Antibody for resisting protein of growth stimulation expression gene 2 and application of antibody

Abstract

The invention discloses an antibody for resisting protein of a growth stimulation expression gene 2 and application of the antibody, and relates to the field of antibodies. The anti-GST2 protein antibody disclosed by the invention comprises a heavy chain complementarity determining region and a light chain complementarity determining region, provides an important raw material source for detection of GST2 protein, and has good activity and sensitivity.

Description

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS

[0002] The present disclosure claims the priority of Chinese patent application No. 202311095438.8 filed with the Chinese Patent Office on August 28, 2023, entitled “An antibody against growth stimulation expression gene 2 protein and its application”, the entire contents of which are incorporated by reference into the present disclosure. Technical Field

[0003] The present invention relates to the field of antibody technology, and in particular to an antibody against growth stimulation expression gene 2 protein and an application thereof. Background Art

[0004] Growth stimulation expressed gene 2 protein (ST2) is a member of the interleukin 1 (IL-1) receptor superfamily, also known as T1, IL1RL1 or Fit1 gene and is located on human chromosome 2q12, about 40KD. The transcription product of the ST2 gene has four subtypes, of which two are most important due to the regulation of different promoters: the first is transmembrane ST2, which consists of three extracellular immunoglobulin G domains, a transmembrane domain and an intracellular domain, and is a membrane receptor member of the interleukin-1 receptor family; the other is soluble ST2. Since soluble ST2 lacks transmembrane and intracellular domains, it can flow freely in the blood, making it detectable in serum.

[0005] Vascular disease (CVD) is the leading cause of death worldwide. Brain natriuretic peptide (BNP) and NT-proBNP are the most famous markers of heart failure (HF). Troponin markers have improved the diagnosis of acute and chronic coronary artery disease. However, a single biomarker can only reflect one aspect of the pathological mechanism of heart failure, and is also affected by kidney function, age, BMI, and pulmonary hypertension, which reduces the accuracy of these biomarkers. With the increasing understanding of the role of ST2 in the cardiovascular system, people have begun to consider the assessment of plasma ST2 levels as a new marker for cardiovascular events, especially those related to heart failure and ischemic heart disease, and clinical conditions. ST2 is not affected by renal function, age, weight, and other indices. ST2 is also considered to be a possible biomarker for patients with acute heart failure and asthma. Studies have also confirmed the role of ST2 in predicting mortality. ST2 can not only be combined with other biomarkers as a prognostic biomarker, but can also be used to monitor the pharmacological response of heart failure. It is related to the recommended therapeutic drugs for heart failure and can provide a reference for the selection of therapeutic drugs and regimens in clinic.

[0006] At present, the main methods used in China to detect growth stimulation expression gene 2 protein include enzyme-linked immunosorbent assay (ELISA), magnetic particle chemiluminescence method, gold label method, etc. Different detection methods require antibodies against growth stimulation expression gene 2 protein.

[0007] Therefore, those skilled in the art have a strong need for anti-GSH2 protein antibodies with good performance. Summary of the invention

[0008] The present application provides an antibody or an antigen-binding fragment thereof against growth stimulating expression gene 2 protein, which provides an important source of raw materials for the detection of growth stimulating expression gene 2 protein and has good activity and sensitivity.

[0009] In order to achieve the above-mentioned object, according to one aspect of the present invention, an antibody or an antigen-binding fragment thereof against growth-stimulating expression gene 2 protein is provided, wherein the antibody or the antigen-binding fragment thereof comprises three complementary determining regions of a heavy chain variable region having an amino acid sequence as shown in any one of SEQ IDNO: 22, 23, 24, and 25, and three complementary determining regions of a light chain variable region having an amino acid sequence as shown in any one of SEQ IDNO: 30, 31, and 32.

[0010] In order to achieve the above object, according to the second aspect of the present invention, an antibody or an antigen-binding fragment thereof against growth-stimulating gene expression 2 protein is provided, wherein the antibody or the antigen-binding fragment thereof comprises the following complementary determining regions:

[0011] HCDR1, which comprises or consists of the amino acid sequence shown in SEQ ID NO: 1;

[0012] HCDR2, which comprises or consists of the amino acid sequence shown in SEQ ID NO: 2;

[0013] HCDR3 comprising or consisting of the amino acid sequence shown in SEQ ID NO: 3, 17 or 18;

[0014] LCDR1, which comprises or consists of the amino acid sequence shown in SEQ ID NO:4;

[0015] LCDR2, which comprises or consists of the amino acid sequence shown in SEQ ID NO:5; and

[0016] LCDR3 comprises or consists of the amino acid sequence shown in SEQ ID NO: 6 or 19.

[0017] In order to achieve the above-mentioned object, according to the third aspect of the present invention, there is provided an antibody or an antigen-binding fragment thereof against growth stimulation expression gene 2 protein, comprising a heavy chain variable region and / or a light chain variable region, wherein the amino acid sequence of the heavy chain variable region is shown in any one of SEQ ID NOs: 22, 23, 24, and 25; and the amino acid sequence of the light chain variable region is shown in any one of SEQ ID NOs: 30, 31, and 32.

[0018] In order to achieve the above-mentioned object, according to the fourth aspect of the present invention, there is provided an antibody or an antigen-binding fragment thereof against growth stimulation expression gene 2 protein, comprising a heavy chain and / or a light chain, wherein the amino acid sequence of the heavy chain is shown in any one of SEQ ID NOs: 26, 27, 28, and 29; and the amino acid sequence of the light chain is shown in any one of SEQ ID NOs: 33, 34, and 35.

[0019] In order to achieve the above object, according to the fifth aspect of the present invention, an antibody conjugate is provided, wherein the antibody conjugate comprises the above antibody or antigen-binding fragment thereof.

[0020] In order to achieve the above object, according to the sixth aspect of the present invention, a reagent or a kit is provided, wherein the reagent or the kit comprises the above-mentioned antibody or antigen-binding fragment thereof or the above-mentioned antibody conjugate.

[0021] In order to achieve the above-mentioned object, according to the seventh aspect of the present invention, a method for detecting growth stimulating expression gene 2 protein is provided, comprising: a) contacting the above-mentioned antibody or its antigen-binding fragment, antibody conjugate, or reagent or kit with the growth stimulating expression gene 2 protein in the sample to be detected under conditions sufficient for an antibody / antigen binding reaction to occur to form an immune complex; and b) detecting the presence of the immune complex, wherein the presence of the complex indicates the presence of the antigen in the test sample.

