Allergy antigen and epitope for the same

Abstract

To provide novel antigens of an allergy to shrimp, methods and kits for diagnosing an allergy to shrimp, pharmaceutical compositions comprising such an antigen, shrimps or processed products of shrimp in which such an antigen is eliminated, and a tester composition for determining the presence or absence of a shrimp antigen in an object of interest.SOLUTION: The present invention provides polypeptides comprising an epitope of an antigen, kits, compositions and methods for diagnosing an allergy, comprising such a polypeptide, pharmaceutical compositions comprising such a polypeptide, and raw materials or processed products in which an antigen comprising such a polypeptide is eliminated or reduced, and further provides a tester composition for determining the presence or absence of an antigen in an object of interest.SELECTED DRAWING: None

Description

[Technical Field]

[0001] The present invention relates to a novel antigen for shrimp allergy. The present invention also relates to a diagnostic kit, diagnostic composition, and diagnostic method for shrimp allergy. The present invention also relates to a pharmaceutical composition containing the antigen, and shrimp or shrimp processed products in which the antigen has been removed or reduced. The present invention further relates to a tester composition for determining the presence or absence of a shrimp antigen in a subject.

[0002] The present invention also relates to a polypeptide comprising an epitope of an antigen. The present invention also relates to an allergy diagnostic kit, diagnostic composition, and diagnostic method comprising the polypeptide. The present invention also relates to a pharmaceutical composition comprising the polypeptide, and a raw material or processed product from which the polypeptide has been removed or reduced. The present invention further relates to a method for producing a processed product from which the polypeptide has been removed or reduced. The present invention still further relates to a tester composition for determining the presence or absence of an antigen comprising the polypeptide in a subject. [Background technology]

[0003] IgE antibodies specific to specific antigens (hereinafter referred to as allergens) are produced in the serum and tissues of allergic patients. The physiological consequences of the interaction between these IgE antibodies and specific antigens trigger an allergic reaction. In a broad sense, an antigen refers to foods and ingredients that cause allergic symptoms, and in a narrow sense, it refers to proteins contained in foods and ingredients (hereinafter referred to as allergen components) that are bound by specific IgE antibodies.

[0004] In conventional allergy test drugs, antigen reagents are often prepared by simply grinding foods or ingredients that are allergen candidates (Patent Document 1). For this reason, it was only possible to detect a positive reaction in an allergy test if the content of the many allergen components contained in the conventional antigen reagent exceeded the threshold at which a positive reaction could be determined for binding with IgE antibodies, and the diagnostic efficiency was not sufficiently high.

[0005] Several allergen components have been identified in candidate foods and ingredients, and test kits have been commercialized. To increase the reliability of allergy testing, comprehensive identification of allergen components is necessary; however, the patient detection rate by measuring the above allergen components is still insufficient. Identifying novel shrimp allergens is extremely important not only for improving the accuracy of diagnostic reagents, but also as targets for hypoallergenic foods, hypoallergenic ingredients, and therapeutic drugs.

[0006] Meanwhile, with regard to the separation and purification of proteins, two-dimensional electrophoresis, in which isoelectric focusing is performed in the first dimension and SDS-PAGE (sodium dodecyl sulfate-polyacrylamide gel electrophoresis) is performed in the second dimension, has recently been used as a method for separating and purifying a variety of proteins from a small amount of sample. The applicants have previously developed two-dimensional electrophoresis methods with high resolution (Patent Documents 2 to 5).

[0007] Allergen-specific IgE antibodies recognize and bind to epitopes, which are specific amino acid sequences in allergen components. However, although there are a few examples in which epitopes have been analyzed for allergen components (Non-Patent Document 1), the current situation is extremely limited. Furthermore, there are currently no allergy diagnostic kits on the market that utilize polypeptides containing epitopes. [Prior art documents] [Patent documents]

[0008] [Patent Document 1] Patent Publication No. 2002-286716 [Patent Document 2] Patent Publication No. 2011-33544 [Patent Document 3] Patent Publication No. 2011-33546 [Patent Document 4] Patent Publication No. 2011-33547 [Patent Document 5] Patent Publication No. 2011-33548 [Non-patent literature]

[0009] [Non-Patent Document 1] Matsuo, H., et al., J. Biol. Chem., (2004), Vol.279, No.13, pp.12135-12140 Summary of the Invention [Problem to be solved by the invention]

[0010] The present invention provides a novel antigen for shrimp allergy. The present invention also provides a method and a diagnostic kit for shrimp allergy. The present invention also provides a pharmaceutical composition containing the antigen, and shrimp or shrimp processed products in which the antigen has been removed or reduced. The present invention further provides a tester composition for determining the presence or absence of a shrimp antigen in a subject.

[0011] The present invention also provides a polypeptide comprising an epitope of an antigen. The present invention also provides an allergy diagnostic kit, diagnostic composition, and diagnostic method comprising the polypeptide. The present invention also provides a pharmaceutical composition comprising the polypeptide, and a raw material or processed product from which an antigen comprising the polypeptide has been removed or reduced. The present invention further relates to a method for producing a processed product from which the antigen has been removed or reduced. The present invention still further provides a tester composition for determining the presence or absence of an antigen comprising the polypeptide in a subject. [Means for solving the problem]

[0012] In order to solve the above problems, the present inventors have conducted extensive research to identify the antigen responsible for shrimp allergy. As a result, they have succeeded in identifying a novel antigen to which IgE antibodies in the serum of patients with shrimp allergy specifically bind. Based on this finding, the present invention has been completed.

[0013] That is, in one aspect, the present invention may be as follows.

[0014] [1] A diagnostic kit for shrimp allergy, comprising at least one of the following proteins (1) to (11): (1) (1A) A protein or a mutant thereof containing the C-terminal portion of myosin heavy chain type 1 or the C-terminal portion of myosin heavy chain type a, which is an antigen for shrimp allergy, and is any one of the following proteins (1A-a) to (1A-e): (1A-a) a protein comprising an amino acid sequence in which one or several amino acids are deleted, substituted, inserted, or added in SEQ ID NO: 2, 45, or 87; (1A-b) a protein comprising an amino acid sequence having 70% or more identity to the amino acid sequence set forth in SEQ ID NO: 2, 45, or 87; (1A-c) A protein comprising an amino acid sequence encoded by a base sequence in which one or several nucleotides are deleted, substituted, inserted or added in SEQ ID NO: 1, 44 or 86; (1A-d) a protein containing an amino acid sequence encoded by a nucleotide sequence having 70% or more identity to the nucleotide sequence shown in SEQ ID NO: 1, 44, or 86; or (1A-e) a protein comprising an amino acid sequence encoded by a nucleic acid that hybridizes under stringent conditions with a nucleic acid consisting of a nucleotide sequence complementary to the nucleotide sequence shown in SEQ ID NO: 1, 44, or 86; or (1B) a protein comprising at least one amino acid sequence selected from the group consisting of SEQ ID NOs: 2 to 43, the group consisting of SEQ ID NOs: 45 to 85, or the group consisting of SEQ ID NOs: 87 to 115; (2) (2A) A protein or a mutant thereof containing the N-terminal portion of myosin heavy chain type 1 or the N-terminal portion of myosin heavy chain type a, which is an antigen for shrimp allergy, and is any one of the following proteins (2A-a) to (2A-e): (2A-a) a protein comprising an amino acid sequence in which one or several amino acids are deleted, substituted, inserted, or added in SEQ ID NO: 117, 141, or 146; (2A-b) a protein comprising an amino acid sequence having 70% or more identity to the amino acid sequence set forth in SEQ ID NO: 117, 141, or 146; (2A-c) A protein comprising an amino acid sequence encoded by a base sequence in which one or several nucleotides are deleted, substituted, inserted, or added in SEQ ID NO: 116, 140, or 145; (2A-d) a protein comprising an amino acid sequence encoded by a nucleotide sequence having 70% or more identity to the nucleotide sequence set forth in SEQ ID NO: 116, 140, or 145; or (2A-e) A protein comprising an amino acid sequence encoded by a nucleic acid that hybridizes under stringent conditions with a nucleic acid consisting of a nucleotide sequence complementary to the nucleotide sequence shown in SEQ ID NO: 116, 140, or 145; or (2B) a protein comprising at least one amino acid sequence selected from the group consisting of SEQ ID NOs: 117 to 139, the group consisting of SEQ ID NOs: 141 to 144, or the group consisting of SEQ ID NOs: 146 to 159; (3) (3A) A protein or a mutant thereof containing the C-terminal portion of myosin heavy chain type 2 or the C-terminal portion of myosin heavy chain type b, which is an antigen for shrimp allergy, and is any one of the following proteins (3A-a) to (3A-e): (3A-a) a protein comprising an amino acid sequence in which one or several amino acids are deleted, substituted, inserted, or added in SEQ ID NO: 161, 179, or 230; (3A-b) a protein comprising an amino acid sequence having 70% or more identity to the amino acid sequence set forth in SEQ ID NO: 161, 179, or 230; (3A-c) A protein comprising an amino acid sequence encoded by a base sequence in which one or several nucleotides are deleted, substituted, inserted, or added in SEQ ID NO: 160, 178, or 229; (3A-d) a protein comprising an amino acid sequence encoded by a nucleotide sequence having 70% or more identity to the nucleotide sequence set forth in SEQ ID NO: 160, 178, or 229; or (3A-e) A protein comprising an amino acid sequence encoded by a nucleic acid that hybridizes under stringent conditions with a nucleic acid consisting of a nucleotide sequence complementary to the nucleotide sequence shown in SEQ ID NO: 160, 178, or 229; or (3B) a protein comprising at least one amino acid sequence selected from the group consisting of SEQ ID NOs: 161 to 177, the group consisting of SEQ ID NOs: 179 to 228, or the group consisting of SEQ ID NOs: 230 to 274; (4) (4A) A protein or a mutant thereof containing the N-terminal portion of myosin heavy chain type 2 or the N-terminal portion of myosin heavy chain type b, which is an antigen for shrimp allergy, and is any one of the following proteins (4A-a) to (4A-e): (4A-a) a protein comprising an amino acid sequence in which one or several amino acids are deleted, substituted, inserted, or added in SEQ ID NO: 276, 300, or 306; (4A-b) a protein comprising an amino acid sequence having 70% or more identity to the amino acid sequence set forth in SEQ ID NO: 276, 300, or 306; (4A-c) A protein comprising an amino acid sequence encoded by a base sequence in which one or several nucleotides are deleted, substituted, inserted, or added in SEQ ID NO: 275, 299, or 305; (4A-d) a protein comprising an amino acid sequence encoded by a nucleotide sequence having 70% or more identity to the nucleotide sequence set forth in SEQ ID NO: 275, 299, or 305; or (4A-e) A protein comprising an amino acid sequence encoded by a nucleic acid that hybridizes under stringent conditions with a nucleic acid consisting of a nucleotide sequence complementary to the nucleotide sequence shown in SEQ ID NO: 275, 299, or 305; or (4B) a protein comprising at least one amino acid sequence selected from the group consisting of SEQ ID NOs: 276 to 298, the group consisting of SEQ ID NOs: 300 to 304, or the group consisting of SEQ ID NOs: 306 to 320; (5) (5A) A glycogen phosphorylase-containing protein or a mutant thereof, which is an antigen for shrimp allergy, is any one of the following proteins (5A-a) to (5A-e): (5A-a) a protein comprising the amino acid sequence of SEQ ID NO: 362 in which one or several amino acids are deleted, substituted, inserted, or added; (5A-b) a protein containing an amino acid sequence having 70% or more identity to the amino acid sequence set forth in SEQ ID NO: 362; (5A-c) a protein comprising an amino acid sequence encoded by a base sequence in which one or several nucleotides in SEQ ID NO: 361 are deleted, substituted, inserted, or added; (5A-d) a protein comprising an amino acid sequence encoded by a nucleotide sequence having 70% or more identity to the nucleotide sequence set forth in SEQ ID NO: 361; or (5A-e) A protein comprising an amino acid sequence encoded by a nucleic acid that hybridizes under stringent conditions with a nucleic acid consisting of a nucleotide sequence complementary to the nucleotide sequence shown in SEQ ID NO: 361; or (5B) a protein comprising at least one amino acid sequence selected from the group consisting of SEQ ID NOs: 321 to 336, the group consisting of SEQ ID NOs: 337 to 360, or the group consisting of SEQ ID NOs: 362 to 379; (6)(6A) Part of hemocyanin subunit L1, hemocyanin or hemocyanin subunit L L), or a mutant thereof, which is an antigen for shrimp allergy, and is any one of the following proteins (6A-a) to (6A-e): (6A-a) A protein comprising an amino acid sequence in which one or several amino acids are deleted, substituted, inserted or added in SEQ ID NO: 381, 399 or 414; (6A-b) a protein comprising an amino acid sequence having 70% or more identity to the amino acid sequence set forth in SEQ ID NO: 381, 399, or 414; (6A-c) A protein comprising an amino acid sequence encoded by a base sequence in which one or several nucleotides are deleted, substituted, inserted, or added in SEQ ID NO: 380, 398, or 413; (6A-d) a protein comprising an amino acid sequence encoded by a nucleotide sequence having 70% or more identity to the nucleotide sequence set forth in SEQ ID NO: 380, 398, or 413; or (6A-e) A protein comprising an amino acid sequence encoded by a nucleic acid that hybridizes under stringent conditions with a nucleic acid consisting of a nucleotide sequence complementary to the nucleotide sequence shown in SEQ ID NO: 380, 398, or 413; or (6B) a protein comprising at least one amino acid sequence selected from the group consisting of SEQ ID NOs: 381 to 397, the group consisting of SEQ ID NOs: 399 to 412, or the group consisting of SEQ ID NOs: 414 to 419; (7) (7A) A protein containing pyruvate kinase 3 or a mutant thereof, which is an antigen for shrimp allergy, and is any one of the following proteins (7A-a) to (7A-e): (7A-a) a protein comprising the amino acid sequence of SEQ ID NO: 421 in which one or several amino acids are deleted, substituted, inserted, or added; (7A-b) a protein containing an amino acid sequence having 70% or more identity to the amino acid sequence set forth in SEQ ID NO: 421; (7A-c) A protein comprising an amino acid sequence encoded by a base sequence in which one or several nucleotides in SEQ ID NO: 420 are deleted, substituted, inserted, or added; (7A-d) a protein comprising an amino acid sequence encoded by a nucleotide sequence having 70% or more identity to the nucleotide sequence set forth in SEQ ID NO: 420; or (7A-e) A protein comprising an amino acid sequence encoded by a nucleic acid that hybridizes under stringent conditions with a nucleic acid consisting of a nucleotide sequence complementary to the nucleotide sequence shown in SEQ ID NO: 420; or (7B) a protein comprising at least one amino acid sequence selected from the group consisting of SEQ ID NOs: 421 to 435 or the group consisting of SEQ ID NOs: 436 to 441; (8) (8A) A protein containing phosphopyruvate hydratase or a mutant thereof, which is an antigen for shrimp allergy, and is any one of the following proteins (8A-a) to (8A-e): (8A-a) a protein comprising an amino acid sequence in which one or several amino acids are deleted, substituted, inserted, or added in SEQ ID NO: 455 or 481; (8A-b) a protein comprising an amino acid sequence having 70% or more identity to the amino acid sequence set forth in SEQ ID NO: 455 or 481; (8A-c) A protein comprising an amino acid sequence encoded by a base sequence in which one or several nucleotides are deleted, substituted, inserted, or added in SEQ ID NO: 454 or 480; (8A-d) a protein comprising an amino acid sequence encoded by a nucleotide sequence having 70% or more identity to the nucleotide sequence set forth in SEQ ID NO: 454 or 480; or (8A-e) A protein comprising an amino acid sequence encoded by a nucleic acid that hybridizes under stringent conditions with a nucleic acid consisting of a nucleotide sequence complementary to the nucleotide sequence shown in SEQ ID NO: 454 or 480; or (8B) a protein comprising at least one amino acid sequence selected from the group consisting of SEQ ID NOs: 442 to 453, the group consisting of SEQ ID NOs: 455 to 479, or the group consisting of SEQ ID NOs: 481 to 484; (9) (9A) A protein containing a mitochondrial ATP synthase subunit alpha precursor or a mutant thereof, which is an antigen for shrimp allergy, and is any one of the following proteins (9A-a) to (9A-e): (9A-a) A protein comprising the amino acid sequence of SEQ ID NO: 486 in which one or several amino acids are deleted, substituted, inserted, or added; (9A-b) A protein containing an amino acid sequence having 70% or more identity to the amino acid sequence set forth in SEQ ID NO: 486; (9A-c) A protein comprising an amino acid sequence encoded by a base sequence in which one or several nucleotides in SEQ ID NO: 485 are deleted, substituted, inserted, or added; (9A-d) a protein comprising an amino acid sequence encoded by a nucleotide sequence having 70% or more identity to the nucleotide sequence set forth in SEQ ID NO: 485; or (9A-e) A protein comprising an amino acid sequence encoded by a nucleic acid that hybridizes under stringent conditions with a nucleic acid consisting of a nucleotide sequence complementary to the nucleotide sequence shown in SEQ ID NO: 485; or (9B) a protein comprising at least one amino acid sequence selected from the group consisting of SEQ ID NOs: 486 to 490, the group consisting of SEQ ID NOs: 491 to 497, or the group consisting of SEQ ID NOs: 498 to 518; (10) (10A) A protein containing troponin I or a mutant thereof, which is an antigen for shrimp allergy, is any one of the following proteins (10A-a) to (10A-e): (10A-a) a protein comprising the amino acid sequence of SEQ ID NO: 520 in which one or several amino acids are deleted, substituted, inserted, or added; (10A-b) a protein containing an amino acid sequence having 70% or more identity to the amino acid sequence set forth in SEQ ID NO: 520; (10A-c) A protein comprising an amino acid sequence encoded by a base sequence in which one or several nucleotides in SEQ ID NO: 519 have been deleted, substituted, inserted, or added; (10A-d) a protein comprising an amino acid sequence encoded by a nucleotide sequence having 70% or more identity to the nucleotide sequence set forth in SEQ ID NO: 519; or (10A-e) A protein comprising an amino acid sequence encoded by a nucleic acid that hybridizes under stringent conditions with a nucleic acid consisting of a nucleotide sequence complementary to the nucleotide sequence shown in SEQ ID NO: 519; or (10B) a protein comprising at least one amino acid sequence selected from the group consisting of SEQ ID NOs: 520 to 527, the group consisting of SEQ ID NOs: 528 to 532, or the group consisting of SEQ ID NOs: 533 to 539; (11) (11A) A protein containing cyclophilin A or a mutant thereof, which is an antigen for shrimp allergy, is any one of the following proteins (11A-a) to (11A-e): (11A-a) A protein comprising an amino acid sequence in which one or several amino acids are deleted, substituted, inserted, or added in SEQ ID NO: 541 or 549; (11A-b) a protein containing an amino acid sequence having 70% or more identity to the amino acid sequence set forth in SEQ ID NO: 541 or 549; (11A-c) A protein comprising an amino acid sequence encoded by a base sequence in which one or several nucleotides in SEQ ID NO: 540 or 548 have been deleted, substituted, inserted or added; (11A-d) A protein comprising an amino acid sequence encoded by a base sequence having 70% or more identity to the base sequence shown in SEQ ID NO: 540 or 548; or (11A-e) A protein comprising an amino acid sequence encoded by a nucleic acid that hybridizes under stringent conditions with a nucleic acid consisting of a nucleotide sequence complementary to the nucleotide sequence shown in SEQ ID NO: 540 or 548; or (11B) a protein comprising at least one amino acid sequence selected from the group consisting of SEQ ID NOs: 541 to 547, the group consisting of SEQ ID NOs: 549 to 553, or the group consisting of SEQ ID NOs: 554 to 557; as an antigen.

[0015] [2] A diagnostic composition for shrimp allergy, comprising at least one of the proteins specified as (1) to (11) in [1] above as an antigen.

[0016] [3] A method for providing an indicator for diagnosing shrimp allergy in a subject, comprising the steps of: (i) contacting a sample obtained from a subject with an antigen, wherein the sample is a solution containing IgE antibodies; (ii) detecting binding of IgE antibodies to the antigen in a sample obtained from the subject; (iii) if binding of the subject's IgE antibodies to the antigen is detected, an indication that the subject is allergic to shrimp is provided; wherein the antigen is at least one of the proteins specified as (1) to (11) in [1] above.

[0017] [4] A pharmaceutical composition comprising at least one of the proteins identified as (1) to (11) in [1] above.

[0018] [5] The pharmaceutical composition according to [4] above for treating shrimp allergy.

[0019] [6] Shrimp or shrimp processed products characterized in that the antigen is removed or reduced, and the antigen is at least one of the proteins specified as (1) to (11) in [1] above.

[0020] [7] A tester composition for determining the presence or absence of a shrimp antigen in a subject, characterized by comprising an antibody that binds to at least one of the proteins specified as (1) to (11) in [1] above.

[0021] [8] A tester composition for determining the presence or absence of an antigen that causes shrimp allergy in a subject, characterized by comprising a primer having a base sequence complementary to a portion of at least one base sequence selected from the group consisting of SEQ ID NOs: 1, 44, 86, 116, 140, 145, 160, 178, 229, 275, 299, 305, 361, 380, 398, 413, 420, 454, 480, 485, 519, 540, and 548.

[0022] The present inventors have also succeeded in finding epitopes for shrimp-derived antigens, including the above antigens.

[0023] Because epitopes have relatively short amino acid sequences, if the same amino acid sequence is present in different allergen components, the IgE antibody can bind to multiple allergen components. As a result of the presence of a common epitope in different allergen components, IgE antibodies from allergy patients bind to both of them, making the antigen cross-reactive. Therefore, the epitopes identified in the present application enable the diagnosis and treatment of allergies, including cross-reactivity, and the detection of multiple allergen components containing the epitope.

[0024] Based on this finding, the present invention has been completed. That is, in another aspect, the present invention may be as follows.

[0025] [Aspect 1] An allergy diagnostic kit comprising at least one of the following polypeptides: (E1)(i) a polypeptide comprising the amino acid sequence of SEQ ID NO: 558-565; (ii) a polypeptide comprising an amino acid sequence of SEQ ID NOs: 558-565, in which one or more amino acid residues corresponding to amino acid residues at positions 4, 5, 7, 8, 10, 11, 12, 13, 14, and 15 of SEQ ID NO: 558 are substituted with any amino acid residue; (E2)(i) a polypeptide comprising the amino acid sequence of SEQ ID NOs: 566-581, 949; (ii) a polypeptide comprising an amino acid sequence of SEQ ID NOs: 566-581, 949, in which one or more amino acid residues corresponding to the 1st, 3rd, 4th, 5th, 6th, 7th, 8th, 9th, 11th, and 12th amino acid residues of SEQ ID NO: 566 are substituted with any amino acid residue; (E3)(i) a polypeptide comprising the amino acid sequence of SEQ ID NOs: 582-585, 950; (ii) a polypeptide comprising an amino acid sequence of SEQ ID NOs: 582-585, 950, in which one or more amino acid residues corresponding to the 9th, 10th, 11th, 12th, and 13th amino acid residues of SEQ ID NO: 582 are substituted with any amino acid residue; (E4) (i) a polypeptide comprising the amino acid sequence of SEQ ID NOs: 586-593, 951; (ii) a polypeptide comprising an amino acid sequence of SEQ ID NOs: 586-593, 951, in which one or more amino acid residues corresponding to the 2nd, 4th, 5th, 6th, 7th, 10th, 11th, and 12th amino acid residues of SEQ ID NO: 586 are substituted with any amino acid residue; (E5) (i) a polypeptide comprising the amino acid sequence of SEQ ID NOs: 594-598, 952; (ii) a polypeptide comprising an amino acid sequence of SEQ ID NOs: 594-598, 952, in which one or more amino acid residues corresponding to the 9th, 11th, and 12th amino acid residues of SEQ ID NO: 594 are substituted with any amino acid residue; (E6)(i), a polypeptide comprising the amino acid sequence of SEQ ID NOs: 599-613, 953, 954; (ii) a polypeptide comprising an amino acid sequence of SEQ ID NOs: 599-613, 953, 954, in which one or more amino acid residues corresponding to the 2nd, 3rd, 4th, 5th, 6th, 7th, 8th, 9th, 10th, 11th, 12th, and 13th amino acid residues of SEQ ID NO: 599 are substituted with any amino acid residue; (E7) (i) a polypeptide comprising the amino acid sequence of SEQ ID NOs: 614-623, 955; (ii) a polypeptide comprising an amino acid sequence of SEQ ID NOs: 614-623, 955, in which one or more amino acid residues corresponding to the 1st, 2nd, 3rd, 4th, 5th, 6th, 7th, 8th, 9th, 10th, 13th, 14th, and 15th amino acid residues of SEQ ID NO: 614 are substituted with any amino acid residue; (E8)(i) a polypeptide comprising the amino acid sequence of SEQ ID NO: 624-633; (ii) a polypeptide comprising an amino acid sequence of SEQ ID NOs: 624-633, in which one or more amino acid residues corresponding to the 3rd, 5th, 6th, 7th, 8th, 10th, 11th, 12th, and 14th amino acid residues of SEQ ID NO: 624 are substituted with any amino acid residue; (E9)(i) a polypeptide comprising the amino acid sequence of SEQ ID NO: 634-639; (ii) a polypeptide comprising an amino acid sequence of SEQ ID NOs: 634-639, in which one or more amino acid residues corresponding to the 4th, 6th, 8th, 10th, and 11th amino acid residues of SEQ ID NO: 634 are substituted with any amino acid residue; (E10)(i) a polypeptide comprising the amino acid sequence of SEQ ID NOs: 640-653, 956, 957; (ii) a polypeptide comprising an amino acid sequence of SEQ ID NOs: 640-653, 956, or 957, in which one or more amino acid residues corresponding to amino acid residues 4, 6, 8, 9, 11, 14, or 15 of SEQ ID NO: 640 are substituted with any amino acid residue; (E11)(i) a polypeptide comprising the amino acid sequence of SEQ ID NO: 654-670; (ii) a polypeptide comprising an amino acid sequence of SEQ ID NO: 654-670, in which one or more amino acid residues corresponding to amino acid residues at positions 2, 4, 5, 6, 7, 8, 9, 10, 12, 13, and 15 of SEQ ID NO: 654 are substituted with any amino acid residue; (E12) a polypeptide comprising the amino acid sequence of SEQ ID NO: 671; (E13) (i) a polypeptide comprising the amino acid sequence of SEQ ID NOs: 672-677, 958; (ii) In SEQ ID NOs: 672-677, 958, one or more amino acid residues corresponding to the 6th, 7th, 8th, 9th, 10th, and 14th amino acid residues of SEQ ID NO: 672 may be substituted with any amino acid residues corresponding to these residues.

