Allergen Detection Kit

Abstract

To provide a simple and convenient kit for detecting an allergen, which can detect a food allergen even when a detection surface for wiping a sample is in a dry state.SOLUTION: A method in the allergen detection kit executes a preliminary selection step of calculating a passage rate of an allergen that has passed through the bottom surface of a resin body in which a developing solution prepared by adding a surfactant to PBS is dropped on the top surface of a resin constituting a wiping portion, and a secondary main selection step of detecting the allergen by using the allergen kit for an allergen-containing developing liquid that has flowed down in the resin and flowed out from the bottom of the resin, thereby selecting a combination of type of allergen, type of resin, and type of developing solution.SELECTED DRAWING: None

Description

[Technical Field]

[0001] The present invention relates to a method for selecting a combination of an allergen type, a resin type, and a developing solution type suitable for an allergen detection kit, the method comprising a preliminary selection step and a main selection step. [Background technology]

[0002] Allergies are harmful immune reactions caused by the ingestion of allergens (hereinafter referred to as "allergens") contained in foods, dust mites, etc., and are known to cause dermatitis, asthma, gastrointestinal disorders, anaphylactic shock, etc. In particular, since anaphylactic shock can be fatal in some cases, various methods for detecting allergens have been proposed.

[0003] Among these, immunoassays that utilize an antigen-antibody reaction to detect a substance to be detected that consists of a specific antigen or antibody are widely used, and in these immunoassays, antibodies or antigens labeled with a labeling substance such as a fluorescent substance are allowed to bind to the substance to be detected in a sample through an immune reaction, and the bound labeled substance is measured. In these immunoassays, competitive reactions and sandwich reactions are widely used, and allergen detection kits that use immunochromatography using sandwich reactions (see, for example, Patent Document 1) are commercially available.

[0004] The present applicant has proposed an immunochromatographic method (see, for example, Patent Document 2) that extracts food allergens using an extraction solution containing a denaturant and a reducing agent, suppressing nonspecific reactions associated with the breakdown of colloidal gold and enabling rapid and accurate allergen detection. The applicant has also proposed an immunochromatographic method (see, for example, Patent Document 3) that does not use 2-mercaptoethanol, which is designated as a toxic substance in Japan, but instead uses a developing solution containing a measurement sample of denatured and undenatured allergens extracted using an anionic surfactant and a thiosulfate, or an anionic surfactant and a nonionic surfactant, and detects allergens based on the presence or absence of gold colloid accumulation. Furthermore, the applicant has proposed an allergen detection method (see, for example, Patent Document 4) that involves subjecting a liquid sample, such as a cleaning solution or a rinsing solution, to immunochromatographic treatment. [Prior art documents] [Patent documents]

[0005] [Patent Document 1] Japanese Patent Application Publication No. 5-010950 [Patent Document 2] Japanese Patent Application Laid-Open No. 2007-278773 [Patent Document 3] International Publication No. WO2010 / 095469 Pamphlet [Patent Document 4] Japanese Patent Application Laid-Open No. 2017-129422 Summary of the Invention [Problem to be solved by the invention]

[0006] As described above, the present inventors have developed a method for rapid and accurate allergen detection by collecting cleaning water or rinse water from food production equipment as a liquid sample and subjecting it to immunochromatography. This method is specialized for detecting allergens in liquid samples by wiping the sample with a cotton swab or other device using a wiping solution or cleaning water. This method is extremely useful in that it allows for checking whether allergens remain after cleaning or rinsing. However, in food production sites, it may be necessary to check for the presence of allergens before cleaning, and some food production processes do not require the use of water. Furthermore, there are some areas that are not suitable for cleaning, such as packaging containers, electrical equipment, and warehouse fixtures. In such areas, it may be difficult to detect allergens using cleaning or rinsing liquid as a liquid sample. Even at home, for example, when attempting to detect mite allergens, wiping the wiped area with water is often inappropriate. Furthermore, kits used to detect allergens are frequently used in food manufacturing sites and the like, and therefore are required to be inexpensive and easy to process. However, it takes time and effort to select the resin for the wiping part and the developing liquid, etc., that can achieve both these characteristics and the accuracy of allergen detection, based on the allergen to be detected from a huge number of candidates.

[0007] The object of the present invention is to provide a simple and easy method for selecting a combination of the type of allergen, type of resin, and type of developing liquid suitable for an allergen detection kit with excellent detection accuracy when designing an allergen detection kit that is used when the detection surface from which the sample is wiped is free of moisture or is preferably in a dry state. [Means for solving the problem]

[0008] The inventors first selected polyethylene resin, known for its low cost, ease of processing, and durability. They then began their investigations using porous polyethylene resin, hoping for improved penetration of the developer. Various food allergens were applied to air-dried aluminum foil to create a detection surface. The dried detection surface was then wiped with dried porous polyethylene resin. Phosphate buffered saline (PBS), available from a commercially available wipe kit, was used as the developer. When the wiped surface of each resin was used as the top surface, the developer was applied to determine whether the developer containing the allergen penetrated the resin and could be recovered from the bottom surface of each resin. However, when the developer consisting of PBS was applied from the top surface of the polyethylene resin, the developer did not penetrate the polyethylene resin at all, and the developer containing the allergen could not be recovered from the bottom surface of the resin.

[0009] Therefore, in addition to polyethylene resin, polyester resin and polyvinyl alcohol resin were also investigated as resins suitable for allergen detection kits, and various developer solutions containing allergens were prepared as test solutions by adding various surfactants at various concentrations to PBS.Then, by combining each resin with each test solution and calculating the permeability of each allergen into the resin as the allergen penetration rate, we came up with the idea of ​​selecting a combination of resin type and developer type suitable for a simple and easy-to-use allergen detection kit for each allergen to be detected.

[0010] The test solution was a 500 μL developer solution containing 5 ppm of allergen, which was prepared by adding various surfactants to phosphate buffered saline at concentrations of 0.01%, 0.05%, 0.1%, 0.5%, or 1.0%. 2A combination of the type of allergen, the type of resin, and the type of developing liquid was selected such that, when dropped onto the top surface of a rectangular resin body 10 mm in height, the passage rate of allergens passing through the bottom surface of the resin body within 10 minutes was 75% or more.

[0011] We investigated whether allergens could be detected by loading the allergen-containing developer that flows down the resin and out the bottom of the resin onto a sample carrier for an immunochromatographic strip used in an allergen detection kit when the combination of allergen type, resin type, and developer type resulted in a passage rate of 75% or more of the allergen that passed through the bottom of the resin body. We found that even when the combination of allergen type, resin type, and developer type resulted in a high passage rate of 75% or more, 80% or more, 85% or more, 90% or more, or 95% or more of the allergen that passed through the bottom of the resin body, it was sometimes impossible to detect the allergen when using a kit loaded with a sample carrier for an immunochromatographic strip. Furthermore, when two types of polyethylene resin with different pore sizes were examined, different results were sometimes obtained.

[0012] Therefore, by carrying out a two-step process in which preliminary selection is performed as a simple first preliminary selection using the resin body, and then the allergen-containing developer liquid that has flowed down through the resin and out the bottom of the resin is supported on a sample carrier of an immunochromatographic strip to perform a second main selection step in which the allergen is detected, it has been confirmed that by going through this two-step process, it is possible to select a combination of the type of allergen, the type of resin, and the type of developer liquid that is suitable for an allergen detection kit with better detection accuracy, for example, capable of detecting allergens at a level of 2 μg / mL (ppm), and this has led to the completion of the present invention.

[0013] That is, the present invention is as specified by the following items. [1] A method for selecting a combination of an allergen type, a resin type, and a developing solution type suitable for an allergen detection kit, comprising the steps of (a) and (b) below. (a) A test solution containing 5 ppm of allergen was prepared by adding 500 μL of a developer solution containing a surfactant to phosphate buffered saline at a concentration of 0.01%, 0.05%, 0.1%, 0.5%, or 1.0%. The test solution was placed in a container with a top and bottom area of ​​35 mm 2 a step of pre-selecting a combination of the type of allergen, the type of resin, and the type of developing liquid, which, when dropped onto the top surface of a rectangular parallelepiped resin body having a height of 10 mm, results in a penetration rate of 75% or more of the allergen through the bottom surface of the resin body within 10 minutes; (b) conducting a test to confirm the detection of allergens in the preselected combinations, and finally selecting a combination of allergen type, resin type, and developer type suitable for the allergen detection kit, which is capable of detecting allergens at a level of 2 μg / mL; [2] The method according to [1] above, wherein the passage rate of allergens that have passed through the bottom surface of the resin body is 85% or more within 10 minutes. [3] The method according to [1] or [2] above, wherein the resin is porous. [4] The method according to any one of the above [1] to [3], wherein the resin is selected from polyester, polyethylene, and polyvinyl alcohol. [5] The method according to any one of [1] to [4] above, wherein the surfactant is polyoxyethylene (10) octylphenyl ether, sodium dodecyl sulfate, or polyoxyethylene sorbitan monolaurate. [6] The method according to any one of [1] to [5] above, wherein the allergen is a food allergen. [7] The method according to [6] above, wherein the food allergen is egg, milk, or wheat. [8] A combination of an allergen type, a resin type, and a developing solution type selected by the following steps (a) and (b) for use in an allergen detection kit. (a) A test solution containing 5 ppm of allergen was prepared by adding 500 μL of a developer solution containing a surfactant to phosphate buffered saline at a concentration of 0.01%, 0.05%, 0.1%, 0.5%, or 1.0%. The test solution was placed in a container with a top and bottom area of ​​35 mm 2 a step of pre-selecting a combination of the type of allergen, the type of resin, and the type of developing liquid, which, when dropped onto the top surface of a rectangular parallelepiped resin body having a height of 10 mm, results in a penetration rate of 75% or more of the allergen through the bottom surface of the resin body within 10 minutes; (b) conducting a test to confirm the detection of allergens in the preselected combinations, and finally selecting a combination of allergen type, resin type, and developer type suitable for the allergen detection kit, which is capable of detecting allergens at a level of 2 μg / mL (ppm); [9] A longitudinal case body having a developing support therein, on which are fixed at predetermined positions a labeled antibody obtained by binding a label to a monoclonal antibody capable of recognizing both denatured and native allergens, and a monoclonal antibody that recognizes both denatured and native allergens and recognizes an epitope different from that of the labeled antibody bound to the label; a lid for the case body having a wiping part made of resin attached to one end; and a developing liquid; In the allergen detection kit, the bottom of the wiping unit is located on one end of the developing support, and the wiping surface is disposed on the lid of the case body in a state where it penetrates the lid and protrudes outward, The test solution was prepared by adding 5 ppm of allergen to 500 μL of a phosphate buffered saline solution containing a surfactant at a concentration of 0.01%, 0.05%, 0.1%, 0.5%, or 1.0%. The test solution was placed in a container with a top and bottom area of ​​35 mm 2the allergen detection kit comprises the steps of: pre-selecting a combination of an allergen type, a resin type, and a developing liquid type that, when dropped onto the top surface of a rectangular resin body 10 mm in height, results in a passage rate of 75% or more of the allergen passing through to the bottom surface of the resin body within 10 minutes; and conducting an allergen detection confirmation test for the pre-selected combination to finally select a combination of an allergen type, a resin type, and a developing liquid type that is suitable for the allergen detection kit and that is capable of detecting allergens at a level of 2 μg / mL; and the allergen detection kit is characterized in that it detects allergens using a wiping part made of a resin selected by the method for selecting a combination of an allergen type, a resin type, and a developing liquid type that is suitable for the allergen detection kit, and the selected developing liquid.

[10] A longitudinal case body having a developing support therein, on which are fixed at predetermined positions a labeled antibody obtained by binding a label to a monoclonal antibody capable of recognizing both denatured and native allergens, and a monoclonal antibody that recognizes both denatured and native allergens and recognizes an epitope different from that of the labeled antibody bound to the label; a lid for the case body having a wiping part made of resin attached to one end; and a developing liquid; In the allergen detection kit, the bottom of the wiping unit is located on one end of the developing support, and the wiping surface is disposed on the lid of the case body in a state where it penetrates the lid and protrudes outward, The allergen detection kit is characterized in that a selected allergen is detected using a wiping part made of a resin selected by the method described in any one of [1] to [7] above and a selected developing liquid.

[11] The kit according to the above [9] or

[10] , wherein the resin is porous.

[12] The kit according to any one of the above [9] to

[11] , wherein the resin is selected from polyester, polyethylene, and polyvinyl alcohol.

[13] The kit according to any one of the above [9] to

[12] , wherein the developing solution contains polyoxyethylene (10) octylphenyl ether, sodium dodecyl sulfate, or polyoxyethylene sorbitan monolaurate.

