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

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.¹.