JP2020056590A - Immunochromatographic diagnostic kit - Google Patents

Abstract

Kind Code: A1 An immunochromatography diagnostic kit is provided which is excellent in releasability and reproducibility of test results. The conjugation is made of a thermoplastic long fiber nonwoven fabric having an average fiber diameter of 10 to 80 μm, a thermocompression bonding area ratio of 1 to 50%, and a basis weight of 30 to 400 g/m 2 . Immunochromatographic diagnostic kit including gate pads. [Selection figure] None

Description

  The present invention relates to an immunochromatographic diagnostic kit. More specifically, the present invention relates to a lateral flow type immunochromatography diagnostic kit including a specific conjugate pad, which has excellent releasability and excellent reproducibility of test results.

  In recent years, simple test reagents and diagnostics that perform various tests in a short time, such as the presence of pathogen infection such as viruses and bacteria, the presence of pregnancy, the presence of cancer markers, the presence of harmful substances such as specific raw materials in food and the presence of residual pesticides, etc. Drugs and diagnostic kits are being developed. These use a specific reaction by each test substance and a substance that specifically reacts with the test substance. In particular, immunoassays using an antigen-antibody reaction between an antigen and an antibody include immunochromatography, turbidimetric immunoassay, enzyme immunoassay, chemiluminescence assay, radioimmunoassay, and assay using surface plasmon resonance. Many measurement methods have been developed. These measurement methods are used for examinations such as illness in hospitals and clinics, and for food examinations in food companies and the like. Above all, the immunochromatographic measurement method has been carried out in a great number of tests because it requires no special equipment, equipment and knowledge, is easy to operate, is inexpensive, and can be rapidly diagnosed. In recent years, pregnancy test drugs and HIV test drugs have been marketed at general pharmacies and can be measured by general consumers. In addition to qualitative tests to check for the presence of test substances, quantitative tests to measure the amount are also available. It is becoming possible.

The measurement principle of the immunochromatography method includes a method called a sandwich method and a method called a competition method. As a measurement format, there is a method called a flow-through type or a lateral flow type. Various substances can be detected as the test substance in the sample. A typical example is a measurement for detecting an antigen by a sandwich method, and the following operations are sequentially performed.
(1) An antibody that specifically binds to an antigen to be tested is immobilized on a predetermined portion of a chromatographic medium such as a nitrocellulose membrane, and a test line (hereinafter referred to as “TL”) is provided at an arbitrary position on the chromatographic medium. ) To form a reaction site.
(2) Prepare a detection reagent in which an antibody that specifically binds to a test substance is supported on a labeling substance such as an enzyme, a labeling substance, a fluorescent labeling substance, or magnetic particles, and apply the detection reagent to a conjugate pad or the like and dry it. Then, a detection reagent-containing part is formed, and an immunochromatography diagnostic kit is formed in combination with the above-mentioned chromatographic medium.
(3) An antigen-containing sample itself or a solution obtained by diluting the same with an arbitrary liquid is dropped at a predetermined position of the immunochromatography diagnostic kit, for example, on a sample pad, and the antigen and the detection reagent are developed on a chromatographic medium. .

  By these operations, the labeling substance is captured via the antigen by the antibody immobilized on the chromatographic medium at the reaction site, and the signal of the labeling substance is detected, thereby performing diagnosis using an immunochromatography diagnostic kit. A general diagnosis is a qualitative diagnosis in which only the presence or absence of an antigen is tested, but in recent years, it has become possible to perform a quantitative diagnosis by visually or mechanically detecting the intensity of the signal. In particular, in the quantitative diagnosis, since the amount of the antigen in the test substance must be accurately detected, reproducibility of the test (value) in immunochromatographic measurement is strongly required.

Means for achieving this include, for example, a method using particles having a small particle size distribution as a labeling substance, and a method using a housing (casing) of a highly accurate immunochromatography diagnostic kit.
As another means, Patent Literature 1 below discloses that the reproducibility of inspection is significantly improved by using a nonwoven fabric made of regenerated cellulose fiber for a sample pad. However, depending on the type of specimen such as blood, a sample pad made of cellulose is often unfavorable, and in order to improve the reproducibility of the test, a method that can be used regardless of the type of the diagnostic system or the specimen is desired. .
On the other hand, in many immunochromatography diagnostic kits, as described above, a labeling substance is applied to a conjugate pad and dried, but the causal relationship between the conjugate pad and the reproducibility of the test has not been particularly described so far. For example, Patent Document 2 below reports that the use of a synthetic fiber nonwoven fabric for a conjugate pad can reduce the inspection time, but does not mention reproducibility at all.

International Publication No. WO 2014/199954 JP 2010-256309 A

  In view of the state of the prior art, the problem to be solved by the present invention is to provide an immunochromatography diagnostic kit which is excellent in releasability and reproducibility of test results.

  The present inventors have conducted intensive studies and repeated experiments to solve the above problems, and as a result, by using a thermoplastic long-fiber nonwoven fabric having a specific thermocompression bonding area ratio as a conjugate pad, the release property is improved. The present inventors unexpectedly found that the reproducibility of the test results was excellent, and completed the present invention.

That is, the present invention is as follows.
[1] A conjugate made of a thermoplastic long-fiber nonwoven fabric composed of thermoplastic long fibers having an average fiber diameter of 10 to 80 μm, having a thermocompression bonding area ratio of 1 to 50%, and a basis weight of 30 to 400 g / m ^2. Immunochromatographic diagnostic kit including gate pad.
[2] The immunochromatographic diagnostic kit according to the above [1], wherein the birefringence of the thermoplastic long fibers constituting the nonwoven fabric is 0.005 to 0.100.
[3] The immunochromatographic diagnostic kit according to [1] or [2], wherein the nonwoven fabric has been previously treated with a surfactant.
[4] The immunochromatographic diagnostic kit according to [3], wherein the water absorption capacity of the nonwoven fabric is 20 to 400%.

  The immunochromatographic diagnostic kit according to the present invention has excellent release properties, excellent reproducibility of test results, and improved speed of diagnosis by using a specific thermoplastic long-fiber nonwoven fabric as a conjugate pad. is there.

1 is a perspective view of an immunochromatography diagnostic kit as one embodiment of the present invention. It is sectional drawing which shows the state where the adhesion surface of a conjugate pad is uneven. It is sectional drawing which shows the state where the adhesion surface of a conjugate pad is uniform. It is a photograph replacing a drawing explaining the phenomenon of "streaks".

Hereinafter, embodiments of the present invention will be described in detail.
The immunochromatography diagnostic kit of the present embodiment is made of thermoplastic long fibers having an average fiber diameter of 10 to 80 μm, and has a thermocompression bonding area ratio of 1 to 50% and a basis weight of 30 to 400 g / m ² . It is characterized by including a conjugate pad made of a long-fiber nonwoven fabric.