[0022] In order to achieve the above object, according to the eighth aspect of the present invention, there is provided a use of the above antibody or antigen-binding fragment thereof, antibody conjugate, reagent or kit in the preparation of a product for detecting growth stimulation expression gene 2 protein.

[0023] In order to achieve the above object, the present invention also provides a nucleic acid, a vector, a cell and a method for preparing the above antibody or its antigen-binding fragment. BRIEF DESCRIPTION OF THE DRAWINGS

[0024] In order to more clearly illustrate the technical solutions of the embodiments of the present invention, the drawings required for use in the embodiments are briefly introduced below. It should be understood that the following drawings only show certain embodiments of the present invention and therefore should not be regarded as limiting the scope. For ordinary technicians in this field, other related drawings can be obtained based on these drawings without creative work.

[0025] Figure 1 The results of reducing SDS-PAGE of Anti-ST2 10F9 Rmb1 to Anti-ST2 10F9 Rmb8 are shown. DETAILED DESCRIPTION

[0026] In the first aspect, an embodiment of the present invention provides an antibody or an antigen-binding fragment thereof against growth stimulation expression gene 2 protein, wherein the antibody or the antigen-binding fragment thereof comprises three complementary determining regions of a heavy chain variable region having an amino acid sequence as shown in any one of SEQ ID NOs: 22, 23, 24, and 25, and three complementary determining regions of a light chain variable region having an amino acid sequence as shown in any one of SEQ ID NOs: 30, 31, and 32.

[0027] It should be noted that HCDR1, HCDR2 and HCDR3 are amino acid sequences consistent with HCDR1, HCDR2 and HCDR3 of the same heavy chain variable region defined in the antibody or antigen-binding fragment thereof described in the first aspect, and LCDR1, LCDR2 and LCDR3 are amino acid sequences consistent with LCDR1, LCDR2 and LCDR3 of the same light chain variable region defined in the antibody or antigen-binding fragment thereof described in the first aspect.

[0028] For example, the HCDR1, HCDR2, and HCDR3 are amino acid sequences consistent with HCDR1, HCDR2, and HCDR3 of the heavy chain variable region shown in SEQ ID NO:22; the LCDR1, LCDR2, and LCDR3 are amino acid sequences consistent with LCDR1, LCDR2, and LCDR3 of the light chain variable region shown in SEQ ID NO:30.

[0029] In the present invention, the term "antibody" is used in the broadest sense and may include full-length monoclonal antibodies, bispecific or multispecific antibodies, and chimeric antibodies, so long as they exhibit the desired biological activity.

[0030] In the present invention, the term "complementarity determining region", "CDR" or "CDRs" refers to the highly variable region of the heavy and light chains of immunoglobulins, and refers to the region containing one or more or even all of the major amino acid residues that play a role in the binding of an antibody or antigen-binding fragment to its recognized antigen or epitope. In a specific embodiment of the present invention, CDRs refers to the highly variable region of the heavy and light chains of the antibody.

[0031] In the present invention, the heavy chain complementarity determining region is represented by HCDR, which includes HCDR1, HCDR2 and HCDR3; the light chain complementarity determining region is represented by LCDR, which includes LCDR1, LCDR2 and LCDR3.

[0032] The definition methods of CDR are well known in the art, and the CDR definition methods include: Kabat definition, Chothia definition, IMGT definition, Contact definition and AbM definition. As described herein, "Kabat definition" refers to the definition system described in Kabat et al., US Pept. of Health and Human Services, "Sequence of Proteins of Immunological Interest" (1983). "Chothia definition" refers to Chothia et al., J Mol Biol 196: 901-917 (1987). There are other CDR definition methods that may not strictly follow one of the above schemes, but will still overlap with at least a portion of the CDR region defined by Kabat, although they may be shortened or extended based on predictions or experimental results of specific residues or residue groups. Exemplary defined CDRs are listed in Table 1 below, and the definitions in different documents are slightly different. Given the variable region amino acid sequence of an antibody, a person skilled in the art can routinely determine which residues contain a specific CDR. It should be noted that CDRs defined by other methods not limited to Table 1 also belong to the scope of protection of the present disclosure.

[0033] Table 1: CDR Definition 1

[0034]

[0035]

[0036] 1The numbering of all CDR definitions in Table 1 is based on the Kabat numbering system (see below), with the amino acid numbering on the heavy chain represented by "H+number" and the amino acid numbering on the light chain represented by "L+number". One of ordinary skill in the art can unambiguously map the Kabat numbering system to any variable region sequence without relying on any experimental data other than the sequence itself. As used herein, "Kabat numbering" refers to the numbering system described by Kabat et al., U.S. Patent. of Health and Human Services, "Sequence of Proteins of Immunological Interest" (1983).

[0037] 2 "AbM" as used in Table 1 with a lowercase "b" refers to CDRs defined by Oxford Molecular's "AbM" antibody modeling software.

[0038] 3 If both H35A and H35B are absent, CDR-H1 ends at position 35; if only H35A is present, CDR-H1 ends at position 35A; if both H35A and H35B are present, CDR-H1 ends at position 35B.

[0039] 4 If both H35A and H35B are absent, CDR-H1 ends at position 32; if only H35A is present, CDR-H1 ends at position 33; if both H35A and H35B are present, CDR-H1 ends at position 34.

[0040] 5 If both H35A and H35B are absent, CDR-H1 ends at position 33; if only H35A is present, CDR-H1 ends at position 34; if both H35A and H35B are present, CDR-H1 ends at position 35.

[0041] According to an embodiment of the present invention, the HCDR1, HCDR2, HCDR3, LCDR1, LCDR2 or LCDR3 is defined by any one system or a combination of multiple systems of Kabat, Chothia, IMGT, AbM or Contact.

[0042] In some optional embodiments of the present invention, the HCDR1, HCDR2, HCDR3, LCDR1, LCDR2 and LCDR3 are defined by the Kabat system.

[0043] In some optional embodiments of the present invention, the HCDR1, HCDR2, HCDR3, LCDR1, LCDR2 and LCDR3 are defined by the Chothia system.

[0044] In some optional embodiments of the present invention, the HCDR1, HCDR2, HCDR3, LCDR1, LCDR2 and LCDR3 are defined by the IMGT system.

[0045] In some optional embodiments of the present invention, the HCDR1, HCDR2, HCDR3, LCDR1, LCDR2 and LCDR3 are defined by the AbM system.

