Kit for detecting antigen and detection method

Abstract

To provide: a kit that can detect an allergen more reliably; and a detection method.SOLUTION: A kit for detecting an antigen comprises a support member, a base layer, and a polypeptide-containing layer disposed thereon. The base layer is a moisture-supplying layer that supplies moisture to the polypeptide-containing layer, or a moisture-permeable layer that permits moisture penetration. The support member or base layer comprises a fluorescent agent or a luminous agent.SELECTED DRAWING: None

Description

[Technical field]

[0001] The present disclosure relates to kits and methods for detecting antigens. [Background technology]

[0002] In recent years, allergy-related diseases have been increasing and become a social problem. In order to prevent allergies, it is important to detect allergens present in the environment. As a method for detecting allergens, a petri dish having a gel containing a polypolypeptide placed on a base layer is placed in a humid box for a predetermined period of time, stained, and the allergen attached to the gel is detected. The gel is decomposed by proteases in the allergens attached to the gel, and the decomposed part of the base layer is exposed, thereby detecting the allergens. [Prior art documents] [Patent documents]

[0003] [Patent Document 1] JP 2012-21980 A Summary of the Invention [Problem to be solved by the invention]

[0004] In the above-mentioned methods, the contrast between the gel layer containing the polypeptide and the base layer may be insufficient, making it difficult to detect allergens.

[0005] An object of the present disclosure is to provide a kit and a detection method that can detect allergens more reliably. [Means for solving the problem]

[0006] The present disclosure includes the following aspects. [1] A kit for detecting an antigen, comprising: a support member; a base layer; and a polypeptide-containing layer disposed thereon; the base layer is a moisture supplying layer that supplies moisture to the polypeptide-containing layer, or a moisture permeable layer that transmits moisture; The support member or base layer contains a fluorescent or phosphorescent material. kit. [2] The kit described in [1] above, wherein the base layer is colored. [3] The kit described in [1] above, wherein the base layer is colorless. [4] The kit described in [1] above, wherein the base layer contains talc or calcium sulfate. [5] The kit according to any one of the above [1] to [4], wherein the moisture supplying layer contains agar, a water-containing polymer, carrageenan, pectin or a water-containing gel. [6] The kit according to any one of the above [1] to [4], wherein the moisture-permeable layer is paper, a woven fabric, a nonwoven fabric, or a sponge. [7] The kit according to any one of the above [1] to [6], wherein the polypeptide-containing layer is colored. [8] The kit according to any one of the above [1] to [7], wherein the polypeptide-containing layer contains gelatin or a synthetic polypeptide. [9] The kit according to any one of the above [1] to [8], wherein the polypeptide-containing layer contains a protease activity promoter.

[10] The kit according to any one of the above [1] to [9], wherein the antigen is a cysteine ​​protease antigen, a serine protease antigen, an aspartic acid protease antigen, or a metalloprotease antigen.

[11] The kit according to any one of the above [1] to

[10] , wherein the polypeptide-containing layer contains a cysteine ​​protease inhibitor, a serine protease inhibitor, an aspartic acid protease inhibitor, or a metalloprotease inhibitor.

[12] The kit according to any one of the above [1] to

[11] , wherein the polypeptide-containing layer comprises a plurality of layers, the uppermost layer of which is colored.

[13] The kit according to any one of the above [1] to

[12] , wherein the support member contains a fluorescent agent or a luminescent agent.

[14] The kit according to any one of [1] to

[12] , wherein the base layer contains a fluorescent agent or a luminescent agent.

[15] The kit described in any one of [1] to

[14] , wherein the support member is a petri dish.

[16] The kit according to any one of the above [1] to

[15] , wherein the base layer and the polypeptide-containing layer are formed in a petri dish.

[17] Placing the kit according to any one of [1] to

[16] above at a detection location; detecting digestion marks on the surface of the kit; A method for detecting an antigen, comprising:

[18] The detection method according to

[17] above, wherein the antigen comprises a cysteine ​​protease antigen, a serine protease antigen, an aspartic acid protease antigen, or a metalloprotease antigen.

[19] The detection method according to

[18] or

[19] above, comprising placing the kit in a detection location and then placing the kit under a water vapor atmosphere.

