An anti-human MBP monoclonal antibody and its application

Abstract

This invention discloses an anti-human MBP monoclonal antibody and its applications. The complementarity-determining region (CDR) of the heavy chain variable region of the monoclonal antibody has the amino acid sequences shown in SEQ ID NO. 1-3, and the CDR of the light chain variable region has the amino acid sequences shown in SEQ ID NO. 4-6. This invention uses a chemically synthesized polypeptide as an immunogen to immunize mice. Through cell fusion, screening, and subcloning, a monoclonal cell line that efficiently secretes anti-human MBP monoclonal antibodies is obtained. The antibody obtained by this invention has high specificity and high binding capacity, effectively targeting and binding MBP protein from serum. It is suitable for detecting MBP protein in blood, thereby evaluating neurological injury diseases that may cause changes in MBP protein concentration, and providing more convenient conditions for the diagnosis of mild traumatic brain injury.

Description

Technical Field

[0001] This invention belongs to the field of antibody preparation technology, specifically relating to an anti-human MBP monoclonal antibody and its application. Background Technology

[0002] Myelin basic protein (MBP) is a strongly basic membrane protein synthesized by oligodendrocytes in the central nervous system and Schwann cells in the peripheral nervous system. With a molecular weight of approximately 18-21 kDa, it is located on the long arm of chromosome 18, region 23 (18q23), and contains 7 exons and 6 introns. As a major component of the white matter myelin sheath in the central nervous system, changes in MBP levels can reflect the degree of damage to the nervous system. Therefore, MBP is a diagnostic indicator of substantial damage to the central nervous system, particularly myelin loss. When the central nervous system is diseased (e.g., by infection, inflammation, tumors, trauma, hemorrhage, edema), changes in myelin basic protein (MBP) in the cerebrospinal fluid can be measured as a basis for clinical diagnosis, disease treatment, and prognosis. In traumatic brain injury (TBI), due to axonal damage or myelin sheath destruction, MBP is released into the cerebrospinal fluid (CSF) or enters the bloodstream through a breached blood-brain barrier (BBB), leading to elevated blood MBP levels.

[0003] Traumatic brain injury (TBI) refers to the brain damage resulting from external trauma, leading to functional disorders. It is a leading cause of disability and death worldwide and a common public health problem. Globally, approximately 69 million people are affected by TBI annually, resulting in global economic losses of up to $400 billion. According to domestic and international research, TBI is believed to cause cognitive and attentional changes, increase the risk of stroke by 50%, and also increase the risk of diseases such as Parkinson's disease and Alzheimer's disease. Numerous clinical studies have shown that myxobin (MBP) can serve as an important biomolecular marker for assessing the severity of traumatic brain injury and prognostic evaluation. Elevated MBP concentrations can predict CT / MRI imaging abnormalities, particularly lesions related to white matter (such as diffuse axonal injury). Furthermore, serum MBP concentrations can remain elevated for several hours to several days after TBI, typically peaking within 24-48 days, providing a sufficient detection window for its use as a biomarker.

[0004] Monoclonal antibodies can specifically recognize and bind to their corresponding antigens. High affinity and strong specificity are essential for achieving highly sensitive detection. There is a need in this field for an antibody against human MBP that can be used to detect MBP levels in serum and analyze related neurological diseases, particularly for the effective diagnosis of mTBI, in order to further confirm the diagnosis and treatment. Summary of the Invention

[0005] The purpose of this invention is to provide an anti-human MBP monoclonal antibody and its application.

[0006] An anti-human MBP monoclonal antibody comprises a heavy chain and a light chain, wherein the heavy chain includes a heavy chain variable region, and the light chain includes a light chain variable region; the heavy chain variable region includes HCDR1, HCDR2, and HCDR3, and the light chain variable region includes LCDR1, LCDR2, and LCDR3; the amino acid sequence of HCDR1 is shown in SEQ ID NO.1; the amino acid sequence of HCDR2 is shown in SEQ ID NO.2; the amino acid sequence of HCDR3 is shown in SEQ ID NO.3; the amino acid sequence of LCDR1 is shown in SEQ ID NO.4; the amino acid sequence of LCDR2 is shown in SEQ ID NO.5; and the amino acid sequence of LCDR3 is shown in SEQ ID NO.6.

