shown by a broken line in the drawing) that connects the plurality of pores 14 arranged on the outermost periphery is a square having a length a on one side. Yes, the length S 3 of the outer circumferential edge 22 of the pore is 4a. Here, the interval between the pores 14 is set to a 0 equivalent to one side of the pore 14. Therefore, S 3 = 20a 0 and S 2 > S 3 × 1/2. The total length S 1 of the portion of the opening of the hollow portion 16 facing the inspection region 20 is the length of the outer periphery 24 of the square inspection region 20 whose one side is A, and S 1 = 4A. is there. Here, since A>a> 3a 0 , S 1 ≧ S 2 is satisfied.
　図５は、第１の実施形態の検査デバイス１と空洞部の形状が異なる第２の実施形態の検査デバイス２の平面図である。図５に示す検査デバイス２において、複数の細孔１４を含む検査領域２０は、複数の（ここでは４つの）孤立した矩形の空洞部１７によって構成されている。複数の細孔１４を含む検査領域２０は図２に示したものと同様であり、検査領域２０内の複数の細孔１４のうち最外周に配置されている複数の細孔１４の空洞部１７に面する部分の長さの合計Ｓ２＝１２ａ０である。また、細孔外周縁２２の長さＳ３＝４ａである。一方、１つの空洞部１７の開口の検査領域２０に面する部分の長さは、矩形状の空洞部１７の長辺の長さＢであり、４つの空洞部１７についての検査領域２０に面する部分の長さの合計Ｓ１は４Ｂである。ここでも、Ｂ＞ａ＞３ａ０であるため、Ｓ１≧Ｓ２を満たす。 FIG. 5 is a plan view of the inspection device 2 of the second embodiment in which the shape of the cavity is different from that of the inspection device 1 of the first embodiment. In the inspection device 2 illustrated in FIG. 5, the inspection region 20 including the plurality of pores 14 is configured by a plurality (here, four) of isolated rectangular cavities 17. The inspection region 20 including the plurality of pores 14 is the same as that shown in FIG. 2, and the cavity 17 of the plurality of pores 14 arranged on the outermost periphery among the plurality of pores 14 in the inspection region 20. The total length S 2 = 12a 0 of the portions facing the surface. Further, the length S 3 = 4a of the outer periphery 22 of the pores. On the other hand, the length of the portion of the opening of one cavity portion 17 facing the inspection region 20 is the length B of the long side of the rectangular cavity portion 17 and faces the inspection region 20 for the four cavity portions 17. total S 1 of length of the portion that is 4B. Again, since B>a> 3a 0 , S 1 ≧ S 2 is satisfied.
　なお、細孔１４の開口形状や空洞部１７の開口形状は矩形に限らない。図６は、細孔１１４および空洞部１１６が共に円形の開口を有し、複数の細孔１１４を含む領域１２０が複数の空洞部１１６により囲まれた場合を示している。このとき、上記のＳ1、Ｓ２およびＳ３は以下のように規定される。 The opening shape of the pores 14 and the opening shape of the hollow portion 17 are not limited to a rectangle. FIG. 6 shows a case where the pore 114 and the cavity 116 both have a circular opening, and the region 120 including the plurality of pores 114 is surrounded by the plurality of cavities 116. At this time, the above S 1 , S 2 and S 3 are defined as follows.