[0026] (E14) a polypeptide comprising the amino acid sequence of SEQ ID NO: 678-680; (E15)(i) a polypeptide comprising the amino acid sequence of SEQ ID NOs: 681-685; (ii) a polypeptide comprising an amino acid sequence of SEQ ID NOs: 681-685, in which one or more amino acid residues corresponding to the 10th amino acid residue of SEQ ID NO: 681 are substituted with any amino acid residue; (E16)(i) a polypeptide comprising the amino acid sequence of SEQ ID NOs: 686-690, 959,960; (ii) a polypeptide comprising an amino acid sequence of SEQ ID NOs: 686-690, 959, and 960, in which one or more amino acid residues corresponding to the 4th, 5th, 6th, 7th, 9th, 10th, and 12th amino acid residues of SEQ ID NO: 686 are substituted with any amino acid residue; (E17) (i) a polypeptide comprising the amino acid sequence of SEQ ID NOs: 691-696, 961; (ii) a polypeptide comprising an amino acid sequence of SEQ ID NOs: 691-696, 961, in which one or more amino acid residues corresponding to the 7th, 10th, and 12th amino acid residues of SEQ ID NO: 691 are substituted with any amino acid residue; (E18)(i) a polypeptide comprising the amino acid sequence of SEQ ID NO: 697-703; (ii) a polypeptide comprising an amino acid sequence of SEQ ID NOs: 697-703, in which one or more amino acid residues corresponding to the 1st, 3rd, and 5th amino acid residues of SEQ ID NO: 697 are substituted with any amino acid residue; (E19) a polypeptide comprising the amino acid sequence of SEQ ID NO: 704; (E20) a polypeptide comprising the amino acid sequence of SEQ ID NO: 705-707; (E21) (i) a polypeptide comprising the amino acid sequence of SEQ ID NOs: 708-716, 962, 963; (ii) a polypeptide comprising an amino acid sequence of SEQ ID NOs: 708-716, 962, 963, in which one or more amino acid residues corresponding to the 1st, 2nd, 4th, 5th, 7th, 8th, 9th, 10th, 12th, 13th, and 15th amino acid residues of SEQ ID NO: 708 are substituted with any amino acid residue; (E22) (i) a polypeptide comprising the amino acid sequence of SEQ ID NOs: 717-724, 964; (ii) a polypeptide comprising an amino acid sequence of SEQ ID NOs: 717-724, 964, in which one or more amino acid residues corresponding to the 2nd, 4th, 5th, 6th, 8th, 9th, 10th, and 12th amino acid residues of SEQ ID NO: 717 are substituted with any amino acid residue; (E23) a polypeptide comprising the amino acid sequence of SEQ ID NO: 725-728; (E24) (i) a polypeptide comprising the amino acid sequence of SEQ ID NOs: 729-740, 965, 966; (ii) a polypeptide comprising an amino acid sequence of SEQ ID NOs: 729-740, 965, 966, in which one or more amino acid residues corresponding to amino acid residues 4, 5, 6, 7, 8, 9, 11, and 12 of SEQ ID NO: 729 are substituted with any amino acid residue; (E25) (i) a polypeptide comprising the amino acid sequence of SEQ ID NOs: 741-749, 967, 968; (ii) a polypeptide comprising an amino acid sequence of SEQ ID NOs: 741-749, 967, 968, in which one or more amino acid residues corresponding to the 3rd, 5th, 6th, 8th, 9th, 10th, 11th, and 13th amino acid residues of SEQ ID NO: 741 are substituted with any amino acid residue; (E26) a polypeptide comprising the amino acid sequence of SEQ ID NO: 750-751; (E27) (i) a polypeptide comprising the amino acid sequence of SEQ ID NOs: 752-764, 969, 970; (ii) a polypeptide comprising an amino acid sequence of SEQ ID NOs: 752-764, 969, 970, in which one or more amino acid residues corresponding to the 3rd, 4th, 5th, 7th, 8th, 9th, 10th, and 11th amino acid residues of SEQ ID NO: 752 are substituted with any amino acid residue; (E28)(i) a polypeptide comprising the amino acid sequence of SEQ ID NO: 765-769; (ii) a polypeptide comprising an amino acid sequence of SEQ ID NOs: 765-769, in which one or more amino acid residues corresponding to the 3rd, 5th, and 6th amino acid residues of SEQ ID NO: 765 are substituted with any amino acid residue; (E29) (i) a polypeptide comprising the amino acid sequence of SEQ ID NOs: 770-777, 971, 972; (ii) a polypeptide comprising an amino acid sequence of SEQ ID NOs: 770-777, 971, 972, in which one or more amino acid residues corresponding to the 1st, 2nd, 5th, 6th, 7th, 8th, 9th, 10th, 11th, and 12th amino acid residues of SEQ ID NO: 770 are substituted with any amino acid residue; (E30) (i) a polypeptide comprising the amino acid sequence of SEQ ID NOs: 778-787, 973; (ii) a polypeptide comprising an amino acid sequence of SEQ ID NOs: 778-787, 973, in which one or more amino acid residues corresponding to the 2nd, 3rd, 4th, 6th, 7th, 8th, 9th, 10th, and 14th amino acid residues of SEQ ID NO: 778 are substituted with any amino acid residue; (E31)(i) a polypeptide comprising the amino acid sequence of SEQ ID NO: 788-792; (ii) a polypeptide comprising an amino acid sequence of SEQ ID NOs: 788-792, in which one or more amino acid residues corresponding to the 4th, 7th, 8th, 9th, 10th, 11th, and 13th amino acid residues of SEQ ID NO: 788 are substituted with any amino acid residue; (E32) (i) a polypeptide comprising the amino acid sequence of SEQ ID NOs: 793-809, 974, 975; (ii) a polypeptide comprising an amino acid sequence of SEQ ID NOs: 793-809, 974, 975, in which one or more amino acid residues corresponding to the 3rd, 4th, 5th, 6th, 7th, 8th, 9th, 10th, 11th, 12th, 13th, 14th, and 15th amino acid residues of SEQ ID NO: 793 are substituted with any amino acid residue; (E33)(i) a polypeptide comprising the amino acid sequence of SEQ ID NO: 810-816; (ii) a polypeptide comprising an amino acid sequence of SEQ ID NOs: 810-816, in which one or more amino acid residues corresponding to the 2nd, 4th, 5th, 6th, 7th, 8th, 9th, 10th, 12th, and 14th amino acid residues of SEQ ID NO: 810 are substituted with any amino acid residue; (E34)(i) a polypeptide comprising the amino acid sequence of SEQ ID NO: 817-825; (ii) a polypeptide comprising an amino acid sequence of SEQ ID NOs: 817-825, in which one or more amino acid residues corresponding to the amino acid residues at positions 2, 4, 5, 7, 8, 9, 11, 12, 13, and 14 of SEQ ID NO: 817 are substituted with any amino acid residue; (E35)(i) a polypeptide comprising the amino acid sequence of SEQ ID NO: 826-832; (ii) a polypeptide comprising an amino acid sequence of SEQ ID NOs: 826-832, in which one or more amino acid residues corresponding to the 2nd, 3rd, 5th, 8th, 9th, 10th, 11th, 12th, and 14th amino acid residues of SEQ ID NO: 826 are substituted with any amino acid residue; (E36) (i) a polypeptide comprising the amino acid sequence of SEQ ID NOs: 833-846, 976; (ii) a polypeptide comprising an amino acid sequence of SEQ ID NOs: 833-846, 976, in which one or more amino acid residues corresponding to the 1st, 2nd, 3rd, 4th, 5th, 6th, 7th, 8th, 9th, 10th, 11th, 12th, 13th, and 14th amino acid residues of SEQ ID NO: 833 are substituted with any amino acid residue; (E37) (i) a polypeptide comprising the amino acid sequence of SEQ ID NOs: 847-857, 977; (ii) a polypeptide comprising an amino acid sequence of SEQ ID NOs: 847-857, 977, in which one or more amino acid residues corresponding to the 5th, 6th, 7th, 9th, 10th, 11th, 12th, 13th, and 14th amino acid residues of SEQ ID NO: 847 are substituted with any amino acid residue; (E38)(i) a polypeptide comprising the amino acid sequence of SEQ ID NO: 858-864; (ii) a polypeptide comprising an amino acid sequence of SEQ ID NOs: 858-864, in which one or more amino acid residues corresponding to the 6th, 7th, 8th, 9th, 13th, and 14th amino acid residues of SEQ ID NO: 858 are substituted with any amino acid residue; (E39) (i) a polypeptide comprising the amino acid sequence of SEQ ID NOs: 865-878, 984, 978; (ii) a polypeptide comprising an amino acid sequence of SEQ ID NOs: 865-878, 984, 978, in which one or more amino acid residues corresponding to amino acid residues 1, 2, 4, 5, 6, 8, 9, 10, 11, and 12 of SEQ ID NO: 865 are substituted with any amino acid residue; (E40) a polypeptide comprising the amino acid sequence of SEQ ID NO: 879-880; (E41) (i) a polypeptide comprising the amino acid sequence of SEQ ID NOs: 881-891, 979; (ii) a polypeptide comprising an amino acid sequence of SEQ ID NOs: 881-891, 979, in which one or more amino acid residues corresponding to the amino acid residues at positions 2, 3, 4, 6, 9, 10, 11, 12, 13, and 15 of SEQ ID NO: 881 are substituted with any amino acid residue; (E42) (i) a polypeptide comprising the amino acid sequence of SEQ ID NOs: 892-907, 980; (ii) a polypeptide comprising an amino acid sequence of SEQ ID NOs: 892-907, 980, in which one or more amino acid residues corresponding to the 1st, 2nd, 3rd, 4th, 5th, 8th, 9th, 10th, 11th, 12th, 13th, 14th, and 15th amino acid residues of SEQ ID NO: 892 are substituted with any amino acid residue; (E43) (i) a polypeptide comprising the amino acid sequence of SEQ ID NOs: 908-911, 981; (ii) a polypeptide comprising an amino acid sequence of SEQ ID NOs: 908-911, 981, in which one or more amino acid residues corresponding to the 7th amino acid residue of SEQ ID NO: 908 are substituted with any amino acid residue; (E44) a polypeptide comprising the amino acid sequence of SEQ ID NO: 912-914; (E45) a polypeptide comprising the amino acid sequence of SEQ ID NO: 915-916; (E46) (i) a polypeptide comprising the amino acid sequence of SEQ ID NOs: 917-927, 982, 983; (ii) a polypeptide comprising an amino acid sequence of SEQ ID NOs: 917-927, 982, 983, in which one or more amino acid residues corresponding to the 4th, 7th, 8th, 9th, 10th, and 15th amino acid residues of SEQ ID NO: 917 are substituted with any amino acid residue; (E47)(i) a polypeptide comprising the amino acid sequence of SEQ ID NO: 928-934; (ii) a polypeptide comprising an amino acid sequence of SEQ ID NOs: 928-934, in which one or more amino acid residues corresponding to the 6th, 7th, 8th, 10th, 12th, and 14th amino acid residues of SEQ ID NO: 928 are substituted with any amino acid residue; (E48)(i) a polypeptide comprising the amino acid sequence of SEQ ID NO: 935-938; (ii) a polypeptide comprising the amino acid sequence of SEQ ID NOs: 935-938, in which one or more amino acid residues corresponding to the 5th and 6th amino acid residues of SEQ ID NO: 935 are substituted with any amino acid residue; (E49)(i) a polypeptide comprising the amino acid sequence of SEQ ID NO: 939-942; (ii) a polypeptide comprising an amino acid sequence of SEQ ID NOs: 939-942, in which one or more amino acid residues corresponding to the 3rd, 6th, 7th, 10th, and 11th amino acid residues of SEQ ID NO: 939 are substituted with any amino acid residue; (E50) (i) a polypeptide comprising the amino acid sequence of SEQ ID NO: 943-948; (ii) A polypeptide comprising an amino acid sequence of SEQ ID NO: 943-948, in which one or more amino acid residues corresponding to the 1st, 2nd, 3rd, 4th, 5th, 8th, 10th, and 11th amino acid residues of SEQ ID NO: 943 are substituted with any amino acid residue.

[0027] [Aspect 2] A composition for diagnosing allergies, comprising at least one polypeptide according to embodiment 1.

[0028] [Aspect 3] A polypeptide that specifically binds to IgE antibodies of an allergy sufferer, the polypeptide being any one of the polypeptides according to aspect 1.

[0029] [Aspect 4] 1. A method for providing an indicator for diagnosing allergy in a subject, comprising the steps of: (i) contacting a sample obtained from a subject with an antigen, wherein the sample is a solution containing IgE antibodies; (ii) detecting binding of IgE antibodies to the antigen in a sample obtained from the subject; (iii) if binding of the subject's IgE antibodies to the antigen is detected, an indication that the subject is allergic is provided; wherein the antigen is at least one of the polypeptides of embodiment 3.

[0030] [Aspect 5] A pharmaceutical composition comprising at least one polypeptide according to embodiment 3.

[0031] [Aspect 6] 6. The pharmaceutical composition according to aspect 5, for treating allergies.

[0032] [Aspect 7] A tester composition for determining the presence or absence of an antigen in a subject, comprising an antibody that binds to at least one of the polypeptides according to embodiment 3.

[0033] [Aspect 8] One of the following primers: (a) a primer comprising a portion of the nucleotide sequence of a nucleic acid encoding the polypeptide of embodiment 3 and / or a portion of the complementary strand thereof; or (b) a primer which is a part of at least one of the nucleotide sequences shown in SEQ ID NO: 1, 44, 86, 116, 140, 145, 160, 178, 229, 275, 299, 305, 361, 380, 398, 413, 420, 454, 480, 485, 519, 540, or 548 and / or a primer which is a part of a sequence complementary to at least one of the nucleotide sequences shown in SEQ ID NO: 1, 44, 86, 116, 140, 145, 160, 178, 229, 275, 299, 305, 361, 380, 398, 413, 420, 454, 480, 485, 519, 540, or 548; A tester composition for determining the presence or absence of an antigen in a subject, comprising:

[0034] [Aspect 9] A method for determining the presence or absence of the polypeptide according to aspect 3 in a raw material or processed product, the method comprising detecting the polypeptide according to aspect 3 in the raw material or processed product.

[0035] [Aspect 10] A raw material or processed product characterized in that an antigen has been removed or reduced, wherein the antigen is at least one of the polypeptides described in aspect 3.

[0036] [Aspect 11] A method for producing a processed product in which antigens have been removed or reduced, the method comprising a step of confirming that the antigens have been removed or reduced during the production of the processed product, wherein the antigen is at least one of the polypeptides described in Aspect 3. [Effects of the Invention]

[0037] The present invention provides a novel antigen for shrimp allergy. Since the present invention has identified a novel allergen component that causes shrimp allergy, it can provide a highly sensitive diagnostic method and diagnostic kit for shrimp allergy, a pharmaceutical composition containing the antigen, shrimp or shrimp processed products in which the antigen has been removed or reduced, and a tester composition for determining the presence or absence of the shrimp antigen in a target substance.

[0038] The present invention also provides a novel polypeptide containing an epitope of an antigen. By utilizing the polypeptide of the present invention, it is possible to provide a highly sensitive diagnostic kit, diagnostic composition, and diagnostic method for allergies, a pharmaceutical composition containing the polypeptide, a tester composition for determining the presence or absence of an antigen containing the polypeptide in a target substance, as well as a raw material or processed product from which the polypeptide has been removed or reduced, and a method for producing such a processed product. [Brief explanation of the drawings]

[0039] [Figure 1] Figure 1 is a photograph of a gel showing the migration pattern of proteins contained in vannamei shrimp by two-dimensional electrophoresis. The bands on the left side of the photograph are molecular weight marker bands, and the numbers on the left side of the photograph are the molecular weight (KDa) of each molecular weight marker. The numbers at the top of the photograph represent the isoelectric points. [Figure 2]Figure 2 shows a photograph of an immunoblot performed using serum from a patient with shrimp allergy against the two-dimensional electrophoresis pattern of proteins contained in vannamei shrimp. Spots 1 to 11, which specifically reacted with IgE antibodies in the serum of the patient with shrimp allergy, are each enclosed in a white frame. [Figure 3] Figure 3 is a photograph of a gel showing the migration pattern of proteins contained in black tiger shrimp by two-dimensional electrophoresis. The bands on the left side of the photograph are molecular weight marker bands, and the numbers on the left side of the photograph are the molecular weight (KDa) of each molecular weight marker. The numbers at the top of the photograph represent the isoelectric point. [Figure 4] Figure 4 shows a photograph of an immunoblot performed using the serum of a shrimp allergy patient against the two-dimensional electrophoresis pattern of proteins contained in black tiger shrimp. Spots 1 to 11, which specifically reacted with IgE antibodies in the serum of the shrimp allergy patient, are each enclosed in a white frame. [Figure 5] Figure 5 is a photograph of a gel showing the migration pattern of proteins contained in kuruma shrimp by two-dimensional electrophoresis. The bands on the left side of the photograph are molecular weight marker bands, and the numbers on the left side of the photograph are the molecular weight (KDa) of each molecular weight marker. The numbers at the top of the photograph represent the isoelectric point. [Figure 6] Figure 6 shows a photograph of an immunoblot performed using serum from a shrimp-allergic patient against the two-dimensional electrophoresis pattern of proteins contained in kuruma shrimp. Spots 1-6 and 8-11, which specifically reacted with IgE antibodies in the shrimp-allergic patient's serum, are each enclosed in a white frame. [Figure 7] FIG. 7 shows the results of examining cross-reactivity by ELISA for peptides having the amino acid sequences of each epitope using sera from patients who are simultaneously allergic to shrimp and wheat or crab. [Figure 8] FIG. 8 shows the results of examining cross-reactivity by ELISA for peptides having the amino acid sequences of each epitope using sera from patients who are simultaneously allergic to shrimp and wheat or crab. [Figure 9]FIG. 9 shows the results of examining cross-reactivity by ELISA for peptides having the amino acid sequences of each epitope using sera from patients who are simultaneously allergic to shrimp and wheat or crab. [Figure 10] FIG. 10 shows the results of examining cross-reactivity by ELISA for peptides having the amino acid sequences of each epitope using sera from patients who are simultaneously allergic to shrimp and wheat or crab. [Figure 11] FIG. 11 shows the results of examining cross-reactivity by ELISA for peptides having the amino acid sequences of each epitope using sera from patients who are simultaneously allergic to shrimp and wheat or crab. [Figure 12] FIG. 12 shows the results of examining cross-reactivity by ELISA for peptides having the amino acid sequences of each epitope using sera from patients who are simultaneously allergic to shrimp and wheat or crab. DETAILED DESCRIPTION OF THE INVENTION

[0040] The present invention will be specifically described below, but the present invention is not limited thereto.

[0041] Unless otherwise defined herein, scientific and technical terms used in connection with the present invention shall have the meanings that are commonly understood by those skilled in the art.

[0042] As used herein, allergy refers to a state in which an organism sensitized to a certain antigen exhibits an adverse hypersensitivity reaction when that antigen is reintroduced. An allergic reaction can occur when the organism comes into contact with or ingests the antigen. Here, "contact" refers to touching an object, and in the case of the human body, it particularly refers to adhesion to the skin or mucous membranes (eyes, lips, etc.). Furthermore, "ingestion" refers to taking something into the body, such as by inhalation or oral administration. Generally, an allergic reaction that occurs when food is ingested is specifically referred to as a food allergy. In a preferred embodiment, the allergy may be a food allergy. In many food allergic diseases, antigen-specific IgE antibodies are produced in the blood and tissues. The IgE antibodies bind to mast cells or basophils. When an antigen specific to the IgE antibody reintroduces itself into the body of an allergic patient, the antigen combines with the IgE antibody bound to mast cells or basophils, resulting in the physiological effects of IgE antibody-antigen interaction. These physiological effects include the release of histamine, serotonin, heparin, eosinophil chemotactic factor, and various leukotrienes. These released substances trigger allergic reactions caused by the combination of IgE antibodies and specific antigens. Specifically, IgE antibodies recognize and bind to epitopes, which are specific amino acid sequences in specific antigens, and allergic reactions caused by these antigens manifest through the above pathways.

[0043] The allergy targeted by the present invention is not particularly limited as long as it is an allergy to an allergen containing the epitope used. In one embodiment, the allergen includes seafood, fruits, vegetables, nuts (seeds), edible grasses, grains, meat, milk, dairy products, etc. ingested by living organisms (especially humans), or parasites that infect living organisms (especially humans).

[0044] Seafood includes, but is not limited to, shrimp, crabs, and the like, which belong to the Decapoda (order Decapoda). Most of what is generally recognized as "crustaceans" are included in the Decapoda (order Decapoda). The Decapoda (order Decapoda) includes the infraorder Brachyura (also known as the infraorder Crabs) and the infraorder Decapoda Anomura (infraorder Hermit Crabs). Of the Decapoda (order Decapoda), all species except the infraorder Brachyura (also known as the infraorder Crabs) and the infraorder Decapoda Anomura (infraorder Hermit Crabs) are collectively referred to as "shrimp." Shrimp will be discussed later. The infraorder Decapoda Brachyura (also known as the infraorder Crabs) includes, but is not limited to, the family Scyllidae (e.g., Erimacrus isenbeckii), the family Scyllidae (e.g., Chionoecetes opilio), and the family Portunidae (Portunus trituberculatus). Without limitation, the Decapoda Anomura (Infraorder Hermit Crabs) includes the family Paralithidae (e.g., Paralithodes camtschaticus).

[0045] Seafood also includes, but is not limited to, squid belonging to the order Pectiniformes and the order Octopidae. Seafood further includes fish belonging to the families Scombridae and Gadidae. Seafood further includes shellfish from the family Veneridae. Squid from the family Pectiniformes include, but are not limited to, the Japanese common squid (Todarodes pacificus), which is also called "true squid." Octopuses from the family Octopidae include the common octopus (Octopus vulgaris). Fish from the family Scombridae include tuna (Thunnus orientalis) and chub mackerel (Scomber japonicas). Fish from the family Gadidae include Pacific cod (Gadus macrocephalus). Shellfish from the family Veneridae include clams, hard clams, and mussels. In one embodiment, they include the clam (Ruditapes philippinarum).

[0046] Non-limiting examples of fruits include fruits belonging to the Actinidiaceae, Bromeliaceae, Anacardiaceae, Cucurbitaceae, Musaceae, Rutaceae, and Rosaceae families. Vegetables include fruits belonging to the Solanaceae, Cucurbitaceae, and Lauraceae families. Nuts (seeds) include nuts (seeds) belonging to the Anacardiaceae, Rosaceae, and Juglandaceae families. Edible herbs include edible herbs belonging to the Asteraceae family. Grains include grains belonging to the Poaceae and Polygonaceae families.

