Use of allergen samples prepared from household waste

Abstract

To provide a measurement kit for evaluating allergen inactivation and a method for evaluating allergen-inactivating substances which enable more accurate evaluation.SOLUTION: A method includes: a reaction step in which an allergen-inactivating substance is reacted with an allergen sample by using an allergen inactivation evaluation kit including an allergen sample prepared from household waste and an extraction solvent made of a buffer solution; a test sample extraction step in which an extraction solvent is added to the reaction product obtained in the reaction step to obtain a test sample extract; a control sample extraction step for adding the extraction solvent to the allergen sample for acquiring a control sample extract; an allergen content measurement step in which the allergen content in the test sample extract and the allergen content in the control sample extract are measured by ELISA; and an allergen inactivation rate calculation step in which the allergen inactivation rate is calculated on the basis of the allergen content in the control sample extract and the allergen content in the test sample extract.SELECTED DRAWING: Figure 1

Description

[Technical Field]

[0001] The present invention relates to a measurement kit for evaluating allergen inactivation and a method for evaluating allergen inactivating substances. [Background technology]

[0002] In recent years, many allergic diseases, including allergic rhinitis, atopic dermatitis, and bronchial asthma, have become a problem. The types of allergens that cause allergic diseases include food, drugs, indoor dirt and dust (house dust, such as pet hair and mites), and pollen. When people with allergies come into contact with allergens, an allergic reaction is triggered, resulting in symptoms such as hives, dermatitis, asthma, and fever.

[0003] In particular, mites live in bedding such as futons and flooring materials that are commonly found in living environments, and it is known that not only living organisms but also pieces of their carcasses and excrement can be allergens.

[0004] As one of the countermeasures for allergic diseases and other allergic symptoms, attempts have been made to develop drugs that inactivate the causative allergens.

[0005] For example, Patent Document 1 below discloses an invention of a method for removing allergens from the environment, characterized by treating the environment with tannic acid. It states that treating house dust containing Dermatophagoides pteronyssinus, or extracts from the grass Phleum pratense or the weed Plantago lanceolata with a 1% aqueous solution of tannic acid can reduce allergenicity in skin reaction tests.

[0006] Furthermore, Patent Document 2 below describes that by binding tannic acid, which is highly safe for the human body, to fibers or textile products containing cellulosic fibers via a crosslinking agent, it is possible to prevent the allergen inactivation activity of tannic acid from decreasing even after repeated washing.

[0007] Meanwhile, the effectiveness of various substances other than tannic acid has been suggested. For example, Patent Document 3 below describes the use of a compound or salt thereof that has at least one guanidino group as a cationic group in the molecule and has surfactant properties. Patent Document 4 below describes the use of a pomegranate leaf extract using a solvent consisting of at least one selected from water and hydrophilic solvents. Patent Document 5 below describes the use of antiallergenic metal components such as silver and zinc. Patent Document 6 below describes the use of a polymer compound having a styrenesulfonic acid unit. [Prior art documents] [Patent documents]

[0008] [Patent Document 1] Japanese Patent Application Publication No. 61-44821 [Patent Document 2] Japanese Patent Application Laid-Open No. 2007-107149 [Patent Document 3] Japanese Patent Application Publication No. 9-157152 [Patent Document 4] Japanese Patent Application Laid-Open No. 2002-370996 [Patent Document 5] Japanese Patent Application Laid-Open No. 2006-241431 [Patent Document 6] Japanese Patent Application Laid-Open No. 2009-155453 Summary of the Invention [Problem to be solved by the invention]

[0009] In general, in the development of allergen inactivators, drugs expected to have inactivating properties are evaluated by reacting them with allergens and measuring the allergen content in the solution after the reaction using ELISA, etc. However, when food residue, animal hair, dead mites, or pollen are used as allergen samples to be reacted with allergen inactivators, immunogenic allergens remain inside these allergen samples, making it impossible to accurately measure the allergen content, and as a result, there is a problem in that the inactivating properties of the test drug cannot be accurately evaluated.

[0010] Therefore, an object of the present invention is to provide a measurement kit for evaluating allergen inactivation and a method for evaluating allergen inactivating substances, which enable more accurate evaluation of the inactivating ability of allergen inactivating agents. [Means for solving the problem]

[0011] The present inventors have conducted extensive research to achieve the above object and have completed the present invention.

