A monoclonal antibody and its application

Abstract

The present invention provides a monoclonal antibody and its use. The monoclonal antibody provided by the present invention has excellent specificity and can target monomeric C-reactive protein (mCRP) without generating an antigen-antibody reaction with pentameric C-reactive protein (pCRP). Therefore, it can be used as an antibody for detecting mCRP. Furthermore, due to this specific targeting effect, the monoclonal antibody of the present invention can bind to mCRP and thereby inhibit the pro-inflammatory effects of mCRP, thus showing potential for targeted therapy.

Description

Technical Field

[0001] The present invention relates to biotechnology, in particular to a monoclonal antibody and application thereof. Background Art

[0002] C-reactive protein (CRP) is a protein (acute protein) that rises sharply in plasma when the body is infected or tissue is damaged. It can activate complement and enhance phagocytosis by phagocytes, thereby playing a conditioning role, capable of clearing invading pathogens and damaged, necrotic, and apoptotic tissue cells. Circulating CRP exists as pentameric CRP (pCRP), which dissociates into monomeric CRP (mCRP) under inflammatory conditions. Current research suggests that the two natural isomers, pCRP and mCRP, have opposite activities in inflammatory responses, with pCRP having weak anti-inflammatory biological activity and mCRP having strong pro-inflammatory biological activity. Therefore, there is an urgent need to obtain monoclonal antibodies targeting mCRP for detection or targeted treatment of mCRP. Summary of the Invention

[0003] The present invention provides a monoclonal antibody, an immunodetection agent and an immunodetection kit, which can target monomeric C-reactive protein (mCRP) without generating antigen-antibody reaction with pentameric C-reactive protein (pCRP) and have excellent specificity.

[0004] The present invention provides a use of the aforementioned monoclonal antibody, immunodetection agent, and immunodetection kit for detecting levels of monomeric C-reactive protein (mCRP). The monoclonal antibody, immunodetection agent, and immunodetection kit of the present invention have excellent specificity and can therefore be used as a detection antibody to detect mCRP levels, achieving good detection results.

[0005] The present invention provides a monoclonal antibody and a pharmaceutical composition, which can specifically bind to monomeric C-reactive protein (mCRP), thereby inhibiting the pro-inflammatory biological activity of mCRP and have the potential for targeted therapy.

[0006] The present invention provides the use of the aforementioned monoclonal antibody and pharmaceutical composition in the preparation of a medicament for treating autoimmune diseases induced by monomeric C-reactive protein (mCRP). The monoclonal antibody and pharmaceutical composition of the present invention can inhibit the pro-inflammatory effects of mCRP through targeted binding to mCRP, thereby having the potential to treat autoimmune diseases.

[0007] The present invention also provides a method for preparing the above-mentioned monoclonal antibody, which can quickly and simply prepare high-purity monoclonal antibodies through hybridoma technology, and can also be used for mass production of monoclonal antibodies.

[0008] The present invention provides a monoclonal antibody that specifically binds to monomeric C-reactive protein (mCRP), comprising the following sequences: HCDR1 as shown in SEQ ID NO: 1, HCDR2 as shown in SEQ ID NO: 2, HCDR3 as shown in SEQ ID NO: 3, LCDR1 as shown in SEQ ID NO: 4, LCDR2 as shown in SEQ ID NO: 5, and LCDR3 as shown in SEQ ID NO: 6.

[0009] The monoclonal antibody as described above, wherein the monoclonal antibody comprises a heavy chain amino acid sequence as shown in SEQ ID NO: 7 and a light chain amino acid sequence as shown in SEQ ID NO: 8.

[0010] The present invention provides an immune detection agent, comprising the above-mentioned monoclonal antibody.

[0011] The present invention provides an immunoassay kit comprising the above-mentioned monoclonal antibody or the above-mentioned immunoassay agent.

[0012] The immunoassay kit as described above, wherein the immunoassay kit further comprises at least one of a standard, a blocking solution, a washing solution, a detection antibody, a chemiluminescent substrate, and a stop solution.

[0013] The present invention provides a use of the above-mentioned monoclonal antibody, the above-mentioned immunodetection agent or the above-mentioned immunodetection kit in detecting the level of monomeric C-reactive protein (mCRP).

[0014] The present invention provides a pharmaceutical composition comprising the above-mentioned monoclonal antibody.

[0015] The present invention provides a use of the above-mentioned monoclonal antibody or the above-mentioned pharmaceutical composition in the preparation of a drug for treating autoimmune diseases induced by monomeric C-reactive protein (mCRP).