[0047] Without limitation, Actinidiaceae fruits include kiwi (Actinidia deliciosa). Bromeliaceae fruits include pineapple (Ananas comosus). Anacardiaceae includes fruits such as mango (Mangifera indica) and nuts (seeds) such as cashew (Anacardium occidentale). Cucurbitaceae includes fruits such as melon (Cucumis melo) and vegetables such as cucumber (Cucumis sativus). Musaceae fruits include banana (Musa acuminate). Rutaceae fruits include orange (Citrus sinensis). Rosaceae includes fruits such as peaches, strawberries, apples, pears, and loquats, as well as nuts (seeds) such as almonds. In one aspect, Rosaceae includes apple (Malus domestica) and almond (Prunus dulcis).

[0048] Vegetables from the Solanaceae family include, but are not limited to, eggplant (Solanum melongena) and tomato (Solanum lycopersicum). Vegetables from the Lauraceae family include avocado (Persea americana). Nuts from the Anacardiaceae family include, but are not limited to, cashews (Anacardium occidentale) and almonds (Prunus dulcis) from the Rosaceae family. Nuts from the Juglandaceae family include walnuts (Juglans regia).

[0049] Non-limiting examples of edible herbs in the Asteraceae family include mugwort (Artemisia indica var. maximowiczii or Artemisia indica). Grains in the Gramineae family include bread wheat (Triticum aestivum). Grains in the Polygonaceae family include, for example, buckwheat (Fagopyrum esculentum) of the Polygonaceae family.

[0050] The type of meat is not particularly limited. In one embodiment, it includes meat from birds (chicken, duck, etc.), pork, beef, lamb, etc. In one embodiment, it is poultry meat. In one embodiment, it is chicken (Gallus gallus) meat.

[0051] The origin of milk is not particularly limited. In one embodiment, it includes milk from cows, goats, sheep, etc. In one embodiment, it is cow's milk (Bos Taurus). Dairy products are processed milk products. Examples include, but are not limited to, butter, cream, cheese, yogurt, and ice cream.

[0052] Parasites include, but are not limited to, parasites of the Anisakidae family, including Anisakis simplex.

[0053] In one embodiment, the allergen is any of the following: Shrimp, hairy crab (Erimacrus isenbeckii), snow crab (Chionoecetes opilio), blue crab (Portunus trituberculatus), red king crab (Paralithodes camtschaticus), Japanese flying squid (Todarodes pacificus), common octopus (Octopus vulgaris), tuna (Thunnus orientalis), chub mackerel (Scomber japonicas), Pacific cod (Gadus macrocephalus), clam (Ruditapes philippinarum), kiwi (Actinidia deliciosa), pineapple (Ananas comosus), mango (Mangifera indica), cashew nuts (Anacardium occidentale), melon (Cucumis melo), banana (Musa acuminate), orange (Citrus sinensis), apple (Malus domestica), almonds (Prunus dulcis), eggplant (Solanum melongena), tomato (Solanum lycopersicum), cucumber (Cucumis sativus), avocado (Persea americana), Chinese walnut (Juglans regia), mugwort (Artemisia indica var. maximowiczii or Artemisia indica), bread wheat (Triticum aestivum), buckwheat (Fagopyrum esculentum), chicken (Gallus gallus) meat, milk (Bos taurus), and anisakis (Anisakis simplex).

[0054] As used herein, shrimp refers to shrimp belonging to the superorder Eucarida. This includes all species within the order Decapoda (Crabs) except for the infraorder Brachyura (also known as the infraorder Carbidae) and the infraorder Anomura (infraorder Hermit Crabs). Shrimp belonging to the superorder Decapoda may be, for example, shrimp belonging to the families Penaeidae, Spinybrinidae, Nephropsidae, Prionidae, Podocarpus, Palaemonidae, and Euphausiidae. Shrimp belonging to the Penaeidae family may be, for example, whitenamei shrimp, black tiger shrimp, or kuruma prawn; shrimp belonging to the Spinybrinidae family may be, for example, spiny lobster or fan shrimp; and shrimp belonging to the Nephropsidae family may be, for example, lobster. Shrimp belonging to the Primacidae family may be, for example, cherry shrimp, shrimp belonging to the Ponurusidae family may be, for example, white shrimp, shrimp belonging to the Palaemonidae family may be, for example, sweet shrimp, botan shrimp, or striped shrimp, and shrimp belonging to the Euphausiidae family may be, for example, krill. The shrimp that is the subject of the present invention may be any of the above shrimp. Preferably, the shrimp that is the subject of the present invention is a shrimp belonging to the family Panaeidae.

[0055] As used herein, shrimp allergy refers to a state in which an allergic reaction occurs due to an antigen, such as a protein contained in shrimp. Shrimp allergy can occur when a person comes into contact with an antigen contained in shrimp or when the person ingests the antigen. Generally, an allergic reaction that occurs when a food is ingested is specifically referred to as a food allergy. The shrimp allergy may be a food allergy.

[0056] As used herein, an antigen is a substance that induces an allergic reaction. When the antigen is a protein contained in a raw material such as a food ingredient, it is also called an allergen component. The antigen is preferably a protein.

[0057] As used herein, a protein is a molecule having a structure in which natural amino acids are linked by peptide bonds. The number of amino acids contained in a protein is not particularly limited. As used herein, the term "polypeptide" also refers to a molecule having a structure in which natural amino acids are linked by peptide bonds. The number of amino acids contained in a polypeptide is not particularly limited. "Polypeptide" is a concept that includes "protein." Furthermore, a polypeptide in which approximately 2 to 50 amino acids are linked by peptide bonds is sometimes specifically called a peptide.

[0058] When an amino acid has optical isomers, the L-isomer is indicated unless otherwise specified. The notation method for the amino acid sequence of a protein, polypeptide, or peptide used herein is based on standard usage and common practice in the art, and is represented by a single-letter notation for the amino acid, with the left-hand direction being the amino-terminal direction and the right-hand direction being the carboxy-terminal direction. In the single-letter notation for an amino acid, X represents any substance having an amino group and a carboxyl group that can bond to the amino acids at both ends, and in particular any of the 20 naturally occurring amino acids.

[0059] The alanine scanning method (or "alanine glycine scanning" method) is a method for site-specifically identifying residues important for the structure and function of a protein by mutating each residue in the protein to alanine (or glycine if the original amino acid is alanine) to create mutants. If the binding ability to a patient's IgE antibody remains even after mutation to alanine (or glycine if the original amino acid is alanine), then that residue is not important for the binding ability to IgE antibody, and the binding ability will remain even if the residue is changed to another amino acid. The binding ability to IgE antibody refers to the detection of the binding and reaction between the target epitope and the IgE antibody. In the sequence listing of the present application, the residue represented by X is an amino acid residue at a site that remains binding to an IgE antibody of an allergic patient even when substituted with alanine (glycine if the original amino acid is alanine) by alanine glycine scanning as shown in Example 4. It is well known to those skilled in the art that such sites are highly likely to retain the binding ability to the IgE antibody even when substituted with any other amino acid. That is, it is a residue that can be substituted not only with alanine and glycine but also with any amino acid residue other than alanine.

[0060] The binding and maintenance of IgE to antigens (epitopes) is important for subsequent allergic reactions, and this binding and maintenance is mediated by the charge, hydrophobic bonds, hydrogen bonds, and aromatic interactions of the epitope. The fact that binding and maintenance are possible even when these are lost by changing to alanine or glycine means that the amino acid is not important.

[0061] Identification of antigens Proteins contained in shrimp were subjected to two-dimensional electrophoresis under the following conditions to identify the antigens causing shrimp allergies.

[0062] For the first-dimension electrophoresis, the gel length was 5-10 cm, the pH range was 3-10, and the pH gradient of the gel in the direction of migration was 0.15-0.3, 0.4-0.7, and 0.15-0.3, where the total length of the gel is 1, the gel length up to pH 5 is a, the gel length from pH 5 to 7 is b, and the gel length at pH 7 or higher is c. Specifically, isoelectric focusing was performed using IPG Gel Immobiline Drystrip (pH 3-10NL) manufactured by GE Healthcare Biosciences, Inc. (hereinafter abbreviated as GE). The electrophoresis apparatus used was an IPGphor manufactured by GE. The upper limit of the current value of the electrophoresis equipment was set to 75 μA per gel, and the voltage program was as follows: (1) a constant voltage step of 300 V was performed up to 750 Vhr (the current change during the 30 minutes of electrophoresis before the end of this step was 5 μA), (2) the voltage was gradually increased to 1000 V over 300 Vhr, (3) the voltage was further gradually increased to 5000 V over 4500 Vhr, and (4) first-dimension isoelectric focusing was then performed at a constant voltage of 5000 V until the total Vhr reached 12000.

[0063] Second-dimensional electrophoresis was performed using polyacrylamide gels with a gel concentration of 3-6% at the base of the electrophoresis direction and a higher gel concentration at the leading edge of the electrophoresis direction. Specifically, SDS-PAGE was performed using a NuPAGE 4-12% Bris-Tris Gels IPG Well Mini 1mm (Life Technologies). The electrophoresis apparatus used was an XCell SureLock Mini-Cell (Life Technologies). The electrophoresis buffer used was 50 mM MOPS, 50 mM Tris base, 0.1% (w / v) SDS, and 1 mM EDTA, and electrophoresis was performed at a constant voltage of 200 V for approximately 45 minutes.

[0064] As a result, when two-dimensional electrophoresis was performed on the proteins of shrimp (vannamei shrimp (Litopenaeus vannamei), black tiger prawn (Penaeus monodon), and kuruma shrimp (Marsupenaeus japonicas)) under the above conditions, the antigens in the following spots 1 to 11 on the gel were found to specifically bind to the IgE antibodies of patients with shrimp allergies (Figures 2, 4, and 6).

[0065] antigen (1) Spot 1 antigen As a result of performing sequence identification by mass spectrometry for spot 1, the amino acid sequences of SEQ ID NOs: 3 to 43 for vannamei shrimp, SEQ ID NOs: 46 to 85 for black tiger prawn, and SEQ ID NOs: 88 to 115 for kuruma prawn were detected.

[0066] Furthermore, the mass data obtained from the mass spectrometer for spot 1 was analyzed by collating it with protein data from the National Center for Biotechnology Information (NCBI). As a result, SEQ ID NOS: 3 to 43 were identified as the C-terminal portion of myosin heavy chain type 1 derived from vannamei shrimp (amino acid sequence: SEQ ID NOS: 2, nucleotide sequence encoding it: SEQ ID NOS: 1), SEQ ID NOS: 46 to 85 were identified as the C-terminal portion of myosin heavy chain type 1 derived from black tiger prawn (amino acid sequence: SEQ ID NOS: 45, nucleotide sequence encoding it: SEQ ID NOS: 44), and SEQ ID NOS: 88 to 115 were identified as the C-terminal portion of myosin heavy chain type a derived from kuruma prawn (amino acid sequence: SEQ ID NOS: 87, nucleotide sequence encoding it: SEQ ID NOS: 86).

[0067] Therefore, in the present application, the antigen of spot 1 may be any of the following (1A-a) to (1A-e) and (1B). (1A-a) A protein comprising the amino acid sequence of SEQ ID NO: 2, 45 or 87 in which one or several amino acids have been deleted, substituted, inserted or added. (1A-b) A protein comprising an amino acid sequence that is 70% or more, preferably 75% or more, 80% or more, 85% or more, 90% or more, 95% or more, 97% or more, 98% or more, or 99% or more identical to the amino acid sequence shown in SEQ ID NO: 2, 45, or 87. (1A-c) A protein comprising an amino acid sequence encoded by a base sequence in which one or several nucleotides in SEQ ID NO: 1, 44 or 86 have been deleted, substituted, inserted or added. (1A-d) A protein comprising an amino acid sequence encoded by a nucleotide sequence having 70% or more, preferably 75% or more, 80% or more, 85% or more, 90% or more, 95% or more, 97% or more, 98% or more, or 99% or more identity to the nucleotide sequence shown in SEQ ID NO: 1, 44, or 86. (1A-e) A protein comprising an amino acid sequence encoded by a nucleic acid that hybridizes under stringent conditions to a nucleic acid consisting of a nucleotide sequence complementary to the nucleotide sequence shown in SEQ ID NO: 1, 44, or 86. (1B) A protein comprising at least one amino acid sequence selected from the group consisting of SEQ ID NOs: 2 to 43, 45 to 85, or 87 to 115, preferably a protein comprising at least 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, 25, 30, 45, 50, or all of the amino acid sequences. More preferably, a protein comprising at least one amino acid sequence selected from the group consisting of SEQ ID NOs: 2 to 5, 7 to 13, 16 to 24, and 26 to 43, the group consisting of SEQ ID NOs: 45, 46, 48, 49, 51 to 55, 58 to 66, and 68 to 85, or the group consisting of SEQ ID NOs: 87 to 90 and 92 to 115, preferably a protein comprising at least 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, 25, 30, 45, or all of the amino acid sequences. Here, the amino acid sequences shown in any of SEQ ID NOs: 2 to 43, 45 to 85, and 87 to 115 may have one or several amino acids deleted, substituted, inserted, or added.

[0068] The above proteins (1A-a) to (1A-e) and (1B) may be proteins that appear as spots in a gel when two-dimensional electrophoresis is performed under the conditions described in the above section "Identification of antigens" with a molecular weight of 80 to 260 kDa, preferably a molecular weight of around 90 to 230 kDa, and more preferably 100 to 200 kDa, and an isoelectric point of 3.0 to 7.0, preferably 4.0 to 6.5, and more preferably 5.0 to 6.0.

[0069] (2) Spot 2 antigen As a result of performing sequence identification by mass spectrometry for spot 2, the amino acid sequences of SEQ ID NOs: 118 to 139 for vannamei shrimp, SEQ ID NOs: 142 to 144 for black tiger shrimp, and SEQ ID NOs: 147 to 159 for kuruma prawn were detected.

[0070] Furthermore, mass data obtained from the mass spectrometer for spot 2 was analyzed by collating it with NCBI protein data. As a result, SEQ ID NOS: 118 to 139 were identified as the N-terminal portion of myosin heavy chain type 1 derived from vannamei shrimp (amino acid sequence: SEQ ID NOS: 117, nucleotide sequence encoding it: SEQ ID NOS: 116), SEQ ID NOS: 142 to 144 were identified as the N-terminal portion of myosin heavy chain type 1 derived from black tiger prawn (amino acid sequence: SEQ ID NOS: 141, nucleotide sequence encoding it: SEQ ID NOS: 140), and SEQ ID NOS: 147 to 159 were identified as the N-terminal portion of myosin heavy chain type a derived from kuruma prawn (amino acid sequence: SEQ ID NOS: 146, nucleotide sequence encoding it: SEQ ID NOS: 145).

[0071] Therefore, in the present application, the antigen of spot 2 may be any of the following (2A-a) to (2A-e) and (2B). (2A-a) A protein comprising an amino acid sequence in which one or several amino acids are deleted, substituted, inserted or added in SEQ ID NO: 117, 141 or 146. (2A-b) A protein comprising an amino acid sequence that has an identity of 70% or more, preferably 75% or more, 80% or more, 85% or more, 90% or more, 95% or more, 97% or more, 98% or more, or 99% or more to the amino acid sequence shown in SEQ ID NO: 117, 141, or 146. (2A-c) A protein comprising an amino acid sequence encoded by a base sequence in which one or several nucleotides in SEQ ID NO: 116, 140 or 145 have been deleted, substituted, inserted or added. (2A-d) A protein comprising an amino acid sequence encoded by a nucleotide sequence that has an identity of 70% or more, preferably 75% or more, 80% or more, 85% or more, 90% or more, 95% or more, 97% or more, 98% or more, or 99% or more to the nucleotide sequence shown in SEQ ID NO: 116, 140, or 145. (2A-e) A protein comprising an amino acid sequence encoded by a nucleic acid that hybridizes under stringent conditions with a nucleic acid consisting of a base sequence complementary to the base sequence shown in SEQ ID NO: 116, 140 or 145. (2B) A protein comprising at least one amino acid sequence selected from the group consisting of SEQ ID NOs: 117 to 139, 141 to 144, or 146 to 159, preferably at least 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, or all of the amino acid sequences. More preferably, a protein comprising at least one amino acid sequence selected from the group consisting of SEQ ID NOs: 117 to 126, 128 to 132, 136, 138, and 139, 141 to 144, or 146 to 159, preferably at least 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, or all of the amino acid sequences. The amino acid sequence represented by any of SEQ ID NOs: 117 to 139, 141 to 144, and 146 to 159 may have one or more amino acids deleted, substituted, inserted, or added.

[0072] The above proteins (2A-a) to (2A-e) and (2B) may be proteins that appear as spots in a gel when two-dimensional electrophoresis is performed under the conditions described in the above section "Identification of antigens" with a molecular weight of 50 to 160 kDa, preferably a molecular weight of around 55 to 150 kDa, more preferably 60 to 130 kDa, and an isoelectric point of 4.5 to 10.0, preferably 5.0 to 9.5, more preferably 5.5 to 9.0.

[0073] (3) Spot 3 antigen Spot 3 was subjected to sequence identification by mass spectrometry, and as a result, the amino acid sequences of SEQ ID NOs: 162 to 177 for vannamei shrimp, SEQ ID NOs: 180 to 228 for black tiger prawn, and SEQ ID NOs: 231 to 274 for kuruma prawn were detected.

[0074] Furthermore, mass data obtained from the mass spectrometer for spot 3 was analyzed by collating it with NCBI protein data. As a result, SEQ ID NOS: 162 to 177 were identified as the C-terminal portion of myosin heavy chain type 2 derived from whitenamei shrimp (amino acid sequence: SEQ ID NOS: 161, nucleotide sequence encoding it: SEQ ID NOS: 160), SEQ ID NOS: 180 to 228 were identified as the C-terminal portion of myosin heavy chain type 2 derived from black tiger prawn (amino acid sequence: SEQ ID NOS: 179, nucleotide sequence encoding it: SEQ ID NOS: 178), and SEQ ID NOS: 231 to 274 were identified as the C-terminal portion of myosin heavy chain type b derived from kuruma prawn (amino acid sequence: SEQ ID NOS: 230, nucleotide sequence encoding it: SEQ ID NOS: 229).

[0075] Therefore, in the present application, the antigen of spot 3 may be any of the following (3A-a) to (3A-e) and (3B). (3A-a) A protein comprising an amino acid sequence in which one or several amino acids are deleted, substituted, inserted or added in SEQ ID NO: 161, 179 or 230. (3A-b) A protein comprising an amino acid sequence that has an identity of 70% or more, preferably 75% or more, 80% or more, 85% or more, 90% or more, 95% or more, 97% or more, 98% or more, or 99% or more to the amino acid sequence shown in SEQ ID NO: 161, 179, or 230. (3A-c) A protein comprising an amino acid sequence encoded by a base sequence in which one or several nucleotides in SEQ ID NO: 160, 178 or 229 have been deleted, substituted, inserted or added. (3A-d) A protein comprising an amino acid sequence encoded by a nucleotide sequence that has an identity of 70% or more, preferably 75% or more, 80% or more, 85% or more, 90% or more, 95% or more, 97% or more, 98% or more, or 99% or more to the nucleotide sequence shown in SEQ ID NO: 160, 178, or 229. (3A-e) A protein comprising an amino acid sequence encoded by a nucleic acid that hybridizes under stringent conditions with a nucleic acid consisting of a nucleotide sequence complementary to the nucleotide sequence shown in SEQ ID NO: 160, 178, or 229. (3B) A protein comprising at least one amino acid sequence selected from the group consisting of SEQ ID NOs: 161 to 177, the group consisting of SEQ ID NOs: 179 to 228, or the group consisting of SEQ ID NOs: 230 to 274, preferably a protein comprising at least 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, 25, 30, 35, 40, 45 or all of the amino acid sequences. More preferably, the protein comprises at least one amino acid sequence selected from the group consisting of SEQ ID NOs: 161, 162, and 164 to 177, the group consisting of SEQ ID NOs: 179 to 181, 183 to 186, 188 to 190, 192 to 212, 214 to 216, and 218 to 228, and the group consisting of SEQ ID NOs: 230 to 233, 235 to 240, 242, 244 to 258, 260 to 262, and 264 to 274, and preferably a protein comprising at least 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, 25, 30, 35, 40, or all of the amino acid sequences. In this regard, the amino acid sequence represented by any of SEQ ID NOs: 161 to 177, 179 to 228, and 230 to 274 may have one or more amino acids deleted, substituted, inserted, or added.

[0076] The above proteins (3A-a) to (3A-e) and (3B) may be proteins that appear as spots in a gel when two-dimensional electrophoresis is performed under the conditions described in the above section "Identification of antigens" with a molecular weight of 80 to 260 kDa, preferably a molecular weight of around 90 to 230 kDa, and more preferably 100 to 200 kDa, and an isoelectric point of 3.0 to 7.0, preferably 4.0 to 6.5, and more preferably 5.0 to 6.0.

[0077] (4) Spot 4 antigen As a result of performing sequence identification by mass spectrometry for spot 4, the amino acid sequences of SEQ ID NOs: 277 to 298 for vannamei shrimp, SEQ ID NOs: 301 to 304 for black tiger prawn, and SEQ ID NOs: 307 to 320 for kuruma prawn were detected.

[0078] Furthermore, mass data obtained from the mass spectrometer for spot 4 was analyzed by collating it with NCBI protein data. As a result, SEQ ID NOs: 277 to 298 were identified as the N-terminal portion of myosin heavy chain type 2 derived from vannamei shrimp (amino acid sequence: SEQ ID NO: 276, nucleotide sequence encoding it: SEQ ID NO: 275), SEQ ID NOs: 301 to 304 were identified as the N-terminal portion of myosin heavy chain type 2 derived from black tiger prawn (amino acid sequence: SEQ ID NO: 300, nucleotide sequence encoding it: SEQ ID NO: 299), and SEQ ID NOs: 307 to 320 were identified as the N-terminal portion of myosin heavy chain type b derived from kuruma prawn (amino acid sequence: SEQ ID NO: 306, nucleotide sequence encoding it: SEQ ID NO: 305).

[0079] Therefore, in the present application, the antigen of spot 4 may be any of the following (4A-a) to (4A-e) and (4B). (4A-a) A protein comprising an amino acid sequence in which one or several amino acids are deleted, substituted, inserted or added in SEQ ID NO: 276, 300 or 306. (4A-b) A protein containing an amino acid sequence that has an identity of 70% or more, preferably 75% or more, 80% or more, 85% or more, 90% or more, 95% or more, 97% or more, 98% or more, or 99% or more to the amino acid sequence shown in SEQ ID NO: 276, 300, or 306. (4A-c) A protein comprising an amino acid sequence encoded by a base sequence in which one or several nucleotides in SEQ ID NO: 275, 299 or 305 have been deleted, substituted, inserted or added. (4A-d) A protein containing an amino acid sequence encoded by a nucleotide sequence that has an identity of 70% or more, preferably 75% or more, 80% or more, 85% or more, 90% or more, 95% or more, 97% or more, 98% or more, or 99% or more to the nucleotide sequence shown in SEQ ID NO: 275, 299, or 305. (4A-e) A protein comprising an amino acid sequence encoded by a nucleic acid that hybridizes under stringent conditions with a nucleic acid consisting of a nucleotide sequence complementary to the nucleotide sequence shown in SEQ ID NO: 275, 299, or 305. (4B) A protein comprising at least one amino acid sequence selected from the group consisting of SEQ ID NOs: 276 to 298, 300 to 304, or 306 to 320, preferably at least 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, or all of the amino acid sequences. More preferably, a protein comprising at least one amino acid sequence selected from the group consisting of SEQ ID NOs: 276 to 293 and 295 to 298, 300 to 304, or 306, 307, and 309 to 319, preferably at least 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, or all of the amino acid sequences. In this regard, the amino acid sequence represented by any of SEQ ID NOs: 276 to 298, 300 to 304, and 306 to 320 may have one or more amino acids deleted, substituted, inserted, or added.

[0080] The above proteins (4A-a) to (4A-e) and (4B) may be proteins that appear as spots in a gel when two-dimensional electrophoresis is performed under the conditions described in the above section "Identification of antigens" with a molecular weight of 50 to 160 kDa, preferably a molecular weight of around 55 to 150 kDa, more preferably 60 to 130 kDa, and an isoelectric point of 4.5 to 10.0, preferably 5.0 to 9.5, more preferably 5.5 to 9.0.

