Hybridoma cell strain secreting anti-reserpine monoclonal antibody and its application

Abstract

The present application relates to a hybridoma cell strain secreting anti-reserpine monoclonal antibody and its application, and belongs to the technical field of drug safety immune detection. The hybridoma cell strain secreting anti-monoclonal antibody has been preserved in the China General Microbiological Culture Collection Center on April 17, 2025, with a preservation number of CGMCC No. 46508, a preservation address of No. 3, Xibaixili, Chaoyang District, Beijing, and a classification designation of monoclonal cell strain. The reserpine monoclonal antibody cell strain obtained by the present application can be used for reserpine analysis and detection in drugs and food, has good detection sensitivity (IC 50 value of 0.579 ng / mL) and specificity (cross rate of less than 1% for related structural analogues), can realize the detection of reserpine in drugs, can assist clinical diagnosis, and has practical application value.

Description

Technical Field

[0001] This invention relates to the field of drug safety immunoassay technology, and in particular to a hybridoma cell line that secretes anti-reserpine monoclonal antibody and its application. Background Technology

[0002] Reserpine, a drug with a long history and wide application, plays a crucial role in the medical and antifungal fields. In clinical medicine, reserpine acts on the central nervous system, effectively lowering blood pressure by influencing sympathetic nerve transmission, making it a classic drug for treating hypertension. Given the various applications of reserpine, content monitoring in different scenarios is crucial. In the pharmaceutical manufacturing process, precisely controlling the content of reserpine-like drugs is key to ensuring stable efficacy and safety. Even slight deviations in content can lead to poor efficacy, ineffective treatment of patients' diseases, or even serious adverse reactions due to improper dosage.

[0003] Existing methods for reserpine content determination, besides ultra-high performance liquid chromatography-mass spectrometry (UPLC-MS / MS), gas chromatography-tandem mass spectrometry (GC-MS / MS), and high performance liquid chromatography (HPLC-UV), also include capillary electrophoresis, which suffers from time-consuming methods, requires specialized equipment and operators, and is costly, making it difficult to meet the needs of real-time, batch detection. Enzyme-linked immunosorbent assay (ELISA) technology has opened up a new path for reserpine detection. With its high sensitivity and fast detection speed, this technology can achieve trace detection of reserpine, and its operation is relatively simple, greatly improving the flexibility and timeliness of detection. It is expected to solve the problems of traditional detection methods and propel reserpine detection technology to new heights. Summary of the Invention

[0004] Therefore, the technical problem to be solved by this invention is to overcome the shortcomings of the aforementioned instruments in the prior art for the detection of reserpine. This invention provides a hybridoma cell line that secretes a monoclonal antibody against reserpine and its application. The antibody secreted by this cell line has high detection sensitivity against reserpine and can be used to establish an immunological detection method for reserpine.

[0005] This invention is achieved through the following technical solution:

[0006] The first objective of this invention is to provide a hybridoma cell line that secretes a monoclonal antibody against reserpine. This hybridoma cell line was deposited on April 17, 2025, at the China General Microbiological Culture Collection Center (CGMCC) with accession number CGMCC No. 46508. The deposit address is No. 3, Courtyard 1, Beichen West Road, Chaoyang District, Beijing, and it is classified as a monoclonal cell line.

[0007] In one embodiment of the present invention, the hybridoma cell line is obtained by immunizing mice with reserpine complete antigen.

[0008] In one embodiment of the present invention, the reserpine complete antigen is obtained by conjugating a reserpine hapten with a carrier protein; the reserpine complete antigen includes a reserpine immunogen and a reserpine-coated antigen.

[0009] In one embodiment of the present invention, the carrier protein is selected from keyhole hemocyanin and / or ovalbumin.

[0010] In one embodiment of the present invention, the structural formula of the reserpine hapten is:

[0011]

[0012] A second objective of this invention is to provide a reserpine monoclonal antibody secreted by the hybridoma cell line described above.