[0046] In some optional embodiments of the present invention, the HCDR1, HCDR2, HCDR3, LCDR1, LCDR2 and LCDR3 are defined by a Contact system.

[0047] In some optional embodiments of the present invention, the HCDR1, HCDR2, HCDR3, LCDR1, LCDR2 and LCDR3 are defined by a combination of Kabat, Chothia, IMGT, AbM or Contact systems.

[0048] According to an embodiment of the present invention, the Kabat numbering positions corresponding to the amino acid sequences of HCDR1, HCDR2, HCDR3, LCDR1, LCDR2 or LCDR3 defined by the Kabat, Chothia, AbM or IMGT systems are as follows:

[0049] CDR Kabat AbM IMGT Chothia HCDR1 H31~H35A H26~H35A H26~H34 H26~H33 HCDR2 H50~H65 H50~H58 H51~H57 H52~H56 HCDR3 H95~H102 H95~H102 H93~H102 H95~H102 LCDR1 L24~L34 L24~L34 L27~L32 L24~L34 LCDR2 L50~L56 L50~L56 L50~L51 L50~L56 LCDR3 L89~L97 L89~L97 L89~L97 L89~L97

[0050] In a second aspect, an embodiment of the present invention provides an antibody or an antigen-binding fragment thereof against growth-stimulating gene expression 2 protein, wherein the antibody or the antigen-binding fragment thereof comprises the following complementary determining regions:

[0051] HCDR1, which comprises or consists of the amino acid sequence shown in SEQ ID NO: 1;

[0052] HCDR2, which comprises or consists of the amino acid sequence shown in SEQ ID NO: 2;

[0053] HCDR3 comprising or consisting of the amino acid sequence shown in SEQ ID NO: 3, 17 or 18;

[0054] LCDR1, which comprises or consists of the amino acid sequence shown in SEQ ID NO:4;

[0055] LCDR2, which comprises or consists of the amino acid sequence shown in SEQ ID NO:5; and

[0056] LCDR3 comprises or consists of the amino acid sequence shown in SEQ ID NO: 6 or 19.

[0057] According to an embodiment of the present invention, the HCDRs and LCDRs are defined by the Kabat system.

[0058] In an optional embodiment, the antibody or antigen-binding fragment thereof comprises a CDR sequence of any of the following numbered combinations:

[0059] serial number HCDR1 HCDR2 HCDR3 LCDR1 LCDR2 LCDR3 1 SEQ ID NO:1 SEQ ID NO:2 SEQ ID NO:3 SEQ ID NO:4 SEQ ID NO:5 SEQ ID NO:6 2 SEQ ID NO:1 SEQ ID NO:2 SEQ ID NO:17 SEQ ID NO:4 SEQ ID NO:5 SEQ ID NO:6 3 SEQ ID NO:1 SEQ ID NO:2 SEQ ID NO:18 SEQ ID NO:4 SEQ ID NO:5 SEQ ID NO:6 4 SEQ ID NO:1 SEQ ID NO:2 SEQ ID NO:3 SEQ ID NO:4 SEQ ID NO:5 SEQ ID NO:19

[0060] In the present invention, the "framework region" or "FR" region includes the heavy chain framework region and the light chain framework region, and refers to the region other than the CDR in the heavy chain variable region and the light chain variable region of the antibody; wherein the heavy chain framework region can be further subdivided into adjacent regions separated by CDR, including HFR1, HFR2, HFR3 and HFR4 framework regions; the light chain framework region can be further subdivided into adjacent regions separated by CDR, including LFR1, LFR2, LFR3 and LFR4 framework regions.

[0061] In the present invention, the heavy chain variable region is obtained by arranging and connecting the following numbered CDRs and FRs in the following combinations: HFR1-HCDR1-HFR2-HCDR2-HFR3-HCDR3-HFR4; the light chain variable region is obtained by arranging and connecting the following numbered CDRs and FRs in the following combinations: LFR1-LCDR1-LFR2-LCDR2-LFR3-LCDR3-LFR4.

[0062] In an optional embodiment, the antibody or antigen-binding fragment thereof of the first aspect or the second aspect further has at least one of HFR1, HFR2, HFR3, HFR4, LFR1, LFR2, LFR3 and LFR4;

[0063] The HFR1 comprises / is SEQ ID NO:7 or an amino acid sequence having at least 80% identity thereto;

[0064] The HFR2 comprises / is SEQ ID NO:8 or an amino acid sequence having at least 80% identity thereto;

[0065] The HFR3 comprises / is SEQ ID NO:9 or an amino acid sequence having at least 80% identity thereto;

[0066] The HFR4 comprises / is SEQ ID NO: 10 or an amino acid sequence having at least 80% identity thereto;

[0067] The LFR1 comprises / is SEQ ID NO:11 or an amino acid sequence having at least 80% identity thereto;

[0068] The LFR2 comprises / is SEQ ID NO: 12 or an amino acid sequence having at least 80% identity thereto;

[0069] The LFR3 comprises / is SEQ ID NO: 13 or an amino acid sequence having at least 80% identity thereto;

[0070] The LFR4 comprises / is SEQ ID NO: 14 or an amino acid sequence having at least 80% identity thereto.

[0071] It should be noted that, in other embodiments, the amino acid sequences of the framework regions of the antibodies or antigen-binding fragments thereof provided by the present invention may be at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identical to the above-mentioned corresponding framework regions (SEQ ID NO: 7, 8, 9, 10, 11, 12, 13 or 14).

[0072] In an optional embodiment, the HFR1 comprises / is represented by the amino acid sequence shown in SEQ ID NO:20.

[0073] In an optional embodiment, the LFR3 comprises / is an amino acid sequence as shown in SEQ ID NO:21.

[0074] In a third aspect, an embodiment of the present invention provides an antibody or an antigen-binding fragment thereof against growth stimulation expression gene 2 protein, comprising a heavy chain variable region and / or a light chain variable region, wherein the amino acid sequence of the heavy chain variable region is shown in any one of SEQ ID NOs: 22, 23, 24, and 25, and the amino acid sequence of the light chain variable region is shown in any one of SEQ ID NOs: 30, 31, and 32.