[0081] (5) Spot 5 antigen Spot 5 was subjected to sequence identification by mass spectrometry, and as a result, the amino acid sequences of SEQ ID NOs: 321 to 336 for vannamei shrimp, SEQ ID NOs: 337 to 360 for black tiger prawn, and SEQ ID NOs: 363 to 379 for kuruma prawn were detected.

[0082] Furthermore, mass data obtained from the mass spectrometer for spot 5 was analyzed by collating it with NCBI protein data. As a result, SEQ ID NOs: 363 to 379 were identified as kuruma shrimp glycogen phosphorylase (amino acid sequence: SEQ ID NO: 362, nucleotide sequence encoding it: SEQ ID NO: 361). SEQ ID NOs: 321 to 336 and 337 to 360 also matched kuruma shrimp glycogen phosphorylase. In other words, since proteins with high homology were detected in vannamei shrimp and black tiger shrimp, it was determined that glycogen phosphorylase or its homologs were shrimp antigens.

[0083] Therefore, in the present application, the antigen of spot 5 may be any of the following (5A-a) to (5A-e) and (5B). (5A-a) A protein comprising an amino acid sequence in which one or several amino acids are deleted, substituted, inserted or added in SEQ ID NO: 362. (5A-b) A protein containing an amino acid sequence that has an identity of 70% or more, preferably 75% or more, 80% or more, 85% or more, 90% or more, 95% or more, 97% or more, 98% or more, or 99% or more to the amino acid sequence shown in SEQ ID NO: 362. (5A-c) A protein comprising an amino acid sequence encoded by a base sequence in which one or more nucleotides in SEQ ID NO: 361 have been deleted, substituted, inserted or added. (5A-d) A protein containing an amino acid sequence encoded by a nucleotide sequence that has an identity of 70% or more, preferably 75% or more, 80% or more, 85% or more, 90% or more, 95% or more, 97% or more, 98% or more, or 99% or more to the nucleotide sequence shown in SEQ ID NO: 361. (5A-e) A protein comprising an amino acid sequence encoded by a nucleic acid that hybridizes under stringent conditions with a nucleic acid consisting of a base sequence complementary to the base sequence shown in SEQ ID NO: 361. (5B) A protein comprising at least one amino acid sequence selected from the group consisting of SEQ ID NOs: 321 to 336, 337 to 360, or 362 to 379, preferably a protein comprising at least 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, or all of the amino acid sequences. More preferably, a protein comprising at least one amino acid sequence selected from the group consisting of SEQ ID NOs: 322 to 326, 329 to 330, and 332 to 336, the group consisting of SEQ ID NOs: 338 to 343, 345, 347 to 349, and 352 to 360, and the group consisting of SEQ ID NOs: 362 to 367, 369, 371, and 373 to 379, preferably a protein comprising at least 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, or all of the amino acid sequences. Here, the amino acid sequences shown in any of SEQ ID NOs: 321 to 336, 337 to 360, and 362 to 379 may have one or several amino acids deleted, substituted, inserted, or added.

[0084] The above proteins (5A-a) to (5A-e) and (5B) may be proteins that appear as spots in a gel when two-dimensional electrophoresis is performed under the conditions described in the above section "Identification of antigens" with a molecular weight of 70 to 160 kDa, preferably a molecular weight of around 75 to 140 kDa, and more preferably 80 to 130 kDa, and an isoelectric point of 5.0 to 9.0, preferably 5.5 to 8.5, and more preferably 6.0 to 8.0.

[0085] (6) Spot 6 antigen Spot 6 was subjected to sequence identification by mass spectrometry, and as a result, the amino acid sequences of SEQ ID NOs: 382 to 397 for vannamei shrimp, SEQ ID NOs: 400 to 412 for black tiger prawn, and SEQ ID NOs: 415 to 419 for kuruma prawn were detected.

[0086] Furthermore, mass data obtained from the mass spectrometer for spot 6 was analyzed by collating it with NCBI protein data. As a result, SEQ ID NOs: 382 to 397 were identified as a portion of hemocyanin subunit L1 derived from whitenamei shrimp (amino acid sequence: SEQ ID NO: 381, nucleotide sequence encoding it: SEQ ID NO: 380), SEQ ID NOs: 400 to 412 were identified as hemocyanin derived from black tiger prawn (amino acid sequence: SEQ ID NO: 399, nucleotide sequence encoding it: SEQ ID NO: 398), and SEQ ID NOs: 415 to 419 were identified as hemocyanin subunit L derived from kuruma shrimp (amino acid sequence: SEQ ID NO: 414, nucleotide sequence encoding it: SEQ ID NO: 413).

[0087] Therefore, in the present application, the antigen of spot 6 may be any of the following (6A-a) to (6A-e) and (6B). (6A-a) A protein comprising an amino acid sequence in which one or several amino acids are deleted, substituted, inserted or added in SEQ ID NO: 381, 399 or 414. (6A-b) A protein containing an amino acid sequence that has an identity of 70% or more, preferably 75% or more, 80% or more, 85% or more, 90% or more, 95% or more, 97% or more, 98% or more, or 99% or more to the amino acid sequence shown in SEQ ID NO: 381, 399, or 414. (6A-c) A protein comprising an amino acid sequence encoded by a base sequence in which one or several nucleotides in SEQ ID NO: 380, 398 or 413 have been deleted, substituted, inserted or added. (6A-d) A protein containing an amino acid sequence encoded by a nucleotide sequence that has an identity of 70% or more, preferably 75% or more, 80% or more, 85% or more, 90% or more, 95% or more, 97% or more, 98% or more, or 99% or more to the nucleotide sequence shown in SEQ ID NO: 380, 398, or 413. (6A-e) A protein comprising an amino acid sequence encoded by a nucleic acid that hybridizes under stringent conditions with a nucleic acid consisting of a nucleotide sequence complementary to the nucleotide sequence shown in SEQ ID NO: 380, 398 or 413. (6B) A protein comprising at least one amino acid sequence selected from the group consisting of SEQ ID NOs: 381 to 397, 399 to 412, or 414 to 419, preferably a protein comprising at least 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, or all of the amino acid sequences. More preferably, a protein comprising at least one amino acid sequence selected from the group consisting of SEQ ID NOs: 381 to 384 and 386 to 397, 399 to 402, 404, and 407 to 412, or 414 to 416, 418, and 419, preferably a protein comprising at least 2, 3, 4, 5, 6, 7, 8, 9, 10, or all of the amino acid sequences. The amino acid sequence represented by any of SEQ ID NOs: 381 to 397, 399 to 412, and 414 to 419 may have one or more amino acids deleted, substituted, inserted, or added.

[0088] The above proteins (6A-a) to (6A-e) and (6B) may be proteins that appear as spots in a gel when two-dimensional electrophoresis is performed under the conditions described in the above section "Identification of antigens" with a molecular weight of 50 to 110 kDa, preferably a molecular weight of around 55 to 100 kDa, more preferably 60 to 90 kDa, and an isoelectric point of 4.0 to 7.0, preferably 4.5 to 6.5, more preferably 5.0 to 6.0.

[0089] (7) Spot 7 antigen Spot 7 was subjected to sequence identification by mass spectrometry, and as a result, the amino acid sequences of SEQ ID NOs: 422 to 435 for vannamei shrimp and SEQ ID NOs: 436 to 441 for black tiger shrimp were detected.

[0090] Furthermore, for spot 7, the mass data obtained from the mass spectrometer was analyzed by collating it with NCBI protein data. As a result, SEQ ID NOs: 422 to 435 were identified as pyruvate kinase 3 derived from vannamei shrimp (amino acid sequence: SEQ ID NO: 421, nucleotide sequence encoding same: SEQ ID NO: 420). SEQ ID NOs: 436 to 441 also matched pyruvate kinase 3 derived from vannamei shrimp. In other words, since proteins with high homology were detected in black tiger shrimp, it was determined that pyruvate kinase 3 or its homologs are shrimp antigens.

[0091] Therefore, in the present application, the antigen of spot 7 may be any of the following (7A-a) to (7A-e) and (7B). (7A-a) A protein comprising an amino acid sequence in which one or several amino acids are deleted, substituted, inserted or added in SEQ ID NO: 421. (7A-b) A protein containing an amino acid sequence that has an identity of 70% or more, preferably 75% or more, 80% or more, 85% or more, 90% or more, 95% or more, 97% or more, 98% or more, or 99% or more to the amino acid sequence shown in SEQ ID NO: 421. (7A-c) A protein comprising an amino acid sequence encoded by a base sequence in which one or more nucleotides in SEQ ID NO: 420 have been deleted, substituted, inserted or added. (7A-d) A protein containing an amino acid sequence encoded by a nucleotide sequence that has an identity of 70% or more, preferably 75% or more, 80% or more, 85% or more, 90% or more, 95% or more, 97% or more, 98% or more, or 99% or more to the nucleotide sequence shown in SEQ ID NO: 420. (7A-e) A protein comprising an amino acid sequence encoded by a nucleic acid that hybridizes under stringent conditions with a nucleic acid consisting of a base sequence complementary to the base sequence shown in SEQ ID NO: 420. (7B) A protein comprising at least one amino acid sequence selected from the group consisting of SEQ ID NOs: 421 to 435 or the group consisting of SEQ ID NOs: 436 to 441, preferably a protein comprising at least 2, 3, 4, 5, 6, 7, 8, 9, 10, or all of the amino acid sequences. In this regard, the amino acid sequence represented by any of SEQ ID NOs: 421 to 435 and 436 to 441 may have one or several amino acids deleted, substituted, inserted, or added.

[0092] The above proteins (7A-a) to (7A-e) and (7B) may be proteins that appear as spots with a molecular weight of 45 to 80 kDa, preferably around 50 to 75 kDa, more preferably 55 to 70 kDa, and an isoelectric point of 4.5 to 9.0, preferably 5.0 to 8.5, more preferably 5.5 to 8.0, in a gel when two-dimensional electrophoresis is performed under the conditions described in the above section "Identification of antigen."

[0093] (8) Spot 8 antigen Spot 8 was subjected to sequence identification by mass spectrometry, and as a result, the amino acid sequences of SEQ ID NOs: 442 to 453 for vannamei shrimp, SEQ ID NOs: 456 to 479 for black tiger shrimp, and SEQ ID NOs: 482 to 484 for kuruma prawn were detected.

[0094] Furthermore, for spot 8, the mass data obtained from the mass spectrometer was analyzed by collating it with NCBI protein data. As a result, SEQ ID NOs: 456 to 479 were identified as phosphopyruvate hydratase derived from black tiger prawn (amino acid sequence: SEQ ID NO: 455, nucleotide sequence encoding it: SEQ ID NO: 454). SEQ ID NOs: 442 to 453 also matched phosphopyruvate hydratase derived from black tiger prawn. In other words, since a protein with high homology in vannamei shrimp was detected, it was determined that phosphopyruvate hydratase or its homolog is a shrimp antigen. SEQ ID NOs: 482 to 484 were identified as phosphopyruvate hydratase derived from kuruma prawn (amino acid sequence: SEQ ID NO: 481, nucleotide sequence encoding it: SEQ ID NO: 480).

[0095] Therefore, in the present application, the antigen of spot 8 may be any of the following (8A-a) to (8A-e) and (8B). (8A-a) A protein comprising an amino acid sequence in which one or several amino acids are deleted, substituted, inserted or added in SEQ ID NO: 455 or 481. (8A-b) A protein containing an amino acid sequence having an identity of 70% or more, preferably 75% or more, 80% or more, 85% or more, 90% or more, 95% or more, 97% or more, 98% or more, or 99% or more to the amino acid sequence shown in SEQ ID NO: 455 or 481. (8A-c) A protein comprising an amino acid sequence encoded by a base sequence in which one or several nucleotides in SEQ ID NO: 454 or 480 have been deleted, substituted, inserted or added. (8A-d) A protein comprising an amino acid sequence encoded by a nucleotide sequence having 70% or more, preferably 75% or more, 80% or more, 85% or more, 90% or more, 95% or more, 97% or more, 98% or more, or 99% or more identity to the nucleotide sequence shown in SEQ ID NO: 454 or 480. (8A-e) A protein comprising an amino acid sequence encoded by a nucleic acid that hybridizes under stringent conditions to a nucleic acid consisting of a nucleotide sequence complementary to the nucleotide sequence shown in SEQ ID NO: 454 or 480. (8B) A protein comprising at least one amino acid sequence selected from the group consisting of SEQ ID NOs: 442 to 453, 455 to 479, or 481 to 484, preferably a protein comprising at least 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, or all of the amino acid sequences. More preferably, a protein comprising at least one amino acid sequence selected from the group consisting of SEQ ID NOs: 442 to 450, 452, and 453, the group consisting of SEQ ID NOs: 455 to 459, 463 to 469, 471 to 473, and 476 to 479, or the group consisting of SEQ ID NOs: 481 to 484, preferably a protein comprising at least 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, or all of the amino acid sequences. Here, the amino acid sequences shown in any of SEQ ID NOs: 442 to 453, 455 to 479, and 481 to 484 may have one or several amino acids deleted, substituted, inserted, or added.

[0096] The above proteins (8A-a) to (8A-e) and (8B) may be proteins that appear as spots with a molecular weight of 35 to 80 kDa, preferably a molecular weight of around 40 to 75 kDa, more preferably 45 to 70 kDa, and an isoelectric point of 4.0 to 8.0, preferably 4.5 to 7.5, more preferably 5.0 to 7.0, in a gel when two-dimensional electrophoresis is performed under the conditions described in the above section "Identification of antigen."

[0097] (9) Spot 9 antigen Spot 9 was subjected to sequence identification by mass spectrometry, and as a result, the amino acid sequences of SEQ ID NOs: 487 to 490 for vannamei shrimp, SEQ ID NOs: 491 to 497 for black tiger shrimp, and SEQ ID NOs: 498 to 518 for kuruma prawn were detected.

[0098] Furthermore, mass data obtained from the mass spectrometer for spot 9 was analyzed by collating it with NCBI protein data. As a result, SEQ ID NOs: 487 to 490 were identified as a mitochondrial ATP synthase subunit alpha precursor derived from vannamei shrimp (amino acid sequence: SEQ ID NO: 486, encoding nucleotide sequence: SEQ ID NO: 485). Furthermore, SEQ ID NOs: 491 to 497 and SEQ ID NOs: 498 to 518 also matched a mitochondrial ATP synthase subunit alpha precursor derived from vannamei shrimp. In other words, because proteins with high homology were detected in black tiger prawn and kuruma prawn, it was determined that a mitochondrial ATP synthase subunit alpha precursor or its homolog is a shrimp antigen.

[0099] Therefore, in the present application, the antigen of spot 9 may be any of the following (9A-a) to (9A-e) and (9B). (9A-a) A protein comprising an amino acid sequence in which one or several amino acids are deleted, substituted, inserted or added in SEQ ID NO: 486. (9A-b) A protein containing an amino acid sequence that has an identity of 70% or more, preferably 75% or more, 80% or more, 85% or more, 90% or more, 95% or more, 97% or more, 98% or more, or 99% or more to the amino acid sequence shown in SEQ ID NO: 486. (9A-c) A protein comprising an amino acid sequence encoded by a base sequence in which one or more nucleotides in SEQ ID NO: 485 have been deleted, substituted, inserted or added. (9A-d) A protein containing an amino acid sequence encoded by a nucleotide sequence that has an identity of 70% or more, preferably 75% or more, 80% or more, 85% or more, 90% or more, 95% or more, 97% or more, 98% or more, or 99% or more to the nucleotide sequence shown in SEQ ID NO: 485. (9A-e) A protein comprising an amino acid sequence encoded by a nucleic acid that hybridizes under stringent conditions with a nucleic acid consisting of a base sequence complementary to the base sequence shown in SEQ ID NO: 485. (9B) A protein comprising at least one amino acid sequence selected from the group consisting of SEQ ID NOs: 486 to 490, 491 to 497, or 498 to 518, preferably a protein comprising at least 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, or all of the amino acid sequences. More preferably, a protein comprising at least one amino acid sequence selected from the group consisting of SEQ ID NOs: 486 to 490, 491 to 497, or the group consisting of SEQ ID NOs: 498 to 501, 503 to 507, 509, and 512 to 518, preferably a protein comprising at least 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, or all of the amino acid sequences. In this regard, the amino acid sequence represented by any of SEQ ID NOs: 486 to 490, 491 to 497, and 498 to 518 may have one or more amino acids deleted, substituted, inserted, or added.

[0100] The above proteins (9A-a) to (9A-e) and (9B) may be proteins that appear as spots with a molecular weight of 40 to 70 kDa, preferably around 45 to 65 kDa, more preferably 50 to 60 kDa, and an isoelectric point of 5.0 to 10.0, preferably 5.5 to 9.5, more preferably 6.0 to 9.0, in a gel when two-dimensional electrophoresis is performed under the conditions described in the above section "Identification of antigen."

[0101] (10) Spot 10 antigen As a result of performing sequence identification by mass spectrometry for spot 10, the amino acid sequences of SEQ ID NOs: 521 to 527 for vannamei shrimp, SEQ ID NOs: 528 to 532 for black tiger prawn, and SEQ ID NOs: 533 to 539 for kuruma prawn were detected.

[0102] Furthermore, for spot 10, the mass data obtained from the mass spectrometer was analyzed by collating it with NCBI protein data. As a result, SEQ ID NOs: 521 to 527 were identified as vannamei shrimp-derived troponin I (amino acid sequence: SEQ ID NO: 520, nucleotide sequence encoding it: SEQ ID NO: 519). SEQ ID NOs: 528 to 532 and 533 to 539 also matched vannamei shrimp-derived troponin I. In other words, since proteins highly homologous to black tiger prawn and kuruma prawn were detected, it was determined that troponin I and its homologs are shrimp antigens.

[0103] Therefore, in the present application, the antigen of spot 10 may be any of the following (10A-a) to (10A-e) and (10B). (10A-a) A protein comprising an amino acid sequence in which one or several amino acids are deleted, substituted, inserted or added in SEQ ID NO: 520. (10A-b) A protein comprising an amino acid sequence that has an identity of 70% or more, preferably 75% or more, 80% or more, 85% or more, 90% or more, 95% or more, 97% or more, 98% or more, or 99% or more to the amino acid sequence set forth in SEQ ID NO: 520. (10A-c) A protein comprising an amino acid sequence encoded by a base sequence in which one or more nucleotides in SEQ ID NO: 519 have been deleted, substituted, inserted or added. (10A-d) A protein comprising an amino acid sequence encoded by a nucleotide sequence that has an identity of 70% or more, preferably 75% or more, 80% or more, 85% or more, 90% or more, 95% or more, 97% or more, 98% or more, or 99% or more to the nucleotide sequence shown in SEQ ID NO: 519. (10A-e) A protein comprising an amino acid sequence encoded by a nucleic acid that hybridizes under stringent conditions with a nucleic acid consisting of a base sequence complementary to the base sequence shown in SEQ ID NO: 519. (10B) A protein comprising at least one amino acid sequence selected from the group consisting of SEQ ID NOs: 520 to 527, 528 to 532, or 533 to 539, preferably a protein comprising at least two, three, four, five, six, or all of the amino acid sequences. More preferably, a protein comprising at least one amino acid sequence selected from the group consisting of SEQ ID NOs: 520 to 525 and 527, 528 to 530 and 532, or 533 to 537 and 539, preferably a protein comprising at least two, three, four, five, or all of the amino acid sequences. Here, the amino acid sequence represented by any of SEQ ID NOs: 520 to 527, 528 to 532, and 533 to 539 may have one or more amino acids deleted, substituted, inserted, or added.

[0104] The above proteins (10A-a) to (10A-e) and (10B) may be proteins that appear as spots in a gel when two-dimensional electrophoresis is performed under the conditions described in the above section "Identification of antigens" with a molecular weight of 10 to 50 kDa, preferably a molecular weight of around 15 to 40 kDa, more preferably 20 to 40 kDa, and an isoelectric point of 7.0 to 11.0, preferably 7.5 to 10.5, more preferably 8.0 to 10.0.

[0105] (11) Spot 11 antigen As a result of performing sequence identification by mass spectrometry for spot 11, the amino acid sequences of SEQ ID NOs: 542 to 547 for vannamei shrimp, SEQ ID NOs: 550 to 553 for black tiger prawn, and SEQ ID NOs: 554 to 557 for kuruma prawn were detected.

[0106] Furthermore, for spot 11, the mass data obtained from the mass spectrometer was analyzed by collating it with the protein data from NCBI. As a result, SEQ ID NOs: 542 to 547 were identified as cyclophilin A derived from vannamei shrimp (amino acid sequence: SEQ ID NO: 541, nucleotide sequence encoding it: SEQ ID NO: 540). SEQ ID NOs: 554 to 557 also yielded hits for cyclophilin A derived from vannamei shrimp. In other words, since proteins highly homologous to kuruma shrimp were detected, it was determined that cyclophilin A or its homologs are shrimp antigens. SEQ ID NOs: 550 to 553 were identified as cyclophilin A derived from black tiger prawn (amino acid sequence: SEQ ID NO: 549, nucleotide sequence encoding it: SEQ ID NO: 548).

[0107] Therefore, in the present application, the antigen of spot 11 may be any of the following (11A-a) to (11A-e) and (11B). (11A-a) A protein comprising an amino acid sequence in which one or several amino acids are deleted, substituted, inserted or added in SEQ ID NO: 541 or 549. (11A-b) A protein containing an amino acid sequence that has an identity of 70% or more, preferably 75% or more, 80% or more, 85% or more, 90% or more, 95% or more, 97% or more, 98% or more, or 99% or more to the amino acid sequence shown in SEQ ID NO: 541 or 549. (11A-c) A protein comprising an amino acid sequence encoded by a nucleotide sequence in which one or several nucleotides have been deleted, substituted, inserted or added in SEQ ID NO: 540 or 548. (11A-d) A protein comprising an amino acid sequence encoded by a nucleotide sequence that has 70% or more, preferably 75% or more, 80% or more, 85% or more, 90% or more, 95% or more, 97% or more, 98% or more, or 99% or more identity to the nucleotide sequence shown in SEQ ID NO: 540 or 548. (11A-e) A protein comprising an amino acid sequence encoded by a nucleic acid that hybridizes under stringent conditions with a nucleic acid consisting of a base sequence complementary to the base sequence shown in SEQ ID NO: 540 or 548. (11B) A protein comprising at least one amino acid sequence selected from the group consisting of SEQ ID NOs: 541 to 547, 549 to 553, or 554 to 557, preferably a protein comprising at least two, three, four, five, or all of the amino acid sequences. More preferably, a protein comprising at least one amino acid sequence selected from the group consisting of SEQ ID NOs: 541 to 543, 545, and 546, 549 to 553, or 554 to 556, preferably a protein comprising at least two, three, or all of the amino acid sequences. Here, the amino acid sequence represented by any of SEQ ID NOs: 541 to 547, 549 to 553, and 554 to 557 may have one or more amino acids deleted, substituted, inserted, or added.

[0108] The above proteins (11A-a) to (11A-e) and (11B) may be proteins that appear as spots in a gel when two-dimensional electrophoresis is performed under the conditions described in the above section "Identification of antigens" with a molecular weight of 10 to 30 kDa, preferably a molecular weight of around 13 to 25 kDa, more preferably 15 to 20 kDa, and an isoelectric point of 7.0 to 11.0, preferably 7.5 to 10.5, more preferably 8.0 to 10.0.

[0109] The antigenic proteins (1) to (11) above and the polypeptides (E1) to (E50) described below also include those in which amino acid residues of the proteins or polypeptides have been modified by phosphorylation, glycosylation, aminoacylation, ring-opening, deamination, or the like.

[0110] Preferably, the antigenic proteins (1) to (11) above and the polypeptides (E1) to (E50) described below are allergens.

[0111] As used herein, the phrase "one or several amino acids have been deleted, substituted, inserted, or added" in reference to an amino acid sequence refers to an amino acid sequence in which one or several amino acids have been deleted, substituted, inserted, and / or added. The term "several amino acids" refers to, but is not limited to, 200 or fewer, 100 or fewer, 50 or fewer, 30 or fewer, 20 or fewer, 15 or fewer, 12 or fewer, 10 or fewer, 8 or fewer, 6 or fewer, 4 or fewer, or 3 or fewer amino acids. Alternatively, the term "several amino acids" refers to 30%, preferably 25%, 20%, 15%, 10%, 5%, 3%, 2%, or 1% of the total amino acid sequence.