[0012] That is, the allergen deactivation evaluation kit of the present invention comprises an allergen sample prepared from household waste and an extraction solvent consisting of a buffer solution.

[0013] In the allergen deactivation evaluation kit of the present invention, the allergen sample preferably consists of fine dust obtained by sieving household waste.

[0014] Furthermore, it is preferable that the allergen contained in the allergen sample is one or more types selected from dust mites, pollen, animal hair, animal excrement, dandruff, mold, food residue, and substances derived from sanitary animals.

[0015] The extraction solvent is preferably one selected from a phosphate buffer solution, a carbonate buffer solution, and an acetate buffer solution.

[0016] Furthermore, it is preferable that the extraction solvent is a phosphate buffer solution when the allergen is a mite, a carbonate buffer solution when the allergen is pollen, and an acetate buffer solution when the allergen contains collagen.

[0017] Furthermore, it is preferable that the allergen sample be selected through an additive recovery test in which a predetermined amount of the target allergen is added to confirm whether or not measurement interfering substances affect the ELISA measurement.

[0018] On the other hand, the method for evaluating an allergen inactivating substance of the present invention is characterized by comprising the following steps: a reaction step using the allergen inactivation evaluation kit to react an allergen sample with an allergen inactivating substance to obtain a test product; a test product extraction step after the reaction step to obtain a test product extract by adding the extraction solvent; a control product extraction step to prepare a control product without reacting the allergen inactivating substance with the allergen sample and obtain a control product extract by adding the extraction solvent; an allergen content measurement step to measure the allergen contents of a solution derived from the test product containing the test product extract and a solution derived from the control product containing the control product extract by ELISA; and an allergen inactivation rate calculation step to calculate the allergen inactivation rate from the allergen content of the solution derived from the test product and the allergen content of the solution derived from the control product.

[0019] In the method for evaluating an allergen inactivating substance of the present invention, it is preferable that the test product extraction step further comprises a step of adding the extraction solvent to the residue from which the test product extract has been collected and collecting the supernatant again to obtain a test product supernatant, and that the control product extraction step further comprises a step of adding the extraction solvent to the residue from which the control product extract has been collected and collecting the supernatant again to obtain a control product supernatant, and that the solution derived from the test product contains the test product supernatant and the solution derived from the control product contains the control product supernatant. [Effects of the Invention]

[0020] According to the present invention, it is possible to provide a measurement kit for evaluating allergen inactivation and a method for evaluating allergen inactivating substances, which enable evaluation with higher accuracy. [Brief explanation of the drawings]

[0021] [Figure 1] FIG. 1 is a schematic diagram showing the test procedure of the method of the present invention. [Figure 2] FIG. 2 is a schematic diagram showing the test procedure for the method of evaluating an allergen inactivating substance when the allergen inactivating substance is used as a solution. [Figure 3] FIG. 3 is a schematic diagram showing the test procedure for the method of evaluating an allergen inactivating substance when the allergen inactivating substance is an airborne substance such as nano-sized charged water particles. DETAILED DESCRIPTION OF THE INVENTION

[0022] The present invention provides an evaluation kit and evaluation method for evaluating the level of inactivation ability of any allergen inactivating substance against an allergen.

[0023] Here, the term "allergen inactivator" may refer to any substance desired by the evaluator, including substances known to have allergen-suppressing activity, such as those with a polar moiety in their chemical structure that inhibits allergens. Examples of such allergen inactivators include water-insoluble polymers with phenolic hydroxyl groups, anionic surfactants, compounds or polymers with hydroxyl groups, and microparticles of metal salts supported on inorganic carriers. More specifically, the allergen inactivator may be a natural extract such as persimmon tannin, tannic acid, catechin, methylated catechin, hydroxybenzoic acid compounds such as dihydroxybenzoic acid and 2,4,6-trihydroxybenzoic acid or their salts, polystyrene compounds such as parahydroxypolystyrene and polystyrene sulfonate, water-insoluble zinc compounds or water-insoluble zinc-metal oxide composite particles, silicon oxide, zinc oxide, aluminum oxide, photocatalytic titanium oxide, or inorganic salt compounds such as calcium salts and strontium salts. Alternatively, it may be applied to evaluate the allergen inactivation ability of substances that are effective for air sterilization, virus removal, decontamination, deodorization, etc., such as nano-sized charged water particles containing OH radicals generated by applying high voltage to water in the air, stable ionized oxygen and water molecules generated by plasma discharge, and microparticulated hypochlorous acid solution generated by electrolyzing saline solution.