　図６に示すように、細孔外周縁１２２は、複数の細孔１１４のうち最外周に配置されている細孔１１４に接する直線と一部の細孔１１４の縁を通る線で表される。この細孔外周縁（最外周縁）１２２に配置されている細孔１１４の空洞部１１６に面する部分の長さは、各細孔１１４の開口の円周のうち空洞部１１６に臨む弧の長さであり、本例では、図６中太線で示す半円もしくは３/４円の円弧部分である。細孔１１４および空洞部１１６の半径をｒであるとき、Ｓ２＝２πｒ×（１/２）×４＋２πｒ×（３/４）×４＝１０πｒである。その領域１２０を囲むように配置された複数の空洞部１１６の開口の検査領域１２０に面する部分の長さは、それぞれ空洞部１１６の開口の円周のうち検査領域１２０に臨む弧の長さであり、本例では、図６中太線で示す半円もしくは１/４円の円弧部分である。そして、これらの合計を空洞部１１６の検査領域２０に面する部分の長さの合計Ｓ１は、Ｓ１＝２πｒ×（１/２）×１２＋２πｒ×（１/４）×４＝１４πｒである。したがって、ここでもＳ1≧Ｓ２を満たす。 As shown in FIG. 6, the outer peripheral edge 122 of the pores is represented by a straight line in contact with the pores 114 arranged on the outermost periphery among the plurality of pores 114 and a line passing through the edges of some of the pores 114. . The length of the portion of the pore 114 facing the cavity 116 disposed on the outer periphery (outermost periphery) 122 of the pore is the length of the arc facing the cavity 116 in the circumference of the opening of each pore 114. In this example, it is a semicircle or 3/4 circular arc portion indicated by a thick line in FIG. When the radius of the pore 114 and the cavity 116 is r, S 2 = 2πr × (1/2) × 4 + 2πr × (3/4) × 4 = 10πr. The lengths of the portions of the openings of the plurality of cavities 116 arranged so as to surround the region 120 facing the inspection region 120 are the lengths of the arcs facing the inspection region 120 in the circumference of the openings of the cavities 116, respectively. In this example, it is a semicircle or a quarter of a circular arc indicated by a thick line in FIG. The total length S 1 of the portions of these cavities 116 facing the inspection region 20 is S 1 = 2πr × (1/2) × 12 + 2πr × (1/4) × 4 = 14πr. . Therefore, S 1 ≧ S 2 is also satisfied here.
　また、空洞部の開口形状が円形以外の特殊な形状である場合、あるいは開口形状が多角形であるが多角形の辺が検査領域に平行に配置されていない場合についても、検査領域の細孔外周縁および外周縁に位置する細孔の空洞部に面する部分の長さ、空洞部の開口の検査領域に面する部分の長さは、図６の場合と同様に規定できる。 Also, if the opening shape of the cavity is a special shape other than a circle, or if the opening shape is a polygon but the sides of the polygon are not arranged parallel to the inspection region, the pores in the inspection region The length of the outer peripheral edge and the length of the portion of the pore located on the outer peripheral edge facing the cavity and the length of the portion of the opening of the cavity facing the inspection region can be defined in the same manner as in FIG.
　図７は、第３の実施形態の検査デバイス３の平面図であり、図８は図７の検査デバイス３のＶIII－ＶIII線断面図である。図７に示す検査デバイス３においては、細孔１４と空洞部１８が縦横に交互に整列配置されている。本検査デバイス３においては、１つの細孔１４が４つの空洞部１８に囲まれた検査領域２８中に配置されている。本実施形態においては、細孔１４と空洞部１８とは同一の開口形状を有している。本実施形態のように、１つの検査領域２８に１つの細孔１４のみが備えられている場合、細孔外周縁は細孔１４の開口周の長さ、すなわち、Ｓ３＝Ｓ２となる。また、細孔１４と空洞部１８の開口形状が同一であるため、細孔１４を囲む４つの空洞部１８の検査領域２８に面する部分の長さの合計は細孔１４の開口周の長さと一致する。すなわち本例では、Ｓ１＝Ｓ２である。 FIG. 7 is a plan view of the inspection device 3 according to the third embodiment, and FIG. 8 is a cross-sectional view taken along line VIII-VIII of the inspection device 3 of FIG. In the inspection device 3 shown in FIG. 7, the pores 14 and the cavities 18 are alternately arranged vertically and horizontally. In the inspection device 3, one pore 14 is arranged in an inspection region 28 surrounded by four cavities 18. In the present embodiment, the pore 14 and the cavity 18 have the same opening shape. When only one pore 14 is provided in one inspection region 28 as in this embodiment, the outer periphery of the pore is the length of the opening circumference of the pore 14, that is, S 3 = S 2. . Further, since the opening shapes of the pore 14 and the cavity portion 18 are the same, the total length of the portions facing the inspection region 28 of the four cavity portions 18 surrounding the pore 14 is the length of the opening circumference of the pore 14. Match. That is, in this example, S 1 = S 2 .
　図８に示すように細孔１４は貫通孔であり、空洞部１８は、板状基材１０の他方の面１２に貫通していない止まり穴として設けられたものである。 As shown in FIG. 8, the pore 14 is a through hole, and the cavity 18 is provided as a blind hole that does not penetrate the other surface 12 of the plate-like substrate 10.