As disclosed in Patent Documents 1 and 2, it is a general method to use a thermoplastic synthetic fiber non-woven fabric for a conjugate pad, and also sells a polyester non-woven conjugate pad from Ahlstrom. Have been.
FIG. 1 is a perspective view of an immunochromatography diagnostic kit as one embodiment. In the embodiment shown in FIG. 1, the immunochromatographic diagnostic kit comprises (a) a sample pad, (b) a conjugate pad (including an antibody sensitizing label), (c) a test line (TL), and (d) a control line. , (E) a nitrocellulose membrane, (f) an absorbent pad, and (g) a mount. However, as shown in FIG. 2, in general, unevenness is present on the surface of the nonwoven fabric (b) used as a conjugate pad, so that the bonding surface with the nitrocellulose membrane (e) becomes uneven. However, there is a problem that the development of the labeling substance is not stable. The present inventors, in order to solve such a problem, as shown in FIG. 3, the fibers constituting the non-woven fabric used as a conjugate pad are thermocompression-bonded with an embossing roll, thereby bonding the fibers to each other. By making the surface flat, the adhesion surface with the nitrocellulose membrane was made uniform. On the other hand, in general, the conjugate pad emphasizes the releasability, so that it is desired to create voids as much as possible as a fiber structure in the nonwoven fabric. Under these circumstances, the present inventors have conducted intensive studies and repeated experiments, and as a result, by applying appropriate thermocompression bonding to the nonwoven fabric used as the conjugate pad, reproduction of the test results of the immunochromatography diagnostic kit using this It was unexpectedly found that there was a great effect on the characteristics, and based on this, the invention was completed.
Patent Documents 1 and 2 have no description, teaching, or suggestion about such a problem or a technology capable of solving the problem.

  The thermoplastic long-fiber nonwoven fabric constituting the conjugate pad used in the immunochromatography diagnostic kit of the present embodiment can be, for example, a long-fiber nonwoven fabric obtained by a spun-pound method or the like. The fibers constituting the nonwoven fabric may be thermoplastic long fibers, for example, polyester fibers such as polyethylene terephthalate, polybutylene terephthalate, copolymerized polyester, low-density polyethylene, high-density polyethylene, polypropylene, copolymerized polyethylene, copolymerized polyethylene, and the like. Examples include synthetic fibers such as polyolefin fibers such as polymerized polypropylene, polyamide fibers such as nylon 6, nylon 66, and copolyamide. These fibers may be used alone or as a composite fiber of two or more kinds, or a low-melting fiber and a high-melting fiber may be mixed. The fibers constituting the nonwoven fabric are preferably polyester fibers or polypropylene fibers from the viewpoint of the release property of the labeling substance.

It is important that the thermocompression bonding area ratio of the thermoplastic long-fiber nonwoven fabric used in the present embodiment is 1 to 50%. The thermocompression bonding here is not particularly limited as long as it is a method in which nonwoven fabric yarns are thermally pressed. The thermoplastic long-fiber nonwoven fabric may be embossed, but may be point-bonded on the surface of the nonwoven fabric fiber. The method of point crimping is not particularly limited, but preferably includes a method using a pair of embossing rolls having an uneven pattern on at least one surface, and a method using a pair of flat rolls having a flat surface. Further, a process such as a needle punching process, a spunlace process, and a felt calendering process may be performed. In the case of performing thermocompression bonding, it is important to perform thermocompression bonding at a thermocompression bonding area ratio of 1 to 50% with respect to the entire area of the nonwoven fabric. The effect of flattening is not obtained, and during immunochromatography, a difference occurs in the flow of the labeling substance between the diagnostic kits, and the reproducibility of the test deteriorates. From the above points, the thermocompression bonding area ratio to the entire area of the nonwoven fabric is preferably 3% or more, more preferably 5% or more. On the other hand, if the thermocompression bonding area ratio exceeds 50%, the structure of the nonwoven fabric becomes crushed, the release property of the labeling substance is rapidly deteriorated, and a flow like a streak occurs when the immunochromatography is developed. As a result, the reproducibility of the inspection deteriorates. From the above points, the thermocompression bonding area ratio to the entire area of the nonwoven fabric is preferably 48% or less, more preferably 45% or less, and further preferably 40% or less.
The “streak” here is a phenomenon as shown in FIG. In other words, when the labeling substance is released from the conjugate pad during immunochromatographic development, if thermocompression is performed too much, "streaks" will occur and the determination line will be uneven, which will rapidly deteriorate the reproducibility of the test. The labeling substance is applied to the conjugate pad and dried. When the labeling substance is dried, if there is an excessive thermocompression bonding portion, a clear line or the like is generated in the internal structure of the nonwoven fabric. Therefore, it is important to control the thermocompression bonding area ratio.

  In the present specification, the term “thermocompression bonding” means that the fibers constituting the nonwoven fabric are not fused by heat by processing at the time of thermocompression bonding, are compressed at a density of 1.05 times or more higher than the base fiber, and are visually observed. Means a state where a difference from the substrate is confirmed. As means for thermocompression bonding, for example, a thermoplastic long-fiber nonwoven fabric is passed between a pair of heating rolls composed of an embossing roll and a flat roll having an uneven surface structure to form a thermocompression bonding portion uniformly dispersed throughout the nonwoven fabric. There is a means for causing the above, but the present invention is not limited to this. As a method of adjusting the area ratio of the thermocompression bonding, for example, there is a means of forming the thermocompression bonding portion using the embossing rolls having different area ratios of the concave and convex portions.

The average fiber diameter of the thermoplastic long-fiber nonwoven fabric used in the present embodiment is 10 to 80 μm. If the average fiber diameter is less than 10 μm, the fibers are too thin, the voids between the fibers of the nonwoven fabric become small, and the labeling substance is clogged during immunochromatographic development, resulting in poor release properties. Further, even after immunochromatographic development is completed, “particles remaining” are generated on the conjugate pad. As a result, the sensitivity is significantly reduced. From the above points, the average fiber diameter of the thermoplastic long-fiber nonwoven fabric is preferably 12 μm or more, and more preferably 15 μm or more. On the other hand, when the average fiber diameter exceeds 80 μm, the uniformity of the fibers of the thermoplastic long-fiber nonwoven fabric significantly deteriorates, and finally, during immunochromatography, a non-uniform flow of the labeling substance occurs, and the reproducibility of the test deteriorates. . From the above points, the average fiber diameter of the thermoplastic long-fiber nonwoven fabric is preferably 75 μm or less, more preferably 70 μm or less, and still more preferably 65 μm or less.
The term “particle remaining” as used herein refers to a phenomenon in which a labeling substance remains on a conjugate pad even after immunochromatographic development is completed. When this “particle remaining” occurs, the particles that should have flowed do not flow, so that the sensitivity is reduced and the reproducibility of the inspection is also deteriorated.

  It is preferable that the thermoplastic long-fiber nonwoven fabric used in the present embodiment does not contain a binder, a polymer binder, and the like. If the conjugate pad contains a polymer binder or the like, it is not preferable because, at the time of immunochromatographic development, there is a possibility that poor development, residual particles, and "false positive" may occur or influence.

