A practical guide to intelligent image-activated cell sorting

Akihiro Isozaki, Hideharu Mikami, Kotaro Hiramatsu, Shinya Sakuma, Yusuke Kasai, Takanori Iino, Takashi Yamano, Atsushi Yasumoto, Yusuke Oguchi, Nobutake Suzuki, Yoshitaka Shirasaki, Taichiro Endo, Takuro Ito, Kei Hiraki, Makoto Yamada, Satoshi Matsusaka, Takeshi Hayakawa, Hideya Fukuzawa, Yutaka Yatomi, Fumihito AraiDino Di Carlo, Atsuhiro Nakagawa, Yu Hoshino, Yoichiroh Hosokawa, Sotaro Uemura, Takeaki Sugimura, Yasuyuki Ozeki, Nao Nitta, Keisuke Goda

Research output: Contribution to journalArticlepeer-review

70 Citations (Scopus)


Intelligent image-activated cell sorting (iIACS) is a machine-intelligence technology that performs real-time intelligent image-based sorting of single cells with high throughput. iIACS extends beyond the capabilities of fluorescence-activated cell sorting (FACS) from fluorescence intensity profiles of cells to multidimensional images, thereby enabling high-content sorting of cells or cell clusters with unique spatial chemical and morphological traits. Therefore, iIACS serves as an integral part of holistic single-cell analysis by enabling direct links between population-level analysis (flow cytometry), cell-level analysis (microscopy), and gene-level analysis (sequencing). Specifically, iIACS is based on a seamless integration of high-throughput cell microscopy (e.g., multicolor fluorescence imaging, bright-field imaging), cell focusing, cell sorting, and deep learning on a hybrid software–hardware data management infrastructure, enabling real-time automated operation for data acquisition, data processing, intelligent decision making, and actuation. Here, we provide a practical guide to iIACS that describes how to design, build, characterize, and use an iIACS machine. The guide includes the consideration of several important design parameters, such as throughput, sensitivity, dynamic range, image quality, sort purity, and sort yield; the development and integration of optical, microfluidic, electrical, computational, and mechanical components; and the characterization and practical usage of the integrated system. Assuming that all components are readily available, a team of several researchers experienced in optics, electronics, digital signal processing, microfluidics, mechatronics, and flow cytometry can complete this protocol in ~3 months.

Original languageEnglish
Pages (from-to)2370-2415
Number of pages46
JournalNature Protocols
Issue number8
Publication statusPublished - Aug 1 2019

All Science Journal Classification (ASJC) codes

  • Biochemistry, Genetics and Molecular Biology(all)


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