[17] The kit includes a fluorescent agent in a base layer, The detection method according to any one of

[18] to

[19] above, comprising irradiating the kit with ultraviolet light in detecting digestion traces on the surface of the kit. Effect of the Invention

[0007] According to the kit and method of the present disclosure, a kit and a detection method capable of detecting allergens more reliably can be provided. [Brief description of the drawings]

[0008] [Figure 1] FIG. 1 shows the results of ultraviolet observation 10 minutes after the start of the test using Kit A in the example. [Diagram 2] FIG. 2 shows the results of ultraviolet observation 30 minutes after the start of the test for Kit A in the example. [Diagram 3] FIG. 3 shows the results of ultraviolet observation 60 minutes after the start of the test for Kit A in the example. [Figure 4]FIG. 4 shows the results of observation under fluorescent light 10 minutes after the start of the test for Kit A in the example. [Diagram 5] FIG. 5 shows the results of observation under fluorescent light 30 minutes after the start of the test for Kit A in the example. [Figure 6] FIG. 6 shows the results of observation under fluorescent light 60 minutes after the start of the test for Kit A in this example. [Figure 7] FIG. 7 shows the results of ultraviolet observation 10 minutes after the start of the test for Kit B in the example. [Figure 8] FIG. 8 shows the results of ultraviolet observation 30 minutes after the start of the test for Kit B in the example. [Figure 9] FIG. 9 shows the results of ultraviolet observation 60 minutes after the start of the test for Kit B in the example. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0009] Hereinafter, the embodiments of the present invention will be described in detail. However, the embodiments shown below are merely examples of kits and detection methods for embodying the technical ideas of the present invention, and the present invention is not limited to the kits and detection methods shown below. The upper and lower limits of the numerical ranges described in this specification can be arbitrarily selected and combined.

[0010] (Antigen detection kit) The present disclosure relates to A kit for detecting an antigen, comprising a support member, a base layer, and a polypeptide-containing layer disposed thereon, the base layer is a moisture supplying layer that supplies moisture to the polypeptide-containing layer, or a moisture permeable layer that transmits moisture; The support member or base layer contains a fluorescent or phosphorescent material. Provide a kit.

[0011] The kit of the present disclosure is placed at a place where the presence of an antigen (e.g., an allergen) is to be tested, and the antigen can be detected by observing the digestion marks, specifically the recesses or holes, formed in the polypeptide-containing layer. Specifically, in the kit of the present disclosure, the antigen-derived protease comes into contact with the polypeptide-containing layer, and the polypeptide in the polypeptide-containing layer is hydrolyzed by the protease activity, forming recesses or holes. In the kit of the present disclosure, the base layer below the polypeptide-containing layer is a moisture supplying layer or a moisture permeable layer, so that the hydrolysis reaction occurs easily. Therefore, the test result can be obtained easily and quickly. In addition, the kit of the present disclosure contains a fluorescent agent or a phosphorescent agent in the support member or base layer, so that the base layer can be easily viewed from the recesses or holes in the polypeptide-containing layer.

[0012] (antigen) Antigens that can be detected by the kits of the present disclosure are antigens that contain proteases (eg, protease-containing allergens).

[0013] The protease antigen contained in the antigen is not particularly limited, but examples thereof include cysteine ​​proteases, serine proteases, aspartic acid proteases, metalloproteases, trypsin-like proteases, chymotrypsin-like proteases, papain-like proteases, elastase-like proteases, peptidase-like proteases, collagenase-like proteases, and the like.

[0014] Protease-containing antigens can be, for example, insects, insect feces, and insect carcasses; and fragments thereof, where insects can be, for example, mites, cockroaches, etc.

[0015] Insects, insect droppings, and insect carcasses; as well as their debris, contain Der f1 / p1, Der f3 / p3, Der f6 / p6, or Der f9 / p9 in mites and Bla g2 in cockroaches.

[0016] The protease-containing antigen may be, for example, a fungus (mold) or a bacterium, etc. Here, the bacterium may be, for example, Bacillus subtilis, Staphylococcus aureus, etc.

[0017] Fungi (molds) and bacteria contain proteases such as Proteinase K in the case of Tritirachium albulim and subtilisin in the case of Bacillus subtilis.

[0018] The protease-containing allergen may be, for example, pollen, etc. Here, the plants from which the pollen is derived may be, for example, cedar, cypress, ragweed, pine, grass, mugwort, white birch (Betula platyphylla), alder, Alnus alpina, and Obayashabushi.