[14] A kit according to any one of [9] to

[13] above, characterized in that the case body and the lid are engaged at the end far from the wiping portion, and the developing support can be removed after measurement by opening and closing the case body and the lid at the end near the wiping portion. [Effects of the Invention]

[0014] According to the method of the present invention, from the vast number of combinations of allergen types, resin types, and developer types, a combination of allergen types, resin types, and developer types that results in an allergen penetration rate of 75% or more through the bottom of the resin body is pre-selected by the very simple method of dripping an allergen-containing developer solution (test solution) onto a resin body, and then the allergen-containing developer solution that flows down through the resin and out the bottom of the resin is supported on a sample carrier of an immunochromatographic strip, thereby performing the final selection to detect the allergen using an allergen kit.This makes it possible to select a combination of allergen types, resin types, and developer types that is suitable for an allergen detection kit inexpensively and with little effort. [Brief explanation of the drawings]

[0015] [Figure 1] This graph shows the rate at which allergens passed through the bottom surface of each of the resin bodies (a) PES, (b) PE1, (c) PE2, and (d) PVA within 10 minutes after 500 μL of a test solution containing 5 ppm egg allergens, prepared by dissolving egg allergens in a PBS solution containing Triton X-100, was dropped onto the top surface of the resin body. [Figure 2] This graph shows the rate at which allergens passed through the bottom surface of each of the resin bodies (a) PES, (b) PE1, (c) PE2, and (d) PVA within 10 minutes after 500 μL of a test solution containing 5 ppm egg allergens, prepared by dissolving egg allergens in a PBS solution containing added SDS, was dropped onto the top surface of the resin body. [Figure 3]This graph shows the rate at which allergens passed through the bottom surface of each of the resin bodies (a) PES, (b) PE1, (c) PE2, and (d) PVA within 10 minutes after 500 μL of a test solution containing 5 ppm egg allergens, prepared by dissolving egg allergens in a PBS solution containing Tween 20, was dropped onto the top surface of the resin body. [Figure 4] This graph shows the rate at which allergens passed through the bottom surface of each of the resin bodies (a) PES, (b) PE1, (c) PE2, and (d) PVA within 10 minutes after 500 μL of a test solution containing 5 ppm of milk allergen, which was prepared by dissolving milk allergens in a PBS solution containing Triton X-100, was dropped onto the top surface of the resin body. [Figure 5] This graph shows the rate at which allergens passed through the bottom surface of each of the resin bodies (a) PES, (b) PE1, (c) PE2, and (d) PVA within 10 minutes after 500 μL of a test solution containing 5 ppm of milk allergens, which was prepared by dissolving milk allergens in a PBS solution containing added SDS, was dropped onto the top surface of the resin body. [Figure 6] This graph shows the rate at which allergens passed through the bottom surface of each of the resin bodies (a) PES, (b) PE1, (c) PE2, and (d) PVA within 10 minutes after 500 μL of a test solution containing 5 ppm of milk allergens, which was prepared by dissolving milk allergens in a PBS solution containing added Tween 20, was dropped onto the top surface of the resin body. [Figure 7] This graph shows the rate at which allergens passed through the bottom surface of each of the resin bodies (a) PES, (b) PE1, (c) PE2, and (d) PVA within 10 minutes after 500 μL of a test solution containing 5 ppm wheat allergens, which was prepared by dissolving wheat allergens in a PBS solution containing Triton X-100, was dropped onto the top surface of the resin body. [Figure 8] This graph shows the rate at which allergens passed through the bottom surface of each of the resin bodies (a) PES, (b) PE1, (c) PE2, and (d) PVA within 10 minutes after 500 μL of a test solution containing 5 ppm wheat allergens, which was prepared by dissolving wheat allergens in a PBS solution containing added SDS, was dropped onto the top surface of the resin body. [Figure 9]This graph shows the rate at which allergens passed through the bottom surface of each of the resin bodies (a) PES, (b) PE1, (c) PE2, and (d) PVA within 10 minutes after 500 μL of a test solution containing 5 ppm wheat allergens, which was prepared by dissolving wheat allergens in a PBS solution containing Tween 20, was dropped onto the top surface of the resin body. [Figure 10] 1 shows a conceptual diagram of the main body case of the kit of the present invention. DETAILED DESCRIPTION OF THE INVENTION

[0016] The method for selecting a combination of the type of allergen, the type of resin, and the type of developer suitable for the allergen detection kit of the present invention is as follows: (a) A test solution containing 5 ppm of allergen in 500 μL of developer solution prepared by adding a surfactant to phosphate buffered saline, the surfactant concentration of which is 0.01%, 0.05%, 0.1%, 0.5%, or 1.0%, is placed on a tray with a top and bottom area of ​​35 mm 2 a step of pre-selecting a combination of the type of allergen, the type of resin, and the type of developing liquid, which, when dropped onto the top surface of a resin body that is a rectangular parallelepiped resin with a height of 10 mm, results in a passage rate of 75% or more of the allergen passing through the bottom surface of the resin body within 10 minutes; (b) conducting an allergen detection confirmation test for the preselected combination and selecting a combination of allergen type, resin type, and developing liquid type suitable for the allergen detection kit that can detect allergens at a level of 2 μg / mL; and the method is not particularly limited as long as it comprises the steps of: (a) conducting an allergen detection confirmation test for the preselected combination and selecting a combination of allergen type, resin type, and developing liquid type that can detect allergens at a level of 2 μg / mL; and the combination of allergen, resin that allows the allergen to pass through the bottom surface at a rate of 75% or more, and developing liquid can be easily obtained by conducting experiments by changing the type of resin that makes up the wiping part and the type of developing liquid (such as the type and concentration of surfactant added to PBS) for each allergen.

[0017] In the step (a) above, the step of preliminarily selecting a combination of the type of allergen, the type of resin, and the type of developing solution is carried out by filling a test solution containing 5 ppm of allergen in 500 μL of a developing solution prepared by adding a surfactant to phosphate buffered saline, the surfactant concentration being 0.01%, 0.05%, 0.1%, 0.5%, or 1.0%. 2 One example of a method for calculating the allergen penetration rate is to select a combination of the type of allergen, the type of resin, and the type of developing liquid that, when dropped onto the top surface of a 10 mm high rectangular parallelepiped resin (resin body), results in an allergen transmittance of 75% or more within 10 minutes.The method for calculating the allergen penetration rate is to collect the developing liquid that has passed through the bottom surface of the resin body to form a collected sample, and determine the concentration of the allergen to be detected in the collected sample using a well-known antigen concentration determination method such as ELISA.Specifically, one example of a method for calculating the penetration rate is the ratio of the concentration of the allergen that has passed through the bottom surface of the resin body, which can be expressed by the following formula.

[0018]

number

[0019] The resin is not particularly limited as long as it is a known resin that allows the allergens to pass through at least 75%, and examples thereof include polyester, polyethylene, polyvinyl alcohol, and the like.

[0020] The polyester is preferably a porous polyester, and the porosity of the porous polyester is, for example, 60 to 90%, preferably 70 to 85%, and more preferably 73 to 83%.

[0021] The polyethylene is preferably porous polyethylene, more preferably porous polyethylene produced by a sintering method, and the average pore size of the porous polyethylene is 50 μm to 250 μm, preferably 75 μm to 225 μm, or 50 μm to 150 μm, preferably 75 μm to 125 μm, or 150 μm to 250 μm, preferably 175 μm to 225 μm. The porosity of the porous polyethylene is 15% to 90%, preferably 20 to 70%, more preferably 30% to 50%.

[0022] The polyvinyl alcohol is preferably porous, with an average pore size of 50 μm to 250 μm, preferably 75 μm to 225 μm, more preferably 100 μm to 200 μm, and even more preferably 125 μm to 175 μm. The porosity of the porous polyvinyl alcohol is 40% to 95%, preferably 60% to 95%, and more preferably 80% to 92%.

[0023] The allergen is not particularly limited as long as it is any antigenic protein or peptide capable of inducing antibody production, and examples thereof include food allergens such as eggs, milk, meat such as beef, fish such as salmon and tuna, crustaceans and mollusks such as shrimp and crab, grains, beans and nuts, fruits, vegetables, brewer's yeast, and gelatin. Among these, αs1 casein, which is the main component of milk allergens, β-lactoglobulin, which is the main component of whey allergens, and onion, which is the main component of egg white allergens, are particularly suitable. These allergens include food allergens such as voalbumin and ovomucoid, gliadin, the main component of wheat allergens, proteins with molecular weights of 24kDa and 76kDa that are the main proteins in buckwheat, AraH1, the main protein in peanuts, soybean 7S globulin contained in soybeans, sesame 11S globulin contained in sesame, and tropomyosin contained in crustaceans, as well as allergens such as house dust, mites and mite excrement, pollen, animal hair, epithelium and excrement, insects, and mold.

[0024] The developing solution is not particularly limited as long as it is a developing solution in which a surfactant has been added to phosphate buffered saline, and 500 μL of the developing solution containing 5 ppm allergen is dropped onto the top surface of the resin body as a test solution, and within 10 minutes, the proportion of allergen that has passed through to the bottom surface of the resin body is 75% or more. Examples of the developing solution include a developing solution containing phosphate buffered saline, and preferred examples include a developing solution containing phosphate buffered saline and polyoxyethylene (10) octylphenyl ether, sodium dodecyl sulfate, or polyoxyethylene sorbitan monolaurate.

[0025] The phosphate buffered saline may include well-known buffer solutions containing sodium chloride, potassium chloride, sodium phosphate, and potassium phosphate, which are commonly used in biological research, and may also include PBS(+) containing calcium and magnesium, and PBS(-) which does not contain calcium or magnesium.

[0026] An example of the polyoxyethylene (10) octylphenyl ether is a nonionic surfactant commonly available under the trade name Triton X-100. Sodium dodecyl sulfate (SDS) is widely known as an anionic surfactant. An example of the polyoxyethylene sorbitan monolaurate is a nonionic surfactant commonly available under the trade name Tween 20.

[0027] Using a combination of allergen, resin and developing solution that results in an allergen passage rate of 75% or more in the pre-selection step of step (a), an allergen detection confirmation test for the pre-selected combination of step (b) is conducted, and taking into account the test results, the process can proceed to the final selection step of a combination of allergen type, resin type and developing solution type that is suitable for the allergen detection kit.

[0028] The step (b) of the present invention, in which a combination of allergen type, resin type and developing solution type suitable for the allergen detection kit is selected, is not particularly limited as long as it is a step of conducting an allergen detection confirmation test for the preselected combination and selecting a combination of allergen type, resin type and developing solution type that can detect the allergen at a level of 2 μg / mL.The allergen detection confirmation test can be performed in the allergen detection kit that is expected to be used, using the combination of allergen type, resin type and developing solution type preselected in step (a).

[0029] The allergen detection kit comprises a longitudinal case body having a developing support therein, fixed at predetermined positions, a labeled antibody obtained by binding a label to a monoclonal antibody capable of recognizing both denatured and native allergens, and a monoclonal antibody that recognizes both denatured and native allergens and recognizes an epitope different from that of the labeled antibody bound to the label; a lid for the case body, one end of which is fitted with a wiping part made of resin; and a developing solution. An example of an allergen detection kit is one in which the bottom of the wiping portion is located on one end of the unfolding support, and the wiping surface is positioned on the lid of the case body with the wiping surface passing through the lid and protruding outward.

[0030] The detection surface to be wiped by the wiping unit is not particularly limited as long as it is a location where allergens may be detected, but examples include dry, moisture-free surfaces at food production sites where allergens may adhere or remain, such as food production equipment, components of the food production equipment, floors, walls, windows, and packaging containers. Other examples include furniture and fixtures such as tables and chairs at home or school, floors, walls, windows, doorknobs, bedding, and curtains. Detection surfaces also include detection surfaces from which the water in cleaning liquid or rinsing liquid has evaporated, but moisture may remain on the detection surface to be wiped as long as the effects of the present invention are achieved.

[0031] The wiping portion made of the above-mentioned resin is not particularly limited as long as its bottom surface is located on one end of the unfolding support, and its wiping surface is inserted through the lid of the case body and engaged with the lid in a state where it protrudes outward, and the wiping portion is made of a resin selected from resins having an allergen penetration rate of 75% or more in the pre-selection step, and for convenience, this also includes cases where the wiping portion has other parts such as a support such as a frame that supports the resin (part).

[0032] The wiping surface of the resin constituting the wiping portion is inserted into the lid of the detection case and engaged with the lid while protruding outward, so that the wiping surface can wipe the detection surface that is the target for detecting the presence or absence of allergens. The method for engaging the resin with the lid of the case body while inserting it through the lid and protruding outward is not particularly limited as long as it is a known engaging means, but a method in which the resin is engaged with a frame perforated in the lid is preferred. For example, engaging methods include using adhesive, hot plate welding, solvent bonding, ultrasonic welding, etc., a method in which the resin constituting the wiping portion is fitted into a frame that forms perforations for engaging the resin by utilizing the elasticity of the resin, or a method in which a groove is formed in the resin and the resin is fitted into at least a part of a frame that forms a perforation.

[0033] The shape of the wiping surface of the resin that constitutes the wiping portion is preferably flat, but it may also be uneven, etc., to accommodate cases where the detection surface is not uniformly flat and has an uneven shape.

[0034] The bottom of the resin constituting the wiping unit is located on one end of the developing support, so that when the detection surface is wiped with the wiping surface of the resin of the wiping unit and developing liquid is dripped from the top surface of the wiping unit, the developing liquid containing allergens permeates the resin of the wiping unit and flows down, and the allergen-containing developing liquid can reach a predetermined position on the developing support via the bottom of the wiping unit. The shape of the bottom of the resin is not particularly limited as long as it allows the allergen-containing developing liquid to reach a predetermined position on the developing support, but it may be a polygon such as a rectangle, or a flat surface such as a circle or ellipse, or it may be a cone or pyramid shape that causes the developing liquid to flow down in a tapered shape.

[0035] A sample carrier capable of supporting the developing solution is preferably placed at a predetermined position on the developing support. An example of the sample carrier is a glass wool sample pad. An immunochromatographic test strip (immunochromatographic strip) can be obtained by sequentially connecting this sample carrier, the gold colloid-labeled antibody carrier, and the developing support, preferably an absorbent such as an absorbent pad that absorbs the developing solution, to the other end of the developing support. The allergen in the developing solution migrates by capillary action or the like and binds to the gold colloid-labeled antibody. This antigen-antibody complex continues to migrate on the developing support by capillary action or the like until it is captured at a predetermined position where a monoclonal antibody against a denatured or undenatured allergen that recognizes a different epitope from the gold colloid-labeled antibody is immobilized. The allergen can be detected based on the presence or absence of a colored test line that appears at the predetermined position.

[0036] Methods for producing the above-mentioned colloidal gold-labeled antibody by conjugating a gold colloid to the monoclonal antibody include, but are not limited to, conventionally known methods. For example, a method in which a solution of a monoclonal antibody dissolved in 2 mM borate buffer (pH 9.0) is added to a colloidal gold solution adjusted to pH 9.0 with 0.2 M potassium carbonate solution, and the mixture is allowed to react at room temperature for 30 minutes. After that, a 10% BSA solution is added, and the mixture is allowed to react for an additional 15 minutes, followed by centrifugation. Alternatively, the above-prepared colloidal gold-labeled antibody can be produced by applying the colloidal gold-labeled antibody to, for example, a glass wool conjugate pad and drying it.

[0037] The developing support can be prepared, for example, by linearly applying a buffer solution containing monoclonal antibodies against denatured and native allergens that recognize different epitopes from those of the gold colloid-labeled antibodies to a nitrocellulose membrane, drying the membrane, and then performing a blocking treatment.

