Anti-adiponectin antibodies, reagents and kits for detecting adiponectin

Abstract

The present invention discloses anti-adiponectin antibodies, reagents and kits for detecting adiponectin, and relates to the field of antibodies. The anti-adiponectin antibodies disclosed in the present invention include heavy chain complementary determining regions and light chain complementary determining regions. The antibodies provide an important source of raw materials for the detection of adiponectin and have good affinity or activity.

Description

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS

[0002] The present invention claims priority to Chinese patent application No. 202211197535.3 filed with the Patent Office of China on September 29, 2022, entitled “Anti-adiponectin antibodies or functional fragments thereof, reagents and kits for detecting adiponectin”, the entire contents of which are incorporated by reference into this disclosure. Technical Field

[0003] The present invention relates to the field of antibody technology, in particular to anti-adiponectin antibodies, reagents and kits for detecting adiponectin. Background Art

[0004] Adiponectin (APN), also known as Acrp30, GBP28, AdipoQ, or adiponectin, is a protein hormone primarily secreted by adipocytes. Human adiponectin monomers consist of 244 amino acids and include three regions: an amino-terminal signal sequence, a collagenous domain, and a carboxyl-terminal globular domain. Its serum concentration ranges from 5 to 30 mg / L, representing approximately 0.01% of total human serum protein.

[0005] The main biological function of adiponectin is to regulate the body's energy metabolism, promote the catabolism of glucose and fat, and improve insulin sensitivity. In recent years, an increasing number of studies have shown that adiponectin levels are closely related to cardiovascular risk factors such as obesity, diabetes, hypertension, hyperlipidemia, atherosclerosis, and insulin resistance, and that adiponectin levels in the serum of patients are significantly lower than those in healthy individuals. Adiponectin has the effects of enhancing insulin sensitivity, resisting atherosclerosis, fighting inflammation, and resisting intimal hyperplasia after vascular injury. Therefore, detecting the level of adiponectin in the blood is of great significance for understanding its physiological functions, regulatory mechanisms, and its relationship with the development and progression of diseases such as type 2 diabetes, coronary heart disease, and hypertension.

[0006] Currently, the main method for detecting adiponectin is fluorescent immunochromatography, an immunological detection method based on the specific reaction between antibodies and antigens, which also uses fluorescence to amplify and display the detected signal. Similar immunological detection methods include radioimmunoassay, enzyme-linked immunosorbent assay, and chemiluminescence. These immunological detection methods all require antibodies specific for adiponectin. Therefore, there is a strong demand in the field for antibodies that effectively bind to and detect adiponectin.

[0007] In view of this, the present invention is proposed. Summary of the Invention

[0008] The purpose of the present invention is to provide anti-adiponectin antibodies, reagents and kits for detecting adiponectin.

[0009] The present invention is achieved in that:

[0010] In a first aspect, an embodiment of the present invention provides an anti-adiponectin antibody, comprising HCDR1, HCDR2, HCDR3 and LCDR1, LCDR2, LCDR3, wherein the HCDR1 / HCDR2 / HCDR3 combination is the same as the HCDR1 / HCDR2 / HCDR3 combination contained in the heavy chain variable region shown in any one of SEQ ID NO:20 to SEQ ID NO:26, and the LCDR1 / LCDR2 / LCDR3 combination is the same as the LCDR1 / LCDR2 / LCDR3 combination contained in the light chain variable region shown in SEQ ID NO:27.

[0011] In alternative embodiments, the CDRs are defined by the Kabat, Chothia, AbM, Contact or IMGT systems.

[0012] In a second aspect, an embodiment of the present invention provides an anti-adiponectin antibody, comprising HCDR1, HCDR2, HCDR3 and LCDR1, LCDR2, LCDR3, wherein the amino acid sequence of HCDR1 comprises SEQ ID NO: 1, or as shown in SEQ ID NO: 1; the amino acid sequence of HCDR2 comprises SEQ ID NO: 2, or as shown in SEQ ID NO: 2; the amino acid sequence of HCDR3 comprises SEQ ID NO: 3, 4, 5 or 6, or as shown in SEQ ID NO: 3, 4, 5 or 6; the amino acid sequences of LCDR1, LCDR2, LCDR3 comprise SEQ ID NOs: 7 to 9 in sequence or are as shown in SEQ ID NOs: 7 to 9 in sequence.

[0013] In a third aspect, an embodiment of the present invention provides an anti-adiponectin antibody, which comprises a heavy chain variable region and a light chain variable region, wherein the heavy chain variable region comprises a sequence structure of HFR1-HCDR1-HFR2-HCDR2-HFR3-HCDR3-HFR4, and the light chain variable region comprises a sequence structure of LFR1-LCDR1-LFR2-LCDR2-LFR3-LCDR3-LFR4, and the amino acid sequences of HCDR1, HCDR2, HCDR3, LCDR1, LCDR2, and LCDR3 are the above-mentioned amino acid sequences of HCDR1, HCDR2, HCDR3, LCDR1, LCDR2, and LCDR3.

[0014] In a fourth aspect, an embodiment of the present invention provides an anti-adiponectin antibody, comprising a heavy chain variable region and / or a light chain variable region, wherein the heavy chain variable region comprises the amino acid sequence shown in any one of SEQ ID NOs: 20 to 26; or, consists of the amino acid sequence shown in any one of SEQ ID NOs: 20 to 26; and the light chain variable region comprises the amino acid sequence shown in SEQ ID NO: 27; or, consists of the amino acid sequence shown in SEQ ID NO: 27.

[0015] In a fifth aspect, an embodiment of the present invention provides an anti-adiponectin antibody, comprising a heavy chain and a light chain; the heavy chain comprises the amino acid sequence shown in any one of SEQ ID NOs: 30 to 36; or, consists of the amino acid sequence shown in any one of SEQ ID NOs: 30 to 36; the light chain comprises the amino acid sequence shown in SEQ ID NO: 37; or, consists of the amino acid sequence shown in SEQ ID NO: 37.

