Bispecific antibodies to feline SAA protein and their uses

Abstract

The present invention discloses a bispecific antibody or a functional fragment thereof of a cat SAA protein, comprising: a first functional domain and a second functional domain that specifically recognize and bind to a cat SAA protein, the amino acid sequences of VH CDR1, CDR2, CDR3, VL CDR1, CDR2 and CDR3 of the first functional domain are shown in SEQ ID NOs: 1 to 6, respectively, and the amino acid sequences of VH CDR1, CDR2, CDR3, VL CDR1, CDR2 and CDR3 of the second functional domain are shown in SEQ ID NOs: 7 to 12, respectively. The present invention also discloses the use of a bispecific antibody or a functional fragment thereof of a cat SAA protein in preparing a drug or a kit for evaluating the degree of inflammation and tissue damage and the prognosis of a cat. The present invention can rapidly and dynamically monitor the concentration of SAA in cat plasma, and has important indicative significance for early diagnosis and treatment of inflammation and tissue damage in cats.

Description

Technical Field

[0001] The present invention relates to the technical field of immunology, and more specifically, to a bispecific antibody of cat SAA protein and its use. Background Art

[0002] The acute-phase response (APR) is a systemic response triggered by local inflammation and can be seen in both acute and chronic inflammation. Serum amyloid (SAA) is the major acute-phase protein produced in large quantities during the APR. Currently, elevated SAA concentrations have become a clinically sensitive marker of inflammation. In addition to characterizing inflammatory factors, SAA may play a homeostatic role in the inflammatory process.

[0003] SAA is the precursor of tissue amyloid protein A and is an acute phase protein produced by liver cells. SAA exists in a constant amount in the serum of normal cats, but it can increase hundreds to thousands of times within 6-8 hours during the acute phase of inflammation or infection. The SAA content in the blood reaches a peak in 24-48 hours, and can quickly drop to normal levels after the pathogens are cleared. It is a sensitive indicator reflecting the body's infection status and inflammation recovery, and is a sensitive marker for diagnosing bacterial or viral infections. SAA elevation is seen in viral, mycoplasma, and bacterial infections, and its sensitivity is higher than CRP; cat CRP does not increase or only slightly increases in bacterial infections, viral and mycoplasma infections, and can provide better reference value for clinical practice.

[0004] In general, high-density lipoproteins (HDLs) are considered to be anti-atherosclerotic. In experiments studying the protein composition of HDL, it was found that the acute phase protein serum amyloid A may be incorporated into HDL particles. Studies have shown that SAA is a strong marker for cardiovascular mortality.

[0005] In veterinary medicine, SAA measurement in the blood can be used to diagnose subclinical inflammation, monitor the effectiveness of treatment in animals with infections or inflammatory conditions, and monitor patients undergoing surgery. Currently, the tools available on the market for detecting SAA concentrations mainly use enzyme-linked immunosorbent assays (ELISA) for quantitative analysis. The problem with the above monitoring methods is that they are time-consuming or the results are not very accurate. Summary of the invention

[0006] An object of the present invention is to solve at least the above problems and / or disadvantages and to provide at least the advantages which will be described hereinafter.

[0007] Another object of the present invention is to provide a bispecific antibody or a functional fragment thereof of a cat SAA protein and uses thereof.

[0008] To this end, the technical solution provided by the present invention is:

[0009] In the first aspect, a bispecific antibody or a functional fragment thereof of a cat SAA protein comprises:

[0010] A first functional domain that specifically recognizes and binds to a cat SAA protein, the first functional domain comprising an scFv1 of an anti-cat SAA bispecific antibody, the amino acid sequences of the heavy chain variable region VH CDR1, CDR2 and CDR3 of the scFv1 being shown in SEQ ID NOs: 1, 2 and 3, respectively, and the amino acid sequences of the light chain variable region VL CDR1, CDR2 and CDR3 of the scFv1 being shown in SEQ ID NOs: 4, 5 and 6, respectively; and

[0011] The second functional domain specifically recognizes and binds to the cat SAA protein, wherein the second functional domain includes the scFv2 region of the anti-cat SAA bispecific antibody, and the amino acid sequences of the heavy chain variable region VH CDR1, CDR2 and CDR3 of the scFv2 are shown in SEQID NOs: 7, 8 and 9, respectively, and the amino acid sequences of the light chain variable region VL CDR1, CDR2 and CDR3 of the scFv2 are shown in SEQID NOs: 10, 11 and 12, respectively.