[0019] Pollen contains many proteins that are highly homologous to pollen allergens of other plants, such as Act c1 homologous protein and Cuc m1 homologous protein in the case of cedar pollen.

[0020] The protease-containing allergen may be, for example, house dust, etc. House dust is a mixture of various allergens, and may be a mixture of the above-mentioned various allergens (insects, insect feces, insect corpses, and their fragments; fungi (molds) and bacteria, etc.; pollen, etc.) in addition to dander and hair derived from humans and non-human animals.

[0021] Among house dust, dander and hair derived from humans and non-human animals may have serine proteases and cysteine ​​proteases derived from mites, molds, and fungi attached to their surfaces.

[0022] (Polypeptide-containing layer) The polypeptide-containing layer is a layer that contains a polypeptide. The polypeptide-containing layer contains a polypeptide that is a protease substrate, and therefore the polypeptide is decomposed by the protease activity derived from the allergen, and a recess or hole is formed by the decomposition. The allergen can be detected based on the presence or absence of such a recess or hole.

[0023] The polypeptide is not particularly limited as long as it can serve as a protease substrate.

[0024] The polypeptide to be used preferably has the following sequence within the molecule: Gln-Ala-Arg Phe-Ser-Arg Ala-Ala-Phe Ala-Ala-Pro-Phe Leu-Leu-Val-Tyr Lys-Phe-Gly-Gly-Ala-Ala Ala-Ala-Ala Ala-Pro-Ala Gly-Pro-Leu-Gly-Pro Gly-Pro-Gly-Gly-Pro-Ala Gly-Pro-Gln-Gly-Ile-Ala-Gly-Gln-Gln

[0025] The polypeptide preferably has a molecular weight of at least 200. The molecular weight of the polypeptide is more preferably 2 million to 10 million, and even more preferably 1 million to 1 million.

[0026] Examples of the polypeptide include gelatin, collagen, keratin, casein, albumin, etc., or a polypeptide having a structure similar to at least a portion of these, or a synthetic polypeptide, etc. The polypeptide is preferably gelatin or a synthetic polypeptide, and more preferably gelatin.

[0027] The polypeptide-containing layer preferably has a certain degree of shape retention. The shape retention of the polypeptide-containing layer makes it easier to retain the shape of the recesses or holes formed in the polypeptide-containing layer, making it possible to quantitatively evaluate the protease activity and thus the amount of antigen. In particular, the size and depth of the recesses or holes make it possible to make a relative comparison of the amount of antigen.

[0028] The polypeptide-containing layer may contain a crosslinking agent, which improves the shape retention of the polypeptide-containing layer.

[0029] The cross-linking agent is not particularly limited as long as it can cross-link peptides together, and examples thereof include glutaraldehyde, glutaraldehyde oligomer, succinaldehyde, and formaldehyde.

[0030] The proportion of the polypeptide contained in the polypeptide-containing layer is not particularly limited, but may be, for example, 70 to 100% by weight, preferably 80 to 90% by weight, of the substrate polypeptide in 100% by weight of the polypeptide-containing layer, calculated on a dry matter basis.

[0031] In one embodiment, the polypeptide-containing layer may contain a protease activator or a protease inhibitor that acts specifically on various proteases contained in the antigen. By containing a protease activity promoter in the polypeptide-containing layer, the hydrolysis reaction of the protease can be promoted, and thus the test time can be shortened. By using a protease activator or a protease inhibitor, it is possible to confirm whether a specific protease is present in the sample, and thus to identify the antigen.

[0032] An example of a protease activator is DTT.

[0033] Protease inhibitors include cysteine ​​protease inhibitors (eg, E-64), serine protease inhibitors (eg, AEBSF), aspartic protease inhibitors, or metalloprotease inhibitors.

[0034] The polypeptide-containing layer may be colored. The polypeptide-containing layer may be colored from the beginning, or may be dyed with a dye.

[0035] The dyes include red dyes, blue dyes, etc., such as Nylon Red, Nylon Blue, Brilliant Blue, Methylene Blue, Lanasin Black, Biebrich Scarlet, etc. The dye may preferably be Biebrich Scarlet.