[0016] In a sixth aspect, an embodiment of the present invention provides an anti-adiponectin antibody, which binds to the same epitope in the amino acid sequence of adiponectin as the antibody described in the previous embodiment; or, the antibody competitively binds to adiponectin with the antibody described in the previous embodiment.

[0017] In a seventh aspect, an embodiment of the present invention provides an anti-adiponectin antibody conjugate, which includes the antibody described in the above embodiment.

[0018] In an eighth aspect, an embodiment of the present invention provides a reagent or a kit, which includes the antibody as described in the above embodiment or the antibody conjugate as described in the above embodiment.

[0019] In a ninth aspect, an embodiment of the present invention provides a method for detecting adiponectin, comprising: contacting the antibody, antibody conjugate, reagent or kit described in the above embodiments with adiponectin in a sample to be tested to form an immune complex.

[0020] In a tenth aspect, an embodiment of the present invention provides an isolated nucleic acid encoding the antibody described in the above embodiment.

[0021] In the eleventh aspect, an embodiment of the present invention provides a vector comprising the isolated nucleic acid described in the preceding embodiment.

[0022] In a twelfth aspect, an embodiment of the present invention provides a cell containing the isolated nucleic acid described in the preceding embodiment or the vector described in the preceding embodiment.

[0023] In a thirteenth aspect, an embodiment of the present invention provides a method for preparing the antibody described in the preceding embodiment, comprising: culturing the cells described in the preceding embodiment.

[0024] In a fourteenth aspect, embodiments of the present invention provide use of the antibodies, antibody conjugates, reagents or kits described in the preceding embodiments in detecting adiponectin or preparing products for detecting adiponectin.

[0025] The present invention has the following beneficial effects:

[0026] The anti-adiponectin antibody disclosed in the present invention comprises the heavy chain complementary determining region and the light chain complementary determining region. The antibody provides an important source of raw materials for the detection of adiponectin and has improved affinity or activity. BRIEF DESCRIPTION OF THE DRAWINGS

[0027] In order to more clearly illustrate the technical solutions of the embodiments of the present invention, the following briefly introduces the drawings required for use in the embodiments. It should be understood that the following drawings only illustrate certain embodiments of the present invention and therefore should not be regarded as limiting the scope. For ordinary technicians in this field, other relevant drawings can be obtained based on these drawings without paying any creative work.

[0028] Figure 1 These are the results of reducing SDS-PAGE of Anti-ADPN 3D15RMb1 to Anti-ADPN 3D15RMb7. DETAILED DESCRIPTION

[0029] To make the purpose, technical solutions and advantages of the embodiments of the present invention clearer, the technical solutions in the embodiments of the present invention are described clearly and completely below. Where specific conditions are not specified in the embodiments, conventional conditions or conditions recommended by the manufacturer are used. Where the manufacturer of the reagents or instruments is not specified, all are conventional products that can be purchased commercially.

[0030] The features and performance of the present invention are further described in detail below with reference to the embodiments.

[0031] In a first aspect, an embodiment of the present invention provides an anti-adiponectin antibody, comprising HCDR1, HCDR2, HCDR3 and LCDR1, LCDR2, LCDR3, wherein the HCDR1 / HCDR2 / HCDR3 combination is the same as the HCDR1 / HCDR2 / HCDR3 combination contained in the heavy chain variable region shown in any one of SEQ ID NO:20 to SEQ ID NO:26, and the LCDR1 / LCDR2 / LCDR3 combination is the same as the LCDR1 / LCDR2 / LCDR3 combination contained in the light chain variable region shown in SEQ ID NO:27.

[0032] In alternative embodiments, the CDRs are defined by the Kabat, Chothia, AbM, Contact, or IMGT numbering systems.

[0033] 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 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 described in the first aspect.

[0034] For example, the HCDR1, HCDR2, and HCDR3 have amino acid sequences consistent with those of HCDR1, HCDR2, and HCDR3 in the heavy chain variable region shown in SEQ ID NO: 20; and the LCDR1, LCDR2, and LCDR3 have amino acid sequences consistent with those of LCDR1, LCDR2, and LCDR3 in the light chain variable region shown in SEQ ID NO: 27.

[0035] In the present invention, the term "antibody" is used in the broadest sense and may include full-length monoclonal antibodies, bispecific antibodies, multispecific antibodies, chimeric antibodies, or antigen-binding fragments, as long as they exhibit the desired biological activity. A full-length monoclonal antibody is composed of a heavy chain variable region (VH), a light chain variable region (VL), a heavy chain constant region (CH), and a light chain constant region (CL). In the present invention, the heavy chain variable region, light chain variable region, heavy chain constant region, and light chain constant region are represented by VH, VL, CH, and CL, respectively.

[0036] Antigen-binding fragments typically have the same binding specificity as the antibody from which they are derived. Those skilled in the art will readily appreciate, based on the disclosure herein, that such antigen-binding fragments can be obtained by methods such as enzymatic digestion (including pepsin or papain) and / or by chemical reduction to cleave disulfide bonds. Based on the structure of the intact antibody disclosed herein, those skilled in the art can readily obtain such antigen-binding fragments.

[0037] The above-mentioned antigen-binding fragments 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.

[0038] In the present invention, the term "complementarity determining region", "CDR" or "CDRs" refers to the highly variable regions 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 contribute to the binding of an antibody or antigen-binding fragment to its recognized antigen or epitope.

[0039] In the present invention, the term "heavy chain complementarity determining region", "HCDR" or "HCDRs" refers to one or more or even all of the complementarity determining regions in the heavy chain variable region. The three HCDRs contained in the heavy chain variable region include HCDR1, HCDR2 and HCDR3.