[0038] Suitable examples of monoclonal antibodies that recognize both the above-mentioned denatured and native allergens include two types of monoclonal antibodies that specifically recognize denatured and native allergens selected from αs1 casein, the main component of milk allergens; β-lactoglobulin, the main component of whey allergens; ovalbumin and ovomucoid, the main components of egg white allergens; gliadin, the main component of wheat allergens; proteins with molecular weights of 24 kDa and 76 kDa, the main proteins in buckwheat; AraH1, the main protein in peanuts; 7S globulin, the main allergen in soybeans; and 11S globulin, the main allergen in sesame.

[0039] More specifically, examples of anti-αs1 casein monoclonal antibodies produced by the inventors include the anti-αs1 casein monoclonal antibody Pas1CN1 produced by hybridoma (FERM-BP-10263) and the anti-αs1 casein monoclonal antibody Pas1CN2 produced by hybridoma (FERM-BP-10264), and examples of anti-β-lactoglobulin monoclonal antibodies include the anti-β-lactoglobulin monoclonal antibody PβLG3 produced by hybridoma (FERM-BP-11237) and the anti-β-lactoglobulin monoclonal antibody PβLG4 produced by hybridoma (FERM-BP-11238).

[0040] Examples of anti-ovalbumin monoclonal antibodies include the anti-ovalbumin monoclonal antibody PDOA3 produced by hybridoma (FERM-BP-11235) and the anti-ovalbumin monoclonal antibody PDOA4 produced by hybridoma (FERM-BP-11236). Examples of anti-ovomucoid monoclonal antibodies include the anti-ovomucoid monoclonal antibody PNOM1 produced by hybridoma (FERM-BP-10279), the anti-ovomucoid monoclonal antibody PNOM2 produced by hybridoma (FERM-BP-10280), the anti-ovomucoid monoclonal antibody PDOM1 produced by hybridoma (FERM-BP-10277), and the anti-ovomucoid monoclonal antibody PDOM2 produced by hybridoma (FERM-BP-10278).

[0041] Examples of the anti-wheat gliadin monoclonal antibody include the anti-wheat gliadin monoclonal antibody PGL1 produced by the hybridoma (FERM-BP-10267) and the anti-wheat gliadin monoclonal antibody PGL2 produced by the hybridoma (FERM-BP-10268).

[0042] Examples of the anti-buckwheat protein monoclonal antibody include the anti-24kDa protein monoclonal antibody PBW5 produced by the hybridoma (FERM-BP-11241), the anti-24kDa protein monoclonal antibody PBW1 produced by the hybridoma (FERM BP-10272), the anti-76kDa protein monoclonal antibody PBW2 produced by the hybridoma (FERM BP-10273), and the anti-76kDa protein monoclonal antibody PBW3 produced by the hybridoma (FERM BP-10274).

[0043] Examples of the anti-peanut Ara h1 protein monoclonal antibody include the anti-Ara h1 protein monoclonal antibody PAh1-5 produced by the hybridoma (FERM-BP-11240) and the anti-Ara h1 protein monoclonal antibody PAh1-4 produced by the hybridoma (FERM-BP-11239).

[0044] Examples of the anti-soybean 7S globulin monoclonal antibody include the anti-soybean 7S globulin monoclonal antibody PDSY1 produced by a hybridoma (NITE BP-02039) and the anti-soybean 7S globulin monoclonal antibody PDSY2 produced by a hybridoma (NITE BP-02040).

[0045] Examples of the anti-sesame 11S globulin monoclonal antibody include the anti-sesame 11S globulin monoclonal antibody PDSE1 produced by the hybridoma (NITE BP-02041) and the anti-sesame 11S globulin monoclonal antibody PDSE2 produced by the hybridoma (NITE BP-02042).

[0046] The amount of allergen that can be detected by the present invention is preferably 2 μg / mL or more as a concentration in the developing solution. Taking into consideration the shape and size of the wiping area and the effort required for wiping, 2 It is desirable to be able to detect 1 μg or more of allergen per sample.

[0047] The longitudinal case body is not particularly limited as long as the unfolding support is arranged inside the case body, but it is preferable that the case lid is engaged with the case at the end far from the wiping portion, and that the unfolding support can be removed after measurement by opening and closing the case body and lid at the end near the wiping portion.For example, the case body and lid can be opened and closed by making it possible to separate a lid opening piece protruding from a part of the peripheral wall of the lid and a main body case opening piece protruding from the main body peripheral wall of the main body case near the lid opening piece with fingers, thereby opening or separating the lid from the main body container.

[0048] The case body and its lid can be made of any material as long as they achieve the effects of the present invention, but transparent to translucent plastic is preferred so that the colored test line can be seen, and a see-through window may be provided in the lid of the case body so that the colored test line can be seen. Figure 10 shows an example of a detection case in which a wiping unit made of resin and a deployment support are disposed inside the case body, and the case lid is engaged with the end far from the wiping unit, and the case body and lid can be opened and closed at the end near the wiping unit.

[0049] The present invention will be explained in more detail below with reference to examples, but the technical scope of the present invention is not limited to these examples. [Example]

[0050] [Reference example] [PBS passing rate for each resin] In conventional swab tests, wiping is performed using a moist wipe, such as a cotton swab soaked in PBS. Therefore, we first checked whether food allergens could pass through the wipe using PBS alone when wiped with a dry polyethylene resin wipe. From this point on, we used Dulbecco's PBS(-) Powder "Nissui" (manufactured by Nissui Pharmaceutical Co., Ltd.), prepared as described in the package insert.

[0051] [Food protein preparation] (Egg protein preparation) Egg protein was prepared from powder produced according to the standard product specifications described in "Testing methods for foods containing allergens (reference) (March 26, 2014, Consumer Affairs Agency)."

[0052] (Preparation of milk proteins) Casein protein was used as the milk protein. Casein protein was prepared from powder prepared according to the standard product specifications described in "Testing Methods for Foods Containing Allergens (Reference) (March 26, 2014, Consumer Affairs Agency)."

[0053] (Wheat protein preparation) Wheat protein was prepared from powder produced according to the standard product specifications described in "Testing methods for foods containing allergens (reference) (March 26, 2014, Consumer Affairs Agency)."

[0054] [PBS passing rate for each resin] We used PBS, which has been used in conventional swab test kits, as the developer to confirm whether the test would be possible when the swab part was made into an integrated kit using polyethylene resin. The results are shown in Table 1 below.

[0055] [Table 1]

[0056] It was found that when PBS alone was used as a developer, allergens could not pass through the wiped area made of polyethylene resin.

[0057] [Example 1] [Step of preselecting a combination of allergen type, resin type, and developer type that results in an allergen penetration rate of 75% or more] (Resin that makes up the wiping part) The following four types of resin were used for the wiping part. (1) Polyester (PES) with a porosity of 79% (+4~-6%) (2) Polyethylene 1, average pore size 100 μm (PE1) (3) Polyethylene 2, average pore size 200 μm (PE2) (4) Polyvinyl alcohol (PVA) with an average pore size of 150 μm and a porosity of 89%

[0058] For the resins making up the wiping part, rectangular parallelepiped resins measuring 5 mm x 7 mm x 10 mm in height were prepared for each of the resins (1) to (4) above, i.e., PES, PE1, PE2, and PVA, and the study was carried out. 500 μL of a developing solution containing 5 ppm allergen was dropped onto the top surface of each resin body, and the percentage of allergen that had passed through to the bottom surface within 10 minutes was calculated to determine the allergen penetration rate.

[0059] The food allergens used were egg protein, milk (casein) protein, or wheat protein. The concentrations of each food allergen protein were calculated using the "Allergen Eye ELISA II Egg" (Primaham Meat Packers) for the egg protein solution, the "Morinaga FASPEK ELISA II Milk (Casein)" (Morinaga Institute of Biological Sciences) for the milk protein solution, and the "Allergen Eye ELISA II Wheat" (Primaham Meat Packers) for the wheat protein solution. Each food allergen protein was diluted to 5 ppm with a developer solution containing PBS and a surfactant to prepare an allergen-containing developer solution (test solution). The surfactants used in this study were Triton X-100 (Sigma) as polyoxyethylene (10) octylphenyl ether, SDS (Fujifilm Wako Pure Chemical Industries) as sodium dodecyl sulfate, and Tween 20 (MP Biomedical) as polyoxyethylene sorbitan monolaurate.

[0060] The allergen penetration rate of each resin was calculated by dropping 500 μL of a developing solution containing 5 ppm of each food allergen protein onto the top surface of each resin body, collecting the solution that passed through the bottom surface of the resin body within 10 minutes, and measuring the concentration of each food allergen protein in the collected sample using the ELISA kit. The percentage of the concentration of each food allergen protein that passed through the bottom surface of the resin body (penetration rate) was calculated using the following formula.

[0061]

number

[0062] (Egg-TritonX-100) Egg protein was diluted to 5 ppm in a developing solution containing PBS and Triton X-100 to prepare an allergen-containing developing solution, and the allergen penetration rates are shown in Figures 1(a) to (d).

[0063] (result) As is clear from Figure 1(a), for polyester resin (PES), the passage rate was 100% when PBS alone was used. The passage rate was 99.7% at a Triton X-100 concentration of 0.01%, 96.8% at 0.05%, 99.9% at 0.1%, 99.3% at 0.5%, and 100% at 1.0%. The passage rate was 95% or higher at Triton X-100 concentrations of 0.01%, 0.05%, 0.1%, 0.5%, and 1.0%. Hereinafter, "%" means % by mass.

[0064] As is clear from Figure 1(b), for polyethylene resin (PE1), the passage rate was 0% when PBS alone or when the Triton X-100 concentration was 0.01%, but was 96.1% at 0.05%, and 100% at 0.1%, 0.5%, and 1.0%, and the passage rate was 95% or more at Triton X-100 concentrations of 0.05%, 0.1%, 0.5%, and 1.0%.

[0065] As is clear from Figure 1(c), for polyethylene resin (PE2), the passage rate was 0% when PBS alone or when the Triton X-100 concentration was 0.01%, but was 98.5% at 0.05%, 100% at 0.1%, 98.9% at 0.5%, and 100% at 1.0%, resulting in a passage rate of 95% or more at Triton X-100 concentrations of 0.05%, 0.1%, 0.5%, and 1.0%.

[0066] As shown in Figure 1(d), for polyvinyl alcohol resin (PVA), the passage rate was 88.7% when PBS was used alone. The passage rates were 100% at Triton X-100 concentrations of 0.01%, 98.5% at 0.05%, 100% at 0.1%, 99.2% at 0.5%, and 100% at 1.0%. The passage rates were 95% or higher at Triton X-100 concentrations of 0.01%, 0.05%, 0.1%, 0.5%, and 1.0%.

[0067] (Egg-TritonX-100 Summary) As shown above, Triton X-100 added to PBS improved the passage rate of egg allergens. When egg allergens were selected as the detection target and PES or PVA was used as the resin, the passage rate was 95% or higher at Triton X-100 concentrations of 0.01%, 0.05%, 0.1%, 0.5%, and 1.0%. When PE was used as the resin, the passage rate was 95% or higher at Triton X-100 concentrations of 0.05%, 0.1%, 0.5%, and 1.0%.

[0068] (Egg-SDS) The allergen penetration rates when egg protein was diluted to 5 ppm in a developing solution containing PBS and SDS to prepare an allergen-containing developing solution are shown in Figures 2(a) to (d).

[0069] As is clear from Figure 2(a), for polyester resin (PES), PBS alone and SDS concentrations of 0.01%, 0.05%, and 0.1% showed a 100% passage rate, 98.0% at 0.5%, and 99.2% at 1.0%, and the passage rate was 95% or higher at SDS concentrations of 0.01%, 0.05%, 0.1%, 0.5%, and 1.0%.

[0070] As is clear from Figure 2(b), for polyethylene resin (PE1), the passage rate was 0% when PBS alone or when the SDS concentration was 0.01%, but 99.3% was observed at 0.05%, and 100% was observed at 0.1%, 0.5%, and 1.0% SDS concentrations, with the passage rate being 95% or higher.

[0071] As is clear from Figure 2(c), for polyethylene resin (PE2), the passage rate was 0% when PBS alone or when the SDS concentration was 0.01%, but 100% was observed when PBS alone, 0.1%, 0.5%, and 1.0%, and the passage rate was 95% or more when SDS concentrations were 0.05%, 0.1%, 0.5%, and 1.0%.

[0072] As shown in Figure 2(d), for polyvinyl alcohol resin (PVA), the passage rate was 88.7% when PBS was used alone. The passage rate was 97.7% when SDS was added at 0.01%, 94.5% at 0.05%, 100% at 0.1%, 97.8% at 0.5%, and 100% at 1.0% SDS. The passage rate was over 90% at SDS concentrations of 0.01%, 0.05%, 0.1%, 0.5%, and 1.0%.

[0073] (Eggs - Summary of SDS) As shown above, adding SDS to PBS improved the passage rate of egg allergens. When egg allergens were selected as the detection target and PES was used as the resin, the passage rate was 95% or higher at SDS concentrations of 0.01%, 0.05%, 0.1%, 0.5%, and 1.0%. When PE was used as the resin, the passage rate was 95% or higher at SDS concentrations of 0.05%, 0.1%, 0.5%, and 1.0%. When PVA was used as the resin, the passage rate was 90% or higher at SDS concentrations of 0.01%, 0.05%, 0.1%, 0.5%, and 1.0%.

[0074] (Egg-Tween 20) The allergen penetration rates when egg protein was diluted to 5 ppm in a developing solution containing PBS and Tween 20 to prepare an allergen-containing developing solution are shown in Figures 3(a) to (d).

[0075] As is clear from Figure 3(a), for polyester resin (PES), the passage rate was 100% when PBS alone was used. The passage rate was 95.3% when Tween 20 was added at a concentration of 0.01%, 100% at 0.05%, 0.1%, and 0.5%, and 98.6% at 1.0%. The passage rate was 95% or higher at Tween 20 concentrations of 0.01%, 0.05%, 0.1%, 0.5%, and 1.0%.

