A detection method for botulinum neurotoxin type B and its application

Abstract

The present invention discloses a detection method and application for type B botulinum neurotoxin. The method of the present invention has a short detection time and is highly sensitive as a qualitative detection method, reaching the picogram level, and has obvious advantages in detecting trace toxin samples. Compared with the animal method using antiserum detection, the use of monoclonal antibodies has high detection specificity, which is conducive to accurate decision-making whether it is used for scientific research detection and food hygiene inspection, and has a small error compared to the animal method. In addition, the characteristic peak of the botulinum neurotoxin double antibody sandwich reaction produced by the present invention can be used for the stability detection of botulinum neurotoxin, and plays a role that cannot be replaced by other methods in the production of botulinum toxin products, quality control, new product development, quality control and monitoring of calibration standards, and market supervision. Therefore, the present invention can be used to guide the establishment of toxin immunoassay standard substances and promote capillary immunoassay technology to become a new standard method for botulinum toxin detection.

Description

Technical Field

[0001] The invention belongs to the field of immunological qualitative detection of trace proteins, and particularly relates to a detection method for type B botulinum neurotoxin and its application. Background Art

[0002] Botulinum toxin is a bacterial exotoxin produced during the growth and reproduction of Clostridium botulinum. It is divided into seven types according to antigenicity, namely A, B, C, D, E, F, and G. Botulinum toxin is highly toxic. Food contaminated with botulinum toxin can cause food poisoning, which can be fatal in severe cases. The types that cause human botulism poisoning are mainly A, B, E, and F. Therefore, botulinum toxin testing is an important part of food hygiene testing. Research on the prevention and control of botulinum toxin is of great significance to national biosafety and social health security. Immunological testing is the main method for botulinum toxin testing.

[0003] The key to clinical rescue of botulism poisoning is to make a correct diagnosis as early as possible, which is related to whether lives can be saved. The diagnosis of symptoms is only a reference. The decisive diagnosis is the qualitative detection of poisoned food. The qualitative detection of poisoned food is the key to diagnosis. The identification of the type of toxin in poisoned food is the key to making a treatment plan. In the field of laboratory research on toxins, in addition to qualitative and identification detection of samples, purity, stability and other aspects are also required.

[0004] The existing conventional technology is firstly the experimental animal method based on immune neutralization of toxins. The defects of this method are: 1) the detection time is long, usually 2-4 days; 2) the use of experimental animals faces pressure in experimental ethics and is bound to be eliminated; 3) there are individual differences in animals, resulting in large experimental errors; 4) the diagnostic serum used for type identification has low sensitivity, and the results are judged by the death and symptoms of mice. The mice need to be continuously observed within 2-4 days. If continuous observation is not possible, the observation of mouse symptoms may be missed, and there are also mice that die for other reasons (non-specific death), so the identification effect of toxins is not good. The second commonly used detection technology is the ELISA sandwich method, but the ELISA method has the following defects: the sensitivity can only reach the ng level, and it cannot detect toxins at the pg level. Botulinum toxin is extremely toxic, and a very small amount of toxin can cause poisoning, and the amount of toxin that produces biological effects is less than the nanogram level. The ELISA sandwich method has a low detection rate for low-dose toxins. Secondly, the ELISA method can only detect the overall molecular structure of the toxin and make a macroscopic judgment. It cannot specifically analyze the structure of each part of the toxin molecule, and cannot detect important properties such as the purity and stability of the toxin, and cannot fully meet the needs of botulinum toxin research.

[0005] Although there are other analytical methods for botulinum toxin, they are only used in highly theoretical fields such as structural analysis of toxins and are not routine methods for clinical and laboratory testing. Summary of the invention

[0006] In order to solve at least some of the technical problems in the above-mentioned prior art, the present invention has developed a capillary immunoelectrophoresis detection technology based on the sandwich method, which can achieve pg-level sensitivity. Specifically, the present invention includes the following contents.