  The term “false positive” as used herein means that a non-specific reaction occurs, and the specimen originally contains no antigen, that is, a positive reaction is observed despite the negative. This “false positive” is a cause of confusion in the clinical setting and is not preferred.

The basis weight of the thermoplastic long-fiber nonwoven fabric used in the present embodiment is 30 to 400 g / m ² . If the basis weight of the thermoplastic long-fiber nonwoven fabric is less than 30 g / m ² , the labeling substance cannot be sufficiently retained, and does not function as a conjugate pad for a general immunochromatographic diagnostic agent. The basis weight of the thermoplastic long-fiber nonwoven fabric is preferably 35 g / m ² or more, more preferably 40 g / m ² or more. On the other hand, if the basis weight of the thermoplastic long-fiber nonwoven fabric exceeds 400 g / m ² , it becomes too bulky and retains a developing solution used during immunochromatographic development, and the developing solution does not flow well. The basis weight of the thermoplastic filament nonwoven fabric, preferably 380 g / ^{m 2} or less, more preferably 350 g / ^{m 2,} more preferably not more than 320 g / ^{m 2.}

  The thermoplastic nonwoven fabric used in the present embodiment may be subjected to a hydrophilic treatment such as a graft treatment or a water-repellent treatment.

  In the thermoplastic nonwoven fabric used in the present embodiment, the fibers constituting the nonwoven fabric preferably have a birefringence (Δn) of 0.005 to 0.100. Although the reason is not clear, there is a relationship between the birefringence and the release property of the labeling substance. That is, when the birefringence is within this range, the release property of the labeling substance becomes excellent. If the birefringence is less than 0.005 or exceeds 0.100, the labeling substance may remain on the conjugate pad even after immunochromatographic development, and the phenomenon that the reproducibility of the test is deteriorated may occur, A phenomenon in which the sensitivity was reduced was confirmed. The lower limit of the birefringence is preferably 0.008 or more, more preferably 0.010 or more. The upper limit of the birefringence is preferably 0.095 or less, more preferably 0.090 or less. With respect to these birefringences, the birefringence can be adjusted by controlling the draft ratio when the spun yarn is drawn and thinned.

  The water absorption ratio of the thermoplastic long-fiber nonwoven fabric used in the present embodiment is preferably 20 to 400%. The “water absorption capacity” here is a value measured for a thermoplastic long-fiber nonwoven fabric pretreated with a surfactant described below. If the water absorption ratio is 20% or less, the absorption of the developing solution flowing from the sample pad during immunochromatographic development is delayed, and the release of the developing solution from the conjugate pad to the nitrocellulose membrane is also delayed. It is not preferable because time is delayed. The lower limit of the water absorption is more preferably 25% or more, and still more preferably 30% or more. On the other hand, when the water absorption ratio exceeds 400%, the release of the developing solution from the conjugate pad to the nitrocellulose membrane is delayed, which not only causes a delay in the inspection time, but also leads to a deterioration in the reproducibility of the inspection. The upper limit of the water absorption capacity is more preferably 370% or less, further preferably 350%, and still more preferably 320% or less. These water absorption ratios can be adjusted by the porosity of the thermoplastic long-fiber nonwoven fabric, the basis weight, and the area ratio of the thermocompression bonding. However, it is adjusted by the type of the surfactant to be subjected to the pretreatment and the concentration of the surfactant. Can be.

It is preferable that the thermoplastic long-fiber nonwoven fabric constituting the conjugate pad used in the present embodiment has been previously treated with a surfactant. The components of the surfactant are not particularly limited, but may be any of a cationic surfactant, an anionic surfactant, and a nonionic surfactant. The nonionic surfactant can be appropriately selected from those usually used. Triton X-100 (registered trademark), Triton X-114, (registered trademark) Brij 58 (registered trademark), Brij 35 (registered trademark) ), Tween 20®, Tween 80®, 1-On-octyl-β-D-glucopyranoside, n-octyl-β-D-thioglucopyranoside, n-dodecyl-β-D-maltopyranoside. N-dodecyl-α-D-maltopyranoside, n-dodecyl-N, N-dimethylamine-N-oxide, isopropyl-β-D-thiogalactoside, sucrose monododecanoic acid, n-octyl-β-D-glucopyranoside, n-dodecyl-β-D-maltopyranoside, n-tridecyl-β-D-maltopyranoside and the like It is below. The anionic surfactant can be appropriately selected from those usually used, and examples thereof include cholic acid, deoxycholic acid, glycolic acid, taurocholic acid, taurodeoxycholic acid, and salts thereof. Further, from the viewpoints of release properties and reproducibility of test results, surfactants having an HLB value of 10 to 20 are preferable.
The concentration of the surfactant required for the pre-treatment (pre-treatment) may be in the range of 0.001% to 10.0000%. Within such a concentration range, the labeling substance flows without any clogging of the labeling substance during immunochromatography and without the generation of residual particles on the conjugate pad. If the pretreatment with the surfactant is not performed, the labeling substance is not uniformly retained in the nonwoven fabric when the labeling substance is impregnated, which may lead to deterioration in the reproducibility of the test.

  In the present specification, the term “labeling substance” refers to a particulate substance that is insoluble in water, a buffer, and the like, and that carries a dye, a dye, and the like. Although the material constituting the particles is not particularly limited, examples of such a labeling substance include metal colloid particles such as gold colloid, platinum colloid, silver colloid, and selenium colloid, styrene latex such as polystyrene latex, and acrylic acid latex. Colored latex particles, colored silica particles composed of a three-dimensional structure composed of silicon atoms and oxygen atoms, colored silica particles colored cellulose, colored cellulose particles colored cellulose, and labeled substances obtained by directly coloring coloring components such as carbon black. , Magnetic particles, and the like. The labeling substance may be fluorescent particles.

The average particle diameter of the “labeling substance” is preferably 10 to 1000 nm.
In the present specification, the “average particle diameter” of the labeling substance refers to a volume average median diameter measured by a dynamic light scattering method. When the average particle diameter is 10 to 1000 nm, the TL becomes deeper when used as an immunochromatography diagnostic kit because the surface area of the particles is large, that is, the analysis sensitivity is increased. If the average particle diameter is too small, the surface area becomes small, which may lower the analytical sensitivity or cause aggregation of particles. From the above points, the average particle diameter of the labeling substance is more preferably 20 nm or more, and still more preferably 30 nm or more. On the other hand, if the average particle size of the labeling substance is too large, the pores of the nitrocellulose membrane will be clogged and the parts that should originally be white after the test will be colored, adversely affecting the judgment of the test result, or the detection limit will be deteriorated. There are cases. From the above points, the average particle diameter of the labeling substance is more preferably 800 nm or less, and still more preferably 600 nm or less. The “average particle size” described here is an average value, and a part of the particle size distribution may be out of the above range.