[0024] Furthermore, the term "inactivation ability" refers to the ability to inactivate allergens known to cause allergies in humans or animals.

[0025] The allergen inactivation evaluation kit of the present invention comprises an allergen sample prepared from household waste and an extraction solvent made of a buffer solution. The allergen inactivation evaluation kit of the present invention enables more accurate evaluation of the inactivation ability of allergen inactivating substances.

[0026] The allergen sample is prepared from household waste, typically from house dust. In this case, it is preferable that the household waste is sieved to obtain fine dust. By sieving the fine dust, large particles contained in the household waste can be removed, and an allergen sample with a high allergen content can be prepared.

[0027] The sieve preferably has an opening of 0.3 mm or less. By using a sieve with this opening, it is possible to remove relatively large dust particles that are not allergens while leaving relatively small dust particles that may be allergens, such as dead mites.

[0028] The allergen sample may also be fine dust obtained by sieving household waste, crushing the fine dust using a known method such as beads, and then sieving the resulting fine dust. Fine dust can be obtained by sieving household waste through a sieve with a mesh size of 0.3 mm to extract the fine dust, crushing the fine dust, and then sieving the resulting fine dust through a sieve with a mesh size of 0.18 mm.

[0029] When the allergen inactivating substance tested using the allergen inactivation evaluation kit is a substance that is effective for air sterilization, virus removal, decontamination, deodorization, etc., such as the above-mentioned charged atomized water particles, stable ionized oxygen-water molecules, or atomized hypochlorous acid aqueous solution, the allergen sample is preferably the above-mentioned fine dust.

[0030] The allergen sample contains an allergen, and the allergens contained are preferably one or more, more preferably two or more, selected from dust mites, pollen, animal hair, animal excrement, dandruff, mold, food residue, and substances derived from sanitary animals. By containing multiple types of allergens, one allergen sample can be used for inactivation ability tests against various allergens.

[0031] Examples of the mites include Dermatophagoides pteronyssinus, Dermatophagoides farinae, Acaridae, Tyrophagus putrescentiae, and Dermoptera spp., with Dermatophagoides pteronyssinus and Dermatophagoides farinae being preferred.

[0032] Examples of the pollen include pollen from plants of the families Cupressaceae, Cupressaceae, Birch, Poaceae, and Asteraceae, and it is preferable that the pollen be from plants of the families Cupressaceae or Cupressaceae.

[0033] Examples of animals from which the animal hair and animal excrement are derived include dogs, cats, birds, rabbits, and the like.

[0034] Examples of the above-mentioned molds include Alternaria (sob mold), Aspergillus (aspergillus), Penicillium (blue mold), and Cladosporium (black mold).

[0035] Examples of foods from which the food residues are derived include wheat, buckwheat, eggs, milk, peanuts, shrimp, crab, abalone, squid, salmon roe, cashew nuts, kiwi fruit, oranges, beef, walnuts, sesame, salmon, mackerel, soybeans, chicken, bananas, pork, matsutake mushrooms, peaches, yams, apples, gelatin, etc. In these cases, the foods may be processed into powders such as wheat flour, buckwheat flour, milk powder, soybean flour, etc.

[0036] Examples of the sanitary animals include insects and rats.

[0037] Examples of methods for appropriately incorporating appropriate allergens into the allergen sample include collecting household waste from a location that is likely to contain the target allergen, such as collecting household waste from a hot and humid location suitable for the proliferation of dust mites and mold, collecting household waste in early spring when there is a lot of pollen in the air, or collecting household waste from a home with pets.Furthermore, household waste collected from multiple locations with different characteristics may be mixed.

[0038] Methods for collecting household waste include vacuuming house dust present in a typical household, wiping it away with a wiper sheet, etc. Furthermore, the allergen sample may further contain, in addition to the household waste, cultures obtained by culturing mites, molds, or sanitary animals, pollen collected directly from plants, commercially available allergen extracts, etc.