[0076] As is clear from Figures 3(b) and 3(c), the penetration rate was 0% for the polyethylene resin (PE1) and the polyethylene resin (PE2), regardless of the concentration of Tween 20 added to PBS.

[0077] As is clear from Figure 3(d), for polyvinyl alcohol resin (PVA), the passage rate was 88.7% when PBS alone was used, 94.5% when the Tween 20 concentration was 0.01%, 93.9% when it was 0.05%, 100% when it was 0.1% and 0.5%, and 97.3% when it was 1.0%. The passage rate was 90% or more at Tween 20 concentrations of 0.01%, 0.05%, 0.1%, 0.5%, and 1.0%.

[0078] (Egg-Tween 20 Summary) When egg allergens were selected as the target and PES was used as the resin, the penetration rate was 95% or higher at Tween 20 concentrations of 0.01%, 0.05%, 0.1%, 0.5%, and 1.0%. When PE was used as the resin, the penetration rate was 0% at any Tween 20 concentration. When PVA was used as the resin, the penetration rate was 90% or higher at Tween 20 concentrations of 0.01%, 0.05%, 0.1%, 0.5%, and 1.0%.

[0079] (Milk-TritonX-100) The allergen penetration rates when milk protein was diluted to 5 ppm in a developing solution containing PBS and Triton X-100 to prepare an allergen-containing developing solution are shown in Figures 4(a) to (d).

[0080] As is clear from Figure 4(a), for polyester resin (PES), the passage rate was 60.7% when PBS alone was used, 89.8% when the Triton X-100 concentration was 0.01%, 95.3% when it was 0.05%, 95.5% when it was 0.1%, 100% when it was 0.5%, and 96.5% when it was 1.0%.The passage rate was 85% or more at Triton X-100 concentrations of 0.01%, 0.05%, 0.1%, 0.5%, and 1.0%.

[0081] As is clear from Figure 4(b), for polyethylene resin (PE1), the passage rate was 0% when PBS alone or when the Triton X-100 concentration was 0.01%, but was 99.4% at 0.05%, 98.2% at 0.1%, 100% at 0.5%, and 95.7% at 1.0%, meaning that the passage rate was 95% or higher at Triton X-100 concentrations of 0.05%, 0.1%, 0.5%, and 1.0%.

[0082] As is clear from Figure 4(c), for polyethylene resin (PE2), the passage rate was 0% when PBS alone or when the Triton X-100 concentration was 0.01%, but was 99.0% at 0.05%, 100% at 0.1% and 0.5%, and 94.9% at 1.0%, meaning that the passage rate was 90% or more at Triton X-100 concentrations of 0.05%, 0.1%, 0.5%, and 1.0%.

[0083] As is clear from Figure 4(d), for polyvinyl alcohol resin (PVA), the passage rate was 15.3% when PBS alone was used and 46.6% when the Triton X-100 concentration was 0.01%, but the passage rates were 87.7% at 0.05%, 86.4% at 0.1%, 93.2% at 0.5%, and 92.8% at 1.0%, meaning that the passage rates were 85% or higher at Triton X-100 concentrations of 0.05%, 0.1%, 0.5%, and 1.0%.

[0084] (Milk-TritonX-100 Summary) When milk allergens were selected as the target and PES resin was used, the penetration rate was 85% or higher at Triton X-100 concentrations of 0.01%, 0.05%, 0.1%, 0.5%, and 1.0%. When PE resin was used, the penetration rate was 90% or higher at Triton X-100 concentrations of 0.05%, 0.1%, 0.5%, and 1.0%. When PVA resin was used, the penetration rate was 85% or higher at Triton X-100 concentrations of 0.05%, 0.1%, 0.5%, and 1.0%.

[0085] (Milk-SDS) The allergen penetration rates when milk protein was diluted to 5 ppm in a developing solution containing PBS and SDS to prepare an allergen-containing developing solution are shown in Figures 5(a) to (d).

[0086] As is clear from Figure 5(a), for polyester resin (PES), the passage rate was 60.7% when PBS alone was used. The passage rates were 95.2% at SDS concentrations of 0.01%, 100% at 0.05% and 0.1%, 99.6% at 0.5%, and 99.0% at 1.0%. The passage rates were over 95% at SDS concentrations of 0.01%, 0.05%, 0.1%, 0.5%, and 1.0%.

[0087] As is clear from Figure 5(b), for polyethylene resin (PE1), the passage rate was 0% when PBS alone or when the SDS concentration was 0.01%, but was 100% at 0.05%, 98.1% at 0.1%, 97.6% at 0.5%, and 98.3% at 1.0%, meaning that the passage rate was 95% or higher at SDS concentrations of 0.05%, 0.1%, 0.5%, and 1.0%.

[0088] As is clear from Figure 5(c), for polyethylene resin (PE2), the passage rate was 0% when PBS alone or when the SDS concentration was 0.01%, but was 100% at 0.05%, 97.7% at 0.1%, and 100% at 0.5% and 1.0%, and the passage rate was 95% or more at SDS concentrations of 0.05%, 0.1%, 0.5%, and 1.0%.

[0089] As is clear from Figure 5(d), for polyvinyl alcohol resin (PVA), PBS alone showed a 15.3% passage rate. At an SDS concentration of 0.01%, the passage rate was 87.1%. At 0.05%, 0.1%, and 0.5%, the passage rate was 100%, and at 1.0%, the passage rate was 99.5%. At SDS concentrations of 0.01%, 0.05%, 0.1%, 0.5%, and 1.0%, the passage rate was 85% or higher.

[0090] (Milk - Summary of SDS) When milk allergens were selected as the target and PES resin was used, the pass rate was 95% or higher at SDS concentrations of 0.01%, 0.05%, 0.1%, 0.5%, and 1.0%. When PE resin was used, the pass rate was 95% or higher at SDS concentrations of 0.05%, 0.1%, 0.5%, and 1.0%. When PVA resin was used, the pass rate was 85% or higher at SDS concentrations of 0.01%, 0.05%, 0.1%, 0.5%, and 1.0%.

[0091] (Milk-Tween 20) The allergen penetration rates when milk protein was diluted to 5 ppm in a developing solution containing PBS and Tween 20 to prepare an allergen-containing developing solution are shown in Figures 6(a) to (d).

[0092] As is clear from Figure 6(a), for polyester resin (PES), the passage rate was 60.7% with PBS alone, 75.6% with a Tween 20 concentration of 0.01%, 76.1% with a Tween 20 concentration of 0.05%, 78.5% with a Tween 20 concentration of 0.1%, 92.4% with a Tween 20 concentration of 0.5%, and 93.0% with a Tween 20 concentration of 1.0%, and the passage rate was 75% or higher at Tween 20 concentrations of 0.01%, 0.05%, 0.1%, 0.5%, and 1.0%.

[0093] As is clear from Figures 6(b) and 6(c), the penetration rate was 0% for the polyethylene resin (PE1) and the polyethylene resin (PE2), regardless of the concentration of Tween 20 added to PBS.

[0094] As is clear from Figure 6(d), for polyvinyl alcohol resin (PVA), the passage rate was 15.3% when PBS alone was used, 42.8% when the Tween 20 concentration was 0.01%, 55.0% when it was 0.05%, 55.4% when it was 0.1%, 83.0% when it was 0.5%, and 78.3% when it was 1.0%. The passage rate was 75% or more at Tween 20 concentrations of 0.5% and 1.0%.

[0095] (Milk-Tween 20 Summary) When milk allergens were selected as the target and PES was used as the resin, the penetration rate was 75% or higher at Tween 20 concentrations of 0.01%, 0.05%, 0.1%, 0.5%, and 1.0%. When PE was used as the resin, the penetration rate was 0% at any Tween 20 concentration. When PVA was used as the resin, the penetration rate was 75% or higher at Tween 20 concentrations of 0.5% and 1.0%.

[0096] (Wheat-TritonX-100) The allergen penetration rates when wheat protein was diluted to 5 ppm in a developing solution containing PBS and Triton X-100 to prepare an allergen-containing developing solution are shown in Figures 7(a) to (d).

[0097] As is clear from Figure 7(a), for polyester resin (PES), the passage rate was 85.1% when PBS alone was used, 100% when TritonX-100 concentrations were 0.01%, 0.05%, and 0.1%, 96.0% at 0.5%, and 100% at 1.0% TritonX-100 concentrations, and the passage rate was 95% or more at TritonX-100 concentrations of 0.01%, 0.05%, 0.1%, 0.5%, and 1.0%.

[0098] As is clear from Figure 7(b), for polyethylene resin (PE1), the passage rate was 0% when PBS alone or when the Triton X-100 concentration was 0.01%, but was 92.2% at 0.05%, 94.1% at 0.1%, and 100% at 0.5% and 1.0%, resulting in a passage rate of 90% or more at Triton X-100 concentrations of 0.05%, 0.1%, 0.5%, and 1.0%.

[0099] As is clear from Figure 7(c), for polyethylene resin (PE2), the passage rate was 0% when PBS alone or when the Triton X-100 concentration was 0.01%, but was 87.1% at 0.05%, 98.9% at 0.1%, and 100% at 0.5% and 1.0%, resulting in a passage rate of 85% or more at Triton X-100 concentrations of 0.05%, 0.1%, 0.5%, and 1.0%.

[0100] As is clear from Figure 7(d), for polyvinyl alcohol resin (PVA), the passage rate was 59.0% when PBS alone was used, 85.7% when the Triton X-100 concentration was 0.01%, 88.2% when it was 0.05%, and 100% when it was 0.1%, 0.5%, and 1.0%.The passage rate was 85% or more at Triton X-100 concentrations of 0.01%, 0.05%, 0.1%, 0.5%, and 1.0%.

[0101] (Wheat - Triton X-100 Summary) When wheat allergens were selected as the target and PES resin was used, the penetration rate was 95% or higher at Triton X-100 concentrations of 0.01%, 0.05%, 0.1%, 0.5%, and 1.0%. When PE resin was used, the penetration rate was 85% or higher at Triton X-100 concentrations of 0.05%, 0.1%, 0.5%, and 1.0%. When PVA resin was used, the penetration rate was 85% or higher at Triton X-100 concentrations of 0.01%, 0.05%, 0.1%, 0.5%, and 1.0%.

[0102] (Wheat-SDS) The allergen penetration rates when wheat protein was diluted to 5 ppm in a developing solution containing PBS and SDS to prepare an allergen-containing developing solution are shown in Figures 8(a) to (d).

[0103] As is clear from Figure 8(a), for polyester resin (PES), the passage rate was 85.1% when PBS alone was used. The passage rates were 86.0% when the SDS concentration was 0.01%, 95.0% when it was 0.05%, 99.4% when it was 0.1%, 92.6% when it was 0.5%, and 98.8% when it was 1.0%. The passage rates were 85% or higher at SDS concentrations of 0.01%, 0.05%, 0.1%, 0.5%, and 1.0%.

[0104] As is clear from Figure 8(b), for polyethylene resin (PE1), the passage rate was 0% when PBS alone or when the SDS concentration was 0.01%, but was 100% at 0.05%, 99.4% at 0.1%, 96.8% at 0.5%, and 100% at 1.0%, and the passage rate was 95% or more at SDS concentrations of 0.05%, 0.1%, 0.5%, and 1.0%.

[0105] As is clear from Figure 8(c), for polyethylene resin (PE2), the passage rate was 0% when PBS alone or when the SDS concentration was 0.01%, but was 100% at 0.05% and 0.1%, 98.3% at 0.5%, and 100% at 1.0%, and the passage rate was 95% or more at SDS concentrations of 0.05%, 0.1%, 0.5%, and 1.0%.

[0106] As is clear from Figure 8(d), for polyvinyl alcohol resin (PVA), PBS alone showed a 59.0% passage rate. At an SDS concentration of 0.01%, the passage rate was 59.5%. At 0.05% and 0.1%, the passage rate was 100%, at 0.5%, the passage rate was 94.5%, and at 1.0%, the passage rate was 97.2%. At SDS concentrations of 0.05%, 0.1%, 0.5%, and 1.0%, the passage rate was over 90%.

[0107] (Wheat - Summary of SDS) When wheat allergens were selected as the target and PES resin was used, the pass rate was 85% or higher at SDS concentrations of 0.01%, 0.05%, 0.1%, 0.5%, and 1.0%. When PE resin was used, the pass rate was 95% or higher at SDS concentrations of 0.05%, 0.1%, 0.5%, and 1.0%. When PVA resin was used, the pass rate was 90% or higher at SDS concentrations of 0.05%, 0.1%, 0.5%, and 1.0%.

[0108] (Wheat-Tween 20) The allergen penetration rates when wheat protein was diluted to 5 ppm in a developing solution containing PBS and Tween 20 to prepare an allergen-containing developing solution are shown in Figures 9(a) to (d).

[0109] As is clear from Figure 9(a), for polyester resin (PES), the passage rate was 85.1% with PBS alone, 92.8% with a Tween 20 concentration of 0.01%, 89.8% with a Tween 20 concentration of 0.05%, 95.1% with a Tween 20 concentration of 0.1%, 98.1% with a Tween 20 concentration of 0.5%, and 99.1% with a Tween 20 concentration of 1.0%, and the passage rate was 85% or higher at Tween 20 concentrations of 0.01%, 0.05%, 0.1%, 0.5%, and 1.0%.

[0110] As is clear from Figures 9(b) and 9(c), the penetration rate was 0% for the polyethylene resin (PE1) and the polyethylene resin (PE2), regardless of the concentration of Tween 20 added to PBS.

[0111] As is clear from Figure 9(d), for polyvinyl alcohol resin (PVA), the passage rate was 59.0% when PBS alone was used, 83.4% when the Tween 20 concentration was 0.01%, 81.4% when it was 0.05%, 88.6% when it was 0.1%, 93.6% when it was 0.5%, and 99.9% when it was 1.0%.The passage rate was 80% or more at Tween 20 concentrations of 0.01%, 0.05%, 0.1%, 0.5%, and 1.0%.

[0112] (Wheat - Tween 20 Summary) When wheat allergens were selected as the target and PES resin was used, the penetration rate was 85% or higher at Tween 20 concentrations of 0.01%, 0.05%, 0.1%, 0.5%, and 1.0%. When PE resin was used, the penetration rate was 0% at any Tween 20 concentration. When PVA resin was used, the penetration rate was 80% or higher at Tween 20 concentrations of 0.01%, 0.05%, 0.1%, 0.5%, and 1.0%.

