Method for manufacturing analysis element and analysis element

Abstract

To provide an analysis element and a manufacturing method thereof that can prevent an object to be analyzed from being unable to be analyzed appropriately.SOLUTION: A manufacturing method of this analysis element 100 includes the steps of forming a mask layer 20, forming a membrane layer 30, exposing a substrate 10, forming an introduction hole 13 and forming a protrusion 14 at the inlet end 13a of the introduction hole 13, and removing the protrusion 14 by wet etching the protrusion 14.SELECTED DRAWING: Figure 13

Description

[Technical Field]

[0001] The present invention relates to a method for manufacturing an analytical element and an analytical element, and more particularly to a method for manufacturing an analytical element for analyzing an analyte by passing an electrolyte and an analyte through pores, and an analytical element. [Background technology]

[0002] BACKGROUND ART Conventionally, analytical elements for analyzing an analyte by passing the analyte through pores (fine holes) have been known (see, for example, Patent Document 1).

[0003] The above-mentioned Patent Document 1 discloses a chip (analysis element) for analyzing a biomolecular sample (analyte) by passing the biomolecular sample through a nanopore (pore). The chip includes a nanopore through which the biomolecular sample passes, and an introduction hole connected to the nanopore and having a tapered shape that gradually narrows from the inlet side toward the outlet side. When analyzing the analyte, an electrolyte solution and the biomolecular sample are introduced into the nanopore through the introduction hole and pass through the nanopore. A voltage is applied to the electrolyte solution by a pair of electrodes, and the current is monitored as the biomolecular sample passes through the nanopore. The biomolecular sample is analyzed based on the change in current as the biomolecular sample passes through the nanopore. [Prior art documents] [Patent documents]

[0004] [Patent Document 1] Japanese Patent Publication No. 2022-12168 Summary of the Invention [Problem to be solved by the invention]

[0005] In the chip described in Patent Document 1, the inlet holes for introducing the electrolyte (electrolyte solution) and the analyte (biomolecular sample) into the pores (nanopores) may be formed by wet etching. In this case, side etching may occur during wet etching. If side etching occurs, a protrusion that blocks the inflow of the electrolyte into the inlet hole may be formed at the inlet end of the inlet hole. Figure 34 shows a conventional analytical element 400 with a protrusion 414. The analytical element 400 includes a substrate 410, a mask layer 420 formed on a first surface 411 of the substrate 410, and a membrane layer 430 having a pore 431 formed on a second surface 412 of the substrate 410 opposite the first surface 411. The substrate 410 also includes an inlet hole 413 for introducing the electrolyte and the analyte into the pore 431. The inlet hole 413 has a tapered shape that gradually narrows from the inlet side toward the outlet side. At end 413a on the inlet side of introduction hole 413, a protrusion 414 is formed that protrudes from the outside toward the inside of introduction hole 413 and is covered with mask layer 420. In this case, protrusion 414 formed at end 413a on the inlet side of introduction hole 413 inhibits the flow of the electrolyte into introduction hole 413, making it difficult for the electrolyte and the analyte to be introduced into pore 431. This poses a problem in that it may not be possible to properly analyze the analyte.

[0006] The present invention has been made to solve the above-mentioned problems, and one object of the present invention is to provide an analytical element and a manufacturing method thereof that can avoid situations where the analysis of the subject to be analyzed cannot be performed properly. [Means for solving the problem]

[0007] In order to achieve the above object, a method for manufacturing an analytical element according to a first aspect of the present invention comprises the steps of: forming a first mask layer on a first surface of a substrate; exposing the substrate by forming a first hole in the first mask layer, the first hole being the inlet side; forming a membrane layer having a pore on the second surface of the substrate opposite the first surface; forming an introduction hole connected to the pore on the outlet side and having a tapered shape that gradually tapers from the inlet side toward the outlet side, and forming a protrusion at the inlet end of the introduction hole that protrudes from the outside toward the inside of the introduction hole and is not covered by the first mask layer; and removing the protrusion by wet etching.

[0008] In the method for manufacturing an analytical element according to the first aspect of the present invention, as described above, the protrusion is removed by wet etching. This allows the removal of a protrusion that blocks the inflow of electrolyte into the inlet end of the introduction hole due to side etching during wet etching. As a result, it is possible to avoid the protrusion formed at the inlet end of the introduction hole from blocking the inflow of electrolyte into the introduction hole. This prevents the electrolyte and the analyte from being difficult to introduce into the pore due to the blocking of the inflow of electrolyte into the introduction hole. As a result, it is possible to avoid the inability to properly analyze the analyte.

[0009] The method for manufacturing an analytical element according to the first aspect preferably includes the steps of: forming a second mask layer including an inner mask portion disposed to cover a portion of the substrate near the inner periphery of the first hole and having a second hole smaller than the first hole, and an outer mask portion covering the first mask layer; wet-etching the portion of the substrate exposed by the second hole to form an introduction hole and leaving the portion of the substrate covered by the inner mask portion to form a protrusion covered by the inner mask portion; and, after forming the introduction hole and the protrusion, removing the second mask layer to expose the first-side surface of the protrusion. With this configuration, after the protrusion is formed, which is covered by the inner mask portion of the second mask layer but not covered by the first mask layer, the second mask layer is removed to expose the first-side surface of the protrusion, making it easy to remove.