[0039] Whether or not the allergens are contained in an allergen sample can be confirmed by known detection methods based on antigen-antibody reactions, such as ELISA (Enzyme-Linked Immunosorbent Assay), sandwich immunoassay, immunoblot, immunoprecipitation, immunochromatography, and immunocap.

[0040] ELISA measurement is a method for quantifying allergen activity with high accuracy, and is also called enzyme immunoassay. ELISA kits are commercially available depending on the allergen used, and allergen activity can be measured using these commercially available products. The measured allergen activity can then be used to determine the amount of allergen contained in the sample.

[0041] However, it is preferable that the allergen sample be one that has been selected through an additive recovery test in which a predetermined amount of the target allergen is added to confirm whether or not any measurement-interfering substances affect the ELISA measurement.

[0042] In general, in ELISA assays, unknown substances other than the target allergen contained in the allergen sample may interfere with the analysis. These unknown substances are the above-mentioned measurement interfering substances. When measurement interfering substances interfere with the analysis, the measurement of allergen activity results in an inaccurate result. This phenomenon is also called matrix interference.

[0043] The spiked recovery test is a test to check for the presence or absence of the above-mentioned matrix interference. Specifically, it can be performed as follows: An allergen sample is divided into two, and a predetermined amount of the target allergen is added to one (hereinafter referred to as the spiked sample). Nothing is added to the other (hereinafter referred to as the unspiked sample), and the allergen amounts in both samples are measured by ELISA. The amount of allergen in the unspiked sample is subtracted from the amount of allergen in the spiked sample, and then divided by the amount of allergen added to the spiked sample, and then multiplied by 100 to calculate the spiked recovery rate (%).

[0044] When there is absolutely no matrix interference, the recovery rate is 100%, and the greater the effect of matrix interference, the greater the deviation of the recovery rate from 100%. Therefore, by selecting an allergen sample based on the calculated recovery rate, ELISA measurements can be performed with greater accuracy. The allergen sample of the present invention preferably has a recovery rate of 90 to 110%.

[0045] The extraction solvent is a buffer solution used to extract allergens from an allergen sample. By adding the extraction solvent to the allergen sample, the allergens are extracted from the allergen sample, allowing for more accurate measurement of the allergen content and more accurate evaluation of the allergen inactivation ability of an allergen inactivating substance.

[0046] Examples of buffer solutions used in the extraction solvent include phosphate buffer, carbonate buffer, acetate buffer, citrate buffer, citrate-phosphate buffer, borate buffer, tartrate buffer, Tris buffer, etc. Among these, a buffer solution selected from phosphate buffer, carbonate buffer, and acetate buffer is preferred. By using a buffer solution selected from phosphate buffer, carbonate buffer, and acetate buffer, allergens can be more easily extracted.

[0047] Furthermore, when the allergen contained in the allergen sample is a dust mite, it is preferable that the extraction solvent be a phosphate buffer solution; when the allergen is pollen, it is preferable that the extraction solvent be a carbonate buffer solution; and when the allergen contains collagen, it is preferable that the extraction solvent be an acetate buffer solution.

[0048] Here, "materials containing collagen" means materials containing animal skin, etc., including animal hair, dandruff, and materials derived from sanitary animals.

[0049] Allergens are more easily extracted if a phosphate buffer solution is used for dust mites, a carbonate buffer solution for pollen, or an acetate buffer solution for substances containing collagen.

[0050] The allergen inactivation evaluation kit of the present invention may contain, as minimum components, the allergen sample and the extraction solvent, and other elements, such as an ELISA kit for use in ELISA measurement, a test vessel such as a test tube or a petri dish for reacting with an allergen inactivating substance, a surfactant for preventing adsorption of an allergen substance, and other elements not described herein.

[0051] Next, the method for evaluating an allergen inactivating substance according to the present invention will be described. As shown in Figure 1, the method for evaluating an allergen inactivating substance according to the present invention comprises the following steps: a reaction step in which an allergen sample is reacted with an allergen inactivating substance using the allergen inactivation evaluation kit to obtain a test product; a test product extraction step in which an extraction solvent is added to obtain a test product extract after the reaction step; a control product extraction step in which an extraction solvent is added to a control sample without treating the allergen inactivating substance to obtain a control product extract; an allergen content measurement step in which the allergen contents of a solution derived from the test product containing the test product extract and a solution derived from a control product containing the control product extract are measured by ELISA; and an allergen inactivation rate calculation step in which the allergen contents of the solution derived from the test product and the solution derived from the control product are calculated.