[0007] The heavy chain further includes the framework regions HFR1, HFR2, HFR3, and HFR4 of the heavy chain variable region; the light chain further includes the framework regions LFR1, LFR2, LFR3, and LFR4 of the light chain variable region; the amino acid sequence of HFR1 is as shown in SEQ ID NO.7, or has more than 80% homology with SEQ ID NO.7; the amino acid sequence of HFR2 is as shown in SEQ ID NO.8, or has more than 80% homology with SEQ ID NO.8;

[0008] The amino acid sequence of HFR3 is as shown in SEQ ID NO. 9, or has more than 80% homology with SEQ ID NO. 9; the amino acid sequence of HFR4 is as shown in SEQ ID NO. 10, or has more than 80% homology with SEQ ID NO. 10; the amino acid sequence of LFR1 is as shown in SEQ ID NO. 11, or has more than 80% homology with SEQ ID NO. 11; the amino acid sequence of LFR2 is as shown in SEQ ID NO. 12, or has more than 80% homology with SEQ ID NO. 12; the amino acid sequence of LFR3 is as shown in SEQ ID NO. 13, or has more than 80% homology with SEQ ID NO. 13; the amino acid sequence of LFR4 is as shown in SEQ ID NO. 13, or has more than 80% homology with SEQ ID NO. 13.

[0009] The amino acid sequence of its heavy chain variable region is shown in SEQ ID NO.15, or has more than 80% sequence similarity with SEQ ID NO.15; the amino acid sequence of its light chain variable region is shown in SEQ ID NO.16, or has more than 80% sequence similarity with SEQ ID NO.16.

[0010] The anti-human MBP monoclonal antibody is a murine-derived antibody.

[0011] The nucleotide sequence expressing the anti-human MBP monoclonal antibody or functional fragment is shown in SEQ ID NO. 17 of the sequence listing, and the nucleotide sequence corresponding to SEQ ID NO. 16 of the light chain is shown in SEQ ID NO. 18 of the sequence listing.

[0012] Cell lines capable of expressing the aforementioned anti-human MBP monoclonal antibody.

[0013] Application of the anti-human MBP monoclonal antibody in the preparation of immunoassay-related detection reagents.

[0014] The preparation of immunoassay-related detection reagents includes the preparation of ELISA kits, immunoassay colloidal gold test strips, or dry immunofluorescence lateral flow chromatography test strips.

[0015] The beneficial effects of this invention are as follows: The anti-human MBP monoclonal antibody prepared by this invention has a high affinity for human MBP and can target and bind to MBP protein in blood for quantitative detection. This invention also provides a cell expressing the anti-human MBP antibody, thereby enabling large-scale preparation and providing more convenient conditions for MBP detection. Attached Figure Description

[0016] Figure 1 This is a high-performance liquid chromatography (HPLC) of peptides.

[0017] Figure 2 This is a mass spectrum of a polypeptide.

[0018] Figure 3 This is for antibody specificity assay. Detailed Implementation

[0019] To facilitate understanding of the present invention, a more comprehensive description will be given below. However, the present invention can be implemented in many different forms and is not limited to the embodiments described herein. Rather, these embodiments are provided to provide a thorough and complete understanding of the disclosure of the present invention.

[0020] Example 1: Peptide selection and synthesis of anti-MBP antibodies

[0021] In this embodiment, 18kD MBP was used for antigen epitope analysis, and the target peptide was selected by using three different tools and methods.

[0022] (1) B cell epitope prediction (BepiPred-3.0): Based on deep learning technology of protein language model, predict possible B cell epitopes in MBP protein.

[0023] Key steps: Input the MBP protein sequence. This method extracts deep features from the protein sequence using a model, trains the model to predict possible B cell epitopes, and generates prediction results. The top 20% of high-scoring regions are selected, with a focus on high-confidence epitopes. Different versions of prediction results are distinguished (e.g., "linepitope top 20%)", and cross-validation is used to improve reliability.