  The immunochromatographic diagnostic kit of the present embodiment is for easily detecting the presence or absence of a test substance in various samples. Examples of the type of the diagnostic kit include a lateral flow type and a flow-through type. The immunochromatographic diagnostic kit of the present embodiment is not particularly limited as long as it uses a labeling substance or a sample pad, but is preferably a lateral flow type. Among the lateral flow types, there are a dipstick type and a cassette type, but these types are not particularly limited. The configuration of the immunochromatography diagnostic kit is not particularly limited, and any configuration generally used in the art may be used. The types of members other than the labeling substance and the sample pad are not particularly limited as long as they are used in the art. For example, as shown in FIG. 1, (b) a conjugate pad (including an antibody-sensitized labeling substance), (E) nitrocellulose membrane, (f) absorbent pad, and (g) mount. Further, some of these members may be omitted as necessary.

The “sample pad” shown in FIG. 1A is a portion of the immunochromatography diagnostic kit which first receives a sample to be measured. Typical sample pads include cellulose filter paper, paper, glass fiber, glass fiber, acrylic fiber, nylon fiber, various fabrics, and the like.
As a sample pad, a nonwoven fabric made of regenerated cellulosic fiber is used as long as it does not adversely affect the desired effect and does not affect the antigen-antibody reaction or the stability of the antibody. It may contain various drugs or powders, or may be a derivative of a part of cellulose. Examples of the agent or powder to be impregnated include a surfactant, a protein, an antibody, a resin, a water-soluble polymer, an antibacterial agent, a preservative, and an antioxidant. Derivatization of cellulose includes carboxymethylation, carboxyethylation, primary amination, secondary amination, tertiary amination, quaternary amination, oxylation, and the like.

  As described above, the sample pad may be subjected to pretreatment as needed. For example, a treatment may be performed in which a buffer, a surfactant, a protein, a reagent for trapping impurities in a sample sample, a preservative, an antibacterial agent, an antioxidant, a moisture absorbent, and the like are previously contained. The shape of the sample pad is not particularly limited. For example, the length of the sample pad (the length of the flow of the liquid) is about 10 to 25 mm in consideration of the connection from the sample liquid and the diagnosis time. There is no problem if the width (perpendicular to the liquid flow) is larger than the width of the conjugate pad. If the width is too narrow, there is a possibility that the test solution will flow from the end of the sample pad. In the following examples, as the size of the sample pad, a length of 20 mm and a width of 5.0 mm were selected.

  The “absorption pad” shown in FIG. 1 (f) is a part that finally absorbs a sample to be measured in immunochromatography. Typical absorbent pads include cellulose filter paper, paper, glass fiber, glass fiber, acrylic fiber, nylon fiber, various fabrics, and the like.

  The “diagnosis method” that can be performed using the immunochromatographic diagnostic kit of the present embodiment is not particularly limited, and includes various diagnoses performed using the immunochromatographic diagnostic kit. The diagnostic object is not particularly limited, and includes various diagnostic objects such as human, animal, food, plant, and other environmental tests. In a general diagnostic procedure, a specimen sample is collected from the test object, if necessary, subjected to pretreatment such as extraction and filtration, dropped on a sample pad, waited for a predetermined time from the start of the test, and The diagnostic result is determined from different colors depending on the presence or absence. Of course, the present invention is not limited to this procedure, and the immunochromatographic diagnosis kit of the present embodiment can be used for diagnosis of the same procedure and principle. A procedure in which a sample sample is filtered in advance is preferable because extraneous foreign substances and impurities can be removed, thereby further speeding up diagnosis and improving diagnosis accuracy.

  Subjects that can be diagnosed using the immunochromatography diagnostic kit of the present embodiment are not particularly limited, but specific examples include the following: cancer markers, hormones, infectious diseases, autoimmunity, Examples include plasma proteins, TDM, coagulation / fibrinolysis, amino acids, peptides, proteins, genes, cells, and the like. More specifically, CEA, AFP, ferritin, β2 microglobulin, PSA, CA19-9, CA125, BFP, elastase 1, pepsinogen 1.2, fecal occult blood, urinary β2 microglobulin, PIVKA-2, urinary BTA , Insulin, E3, HCG, HPL, LH, HCV antigen, HBs antigen, HBs antibody, HBc antibody, HBe antigen, HBe antibody, HTLV-1 antibody, HIV antibody, toxoplasma antibody, syphilis, ASO, influenza A antigen, A Type influenza antibody, influenza type B antigen, influenza type B antibody, rota antigen, adenovirus antigen, rota adenovirus antigen, group A streptococcus, group B streptococcus, candida antigen, CD bacterium, cryptolocus antigen, cholera, medulla Neisseria meningitidis antigen, granulosa elastase, helicopter Bacter pylori antibody, O157 antibody, O157 antigen, Leptospira antibody, Aspergillus antigen, MRSA, RF, total IgE, LE test, CRP, IgG, A, M, IgD, transferrin, urinary albumin, urinary transferrin, myoglobin, C3. C4, SAA, LP (a), α1-AC, α1-M, haptoglobin, microtransferrin, APR score, FDP, D-dimer, plasminogen, AT3, α2PI, PIC, PAI-1, protein C, coagulation X3 Factor, type IV collagen, hyaluronic acid, GHbA1c, other various antigens, various antibodies, various viruses, various bacteria, various amino acids, various peptides, various proteins, various DNAs, various cells, various allergens, various pesticides, various harmful substances .

  In the immunochromatography diagnostic kit of the present embodiment, the labeling substance needs to carry a substance that specifically binds to the analyte, for example, an antibody, but the carrying method is not particularly limited. For example, loading by physical adsorption, loading by covalent bonding, loading by a combination thereof, and the like are included. The type and amount of the substance to be supported are not particularly limited. As the type of the substance to be carried, an antibody is the most common and preferable. In addition, as a method of loading, from the viewpoint of easiness, loading by physical adsorption is preferable, and from the viewpoint of stability and performance, loading by covalent bonding is preferable.

  The chromatographic medium used in the immunochromatographic diagnostic kit of the present embodiment is not particularly limited, and various commonly used chromatographic media can be used. Specifically, FIG. A nitrocellulose membrane represented by

[Method for producing thermoplastic long-fiber nonwoven fabric]
The thermoplastic long-fiber nonwoven fabric constituting the conjugate pad of the immunochromatography diagnostic kit of the present embodiment can be efficiently produced by a spunbond method. That is, the thermoplastic resin is heated and melted and discharged from a spinneret, and the obtained spun yarn is cooled using a known cooling device, and drawn and thinned by a suction device such as an air sucker. Subsequently, after the yarn group discharged from the suction device is spread, it is deposited on a conveyor to form a web. Next, a predetermined thermoplastic long-fiber nonwoven fabric can be produced by partially performing thermocompression bonding on the web formed on the conveyor using a partial thermocompression bonding device such as a heated embossing roll.

