Background. This study aimed to explore the zearalenone (ZEN) immunogen synthesis method, immunogenicity, and antibody characteristics and to lay a foundation for the establishment of immunoassay methods for ZEN single residue and ZEN and its analogs total residue. Methods. Based on the molecular structure and active sites of ZEN, oxime active ester (OAE), condensation mixed anhydride (CMA), formaldehyde (FA), and 1,4-butanediol diglycidyl ether method (BDE) were designed and used for immunogen (ZEN-BSA) synthesis. The immunogens were identified by infrared (IR) and ultraviolet (UV) spectra and gel electrophoresis (SDS-PAGE) and were then used to immunize Balb/c mice to prepare ZEN polyclonal antibody (ZEN pAb). The titers and sensitivity of the ZEN pAb were determined by indirect noncompetitive ELISA (inELISA) and indirect competitive ELISA (icELISA), respectively, and its specificity was assessed by the cross-reaction test (CR). Results. ZEN-BSA was successfully synthesized, and the molecular binding ratios of ZEN to BSA were 17.2 : 1 (OAE), 14.6 : 1 (CMA), 9.7 : 1 (FA), and 8.3 : 1 (BDE), respectively. The highest inELISA titers of ZEN pAb of each group were 1 : (6.4 × 10³) (OAE), 1 : (3.2 × 10³) (CMA), 1 : (1.6 × 10³) (FA), and 1 : (1.6 × 10³) (BDE), respectively. The 50% inhibition concentrations (IC50) for ZEN by icELISA of each group were 11.67 μg/L (OAE), 16.29 μg/L (CMA), 20.92 μg/L (FA) and 24.36 μg/L (BDE), respectively. ZEN pAb from the mice immunized with ZEN-BSA (OAE) and ZEN-BSA (CMA) had class broad specificity to ZEN and its analogs. The CRs of ZEN pAb with α-ZAL, β-ZAL, α-ZOL, β-ZOL, and ZON were 36.53%, 16.98%, 64.33%, 20.16%, and 10.66%, respectively. ZEN pAb from the mice immunized with ZEN-BSA (FA) and ZEN-BSA (BDE) had high specificity for ZEN. The CRs of ZEN pAb with its analogs were all less than 1.0%. Conclusion. This study demonstrated that the preparation of the class broad-specificity antibodies of ZEN and its analogs can be achieved by immunizing animals with the immunogen ZEN-BSA prepared by the OAE method, while the preparation of highly specific antibodies can be achieved by immunizing animals with the immunogen ZEN-BSA prepared by the FA method. These findings lay the material and technical foundation for immunoassay of ZEN single residue and ZEN and its analogs total residue.

中文翻译：

---

玉米赤霉烯酮免疫原的合成及抗体特性比较分析

背景。本研究旨在探讨玉米赤霉烯酮（ZEN）的免疫原合成方法、免疫原性及抗体特性，为建立ZEN单残基和ZEN及其类似物总残基的免疫测定方法奠定基础。方法. 基于ZEN的分子结构和活性位点，设计了肟活性酯（OAE）、缩合混合酸酐（CMA）、甲醛（FA）和1,4-丁二醇二缩水甘油醚法（BDE）等方法用于免疫原（ZEN -BSA) 合成。通过红外（IR）和紫外（UV）光谱和凝胶电泳（SDS-PAGE）鉴定免疫原，然后用于免疫Balb / c小鼠以制备ZEN多克隆抗体（ZEN pAb）。ZEN pAb的滴度和敏感性分别通过间接非竞争性ELISA（inELISA）和间接竞争性ELISA（icELISA）测定，其特异性通过交叉反应试验（CR）评估。结果. ZEN-BSA合成成功，ZEN与BSA的分子结合比分别为17.2:1（OAE）、14.6:1（CMA）、9.7:1（FA）和8.3:1（BDE）。各组ZEN pAb的ELISA效价最高分别为1：（6.4×10 ³）（OAE）、1：（3.2×10 ³）（CMA）、1：（1.6×10 ³）（FA）和1： (1.6 × 10 ³ ) (BDE)。各组icELISA对ZEN的50%抑制浓度（IC50）分别为11.67  μ g/L（OAE）、16.29  μ g/L（CMA）、20.92  μ g/L（FA）和24.36  μ克/升（BDE），分别。来自用 ZEN-BSA (OAE) 和 ZEN-BSA (CMA) 免疫的小鼠的 ZEN pAb 对 ZEN 及其类似物具有广泛的特异性。ZEN pAb与α -ZAL、β -ZAL、α -ZOL、β -ZOL和ZON的CR分别为36.53%、16.98%、64.33%、20.16%和10.66%。来自用 ZEN-BSA (FA) 和 ZEN-BSA (BDE) 免疫的小鼠的 ZEN pAb 对 ZEN 具有高特异性。ZEN pAb 及其类似物的 CR 均小于 1.0%。结论. 本研究表明，用OAE法制备的免疫原ZEN-BSA对动物进行免疫可以制备ZEN及其类似物类广谱特异性抗体，而用OAE法制备的免疫原ZEN-BSA对动物进行免疫可以制备高特异性抗体。 FA法制备的免疫原ZEN-BSA。这些发现为ZEN单残基和ZEN及其类似物总残基的免疫测定奠定了物质和技术基础。