Zhao Wenjie, Zhang Lingqian, Ye Yifei, Li Yuang, Luan Xiaofeng, Liu Jinlong, Cheng Jie, Zhao Yang, Li Mingxiao, Huang Chengjun
Institute of Microelectronics, Chinese Academy of Sciences, Beijing, People's Republic of China.
School of Future Technology, University of Chinese Academy of Sciences, Beijing, People's Republic of China.
Analyst. 2021 Sep 27;146(19):5962-5972. doi: 10.1039/d1an01061a.
Tumor-derived exosomes have been recognized as potential biomarkers for cancer diagnosis because they are actively involved in cancer progression and metastasis. However, progress in practical exosome analysis is still slow due to the limitation in exosome isolation and detection. The development of microfluidic devices has provided a promising analytical platform compared with traditional methods. In this study, we develop an exosome isolation and detection method based on a microfluidic device (ExoDEP-chip), which realized microsphere mediated dielectrophoretic isolation and immunoaffinity detection. Exosomes were firstly isolated by binding to antibodies pre-immobilized on the polystyrene (PS) microsphere surface and were further detected using fluorescently labeled antibodies by fluorescence microscopy. Single microspheres were then trapped into single microwells under the DEP force in the ExoDEP-chip. A wide range from 1.4 × 10 to 1.4 × 10 exosomes per mL with a detection limit of 193 exosomes per mL was obtained. Through monitoring five proteins (CD81, CEA, EpCAM, CD147, and AFP) of exosomes from three different cell lines (A549, HEK293, and HepG2), a significant difference in marker expression levels was observed in different cell lines. Therefore, this method has good prospects in exosome-based tumor marker detection and cancer diagnosis.
肿瘤来源的外泌体已被公认为癌症诊断的潜在生物标志物，因为它们积极参与癌症的进展和转移。然而，由于外泌体分离和检测的局限性，实际外泌体分析的进展仍然缓慢。与传统方法相比，微流控装置的发展提供了一个有前景的分析平台。在本研究中，我们开发了一种基于微流控装置（ExoDEP芯片）的外泌体分离和检测方法，该方法实现了微球介导的介电电泳分离和免疫亲和检测。外泌体首先通过与预先固定在聚苯乙烯（PS）微球表面的抗体结合进行分离，然后使用荧光标记抗体通过荧光显微镜进行进一步检测。然后，在ExoDEP芯片中，单个微球在介电电泳力作用下被捕获到单个微孔中。获得了每毫升1.4×10至1.4×10个外泌体的宽范围，检测限为每毫升193个外泌体。通过监测来自三种不同细胞系（A549、HEK293和HepG2）的外泌体的五种蛋白质（CD81、CEA、EpCAM、CD147和AFP），观察到不同细胞系中标志物表达水平存在显著差异。因此，该方法在基于外泌体的肿瘤标志物检测和癌症诊断方面具有良好的前景。