[0010] The method for manufacturing an analytical element according to the first aspect preferably includes the steps of: forming an introduction hole by wet etching the portion of the substrate exposed by the first hole, and forming a protrusion covered with the first mask layer by leaving the portion of the substrate covered with the first mask layer; and, after forming the introduction hole and the protrusion, removing the first mask layer on the protrusion to expose the first-side surface of the protrusion. With this configuration, after the protrusion covered with the first mask layer is formed, the first mask layer is removed to expose the first-side surface of the protrusion, making it easy to remove the protrusion.

[0011] The method for manufacturing an analytical element according to the first aspect preferably includes the steps of forming a depressed portion in the first mask layer that is thinner than other portions, forming a first hole that is positioned inside the depressed portion and is smaller than the depressed portion, and exposing the substrate, and wet-etching the portion of the substrate exposed by the first hole and the depressed portion to form an introduction hole and to form a protruding portion whose surface on the first surface side is exposed by removing the depressed portion. With this configuration, the protruding portion whose surface on the first surface side is exposed is formed by removing the depressed portion, and therefore the protruding portion can be easily removed.

[0012] In the above-described configuration for forming the second mask layer, the step of forming the second mask layer preferably includes the steps of: forming the second mask layer on the first mask layer and on the portions of the substrate exposed by the first holes; and dry-etching or wet-etching the second mask layer to form an inner mask portion having the second holes in the second mask layer. With this configuration, dry-etching or wet-etching can easily remove the inner portion of the second mask layer, and therefore the inner mask portion having the second holes can easily be formed in the second mask layer.

[0013] In the above-described configuration for forming the second mask layer, the first mask layer and the second mask layer are preferably formed of different materials, and the step of exposing the surface of the protrusion on the first side includes wet-etching the second mask layer, including the inner mask portion and the outer mask portion, using an etching solution that does not remove the first mask layer. This configuration allows the second mask layer to be selectively removed while the first mask layer remains properly. The inner mask portion is removed to form a protrusion with the surface on the first side (the surface of the substrate) exposed. As a result, the step of removing the protrusion can be performed properly.

[0014] In the method for manufacturing an analytical element according to the first aspect, the step of removing the protrusion preferably includes the step of wet-etching the protrusion to form a tapered shape at the inlet end of the introduction hole, the tapered shape having an inclination angle greater than the inclination angle of the introduction hole with respect to an imaginary line perpendicular to the first surface. With this configuration, the protrusion can be appropriately removed, and therefore it is possible to appropriately avoid the protrusion formed at the inlet end of the introduction hole from interfering with the flow of the electrolyte into the introduction hole.

[0015] In order to achieve the above object, an analytical element according to a second aspect of the present invention comprises a substrate, a mask layer formed on a first surface of the substrate and having a hole on the inlet side, and a membrane layer formed on a second surface opposite the first surface of the substrate and having a pore on the outlet side through which the electrolyte and the analyte pass, wherein the substrate includes an inlet hole for introducing the electrolyte and the analyte into the pore, and the inlet hole is connected to the pore on the outlet side and has a tapered shape that gradually tapers from the inlet side toward the outlet side, and includes an end portion located on the inlet side and having a tapered shape with an inclination angle greater than the inclination angle of the tapered shape of the inlet hole with respect to an imaginary line perpendicular to the first surface.

[0016] In the analytical element according to the second aspect of the present invention, as described above, the introduction hole is connected to the pore on the outlet side, has a tapered shape that gradually narrows from the inlet side toward the outlet side, and includes an end portion located on the inlet side and having a tapered shape with an inclination angle greater than the inclination angle of the tapered shape of the introduction hole with respect to an imaginary line perpendicular to the first surface. Thus, by wet-etching the protruding portion, the protruding portion is removed, resulting in an analytical element in which the introduction hole is located on the inlet side and includes an end portion having a tapered shape with an inclination angle greater than the inclination angle of the tapered shape of the introduction hole with respect to an imaginary line perpendicular to the first surface. As a result, an analytical element can be provided that can avoid problems that prevent the analysis of the analyte from being unable to be performed appropriately, similar to the manufacturing method of the analytical element according to the first aspect. [Effects of the Invention]

[0017] According to the present invention, it is possible to avoid the situation where the analysis target cannot be analyzed appropriately, as described above. [Brief explanation of the drawings]

