Preparation and evaluation method of scolospora toxin gene engineering antibody
By constructing a genetically engineered antibody against *Strombyx moritoxin* using RT-PCR and overlap PCR techniques, the problem of insufficient detection sensitivity in existing technologies has been solved, enabling rapid and accurate determination of *Strombyx moritoxin*.
CN121673402APending Publication Date: 2026-03-17JINAN UNIVERSITY
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Patent Information
- Application Number
- CN202511977057.1
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Priority Date
- 2025-07-03
- Filing Date
- 2025-12-25
- Publication Date
- 2026-03-17
AI Technical Summary
Technical Problem
Existing technologies have limited means of monitoring Cyclopsacrin, lacking highly sensitive detection methods, making it difficult to achieve rapid and accurate determination.
Method used
The variable region gene fragment of the *Cyclophorus toxin* genetically engineered antibody was obtained using RT-PCR, degenerate primers, and overlap PCR. The scFv gene was constructed and ligated with the mouse Fc segment. The MN8 (scFv-Fc) antibody was formed by overlap PCR and identified and detected using direct competitive ELISA.
Benefits of technology
This improved the stability and sensitivity of the immunoassay method for Cyclopsacinitoxins, providing a basis for rapid and accurate determination.
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Abstract
The preparation and evaluation method comprises the following steps: 1, extracting a heavy-chain DNA fragment and a light-chain DNA fragment, carrying out recombinant transformation by using an overlapping PCR method to construct an scFv gene, 2, constructing an expression vector, culturing the expression vector, carrying out sequencing identification to obtain an scFv bacterial solution, recombining the scFv bacterial solution with a mouse Fc fragment, and identifying a sequence to obtain an MN8 bacterial solution, the method comprises the following steps: 1, preparing an MN8 bacterial liquid, 2, extracting the MN8 bacterial liquid, 3, carrying out inoculated culture on the MN8 bacterial liquid to obtain an MN8 genetically engineered antibody bacterial liquid, 4, extracting MN8 genetically engineered antibody bacterial liquid plasmids and carrying out transfection expression purification, and 5, carrying out antibody titer and thermal stability determination on the MN8 genetically engineered antibody. The scFv single-chain antibody obtained by recombination according to the method has the advantages of small molecular weight, high penetrability and the like, the stability and the sensitivity of an immunoassay method of the scolosporins can be improved, and a certain basis is provided for rapid determination of the scolosporins.
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