[0052] According to the method for evaluating an allergen inactivating substance of the present invention, the inactivating ability of an allergen inactivating substance can be evaluated with higher accuracy.

[0053] (Reaction step) An allergen inactivator is added to an allergen sample to obtain a test sample. The method for adding the allergen inactivator to the allergen sample is not particularly limited, and examples thereof include, when the allergen inactivator is used in the form of a solution, placing the allergen sample in a test container such as a test tube or a petri dish and then adding the allergen inactivator, or uniformly scattering the allergen sample on a felt fabric to allow it to adhere, and then spraying the allergen inactivator. When the allergen inactivator is an airborne substance such as nano-sized charged water particles, examples thereof include, when the allergen inactivator is an airborne substance, placing the allergen sample in a petri dish after subjecting it to an antistatic treatment, vibrating the dish to uniformly coat the allergen sample, and then spraying the allergen inactivator onto the dish.

[0054] After adding an allergen deactivator to an allergen sample, a step of allowing the allergen deactivator to stand for a certain period of time to promote the reaction of the allergen deactivator may be carried out. The reaction temperature for the reaction is not particularly limited, and may be adjusted to a temperature below the temperature at which proteins denature, for example, within the range of 5°C to 30°C, preferably 10°C to 25°C. The reaction time is also not particularly limited, and may be adjusted, for example, between 5 minutes and 24 hours. If the reaction time is set long, the water content in the test sample may decrease due to volatilization or evaporation, which may change the allergen concentration in the test sample, so it is preferable to keep the test container sealed.

[0055] (Test sample extraction process) An extraction solvent is added to the test sample obtained in the reaction step to extract the allergen from the test sample, thereby obtaining a test sample extract. The test sample extract is preferably the supernatant obtained by adding the extraction solvent to the test sample, recovering the liquid, and then centrifuging. When the allergen inactivating substance is used in the form of a solution, the extraction solvent may be added after centrifuging the test sample and recovering the resulting supernatant with a pipette, or the extraction solvent may be added without recovering the supernatant. When recovering the supernatant, the extraction solvent can be added to the residue from which the supernatant has been recovered, followed by stirring, allowing to stand, and then centrifuging in the same manner.

[0056] Furthermore, if the reaction step is carried out by spraying an allergen inactivating substance onto a felt fabric, the test sample extraction step can be carried out by immersing the felt fabric in an extraction solvent.If the allergen inactivating substance is an airborne substance such as nano-sized charged water particles and the reaction step is carried out by spraying it onto a petri dish, the test sample extraction step can be carried out by rinsing the petri dish with the extraction solvent, recovering the solution, and centrifuging it.

[0057] The amount of extraction solvent to be added is not particularly limited, but is preferably 10 to 1,000 times, and more preferably 50 to 750 times, the mass of the allergen sample. After adding the extraction solvent, the mixture may be stirred using a stirrer such as a rotator and left to stand for a certain period of time to promote the reaction of the allergen inactivating substance and the extraction of the allergen. The temperature during the standing period can be adjusted in the same manner as in the reaction step, and it is preferable to seal the test container. Furthermore, to obtain a test sample extract, the reaction of the allergen inactivating substance may be stopped by cooling after standing for a certain period of time.

[0058] Preferably, the test sample extraction step further comprises the step of adding an extraction solvent to the residue from which the test sample extract has been collected, and then collecting the supernatant again to obtain a test sample supernatant. The step of collecting the supernatant again can be performed by adding an extraction solvent to the residue from which the test sample extract has been collected, stirring, allowing it to stand, and then similarly centrifuging. By further including the step of collecting the supernatant again, it is possible to extract a larger amount of allergens, and more accurately evaluate the inactivation ability of the allergen inactivating substance.

[0059] (Control product extraction process) A control sample is prepared by extracting the allergen from the allergen sample without treating it with an allergen inactivating substance in the same manner as in the test sample extraction step, to obtain a control sample extract. The control sample may be prepared by adding distilled water or an extraction solvent in the same amount as the allergen inactivating substance added in the reaction step to the allergen sample in the same manner as in the reaction step. When adding the extraction solvent to the control sample, the control sample may be centrifuged, the supernatant obtained is recovered with a pipette, or the extraction solvent may be added without recovering the supernatant. The supernatant may be recovered in the same manner as in the test sample extraction step.