[0018] [Figure 1] FIG. 1 is a schematic perspective view showing an analysis element according to a first embodiment. [Figure 2] FIG. 1 is a schematic cross-sectional view showing an analytical element according to a first embodiment. [Figure 3]FIG. 1 is a schematic diagram (1) illustrating a manufacturing process of the analytical element according to the first embodiment. [Figure 4] FIG. 2 is a schematic diagram (2) illustrating the manufacturing process of the analytical element according to the first embodiment. [Figure 5] FIG. 3 is a schematic diagram (3) for explaining the manufacturing process of the analytical element according to the first embodiment. [Figure 6] FIG. 4 is a schematic diagram (4) illustrating the manufacturing process of the analytical element according to the first embodiment. [Figure 7] FIG. 5 is a schematic diagram (5) for explaining the manufacturing process of the analytical element according to the first embodiment. [Figure 8] FIG. 6 is a schematic diagram (6) illustrating the manufacturing process of the analytical element according to the first embodiment. [Figure 9] FIG. 7 is a schematic diagram (7) for explaining the manufacturing process of the analytical element according to the first embodiment. [Figure 10] FIG. 8 is a schematic diagram (8) for explaining the manufacturing process of the analytical element according to the first embodiment. [Figure 11] FIG. 9 is a schematic diagram (9) for explaining the manufacturing process of the analytical element according to the first embodiment. [Figure 12] FIG. 1 is a schematic diagram (10) illustrating the manufacturing process of the analytical element according to the first embodiment. [Figure 13] FIG. 11 is a schematic diagram (11) illustrating the manufacturing process of the analytical element according to the first embodiment. [Figure 14] FIG. 10 is a schematic diagram (1) illustrating the manufacturing process of the analytical element according to the second embodiment. [Figure 15] FIG. 10 is a schematic diagram (2) illustrating the manufacturing process of the analytical element according to the second embodiment. [Figure 16] FIG. 10 is a schematic diagram (3) illustrating the manufacturing process of the analytical element according to the second embodiment. [Figure 17] FIG. 10 is a schematic diagram (4) illustrating the manufacturing process of the analytical element according to the second embodiment. [Figure 18] FIG. 5 is a schematic diagram (5) for explaining the manufacturing process of the analytical element according to the second embodiment. [Figure 19]FIG. 6 is a schematic diagram (6) for explaining the manufacturing process of the analytical element according to the second embodiment. [Figure 20] FIG. 7 is a schematic diagram (7) for explaining the manufacturing process of the analytical element according to the second embodiment. [Figure 21] FIG. 8 is a schematic diagram (8) for explaining the manufacturing process of the analytical element according to the second embodiment. [Figure 22] FIG. 9 is a schematic diagram (9) for explaining the manufacturing process of the analytical element according to the second embodiment. [Figure 23] FIG. 10 is a schematic diagram (10) illustrating the manufacturing process of the analytical element according to the second embodiment. [Figure 24] FIG. 10 is a schematic diagram (1) illustrating the manufacturing process of the analytical element according to the third embodiment. [Figure 25] FIG. 10 is a schematic diagram (2) illustrating the manufacturing process of the analytical element according to the third embodiment. [Figure 26] FIG. 10 is a schematic diagram (3) for explaining the manufacturing process of the analytical element according to the third embodiment. [Figure 27] FIG. 10 is a schematic diagram (4) illustrating the manufacturing process of the analytical element according to the third embodiment. [Figure 28] FIG. 5 is a schematic diagram (5) for explaining the manufacturing process of the analytical element according to the third embodiment. [Figure 29] FIG. 6 is a schematic diagram (6) illustrating the manufacturing process of the analytical element according to the third embodiment. [Figure 30] FIG. 7 is a schematic diagram (7) for explaining the manufacturing process of the analytical element according to the third embodiment. [Figure 31] FIG. 8 is a schematic diagram (8) illustrating the manufacturing process of the analytical element according to the third embodiment. [Figure 32] FIG. 9 is a schematic diagram (9) for explaining the manufacturing process of the analytical element according to the third embodiment. [Figure 33] FIG. 10 is a schematic diagram (10) illustrating the manufacturing process of the analytical element according to the third embodiment. [Figure 34] FIG. 1 is a schematic cross-sectional view illustrating a conventional analytical element. DETAILED DESCRIPTION OF THE INVENTION

[0019] Hereinafter, an embodiment of the present invention will be described with reference to the drawings.

[0020] [First embodiment] (Configuration of analytical element) The configuration of an analyzing element 100 according to a first embodiment of the present invention will be described with reference to FIGS.

[0021] 1 and 2, the analytical element 100 is an element for analyzing an analytical target such as a particle. The analytical element 100 includes a substrate 10, a mask layer 20, and a membrane layer 30. The mask layer 20 is an example of the "first mask layer" in the claims.

[0022] The substrate 10 is a silicon substrate.

[0023] The mask layer 20 is formed on the first surface 11 of the substrate 10. The mask layer 20 has holes 21 that serve as inlet holes. The material of the mask layer 20 is SiN. The holes 21 are an example of the "first holes" in the claims.

[0024] The membrane layer 30 is formed on the second surface 12 of the substrate 10, opposite the first surface 11. The membrane layer 30 has pores 31 on the outlet side through which the electrolyte and the analyte pass. The material of the membrane layer 30 is SiN. The pores 31 are smaller than the holes 21.

[0025] The substrate 10 includes an inlet 13 for introducing an electrolyte and an analyte into the pore 31. The inlet 13 is connected to the pore 31 on the outlet side and has a tapered shape that gradually narrows from the inlet side toward the outlet side. The inlet 13 includes an end 13a that is disposed on the inlet side and has a tapered shape with an inclination angle θ2 that is larger than the inclination angle θ1 of the tapered shape of the inlet 13 with respect to an imaginary line L that is perpendicular to the first surface 11.