[0013] In one embodiment of the present invention, sterile paraffin oil was injected into the peritoneal cavity of an immunized animal, followed by a hybridoma cell line with accession number CGMCC No. 46508. Ascites fluid was collected after injection, purified, and the reserpine monoclonal antibody was obtained and stored at low temperature.

[0014] A third objective of this invention is to provide a composition for detecting reserpine, the composition comprising the hybridoma cell line and / or the reserpine monoclonal antibody.

[0015] A fourth objective of this invention is to provide a kit for detecting reserpine, the kit comprising one or more of the hybridoma cell line, the reserpine monoclonal antibody, and the composition described herein.

[0016] In one embodiment of the present invention, the kit is selected from enzyme-linked immunosorbent assay (ELISA) kits, fluorescence immunoassay kits, or chemiluminescent immunoassay kits.

[0017] A fifth objective of this invention is to provide a test strip for detecting reserpine, the test strip comprising one or more of the hybridoma cell line, the reserpine monoclonal antibody, and the composition described above.

[0018] The sixth objective of this invention is to provide the application of the hybridoma cell line, the reserpine monoclonal antibody, the composition, the kit, or the test strip in the detection of reserpine, particularly in the analysis and detection of reserpine residues in food.

[0019] The seventh objective of this invention is to provide a method for preparing the hybridoma cell line that secretes anti-reserpine monoclonal antibody, comprising the following steps:

[0020] S1. Preparation of reserpine hapten;

[0021] S2. Prepare reserpine immunogen and reserpine-coated antigen, and prepare the obtained reserpine immunogen into Freund's complete adjuvant containing antigen and Freund's incomplete adjuvant containing antigen.

[0022] S3. Perform initial immunization, booster immunization, and final immunization on immunized animals. The initial immunization uses Freund's complete adjuvant containing antigen as described in S2, the booster immunization uses Freund's incomplete adjuvant containing antigen as described in S2, and the final immunization uses reserpine immunogen as described in S2.

[0023] S4. Take spleen cells and myeloma cells from the immunized animals after the sprint immunization in S3 and perform cell fusion to obtain the hybridoma cell line of the anti-reserpine monoclonal antibody.

[0024] In one embodiment of the present invention, step S1, the preparation method of the reserpine hapten includes the following steps: dissolving 4-hydroxycyclohexane-1-carboxylic acid and bromomethylbenzene in a solvent, adding potassium carbonate and heating in a water bath to remove the organic solvent, washing, and recrystallizing to obtain colorless needle-like crystals. Subsequently, dissolving the colorless needle-like crystals with 4-dimethylaminopyridine in an organic solvent, adding 3,4,5-trimethoxybenzoyl chloride, reacting at room temperature, washing, and recrystallizing to obtain yellow needle-like crystals. Adding this yellow needle-like crystals to ethyl acetate and NaOH, heating in a water bath again, washing, and recrystallizing to obtain a white solid.

[0025] In one embodiment of the present invention, step S2, the method for preparing the reserpine immunogen and the coating antigen includes the following steps: the reserpine hapten is coupled with the carrier protein KLH and the coating antigen reserpine-OVA respectively by the carbodiimide method to prepare the immunogen reserpine-KLH and the coating antigen reserpine-OVA.

[0026] In one embodiment of the present invention, the preparation method of the reserpine immunogen and the coating antigen specifically includes the following steps: dissolving the reserpine hapten in DMF, adding N-hydroxysuccinimide and 1-ethylcarbodiimide hydrochloride, and stirring at room temperature to react, which is called solution A; weighing the carrier protein and adding it to a buffer solution, which is called solution B; slowly adding solution A dropwise to solution B while stirring, and reacting at room temperature to obtain a mixture; dialysis to obtain the reserpine immunogen and the coating antigen.