[0024] BepiPred-3.0 Analysis Results:

[0025] MASQKRPSQRHGSKYLATA STMDHARHGFLPRHRD TGILDSIGRFFGG DRGAPKRGSGKDSHHP A RTA HYGSLPQKSHGRTQD ENPVVHFFKNIVTP RTPPPSQGKG RGLSLSRFSWGA EGQRPGFGYGGRASDYKSAHKGFK GVDAQGTLSKIFKLGG RDSRSGSPMARR.

[0026] (2) Linear epitope analysis (Bepitope): Predicts the location of linear epitopes (continuous epitopes) in conjunction with the diverse structures of amino acid side chain backbones.

[0027] Key steps: By comparing different algorithms (such as Emini, Pellequer, Karplus, etc.), different scores were provided for each predicted epitope. These epitope predictions included continuous segments of the protein sequence and their corresponding amino acid positions. For example, MASQKRPSQRHGSK (Emini accessibility), LPRHRDTGILDSIGRF (Pellequer translocation), etc., and by combining the results of these different methods, more accurate epitope predictions were obtained.

[0028] Bepitope software analysis:

[0029] MASQKRPSQRHGSK YLATASTMDHARHGFL PRHRDTGILDSIGRFF GGDRGAPK RGSGKDSHHPARTA HY GSLPQ KSHGRTQDENPVVHFFKNIVTPRTPPSQGKG RGLSLSRF SWGAEGQRPGFGYG GRASDYK SAHKGFKGV DAQ GTLSKIFKLGGRDSRSGSPMARR.

[0030] (3) Structural and solvent accessibility analysis (Scratch): Predict protein secondary structure (helix, turn, coil) and solvent accessibility.

[0031] Key steps: Utilize neural network models to predict the structural states of protein segments, such as helices, turns, and coils, including relative solvent accessibility at different exposure levels (e.g., predictions at 25% and 30% exposure thresholds) and predictions of multilayer structures (e.g., "C" for coil and "H" for helix).

[0032] Scratch Structure Analysis:

[0033] MASQKRPSQRHGSK YLATASTMDHARHGFLPRHRDTGILDSIGRFFGG DRGAPKRGSGKDSHHPARTA HYGSLPQKSHGRTQD ENPVVHFFKNIV TPRTPPPSQGKGRG LSLSRFSWGAEG QRPGFGYGGR ASDYKSAHKG FKG VDAQGT LSKIF KLGGRDSRSGSPMARR.

[0034] (4) Comprehensive analysis and selection of target peptides

[0035] We will cross-align the high-binding fragments obtained from machine learning models and different computational methods, and retain the fragments supported by all three tools to finally determine the peptide we want to select.

[0036] Final choice: C-DRGAPKRGSGKDSHHPART: High resolution region predicted by multiple tools, with good solvent accessibility.

[0037] 1. Peptide synthesis and detection

[0038] The synthesis and testing of the peptides were handled by Suzhou Bio-Long Technology Co., Ltd.

[0039] C-DRGAPKRGSGKDSHHPART. For example... Figure 1 As shown, peak 4 (8.845 min) is the most prominent peak, accounting for the largest proportion of the sample (concentration of 95.38%). The concentrations of other peaks are much lower, with peaks 5 and 3 being the next most significant, but still far lower than peak 4. This indicates that the sample contains the major component represented by peak 4, as well as small amounts of impurities or other components represented by the other peaks.

[0040] like Figure 2As shown, each peak in the spectrum corresponds to these different ionization states. The mass-to-charge ratio (m / z) of each charged substance will be the molecular weight divided by the charge, resulting in a molecular weight of 2133.31 Da for the sample.

[0041] 2. KLH Coupling

[0042] Small molecule peptides were used as antigens to prepare antibodies through animal immunization. 10 mg of the KLH-conjugated peptide was used for immunization.

[0043] Example 2 Animal Immunization Program

[0044] Experimental animals: Five 7-week-old female BaLB / c mice were selected for immunization; the antigen and adjuvant were mixed at a dosage of 100 μL per dose, with any shortfall made up with physiological saline; the primary immunization was administered on day 0 via injection into the gastrocnemius muscle of the left hind limb, 100 μL / mouse. Booster immunizations were administered every 14 days.