[0075] In an optional embodiment, the heavy chain variable region and the light chain variable region described in the first aspect or the third aspect are selected from any combination of the following:

[0076] combination Heavy chain variable region Light chain variable region 1 SEQ ID NO:22 SEQ ID NO:30 2 SEQ ID NO:23 SEQ ID NO:30 3 SEQ ID NO:24 SEQ ID NO:30 4 SEQ ID NO:25 SEQ ID NO:30 5 SEQ ID NO:22 SEQ ID NO:32 6 SEQ ID NO:22 SEQ ID NO:31 7 SEQ ID NO:25 SEQ ID NO:32 8 SEQ ID NO:25 SEQ ID NO:31 .

[0077] In an optional embodiment, the antibody or antigen-binding fragment thereof described in the first aspect, the second aspect, or the third aspect further comprises a constant region.

[0078] In an alternative embodiment, the constant region includes a heavy chain constant region and / or a light chain constant region.

[0079] In an optional embodiment, the heavy chain constant region is selected from any one of the heavy chain constant regions of IgG, IgA, IgM, IgE, and IgD, or a combination of multiple constant region segments.

[0080] In an alternative embodiment, the heavy chain constant region includes CH1 of IgG, a hinge region of IgG, CH2 of IgM, CH3 of IgM and / or CH4 of IgM.

[0081] In an alternative embodiment, the IgG is selected from IgG1, IgG2, IgG3 or IgG4.

[0082] In an alternative embodiment, the light chain constant region is selected from a kappa-type or a lambda-type light chain constant region.

[0083] In an alternative embodiment, the constant region is derived from a species of cow, horse, dairy cow, pig, sheep, rat, mouse, dog, cat, rabbit, donkey, deer, mink, chicken, duck, goose, turkey, fighting cock or human.

[0084] In an alternative embodiment, the species origin of the constant region is rabbit.

[0085] In an optional embodiment, the heavy chain constant region sequence (CH) is as shown in SEQ ID NO:15, and the light chain constant region (CL) sequence is as shown in SEQ ID NO:16.

[0086] It should be noted that, in other embodiments, the constant region sequence may have at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identity with the above-mentioned constant region (SEQ ID NO: 15 or 16).

[0087] In an optional embodiment, the antigen-binding fragment is selected from any one of F(ab)2, F(ab')2, Fab', Fab, Fv and scFv of the antibody.

[0088] The antigen-binding fragments of the above antibodies usually have the same binding specificity as the antibodies from which they are derived. It is easy for those skilled in the art to understand based on the contents described in the present invention that the antigen-binding fragments of the above antibodies can be obtained by methods such as enzymatic digestion (including pepsin or papain) and / or by chemical reduction to split disulfide bonds. Based on the structure of the complete antibody disclosed in the present invention, those skilled in the art can easily obtain the above antigen-binding fragments.

[0089] The antigen-binding fragments of the above antibodies can also be synthesized by recombinant genetic techniques also known to those skilled in the art or by, for example, an automatic peptide synthesizer, such as those sold by Applied BioSystems and the like.

[0090] In a fourth aspect, the present invention provides an antibody or an antigen-binding fragment thereof against growth stimulation expression gene 2 protein, comprising a heavy chain and / or a light chain, wherein the amino acid sequence of the heavy chain is shown in any one of SEQ ID NOs: 26, 27, 28, and 29, and the amino acid sequence of the light chain is shown in any one of SEQ ID NOs: 33, 34, and 35.

[0091] In an optional embodiment, the antibody or antigen-binding fragment thereof of the first, second, third or fourth aspect above comprises any combination of the following heavy chains and light chains:

[0092] combination Heavy chain Light chain 1 SEQ ID NO:26 SEQ ID NO:33 2 SEQ ID NO:27 SEQ ID NO:33 3 SEQ ID NO:28 SEQ ID NO:33 4 SEQ ID NO:29 SEQ ID NO:33 5 SEQ ID NO:26 SEQ ID NO:35 6 SEQ ID NO:26 SEQ ID NO:34 7 SEQ ID NO:29 SEQ ID NO:35 8 SEQ ID NO:29 SEQ ID NO:34

[0093] In a fifth aspect, the present invention provides an antibody conjugate, wherein the antibody conjugate comprises the above-mentioned antibody or antigen-binding fragment thereof.

[0094] In an optional embodiment, the above-mentioned antibody conjugate further comprises biotin or a biotin derivative conjugated to the antibody or antigen-binding fragment thereof.

[0095] In an optional embodiment, the antibody conjugate further comprises a label coupled to the antibody or antigen-binding fragment thereof.

[0096] In an optional embodiment, the above-mentioned marker refers to a class of substances with properties that can be directly observed by the naked eye or detected or detected by an instrument, such as luminescence, color development, radioactivity, etc., through which qualitative or quantitative detection of the corresponding target can be achieved.

[0097] In an alternative embodiment, the label includes but is not limited to fluorescent dyes, enzymes, radioactive isotopes, chemiluminescent agents and nanoparticle labels.

[0098] In actual use, those skilled in the art can select a suitable marker according to the detection conditions or actual needs. No matter which marker is used, it belongs to the protection scope of the present invention.

[0099] In an optional embodiment, the fluorescent dye is not limited to fluorescein dyes and their derivatives (for example, including but not limited to fluorescein isothiocyanate (FITC), hydroxyfluorescein (FAM), tetrachlorofluorescein (TET), etc. or their analogs), rhodamine dyes and their derivatives (for example, including but not limited to red rhodamine (RBITC), tetramethylrhodamine (TAMRA), rhodamine B (TRITC), etc. or their analogs), Cy series dyes and their derivatives (for example, including but not limited to Cy2, Cy3, Cy3B, Cy3.5, Cy3.7, Cy3.8, Cy3.9, Cy3.10, Cy3.11, Cy3.12, Cy3.13, Cy3.14, Cy3.15, Cy3.16, Cy3.17, Cy3.18, Cy3.19, Cy3.20, Cy3.21, Cy3.22, Cy3.23, Cy3.24, Cy3.25, Cy3.26, Cy3.27, Cy3.28, Cy3.29, Cy3.30, Cy3.31, Cy3.32, Cy3.33, Cy3.34, Cy3.35, Cy3.36, Cy3.37, Cy3.38, Cy3.39, Cy3.40, Cy3.41, Cy3.42, Cy3.43, Cy3.44, Cy3.45, Cy3.46, Cy3.47, Cy3.48, Cy3.49, Cy3.50, Cy3.51, Cy3.52, Cy3.53, Cy3.54, Cy3.55, Cy3.56, Cy3.57, Cy3.58, Cy3.59, Cy3.59, Cy3.59, Cy3.59, Cy3.59, Cy3.5 y5, Cy5.5, Cy3, etc. or their analogs), Alexa series dyes and their derivatives (for example, including but not limited to AlexaFluor350, 405, 430, 488, 532, 546, 555, 568, 594, 610, 33, 647, 680, 700, 750, etc. or their analogs) and protein dyes and their derivatives (for example, including but not limited to phycoerythrin (PE), phycocyanin (PC), allophycocyanin (APC), pre-chlorophyll protein (preCP), etc.).