[0113] [Example 2] [Step of selecting a combination of allergen type, resin type, and developing liquid type] Next, the main selection was carried out using the wiper-integrated kit.

[0114] [Production of a kit with an integrated wiping unit] (Preparation of immunochromatographic strips for detecting ovalbumin) 1) Preparation of colloidal gold-labeled antibodies A PDOA3 monoclonal antibody solution was prepared at 1 mg / mL in 2 mM borate buffer (pH 9.0). 500 μL of the PDOA3 monoclonal antibody solution was added to 5 mL of gold colloid solution (Sigma) previously adjusted to pH 9.0 with 0.2 M potassium carbonate solution, and the mixture was incubated at room temperature for 30 minutes. 635 μL of a 10% BSA solution was then added, and the mixture was incubated for an additional 15 minutes. After centrifugation, the mixture was adjusted to OD525 = 1.0 with a 1% BSA solution. 68 μL / cm was applied to a glass wool conjugate pad. 2 The coating was applied so as to become a uniform layer, and then allowed to dry.

[0115] 2) Preparation of antibody-immobilized membrane A 4 mg / mL solution of PDOA4 monoclonal antibody was prepared in phosphate buffered saline (PBS), applied linearly to a nitrocellulose membrane, and allowed to dry. The membrane was then blocked with TBS containing 0.1% bovine gelatin at 37°C for 1 hour, washed with TBS, and then dried.

[0116] 3) Assembly of immunochromatographic strips In addition to the antibody-immobilized membrane, a glass wool sample pad as a sample carrier and a glass wool absorbent pad for absorbing liquid samples were separately prepared, and the sample pad, antibody-immobilized membrane, and absorbent pad were attached in that order to create an immunochromatographic strip.

[0117] (Preparation of immunochromatographic strips for casein detection) An immunochromatographic strip for detecting casein was prepared using the same procedure as for preparing the immunochromatographic strip for detecting ovalbumin described above, except that a Pas1CN1 monoclonal antibody solution was prepared in the preparation of the gold colloid-labeled antibody described above in 1) above, and a Pas1CN2 monoclonal antibody solution was prepared in the preparation of the antibody-immobilized membrane described above in 2).

[0118] (Preparation of immunochromatographic strips for detecting wheat gliadin) An immunochromatographic strip for detecting wheat gliadin was prepared using the same procedure as for preparing the immunochromatographic strip for detecting ovalbumin described above, except that a PGL1 monoclonal antibody solution was prepared in the preparation of the gold colloid-labeled antibody described above in 1) above, and a PGL2 monoclonal antibody solution was prepared in the preparation of the antibody-immobilized membrane described above in 2).

[0119] [Production of a kit with an integrated wiping unit] A main case was prepared in which the lid was engaged at the end distal from the wiping portion, and the lid and the case body were opened and closed by a lid release piece and a main case release piece at the end proximal to the wiping portion. The immunochromatographic strips for detecting food allergens, for ovalbumin, casein, and wheat gliadin, were placed in the longitudinal main case with the sample pad facing the wiping portion. Any of the materials (1) to (4) above was used as the resin for the wiping portion, and its bottom was positioned on one end of the developing support and in contact with the sample carrier portion of each immunochromatographic strip. The wiping surface was inserted through the lid of the main case and protruded outward, and the resin used for the wiping portion was a rectangular parallelepiped 10 mm high with a 5 mm x 7 mm flat surface (top surface) serving as the wiping surface.

[0120] [Swab test using the kit of the present invention] Using the above-mentioned wipe-integrated kit, the above-mentioned three types of surfactants, Triton X-100, SDS, and Tween 20, were added to the developing solution to confirm whether allergens could be detected by combining the above-mentioned types of allergens, resins, and developing solutions.

[0121] (Creating the detection surface) Aluminum foil was cut into 10cm squares, and 200μL of a solution containing 20ppm, 10ppm, or 5ppm of various allergen proteins was dropped onto the foil to determine the concentration of 4μg, 2μg, or 1μg / 100cm. 2 The food allergen protein was applied to the detection surface, which was then left to dry completely at room temperature, to prepare a detection surface coated with egg protein, casein protein, or wheat protein.

[0122] (Test using a kit) The kit was held with the index finger from the back with the wiping section facing downward, and the wiping section was pressed horizontally against the detection surface without tilting the kit. While pressing the wiping section against the detection surface, the entire allergen applied to the detection surface was wiped off by moving the wiping section back and forth 10 times in both the vertical and horizontal directions. PBS containing Triton X-100, SDS, or Tween 20 (as shown below) was dropped dropwise onto the wiping surface of the wiping section, totaling 500 μL, using a micropipette. The allergen-containing developer solution was allowed to flow down to the bottom of the wiping surface, and the allergen-containing developer solution that reached the sample carrier was developed on the immunochromatographic strip. After 10 minutes, the presence or absence of a test line was visually determined. The allergen concentrations in 500 μL of developing solution containing 4 μg, 2 μg, or 1 μg of allergen are 8 μg / mL, 4 μg / mL, and 2 μg / mL (ppm), respectively.

[0123] [Book Selection] (PBS+Triton X-100) Four micrograms of each of the food allergen proteins listed below was applied to the detection surface, and the developer solution was prepared by adding 0.01%, 0.05%, 0.1%, 0.5%, or 1.0% Triton X-100 to PBS. The results are shown in Tables 2-1 to 2-3. Tests in which the developer solution did not penetrate and testing was not possible were marked "untestable." Tests in which testing was possible were marked with +, +w, and +- in descending order of strength of the test line, and negative results were marked with -.

[0124] [Table 2-1]

[0125] (result) As is clear from Table 2-1, when 4 μg of eggs were applied, if the resin used was PES, the results were positive at concentrations of 0.01%(+), 0.05%(+), 0.1%(+), 0.5%(+), and 1.0%(+). In the case of PE1, the positive results were 0.05% (+), 0.1% (+), and 0.5% (+). In the case of PE2, the positive results were 0.05% (+), 0.1% (+), 0.5% (+), and 1.0% (+-). In the case of PVA, the results were positive at concentrations of 0.01% (+), 0.05% (+), 0.1% (+), 0.5% (+), and 1.0% (+-).

[0126] Therefore, when detecting 4 μg of eggs, PES and PVA were used as developing solutions and Triton X-100 concentrations in PBS were 0.01%, 0.05%, 0.1%, 0.5%, and 1.0%. PE1 was used as developing solutions and Triton X-100 concentrations in PBS were 0.05%, 0.1%, and 0.5%. PE2 was used as developing solutions and Triton X-100 concentrations in PBS were 0.05%, 0.1%, 0.5%, and 1.0%.

[0127] [Table 2-2]

[0128] (result) As is clear from Table 2-2, when 4 μg of milk was applied, if the resin used was PES, the results were positive at concentrations of 0.01% (+), 0.05% (+), 0.1% (+), and 0.5% (+-). In the case of PE1, the results were positive at concentrations of 0.05% (+) and 0.1% (+). In the case of PE2, the results were positive at concentrations of 0.05% (+), 0.1% (+w), and 0.5% (+w). In the case of PVA, the results were positive at concentrations of 0.01% (+), 0.05% (+), 0.1% (+), and 0.5% (+-).

[0129] Therefore, when detecting 4 μg of milk, it was confirmed that PES and PVA could detect it when Triton X-100 was added to PBS at concentrations of 0.01%, 0.05%, 0.1%, and 0.5%, PE1 could detect it when Triton X-100 was added to PBS at concentrations of 0.05% and 0.1%, and PE2 could detect it when Triton X-100 was added to PBS at concentrations of 0.05%, 0.1%, and 0.5%.

[0130] [Table 2-3]

[0131] As is clear from Table 2-3, when 4 μg of wheat was applied, if the resin used was PES, the results were positive at concentrations of 0.01%(+), 0.05%(+), 0.1%(+), 0.5%(+), and 1.0%(+). In the case of PE1, the results were positive at concentrations of 0.05% (+), 0.1% (+), 0.5% (+), and 1.0% (+). In the case of PE2, the results were positive at concentrations of 0.05% (+), 0.1% (+), 0.5% (+), and 1.0% (+). In the case of PVA, the results were positive at concentrations of 0.01%(+), 0.05%(+), 0.1%(+), 0.5%(+), and 1.0%(+).

[0132] Therefore, when detecting 4 μg of wheat, it was confirmed that PES and PVA could detect it when Triton X-100 was added to PBS at concentrations of 0.01%, 0.05%, 0.1%, 0.5%, and 1.0%, while PE1 and PE2 could detect it when Triton X-100 was added to PBS at concentrations of 0.05%, 0.1%, 0.5%, and 1.0%.

[0133] (TritonX-100-Summary) To summarize these results, swab tests were possible on all materials when Triton X-100 was used as the developer. Furthermore, negative results were sometimes observed when the Triton X-100 concentration was high. This was thought to be due to Triton X-100 adversely affecting the antigen-antibody reaction of the kit, so it was necessary to select an appropriate concentration for each allergen.

[0134] (PBS+SDS) 4 μg of each of the food allergen proteins listed below was applied to the detection surface, and the developing solution was prepared by adding 0.01%, 0.05%, 0.1%, 0.5%, or 1.0% SDS to PBS. The results are shown in Tables 3-1 to 3.

[0135] [Table 3-1]

[0136] (result) As is clear from Table 3-1, when 4 μg of eggs were applied, if the resin used was PES, the results were positive at concentrations of 0.01%(+), 0.05%(+), 0.1%(+), 0.5%(+), and 1.0%(+). In the case of PE1, the results were positive at concentrations of 0.05% (+), 0.1% (+), 0.5% (+), and 1.0% (+). In the case of PE2, the results were positive at concentrations of 0.05% (+), 0.1% (+), 0.5% (+), and 1.0% (+). In the case of PVA, the results were positive at concentrations of 0.01%(+), 0.05%(+), 0.1%(+), 0.5%(+), and 1.0%(+).

[0137] Therefore, when detecting 4 μg of eggs, it was confirmed that PES and PVA could detect eggs when the concentrations of SDS added to PBS were 0.01%, 0.05%, 0.1%, 0.5%, and 1.0%, and that PE1 and PE2 could detect eggs when the concentrations of SDS added to PBS were 0.05%, 0.1%, 0.5%, and 1.0%.

[0138] [Table 3-2]

[0139] (result) As is clear from Table 3-2, when 4 μg of milk was applied, if the resin used was PES, the results were positive at concentrations of 0.01% (+) and 0.05% (+). In the case of PE1, the results were positive at concentrations of 0.05% (+w), 0.1% (+w), and 0.5% (+-). In the case of PE2, the results were positive at concentrations of 0.05% (+) and 0.1% (+). In the case of PVA, the results were positive at concentrations of 0.01% (+) and 0.05% (+).

[0140] Therefore, when detecting 4 μg of milk, it was confirmed that PES and PVA could detect it when the SDS concentration added to PBS was 0.01% and 0.05%, PE1 could detect it when the SDS concentration added to PBS was 0.05%, 0.1%, and 0.5%, and PE2 could detect it when the SDS concentration added to PBS was 0.05% and 0.1%.

[0141] [Table 3-3]

[0142] As is clear from Table 3-3, when 4 μg of wheat was applied, if the resin used was PES, the results were positive at concentrations of 0.01%(+), 0.05%(+), 0.1%(+), 0.5%(+), and 1.0%(+). In the case of PE1, the results were positive at concentrations of 0.05% (+), 0.1% (+), 0.5% (+), and 1.0% (+). In the case of PE2, the results were positive at concentrations of 0.05% (+), 0.1% (+), 0.5% (+), and 1.0% (+). In the case of PVA, the results were positive at concentrations of 0.01%(+), 0.05%(+), 0.1%(+), 0.5%(+), and 1.0%(+).

[0143] Therefore, when detecting 4 μg of wheat, it was confirmed that PES and PVA could detect it when the concentrations of SDS added to PBS were 0.01%, 0.05%, 0.1%, 0.5%, and 1.0%, and that PE1 and PE2 could detect it when the concentrations of SDS added to PBS were 0.05%, 0.1%, 0.5%, and 1.0%.

[0144] (SDS-Summary) To summarize these results, swab tests were possible for all materials when SDS was used as a developer. Furthermore, negative results were sometimes observed when the SDS concentration was high. This was thought to be due to the adverse effect of SDS on the antigen-antibody reaction of the kit, so it was necessary to select an appropriate concentration for each allergen.

[0145] (PBS+Tween 20) 4 μg of each of the food allergen proteins listed below was applied to the detection surface, and the developing solution was prepared by adding 0.01%, 0.05%, 0.1%, 0.5%, or 1.0% Tween 20 to PBS. The results are shown in Tables 4-1 to 4-3.

[0146] [Table 4-1]

[0147] (result) As is clear from Table 4-1, when 4 μg of eggs were applied, if the resin used was PES, the results were positive at concentrations of 0.01%(+), 0.05%(+), 0.1%(+), 0.5%(+), and 1.0%(+). In the case of PE1 and PE2, it was impossible to test at any of the concentrations of 0.01%, 0.05%, 0.1%, 0.5%, and 1.0%. In the case of PVA, the results were positive at concentrations of 0.01%(+), 0.05%(+), 0.1%(+), and 0.5%(+).

[0148] For PE1 and PE2, detection was impossible at any concentration, and it was confirmed that the developing solution containing Tween 20 is not suitable for use in detecting 4 μg of eggs.

[0149] [Table 4-2]

[0150] (result) As is clear from Table 4-2, when 4 μg of milk was applied, if the resin used was PES, the results were positive at concentrations of 0.01% (+), 0.05% (+w), 0.1% (+w), 0.5% (+), and 1.0% (+). In the case of PE1 and PE2, it was impossible to test at any of the concentrations of 0.01%, 0.05%, 0.1%, 0.5%, and 1.0%. In the case of PVA, the results were positive at concentrations of 0.01%(+), 0.05%(+), 0.1%(+), 0.5%(+), and 1.0%(+).

