Manufacturing method of microchip for liquid sample analysis

Abstract

A method for manufacturing a microchip for analyzing a component in a liquid sample by passing the sample through a flow path provided therein and causing reaction at a reaction part provided to a part of the flow path, the method comprising: a step (A) for preparing a substrate (1) having, on the surface thereof, a groove (11) which serves as the flow path and the reaction part (14) provided at a part between the two ends of the groove; a step (B) for applying a tackifier or an adhesive on a region other than where the groove is provided on the surface of the substrate; a step (C) for preparing a film (2) having a partial region (21) on which a reactive substance is applied; and a step (D) for adhering the film onto the substrate such that the groove on the substrate is covered with the film to form the flow path and such that the reaction part on the surface of the substrate having the tackifier or the adhesive applied thereon overlaps the region of the film on which the reactive substance is applied.

Description

[Technical Field]

[0001] The present invention relates to a method for manufacturing a microchip for analyzing liquid samples. [Background technology]

[0002] It is known that a liquid sample such as blood is introduced into a flow channel in a microchip and reacted with an antibody or the like in a reaction section provided midway through the flow channel to analyze the components in the liquid sample. A known method for producing such a microchip involves bonding a substrate having grooves formed on the surface thereof to a film with an adhesive (Patent Documents 1 and 2). However, conventional methods involve placing beads onto which antibodies or other substances to be used in the reaction are immobilized in the reaction section of the channel, which requires cost and time to manufacture the microchip, so a simpler method of manufacturing it was desired. [Prior art documents] [Patent documents]

[0003] [Patent Document 1] Patent Publication No. 2008-232939 [Patent Document 2] Patent Publication No. 2008-175795 Summary of the Invention [Problem to be solved by the invention]

[0004] An object of the present invention is to provide a method for easily and inexpensively manufacturing a microchip for analyzing components in a liquid sample by passing the liquid sample through a flow path provided inside the microchip and causing a reaction in a reaction section provided in a part of the flow path. [Means for solving the problem]

[0005] The present inventors conducted extensive research to solve the above-mentioned problems. As a result, they found that a microchip can be easily manufactured by preparing a substrate having a groove on its surface that serves as a flow path and a reaction site located partially between the ends of the groove, applying at least one of an adhesive and a pressure-sensitive adhesive to the area of ​​the substrate surface other than the groove, and preparing a film having a reaction substance applied to a portion of the substrate surface, and bonding the film to the substrate so that the groove on the substrate is covered with the film to form a flow path, and the reaction site on the adhesive and / or pressure-sensitive adhesive-coated surface of the film overlaps with the area of ​​the film coated with the reaction substance. The obtained microchip does not leak liquid and can be suitably used for analyzing components in liquid samples. Furthermore, they found conditions, such as the type of adhesive and pressure-sensitive adhesive, for efficiently bonding the substrate and the film, which led to the completion of the present invention.

[0006] That is, the present invention provides a method for manufacturing a microchip for analyzing components in a sample by passing a liquid sample through a flow path provided inside the microchip and causing a reaction in a reaction section provided in a part of the flow path, the method comprising: A substrate is prepared on its surface, the substrate having a groove serving as a flow path and a reaction area located between both ends of the groove; applying at least one of an adhesive and a pressure-sensitive adhesive to areas other than the grooves on the surface of the substrate on which the grooves are formed; providing a film having a reactive material applied to some areas; and The manufacturing method includes a step of laminating a film onto the substrate so that a groove on the substrate is covered with the film to form a flow path, and a reactive portion on at least one of the adhesive and pressure-sensitive adhesive application surfaces of the substrate overlaps with an area of ​​the film to which the reactive substance is applied.

[0007] Here, the substrate is preferably made of any one of plastic, silicone, and glass. The film is a film of cycloolefin polymer (COP), cycloolefin copolymer (COC), polymethyl methacrylate (PMMA), polystyrene (PS), polycarbonate (PC), or polyethylene terephthalate (PET). It is preferable. The reactant is preferably an antibody, an enzyme, a nucleic acid, or a bead containing any of them. The adhesive and pressure sensitive adhesive are preferably UV curable. The adhesive and pressure sensitive adhesive are preferably applied to the areas of the base material other than the grooves by screen printing. The surface of the substrate may be hydrophilically treated, and at least one of an adhesive and a pressure-sensitive adhesive may be applied to the hydrophilically treated surface. In one aspect of the present invention, the substrate or film may have through holes that serve as an inlet and an outlet at positions corresponding to both ends of the flow channel formed by bonding the substrate and film together, sandwiching the reaction section. In one aspect of the present invention, the substrate, the surface of which is coated with at least one of an adhesive and a pressure-sensitive adhesive, may be bonded to the film after a stirring bar is placed in the depression that will become the reaction area. In one embodiment of the present invention, a mixture of adhesive and pressure-sensitive adhesive may be applied to the area of ​​the substrate other than the grooves. In one aspect of the present invention, an adhesive is applied to the inner region of the substrate excluding the outer peripheral portion, excluding the groove that becomes the flow path, and a pressure-sensitive adhesive is applied to the region of the film that corresponds to the outer peripheral portion of the substrate when they are bonded together, and the two are bonded together with the adhesive or pressure-sensitive adhesive coated surface facing inward. In one aspect of the present invention, the film may be subjected to a hydrophilization treatment in the area where the reactive substance is to be applied, and the reactive substance may be applied to the hydrophilized area. Then, the film may be bonded to a base film having at least a portion of the grooves that has been hydrophilized. [Effects of the Invention]

[0008] According to the present invention, a microchip for analyzing components in a liquid sample can be produced simply and inexpensively. [Brief explanation of the drawings]

[0009] [Figure 1] 1 shows an embodiment of the microchip of the present invention, where A shows the substrate of the microchip (before adhesive application), B shows the substrate of the microchip (after adhesive application), C shows the film of the microchip, and D shows the completed microchip. [Figure 2] 1 shows a microchip according to Example 6 of the present invention, in which A shows the substrate of the microchip (before adhesive application), B shows the substrate of the microchip (after adhesive application), C shows the film of the microchip, and D shows the completed microchip. [Figure 3] 1 shows a microchip according to Example 1 of the present invention, in which A shows the substrate of the microchip (before adhesive application), B shows the substrate of the microchip (after adhesive application), C shows the film of the microchip, and D shows the completed microchip. [Figure 4] 1 shows a microchip according to Example 5 of the present invention, in which A shows the substrate of the microchip (before adhesive application), B shows the substrate of the microchip (after adhesive application), C shows the film of the microchip (after adhesive application), and D shows the completed microchip. DETAILED DESCRIPTION OF THE INVENTION

[0010] The manufacturing method of the present invention is a method for manufacturing a microchip for analyzing components in a sample by passing a liquid sample through a flow path provided inside the microchip and causing a reaction in a reaction section provided in a part of the flow path.

