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 an allergen inactivating substance. [Background technology]

[0002] In recent years, many allergies, including allergic rhinitis, atopic dermatitis, and bronchial asthma, have become a problem. The types of allergens that cause allergies include food, drugs, indoor garbage and dust (house dust = pet hair, mites, etc.), and pollen. When a person suffering from an allergy comes into contact with an allergen, an allergic reaction is induced, resulting in hives, dermatitis, asthma, fever, and other symptoms.

[0003] In particular, mites inhabit bedding such as futons and flooring 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 against allergic diseases and other allergic symptoms, attempts have been made to develop drugs that inactivate the causative allergens.

[0005] For example, the following Patent Document 1 discloses an invention of a method for removing allergens from the environment, characterized by treating the environment with tannic acid. It also describes that when house dust containing Dermatophagoides pteronyssinus, or extracts from the grass Phleum pratense or the weed Plantago lanceolata are treated with a 1% aqueous solution of tannic acid, allergenicity in a skin reaction test can be reduced.

[0006] In addition, the following Patent Document 2 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] On the other hand, the effectiveness of various substances other than the above-mentioned tannic acid has been suggested. For example, the following Patent Document 3 describes the use of a compound or its salt having at least one guanidino group as a cationic group in the molecule and having surface activity. The following Patent Document 4 describes the use of a pomegranate leaf extract using a solvent consisting of at least one selected from water and hydrophilic solvents. The following Patent Document 5 describes the use of anti-allergenic metal components such as silver and zinc. The following Patent Document 6 describes the use of a polymer compound having a styrene sulfonic acid unit. [Prior art documents] [Patent documents]

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

[0009] In general, in the development of allergen inactivating agents, in order to evaluate a drug that is expected to have an inactivating ability, it is reacted with an allergen and the allergen content in the solution after the reaction is measured by ELISA, etc. However, when food residue, animal hair, dead mites, or pollen are used as allergen samples to be reacted with an allergen inactivating agent, immunogenic allergen substances remain inside the allergen samples, making it impossible to accurately measure the allergen content, and as a result, there is a problem that the inactivating ability 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 an allergen inactivating substance, which enable more accurate evaluation of the inactivating ability of an allergen inactivating agent. [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 made of a buffer solution.

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

[0014] In addition, the allergen contained in the allergen sample is preferably one or more types selected from dust mites, pollen, animal hair, animal excrement, dandruff, mold, food residues, and substances derived from sanitary animals.

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

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

[0017] Furthermore, it is preferable that the allergen sample is 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 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 of reacting an allergen sample with an allergen inactivating substance to obtain a test product using the allergen inactivation evaluation kit described above; a test product extraction step of obtaining a test product extract by adding the extraction solvent after the reaction step; a control product extraction step of preparing a control product without reacting the allergen inactivating substance with the allergen sample and obtaining a control product extract by adding the extraction solvent; an allergen content measurement step of measuring the allergen content 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, respectively; and an allergen inactivation rate calculation step of calculating an 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 process further comprises a step of adding the extraction solvent to the residue from which the test product extract was collected and collecting the supernatant again to obtain a test product supernatant, and that the control product extraction process further comprises a step of adding the extraction solvent to the residue from which the control product extract was 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. Effect 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 an allergen inactivating substance, which enable evaluation with higher accuracy. [Brief description of the drawings]

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

[0022] The present invention provides an evaluation kit and an evaluation method for evaluating the degree of ability of any allergen inactivating substance to inactivate an allergen.

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

[0024] Moreover, the term "inactivation ability" refers to the ability to inactivate an allergen that is known to cause allergies in humans or animals.

[0025] The allergen inactivation evaluation kit of the present invention includes 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 the inactivation ability of an allergen inactivating substance to be evaluated more accurately.

[0026] The allergen sample is prepared from household waste, typically from indoor dust in a general household. In this case, it is preferable that the household waste is sieved to obtain fine dust. By sieving the fine dust, large dust particles contained in the household waste can be removed, and an allergen sample having 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 such as dead mites that can be allergens.

[0028] The allergen sample may be fine dust obtained by sieving household waste, crushing the fine dust using a known method such as beads, and then sieving the fine dust. The method for obtaining fine dust is to sieve household waste with a sieve having a mesh size of 0.3 mm to extract fine dust, crush the dust, and then sieve the fine dust with a sieve having a mesh size of 0.18 mm.

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

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

[0031] Examples of the above mites include Dermatophagoides pteronyssinus, Dermatophagoides farinae, Acaridae, Tyrophagus putrescentiae, and Dermoptera, and it is preferable that the mites are Dermatophagoides pteronyssinus or Dermatophagoides farinae.