[0012] Preferably, in the bispecific antibody or functional fragment thereof of the feline SAA protein, the amino acid sequence of the heavy chain variable region VH of the scFv1 is as shown in SEQ ID NO:13, and the amino acid sequence of the light chain variable region VL of the scFv1 is as shown in SEQ ID NO:14.

[0013] Preferably, in the bispecific antibody or functional fragment thereof of the feline SAA protein, the amino acid sequence of the heavy chain variable region VH of the scFv2 is as shown in SEQ ID NO:15, and the amino acid sequence of the light chain variable region VL of the scFv2 is as shown in SEQ ID NO:16.

[0014] Preferably, in the bispecific antibody of cat SAA protein or its functional fragment, the heavy chain variable region and the light chain variable region of the scFv1 are connected by a Linker, and the heavy chain variable region and the light chain variable region of the scFv2 are also connected by a Linker, and the Linker is (G4S)n.

[0015] Furthermore, in the bispecific antibody of cat SAA protein or its functional fragment, n is 3.

[0016] In the second aspect, a gene encoding the bispecific antibody or a functional fragment thereof of the above-mentioned feline SAA protein.

[0017] The third aspect includes a recombinant vector containing the above gene.

[0018] In a fourth aspect, a composition for rapid diagnosis of inflammation and tissue damage in cats comprises the above-mentioned bispecific antibody to the feline SAA protein or a functional fragment thereof.

[0019] A fifth aspect is a kit, which comprises the above-mentioned bispecific antibody to the feline SAA protein or a functional fragment thereof, and the kit is a kit for rapid in vitro detection of feline SAA by latex-enhanced immunosorbent assay or chemiluminescent immunosorbent assay.

[0020] Preferably, in the kit, the kit is used to detect SAA protein in cat plasma.

[0021] In a sixth aspect, the bispecific antibody or functional fragment thereof of the above-mentioned cat SAA protein is used in the preparation of a drug or a kit for assessing the degree of inflammation and tissue damage and the prognosis of a cat.

[0022] The present invention has at least the following beneficial effects:

[0023] The present invention first screens out two monoclonal antibodies with high affinity for cat SAA protein, then obtains a genetically recombinant cat SAA bispecific antibody through genetic engineering means, and uses it for detecting cat SAA by LEIA method. The kit of this method has higher sensitivity and wider detection range, and can be used for different clinical diagnosis needs.

[0024] The present invention can rapidly and dynamically monitor the SAA concentration in cat plasma, and has important indicative significance for the early diagnosis and treatment of inflammation and tissue damage in cats.

[0025] Other advantages, objectives and features of the present invention will be embodied in part through the following description, and in part will be understood by those skilled in the art through study and practice of the present invention. BRIEF DESCRIPTION OF THE DRAWINGS

[0026] Figure 1 Schematic diagram of the structure of the anti-cat SAA bispecific antibody in an embodiment of the present invention.

[0027] Figure 2 This is the calibration curve of the cat SAA assay kit prepared with the cat SAA bispecific antibody in the embodiment of the present invention. The cat SAA antigen is prepared into a series of calibrators with calf serum, and the concentrations are 0 mg / L, 10 mg / L, 27 mg / L, 70 mg / L, 160 mg / L, and 400 mg / L, respectively.

[0028] Figure 3 This is a sample conformity curve of the immunoturbidimetric platform of the cat SAA antigen determination kit in the embodiment of the present invention. DETAILED DESCRIPTION

[0029] The present invention is described in further detail below so that those skilled in the art can implement it according to the description.

[0030] The present invention is described below with reference to specific examples. Those skilled in the art will appreciate that the experimental methods in the following examples are conventional methods unless otherwise specified; the raw materials, reagents, materials, etc. used in the following examples are commercially available products unless otherwise specified.