　上記各実施形態においては、空洞部１６、１７、１８は板状基材１０の一方の面１１から他方の面１２の手前まで延びて形成されるものとして説明したが、図９に設計変更例の検査デバイス４の断面図を示すように、空洞部１１８は、板状基材１０に一方の面１１から他方の面１２に貫通する貫通孔４０を設けた後に、その他方の面１２側の開口を板状基材１０とは異なる材質からなる閉塞部材４２で塞ぐようにして形成されていてもよい。すなわち、図９においては、空洞部１１８は板状基材１０に設けられた貫通孔４０と閉塞部材４２とにより構成されている。 In each of the above embodiments, the hollow portions 16, 17, and 18 have been described as extending from one surface 11 of the plate-like base material 10 to the front of the other surface 12, but FIG. 9 shows a design change example. As shown in the sectional view of the inspection device 4, the cavity 118 is formed on the plate-like substrate 10 after the through-hole 40 penetrating from one surface 11 to the other surface 12 is provided, and then on the other surface 12 side. The opening may be formed so as to be closed with a closing member 42 made of a material different from that of the plate-like substrate 10. That is, in FIG. 9, the cavity 118 is constituted by the through hole 40 and the closing member 42 provided in the plate-like base material 10.
　また、図１０にさらに別の設計変更例の検査デバイス５の断面図を示すように、空洞部１１８は板状基材１０の一方の面１１側の開口が板状基材１０とは異なる材質の第２の閉塞部材４４で塞がれていても構わない。すなわち、空洞部１１８は閉じた空間であってもよい。空洞部内部は真空であってもよいし、屈折率が略１．０である空気などの気体が封じ込められていてもよい。 Further, as shown in FIG. 10 which is a cross-sectional view of the inspection device 5 of another design modification example, the cavity 118 is made of a material whose opening on the one surface 11 side of the plate-like substrate 10 is different from that of the plate-like substrate 10. The second closing member 44 may be closed. That is, the cavity 118 may be a closed space. The inside of the cavity may be a vacuum, or a gas such as air having a refractive index of approximately 1.0 may be enclosed.
　なお、空洞部１１８を構成する閉塞部材４２および第２の閉塞部材４４を構成する材料は、特に制限されないが、金属もしくは樹脂材料など、インクジェット装置により滴下後、硬化して閉塞部材として構成されうる材料が好適である。 The material constituting the closing member 42 and the second closing member 44 constituting the hollow portion 118 is not particularly limited, but it can be configured as a closing member by being hardened after being dropped by an ink jet device, such as a metal or a resin material. Material is preferred.
　なお、図７～図１０では、空洞部に囲まれた検査領域に１つの細孔を含む検査デバイスについて説明したが、検査デバイスとしては、空洞部に囲まれた検査領域には複数の細孔を有することがより好ましい。 7 to 10, the inspection device including one pore in the inspection region surrounded by the cavity has been described. However, as the inspection device, a plurality of pores are included in the inspection region surrounded by the cavity. It is more preferable to have.
　次に、上記各実施形態で示された本発明の検査デバイスの効果について図１１を参照して説明する。
　図１１は検査デバイス１の一部を拡大した模式図である。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１１中では省略している。）に、被検物質を結合させ、被検物質にさらに酵素により標識された標識物質を結合させ、標識物質を触媒とした化学発光を検出することにより被検物質の有無を検出する、あるいは被検物質の有無および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。なお、空洞部１６には溶液６１が入り込まないようにして用いられ、空洞部１６は空気層を構成する。 Next, the effect of the inspection device of the present invention shown in the above embodiments will be described with reference to FIG.
FIG. 11 is an enlarged schematic view of a part of the inspection device 1. In this inspection device 1, a test substance is bound to a capture substance (not shown in FIG. 11) fixed to the inner wall surface 14a of the pore 14, and the test substance is further labeled with an enzyme. It is used in a test method for detecting the presence or absence of a test substance by detecting the chemiluminescence using a labeling substance as a catalyst by binding the substance or detecting the presence and quantity of the 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. In addition, it uses so that the solution 61 may not enter into the cavity part 16, and the cavity part 16 comprises an air layer.