[0007] In a first aspect of the present invention, a method for detecting botulinum neurotoxin type B is provided, which comprises forming a sandwich complex with a capture antibody, a test sample and a detection antibody in a capillary and catalyzing a detection liquid to produce a signal that can be captured by a detector, thereby determining the presence of botulinum neurotoxin type B and / or quantifying botulinum neurotoxin type B, wherein the capture antibody is capable of targeting botulinum neurotoxin type B, and the capture antibody comprises heavy chain CDR1-3 having amino acid sequences as shown in GFAFSSYD, ISSGGSST and ARQAFYTYDGTAMDY, respectively, and / or light chain CDR1-3 having amino acid sequences as shown in QSIVHNNGDTY, KVS and FQGSHIPWT, respectively.

[0008] In certain embodiments, the method for detecting botulinum neurotoxin type B according to the present invention comprises the following steps:

[0009] (1) adding the capture antibody, the sample to be tested, the detection antibody and the detection solution to the detection plate of the capillary immunoelectrophoresis device respectively;

[0010] (2) allowing the sample to be tested to bind to the capture antibody in the capillary assembly and then further bind to the detection antibody to form a complex, and pass through the capillary in different zones;

[0011] (3) allowing the detection antibody to catalyze the detection solution and determining the presence and / or quantification of botulinum neurotoxin type B based on the fluorescent signal.

[0012] In certain embodiments, according to the method for detecting botulinum neurotoxin type B of the present invention, the detection antibody has any one of the amino acid sequences shown in (I)-(III):

[0013] (I) comprising the heavy chain amino acid sequence shown in SEQ ID NO.1 and / or the light chain amino acid sequence shown in SEQ ID NO.2;

[0014] (II) an amino acid sequence that has at least 90% homology to the amino acid sequence shown in (I) and has the same function;

[0015] (III) an amino acid sequence obtained by modifying, replacing, deleting or adding one or more amino acids to the amino acid sequence shown in (I) or (II) and having the same function.

[0016] In certain embodiments, according to the method for detecting botulinum neurotoxin type B of the present invention, the detection antibody is an enzyme-labeled antibody, and the enzyme includes horseradish peroxidase, alkaline phosphatase or β-galactosidase.

[0017] In certain embodiments, according to the method for detecting botulinum neurotoxin type B of the present invention, the detection solution includes a chemiluminescent agent or a color developer, the detection solution corresponding to the horseradish peroxidase includes at least one of luminol, o-phenylenediamine, tetramethylbenzidine, 5-aminosalicylic acid and 2,2-azino-bis(3-ethyl-benzothiazole-6-sulfonic acid) diammonium salt, the detection solution corresponding to the alkaline phosphatase includes at least one of AMPPD, acridinium ester and 4-methylumbelliferyl phosphate, and the detection solution corresponding to the β-galactosidase includes at least one of 5-bromo-4-chloro-3-indole-β-D-galactoside, p-nitrophenyl phosphate and 4-methylumbelliferyl-RD galactoside.

[0018] In certain embodiments, according to the method for detecting botulinum neurotoxin type B of the present invention, the concentration of the capture antibody and the detection antibody is 0.1-50 mg / mL.

[0019] The second aspect of the present invention provides a capillary immunoelectrophoresis detection kit for botulinum neurotoxin type B, comprising:

[0020] I. A capture antibody solution, wherein the capture antibody is capable of targeting botulinum neurotoxin type B, and the capture antibody comprises heavy chain CDR1-3 having amino acid sequences as shown in GFAFSSYD, ISSGGSST, and ARQAFYTYDGTAMDY, respectively, and / or light chain CDR1-3 having amino acid sequences as shown in QSIVHNNGDTY, KVS, and FQGSHIPWT, respectively;

[0021] II. Detection antibody solution;

[0022] III. Test fluid.

[0023] In certain embodiments, according to the capillary immunoelectrophoresis detection kit for botulinum neurotoxin type B of the present invention, the detection solution is a mixed solution of equal volumes of luminol and hydrogen peroxide.

[0024] In certain embodiments, according to the capillary immunoelectrophoresis detection kit for botulinum neurotoxin type B according to the present invention, the detection kit further comprises 5×Master Mix and Sample Buffer.

[0025] The third aspect of the present invention provides an application of the method according to the present invention or the kit, wherein the application includes the production of botulinum neurotoxin type B or the quality control and monitoring of botulinum neurotoxin type B standards.