[0031] The present invention relates to a bispecific antibody binding to cat SAA and its use, a cat SAA latex enhanced immunoturbidimetric detection kit for veterinary clinical diagnosis comprising the bispecific antibody, and a nucleic acid encoding the antibody or its antigen-binding fragment, a vector carrying the nucleic acid, and a method for preparing a cat SAA bispecific antibody or its antigen-binding fragment using the vector and a host cell. Rapid dynamic monitoring of cat plasma SAA concentration has important indicative significance for early diagnosis and treatment of cat inflammation and tissue damage.

[0032] The present invention provides a bispecific antibody or a functional fragment thereof of a cat SAA protein, comprising:

[0033] A first functional domain that specifically recognizes and binds to a cat SAA protein, the first functional domain comprising an scFv1 of an anti-cat SAA bispecific antibody, the amino acid sequences of the heavy chain variable region VH CDR1, CDR2 and CDR3 of the first functional domain scFv1 are shown in SEQ ID NOs: 1, 2 and 3, respectively, and the amino acid sequences of the light chain variable region VL CDR1, CDR2 and CDR3 of the first functional domain scFv1 are shown in SEQ ID NOs: 4, 5 and 6, respectively; and

[0034] The second functional domain that specifically recognizes and binds to the cat SAA protein comprises the scFv2 region of the anti-cat SAA bispecific antibody, the amino acid sequences of the heavy chain variable region VH CDR1, CDR2 and CDR3 of the second functional domain scFv2 are shown in SEQ ID NOs: 7, 8 and 9, respectively, and the amino acid sequences of the light chain variable region VL CDR1, CDR2 and CDR3 of the second functional domain scFv2 are shown in SEQ ID NOs: 10, 11 and 12, respectively.

[0035] Serial Number Amino acid sequence SEQ ID NO:1 GYTFSHTYG SEQ ID NO:2 IYPYDG SEQ ID NO:3 ARNDHYAVMDY SEQ ID NO:4 QNVGSFH SEQ ID NO:5 KVS SEQ ID NO:6 QHFSWSAPYT SEQ ID NO:7 GSAFSDNYW SEQ ID NO:8 ITSRGLYT SEQ ID NO:9 AIFYGHSSFTMDY SEQ ID NO:10 QNHIVAW SEQ ID NO:11 GAK SEQ ID NO:12 FRGSPTPYT

[0036] In one of the technical solutions of the present invention, preferably, the amino acid sequence of the heavy chain variable region VH of the first functional domain scFv1 is as shown in SEQ ID NO:13, and the amino acid sequence of the light chain variable region VL of the first functional domain scFv1 is as shown in SEQ ID NO:14.

[0037] SEQ ID NO:13

[0038] EVQLMQSGELAKPGASVDKMSCKASGYTFSHTYGIEWLRQRPIQGLEWIGCIYPYDGGTTYNPSLTSRTAITRDTSKNQFFLEMNSLTAEDTATYYCARNDHYAVMDYWGQGTSVTVSS

[0039] SEQ ID NO:14

[0040] DIVLPQSPASLAVSLGQPVTISCRASQNVGSFHLAWYQQKPGKSPKLLIYKVSESISG IPARFSGSGSGTDFTLSIDSVESEDLAAYYCQHFSWSAPYTFGGGTNLEIK

[0041] In one of the technical solutions of the present invention, preferably, the amino acid sequence of the heavy chain variable region VH of the second functional domain scFv2 is as shown in SEQ ID NO:15, and the amino acid sequence of the light chain variable region VL of the second functional domain scFv2 is as shown in SEQ ID NO:16.