　検査領域２０の細孔１４を構成する基材は光透過性を有するので、シリコンの不透過な基材により構成された細孔１４と比較して、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、光透過性を有する板状基材１０の屈折率と検査用溶液の屈折率は１．３～１．６程度の範囲で同等であるのに対して、空気層の屈折率は略１．０である。すなわち、板状基材１０と空気層との屈折率差は非常に大きいため、界面で大きな反射が生じる。また、板状基材１０は空気層よりの屈折率が大きいため、板状基材１０側から空気層に向かって臨界角以上の入射角θ１で界面Ｃ１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。また、板状基材１０側から界面Ｃ１に臨界角より小さい入射角θ２で入射した場合であっても、入射光の一部は界面Ｃ１で正反射して板状基材１０の表面側に向かうこととなる。また、界面Ｃ１を透過した光は空気層から空洞部１６を構成する板状基材１０に入射し、この界面Ｃ２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、検査領域２０の周囲に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、他方の面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。 Since the base material forming the pores 14 in the inspection region 20 has light transmittance, the surface of the light generated in the pores 14 is compared with the pores 14 formed of the base material impermeable to silicon. The exit aperture ratio increases. In addition, the refractive index of the light-transmitting plate-like substrate 10 and the refractive index of the test solution are equivalent in the range 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 plate-like substrate 10 and the air layer is very large, a large reflection occurs at the interface. Further, since the plate-like substrate 10 has a higher refractive index than the air layer, light incident on the interface C 1 at an incident angle θ 1 greater than the critical angle from the plate-like substrate 10 side toward the air layer is totally reflected. Then, it goes to the surface side of the plate-like substrate 10. Even when the incident light is incident on the interface C 1 from the plate-like substrate 10 side at an incident angle θ 2 smaller than the critical angle, a part of the incident light is specularly reflected at the interface C 1 and It will go to the surface side. Further, the light transmitted through the interface C 1 is incident on the plate-like base material 10 constituting the cavity 16 from the air layer, and at least a part of the light is regularly reflected at the interface C 2 , and the surface of the plate-like base material 10 is again formed. Head to the side. Thus, by providing an air layer around the inspection region 20, a function of condensing light generated in the pores 14 toward the surface of the plate-like substrate 10 is provided. 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. In addition, here, when not distinguishing one surface of the plate-shaped base material 10 and the other surface, it is referred to as the surface of the base material (the surface of the inspection device).
　本発明の検査デバイスでは、板状基材と空気層との界面で光量をほとんど減衰させることなく反射させて、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１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。 In the inspection device of the present invention, the light amount can be reflected at the interface between the plate-like base material and the air layer with almost no attenuation, and the light can be condensed on the surface side of the device. The light extraction efficiency can be greatly improved as compared with the case where a frame made of silicon is provided.
　次に、本発明の検査デバイスの作製方法を説明する。 Next, a method for producing the inspection device of the present invention will be described.
　複数の細孔および空洞部を備えたＳｉＯ２からなる板状基材は下のようにして作製できる。
　まず、Ｓｉからなる板状基材を用意し、その一方の面側からウェットエッチングもしくはドライエッチングにより細孔および空洞部を形成する。例えば、ＭＥＭＳ（Micro Electro Mechanical System）技術において公知のマイクロポーラスＳｉの製造プロセスを用いることができる。なお、図２あるいは図５に示したような空洞部と細孔を作製する場合には、細孔をエッチング形成する際には、細孔形成領域以外にはマスクをし、空洞部をエッチング形成する際には、空洞部形成領域以外にはマスクをして順次形成すればよい。空洞部は、一方の面側からエッチングを行い、他面の手前でエッチングを止めることにより形成することができる。 A plate-like substrate made of SiO 2 having a plurality of pores and cavities can be produced as follows.
First, a plate-like substrate made of Si is prepared, and pores and cavities are formed from one surface side by wet etching or dry etching. For example, a microporous Si manufacturing process known in MEMS (Micro Electro Mechanical System) technology can be used. In the case where the cavity and the pore as shown in FIG. 2 or FIG. 5 are manufactured, when the pore is formed by etching, a mask is provided outside the pore formation region and the cavity is formed by etching. In this case, the mask may be formed sequentially with a mask other than the cavity forming region. The cavity can be formed by etching from one side and stopping the etching before the other side.