[0060] Preferably, the test product extraction step further comprises a step of adding an extraction solvent to the residue from which the control product extract has been collected, and then collecting the supernatant again to obtain a control product supernatant. The step of collecting the supernatant again can be performed in the same manner as the test product extraction step. By further including the step of collecting the supernatant again, it is possible to extract a larger amount of allergens, and more accurately evaluate the inactivation ability of the allergen inactivating substance.

[0061] (Allergen content measurement process) A solution derived from the test product containing the test product extract and a solution derived from the control product containing the control product extract are collected and their allergen activity is evaluated by ELISA. The amount of allergen contained in each solution can be calculated based on the measured allergen activity (antigenicity). The solution derived from the test product contains the test product extract and, if the test product is centrifuged and a supernatant is collected in the test product extraction step, also contains the supernatant. The solution derived from the control product contains the control product extract and, if the control product is centrifuged and a supernatant is collected in the control product extraction step, also contains the supernatant. Furthermore, the solution derived from the test product and the solution derived from the control product preferably contain a supernatant obtained by adding an extraction solvent to the residue from which the test product extract or the control product extract is collected. For evaluation by ELISA, commercially available ELISA kits can be used depending on the target allergen. The test method may follow known techniques.

[0062] (Allergen inactivation rate calculation process) The allergen inactivation rate is calculated from the allergen content of the solution derived from the test product and the allergen content of the solution derived from the control product calculated in the allergen content measurement step. The allergen inactivation rate can be calculated, for example, by the following formula. Inactivation rate (%) = 100 - (allergen content of the solution derived from the test product / allergen content of the solution derived from the control product) x 100

[0063] A method for determining whether an allergen inactivating substance has inactivation ability is to determine whether it has inactivation ability when the inactivation rate is 95% or higher, and whether it has inactivation ability when it is less than 95%. The threshold for the inactivation rate to confirm the inactivation ability may be 90% or higher, or 80% or higher. The threshold for the inactivation rate to deny the inactivation ability may be 50% or lower, or 25% or lower.

[0064] An example of the test procedure for the method of evaluating allergen inactivating substances, from the reaction step to the allergen inactivation rate calculation step, will be described with reference to FIGS. 2 and 3.

[0065] First, the case where the allergen inactivating substance is used as a solution will be described with reference to FIG.

[0066] In the reaction step, a predetermined amount of allergen sample 1 and allergen inactivating substance (solution) 2 are placed in one test tube, stirred, and allowed to stand to obtain test sample 11. In the test sample extraction step, test sample 11 is centrifuged to collect supernatant 12, and then extraction solvent 3 is added to residue 13 from which supernatant 12 has been collected, and the mixture is stirred and allowed to stand. Further centrifugation is performed to collect the supernatant, test sample extract 14. In the control sample extraction step, a predetermined amount of allergen sample 1 and distilled water are placed in another test tube, stirred, and allowed to stand to obtain control sample 21. Next, control sample 21 is centrifuged to collect supernatant 22, and then extraction solvent 3 is added to residue 23 from which supernatant 22 has been collected, stirred, allowed to stand, and further centrifugation is performed to collect the supernatant, control sample extract 24.

[0067] In the allergen content measurement step, the allergen contents of supernatant 12, test product extract 14, supernatant 22, and control product extract 24 are each measured by ELISA. Here, supernatant 12 and test product extract 14 are solutions derived from test product 11, and supernatant 22 and control product extract 24 are solutions derived from control product 21. Next, in the allergen inactivation rate calculation step, the inactivation rate is calculated from the allergen contents of supernatant 12, test product extract 14, supernatant 22, and control product extract 24 using the following formula: Inactivation rate (%) = 100 - (total amount of allergen content in supernatant 12 and test product extract 14 / total amount of allergen content in supernatant 22 and control product extract 24) x 100

[0068] In the manner described above, when the allergen inactivating substance is used in the form of a solution, the method for evaluating the allergen inactivating substance can be carried out.

[0069] Next, a case where the allergen inactivating substance is an airborne substance such as nano-sized charged water particles will be described with reference to FIG.

