赭曲霉毒素A时间分辨荧光侧流层析试纸条的研究

Study of time-resolved fluorescence lateral flow chromatographic test strips of ochratoxin A

中文关键词:  赭曲霉毒素A  时间分辨荧光纳米微球  荧光定量免疫层析  快速定量检测

英文关键词:ochratoxin A  time-resolved fluorescent microsphere  fluorescence quantitative immunochromatography  rapid quantitative detection.

中文摘要:

      目的 构建一种基于时间分辨荧光纳米微球的赭曲霉毒素A (ochratoxin A, OTA)侧流层析试纸条。方法 基于免疫层析原理, 以时间分辨荧光纳米微球为信号探针, 降低非特异性荧光的干扰, 提高检测灵敏度, 并通过优化样品提取液和样品稀释液, 进一步提高现场检测OTA的灵敏度和准确性。结果 OTA在1.0~50.0 μg/kg范围内, T线和C线荧光强度比值与OTA浓度的对数线性关系良好, 相关系数r2为0.9981~0.9998。不同基质中OTA的检出限和定量限分别为0.401~0.614 μg/kg和0.973~1.617 μg/kg, 加标回收率为89.53%~118.37%, 相对标准偏差(relative standard deviations, RSDs)小于12% (n=3), 且与将呕吐毒素、伏马菌素B1、黄曲霉毒素B1、玉米赤霉烯酮和T-2毒素的交叉反应率均小于5%, 特异性良好。OTA侧流层析试纸条基于荧光定量快速检测技术平台可在8 min内快速准确地定量测定出待测样本中OTA含量。结论 本研究所制备的时间分辨荧光侧流层析试纸条可实现玉米、小麦和饲料中OTA的快速定量检测, 并具有成本低、灵敏度高、操作简便、准确高、重复性好、特异性好的优点, 可满足国内外OTA检测的技术要求, 为真菌毒素的快检技术的发展提供了技术支撑。

英文摘要:

      Objective To establish a time-resolved fluorescent microsphere-based lateral flow chromatographic test strip for ochratoxin A (OTA). Methods Based on the principle of immunochromatography, time-resolved fluorescent nanospheres were used as signal probes to reduce the interference of non-specific fluorescence and improve the detection sensitivity. Furthermore, the sensitivity and accuracy of the on-site detection of OTA were improved by optimizing the sample extract and sample diluent. Results The ratio of the T-line and C-line fluorescence intensity had a good log-linear relationship with the OTA concentration in the range of 1.0?50.0 μg/kg, and the correlation coefficient r2 was in the range of 0.9981?0.9998. The limits of detection and limits of quantitation of OTA in different matrices were in the range of 0.401?0.614 μg/kg and 0.973?1.617 μg/kg, respectively. The recoveries ranged from 89.53% to 118.37% with relative standard deviations (RSDs) all less than 12% (n=3), and the cross-reaction rates with deoxynivalenol, fumonisin B1, aflatoxin B1, zearalenone and T-2 toxin were all less than 5%, showing good specificity. The OTA lateral flow chromatographic test strips based on the fluorescent quantitative rapid detection technology platform could quickly and accurately quantify the content of OTA in samples within 8 minutes. Conclusion The time-resolved fluorescence lateral flow chromatographic test strip prepared in this study can realize the rapid quantitative detection of OTA in corn, wheat and feed, with the advantages of low cost, high sensitivity, simple operation, high accuracy, good repeatability and good specificity, which meets the technical requirements of OTA detection at home and abroad, and provides technical supports for the development of mycotoxins rapid detection technology.