[0042] SEQ ID NO:15

[0043] QVHLNQSAAELARPGASVRTSCKASGSAFSDNYWVSWIRQFPGNKLEWLGYITSRGLYTNYSQKFKGKATLTADTSSSTAYMNISSLTSEDSAVYFCAIFYGHSSFTMDYWGQGTTLTVSS

[0044] SEQ ID NO:16

[0045] DIVMAQSHKFDSTSVGDRVSITFKASQNHIVAWIHWYQQRTNGSPRLLVKGAKNLE SGIPARFSGSGSRTDFTLTISPVEADDVATYYCFRGSPTPYTFGGGTKLEIK

[0046] In one of the technical solutions of the present invention, preferably, the heavy chain variable region and the light chain variable region of the first functional domain scFv1 are connected by a Linker, and the heavy chain variable region and the light chain variable region of the second functional domain scFv2 are also connected by a Linker, and the Linker is (G4S)n.

[0047] In the above scheme, preferably, n is 3.

[0048] The present invention also provides a gene encoding the bispecific antibody or a functional fragment thereof of the above-mentioned feline SAA protein.

[0049] The present invention also provides a recombinant vector comprising the above gene.

[0050] The present invention also provides a composition for rapid diagnosis of inflammation and tissue damage in cats, which comprises the bispecific antibody or a functional fragment thereof of the above-mentioned cat SAA protein.

[0051] The present invention also provides a kit, which comprises the bispecific antibody or functional fragment of the cat SAA protein, and is a kit for rapid in vitro detection of cat SAA by latex enhanced immunoassay or chemiluminescence immunoassay.

[0052] In the above scheme, preferably, the kit is used to detect SAA protein in cat plasma.

[0053] The present invention also provides the use of the bispecific antibody of the cat SAA protein in preparing a drug or a kit for evaluating the degree of inflammation and tissue damage and the prognosis of a cat.

[0054] A bispecific antibody, or a variant thereof, or a functional fragment thereof, that binds to a feline SAA protein, comprising:

[0055] a. a first functional domain that specifically recognizes and binds to a cat SAA protein, which includes an scFv1 region of an anti-cat SAA-specific antibody; and

[0056] b. The second functional domain that specifically recognizes and binds to the feline SAA protein comprises the scFv2 region of the anti-feline SAA specific antibody.

[0057] Preferably, the functional domain that specifically recognizes and binds to cat SAA protein is composed of a single-chain antibody scFv1 and a single-chain antibody scFv2 connected in series via an Fc region and a Linker.

[0058] Preferably, the anti-feline SAA protein bispecific antibody, or its variant, or its functional fragment is a chimeric antibody, a humanized antibody, or a fully humanized antibody.

[0059] Preferably, the amino acid sequence of the heavy chain variable region of the scFv1 that specifically recognizes and binds to feline SAA protein is as shown in SEQ ID NO:13; the amino acid sequence of the light chain variable region is as shown in SEQ ID NO:14.

[0060] Preferably, the amino acid sequence of the heavy chain variable region of the scFv2 that specifically recognizes and binds to feline SAA protein is as shown in SEQ ID NO:15; the amino acid sequence of the light chain variable region is as shown in SEQ ID NO:16.

[0061] Its first functional domain scFv1 is formed by connecting SEQ ID NO: 13 and SEQ ID NO: 14 via a linker, and its second functional domain scFv2 is formed by connecting SEQ ID NO: 15 and SEQ ID NO: 16 via a linker. The linker is (G4S)n, and n is preferably 3.

[0062] The bispecific antibody, or variant thereof, or functional fragment thereof described in the present invention may be a full-length antibody, or only contain two antigen domain binding parts, such as a single-chain antibody (scFv), or a fusion protein consisting of a full-length antibody and an antigen binding part.

[0063] Under the premise of not substantially affecting the affinity of the antibody, researchers in the field can replace, add and / or delete one or more (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10) amino acids in the sequence of the present invention to obtain variants of the sequence of the antibody or its functional fragment. They are all considered to be included in the scope of protection of the present invention.

[0064] The feline SAA assay kit comprising the feline SAA bispecific antibody has a short detection time, high sensitivity, and a wide detection linearity, and can be used for different clinical diagnostic needs.