[0040] In the present invention, the term "light chain complementarity determining region", "LCDR" or "LCDRs" refers to one or more or even all complementarity determining regions in the light chain variable region. The three LCDRs contained in the light chain variable region include LCDR1, LCDR2 and LCDR3.

[0041] Definitions of CDRs are known in the art and may be defined, for example, according to the Kabat, Chothia, AbM, Contact, or IMGT systems. Other definitions of CDRs may not strictly follow one of these methods but may still overlap at least partially with the Kabat-defined CDRs, although they may be shortened or lengthened based on predictions or experimental results for particular residues or groups of residues. In the present invention, CDRs may refer to those defined by any method known in the art, including combinations of known methods.

[0042] As used herein, the "Kabat definition" refers to the definition system described in Kabat et al., U. S. Pat. of Health and Human Services, "Sequence of Proteins of Immunological Interest" (1983). The "Chothia definition" refers to Chothia et al., J Mol Biol 196:901-917 (1987). Exemplary defined CDRs are listed in Table 1 below, and the definitions in different literature vary slightly. Given the variable region amino acid sequence of an antibody, one skilled in the art can routinely determine which residues comprise a particular CDR. It should be noted that CDRs defined by other methods other than those in Table 1 also fall within the scope of protection of the present disclosure.

[0043] Table 1: CDR Definition 1

[0044] CDR Kabat <![CDATA[AbM 2 ]]> IMGT Chothia HCDR1 <![CDATA[H31~H35 3 ]]> <![CDATA[H26~H35 3 ]]> <![CDATA[H26~H33..5 5 ]]> <![CDATA[H26~H32..34 4 ]]> 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

[0045] 1The numbering of all CDR definitions in Table 1 is based on the Kabat numbering system (see below), with amino acid numbers on the heavy chain represented by "H+number" and amino acid numbers on the light chain represented by "L+number." One of ordinary skill in the art can unambiguously assign this 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 in Kabat et al., U.S. Pat. of Health and Human Services, "Sequence of Proteins of Immunological Interest" (1983).

[0046] 2 "AbM" as used in Table 1 with a lowercase "b" refers to CDRs defined by Oxford Molecular's "AbM" antibody modeling software.

[0047] 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.

[0048] 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.

[0049] 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.

[0050] According to an embodiment of the present invention, the HCDR1, HCDR2, HCDR3, LCDR1, LCDR2 or LCDR3 is defined by any one of the Kabat, Chothia, IMGT, AbM or Contact systems or a combination of multiple systems.

[0051] In some optional embodiments of the present invention, the HCDR1, HCDR2, HCDR3, LCDR1, LCDR2 and LCDR3 are defined by the Kabat system.

[0052] In some optional embodiments of the present invention, the HCDR1, HCDR2, HCDR3, LCDR1, LCDR2 and LCDR3 are defined by the Chothia system.

[0053] In some optional embodiments of the present invention, the HCDR1, HCDR2, HCDR3, LCDR1, LCDR2 and LCDR3 are defined by an IMGT system.

[0054] In some optional embodiments of the present invention, the HCDR1, HCDR2, HCDR3, LCDR1, LCDR2 and LCDR3 are defined by the AbM system.

[0055] In some optional embodiments of the present invention, the HCDR1, HCDR2, HCDR3, LCDR1, LCDR2 and LCDR3 are defined by a Contact system.

[0056] 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.

[0057] 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:

[0058] CDR Kabat AbM IMGT Chothia HCDR1 H31~H35 H26~H35 H26~H33 H26~H32 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

[0059] In a second aspect, an embodiment of the present invention provides an anti-adiponectin antibody, comprising HCDR1, HCDR2, HCDR3 and LCDR1, LCDR2, LCDR3, wherein the amino acid sequence of HCDR1 comprises SEQ ID NO: 1, or as shown in SEQ ID NO: 1; the amino acid sequence of HCDR2 comprises SEQ ID NO: 2, or as shown in SEQ ID NO: 2; the amino acid sequence of HCDR3 comprises SEQ ID NO: 3, 4, 5 or 6, or as shown in SEQ ID NO: 3, 4, 5 or 6; the amino acid sequences of LCDR1, LCDR2, LCDR3 comprise SEQ ID NOs: 7 to 9 in sequence or are as shown in SEQ ID NOs: 7 to 9 in sequence.

[0060] According to an embodiment of the present invention, the HCDR1, HCDR2 and HCDR3 are, respectively, the amino acid sequences of positions 31 to 35 (SEQ ID No: 1), 50 to 65 (SEQ ID No: 2), and 95 to 100C (SEQ ID No: 3, SEQ ID No: 4, SEQ ID No: 5, or SEQ ID No: 6) of the heavy chain variable region according to Kabat numbering; the LCDR1, LCDR2 and LCDR3 include or are, respectively, the amino acid sequences of positions 24 to 34 (SEQ ID No: 7), 50 to 56 (SEQ ID No: 8), and 89 to 95 (SEQ ID No: 9) of the light chain variable region according to Kabat numbering.

[0061] In the present invention, the term "framework region" or "FRs" refers to the region of the heavy and light chain variable regions of an antibody, excluding CDRs, including the heavy chain framework region and the light chain framework region. The heavy chain framework region can be further subdivided into adjacent regions separated by HCDRs, including the HFR1, HFR2, HFR3, and HFR4 framework regions; the light chain framework region can be further subdivided into adjacent regions separated by LCDRs, including the LFR1, LFR2, LFR3, and LFR4 framework regions. In the present invention, the heavy chain framework region is represented by HFR or HFRs; the light chain framework region is represented by LFR or LFRs.