[0016] The use as described above, wherein the autoimmune disease includes at least one of lupus nephritis and rheumatoid arthritis.

[0017] The present invention also provides a method for preparing a monoclonal antibody, comprising the following steps:

[0018] Mice were immunized with monomeric C-reactive protein (mCRP) to obtain immune mice;

[0019] The spleen cells of the immunized mice were fused with myeloma cells to obtain hybridoma cells;

[0020] The secretory products of the hybridoma cells are separated and purified to obtain monoclonal antibodies.

[0021] The present invention provides a monoclonal antibody that specifically binds to monomeric C-reactive protein (mCRP). This antibody specifically targets monomeric C-reactive protein (mCRP) without cross-reacting with pentameric C-reactive protein (pCRP). It exhibits excellent specificity and can be used as a detection antibody. Furthermore, the monoclonal antibody of the present invention specifically binds to monomeric C-reactive protein (mCRP) and can inhibit the pro-inflammatory effects of mCRP, thus offering potential as a targeted treatment for autoimmune diseases. BRIEF DESCRIPTION OF THE DRAWINGS

[0022] Figure 1 This is a graph showing the specificity of the monoclonal antibody in Example 2;

[0023] Figure 2 This is a fitting curve obtained when the monoclonal antibody in Example 2 is used as the detection antibody at a concentration of 5 μg / ml;

[0024] Figure 3 This is a fitting curve diagram obtained when the monoclonal antibody in Example 2 is used as the detection antibody at a concentration of 1 μg / ml. DETAILED DESCRIPTION

[0025] In order to enable those skilled in the art to better understand the scheme of the present invention, the present invention is further described in detail below. The specific embodiments listed below are only for describing the principles and features of the present invention. The examples are only used to explain the present invention and are not intended to limit the scope of the present invention. Based on the embodiments of the present invention, all other embodiments obtained by those of ordinary skill in the art without making creative efforts are within the scope of protection of the present invention.

[0026] In a first aspect, the present invention provides a monoclonal antibody that specifically binds to monomeric C-reactive protein (mCRP), comprising the following sequence: HCDR1 as shown in SEQ ID NO: 1, HCDR2 as shown in SEQ ID NO: 2, HCDR3 as shown in SEQ ID NO: 3, LCDR1 as shown in SEQ ID NO: 4, LCDR2 as shown in SEQ ID NO: 5, and LCDR3 as shown in SEQ ID NO: 6.

[0027] The amino acid sequences of the complementarity determining regions (CDRs) of the monoclonal antibodies of the present invention are shown in Table 1:

[0028] Table 1

[0029] HCDR1 SYWMQ (SEQ ID NO: 1) HCDR2 AIYPGDGDTRYTQKFKG(SEQ ID NO:2) HCDR3 WTSSHYFDY (SEQ ID NO: 3) LCDR1 RSGQSLVHSNGDTYLH (SEQ ID NO: 4) LCDR2 KVSNRFS (SEQ ID NO: 5) LCDR3 SQNTHVPPT (SEQ ID NO: 6)

[0030] Monoclonal antibodies can be divided into constant and variable regions. Within the variable region, a small portion of amino acid residues varies significantly, making the composition and order of these amino acid residues more susceptible to variation. Because this region can form precise complementarity with the antigenic determinant in terms of spatial structure, it is called the complementarity determining region (CDR). The heavy chain (H chain) and light chain (L chain) each have three complementarity determining regions in their variable regions: HCDR1, HCDR2, and HCDR3 for the heavy chain, and LCDR1, LCDR2, and LCDR3 for the light chain.

[0031] It will be appreciated that HCDR1 comprises the amino acid sequence as shown in SEQ ID NO: 1, or an amino acid sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity to SEQ ID NO: 1, or an amino acid sequence in which one or more amino acid residues are substituted and / or deleted and / or added compared to SEQ ID NO: 1.

[0032] HCDR2 comprises the amino acid sequence as shown in SEQ ID NO: 2, or an amino acid sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity to SEQ ID NO: 2, or an amino acid sequence which has one or several amino acid residues substituted and / or deleted and / or added compared to SEQ ID NO: 2.

[0033] HCDR3 comprising the amino acid sequence as shown in SEQ ID NO: 3, or an amino acid sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity to SEQ ID NO: 3, or an amino acid sequence having one or several amino acid residues substituted and / or deleted and / or added compared to SEQ ID NO: 3.

