使用荧光激活的非膜凝聚物分离法在HeLa细胞中分离应激颗粒的实验方案。

Protocol to isolate stress granules in HeLa cells using fluorescence-activated non-membrane condensate isolation.

作者信息

Zhou Yilong, Shvedunova Maria, Akhtar Asifa

机构信息

Max Planck Institute of Immunobiology and Epigenetics, 79108 Freiburg, Germany; Cancer Science Institute of Singapore, National University of Singapore, Singapore 117599, Singapore.

Max Planck Institute of Immunobiology and Epigenetics, 79108 Freiburg, Germany.

出版信息

STAR Protoc. 2025 Jul 26;6(3):103987. doi: 10.1016/j.xpro.2025.103987.

DOI:10.1016/j.xpro.2025.103987

PMID:40716059

原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC12319554/

Abstract

Various stress stimuli induce cytosolic stress granules (SGs) in mammalian cells, which are composed of RNA and RNA-binding proteins and have important physiological functions. Here, we present a protocol for isolating SGs using fluorescence-activated non-membrane condensate isolation (FANCI). We describe steps for seeding and stressing G3BP1-mCherry HeLa cells. We then detail procedures for purifying SGs by FANCI with flow cytometry. This protocol has potential application in studying SGs from other cells in the future. For complete details on the use and execution of this protocol, please refer to Zhou et al..

摘要

多种应激刺激可在哺乳动物细胞中诱导胞质应激颗粒（SGs）形成，应激颗粒由RNA和RNA结合蛋白组成，具有重要的生理功能。在此，我们展示了一种使用荧光激活非膜凝聚物分离（FANCI）来分离应激颗粒的方法。我们描述了接种和应激处理G3BP1- mCherry HeLa细胞的步骤。然后详细说明了通过流式细胞术利用FANCI纯化应激颗粒的程序。该方法未来在研究来自其他细胞的应激颗粒方面具有潜在应用价值。有关该方法的使用和实施的完整详细信息，请参考Zhou等人的研究。