Single-cell analysis to investigate cellular heterogeneity and cell-to-cell interactions is a crucial compartment to answer key questions in important biological mechanisms. Droplet-based microfluidics appears to be the ideal platform for such a purpose because the compartmentalization of single cells into microdroplets offers unique advantages of enhancing assay sensitivity, protecting cells against external stresses, allowing versatile and precise manipulations over tested samples, and providing a stable microenvironment for long-term cell proliferation and observation. The present Review aims to give a preliminary guidance for researchers from different backgrounds to explore the field of single-cell encapsulation and analysis. A comprehensive and introductory overview of the droplet formation mechanism, fabrication methods of microchips, and a myriad of passive and active encapsulation techniques to enhance single-cell encapsulation efficiency were presented. Meanwhile, common methods for single-cell analysis, especially for long-term cell proliferation, differentiation, and observation inside microcapsules, are briefly introduced. Finally, the major challenges faced in the field are illustrated, and potential prospects for future work are discussed.
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