Lab of Biosystem and Microanalysis, State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai 200237, China.
Department of Laboratory Medicine, Renji Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai 200127, China.
Anal Chem. 2022 May 31;94(21):7703-7712. doi: 10.1021/acs.analchem.2c01187. Epub 2022 May 16.
Exosomes are lipid bilayer extracellular vesicles secreted by various types of cells and inherit abundant molecular information from parental cells. Tumor-derived exosomes have been widely recognized as noninvasive biomarkers for early cancer diagnosis and surveillance, but the separation of intact exosomes and detection of exosomal proteins remain challenging. Herein, we proposed a microfluidic chip for specific exosome isolation, integrated with sensitive quantification by a novel PTCDI-aptamer signal switch strategy. To enhance the capture efficiency, an alternating drop-shaped micropillar array was designed to assist the capture of tumor-derived exosomes by Tim4-modified magnetic beads (Tim4 beads) on the chip. Following capture, a chelating agent can easily elute intact exosomes which were further used for profiling exosomal surface proteins by the multiplexed fluorescence turn-on approach. Profiting from the efficient on-chip enrichment of the Tim4 beads and superior fluorescence signal transduction strategy, the detection limit of the analysis platform for HepG2 exosomes is as low as 8.69 × 10 particles/mL with a wide linear range spanning 6 orders of magnitude. Meanwhile, the proposed platform could recognize subtle changes in protein levels on the exosomal surface from various cell lines. More importantly, this strategy is successfully applied to analyze exosomes in human serum to distinguish liver cancer patients from healthy individuals. Combined analysis of different types of biomarkers on the exosomal membrane surface can greatly improve the accuracy of cancer type identification and disease monitoring. We hope that this convenient, rapid, and sensitive platform may become a powerful tool in the field of exosome analysis and early cancer screening.
外泌体是各种类型细胞分泌的具有双层脂膜的细胞外囊泡，从亲代细胞中继承了丰富的分子信息。肿瘤来源的外泌体已被广泛认为是用于癌症早期诊断和监测的非侵入性生物标志物，但完整外泌体的分离和外泌体蛋白的检测仍然具有挑战性。在此，我们提出了一种用于特定外泌体分离的微流控芯片，集成了通过新型 PTCDI-适体信号开关策略进行的敏感定量检测。为了提高捕获效率，设计了交替的滴形微柱阵列，以辅助 Tim4 修饰的磁性珠（Tim4 珠）在芯片上捕获肿瘤来源的外泌体。捕获后，螯合剂可以轻松洗脱完整的外泌体，然后通过多重荧光开启方法对外泌体表面蛋白进行分析。得益于 Tim4 珠在芯片上的高效富集和优越的荧光信号转导策略，该分析平台对 HepG2 外泌体的检测限低至 8.69×10 个颗粒/mL，线性范围跨越 6 个数量级。同时，该平台能够识别来自各种细胞系的外泌体表面蛋白水平的微小变化。更重要的是，该策略成功应用于分析人血清中外泌体，以区分肝癌患者和健康个体。对外泌体膜表面不同类型生物标志物的联合分析可以大大提高癌症类型识别和疾病监测的准确性。我们希望这个方便、快速和灵敏的平台可以成为外泌体分析和早期癌症筛查领域的有力工具。
