Preparation and characterisation of monoclonal antibodies against deoxynivalenol

Abstract

To prepare broad-spectrum and specific monoclonal antibodies (mAb) against deoxynivalenol (DON), DON-BSA was synthesised by the carbonyl diimidazole method as an artificial antigen to immunise Balb/C mice, and the coated DON-OVA was synthesised by the carbodiimide method to detect anti-DON antibodies. Monoclonal antibodies were screened by indirect ELISA and indirect competitive ELISA. A hybridoma cell line (4F3) capable of stably secreting DON antibody was obtained. The titre of antibody in the culture supernatant was 1:1.28 Â 10 3 , and the titre of the ascites antibody was 1:3.2 Â 10 5 . The monoclonal antibody homology was IgG1/k. The half inhibitory concentration (IC 50 ) of DON was 9.84 ng/mL. The cross-reaction rates with 3-Ac-DON and 15-Ac-DON were 60.44% and 52.04%, respectively, but no cross-reaction with other mycotoxins was observed. The results showed that the anti-DON monoclonal antibodies prepared in this experiment could recognise not only DON but also 3-Ac-DON and 15-Ac-DON, which could provide materials for the next step to establish a method to detect DON and its similar compounds. HIGHLIGHTSThis study demonstrated that an artificial antigen was synthesised by carbonyl diimidazole method, and a novel broad-spectrum, high affinity monoclonal antibody was developed. The anti-DON mAb developed in this study has better sensitivity, moreover the antibody was sensitive to DON and its derivatives. Therefore, it could be used for simultaneous monitoring of three mycotoxins. Results of this study will further lay the foundation for the establishment of immunological assays for the total amount of similar compounds.

Synthesis of Zearalenone Immunogen and Comparative Analysis of Antibody Characteristics

“…e ZEN pAb titers were tested through indirect noncompetitive ELISA (inELISA) using the procedure described by Han et al [34]. e coating antigen ZEN-OVA was diluted in CBS at 2 μg/mL, and 100 mL/well was added to the 96-well microplate and incubated at 37.8°C for 2 h. After washing with PBST three times, unbound active sites were blocked with 250 μL/well of blocking buffer at 37.8°C for 1 h or 4°C overnight.…”

Comparison of Synthetic Methods and Identification of Several Artificial Antigens of Deoxynivalenol

“…The results showed that the CDI method described by Maragos et al [ 28 ] and the improved method had the best effects, and the 3-hydroxy group on the DON molecule had a greater chance of reacting. Therefore, in this study, the hydroxyl group at the C3 position of the DON molecule was successfully covalently coupled to the amino group of the carrier protein BSA by the CDI activation method to obtain the artificial antigen DON-BSA without blocking the hydroxyl group at any site, and the imidazole group was directly introduced to achieve coupling with the carrier protein BSA [ 29 ]. The artificial antigen synthesised by this method was not very different from that produced by the preparation method reported by Lee et al [ 30 ] in the previous literature.…”

Immunological quantitative detection of dicofol in medicinal materials

“…DICH was coupled with KLH as the immunogen (DICH-KLH) using the carbodiimide method. 20–22 Briefly, 10.0 mg of DICH and 12.0 mg of N -hydroxysuccinimide were dissolved in 400 μL of DMF. Then, 24.0 mg of 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride dissolved in 50 μL of MES (0.1 mol L −1 , pH 4.0) was added with continuous stirring for 8 h at room temperature.…”