[Method for preparing immunochromatographic diagnostic kit]
Hereinafter, an example of a method for producing an immunochromatographic diagnostic kit will be described.
A dispersion of the labeling substance adjusted to a predetermined concentration is prepared, a buffer solution and an antibody are added, and the mixture is stirred for a certain period of time while adjusting the temperature to adsorb the antibody to the labeling substance. After stirring for a certain period of time, the coloring cellulose particles are blocked by further adding a blocking agent and stirring for a certain period of time while controlling the temperature. As the blocking agent, various blocking agents can be used according to the composition of the test substance, the specimen, or the solution for diluting the specimen. In order to wash the labeling substance after the antibody adsorption and blocking, centrifugation is performed to separate the supernatant containing the excess antibody and the blocking agent from the precipitated particles, and the supernatant is removed by decantation. A liquid such as a buffer solution is added to the sedimented particles, and a dispersion treatment is performed by ultrasonic waves or the like as necessary. Washing by a series of operations including sedimentation by centrifugation, removal of supernatant, and addition of liquid is performed as many times as necessary to prepare a dispersion containing particles having been subjected to antibody adsorption and blocking at a predetermined concentration. If necessary, proteins, surfactants, sugars such as sucrose and trehalose are added to the dispersion, and a certain amount of the obtained solution is applied to a thermoplastic long-fiber nonwoven fabric constituting the conjugate pad, dried, and detected. Adjust the reagent-containing part. In addition, a buffer solution, a surfactant, a protein, a reagent for trapping impurities in a specimen sample, a preservative, an antibacterial agent, an antioxidant, a hygroscopic agent, and the like are applied to the regenerated cellulose continuous filament nonwoven fabric as necessary, Allow to dry and prepare sample pad. Further, a nitrocellulose membrane chromatography medium having an antibody immobilized at a predetermined position, and an absorption pad made of cellulose filter paper for absorbing a specimen are prepared. These are immobilized on a sheet having an adhesive portion called a backing sheet, and cut into a predetermined size to produce an immunochromatographic diagnostic kit (see FIG. 1).

Hereinafter, the present invention will be specifically described with reference to Examples and Comparative Examples, but the present invention is not limited to Examples.
First, methods for measuring various physical properties and methods for measuring the results of immunochromatographic diagnosis used in Examples and the like will be described. Unless otherwise specified, all operations were performed in an environment at a temperature of 23 ° C. and a relative humidity of 55% RH.

[Thermo-compression bonding area ratio (%)]
A 1 cm square test piece was sampled and photographed with an electron microscope. The area of the thermocompression bonding portion was measured from each photograph, and the average value was defined as the area of the thermocompression bonding portion. Further, the pitch of the pattern of the thermocompression bonding portion was measured in the MD direction and the CD direction, and the ratio of the thermocompression bonding area per unit area of the nonwoven fabric was calculated as the thermocompression bonding area ratio based on these values.

[Average fiber diameter (μm)]
Using a microscope microscope (VH-8000) manufactured by KEYENCE CORPORATION, the diameter of the fiber was magnified 1000 times, and the fiber diameter was measured.

[Eye weight (g / m ² )]
After drying a nonwoven fabric having an area of 0.5 m ² or more to a constant weight at 105 ° C., it is left in a constant temperature room at 20 ° C. and 65% RH for 16 hours or more to measure its weight, and the weight per unit area of the nonwoven fabric is measured. The weight was measured.

[Birefringence (Δn)]
Using a BH2-type polarizing microscope compensator manufactured by OLYMPUS, the birefringence of the fiber immediately after drawing was determined from the retardation and the fiber diameter by a normal interference fringe method.

[Water absorption ratio (%)]
The thermoplastic long-fiber nonwoven fabric pretreated (pretreated) with a surfactant was left in a room controlled at 20 ° C. and 65% RH for 15 hours to control the humidity, and a sample cut into a 10 cm square was weighed (W1). (G)). Next, the sample was placed on a wire mesh having a wire diameter of 0.5 mm and 10 mesh, and immersed together with the wire mesh in water at 20 ° C. for 30 seconds. After that, the sample was left in the air for 10 minutes while being kept horizontal on a wire mesh to drain water, weighed again (W2 (g)), and the water absorption ratio (%) was determined by the following equation.
Water absorption ratio (%) = {(W2−W1) / W1} × 100

[Thickness (mm)]
The thickness was measured in a thickness test according to JIS-L1096 with a load of 1.96 kPa (unit: mm).

[Particles remaining in immunochromatography diagnostic kit]
Thirty minutes after the start of immunochromatographic development, it was judged as "x" when coloring was observed on the conjugate pad, and as "〇" when no coloring was seen.

[Streak of immunochromatography diagnostic kit]
At the time of immunochromatographic development, when a dark streak occurred on the nitrocellulose membrane, it was judged as "x", and when no streak was seen, it was judged as "〇".

[Diagnosis time of immunochromatographic diagnosis kit (TL coloring time)]
The immunochromatographic diagnostic kit cut into a width of 5 mm was placed in a plastic housing. The obtained diagnostic kit containing a housing was measured using an immunochromatographic reader C10066-10 manufactured by Hamamatsu Photonics. Apparatus settings were made according to the color of the particles used. Human chorionic gonadotropin (hereinafter referred to as "hCG") is used as a test substance, and hCG is a 66 mM phosphate buffer (pH 7.4) containing 1% by weight of bovine serum albumin (hereinafter referred to as "BSA"). (Hereinafter referred to as "PBS") to prepare a positive sample having an hCG concentration of 10 mIU / ml. 120 µl of this positive sample was dropped on the sample dropping part of the diagnostic kit, and thereafter, measurement was performed every 20 seconds with an immunochromatographic reader, and the change over time in TL was measured. The reason for setting every 20 seconds here is that it takes less than 20 seconds for one measurement. The time (between colors) when the color intensity (unit: mABS) of the TL obtained by the immunochromatography reader became 20 mABS or more was measured. Here, the reason why the value of 20 mABS is used is that the presence of TL can be visually confirmed when the value exceeds 20 mABS although there is an individual difference. This measurement was performed five times, and the average time was taken as the diagnosis time (TL coloring time (second)).

[Reproducibility of immunochromatographic diagnostic kit (TL intensity, TL intensity standard deviation,% CV)]
In the same manner as described above, 120 µl of the positive specimen was dropped on the sample dropping part of the diagnostic kit, and the color intensity of TL after 15 minutes was measured with an immunochromatographic reader. This measurement was performed 20 times, and the average value of the obtained values was defined as the TL intensity, and the standard deviation thereof was defined as the TL intensity standard deviation. % CV, which is an index representing reproducibility, is expressed by the following equation (1):
% CV = {TL intensity standard deviation / TL intensity} × 100 (1)
Was calculated by

[Detection limit of immunochromatography diagnostic kit]
hCG concentration 3.20 mIU / ml, 1.60 mIU / ml, 0.80 mIU / ml, 0.40 mIU / ml, 0.20 mIU / ml, 0.10 mIU / ml, 0.05 mIU / ml, 0.025 mIU / ml , And a positive sample was prepared by gradually thinning the sample. In the same manner as described above, 120 μl was dropped into the sample dropping portion of the diagnostic kit, and the color development intensity of TL after 15 minutes was measured with an immunochromatographic reader. This measurement was performed five times at each concentration, and when the average value of the obtained values was equal to or higher than the value obtained when the negative sample was measured + 20 mABS, it was determined to be positive, and in the following cases, it was considered to be below the detection limit. The lower limit of hCG concentration at which this positive determination was obtained was taken as the detection limit.

