[Progress and prospect of separation and analysis of single-cell and single-particle exosomes]

Exosomes are nanoscale vesicles secreted by cells and are encapsulated in lipid bilayers. They play crucial roles in cell communication and are involved in a variety of physiological and pathological processes, including immune regulation, angiogenesis, and tumor initiation and metastasis. Exosomes carry a variety of biomolecules from maternal cells and are therefore important vehicles for discovering disease markers. Traditional detection methods only provide average cell-population information for a given sample and cannot establish clear relationships between the biological functions of exosomes and subtype owing to the significant heterogeneity associated with exosomes from different cell subsets. Therefore, characterizing exosomes at the single-cell and single-particle levels requires exosome specificities to be further explored and the characteristics of various exosome subtypes to be distinguished. Commonly used single-particle exosome characterization technologies include flow cytometry, super-resolution microscopy, atomic force microscopy, surface-enhanced Raman spectroscopy, proximity barcoding assay and MS. In this paper, we summarize recent advances in the separation and characterization of single-cell exosomes based on microfluidics and provide future applications prospects for emerging technologies (such as Olink proteomics, click chemistry, and molecular imprinting) for studying single-cell and single-particle exosomes.