　また、空洞部の径が細孔の径よりも小さい場合には、細孔と空洞部のエッチングを同時に行ってもよい。径の小さい空洞部のエッチングスピードは細孔部分のエッチングスピードより遅くなるので、細孔部分は他方の面を貫通するが、空洞部は貫通していない状態とすることが可能である。なお、この原理を利用して、細孔の径よりも小さい径の未貫通の穴を複数形成したのち、穴同士をつなげるように横方向にエッチングして大きな空洞部を形成することもできる。 Further, when the diameter of the cavity is smaller than the diameter of the pore, the pore and the cavity may be etched at the same time. Since the etching speed of the cavity portion having a small diameter is slower than the etching speed of the pore portion, the pore portion penetrates the other surface, but the cavity portion can be left unpenetrated. In addition, using this principle, after forming a plurality of non-through holes having a diameter smaller than the diameter of the pores, a large cavity can be formed by etching in the lateral direction so as to connect the holes.
　上記のようにして形成した細孔、空洞部を有するＳｉからなる基材を酸化処理することによりＳｉＯ２に変化させることにより、基材全体を透明化することができる。酸化処理としては、例えば温度１１００℃で所定時間熱酸化処理を行う等である。熱酸化処理時間は板状基材の大きさ、形状および所望の酸化処理領域に応じて適宜調整すればよい。 The whole base material can be made transparent by changing the base material made of Si having pores and cavities formed as described above to SiO 2 by oxidation treatment. As the oxidation treatment, for example, thermal oxidation treatment is performed at a temperature of 1100 ° C. for a predetermined time. The thermal oxidation treatment time may be appropriately adjusted according to the size and shape of the plate-like substrate and the desired oxidation treatment region.
　なお、図９に示したように空洞部１１８を貫通孔４０と閉塞部材４２により構成する場合には、まず、空洞部の一部を構成する貫通孔４０を細孔１４と同時にエッチングにより形成する。その後、上記の熱酸化処理によりＳｉ基材をＳｉＯ２化させた後、空洞部の一部を構成する貫通孔４０の他方の面側の開口を基材とは異なる材質の閉塞部材４２で塞ぐ。これにより他方の面側を閉塞させた空洞部１１８を形成することができる。閉塞部材４２は貫通孔４０の他方の面側の開口を塞ぐことができればよく、貫通孔４０内部に一部侵入して形成されていても構わない。閉塞部材４２は、例えば、インクジェット装置を用いて貫通孔の他方の面側の開口を塞ぐように金属もしくは樹脂材料を滴下させ、固化あるいは硬化して形成することができる。 In the case where the cavity 118 is constituted by the through hole 40 and the closing member 42 as shown in FIG. 9, first, the through hole 40 constituting a part of the cavity is formed by etching simultaneously with the pore 14. . Thereafter, the Si base material is changed to SiO 2 by the above thermal oxidation treatment, and then the opening on the other surface side of the through hole 40 constituting a part of the cavity is closed with a closing member 42 made of a material different from the base material. . As a result, the cavity 118 with the other surface closed can be formed. The blocking member 42 only needs to be able to block the opening on the other surface side of the through hole 40, and may be formed so as to partially enter the through hole 40. The closing member 42 can be formed, for example, by dripping a metal or a resin material so as to close the opening on the other surface side of the through hole using an ink jet device, and solidifying or curing.
　一方、複数の細孔および空洞部を備えたＡｌ２Ｏ３からなる板状基材は次のようにして作製できる。
　Ａｌからなる基材を用意し、この基材に対して、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処理およびポワワイド処理を施すことにより、多孔性アルミナからなる板状基材を作製することができる。まず、Ａｌ基材を陽極酸化処理して多孔性アルミナの被膜を形成する。さらに、ポワワイド処理を行って微細孔の径を広げることにより、所望の直径の細孔を形成する。 On the other hand, a plate-like substrate made of Al 2 O 3 having a plurality of pores and cavities can be produced as follows.
By preparing a base material made of Al and subjecting this base material to known anodization treatment and power wide treatment described in, for example, JP-A-2015-132737, JP-A-2014-198453, etc., A plate-like substrate made of porous alumina can be produced. First, an Al base material is anodized to form a porous alumina coating. Furthermore, a pore having a desired diameter is formed by performing a pore wide process to widen the diameter of the micropore.