[0027] In one embodiment of the present invention, in step S2, the antigen-containing Freund's complete adjuvant is an emulsion of equal volume of Freund's complete adjuvant and reserpine immunogen.

[0028] In one embodiment of the present invention, in S2, the antigen-containing Freund's incomplete adjuvant is an emulsion of equal volume of Freund's incomplete adjuvant and reserpine immunogen.

[0029] In one embodiment of the present invention, in step S3, the entire immunization process includes one initial immunization, three to five booster immunizations, and one sprint immunization.

[0030] In one embodiment of the present invention, in step S3, the interval between the first immunization and the booster immunization is 28-31 days, the interval between booster immunizations is 20-22 days, and the interval between booster immunization and the sprint immunization is 18-21 days.

[0031] In one embodiment of the present invention, in step S3, the initial immunization and booster immunization are administered to the immunized animal via subcutaneous injection on the back; the sprint immunization is administered to the immunized animal via intraperitoneal injection.

[0032] In one embodiment of the present invention, in step S3, blood is collected from the immunized animals during the booster immunization process, and the serum titer and inhibition rate are detected by indirect competitive enzyme-linked immunosorbent assay (icELISA) to screen out immunized animals with high titer and good inhibition.

[0033] In one embodiment of the present invention, in step S3, blood is collected on days 6-8 after the completion of the booster immunization process.

[0034] In one embodiment of the present invention, in step S4, cell fusion involves culturing the fused cells in HAT medium, detecting positive cell wells using indirect ELISA, further determining the inhibitory effect of positive cell wells using indirect competitive ELISA, and subcloning the positive cell wells with the best inhibition using limiting dilution to obtain hybridoma cell lines.

[0035] In one embodiment of the present invention, the number of subcloning operations is 2-3.

[0036] In one embodiment of the present invention, in step S4, the cell fusion is performed by the polyethylene glycol (PEG 1500) method.

[0037] In one embodiment of the present invention, in step S4, the cell fusion is performed 3 days after the end of the sprint immunization.

[0038] The technical solution of the present invention has the following advantages compared with the prior art:

[0039] The monoclonal antibody secreted by the tumor cell line provided by this invention exhibits good specificity (cross-linking rate with related structural analogs less than 1%) and detection sensitivity (IC50) for reserpine. 50 The values ​​were 0.579 ng / mL, which can be used to detect the reserpine content in blood drug concentration, laying the foundation for obtaining the best therapeutic effect in specific populations and having practical application value.

[0040] Preservation of biological materials:

[0041] A hybridoma cell line, DHS, secreting a monoclonal antibody against reserpine, has been deposited at the China General Microbiological Culture Collection Center (CGMCC), located at No. 3, Courtyard 1, Beichen West Road, Chaoyang District, Beijing, Institute of Microbiology, Chinese Academy of Sciences. It has been classified as a monoclonal cell line, deposited on April 17, 2025, with accession number CGMCC No. 46508. Attached Figure Description

[0042] To make the content of this invention easier to understand, the invention will be further described in detail below with reference to specific embodiments and accompanying drawings, wherein...

[0043] Figure 1 This is the standard curve of the inhibition of reserpine by the reserpine monoclonal antibody of the present invention. Detailed Implementation

[0044] The present invention will be further described below with reference to the accompanying drawings and specific embodiments, so that those skilled in the art can better understand and implement the present invention. However, the embodiments described are not intended to limit the present invention.

[0045] Unless otherwise specified, the experimental methods used in the following examples are conventional methods, and the materials and reagents used are commercially available.