[0112] Among the above, the substitution is preferably a conservative substitution. A conservative substitution is a replacement of a specific amino acid residue with a residue having similar physicochemical characteristics, but any substitution may be made as long as it does not substantially change the structural characteristics of the original sequence, for example, any substitution may be made as long as the substituted amino acid does not disrupt the helix present in the original sequence or other types of secondary structure that characterize the original sequence. Below, conservative substitutions of amino acid residues are categorized by substitutable residues and exemplified, but the substitutable amino acid residues are not limited to those listed below. Group A: leucine, isoleucine, valine, alanine, methionine Group B: aspartic acid, glutamic acid Group C: asparagine, glutamine D group: lysine, arginine, Group E: serine, threonine Group F: phenylalanine, tyrosine

[0113] In the case of non-conservative substitutions, one member of the above-mentioned classes can be exchanged for another member of the other classes. For example, amino acids in the above-mentioned groups B, D, and E may be substituted with amino acids from other groups to eliminate inadvertent glycosylation. Alternatively, cysteines may be deleted or substituted with other amino acids to prevent folding into a protein in a tertiary structure. Alternatively, amino acids may be substituted taking into account the hydropathic index of amino acids, which is an index of hydrophobicity / hydrophilicity for amino acids (J. Kyte and R. Doolittle, J. Mol. Biol., Vol. 157, pp. 105-132, 1982), to maintain a balance of hydrophilicity / hydrophobicity or to increase hydrophilicity for easier synthesis.

[0114] In another embodiment, substitution with an amino acid that is less sterically hindering than the original amino acid, for example, substitution of group F with group A, B, C, D, or E, or substitution of a charged amino acid with an uncharged amino acid, for example, substitution of group B with group C, may be performed. This may improve the binding affinity to IgE antibodies.

[0115] As used herein, the percent identity between two amino acid sequences can be determined by visual inspection and mathematical calculation. Percent identity can also be determined using a computer program. Examples of such computer programs include BLAST and ClustalW. In particular, the various conditions (parameters) for identity searches using the BLAST program are described in Altschul et al. (Nucl. Acids. Res., 25, pp. 3389-3402, 1997) and are publicly available from the websites of NCBI and the DNA Data Bank of Japan (DDBJ) (BLAST Manual, Altschul et al., NCB / NLM / NIH, Bethesda, MD 20894; Altschul et al.). Percent identity can also be determined using genetic information processing software programs such as GENETYX Ver. 7 (Genetyx), DNASIS Pro (Hitachi Software), and Vector NTI (Infomax).

[0116] As used herein, the phrase "one or several nucleotides deleted, substituted, inserted, or added" in reference to a base sequence refers to a base sequence in which one or several nucleotides have been deleted, substituted, inserted, and / or added. The term "several nucleotides" refers, but is not limited to, to 600 or less, 300 or less, 150 or less, 100 or less, 50 or less, 30 or less, 20 or less, 15 or less, 12 or less, 10 or less, 8 or less, 6 or less, 4 or less, or 3 or less nucleotides. Alternatively, "several nucleotides" refers to 30%, preferably 25%, 20%, 15%, 10%, 5%, 3%, 2%, or 1% of the total length of the base sequence. It is preferred that the deletion, substitution, insertion, or addition of the nucleotides does not result in a frameshift in the amino acid-encoding sequence.

[0117] As used herein, the percent identity between two nucleotide sequences can be determined by visual inspection and mathematical calculation. Percent identity can also be determined using a computer program. Examples of such sequence comparison computer programs include the BLASTN program (Altschul et al. (1990) J. Mol. Biol. 215: 403-10): version 2.2.7, available from the U.S. National Library of Medicine website: https: / / blast.ncbi.nlm.nih.gov / Blast.cgi, or the WU-BLAST2.0 algorithm. Standard default parameter settings for WU-BLAST2.0 can be found at the following internet site: http: / / blast.wustl.edu.

[0118] As used herein, "under stringent conditions" refers to hybridization under moderately or highly stringent conditions. Specifically, moderately stringent conditions can be easily determined by those skilled in the art, for example, based on the length of the DNA. Basic conditions are set forth in Sambrook et al., Molecular Cloning: A Laboratory Manual, 3rd ed., Chapters 6-7, Cold Spring Harbor Laboratory Press, 2001. Moderately stringent conditions are preferably hybridization conditions of 1xSSC to 6xSSC at 42°C to 55°C, more preferably 1xSSC to 3xSSC at 45°C to 50°C, and most preferably 2xSSC at 50°C. When the hybridization solution contains, for example, about 50% formamide, a temperature 5 to 15°C lower than the above temperatures is used. Washing conditions include 0.5×SSC to 6×SSC and 40°C to 60°C. During hybridization and washing, generally, 0.05% to 0.2%, preferably about 0.1%, SDS may be added. Highly stringent conditions can also be easily determined by those skilled in the art, for example, based on the length of the DNA. Generally, highly stringent (highly stringent) conditions include hybridization and / or washing at a higher temperature and / or lower salt concentration than moderately stringent conditions. For example, hybridization conditions include 0.1×SSC to 2×SSC and 55°C to 65°C, more preferably 0.1×SSC to 1×SSC and 60°C to 65°C, and most preferably 0.2×SSC and 63°C. Washing conditions include 0.2×SSC to 2×SSC, 50°C to 68°C, and more preferably 0.2×SSC, 60°C to 65°C.

[0119] The antigen may be obtained from shrimp by isolating and purifying it using a combination of protein purification methods well known to those skilled in the art, or by expressing the antigen as a recombinant protein using gene recombination techniques well known to those skilled in the art, and then isolating and purifying it using protein purification methods well known to those skilled in the art.

[0120] Examples of protein purification methods include methods that utilize solubility, such as salting out and solvent precipitation; methods that utilize differences in molecular weight, such as dialysis, ultrafiltration, gel filtration and SDS-PAGE; methods that utilize charge, such as ion exchange chromatography and hydroxylapatite chromatography; methods that utilize specific affinity, such as affinity chromatography; methods that utilize differences in hydrophobicity, such as reversed-phase high-performance liquid chromatography; and methods that utilize differences in isoelectric point, such as isoelectric focusing.

[0121] Proteins are prepared by recombinant DNA technology by preparing an expression vector containing a nucleic acid encoding an antigen, introducing the expression vector into suitable host cells by gene transfer or transformation, culturing the host cells under conditions suitable for expression of the recombinant protein, and recovering the recombinant protein expressed in the host cells.

[0122] A "vector" is a nucleic acid that can be used to introduce a nucleic acid linked thereto into a host cell, and an "expression vector" is a vector that can induce the expression of a protein encoded by the nucleic acid introduced by the vector. Vectors include plasmid vectors, viral vectors, etc. Those skilled in the art can select an appropriate expression vector for expressing a recombinant protein depending on the type of host cell to be used.

[0123] A "host cell" is a cell that is transfected or transformed with a vector. Those skilled in the art can appropriately select a host cell depending on the vector used. The host cell can be derived from a prokaryote, such as E. coli. When a prokaryote such as E. coli is used as a host, the antigen of the present invention may contain an N-terminal methionine residue to facilitate recombinant protein expression in the prokaryote. This N-terminal methionine can be cleaved from the recombinant protein after expression. Alternatively, the host cell can be a cell derived from a eukaryote, such as a unicellular eukaryote such as yeast, a plant cell, an animal cell (e.g., a human cell, a monkey cell, a hamster cell, a rat cell, a mouse cell, or an insect cell), or a silkworm.

[0124] Gene transfer or transformation of an expression vector into a host cell can be carried out appropriately by methods known to those skilled in the art. Furthermore, those skilled in the art can express a recombinant protein by culturing the host cells under appropriate conditions for recombinant protein expression, depending on the type of host cell. The host cells expressing the recombinant protein can then be homogenized, and the antigen expressed as the recombinant protein can be isolated and purified from the resulting homogenate by appropriately combining the above-mentioned protein purification methods. Antigens can also be prepared by introducing the above-mentioned expression vector, synthesized double-stranded DNA, or mRNA transcribed therefrom into a cell-free protein synthesis system, expressing the vector, and then isolating and purifying the expressed protein.

[0125] Diagnostic kits and methods (1) The present invention provides a method for providing an indicator for diagnosing shrimp allergy in a subject, the method comprising the steps of: (i) contacting a sample obtained from a subject with an antigen, wherein the sample is a solution containing IgE antibodies; (ii) detecting binding of IgE antibodies to the antigen in a sample obtained from the subject; (iii) if binding of the subject's IgE antibodies to the antigen is detected, an indication that the subject is allergic to shrimp is provided; wherein the antigen is at least one of the proteins identified as antigens (1) to (11) above.

[0126] As used herein, "diagnosis" generally includes not only a (definitive) diagnosis by a doctor, but also simple "detection," including possibility.

[0127] A sample obtained from a subject is a solution containing IgE antibodies collected from a subject. Examples of such solutions include blood, saliva, sputum, nasal discharge, urine, sweat, and tears. The sample obtained from a subject may be pretreated to increase the IgE antibody concentration in the sample before contacting it with an antigen. Pretreatment of the sample may include, for example, obtaining serum or plasma from the blood. Furthermore, the Fab portion, which is the binding portion to the antigen, may be purified. In a particularly preferred embodiment, the above step (i) is carried out by contacting the antigen with IgE antibodies in serum obtained from the subject.

[0128] The IgE antibody may be the IgE antibody itself, or may be a mast cell to which the IgE antibody is bound.

[0129] Contact between a sample obtained from a subject and an antigen and detection of its binding can be performed using known methods. Examples of such methods include ELISA (Enzyme-Linked Immunosorbent Assay), sandwich immunoassay, immunoblotting, immunoprecipitation, and immunochromatography. All of these methods involve contacting the subject's IgE antibody with the antigen to allow it to bind, allowing an enzyme-labeled secondary antibody to act on the IgE antibody specifically bound to the antigen, and then adding an enzyme substrate (usually a colorimetric or luminescent reagent) to detect the product of the enzyme reaction, thereby detecting the binding between the antigen and the subject's IgE antibody. Alternatively, fluorescently labeled secondary antibodies can be detected. Alternatively, surface plasmon resonance (SPR), or other measurement methods capable of evaluating antigen-IgE antibody binding, can also be used. Multiple antigen-specific IgE antibodies may be mixed.

[0130] The antigen may be in a state where the isolated antigen is immobilized on a carrier. In this case, ELISA, sandwich immunoassay, immunochromatography, surface plasmon resonance, etc. can be used in the above steps (i) and (ii). In the above step (i), it is carried out by contacting a sample obtained from a subject with the surface on which the antigen is immobilized. The isolated antigen may be obtained by separating and purifying it by combining protein purification methods well-known to those skilled in the art from shrimp, or by preparing it by genetic recombination technology. Also, it may be one to which an antibody is attached.

[0131] The antigen may not be immobilized on a carrier. In this case, flow cytometry, etc. can be used in the above steps (i) and (ii), and the presence of the antigen to which the antibody is bound can be confirmed by laser light. For example, a basophil activation test (BAT), etc. can be mentioned. Also, a histamine release test (HRT) for examining whether histamine is released by further contacting the antigen with blood cells in a sample can also be mentioned.

[0132] Further, the antigen may be transferred from a state separated by two-dimensional electrophoresis and detected by immunoblotting. Two-dimensional electrophoresis is a technique for separating a protein sample by performing isoelectric focusing electrophoresis in the first dimension and SDS-PAGE in the second dimension. In this case, the conditions for two-dimensional electrophoresis are not particularly limited as long as the antigen of the present invention can be separated. For example, the conditions for two-dimensional electrophoresis described in the item of "identification of antigen" above can be used. Alternatively, the electrophoresis conditions can also be determined referring to the descriptions of Patent Documents 1 to 4 above. For example, as follows: (A) As the isoelectric focusing electrophoresis gel in the first dimension, the gel length is within the range of 5 to 10 cm, the pH range of the gel is 3 to 10, and when the gel length up to pH 5 is a, the gel length from pH 5 to 7 is b, and the gel length of pH 7 or more is c, the relationships of "a < b" and "b > c" are satisfied; (B) In the case of (A), when the total length of the gel is 1, a is in the range of 0.15 to 0.3, b is in the range of 0.4 to 0.7, and c is in the range of 0.15 to 0.3; (C) In the first-dimensional isoelectric focusing, a constant voltage step is performed by applying a constant voltage within a range of 100 V to 600 V to each gel containing a sample, and after the electrophoretic change amplitude per 30 minutes of electrophoresis reaches a range of 5 μA, a voltage increase step is started in which the voltage is increased from the constant voltage; (D) In the case of (C), the final voltage of the voltage increasing step is set to be within the range of 3000V to 6000V; (E) the length of the first-dimension isoelectric focusing gel in the longitudinal direction is 5 to 10 cm, and the gel concentration at the base end of the second-dimension electrophoresis gel in the migration direction is 3 to 6%; and (F) In the case of (E), the gel concentration at the leading end of the second-dimensional electrophoresis gel in the migration direction is set higher than the gel concentration at the base end in the migration direction; Two-dimensional electrophoresis can be performed under conditions that satisfy at least one selected from the group consisting of:

[0133] The antigens (1) to (11) above are antigens that specifically bind to IgE antibodies of patients allergic to shrimp. Therefore, when binding between the subject's IgE antibodies and the antigens is detected, an indicator that the subject is allergic to shrimp is provided.

[0134] The present invention also provides a diagnostic kit for shrimp allergy, which comprises at least one of the antigens (1) to (11) above. The diagnostic kit of the present invention may be used in the above-mentioned method for providing an indicator for diagnosing shrimp allergy, or in the diagnostic method described below. In addition to comprising at least one of the antigens (1) to (11) above, the diagnostic kit of the present invention may also comprise an enzyme-labeled anti-IgE antibody and a chromogenic or luminescent substrate that serves as a substrate for the enzyme. A fluorescently labeled anti-IgE antibody may also be used. In the diagnostic kit of the present invention, the antigen may be provided in a state immobilized on a carrier. The diagnostic kit of the present invention may also be provided together with instructions on the diagnostic procedure or a package containing such instructions.

[0135] In another embodiment, the diagnostic kit includes a companion diagnostic for shrimp allergy. The companion diagnostic is used to identify patients who are expected to benefit from a drug or who are at risk of serious side effects from the drug, or to examine the responsiveness of the drug to optimize treatment with the drug. Here, optimization of treatment includes, for example, determining the dosage and administration, deciding when to discontinue administration, and identifying which allergen component is used to induce immune tolerance.

[0136] The present invention also provides a diagnostic composition for shrimp allergy, which contains at least one of the antigens (1) to (11) above. The diagnostic composition of the present invention can be used in the diagnostic methods described below. The diagnostic composition of the present invention may contain, as needed, pharmaceutically acceptable carriers and additives that are commonly used together with the antigen of the present invention.

[0137] In one aspect, the present invention provides a method for diagnosing shrimp allergy in a subject, comprising the steps of: (i) contacting a sample obtained from a subject with an antigen; (ii) detecting binding of IgE antibodies to the antigen in a sample obtained from the subject; (iii) if binding of the subject's IgE antibody to the antigen is detected, determining that the subject is allergic to shrimp; wherein the antigen is at least one of the proteins identified as antigens in (1) to (11) above, wherein steps (i) and (ii) are carried out as described for each step of the method for providing an indicator for diagnosing shrimp allergy.

[0138] In another aspect, the present invention provides a method for diagnosing shrimp allergy in a subject, the method comprising administering to the subject at least one of the antigens (1) to (11) above. The method may be performed in the form of a skin test, characterized by applying the antigen to the skin. Examples of skin tests include a prick test, in which a diagnostic composition is applied to the skin and then a small scratch is made to penetrate the skin without causing bleeding, allowing the antigen to penetrate the skin and observe a skin reaction; a scratch test, in which a diagnostic composition is applied and then a small scratch is made to observe a reaction; a patch test, in which a diagnostic composition in the form of a cream or ointment is applied to the skin and a reaction is observed; and an intradermal test, in which an antigen is administered intradermally and a reaction is observed. If a skin reaction, such as swelling, occurs in the area where the antigen was applied, the subject is diagnosed as having shrimp allergy. The amount of antigen applied to the skin may be, for example, 100 μg or less per administration.

[0139] In the diagnosis of allergies, a challenge test is often performed to identify the antigen. At least one of the antigens (1) to (11) above can be used as an active ingredient in a challenge test to diagnose shrimp allergy. The antigen protein used in the challenge test may be an expressed and purified protein, or may be expressed in a raw material or processed product, such as pollen rice, in which a cedar pollen antigen gene is transformed into rice and the antigen protein is expressed in the rice.

[0140] In one embodiment, the above-mentioned diagnostic composition and diagnostic kit can be used for prick tests, scratch tests, patch tests, intradermal tests, and the like.

[0141] In another aspect, the present invention provides at least one of the antigens (1) to (11) above for use in diagnosing shrimp allergy, including providing at least one of the antigens (1) to (11) above in a mixture with a known antigen.

[0142] In yet another aspect, the present invention provides the use of at least one of the antigens (1) to (11) above in the manufacture of a composition for diagnosing shrimp allergy.

[0143] Pharmaceutical Compositions and Treatment Methods (1) The present invention provides a pharmaceutical composition containing at least one of the antigens (1) to (11) above.

[0144] In one embodiment, the pharmaceutical composition is used to treat shrimp allergy. As used herein, "treatment of allergy" refers to increasing the limit of the amount of antigen that can be ingested into the body without causing symptoms, ultimately aiming to achieve a state (remission) in which symptoms do not appear with normal intake of the antigen.

[0145] The present invention also provides a method for treating shrimp allergy, which comprises administering at least one of the antigens (1) to (11) above to a patient in need of treatment for shrimp allergy.

[0146] In another aspect, the present invention provides at least one of the antigens (1) to (11) above for use in treating shrimp allergy. In yet another aspect, the present invention provides use of at least one of the antigens (1) to (11) above for the manufacture of a medicament for treating shrimp allergy.

[0147] In the treatment of allergies, hyposensitization therapy is often performed, with the goal of inducing immune tolerance by administering an antigen to the patient. At least one of the antigens (1) to (11) above can be used as an active ingredient in hyposensitization therapy for shrimp allergies. Here, the antigen protein used in hyposensitization therapy may be an expressed and purified protein, or may be expressed in a raw material or processed product, such as pollen rice, in which a cedar pollen antigen gene is transformed into rice and the antigen protein is expressed in the rice.

[0148] The pharmaceutical compositions of the present invention can be administered by any conventional route of administration, including, for example, oral, sublingual, transdermal, intradermal, subcutaneous, intravascular, intranasal, intramuscular, intraperitoneal, and rectal administration.

[0149] The pharmaceutical composition of the present invention can be prepared by adding commonly used pharmaceutically acceptable adjuvants, excipients, or various additives (e.g., stabilizers, solubilizers, emulsifiers, buffers, preservatives, colorants, etc.) together with the antigen of the present invention in a conventional manner, as needed. The dosage form of the pharmaceutical composition can be appropriately selected by those skilled in the art depending on the route of administration. For example, it may be in the form of tablets, capsules, troches, sublingual tablets, injections, nasal sprays, poultices, liquids, creams, lotions, suppositories, etc. The dosage, frequency of administration, and / or administration period of the pharmaceutical composition of the present invention can be appropriately selected by a physician depending on the route of administration, symptoms, and patient characteristics such as age and body weight. For example, in the case of adults, the composition may be administered at a dose of 100 μg or less per administration. The administration interval may be, for example, daily, once a week, twice a month, or once every three months. The administration period may be, for example, from several weeks to several years. A method of gradually increasing the dosage during the administration period may also be used.

[0150] Tester Composition (1) The present invention provides a tester composition containing an antibody against at least one of the antigens (1) to (11) above.

[0151] The antibody can be produced by a conventional method. For example, it may be produced by immunizing a mammal such as a rabbit with the antigens (1) to (11) above. The antibody may be an Ig antibody, a polyclonal antibody, a monoclonal antibody, or an antigen-binding fragment thereof (e.g., Fab, F(ab')2, Fab').

[0152] In the tester composition, the antibody may be provided in a form bound to a carrier. The carrier is not particularly limited as long as it can be used to detect antibody-antigen binding. Any carrier known to those skilled in the art can be used.

[0153] The following methods can be used to check whether or not an antigen is contained. A tester composition containing the prepared Ig antibody is brought into contact with a sample obtained from a raw material, processed product, etc., and the binding between the Ig antibody and the antigen in the sample is detected using, for example, ELISA, and if binding between the Ig antibody and the antigen is detected, it is determined that the target raw material, processed product, etc. contains the antigen. A method in which raw materials or processed products are soaked in filter paper or the like, and an antibody solution is reacted to detect the antigens contained therein.

[0154] In another aspect of the present invention, there is provided a tester composition for determining the presence or absence of a shrimp allergy antigen in a subject, characterized in that it comprises a primer having a nucleotide sequence complementary to at least a portion of the nucleotide sequence shown in SEQ ID NO: 1, 44, 86, 116, 140, 145, 160, 178, 229, 275, 299, 305, 361, 380, 398, 413, 420, 454, 480, 485, 519, 540, or 548. Non-limiting examples of the primers include a base sequence complementary to a portion of the 3' end or central portion of at least one of the base sequences set forth in SEQ ID NO: 1, 44, 86, 116, 140, 145, 160, 178, 229, 275, 299, 305, 361, 380, 398, 413, 420, 454, 480, 485, 519, 540, or 548, preferably 12 residues, 15 bases, 20 bases, or 25 bases. In particular, when targeting mRNA, the primer has a complementary primer to the poly(A) tail. In a preferred embodiment, the tester composition containing the above-mentioned primers may further contain a primer containing a base sequence of the 5'-terminal portion of at least one of the base sequences shown in SEQ ID NO: 1, 44, 86, 116, 140, 145, 160, 178, 229, 275, 299, 305, 361, 380, 398, 413, 420, 454, 480, 485, 519, 540, or 548, preferably a base sequence consisting of 12 bases, 15 bases, 20 bases, or 25 bases.

[0155] For example, the presence or absence of the antigen can be determined by using DNA or mRNA obtained from shrimp as a template and the complementary primers to amplify cDNA by PCR (Polymerase Chain Reaction), including RT-PCR (Reverse Transcription-Polymerase Chain Reaction), and comparing the sequence of the amplified cDNA with SEQ ID NO: 1, 44, 86, 116, 140, 145, 160, 178, 229, 275, 299, 305, 361, 380, 398, 413, 420, 454, 480, 485, 519, 540, or 548. Examples of PCR amplification methods include the RACE (Rapid Amplification of cDNA End) method. In this case, when the amplified cDNA is compared with SEQ ID NO: 1, 44, 86, 116, 140, 145, 160, 178, 229, 275, 299, 305, 361, 380, 398, 413, 420, 454, 480, 485, 519, 540 or 548, if there is a point mutation encoding the same amino acid, or if there is a point mutation encoding the same amino acid in the base sequence of the amplified cDNA, Even if there is an insertion, deletion, substitution, or addition of bases in the base sequence of SEQ ID NO: 54, 480, 485, 519, 540, or 548, the antigen is determined to be present if the amino acid sequence encoded by the cDNA has an identity of 70% or more, preferably 80, 90, 95, 98, or 99% or more, to the amino acid sequence of SEQ ID NO: 2, 45, 87, 117, 141, 146, 161, 179, 230, 276, 300, 306, 362, 381, 399, 414, 421, 455, 481, 486, 520, 541, or 549.

[0156] In one embodiment, the tester composition is used to examine whether or not an antigen is present in a target object, such as a food ingredient (shrimp) or in a food production line. The tester composition may be used by manufacturers to inspect the quality of production lines and products before shipping, or by consumers themselves to check whether or not the target raw materials or processed products contain the antigen.

[0157] Method for determining the presence or absence of antigen (1) The present invention includes a method for determining the presence or absence of any of the above antigens (1) to (11) in a target substance, which comprises contacting a raw material or processed product (including a liquid) with an antibody against at least one of the above antigens (1) to (11).

[0158] The raw material may be a food ingredient, a cosmetic raw material, a pharmaceutical raw material, etc. The processed product may be an edible processed product, a cosmetic, a pharmaceutical, etc.

[0159] The antibody, the method for producing the antibody, the method for contacting the antibody with raw materials or processed products, the binding of the antibody with the antigen, etc. are as described above in "Tester composition (1)."

[0160] Allergen-free foods, etc. (1) The present invention provides shrimp or shrimp processed products characterized in that at least one of the above antigens (1) to (11) has been removed or reduced.

[0161] The method for removing or reducing the antigens of the present invention in shrimp or shrimp processed products is not limited, and the removal or reduction of the antigens may be carried out by any method as long as the antigens of the present invention are removed or reduced.

[0162] For example, shrimp in which the antigens of the present invention have been removed or reduced may be prepared by using genetic modification techniques to modify the expression of the antigens of the present invention.