[Example 1]
[Preparation of polyester long fiber nonwoven fabric]
A polyester resin having an intrinsic viscosity (IV) of 0.8 and a melting point of 247 ° C. is supplied to a conventional melt spinning apparatus and melted at 275 ° C., and a spinning speed of 4500 m / min from a spinneret having a circular cross-section spinning hole. The polyester fiber was melt-spun at a draft ratio of 2120 to obtain a polyester filament having a fiber diameter of 25 μm. Next, the fibers are spread and dispersed by using a dispersing apparatus (inclination angle with respect to the flat filaments of 4 °) for controlling a flat air flow to produce a web having a basis weight of 120 g / m ^2. , A polyester long-fiber nonwoven fabric was obtained by partial thermocompression bonding at a thermocompression bonding area ratio of 15%. The thickness was 0.03 mm.

[Preparation of antibody-sensitized colored cellulose particles]
120 μl of colored cellulose particles (trade name: NanoAct, manufactured by Asahi Kasei Corporation, average particle diameter: 352 nm, particle concentration: 1.0%) were put into a 15 ml centrifuge tube, and further, 240 μl of Tris buffer (50 mM, pH 7.0) was added to 0.1 ml. 120 μl of 1% anti-hCG-α mouse antibody (10-C25C, manufactured by Fitzgerald) was added, and the mixture was vortexed for 10 seconds. Subsequently, it was placed in a dryer adjusted to 37 ° C. and allowed to stand for 120 minutes. Subsequently, 14.4 ml of a blocking solution (100 mM boric acid, pH 8.5) containing 1.0% by weight of casein (030-01505, manufactured by Wako Pure Chemical Industries, Ltd.) was added, and the mixture was further dried in a dryer at 37 ° C. for 60 minutes. It was left still. Subsequently, using a centrifugal separator (manufactured by Kubota Shoji Co., Ltd., 6200) and a centrifugal rotor (manufactured by Kubota Shoji Co., Ltd., AF-5008C), centrifugation was performed at 10,000 g for 15 minutes to precipitate the sensitized particles, and the supernatant was removed. Removed. Next, 14.4 ml of a borate buffer (50 mM boric acid, pH 10.0) was added, and the mixture was treated with an ultrasonic dispersing machine (UH-50, manufactured by SMT Corporation) for 10 seconds. Subsequently, centrifugation was performed at 10,000 g for 15 minutes to precipitate the sensitized particles, and then the supernatant was removed. Subsequently, 0.6 g of sucrose (manufactured by Wako Pure Chemical Industries, Ltd., 196-0015) and 0.8 g of a 1.0% by weight casein blocking solution were added, and a borate buffer (50 mM boric acid, PH 10.0) was used. The weight was adjusted to 4.0 g, and an antibody-sensitized colored cellulose particle dispersion of 0.03% by weight was prepared and treated with an ultrasonic disperser for 10 seconds.

[Impregnation and drying of conjugate pad with labeling substance]
A polyethylene conjugate pad (Pall, 6613) was immersed in a large excess of 0.10% by weight of Tween-20 (registered trademark, Sigma-Aldrich, T2700) to remove excess liquid and then at 50 ° C. Dried for 60 minutes. Subsequently, it was cut into a shape having a height of 10 mm and a length of 300 mm. Subsequently, using a micropipette, 0.02% by weight of an antibody-sensitized colored cellulose particle dispersion was uniformly applied to 1020 μl, and dried at 50 ° C. for 60 minutes.

[Pretreatment of sample pad]
A sample pad (Millipore, C048) was washed with a large excess of 2.0% by weight of BSA (Sigma-Aldrich, A7906) and 2.0% by weight of PBS buffer containing Tween-20®. (66 mM, pH 7.4), dried after removing excess liquid at 50 ° C. for 60 minutes, and then cut into a shape having a height of 18 mm and a length of 300 mm.

[Preparation of nitrocellulose membrane coated with capture antibody]
A nitrocellulose membrane (Millipore, SHF0900425) was cut into a shape having a height of 25 mm and a length of 300 mm. Using a liquid coating apparatus (Musashi Engineering Co., Ltd., 300DS), a PBS solution (66 mM, pH 7.4) containing 0.1% by weight of anti-hCG-β mouse antibody (Mixix Biochemical Co., Ltd., 6601) was added at 0.1 μl / mm. It was applied to a portion having a height of 7 mm in a ratio. Subsequently, a PBS solution (66 mM, pH 7.4) containing 0.1% by weight of an anti-mouse-rabbit antibody (Z0259, manufactured by Daco) was applied to a portion having a height of 12 mm at a rate of 0.1 μl / mm. Subsequently, it was dried at 37 ° C. for 30 minutes.

[Preparation of immunochromatographic diagnostic kit]
A nitrocellulose membrane coated with a capture antibody, an absorption pad (C083, manufactured by Millipore), a conjugate pad containing a labeling substance, and a sample pad were attached to a backing card (AD9020, manufactured by Adhesives Research). Then, it was cut to a width of 5 mm by a cutter to obtain an immunochromatographic diagnostic kit having a width of 5 mm and a height of 60 mm.

[Performance evaluation of immunochromatographic diagnostic kit]
The performance of the obtained immunochromatographic diagnostic kit was evaluated. The results are shown in Tables 1 and 2 below, together with the physical properties of the obtained nonwoven fabric and the physical properties of the labeling substance.

[Example 2]
An immunochromatography diagnostic kit was prepared in the same manner as in Example 1 except that the thermocompression bonding area ratio of the thermoplastic long-fiber nonwoven fabric was 5%, and its performance was evaluated. The results are shown in Tables 1 and 2 below, together with the physical properties of the obtained nonwoven fabric and the physical properties of the labeling substance.

[Example 3]
An immunochromatographic diagnostic kit was prepared in the same manner as in Example 1 except that the thermocompression bonding area ratio of the thermoplastic long-fiber nonwoven fabric was 25%, and the performance was evaluated. The results are shown in Tables 1 and 2 below, together with the physical properties of the obtained nonwoven fabric and the physical properties of the labeling substance.

[Example 4]
An immunochromatographic diagnostic kit was prepared in the same manner as in Example 1 except that the thermocompression bonding area ratio of the thermoplastic long-fiber nonwoven fabric was 45%, and the performance was evaluated. The results are shown in Tables 1 and 2 below, together with the physical properties of the obtained nonwoven fabric and the physical properties of the labeling substance.

[Example 5]
An immunochromatographic diagnostic kit was prepared in the same manner as in Example 1 except that the thermoplastic long-fiber nonwoven fabric was spun so that the average fiber diameter became 13 μm, and its performance was evaluated. The results are shown in Tables 1 and 2 below, together with the physical properties of the obtained nonwoven fabric and the physical properties of the labeling substance.

[Example 6]
An immunochromatography diagnostic kit was prepared in the same manner as in Example 1 except that the thermoplastic long-fiber nonwoven fabric was spun so as to have an average fiber diameter of 20 μm, and its performance was evaluated. The results are shown in Tables 1 and 2 below, together with the physical properties of the obtained nonwoven fabric and the physical properties of the labeling substance.