[0011] The liquid sample is not particularly limited as long as it can be passed through the microchip, and examples thereof include liquid samples obtained from living organisms such as blood and urine, or diluted solutions thereof, extracts from living organisms such as plants and animals, naturally occurring water such as rivers, oceans, and rainfall, cleaning solutions, waste liquids, etc. The components in the sample are also not particularly limited, and examples thereof include proteins, nucleic acids, low molecular weight compounds, sugars, etc.

[0012] The manufacturing method of the present invention comprises: A substrate is prepared on its surface, the substrate having a groove serving as a flow path and a reaction area located between both ends of the groove; applying adhesive and pressure-sensitive adhesive to areas of the grooved surface of the substrate other than the grooves; providing a film having a reactant applied to some areas; The method includes a step of laminating a film onto the substrate so that the groove on the substrate is covered with the film to form a flow path, and the reactive portion on the adhesive and pressure-sensitive adhesive-coated surface of the substrate overlaps with the area on the film where the reactive substance is coated. In the manufacturing method of the present invention, a second substrate having no channel grooves formed on its surface may be used instead of the film. In this case, the following description of the film can be applied to the second substrate as is.

[0013] The manufacturing method of the liquid sample analysis microchip of the present invention will be described below with reference to the drawings. However, the following is merely an example, and the manufacturing method of the present invention and the microchip obtained thereby are not limited to the following aspects.

[0014] FIG. 1 is a conceptual diagram showing an example of the configuration of a microchip 10. As shown in FIG. FIG. 1A is a plan view of a substrate 1 on whose surface a groove is dug to become a flow channel 11 of a microchip 10. A through-hole is provided on the first end side of the groove to become an inlet 12 for a liquid sample, and the other A through-hole is provided at the end side to serve as the outlet 13. In addition, a depression to serve as the reaction section 14 is provided midway along the groove, i.e., in a part between the through-hole to serve as the inlet 12 and the through-hole to serve as the outlet (discharge port) 13.

[0015] Two or more flow paths may be provided. The flow path may have any shape, either linear or curved. The flow path may also have a branch. In this case, two or more inlets, reaction sections, and / or air holes may be provided. For example, two inlets may be provided, with a liquid sample flowing into the first flow path through the first inlet and a reaction substrate liquid flowing into the second flow path through the second inlet. A reaction section may be provided at the junction of the first and second flow paths, with a junction flow path and an outlet (discharge port) provided downstream. The inlet and outlet may be provided on either the substrate or the film. For example, a groove serving as a flow path may be provided on the substrate, and a film having holes at positions overlapping both ends of the groove may be prepared and attached to the substrate. Alternatively, one of the holes serving as the inlet and outlet may be provided in the substrate, and the other in the film.

[0016] The cross-sectional shape of the groove that becomes the flow path may be any shape, such as a concave, U-shape, or V-shape. The depth of the groove that becomes the flow path is preferably 10 to 500 μm, and the width is preferably 10 μm to 3 mm. The length of the portion that corresponds to the flow path is, for example, 3 mm to 5 cm. The width of the groove may be constant or may vary, and the depth of the groove may be constant or may vary.

[0017] The depression that serves as the reaction section may be large enough to store the liquid sample introduced from the inlet and react with the reactant contained in the reaction section, and its shape is not particularly limited, but may be, for example, cylindrical or prismatic, and by increasing the area and depth, it is possible to store a larger amount of liquid sample. The area of ​​the depression may be, for example, 0.1 to 50 mm 2 In the case of a circular reaction section, the diameter is, for example, 0.2 to 6 mm. However, the area may vary depending on the depth of the groove, and may be, for example, a cone-shaped depression. The depth of the depression is preferably deeper than the depth of the groove that becomes the flow path, and is, for example, 20 μm to 3 mm.

[0018] When the reaction zone extends relative to the flow path, for example, in a cylindrical or prismatic shape, air may easily accumulate in the reaction zone. In such cases, the flow direction can be controlled by hydrophilizing all or part of the film and / or substrate (such as the groove that forms the flow path in the substrate or the portion of the film that covers the flow path), thereby preventing air bubbles from remaining in the cylindrical or prismatic reaction zone. The hydrophilization treatment may be performed on the portion of the substrate that corresponds to the reaction zone and the portion of the film that covers the reaction zone.

[0019] Furthermore, if the reaction between the reactant and the sample proceeds rapidly, if the flow rate of the sample in the reaction section is very slow, or if the movement of the sample pauses or moves back and forth in the reaction section, there is no need to store the liquid sample in the reaction section, and the reaction section may be the same depth as the flow channel. In other words, there is no need to provide a recess, and the width of the flow channel may be increased without providing a recess. The reaction section may also be the same width as the flow channel.

[0020] Widening the channel width and providing a depression is suitable for promoting the reaction by mixing the sample and reactants with a stirrer. On the other hand, widening the channel width without changing the depth increases the contact area with the reactants, making it suitable for dissolving and diffusing the reactants without stirring, and can be selected according to the purpose of the test.

[0021] A wide portion serving as a waste liquid (solution) reservoir may also be provided downstream of the flow channel. That is, in one embodiment of the present invention, the flow channel 11 has a shape such that the waste liquid reservoir is connected to an end on the side different from the inlet end. This allows the liquid sample that has passed through the flow channel to be retained in the waste liquid reservoir. The solution reservoir may also be provided upstream of the flow channel. Furthermore, by providing a through-hole (which may be on the substrate side or the film side) in a part of the waste liquid storage section, it can be made to function as an air hole. An absorbent material of a size that can be accommodated in the waste liquid storage section can also be placed in the waste liquid storage section. Examples of absorbent materials include sponge and cloth. The depth of the groove corresponding to the waste liquid storage section is preferably deeper than the depth of the groove corresponding to the flow path in order to store a large amount of waste liquid.

[0022] The size of the through-hole that serves as the inlet 12 may be any size that allows a liquid sample such as blood to be injected using a microsyringe, etc. For example, the diameter is 0.2 to 3 mm. The size of the through-hole that serves as outlet 13 is not particularly limited as long as it functions as an outlet for the liquid sample, and is, for example, 0.2 to 2 mm in diameter.

[0023] Microchip materials that can be used include metal, glass, plastic, silicone, etc., but from the viewpoint of detecting reactions by luminescence, color development, or visual inspection, transparent materials are preferred, and transparent plastics are more preferred. Examples include polyethylene, polypropylene, polystyrene, polymethyl methacrylate, cycloolefin polymer, cycloolefin copolymer, polyphenylene oxide, polyethylene terephthalate, polyethylene naphthalate, polycarbonate, polyamide, polyimide, phenolic resin, epoxy resin, polyvinylidene chloride, polyvinyl chloride, ABS resin, and poly(2-methoxyethyl acrylate) (PMEA) resin.