[0026] The method of the present invention has a short detection time and can detect results within 6 hours. At the same time, as a qualitative detection method, it has a high sensitivity, reaching the picogram level, which is 1-2 orders of magnitude higher than other methods, and has obvious advantages in detecting trace toxin samples. Compared with the animal method using antiserum detection, the use of monoclonal antibodies has high detection specificity, which is conducive to accurate decision-making whether it is used for scientific research detection or food hygiene inspection, and has a small error compared to the animal method. In addition, the characteristic peak of the botulinum neurotoxin double antibody sandwich reaction produced by the present invention can be used for the stability detection of botulinum toxin and botulinum neurotoxin, and plays a role that cannot be replaced by other methods in the production of botulinum toxin products, quality control, new product development, quality control and monitoring of calibration standards, and market supervision. Therefore, the present invention can be used to guide the establishment of toxin immunoassay standard substances and promote capillary immunoassay technology to become a new standard method for botulinum toxin detection. BRIEF DESCRIPTION OF THE DRAWINGS

[0027] Figure 1-5 The characteristic spectrum of the capillary immunoelectrophoresis sandwich reaction in Example 1 is shown.

[0028] Figure 6-11 The characteristic spectrum of the capillary immunoelectrophoresis sandwich reaction in Example 2 is shown.

[0029] Fig.12 The results of qualitative detection of botulinum neurotoxin type B by ELISA sandwich method used in comparative examples are shown. DETAILED DESCRIPTION

[0030] Various exemplary embodiments of the present invention will now be described in detail. This detailed description should not be considered as limiting the present invention, but should be understood as a more detailed description of certain aspects, features, and embodiments of the present invention.

[0031] It should be understood that the terms described in the present invention are only for describing special embodiments and are not intended to limit the present invention. In addition, for the numerical range in the present invention, it should be understood that the upper and lower limits of the scope and each intermediate value therebetween are specifically disclosed. Each smaller range between the intermediate value in any stated value or stated range and any other stated value or intermediate value in the described range is also included in the present invention. The upper and lower limits of these smaller ranges can be independently included or excluded in the scope.

[0032] Unless otherwise indicated, all technical and scientific terms used herein have the same meanings as commonly understood by those skilled in the art to which the invention belongs. Although the present invention describes only preferred methods and materials, any methods and materials similar or equivalent to those described herein may also be used in the implementation or testing of the present invention. All documents mentioned in this specification are incorporated by reference to disclose and describe the methods and / or materials related to the documents. In the event of a conflict with any incorporated document, the content of this specification shall prevail.

[0033] Method for detecting botulinum neurotoxin type B

[0034] Conventional capillary immunoelectrophoresis detection technology uses an indirect method for immunological reaction, and there is no precedent for sandwich detection. The indirect method is not suitable for detecting botulinum toxin antigens, so there is currently no capillary immunoelectrophoresis sandwich method for detecting botulinum toxin. The present invention has developed a capillary immunoelectrophoresis detection technology based on the sandwich method, and surprisingly found that the method can achieve pg-level sensitivity. Therefore, in one aspect of the present invention, a method for detecting botulinum neurotoxin type B is provided, comprising forming a sandwich complex with a capture antibody, a test sample and a detection antibody in a capillary and catalyzing a detection liquid to produce a signal that can be captured by a detector, thereby determining the presence of botulinum neurotoxin type B and / or quantifying botulinum neurotoxin type B, wherein the capture antibody is capable of targeting botulinum neurotoxin type B, and the capture antibody comprises heavy chain CDR1-3 having amino acid sequences as shown in GFAFSSYD, ISSGGSST and ARQAFYTYDGTAMDY, respectively, and / or light chain CDR1-3 having amino acid sequences as shown in QSIVHNNGDTY, KVS and FQGSHIPWT, respectively.