[0065] The Feline SAA Assay Kit can quickly diagnose the degree of inflammation and tissue damage in cats. Its functions are as follows:

[0066] 1) Auxiliary diagnostic indicators for bacterial and viral infections, and new indicators for efficacy monitoring;

[0067] 2) SAA is more sensitive than CRP in viral and bacterial infections, increases earlier, and decreases faster during recovery;

[0068] 3) The clinical application range is wider, including infectious diseases, cardiovascular diseases, tumors, amyloidosis and other diseases

[0069] SAA increased;

[0070] 4)SAA is more suitable for the diagnosis of infectious diseases in kittens and is a sensitive indicator for early detection of viral infections in cats.

[0071] In order to enable those skilled in the art to better understand the technical solution of the present invention, the following embodiments are provided for illustration:

[0072] Example 1 Obtaining bispecific antibodies against feline SAA

[0073] Immunization of animals

[0074] After 1 ml of 1 mg / ml cat SAA antigen was evenly mixed with an equal volume of Freund's complete adjuvant, multiple subcutaneous injections were performed into the abdomen of 12 4-6 week old BALB / C mice. One week later, 1 ml of cat SAA protein was evenly mixed with an equal volume of Freund's incomplete adjuvant and multiple subcutaneous injections were performed into the abdomen of the mice. In the third week, the above steps were repeated. In the fourth week, the above steps were repeated. Three days after the fourth injection, 20 μl of blood was collected from the tip of the tail of the mouse, allowed to stand at room temperature for 2.5 hours, centrifuged at 25°C and 11000 rpm for 15 minutes, the supernatant was taken, and the titer was tested by ELISA. If the titer test is available, whole blood is taken from the eye, allowed to stand at room temperature for 2 hours, centrifuged at 25°C and 7000 rpm for 10 minutes, the supernatant is taken, and stored at -80°C.

[0075] Hybridoma preparation

[0076] The mice were killed by dislocation and disinfected with 75% alcohol. The spleen cells of the mice were aseptically obtained in a clean bench and ground into a cell suspension. 1.5×10 8 splenocytes and 5×10 7 Myeloma (SP2 / 0) cells were fused with 50% PEG as a fusion agent, cultured in HAT selection medium for 4-10 days, and then cultured in HT medium at 5% CO2 and 37°C.

[0077] Positive clone screening and epitope identification

[0078] ELISA reaction was used to screen two clones of monoclonal hybridoma cells that could secrete antibodies that bind to feline SAA antigen. The epitope was identified using the sandwich method, and the first antibody was captured and fixed to the surface of the ELISA plate. The feline SAA antigen was then injected and bound to the first antibody. The second antibody labeled with HRP was then injected. The second antibody either binds to the feline SAA antigen, producing a "sandwich" binding, or is blocked by the first antibody and does not bind to the feline SAA antigen. The results show that the second antibody can bind to the feline SAA antigen in the presence of the first antibody, so they have different epitopes and therefore belong to different bins.

[0079] Example 2 Structural design of bispecific antibodies binding to feline SAA

[0080] The present invention provides a structural model of a bispecific antibody that can specifically bind to two different epitopes of a cat SAA protein at the same time. The structure of a main example of such an antibody is as follows: Figure 1As shown, a bispecific antibody composed of the scFv1 region of an anti-cat SAA specific antibody (VH1-Linker-VL1 or VL1-Linker-VH1) - the Fc region of mouse IgG1 - the scFv2 region of an anti-cat SAA specific antibody - Linker - (VH2-Linker-VL2 or VL2-Linker-VH2).

[0081] Variants and functional fragments of feline SAA bispecific antibodies include, but are not limited to:

[0082] ① VH1-Linker-VL1-Fc region of mIgG1-Linker-VH2-Linker-VL2;

[0083] ②VH1-Linker-VL1-Fc region of mIgG1-Linker-VL2-Linker-VH2;

[0084] ③VL1-Linker-VH1-Fc region of mIgG1-Linker-VH2-Linker-VL2;

[0085] ④VL1-Linker-VH1-Fc region of mIgG1-Linker-VL2-Linker-VH2.

[0086] Example 3 Construction of anti-cat SAA bispecific antibody expression vector

[0087] The Fc region-Linker-VH2-Linker-VL2 encoding gene of VH1-Linker-VL1-mIgG1 was connected to the plasmid pTT5 through the HindIII / EcoRI cloning site to obtain the vector pTT5-fSAA-bsAb expressing the bispecific antibody.