[0024] The grooves and holes in the microchip substrate can be made by cutting with a blade or laser beam, but if the microchip is made of plastic, they can also be formed by injection molding, which is preferable because it allows for the efficient production of microchips of consistent quality.

[0025] The hydrophilization treatment is preferably carried out by applying a hydrophilization reagent or by plasma treatment. Examples of the hydrophilization reagent include S-1570 (sucrose fatty acid ester: Mitsubishi Chemical Foods Corporation), LWA-1570 (sucrose laurate: Mitsubishi Chemical Foods Corporation), Poem DL-100 (diglycerin monolaurate: Riken Vitamin Co., Ltd.), and Rikemal A (sucrose laurate: Mitsubishi Chemical Foods Corporation). Examples of suitable surfactants include nonionic surfactants such as sugar fatty acid esters (Riken Vitamin Co., Ltd.), Cera Aqua NS235-N1 (Shima Trading Co., Ltd.), Aminoion (Nippon Nyukazai Co., Ltd.), LAMBIC-771W (Osaka Organic Chemical Industry Ltd.), LAMBIC-1000W (Osaka Organic Chemical Industry Ltd.), SPRA-101 (Tokyo Ohka Kogyo Co., Ltd.), and SPRA-202 (Tokyo Ohka Kogyo Co., Ltd.). Specific conditions include those in which the water contact angle of the substrate surface is 55° or less.

[0026] 1C is a plan view of the film 2. Transparent plastic is preferable as the material of the film, and examples thereof include the materials mentioned above, with PET resin, COP resin, COC resin, PS resin, PC resin, and PMMA resin being more preferable. The thickness of the film is, for example, preferably 50 to 200 μm, more preferably 100 to 200 μm.

[0027] A reactive substance is coated on the film in the area that overlaps with the reaction section 14 on the flow path 11 when the film is laminated with the substrate 1, and when this coated section 21 is laminated with the substrate 1, the reactive substance is accommodated in the reaction section.

[0028] The reactive substance may be any substance that reacts with the target (detection target) component in the liquid sample and can be appropriately selected depending on the type of target substance. Examples of the reactivity of a reactive substance include biological reactions and chemical reactions, with biological reactions including binding reactions. Reactive substances include proteins (including peptides), sugars, nucleic acids, low-molecular-weight compounds, and the like. Examples include substances such as antibodies that specifically bind to the target substance, and blood coagulation factors such as enzyme proteins and PT reagents that use the target substance as a substrate. Furthermore, if the target substance is a nucleic acid, nucleic acid probes and polymerases (nucleic acid amplification enzymes) that amplify nucleic acids may also be used.

[0029] The reactive substance may be two or more types, and two or more types of reactive substances may be coated on the film. Furthermore, a substance other than the reactive substance may also be coated on the film. For example, if the reactive substance is an enzyme, the enzyme substrate or buffer may also be coated on the film. Such substrates, buffers, etc. may be contained in a recess or other portion that will serve as the reaction portion on the substrate side. Furthermore, when two types of reactive substances are used, one type may be coated on a film, and the other type may be contained in a recess or other portion that will serve as the reaction portion on the substrate side. By separately coating the reactive substances on the substrate reaction portion and the film, it is possible to prevent aggregation or reaction during microchip production by coating a reagent that will react or aggregate when mixed, or two types of reagents that will react like an enzyme and a substrate, on the substrate and the film and laminating them together so that they overlap.

[0030] Alternatively, enzymes or antibodies may be immobilized on microbeads as reactants and then coated onto the film. By immobilizing the reactants on microbeads and then coating them, the contact area between the liquid sample and the reactants increases, which can accelerate the reaction.

[0031] The coating amount of the reactive substance can be appropriately set depending on the type of reactive substance, but for example, it is 1 to 10,000 μg / cm 2 The reactive material may be multi-coated.

[0032] The coating of the reactive substance can be appropriately selected depending on the type of reactive substance, and known methods can be used. For example, a method in which a solution of the reactive substance is prepared, spotted on a predetermined position on the film, and then dried naturally or under reduced pressure can be mentioned.

[0033] When plastic is used as the film material, the area on the film where the reactant is to be coated can be precisely coated with a hydrophilizing reagent by inkjet printing or a dispenser, followed by hydrophilization treatment. The reactant solution can then be dropped onto the desired hydrophilized area using a dispenser such as a pipette or syringe, allowing the reactant solution to spread evenly over the pre-hydrophilized area on the film. The applied reactant solution is preferably coated onto the reactant by natural drying, drying under reduced pressure, or freeze-drying. The hydrophilization treatment of the film for precise application of the reactant aqueous solution is not particularly limited, but it is preferable that the contact angle is 55° or less, preferably 40° or less, so that the dropped reactant aqueous solution spreads well over the pre-hydrophilized area.

[0034] Alternatively, a reactive functional group can be introduced into the target region on the film surface, and then reacted with a functional group of the reactive substance, thereby achieving stable immobilization through a covalent bond.

[0035] 1D is a plan view of microchip 10 obtained by bonding substrate 1 and film 2 together so that the grooved surface of substrate 1 and the surface coated with a reactive substance of film 2 are in contact with each other. Dashed lines indicate that flow channel 11, reaction section 14, etc. are present inside microchip 10.

[0036] By laminating and bonding the film 2 onto the substrate 1, the upper portions of the grooves and depressions that will become the flow channels and reaction areas are covered with the film, forming the flow channels through which the liquid sample passes and the reaction areas where the reactions take place. Furthermore, by laminating a film, one side of the through-hole is sealed, and only the side of the base material that is not laminated with the film becomes an opening, which functions as an inlet and an outlet. That is, a liquid sample introduced through an inlet reacts with a reactant in the reaction section and is then discharged through an outlet. The target substance in the sample can be measured by observing or detecting the reaction in the reaction section. Examples of reactions include, but are not limited to, color reactions, luminescence reactions, amplification reactions, and agglutination reactions.