[0163] Genetic modification techniques can be any technique known to those skilled in the art. For example, Oishi et al. (Scientific Reports, Vol. 6, Article number: 23980, 2016, doi:10.1038 / srep23980) describes the application of the genome editing technique CRISPER / Cas9 to chicken primordial germ cells to obtain ovomucoid gene-deficient individuals. A similar technique can also be used to obtain shrimp from which the antigens of the present invention have been removed.

[0164] Alternatively, shrimp with reduced antigen content of the present invention may be obtained by crossbreeding with shrimp that do not contain or have low antigen content by artificial insemination. Artificial crossbreeding of shrimp can be carried out by conventional methods, such as the method described in "Maturation, Spawning, and Egg Collection Techniques of Kuruma Shrimp" (edited by Takuji Okumura and Katsutoshi Suito), Aichi Prefectural Fisheries Promotion Fund, 2014, by the Fisheries Engineering Research Institute, National Research and Development Agency, Fisheries Research and Education Agency.

[0165] The shrimp processed product of the present invention from which antigens have been removed or reduced may be a processed product made from shrimp from which antigens of the present invention have been removed or reduced. When using normal shrimp as the raw material, treatment to remove or reduce the antigens of the present invention is carried out before or after preparation of the shrimp processed product. Methods for removing or reducing the antigens of the present invention in shrimp processed products made from normal shrimp include methods for removing protein components from the raw material or processed product, such as high-pressure treatment, elution with a neutral salt solution, or high-temperature steam, and methods for hydrolysis, denaturation, or amino acid changes (chemical modification or elimination of side chains, etc.) by heat treatment or acid treatment.

[0166] Manufacturing method for allergen-free processed products (1) The present invention provides a method for producing shrimp or shrimp processed products from which antigens have been removed or reduced, the method comprising a step of confirming that the antigens have been removed or reduced during the production of the processed product, wherein the antigen is at least one of the antigens (1) to (11) above.

[0167] The step of confirming that the antigens have been removed or reduced during the production process of shrimp or shrimp processed products from which the antigens have been removed or reduced may be carried out by confirming whether or not the antigens are present using the method described above in the section "Tester (1)."

[0168] Furthermore, shrimp or shrimp processed products from which antigens have been removed or reduced may be produced by the methods described above in the section "Allergen-removed foods, etc."

[0169] antigen epitope For the antigens and arginine kinase identified as shown in Examples 1-3, epitopes and amino acids within the epitopes that are important for binding to IgE antibodies from allergic patients were identified as shown in Example 4.

[0170] The present invention provides polypeptides comprising amino acid sequences (E1) to (E50) that contain or consist of any of the amino acid sequences of SEQ ID NOs: 558-984 listed in Table 3 below, as polypeptides comprising amino acid sequences that specifically bind to IgE antibodies of allergy patients. (E1) to (E50) are amino acid sequences that bind to IgE antibodies derived from the proteins listed under "Origin" in Table 3 (hereinafter, these may be referred to as "epitopes").

[0171] Myosin heavy chain type 1 derived: (E1)-(E4), (E26)-(E43) Myosin heavy chain type 2 derived: (E5)-(E11) Glycogen phosphorylase derived: (E12), (E13) Hemocyanin subunit L1 derived: (E14), (E15), (E44) Pyruvate kinase 3 derived: (E16), (E17), (E45) Phosphopyruvate hydratase derived: (E18), (E46), (E47) Mitochondrial ATP synthase subunit alpha precursor derived: (E19), (E20) Troponin I derived: (E21), (E48), (E49) Cyclophilin A derived: (E22), (E23), (E50) Arginine kinase derived: (E24), (E25)

[0172] Polypeptides containing the amino acid sequences (E1)-(E50) above may be prepared by chemical synthesis techniques such as solid-phase peptide synthesis. Alternatively, polypeptides containing epitopes may be expressed as recombinant polypeptides using genetic recombination techniques well known to those skilled in the art, followed by isolation and purification using protein production methods well known to those skilled in the art. Two or more types of polypeptides may be linked in combination, or one type of epitope may be linked repeatedly. In this case, binding to Ig antibodies is generally improved.

[0173] The length of a polypeptide containing the amino acid sequence of (E1)-(E50) is not particularly limited. In preferred embodiments, the length of a polypeptide containing the amino acid sequence of (E1)-(E50) may be 500 amino acids or less, 300 amino acids or less, 200 amino acids or less, 100 amino acids or less, 50 amino acids or less, 30 amino acids or less, 20 amino acids or less, 15 amino acids or less, 10 amino acids or less, or 5 amino acids or less. When a polypeptide is formed by repeatedly linking one or more of the amino acid sequences of (E1)-(E50) once or twice, in preferred embodiments, the length of the amino acid sequence portion may be 1,000 amino acids or less, 750 amino acids or less, 500 amino acids or less, 250 amino acids or less, 100 amino acids or less, 75 amino acids or less, 50 amino acids or less, 30 amino acids or less, 15 amino acids or less, 10 amino acids or less, or 5 amino acids or less. The number of amino acid residues described in preferred embodiments as the length of the polypeptide refers to the sum of the lengths of the sequences before and after the spacer (excluding the spacer).

[0174] Among SEQ ID NOS: 558-984 listed in Table 3, SEQ ID NOS: 558, 566, 582, 586, 594, 599, 614, 624, 634, 640, 654, 671, 672, 678, 681, 686, 691, 697, 704, 705, 708, 717, 725, 729, 741, 750, 752, 765, 770, 778, 788, 793 listed in the "Common 15-residue sequence" of Table 3 Each of the 50 sequences, 810, 817, 826, 833, 847, 858, 865, 879, 881, 892, 908, 912, 915, 917, 928, 935, 939, and 943, is a common 15-amino acid residue sequence identified by overlapping epitope mapping as an epitope that binds to IgE antibodies in each of the epitopes (E1) to (E50). In one aspect, the present invention relates to a polypeptide comprising or consisting of these amino acid sequences. The epitope sequence contained in the polypeptide of the present invention may be the entire 15-amino acid residue common epitope, or a part thereof. The epitope sequence may be 4 or more amino acid residues, 5 or more amino acid residues, 6 or more amino acid residues, 7 or more amino acid residues, 8 or more amino acid residues, 9 or more amino acid residues, 10 or more amino acid residues, 11 or more amino acid residues, 12 or more amino acid residues, 13 or more amino acid residues, or 14 or more amino acid residues.

[0175] In the Examples herein, for example, epitope cross-reactivity was confirmed for many polypeptides consisting of four amino acid residues (e.g., SEQ ID NOS: 587, 593, 595, 636, 655, 662, 663, 679, 707, 726, 727, 794, 880, etc.). Furthermore, epitope cross-reactivity was confirmed for many epitopes consisting of five or more amino acid residues. Thus, epitope sequences containing at least four amino acid residues are useful for detecting cross-reactivity with various antigens.

[0176] Therefore, the present invention also encompasses polypeptides comprising or consisting of the "consensus 15-residue sequence" of SEQ ID NOs: 558-984, i.e., amino acid sequences other than SEQ ID NOs: 558, 566, 582, 586, 594, 599, 614, 624, 634, 640, 654, 671, 672, 678, 681, 686, 691, 697, 704, 705, 708, 717, 725, 729, 741, 750, 752, 765, 770, 778, 788, 793, 810, 817, 826, 833, 847, 858, 865, 879, 881, 892, 908, 912, 915, 917, 928, 935, 939, and 943.

[0177] In Table 3, "preferred" sequences are shorter partial sequences of the "15-residue consensus sequence" that can function as an epitope. "Key" sequences indicate sequences that are considered to be particularly important among the "15-residue consensus sequence" or "key" sequences. Among the "key" sequences, amino acid sequences designated with "X" are amino acid residues that have been confirmed by alanine-glycine scanning to retain their binding to IgE antibodies even when changed to any alanine (or glycine if the original amino acid residue is alanine). Thus, X is any amino acid residue, preferably alanine (or glycine). Note that "key" sequences that do not contain a sequence designated with "X" are those in which no amino acid residues confirmed to retain their binding to IgE antibodies were found by alanine-glycine scanning.

[0178] With respect to the epitope (E1), the amino acid residues designated as X in SEQ ID NOs: 560, 562, 564, and 565 are amino acid residues that have been confirmed to retain IgE antibody binding even when altered. Thus, in SEQ ID NOs: 558-565, one or more amino acid residues corresponding to the 4th, 5th, 7th, 8th, 10th, 11th, 12th, 13th, 14th, and 15th amino acid residues of SEQ ID NO: 558 may be substituted with any amino acid residue. In the present invention, preferably, one or more amino acid residues designated as X in the "key sequence" corresponding to each "preferred sequence" may be substituted with any amino acid residue. For example, in one embodiment, the key sequence corresponding to the preferred sequence SEQ ID NO: 559 is SEQ ID NO: 560. Preferably, in SEQ ID NO: 559, the 2nd, 6th, 7th, 8th, and 9th amino acid residues designated as X in SEQ ID NO: 560 (corresponding to the 8th, 12th, 13th, 14th, and 15th amino acid residues of SEQ ID NO: 558) may be substituted with any amino acid residue. Alternatively, in another embodiment, the key sequence corresponding to the preferred sequence SEQ ID NO: 563 is SEQ ID NO: 564. Preferably, in SEQ ID NO: 563, the 1st, 2nd, 4th, 5th, 7th, and 8th amino acid residues that are X in SEQ ID NO: 564 (corresponding to the 4th, 5th, 7th, 8th, 10th, and 11th amino acid residues in SEQ ID NO: 558) are amino acid residues that can be optionally substituted. The same applies hereinafter to other "preferred sequences," "key sequences," and (E2)-(E50) in (E1).

[0179] The number of amino acid residues that may be substituted is not limited, but is preferably 6 or less, 5 or less, 4 or less, 3 or less, 2 or less, or 1 or less. The same applies hereinafter to (E2) to (E50).

[0180] Information about the epitopes (E1)-(E50) is summarized in Table 1.

[0181] [Table 1]

[0182] The sequences listed in the "Synthetic Sequence" and / or "SEQ ID NO:" columns to the right of Graph Nos. 1-114 in Table 3 are sequences for which epitope cross-reactivity was confirmed in Example 5. "Epitope cross-reactivity was confirmed" means that the IgE antibodies in the serum of each allergic patient showed higher binding affinity than the IgE antibodies in the serum of a non-allergic subject (specifically, greater than "1" in Figures 7-12). Thus, in one aspect, the polypeptides of the present invention include polypeptides comprising or consisting of these amino acid sequences. In one aspect, the IgE antibodies in the serum of each allergic patient show binding affinity to the polypeptides of the present invention that is 1.05-fold or more, 1.10-fold or more, 1.15-fold or more, 1.20-fold or more, or 1.25-fold or more higher than the IgE antibodies in the serum of a non-allergic subject.

[0183] As used herein, "polypeptides comprising the amino acid sequences of (E1) to (E50)" include polypeptides comprising or consisting of any of the amino acid sequences of SEQ ID NOs: 558-984, as well as polypeptides in which amino acid residues have been substituted as described above. "Comprising any of the amino acid sequences of SEQ ID NOs: 558-984" means that any other amino acid sequence may be included as long as it does not affect the binding of any of the amino acid sequences of SEQ ID NOs: 558-984 (including the substituted forms) to IgE antibodies (i.e., their function as epitopes).

[0184] Diagnostic kits and methods (2) The present invention provides a method for providing an indicator for diagnosing allergies in a subject, comprising the steps of: (i) contacting a sample obtained from a subject with an antigen, wherein the sample is a solution containing IgE antibodies; (ii) detecting binding of IgE antibodies to the antigen in a sample obtained from the subject; (iii) if binding of the subject's IgE antibodies to the antigen is detected, an indication that the subject is allergic is provided; wherein the antigen is at least one polypeptide comprising the amino acid sequence of (E1)-(E50), or a polypeptide in which two or more polypeptides comprising the amino acid sequences of (E1)-(E50) are linked together with or without a spacer.

[0185] Hereinafter, a polypeptide that is at least one of the polypeptides containing the amino acid sequence of (E1)-(E50) above, or a polypeptide in which two or more polypeptides containing the amino acid sequence of (E1)-(E50) above are linked with or without a spacer, may be referred to as an "antigen containing (E1)-(E50) above." The type of spacer is not particularly limited, and any spacer commonly used by those skilled in the art for linking multiple peptides can be used. The spacer may be, for example, a hydrocarbon chain such as Acp(6)-OH, an amino acid chain, or other polypeptide.

[0186] When polypeptides containing the amino acid sequences of (E1) to (E50) are linked with or without a spacer, the number of linked polypeptides is not particularly limited. In one embodiment, the number is 2 or more, 3 or more, 4 or more, 5 or more, 6 or more, 8 or more, 10 or more, or 15 or more. In another embodiment, the number is 30 or less, 20 or less, 15 or less, 10 or less, 8 or less, 6 or less, 5 or less, 3 or less, or 2 or less.

[0187] The polypeptides containing the amino acid sequences (E1)-(E50) may be composed of repeated identical polypeptides or multiple linked polypeptides containing different amino acid sequences. Even when multiple linked polypeptides containing the amino acid sequences (E1)-(E50) are combined, they can be used in the methods, kits, and compositions of the present invention as the polypeptide of the present invention.

[0188] The sample obtained from the subject is as described above in the section "Diagnostic kit and diagnostic method (1)."

[0189] Contact between a sample obtained from a subject and the polypeptide and detection of its binding can be performed by known methods described in the above section "Diagnostic Kit / Diagnostic Method (1)," such as ELISA (Enzyme-Linked Immunosorbent Assay), sandwich immunoassay, immunoblot, immunoprecipitation, immunochromatography, etc.

[0190] The polypeptide comprising the amino acid sequence (E1)-(E50) may be immobilized on a carrier. In this case, ELISA, sandwich immunoassay, immunochromatography, surface plasmon resonance, or the like can be used in steps (i) and (ii). Step (i) is performed by contacting a sample obtained from a subject with a surface on which the polypeptide comprising the amino acid sequence (E1)-(E50) is immobilized. Alternatively, the subject's IgE antibody may be immobilized on a carrier, and the binding of the antibody to the polypeptide comprising the amino acid sequence (E1)-(E50) may be detected by the above-described method. A spacer or a tag such as biotin may be attached to the N-terminus or C-terminus of the polypeptide to facilitate binding to the carrier, to provide a space between the carrier and the polypeptide, or to facilitate antibody contact with the polypeptide. In the case of binding to biotin, the carrier preferably contains avidin.

[0191] The polypeptide containing the amino acid sequence (E1)-(E50) may not be immobilized on a support. In this case, flow cytometry or the like can be used in steps (i) and (ii) above, and the presence of the polypeptide containing the amino acid sequence (E1)-(E50) bound to an IgE antibody can be confirmed using laser light. Examples of this method include the basophil activation test (BAT), which detects the surface antigen CD203c that appears when basophils are activated by contact with a polypeptide containing the amino acid sequence (E1)-(E50). Another example is the histamine release test (HRT), which examines whether a blood cell in a sample releases histamine by further contacting the polypeptide containing the amino acid sequence (E1)-(E50) with the blood cell.

[0192] A polypeptide comprising the amino acid sequence (E1)-(E50) is an antigen that specifically binds to the IgE antibody of an allergic patient. Therefore, detection of binding between a subject's IgE antibody and the antigen provides an indication that the subject is allergic, including cross-reactivity. When synthesizing a polypeptide comprising the amino acid sequence (E1)-(E50), sequences may be added before and after the epitope to facilitate synthesis using Escherichia coli, for example, to increase the sequence length. Even in such cases, detection of binding between a subject's IgE antibody and the amino acid sequence (E1)-(E50) provides an indication that the subject is allergic, including cross-reactivity. Therefore, any sequence may be added before or after the amino acid sequence (E1)-(E50), which is the epitope.

[0193] The present invention also provides a diagnostic kit for allergies, comprising at least one polypeptide comprising the amino acid sequence of (E1)-(E50). The diagnostic kit of the present invention may be used in the above-mentioned method for providing an indicator for diagnosing allergies, or in the diagnostic method described below. In addition to comprising at least one polypeptide comprising the amino acid sequence of (E1)-(E50), the diagnostic kit of the present invention may also comprise an enzyme-labeled anti-IgE antibody and a chromogenic or luminescent substrate that serves as a substrate for the enzyme. A fluorescently labeled anti-IgE antibody may also be used. In the diagnostic kit of the present invention, the polypeptide comprising the amino acid sequence of (E1)-(E50) may be provided in a state immobilized on a carrier. The diagnostic kit of the present invention may also be provided together with instructions on the diagnostic procedure or a package containing such instructions.

[0194] In another embodiment, the diagnostic kit includes a companion diagnostic for allergies. The companion diagnostic is used to identify patients who are expected to benefit from a drug or who are at risk of serious side effects from the drug, or to examine the responsiveness of the drug to optimize treatment with the drug. Here, optimization of treatment includes, for example, determining the dosage and administration, deciding whether to discontinue administration, and identifying which allergen component is used to induce immune tolerance.

[0195] The present invention also provides a composition for diagnosing allergies, comprising at least one polypeptide comprising the amino acid sequence of any of (E1)-(E50). The diagnostic composition of the present invention can be used in the diagnostic methods described below. The diagnostic composition of the present invention may, as needed, contain pharmaceutically acceptable carriers or additives that are commonly used together with the polypeptide of the present invention.

[0196] In one aspect, the present invention provides a method for diagnosing allergy in a subject, comprising the steps of: (i) contacting a sample obtained from a subject with an antigen; (ii) detecting binding of IgE antibodies to the antigen in a sample obtained from the subject; (iii) determining that the subject is allergic if binding of the subject's IgE antibody to the antigen is detected; wherein the antigen is at least one of the polypeptides identified as comprising any one of the amino acid sequences (E1)-(E50) above, wherein steps (i) and (ii) are carried out as described for each step of the method for providing an indicator for diagnosing allergies.

[0197] In another aspect, the present invention provides a method for diagnosing allergies in a subject, the method comprising administering to the subject at least one polypeptide comprising the amino acid sequence (E1)-(E50). The method may be performed in the form of a skin test, which involves applying a polypeptide comprising the amino acid sequence (E1)-(E50) to the skin. Skin tests include a prick test, in which a diagnostic composition is applied to the skin, followed by a small scratch (not enough to cause bleeding) to penetrate the polypeptide comprising the amino acid sequence (E1)-(E50) into the skin and observe a skin reaction; a scratch test, in which a diagnostic composition is applied and then the skin is slightly scratched to observe a reaction; a patch test, in which a diagnostic composition in the form of a cream or ointment is applied to the skin and observe a reaction; and an intradermal test, in which a polypeptide comprising the amino acid sequence (E1)-(E50) is intradermally administered and observe a reaction. If a skin reaction, such as swelling, occurs in the area where the polypeptide comprising the amino acid sequence (E1)-(E50) was applied, the subject is diagnosed as having an allergy. Here, the amount of the polypeptide to be applied to the skin may be, for example, 100 μg or less per application.

[0198] In the diagnosis of allergies, oral challenge tests are often performed to identify the antigen, verify antigen intake, and verify the severity of symptoms. At least one of the polypeptides containing the amino acid sequence (E1)-(E50) above can be used as an active ingredient in oral challenge tests to diagnose allergies. The polypeptide used in oral challenge tests may be an expressed and purified polypeptide, or it may be expressed in raw materials or processed products, such as pollen rice, in which a cedar pollen antigen gene is transformed into rice and the polypeptide is expressed in the rice.

[0199] In one embodiment, the above-mentioned diagnostic composition and diagnostic kit can be used for prick tests, scratch tests, patch tests, intradermal tests, and the like.

[0200] In another aspect, the present invention provides at least one polypeptide comprising the amino acid sequence of any of (E1)-(E50) above for use in diagnosing allergies.

[0201] In yet another aspect, the present invention provides the use of at least one polypeptide comprising the amino acid sequence of any one of (E1) to (E50) above in the manufacture of a diagnostic agent for allergies.

[0202] In this section, the allergy to be diagnosed may be an allergy to a polypeptide comprising the amino acid sequence of (E1)-(E50). That is, the diagnosis of allergy, including the detection of allergy and the provision of a diagnostic indicator, may diagnose not only allergy to a single polypeptide comprising the amino acid sequence of (E1)-(E50), but also allergy including cross-reactivity.

[0203] Pharmaceutical Compositions and Treatment Methods (2) The present invention provides pharmaceutical compositions containing at least one of the polypeptides comprising the amino acid sequence of (E1)-(E50). In one aspect, the pharmaceutical compositions are used to treat allergies. Treating allergies involves increasing the limiting amount of polypeptide that does not cause allergies when taken into the body, ultimately aiming to achieve a state (remission) in which allergies do not occur with normal intake of the polypeptide.

[0204] The present invention also provides a method for treating allergies, which comprises administering to a patient in need of such treatment at least one of the polypeptides comprising the amino acid sequence of (E1)-(E50) above.

[0205] In another aspect, the present invention provides at least one polypeptide comprising the amino acid sequence of any of (E1)-(E50) above for use in treating allergies. In yet another aspect, the present invention provides use of at least one polypeptide comprising the amino acid sequence of any of (E1)-(E50) above for the manufacture of a medicament for treating allergies.

[0206] In the treatment of allergies, hyposensitization therapy is often performed, with the goal of inducing immune tolerance by administering an antigen to the patient. At least one of the polypeptides containing the amino acid sequence (E1)-(E50) above can be used as an active ingredient in hyposensitization therapy for allergies. Here, the antigen used in hyposensitization therapy may be an expressed and purified polypeptide, or it may be expressed in a raw material or processed product, such as pollen rice.

[0207] The administration route, dosage, number of doses and / or duration of administration, other ingredients contained in the pharmaceutical composition, and dosage form of the pharmaceutical composition of the present invention can be those described above in the section "Pharmaceutical Compositions and Treatment Methods (1)." When using a polypeptide containing the amino acid sequence of (E1)-(E50) above, the dosage may be, for example, 100 μg or less per dose for an adult.

[0208] In this section, the allergy to be treated may be an allergy to a polypeptide comprising the amino acid sequence of any one of (E1)-(E50). That is, the allergy treatment may be not only an allergy to a single polypeptide comprising the amino acid sequence of any one of (E1)-(E50), but also an allergy including cross-reactive allergies.

[0209] Tester Composition (2) The present invention provides a tester composition comprising an antibody against at least one of the polypeptides comprising the amino acid sequences (E1)-(E50) above.

[0210] The antibody can be produced by conventional methods. For example, it may be produced by immunizing a mammal such as a rabbit with a polypeptide containing the amino acid sequence of any one of (E1)-(E50). The antibody may be an Ig antibody, a polyclonal antibody, a monoclonal antibody, or an antigen-binding fragment thereof (e.g., Fab, F(ab')2, Fab').

[0211] Furthermore, in the above-mentioned tester composition, the antibody may be provided in a form bound to a carrier. The carrier is not particularly limited as long as it can be used to detect the binding between the antibody and the polypeptide comprising the amino acid sequence of (E1)-(E50). Any carrier known to those skilled in the art can be used. Furthermore, it is preferable that the antibody against the polypeptide comprising the amino acid sequence of (E1)-(E50) is an antibody against a polypeptide having an amino acid sequence identical to the epitope and important amino acids described in the above section "Epitope of the antigen." This allows for the preparation of a tester composition that can detect cross-reactivity.

[0212] The following method can be used to examine whether or not a polypeptide containing any of the amino acid sequences (E1) to (E50) is contained.

[0213] A method for determining whether a polypeptide containing the amino acid sequence (E1)-(E50) is contained in a target raw material, processed product, etc., by contacting a tester composition containing the antibody thus prepared with a sample obtained from the raw material, processed product, etc., and detecting the binding between the antibody and the polypeptide containing the amino acid sequence (E1)-(E50) in the sample using, for example, ELISA, and determining that the polypeptide containing the amino acid sequence is contained in the target raw material, processed product, etc., if the binding between the antibody and the polypeptide containing the amino acid sequence (E1)-(E50) is detected. (The "method for determining whether a polypeptide is contained" includes determining that the polypeptide has been removed or reduced if the binding between the antibody and the polypeptide containing the amino acid sequence (E1)-(E50) is reduced.) A method in which raw materials, processed products, etc. are soaked in filter paper, etc., and an antibody solution is reacted to detect polypeptides containing the amino acid sequences (E1)-(E50) contained therein.