[0026] When an analyte is analyzed using the analytical element 100, the electrolyte and the analyte are introduced into the pores 31 through the holes 21 and the introduction holes 13. A voltage is applied to the electrolyte by a pair of electrodes, and the current is measured when the analyte passes through the pores 31. The analyte is analyzed based on the change in current when the analyte passes through the pores 31. By analyzing the analyte using the analytical element 100, characteristics of the analyte, such as the number of particles, particle size, particle size distribution, and particle shape distribution, are obtained.

[0027] (Method of manufacturing an analytical element) A method for manufacturing the analytical element 100 will be described with reference to FIGS.

[0028] As shown in FIG. 3, a mask layer 20 is formed on a first surface 11 of a substrate 10, which is a silicon substrate. A membrane layer 30 is formed on a second surface 12 of the substrate 10, opposite the first surface 11. The process of forming the mask layer 20 and the process of forming the membrane layer 30 are performed as a common process. The material of the mask layer 20 and the membrane layer 30 is SiN. Then, as shown in FIGS. 4 to 6, the mask layer 20 and the membrane layer 30 are dry-etched or wet-etched to form holes 21 in the mask layer 20 and pores 31 in the membrane layer 30.

[0029] Specifically, as shown in FIG. 4, a resist layer 50 having holes 51 corresponding to the holes 21 is formed on the mask layer 20, and a resist layer 60 having pores 61 corresponding to the pores 31 is formed on the membrane layer 30. Then, as shown in FIG. 5, the portions of the mask layer 20 exposed by the holes 51 and the portions of the membrane layer 30 exposed by the pores 61 are removed by dry etching using an etching gas or wet etching using an etching solution. This forms the mask layer 20 having the holes 21 and the membrane layer 30 having the pores 31. Then, as shown in FIG. 6, the resist layer 50 and the resist layer 60 are removed. This forms the holes 21 on the inlet side in the mask layer 20, thereby exposing the substrate 10, as shown in FIG. 6. Furthermore, as shown in FIG. 6, a membrane layer 30 having the pores 31 on the outlet side is formed on the second surface 12 of the substrate 10, opposite the first surface 11.

[0030] Then, as shown in FIG. 7, a mask layer 40 is formed. Specifically, the mask layer 40 is laminated (formed) on the mask layer 20 and the portions of the substrate 10 exposed by the holes 21, for example, by sputtering. At this time, an outer mask portion 42 is formed on the mask layer 40. The material of the mask layer 40 contains any of SiO2, Cr, Pt, Au, Ag, Cu, and Ta. The mask layer 40 may be two or more layers (e.g., two layers of Cr / Au), and the thickness may not be uniform. The mask layer 20 and the mask layer 40 are formed of different materials. Then, as shown in FIGS. 8 to 10, the mask layer 40 is dry-etched or wet-etched to form an inner mask portion 41 having holes 41a in the mask layer 40. The mask layer 40 is an example of a "second mask layer" in the claims.

[0031] Specifically, as shown in FIG. 8, a resist layer 70 having holes 71 corresponding to the holes 41a is formed on the mask layer 40. Then, as shown in FIG. 9, the portions of the mask layer 40 exposed by the holes 71 are removed by dry etching using an etching gas or wet etching using an etching solution. This forms the mask layer 40 including an inner mask portion 41 having the holes 41a and an outer mask portion 42. Then, as shown in FIG. 10, the resist layer 70 is removed. This forms the mask layer 40 including the inner mask portion 41 having the holes 41a smaller than the holes 21 and arranged to cover the portions of the substrate 10 near the inner peripheries of the holes 21, and the outer mask portion 42 covering the mask layer 20, as shown in FIG.

[0032] As shown in FIG. 11 , the portion of the substrate 10 exposed by the hole 41a is wet-etched to form the introduction hole 13, which is connected to the pore 31 at the outlet side and has a tapered shape that gradually tapers from the inlet side to the outlet side. Furthermore, the portion of the substrate 10 covered by the inner mask portion 41 remains, forming a protrusion 14, covered by the inner mask portion 41, that protrudes from the outside of the introduction hole 13 toward the inside at the inlet end 13a of the introduction hole 13. Anisotropic wet etching is then performed to form the introduction hole 13, which has a tapered shape that gradually tapers from the inlet side to the outlet side. For example, KOH or TMAH (tetramethyl ammonium hydroxide) is used as an etchant for the anisotropic wet etching. Furthermore, side etching occurs during the anisotropic wet etching, resulting in the formation of the protrusion 14 at the inlet end 13a of the introduction hole 13.

[0033] Then, as shown in FIG. 12 , the mask layer 40 is removed to expose the surface of the protrusion 14 on the first surface 11 side. Specifically, the mask layer 40, including the inner mask portion 41 and the outer mask portion 42, is wet-etched using an etching solution that does not remove the mask layer 20, thereby removing the mask layer 40 including the inner mask portion 41 and the outer mask portion 42. The etching solution used for the wet etching is an acidic solution such as hydrofluoric acid, aqua regia, a Cr etching solution, or an Au etching solution. As shown in FIG. 12 , an introduction hole 13 is formed, which is connected to the pore 31 at the outlet side and has a tapered shape that gradually tapers from the inlet side toward the outlet side. Furthermore, a protrusion 14 that is not covered by the mask layer 20 and extends from the outside of the introduction hole 13 toward the inside is formed at the inlet end 13a of the introduction hole 13.