[0037] To bond the film 2 onto the substrate 1, an adhesive and / or a pressure sensitive adhesive is used. Examples of adhesives include (meth)acrylic resin adhesives, natural rubber adhesives, urethane resin adhesives, ethylene-vinyl acetate resin emulsion adhesives, ethylene-vinyl acetate resin adhesives, epoxy resin adhesives, vinyl chloride resin solvent-based adhesives, chloroprene rubber adhesives, cyanoacrylate adhesives, silicone adhesives, styrene-butadiene rubber solvent-based adhesives, nitrile rubber adhesives, nitrocellulose adhesives, phenolic resin adhesives, modified silicone adhesives, polyester adhesives, polyamide adhesives, polyimide adhesives, olefin resin adhesives, vinyl acetate resin emulsion-based adhesives, polystyrene resin solvent-based adhesives, polyvinyl alcohol adhesives, polyvinylpyrrolidone resin adhesives, polyvinyl butyral adhesives, polybenzimidazole adhesives, polymethacrylate resin solvent-based adhesives, melamine resin adhesives, urea resin adhesives, and resorcinol adhesives. The adhesives can be used alone or in combination of two or more. Examples of the adhesive include rubber-based adhesives, (meth)acrylic-based adhesives, silicone-based adhesives, urethane-based adhesives, vinyl alkyl ether-based adhesives, polyvinyl alcohol-based adhesives, polyvinylpyrrolidone-based adhesives, polyacrylamide-based adhesives, cellulose-based adhesives, etc. Such adhesives may be used alone or in combination of two or more. The adhesive or pressure-sensitive adhesive is preferably a photocurable adhesive (either radically reactive or cationic polymerizable), and more preferably a UV-curable adhesive. UV-curable adhesives or pressure-sensitive adhesives can be applied after the coating process, and the curing reaction can be rapidly initiated to bond the substrates. Examples of UV-curable adhesives include acrylic UV-curable adhesives such as UVX-8204 (manufactured by Denka Co., Ltd.), UVX-8400 (manufactured by Denka Co., Ltd.), SX-UV100A (manufactured by Cemedine Co., Ltd.), SX-UV200 (manufactured by Cemedine Co., Ltd.), BBX-UV300 (manufactured by Cemedine Co., Ltd.), U-1340 (Chemitech Co., Ltd.), U-1455B (Chemitech Co., Ltd.), U-1558B (Chemitech Co., Ltd.), Aronix UV-3000 (Toagosei Co., Ltd.), TB3094 (ThreeBond Co., Ltd.), and Hitaloid 7975D (Hitachi Chemical Co., Ltd.). The UV-curable adhesive is preferably an acrylic UV-curable adhesive such as UV-3630ID80 (Mitsubishi Chemical Corporation), UX-3204 (Nippon Kayaku Co., Ltd.), or Finetack RX-104 (DIC Corporation). Acrylic UV-curable adhesives and adhesives exhibit good adhesion to a wide range of plastic materials and rapidly develop strength after UV irradiation. The viscosity of the adhesive or adhesive used to bond the film 2 to the substrate 1 is preferably, for example, 2,000 to 31,000 mPa·s.

[0038] The adhesive and pressure-sensitive adhesive are applied to positions on the substrate surface other than the grooves. For example, as shown in B in Figure 1, the adhesive and pressure-sensitive adhesive are preferably applied to areas on the substrate surface other than the flow path and reaction section. To more accurately apply the adhesive and pressure-sensitive adhesive to areas other than the grooves, the adhesive and pressure-sensitive adhesive are preferably applied using a printing technique, and screen printing is particularly preferred. By using screen printing, even when the adhesive and pressure-sensitive adhesive are filled into a plate covering the entire surface of the substrate, the adhesive and pressure-sensitive adhesive are transferred to areas other than the grooves that come into contact with the screen printing plate, but are not transferred to grooves that do not come into contact. This makes it possible to apply the adhesive and pressure-sensitive adhesive well to areas other than the grooves. The thickness of the applied adhesive or pressure-sensitive adhesive is preferably 5 to 15 μm. To control the thickness of the adhesive or pressure-sensitive adhesive, the number of meshes per inch of the screen is preferably, for example, 500 to 730. The opening rate of the mesh is preferably, for example, 39 to 47%. The thickness of the mesh is preferably, for example, 15 to 28 μm. As a result, the thickness of the applied adhesive or pressure-sensitive adhesive is preferably 5 to 15 μm.

[0039] Other methods for applying adhesives and pressure sensitive adhesives to the substrate include inkjet printing, gravure printing, and using a dispenser to precisely apply the adhesive to the outside of the flow path. With these application techniques, if adhesive or pressure-sensitive adhesive is dispensed onto a groove, the adhesive will be applied inside the groove, changing the shape of the flow path. Therefore, it is necessary to apply the adhesive or pressure-sensitive adhesive to areas other than the groove by capturing an image of the groove position on the substrate, or by fixing the positions of the printing stage and substrate and programming the printer or dispenser to apply the adhesive or pressure-sensitive adhesive to areas other than the groove.

[0040] Alternatively, the surface of the substrate may be hydrophilized before being coated with the adhesive or pressure-sensitive adhesive. The hydrophilization treatment is preferably a plasma treatment or a corona treatment. Good bonding can be achieved by ensuring that the substrate does not repel the adhesive or pressure-sensitive adhesive, that the adhesive or pressure-sensitive adhesive wets and spreads on the substrate, and that the adhesive or pressure-sensitive adhesive does not flow into the flow path.

[0041] Furthermore, in order to improve the internal pressure strength and peel strength of the microchip and to reduce elution into the flow path, an adhesive is applied to the inner region (and the region other than the grooves) excluding the outer peripheral portion of the substrate surface (for example, an outer peripheral region having a width of 1 to 5 mm), and a pressure-sensitive adhesive is applied to the outer peripheral portion (for example, an outer peripheral region having a width of 1 to 5 mm) of a film that will be joined to the substrate with the molded grooves, and the two are joined to produce a microchip. For the inner region, including the periphery of the groove on the substrate surface, it is preferable to select a UV-curable adhesive, particularly a radically reactive acrylic UV-curable adhesive. Radical-reactive acrylic UV-curable adhesives can be completely cured by performing UV irradiation in a nitrogen-filled environment, suppressing curing inhibition by oxygen. This improves the internal pressure strength of the flow path. Furthermore, by completely curing the adhesive and completing the polymerization reaction of the polymer contained in the adhesive, it is possible to reduce the elution of adhesive-derived components into the flow path. While there are no particular limitations on the method for creating a nitrogen-filled environment, using a nitrogen-substituted box consisting of components made of UV-transparent materials such as intake valves, exhaust valves, relief valves, and glass is preferable, as it allows for easy UV irradiation in a nitrogen atmosphere. A UV-curable adhesive can be selected for the outer periphery. UV-curable adhesives do not easily peel off even when subjected to external physical stress, and can provide the microchip with peel strength. Even if peeling does occur, they can be re-adhered by applying pressure with the fingers, etc. Even when applying adhesive to the inner region including the periphery of the groove on the surface of the substrate and applying a pressure sensitive adhesive to the outer periphery of the substrate, the adhesive can be precisely applied to the region other than the groove by screen printing. There are no particular restrictions on the method for applying the adhesive. After the steps of applying the adhesive and adhesive, the pieces can be aligned so that the coated areas do not overlap, and then irradiated with UV light, allowing for efficient production.

[0042] Alternatively, an adhesive or pressure-sensitive adhesive can be applied to the surface of the substrate, a stirrer bar can be placed in the depression that will become the reaction zone, and then the substrate and film can be attached. This allows the stirrer bar to be housed within the reaction zone, and by driving the stirrer bar with an external magnetic force or other force, the reaction between the reactive substance and the target substance in the liquid sample can proceed efficiently. The stirrer bar can also be hydrophilized, which can prevent air bubbles from accumulating around the stirrer.

