Versatile, Facile and Low-Cost Single-Cell Isolation, Culture and Sequencing by Optical Tweezer-Assisted Pool-Screening

Real-time image-based sorting of target cells in a precisely indexed manner is desirable for sequencing or cultivating individual human or microbial cells directly from clinical or environmental samples, however versatility of existing methods is limited as they are usually not broadly applicable to all cell sizes. Here, an Optical tweezer-assisted Pool-screening and Single-cell Isolation (OPSI) system is established for precise, indexed isolation of individual bacterial, yeast or human-cancer cells. A controllable static flow field that acts as cell-pool is achieved in a microfluidics chip, to enable precise and readily screening of cells of 1 to 40 um in size by bright-field, fluorescence, or Raman imaging. The target cell is then captured by a 1064 nm optical tweezer and deposited as one-cell-harboring nanoliter microdroplets in a one-cell-one-tube manner. For bacterial, yeast and human cells, OPSI achieves >99.7% target-cell sorting purity and10-fold elevated speed of 10~20 cells/min. Moreover, OPSI-based one-cell RNA-seq of human cancer cells yields high quality and reproducibility of single-cell transcriptome profiles. The versality, facileness, flexibility, modularized design, and low cost of OPSI suggest its broad applications for image-based sorting of target cells.