　そのようにして形成した多数の細孔のうち、空洞部として利用する細孔については、上記ＳｉＯ２からなるデバイスの場合と同様に、他方の面側の開口を覆う閉塞部材を設ければよい。 Of the large number of pores thus formed, the pores used as the cavity may be provided with a blocking member that covers the opening on the other surface side as in the case of the device made of SiO 2. .
　以上のようにして、作製された、複数の細孔および空洞部を備えた板状基材の細孔の内壁面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固定化することにより、検査デバイスを作製することができる。
　細孔の内壁面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１あるいは特許文献２等に開示されている方法を用いることができる。 As described above, the trapping substance, which is a specific binding substance that specifically binds to the test substance, is fixed to the inner wall surface of the pores of the plate-like substrate having a plurality of pores and cavities produced as described above. Thus, an inspection device can be manufactured.
As a method for fixing the trapping substance to the inner wall surface of the pore, a known method can be applied without particular limitation. For example, the method disclosed in Patent Document 1 or Patent Document 2 described above 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１２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り、ここでは、検査デバイス１の一方の面１１側に配置されているが、他方の面１２側に配置されるものであってもよい。検査装置５０においては検査デバイス１に限らず、上記のいずれの検査デバイスを備えてもよい。 FIG. 12 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 and is disposed on one surface 11 side of the inspection device 1 here, but is disposed on the other surface 12 side. It may be. The inspection apparatus 50 is not limited to the inspection device 1 and may include any of the inspection devices described above.
　図１３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. 13 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、空洞部１６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側から空洞部１６に浸入することはない。そして、既述の空洞部１６を備えた効果を得るために、空洞部１６が開口されている一方の面１１からも溶液が空洞部に入りこまないように溶液供給部６０は溶液の供給を制御する。図１３に示すように、溶液供給部６０は、減加圧空間部６６の一部に空洞部１６の開口を覆う蓋部６７を備えた、一方の面１１の細孔１４の開口から溢れた溶液が空洞部１６に入り込まないように構成であってもよい。 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. The cavity 16 is not open on the other surface 12 side, so that the supplied solution does not enter the cavity 16 from the other surface 12 side. In order to obtain the effect provided with the cavity 16 described above, the solution supply unit 60 supplies the solution so that the solution does not enter the cavity from one surface 11 where the cavity 16 is opened. Control. As shown in FIG. 13, the solution supply unit 60 was provided with a lid portion 67 that covers the opening of the cavity portion 16 in a part of the depressurizing space portion 66, and overflowed from the opening of the pores 14 on one surface 11. The configuration may be such that the solution does not enter the cavity 16.
　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、もしくは標識が反応液の基質として作用することで生じる発光反応による発光（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 inspection device and detected by the photodetector is, for example, fluorescence generated when the label attached to the test substance is excited, or luminescence generated by the label acting as a substrate of the reaction solution It is an optical signal resulting from a label, such as luminescence by reaction (hereinafter referred to as “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１４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. 14 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、上記では、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に結合させることとしたが、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 above description, the labeling substance 35 is bound to the test substance 32 after the test substance 32 is bound to the capture substance 30. However, the sample liquid and the labeling solution are mixed in advance outside the test apparatus 50. Thus, the test substance 32 and the labeling substance 35 may be combined to 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が触媒として作用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、空洞部１６は空気層を構成している。 Finally, the cleaning liquid is discharged, and the optical signal from the luminescence reaction in which the label F acts as a catalyst is detected by the photodetector 70 in a state where the inspection solution such as the buffer solution is filled in the pores 14 ( S4). When the light emitted from the inspection device 1 is detected by the photodetector 70, the cavity 16 forms an air layer.
　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に付与された酵素標識が反応液中の物質の化学反応を触媒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. Then, in the photodetector 70, by supplying this reaction solution to the pores 14, the enzyme label imparted to the test substance 32 captured by the capture material 30 in the pores 14 becomes the substance in the reaction solution. Luminescence generated by catalyzing a chemical reaction 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.