[0062] In the present invention, the heavy chain variable region is formed by connecting the following numbered HCDRs and HFRs in the following arrangement: HFR1-HCDR1-HFR2-HCDR2-HFR3-HCDR3-HFR4; the light chain variable region is formed by connecting the following numbered LCDRs and LFRs in the following arrangement: LFR1-LCDR1-LFR2-LCDR2-LFR3-LCDR3-LFR4. In the present invention, "-" represents a peptide bond.

[0063] In an alternative embodiment, the antibody further comprises HFR1, HFR2, HFR3, HFR4, LFR1, LFR2, LFR3 and LFR4.

[0064] In an optional embodiment, the HFR1 to HFR4 include SEQ ID NO:10 to SEQ ID NO:13, or an amino acid sequence with at least 80% identity thereto; or, are shown in sequence as SEQ ID NO:10 to SEQ ID NO:13, or an amino acid sequence with at least 80% identity thereto; the LFR1 to LFR4 include SEQ ID NO:14 to SEQ ID NO:17, or an amino acid sequence with at least 80% identity thereto; or, are shown in sequence as SEQ ID NO:14 to SEQ ID NO:17, or an amino acid sequence with at least 80% identity thereto.

[0065] In alternative embodiments, the framework region amino acid sequence of the antibody 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 framework regions.

[0066] In an optional embodiment, the amino acid sequence of HFR1 is shown in SEQ ID NO:18.

[0067] In an optional embodiment, the amino acid sequence of HFR3 is shown in SEQ ID NO:19.

[0068] In a third aspect, an embodiment of the present invention provides an anti-adiponectin antibody, which comprises a heavy chain variable region and a light chain variable region, wherein the heavy chain variable region comprises a sequence structure of HFR1-HCDR1-HFR2-HCDR2-HFR3-HCDR3-HFR4, and the light chain variable region comprises a sequence structure of LFR1-LCDR1-LFR2-LCDR2-LFR3-LCDR3-LFR4, and the amino acid sequences of the HCDR1, HCDR2, HCDR3, LCDR1, LCDR2, and LCDR3 are the above-mentioned amino acid sequences of HCDR1, HCDR2, HCDR3, LCDR1, LCDR2, and LCDR3, and the amino acid sequences of the HFR1, HFR2, HFR3, HFR4, LFR1, LFR2, LFR3, and LFR4 are the above-mentioned amino acid sequences of HFR1, HFR2, HFR3, HFR4, LFR1, LFR2, LFR3, and LFR4.

[0069] In a fourth aspect, embodiments of the present invention provide an anti-adiponectin antibody, comprising a heavy chain variable region and / or a light chain variable region, wherein the heavy chain variable region comprises or consists of the amino acid sequence of any one of SEQ ID NOs: 20 to 26. In an alternative embodiment, the light chain variable region comprises or consists of the amino acid sequence of SEQ ID NO: 27.

[0070] In an alternative embodiment, the antibody further comprises a constant region.

[0071] In alternative embodiments, the constant region comprises a heavy chain constant region and / or a light chain constant region.

[0072] In an optional embodiment, the heavy chain constant region is selected from the heavy chain constant region of any one of IgG1, IgG2, IgG3, IgG4, IgA, IgM, IgE and IgD; and the light chain constant region is selected from the κ or λ type light chain constant region.

[0073] In an optional embodiment, the species origin of the constant region is any one of cow, horse, pig, sheep, goat, rat, mouse, dog, cat, rabbit, donkey, deer, mink, chicken, duck, goose and human.

[0074] In an alternative embodiment, the species origin of the constant region is mouse.

[0075] In an alternative embodiment, the heavy chain constant region comprises an amino acid sequence as shown in SEQ ID NO: 28, or having at least 80% identity thereto; or, consists of an amino acid sequence as shown in SEQ ID NO: 28, or having at least 80% identity thereto. In an alternative embodiment, the light chain constant region comprises an amino acid sequence as shown in SEQ ID NO: 29, or having at least 80% identity thereto; or, consists of an amino acid sequence as shown in SEQ ID NO: 29, or having at least 80% identity thereto.

[0076] Specifically, the constant region sequence can 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 constant region (SEQ ID NO: 28 or 29).

[0077] In an optional embodiment, the antibody includes a heavy chain and a light chain; the heavy chain contains a sequence structure of VH-CH, the light chain contains a sequence structure of VL-CL, and the amino acid sequences of VH, VL, CH, and CL are the amino acid sequences of VH, VL, CH, and CL described in any of the above embodiments.

[0078] In a fifth aspect, an embodiment of the present invention provides an anti-adiponectin antibody, which comprises a heavy chain and / or a light chain, and the antibody includes a heavy chain and a light chain; the heavy chain includes the amino acid sequence shown in any one of SEQ ID NOs: 30 to 36; or, consists of the amino acid sequence shown in any one of SEQ ID NOs: 30 to 36; the light chain includes the amino acid sequence shown in SEQ ID NO: 37; or, consists of the amino acid sequence shown in SEQ ID NO: 37.

[0079] On the other hand, an embodiment of the present invention provides an anti-adiponectin antibody, which binds to the same epitope in the adiponectin amino acid sequence as any of the antibodies described above; or, the antibody competes with any of the antibodies described above for binding to adiponectin.

[0080] In an alternative embodiment, the antibody has a KD ≤ 5.87 × 10 -7 M binds adiponectin with high affinity.

[0081] In an alternative embodiment, the antibody has a KD ≤ 10 -8 M, KD≤10 -9 M, KD≤10 -10 M, KD≤10 -11 M or KD≤10 -12 M binds adiponectin with high affinity.

[0082] In an alternative embodiment, the antibody has a KD ≤ 4.83 × 10 -9 M binds adiponectin with high affinity.