[0151] PE1 and PE2 were not detectable at any concentration, confirming that the developer containing Tween 20 is not suitable for use in detecting 4 μg of milk.

[0152] [Table 4-3]

[0153] (result) As is clear from Table 4-3, when 4 μg of wheat was applied, if the resin used was PES, the results were positive at concentrations of 0.01%(+), 0.05%(+), 0.1%(+), 0.5%(+), and 1.0%(+). In the case of PE1 and PE2, it was impossible to test at any of the concentrations of 0.01%, 0.05%, 0.1%, 0.5%, and 1.0%. In the case of PVA, the results were positive at concentrations of 0.01%(+), 0.05%(+), 0.1%(+), 0.5%(+), and 1.0%(+).

[0154] For PE1 and PE2, detection was impossible at any concentration, and it was confirmed that the developer containing Tween 20 is not suitable for use in detecting 4 μg of wheat.

[0155] (PBS+Triton X-100) Tables 5-1 to 5-3 show the results when 2 μg of each of the food allergen proteins listed below was applied to the detection surface and the developing solution was prepared by adding 0.01%, 0.05%, 0.1%, 0.5%, or 1.0% Triton X-100 to PBS.

[0156] [Table 5-1]

[0157] (result) As is clear from Table 5-1, when 2 μg of eggs were applied, if the resin used was PES, the results were positive at concentrations of 0.01%(+), 0.05%(+), 0.1%(+), 0.5%(+), and 1.0%(+). In the case of PE1, the results were positive at concentrations of 0.05% (+), 0.1% (+), 0.5% (+), and 1.0% (+). In the case of PE2, the results were positive at concentrations of 0.05% (+), 0.1% (+), 0.5% (+w), and 1.0% (+). In the case of PVA, the results were positive at concentrations of 0.01%(+), 0.05%(+), 0.1%(+), 0.5%(+), and 1.0%(+).

[0158] Therefore, when detecting 2 μg of eggs, it was confirmed that PES and PVA could detect them when Triton X-100 was added to PBS at concentrations of 0.01%, 0.05%, 0.1%, 0.5%, and 1.0%, while PE1 and PE2 could detect them when Triton X-100 was added to PBS at concentrations of 0.05%, 0.1%, 0.5%, and 1.0%.

[0159] [Table 5-2]

[0160] (result) As is clear from Table 5-2, when 2 μg of milk was applied, if the resin used was PES, the results were positive at concentrations of 0.01% (+), 0.05% (+-), 0.1% (+-), and 0.5% (+-). In the case of PE1, the results were positive at concentrations of 0.05% (+-) and 0.1% (+-). In the case of PE2, the results were positive at concentrations of 0.05% (+-) and 0.1% (+w). In the case of PVA, the results were positive at concentrations of 0.01% (±), 0.05% (±), 0.1% (±), and 0.5% (±).

[0161] Therefore, when detecting 2 μg of milk, it was confirmed that PES and PVA could detect it when Triton X-100 was added to PBS at concentrations of 0.01%, 0.05%, 0.1%, and 0.5%, and that PE1 and PE2 could detect it when Triton X-100 was added to PBS at concentrations of 0.05% and 0.1%.

[0162] [Table 5-3]

[0163] As is clear from Table 5-3, when 2 μg of wheat was applied, if the resin used was PES, the results were positive at concentrations of 0.01%(+), 0.05%(+), 0.1%(+), 0.5%(+), and 1.0%(+). In the case of PE1, the results were positive at concentrations of 0.05% (+), 0.1% (+), 0.5% (+), and 1.0% (+). In the case of PE2, the results were positive at concentrations of 0.05% (+), 0.1% (+), 0.5% (+), and 1.0% (+). In the case of PVA, the results were positive at concentrations of 0.01%(+), 0.05%(+), 0.1%(+), 0.5%(+), and 1.0%(+).

[0164] Therefore, when detecting 2 μg of wheat, it was confirmed that PES and PVA could detect it when Triton X-100 was added to PBS at concentrations of 0.01%, 0.05%, 0.1%, 0.5%, and 1.0%, while PE1 and PE2 could detect it when Triton X-100 was added to PBS at concentrations of 0.05%, 0.1%, 0.5%, and 1.0%.

[0165] (PBS+SDS) 2 μg of each of the food allergen proteins listed below was applied to the detection surface, and the developing solution was prepared by adding 0.01%, 0.05%, 0.1%, 0.5%, or 1.0% SDS to PBS. The results are shown in Tables 6-1 to 6-3.

[0166] [Table 6-1]

[0167] (result) As is clear from Table 6-1, when 2 μg of eggs were applied, if the resin used was PES, the results were positive at concentrations of 0.01%(+), 0.05%(+), 0.1%(+), 0.5%(+), and 1.0%(+). In the case of PE1, the results were positive at concentrations of 0.05% (+), 0.1% (+), 0.5% (+), and 1.0% (+). In the case of PE2, the results were positive at concentrations of 0.05% (+), 0.1% (+), 0.5% (+), and 1.0% (+-). In the case of PVA, the results were positive at concentrations of 0.01%(+), 0.05%(+), 0.1%(+), 0.5%(+), and 1.0%(+).

[0168] Therefore, when detecting 2 μg of eggs, it was confirmed that PES and PVA could detect eggs when the concentrations of SDS added to PBS were 0.01%, 0.05%, 0.1%, 0.5%, and 1.0%, and that PE1 and PE2 could detect eggs when the concentrations of SDS added to PBS were 0.05%, 0.1%, 0.5%, and 1.0%.

[0169] [Table 6-2]

[0170] (result) As is clear from Table 6-2, when 2 μg of milk was applied, if the resin used was PES, the results were positive at concentrations of 0.01% (+) and 0.05% (+w). In the case of PE1, 0.05% (+w) was positive. In the case of PE2, the positive results were 0.05% (+) and 0.1% (+). In the case of PVA, the results were positive at concentrations of 0.01% (+) and 0.05% (+).

[0171] Therefore, when detecting 2 μg of milk, it was confirmed that PES and PVA could detect it when the concentration of SDS added to PBS was 0.01% and 0.05%, PE1 could detect it when the concentration of SDS added to PBS was 0.05%, and PE2 could detect it when the concentration of SDS added to PBS was 0.05% and 0.1%.

[0172] [Table 6-3]

[0173] As is clear from Table 6-3, when 2 μg of wheat was applied, if the resin used was PES, the results were positive at concentrations of 0.01%(+), 0.05%(+), 0.1%(+), 0.5%(+), and 1.0%(+). In the case of PE1, the results were positive at concentrations of 0.05% (+), 0.1% (+), 0.5% (+), and 1.0% (+). In the case of PE2, the results were positive at concentrations of 0.05% (+), 0.1% (+), 0.5% (+), and 1.0% (+). In the case of PVA, the results were positive at concentrations of 0.01%(+), 0.05%(+), 0.1%(+), 0.5%(+), and 1.0%(+).

[0174] Therefore, when detecting 2 μg of wheat, it was confirmed that PES and PVA could detect it when the concentrations of SDS added to PBS were 0.01%, 0.05%, 0.1%, 0.5%, and 1.0%, and that PE1 and PE2 could detect it when the concentrations of SDS added to PBS were 0.05%, 0.1%, 0.5%, and 1.0%.

[0175] (PBS+Tween 20) 2 μg of each of the food allergen proteins listed below was applied to the detection surface, and the developing solution was prepared by adding 0.01%, 0.05%, 0.1%, 0.5%, or 1.0% Tween 20 to PBS. The results are shown in Tables 7-1 to 7-3.

[0176] [Table 7-1]

[0177] (result) As is clear from Table 7-1, when 2 μg of eggs were applied, if the resin used was PES, the results were positive at concentrations of 0.01%(+), 0.05%(+), 0.1%(+), 0.5%(+), and 1.0%(+). In the case of PE1 and PE2, it was impossible to test at any of the concentrations of 0.01%, 0.05%, 0.1%, 0.5%, and 1.0%. In the case of PVA, the results were positive at concentrations of 0.01%(+), 0.05%(+), 0.1%(+), 0.5%(+), and 1.0%(+).

[0178] For PE1 and PE2, detection was impossible at any concentration, and it was confirmed that the developing solution containing Tween 20 is not suitable for use in detecting 2 μg of eggs.

[0179] [Table 7-2]

[0180] (result) As can be seen from Table 7-2, when 2 μg of milk was applied, if the resin used was PES, the results were positive at concentrations of 0.01% (+), 0.05% (+-), 0.1% (+-), 0.5% (+), and 1.0% (+-). In the case of PE1 and PE2, it was impossible to test at any of the concentrations of 0.01%, 0.05%, 0.1%, 0.5%, and 1.0%. In the case of PVA, the results were positive at concentrations of 0.5% (+w) and 1.0% (+-).

[0181] PE1 and PE2 were not detectable at any concentration, confirming that the developer containing Tween 20 is not suitable for use in detecting 2 μg of milk.

[0182] [Table 7-3]

[0183] (result) As is clear from Table 7-3, when 2 μg of wheat was applied, if the resin used was PES, the results were positive at concentrations of 0.01%(+), 0.05%(+), 0.1%(+), 0.5%(+), and 1.0%(+). In the case of PE1 and PE2, it was impossible to test at any of the concentrations of 0.01%, 0.05%, 0.1%, 0.5%, and 1.0%. In the case of PVA, the results were positive at concentrations of 0.01%(+), 0.05%(+), 0.1%(+), 0.5%(+), and 1.0%(+).

[0184] PE1 and PE2 were not detectable at any concentration, confirming that the developer containing Tween 20 is not suitable for use in detecting 2 μg of wheat.

[0185] (PBS+Triton X-100) Tables 8-1 to 8-3 show the results when 1 μg of each of the food allergen proteins listed below was applied to the detection surface and the developing solution was prepared by adding 0.01%, 0.05%, 0.1%, 0.5%, or 1.0% Triton X-100 to PBS.

[0186] [Table 8-1]

[0187] (result) As is clear from Table 8-1, when 1 μg of eggs was applied, if the resin used was PES, the results were positive at concentrations of 0.01% (+), 0.05% (+), 0.1% (+), 0.5% (+), and 1.0% (+-). In the case of PE1, 0.1% (+) and 0.5% (+-) were positive. In the case of PE2, the results were positive in 0.05% (+), 0.1% (+), 0.5% (+), and 1.0% (+w). In the case of PVA, the results were positive at concentrations of 0.01% (+-), 0.05% (+), 0.1% (+), and 0.5% (+).

[0188] Therefore, when detecting 1 μg of eggs, it was confirmed that in the developing solution, PES was capable of detection when Triton X-100 was added to PBS at concentrations of 0.01%, 0.05%, 0.1%, 0.5%, and 1.0%, in PE1 it was capable of detection when Triton X-100 was added to PBS at concentrations of 0.1% and 0.5%, in PE2 it was capable of detection when Triton X-100 was added to PBS at concentrations of 0.05%, 0.1%, 0.5%, and 1.0%, and in PVA it was capable of detection when Triton X-100 was added to PBS at concentrations of 0.01%, 0.05%, 0.1%, and 0.5%.

[0189] [Table 8-2]

[0190] (result) As is clear from Table 8-2, when 1 μg of milk was applied, if the resin used was PES, the results were positive at concentrations of 0.01% (+), 0.05% (+-), 0.1% (+w), and 0.5% (+-). In the case of PE1, 0.05% (+-) and 0.1% (+-) were positive. In the case of PE2, 0.05% (+-) were positive. In the case of PVA, the results were positive at concentrations of 0.05% (+ / -) and 0.1% (+ / -).

[0191] Therefore, when detecting 1 μg of milk, it was confirmed that PES could detect it when Triton X-100 was added to PBS at concentrations of 0.01%, 0.05%, 0.1%, and 0.5%, PE1 could detect it when Triton X-100 was added to PBS at concentrations of 0.05% and 0.1%, PE2 could detect it when Triton X-100 was added to PBS at a concentration of 0.05%, and PVA could detect it when Triton X-100 was added to PBS at concentrations of 0.05% and 0.1%.

[0192] [Table 8-3]

[0193] As is clear from Table 8-3, when 1 μg of wheat was applied, if the resin used was PES, the results were positive at concentrations of 0.01%(+), 0.05%(+), 0.1%(+), 0.5%(+), and 1.0%(+). In the case of PE1, the results were positive at concentrations of 0.05% (+), 0.1% (+), 0.5% (+), and 1.0% (+). In the case of PE2, the results were positive at concentrations of 0.05% (+), 0.1% (+), 0.5% (+), and 1.0% (+). In the case of PVA, the results were positive at concentrations of 0.01%(+), 0.05%(+), 0.1%(+), 0.5%(+), and 1.0%(+).

[0194] Therefore, when detecting 1 μg of wheat, it was confirmed that PES and PVA could detect it when Triton X-100 was added to PBS at concentrations of 0.01%, 0.05%, 0.1%, 0.5%, and 1.0%, while PE1 and PE2 could detect it when Triton X-100 was added to PBS at concentrations of 0.05%, 0.1%, 0.5%, and 1.0%.

[0195] (PBS+SDS) 1 μg of each of the food allergen proteins listed below was applied to the detection surface, and the developing solution was prepared by adding 0.01%, 0.05%, 0.1%, 0.5%, or 1.0% SDS to PBS. The results are shown in Tables 9-1 to 9-3.

[0196] [Table 9-1]

[0197] (result) As is clear from Table 9-1, when 1 μg of eggs was applied, if the resin used was PES, the results were positive at concentrations of 0.01%(+), 0.05%(+), 0.1%(+), 0.5%(+), and 1.0%(+). In the case of PE1, the results were positive in 0.05% (+w), 0.1% (+), 0.5% (+), and 1.0% (+w). In the case of PE2, the positive results were 0.05% (+), 0.1% (+), 0.5% (+), and 1.0% (+-). In the case of PVA, the results were positive at concentrations of 0.01%(+), 0.05%(+), 0.1%(+), 0.5%(+), and 1.0%(+).