[0045] Example 3: Determination of serum titer in animals

[0046] On day 35, orbital blood was collected from mice and serum was separated. Serum titer was determined by ELISA, and the best-immunized mice were selected for cell fusion.

[0047] The ELISA procedure is as follows:

[0048] (1) Coating antigen: Dilute the antigen (peptide or MBP protein) with coating solution, and coat at 2 mg / mL (peptide), 1 mg / mL (protein), 100 mL / well, overnight at 4℃; wash 3 times with washing solution, 300 mL / well, and pat dry.

[0049] (2) Blocking: 200 mL / well of blocking solution, incubate at 37℃ for 2 h; wash 3 times with washing solution, 300 mL / well, and pat dry.

[0050] (3) Add primary antibody blocking solution to mouse serum at a ratio of 1:1000, then dilute twice and add 100 mL / well. Incubate at 37°C for 1 h. Wash 3 times with washing solution, 300 mL / well, and pat dry.

[0051] (4) Dilute the secondary antibody with antibody diluent at a ratio of 1:10000, 100 mL / well, and incubate at 37 °C for 1 h; wash 3 times with washing buffer, 300 mL / well, and pat dry.

[0052] (5) Color development: 100 mL / well color development solution, incubate at room temperature for 10 min.

[0053] Table 1. Serum titer determination in mice

[0054] serum dilution BDS2134-#1 BDS2134-#2 BDS2134-#3 BDS2134-#4 BDS2134-#5 200 0.8629 3.1327 3.0183 3.487 0.1681 2000 0.126 2.3265 2.4456 1.1531 0.0249 20000 0.0302 0.3488 0.7702 0.1413 0.0246 0 0.0152 0.0108 0.0372 0.0421 0.0141

[0055] Intraperitoneal shock: On day 42, BDS2134-#3 mice were finally immunized by intraperitoneal injection (PBS + antigen 100 mL / mouse) to induce the maximum immune response.

[0056] Example 4: Preparation of anti-human MBP monoclonal antibody

[0057] 1. Cell fusion

[0058] Mouse spleens were fused with Sp2 / 0 cells using the PEG method; the fused cells were then screened and cultured in a semi-solid medium containing HAT.

[0059] (1) Gently pipette the healthy Sp2 / 0 cells off the culture flask and place them into a 50ml centrifuge tube.

[0060] (2) Blood was collected from the eyes of the mice, and they were then killed by pulling their necks. The mice were then soaked in 75% alcohol for 5 minutes.

[0061] (3) Pour a small amount of serum-free IMDM into a petri dish, place the cell sieve and syringe core into the petri dish, remove the mouse spleen with scissors and tweezers, and place it on the cell sieve. Gently crush the spleen thoroughly with the syringe core, aspirate the ground cells into a centrifuge tube containing Sp2 / O, and centrifuge at 1500 rpm for 10 min.

[0062] (4) Remove the thymus from the mouse with scissors and tweezers and crush it. Put the crushed cells into a 15ml centrifuge tube, add 2ml HAT and 1ml HT, and put it in an incubator for later use.

[0063] (5) Discard the supernatant after centrifuging the spleen cells. Carefully and gently mix the centrifuged cells with Sp2 / 0 cells using serum-free IMDM, with a Sp2 / 0:spleen cell ratio of 1:3 to 1:10; centrifuge at 1500 rpm for 10 min.

[0064] (6) Discard as much of the cell supernatant as possible after centrifugation. Tap the centrifuge tube to mix the cells thoroughly, place the centrifuge tube in 37°C warm water, and slowly add 1 ml of PEG over 1 minute. After adding, let it stand in warm water for 1 minute. Then slowly add 8 ml of serum-free IMDM over 2 minutes, and centrifuge at 1000 rpm for 10 minutes.

[0065] (7) Discard the supernatant, add 10 ml of serum, carefully blow the cells evenly, then add 5 ml of Hybridoma Feeder, the previously prepared thymocyte mixture (2 ml thymus + 2 ml HAT + 1 ml HT), and 25 ml of semi-solid culture medium, and mix thoroughly. Then pour evenly into 30 cell culture dishes. Place the cell culture dishes in a humidified box and incubate in an incubator for 11-14 days. When the clones are visible to the naked eye, use a stereomicroscope to pick out the clones.