[0083] In an optional embodiment, the antibody is selected from any one of F(ab')2, Fab', Fab, Fv and scFv.

[0084] On the other hand, an embodiment of the present invention further provides an anti-adiponectin antibody conjugate, which includes the antibody described in the above embodiment.

[0085] In an optional embodiment, the antibody conjugate further comprises biotin or a biotin derivative conjugated to the antibody.

[0086] In an optional embodiment, the antibody conjugate further comprises a label coupled to the antibody.

[0087] 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.

[0088] In an optional embodiment, the label is selected from at least one of fluorescent dyes, enzymes, radioisotopes, chemiluminescent reagents and nanoparticle labels.

[0089] 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 falls within the scope of protection of the present invention.

[0090] In an optional embodiment, the fluorescent dyes include but are 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, C 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), peridinin-chlorophyll protein (preCP), etc.).

[0091] In alternative embodiments, the enzyme includes, but is not limited to, horseradish peroxidase, alkaline phosphatase, β-galactosidase, glucose oxidase, carbonic anhydrase, acetylcholinesterase, and glucose-6-phosphate deoxidase.

[0092] 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.

[0093] In an optional embodiment, the chemiluminescent reagent includes but is not limited to luminol and its derivatives, lucigenin, crustacean fluorescein and its derivatives, ruthenium bipyridine and its derivatives, acridinium esters and their derivatives, dioxetanes and their derivatives, lophanes and their derivatives, and peroxalates and their derivatives.

[0094] 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.

[0095] In alternative embodiments, the colloid includes, but is not limited to, colloidal metals, colloidal selenium, disperse dyes, dye-labeled microspheres, and latex.

[0096] In an optional embodiment, the colloidal metal includes but is not limited to colloidal gold or colloidal silver.

[0097] In an optional embodiment, the colloidal metal is colloidal gold.

[0098] In an optional embodiment, the antibody conjugate further comprises a solid phase carrier coupled to the antibody. In the antibody conjugate, the antibody is coupled to the solid phase carrier.

[0099] In an alternative embodiment, the solid support is selected from microspheres, plates, and membranes.

[0100] In an optional embodiment, the solid phase includes but is not limited to magnetic microspheres, plastic microspheres, plastic microparticles, microplates, glass, capillaries, nylon and nitrocellulose membranes.

[0101] In an optional embodiment, the solid phase carrier is a nitrocellulose membrane.

[0102] On the other hand, an embodiment of the present invention further provides a reagent or kit, which includes the antibody as described in any of the foregoing embodiments or the antibody conjugate as described in any of the foregoing embodiments.

[0103] In an alternative embodiment, the reagent or kit has improved detection sensitivity or specificity.

[0104] On the other hand, an embodiment of the present invention further provides a method for detecting adiponectin, comprising:

[0105] The antibody described in any of the preceding embodiments, or the antibody conjugate described in any of the preceding embodiments, or the reagent or kit described in any of the preceding embodiments is contacted with adiponectin in the sample to be detected to form an immune complex.

[0106] In a preferred embodiment, the immune complex further comprises a second antibody that binds to the antibody.

[0107] In a preferred embodiment, the immune complex further comprises a second antibody, which binds to adiponectin.

[0108] On the other hand, embodiments of the present invention further provide use of the antibody or functional fragment described in any of the foregoing embodiments, or the antibody conjugate described in any of the foregoing embodiments, or the reagent or kit described in any of the foregoing embodiments in detecting adiponectin or preparing a product for detecting adiponectin.

[0109] On the other hand, an embodiment of the present invention further provides the use of the antibody or functional fragment described in any of the preceding embodiments, or the antibody conjugate described in any of the preceding embodiments, or the reagent or kit described in any of the preceding embodiments in the preparation of a product having at least one of the following uses, wherein the uses include: diagnosing or assisting in the diagnosis of diseases related to adiponectin metabolism, and predicting or assisting in the prediction of the prognosis and efficacy of diseases related to adiponectin metabolism.

[0110] In an alternative embodiment, the adiponectin metabolism-related disease includes cardiovascular disease.

[0111] In an optional embodiment, the cardiovascular disease is at least one of diabetes, hypertension, hyperlipidemia, atherosclerosis, and coronary heart disease.

[0112] In an alternative embodiment, the product comprises a reagent or a kit.

[0113] On the other hand, an embodiment of the present invention further provides an isolated nucleic acid encoding the antibody described in any of the above embodiments.

[0114] On the other hand, an embodiment of the present invention further provides a vector comprising the isolated nucleic acid described in any of the aforementioned embodiments.

[0115] On the other hand, an embodiment of the present invention further provides a cell comprising the isolated nucleic acid described in any of the foregoing embodiments or the vector described in any of the foregoing embodiments.

[0116] On the other hand, an embodiment of the present invention further provides a method for preparing the antibody as described in any of the aforementioned embodiments, comprising: culturing the cell as described in any of the aforementioned embodiments.

[0117] Based on the amino acid sequence of the antibody disclosed in the present invention, those skilled in the art can easily conceive of using genetic engineering technology or other technologies (chemical synthesis, recombinant expression) to prepare the antibody. For example, the antibody can be isolated and purified from the culture product of recombinant cells that can recombinantly express the antibody as described in any of the above items. This is easy for those skilled in the art to achieve. Based on this, regardless of the technology used to prepare the antibody of the present invention, it falls within the scope of protection of the present invention.

[0118] To make the purpose, technical solutions and advantages of the embodiments of the present invention clearer, the technical solutions in the embodiments of the present invention are described clearly and completely below. Where specific conditions are not specified in the embodiments, conventional conditions or conditions recommended by the manufacturer are used. Where the manufacturer of the reagents or instruments is not specified, all are conventional products that can be purchased commercially.