　１，２，３，４，５　　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
　１４，１１４　細孔
　１４ａ　細孔の内壁面
　１６，１７，１８，１１６，１１８　空洞部
　２０，２８，１２０　検査領域
　２２，１２２　細孔外周縁
　２４　検査領域の外周
　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
　４０　貫通孔
　４２　閉塞部材
　４４　第２の閉塞部材
　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
　Ｆ　標識 1, 2, 3, 4, 5 Inspection device 10 Plate-like substrate 11 One surface of plate-like substrate (inspection device) 12 Other surface of plate-like substrate (inspection device) 14, 114 Pore 14a Pore Inner surface 16, 17, 18, 116, 118 cavity 20, 28, 120 inspection region 22, 122 outer periphery of pore 24 outer periphery of inspection region 30 capture substance 32 test substance 33 specifically bind to test substance Substance 35 Labeling substance 40 Through-hole 42 Closing member 44 Second closing member 50 Inspection device 60 Solution supply part 61 Solution for inspection 62 Reservoir part 64 Pipette part 66 Depressurized space part 68 Pump 70 Photo detector F Marker

Claims (12)

  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、
    　前記細孔の深さ方向に延び、少なくとも前記他方の面に非貫通であり、前記一方の面から見た場合に前記複数の細孔の少なくとも一部の細孔を含む領域の周囲に形成された空洞部を有し、
    　前記少なくとも一部の細孔を含む領域を構成する前記板状基材の部分が光透過性を有し、
    　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,
    It extends in the depth direction of the pores, is at least non-penetrating on the other surface, and is formed around a region including at least some of the plurality of pores when viewed from the one surface. Having a hollow portion,
    A portion of the plate-like substrate constituting the region including the at least some pores has light transmittance;
    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. 　前記板状基材はＳｉＯ２またはＡｌ２Ｏ３からなる請求項１記載の検査デバイス。 The inspection device according to claim 1, wherein the plate-like substrate is made of SiO 2 or Al 2 O 3 .
  3. 　前記一方の面からみた場合に、前記複数の細孔は、前記空洞部により区画された複数の領域に配置されており、
    　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または２に記載の検査デバイス。     When viewed from the one surface, the plurality of pores are disposed in a plurality of regions partitioned by the cavity,
    A trapping substance of the same type is bound as the trapping substance to the inner wall surface of the pore in one of the plurality of areas,
    The inspection device according to claim 1, wherein the plurality of regions have regions in which the combined capture substances are different from each other.
  4. 　前記空洞部が、前記板状基材の前記一方の面から前記他方の面の手前まで延びて形成されてなり、前記他方の面に非貫通とされている請求項１から３のいずれか１項記載の検査デバイス。 4. The device according to claim 1, wherein the hollow portion is formed to extend from the one surface of the plate-like base material to the front of the other surface, and is non-penetrating on the other surface. Inspection device according to item.
  5. 　前記空洞部が、前記板状基材の前記一方の面から前記他方の面に貫通して設けられた貫通孔の前記他方の面側の開口が前記板状基材とは異なる材質の部材により塞がれてなり、前記他方の面に非貫通とされている請求項１から３のいずれか１項記載の検査デバイス。 The hollow portion is formed of a member made of a material different from that of the plate-like base material in which the opening on the other surface side of the through hole provided so as to penetrate from the one surface of the plate-like base material to the other surface is provided. The inspection device according to claim 1, wherein the inspection device is blocked and non-penetrated on the other surface.
  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６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 6, wherein an equivalent circle diameter of the opening region on the one surface of the pore is 1 µm to 100 µm.
  8. 　前記板状基材の厚さは１００μｍ～２０００μｍである請求項１から７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 thickness of the plate-like substrate is 100 µm to 2000 µm.
  9. 　請求項１から８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または前記他方の面の側に配置され、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 8,
    A solution supply unit for supplying a test solution into the pores of the test device;
    An inspection apparatus comprising: a photodetector that is disposed on the one surface or the other surface of the inspection device and detects light emitted from the inspection device.
  10. 　請求項１から８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 pores of the test device according to any one of claims 1 to 8, 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.
  11. 　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１０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 10, wherein as the emitted light, light generated when the substrate in the reaction solution is catalyzed by the enzyme label is detected.
  12. 　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１０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 10, wherein fluorescence emitted from the labeling substance by irradiation with the excitation light is detected as the emitted light.
PCT/JP2017/010091 2016-03-30 2017-03-14 Inspection device, inspection apparatus and inspection method WO2017169714A1 (en)

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JP2003066042A (en) * 2001-08-27 2003-03-05 Nippon Shokubai Co Ltd Biochip
JP2004286579A (en) * 2003-03-20 2004-10-14 Kanagawa Acad Of Sci & Technol Dna analyzing array, and dna analysis system and analysis method using the same
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