[0100] In alternative embodiments, the enzyme includes, but is not limited to, horseradish peroxidase, alkaline phosphatase, β-galactosidase, glucose oxidase, carbonic anhydrase, acetylcholinesterase, and 6-phosphoglucose deoxygenase.

[0101] In an optional embodiment, the radioactive isotopes include but are not limited to 212Bi, 131I, 111In, 90Y, 186Re, 211At, 125I, 188Re, 153Sm, 213Bi, 32P, 94mTc, 99mTc, 203Pb, 67Ga, 68Ga, 43Sc, 47Sc, 110mIn, 97Ru, 62Cu, 64Cu, 67Cu, 68Cu, 86Y, 88Y, 121Sn, 161Tb, 166Ho, 105Rh, 177Lu, 172Lu and 18F.

[0102] In an optional embodiment, the chemiluminescent reagent includes but is not limited to luminol and its derivatives, lucigenin, crustacean fluorescein and its derivatives, bipyridine ruthenium and its derivatives, acridinium esters and their derivatives, dioxetanes and their derivatives, lophanes and their derivatives, and peroxyoxalates and their derivatives.

[0103] In an optional embodiment, the nanoparticle markers include, but are not limited to, nanoparticles, colloids, organic nanoparticles, magnetic nanoparticles, quantum dot nanoparticles, and rare earth complex nanoparticles.

[0104] In alternative embodiments, the colloid includes, but is not limited to, colloidal metals, disperse dyes, dye-labeled microspheres, and latex.

[0105] In an alternative embodiment, the colloidal metal includes, but is not limited to, colloidal gold, colloidal silver, and colloidal selenium.

[0106] In an optional embodiment, the colloidal metal is colloidal gold.

[0107] In an optional embodiment, the above-mentioned antibody conjugate further includes a solid phase carrier coupled to the antibody or antigen-binding fragment thereof.

[0108] In an optional embodiment, the solid phase carrier includes but is not limited to magnetic microspheres, plastic microspheres, plastic microparticles, microwell plates, glass, capillaries, nylon and nitrocellulose membranes.

[0109] In a sixth aspect, the present invention provides a reagent or a kit, wherein the reagent or the kit comprises the above-mentioned antibody or antigen-binding fragment thereof or the above-mentioned antibody conjugate.

[0110] As mentioned above, the antibodies or antigen binding fragments thereof in some embodiments or examples of the present invention can effectively bind to the growth stimulating expression gene 2 protein, and therefore, the reagent or kit containing the antibody or antigen binding fragments thereof can effectively detect the growth stimulating expression gene 2 protein qualitatively or quantitatively. The reagent or kit provided by the present invention can be used, for example, for detection involving the specific binding performance of the growth stimulating expression gene 2 protein and its antibody, such as immunoblotting and immunoprecipitation. As mentioned above, the antibodies or antigen binding fragments thereof in some embodiments or examples of the present invention have higher binding activity or affinity with the growth stimulating expression gene 2 protein, and therefore, the reagent or kit containing the antibody or antigen binding fragments thereof has higher detection sensitivity or specificity.

[0111] In a seventh aspect, the present invention provides a method for detecting growth stimulatory expression gene 2 protein, comprising: a) contacting the above-mentioned antibody or antigen-binding fragment thereof, antibody conjugate, reagent or kit with growth stimulatory expression gene 2 protein in a sample to be detected to form an immune complex under conditions sufficient for an antibody / antigen binding reaction to occur; and b) detecting the presence of the immune complex, wherein the presence of the complex indicates the presence of the antigen in the test sample;

[0112] In an alternative embodiment, the immune complex further comprises a second antibody that binds to the antibody or antigen-binding fragment thereof.

[0113] In an optional embodiment, the immune complex further comprises a second antibody, which binds to the growth stimulation expression gene 2 protein.

[0114] In an eighth aspect, the present invention provides use of the above-mentioned anti-Growth Stimulatory Expression Gene 2 protein antibody or its antigen-binding fragment, antibody conjugate or the above-mentioned reagent or kit in the preparation of a product for detecting Growth Stimulatory Expression Gene 2 protein.

[0115] In a ninth aspect, the present invention provides a nucleic acid molecule encoding the above-mentioned antibody or antigen-binding fragment thereof.

[0116] In a tenth aspect, the present invention provides a vector containing the above-mentioned nucleic acid molecule.

[0117] In an eleventh aspect, the present invention provides a cell containing the above-mentioned vector.

[0118] In a twelfth aspect, the present invention provides a method for preparing an anti-Growth Stimulation Expression Gene 2 protein antibody or an antigen-binding fragment thereof, comprising: culturing the cells as described above.

[0119] Based on the amino acid sequence of the anti-growth stimulatory expression gene 2 protein antibody or its antigen-binding fragment disclosed in the present invention, it is easy for those skilled in the art to think of using genetic engineering technology or other technologies (chemical synthesis, recombinant expression) to prepare the anti-growth stimulatory expression gene 2 protein antibody or its antigen-binding fragment, for example, separating and purifying the antibody or its antigen-binding fragment from the culture product of a recombinant cell that can recombinantly express the antibody or its antigen-binding fragment as described in any of the above items, which is easy to achieve for those skilled in the art. Based on this, no matter what technology is used to prepare the anti-growth stimulatory expression gene 2 protein antibody or its antigen-binding fragment of the present invention, it belongs to the protection scope of the present invention.

[0120] In order to make the purpose, technical scheme and advantages of the embodiments of the present invention clearer, the technical scheme in the embodiments of the present invention will be described clearly and completely below. If the specific conditions are not specified in the embodiments, they are carried out according to conventional conditions or conditions recommended by the manufacturer. If the manufacturer of the reagents or instruments used is not specified, they are all conventional products that can be purchased commercially.

[0121] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as those generally understood by those of ordinary skill in the art to which the present disclosure belongs. Although any methods and materials similar or equivalent to those described herein can be used for the practice or testing of the preparations or unit doses herein, some methods and materials are now described. Unless otherwise stated, the techniques adopted or considered herein are standard methods. Materials, methods and examples are illustrative and non-restrictive only.