[0070] After two petri dishes are treated to prevent static electricity, allergen sample 1 is placed inside and the dishes are vibrated to evenly coat the allergen sample 1. In the reaction step, a predetermined amount of allergen inactivating substance 2 is sprayed onto one of the dishes and left to stand to obtain test sample 11. In the test sample extraction step, extraction solvent 3 is added to test sample 11, and the dish is rinsed to recover solution 15. Solution 15 is centrifuged, and the supernatant, test sample extract 14, is recovered. In the control sample extraction step, extraction solvent 3 is added to the other petri dish, and the dish is rinsed to recover solution 25. Solution 25 is centrifuged, and the supernatant, control sample extract 24, is recovered.

[0071] In the allergen content measurement step, the allergen contents of the test product extract 14 and the control product extract 24 are measured by ELISA. Here, the test product extract 14 is a solution derived from the test product 11, and the control product extract 24 is a solution derived from the control product 21. Next, in the allergen inactivation rate calculation step, the inactivation rate is calculated from the allergen contents of the test product extract 14 and the control product extract 24 using the following formula: Inactivation rate (%) = 100 - (allergen content in test product extract 14 / allergen content in control product extract 24) x 100

[0072] In this manner, when the allergen inactivating substance is an airborne substance such as nano-sized charged water particles, the method for evaluating the allergen inactivating substance can be carried out.

[0073] The present invention is not limited to the above-described embodiments, and various combinations and modifications within the scope of the disclosure in this specification are possible, and such embodiments are also included in the technical scope of the present invention. [Example]

[0074] The method of the present invention will now be described more specifically with reference to examples. [Creating allergen samples] Indoor dust from an ordinary household was collected by vacuuming, and the indoor dust was passed through a sieve with 0.3 mm openings to collect fine dust, which was used as an allergen sample.

[0075] [Test Example 1. Comparison of allergen protein extraction amount depending on the type of extraction solvent] (A) 20 mg of allergen sample was placed in a test tube, 1 mL of 0.02 M PBS-T (phosphate buffer solution) was added, and the mixture was stirred using a rotator. The tube was then sealed and left at room temperature for 30 minutes. The tube was then cooled to 4°C and centrifuged at 8000 rpm for 10 minutes. The supernatant obtained after centrifugation was removed with a pipette and designated as Supernatant A. (a) 20 mg of allergen sample was placed in a test tube, 1 mL of distilled water was added, and the mixture was stirred using a rotator. The tube was then sealed and left at room temperature for 30 minutes. The tube was then cooled to 4°C and centrifuged at 8000 rpm for 10 minutes. The supernatant obtained after centrifugation was removed with a pipette and designated as Supernatant B. (c) The allergen activity of supernatants A and B was measured by ELISA, and the amount of allergen (ng) contained in each supernatant was calculated. A mite allergen (Der f 1) ELISA kit (product number 10205, ITEA) was used for the measurement. The results are shown in Table 1.

[0076] [Table 1]

[0077] As shown in Table 1, it was found that supernatant A, which used a buffer solution as the extraction solvent, contained more allergens than supernatant B, which used distilled water as the extraction solvent. Therefore, it was found that using a buffer solution as the extraction solvent allows for the extraction of more allergens, making it useful for evaluating inactivation ability.

[0078] [Test Example 2. Allergen inactivation test of hypochlorous acid aqueous solution] The test was carried out as follows, following the test procedure explained in Figure 2. (A) 20 mg of the allergen sample was placed in a test tube, and 15 mL of hypochlorous acid solution was added. The mixture was then stirred using a rotator and allowed to stand in a sealed state at 30°C for 30 minutes. The test tube was then cooled to 4°C and centrifuged at 8000 rpm for 10 minutes. The supernatant obtained after centrifugation was removed with a pipette and designated as Supernatant A. This Supernatant A corresponds to Supernatant 12 in Figure 2. (a) 15 mL of 0.02 M PBS-T was added to the residue from which supernatant A was removed, and the mixture was stirred using a rotator. The test tube was then sealed and left at room temperature for 60 minutes. The test tube was then cooled to 4°C and centrifuged at 8000 rpm for 10 minutes. The supernatant obtained after centrifugation was removed with a pipette and designated as supernatant B. This supernatant B corresponds to test article extract 14 in Figure 2. (c) 20 mg of the allergen sample was placed in a test tube, and 15 mL of distilled water was added. The mixture was then stirred using a rotator and allowed to stand at 30°C for 30 minutes in a sealed state. The test tube was then cooled to 4°C and centrifuged at 8000 rpm for 10 minutes. The supernatant obtained after centrifugation was removed with a pipette and designated as Supernatant C. This Supernatant C corresponds to Supernatant 22 in Figure 2. (d) 15 mL of 0.02 M PBS-T was added to the residue from which supernatant C was removed, and the mixture was stirred using a rotator. The test tube was then sealed and left at room temperature for 60 minutes. The test tube was then cooled to 4°C and centrifuged at 8000 rpm for 10 minutes. The supernatant obtained after centrifugation was removed with a pipette and designated as supernatant D. This supernatant D corresponds to control extract 24 in Figure 2. (e) The allergen activity of each of the supernatants A to D was measured by ELISA in the same manner as in Test Example 1, and the amount of allergen (ng) contained in each supernatant was calculated. The results are shown in Table 2.