[0034] Then, as shown in FIG. 13, the protrusion 14 is removed by wet etching. Specifically, by wet etching the protrusion 14, a tapered shape is formed at the inlet end 13a of the introduction hole 13 with an inclination angle θ2 (see FIG. 2) larger than the inclination angle θ1 (see FIG. 2) of the introduction hole 13 relative to an imaginary line L (see FIG. 2) perpendicular to the first surface 11. In this case, the same etching liquid as that used to form the introduction hole 13 and the protrusion 14 may be used, or a different etching liquid may be used. Note that while FIGS. 2 and 13 show an example in which the inlet end 13a of the introduction hole 13 has a single inclination, the inlet end 13a of the introduction hole 13 may have two or more inclinations. In this manner, the analytical element 100 is manufactured.

[0035] (Effects of the first embodiment) In the first embodiment, the following effects can be obtained.

[0036] In the first embodiment, as described above, the protrusion 14 is removed by wet etching the protrusion 14. This makes it possible to remove the protrusion 14 even when the protrusion 14, which inhibits the inflow of the electrolyte into the inlet hole 13, is formed at the inlet end 13a of the inlet hole 13 due to side etching during wet etching. As a result, it is possible to avoid the protrusion 14 formed at the inlet end 13a of the inlet hole 13 from inhibiting the inflow of the electrolyte into the inlet hole 13. This makes it possible to avoid the electrolyte and the analyte being difficult to introduce into the pore 31 due to the inhibition of the inflow of the electrolyte into the inlet hole 13. As a result, it is possible to avoid the inability to properly analyze the analyte.

[0037] In the first embodiment, as described above, the method for manufacturing the analytical element 100 includes the steps of: forming a mask layer 40 including an inner mask portion 41 having a hole 41a smaller than the hole 21 and arranged to cover a portion of the substrate 10 near the inner periphery of the hole 21; and an outer mask portion 42 covering the mask layer 20; wet-etching the portion of the substrate 10 exposed by the hole 41a to form an introduction hole 13 and leaving the portion of the substrate 10 covered by the inner mask portion 41, thereby forming a protrusion 14 covered by the inner mask portion 41; and, after forming the introduction hole 13 and the protrusion 14, removing the mask layer 40 to expose the surface of the protrusion 14 facing the first surface 11. As a result, after the protrusion 14 is formed, which is covered by the inner mask portion 41 of the mask layer 40 but not covered by the mask layer 20, removing the mask layer 40 exposes the surface of the protrusion 14 facing the first surface 11, allowing the protrusion 14 to be easily removed.

[0038] In the first embodiment, as described above, the step of forming the mask layer 40 includes the steps of forming the mask layer 40 on the mask layer 20 and the portions of the substrate 10 exposed by the holes 21, and dry-etching or wet-etching the mask layer 40 to form the inner mask portions 41 having the holes 41a in the mask layer 40. In this way, if dry-etching or wet-etching is used, the inner portions of the mask layer 40 can be easily removed, and therefore the inner mask portions 41 having the holes 41a can be easily formed in the mask layer 40.

[0039] In the first embodiment, as described above, the mask layer 20 and the mask layer 40 are formed of different materials, and the step of exposing the surface of the protrusion 14 on the first surface 11 side includes a step of wet-etching the mask layer 40, including the inner mask portion 41 and the outer mask portion 42, using an etching solution that does not remove the mask layer 20. This allows the mask layer 40 to be selectively removed, while the mask layer 20 remains appropriately. The inner mask portion 41 is removed, and the protrusion 14 is formed with the surface on the first surface 11 side (the surface of the substrate 10) exposed. As a result, the step of removing the protrusion 14 can be performed appropriately.

[0040] In the first embodiment, as described above, the step of removing protrusion 14 includes a step of wet-etching protrusion 14 to form a tapered shape at inlet end 13a of introduction hole 13 with an inclination angle θ2 that is larger than the inclination angle θ1 of introduction hole 13 with respect to virtual line L that is orthogonal to first surface 11. This allows protrusion 14 to be appropriately removed, and therefore it is possible to appropriately prevent protrusion 14 formed at inlet end 13a of introduction hole 13 from hindering the flow of the electrolyte into introduction hole 13.

[0041] [Second embodiment] Next, a method for manufacturing the analyzing element 100 according to the second embodiment of the present invention will be described with reference to Figures 14 to 23. Note that the same components as those in the first embodiment are designated by the same reference numerals and will not be described again.

[0042] (Method of manufacturing an analytical element) As shown in FIG. 14, a mask layer 20 is formed on a first surface 11 of a substrate 10, which is a silicon substrate. A membrane layer 30 is formed on a second surface 12 of the substrate 10, opposite the first surface 11. The process of forming the mask layer 20 and the process of forming the membrane layer 30 are performed as a common process. The material of the mask layer 20 and the membrane layer 30 is SiN. Then, as shown in FIGS. 15 to 17, the mask layer 20 and the membrane layer 30 are dry-etched or wet-etched to form holes 21 in the mask layer 20 and pores 31 in the membrane layer 30.