[0066] 2. Selecting clones

[0067] Plate the cells using 20% ​​newborn calf serum + HT + 10% Hybridoma Feeder, 150 μL / well, and pick 1200 clones from BDS2134#3.

[0068] 3. Monoclonal cell screening

[0069] Using "BDS2134#3" plates, the selected monoclonal antibodies were initially screened using an indirect ELISA method. Antigen was coated at a concentration of 1 μg / mL (100 μL / well) onto 96-well microplates. PBS-T-Casein was used as a blank control, Sp2 / 0 cell supernatant as a negative control, and mouse positive serum diluted 1:1000 in PBS-T-Casein as a positive control. Forty-six positive hybridoma cell lines were obtained and subsequently expanded into larger cultures.

[0070] Twelve positive hybridoma subclones were selected and coated with "BDS2134#3 polypeptide". A second ELISA screening was performed on the selected single clones (12 BDS2134#3 clones in total). The culture was then expanded to a larger culture size for further screening: 2 μg / ml antigen was coated, 100 μL of cell supernatant was added, and the rest of the steps were the same as above. The final subclonal cell line, BDS2134#3-1, was selected.

[0071] Table 2 Monoclonal cell screening

[0072] Cell line number OD 1 4.4067 3 4.3848 4 2.4944 6 2.4427 10 2.3952 11 2.9606 12 2.6906 13 2.3953 17 2.636 18 2.4096 19 3.1852 20 2.2623 null 0.0099 Positive 3.2507

[0073] Example 5 Monoclonal Antibody Analysis

[0074] 1. Evaluation of monoclonal antibody specificity

[0075] The purpose of this study in preparing MBP antibodies was to detect the MBP protein content in serum. Therefore, human serum, rat serum, and mouse serum were used to evaluate the specificity of the prepared antibodies. MBP protein was added to human serum as a positive control, and serum-free coated wells were used as blank controls. The prepared antibody BDS2134#3-1 was used as the primary antibody, and the antibody specificity was evaluated using ELISA.

[0076] result( Figure 3 The results show that the antibody of the present invention has a low background in the serum of different species, but has a high affinity only for positive samples, indicating that it has good specificity.

[0077] 2. Monoclonal antibody titer determination

[0078] The titer of the prepared BDS2134#3-1 antibody was detected using ELISA, and compared with that of a commercially available antibody. The antigen was coated onto a 96-well ELISA plate at a concentration of 1 μg / mL, 100 μL / well (100 ng of antigen per well), with a blank negative control included. Both the purified antibody and the commercially available antibody were diluted to 1 mg / mL and then diluted with PBS at a 1:10 ratio. 3 1:10 4 1:10 5 1:10 6 1:10 7 Serial dilutions were performed, with 100 μL / well as the primary antibody. The remaining ELISA procedures were the same as for mouse serum titer determination. The OD value was measured using a microplate reader. 450 value.

[0079] The results (Table 3) show that the monoclonal antibody prepared in Example 4 has a titer comparable to that of commercially available antibodies.

[0080] Table 3 Antibody titer determination

[0081] Dilution factor BDS2134#3-1 antibody OD450 Commercial antibody OD450 <![CDATA[1:10 3 ]]> 2.3749 3.0268 <![CDATA[1:10 4 ]]> 2.0488 2.3479 <![CDATA[1:10 5 ]]> 1.0938 1.5569 <![CDATA[1:10 6 ]]> 0.2083 0.3481 <![CDATA[1:10 7 ]]> 0.0842 0.1155 Negative 0.0569 0.0611

[0082] 3. Identification of monoclonal cell line subtypes

[0083] The BDS2134#3-1 antibody was identified as an IgG1 antibody using a monoclonal antibody subtype identification kit, coated with antigen in a 96-well ELISA plate. The procedure was performed according to the kit instructions. The results showed that the antibody subtype was IgG1.