[0119] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as those generally understood by those skilled in the art to which this disclosure belongs. Although any methods and materials similar or equivalent to those described herein can be used in the practice or testing of the preparations or unit doses herein, some methods and materials are now described. Unless otherwise indicated, the techniques employed or contemplated herein are standard methods. Materials, methods, and examples are illustrative and non-limiting only.

[0120] 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 person skilled in the art. The technique is fully explained in the literature, for example, in Molecular Cloning: A Laboratory Manual, 2nd ed. (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); and PCR: The Polymerase Chain Reaction. 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.

[0121] The features and performance of the present invention are further described in detail below with reference to the embodiments.

[0122] Example 1 Preparation of Anti-ADPN 3D15 Monoclonal Antibody

[0123] In this example, restriction endonucleases and Prime Star DNA polymerase were purchased from Takara. MagExtractor-RNA extraction kit was purchased from TOYOBO. BD SMART TMThe RACE cDNA Amplification Kit was purchased from Takara. The pMD-18T vector was also purchased from Takara. The plasmid extraction kit was purchased from Tiangen. Primer synthesis and gene sequencing were performed by a gene sequencing company.

[0124] 1. Expression Plasmid Construction

[0125] In this example, restriction endonucleases and Prime Star DNA polymerase were purchased from Takara. MagExtractor-RNA extraction kit was purchased from TOYOBO. BD SMART TM The RACE cDNA Amplification Kit was purchased from Takara. The pMD-18T vector was also purchased from Takara. The plasmid extraction kit was purchased from Tiangen. Primer synthesis and gene sequencing were performed by a gene sequencing company.

[0126] 1.1 Anti-ADPN 3D15 Antibody Gene Preparation

[0127] mRNA was extracted from a hybridoma cell line secreting the Anti-ADPN 3D15 monoclonal antibody. DNA was obtained by RT-PCR and inserted into the pMD-18T vector. The DNA was then transformed into DH5α competent cells. After colonies were grown, four positive clones each for the Heavy Chain and Light Chain genes were collected and sent to a gene sequencing company for sequencing.

[0128] 1.2 Sequence Analysis of the Anti-ADPN 3D15 Antibody Variable Region Gene

[0129] The gene sequences obtained by the above sequencing were placed in the Kabat antibody database for analysis, and VNTI11.5 software was used for analysis to determine that the genes amplified by the heavy chain and light chain primer pairs were correct. Among them, in the gene fragment amplified by the Light Chain, the VL gene sequence was 321 bp, preceded by a 57 bp leader peptide sequence; in the gene fragment amplified by the Heavy Chain primer pair, the VH gene sequence was 363 bp, belonging to the VH1 gene family, and preceded by a 57 bp leader peptide sequence.

[0130] 1.3 Construction of recombinant antibody expression plasmid

[0131] pcDNA TM 3.4 Vector is a constructed recombinant antibody eukaryotic expression vector, which has been modified to introduce multiple cloning restriction enzyme sites and is subsequently referred to as the 3.4A expression vector. Based on the sequencing results of the antibody variable region genes in the above-mentioned pMD-18T, specific primers for the VL and VH genes of the Anti-ADPN 3D15 antibody were designed, with restriction endonuclease sites and protective bases at both ends, respectively. PCR amplification was used to amplify the 0.72KB Light Chain gene fragment and the 1.42kb Heavy Chain gene fragment.

[0132] 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 Light Chain gene were respectively connected to the 3.4A expression vector to obtain the recombinant expression plasmids of Heavy Chain and Light Chain, respectively.

[0133] 2. Recombinant Antibody Sample Preparation

[0134] Resuscitate HEK293 cells in advance and subculture them into 200 ml system to make the cell density reach 3-5×10 6 cells / ml, cell viability>95%; wash the cells by centrifugation, resolubilize with culture medium, and adjust the cell density to 2.9×10 6 cells / ml, as the cell dilution. Use culture medium to prepare plasmid DNA and transfection reagent dilution, respectively. Add the transfection reagent dilution to the plasmid DNA dilution, mix well, and let it stand at room temperature for 15 minutes; slowly add the mixture to the cell dilution within 1 minute, mix well, take samples and count, record and observe the viability of the cells after transfection, and place them in a 35°C constant temperature incubator for culture, speed of 120 rpm, CO2 content of 8%, and centrifuge after 13 days. The centrifuged supernatant is affinity purified using a protein A affinity chromatography column. Take 6μg of the purified antibody for reducing SDS-PAGE, and the electrophoresis is shown in the figure. After reducing SDS-PAGE, two bands are shown, one with Mr of 50KD (heavy chain) and the other with Mr of 28KD (light chain).

[0135] 3. Affinity and activity optimization

[0136] While the Anti-ADPN 3D15 monoclonal antibody prepared above possessed the ability to bind to the adiponectin antigen, its affinity and activity were suboptimal. Therefore, the applicants performed targeted mutagenesis on the antibody's light and heavy chain CDRs. Specifically, computer simulations were performed on the antibody variable region structure, the structure of the antigen-antibody variable region complex, key amino acid analysis, and mutation design. Bidirectional primers covering the mutation sites were designed and synthesized based on the mutagenesis strategy. Primers were synthesized at both ends of the target DNA, and a high-fidelity PCR reaction was performed. The PCR product was cloned into a vector, and the mutant antibodies were prepared according to the method of step 2 above. After screening, monoclonal antibodies with significantly improved affinity and activity were obtained and designated Anti-ADPN 3D15RMb1 to Anti-ADPN 3D15RMb7. Their heavy and light chain amino acid sequences are shown in the table below.