[0088] Example 4 Preparation of anti-cat SAA bispecific antibody

[0089] Expression of feline SAA bispecific antibody using HEK293F host cells

[0090] The cell density was diluted to 2.5 × 10 6 / ml, place the shake flask in a 5% CO2 constant temperature shaker, and start transfection after constant temperature shaking and culture at 37℃, 150rpm for 10 minutes. Prepare two 50ml sterile centrifuge tubes, add 10ml OPM and 200μg sterile plasmid DNA to one of them, and gently blow to mix; take another centrifuge tube, add 10ml OPM and 0.5ml PEI transfection reagent, and gently blow to mix; transfer all the liquid in the centrifuge tube containing the transfection reagent to the centrifuge tube containing the plasmid, and gently blow to mix; let it stand at room temperature for 10 minutes to prepare the plasmid-vector complex; remove the cells from the constant temperature shaker, add the prepared plasmid-vector complex while shaking, and put it back in the CO2 constant temperature shaker for shaking culture.

[0091] 1.2 ml of cell protein expression enhancer VPA was added 24 hours after transfection, and glucose was added to increase the expression level of the product. The product expression level was measured on the 6th day after transfection.

[0092] Antibody purification

[0093] The cell culture supernatant was centrifuged at 6000 rpm for 40 min, the supernatant was collected and filtered with a 0.22 μm filter, then purified by protein A affinity chromatography column, desalted and replaced with buffer, and stored in aliquots.

[0094] Example 5 Development of a Feline SAA Detection Kit

[0095] Calibrators for the Feline SAA Detection Kit: Prepare a series of calibrators using feline SAA antigen with calf serum, with concentrations of 0 mg / L, 10 mg / L, 27 mg / L, 70 mg / L, 160 mg / L, and 400 mg / L. The calibration curve is as follows: Figure 2 shown.

[0096] The detection principle of a cat SAA assay kit is:

[0097] The kit uses latex enhanced immunoturbidimetry. The basic principle is to cross-link the feline SAA bispecific antibody to the latex particles, and react with the feline SAA in the sample to be tested to form a large antigen-antibody complex. The turbidity increases, and the concentration of feline SAA is proportional to the turbidity formed. At a wavelength of 570nm, the feline SAA antigen content in the sample can be calculated by comparing the detected turbidity with the standard curve. The feline SAA assay kit containing the feline SAA bispecific antibody has high sensitivity and a wide detection linearity, and can be used for different clinical diagnostic needs.

[0098] A method of using a cat SAA antigen assay kit is as follows:

[0099] The cat SAA antigen kit was tested with a fully automatic biochemical analyzer 7180 with the following parameters: first add 3 μl of the sample, then add 150 μl of reagent R1, incubate at 37°C for 5 minutes, add 50 μl of reagent R2, read the absorbance A1 after 5 minutes, and calculate the absorbance difference △A=A1-A0 (starting value); use the matching calibrator for multi-point calibration, obtain the calibration curve and perform linear fitting, then the sample concentration (in mg / L) can be calculated on the calibration curve through the absorbance difference detected.

[0100] Example 6 Cat SAA Antigen Detection Kit Performance Test

[0101] One of the methods for testing the performance of a detection kit is sample compliance determination.

[0102] The sample conformity rate can also be called clinical comparative analysis, which reflects the degree of consistency between the measured value of the clinically assigned sample and the theoretical value of the clinically assigned sample by the diagnostic kit. As the core raw material of in vitro diagnostic reagents, the performance of antibodies directly affects the quality of in vitro diagnostic reagent products. As one of the most important indicators for verifying the performance of antibody raw materials, the sample conformity rate is the part that in vitro diagnostic reagent companies pay the most attention to when screening raw materials. To assess the sample conformity rate of a new antibody raw material, an in vitro diagnostic kit (colloidal gold, immunoturbidimetry and other platforms) is usually prepared with the antibody as the raw material, and the actual clinical specimens are tested using the kit, and the test results are statistically analyzed and analyzed for comprehensive assessment and evaluation. Through standardized comparative studies, analysis, and summarization of the relationship between the test results of the kit and the theoretical values ​​of clinical samples, the performance of the antibody raw material can be indirectly inferred. The closer the results of the two are, the better the consistency, and the better the performance of the antibody.

