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Hybridoma cell strain for secreting nitrovin monoclonal antibody and application of hybridoma cell strain

A technology based on hybridoma cell lines and nifurene hydrazone alone, which is applied in the fields of resistance to vector-borne diseases, analytical materials, climate change adaptation, etc., and can solve problems such as inapplicability to rapid detection of a large number of samples, long detection time, and complicated processing

Active Publication Date: 2021-05-18
JIANGNAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, these methods have shortcomings such as complicated sample pretreatment and long detection time, and are not suitable for rapid detection of a large number of samples. In order to protect the interests of consumers, it is necessary to establish an efficient and rapid detection method for nifurazene

Method used

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  • Hybridoma cell strain for secreting nitrovin monoclonal antibody and application of hybridoma cell strain
  • Hybridoma cell strain for secreting nitrovin monoclonal antibody and application of hybridoma cell strain
  • Hybridoma cell strain for secreting nitrovin monoclonal antibody and application of hybridoma cell strain

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0048] Example 1: Synthesis of Nifurene Hydrazone Hapten

[0049] The structure of the nifurene hydrazone hapten after the derivation of the present invention is as follows:

[0050]

[0051] The derivation process is as follows: Nifurene hydrazone is used as a raw material, and a carboxyl group is introduced through succinic anhydride to obtain a hapten. Dissolve 30 mg of nifurazone in 10 mL of anhydrous pyridine, add 10 mg of succinic anhydride, reflux at 60°C for 6 hours, and blow dry the reaction solution with nitrogen to obtain the nifurazene derivative, which is the nifurazene hapten NTV-COOH.

Embodiment 2

[0052] Example 2: Synthesis of Nifurene Hydrazone Complete Antigen

[0053] Weigh 3.45mg nifurene hydrazone hapten (NTV-COOH), 1.73mg N-hydroxysuccinimide (NHS), dissolve in 300μL N,N-dimethylformamide (DMF), stir at room temperature for 10min ; Then weigh 2.88mg 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (EDC), fully dissolve it with 100 μL DMF, add it to the NTV-COOH solution, and stir at room temperature Reaction 4-6h (called A solution). Take 6mg of KLH, dilute it to 3mg / mL with 0.01M carbonate buffer (CBS) (referred to as solution B), then slowly add solution A to solution B drop by drop, react overnight at room temperature; then use 0.01M PBS solution Dialyze to remove the unreacted small molecule hapten to obtain the complete antigen NTV-COOH-KLH of nifurene hydrazone, which is identified by ultraviolet absorption scanning method.

Embodiment 3

[0054] Embodiment 3: the synthesis of nifurene hydrazone coating former

[0055] Dissolve 5.52 mg of nifurene hydrazone hapten (NTV-COOH) and 2.8 mg of N-hydroxysuccinimide (NHS) in 300 μL of anhydrous N,N-dimethylformamide (DMF), and stir at room temperature for 10 min , to obtain a nifurene hydrazone hapten (NTV-COOH) solution; 4.6 mg of 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (EDC) was dissolved in 100 μL of anhydrous DMF After that, it was added to the NTV-COOH solution, stirred at room temperature and reacted for 4-6h to obtain A2 solution; 6mg of chicken ovalbumin (OVA) was diluted with 0.01mmol / L carbonate buffer solution (CBS) with 1mL concentration to obtain Solution B2; slowly add solution A2 dropwise into solution B2 for reaction to obtain a reaction solution; dialyze the reaction solution with PBS solution to remove unreacted small molecule hapten to obtain nifurene hydrazone-coated original (NTV-COOH -OVA).

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Abstract

The invention discloses a hybridoma cell strain for secreting a nitrovin monoclonal antibody and application of the hybridoma cell strain, and belongs to the field of food safety immunodetection. The hybridoma cell strain FQJ 5G11 for secreting the nitrovin monoclonal antibody is preserved in the China General Microbiological Culture Collection Center (CGMCC), is classified and named as a monoclonal cell strain, and has a preservation date of September 27, 2020 and a preservation number of CGMCC No.20784. The method comprises the following steps of synthesizing a nitrovin hapten, preparing a nitrovin complete antigen, mixing and emulsifying the nitrovin hapten and the nitrovin complete antigen with an equivalent Freund's adjuvant completely, immunizing a BALB / c mouse through back subcutaneous injection, and performing screening by an indirect competitive enzyme-linked immunosorbent assay and three times of subcloning to obtain the hybridoma cell strain FQJ 5G11. The obtained nitrovin monoclonal antibody cell strain can be used for immunoassay detection, and has relatively good detection sensitivity and specificity for nitrovin.

Description

technical field [0001] The invention relates to a hybridoma cell strain secreting nifurene hydrazone monoclonal antibody and an application thereof, which belongs to the field of food safety immune detection. Background technique [0002] Nitrovin is an antibiotic that can promote the growth of animals. It can effectively improve the digestion and absorption of nutrients in the animal digestive tract by increasing the permeability of the intestinal mucosa of animals; It can effectively inhibit the utilization of glucose by pathogenic microorganisms in the digestive tract of animals, hinder their metabolic process, thereby inhibit and kill a variety of original microorganisms, reduce the chance of animal infection and improve disease resistance. There are certain applications in farming. Due to the characteristics of antibiotic resistance, residual and superinfection, the disadvantages and harms brought about have become a major challenge for human beings. Announcement of t...

Claims

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Application Information

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IPC IPC(8): C12N5/20C07K16/44C07K14/795C07K14/77C07D307/70G01N33/577G01N33/53C12R1/91
CPCC07K16/44C07K14/795C07K14/77C07D307/70G01N33/577G01N33/9446Y02A40/70
Inventor 胥传来路倩倩匡华徐丽广孙茂忠刘丽强宋珊珊吴晓玲郝昌龙胡拥明
Owner JIANGNAN UNIV
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