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

【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 (Ochratoxin A,OTA)和玉米赤霉烯酮(Zeralenone,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。

[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.