[0032] Examples of the above-mentioned pollen include pollen of plants of the families Cupressaceae, Cupressaceae, Betulaceae, Poaceae, and Asteraceae, and pollen of plants of the families Cupressaceae or Cupressaceae is preferable.

[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 molds include Alternaria (soy mold), Aspergillus (aspergillus), Penicillium (blue mold), and Cladosporium (black mold).

[0035] Examples of foods from which the above 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 this case, the above foods may be processed into powder form, such as wheat flour, buckwheat flour, milk powder, soybean powder, etc.

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

[0037] Examples of methods for appropriately incorporating an appropriate allergen 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 mites and mold, collecting household waste in early spring when there is a lot of pollen in the air, collecting household waste from a home where pets are kept, etc. Also, household waste collected from multiple locations with different characteristics may be mixed.

[0038] Examples of methods for collecting household waste include a method of sucking up indoor dust present in a room of an ordinary household using a vacuum cleaner, a method of wiping it off using a wiper sheet, etc. Furthermore, the allergen sample product may further contain, in addition to the household waste, cultures obtained by culturing mites, molds, and sanitary animals, pollen directly collected from plants, and commercially available allergen extracts.

[0039] Whether the allergens are contained in the 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 amount of allergen contained in the sample can be measured based on the measured allergen activity.

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

[0042] That is, in general, in ELISA measurement, 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 interference substances. When the measurement interference 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 specified amount of the target allergen is added to one (hereinafter referred to as spiked sample). Nothing is added to the other (hereinafter referred to as additive-free sample), and the amount of allergen in both samples is measured by ELISA. The amount of allergen in the additive-free 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 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 according to the calculated recovery rate, the ELISA measurement can be performed with higher accuracy. The allergen sample in 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 allergen samples. By adding the extraction solvent to the allergen sample, the allergens are extracted from the allergen sample, so that the content of the allergens can be measured more accurately and the allergen inactivation ability of the allergen inactivating substance can be evaluated more accurately.

[0046] Examples of the buffer solution used in the extraction solvent include phosphate buffer, carbonate buffer, acetate buffer, citrate buffer, citrate phosphate buffer, borate buffer, tartrate buffer, Tris buffer, etc., and among them, one selected from phosphate buffer, carbonate buffer, and acetate buffer is preferable. By using one 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 mite, it is preferable that the extraction solvent is a phosphate buffer solution; when the allergen is pollen, it is preferable that the extraction solvent is a carbonate buffer solution; and when the allergen contains collagen, it is preferable that the extraction solvent is an acetate buffer solution.

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

[0049] Allergens are more easily extracted by using a phosphate buffer solution for dust mites, a carbonate buffer solution for pollen, and an acetate buffer solution for collagen-containing substances.

[0050] The allergen inactivation evaluation kit of the present invention may contain other elements other than the above allergen sample and the above extraction solvent as minimum components, such as an ELISA kit used in ELISA measurement, a test container 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, etc., and may also contain elements not described in this specification.

[0051] Next, the method for evaluating the allergen inactivating substance according to the present invention will be described. As shown in Fig. 1, the method for evaluating the allergen inactivating substance according to the present invention includes a reaction step of reacting the allergen sample with the allergen inactivating substance to obtain a test product using the allergen inactivation evaluation kit, a test product extraction step of obtaining a test product extract by adding an extraction solvent after the reaction step, a control product extraction step of obtaining a control product extract by adding an extraction solvent to the allergen sample without acting the allergen inactivating substance on it, an allergen content measurement step of measuring the allergen content 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, and an allergen inactivation rate calculation step of calculating 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.

[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 process) An allergen inactivating substance is added to an allergen sample to obtain a test sample. The method of adding an allergen inactivating substance to an allergen sample is not particularly limited, and for example, when the allergen inactivating substance is used in a solution state, the allergen sample can be placed in a test container such as a test tube or a petri dish, and then the allergen inactivating substance can be added, or the allergen sample can be uniformly sprayed and attached to a felt fabric, and then the allergen inactivating substance can be sprayed. When the allergen inactivating substance is an airborne substance such as nano-sized charged fine water particles, the allergen sample can be placed in a petri dish after antistatic treatment, and the petri dish can be further vibrated to uniformly apply the allergen sample, and then the allergen inactivating substance can be sprayed on the petri dish.