[0034] LCDR1, which comprises the amino acid sequence as shown in SEQ ID NO:4, or an amino acid sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity with SEQ ID NO:4, or an amino acid sequence which has one or more amino acid residues substituted and / or deleted and / or added compared to SEQ ID NO:4.

[0035] LCDR2 comprises the amino acid sequence as shown in SEQ ID NO:5, or an amino acid sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity to SEQ ID NO:5, or an amino acid sequence which has one or more amino acid residues substituted and / or deleted and / or added compared to SEQ ID NO:5.

[0036] LCDR3 comprising the amino acid sequence as shown in SEQ ID NO:6, or an amino acid sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity to SEQ ID NO:6, or an amino acid sequence having one or more amino acid residues substituted and / or deleted and / or added compared to SEQ ID NO:6.

[0037] The monoclonal antibodies provided by the present invention have excellent specificity and can target monomeric C-reactive protein without producing an antigen-antibody reaction with pentameric C-reactive protein. Monomeric C-reactive protein (mCRP) is a dissociated form of C-reactive protein (CRP). Normally, CRP exists in the form of a cyclic pentameric globular protein composed of 5 identical subunits, also known as pentameric CRP (pCRP); under inflammatory conditions, pCRP dissociates into mCRP and exerts a pro-inflammatory effect. The monoclonal antibodies of the present invention can accurately target the 199-206 amino acid epitope of mCRP, i.e., mCRP199-206. mCRP199-206 has a relatively strong pro-inflammatory biological activity and is an ideal target for treating inflammation-related diseases. It is understandable that the monoclonal antibodies of the present invention have the ability to target mCRP199-206, i.e., have the ability to target mCRP.

[0038] Therefore, the monoclonal antibody of the present invention specifically binds to mCRP but not to pCRP, has excellent specificity, and can be used as a detection antibody for mCRP. At the same time, due to this specific targeting effect, the monoclonal antibody of the present invention can bind to mCRP and thereby inhibit the pro-inflammatory effect of mCRP, thus having the potential for targeted therapy.

[0039] In a specific embodiment, the monoclonal antibody comprises a heavy chain amino acid sequence as shown in SEQ ID NO:7 and a light chain amino acid sequence as shown in SEQ ID NO:8.

[0040] It is understood that, based on the knowledge of the above-mentioned amino acid sequence of the monoclonal antibody of the present invention, those skilled in the art can obtain a polynucleotide encoding the monoclonal antibody, insert the polynucleotide into an expression vector to obtain a recombinant expression vector, then transform the recombinant expression vector into a host cell, and then isolate and purify the expression product of the host cell to obtain the monoclonal antibody of the present invention.

[0041] Therefore, the present invention provides a polynucleotide encoding the above-mentioned monoclonal antibody; the present invention provides a recombinant expression vector comprising the above-mentioned polynucleotide; the present invention also provides a host cell comprising the above-mentioned recombinant expression vector, which is a host cell for expressing exogenous proteins, such as the 293 cell line.

[0042] The second aspect of the present invention provides an immunodetection agent comprising the above-mentioned monoclonal antibody.

[0043] Since the immunodetection agent provided by the present invention includes the above-mentioned monoclonal antibody, it can specifically bind to mCRP, thereby detecting the level of mCRP.

[0044] The third aspect of the present invention provides an immunoassay kit, comprising the above-mentioned monoclonal antibody or the above-mentioned immunoassay agent.

[0045] Since the immunoassay kit provided by the present invention includes the above-mentioned monoclonal antibody or the above-mentioned immunoassay agent, it can specifically target mCRP to detect the level of mCRP.

[0046] Furthermore, the immunoassay kit further comprises at least one of a standard, a blocking solution, a washing solution, a detection antibody, a chemiluminescent substrate, and a stop solution.

