Hapten and artificial antigen of monepantel sulfone as well as preparation method and application of hapten and artificial antigen
An artificial antigen and hapten technology, which is applied in the field of food safety testing, can solve the problems of expensive instruments, complex pretreatment process operators, and unsuitability for rapid screening of a large number of samples, and achieves low price, high sensitivity, simple and efficient synthesis method Effect
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Embodiment 1
[0034] Example 1, Preparation and Characterization of Monepal Sulfone Hapten
[0035] Weigh 100 mg of p-trifluoromethylthiobenzoic acid and dissolve in 10 mL of glacial acetic acid, add 20 mL of 37% hydrogen peroxide dropwise while stirring, and react at room temperature 25°C for 24 hours. The reaction solution was diluted with pure water and then extracted with dichloromethane. The extract was spin-dried and washed with pure water. After washing, 86 mg of white solid was obtained by suction filtration, which was a monetyl sulfone hapten and was identified by mass spectrometry.
[0036] The reaction equation is as follows:
[0037]
[0038] Mass spectrometry results such as figure 1 As shown, the molecular weight of 4-((trifluoromethyl)sulfonyl)benzoic acid is 254.18, and the parent ion fragment obtained in negative ion mode is m / z=253.18, and there is a characteristic parent ion of m / z=253.0 in the mass spectrum Fragments, indicating successful synthesis of the monetal s...
Embodiment 2
[0039] Example 2, Preparation and Characterization of Monetal Sulfone Artificial Antigen
[0040] In the preparation methods of the immunogen and the coating source, the difference lies in the type of carrier protein used. The carrier protein of the immunogen is mainly KLH, the carrier protein of the coating source is mainly BSA and OVA, and the coupling method used is the active ester method.
[0041] Weigh 20 mg of the prepared monetyl sulfone hapten (MOPS), 26.2 mg of N-hydroxysuccinimide (NHS), 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide Hydrochloride (EDC) 43.6mg, dissolved in 1.5mL N,N-dimethylformamide (DMF), stirred at room temperature for 6 hours to obtain liquid A; take 50mg BSA (50mg OVA or 20mg KLH) with 0.05M Dissolve CB to obtain liquid B; slowly drop liquid A into liquid B drop by drop, stir and react overnight at room temperature to obtain a mixture of conjugates; dialyze the mixture in 0.01M PBS for three days to finally obtain monepal Sulfone artificial an...
Embodiment 3
[0045] Embodiment 3, the preparation of monetal sulfone monoclonal antibody
[0046] The artificial antigen MOPS-KLH prepared in Example 2 was used to immunize six 6-8 week old BALB / c female mice. Each immunogen was diluted to 1 mg / mL with 0.01 M PBS, and emulsified with an equal amount of Freund's adjuvant to form a water-in-oil structure. Except Freund's complete adjuvant was used for the first immunization, Freund's incomplete adjuvant was used for the rest of the booster immunization, the immunization dose was 100 μg / mouse, the injection dose was 0.2 mL / mouse, and the mice were inoculated at multiple points on the back of the neck. After 4 weeks, booster immunization was carried out and immunization was given once every 3 weeks, for a total of 2 additional immunizations, which were changed to multi-point subcutaneous injections on the back of the neck. Afterwards, a booster immunization was carried out every three weeks, and Freund's complete adjuvant was replaced with Fr...
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