Anti-natamycin monoclonal antibody hybridoma cell strain and application thereof

A hybridoma cell line and monoclonal antibody technology, applied in biochemical equipment and methods, instruments, microorganisms, etc., can solve the problems of expensive instruments, time-consuming, and relatively high requirements for experimental conditions, and achieve good detection sensitivity and specificity Effect

Active Publication Date: 2015-04-29
JIANGNAN UNIV
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Problems solved by technology

The biological potency determination method has high requirements on the experimental environment factors (temperature, etc.), material factors (culture medium, etc.), operating technology and other conditions, and the operation process is relatively cumbersome; the microplate bioassay method is suitable for large quantities of fermentation broth or samples detection; UV spectrophotometry is generally used as a routine control for natamycin detection, if it is used for quantitative analysis, the requirements for experimental con

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  • Anti-natamycin monoclonal antibody hybridoma cell strain and application thereof

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Embodiment 1

[0020] Example 1: Preparation of hybridoma cell line M

[0021] 1. Antigen synthesis

[0022] Weigh 20 mg of natamycin and dissolve it in 3 mL of DMF; weigh 100 mg of BSA and dissolve it in 5 mL of PBS; add 50 μL of 25% glutaraldehyde solution drop by drop to the natamycin solution, and store it in the dark at room temperature Stir and activate for 30 min; add the activated natamycin dropwise to BSA, and stir and react at room temperature for 4 h in the dark. After the reaction, unreacted natamycin was removed by dialysis with 0.01M PBS for three days to obtain a conjugate of natamycin and BSA, which was characterized by ultraviolet light.

[0023] 2. mouse immunization

[0024] Healthy BALB / c mice aged 6-8 weeks were selected for immunization. Natamycin antigen was mixed and emulsified with an equal amount of Freund's adjuvant, and BALB / c mice were immunized by subcutaneous injection in the back. Complete Freund's adjuvant was used for the first immunization, followed by...

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Abstract

The invention discloses an anti-natamycin monoclonal antibody hybridoma cell strain and application thereof, and belongs to the technical field of food safety immunological detection. The scheme adopted by the invention is as follows: mixing and emulsifying natamycin complete antigen with an equal amount of complete freund's adjuvant, and then immunizing BALB/c mouse through subcutaneous injection on the back of the mouse, wherein the first immunization uses the complete freund's adjuvant, and subsequent immunization uses an incomplete freund's adjuvant; fusing splenocytes of the mouse with a high titre and a low IC50 value and myeloma cells of the mouse through a PEG method; performing indirect competitive ELISA screening and three times of subcloning to obtain a hybridoma (monoclonal) cell strain. Monoclonal antibodies secreted by the hybridoma (monoclonal) cell strain has relatively good specificity and detection sensitivity (the value of IC50 is 2.2 microgram/L) for natamycin, and can be used for detecting residual natamycin in food safety.

Description

technical field [0001] The invention discloses an anti-natamycin monoclonal antibody hybridoma cell line and application thereof, belonging to the technical field of food safety immunology detection, and relates to a hybridoma cell line M and an anti-natamycin monoclonal antibody produced therefrom. Background technique [0002] Natamycin is a natural, broad-spectrum, efficient and safe inhibitor of filamentous fungi such as yeast and mold, belonging to polyene macrolides. It not only inhibits fungi, but also prevents the production of mycotoxins. It can be widely used in food preservation and antifungal treatment. Natamycin has no inhibitory effect on bacteria, so it does not affect the natural ripening process of yogurt, cheese, raw ham, dry sausage. In June 1982, the U.S. FDA officially approved natamycin as a food preservative. In 1996, Natamycin was officially approved by China Food Additives Standardization Chemical Technical Committee as a food preservative. [00...

Claims

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Application Information

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IPC IPC(8): C12N5/20C07K16/44G01N33/577
Inventor 刘丽强陈燕妮胥传来匡华徐丽广马伟宋珊珊吴晓玲
Owner JIANGNAN UNIV
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