[0047] Among them, the standard is a sample containing a target protein of known concentration, and a standard curve can be obtained by the standard. For example, in the present invention, the standard can be a sample containing a known concentration of mCRP; the blocking solution is a reagent used to block the blank positions on the surface of the solid phase carrier to avoid nonspecific binding, for example, it can be a bovine serum albumin solution, a skim milk powder solution, a casein solution, etc.; the washing solution is a reagent used to remove unbound reagents to reduce background, for example, it can be Tris buffered saline (TBS), Tris buffered saline with Tween 20 (TBST), phosphate buffered saline (PBS), etc. Buffered Saline (PBS), etc.; the detection antibody is a secondary antibody that carries a label and can specifically bind to the monoclonal antibody of the present invention, and the label can be an enzyme, such as horseradish peroxidase (HRP), or a fluorescent group; the chemiluminescent substrate is a reagent that can react with the label to emit light, for example, it can be the chemiluminescent substrate 3,3',5,5'-tetramethylbenzidine (TMB) of HRP, which can be oxidized by hydrogen peroxide (H2O2) under the catalysis of HRP to generate a blue product with maximum absorption values ​​at 370nm and 620-650nm; the stop solution is a reagent that can terminate the reaction between the chemiluminescent substrate and the label, for example, it can be 2M sulfuric acid, which can terminate the reaction between TMB and H2O2, causing the blue product to turn yellow with a maximum absorption value at 450nm.

[0048] A fourth aspect of the present invention provides a use of the above-mentioned monoclonal antibody, the above-mentioned immunodetection agent or the above-mentioned immunodetection kit in detecting the level of monomeric C-reactive protein (mCRP).

[0049] The monoclonal antibody, immunodetection agent and immunodetection kit of the present invention have excellent specificity, can be used as detection antibodies to detect the level of mCRP, and have good detection effects.

[0050] In one embodiment of the present invention, the monoclonal antibody of the present invention is used as a detection antibody, and when its working concentration is 5 μg / ml, the R-squared of the fitting curve is 0.97, and the fitting effect is good; when its working concentration is 1 μg / ml, the R-squared of the fitting curve is 0.97, and the fitting effect is good.

[0051] It is understood that the present invention is not limited to the method of detecting monomeric C-reactive protein (mCRP) using the monoclonal antibody, immunodetection agent, or immunodetection kit of the present invention. For example, the level of mCRP can be detected by enzyme-linked immunosorbent assay (ELISA), enzyme-linked immunospot assay (ELISPOT), micro-sample multi-index flow cytometry (CBA), liquid chip technology, etc.

[0052] In one embodiment of the present invention, the monoclonal antibody of the present invention is used to detect monomeric C-reactive protein (mCRP) using ELISA technology. Specifically, the ELISA plate is first coated with mCRP protein, washed and blocked, and then washed again. The monoclonal antibody of the present invention is added to the ELISA plate for incubation. After washing, the secondary antibody is added for incubation, washed again, and TMB substrate solution is added for color development. Finally, hydrochloric acid solution is added to terminate the reaction and the absorbance is read at 450 nm. The level of mCRP is obtained based on the absorbance and the standard curve.

[0053] Furthermore, detecting the level of mCRP refers to detecting the presence or absence of mCRP, or detecting the level of mCRP.

[0054] The fifth aspect of the present invention provides a pharmaceutical composition comprising the above-mentioned monoclonal antibody.

[0055] The monoclonal antibodies provided in the first aspect of the present invention are capable of specifically binding to monomeric C-reactive protein (mCRP), thereby inhibiting the pro-inflammatory biological activity of mCRP and possessing potential for targeted therapy. Therefore, by including the monoclonal antibodies provided in the first aspect of the present invention, the pharmaceutical compositions provided in the present invention also possess potential for targeted therapy. Furthermore, the monoclonal antibodies of the present invention can be formulated into freeze-dried or liquid pharmaceutical compositions.

[0056] To further enhance the immune effect of the pharmaceutical composition, the pharmaceutical composition may also include a pharmaceutically acceptable carrier, such as a stabilizer, preservative, isotonic agent, and the like. Specifically, when the pharmaceutical composition is a freeze-dried preparation, the pharmaceutically acceptable carrier may be mannitol, lactose, sucrose, human albumin, or the like; when the pharmaceutical composition is a liquid preparation, the pharmaceutically acceptable carrier may be physiological saline, water for injection, phosphate buffer, aluminum hydroxide, or the like.

[0057] A sixth aspect of the present invention provides a use of the above-mentioned monoclonal antibody or the above-mentioned pharmaceutical composition in the preparation of a drug for treating autoimmune diseases induced by monomeric C-reactive protein (mCRP).

[0058] Studies have shown that monoclonal antibodies and pharmaceutical compositions can inhibit the pro-inflammatory effects of mCRP through targeted binding to mCRP, thereby having the potential to treat autoimmune diseases.

[0059] Furthermore, the autoimmune disease includes at least one of lupus nephritis and rheumatoid arthritis.