[0084] Example 6 Sequencing of anti-human MBP antibody

[0085] To identify the MBP-134-3 antibody sequence, total RNA was reverse transcribed into cDNA using antisense primers according to the Hiscript III reverse transcriptase technique manual. Then, VH and VL antibody fragments were amplified according to Biotyscience's standard operating procedure (SOP). The PCR fragments were cloned into the TA / Blunt-Zero cloning vector. Colony PCR was performed to screen clones, and at least three positive clones were sequenced.

[0086] The sequencing results are as follows:

[0087] Heavy chain variable region nucleotide sequence:

[0088] CAGGTGCAGCTGAAGCAGTCAGGACCTGGCCTAGTGCAGCCCTCACAGAGCCTGTCCATCACCTGCACAGTCTCTGGTTTCTCATTAACTACCTATGGTGTACTCTGGGTTCGCCAGTCTCCAGGAAAGGGTCTGGAGTGGCTGGGAGTGATATGGAGTGATGGAA ACACAGACTATAATGCAGCTTTCATATCCAGACTGAGCATCAGCAAGGACAATTCCAAGAGCCAAGTTTTCTTTAAAATGAACAGTCTTCAAGGTGATGACACAGCCGTATATTACTGTGTCAGAACTGGTAGCCACGATGGGGACTACTGGGGTCAAGGAACCTC TGTCACCGTCTCCTCA (SEQ ID NO. 17).

[0089] Heavy chain variable region amino acid sequence:

[0090] QVQLKQSGPGLVQPSQSLSITCTVSGFSLT TYGVL WVRQSPGKGLEWLG VIWSDGNTDYNAAFIS RLSISKDNSKSQVFFKMNSLQGDDTAVYYCVR TGSHDGDY WGQGTSVTVSS (SEQ ID NO.15), the underlined region is the complementarity determination region (CDR);

[0091] in:

[0092] HCDR1:TYGVL (SEQ ID NO.1);

[0093] HCDR2: VIWSDGNTDYNAAFIS (SEQ ID NO.2);

[0094] HCDR3:TGSHDGDY (SEQ ID NO.3);

[0095] HFR1: QVQLKQSGPGLVQPSQSLSITCTVSGFSLT (SEQ ID NO.7);

[0096] HFR2: WVRQSPGKGLEWLG (SEQ ID NO.8);

[0097] HFR3: RLSISKDNSKSQVFFKMNSLQGDDTAVYYCVR (SEQ ID NO.9);

[0098] HFR4: WGQGTSVTVSS (SEQ ID NO.10);

[0099] Light chain variable region nucleotide sequence:

[0100] GATATCCAGATGACACAGACTACATCCTCCCTGTCTGCCTCTCTGGGAGACAGAGTCACCATCAGTTGCAGGACAAGTCAGGATATTAGTAATTATTTAAACTGGTATCAGCAGAAACCGGATGGAACTGTTAAACTCCTGATCTATTATACATCAAGATTA CACTCAGGAGTCCCATCAAGGTTCAGTGGCAGTGGGTCTGGGACAGATTATTCTCTCACCATCAGCAACCTGGAACCTGAAGATATTGCCACTTACTATTGTCAGCAGTATAGTAAGCTTCCTCCCACGTTCGGTGCTGGGACCAAGCTGGAGCTGAAA (SEQ ID NO.18);

[0101] Light chain variable region amino acid sequence:

[0102] DIQMTQTTSSLSASLGDRVTISC RTSQDISNYLN WYQQKPDGTVKLLIY YTSRLHS GVPSRFSGSGSGTDYSLTISNLEPEDIATYYC QQYSKLPPT FGAGTKLELK (SEQ ID NO.16); the underlined area indicates the complementarity determination region (CDR).

[0103] in:

[0104] LCDR1:RTSQDISNYLN(SEQ ID NO.4);

[0105] LCDR2: YTSRLHS (SEQ ID NO.5);

[0106] LCDR3: QQYSKLPPT (SEQ ID NO.6);

[0107] LFR1: DIQMTQTTSSLSASLGDRVTISC (SEQ ID NO.11);

[0108] LFR2: WYQQKPDGTVKLLIY (SEQ ID NO.12);

[0109] LFR3: GVPSRFSGSGSGTDYSLTISNLEPEDIATYYC (SEQ ID NO.13)

[0110] LFR4: FGAGTKLELK (SEQ ID NO. 14).

