Ultrasensitive detection is crucial for the early warning and intervention of risk factors, ultimately benefiting the environment and human health. Low levels of ochratoxin A (OTA) present a hidden yet significant threat, and rapid detection via high-performing biosensors is therefore essential. A cascade isothermal amplification aptasensor (CIA-aptasensor) was designed for OTA detection. On the surface of a magnetic bead probe, the OTA level was converted into positively correlated trigger cDNA through its competitive binding with OTA-Apt. The released trigger cDNA activated catalytic hairpin assembly followed by coupling with a hybridization chain reaction to achieve CIA. After adding graphene oxide and SYBR Green I, the background interference was eliminated to specifically obtain OTA-related fluorescence. The ultrasensitive limit of detection was 0.22 pg mL, an improvement of 1368-fold over conventional enzyme-linked aptamer sorbent assay by the same OTA-Apt, demonstrating satisfactory reliability and practicability. Thus, the CIA-aptasensor provides an enzyme- and label-free simplified homogeneous system with minimal background interference using isothermal conditions. This study provides a polymerase chain reaction–like approach for enhancing the sensitivity and performance of a biosensor, which could be extended for the application of CIA and label-free signaling strategy to other risk factors.

中文翻译：

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用于超灵敏检测赭曲霉毒素 A 的无酶和无标记级联等温扩增适体传感器


超灵敏检测对于危险因素的早期预警和干预至关重要，最终有利于环境和人类健康。低水平的赭曲霉毒素 A (OTA) 构成了隐藏但重大的威胁，因此通过高性能生物传感器进行快速检测至关重要。设计了级联等温扩增适体传感器（CIA-aptasensor）用于OTA检测。在磁珠探针表面，OTA水平通过与OTA-Apt竞争性结合而转化为正相关的触发cDNA。释放的触发 cDNA 激活催化发夹组装，然后与杂交链式反应偶联以实现 CIA。添加氧化石墨烯和SYBR Green I后，消除背景干扰，特异性获得OTA相关荧光。相同OTA-Apt的超灵敏检测限为0.22 pg mL，比传统酶联适体吸附剂检测提高了1368倍，具有良好的可靠性和实用性。因此，CIA-适体传感器提供了一种无酶和无标记的简化均质系统，使用等温条件将背景干扰降至最低。这项研究提供了一种类似于聚合酶链反应的方法，用于增强生物传感器的灵敏度和性能，该方法可以扩展到将 CIA 和无标记信号传导策略应用于其他风险因素。