[0060] Lupus nephritis refers to systemic lupus erythematosus (SLE) combined with immune damage of varying renal pathology, accompanied by significant clinical manifestations of renal impairment. Its pathogenesis is associated with immune abnormalities such as immune complex formation, immune cells, and cytokines. Rheumatoid arthritis is an autoimmune disease with synovitis as its pathological basis, which can ultimately lead to joint deformities. Its etiology may also be closely related to autoimmunity, genetics, and microbial infection. Studies have shown that antibodies targeting mCRP can rescue mouse models of lupus nephritis or rheumatoid arthritis.

[0061] A seventh aspect of the present invention provides a method for preparing the above-mentioned monoclonal antibody, comprising the following steps:

[0062] Mice were immunized with monomeric C-reactive protein (mCRP) to obtain immune mice;

[0063] The spleen cells of the immunized mice were fused with myeloma cells to obtain hybridoma cells;

[0064] The secretory products of the hybridoma cells are separated and purified to obtain monoclonal antibodies.

[0065] The present invention first immunizes mice with monomeric C-reactive protein (mCRP). Splenocytes from these immunized mice are then fused with myeloma cells and screened to produce hybridoma cells. Finally, the secretory products of the hybridoma cells are isolated and purified to produce monoclonal antibodies. This invention utilizes hybridoma technology to rapidly and easily produce high-purity monoclonal antibodies, and can also be used for mass production of monoclonal antibodies.

[0066] It is understood that, considering the instability of hybridoma cells, antibody variable region sequences can be obtained by PCR to preserve the desired hybridoma variable region sequences, and antibody production can be carried out using other host cells that highly express antibodies. The present invention is not limited to the method for obtaining antibody light and heavy chain variable region sequences. For example, the method can be: extracting total RNA from hybridoma cells, using this as a template to synthesize first-strand cDNA, and then performing subsequent PCR amplification using the first-strand cDNA as a template to obtain nucleic acids of the antibody light and heavy chain variable regions corresponding to the hybridoma cells, performing agarose gel electrophoresis, excising the gel to recover TA clones, and performing Sanger sequencing to obtain the antibody variable region sequences.

[0067] The technical solutions of this application will be further explained below with reference to specific examples. Experimental methods in the following examples, where specific conditions are not specified, were generally performed under conventional conditions or as recommended by the manufacturer. All reagents used, unless otherwise specified, were commercially available or publicly available.

[0068] Example 1: Obtaining monoclonal antibodies

[0069] Three 6-8 week old BALB / c mice were immunized five times. The primary immunization used a keyhole limpet hemocyanin (KLH)-mCRP199-206 peptide-coupled immunogen at a dose of 70 μg per mouse. The initial immunization was performed as follows: 70 μg of the immunogen was added to 500 μL with sterile water. The mixture was then mixed with 600 μL of complete Freund's adjuvant (CFA). The mixture was sonicated using an ultrasonic cell disruptor to obtain a milky mixture, which was used for the primary immunization of the mice.

[0070] Four booster immunizations were administered on days 3, 7, 9, and 11 after the primary immunization. The booster immunizations used the same immunogen as the primary immunization, with a dose of 140 μg per mouse. The procedure was as follows: 140 μg of the immunogen was taken, sterile water was added to 1 mL, and the mixture was then mixed with 1 mL of AD11.15 adjuvant. The resulting mixture was used to boost the mice. AD11.15 adjuvant is manufactured by ABMAX Biotechnologies, Inc., with the product number AD11.15.

[0071] On the 14th day after the initial immunization, tail blood was collected from mice. On the 17th day, splenic lymphocytes were isolated from the immunized mice and fused with SP2 / 0 myeloma cells using polyethylene glycol (PEG). On the 12th day after fusion, hybridoma cells were harvested and cultured in 96-well plates. Seven days later, the supernatant from the 96-well plates was diluted at a 1:1 volume ratio using phosphate buffered saline containing 5% (w / v) skim milk. The supernatant from the 96-well plates was then evaluated by indirect ELISA to screen for monoclonal cell lines that secreted and recognized the antigen mCRP199-206. Finally, the infinite dilution method was used for monoclonal antibody screening to obtain the monoclonal antibody PUF002. The monoclonal antibody PUF002 includes HCDR1 as shown in SEQ ID NO:1, HCDR2 as shown in SEQ ID NO:2, HCDR3 as shown in SEQ ID NO:3, LCDR1 as shown in SEQ ID NO:4, LCDR2 as shown in SEQ ID NO:5, and LCDR3 as shown in SEQ ID NO:6, its heavy chain amino acid sequence is shown in SEQ ID NO:7, and its light chain amino acid sequence is shown in SEQ ID NO:8.

[0072] Example 2: Verification of the detection effect of monoclonal antibodies

[0073] The specificity of the monoclonal antibody PUF002 was verified by enzyme-linked immunosorbent assay (ELISA). PUF002 was used to coat the ELISA plate at 0.1 μg / ml, washed, and blocked. After washing again, different concentrations of mCRP199-206 protein or pCRP protein were added to the ELISA plate for incubation. The plate was then washed and incubated with horseradish peroxidase (HRP)-labeled goat anti-mCRP antibody. The plate was washed again and 3,3',5,5'-tetramethylbenzidine (TMB) substrate solution was added for color development. Finally, hydrochloric acid solution was added to terminate the reaction and the absorbance was read at 450 nm. The results are shown in Figure 2. Figure 1 shown.

[0074] The detection effect of monoclonal antibody PUF002 as a detection antibody was verified by ELISA. The ELISA plate was coated with different concentrations of mCRP199-206 protein, washed and blocked, and then washed again. 5μg / ml or 1μg / ml of PUF002 was added to the ELISA plate for incubation. The plate was washed and incubated with HRP-labeled anti-mouse IgG antibody. The plate was washed again and 3,3',5,5'-tetramethylbenzidine (TMB) substrate solution was added for color development. Finally, hydrochloric acid solution was added to terminate the reaction and the absorbance was read at 450nm. The results are shown as follows: Figure 2 or 3, where Figure 2 The concentration of PUF002 was 5μg / ml, Figure 3 The concentration of PUF002 was 1 μg / ml.

[0075] like Figure 1 As shown, the monoclonal antibody PUF002 of the present invention can specifically recognize mCRP199-206 but not pCRP, and has excellent specificity.

[0076] like Figure 2 As shown, the monoclonal antibody PUF002 of the present invention was used as a detection antibody. When its working concentration was 5 μg / ml, the R-squared of the fitting curve was 0.97, and the fitting effect was good, indicating that the monoclonal antibody PUF002 can be used as a detection antibody to detect the level of mCRP199-206 with good detection effect and has the potential for targeted therapy.

[0077] like Figure 3 As shown, the monoclonal antibody PUF002 of the present invention was used as a detection antibody. When its working concentration was 1 μg / ml, the R-square of the fitting curve was 0.97, and the fitting effect was good, indicating that the monoclonal antibody PUF002 can be used as a detection antibody to detect the level of mCRP199-206 with good detection effect and has the potential for targeted therapy.

[0078] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, rather than to limit it. Although the present invention has been described in detail with reference to the above embodiments, those skilled in the art should understand that they can still modify the technical solutions described in the above embodiments, or replace some or all of the technical features therein with equivalents. However, these modifications or replacements do not cause the essence of the corresponding technical solutions to deviate from the scope of the technical solutions of the embodiments of the present invention.

Claims

1. A monoclonal antibody, characterized in that The monoclonal antibody comprises the following sequences: HCDR1 as shown in SEQ ID NO: 1, HCDR2 as shown in SEQ ID NO: 2, HCDR3 as shown in SEQ ID NO: 3, LCDR1 as shown in SEQ ID NO: 4, LCDR2 as shown in SEQ ID NO: 5, and LCDR3 as shown in SEQ ID NO:

6.

2. The monoclonal antibody according to claim 1, characterized in that The monoclonal antibody comprises a heavy chain variable region amino acid sequence as shown in SEQ ID NO: 7 and a light chain variable region amino acid sequence as shown in SEQ ID NO:

8.

3. An immunodetection agent, characterized in that The monoclonal antibody comprises the monoclonal antibody according to any one of claims 1 to 2.

4. An immunoassay kit, characterized in that: The method comprises the monoclonal antibody according to any one of claims 1 to 2 or the immunodetection agent according to claim 3.

5. The immunoassay kit according to claim 4, characterized in that The immunoassay kit further comprises at least one of a standard, a blocking solution, a washing solution, a detection antibody, a chemiluminescent substrate, and a stop solution.

6. Use of the monoclonal antibody according to any one of claims 1 to 2, the immunodetection agent according to claim 3, or the immunodetection kit according to any one of claims 4 to 5 in detecting monomeric C-reactive protein (mCRP) levels for non-diagnostic purposes.

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