[0043] The present invention will be specifically described below with reference to examples, but the present invention is not limited to the following embodiments. [Example]

[0044] <Microchip production 1> We investigated the presence or absence of solvents in the adhesive or pressure sensitive adhesive applied to the microchip, as well as the curing method. A substrate 201 (MCC Advanced Moldings Co., Ltd. injection molded product: COP resin) (size: 59.4 × 26.2 mm, thickness: 3.0 mm) shown in A of FIG. 3 was prepared. In the substrate 201, the length of the flow path 211 was 33.6 mm, the depth was 80 μm, and the width of the inlet portion was 1.2 mm and the width of the narrowed portion was 0.3 mm. The length of the waste liquid storage portion 212 was 16.5 mm, the depth was 2.2 mm, and the width was 20.2 mm. In the substrate 201, the hole that became the inlet 213 was a through-hole with an inner diameter of 2 mm and a circular cross section. On the other hand, the hole that became the air hole 214 was a through-hole with an inner diameter of 1 mm and a circular cross section. The film C in Figure 3 was a COP film (size 70 × 50 mm, thickness 100 μm). To bond the substrate 201 and film 202, UVX-8204, a solvent-free, radically reactive acrylic UV-curable adhesive, or a radically reactive acrylic UV-curable pressure-sensitive adhesive containing ethyl acetate as a diluent, was used. As shown in Figure 3B, the adhesive or pressure-sensitive adhesive was applied to the surface of the substrate 201 where the flow path and solution reservoir were provided by the following method. The adhesive or pressure-sensitive adhesive was applied by screen printing to the surface of the substrate 201 where the flow path and solution reservoir were provided. The screen used had a mesh count of 640 and an opening rate of 39%. The thickness of the adhesive or pressure sensitive adhesive applied was approximately 7 μm. The adhesive or pressure-sensitive adhesive coated surface of the substrate 201 was laminated with the film 202, and a UV-LED light source was used to irradiate it with ultraviolet light of 365 nm wavelength for 10-20 seconds, thereby initiating the curing reaction of the adhesive and bonding the film 202 onto the substrate 201 (Figure 3D).

[0045] <Microchip Evaluation 1> When microchip 200 was produced, it was found that when an acrylic UV-curable adhesive containing butyl acetate as a diluent was used, the solvent evaporated on the screen, gradually increasing the viscosity of the adhesive. As a result, approximately five minutes after the adhesive was placed on the screen, the mesh of the screen became clogged, making it impossible to apply the adhesive. This suggests that adhesives and adhesives containing solvents as diluents are not suitable for producing microchips by screen printing. On the other hand, when a solvent-free acrylic UV-curing adhesive was used, the mesh of the screen did not become clogged even about five hours after the adhesive was placed on the screen, making it possible to apply the adhesive continuously and uniformly.In addition, by using a UV-curing adhesive or pressure-sensitive adhesive, the curing reaction does not begin on the screen, but only when a specific wavelength of UV light is irradiated onto the microchip coated with the adhesive or pressure-sensitive adhesive, improving workability. Furthermore, when distilled water was pumped through the channels of the fabricated microchip, it was observed that the distilled water flowed only through the channel grooves without leaking out of the channels. These results demonstrate that continuous production of microchips is possible by applying a solvent-free acrylic UV-curing adhesive to the areas of the substrate other than the flow path formed by screen printing, then bonding it to a film and irradiating it with UV light. [Example]

[0046] <Microchip production 2> The optimal film thickness of the adhesive to be applied to the microchip was investigated. The film thickness of the adhesive was controlled by the mesh number of the screen, the opening rate, and the printing speed. Except for the screen used for applying the adhesive, the same method as described in <Fabrication of Microchip 1> in Example 1 was used. The adhesive was applied as follows. The adhesive UVX-8204 was applied by screen printing to the periphery of the flow path of the substrate 201. The implementation conditions were as follows: the screen had a mesh number of 730, an opening rate of 39%, a printing speed of 300 mm / s, and a film thickness of approximately 3 μm; ,Opening rate 39%, printing speed 200mm / s, film thickness is about 5μm, The mesh count is 640, the opening rate is 39%, and the printing speed is 200 mm / s. The film thickness is approximately 1 The mesh number is 400, the opening rate is 49%, and the printing speed is 300 mm / s, resulting in a film thickness of approximately 15 μm. The mesh number is 400, the opening rate is 49%. The printing speed was 200 mm / s, and the film thickness was approximately 18 μm.

[0047] <Microchip Evaluation 2> The mesh number is 730, the opening rate is 39%, and the printing speed is 300 mm / s. When microchip 200 was fabricated under the condition that the thickness was 3 μm, many voids were observed around the flow channel and around the periphery of the microchip. This is thought to be because the adhesive film was thin, making it susceptible to minute shape abnormalities on the substrate surface. The mesh number is 730, the opening rate is 39%, and the printing speed is 200 mm / s. The conditions were as follows: the mesh number was 640, the opening ratio was 39%, and the printing speed was 200 mm / s, resulting in a film thickness of approximately 10 μm; the mesh number was 400, the opening ratio was 39%, and the printing speed was 200 mm / s, resulting in a film thickness of approximately 10 μm; The printing rate was 49%, the printing speed was 300 mm / s, and the film thickness was approximately 15 μm. As a result of fabricating the microchip 200, it was possible to achieve good bonding around the flow channel and the outer periphery in all cases. The mesh number is 400, the opening rate is 49%, and the printing speed is 200 mm / s. The film thickness is approximately When microchip 200 was fabricated using a screen plate with a thickness of 18 μm, the adhesive film was so thick that it flowed into the narrowed part of flow channel 211, making it impossible to send liquid to the flow channel. On the other hand, when distilled water was pumped into the flow channel of a microchip fabricated under conditions that allowed for good bonding, it was observed that the distilled water flowed only within the channel groove without leaking out of the channel. Next, the internal pressure strength against the pressure inside the channel was measured for the microchips bonded under the conditions of a film thickness of approximately 10 μm and a film thickness of approximately 15 μm. To measure the internal pressure strength, a small hole was drilled from the film side into the narrowed part of the channel 211 of the microchip 200, and an epoxy resin was poured in and hardened to block the flow. Distilled water was then continuously pumped using a pressure pump, and the peak pressure when the channel 211 was destroyed and the distilled water leaked out of the channel was read with a pressure sensor. The strength measurement results showed that the microchips exhibited pressure resistance up to internal pressures of 526 kPa and 643 kPa under the conditions of a film thickness of approximately 10 μm and a film thickness of approximately 15 μm, respectively. These results show that, although it depends on the shape of the flow channel and the surface condition of the microchip, by applying the adhesive and pressure-sensitive adhesive to a film thickness of 5 to 15 μm, it is possible to suppress the occurrence of voids and the inflow of adhesive into the flow channel groove, thereby enabling the bonding of microchips with good liquid transport into the flow channel and excellent pressure resistance. [Example]