[0214] Another aspect of the present invention provides a tester composition for determining the presence or absence of a polypeptide containing any of the amino acid sequences (E1)-(E50) of an allergen in a subject, the composition comprising primers corresponding to the polypeptide having an amino acid sequence in which the epitope and key amino acids are identical. The primers may be designed, for example and without limitation, to contain a portion of the nucleotide sequence of a nucleic acid encoding the amino acid sequence specified in (E1)-(E50) above, or its complementary strand. Alternatively, the primers may be designed to correspond to the nucleotide sequence of a region upstream of the portion encoding the polypeptide having an amino acid sequence in which the epitope and key amino acids are identical, or the complementary strand of a region downstream of the portion encoding the polypeptide having an amino acid sequence in which the epitope and key amino acids are identical, in a nucleic acid encoding a protein containing the polypeptide having an amino acid sequence in which the epitope and key amino acids are identical, which is the amino acid sequence specified in (E1)-(E50) above. Examples of such primers include primers that are at least a portion of the nucleotide sequences set forth in SEQ ID NOs: 1, 44, 86, 116, 140, 145, 160, 178, 229, 275, 299, 305, 361, 380, 398, 413, 420, 454, 480, 485, 519, 540, or 548, and / or primers that are a portion of a sequence complementary to at least one of the nucleotide sequences set forth in SEQ ID NOs: 1, 44, 86, 116, 140, 145, 160, 178, 229, 275, 299, 305, 361, 380, 398, 413, 420, 454, 480, 485, 519, 540, or 548. The location of the epitope in the full-length antigen sequence is as specified in Table 3 in Example 4 below. In particular, when targeting mRNA, a complementary primer to the polyA tail may be included.

[0215] For example, DNA or mRNA obtained from a sample is used as a template, and the primers are used to amplify DNA by PCR (Polymerase Chain Reaction), including RT-PCR. The presence or absence of an antigen containing (E1)-(E50) is determined by determining whether the amplified DNA sequence contains a nucleic acid encoding the amino acid sequence specified by (E1)-(E50) above. Examples of methods for PCR amplification of mRNA include the RACE method. If one of the amino acid sequences encoded by three possible open reading frames in the amplified DNA contains the amino acid sequence specified by (E1)-(E50) above, the antigen is determined to be present. If DNA is not amplified, the antigen is determined to be absent.

[0216] In one embodiment, the tester composition is used to examine whether or not a target substance, such as a raw material or a processed product production line, contains a polypeptide comprising the amino acid sequence (E1)-(E50). The raw material may be a food ingredient, a cosmetic raw material, a pharmaceutical raw material, etc. The processed product may be an edible processed product, a cosmetic product, a pharmaceutical product, etc. The tester composition may be used to search for biological species contained in raw materials, for quality inspection by manufacturers on production lines and before shipping, or for consumers or users themselves to check whether or not target raw materials or processed products contain antigens.

[0217] Polypeptides How to determine the presence or absence of (2) The present invention includes a method for determining the presence or absence of a polypeptide comprising the amino acid sequence of any of (E1)-(E50) in a raw material or processed product, which method comprises detecting a polypeptide comprising the whole or part of the amino acid sequence of a polypeptide comprising the amino acid sequence of any of (E1)-(E50) in the raw material or processed product.

[0218] In one aspect, the method of the present invention comprises a step of determining the presence or absence of a polypeptide comprising the amino acid sequence of (E1)-(E50) in a target substance, which step comprises contacting a raw material or processed product (including a liquid) with an antibody against at least one of the polypeptides comprising the amino acid sequence of (E1)-(E50).

[0219] The antibody, raw material / processed product definition, method for producing the antibody, method for contacting the antibody with the raw material / processed product, and binding of the antibody with the antigen are as described above in "Tester composition (2)."

[0220] Alternatively, the method for determining the presence or absence of the antigen also includes detecting an epitope portion of a polypeptide comprising the amino acid sequence (E1)-(E50) contained in the antigen. The "epitope portion" preferably comprises four or more, six or more, or eight or more amino acid residues. The epitope portion can be detected using known methods for detecting a specific amino acid sequence in a portion of a polypeptide. For example, a method may be used in which proteins from a target raw material or processed product (e.g., foodstuff) are cleaved with a digestive enzyme for antigen removal treatment, followed by separation by HPLC or the like, and the peak of a given epitope peptide is measured to determine whether the antigen removal treatment has reduced the peak. Alternatively, the presence or absence of an antigen comprising a polypeptide comprising the amino acid sequence (E1)-(E50) in a target substance may be determined using an antibody that recognizes a portion of the polypeptide comprising the amino acid sequence (E1)-(E50).

[0221] Allergen-removing ingredients, etc. (2) The present invention provides raw materials or processed products characterized in that at least one of the polypeptides comprising the amino acid sequences (E1) to (E50) above has been removed or reduced.

[0222] The method for removing or reducing the antigens of the present invention in raw materials or processed products is not limited. The antigens may be removed or reduced by any method as long as the polypeptides containing the amino acid sequences (E1) to (E50) are removed or reduced. For example, the methods described in the above section "Allergen-free foods, etc." may be used.

[0223] "At least one of the polypeptides comprising the amino acid sequences (E1)-(E50) has been removed or reduced" may be achieved by removing or reducing the entire amino acid sequence, or by truncating or removing the amino acid sequence portions (E1)-(E50) from the antigen protein. "Removed" includes deletion and modification of all or part of the sequence portions specified in (E1)-(E50) above.

[0224] For example, a raw material from which the polypeptide comprising the amino acid sequence of (E1)-(E50) has been removed or reduced may be prepared using genetic modification techniques to eliminate the expression of the polypeptide comprising the amino acid sequence of (E1)-(E50). Any genetic modification knockout technique known to those skilled in the art can be used.

[0225] The processed product from which the polypeptides comprising the amino acid sequences (E1)-(E50) have been removed or reduced may be a processed product made from raw materials from which the polypeptides comprising the amino acid sequences (E1)-(E50) have been removed or reduced, such as powdered milk made from protein digests. When using normal raw materials, treatment to remove or reduce the polypeptides comprising the amino acid sequences (E1)-(E50) is carried out before, during, or after the preparation of the processed product. "Preparation of a processed product" refers to the preparation of a processed food product (e.g., a shrimp product, such as fried shrimp) from a food raw material (e.g., shrimp).

[0226] The methods described in the above section on "Allergen-Free Foods, etc." may be used to remove or reduce the amount of polypeptides containing the amino acid sequence (E1)-(E50) in processed foods made from ordinary raw materials. Examples of methods for cleaving polypeptides containing the amino acid sequence (E1)-(E50) include cleavage using specific digestive enzymes.

[0227] Manufacturing method for allergen-free processed products (2) The present invention provides a method for producing a processed product in which at least one polypeptide comprising the amino acid sequence (E1)-(E50) has been removed or reduced, the method comprising a step of confirming that the antigen has been removed or reduced during the production of the processed product.

[0228] In this production method, the removal or reduction of polypeptides comprising the amino acid sequence of (E1)-(E50) means that at least one of the polypeptides comprising the amino acid sequence of (E1)-(E50)(E1)-(E50) is removed or reduced, or the sequence portion specified by (E1)-(E50) is cleaved or removed from the antigen.

[0229] The method for confirming that polypeptides have been removed or reduced during the manufacturing process of a processed product is not particularly limited, and any method capable of detecting at least one of the polypeptides comprising the amino acid sequence (E1)-(E50) above may be used. For example, the presence or absence of a polypeptide in a processed product may be confirmed by measuring the binding of a sample containing materials resulting from the manufacturing process of the processed product to an antibody against at least one of the polypeptides comprising the amino acid sequence (E1)-(E50). Details of such a method are as described above in the section "Diagnostic Kit / Diagnostic Method (2)." That is, in the manufacturing method, the "subject's IgE antibody" in the section "Diagnostic Kit / Diagnostic Method (2)" above can be replaced with "antibody against at least one of the polypeptides comprising the amino acid sequence (E1)-(E50) above," and the "antigen" and "polypeptide" in the section "Diagnostic Kit / Diagnostic Method (2)" above can be replaced with "sample containing materials resulting from the manufacturing process of the processed product." The method described above in the section "Diagnostic Kit / Diagnostic Method (2)" above can be used to confirm that antigens have been removed or reduced during the manufacturing process of a processed product. In addition, the tester composition described above in the section "Tester composition (2)" can also be used. [Example]

[0230] Examples of the present invention will be described below, but the technical scope of the present invention is not limited to these examples.

[0231] Example 1: Protein pattern confirmation The proteins contained in shrimp (vannamei shrimp, black tiger shrimp, and kuruma prawn) were investigated using the following two-dimensional electrophoresis method.

[0232] Protein extraction The shrimp protein was extracted and purified as follows: Urea buffer was added to the shrimp meat to extract the protein. The composition of the urea buffer was as follows: 30mM Tris 2M thiourea 7M Urea 4% (w / v) CHAPS: 3-[(3-cholamidopropyl)dimethylammonio]propanesulfonate A suitable amount of dilute hydrochloric acid Distilled water was added to the solution to bring the total volume to 100 mL, and the pH was 8.5.

[0233] Then, two precipitation procedures were performed using the 2D-CleanUP kit (GE). In the first precipitation procedure, TCA (trichloroacetic acid) was added to the recovered protein extract, and the resulting precipitate (TCA precipitate) was collected. In the second precipitation procedure, acetone was added to the recovered TCA precipitate, and the resulting precipitate (sample) was collected.

[0234] Preparation of sample solution A portion of the obtained sample (50 μg in terms of protein weight) was dissolved in 150 μl of DeStreak Rehydration Solution (GE), a swelling buffer for the first-dimension isoelectric focusing gel, to prepare the sample solution for first-dimension isoelectric focusing (swelling sample solution). The composition of DeStreak Rehydration Solution is as follows: 7M thiourea 2M urea 4% (w / v) CHAPS 0.5% (v / v) IPG buffer; GE Appropriate amount of BPB (bromophenol blue)

[0235] Permeation of the sample into the first-dimension isoelectric focusing gel A first-dimension isoelectric focusing gel (GE: IPG Gel Immobiline Drystrip (pH 3-10NL)) was immersed in 140 μl of the specimen solution for first-dimension isoelectric focusing (specimen solution for swelling) and allowed to soak overnight at room temperature.

[0236] In this example, an IPGphor manufactured by GE was used as the electrophoresis device.

[0237] The electrophoresis tray was filled with silicone oil, and water-moistened filter paper was placed on both ends of the gel permeated with the sample. The gel was then set on the electrophoresis tray so that it was covered with silicone oil, and the electrodes were set with the filter paper sandwiched between the gel and the tray.

[0238] The upper limit of the current value of the isoelectric focusing device was set to 75 μA per gel, and the voltage program was as follows: (1) a constant voltage step of 300 V was performed up to 750 Vhr (the current change during the 30 minutes of electrophoresis before the end of this step was 5 μA), (2) the voltage was gradually increased to 1000 V over 300 Vhr, (3) the voltage was further gradually increased to 5000 V over 4500 Vhr, and (4) the first-dimension isoelectric focusing was then performed at a constant voltage of 5000 V until the total Vhr reached 12000.

[0239] SDS equilibration of isoelectric focusing gels After the first-dimension isoelectric focusing, the gel was removed from the isoelectric focusing apparatus and immersed in an equilibration buffer containing a reducing agent and shaken at room temperature for 15 minutes. The composition of the equilibration buffer containing a reducing agent was as follows: 100 mM Tris-HCl (pH 8.0) 6M urea 30% (v / v) glycerol 2% (w / v) SDS 1% (w / v) DTT

[0240] Next, the equilibration buffer containing the reducing agent was removed, and the gel was immersed in an equilibration buffer containing an alkylating agent and shaken at room temperature for 15 minutes to obtain an SDS-equilibrated gel. The composition of the equilibration buffer containing the alkylating agent was as follows: 100 mM Tris-HCl (pH 8.0) 6M urea 30% (v / v) glycerol 2% (w / v) SDS 2.5% (w / v) iodoacetamide

[0241] Second dimension SDS-PAGE In this example, the electrophoresis apparatus used was an XCell SureLock Mini-Cell manufactured by Life Technologies, Inc. The second-dimensional electrophoresis gel used was NuPAGE 4-12% Bis-Tris Gels manufactured by Life Technologies, Inc. The electrophoresis buffer solution with the following composition was prepared and used. 50mM MOPS 50mM Tris base 0.1% (w / v) SDS 1mM EDTA

[0242] In this example, an adhesive agarose solution was used in which 0.5% (w / v) Agarose S (manufactured by Nippon Gene Co., Ltd.) and an appropriate amount of BPB (bromophenol blue) were dissolved in the electrophoresis buffer.

[0243] The wells of the SDS-PAGE were thoroughly washed with the above-mentioned electrophoresis buffer, and the washing buffer was then removed. A fully dissolved adhesive agarose solution was then added to the wells. The SDS-equilibrated gel was then immersed in the agarose, and the SDS-equilibrated gel and the second-dimensional electrophoresis gel were brought into close contact with each other using tweezers. After confirming that the agarose had solidified sufficiently with both gels in close contact, electrophoresis was performed at a constant voltage of 200 V for approximately 45 minutes.

[0244] Fluorescent staining of gels The gel was fluorescently stained using SYPRO Ruby (Life Technologies).

[0245] First, the sealed container to be used was thoroughly washed with 98% (v / v) ethanol. After electrophoresis, the second-dimensional electrophoresis gel was removed from the SDS-PAGE machine and placed in the washed sealed container. It was then immersed in an aqueous solution containing 50% (v / v) methanol and 7% (v / v) acetic acid for 30 minutes twice. The aqueous solution was then replaced with water and the gel was immersed for 10 minutes. Next, the second-dimensional electrophoresis gel was immersed in 40 ml of SYPRO Ruby and shaken overnight at room temperature. The SYPRO Ruby was then removed, and the second-dimensional electrophoresis gel was washed with water and then shaken for 30 minutes in an aqueous solution containing 10% (v / v) methanol and 7% (v / v) acetic acid. The aqueous solution was then replaced with water and shaken for at least 30 minutes.

[0246] analysis The second-dimensional electrophoresis gels that had undergone the above series of processes were subjected to fluorescent image scanning using a Typhoon 9500 (GE). The results of two-dimensional electrophoresis of proteins contained in shrimp flesh are shown in Figure 1 (vannamei shrimp), Figure 3 (black tiger prawn), and Figure 5 (kuruma prawn). Molecular weight marker bands can be seen on the left side of the gel photographs in each figure, and the position of the bands indicates a specific molecular weight (kDa).

[0247] Example 2: Antigen confirmation by immunoblotting Antigen confirmation by immunoblotting was performed for vannamei shrimp, black tiger prawn, and kuruma prawn by following the procedure described in Example 1 up to the "second-dimensional SDS-PAGE" step, followed by the following steps of "transfer to membrane," "immunoblot," and "analysis."

[0248] Transfer to membrane The transfer onto the membrane was carried out using the following transfer device and transfer buffer. Transfer device: XCell SureLock Mini-Cell and XCell II Blot Module (Life Technologies) Transfer buffer: NuPAGE transfer buffer (x20) (Life Technologies) was diluted 20 times with milliQ water and used.

[0249] Specifically, proteins in the two-dimensional electrophoresis gel were transferred to a membrane (PVDF membrane) according to the following procedure.

[0250] (1) The PVDF membrane was immersed in 100% methanol, then in milliQ water, and then transferred to a transfer buffer solution to hydrophilize the PVDF membrane.

[0251] (2) The sponge, filter paper, gel after second-dimensional SDS-PAGE, hydrophilic PVDF membrane, filter paper, and sponge were placed in this order, and a constant voltage of 30 V was applied for 1 hour in a transfer device.

[0252] Immunoblot Immunoblots of the membranes were performed using sera from patients allergic to shrimp or sera from non-shrimp-allergic subjects as primary antibodies.

[0253] Membrane immunoblotting was performed according to the following procedure. (1) The transferred membrane was shaken in a 5% skim milk / PBST solution (PBS buffer containing 0.1% of the nonionic surfactant Tween 20) at room temperature for 1 hour. (2) The primary antibody was left to stand in a 5% serum / 5% skim milk / PBST solution at room temperature for 1 hour. (3) Wash with PBST solution (5 minutes x 3 times). (4) Anti-human IgE-HRP (horseradish peroxidase) was used as a secondary antibody and the sample was left to stand at room temperature for 1 hour in a solution diluted 5000 times with 5% skim milk / PBST solution. (5) Wash with PBST solution (5 minutes x 3 times). (6) The sections were left to stand for 5 minutes in Pierce Western Blotting Substrate Plus (Thermo).

[0254] analysis The membrane subjected to the above series of treatments was subjected to fluorescent image scanning using Typhoon 9500 (manufactured by GE).

[0255] Immunoblots using the sera of shrimp-allergic patients were compared with immunoblots using the sera of control subjects without shrimp allergies. Immunoblots using the sera of shrimp-allergic patients for shrimp proteins (Figure 2 for vannamei shrimp, Figure 4 for black tiger prawn, and Figure 6 for kuruma prawn) detected 11 spots that were different from those detected using the sera of non-shrimp-allergic subjects and that were different from known shrimp allergen proteins.

[0256] The molecular weights and isoelectric points of the above 11 spots are as follows (Figs. 2, 4, and 6). Spot 1: Molecular weight 100-200 kDa, pI 5.0-6.0 Spot 2: Molecular weight 60-130 kDa, pI 5.5-9.0 Spot 3: Molecular weight 100-200 kDa, pI 5.0-6.0 Spot 4: Molecular weight 60-130 kDa, pI 5.5-9.0 Spot 5: Molecular weight 80-130 kDa, pI 6.0-8.0 Spot 6: Molecular weight 60-90 kDa, pI 5.0-6.0 Spot 7: molecular weight 55-70 kDa, pI 5.5-8.0 Spot 8: Molecular weight 45-70 kDa, pI 5.0-7.0 Spot 9: Molecular weight 50-60 kDa, pI 6.0-9.0 Spot 10: molecular weight 20-40 kDa, pI 8.0-10.0 Spot 11: molecular weight 15-20 kDa, pI 8.0-10.0

[0257] Example 3: Mass spectrometry and antigen identification The antigens that produced the above three spots were subjected to identification of the amino acid sequences by mass spectrometry.

[0258] Specifically, protein extraction and mass spectrometry were performed according to the following procedure. (1) For shrimp (vannamei shrimp, black tiger shrimp, and kuruma shrimp), protein extraction, two-dimensional electrophoresis, and membrane transfer were performed according to the procedures in Examples 1 and 2, and the shrimp were stained with 0.008% Direct Blue / 40% ethanol / 10% acetic acid by shaking. (2) The sections were then decolorized by treating them with 40% ethanol and 10% acetic acid for 5 minutes three times, washed with water for 5 minutes, and air-dried. (3) The desired spot was cut out with a clean cutter blade and placed in a centrifuge tube. After hydrophilizing the membrane with 50 μL of methanol, it was washed twice with 100 μL of water and then centrifuged. 20 μL of 20 mM NH4HCO3·50% acetonitrile was added. (4) 1 μL of 1 pmol / μL lysyl endopeptidase (WAKO) was added, and the mixture was left to stand at 37°C for 60 minutes. The solution was then collected in a new centrifuge tube. 20 μL of 20 mM NH₄HCO₃·70% acetonitrile was added to the membrane, which was then left to soak for 10 minutes at room temperature. The membrane was then further collected. The membrane was then dissolved in 10 μL of 0.1% formic acid and 4% acetonitrile, and transferred to a tube. (5) The collected solution was dried under reduced pressure, dissolved in 15 μl of solution A (0.1% formic acid, 4% acetonitrile solution), and subjected to mass spectrometry (ESI-TOF6600, manufactured by AB Sciex). (6) Proteins were identified based on the mass data obtained from the mass spectrometer by searching NCBI.

[0259] result Mass spectrometry was performed on each spot, and the following amino acid sequences were detected.

[0260] [Table 2]

[0261] Furthermore, the mass data obtained from the mass spectrometer for each spot was analyzed by NCBI, and each spot was identified as the following protein.

[0262] Spot 1 : The C-terminal portion of myosin heavy chain type 1 (NCBI protein accession number BAM65721.1, GenBank DNA accession number AB758443.1) from the whiteleg shrimp (amino acid sequence: SEQ ID NO: 2, encoding nucleotide sequence: SEQ ID NO: 1) - The C-terminal portion of myosin heavy chain type 1 (NCBI protein accession number BAM65719.1, GenBank DNA accession number AB758441.1) from black tiger shrimp (amino acid sequence: SEQ ID NO: 45, encoding nucleotide sequence: SEQ ID NO: 44) - The C-terminal portion of myosin heavy chain type a (NCBI protein accession number BAK61429.1, GenBank DNA accession number AB613205.1) from Kuruma shrimp (amino acid sequence: SEQ ID NO: 87, encoding nucleotide sequence: SEQ ID NO: 86) Spot 2 : The N-terminal portion of myosin heavy chain type 1 (NCBI protein accession number BAM65721.1, GenBank DNA accession number AB758443.1) from the white shrimp (amino acid sequence: SEQ ID NO: 117, encoding nucleotide sequence: SEQ ID NO: 116) The N-terminal portion of myosin heavy chain type 1 (NCBI protein accession number BAM65719.1, GenBank DNA accession number AB758441.1) from black tiger prawn (amino acid sequence: SEQ ID NO: 141, encoding nucleotide sequence: SEQ ID NO: 140) from myosin heavy chain type a (NCBI protein accession number BAK61429.1, GenBank DNA accession number AB613205.1) from kuruma prawn (amino acid sequence: SEQ ID NO: 146, encoding nucleotide sequence: SEQ ID NO: 145) Spot 3 : The C-terminal portion of myosin heavy chain type 2 (NCBI protein accession number BAM65722.1, GenBank DNA accession number AB758444.1) from the whiteleg shrimp (amino acid sequence: SEQ ID NO: 161, encoding nucleotide sequence: SEQ ID NO: 160) The C-terminal portion of myosin heavy chain type 2 (NCBI protein accession number BAM65720.1, GenBank DNA accession number AB758442.1) from black tiger prawn (amino acid sequence: SEQ ID NO: 179, encoding nucleotide sequence: SEQ ID NO: 178) and the C-terminal portion of myosin heavy chain type b (NCBI protein accession number BAK61430.1, GenBank DNA accession number AB613206.1) from kuruma prawn (amino acid sequence: SEQ ID NO: 230, encoding nucleotide sequence: SEQ ID NO: 229). Spot 4 : The N-terminal portion of myosin heavy chain type 2 (NCBI protein accession number BAM65722.1, GenBank DNA accession number AB758444.1) from the white shrimp (amino acid sequence: SEQ ID NO: 276, encoding nucleotide sequence: SEQ ID NO: 275) The N-terminal portion of myosin heavy chain type 2 (NCBI protein accession number BAM65720.1, GenBank DNA accession number AB758442.1) from black tiger prawn (amino acid sequence: SEQ ID NO: 300, encoding nucleotide sequence: SEQ ID NO: 299). The N-terminal portion of myosin heavy chain type b (NCBI protein accession number BAK61430.1, GenBank DNA accession number AB613206.1) from kuruma prawn (amino acid sequence: SEQ ID NO: 306, encoding nucleotide sequence: SEQ ID NO: 305). Spot 5 : - For Kuruma shrimp, glycogen phosphorylase (NCBI protein accession number BAJ23879.1, GenBank DNA accession number AB596876.1) (amino acid sequence: SEQ ID NO: 362, encoding nucleotide sequence: SEQ ID NO: 361) For vannamei shrimp and black tiger shrimp, the NCBI protein accession number BAJ23879.1 was found to be a hit, indicating that vannamei shrimp and black tiger shrimp have glycogen phosphorylase homologs that could be an antigen for shrimp allergy. Spot 6 : A portion of the hemocyanin subunit L1 from the white shrimp (NCBI protein accession number AHY86471.1, GenBank DNA accession number KF193058.1) (amino acid sequence: SEQ ID NO: 381, encoding nucleotide sequence: SEQ ID NO: 380) Black tiger prawn hemocyanin (NCBI protein accession number AEB77775.1, GenBank DNA accession number JF357966.1) (amino acid sequence: SEQ ID NO: 399, encoding nucleotide sequence: SEQ ID NO: 398) - For Kuruma shrimp, hemocyanin subunit L (NCBI protein accession number ABR14693.1, GenBank DNA accession number EF375711.1) (amino acid sequence: SEQ ID NO: 414, encoding nucleotide sequence: SEQ ID NO: 413) Spot 7 : - Pyruvate kinase 3 from the white shrimp (NCBI protein accession number ABY66598.1, GenBank DNA accession number EU216039.1) (amino acid sequence: SEQ ID NO: 421, encoding nucleotide sequence: SEQ ID NO: 420) For black tiger shrimp, the NCBI protein accession number ABY66598.1 was found, indicating that a pyruvate kinase 3 homolog exists in black tiger shrimp and that it is an antigen for shrimp allergy. Spot 8 : Black tiger shrimp phosphopyruvate hydratase (NCBI protein accession number AAC78141.1, GenBank DNA accession number AF100985.1) (amino acid sequence: SEQ ID NO: 455, encoding nucleotide sequence: SEQ ID NO: 454) -Kuruma shrimp phosphopyruvate hydratase (NCBI protein accession number ALL27930.1, GenBank DNA accession number KR184189.1) (amino acid sequence: SEQ ID NO: 481, encoding nucleotide sequence: SEQ ID NO: 480) For vannamei shrimp, the NCBI protein accession number AAC78141.1 was found to be a hit, indicating that vannamei shrimp contains a homologue of phosphopyruvate hydratase, which may be an antigen for shrimp allergy. Spot 9 : - Mitochondrial ATP synthase subunit alpha precursor from the white shrimp (NCBI protein accession number ADC55251.1, GenBank DNA accession number GQ848643.3) (amino acid sequence: SEQ ID NO: 486, encoding nucleotide sequence: SEQ ID NO: 485) For black tiger prawn and kuruma prawn, the NCBI protein accession number ADC55251.1 was found to be a hit. This indicates that black tiger prawn and kuruma prawn contain a homologue of the mitochondrial ATP synthase subunit alpha precursor, which may be an antigen for shrimp allergy. Spot 10 : Troponin I from the white shrimp (NCBI protein accession number AFW99839.1, GenBank DNA accession number JX683730.1) (amino acid sequence: SEQ ID NO: 520, encoding nucleotide sequence: SEQ ID NO: 519) For black tiger prawn and kuruma prawn, the NCBI protein accession number AFW99839.1 was found to be a hit, indicating that black tiger prawn and kuruma prawn have troponin I homologs that may be an antigen for shrimp allergy. Spot 11 : - For the white shrimp, cyclophilin A (NCBI protein accession number AEP83534.1, GenBank DNA accession number JN546074.1) (amino acid sequence: SEQ ID NO: 541, encoding nucleotide sequence: SEQ ID NO: 540) Black tiger prawn: Cyclophilin A from black tiger prawn (NCBI protein accession number AGS46493.1, GenBank DNA accession number KF214635.1) (amino acid sequence: SEQ ID NO: 549, encoding nucleotide sequence: SEQ ID NO: 548) For kuruma shrimp, the NCBI protein accession AEP83534.1 was found to be a hit, indicating that kuruma shrimp contains a cyclophilin A homologue, which may be an antigen for shrimp allergy.