[0046] (1) The culture media involved in the following examples are as follows:

[0047] RPMI-1640 medium (mg / L): L-arginine 290, L-asparagine 50, L-aspartic acid 20, L-cysteine ​​dihydrochloride 65.15, L-glutamic acid 20, glycine 10, L-histidine 15, L-hydroxyproline 20, L-isoleucine 50, L-leucine 50, L-lysine hydrochloride 40, L-methionine 15, L-phenylalanine 15, L-proline 20, L-serine 30, L-threonine 20, L-tryptophan 5, L-tyrosine 23. 19. L-Valine 20. Para-aminobenzoic acid 1. Calcium nitrate 100. Anhydrous magnesium sulfate 48.84. Anhydrous sodium dihydrogen phosphate 676.13. Potassium chloride 400. Sodium chloride 6000. Glucose 2000. Reduced glutathione 1. Phenol red 5. L-glutamine 300. Biotin 0.2. D-calcium pantothenate 0.25. Folic acid 1. I-inositol 35. Nicotinamide 1. Choline chloride 3. Pyridoxine hydrochloride 1. Riboflavin 0.2. Thiamine hydrochloride 1. Vitamin B12 0.005. Sodium bicarbonate 2000.

[0048] (2) The reagents involved in the following examples are as follows:

[0049] Carbonate buffer (CBS): Weigh 1.59g of Na2CO3 and 2.93g of NaHCO3, dissolve them separately in a small amount of double-distilled water and mix them together. Add double-distilled water to about 800mL and mix well. Adjust the pH to 9.6 and add double-distilled water to make up to 1000mL. Store at 4℃ for later use.

[0050] Phosphate-buffered saline (PBS): 8.00g NaCl, 0.2g KCl, 0.2g KH2PO4, 2.9g Na2HPO4·12H2O, dissolved in 800mL pure water, pH adjusted to 7.2-7.4 with NaOH or HCl, and then brought to a final volume of 1000mL with ultrapure water.

[0051] Washing buffer (PBST): Add 0.5 mL of Tween-20 to 1000 mL of 0.01 mol / L pH 7.4 PBS solution;

[0052] PBST: PBS containing 0.05% Tween-20;

[0053] Antibody diluent: a washing solution containing 0.1% gelatin;

[0054] TMB colorimetric solution: Solution A: 18.43g Na2HPO4·12H2O, 9.33g citric acid, diluted to 1000mL with pure water; Solution B: 60mg TMB dissolved in 100mL ethylene glycol. Mix solutions A and B in a volume ratio of 5:1 to obtain the TMB colorimetric solution. Mix fresh before use.

[0055] (3) The detection methods involved in the following embodiments are as follows:

[0056] Reserpine inhibition rate assay: The optimal antigen and antibody concentrations for icELISA were selected using a checkerboard assay. The antigen was diluted to 1, 0.3, 0.1, and 0.03 μg / mL with carbonate buffer (CBS), and the antibody was diluted to 1, 0.3, 0.1, and 0.03 μg / mL with antibody dilution buffer. After selecting the optimal operating point, reserpine standards were diluted to concentrations of 100, 33.33, 11.11, 3.70, 1.23, 0.41, and 0.14 ng / mL, following the icELISA procedure. Finally, an inhibition curve of reserpine was obtained using Origin 2024, and the IC50 was calculated. 50 .

[0057] Example:

[0058] This embodiment provides a method for preparing a hybridoma cell line that secretes an anti-reserpine monoclonal antibody, specifically including the following steps:

[0059] (1) Preparation of hapten

[0060] 200 mg of 4-hydroxycyclohexane-1-carboxylic acid and 100 mg of bromomethylbenzene were dissolved in a solvent, and 1 mmol of potassium carbonate was added. The mixture was heated in a water bath at 50 °C for 6 h to remove the organic solvent. The solution was then washed alternately with deionized water and 1 M HCl, and recrystallized with ethanol to obtain colorless needle-like crystals. Subsequently, 250 mg of the colorless needle-like crystals were dissolved in tetrahydrofuran with 300 mg of 4-dimethylaminopyridine, and 300 mg of 3,4,5-trimethoxybenzoyl chloride was added. The mixture was reacted at room temperature for 12 h, washed three times with deionized water and 1 M HCl, and recrystallized with ethanol to obtain yellow needle-like crystals. These yellow needle-like crystals were then added to ethyl acetate and 1 M NaOH, and heated in a water bath at 60 °C for 4 h to remove the organic solvent. The solution was washed three times with deionized water and 1 M HCl, and recrystallized with ethanol to obtain a white solid. The reaction equation is as follows:

[0061]

[0062] (2) Preparation of immunogens

[0063] Weigh 8.4 mg of reserpine hapten RSP-COOH and dissolve it in 400 μL of DMF. Add 5.2 mg of N-hydroxysuccinimide under stirring at room temperature and react for 30 min. Then add 12.9 mg of 1-ethylcarbodiimide hydrochloride and react at room temperature for 4 h. The resulting mixture is called solution A. Then weigh 10 mg of keyhole hemocyanin (KLH) and dissolve it in 2 mL of carbonate buffer, which is called solution B. Slowly add solution A to solution B and react at room temperature for 12 h with stirring. Dialyze the mixture with phosphate-buffered saline (PBS) for 3 days to obtain the conjugate RSP-COOH-BSA, which is stored at -20 °C for later use.

[0064] (3) Preparation of coating antigen

[0065] Weigh 4.9 mg of the prepared RSP-COOH and dissolve it in 400 μL of DMF. Add 4.3 mg of N-hydroxysuccinimide and react for 30 min under stirring at room temperature. Then add 5.8 mg of 1-ethylcarbodiimide hydrochloride and react for 4 h. The resulting mixture is called solution A. Then weigh 10 mg of ovalbumin (OVA) and dissolve it in 2 mL of carbonate buffer, which is called solution B. Slowly add solution A to solution B and react at room temperature under stirring for 12 h. Dialyze the mixture with 0.01 mol / L phosphate-buffered saline (PBS) for 3 days to obtain the conjugate RSP-COOH-OVA. Store at -20 °C for later use.

[0066] (4) Immunization of mice: Healthy BALB / c mice aged 6-8 weeks were selected for immunization. Reserpine immunogen was mixed with an equal amount of Freund's adjuvant and emulsified, and then injected subcutaneously into the back of each BALB / c mouse. The first immunization used complete Freund's adjuvant, and subsequent immunizations used incomplete Freund's adjuvant. There was a 28-day interval between the first and second booster immunizations, and a 21-day interval between multiple booster immunizations. Blood was collected 7 days after the third immunization (5 μL of blood was collected from the mouse tail + 995 μL of antibody diluent = antiserum). The serum titer and inhibition of the mice were measured using icELISA. Mice with high titers and good inhibition were selected for a flush immunization 21 days after the fifth immunization, administered intraperitoneally. The flush immunization dose was required to be halved and contained no adjuvant.

[0067] (5) Cell fusion: Three days after the sprint immunization, cell fusion was performed using the conventional PEG (polyethylene glycol, molecular weight 1500) method. The specific steps are as follows:

[0068] a. After euthanizing the mice by enucleation and cervical dislocation-assisted carbon dioxide euthanasia, immediately disinfect the mice in 75% alcohol for about 5 minutes. Under aseptic conditions, remove the spleens of the mice, grind them moderately with the rubber tip of a syringe and pass them through a 200-mesh cell sieve to obtain a spleen cell suspension. Collect the suspension and centrifuge it (1200 rpm, 8 minutes). Wash the spleen cells three times with RPMI-1640 medium. After the last centrifugation, dilute the spleen cells to a certain volume, count them, and set them aside for later use.

[0069] b. Collection of murine myeloma SP2 / 0 cells: 7-10 days before fusion, SP2 / 0 tumor cells were cultured in RPMI-1640 medium containing 10% FBS (fetal bovine serum) in a 5% CO2 incubator. The required number of SP2 / 0 tumor cells before fusion was (1-4) × 10⁻⁶. 7 To ensure that SP2 / 0 tumor cells are in the logarithmic growth phase before fusion. During fusion, tumor cells are collected, suspended in RPMI-1640 basal culture medium, and cell counting is performed.

