Hybridoma cell strain capable of secreting pyridaben monoclonal antibody and application of hybridoma cell strain

A hybridoma cell line and monoclonal antibody technology, applied in the field of immunoassay, can solve the problems of expensive equipment, high solvent consumption, etc., and achieve the effect of good detection sensitivity

Active Publication Date: 2021-01-29
JIANGNAN UNIV
View PDF1 Cites 5 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Despite the high sensitivity and specificity of these chromatography-based methods, there are some disadvantages such as the need for thorough sample cleanup, high solvent consumption, expensive equipment, and skilled technicians

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • Hybridoma cell strain capable of secreting pyridaben monoclonal antibody and application of hybridoma cell strain

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0046] Example 1: Synthesis of Pyridaben Hapten

[0047] Since the small molecule of pyridaben is not immunogenic and cannot stimulate the immune response of mice to produce antibodies, it is necessary to couple pyridaben to the protein through protein linkage technology to obtain immunogenicity; protein coupling Active groups commonly used in technology include amino, carboxyl, hydroxyl, mercapto, etc. Since the molecular structure of pyridaben does not contain these active groups, it needs to be derivatized.

[0048] Weigh 7.30 g (20 mmol) of pyridaben into a 100 mL three-neck flask, dissolve it in 20 mL DMSO, and then add 2.25 g (40 mmol) of KOH. Another 2.10 g (20 mmol) of β-mercaptopropionic acid was weighed, dissolved in 10 mL DMSO and transferred to a 50 mL constant pressure dropping funnel. Slowly add the β-mercaptopropionic acid solution dropwise into the three-necked flask with stirring, and slowly raise the temperature to 100°C on the oil bath, keep the temperature...

Embodiment 2

[0049] Example 2: Synthesis of Pyridaben Complete Antigen

[0050] Weigh 5.5 mg of pyridaben hapten and 4.2 mg of N-hydroxysuccinimide (NHS) prepared in Example 1, dissolve them in 300 μL of N,N-dimethylformamide (DMF), and stir at room temperature for 10 minutes 6.9mg 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (EDC) was weighed again, after fully dissolving with 100 μL DMF, it was added to the pyridaben hapten solution, Stir the reaction at room temperature for 6-8 h (referred to as liquid A). Take 8mg of BSA, dilute it to 4mg / 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 Dialyzed to remove unreacted small molecular hapten to obtain the complete antigen, which was identified by UV absorption scanning method.

Embodiment 3

[0051] Embodiment 3: the synthesis of pyridaben coating former

[0052] Dissolve 3.0 mg of pyridaben hapten and 2.3 mg of N-hydroxysuccinimide (NHS) in 300 μL of anhydrous N,N-dimethylformamide (DMF), and stir at room temperature for 10 minutes to obtain pyridaben hapten Solution: Dissolve 3.8mg of 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (EDC) in 100μL of anhydrous DMF and add it to the pyridaben hapten solution at room temperature Stir and react for 6-8 hours to obtain liquid A; dilute 10 mg of chicken ovalbumin (OVA) with 1 mL of carbonate buffer solution (CBS) with a concentration of 0.01 mmol / L to obtain liquid B; slowly add liquid A drop by drop React in solution B to obtain a reaction solution; dialyze the reaction solution with PBS solution to remove unreacted small molecule hapten to obtain the coating original.

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to view more

PUM

No PUM Login to view more

Abstract

The invention discloses a hybridoma cell strain capable of secreting a pyridaben monoclonal antibody and an application of the hybridoma cell strain, and belongs to the technical field of immunodetection. The invention discloses a hybridoma cell strain ABC05 capable of secreting a pyridaben monoclonal antibody, which is preserved in China General Microbiological Culture Collection Center and is classified and named as a monoclonal cell strain on November 28, 2019, and the preservation number is CGMCC No.19174. The BALB / c mouse with the complete antigen of pyridaben is subjected to immunizing.The spleen cells of a high-titer and low-IC50 mouse are fused with myeloma cells of the mouse through a PEG method, the fused hybridoma cells are cultured through a culture medium, then cells are screened through an indirect competitive enzyme-linked immunosorbent assay method, subcloning is conducted three times, and finally the monoclonal antibody hybridoma cell strain is obtained. The monoclonal antibody secreted by the cell strain has good detection sensitivity (the IC50 value is 9.09 ng / mL) to pyridaben, and can be used for residue detection of pyridaben in food.

Description

technical field [0001] The invention relates to a hybridoma cell strain secreting pyridaben monoclonal antibody and application thereof, belonging to the technical field of immune detection. Background technique [0002] The English common name of pyridaben is: Pyridaben, the Chinese name is also known as: Altair, Tadaben, Pyridaben, and the chemical name: 2-tert-butyl-5-(4-tert-butylbenzylsulfur)-4- Chloro-3 (dihydro) pyridazinone; mechanism of action: pyridaben belongs to the class of pyridazinones to kill insects and mites. It has strong contact killing, but it has no fumigation, systemic and conduction effects. It mainly inhibits the synthesis of glutamate dehydrogenase in muscle tissue, nerve tissue and chromosome I of electron transfer system, thereby exerting insecticidal effect. Main application: It has obvious control effects on all plant-eating pest mites, such as Panonychia, Tetranychus, Eriodous mites, and Clawed mites, etc., and is effective in different growth...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to view more

Application Information

Patent Timeline
no application Login to view more
IPC IPC(8): C12N5/20C07K16/44C07K14/765C07D237/18G01N33/577C12R1/91
CPCC07K16/44C07K14/765C07D237/18G01N33/577G01N33/94
Inventor 胥传来刘杰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
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Try Eureka
PatSnap group products

Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic.

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap