Exosomes are cell-derived nanovesicles that have recently gained popularity as potential biomarkers in liquid biopsies due to the large amounts of molecular cargo they carry, such as nucleic acids and proteins. However, most existing exosome-based analytical sensing methods struggle to achieve high sensitivity and high selectivity simultaneously. In this work, we present an electrochemical micro-aptasensor for the highly sensitive detection of exosomes by integrating a micropatterned electrochemical aptasensor and a hybridization chain reaction (HCR) signal amplification method. Specifically, exosomes are enriched on CD63 aptamer-functionalized electrodes and then recognized by HCR products with avidin-horseradish peroxidase (HRP) attached using EpCAM aptamers as bridges. Subsequently, the current signal that is generated through the enzyme reaction between the HRP enzyme and 3,3’,5,5’-tetramethylbenzidine (TMB)/H₂O₂ directly correlates to the amount of bound HRP on the HCR products and thus to the number of target exosomes. By introducing anti-EpCAM aptamers, micro-aptasensors can detect cancerous exosomes with high specificity. Due to the micropatterned electrodes and HCR dual-amplification strategy, the micro-aptasensors achieve a linear detection response for a wide range of exosome concentrations from 2.5×10³ to 1×10⁷ exosomes/mL, with a detection limit of 5×10² exosomes/mL. Moreover, our method successfully detects lung cancer exosomes in serum samples of early-stage and late-stage lung cancer patients, showcasing the great potential for early cancer diagnosis.

外泌体是细胞衍生的纳米囊泡，由于它们携带大量的分子货物，例如核酸和蛋白质，最近作为液体活检中的潜在生物标志物而广受欢迎。然而，大多数现有的基于外泌体的分析传感方法难以同时实现高灵敏度和高选择性。在这项工作中，我们通过集成微图案电化学适体传感器和杂交链式反应 (HCR) 信号放大方法，提出了一种用于高灵敏度检测外泌体的电化学微适体传感器。具体来说，外泌体在 CD63 适体功能化电极上富集，然后被带有亲和素-辣根过氧化物酶 (HRP) 的 HCR 产品识别，并使用 EpCAM 适体作为桥接。随后，₂ O ₂与 HCR 产物上结合的 HRP 的量直接相关，因此与目标外泌体的数量相关。通过引入抗 EpCAM 适体，微适体传感器可以高特异性地检测癌性外泌体。由于微图案电极和HCR双放大策略，微适配体传感器在2.5×10 ³到1×10 ⁷外泌体/mL的广泛外泌体浓度范围内实现了线性检测响应，检测限为5×10 ^{2 个}外泌体/mL。此外，我们的方法成功地检测了早期和晚期肺癌患者血清样本中的肺癌外泌体，显示了早期癌症诊断的巨大潜力。