[0048] <Microchip production 3> The optimum viscosity of the adhesive to be applied to the microchip was investigated. Except for the type of adhesive, the procedure was the same as that described in <Fabrication of Microchip 1> in Example 1. The adhesives used were SX-UV100A with a viscosity of 35,000 mPa·s, SX-UV100A diluted with butyl acetate with a viscosity of 31,000 mPa·s, UVX-8204 with a viscosity of 16,000 mPa·s, UVX-8400 with a viscosity of 8,300 mPa·s, U-1455B with a viscosity of 2,000 mPa·s, and NOA60 with a viscosity of 300 mPa·s. The screen used had a mesh count of 640, an opening rate of 39%, and a film thickness of approximately 10 μm. When each adhesive was applied to the substrate 201, it formed a fine uneven shape due to the mesh structure, but this gradually smoothed out (leveled) over time. After leveling, the substrate was attached to a film to produce a microchip 200, and its appearance was observed.

[0049] <Microchip Evaluation 3> At a viscosity of 35,000 mPa·s, numerous scratches occurred across the entire microchip, and the adhesive After lamination, voids were observed, which is thought to be due to insufficient transfer from the screen to the microchip due to the high viscosity of the adhesive. Viscosity 31,000mPa·s, 16,000mPa·s, 8,300mPa·s, 2,0 When distilled water was pumped through the channel of the fabricated microchip, it was observed that the distilled water flowed only through the channel groove without leaking out of the channel. At a viscosity of 300 mPa·s, the adhesive flowed into the narrowed portion of the channel 211 immediately after printing, making it impossible to transfer the liquid to the fabricated microchip. These results show that good screen printing is possible when the viscosity of the adhesive or pressure-sensitive adhesive is set to 2,000 to 31,000 mPa·s. [Example]

[0050] <Microchip production 4> Adhesive or pressure sensitive adhesive was applied to the microchip, and the peel strength of the fabricated chips was compared. The same method as described in <Fabrication of Microchip 1> in Example 1 was used for bonding, except that a pressure sensitive adhesive was used instead of an adhesive. A radical reactive acrylic UV curable adhesive was used to bond the substrate 201 and the film 202. The viscosity was 9,500 mPa·s. An adhesive was applied to the surface where the flow path and the solution storage section were provided by the following method: A UV-curable adhesive was applied by screen printing to the surface where the flow path and the solution storage section of the substrate 201 were provided. The screen used had a mesh count of 640, an opening ratio of 39%, and a film thickness of approximately 10 μm. The substrate 201 coated with the adhesive was dried at 95° C. for 15 minutes to remove the solvent contained in the adhesive. The solution reservoir on the adhesive-coated surface of the substrate 201 was laminated with the film 202, and a UV-LED light source was used to irradiate it with ultraviolet light of 365 nm wavelength for 10-20 seconds, initiating the adhesive curing reaction and bonding the film 202 onto the substrate 201 (Figure 3D).

[0051] <Microchip Evaluation 4> Observation of the fabricated microchip 200 revealed no inflow of adhesive into the flow channel. Furthermore, when distilled water was pumped into the flow channel, it was observed that the distilled water flowed only through the flow channel without leaking out of the flow channel. These results demonstrate that it is possible to manufacture microchips by applying a UV-curable adhesive to the areas of the substrate other than the flow path by screen printing, and then bonding it to a film.

[0052] The peel strength between the substrate 201 and film 202 of the fabricated microchip 200 was measured. The peel strength was measured by a 90° peel test using a small tabletop tester EZ-L (Shimadzu Corporation). As a result, the peel strength of the microchip fabricated with the UV-curable adhesive was 1.1 N / 26.2 mm, while the peel strength of the microchip 200 fabricated with the UV-curable pressure-sensitive adhesive was 3.0 N / 26.2 mm. Furthermore, The microchip 200, which was peeled off from the substrate 201 and film 202 and then re-applied with pressure, had a peel strength of 0.7 N / 26.2 mm. As a result of this experiment, it was observed that the distilled water flowed only through the channel groove without leaking out of the channel. These results show that the use of UV-curable adhesives makes it possible to improve the peel strength of microchips and also to re-form flow channels by re-adhesion after peeling. Although no reaction section is provided in the examples, the microchip of the present invention can be obtained by providing a reaction section in the middle of the flow channel. [Example]

[0053] <Microchip production 5> An adhesive was applied to the periphery of the flow path of the substrate, and a pressure-sensitive adhesive was applied to the outer periphery, to fabricate the microchip 300. Except for the areas where the adhesive and pressure-sensitive adhesive were applied, the process was carried out in the same manner as described in <Fabrication of Microchip 4> in Example 4. The adhesive was applied as follows. The adhesive UVX-8204 was applied by screen printing to the adhesive application area 315 around the flow path of the substrate 301 (A in FIG. 4). The area around the flow path on the substrate 301 was a 59.4 mm × 26.2 mm area located 3 mm inward from the short side of the substrate 301 on the waste liquid storage section 312 side, 1 mm inward from the short side on the hole side that would become the inlet 313, and 3 mm inward from the long sides on both sides (B in FIG. 4). The screen used had a mesh count of 640, an opening rate of 39%, and a theoretical film thickness of approximately 10 μm. The adhesive was applied as follows. Adhesive was applied to adhesive application area 303 on the outer periphery of film 302 using a small brush designed for applying adhesives and adhesives. The area near the outer periphery of the film was the outer region of a 59.4 mm × 26.2 mm rectangle, which was 3 mm inward from the short side of film 302 that corresponds to the waste liquid storage area 312 side of substrate 301 when bonded, 1 mm inward from the short side that corresponds to the hole side that becomes inlet 313, and 3 mm inward from the long sides on both sides (C in FIG. 4). The adhesive-coated portion 315 of the substrate 301 and the pressure-sensitive adhesive-coated portion 303 of the film 302 were attached together so as not to overlap. Next, a metal halide light source was used to irradiate ultraviolet light with a continuous wavelength distribution of 254 to 450 nm for 10 to 20 seconds, which initiated the curing reaction of the adhesive and pressure-sensitive adhesive, thereby bonding the film 302 onto the substrate 301 (Fig. 4D).

