Dataset Information

Versatile phenotype-activated cell sorting.

ABSTRACT: Unraveling the genetic and epigenetic determinants of phenotypes is critical for understanding and re-engineering biology and would benefit from improved methods to separate cells based on phenotypes. Here, we report SPOTlight, a versatile high-throughput technique to isolate individual yeast or human cells with unique spatiotemporal profiles from heterogeneous populations. SPOTlight relies on imaging visual phenotypes by microscopy, precise optical tagging of single target cells, and retrieval of tagged cells by fluorescence-activated cell sorting. To illustrate SPOTlight's ability to screen cells based on temporal properties, we chose to develop a photostable yellow fluorescent protein for extended imaging experiments. We screened 3 million cells expressing mutagenesis libraries and identified a bright new variant, mGold, that is the most photostable yellow fluorescent protein reported to date. We anticipate that the versatility of SPOTlight will facilitate its deployment to decipher the rules of life, understand diseases, and engineer new molecules and cells.

SUBMITTER: Lee J

PROVIDER: S-EPMC7608836 | biostudies-literature | 2020 Oct

REPOSITORIES: biostudies-literature

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Versatile phenotype-activated cell sorting.

Lee Jihwan J Liu Zhuohe Z Suzuki Peter H PH Ahrens John F JF Lai Shujuan S Lu Xiaoyu X Guan Sihui S St-Pierre François F

Science advances 20201023 43

Unraveling the genetic and epigenetic determinants of phenotypes is critical for understanding and re-engineering biology and would benefit from improved methods to separate cells based on phenotypes. Here, we report SPOTlight, a versatile high-throughput technique to isolate individual yeast or human cells with unique spatiotemporal profiles from heterogeneous populations. SPOTlight relies on imaging visual phenotypes by microscopy, precise optical tagging of single target cells, and retrieval ...[more]

PMID: 33097540

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Project description:Reactive oxygen species (ROS) play a crucial role in cancer formation and development, especially cancer metastasis. However, lack of a precise tool, which could accurately distinguish specific types of ROS, restricts an in-depth study of ROS in cancer development and progression. Herein, we designed smart and versatile fluorescent Ag nanoclusters (AgNCs) for sensitive and selective detection of different species of ROS in cells and tissues. Methods: Firstly, dual-emission fluorescent AgNCs was synthesized by using bovine serum albumin (BSA) to sense different types of ROS (H2O2, O2•-, •OH). The responsiveness of the AgNCs to different species of ROS was explored by fluorescence spectrum, hydrodynamic diameter, and so on. Furthermore, dual-emission fluorescent AgNCs was used to sense ROS in tumor with different degrees of differentiation. Finally, the relationship between specific types of ROS and tumor cell invasion was explored by cell migration ability and the expression of cell adhesion and EMT markers. Results: This dual-emission fluorescent AgNCs possessed an excellent ability to sensitively and selectively distinguish highly reactive oxygen species (hROS, including O2•-and •OH) from moderate reactive oxygen species (the form of H2O2), and exhibited no fluoresence and green fluorescence, respectively. The emission of AgNCs is effective in detecting cellular and tissular ROS. When cultured with AgNCs, malignant tumor cells exhibit non-fluorescence, while the benign tumor emits green and reduced red light and the normal cells appear in weak green and bright red fluorescence. We further verified that not just H2O2 but specific species of ROS (O2•-and •OH) were involved in cell invasion and malignant transformation. Our study warrants further research on the role of ROS in physiological and pathophysiological processes. Conclusion: Taken together, AgNCs would be a promising approach for sensing ROS, and offer an intelligent tool to detect different kinds of ROS in tumors.

Dataset Information

Versatile phenotype-activated cell sorting.

Publications

Versatile phenotype-activated cell sorting.

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