[0070] c. Fusion process (7 min). At min 1, add 1 mL of PEG 1500 dropwise to the cells, gradually increasing the speed. At min 2, allow to stand. At min 3 and min 4, add 1 mL of RPMI-1640 medium dropwise over 1 min. At min 5 and min 6, add 2 mL of RPMI-1640 medium dropwise over 1 min. At min 7, add 1 mL of RPMI-1640 medium dropwise every 10 seconds. Then incubate at 37°C for 5-8 min. Centrifuge (800 rpm, 8 min), discard the supernatant, and resuspend in RPMI-1640 selection medium containing 20% ​​fetal bovine serum and 2% 50×HAT. Add 200 μL / well to a 96-well cell plate and incubate at 37°C in a 5% CO2 incubator.

[0071] (6) Cell screening and cell line establishment: On day 3 of cell fusion, the fused cells were partially screened using RPMI-1640 medium, and on day 5, the medium was completely replaced with RPMI-1640 transition medium containing 20% ​​fetal bovine serum and 1% 100×HT. On day 7, the cell supernatant was collected for screening. The screening process consisted of two steps: first, positive cell wells were selected using icELISA; second, reserpine was used as a standard, and the inhibitory effect on positive cells was determined using icELISA. Cell wells that showed good inhibition of reserpine standard were selected, and subcloning was performed using the limiting dilution method. The same method was used for detection, repeated three times, to obtain the cell line.

[0072] Test example:

[0073] (1) Preparation and identification of monoclonal antibodies

[0074] BALB / c mice aged 6-8 weeks were injected intraperitoneally with 0.5 mL of sterile paraffin oil; 7 days later, each mouse was injected intraperitoneally with 1×10 6 Hybridoma cells were used, and ascites fluid was collected starting on day 7. The ascites fluid was purified using the caprylic acid-ammonium sulfate method. Under slightly acidic conditions, caprylic acid precipitates other proteins in the ascites fluid besides IgG immunoglobulins. The precipitate was then discarded by centrifugation. IgG-type monoclonal antibodies were then precipitated with an equal volume of saturated ammonium sulfate solution, centrifuged, and the supernatant was discarded. The precipitate was dissolved in 0.01M PBS solution (pH 7.4), dialyzed to desalt, and finally the purified monoclonal antibodies were stored at -20°C.

[0075] The IC50 of the monoclonal antibody against reserpine was determined using an indirect competitive ELISA method. 50 The concentration was 0.579 ng / mL, and its IC50 for analogues was verified. 50 and cross-reactivity rate, with a cross-reactivity rate of less than 1% for related analogues, where cross-reactivity rate = (IC50 of reserpine) / (IC50 of reserpine) 50 ICs of similar types 50 The cross-reactivity rate (CRR) is calculated as 100%, indicating that this antibody has high sensitivity and specificity to reserpine, as shown in Table 1.

[0076] Table 1. IC50 of monoclonal antibodies against reserpine, its functional analogues diserpine, amarine, licorice, and serlimine. 50 and cross-reactivity rate

[0077] <![CDATA[IC 50 (ng / mL)]]> Cross-reactivity rate (%) reserpine 0.579 100 Disheping >58 <1 Amarin >58 <1 Licarin >58 <1 reserpine >58 <1

[0078] (2) Antibody application

[0079] The monoclonal antibody prepared from hybridoma cell lines via in vivo ascites fluid was used in a reserpine spiking and recovery assay. The specific steps are as follows:

[0080] a. Coating: The original reserpine-OVA was serially diluted with 0.05M pH 9.6 carbonate buffer starting from 1 μg / mL, 100 μL / well, and reacted at 37℃ for 2 h;

[0081] b. Washing: Pour off the solution in the plate and wash with washing solution 3 times, 3 minutes each time;

[0082] c. Sealing: After patting dry, add 200 μL / well sealing solution and react at 37℃ for 2 h. Wash and dry for later use.