[0043] Specifically, as shown in FIG. 15 , a resist layer 50 having holes 51 corresponding to the holes 21 is formed on the mask layer 20, and a resist layer 60 having pores 61 corresponding to the pores 31 is formed on the membrane layer 30. Then, as shown in FIG. 16 , the portions of the mask layer 20 exposed by the holes 51 and the portions of the membrane layer 30 exposed by the pores 61 are removed by dry etching using an etching gas or wet etching using an etching solution. This forms the mask layer 20 having the holes 21 and the membrane layer 30 having the pores 31. Then, as shown in FIG. 17 , the resist layer 50 and the resist layer 60 are removed. This forms the holes 21 on the inlet side in the mask layer 20, exposing the substrate 10, as shown in FIG. 17 . Furthermore, as shown in FIG. 17 , a membrane layer 30 having the pores 31 on the outlet side is formed on the second surface 12 of the substrate 10, opposite the first surface 11.

[0044] As shown in FIG. 18 , the portion of the substrate 10 exposed by the hole 21 is wet-etched to form the introduction hole 13, which is connected to the pore 31 at the outlet side and has a tapered shape that gradually tapers from the inlet side to the outlet side. Furthermore, the portion of the substrate 10 covered with the mask layer 20 remains, forming a protrusion 14 covered with the mask layer 20 at the inlet end 13a of the introduction hole 13, which protrudes from the outside to the inside of the introduction hole 13. Anisotropic wet etching is performed to form the introduction hole 13, which has a tapered shape that gradually tapers from the inlet side to the outlet side. For example, KOH or TMAH is used as the etching solution for the anisotropic wet etching. Furthermore, side etching occurs during the anisotropic wet etching, resulting in the formation of the protrusion 14 at the inlet end 13a of the introduction hole 13.

[0045] Then, as shown in FIG. 19 , a resist layer 270 is formed on the mask layer 20 and inside the introduction holes 13 by, for example, spray coating. Then, as shown in FIG. 20 , a resist layer 270 having holes 271 larger than the holes 21 is formed on the mask layer 20 by patterning, thereby exposing the mask layer 20 above the protrusions 14. Then, as shown in FIG. 21 , the mask layer 20 above the protrusions 14 is removed to expose the surface of the protrusions 14 on the first surface 11 side. Specifically, the mask layer 20 is dry-etched using an etching gas to remove the mask layer 20 above the protrusions 14. Furthermore, the holes 21 are widened by removing the mask layer 20 above the protrusions 14. Then, as shown in FIG. 22 , the resist layer 270 is removed. This forms an introduction hole 13 that is connected to the pore 31 on the outlet side and has a tapered shape that gradually narrows from the inlet side toward the outlet side, as shown in Figure 22, and also forms a protrusion 14 that is not covered by the mask layer 20 and extends from the outside toward the inside of the introduction hole 13 at the inlet end 13a of the introduction hole 13.

[0046] Then, as shown in FIG. 23, the protrusion 14 is removed by wet etching. Specifically, by wet etching the protrusion 14, a tapered shape is formed at the inlet end 13a of the introduction hole 13, with an inclination angle θ2 (see FIG. 2) larger than the inclination angle θ1 (see FIG. 2) of the introduction hole 13 relative to an imaginary line L (see FIG. 2) perpendicular to the first surface 11. At this time, the etching liquid used may be the same as or different from the etching liquid used to form the introduction hole 13 and the protrusion 14. In this manner, the analytical element 100 is manufactured.

[0047] (Effects of the second embodiment) In the second embodiment, as described above, the protrusions 14 are removed by wet etching the protrusions 14. This makes it possible to avoid the situation where the analysis target cannot be properly analyzed, as in the first embodiment.

[0048] In the second embodiment, as described above, the manufacturing method of the analytical element 100 includes the steps of wet-etching the portions of the substrate 10 exposed by the holes 21 to form the introduction holes 13 and leaving the portions of the substrate 10 covered with the mask layer 20, thereby forming the protrusions 14 covered with the mask layer 20, and, after forming the introduction holes 13 and the protrusions 14, removing the mask layer 20 on the protrusions 14 to expose the surfaces of the protrusions 14 on the first surface 11 side. As a result, after the protrusions 14 covered by the mask layer 20 are formed, the mask layer 20 is removed to expose the surfaces of the protrusions 14 on the first surface 11 side, so that the protrusions 14 can be easily removed.

[0049] Other effects of the second embodiment are the same as those of the first embodiment.

[0050] [Third embodiment] Next, a method for manufacturing the analyzing element 100 according to the third embodiment of the present invention will be described with reference to Figures 24 to 33. Note that the same components as those in the first embodiment are designated by the same reference numerals and description thereof will be omitted.

[0051] (Method of manufacturing an analytical element) As shown in FIG. 24, a mask layer 20 is formed on a first surface 11 of a substrate 10, which is a silicon substrate. A membrane layer 30 is formed on a second surface 12 of the substrate 10, opposite the first surface 11. The process of forming the mask layer 20 and the process of forming the membrane layer 30 are performed as a common process. The material of the mask layer 20 and the membrane layer 30 is SiN. Then, as shown in FIGS. 25 to 27, the mask layer 20 and the membrane layer 30 are dry-etched or wet-etched to form recesses 321 in the mask layer 20 that are thinner than other portions, and to form pores 31 in the membrane layer 30.