[0036] In this specification, "colored" means having some color (including white), and colored may be transparent or opaque. For example, it is sufficient that the color is such that the presence of a polypeptide-containing layer can be recognized by visual inspection.

[0037] The thickness of the polypeptide-containing layer may be preferably 1 μm to 3 mm, more preferably 3 μm to 20 μm. The smaller the thickness of the polypeptide-containing layer, the easier it is to form holes in the polypeptide-containing layer, making detection quicker and easier. Also, the smaller the thickness of the polypeptide-containing layer, the darker the color of the polypeptide-containing layer, making it easier to detect recesses or holes.

[0038] The polypeptide-containing layer may be a single layer or multiple layers, for example two layers.

[0039] In one embodiment, the polypeptide-containing layer may have multiple layers, the top layer of which may be colored. For example, the polypeptide-containing layer may have a two-layer structure, with the bottom layer being colorless and the top layer being colored.

[0040] (Base layer) The base layer is a moisture-supplying layer that supplies moisture to the polypeptide-containing layer, or a moisture-permeable layer that transmits moisture to the polypeptide-containing layer.

[0041] The kit of the present disclosure has a base layer as described above, which promotes hydrolysis of the polypeptide in the polypeptide-containing layer, allowing for easy and rapid detection of the antigen.

[0042] The moisture supplying layer can retain moisture and supply the retained moisture to the polypeptide-containing layer, which promotes the hydrolysis reaction by the protease in the polypeptide-containing layer.

[0043] The moisture supply layer preferably contains agar, a water-containing polymer, carrageenan, pectin, or a water-containing gel (e.g., polyvinyl alcohol, polyacrylamide). These may be used in combination of two or more. The moisture supply layer more preferably contains agar.

[0044] The moisture supply layer may be a porous body containing water, such as paper (e.g., filter paper), woven fabric, nonwoven fabric, sponge, etc.

[0045] The thickness of the moisture supply layer is not particularly limited as long as it can hold a sufficient amount of moisture, but is preferably 1 mm or more, more preferably 3 mm or more, for example, 5 mm or more, or 10 mm or more. The upper limit of the thickness of the moisture supply layer is not particularly limited, and can be, for example, 20 mm or less, preferably 10 mm or less.

[0046] The moisture-permeable layer allows external moisture to permeate to the polypeptide-containing layer. The moisture that has permeated the moisture-permeable layer promotes the hydrolysis reaction caused by the protease in the polypeptide-containing layer.

[0047] The moisture-permeable layer is preferably a layer that actively takes in moisture from the outside and supplies it to the polypeptide-containing layer. For example, the moisture-permeable layer is preferably a layer that exerts capillary force.

[0048] The moisture-permeable layer may preferably be paper (e.g., filter paper), a woven fabric, a nonwoven fabric, or a sponge. Particularly preferably, the moisture-permeable layer is filter paper.

[0049] The base layer may contain a fluorescent agent or a luminescent agent. By containing a fluorescent agent or a luminescent agent in the base layer, the contrast with the polypeptide-containing layer is increased, and as a result, the depressions or holes can be easily detected.

[0050] The fluorescent agent has a fluorescent effect of absorbing ultraviolet rays from sunlight or artificial light, converting them into visible light, and radiating them. The fluorescent agent is preferably a fluorescent whitening agent. The fluorescent whitening agent can increase the whiteness and blueness of the base layer.

[0051] Examples of fluorescent agents include 7-diethylamino-4-methylcoumarin, 4,4'-bis(2-sulfostyryl)biphenyl disodium, dialkylaminocoumarin, bisdimethylaminostilbene, bismethylaminostilbene, 4-alkoxy-1,8-naphthalenedicarboxylic acid-N-alkylimide, bisbenzoxazolylethylene, 1,4-bis(2-benzoxazolyl)naphthalene, and dialkylstilbene.

