The cytoplasm is densely packed with macromolecules and organelles, displaying viscoelastic properties at various scales. How biochemical reactions function efficiently enough in a seemingly jammed environment remains elusive. Cell-free Xenopus laevis extracts represent a powerful system for investigating the biochemistry and biophysics of living systems. Here we present a protocol for characterizing macromolecular diffusion in self-organizing cytoplasmic extracts using fluorescence correlation spectroscopy (FCS), which measures the motions on a distance scale of ~200 nm. The method can also be used to characterize diffusion in the cytoplasm as it progresses through different phases of the cell cycle.
Keywords: Cell extracts; Cytoplasm; Diffusion; Fluorescence correlation spectroscopy; Self-organization.
© 2024. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.