[0052] Specifically, as shown in FIG. 25 , a resist layer 50 having holes 51 corresponding to the depressions 321 is formed on the mask layer 20, and a resist layer 60 having pores 61 corresponding to the pores 31 is formed on the membrane layer 30. Then, as shown in FIG. 26 , the portions of the mask layer 20 exposed by the holes 51 are partially removed by dry etching using an etching gas or wet etching using an etching solution so that the portions are thinner than the other portions. This forms the mask layer 20 having the depressions 321. Furthermore, the portions of the membrane layer 30 exposed by the pores 61 are removed by dry etching using an etching gas or wet etching using an etching solution. This forms the membrane layer 30 having the pores 31. Then, as shown in FIG. 27 , the resist layer 50 and the resist layer 60 are removed. This forms the depressions 321 in the mask layer 20 that are thinner than the other portions, as shown in FIG. 27 . As shown in FIG. 27, a membrane layer 30 having pores 31 on the outlet side is formed on the second surface 12 of the substrate 10 opposite to the first surface 11.

[0053] Then, as shown in FIG. 28, a resist layer 370 is formed on the mask layer 20 having the depression 321. Then, as shown in FIG. 29, patterning is performed to form the resist layer 370 having a hole 371 smaller than the depression 321, thereby exposing the depression 321. Then, as shown in FIG. 30, a hole 21 is formed that is located inside the depression 321, is smaller than the depression 321, and exposes the substrate 10. At this time, dry etching is performed using an etching gas to remove the portion of the depression 321 exposed by the hole 371. Then, as shown in FIG. 31, the resist layer 370 is removed. Thereby, as shown in FIG. 31, the inlet side hole 21 is formed in the mask layer 20, thereby exposing the substrate 10.

[0054] As shown in FIG. 32 , an introduction hole 13 is formed, connected to the pore 31 at the outlet side and tapered from the inlet side toward the outlet side. A protrusion 14, not covered by the mask layer 20, is formed at the inlet end 13a of the introduction hole 13, extending from the outside toward the inside of the introduction hole 13. Specifically, the introduction hole 13 is formed by wet-etching the portion of the substrate 10 exposed by the hole 21 and the recess 321. The recess 321 is then removed to form the protrusion 14, exposing the surface on the first surface 11 side. Anisotropic wet etching is then performed to form the introduction hole 13, tapering from the inlet side toward the outlet side. For example, KOH or TMAH is used as the etchant for the anisotropic wet etching. Furthermore, side etching occurs during the anisotropic wet etching, resulting in the formation of the protrusion 14 at the inlet end 13a of the introduction hole 13. The recess 321 is removed due to differences in the thickness of the mask layer 20 and differences in etching rate. The thickness of the entire mask layer 20 is reduced, and the recessed portion 321, which was thinner than the other portions of the mask layer 20, is completely removed. In addition, by removing the recessed portion 321, the hole 21 is enlarged.

[0055] Then, as shown in FIG. 33, the protrusion 14 is removed by wet etching. Specifically, by wet etching the protrusion 14, a tapered shape is formed at the inlet end 13a of the introduction hole 13, with an inclination angle θ2 (see FIG. 2) larger than the inclination angle θ1 (see FIG. 2) of the introduction hole 13 relative to an imaginary line L (see FIG. 2) perpendicular to the first surface 11. At this time, the etching liquid used may be the same as or different from the etching liquid used to form the introduction hole 13 and the protrusion 14. In this manner, the analytical element 100 is manufactured.

[0056] (Effects of the third embodiment) In the third embodiment, as described above, the protrusions 14 are removed by wet etching the protrusions 14. This makes it possible to avoid the situation where the analysis target cannot be properly analyzed, as in the first embodiment.

[0057] In the third embodiment, as described above, the manufacturing method of analytical element 100 includes the steps of forming depressed portions 321 in mask layer 20 that are thinner than other portions, forming holes 21 that are positioned inside depressed portion 321 and are smaller than depressed portion 321, exposing substrate 10, and wet-etching the portion of substrate 10 exposed by hole 21 and depressed portion 321 to form introduction holes 13 and to form protruding portions 14 whose surfaces on the first surface 11 side are exposed by removing depressed portions 321. In this way, protruding portions 14 whose surfaces on the first surface 11 side are exposed are formed by removing depressed portions 321, and therefore protruding portions 14 can be easily removed.

[0058] Other effects of the third embodiment are the same as those of the first embodiment.

[0059] (Variation) The embodiments disclosed herein should be considered to be illustrative and not restrictive in all respects. The scope of the present invention is defined by the claims rather than the above description of the embodiments, and further includes all modifications (variations) within the meaning and scope of the claims.

[0060] For example, in the first embodiment described above, the mask layer (second mask layer) 40 is formed to include the inner mask portion 41 having the holes (second holes) 41a by dry etching or wet etching, but the present invention is not limited to this. In the present invention, the second mask layer including the inner mask portion and the outer mask portion having the second holes may be directly formed by sputtering or the like.