[0083] d. Sample addition: Serially dilute the antiserum (antiserum obtained by diluting mouse tail blood with antibody diluent) starting from 1:1000 and add it to each well of the coated sample at 100 μL / well. Incubate at 37°C for 30 min. After thorough washing, add 1:3000 diluted HRP-goat anti-mouse IgG at 100 μL / well and incubate at 37°C for 30 min.

[0084] e. Color development: Remove the microplate, wash it thoroughly, add 100 μL of TMB color development solution to each well, and react at 37°C in the dark for 15 min.

[0085] f. Termination and Measurement: Add 50 μL of stop solution to each well to terminate the reaction, and then measure the OD450 value of each well using a microplate reader.

[0086] The standard curve of reserpine monoclonal antibody inhibition against reserpine is as follows: Figure 1 As shown, the IC50 of the monoclonal antibody against reserpine was determined using icELISA. 50 The concentration was 0.579 ng / mL, indicating good sensitivity to reserpine and suitable for use in the immunoassay of reserpine.

[0087] Regarding recovery rates, biological samples were analyzed by liquid chromatography-tandem mass spectrometry (LC-MS / MS). Reserpine residues were not detected in either sample by LC-MS / MS. Therefore, in this implementation case, recovery was determined by adding reserpine. Specifically, as shown in Table 2, the average recovery rate of LC-MS / MS was 97.8–113.5%, with a relative standard deviation (RSD) of 2.43–6.72%; the recovery rate of the indirect competitive ELISA method was 96.5–105.5%, with an RSD of 2.35–6.45%. This indicates that the method for detecting reserpine based on reserpine monoclonal antibodies is relatively accurate and can be used for immunoassay detection of reserpine residues in plasma and urine.

[0088] Table 2 shows the recovery rates of reserpine in urine and plasma from the addition of monoclonal antibodies.

[0089]

[0090] Obviously, the above embodiments are merely illustrative examples for clear explanation and are not intended to limit the implementation. Those skilled in the art will recognize that other variations or modifications can be made based on the above description. It is neither necessary nor possible to exhaustively list all possible implementations here. However, obvious variations or modifications derived therefrom are still within the scope of protection of this invention.

Claims

1. A hybridoma cell line that secretes a monoclonal antibody against reserpine, characterized in that, The hybridoma cell line was deposited on April 17, 2025, at the China General Microbiological Culture Collection Center (CGMCC) with accession number CGMCC No. 46508. The deposit address is No. 3, No. 1 Beichen West Road, Chaoyang District, Beijing, and it is classified as a monoclonal cell line.

2. Reserpine monoclonal antibody, characterized in that, Produced by the hybridoma cell line described in claim 1.

3. A composition for detecting reserpine, characterized in that, The composition comprises the hybridoma cell line of claim 1 and / or the reserpine monoclonal antibody of claim 2.

4. A reagent kit for detecting reserpine, characterized in that, The kit comprises one or more of the hybridoma cell line of claim 1, the reserpine monoclonal antibody of claim 2, and the composition of claim 3.

5. The reagent kit according to claim 4, characterized in that, The kit is selected from enzyme-linked immunosorbent assay (ELISA) kits, fluorescence immunoassay kits, or chemiluminescence immunoassay kits.

6. A test strip for detecting reserpine, characterized in that, The test strip comprises one or more of the hybridoma cell line of claim 1, the reserpine monoclonal antibody of claim 2, and the composition of claim 3.

7. The use of the hybridoma cell line of claim 1, the reserpine monoclonal antibody of claim 2, the composition of claim 3, the kit of claim 4 or 5, or the test strip of claim 6 in the detection of reserpine.

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