[0263] Example 4: Epitope Identification Epitopes of shrimp allergen components Epitopes were identified for shrimp allergen components using the following procedure.

[0264] (A) Epitope mapping (1) Epitope mapping was performed using a library of overlapping peptides (15 amino acids in length) corresponding to the amino acid sequences identified as shrimp allergens and the amino acid sequence of arginine kinase. Specifically, a library of overlapping peptides was prepared based on the amino acid sequences of SEQ ID NOs: 117, 141, 146, 2, 45, 87, 276, 161, 230, 179, 362, 381, 399, 421, 455, 481, 486, 520, 541, 300, 306, and arginine kinase.

[0265] Each synthesized peptide was shifted by 10 amino acids, i.e., each peptide overlaps with the previous and subsequent peptides by 5 amino acids each.

[0266] To prepare peptide arrays, we used Intavis CelluSpots™ technology. The peptide arrays were prepared by the following procedure: (1) synthesizing the peptides of interest on amino-modified cellulose discs using an automated synthesizer (Intavis MultiPep RS), (2) dissolving the amino-modified cellulose discs to obtain a cellulose-bound peptide solution, and (3) spotting the cellulose-bound peptides onto coated glass slides. Details of each procedure are as follows:

[0267] (1) Peptide synthesis Peptide synthesis was performed stepwise on amino-modified cellulose discs in a 384-well synthesis plate using 9-fluorenylmethoxycarbonyl (Fmoc) chemistry. Specifically, amino acids bearing Fmoc groups were activated with a solution of N,N'-diisopropylcarbodiimide (DIC) and 1-hydroxybenzotriazole (HOBt) in dimethylformamide (DMF) and added dropwise to the cellulose discs to couple the Fmoc-conjugated amino acids to the amino groups on the cellulose discs. The unreacted amino groups were capped with acetic anhydride and washed with DMF. The Fmoc groups were then removed from the amino groups of the amino acids bound to the cellulose discs by treating with piperidine and washing with DMF. The amino termini were then extended by repeating the coupling, capping, and Fmoc removal steps.

[0268] (2) Dissolution of amino-modified cellulose discs The peptide-bound cellulose disks obtained in the above "(1) Peptide Synthesis" section were transferred to a 96-well plate and treated with a side-chain deprotection mixture of trifluoroacetic acid (TFA), dichloromethane, triisopropylsilane (TIPS), and water to deprotect the amino acid side chains. The deprotected cellulose-bound peptides were then dissolved in a mixture of TFA, trifluoromethanesulfonic acid (TFMSA), TIPS, and water, precipitated with tetrabutyl methyl ether (TBME), resuspended in dimethyl sulfoxide (DMSO), and mixed with a mixture of NaCl, sodium citrate, and water to obtain a peptide solution for slide spotting.

[0269] (3) Spotting of cellulose-binding peptide solution The peptide solution for slide spotting obtained in the above "(2) Dissolution of amino-modified cellulose disc" was spotted onto an Intavis CelluSpots™ slide using an Intavis slide spotting robot, and then dried to prepare a peptide array.

[0270] Using the peptide array, whether or not each peptide fragment binds to IgE antibodies in the serum of shrimp allergy patients through an antigen-antibody reaction was measured according to the following procedure.

[0271] (1) The peptide was shaken in Pierce Protein-Free (PBS) Blocking Buffer (Thermo) at room temperature for 1 hour.

[0272] (2) The cells were shaken overnight at 4°C in 2% serum / Pierce Protein-Free (PBS) Blocking Buffer (Thermo).

[0273] (3) Washing was performed with PBST (PBS buffer containing 3% of the nonionic surfactant Tween 20) for 5 minutes (×3 times).

[0274] (4) Anti-human IgE antibody-HRP (1:20,000, Pierce Protein-Free (PBS) Blocking Buffer (Thermo)) was added, and the mixture was shaken at room temperature for 1 hour.

[0275] (5) Wash with PBST for 5 minutes (x 3 times).

[0276] (6) Pierce ECL Plus Western Blotting Substrate (Thermo) was added and the mixture was shaken at room temperature for 5 minutes.

[0277] (7) Using an Amersham Imager 600, the chemiluminescence of the peptides treated in the above (1) to (6) was measured.

[0278] The chemiluminescence intensity of the images obtained by the measurement in (7) above was quantified using ImageQuant TL (GE Healthcare). The second highest value among the quantified values from the images obtained using the serum of five non-shrimp allergic subjects was defined as the N2nd value, and peptides with a value of 35,000 or more, calculated by subtracting the N2nd value of each peptide from the quantified values from the images obtained using the serum of 37 patients, were determined to be peptides that bound to patient-specific IgE antibodies.

[0279] (B) Epitope mapping (2): overlapping The sequences of peptides to which serum IgE antibodies bound specifically to patients by (A) above (SEQ ID NOs: 558, 566, 582, 586, 594, 599, 614, 624, 634, 640, 654, 671, 672, 678, 681, 686, 691, 697, 704, 705, 708, 717, 725, 729, 741, 750, 752, 765, 770, 778, 788, 793, 810, 817, 826, 833, Based on the sequences of the peptides (847, 858, 865, 879, 881, 892, 908, 912, 915, 917, 928, 935, 939 and 943), a library of overlapping peptide fragments (length: 10 amino acids) was created using a sequence that added the sequences before and after the peptide in the amino acid sequence of the allergen component containing the peptide sequence, and epitope mapping was performed.

[0280] Each synthesized peptide was shifted by one amino acid, i.e., each peptide overlaps with the previous and subsequent peptides by 9 amino acids.

[0281] The library was prepared using the same procedure as in (A) above, and each peptide fragment was assayed for binding to IgE antibodies in patient serum using the same method. The control values were determined by digitizing the images obtained from the results of performing only steps (1), (4), and (6) described in the column for spots of cellulose-bound peptide solution in (3) above. The control values for each peptide were calculated by subtracting the digitized values from the images obtained from the results using patient serum. These values were compared with the values obtained from the peptides used as the basis for overlapping. Peptides whose binding to patient IgE antibodies was lost or significantly reduced by peptides shifted by one amino acid were determined to have no IgE antibody binding ability.

[0282] As in (A) above, the amount of chemiluminescence was quantified for the images obtained by measurement. The quantified values from the images obtained from the results (secondary antibody measurement values) when only (1), (4) to (6) described in the column for the spot of cellulose-binding peptide solution in (3) above were performed were used as control values, and the values quantified from the images obtained from the results using patient serum were subtracted from the control values for each peptide. The overlapping sequences (SEQ ID NOs: 558, 566, 582, 586, 594, 599, 614, 624, 634, 640, 654, 671, 672, 678, 681, 686, 691, 697, 704, 705, 708, 717, 725, 729, 741, 750, 752, 765) were used as the basis for the overlapping. , 770, 778, 788, 793, 810, 817, 826, 833, 847, 858, 865, 879, 881, 892, 908, 912, 915, 917, 928, 935, 939, and 943) was taken as 100%, and a binding affinity of less than 30% was determined to indicate no binding to IgE antibodies, a binding affinity of 30% to but less than 50% was determined to indicate poor binding to IgE antibodies but still binding to IgE antibodies, a binding affinity of 50% to but less than 70% was determined to indicate somewhat poor binding to IgE antibodies but still binding to IgE antibodies, and a binding affinity of 70% or more was determined to indicate no difference in binding to IgE antibodies or good binding to IgE antibodies.

[0283] This analysis identified regions in the overlapping sequences that were important for binding to the patient's IgE antibodies.

[0284] (C) Epitope mapping (3): Alanine glycine scanning For the amino acid sequence identified in (A) above, a library of peptide fragments was prepared by substituting one amino acid at a time from the amino-terminus with alanine (or glycine if the original amino acid was alanine) using a technique called alanine glycine scanning (Non-Patent Document 5), and the binding of IgE antibodies in patient serum was measured using the same technique as above. Amino acids at positions where binding to the patient's IgE antibody was lost or significantly reduced upon alanine glycine substitution were determined to be amino acids important for the expression of the original antigenicity or amino acids that affect the expression of the original antigenicity, while amino acids where binding to the patient's IgE antibody was not lost or significantly reduced were determined to be amino acids that are not important for the expression of the original antigenicity and can be substituted.

[0285] As in (A) above, the chemiluminescence intensity was quantified for the images obtained by measurement. The quantified value from the image obtained from the secondary antibody measurement was used as the control value, and the difference between the quantified value from the image obtained from the results using the serum of 37 patients and the control value for each peptide was calculated. The sequences based on the alanine glycine scan (sequence numbers 558, 566, 582, 586, 594, 599, 614, 624, 634, 640, 654, 671, 672, 678, 681, 686, 691, 697, 704, 705, 708, 717, 725, 729, 741, 750, 752, 765, 770, 778, 788, 793, 810, 820, 830, 840, 850, 860, 870, 880, 890, 900, 910, 920, 930, 940, 950, 960, 970, 980, 990, 991, 1000, 1010, 1020, 1030, 1040, 1050, 1060, 1070, 1080, 1090, 1100, 1111, 1120, 1130, 1140, 1150, 1160, 1170, 1180, 1190, 1200, 1210, 1220, 1230, 1240, 125 When the value obtained by the above methods (17, 826, 833, 847, 858, 865, 879, 881, 892, 908, 912, 915, 917, 928, 935, 939, and 943) was taken as 100%, a peptide with a binding affinity of less than 30% was judged to have no binding affinity to IgE antibodies, a peptide with a binding affinity of 30% to less than 50% was judged to have poor binding affinity to IgE antibodies but still have binding affinity, a peptide with a binding affinity of 50% to less than 70% was judged to have somewhat poor binding affinity to IgE antibodies but still have binding affinity, and a peptide with a binding affinity of 70% or more was judged to have no difference in binding affinity to IgE antibodies or have good binding affinity to IgE antibodies.

[0286] Analysis of the results of (A) to (C) revealed that consensus sequences important for the expression of original antigenicity were found in the regions of the sequences used as the basis for the alanineglycine scan that are important for binding to the patient's IgE antibodies. Sequences were identified for all 50 epitopes, and the results are summarized in Table 3 below.

[0287] [Table 3-1]

[0288] [Table 3-2]

[0289] [Table 3-3]

[0290]

Table 3-4

[0291]

Table 3-5

[0292]

Table 3-6

[0293]

Table 3-7

[0294]

Table 3-8

[0295]

Table 3-9

[0296]

Table 3-10

[0297]

Table 3-11

[0298]

Table 3-12

[0299]

Table 3-13

[0300]

Table 3-14

[0301]

Table 3-15

[0302]

Table 3-16

[0303]

Table 3-17

[0304]

Table 3-18

[0305]

Table 3-19

[0306]

Table 3-20

[0307]

Table 3-21

[0308]

Table 3-22

[0309]

Table 3-23

[0310]

Table 3-24

[0311]

Table 3-25

[0312]

Table 3-26

[0313]

Table 3-27

[0314]

Table 3-28

[0315]

Table 3-29

[0316]

Table 3-30

[0317]

Table 3-31

[0318]

Table 3-32

[0319]

Table 3-33

[0320]

Table 3-34

[0321]

Table 3-35

[0322]

Table 3-36

[0323]

Table 3-37

[0324]

Table 3-38

[0325]

Table 3-39

[0326]

Table 3-40

[0327]

Table 3-41

[0328]

Table 3-42

[0329]

Table 3-43

[0330]

Table 3-44

[0331] [Table 3-45]

[0332] [Table 3-46]

[0333] [Table 3-47]

[0334] [Table 3-48]

[0335] [Table 3-49]

[0336] [Table 3-50]

[0337] Furthermore, interview information was obtained indicating that each patient was allergic to the foods listed in Table 4 below.

[0338] [Table 4]

[0339] Furthermore, IgE antibodies from patients shown in Table 5 below, including those shown in Table 3, showed binding to the common 15-residue sequence of (E1)-(E50).

[0340] [Table 5]

[0341] Example 5: Confirmation of epitope cross-reactivity Each epitope sequence found for each shrimp protein in Table 3 above (SEQ ID NOs: 559, 561, 563, 567, 569, 571, 573, 576, 578, 580, 583, 585, 587, 588, 591, 593, 595, 596, 597, 600, 602, 604, 606, 608, 611, 612, 615, 617, 619, 621, 623, 625, 626, 630, 632, 635, 636, 637, 639, 641, 643, 645, 646, 647, 649, 651, 653, 655, 656, 658, 660, 662, 663, 664, 665, 666, 667, 668, 669, 670, 671, 672, 673, 674, 675, 676, 677, 678, 679, 680, 681, 682, 683, 684, 685, 686, 687, 688, 689, 690, 691, 692, 693, 694, 695, 700, 701, 702, 703, 704, 705, 706, 707, 708, 709, 710, 711, 712 64, 666, 668, 669, 673, 676, 679, 680, 682, 683, 684, 687, 689, 692, 694, 695, 698, 700, 702, 703, 706, 707, 709, 711, 713, 715, 719, 721, 723, 724, 726, 727 7, 728, 730, 732, 734, 736, 738, 742, 744, 746, 748, 751, 753, 755, 757, 759, 761, 763, 764, 766, 767, 768, 771, 772, 774, 776, 779, 781, 783, 785, 787, 789 , 791, 794, 795, 797, 799, 801, 803, 805, 808, 811, 813, 815, 818, 820, 822, 824, 827, 829, 831, 834, 836, 838, 840, 842, 845, 848, 850, 851, 852, 854, 856, 859, 861, 863, 866, 868, 870, 872, 875, 877, 880, 882, 884, 885, 887, 889, 893, 895, 897, 899, 900, 902, 904, 906, 909, 910, 913, 914, 916, 918, 920, 921, 9 23, 926, 929, 931, 933, 936, 937, 940, 942, 944, 946 and 947), in which amino acids other than those important for maintaining binding to IgE antibodies are replaced with any amino acid (X) (SEQ ID NOs: 560, 562, 564, 565, 568, 570, 572, 574, 575, 577, 579, 581, 584, 589, 590, 592, 598, 601, 603, 605, 607, 609, 610, 613, 616, 618, 620, 622, 627, 629, 631, 633, 636, 638, 642, 644, 648,650, 652, 657, 659, 661, 665, 667, 670, 674, 675, 677, 685, 688, 690, 693, 696, 701, 710, 712, 714, 716, 720, 722, 731, 733, 735, 737, 739, 740, 743, 745, 747, 749, 754, 756, 758, 760, 762, 769, 773, 775, 777, 780, 782, 784, 786, 790, 792, 796, 798, 800, 802, 804, 806, 807, 809, 812, 814, 816, 819, 821, 823, 825, 828, 830, 832, 835, 837, 839, 841, NCBI searches were conducted to find proteins with the same sequences as the allergen foods that each patient had at the same time for the allergens (843, 844, 846, 849, 853, 855, 857, 860, 862, 864, 867, 869, 871, 873, 874, 876, 878, 984, 883, 886, 888, 890, 891, 894, 896, 898, 901, 903, 905, 907, 911, 919, 922, 924, 925, 927, 930, 932, 934, 938, 941, 945, and 948). As a result, for example, the amino acid sequences contained in the sequences of each food listed in the "Cross-reactivity confirmed foods" column in Tables 3-1 to 3-50 were found to be hits.

[0342] The peptides containing the amino acid sequences listed in the "Synthetic Sequence" and "SEQ ID NO" columns in Table 3 were tested by ELISA for their binding to IgE antibodies from patients allergic to each of the foods listed in the "Foods tested for cross-reactivity" columns in Tables 3-1 to 3-50. The peptides were synthesized using the Fmoc method so that they were biotinylated at the N-terminus.

[0343] Specifically, ELISA was performed according to the following procedure.

[0344] (1) The biotinylated peptide was prepared in PBST (0.1% Tween 20) to a concentration of 10 μg / mL.

[0345] (2) 20 μL of each peptide solution was added to each well of a streptavidin-coated 384-well plate and shaken at room temperature for 1 hour. After recovering the solution, the plate was washed five times with PBS.

[0346] (3) 40 μL of Pierce Protein-Free (PBS) Blocking Buffer (Thermo) was added, and the mixture was shaken at room temperature for 1 hour. After removing the solution, the mixture was washed five times with PBST.

[0347] (4) 20 μL of 2% serum / Canget Signal Solution I (TOYOBO) was added, and the mixture was shaken at room temperature for 1 hour. After removing the solution, the mixture was washed five times with PBST.

[0348] (5) 20 μL of secondary antibody dilution solution (1:10000, Canget Signal Solution II (TOYOBO)) was added, and the mixture was shaken at room temperature for 1 hour. After removing the solution, the mixture was washed five times with PBST.

[0349] (6) 20 μL of 1-Step Ultra TMB-ELISA (Thermo) was added and the mixture was shaken at room temperature for 15 minutes.

[0350] (7) 20 μL of 2M H2SO4 was added, and the absorbance at 450 nm was measured.

[0351] Peptides having these amino acid sequences were prepared in the same manner as in Example 4 (A), and the binding of IgE antibodies in the sera of allergic patients and non-allergic subjects was measured. The sera of two non-allergic subjects were measured, and the values obtained by dividing the measured values by the average were expressed.

[0352] The results are shown in Figures 7-12. The vertical axis of each graph is "absorbance in allergic patients / absorbance in non-allergic patients (healthy individuals)." When there are two bar graphs for the same sequence, different shades of the graphs indicate different patients. For example, graphs No. 8 and No. 10 in Figure 7 show the results for different patients for a peptide with the same sequence. The names of food ingredients listed in each graph are the names of food ingredients containing polypeptides containing the amino acid sequence that forms the basis of each synthetic sequence used.

[0353] As is clear from Figures 7-12, all polypeptides having the amino acid sequences shown in Graph Nos. 1-114 exhibited higher binding affinity (greater than "1") to the IgE antibodies of each allergic patient than to the IgE antibodies in the serum of non-allergic subjects, confirming the cross-reactivity of these polypeptides. This indicates that these epitopes can be used to detect cross-reactivity with antigens other than shrimp. Furthermore, it supports the idea that the "X" moiety can be any amino acid residue.

Claims

1. A polypeptide that specifically binds to the IgE antibody of a shrimp allergy patient, and is any one of the following polypeptides: (E21) (i) a polypeptide comprising the amino acid sequence of SEQ ID NOs: 708-713, 715, 716, 962, 963; (ii) a polypeptide comprising an amino acid sequence in which one or more amino acid residues corresponding to the 1st, 2nd, 4th, 5th, 7th, 8th, 9th, 10th, 12th, 13th, and 15th amino acid residues of SEQ ID NO: 708 are substituted with any amino acid residue; (E48) (i) a polypeptide comprising the amino acid sequence of SEQ ID NO: 935-938; (ii) a polypeptide comprising an amino acid sequence in which one or more amino acid residues corresponding to the 5th and 6th amino acid residues of SEQ ID NO: 935-938 are substituted with any amino acid residue; (E49) (i) a polypeptide comprising the amino acid sequence of SEQ ID NO: 939-942; (ii) a polypeptide comprising an amino acid sequence in which one or more amino acid residues corresponding to the 3rd, 6th, 7th, 10th, and 11th amino acid residues of SEQ ID NO: 939 are substituted with any amino acid residue; (E22) (i) a polypeptide comprising the amino acid sequence of SEQ ID NO: 717-724, 964; (ii) a polypeptide comprising an amino acid sequence in which one or more amino acid residues corresponding to the 2nd, 4th, 5th, 6th, 8th, 9th, 10th, and 12th amino acid residues of SEQ ID NO: 717 are substituted with any amino acid residue; (E23) a polypeptide comprising the amino acid sequence of SEQ ID NO: 725-728; (E50) (i) a polypeptide comprising the amino acid sequence of SEQ ID NO: 943-948; (ii) a polypeptide comprising an amino acid sequence of SEQ ID NOs: 943-948, in which one or more amino acid residues corresponding to the 1st, 2nd, 3rd, 4th, 5th, 8th, 10th, and 11th amino acid residues of SEQ ID NO: 943 are substituted with any amino acid residue; (E19) a polypeptide comprising the amino acid sequence of SEQ ID NO: 704; (E20) A polypeptide comprising the amino acid sequence of SEQ ID NO: 705-707.

2. A diagnostic kit for shrimp allergy comprising at least one polypeptide according to claim 1.

3. A diagnostic composition for shrimp allergy, said diagnostic composition comprising at least one of the polypeptides described in claim 1.

4. 1. A method for providing an indicator for diagnosing shrimp allergy in a subject, comprising the steps of: (i) contacting a sample obtained from a subject with an antigen, wherein the sample is a solution containing IgE antibodies; (ii) detecting binding of IgE antibodies to the antigen in a sample obtained from the subject; (iii) if binding of the subject's IgE antibodies to the antigen is detected, an indication that the subject is allergic to shrimp is provided; wherein the antigen is at least one of the polypeptides of claim 1.

5. A pharmaceutical composition comprising at least one of the polypeptides of claim 1.

6. A pharmaceutical composition according to claim 5 for treating shrimp allergy.

7. A tester composition for determining the presence or absence of a shrimp antigen in a subject, comprising an antibody that binds to at least one of the polypeptides of claim 1.

8. One of the following primers: (a) a primer comprising a portion of the base sequence of a nucleic acid encoding the polypeptide of claim 1 and / or a portion of its complementary strand; or (b) a primer which is a part of at least one of the nucleotide sequences represented by SEQ ID NOs: 485, 519, 540, or 548 and / or a primer which is a part of a sequence complementary to at least one of the nucleotide sequences represented by SEQ ID NOs: 485, 519, 540, or 548; A tester composition for determining the presence or absence of a shrimp antigen in a subject, comprising:

9. A method for determining the presence or absence of the polypeptide described in claim 1 in a raw material or processed product, wherein the raw material or processed product is shrimp or a shrimp processed product, and the method comprises detecting the polypeptide described in claim 1 in the raw material or processed product.

10. A raw material or processed product characterized by having an antigen removed or reduced, wherein the raw material or processed product is shrimp or a shrimp processed product, and the antigen is at least one of the polypeptides described in claim 1.

11. A method for producing a processed product in which antigens have been removed or reduced, the method comprising a step of confirming that the antigens have been removed or reduced during the production process of the processed product, wherein the antigen is at least one of the polypeptides described in claim 1.

Images