[0035] The detection method of the present invention can be performed by capillary immunoelectrophoresis using equipment known in the art, such as SimpleWestern TM Protein Simple Jess multifunctional fully automatic protein blotting quantitative analysis system of Technology, which includes the detection plate, capillary assembly and detector used in the present invention. Those skilled in the art will understand that as long as it can be achieved that a sandwich complex is formed in the capillary with the capture antibody, the sample to be tested and the detection antibody and the detection liquid is catalyzed to produce a signal that can be captured by the detector, other types of capillary immunoelectrophoresis instruments or even microfluidic devices containing capillaries can also be used.

[0036] In a specific embodiment, the solution of the capture antibody is configured as a PBS buffer with a concentration of 0.1-50 mg / mL, preferably 1-10 mg / mL, at pH 7.0-7.2 without glycerol, and the solution of the detection antibody is configured as a PBS buffer with a concentration of 0.1-50 mg / mL, preferably 1-10 mg / mL, at pH 7.0-7.2, containing 30-70% (preferably 40-60%) glycerol.

[0037] In the present invention, the working solution of the capture antibody and the detection antibody can be prepared using Master Mix and Sample Buffer, and the above buffer can use the supporting reagents of the capillary immunoelectrophoresis instrument or be prepared by oneself, which is not particularly limited. In a specific embodiment, the Sample Buffer is the buffer of Protein Simple's catalog number 102660, and the Master Mix is ​​the buffer of Protein Simple's catalog number 87411.

[0038] In the present invention, when adding samples to the test plate, a ladder (i.e., a marker, a molecular weight standard) is added to the first well of the first row of the sample array, the other wells are added with the configured capture antibody solution, the second row is added with an antibody diluent, the first well of the third row is added with an antibody diluent, the other wells are test samples containing type B botulinum neurotoxin, the first well of the fourth row is Streptavidin-HRP, the other wells are detection antibodies, i.e., enzyme-labeled antibodies (HRP-labeled anti-type B botulinum neurotoxin monoclonal antibodies), the fifth row is detection liquid, i.e., luminescent liquid, and the subsequent arrays can be added with Wash Buffer. The above-mentioned ladder, antibody diluent, buffer, luminescent liquid, and Wash Buffer can use the supporting reagents of the capillary immunoelectrophoresis instrument or be self-configured, and there is no particular limitation on this.

[0039] The botulinum neurotoxin type B detected by the method of the present invention has a characteristic peak of 140±6 kDa.

[0040] Detection Kits

[0041] The present invention further provides a detection kit, which is used for capillary immunoelectrophoresis detection (or quantification) of botulinum neurotoxin type B, and the detection kit comprises the detection antibody and capture antibody described in the present invention, and instructions on how to implement the capillary immunoelectrophoresis detection method of the present invention for botulinum neurotoxin type B.

[0042] Herein, the term "kit" refers to a combination of reagents and other materials. The kit is expected to include reagents, such as buffers, protein stabilizing agents, signal generating systems (e.g., fluorescent signal generating systems), antibodies or their antigen binding fragments, control proteins, and test containers (e.g., microtiter plates, etc.). The term "kit" is not limited to a specific combination of reagents and / or other materials, for example, the kit may also include instructions for using the reagents. The kit may be packaged in any suitable manner, typically having components in a single container or (if necessary) in multiple containers and instructions for performing detection instructions. The kit may be prepared by a variety of methods known in the art.

[0043] In certain embodiments, the kit may further include at least one of a cleaning solution, a substrate solution, a diluent, and a calibration solution. The composition of the cleaning solution is not particularly limited, and examples thereof include, but are not limited to, a buffer, a surfactant, and a preservative. The substrate solution may use a known substrate, and examples thereof include, but are not limited to, a chromogenic substrate, a fluorescent substrate, a luminescent substrate, and the like. The composition of the diluent is not particularly limited, and examples thereof include, but are not limited to, a buffer, a surfactant, and the like. The composition of the calibration solution is not particularly limited, and examples thereof include, but are not limited to, a BSA solution, a trehalose solution, an animal serum, and the like.