[0137] Table 2 Antibody sequences

[0138] Sample name Heavy chain sequence number Light chain number Anti-ADPN 3D15RMb1 SEQ ID NO:30 SEQ ID NO:37 Anti-ADPN 3D15RMb2 SEQ ID NO:31 SEQ ID NO:37 Anti-ADPN 3D15RMb3 SEQ ID NO:32 SEQ ID NO:37 Anti-ADPN 3D15RMb4 SEQ ID NO:33 SEQ ID NO:37 Anti-ADPN 3D15RMb5 SEQ ID NO:34 SEQ ID NO:37 Anti-ADPN 3D15RMb6 SEQ ID NO:35 SEQ ID NO:37 Anti-ADPN 3D15RMb7 SEQ ID NO:36 SEQ ID NO:37

[0139] Example 2 Affinity Analysis

[0140] Purified antibodies were diluted in advance, and adiponectin antigen (purchased from HYTEST, 8AN7) was serially diluted. Binding and dissociation curves of the antigen and antibody were measured on a Biacore 8K+ instrument using a CM5 chip pre-coupled with goat anti-mouse IgG. The instrument automatically fitted the affinity constant, association rate, and dissociation rate. The results showed that the KD values of Anti-ADPN 3D15RMb1 to Anti-ADPN 3D15RMb7 were all within 1.0×10 -9 M to 1.0×10 -12 M, better than the control (KD represents the equilibrium dissociation constant, i.e., affinity constant; ka represents the association rate; kd represents the dissociation rate).

[0141] Table 3 Affinity analysis data

[0142] Sample name KD(M) ka kd Control Antibody 5.87E-07 1.53E+03 8.98E-04 Anti-ADPN 3D15RMb1 3.26E-12 3.84E+05 1.25E-06 Anti-ADPN 3D15RMb2 2.07E-10 3.26E+05 6.76E-05 Anti-ADPN 3D15RMb3 3.78E-10 3.25E+05 1.23E-04 Anti-ADPN 3D15RMb4 3.47E-10 3.69E+05 1.28E-04 Anti-ADPN 3D15RMb5 8.38E-10 2.35E+05 1.97E-04 Anti-ADPN 3D15RMb6 5.51E-10 2.43E+05 1.34E-04 Anti-ADPN 3D15RMb7 5.95E-10 2.25E+05 1.34E-04

[0143] Example 3 Activity Identification

[0144] The coating solution (main component NaHCO3) was diluted with adiponectin antigen (purchased from HYTEST, 8AN7) 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; 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-mouse 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; color development solution A (50uL / well) and color development solution B (50uL / well) were added for 10min; stop solution was added, 50uL / well; the OD value was read at 450nm (reference 630nm) on the microplate reader, and the results showed that Anti-ADPN The activity readouts of 3D15RMb1 to Anti-ADPN 3D15RMb7 were all better than those of the control.

[0145] Table 4 Activity data

[0146] Concentration (ng / ml) 7.81 3.91 1.95 0.98 0.49 0.00 Control Antibody 1.321 0.783 0.401 0.213 0.126 0.038 Anti-ADPN 3D15RMb1 1.950 1.376 0.771 0.372 0.202 0.032 Anti-ADPN 3D15RMb2 1.843 1.470 0.754 0.416 0.234 0.043 Anti-ADPN 3D15RMb3 1.896 1.430 0.799 0.417 0.242 0.035 Anti-ADPN 3D15RMb4 2.084 1.648 0.950 0.540 0.311 0.041 Anti-ADPN 3D15RMb5 2.071 1.445 0.787 0.392 0.190 0.003 Anti-ADPN 3D15RMb6 1.963 1.451 0.790 0.446 0.219 0.012 Anti-ADPN 3D15RMb7 1.987 1.404 0.812 0.406 0.206 0.010

[0147] Example 4 Antibody Stability Assessment

[0148] The above-mentioned antibody was placed at 4°C (refrigerator), -80°C (freezer), and 37°C (incubator) for 21 days. Samples were collected at 7, 14, and 21 days for status observation, and the 21-day sample was tested for activity. The results showed that the protein status of Anti-ADPN 3D15RMb4 antibody did not change significantly under all three test conditions for 21 days, and the activity did not decrease with increasing test temperature, indicating that the expressed antibody is stable. Table 5 below shows the OD results of the enzyme immunoassay activity assay after 21 days of testing.

[0149] Table 5 Stability data

[0150] Sample concentration (ng / ml) 3.91 1.95 0.00 4℃, 21-day samples 1.668 0.921 0.033 -80℃, 21-day sample 1.627 0.992 0.031 37℃, 21-day samples 1.661 0.937 0.031

[0151] The foregoing description is merely a preferred embodiment of the present invention and is not intended to limit the present invention. Those skilled in the art will readily appreciate that various modifications and variations of the present invention are possible. Any modifications, equivalent substitutions, or improvements made within the spirit and principles of the present invention are intended to be within the scope of protection of the present invention.

[0152] Some of the amino acid sequences involved in this application are shown in Table 6 below:

[0153]

[0154]

[0155]

[0156]

Claims

1. An anti-adiponectin antibody comprising HCDR1, HCDR2, HCDR3 and LCDR1, LCDR2, LCDR3, characterized in that: The HCDR1 / HCDR2 / HCDR3 combination is the same as the HCDR1 / HCDR2 / HCDR3 combination contained in the heavy chain variable region shown in any one of SEQ ID NO:20 to SEQ ID NO:26, and the LCDR1 / LCDR2 / LCDR3 combination is the same as the LCDR1 / LCDR2 / LCDR3 combination contained in the light chain variable region shown in SEQ ID NO:27; the HCDR1, HCDR2, HCDR3 and LCDR1, LCDR2, LCDR3 are defined by the Kabat, Chothia, AbM, Contact or IMGT systems.