[0061] In the first embodiment, the mask layer 20 (first mask layer) and the mask layer 40 (second mask layer) are formed of different materials, but the present invention is not limited to this. In the present invention, the first mask layer and the second mask layer may be formed of the same material.

[0062] In addition, in the first to third embodiments, the mask layer 20 (first mask layer) is made of SiN, but the present invention is not limited to this. In the present invention, the first mask layer may be made of a material other than SiN.

[0063] In the first embodiment, the material of the mask layer 40 (second mask layer) includes any one of SiO2, Cr, Pt, Au, Ag, Cu, and Ta, but the present invention is not limited to this. In the present invention, the material of the second mask layer may include elements other than SiO2, Cr, Pt, Au, Ag, Cu, and Ta.

[0064] Furthermore, in the first to third embodiments, examples have been shown in which one introduction hole 13 and one pore 31 are provided, but the present invention is not limited to this. In the present invention, two or more introduction holes and two or more pores may be provided.

[0065] Furthermore, in the first to third embodiments, examples have been shown in which the introduction hole 13 is connected to the pore 31 (the introduction hole 13 is penetrated all the way to the pore 31) by one wet etching, but the present invention is not limited to this. In the present invention, the introduction hole may be connected to the pore (the introduction hole may be penetrated all the way to the pore) by two or more wet etchings. For example, the introduction hole may be connected to the pore (the introduction hole may be penetrated all the way to the pore) by a second wet etching that removes the protruding portion. [Explanation of symbols]

[0066] 10 Substrate 11 Page 1 12 Side 2 13 Inlet hole 13a Inlet end of introduction hole 14 Protrusion 20 Mask layer (first mask layer) 21 hole (1st hole) 30 membrane layer 31 Pore 40 Mask layer (second mask layer) 41 Inner mask part 41a hole (2nd hole) 42 Outer mask part 100 analytical element 321 Depression L Virtual Line θ1, θ2 Tilt angle

Claims

1. forming a first mask layer on a first surface of the substrate; forming a first hole serving as an entrance side in the first mask layer to expose the substrate; forming a membrane layer having pores on the outlet side on a second surface of the substrate opposite to the first surface; forming an introduction hole connected to the pore at the outlet side and having a tapered shape that gradually narrows from the inlet side toward the outlet side, and forming a protrusion at the inlet side end of the introduction hole that protrudes from the outside toward the inside of the introduction hole and is not covered by the first mask layer; and removing the protrusions by wet etching the protrusions.

2. forming a second mask layer including an inner mask portion having a second hole smaller than the first hole, the second mask portion being disposed to cover a portion of the substrate near an inner periphery of the first hole, and an outer mask portion covering the first mask layer; wet-etching the portion of the substrate exposed by the second hole to form the introduction hole, and leaving the portion of the substrate covered with the inner mask portion to form the protrusion covered with the inner mask portion; 2. The method for manufacturing an analytical element according to claim 1, further comprising the step of: after forming the introduction hole and the protrusion, removing the second mask layer to expose a surface of the protrusion on the first surface side.

3. wet etching the portion of the substrate exposed by the first hole to form the introduction hole, and leaving the portion of the substrate covered with the first mask layer to form the protrusion covered with the first mask layer; and after forming the introduction hole and the protrusion, removing the first mask layer on the protrusion to expose a surface of the protrusion on the first surface side.

4. forming a recessed portion in the first mask layer that is thinner than other portions; forming the first hole, which is disposed inside the recessed portion, is smaller than the recessed portion, and through which the substrate is exposed; 2. The method for manufacturing an analytical element according to claim 1, further comprising the steps of: wet-etching the portion of the substrate exposed by the first hole and the recessed portion to form the introduction hole; and removing the recessed portion to form the protrusion with the surface on the first surface side exposed.

5. The step of forming the second mask layer includes: forming a second mask layer over the first mask layer and the portion of the substrate exposed by the first hole; and forming the inner mask portion having the second hole in the second mask layer by dry etching or wet etching the second mask layer.

6. the first mask layer and the second mask layer are formed of different materials; 3. The method for manufacturing an analytical element according to claim 2, wherein the step of exposing the surface of the protrusion on the first surface side includes a step of wet-etching the second mask layer including the inner mask portion and the outer mask portion using an etching solution that does not remove the first mask layer.

7. 2. The method for manufacturing an analytical element according to claim 1, wherein the step of removing the protrusion includes a step of wet-etching the protrusion to form a tapered shape at the inlet end of the introduction hole having an inclination angle greater than an inclination angle of the introduction hole with respect to an imaginary line perpendicular to the first surface.

8. A substrate; a mask layer formed on the first surface of the substrate and having an inlet hole; a membrane layer formed on a second surface of the substrate opposite to the first surface, the membrane layer having pores serving as outlet sides through which an electrolyte and an analyte pass; the substrate includes an introduction hole for introducing the electrolyte and the analyte into the pore; the introduction hole is connected to the pore on the outlet side, has a tapered shape that gradually narrows from the inlet side toward the outlet side, and includes an end portion that is disposed on the inlet side and has a tapered shape with an inclination angle that is larger than the inclination angle of the tapered shape of the introduction hole with respect to an imaginary line perpendicular to the first surface.

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