[0044] Example 1

[0045] 1. Materials

[0046] 1. Capture antibody and enzyme-labeled antibody

[0047] The antibody pair is the most critical material for detecting botulinum toxin molecules. The antibody pair used in the present invention is screened from several monoclonal antibodies against botulinum neurotoxin type B using capillary immunoassay technology. The capture antibody includes anti-botulinum neurotoxin monoclonal antibody B1 (which contains heavy chain CDR1-3 with amino acid sequences such as GFAFSSYD, ISSGGSST and ARQAFYTYDGTAMDY, and light chain CDR1-3 with amino acid sequences such as QSIVHNNGDTY, KVS and FQGSHIPWT), which is homemade by the company and purified by protein A affinity chromatography, with a concentration of 1 mg / mL, in a PBS buffer with a pH of 7.0-7.2, without glycerol, and stored at -80°C. The detection antibody is HRP-labeled anti-botulinum neurotoxin type B monoclonal antibody A1 (anti-botulinum neurotoxin type B monoclonal antibody A1 contains the heavy chain amino acid sequence shown in SEQ ID NO.1 and the light chain amino acid sequence shown in SEQ ID NO.2). It is a self-made monoclonal antibody of our company and purified by affinity chromatography with a purity of more than 90%. It is labeled with HRP by Jiangsu Feiya HRP coupling kit. The concentration of the enzyme-labeled antibody is 1 mg / mL, present in PBS buffer with a pH of 7.0-7.2, containing 50% glycerol, and stored at -20°C.

[0048] 2. The capillary immunoelectrophoresis instrument is a Proteinsimple Jess product.

[0049] 2. Methods

[0050] 1. Reagent Configuration

[0051] Prepare 5×Master Mix (i.e., Loading Buffer): add 20 μL of water for injection, 20 μL of 10×Sample Buffer, pipette evenly, and set aside for use.

[0052] Prepare Ladder (i.e., Marker, molecular weight standard): add 20 μL of water for injection, pipette and mix well, and set aside for use.

[0053] Capture antibody: anti-botulinum neurotoxin type B monoclonal antibody B1.

[0054] Antigen: botulinum neurotoxin type B, original concentration 2.3 mg / mL, serial dilutions 10 μg, 100 ng, 10 ng, 1 ng, 100 pg.

[0055] Enzyme-labeled antibody: HRP-labeled anti-botulinum neurotoxin type B monoclonal antibody A1, 60-fold dilution.

[0056] Prepare various types of capture antibodies: prepare each well according to the table below.

[0057] Final concentration (μg / μL) Sample stock solution (μL) 5×Master Mix (μL) 0.1×Sample Buffer (μL) Total volume (μL) 0.5 0.5×4.5 / y 0.9 4.5-(0.5×4.5 / y)-0.9 4.5

[0058] 2. Detection

[0059] Vortex and centrifuge the prepared capture antibody and place it on ice for later use.

[0060] The order of loading samples on the detection plate: The order of loading samples on the detection plate, row A: A1 is Ladder, 5μL, and the remaining wells are configured capture antibodies, with a volume of 3μL; row B: Antibody Diluent II (antibody diluent), with a volume of 10μL; row C: C1 is Antibody Diluent II, and the remaining wells are botulinum neurotoxin type B, with a volume of 10μL; row D: D1 is Streptavidin-HRP, and the remaining wells are HRP-labeled anti-botulinum neurotoxin type B monoclonal antibody A1, with a volume of 10μL; row E: luminescent liquid, with a volume of 15μL.

[0061] The reaction plate was centrifuged at 4000 rpm for 5 minutes, 500 μL WashBuffer was added to each grid of the three rows adjacent to row F, the sealing film was torn open, the reaction plate was placed in Proteinsimple Jess, the capillary assembly was inserted, and the reaction program installed in the computer was started.

[0062] 3. Test results

[0063] The results are as follows Figure 1-5 As shown in the figure, the characteristic graph of the capillary immunoelectrophoresis sandwich reaction of botulinum neurotoxin type B and anti-botulinum neurotoxin type B monoclonal antibody is a peak of about 140 kDa, and 100 pg of botulinum neurotoxin type B was detected. In addition, the peak area and peak height of the reaction peak can reflect the amount of neurotoxin. When the neurotoxin is degraded, the peak area value decreases. At the same time, the degradation of the toxin may cause impurities other than the characteristic peak, thereby determining the stability or degradation degree and purity of the toxin.