[0052] The fluorescent agent may be a commercially available fluorescent agent. Commercially available fluorescent agents include, for example, Super Glo DF-300 (manufactured by Marktec), Kayahor 3BS Liq., Kayahor TAC Liq., Kayaphor HBC Liq., Kayalight B, Kayaphor AS 150, Kayaphor CR 200, Kayaphor JB Liq., Mikawhite ATN conc. (manufactured by Nippon Kayaku Co., Ltd.), TINOPAL OB, TINOPAL NFW. Liqid (manufactured by BASF), Hakkol P, Hakkol PHD, Hakkol RF, Hakkol PSR, Hakkol PSR-B, Hakkol CHP-B, Hakkol PY-2000, Hakkol PYB-D, Hakkol BE conc, Hakkol SG conc 150, Hakkol TH-100, Hakkol S-100, Hakkol RX-1, Hakkol OW-11, Hakkol OW-10extra, Hakkol KIP H / C, Hakkol SF, Hakkol JC, and Hakkol PMS H / C (manufactured by Showa Chemical Industry Co., Ltd.), Nikkafluor SB conc, Nikkafluor KB, Nikkafluor OB, and Nikkabright CX H / C (manufactured by Nippon Chemical Industry Co., Ltd.).

[0053] The fluorescent agents may be used alone or in combination.

[0054] The luminous agent is a substance that absorbs sunlight or artificial light and continues to emit light for a predetermined period of time after the absorption. The luminous agent can cause the base layer to emit light, thereby enhancing the contrast with the polypeptide-containing layer.

[0055] Examples of the luminescent agent include sulfide-based luminescent agents and aluminate-based luminescent agents. Specific examples of the sulfide-based luminescent agents include CaS:Bi, CaSrS:Bi, and ZnCdS:Cu. Specific examples of the aluminate-based luminescent agents include RMg2Al. 16 O 27 :Eu, RMg2Al16O 27 :Eu,Mn,RMg2Al10 O 17 :Eu, RMg2Al 10 O 17 :Eu,Mn (each R is independently Sr or Ba), SrAl2O4:Eu, SrAl2O4:Eu,Dy, Sr4Al 14 O 25 :Eu, Sr4Al 14 O 25 :Eu,Dy, CaAl2O4:Eu,Nd, etc.

[0056] The luminescent agents may be used alone or in combination.

[0057] In one embodiment, the base layer can be colored. In another embodiment, the base layer can be colorless.

[0058] The base layer may include talc or calcium sulfate, which can give the base layer a white color.

[0059] (Support member) The support member is a member that supports the polypeptide-containing layer and the base layer.

[0060] The support member is particularly limited as long as it is a member capable of supporting the polypeptide-containing layer and the base layer, and may be a petri dish, a plate, paper, etc. Also, it may be a combination of two or more of a petri dish, a sheet, a plate, paper, etc. For example, it may be a combination of a petri dish and a sheet.

[0061] In one embodiment, the support member may also serve as the base layer, for example the support member and base layer may be a sheet of filter paper.

[0062] The support member may contain a fluorescent agent or a luminous agent. For example, when the support member is a petri dish, the surface of the support member may be coated with a fluorescent agent or a luminous agent. Alternatively, a transparent petri dish may be combined with a fluorescent or luminous sheet. When the support member contains a fluorescent agent or a luminous agent, the base layer is preferably transparent.

[0063] (Method of detecting antigen) The present disclosure relates to placing the kit of the present disclosure at a detection site; Detecting traces of digestion on the surface of the kit; Includes.

[0064] (Place the kit at the detection location) In the detection method of the present disclosure, the kit is placed at a detection site, which means a location where it is desired to confirm the presence of an antigen.

[0065] When the kit of the present disclosure has a moisture supply layer, the hydrolysis reaction by the protease can proceed without any special operation, except for placing the kit at the detection site.

[0066] When the kit of the present disclosure has a moisture-permeable layer, for example, hydrolysis reaction by protease can proceed simply by contacting the base layer with water.

[0067] In one embodiment, the kit may be placed in a water vapor atmosphere after being placed at the detection location. In another embodiment, the kit may be placed in a water vapor atmosphere after being placed at the detection location, and then the kit may be placed in a location where water is present, for example, on a sponge impregnated with water. By placing the kit in a water vapor atmosphere or in a location where water is present, the hydrolysis reaction by the protease is promoted.

[0068] The kit of the present disclosure may be placed at the detection site, but alternatively, dust can be collected with an air sampler and the resulting dust can be inoculated into the kit to detect digestion traces.

[0069] (Detecting traces of digestion on the surface of the kit) In the detection method of the present disclosure, traces of digestion on the surface of the kit are detected. The term "digestion traces" refers to depressions or holes in the polypeptide-containing layer that are generated by a hydrolysis reaction caused by a protease.

