Chen Xincheng, He Zhenyun, Jiao Sujia, Sun Zhichang, Zhang Sihang, Liu Xing
School of Food Science and Engineering, Key Laboratory of Tropical Fruits and Vegetables Quality and Safety for State Market Regulation Hainan University, Haikou 570228, China.
School of Food Science and Engineering, Key Laboratory of Tropical Fruits and Vegetables Quality and Safety for State Market Regulation Hainan University, Haikou 570228, China; College of International Tourism, Hainan College of Economics and Business, Haikou 571127, China.
J Hazard Mater. 2025 Aug 15;494:138539. doi: 10.1016/j.jhazmat.2025.138539. Epub 2025 May 10.
Ochratoxin A (OTA) seriously threatens public health and agricultural ecosystems, necessitating sensitive detection methods. In view of that traditional immunoassays rely on animal-derived antibodies and toxic substrates, we developed a dual-mode nanosensor integrating colorimetric and fluorescent detection via alkaline phosphatase (ALP)-mediated silver nanoparticles (AgNPs) growth and inner filter effect (IFE). The Ag-triggered aggregation-induced emission (AIE) enhancement of gold nanoclusters (AuNCs) was investigated for constructing a dual-mode nanosensor. The nanosensor employs an OTA-specific nanobody-ALP fusion (Nb-ALP) to dephosphorylate ascorbic acid 2-phosphate into ascorbic acid (AA), which reduces Ag⁺ to abundant AgNPs with NaBH-reduced AgNPs seeds. AgNPs amplify plasmonic absorbance for colorimetric ALP detection, while quenching Ag⁺@AuNC fluorescence via IFE for fluorescent detection. The nanosensor was further combined with the Nb-ALP-based immunoassay to develop a dual-mode nanosensor-powered enzyme immunoassay (DMN-EIA). The DMN-EIA exhibited detection limits of 0.14 ng/mL (colorimetric) and 0.28 ng/mL (fluorescent) with high selectivity for OTA, with spike recovery rates of 93.5 %-108.7 % and relative standard deviations not exceeding 20 %. Furthermore, the colorimetric (R = 0.96) and fluorescent DMN-EIA (R = 0.93) correlated well with high-performance liquid chromatography in detection of ten real pepper samples. Therefore, the developed dual-mode nanosensor and DMN-EIA represent reliable and promising tools for detecting OTA.
赭曲霉毒素A（OTA）严重威胁公众健康和农业生态系统，因此需要灵敏的检测方法。鉴于传统免疫分析依赖动物源抗体和有毒底物，我们开发了一种双模式纳米传感器，通过碱性磷酸酶（ALP）介导的银纳米颗粒（AgNPs）生长和内滤效应（IFE）实现比色和荧光检测。研究了银触发的金纳米团簇（AuNCs）聚集诱导发光（AIE）增强，以构建双模式纳米传感器。该纳米传感器采用OTA特异性纳米抗体-ALP融合蛋白（Nb-ALP）将抗坏血酸2-磷酸去磷酸化为抗坏血酸（AA），AA与硼氢化钠还原的AgNPs种子一起将Ag⁺还原为大量AgNPs。AgNPs增强等离子体吸收用于比色ALP检测，同时通过IFE淬灭Ag⁺@AuNC荧光用于荧光检测。该纳米传感器进一步与基于Nb-ALP的免疫分析相结合，开发了一种双模式纳米传感器驱动的酶免疫分析（DMN-EIA）。DMN-EIA对OTA具有高选择性，比色检测限为0.14 ng/mL，荧光检测限为0.28 ng/mL，加标回收率为93.5 %-108.7 %，相对标准偏差不超过20 %。此外，在检测十个实际辣椒样品时，比色（R = 0.96）和荧光DMN-EIA（R = 0.93）与高效液相色谱法具有良好的相关性。因此，所开发的双模式纳米传感器和DMN-EIA是检测OTA的可靠且有前景的工具。