[0064] Example 2

[0065] This example demonstrates an improvement in the qualitative detection of botulinum neurotoxin type B by capillary immunoelectrophoresis sandwich assay.

[0066] The capture antibody used was anti-botulinum neurotoxin type B monoclonal antibody B1; the antigen to be tested: botulinum neurotoxin type B, original concentration 2.3 mg / mL, serially diluted to 1 μg, 100 ng, 10 ng, 1 ng, 100 pg, 10 pg; enzyme-labeled antibody: HRP-labeled anti-botulinum neurotoxin type B monoclonal antibody A1, 30-fold dilution.

[0067] Test results such as Figure 6-11 As can be seen from the figure, the characteristic graph of the capillary immunoelectrophoresis sandwich reaction of botulinum neurotoxin type B and anti-botulinum neurotoxin type B monoclonal antibody is about 140kDa peak, and 10pg of botulinum neurotoxin type B was detected. In addition, the peak area and peak height of the reaction peak can reflect the amount of neurotoxin. When the neurotoxin decomposes, the peak area value decreases. At the same time, the degradation of the toxin may cause the appearance of impurities other than the characteristic peak, thereby determining the stability and purity of the toxin.

[0068] Comparative Example

[0069] The following shows the qualitative detection of botulinum neurotoxin type B by ELISA sandwich method.

[0070] 1. Materials

[0071] Capture antibody: anti-botulinum neurotoxin type B monoclonal antibody B1.

[0072] Antigen: Botulinum neurotoxin type B, original concentration 2.3 mg / mL.

[0073] Enzyme-labeled antibody: HRP-labeled anti-botulinum neurotoxin type B monoclonal antibody A1.

[0074] 2. Methods

[0075] (1) Capture antibody coating

[0076] Use commercial coating solution (Solebol) to dilute type B monoclonal antibody B1, according to the optimal concentration of 5 μg / mL determined by square array titration method, and coat 7 wells: A1-A7, 100 μL / well, at 4°C overnight.

[0077] (2) Closed

[0078] 5 mL of PBST (Shengang) was added with 0.1 g of BSA (BBI) to prepare blocking solution. The ELISA plate was washed with a plate washer and 100 μL of blocking solution was added to each well at 37°C for 1 h.

[0079] (3) Adding antigen

[0080] Botulinum neurotoxin type B, 2.3 mg / mL, was diluted with PBST to 23 μg / mL, 2.3 μg / mL, 230 ng / mL, 23 ng / mL, 2.3 ng / mL, and 230 pg / mL, and the ELISA plate was washed with a plate washer.

[0081] Toxins of various dilutions were added to wells A1-A6 in sequence, and PBST was added to well A7, 100 μL / well, incubated at 37°C for 1 h.

[0082] (4) Add enzyme-labeled antibody

[0083] HRP-labeled type B monoclonal antibody A1 was diluted with antibody diluent (Borsi) and diluted 1000 times according to the optimal dilution factor determined by the square array titration method. The ELISA plate was washed with a plate washer, and 1000-fold diluted ELISA antibody was added to each well, 100 μL / well, and incubated at 37°C for 1 h.

[0084] (5) Color rendering

[0085] The ELISA plate was washed with a plate washer, and 100 μL / well of colorimetric solution (Maokang Biotechnology) was added at 37°C for 5 min. 100 μL / well of stop solution (Maokang Biotechnology) was added, and the absorbance value at 620 nm was read using a Thermo Multoscan Go ELISA reader.

[0086] 3. Test results

[0087] Test results such as Fig.12 As shown, the antigen concentration corresponding to the weak positive well A4 is 23ng / mL, and the antigen concentration corresponding to the negative well A5 is 2.3ng / mL. The antibody pair composed of anti-type B neurotoxin monoclonal antibody B1 and enzyme-labeled antibody A1 has a sensitivity of more than 2.3ng / mL for sandwich ELISA qualitative detection of type B neurotoxin, which is difficult to achieve pg-level sensitivity. This method is used for qualitative detection results that are negative or positive, and cannot generate detailed immune reaction complex peak diagrams, and cannot make deeper analysis of the purity and stability of the toxin.