[0070] The kit of the present disclosure includes a fluorescent agent or a luminescent agent in the support member or base layer. Therefore, the detection of digestion marks on the surface of the kit includes irradiating light, such as visible light or ultraviolet light, preferably ultraviolet light. According to this method, the base layer emits light, making it easy to detect digestion marks. In addition, even if the base layer is colorless and transparent, digestion marks can be easily detected.

[0071] The method for detecting the recesses or holes is usually performed with the naked eye, but is not particularly limited thereto, and for example, the recesses or holes may be magnified and observed using a telescope, an overhead projector, a scanner, etc. Alternatively, the recesses or holes can be detected by acquiring an image of the polypeptide-containing layer and analyzing the image.

[0072] According to the method of the present disclosure, the amount of antigen in a sample can be quantitatively evaluated by summing up the bottom surface area of ​​the recesses and holes formed in the polypeptide-containing layer.Also, the amount of antigen in a sample can be quantitatively evaluated by summing up the number of recesses and holes formed in the polypeptide-containing layer.Alternatively, the amount of antigen in a sample can be quantitatively evaluated based on both the sum of the bottom surface area of ​​the recesses and holes and the total number of recesses and holes.

[0073] The bottom area of ​​the recess may be measured, for example, by binarizing a photographed image of the polypeptide-containing layer by computer processing, and measuring the area of ​​the region corresponding to the bottom of the recess or hole as the number of pixels.

[0074] The kit and detection method of the present disclosure have been described in detail above. Note that the kit and detection method of the present disclosure are not limited to those exemplified above. EXAMPLES

[0075] The present disclosure will be described in further detail below with reference to examples, but the present disclosure is not limited to the following examples.

[0076] Example 1 An agar solution was prepared by mixing 50 mL of 1 M Tris-HCl aqueous solution, 1 g of agar, 5 g of talc, and 0.05 g of 7-diethylamino-4-methylcoumarin (fluorescent brightening agent A). The resulting agar solution was added to a plastic petri dish with a diameter of 60 mm and allowed to stand to gel, forming a base layer. A gelatin solution was prepared by mixing 10 mL of 1 M Tris-HCl aqueous solution, 0.5 g of gelatin, 20 μL of 1 M DTT, 10 μL of 25% glutaraldehyde, 3 mL of ethanol, and 0.03 g of Biebrich scarlet. The gelatin solution was added onto the base layer to form a polypeptide-containing layer, and antigen detection kit A was prepared. The resulting base layer had a thickness of 8 mm and the polypeptide-containing layer had a thickness of 10 μm.

[0077] Example 2 An antigen detection kit B was prepared in the same manner as in Example 1, except that talc was not added in forming the base layer.

[0078] Example 3 An antigen detection kit C was prepared in the same manner as in Example 1, except that 4,4'-bis(2-sulfostyryl)biphenyl disodium (fluorescent brightener B) was used instead of 7-diethylamino-4-methylcoumarin in forming the base layer.

[0079] Example 4 An antigen detection kit D was prepared in the same manner as in Example 3, except that talc was not added in forming the base layer.

[0080] Comparative Example 1 An antigen detection kit a was prepared in the same manner as in Example 1, except that no fluorescent whitening agent was added in forming the base layer.

[0081] (Experimental Example) An antigen (serine protease solution) was attached onto the antigen detection kit obtained above. After addition, the peptide-containing layer was observed for the presence or absence of digestion marks 10, 30, and 60 minutes later. Observation was performed under fluorescent light and ultraviolet light. The digestion marks were evaluated for visibility. The evaluation criteria are as follows. For reference, Kit A exposed to ultraviolet light for 10, 30, and 60 minutes, fluorescent light for 10, 30, and 60 minutes, and Kit B exposed to ultraviolet light for 10, 30, and 60 minutes are shown in Figures 1 to 9, respectively. 1: It was difficult to visually observe the digestive traces. 2: Digestion marks were visible. 3: Digestion marks were clearly visible. [Table 1]