[0054] After adding the allergen inactivator to the allergen sample, a step of allowing the allergen inactivator to stand for a certain period of time to promote the reaction of the allergen inactivator 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 the protein denatures, 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. When the reaction time is set long, the moisture in the test sample may be reduced by volatilization or evaporation, which may cause a change in the allergen concentration in the test sample, so it is preferable to seal the test container.

[0055] (Test sample extraction process) An extraction solvent is added to the test sample obtained in the above reaction step to extract the allergen from the test sample, thereby obtaining a test sample extract. The test sample extract is preferably a supernatant obtained by adding an 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 the supernatant obtained by centrifuging the test sample and recovering it with a pipette or the like, or the extraction solvent may be added without recovering the supernatant. When recovering the supernatant, the extraction solvent may be added to the residue from which the supernatant has been recovered, followed by stirring, leaving it to stand, and 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 and recovering and centrifuging the solution.

[0057] The amount of the extraction solvent to be added is not particularly limited, but is preferably 10 to 1000 times, more preferably 50 to 750 times, the mass of the allergen sample. After the extraction solvent is added, the mixture may be stirred using a stirrer such as a rotator and left for a certain period of time to promote the reaction of the allergen inactivating substance and the extraction of the allergen substance. The temperature during the time of leaving the mixture may be adjusted in the same manner as in the reaction step, and it is preferable to seal the test container. In addition, when obtaining a test sample extract, the reaction of the allergen inactivating substance may be stopped by leaving the mixture for a certain period of time and then cooling the mixture.

[0058] The test sample extraction step preferably further comprises a step of adding an extraction solvent to the residue from which the test sample extract has been collected, and 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, leaving the mixture to stand, and centrifuging the mixture in the same manner. By further comprising a step of collecting the supernatant again, more allergen substances can be extracted, and the inactivation ability of the allergen inactivating substance can be evaluated more accurately.

[0059] (Control product extraction process) The allergen sample is used as a control without reacting with the allergen inactivating substance, and the allergen substance is extracted from the allergen sample in the same manner as in the above test sample extraction step to obtain a control sample extract. The control may be obtained by adding distilled water or an extraction solvent to the allergen sample in the same amount as the allergen inactivating substance added in the above reaction step in the same manner as in the above reaction step. When adding an extraction solvent to the control, the control may be centrifuged to recover the supernatant with a pipette or the like, and then the extraction solvent may be added, or the extraction solvent may be added without recovering the supernatant. The method for recovering the supernatant may be the same as in the above test sample extraction step.

[0060] The test product extraction step preferably further comprises a step of adding an extraction solvent to the residue from which the control product extract is collected, and collecting the supernatant again to obtain the 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 more allergen substances, and to more accurately evaluate the inactivation ability of the allergen inactivating substance.

[0061] (Allergen content measurement process) 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 are collected, and the allergen activity is evaluated by the ELISA method. 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 when the test product is centrifuged to collect the supernatant in the test product extraction step, the solution derived from the control product contains the control product extract, and when the control product is centrifuged to collect the supernatant in the control product extraction step, the solution derived from the control product contains the control product extract. Furthermore, the solution derived from the test product and the solution derived from the control product preferably contain the supernatant obtained by adding an extraction solvent to the residue from which the test product extract or the control product extract is collected. In the evaluation by the ELISA method, a commercially available ELISA kit can be used depending on the target allergen. The test method may be according to a known method.

[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] The method for determining whether or not an allergen inactivating substance has the inactivation ability is to determine that the allergen inactivating substance has the inactivation ability when the inactivation rate is 95% or more, and to determine that the allergen inactivating substance does not have the inactivation ability when the inactivation rate is less than 95%. The threshold value of the inactivation rate for confirming the inactivation ability may be 90% or more, or 80% or more. The threshold value of the inactivation rate for denying the inactivation ability may be 50% or less, or 25% or less.

[0064] An example of the test procedure for the evaluation method of an allergen inactivating substance, 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 is collected, and the mixture is stirred and allowed to stand. The mixture is further centrifuged to collect the supernatant, test sample extract 14. In addition, 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 is collected, stirred, allowed to stand, and further centrifuged to collect the supernatant, control sample extract 24.

[0067] In an allergen content measuring step, the allergen contents of the supernatant 12, the test product extract 14, the supernatant 22, and the control product extract 24 are each measured by ELISA. Here, the supernatant 12 and the test product extract 14 are solutions derived from the test product 11, and the supernatant 22 and the control product extract 24 are solutions derived from the control product 21. Next, in an allergen inactivation rate calculating step, the inactivation rate is calculated from the allergen contents of the supernatant 12, the test product extract 14, the supernatant 22, and the control product extract 24 by 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] A case in which the allergen inactivating substance is an airborne substance such as nano-sized electrostatically charged water particles will be described with reference to FIG.

