At the single-cell level, cellular parameters, gene expression and cellular function are assayed on an individual but not population-average basis. Essential to observing and analyzing the heterogeneity and behavior of these cells/clusters is the ability to prepare and manipulate individuals. Here, we demonstrate a versatile microsystem, a stereo acoustic streaming tunnel, which is triggered by ultrahigh-frequency bulk acoustic waves and highly confined by a microchannel. We thoroughly analyze the generation and features of stereo acoustic streaming to develop a virtual tunnel for observation, pretreatment and analysis of cells for different single-cell applications. 3D reconstruction, dissociation of clusters, selective trapping/release, in situ analysis and pairing of single cells with barcode gel beads were demonstrated. To further verify the reliability and robustness of this technology in complex biosamples, the separation of circulating tumor cells from undiluted blood based on properties of both physics and immunity was achieved. With the rich selection of handling modes, the platform has the potential to be a full-process microsystem, from pretreatment to analysis, and used in numerous fields, such as in vitro diagnosis, high-throughput single-cell sequencing and drug development.