赭曲霉毒素A和玉米赤霉烯酮-二联胶体金免疫层析试纸条的制备及应用

Dual flow immunochromatographic assay for simultaneous determination of ochratoxin A and zearalenone in cereal and feed samples

摘要:

【背景】真菌毒素为真菌的有毒次级代谢产物，混合污染时毒性显著增强，可对人类和动物健康造成严重伤害。制备二联胶体金免疫层析试纸条，实现对常见真菌毒素混合污染的快速监测，具有重要意义。【目的】制备赭曲霉毒素A (OTA)和玉米赤霉烯酮(ZEN)金标单克隆抗体，基于免疫层析原理，采用竞争反应模式，建立二联胶体金免疫层析检测法用于污染样品中OTA和ZEN的同时快速检测。【方法】采用柠檬酸钠还原法制备胶体金颗粒，并标记获得两种真菌毒素金标单克隆抗体，通过优化相关条件，建立稳定的二联胶体金免疫层析检测方法，用于同时检测谷物和饲料样品中的OTA和ZEN。【结果】制备的OTA和ZEN二联胶体金试纸条对OTA和ZEN的检测限分别为0.625 ng/mL和1.25 ng/mL，且与谷物和饲料中其它真菌毒素(黄曲霉毒素B1、伏马毒素B1、桔青霉毒素、展青霉毒素和呕吐毒素)均无交叉反应，人工添加试验结果准确。对天然样本检测结果表明该方法与LC-MS/MS一致性良好。【结论】本研究制备的二联胶体金试纸条可用于实际样品中OTA和ZEN的同时快速筛查。

Abstract:

[Background] Mycotoxins produced by different species of fungi may coexist in single cereal or feed samples, which could become highly toxic for humans and animals. In this context, a dual flow immunochromatographic assay for rapid monitoring mixed mycotoxin contamination has become a necessity. [Objective] Development of the colloidal gold dual immunoassay for simultaneous detection of ochratoxin A and zearalenone in cereal and feed samples. [Methods] Mycotoxins monoclonal antibodies were labeled with colloidal gold nanoparticles. After optimizing the materials, reagents as well as the experimental conditions, a dual flow immunochromatographic assay was developed and tested for determining the ochratoxin A and zearalenone in cereal and feed samples. [Results] The detection limit for ochratoxin A and zearalenone was 0.625 ng/mL and 1.25 ng/mL, respectively. Naturally contaminated cereal and feed samples were analyzed using both immunochromatographic assay and LC-MS/MS, and showed a good agreement between these two methods. [Conclusion] This immunoassay could detect ochratoxin A and zearalenone in cereal and feed samples rapidly and accurately, more easily and at a lower cost.