[0198] Therefore, when detecting 1 μg of eggs, it was confirmed that PES and PVA could detect eggs when the concentrations of SDS added to PBS were 0.01%, 0.05%, 0.1%, 0.5%, and 1.0%, and that PE1 and PE2 could detect eggs when the concentrations of SDS added to PBS were 0.05%, 0.1%, 0.5%, and 1.0%.

[0199] [Table 9-2]

[0200] (result) As is clear from Table 9-2, when 1 μg of milk was applied, if the resin used was PES, the results were positive at concentrations of 0.01% (+w) and 0.05% (+-). In the case of PE1, the test was either impossible or negative at all concentrations. In the case of PE2, 0.05% (+-) were positive. In the case of PVA, the results were positive at concentrations of 0.01% (+w) and 0.05% (+-).

[0201] Therefore, when detecting 1 μg of milk, it was confirmed that PES and PVA could detect it when the concentration of SDS added to PBS was 0.01% and 0.05%, and that PE2 could detect it when the concentration of SDS added to PBS was 0.05%. However, PE1 was either undetectable or negative at all concentrations.

[0202] [Table 9-3]

[0203] As is clear from Table 9-3, when 1 μg of wheat was applied, if the resin used was PES, the results were positive at concentrations of 0.01%(+), 0.05%(+), 0.1%(+), 0.5%(+), and 1.0%(+). In the case of PE1, the results were positive at concentrations of 0.05% (+), 0.1% (+), 0.5% (+), and 1.0% (+). In the case of PE2, the results were positive at concentrations of 0.05% (+), 0.1% (+), 0.5% (+), and 1.0% (+). In the case of PVA, the results were positive at concentrations of 0.01%(+), 0.05%(+), 0.1%(+), 0.5%(+), and 1.0%(+).

[0204] Therefore, when detecting 1 μg of wheat, it was confirmed that PES and PVA could detect it when the concentrations of SDS added to PBS were 0.01%, 0.05%, 0.1%, 0.5%, and 1.0%, and that PE1 and PE2 could detect it when the concentrations of SDS added to PBS were 0.05%, 0.1%, 0.5%, and 1.0%.

[0205] (PBS+Tween 20) 1 μg of each of the food allergen proteins listed below was applied to the detection surface, and the developing solution was prepared by adding 0.01%, 0.05%, 0.1%, 0.5%, or 1.0% Tween 20 to PBS. The results are shown in Tables 10-1 to 10-3.

[0206] [Table 10-1]

[0207] (result) As is clear from Table 10-1, when 1 μg of eggs was applied, if the resin used was PES, the results were positive at concentrations of 0.01% (+), 0.05% (+), 0.1% (+), 0.5% (+-), and 1.0% (+-). In the case of PE1 and PE2, it was impossible to test at any of the concentrations of 0.01%, 0.05%, 0.1%, 0.5%, and 1.0%. In the case of PVA, the results were positive at concentrations of 0.01% (+), 0.05% (+), 0.1% (+), 0.5% (+w), and 1.0% (+-).

[0208] For PE1 and PE2, detection was impossible at any concentration, and it was confirmed that the developing solution containing Tween 20 is not suitable for use in detecting 1 μg of eggs.

[0209] [Table 10-2]

[0210] (result) As is clear from Table 10-2, when 1 μg of milk was applied, if the resin used was PES, the results were positive at concentrations of 0.01% (+-), 0.05% (+-), 0.1% (+-), 0.5% (+), and 1.0% (+w). In the case of PE1 and PE2, it was impossible to test at any of the concentrations of 0.01%, 0.05%, 0.1%, 0.5%, and 1.0%. In the case of PVA, the results were positive at concentrations of 0.5% (+-) and 1.0% (+-).

[0211] PE1 and PE2 were not detectable at any concentration, confirming that the developer containing Tween 20 is not suitable for use in detecting 1 μg of milk.

[0212] [Table 10-3]

[0213] (result) As is clear from Table 10-3, when 1 μg of wheat was applied, if the resin used was PES, the results were positive at concentrations of 0.01%(+), 0.05%(+), 0.1%(+), 0.5%(+), and 1.0%(+). In the case of PE1 and PE2, it was impossible to test at any of the concentrations of 0.01%, 0.05%, 0.1%, 0.5%, and 1.0%. In the case of PVA, the results were positive at concentrations of 0.01%(+), 0.05%(+), 0.1%(+), 0.5%(+), and 1.0%(+).

[0214] PE1 and PE2 were not detectable at any concentration, confirming that the developer containing Tween 20 is not suitable for use in detecting 1 μg of wheat.

[0215] [Results of this selection process] Taking into account the results of the main selection process described above, as well as the results of the preliminary selection, we conducted a method to select a combination of allergen type, resin type, and developing solution type suitable for the allergen detection kit of the present invention, focusing on whether allergens can be detected at a level of 2 μg / mL. The results are shown below.

[0216] (Egg+PES+TritonX-100) When egg allergens were selected as the detection target, PES was selected as the resin, and a PBS solution containing Triton X-100 was selected as the developing solution, it was confirmed in the preliminary selection process that the passage rate of egg allergens was 95% or higher at Triton X-100 concentrations ranging from 0.01% to 1.0%. Even when 4 μg, 2 μg, or 1 μg of egg allergen was applied to the detection surface and the above allergen detection confirmation test was performed, the allergen was detectable at Triton X-100 concentrations of 0.01% to 1.0%. Taking these results into consideration, it was confirmed that in this selection process, when egg allergens are selected as the detection target, it is preferable to select a combination of PES as the resin and a PBS solution containing 0.01% to 1.0% Triton X-100 as the developing solution.

[0217] (egg + PES + SDS) In the preliminary selection process, it was confirmed that when egg allergens were the detection target, PES was selected as the resin, and a PBS solution containing added SDS was selected as the developing solution, the passage rate of egg allergens was 95% or higher at SDS concentrations of 0.01% to 1.0%. Even when 4 μg, 2 μg, or 1 μg of egg allergen was applied to the detection surface and the above allergen detection confirmation test was carried out, the allergen was detectable at SDS concentrations of 0.01% to 1.0%. Taking these results into consideration, it was confirmed that in this selection process, when egg allergens are selected as the detection target, it is preferable to select a combination of PES as the resin and a PBS solution containing 0.01% to 1.0% SDS as the developing solution.

[0218] (Egg + PES + Tween 20) In the preliminary selection process, it was confirmed that when egg allergens were the detection target, PES was selected as the resin, and a PBS solution containing Tween 20 was selected as the developing solution, the passage rate of egg allergens was 95% or higher at Tween 20 concentrations of 0.01% to 1.0%. Even when 4 μg, 2 μg, or 1 μg of egg allergen was applied to the detection surface and the above allergen detection confirmation test was carried out, the allergen was detectable at Tween 20 concentrations of 0.01% to 1.0%. Taking these results into consideration, it was confirmed that in this selection process, when egg allergens are selected as the detection target, it is preferable to select a combination of PES as the resin and a PBS solution containing 0.01% to 1.0% Tween 20 as the developing solution.

[0219] (Egg+PE+TritonX-100) When egg allergens were used as the detection target, PE was selected as the resin, and a PBS solution containing Triton X-100 was selected as the developing solution, it was confirmed in the preliminary selection process that the passage rate of egg allergens was 95% or higher at Triton X-100 concentrations of 0.05% to 1.0%. However, when 4 μg, 2 μg, or 1 μg of egg allergen was applied to the detection surface and the above-mentioned allergen detection confirmation test was conducted, it was confirmed that when 4 μg of egg allergen was applied to the detection surface, the allergen could not be detected by polyethylene resin (PE1) with an average pore size of 100 μm at a Triton X-100 concentration of 1.0%. Furthermore, when 1 μg was applied, it was confirmed that allergens could not be detected in polyethylene resin (PE1) with an average pore size of 100 μm at Triton X-100 concentrations of 0.05% and 1.0%. Taking these results into consideration, it was confirmed that when egg allergens are selected as the detection target in this selection process, it is preferable to select a combination of polyethylene resin with an average pore size of 100 μm as the resin and a PBS solution containing Triton X-100 at a concentration of 0.1% to 0.5% as the developing solution. On the other hand, when polyethylene resin with an average pore size of 200 μm is selected, it was confirmed that a combination of PBS solution containing Triton X-100 at a concentration of 0.05% to 1.0% can be selected as the developing solution.

[0220] (egg + PE + SDS) In the preliminary selection process, it was confirmed that when egg allergens were the detection target, PE was selected as the resin, and a PBS solution containing added SDS was selected as the developing solution, the passage rate of egg allergens was 95% or higher at all SDS concentrations between 0.05% and 1.0%. Even when 4 μg, 2 μg, or 1 μg of egg allergen was applied to the detection surface and the above allergen detection confirmation test was carried out, the allergen was detectable at SDS concentrations of 0.05% to 1.0%. Taking these results into consideration, it was confirmed that in this selection process, when egg allergens are selected as the detection target, it is preferable to select a combination of PE as the resin and a PBS solution containing 0.05% to 1.0% SDS as the developing solution.

[0221] (Egg + PE + Tween 20) When egg allergens were selected as the detection target, PE was selected as the resin, and a PBS solution containing Tween 20 was selected as the developing solution, the passage rate of egg allergens was confirmed to be 0% (not passing) at all Tween 20 concentrations from 0.01% to 1.0% in the preliminary selection process, and they were also unable to be tested at any concentration in the main selection process.

[0222] (Egg+PVA+TritonX-100) When egg allergens were used as the detection target, PVA was selected as the resin, and a PBS solution containing Triton X-100 was selected as the developing solution, it was confirmed in the preliminary selection process that the passage rate of egg allergens was 95% or higher at Triton X-100 concentrations ranging from 0.01% to 1.0%. However, when 4 μg, 2 μg, or 1 μg of egg allergen was applied to the detection surface and the above-mentioned allergen detection confirmation test was performed, it was confirmed that when 1 μg of egg allergen was applied to the detection surface, the allergen could not be detected by PVA at a Triton X-100 concentration of 1.0%. Taking these results into consideration, it was confirmed that when egg allergens are selected as the detection target in this selection process, it is preferable to select a combination of PVA as the resin and a PBS solution containing Triton X-100 at a concentration of 0.01% to 0.5% as the developing solution.

[0223] (egg + PVA + SDS) In the preliminary selection process, it was confirmed that when egg allergens were the detection target, PVA was selected as the resin, and a PBS solution containing added SDS was selected as the developing solution, the passage rate of egg allergens was 90% or higher at SDS concentrations of 0.01% to 1.0%. Even when 4 μg, 2 μg, or 1 μg of egg allergen was applied to the detection surface and the above allergen detection confirmation test was carried out, the allergen was detectable at SDS concentrations of 0.01% to 1.0%. Taking these results into consideration, it was confirmed that in this selection process, when egg allergens are selected as the detection target, it is preferable to select a combination of PVA as the resin and a PBS solution containing 0.01% to 1.0% SDS as the developing solution.

[0224] (Egg + PVA + Tween 20) In the preliminary selection process, it was confirmed that when egg allergens were selected as the detection target, PVA was selected as the resin, and a PBS solution containing Tween 20 was selected as the developing solution, the passage rate of egg allergens was 90% or higher at Tween 20 concentrations of 0.01% to 1.0%. However, when the above-mentioned allergen detection confirmation test was carried out, it was confirmed that when 4 μg of egg allergen was applied to the detection surface, the allergen could not be detected by PVA at a Tween 20 concentration of 1.0%. Taking these results into consideration, it was confirmed that when egg allergens are selected as the detection target in this selection process, it is preferable to select a combination of PVA as the resin and a PBS solution containing 0.01% to 0.5% Tween 20 as the developing solution.

[0225] (Milk+PES+TritonX-100) When milk allergens were selected as the detection target, PES was selected as the resin, and a PBS solution containing Triton X-100 was selected as the developing solution, it was confirmed in the preliminary selection process that the passage rate of milk allergens was 85% or higher at Triton X-100 concentrations ranging from 0.01% to 1.0%. However, when 4 μg, 2 μg, and 1 μg of milk allergen were applied to the detection surface, it was confirmed that the allergen could not be detected by PES at a Triton X-100 concentration of 1.0%. Taking these results into consideration, it was confirmed that when milk allergens are selected as the detection target in this selection process, it is preferable to select a combination of PES as the resin and a PBS solution containing Triton X-100 at a concentration of 0.01% to 0.5% as the developing solution.

[0226] (Milk+PES+SDS) In the preliminary selection process, it was confirmed that when milk allergens were the detection target, PES was selected as the resin, and a PBS solution containing SDS was selected as the developing solution, the passage rate of milk allergens was 95% or higher at SDS concentrations of 0.01% to 1.0%. However, when 4 μg, 2 μg, and 1 μg of milk allergen were applied to the detection surface, it was confirmed that the allergen could not be detected by PES at SDS concentrations of 0.1%, 0.5%, and 1.0%. Taking these results into consideration, it was confirmed that when milk allergens are selected as the detection target in this selection process, it is preferable to select a combination of PES as the resin and a PBS solution containing 0.01% to 0.05% SDS as the developing solution.

[0227] (Milk + PES + Tween 20) In the preliminary selection process, it was confirmed that when milk allergens were the detection target, PES was selected as the resin, and a PBS solution containing Tween 20 was selected as the developing solution, the passage rate of milk allergens was 75% or higher at Tween 20 concentrations of 0.01% to 1.0%. Even when 4 μg, 2 μg, or 1 μg of milk allergen was applied to the detection surface and the above-mentioned allergen detection confirmation test was performed, the allergen was detectable at Tween 20 concentrations of 0.01% to 1.0%. Taking these results into consideration, it was confirmed that in this selection process, when milk allergens are selected as the detection target, it is preferable to select a combination of PES as the resin and a PBS solution containing Tween 20 at a concentration of 0.01% to 1.0% as the developing solution.