[0088] 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 them; 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 of the technical features therein by equivalents. However, these modifications or replacements do not deviate the essence of the corresponding technical solutions from the spirit and scope of the technical solutions of the embodiments of the present invention.

Claims

1. A method for detecting botulinum neurotoxin type B, characterized in that: The method comprises forming a sandwich complex with a capture antibody, a test sample and a detection antibody in a capillary and catalyzing a detection liquid to produce a signal that can be captured by a detector, thereby determining the presence of botulinum neurotoxin type B and / or quantifying botulinum neurotoxin type B, wherein the capture antibody is capable of targeting botulinum neurotoxin type B, and the capture antibody comprises heavy chain CDR1-3 having amino acid sequences as shown in GFAFSSYD, ISSGGSST and ARQAFYTYDGTAMDY, respectively, and light chain CDR1-3 having amino acid sequences as shown in QSIVHNNGDTY, KVS and FQGSHIPWT, respectively.

2. The method for detecting botulinum neurotoxin type B according to claim 1, characterized in that: The method comprises the following steps: (1) adding the capture antibody, the sample to be tested, the detection antibody and the detection solution to the detection plate of the capillary immunoelectrophoresis device respectively; (2) allowing the sample to be tested to bind to the capture antibody in the capillary assembly and then further bind to the detection antibody to form a complex, and pass through the capillary in different zones; (3) allowing the detection antibody to catalyze the detection solution, and determining the presence and / or quantification of botulinum neurotoxin type B based on the signal captured by the detector.

3. The method for detecting botulinum neurotoxin type B according to claim 1, characterized in that: The detection antibody has a heavy chain amino acid sequence shown in SEQ ID NO.1 and a light chain amino acid sequence shown in SEQ ID NO.

2.

4. The method for detecting botulinum neurotoxin type B according to claim 1, characterized in that: The detection antibody is an enzyme-labeled antibody, and the enzyme used for labeling includes horseradish peroxidase, alkaline phosphatase or beta-galactosidase.

5. The method for detecting botulinum neurotoxin type B according to claim 4, characterized in that: The detection liquid includes a chemiluminescent agent or a color developer, the detection liquid corresponding to the horseradish peroxidase includes at least one of luminol, o-phenylenediamine, tetramethylbenzidine, 5-aminosalicylic acid and 2,2-azino-bis(3-ethyl-benzothiazole-6-sulfonic acid) diammonium salt, the detection liquid corresponding to the alkaline phosphatase includes at least one of AMPPD, acridinium ester and 4-methylumbelliferyl phosphate, and the detection liquid corresponding to the β-galactosidase includes at least one of 5-bromo-4-chloro-3-indole-β-D-galactoside, p-nitrophenyl phosphate and 4-methylumbelliferyl-RD galactoside.

6. The method for detecting botulinum neurotoxin type B according to claim 1, characterized in that: The concentration of the capture antibody and the detection antibody is 0.1-50 mg / mL.

7. A capillary immunoelectrophoresis detection kit for botulinum neurotoxin type B, characterized in that: include: I. A capture antibody solution, wherein the capture antibody is capable of targeting botulinum neurotoxin type B, and the capture antibody comprises heavy chain CDR1-3 having amino acid sequences as shown in GFAFSSYD, ISSGGSST, and ARQAFYTYDGTAMDY, respectively, and light chain CDR1-3 having amino acid sequences as shown in QSIVHNNGDTY, KVS, and FQGSHIPWT, respectively; II. Detection antibody solution; III. Test fluid.

8. The capillary immunoelectrophoresis detection kit for botulinum neurotoxin type B according to claim 7, characterized in that: The detection solution is a mixed solution of luminol and hydrogen peroxide.

9. The capillary immunoelectrophoresis detection kit for botulinum neurotoxin type B according to claim 7, characterized in that: The detection kit further includes Master Mix and Sample Buffer.

10. The method according to any one of claims 1 to 6, or the use of the kit according to any one of claims 7 to 9, characterized in that: The application includes the production of botulinum neurotoxin type B or the quality control and monitoring of botulinum neurotoxin type B standards.

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