[Example 7]
An immunochromatographic diagnostic kit was prepared in the same manner as in Example 1 except that the thermoplastic long-fiber nonwoven fabric was spun so as to have an average fiber diameter of 75 μm, and its performance was evaluated. The results are shown in Tables 1 and 2 below, together with the physical properties of the obtained nonwoven fabric and the physical properties of the labeling substance.

Example 8
An immunochromatographic diagnostic kit was prepared in the same manner as in Example 1 except that the thermoplastic long-fiber nonwoven fabric was spun so that the thermocompression bonding area ratio was 10% and the basis weight was 30 g / m ² , and its performance was evaluated. The results are shown in Tables 1 and 2 below, together with the physical properties of the obtained nonwoven fabric and the physical properties of the labeling substance.

[Example 9]
An immunochromatographic diagnostic kit was prepared in the same manner as in Example 1 except that the thermoplastic long-fiber nonwoven fabric was spun so as to have a thermocompression bonding rate of 12% and a basis weight of 50 g / m ² , and evaluated its performance. The results are shown in Tables 1 and 2 below, together with the physical properties of the obtained nonwoven fabric and the physical properties of the labeling substance.

[Example 10]
An immunochromatographic diagnostic kit was prepared in the same manner as in Example 1 except that the thermoplastic long-fiber nonwoven fabric was spun so as to have a thermocompression bonding ratio of 37% and a basis weight of 300 g / m ² , and evaluated its performance. The results are shown in Tables 1 and 2 below, together with the physical properties of the obtained nonwoven fabric and the physical properties of the labeling substance.

[Example 11]
An immunochromatographic diagnostic kit was prepared in the same manner as in Example 1 except that the thermoplastic long-fiber nonwoven fabric was spun so that the thermocompression bonding ratio was 43% and the basis weight was 380 g / m ² , and the performance was evaluated. The results are shown in Tables 1 and 2 below, together with the physical properties of the obtained nonwoven fabric and the physical properties of the labeling substance.

[Examples 12 to 15]
An immunochromatographic diagnostic kit was prepared in the same manner as in Example 1 except that the spinning speed and the draft ratio were changed in the method for producing a thermoplastic long-fiber nonwoven fabric, and its performance was evaluated. The results are shown in Tables 1 and 2 below, together with the physical properties of the obtained nonwoven fabric and the physical properties of the labeling substance.

[Example 16]
An immunochromatographic diagnostic kit was prepared in the same manner as in Example 1 except that the concentration of Tween 20 (registered trademark) was changed to 8% in the pretreatment of the thermoplastic long-fiber nonwoven fabric, and its performance was evaluated. The results are shown in Tables 1 and 2 below, together with the physical properties of the obtained nonwoven fabric and the physical properties of the labeling substance.

[Example 17]
An immunochromatographic diagnostic kit was prepared in the same manner as in Example 1 except that the concentration of Tween 20 (registered trademark) was set to 0.05% in the pretreatment of the thermoplastic long-fiber nonwoven fabric, and its performance was evaluated. The results are shown in Tables 1 and 2 below, together with the physical properties of the obtained nonwoven fabric and the physical properties of the labeling substance.

[Example 18]
An immunochromatography diagnostic kit was prepared in the same manner as in Example 1 except that Triton X-45 (registered trademark) was used as a surfactant in the pretreatment of the thermoplastic long-fiber nonwoven fabric, and its performance was evaluated. The results are shown in Tables 1 and 2 below, together with the physical properties of the obtained nonwoven fabric and the physical properties of the labeling substance.

[Example 19]
An immunochromatographic diagnostic kit was prepared in the same manner as in Example 1 except that Tergitol NP-40 (registered trademark) was used as a surfactant in the pretreatment of the thermoplastic long-fiber nonwoven fabric, and its performance was evaluated. The results are shown in Tables 1 and 2 below, together with the physical properties of the obtained nonwoven fabric and the physical properties of the labeling substance.

[Example 20]
An immunochromatographic diagnostic kit was prepared in the same manner as in Example 1 except that the antibody-sensitized gold colloid particles were used, and the performance was evaluated. The results are shown in Tables 1 and 2 below, together with the physical properties of the obtained nonwoven fabric and the physical properties of the labeling substance.

The antibody-sensitized gold colloid particles were prepared as follows.
600 μl of a phosphate buffer (50 mM, pH 7.0) was added to 2500 μl of gold colloid particle suspension (manufactured by Tanaka Kikinzoku Co., average particle diameter 40 nm, particle concentration 0.006 wt%, average particle diameter 40 nm), and further anti-hCG 200 μl of a 0.1% aqueous solution of -α mouse antibody (manufactured by Fitzgerald, monoclonal antibody) is added, and the mixture is vortexed. Subsequently, the mixture was stirred at 25 ° C. for 10 minutes while controlling the temperature. 300 μl of a 1% aqueous PEG solution and 600 μl of a 10% aqueous BSA solution (pH 9.0, containing 50 mM boric acid) were added to the above suspension, and the mixture was vortexed. Thereafter, a centrifugation operation (10000 g, 30 minutes) was performed, and the supernatant was removed. To the residue was added 11000 μl of a 1% BSA aqueous solution (0.05% PEG, 150 mM NaCl, pH 8.2, containing 20 mM Tris), and the mixture was vortexed. Thereafter, a centrifugation operation (10000 g, 30 minutes) was performed, and the supernatant was removed. To the residue, 900 μl of a 1% BSA aqueous solution (0.05% PEG, 150 mM NaCl, pH 8.2, containing 20 mM Tris) was added, and sonication was performed for 30 seconds. Further, 1020 μl of the antibody-sensitized gold colloid particle dispersion was uniformly applied and dried at 50 ° C. for 60 minutes.

[Example 21]
An immunochromatography diagnostic kit was prepared in the same manner as in Example 1 except that the thermoplastic resin of the thermoplastic long-fiber nonwoven fabric was changed to polypropylene, and its performance was evaluated. The results are shown in Tables 1 and 2 below, together with the physical properties of the obtained nonwoven fabric and the physical properties of the labeling substance.

[Example 22]
An immunochromatographic diagnostic kit was prepared in the same manner as in Example 1 except that the thermoplastic resin of the thermoplastic long-fiber nonwoven fabric was changed to nylon 66, and its performance was evaluated. The results are shown in Tables 1 and 2 below, together with the physical properties of the obtained nonwoven fabric and the physical properties of the labeling substance.

[Example 23]
An immunochromatographic diagnostic kit was prepared in the same manner as in Example 1, except that the thermoplastic long-fiber nonwoven fabric was manufactured without drafting and spun so that the birefringence was 0.001 or less (ND). And evaluated its performance. The results are shown in Tables 1 and 2 below, together with the physical properties of the obtained nonwoven fabric and the physical properties of the labeling substance.

