Raman image-activated cell sorting

Nao Nitta,Hiroshi Karakawa,Yasutaka Kitahama,Atsuhiro Nakagawa,Hiroshi Watarai,Takeichiro Sekiya,Takashi Yamano,Yuki Inoue,Hiroshi Tezuka,Masaaki Inaba ,Takanori Iino,Masayuki Yazawa,Takeshi Sugimura ,Hideharu Mikami,Hideya Fukuzawa,Minoru Oikawa,Kei Hiraki,Satoshi Matsusaka,Tomohisa Hasunuma,Shinya Sakuma,Yuichi Kato,Kazumasa Hiramatsu ,Seizo Tanaka ,Mai Yamagishi,Takuro Ito,Shunsuke Uemura ,Yoichiroh Hosokawa,Takeshi Hayakawa,Takuya Asai,Tadataka Ota,Nobutake Suzuki,Yu Hoshino,Cheng Lei,Kiyotaka Shiba,Yasuyuki Ozeki,Dino Di Carlo,Dinghuan Deng,Yusuke Yonamine,Akihiro Isozaki,Yusuke Kasai,Keiji Numata,Suguru Ueno ,Fumihito Arai,Masataka Kajikawa,Yoshitaka Shirasaki,Yuta Suzuki,Misa Hase,Hirofumi Kobayashi,Keisuke Goda

doi:10.1038/s41467-020-17285-3

Abstract

The advent of image-activated cell sorting and imaging-based cell picking has advanced our knowledge and exploitation of biological systems in the last decade. Unfortunately, they generally rely on fluorescent labeling for cellular phenotyping, an indirect measure of the molecular landscape in the cell, which has critical limitations. Here we demonstrate Raman image-activated cell sorting by directly probing chemically specific intracellular molecular vibrations via ultrafast multicolor stimulated Raman scattering (SRS) microscopy for cellular phenotyping. Specifically, the technology enables real-time SRS-image-based sorting of single live cells with a throughput of up to ~100 events per second without the need for fluorescent labeling. To show the broad utility of the technology, we show its applicability to diverse cell types and sizes. The technology is highly versatile and holds promise for numerous applications that are previously difficult or undesirable with fluorescence-based technologies.

Highlights

The advent of image-activated cell sorting and imaging-based cell picking has advanced our knowledge and exploitation of biological systems in the last decade
The basic operation of the Raman imageactivated cell sorter (RIACS) is as follows: suspended cells in a sample tube injected into the RIACS are focused by the hydrodynamic and acoustic focusers into a single stream, detected by the event detector, imaged by the stimulated Raman scattering (SRS) microscope, analyzed by the real-time Raman image processor, and sorted into a collection or waste tube by the dual-membrane push-pull cell sorter triggered by decision signals generated by the image processor
The capabilities of the RIACS can be enhanced in multiple directions

Summary

Introduction

The advent of image-activated cell sorting and imaging-based cell picking has advanced our knowledge and exploitation of biological systems in the last decade They generally rely on fluorescent labeling for cellular phenotyping, an indirect measure of the molecular landscape in the cell, which has critical limitations. We anticipate that the ability to sort cells based on unlabeled yet molecularly specific images with high throughput will significantly extend the utility and applicability of image-activated cell sorting and is, expected to further advance single-cell biology and applications In this Article, we demonstrate Raman image-activated cell sorting by directly measuring chemically specific intracellular molecular vibrations via coherent Raman scattering for cellular phenotyping without the need for fluorescent labeling. The RIACS is a highly versatile platform technology and holds promise for numerous applications that are previously difficult or undesirable with fluorescence-based singlecell sorting or isolation technologies[24]

Methods

Results

Conclusion