[0122] The practice of the present invention will employ, unless otherwise indicated, conventional techniques of cell biology, molecular biology (including recombinant techniques), microbiology, biochemistry, and immunology, which are within the capabilities of a skilled artisan. This technique is fully explained in the literature, such as Molecular Cloning: A Laboratory Manual, 2nd Edition (Sambrook et al., 1989); Oligonucleotide Synthesis (MJ Gait, ed., 1984); Animal Cell Culture (RI Freshney, ed., 1987); Methods in Enzymology (Academic Press, Inc.); Handbook of Experimental Immunology (DM Weir and CC Blackwell, eds.); Gene Transfer Vectors for Mammalian Cells (JM Miller and MP Calos, eds., 1987); Current Protocols in Molecular Biology (FM Ausubel et al., eds., 1987); PCR: The Polymerase Chain Reaction (PCR: The Polymerase Chain Reaction) (Academic Press, Inc.); Reaction" (Mullis et al., eds., 1994); and Current Protocols in Immunology (JE Coligan et al., eds., 1991), each of which is expressly incorporated herein by reference.

[0123] The features and performance of the present invention are further described in detail below in conjunction with the embodiments.

[0124] Example 1 Antibody Preparation

[0125] 1. Discovery of rabbit monoclonal antibody against ST2 recombinant protein

[0126] Rabbit monoclonal antibodies against ST2 recombinant protein can be obtained by hybridoma technology, phage display technology or single B cell cloning technology to obtain antibody sequences. The specific implementation method of phage display technology is as follows:

[0127] (1) Rabbit immunization

[0128] The emulsified preparation of ST2 recombinant protein prepared with incomplete Freund's adjuvant was injected subcutaneously to stimulate New Zealand white rabbits aged 4 to 6 weeks to produce an immune response, and the pre- and post-immunization sera were collected on days 0, 14, 28, 42 and 69 for titer testing; rabbits with appropriate titers were selected, the spleens were removed, and spleen cell suspensions were prepared.

[0129] (2) Construction of phage library

[0130] RNA was extracted from rabbit spleen cells and reverse transcribed into cDNA. Specifically designed rabbit antibody gene amplification primers were used to amplify VH and VL gene fragments using cDNA as templates. Then, the VH and VL gene fragments were inserted into the phage vector V02 in sequence by restriction ligation. Finally, the ligated phage plasmid was electrotransformed into TG1 competent cells. The next day, monoclonal colonies were selected for PCR identification and antibody gene sequencing to evaluate the quality of the phage library. After passing the test, the phage library was screened.

[0131] (3) Screening of phage libraries

[0132] The phage library TG1 bacterial liquid was inoculated into a shake flask, and when the appropriate bacterial liquid concentration (OD600 was 0.8-1.0) was cultivated, a helper phage was added to infect for 1 hour, and then, the culture was continued overnight. The bacterial liquid was collected the next day for centrifugation, the supernatant was taken, and the phage library displayed was purified by the method of salting out precipitation. The phage library was selected by the method of magnetic bead selection for 3-4 rounds of selection, and then, the monoclonal phage infection colony was selected for expression of the antibody supernatant, and then the monoclonal phage antibody expression supernatant was screened and identified by the ELISA screening method to obtain the monoclonal phage of the anti-ST2 recombinant protein.

[0133] (4) Phage antibody gene sequencing:

[0134] The monoclonal phage of anti-ST2 recombinant protein was sequenced for antibody gene, and duplicate and invalid sequences were removed through sequence analysis to obtain a unique rabbit monoclonal antibody sequence. The obtained sequence was verified by recombinant expression.

[0135] 2. Expression Plasmid Construction

[0136] pcDNA TM 3.4 vector is a recombinant antibody eukaryotic expression vector constructed, which has been modified to introduce multiple cloning restriction sites and is hereinafter referred to as 3.4A expression vector; based on the variable region genes obtained in the above experiment, VL and VH gene-specific primers were designed, with restriction endonuclease sites and protective bases at both ends, respectively, and 0.71Kb Light Chain gene fragment and 1.41Kb Heavy Chain gene fragment were amplified by PCR amplification method.

[0137] The Heavy Chain and Light Chain gene fragments were double-digested with restriction endonucleases, and the 3.4A vector was double-digested with restriction endonucleases. After the fragments and vectors were purified and recovered, the Heavy Chain gene and the Light Chain gene were respectively connected to the 3.4A expression vector to obtain the recombinant expression plasmids of the Heavy Chain and Light Chain, respectively.

[0138] 3. Recombinant Antibody Sample Preparation

[0139] Resuscitate HEK293 cells in advance and subculture them to 200 ml system to make the cell density reach 3-5×10 6 cells / ml, cell viability>95%; wash the cells by centrifugation, re-dissolve with culture medium, and adjust the cell density to 2.9×10 6 cells / ml, as cell diluent. Prepare plasmid DNA and transfection reagent diluents with culture medium respectively. Add transfection reagent diluent to plasmid DNA diluent, mix well and let stand at room temperature for 15 minutes; slowly add the mixture to cell diluent within 1 minute, mix well and take samples to count, record and observe the viability of cells after transfection, and place them in a 35℃ constant temperature incubator for culture, with a speed of 120rmp and a CO2 content of 8%. Centrifuge and collect samples after 13 days. Affinity purify the supernatant of centrifugation with proteinA affinity chromatography column. Take 6μg of purified antibody for reducing SDS-PAGE, and the electrophoresis is shown in the figure. Two bands are shown after reducing SDS-PAGE, one Mr is 50KD (heavy chain) and the other Mr is 28KD (light chain).

[0140] The obtained antibody was named Anti-ST2 10F9Rmb1. Anti-ST2 10F9Rmb1 was mutated to obtain a mutant antibody. The sequences of the heavy chain (H) and light chain (L) of the above antibody are shown in the following table:

[0141] Table 2: Antibody sequences

[0142] Antibody Name Heavy chain Light chain Anti-ST2 10F9Rmb1 SEQ ID NO:26 SEQ ID NO:33 Anti-ST2 10F9Rmb2 SEQ ID NO:27 SEQ ID NO:33 Anti-ST2 10F9Rmb3 SEQ ID NO:28 SEQ ID NO:33 Anti-ST2 10F9Rmb4 SEQ ID NO:29 SEQ ID NO:33 Anti-ST2 10F9Rmb5 SEQ ID NO:26 SEQ ID NO:35 Anti-ST2 10F9Rmb6 SEQ ID NO:26 SEQ ID NO:34 Anti-ST2 10F9Rmb7 SEQ ID NO:29 SEQ ID NO:35 Anti-ST2 10F9Rmb8 SEQ ID NO:29 SEQ ID NO:34

[0143] Example 2 Performance testing of antibodies

[0144] 1. Activity identification

[0145] The coating solution (main component NaHCO3) was used to dilute ST2 recombinant antigen (from Feipeng Bio) to 1ug / ml, 100uL per well, and incubated at 4°C overnight; the next day, the cells were washed twice with washing solution (main component Na2HPO4+NaCl), and patted dry; blocking solution (20% BSA+80% PBS) was added, 120uL per well, incubated at 37°C for 1h, and patted dry; the diluted purified antibody and control antibody were added, 100uL / well, incubated at 37°C for 30min; the cells were washed 5 times with washing solution, and patted dry; goat anti-rabbit IgG-HRP was added, 100uL per well, incubated at 37°C for 30min; the cells were washed 5 times with washing solution, and patted dry; colorimetric solution A (50uL / well) and colorimetric solution B (50uL / well) were added, and the cells were incubated for 10min; stop solution was added, 50uL / well; OD value was read at 450nm (reference 630nm) on the microplate reader.

[0146] Note: Solution A (main ingredients: citric acid + sodium acetate + acetanilide + urea peroxide); Solution B (main ingredients: citric acid + EDTA·2Na + TMB + concentrated HCl); Stop solution (EDTA·2Na + concentrated H2SO4)

[0147] Table 3: Activity data

[0148] Concentration (ng / ml) 3.13 1.56 0.78 0.39 0.2 0 Comparison 1.203 0.834 0.419 0.292 0.039 0.012 Anti-ST2 10F9Rmb1 1.717 1.137 0.716 0.406 0.239 0.016 Anti-ST2 10F9Rmb2 1.687 1.189 0.762 0.427 0.245 0.017 Anti-ST2 10F9Rmb3 1.498 0.934 0.598 0.325 0.194 0.011 Anti-ST2 10F9Rmb4 1.647 1.086 0.686 0.385 0.24 0.018 Anti-ST2 10F9Rmb5 1.566 0.998 0.633 0.357 0.212 0.016 Anti-ST2 10F9Rmb6 1.613 1.03 0.671 0.369 0.211 0.008 Anti-ST2 10F9Rmb7 1.745 1.201 0.752 0.424 0.262 0.019 Anti-ST2 10F9Rmb8 1.757 1.071 0.641 0.492 0.275 0.018

[0149] 2. Stability assessment

[0150] The above antibodies were placed at 4°C (refrigerator), -80°C (refrigerator), and 37°C (constant incubator) for 21 days, and samples were taken for 7 days, 14 days, and 21 days for status observation, and the activity of the 21-day samples was tested. The results showed that there was no obvious protein state change in the antibodies under the three test conditions for 21 days, and the activity did not show a downward trend with the increase of the test temperature, indicating that the above antibodies are stable. Table 5 below shows the OD results of the enzyme immunoassay activity test of the antibody Anti-ST2 10F9Rmb7 for 21 days.

[0151] Table 4: Stability data

[0152] Sample concentration (ng / ml) 1.56 0.78 0.00 4℃, 21-day samples 1.271 0.779 0.011 -80℃, 21 days sample 1.238 0.735 0.011 37℃, 21 days sample 1.226 0.748 0.012

[0153] 3. Chemiluminescence platform performance test

[0154] 3.1 Antibody coating process

[0155] 10mg / mL carboxyl magnetic beads, washed 3 times with MES buffer. Resuspend the magnetic beads with MES buffer, add EDC with a final concentration of 1mg / mL, shake at 25℃, and react for 30min. Resuspend the magnetic beads with MES buffer, add antibody Anti-ST2-A (from Feipeng Bio) with a final concentration of 0.2mg / mL, use this solution as the magnetic particle working solution, shake at 25℃, and react for 120min. Store in Tris buffer at 2-8℃.

[0156] 3.2 Antibody labeling process

[0157] 3mg / ml of antibodies Anti-ST2 10F9Rmb1, Anti-ST2 10F9Rmb2, Anti-ST210F9Rmb3, Anti-ST210F9Rmb4, Anti-ST2 10F9Rmb5, Anti-ST2 10F9Rmb6, Anti-ST210F9Rmb7, Anti-ST2 10F9Rmb8, and control antibody Anti-ST2-B were replaced with PBS (100mM PB, 50mM sodium chloride, pH8.0) using Zeba desalting columns (10K MWCO). Acridinium ester was prepared into a 4mM solution using DMSO. 10eq of acridinium ester was added to the Anti-cTnI-B and Anti-cTnI-11F13-Rmb antibody solutions, respectively, and reacted at 25°C for 2 hours. The excess reagents were removed by desalting, and the acridinium ester-modified antibody was used as the acridinium ester working solution and stored at 4°C for future use.

[0158] 3.3 Detection process

[0159] The test was performed on the Yingkai shine i2910 fully automatic chemiluminescence immunoassay analyzer using the double antibody sandwich method, that is, 20 μL of sample (quality control product containing ST2 at different concentrations), 50 μL of magnetic particle working solution and 50 μL of acridinium ester working solution were added to the instrument in sequence, mixed and incubated for 10 minutes, and the reaction mixture was rinsed after incubation, and the pre-excitation solution and excitation solution were added to detect the relative luminescence intensity (RLU).

[0160] 3.4 Test results

[0161] The detection data of calibrators of different concentrations are shown in Table 5. The results show that on the chemiluminescence platform, the detection binding activity and sensitivity of antibodies Anti-ST210F9Rmb1 to Anti-ST210F9Rmb8 are better than those of the control antibody.

[0162] Table 5: Chemiluminescence platform performance data

[0163]

[0164] Some of the amino acid sequences involved in this application are shown in Table 6:

[0165] Table 6: Amino Acid Sequence Listing

[0166]

[0167]

[0168]

[0169] The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. For those skilled in the art, the present invention may have various modifications and variations. Any modification, equivalent replacement, improvement, etc. made within the spirit and principle of the present invention shall be included in the protection scope of the present invention.