[0054] <Microchip Evaluation 5> The peel strength between substrate 301 and film 302 of fabricated microchip 300 was measured. As a result, the peel strength of microchip 300 was 7.0 N / 26.2 mm. Furthermore, the bond between substrate 301 and film 302 of microchip 300 was peeled off, and then pressure was applied again to adhere, and the peel strength of microchip 300 after pressure was applied again to adhere was 4.3 N / 26.2 mm. These results show that the peel strength of the microchip can be improved by fabricating the microchip using an adhesive around the flow path of the substrate and a pressure sensitive adhesive around the outer periphery. Furthermore, by using a UV-curable adhesive to bond the periphery of the flow channel and irradiating it with UV light in a nitrogen-filled environment, it is possible to suppress the inhibition of adhesive curing by oxygen and achieve complete curing. This makes it possible to increase the molecular weight of the adhesive polymer, which is expected to reduce the elution of small molecules from the adhesive into the flow channel. [Example]

[0055] <Microchip Fabrication 6> A substrate 101 (Zeon Corporation: COP resin) (size: 57 × 24 mm, thickness: 1 mm) shown in Figure 2A was prepared. The substrate 101 had two opposing channels 111 and 112. Channel 111 was a linear channel measuring 19 mm in length, 75 μm in depth, and 250 μm in width, which branched into two channels measuring 10 mm in length, 75 μm in depth, and 250 μm in width. The branched channel had a bent structure at a point 5 mm long of the total length of 10 mm. The linear channel and the branched channel had solution reservoirs 113 and 114 at the ends, respectively. Solution reservoir 113 at the end of the linear channel had a length of 11.5 mm, a depth of 100 μm, and a width of 4 mm. Solution reservoirs 114 at the ends of the branched channels both had a length of 5 mm, a depth of 100 μm, and a width of 3 mm. Flow path 112 has a structure in which a straight flow path with a length of 22 mm, a depth of 75 μm, and a width of 250 μm branches into two flow paths with a length of 12 mm, a depth of 75 μm, and a width of 250 μm. The ends of the straight flow path and the branched flow path have solution reservoirs 115 and 116, respectively. Solution reservoir 115 at the end of the straight flow path has a length of 10 mm, a depth of 100 μm, and a width of 3 mm. Solution reservoirs 116 at the ends of the branched flow paths both have a length of 4 mm, a depth of 100 μm, and a width of 3 mm. The film 102 in FIG. 2C was a COP film (size: 57×24 mm, thickness: 100 μm). For the film, Seiken Trepan (Kai Industries Co., Ltd.) was used, and φ2 mm through holes were drilled in a total of six places (three places x two) to align with the solution reservoirs of the substrate, and these were used as inlets 117 and air holes 118. The adhesive UVX-8204 was used to bond the substrate 101 and the film 102. As shown in FIG. 2B, the adhesive UVX-8204 was applied to the surface of the substrate 101 on which the flow path and solution storage section were provided, using the following method. The adhesive UVX-8204 was applied by screen printing to the surface of the substrate 101 on which the flow path and solution storage section were provided. The screen used had a mesh count of 730 and an opening rate of 39%. The adhesive was applied to a thickness of approximately 5 μm. The film was attached to the substrate 101 so that the through-holes in the film overlapped with the solution reservoir on the adhesive-coated surface of the substrate 101. Next, a metal halide light source was used to irradiate the film with ultraviolet light having a continuous wavelength distribution of 254 to 450 nm for 10 to 20 seconds, which initiated the curing reaction of the adhesive and bonded the film 102 onto the substrate 101 (Fig. 2D).

[0056] <Microchip Evaluation 6> Observation of the fabricated microchip 100 revealed no inflow of adhesive into the flow channel. Furthermore, when distilled water was pumped into the flow channel, it was observed that the distilled water flowed only through the flow channel without leaking out of the flow channel. These results demonstrate that it is possible to manufacture microchips with flow channel grooves of multiple shapes by applying a UV-curable adhesive to the area of ​​the substrate other than the flow channel by screen printing and then bonding it to a film. Note that although no reaction zones are provided in the reference example, the microchip of the present invention can also be obtained by providing any number of reaction zones in any area along the flow channel. [Example]

[0057] <Microchip production 7> A substrate 1 (Mitsubishi Chemical Corporation: acrylic resin) (size: 3.5 × 1.5 mm, thickness: 3 mm) shown in A of Fig. 1 was prepared. In the substrate 1, the length of the flow channel 11 was 7 mm, the depth was about 1 mm, and the width was 0.3 mm, and the reaction area was a circle with a diameter of 6 mm and a depth of about 1.8 mm. In addition, in the substrate 1, the holes serving as the inlet and outlet were through-holes with an inner diameter of 2 mm and a circular cross section.

[0058] Film 2 was a COP film (size 3.5 x 1.5 mm, thickness 100 μm), and when laminated with substrate 1, the area corresponding to the reaction section of flow channel 11 was coated with a hydrophilic reagent solution, S-1570. The concentration of the coated S-1570 and the coating method are as follows: A solution of S-1570 with a concentration of 0.1 wt% was placed in the region corresponding to the reaction section of the flow channel on the substrate 1. 1 μl was applied. The applied area was 12.56 mm 2 (diameter 4 mm), and the amount applied per area is 0.8 μl / mm 2 is. The applied hydrophilic reagent was allowed to dry naturally at room temperature for approximately 6 hours, and this was used as a hydrophilic treated film.

[0059] 12 μl of PT reagent (Sysmex Corporation) was dropped into the hydrophilic treated area. The applied PT reagent solution was spread evenly over the entire hydrophilized area (diameter 4 mm). The applied PT reagent was then dried at room temperature.

[0060] A stirring bar (length 5 mm, diameter 1 mm) was placed in the reaction section of the substrate 1 before bonding with an adhesive.

[0061] The adhesive UVX-8204 was used to bond the substrate 1 and the film 2. The adhesive UVX-8204 was applied to the surface of the substrate 1 on which the flow path and the reaction section were provided, by the following method. The adhesive UVX-8204 was applied by screen printing to the surface of the substrate 1 on which the flow path and reaction section were provided. The mesh number of the screen used was 730, the opening rate was 39%, and the adhesive coating thickness was approximately 5 μm. The substrate 1 and film 2 were attached together so that the reaction area on the adhesive-coated surface of the substrate 1 and the PT reagent-coated surface of the film 2 overlapped. Next, a metal halide light source was used to irradiate the film with ultraviolet light having a continuous wavelength distribution of 254 to 450 nm for 10-20 seconds to initiate the curing reaction of the adhesive, thereby bonding the film onto the substrate 1. The obtained microchip was left to stand at room temperature for 24 hours and then used in a blood coagulation test.

[0062] <Microchip Evaluation 7> The blood clotting time was evaluated using the fabricated microchip. Human standard plasma (SIEMENS) anticoagulated with sodium citrate and unfractionated plasma 50 μl of human standard plasma to which heparin (Mochida Pharmaceutical Co., Ltd.) was added at 1 U / mL was injected through the inlet and filled into the reaction section. The reaction section of the microchip was placed on a magnetic stirrer, and the stirring bar enclosed in the reaction section was rotated at a rotation speed of approximately 100 rpm. This caused the PT reagent coated on the film to mix with the plasma, initiating the coagulation reaction. As a fibrin clot formed, resistance to the stirring bar increased, causing the rotation speed to slow down and stop. The time from when the stirring bar started to rotate until it stopped was taken as the coagulation time.