2. An anti-adiponectin antibody, characterized in that The antibody comprises HCDR1, HCDR2, HCDR3 and LCDR1, LCDR2, LCDR3, the amino acid sequence of the HCDR1 is shown in SEQ ID NO: 1; the amino acid sequence of the HCDR2 is shown in SEQ ID NO: 2; the amino acid sequence of the HCDR3 is shown in SEQ ID NO: 3, 4, 5 or 6; the amino acid sequences of the LCDR1, LCDR2, LCDR3 are shown in SEQ ID NOs: 7 to 9, respectively.

3. The antibody according to any one of claims 1 to 2, further comprising HFR1, HFR2, HFR3, HFR4, LFR1, LFR2, LFR3 and LFR4; characterized in that, The HFR1 to HFR4 are amino acid sequences having at least 80% identity with SEQ ID NO: 10 to SEQ ID NO: 13, respectively; the LFR1 to LFR4 are amino acid sequences having at least 80% identity with SEQ ID NO: 14 to SEQ ID NO: 17, respectively.

4. The antibody according to claim 3, characterized in that The HFR1 to HFR4 are shown in SEQ ID NO: 10 to SEQ ID NO: 13, and the LFR1 to LFR4 are shown in SEQ ID NO: 14 to SEQ ID NO: 17, respectively.

5. An anti-adiponectin antibody, characterized in that The antibody comprises a heavy chain variable region and a light chain variable region, the heavy chain variable region comprises a sequence structure of HFR1-HCDR1-HFR2-HCDR2-HFR3-HCDR3-HFR4, the light chain variable region comprises a sequence structure of LFR1-LCDR1-LFR2-LCDR2-LFR3-LCDR3-LFR4, the amino acid sequence of the HCDR1, HCDR2, HCDR3, LCDR1, LCDR2, and LCDR3 is the amino acid sequence of the HCDR1, HCDR2, HCDR3, LCDR1, LCDR2, and LCDR3 according to any one of claims 1 to 2, and the amino acid sequence of the HFR1, HFR2, HFR3, HFR4, LFR1, LFR2, LFR3, and LFR4 is the amino acid sequence of HFR1, HFR2, HFR3, HFR4, LFR1, LFR2, LFR3, and LFR4 according to claim 3.

6. An anti-adiponectin antibody, characterized in that The antibody comprises a heavy chain variable region and 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: 20 to 26; and the amino acid sequence of the light chain variable region is shown in SEQ ID NO:

27.

7. The antibody according to any one of claims 1 to 2, 4 to 6, characterized in that The antibody also comprises a constant region.

8. The antibody according to claim 7, characterized in that The constant region includes a heavy chain constant region and a light chain constant region.

9. The antibody according to claim 8, characterized in that The heavy chain constant region is selected from the heavy chain constant region of any one of IgG1, IgG2, IgG3, IgG4, IgA, IgM, IgE and IgD; the light chain constant region is selected from the κ type or λ type light chain constant region.

10. The antibody according to claim 7, characterized in that The species origin of the constant region is any one of cattle, horses, pigs, sheep, goats, rats, mice, dogs, cats, rabbits, donkeys, deer, minks, chickens, ducks, geese and humans.

11. The antibody according to claim 7, characterized in that The species origin of the constant region is mouse.

12. The antibody according to claim 8, characterized in that The heavy chain constant region is an amino acid sequence at least 80% identical to SEQ ID NO: 28, and the light chain constant region is an amino acid sequence at least 80% identical to SEQ ID NO:

29.

13. The antibody according to claim 12, characterized in that The amino acid sequence of the heavy chain constant region is shown in SEQ ID NO: 28, and the amino acid sequence of the light chain constant region is shown in SEQ ID NO:

29.

14. An anti-adiponectin antibody, characterized in that The antibody comprises a heavy chain and a light chain; the amino acid sequence of the heavy chain is shown in any one of SEQ ID NOs: 30 to 36; the amino acid sequence of the light chain is shown in SEQ ID NO:

37.

15. The antibody according to any one of claims 1 to 2, 4 to 6, and 8 to 14, characterized in that The antibody has a KD of ≤5.87×10 -7 M binds adiponectin with high affinity.

16. The antibody according to any one of claims 1 to 2, 4 to 6, which is selected from any one of F(ab')2, Fab', Fab, Fv and scFv.

17. An anti-adiponectin antibody conjugate, characterized in that: The invention comprises the antibody according to any one of claims 1 to 16 and a solid phase carrier or a label coupled to the antibody.

18. The antibody conjugate according to claim 17, characterized in that The label is selected from at least one of fluorescent dyes, enzymes, radioisotopes, chemiluminescent reagents, nanoparticle labels or biotin.

19. A reagent or kit, characterized in that: It comprises the antibody according to any one of claims 1 to 16 or the antibody conjugate according to any one of claims 17 to 18.

20. Use of the antibody according to any one of claims 1 to 16, the antibody conjugate according to any one of claims 17 to 18, or the reagent or kit according to claim 19 in the preparation of a product for detecting adiponectin, characterized in that: It includes: The antibody according to any one of claims 1 to 16, the antibody conjugate according to any one of claims 17 to 18, or the reagent or kit according to claim 19 is contacted with adiponectin in a sample to be detected to form an immune complex. The use according to claim 20 , wherein the immune complex further comprises a second antibody, and the second antibody binds to the antibody.

22. The use according to claim 21, wherein the immune complex further comprises a second antibody, and the second antibody binds to adiponectin.

23. An isolated nucleic acid, characterized in that The nucleic acid encodes the antibody according to any one of claims 1 to 16.

24. A carrier, characterized in that The vector contains the nucleic acid according to claim 23.

25. A cell, characterized in that The cell contains the nucleic acid of claim 23 or the vector of claim 24.

26. A method for preparing the antibody according to any one of claims 1 to 16, comprising: Cultivate the cell of claim 25.

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