[0111] The embodiments described above are merely illustrative of several implementations of the present invention, and while the descriptions are relatively specific and detailed, they should not be construed as limiting the scope of the invention patent. It should be noted that those skilled in the art can make various modifications and improvements without departing from the concept of the present invention, and these all fall within the protection scope of the present invention. Therefore, the protection scope of this invention patent should be determined by the appended claims.

Claims

1. A monoclonal antibody against human MBP, characterized in that, The device comprises a heavy chain and a light chain, wherein the heavy chain includes a heavy chain variable region, and the light chain includes a light chain variable region; the heavy chain variable region includes HCDR1, HCDR2, and HCDR3, and the light chain variable region includes LCDR1, LCDR2, and LCDR3; the amino acid sequence of HCDR1 is shown in SEQ ID NO.1 of the sequence listing; the amino acid sequence of HCDR2 is shown in SEQ ID NO.2 of the sequence listing; the amino acid sequence of HCDR3 is shown in SEQ ID NO.3 of the sequence listing; the amino acid sequence of LCDR1 is shown in SEQ ID NO.4 of the sequence listing; the amino acid sequence of LCDR2 is shown in SEQ ID NO.5 of the sequence listing; and the amino acid sequence of LCDR3 is shown in SEQ ID NO.6 of the sequence listing.

2. The anti-human MBP monoclonal antibody according to claim 1, characterized in that, The heavy chain further includes the framework regions HFR1, HFR2, HFR3, and HFR4 of the heavy chain variable region; the light chain further includes the framework regions LFR1, LFR2, LFR3, and LFR4 of the light chain variable region; the amino acid sequence of HFR1 is as shown in SEQ ID NO.7, or has more than 80% homology with SEQ ID NO.7; the amino acid sequence of HFR2 is as shown in SEQ ID NO.8, or has more than 80% homology with SEQ ID NO.8; The amino acid sequence of HFR3 is as shown in SEQ ID NO. 9, or has more than 80% homology with SEQ ID NO. 9; the amino acid sequence of HFR4 is as shown in SEQ ID NO. 10, or has more than 80% homology with SEQ ID NO. 10; the amino acid sequence of LFR1 is as shown in SEQ ID NO. 11, or has more than 80% homology with SEQ ID NO. 11; the amino acid sequence of LFR2 is as shown in SEQ ID NO. 12, or has more than 80% homology with SEQ ID NO. 12; the amino acid sequence of LFR3 is as shown in SEQ ID NO. 13, or has more than 80% homology with SEQ ID NO. 13; the amino acid sequence of LFR4 is as shown in SEQ ID NO. 13, or has more than 80% homology with SEQ ID NO.

13.

3. The anti-human MBP monoclonal antibody according to claim 1, characterized in that, The amino acid sequence of its heavy chain variable region is shown in SEQ ID NO.15, or has more than 80% sequence similarity with SEQ ID NO.15; the amino acid sequence of its light chain variable region is shown in SEQ ID NO.16, or has more than 80% sequence similarity with SEQ ID NO.

16.

4. The anti-human MBP monoclonal antibody according to claim 1, characterized in that, The anti-human MBP monoclonal antibody is a murine-derived antibody.

5. A nucleic acid molecule expressing the anti-human MBP monoclonal antibody according to any one of claims 1-4, wherein the nucleic acid molecule corresponding to the heavy chain SEQ ID NO.15 is shown in the sequence listing SEQ ID NO.17, and the nucleic acid molecule corresponding to the light chain SEQ ID NO.16 is shown in the sequence listing SEQ ID NO.

18.

6. A cell line capable of expressing the anti-human MBP monoclonal antibody of claim 1.

7. The use of the anti-human MBP monoclonal antibody as described in claim 1 in the preparation of immunoassay-related detection reagents.

8. The application of the anti-human MBP monoclonal antibody according to claim 7 in the preparation of immunoassay-related detection reagents, characterized in that, The preparation of immunoassay-related detection reagents includes the preparation of ELISA kits, immunoassay colloidal gold test strips, or dry immunofluorescence lateral flow chromatography test strips.

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