[0228] (Milk+PE+TritonX-100) In the preliminary selection process, it was confirmed that when milk allergens were the detection target, PE was selected as the resin, and a PBS solution containing Triton X-100 was selected as the developing solution, the passage rate of milk allergens was 90% or higher at Triton X-100 concentrations of 0.05% to 1.0%. However, when 4 μg, 2 μg, or 1 μg of milk allergen was applied to the detection surface and the above-mentioned allergen detection confirmation test was conducted, it was confirmed that when 4 μg, 2 μg, or 1 μg of milk allergen was applied to the detection surface of a polyethylene resin (PE1) with an average pore size of 100 μm, the allergen could not be detected at Triton X-100 concentrations of 0.5% and 1.0%. Furthermore, it was confirmed that when 4 μg of milk allergen was applied to the detection surface of polyethylene resin (PE2) with an average pore size of 200 μm, the allergen could not be detected at a Triton X-100 concentration of 1.0%, when 2 μg of milk allergen was applied to the detection surface, at Triton X-100 concentrations of 0.5% and 1.0%, and when 1 μg of milk allergen was applied to the detection surface, at Triton X-100 concentrations of 0.1%, 0.5%, and 1.0%. Taking these results into consideration, when cow's milk allergens are selected as the detection target in this selection process, it is confirmed that the combination of a polyethylene resin with an average pore size of 100 μm as the resin and a PBS solution containing 0.05% to 0.1% Triton X-100 as the developing solution is suitable. It is also confirmed that the combination of a polyethylene resin with an average pore size of 200 μm as the resin and a PBS solution containing 0.05% Triton X-100 as the developing solution is suitable.

[0229] (milk+PE+SDS) In the preliminary selection process, it was confirmed that when milk allergens were the detection target, PE was selected as the resin, and a PBS solution containing added SDS was selected as the developing solution, the passage rate of milk allergens was 95% or higher at SDS concentrations of 0.05% to 1.0%. However, with polyethylene resin (PE1) with an average pore size of 100 μm, it was confirmed that when 4 μg of milk allergen was applied to the detection surface, the allergen could not be detected at an SDS concentration of 1.0%, when 2 μg of milk allergen was applied to the detection surface, at SDS concentrations of 0.1%, 0.5%, and 1.0%, and when 1 μg of milk allergen was applied to the detection surface, at SDS concentrations of 0.05%, 0.1%, 0.5%, and 1.0%. Furthermore, it was confirmed that when 4 μg and 2 μg of milk allergen were applied to the detection surface of polyethylene resin (PE2) with an average pore size of 200 μm, the allergen could not be detected at SDS concentrations of 0.5% and 1.0%, and when 1 μg of milk allergen was applied to the detection surface, the allergen could not be detected at SDS concentrations of 0.1%, 0.5%, and 1.0%. Taking these results into consideration, it was confirmed that when milk allergens are selected as the detection target in this selection process, it is preferable to select a combination of polyethylene resin with an average pore size of 200 μm as the resin and a PBS solution containing 0.05% SDS as the developing solution.

[0230] (Milk + PE + Tween 20) When milk allergens were selected as the detection target, PE was selected as the resin, and a PBS solution containing Tween 20 was selected as the developing solution, the passage rate of milk allergens was confirmed to be 0% (not passing) at all Tween 20 concentrations from 0.01% to 1.0% in the preliminary selection process, and they were also unable to be tested at any concentration in the main selection process.

[0231] (Milk + PVA + Triton X-100) When milk allergens were used as the detection target, PVA was selected as the resin, and a PBS solution containing Triton X-100 was selected as the developing solution, it was confirmed in the preliminary selection process that the passage rate of milk allergens was 85% or higher at Triton X-100 concentrations of 0.05% to 1.0%. However, when 4 μg, 2 μg, or 1 μg of milk allergen was applied to the detection surface and the above-mentioned allergen detection confirmation test was performed, it was confirmed that when 4 μg and 2 μg of milk allergen was applied to the detection surface, the allergen could not be detected by PVA at a Triton X-100 concentration of 1.0%, and when 1 μg of milk allergen was applied to the detection surface, the allergen could not be detected at Triton X-100 concentrations of 0.5% and 1.0%. Taking these results into consideration, it was confirmed that when milk allergens are selected as the detection target in this selection process, it is preferable to select a combination of PVA as the resin and a PBS solution containing Triton X-100 at a concentration of 0.05% to 0.1% as the developing solution.

[0232] (Milk + PVA + SDS) In the preliminary selection process, it was confirmed that when milk allergens were the detection target, PVA was selected as the resin, and a PBS solution containing SDS was selected as the developing solution, the passage rate of milk allergens was 85% or higher at SDS concentrations of 0.01% to 1.0%. However, when 4 μg, 2 μg, or 1 μg of milk allergen was applied to the detection surface and the above-mentioned allergen detection confirmation test was performed, it was confirmed that when 4 μg, 2 μg, and 1 μg of milk allergen was applied to the detection surface, the allergen could not be detected by PVA at SDS concentrations of 0.1%, 0.5%, and 1.0%. Taking these results into consideration, it was confirmed that when milk allergens are selected as the detection target in this selection process, it is preferable to select a combination of PVA as the resin and a PBS solution containing 0.01% to 0.05% SDS as the developing solution.

[0233] (Milk + PVA + Tween 20) In the preliminary selection process, it was confirmed that when milk allergens were used as the detection target, PVA was selected as the resin, and a PBS solution containing Tween 20 was selected as the developing solution, the passage rate of milk allergens was 75% or more at Tween 20 concentrations of 0.5% to 1.0%. When 4 μg, 2 μg, or 1 μg of milk allergen was applied to the detection surface and the above-mentioned allergen detection confirmation test was conducted, it was confirmed that the allergen could not be detected by PVA at Tween 20 concentrations of 0.01%, 0.05%, and 0.1% when 2 μg and 1 μg were applied. Taking these results into consideration, it was confirmed that when milk allergens are selected as the detection target in this selection process, it is preferable to select a combination of PVA as the resin and a PBS solution containing 0.5% to 1.0% Tween 20 as the developing solution.

[0234] (Wheat+PES+TritonX-100) When wheat allergens were used as the detection target, PES was selected as the resin, and a PBS solution containing Triton X-100 was selected as the developing solution, the passage rate of wheat allergens was confirmed to be 95% or higher at Triton X-100 concentrations of 0.01% to 1.0% in a preliminary selection process. Even when 4 μg, 2 μg, or 1 μg of wheat allergen was applied to the detection surface and the above allergen detection confirmation test was performed, the allergens could be detected at Triton X-100 concentrations of 0.01% to 1.0%. Taking these results into consideration, it was confirmed that in this selection process, when wheat allergens are selected as the detection target, it is preferable to select a combination of PES as the resin and a PBS solution containing Triton X-100 at a concentration of 0.01% to 1.0% as the developing solution.

[0235] (wheat+PES+SDS) When wheat allergens were used as the detection target, PES was selected as the resin, and a PBS solution containing SDS was selected as the developing solution, it was confirmed in the preliminary selection process that the passage rate of wheat allergens was 85% or higher at SDS concentrations of 0.01% to 1.0%. Even when 4 μg, 2 μg, or 1 μg of wheat allergen was applied to the detection surface and the above allergen detection confirmation test was performed, the allergen was detectable at SDS concentrations of 0.01% to 1.0%. Taking these results into consideration, it was confirmed that in this selection process, when wheat allergens are selected as the detection target, it is preferable to select a combination of PES as the resin and a PBS solution containing 0.01% to 1.0% SDS as the developing solution.

[0236] (wheat+PES+Tween20) When wheat allergens were selected as the detection target, PES was selected as the resin, and a PBS solution containing Tween 20 was selected as the developing solution, it was confirmed in the preliminary selection process that the passage rate of wheat allergens was 85% or higher at Tween 20 concentrations of 0.01% to 1.0%. Even when 4μg, 2μg, or 1μg of wheat allergen was applied to the detection surface and the above allergen detection confirmation test was performed, the allergen could be detected at Tween 20 concentrations of 0.01% to 1.0%. Taking these results into consideration, it was confirmed that in this selection process, when wheat allergens are selected as the detection target, it is preferable to select a combination of PES as the resin and a PBS solution containing Tween 20 at a concentration of 0.01% to 1.0% as the developing solution.

[0237] (Wheat+PE+TritonX-100) When wheat allergens were used as the detection target, PE was selected as the resin, and a PBS solution containing Triton X-100 was selected as the developing solution, it was confirmed in the preliminary selection process that the passage rate of wheat allergens was 85% or higher at Triton X-100 concentrations of 0.05% to 1.0%. Even when 4 μg, 2 μg, or 1 μg of wheat allergen was applied to the detection surface and an allergen detection confirmation test was performed, the allergen was detectable at Triton X-100 concentrations of 0.05% to 1.0%. Taking these results into consideration, it was confirmed that in this selection process, when wheat allergens are selected as the detection target, it is preferable to select a combination of PE as the resin and a PBS solution containing Triton X-100 at a concentration of 0.05% to 1.0% as the developing solution.

[0238] (wheat+PE+SDS) When wheat allergens were used as the detection target, PE was selected as the resin, and a PBS solution containing SDS was selected as the developing solution, it was confirmed in the preliminary selection process that the passage rate of wheat allergens was 95% or higher at SDS concentrations of 0.05% to 1.0%. Even when 4 μg, 2 μg, or 1 μg of wheat allergen was applied to the detection surface and an allergen detection confirmation test was performed, the allergen was detectable at SDS concentrations of 0.05% to 1.0%. Taking these results into consideration, it was confirmed that in this selection process, when wheat allergens are selected as the detection target, it is preferable to select a combination of PES as the resin and a PBS solution containing 0.05% to 1.0% SDS as the developing solution.

[0239] (wheat+PE+Tween20) When wheat allergens were selected as the detection target, PE was selected as the resin, and a PBS solution containing Tween 20 was selected as the developing solution, the passage rate of wheat allergens was confirmed to be 0% (not passing) at all Tween 20 concentrations from 0.01% to 1.0% in the preliminary selection process, and they were also unable to be tested at any concentration in the main selection process.

[0240] (Wheat+PVA+TritonX-100) When wheat allergens were used as the detection target, PVA was selected as the resin, and a PBS solution containing Triton X-100 was selected as the developing solution, it was confirmed in the preliminary selection process that the passage rate of wheat allergens was 85% or higher at Triton X-100 concentrations ranging from 0.01% to 1.0%. Even when 4 μg, 2 μg, or 1 μg of wheat allergen was applied to the detection surface and the above allergen detection confirmation test was performed, the allergen was detectable at Triton X-100 concentrations of 0.01% to 1.0%. Taking these results into consideration, it was confirmed that in this selection process, when wheat allergens are selected as the detection target, it is preferable to select a combination of PVA as the resin and a PBS solution containing Triton X-100 at a concentration of 0.01% to 1.0% as the developing solution.

[0241] (wheat+PVA+SDS) When wheat allergens were used as the detection target, PVA was selected as the resin, and a PBS solution containing SDS was selected as the developing solution, it was confirmed in the preliminary selection process that the passage rate of wheat allergens was 90% or higher at SDS concentrations of 0.05% to 1.0%. Even when 4 μg, 2 μg, or 1 μg of wheat allergen was applied to the detection surface and the above allergen detection confirmation test was performed, the allergen was detectable at SDS concentrations of 0.01% to 1.0%. Taking these results into consideration, it was confirmed that in this selection process, when wheat allergens are selected as the detection target, it is preferable to select a combination of PVA as the resin and a PBS solution containing 0.01% to 1.0% SDS as the developing solution.

[0242] (wheat+PVA+Tween20) When wheat allergens were used as the detection target, PVA was selected as the resin, and a PBS solution containing Tween 20 was selected as the developing solution, it was confirmed in the preliminary selection process that the passage rate of wheat allergens was 80% or higher at Tween 20 concentrations of 0.01% to 1.0%. Even when 4 μg, 2 μg, or 1 μg of wheat allergen was applied to the detection surface and the above allergen detection confirmation test was carried out, the allergen was detectable at Tween 20 concentrations of 0.01% to 1.0%. Taking these results into consideration, it was confirmed that in this selection process, when wheat allergens are selected as the detection target, it is preferable to select a combination of PVA as the resin and a PBS solution containing Tween 20 at a concentration of 0.01% to 1.0% as the developing solution.

[0243] (summary) As described above, when carrying out the method of the present invention, by carrying out a preliminary selection and taking into account the results of the main selection, it has become clear that it is possible to clarify the range of combinations of allergen types, resin types, and developing solution types that are highly suitable for accurate allergen detection kits. [Industrial Applicability]

[0244] The allergen detection kit of the present invention, which can rapidly and accurately detect allergens remaining in dry areas at food production sites where food is dried, is particularly useful in the food industry. [Explanation of symbols]

[0245] 1 Main unit case 2 Lid 3 Wiping section 4 Wiping surface 5 Far end of main case 6 Near end of main body case 7 Main body case opening piece 8 Lid release piece

Claims

1. A method for selecting a combination of an allergen type, a resin type, and a developing solution type suitable for an allergen detection kit, comprising the steps of (a) and (b) below: (a) A test solution containing 5 ppm of allergen was prepared by adding 500 μL of a developer solution prepared by adding a surfactant to phosphate buffered saline at a concentration of 0.01%, 0.05%, 0.1%, 0.5%, or 1.0% to a test piece having a top and bottom area of ​​35 mm ２ a step of pre-selecting a combination of the type of allergen, the type of resin, and the type of developing liquid, which, when dropped onto the top surface of a rectangular parallelepiped resin body having a height of 10 mm, results in a penetration rate of 75% or more of the allergen through the bottom surface of the resin body within 10 minutes; (b) conducting an allergen detection confirmation test for the preselected combination, and finally selecting a combination of the type of allergen, the type of resin, and the type of developer suitable for the allergen detection kit, which is capable of detecting the allergen at a level of 2 μg / mL;

2. 2. The method according to claim 1, wherein the rate of allergens passing through the bottom surface of the resin body is 85% or more within 10 minutes.

3. 3. The method according to claim 1, wherein the resin is porous.

4. 4. The method according to claim 1, wherein the resin is selected from the group consisting of polyester, polyethylene, and polyvinyl alcohol.

5. 5. The method according to claim 1, wherein the surfactant is polyoxyethylene (10) octylphenyl ether, sodium dodecyl sulfate, or polyoxyethylene sorbitan monolaurate.

6. 6. The method according to any one of claims 1 to 5, wherein the allergen is a food allergen.

7. 7. The method of claim 6, wherein the food allergen is egg, milk, or wheat.

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