[0082] Example 5 A luminescent agent (Shin-Roihi's compound luminescent paint) was applied to the bottom of a 90 mm diameter plastic petri dish. An agar solution was prepared by mixing 50 mL of 1 M Tris-HCl aqueous solution and 1 g of agar. The obtained agar solution was added to the above-mentioned petri dish and allowed to stand to gel, forming a base layer. A gelatin solution was prepared by mixing 10 mL of 1 M Tris-HCl aqueous solution, 0.5 g of gelatin, 20 μL of 1 M DTT, 10 μL of 25% glutaraldehyde, 3 mL of ethanol, and 0.03 g of Biebrich scarlet. The gelatin solution was added onto the base layer to form a polypeptide-containing layer, and antigen detection kit E was prepared. The resulting base layer had a thickness of 8 mm and the polypeptide-containing layer had a thickness of 10 μm.

[0083] Example 6 An antigen detection kit F was prepared in the same manner as in Example 5, except that a luminescent sheet was attached to the bottom surface of the petri dish instead of applying a luminescent agent.

[0084] Example 7 An antigen detection kit G was prepared in the same manner as in Example 5, except that a luminescent agent was added to the agar solution instead of applying the luminescent agent to the bottom surface of the petri dish.

[0085] Example 8 An antigen detection kit H was prepared in the same manner as in Example 5, except that a luminescent sheet was placed under the petri dish instead of applying a luminescent agent to the bottom surface of the petri dish.

[0086] (Experimental Example) An antigen (serine protease solution) was attached onto the antigen detection kit obtained above. After addition, the peptide-containing layer was observed in a dark place for the presence or absence of digestion marks 10 minutes, 30 minutes, and 60 minutes later. The visibility of the digestion marks was evaluated. The evaluation criteria were as follows. 1: It was difficult to visually observe the digestive traces. 2: Digestion marks were visible. 3: Digestion marks were clearly visible. [Table 2] [Industrial Applicability]

[0087] The antigen detection kit and detection method of the present disclosure are preferably used for detecting antigens, particularly allergens.

Claims

1. A kit for detecting an antigen, comprising a support member, a base layer, and a polypeptide-containing layer disposed thereon, the base layer is a moisture supplying layer that supplies moisture to the polypeptide-containing layer, or a moisture permeable layer that transmits moisture; The support member or base layer contains a fluorescent or phosphorescent material. kit.

2. The kit of claim 1 , wherein the base layer is colored.

3. The kit of claim 1 , wherein the base layer is colorless.

4. The kit of claim 1 , wherein the base layer comprises talc or calcium sulfate.

5. The kit of claim 1 , wherein the moisture supplying layer comprises agar, a hydrous polymer, carrageenan, pectin, or a hydrous gel.

6. The kit of claim 1 , wherein the moisture-permeable layer is paper, a woven fabric, a nonwoven fabric, or a sponge.

7. The kit of claim 1 , wherein the polypeptide-containing layer is colored.

8. The kit of claim 1 , wherein the polypeptide-containing layer comprises gelatin or a synthetic polypeptide.

9. The kit of claim 1 , wherein the polypeptide-containing layer comprises a protease activity promoter.

10. 2. The kit of claim 1, wherein the antigen is a cysteine ​​protease antigen, a serine protease antigen, an aspartic protease antigen, or a metalloprotease antigen.

11. 10. The kit of claim 1, wherein the polypeptide-containing layer comprises a cysteine ​​protease inhibitor, a serine protease inhibitor, an aspartic protease inhibitor, or a metalloprotease inhibitor.

12. The kit of claim 1 , wherein the polypeptide-containing layer comprises multiple layers, the top layer being colored.

13. The kit of claim 1 , wherein the support member comprises a fluorescent or phosphorescent agent.

14. The kit of claim 1 , wherein the base layer comprises a fluorescent or phosphorescent agent.

15. The kit of claim 1 , wherein the support member is a petri dish.

16. Placing the kit according to any one of claims 1 to 13 at a detection site; detecting digestion marks on the surface of the kit; A method for detecting an antigen, comprising:

17. 17. The method of claim 16, wherein the antigen comprises a cysteine ​​protease antigen, a serine protease antigen, an aspartic protease antigen, or a metalloprotease antigen.

18. The detection method according to claim 16, comprising placing the kit in a water vapor atmosphere after placing the kit at a detection location.

19. The detection method according to claim 16, wherein detecting digestion traces on the surface of the kit comprises irradiating with ultraviolet light.

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