Exosomes are a class of cell-secreted, nano-sized extracellular vesicles with a bilayer membrane structure of 30–150 nm in diameter. Their discovery and application have brought breakthroughs in numerous areas, such as liquid biopsies, cancer biology, drug delivery, immunotherapy, tissue repair, and cardiovascular diseases. Isolation of exosomes is the first step in exosome-related research and its applications. Standard benchtop exosome separation and sensing techniques are tedious and challenging, as they require large sample volumes, multi-step operations that are complex and time-consuming, requiring cumbersome and expensive instruments. In contrast, microfluidic platforms have the potential to overcome some of these limitations, owing to their high-precision processing, ability to handle liquids at a microscale, and integrability with various functional units, such as mixers, actuators, reactors, separators, and sensors. These platforms can optimize the detection process on a single device, representing a robust and versatile technique for exosome separation and sensing to attain high purity and high recovery rates with a short processing time. Herein, we overview microfluidic strategies for exosome isolation based on their hydrodynamic properties, size filtration, acoustic fields, immunoaffinity, and dielectrophoretic properties. We focus especially on advances in label-free isolation of exosomes with active biological properties and intact morphological structures. Further, we introduce microfluidic techniques for the detection of exosomal proteins and RNAs with high sensitivity, high specificity, and low detection limits. We summarize the biomedical applications of exosome-mediated therapeutic delivery targeting cancer cells. To highlight the advantages of microfluidic platforms, conventional techniques are included for comparison. Future challenges and prospects of microfluidics towards exosome isolation applications are also discussed. Although the use of exosomes in clinical applications still faces biological, technical, regulatory, and market challenges, in the foreseeable future, recent developments in microfluidic technologies are expected to pave the way for tailoring exosome-related applications in precision medicine.

外泌体是一类细胞分泌的、纳米尺寸的细胞外囊泡，具有直径为30-150 nm的双层膜结构。它们的发现和应用在液体活检、癌症生物学、药物输送、免疫治疗、组织修复和心血管疾病等众多领域带来了突破。外泌体的分离是外泌体相关研究及其应用的第一步。标准台式外泌体分离和传感技术既繁琐又具有挑战性，因为它们需要大量样品、复杂且耗时的多步骤操作，需要笨重且昂贵的仪器。相比之下，微流控平台具有克服其中一些限制的潜力，因为它们具有高精度处理能力、处理微尺度液体的能力以及与各种功能单元（例如混合器、执行器、反应器、分离器和传感器）的集成性。这些平台可以优化单个设备上的检测过程，代表了一种强大且通用的外泌体分离和传感技术，可在较短的处理时间内获得高纯度和高回收率。在此，我们根据外泌体的流体动力学特性、尺寸过滤、声场、免疫亲和性和介电泳特性概述了外泌体分离的微流体策略。我们特别关注具有活性生物学特性和完整形态结构的外泌体的无标记分离方面的进展。此外，我们引入了微流体技术，用于检测外泌体蛋白和 RNA，具有高灵敏度、高特异性和低检测限。我们总结了外泌体介导的靶向癌细胞的治疗递送的生物医学应用。为了突出微流控平台的优势，包括传统技术进行比较。还讨论了微流控外泌体分离应用的未来挑战和前景。尽管外泌体在临床应用中的使用仍面临生物学、技术、监管和市场方面的挑战，但在可预见的未来，微流控技术的最新发展有望为精准医疗中外泌体相关应用的定制铺平道路。