Claims

1. An antibody or an antigen-binding fragment thereof against growth-stimulating gene expression 2 protein, wherein the antibody or the antigen-binding fragment thereof comprises three complementary determining regions of a heavy chain variable region having an amino acid sequence as shown in any one of SEQ ID NOs: 22, 23, 24, and 25, and three complementary determining regions of a light chain variable region having an amino acid sequence as shown in any one of SEQ ID NOs: 30, 31, and 32.

2. The antibody or antigen-binding fragment thereof according to claim 1, characterized in that: The complementarity determining regions of the variable regions are defined by any one of the Kabat, Chothia, IMGT, AbM or Contact systems or a combination of multiple systems.

3. An antibody or antigen-binding fragment thereof against growth-stimulating gene expression 2 protein, characterized in that: The antibody or antigen-binding fragment thereof comprises the following complementarity determining regions: HCDR1, which comprises or consists of the amino acid sequence shown in SEQ ID NO: 1; HCDR2, which comprises or consists of the amino acid sequence shown in SEQ ID NO: 2; HCDR3 comprising or consisting of the amino acid sequence shown in SEQ ID NO: 3, 17 or 18; LCDR1, which comprises or consists of the amino acid sequence shown in SEQ ID NO:4; LCDR2, which comprises or consists of the amino acid sequence shown in SEQ ID NO:5; and LCDR3, which comprises or consists of the amino acid sequence shown in SEQ ID NO: 6 or 19; Optionally, the antibody or antigen-binding fragment thereof comprises a CDR sequence of any of the following numbered combinations: ； Optionally, the antibody or antigen-binding fragment thereof further has at least one of HFR1, HFR2, HFR3, HFR4, LFR1, LFR2, LFR3 and LFR4; The HFR1 comprises SEQ ID NO:7 or an amino acid sequence having at least 80% identity thereto; The HFR2 comprises SEQ ID NO: 8 or an amino acid sequence having at least 80% identity thereto; The HFR3 comprises SEQ ID NO:9 or an amino acid sequence having at least 80% identity thereto; The HFR4 comprises SEQ ID NO: 10 or an amino acid sequence having at least 80% identity thereto; The LFR1 comprises SEQ ID NO: 11 or an amino acid sequence having at least 80% identity thereto; The LFR2 comprises SEQ ID NO: 12 or an amino acid sequence having at least 80% identity thereto; The LFR3 comprises SEQ ID NO: 13 or an amino acid sequence having at least 80% identity thereto; The LFR4 comprises SEQ ID NO: 14 or an amino acid sequence having at least 80% identity thereto.

4. An antibody or antigen-binding fragment thereof against growth-stimulating gene expression 2 protein, comprising a heavy chain variable region and / or a light chain variable region, characterized in that: The amino acid sequence of the heavy chain variable region is shown in any one of SEQ ID NOs: 22, 23, 24, and 25; the amino acid sequence of the light chain variable region is shown in any one of SEQ ID NOs: 30, 31, and 32; Optionally, the combination of the heavy chain variable region and the light chain variable region is selected from any one of the following combinations: ； Optionally, the antibody or antigen-binding fragment thereof further comprises a constant region; Optionally, the constant region includes a heavy chain constant region and / or a light chain constant region; Optionally, the heavy chain constant region is selected from any one of the heavy chain constant regions of IgG, IgA, IgM, IgE, and IgD, or a combination of multiple constant region segments; Optionally, the heavy chain constant region includes CH1 of IgG, hinge region of IgG, CH2 of IgM, CH3 of IgM and / or CH4 of IgM; Optionally, the species of the constant region is cattle, horse, pig, sheep, goat, rat, mouse, dog, cat, rabbit, donkey, deer, mink, chicken, duck, goose or human; Optionally, the species origin of the constant region is rabbit; Optionally, the heavy chain constant region sequence is as shown in SEQ ID NO: 15 or has at least 80% identity thereto; Optionally, the light chain constant region sequence is as shown in SEQ ID NO: 16 or has at least 80% identity thereto; Optionally, the antigen-binding fragment is selected from any one of F(ab')2, Fab', Fab, Fv and scFv of the antibody.

5. An antibody or antigen-binding fragment thereof against growth-stimulating gene expression 2 protein, comprising a heavy chain and / or a light chain, characterized in that: The amino acid sequence of the heavy chain is shown in any one of SEQ ID NOs: 26, 27, 28, and 29; the amino acid sequence of the light chain is shown in any one of SEQ ID NOs: 33, 34, and 35.

6. An antibody conjugate, characterized in that: The antibody conjugate comprises the antibody or antigen-binding fragment thereof according to any one of claims 1 to 5; Optionally, the antibody conjugate further comprises biotin or a biotin derivative conjugated to the antibody or antigen-binding fragment thereof; Optionally, the antibody conjugate further comprises a label coupled to the antibody or antigen-binding fragment thereof; Optionally, the label is selected from fluorescent dyes, enzymes, radioisotopes, chemiluminescent agents and nanoparticle labels; Optionally, the antibody conjugate further comprises a solid phase carrier coupled to the antibody or antigen-binding fragment thereof.

7. A reagent or a kit, characterized in that: The reagent or kit comprises the antibody or antigen-binding fragment thereof according to any one of claims 1 to 5 or the antibody conjugate according to claim 6.

8. A method for detecting growth stimulation expression gene 2 protein, characterized in that: include: a) contacting the antibody or antigen-binding fragment thereof according to any one of claims 1 to 5, the antibody conjugate according to claim 6, or the reagent or kit according to claim 7 with the growth stimulating expression gene 2 protein in the sample to be detected to form an immune complex under conditions sufficient for an antibody / antigen binding reaction to occur; and b) detecting the presence of the immune complex, the presence of the complex indicating the presence of the antigen in the test sample; Optionally, the immune complex further comprises a second antibody, which binds to the antibody or antigen-binding fragment thereof; Optionally, the immune complex further comprises a second antibody, which binds to the growth stimulation expression gene 2 protein.

9. Use of the antibody or antigen-binding fragment thereof according to any one of claims 1 to 5, the antibody conjugate according to claim 6, or the reagent or kit according to claim 7 in the preparation of a product for detecting growth stimulating expression gene 2 protein.

10. A nucleic acid, a vector, a cell or a method for preparing the antibody or antigen-binding fragment thereof according to any one of claims 1 to 5, wherein the nucleic acid encodes the antibody or antigen-binding fragment thereof according to any one of claims 1 to 5; the vector contains a nucleic acid encoding the antibody or antigen-binding fragment thereof according to any one of claims 1 to 5; the cell contains the above-mentioned nucleic acid or vector; and the method comprises the above-mentioned cell.

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