[0103] The concentration of cat SAA samples ranged from 4.23 to 518 mg / L, with a total of 25 cases. A scatter plot was drawn with the measured value as the ordinate and the theoretical value as the abscissa, and a linear regression curve was fitted using a computer, and R was calculated. 2 .like Figure 3 The sample compliance rate R of the cat SAA antigen test kit shown 2 >99%.

[0104] The number of modules and processing scales described here are used to simplify the description of the present invention. Applications, modifications and variations of the present invention are obvious to those skilled in the art.

[0105] Although the embodiments of the present invention have been disclosed as above, they are not limited to the applications listed in the specification and the implementation modes, and they can be fully applied to various fields suitable for the present invention. For those familiar with the art, additional modifications can be easily implemented. Therefore, without departing from the general concept defined by the claims and the scope of equivalents, the present invention is not limited to the specific details and the illustrations shown and described herein.

Claims

1. A bispecific antibody against a feline SAA protein, characterized in that: include: A first functional domain that specifically recognizes and binds to a cat SAA protein, the first functional domain comprising an scFv1 of an anti-cat SAA bispecific antibody, the amino acid sequences of the heavy chain variable region VH CDR1, CDR2 and CDR3 of the scFv1 being shown in SEQ ID NOs: 1, 2 and 3, respectively, and the amino acid sequences of the light chain variable region VL CDR1, CDR2 and CDR3 of the scFv1 being shown in SEQ ID NOs: 4, 5 and 6, respectively; and, A second functional domain that specifically recognizes and binds to a cat SAA protein, wherein the second functional domain comprises an scFv2 region of an anti-cat SAA bispecific antibody, wherein the amino acid sequences of the heavy chain variable region VH CDR1, CDR2 and CDR3 of the scFv2 are shown in SEQ ID NOs: 7, 8 and 9, respectively, and the amino acid sequences of the light chain variable region VL CDR1, CDR2 and CDR3 of the scFv2 are shown in SEQ ID NOs: 10, 11 and 12, respectively.

2. The bispecific antibody of cat SAA protein according to claim 1, characterized in that The amino acid sequence of the heavy chain variable region VH of the scFv1 is shown in SEQ ID NO: 13, and the amino acid sequence of the light chain variable region VL of the scFv1 is shown in SEQ ID NO:

14.

3. The bispecific antibody of cat SAA protein according to claim 1, characterized in that The amino acid sequence of the heavy chain variable region VH of the scFv2 is shown in SEQ ID NO: 15, and the amino acid sequence of the light chain variable region VL of the scFv2 is shown in SEQ ID NO:

16.

4. The bispecific antibody of cat SAA protein according to claim 1, characterized in that The heavy chain variable region and the light chain variable region of the scFv1 are connected via a Linker, and the heavy chain variable region and the light chain variable region of the scFv2 are also connected via a Linker, and the Linker is (G4S)n, where n is 3.

5. A gene encoding a bispecific antibody against the feline SAA protein according to any one of claims 1 to 4.

6. A recombinant vector comprising the gene according to claim 5.

7. A composition for rapid diagnosis of inflammation and tissue damage in cats, characterized in that: It comprises the bispecific antibody against the feline SAA protein according to any one of claims 1 to 4.

8. A kit, characterized in that The kit comprises the bispecific antibody of the feline SAA protein as claimed in any one of claims 1 to 4, and the kit is a kit for rapid in vitro detection of feline SAA by latex enhanced immunoassay or chemiluminescence immunoassay.

9. The kit according to claim 8, characterized in that The kit is used for detecting SAA protein in cat plasma.

10. Use of the bispecific antibody of the feline SAA protein according to any one of claims 1 to 4 in the preparation of a drug or a kit for assessing the degree of inflammation and tissue damage and the prognosis of a feline.

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