[0063] The clotting time of standard plasma without heparin was 35 seconds, whereas that of standard plasma with heparin 1 U / ml was 35 seconds. The clotting time of plasma containing the compound was 1 minute 14 seconds. From the above, it was found that this microchip can be used to evaluate coagulation using plasma. [Example]

[0064] <Microchip production 8> A two-agent encapsulated microchip was fabricated in which different reagents were coated on the substrate reaction area and the film, respectively. Except for the coating of the reagents, the method was the same as that described in <Fabrication of Microchip 2> in Example 1. The reagent coating was carried out as follows.

[0065] 3.3 μl of In-tem reagent (Tem Innovations GmbH), which activates intrinsic blood coagulation, was dropped onto the hydrophilized area of ​​film 2. The In-tem reagent spread evenly throughout the coated area, and the film was then dried at room temperature. On the other hand, 3.3 μl of Star-tem reagent (Tem Innovations GmbH) was applied to the reaction area of ​​the substrate 1. After the Star-tem reagent (calcium chloride) had dried in the reaction zone of the substrate 1, a stirring bar (length 5 mm, diameter 1 mm) was placed therein. Next, adhesive UVX-8204 was applied, and the substrate 1 and the film were bonded by lamination and curing with ultraviolet light in the same manner as in Example 1. The obtained microchip was left to stand at room temperature for 24 hours and then used in a blood coagulation test.

[0066] <Microchip Evaluation 8> 50 μl of whole blood from a healthy donor collected in a vacuum blood collection tube (Terumo Corporation) containing 3.1% sodium citrate and blood supplemented with 0.5 U / ml of unfractionated heparin (Mochida Pharmaceutical) were injected into the inlet of the microchip obtained above, filling the reaction zone. The reaction zone of the microchip was placed on a magnetic stirrer, and the stirring bar enclosed in the reaction zone was rotated at approximately 100 rpm. This mixed the in-tem reagent coated on the film, the Star-tem reagent coated on the reaction zone, and the whole blood, initiating the clotting reaction. As the clotting reaction progressed, resistance to the stirring bar increased, causing the rotation speed to slow down and stop. The time from when the stirring bar started to rotate until it stopped was defined as the clotting time. The clotting time of healthy whole blood without heparin was 2 minutes 9 seconds, whereas the clotting time of whole blood containing 0.5 U / ml of heparin was 7 minutes 52 seconds. It is known that Int-tem and Start-tem reagents aggregate when mixed. By coating each agent so that they overlap in the reaction area of ​​the film and substrate, and stirring them in the reaction area during analysis, it was possible to create a two-agent encapsulated microchip that can analyze blood coagulation. [Explanation of symbols]

[0067] 10 Microchip, 1 Base material, 11 Flow channel, 12 Inlet, 13 Outlet, 14 Reaction section, 2 Film, 21 Reactant coating section

[0068] 100: Microchip; 101: Base material; 111, 112: Flow channel; 113, 114, 115, 116: Solution reservoir; 102: Film; 117: Inlet; 118: Air hole

[0069] 200: Microchip; 201: Substrate; 211: Flow path; 212: Waste liquid storage section; 213: Inlet; 214: Air hole; 202: Film

[0070] 300: Microchip, 301: Base material, 311: Flow path, 312: Waste liquid storage section, 313: Inlet port, 314: Air hole, 315: Adhesive application section, 302: Film, 303: Adhesive application section

Claims

1. A method for manufacturing a microchip for analyzing components in a sample by passing a liquid sample through a flow path provided inside the microchip and causing a reaction in a reaction region provided in a part of the flow path, the method comprising the steps of: preparing a substrate having a groove on its surface that serves as the flow path and a reaction region located in a part between both ends of the groove; applying an adhesive and / or a pressure sensitive adhesive to areas of the grooved surface of the substrate other than the grooves; providing a film having a reactive material applied to some areas; and A step of laminating a film onto a substrate so that the groove on the substrate is covered with the film to form a flow path, and the reactive portion of the adhesive and / or pressure-sensitive adhesive-applied surface of the substrate overlaps with the area of ​​the film to which the reactive substance is applied. wherein a method for applying the adhesive and / or pressure-sensitive adhesive to areas of the substrate other than the grooves is screen printing, the thickness of the adhesive and / or pressure-sensitive adhesive applied by screen printing is 5 to 15 μm, and the viscosity of the adhesive and / or pressure-sensitive adhesive applied by screen printing is 2,000 to 31,000 mPa s.

2. 10. The method of claim 1, wherein the substrate is one of plastic, silicone, and glass. Manufacturing method.

3. 3. The method according to claim 1, wherein the substrate or film has through holes serving as an inlet and an outlet at both ends of the channel formed by bonding the substrate and the film together, the through holes sandwiching the reaction section. The manufacturing method described in

4. The method according to any one of claims 1 to 3, wherein the surface of the substrate is hydrophilically treated, and an adhesive and / or a pressure-sensitive adhesive is applied to the hydrophilically treated surface.

5. The manufacturing method according to any one of claims 1 to 4, wherein the adhesive and / or pressure-sensitive adhesive is a UV-curable adhesive and / or pressure-sensitive adhesive.

6. The substrate having an adhesive and / or a pressure sensitive adhesive applied to its surface is attached to the film after a stirrer is placed at a position to be the reaction zone. The manufacturing method described above.

7. The method according to any one of claims 1 to 6, wherein the film is made of cycloolefin polymer (COP), cycloolefin copolymer (COC), polymethyl methacrylate (PMMA), polystyrene (PS), polycarbonate (PC), or polyethylene terephthalate (PET).

8. The method according to any one of claims 1 to 7, wherein the thickness of the film is 50 to 200 µm.

9. The method according to any one of claims 1 to 8, wherein the reactant is an antibody, an enzyme, a nucleic acid, a blood coagulation factor, or a bead containing any of them.

10. The manufacturing method according to any one of claims 1 to 9, wherein the film has been subjected to a hydrophilization treatment in an area where the reactive substance is to be applied, and the reactive substance is applied onto the hydrophilization-treated area.

11. The manufacturing method according to claim 10 , wherein the film is bonded to a substrate in which at least a portion of the groove is made hydrophilic.

12. The manufacturing method according to any one of claims 1 to 11, wherein an adhesive is applied to an inner region of the substrate excluding the outer peripheral portion, excluding a groove that becomes a flow path, and a pressure-sensitive adhesive is applied to the outer peripheral portion of the film, and the substrate and the film are bonded together with the adhesive and pressure-sensitive adhesive coated surfaces facing inward.

Images