[0079] [Table 2]

[0080] Supernatants A and B are solutions derived from the test product, and Supernatants C and D are solutions derived from the control product. Therefore, the inactivation rate was calculated from the allergen amount in each supernatant shown in Table 2: inactivation rate = 100 - (supernatant A + supernatant B / supernatant C + supernatant D) x 100 = 100 - (1.79 + 1.76 / 73.8 + 16.7) x 100 = 96.1%. Since the inactivation rate was 95% or higher, it was confirmed that the hypochlorous acid aqueous solution has the ability to inactivate allergens. Therefore, it was found that the kit of the present invention, which includes an allergen sample prepared from household waste and an extraction solvent consisting of a buffer solution, can be used as an allergen inactivation evaluation kit. [Explanation of symbols]

[0081] 1 Allergen sample 2 Allergen inactivating substances 3. Extraction solvent 11 Test item 14 Test product extract 21 Control 24 Control extract

Claims

1. An allergen inactivation evaluation kit comprising an allergen sample (excluding cases where the allergen sample is pollen) prepared from household waste and an extraction solvent consisting of a buffer solution, wherein the allergen sample is selected from those with an addition recovery rate of 90 to 110% after undergoing an addition recovery test in which a predetermined amount of the target allergen is added to check whether measurement interfering substances have any effect on ELISA measurement.

2. 2. The allergen inactivation evaluation kit according to claim 1, wherein the allergen sample is made of fine dust obtained by sieving household waste.

3. The allergen inactivation evaluation kit according to claim 1, wherein the allergen contained in the allergen sample is one or more of the following selected from the group consisting of dust mites, animal hair, animal excrement, dandruff, mold, food residue, and substances derived from sanitary animals.

4. The allergen inactivation evaluation kit according to claim 1 , wherein the extraction solvent is one selected from the group consisting of a phosphate buffer solution, a carbonate buffer solution, and an acetate buffer solution.

5. 4. The allergen inactivation evaluation kit according to claim 3, wherein the extraction solvent is a phosphate buffer solution when the allergen is a mite, and an acetate buffer solution when the allergen contains collagen.

6. Using the allergen inactivation evaluation kit according to any one of claims 1 to 5, a reaction step of reacting the allergen sample with an allergen inactivating substance to obtain a test sample; After the reaction step, a test sample extraction step of obtaining a test sample extract by adding the extraction solvent; a control product extraction step in which the allergen sample is treated with the allergen inactivating substance to prepare a control product, and the control product extract is obtained by adding the extraction solvent; an allergen content measuring step of measuring the allergen contents of the solution derived from the test product containing the test product extract and the solution derived from the control product containing the control product extract by ELISA, respectively; A method for evaluating an allergen inactivating substance, characterized by including an allergen inactivation rate calculation step for calculating the allergen inactivation rate from the allergen content of a solution derived from the test product and the allergen content of a solution derived from the control product.

7. The test sample extraction step further includes a step of adding the extraction solvent to the residue from which the test sample extract has been collected, and then collecting the supernatant again to obtain a test sample supernatant; The control product extraction step further includes a step of adding the extraction solvent to the residue from which the control product extract has been collected, and collecting the supernatant again to obtain a control product supernatant; the solution derived from the test article comprises the test article supernatant; The method for evaluating an allergen inactivating substance according to claim 6, wherein the solution derived from the control product contains the supernatant of the control product.

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