[0070] After two petri dishes are subjected to antistatic treatment, allergen sample 1 is placed in them, and the dishes are further vibrated to evenly coat allergen sample 1. In a 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 a 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 to recover the supernatant, test sample extract 14. In a 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 to recover the supernatant, control sample extract 24.

[0071] In an allergen content measuring 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 an allergen inactivation rate calculating step, the inactivation rate is calculated from the allergen contents of the test product extract 14 and the control product extract 24 by the following formula. Inactivation rate (%) = 100 - (allergen content in test extract 14 / allergen content in control extract 24) x 100

[0072] In the manner described above, 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. EXAMPLES

[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 and left to stand at room temperature 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 by centrifugation was removed with a pipette and designated as Supernatant A. (a) 20 mg of the allergen sample was placed in a test tube, 1 mL of distilled water was added, and the mixture was stirred using a rotator and left to stand at room temperature 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 by centrifugation was removed with a pipette and designated as Supernatant B. (c) The allergen activity of the supernatants A and B was measured by ELISA, and the amount of allergen (ng) contained in each supernatant was calculated. For the measurement, a mite allergen (Der f 1) ELISA kit (product No. 10205, ITEA) was used. 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 more allergen substances to be extracted, which is 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 FIG. (A) 20 mg of the allergen sample was placed in a test tube, and 15 mL of an aqueous solution of hypochlorous acid was added. The mixture was then stirred using a rotator and the test tube was sealed and left to stand 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 by centrifugation was removed with a pipette and designated as Supernatant A. This Supernatant A corresponds to Supernatant 12 in FIG. 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 by centrifugation was removed with a pipette and designated as supernatant B. This supernatant B corresponds to the test product extract 14 in FIG. 2. (c) 20 mg of the allergen sample was placed in a test tube, 15 mL of distilled water was added, and the mixture was stirred using a rotator and allowed to stand for 30 minutes at 30°C in a sealed state. The test tube was then cooled to 4°C and centrifuged at 8000 rpm for 10 minutes. The supernatant obtained by centrifugation was removed with a pipette and designated as Supernatant C. This Supernatant C corresponds to Supernatant 22 in FIG. 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 by centrifugation was removed with a pipette and designated as supernatant D. This supernatant D corresponds to the control extract 24 in FIG. 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 of the supernatants was calculated. The results are shown in Table 2.

[0079] [Table 2]

[0080] The supernatants A and B are solutions derived from the test product, and the supernatants C and D are solutions derived from the control product. Therefore, the inactivation rate was calculated from the amount of allergen in each supernatant shown in Table 2, and the inactivation rate was 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 more, it was confirmed that the aqueous hypochlorous acid 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 made of a buffer solution, can be used as an allergen inactivation evaluation kit. [Explanation of symbols]

[0081] 1 Allergen sample 2 Allergen inactivation substances 3. Extraction Solvent 11 Test items 14 Test sample extract 21 Control 24 Control extract

Claims

1. An allergen inactivation evaluation kit comprising an allergen sample prepared from household waste and an extraction solvent consisting of a buffer solution.

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

3. The allergen inactivation evaluation kit according to claim 1, wherein the allergen contained in the allergen sample is one or more types selected from dust mites, pollen, animal hair, animal excrement, dandruff, mold, food residues, 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, a carbonate buffer, and an acetate buffer.

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

6. The allergen inactivation evaluation kit according to claim 1, wherein the allergen sample is selected through an additive recovery test in which a predetermined amount of a target allergen is added to confirm whether measurement interfering substances do not affect the ELISA measurement.

7. Using the allergen inactivation evaluation kit according to any one of claims 1 to 6, a reaction step of reacting the allergen sample with an allergen inactivating substance to obtain a test sample; A test sample extraction step of obtaining a test sample extract by adding the extraction solvent after the reaction step; a control product extraction step of extracting the allergen sample without treating it with the allergen inactivating substance, and extracting the control product by adding the extraction solvent to the allergen sample; an allergen content measuring step of measuring the allergen content 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; A method for evaluating an allergen inactivating substance, comprising: an allergen inactivation rate calculation step of calculating an 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.

8. The test sample extraction step further includes a step of adding the extraction solvent to the residue from which the test sample extract is collected, and 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 is 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 7 , wherein the solution derived from the control product contains the supernatant of the control product.

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