[Example 24]
An immunochromatographic diagnostic kit was prepared in the same manner as in Example 1 except that the draft ratio was changed and the birefringence was 0.160 during the production of the thermoplastic long-fiber nonwoven fabric, and the performance was evaluated. did. The results are shown in Tables 1 and 2 below, together with the physical properties of the obtained nonwoven fabric and the physical properties of the labeling substance.

[Example 25]
An immunochromatographic diagnostic kit was prepared in the same manner as in Example 1 except that the draft ratio was changed and the birefringence was adjusted to 0.120 during production of the thermoplastic long-fiber nonwoven fabric, and the performance was evaluated. did. The results are shown in Tables 1 and 2 below, together with the physical properties of the obtained nonwoven fabric and the physical properties of the labeling substance.

[Comparative Example 1]
An immunochromatographic diagnostic kit was prepared in the same manner as in Example 1 except that a thermoplastic long-fiber nonwoven fabric not subjected to thermocompression bonding was used, and its performance was evaluated. The results are shown in Tables 1 and 2 below, together with the physical properties of the obtained nonwoven fabric and the physical properties of the labeling substance.
As is clear from Tables 1 and 2, there was no particle remaining, no streak was generated, and although the release was good, the% CV was greatly reduced.

[Comparative Example 2]
An immunochromatography diagnostic kit was prepared in the same manner as in Example 1 except that the thermoplastic long-fiber nonwoven fabric was spun so as to have a thermocompression bonding ratio of 55%, and its performance was evaluated. The results are shown in Tables 1 and 2 below, together with the physical properties of the obtained nonwoven fabric and the physical properties of the labeling substance.
As is clear from Tables 1 and 2, streaks were strongly generated during immunochromatographic development. In addition, since particles remained, a significant decrease in sensitivity and a decrease in% CV occurred.

[Comparative Example 3]
An immunochromatographic diagnostic kit was prepared in the same manner as in Example 1 except that the thermoplastic long-fiber nonwoven fabric was spun so that the basis weight was 420 g / m ² , and its performance was evaluated. The results are shown in Tables 1 and 2 below, together with the physical properties of the obtained nonwoven fabric and the physical properties of the labeling substance.
As is clear from Tables 1 and 2, particles remained, resulting in a decrease in% CV and a decrease in sensitivity.

[Comparative Example 4]
An immunochromatographic diagnostic kit was prepared in the same manner as in Example 1 except that the thermoplastic long-fiber nonwoven fabric was spun so that the average fiber diameter became 7 μm, and its performance was evaluated. The results are shown in Tables 1 and 2 below, together with the physical properties of the obtained nonwoven fabric and the physical properties of the labeling substance.
As is clear from Tables 1 and 2, particles remained during the development of the immunochromatography, resulting in a significant decrease in sensitivity.

[Comparative Example 5]
An immunochromatographic diagnostic kit was prepared in the same manner as in Example 1 except that the thermoplastic long-fiber nonwoven fabric was spun so that the average fiber diameter became 100 μm, and its performance was evaluated. The results are shown in Tables 1 and 2 below, together with the physical properties of the obtained nonwoven fabric and the physical properties of the labeling substance.
As is clear from Tables 1 and 2, a large decrease in% CV has occurred.

[Comparative Example 6]
As described in Example 1 of Patent Document 2, the same as Example 1 except that a non-thermoplastic nonwoven fabric with a melt blow method, no thermocompression bonding, an average fiber diameter of 2.2 μm, and a basis weight of 90 g / m ² was used. An immunochromatographic diagnostic kit was prepared by the method and its performance was evaluated. The results are shown in Tables 1 and 2 below, together with the physical properties of the obtained nonwoven fabric and the physical properties of the labeling substance. The birefringence of the fiber of the obtained nonwoven fabric could not be measured because the crystal orientation was not made. Therefore, the birefringence is estimated to be less than 0.001. As is clear from Tables 1 and 2, particles remained, resulting in a decrease in% CV and a decrease in sensitivity.

[Comparative Example 7]
As described in Comparative Example 3 of Patent Document 2, the same as Example 1 except that a thermoplastic long-fiber nonwoven fabric having a thermocompression bonding area ratio of 52%, an average fiber diameter of 16 μm, and a basis weight of 100 g / m ² was used by a spun bond method. An immunochromatographic diagnostic kit was prepared in the same manner, and its performance was evaluated. The results are shown in Tables 1 and 2 below, together with the physical properties of the obtained nonwoven fabric and the physical properties of the labeling substance. As is clear from Tables 1 and 2, streaks were strongly generated during immunochromatographic development. In addition, since residual particles were generated, a significant decrease in sensitivity and a large decrease in% CV occurred.

[Comparative Example 8]
An immunochromatography diagnostic kit was prepared in the same manner as in Example 1 except that a commercially available polyester conjugate pad (trade name: 6613, manufactured by Ahlstrom) was used, and its performance was evaluated. The results are shown in Tables 1 and 2 below, together with the physical properties of the obtained nonwoven fabric and the physical properties of the labeling substance. As is clear from Tables 1 and 2, although the sensitivity of the immunochromatography was almost the same as that of Example 1, the% CV was lowered.

In Examples 1 to 25, by controlling the thermocompression bonding area ratio, the average fiber diameter, the basis weight, and the birefringence within a predetermined range, streaks occurred during development as shown in Tables 1 and 2. Without performing the procedure, the TL developed within 60 seconds, the analysis sensitivity was high, and the immunochromatography diagnostic kit excellent in the reproducibility of the result could be obtained.
On the other hand, in Comparative Examples 1 to 8 in which any one of the thermocompression bonding area ratio, the average fiber diameter, the basis weight, and the birefringence is out of the predetermined range, particles remain, streaks, TL coloring time exceeds 60 seconds, and low analytical sensitivity. One of the bad reproducibility occurred.

  The immunochromatographic diagnostic kit according to the present invention has excellent release properties, excellent reproducibility of test results, and improved speed of diagnosis by using a specific thermoplastic long-fiber nonwoven fabric as a conjugate pad. Because of this, it can be widely used for testing for diseases at hospitals and clinics, and for food testing at food companies and the like.

(A) Sample pad (b) Conjugate pad (including antibody sensitizing labeling substance)
(C) Test line (TL)
(D) Control line (e) Nitrocellulose membrane (f) Absorption pad (g) Backing paper

Claims (4)

A conjugate pad made of a thermoplastic long-fiber nonwoven fabric which is composed of thermoplastic long fibers having an average fiber diameter of 10 to 80 μm, has a thermocompression bonding area ratio of 1 to 50%, and has a basis weight of 30 to 400 g / m ^2. Immunochromatographic diagnostic kit containing.   The immunochromatographic diagnostic kit according to claim 1, wherein the thermoplastic long fibers constituting the nonwoven fabric have a birefringence of 0.005 to 0.100.   The immunochromatography diagnostic kit according to claim 1 or 2, wherein the nonwoven fabric has been previously treated with a surfactant.   The immunochromatography diagnostic kit according to claim 3, wherein the water absorption capacity of the nonwoven fabric is 20 to 400%.