Cell Biology and Biophysics Unit, European Molecular Biology Laboratory, Heidelberg, Germany.
Nat Biotechnol. 2011 Aug 7;29(9):835-9. doi: 10.1038/nbt.1928.
Diffusion processes and local dynamic equilibria inside cells lead to nonuniform spatial distributions of molecules, which are essential for processes such as nuclear organization and signaling in cell division, differentiation and migration. To understand these mechanisms, spatially resolved quantitative measurements of protein abundance, mobilities and interactions are needed, but current methods have limited capabilities to study dynamic parameters. Here we describe a microscope based on light-sheet illumination that allows massively parallel fluorescence correlation spectroscopy (FCS) measurements and use it to visualize the diffusion and interactions of proteins in mammalian cells and in isolated fly tissue. Imaging the mobility of heterochromatin protein HP1α (ref. 4) in cell nuclei we could provide high-resolution diffusion maps that reveal euchromatin areas with heterochromatin-like HP1α-chromatin interactions. We expect that FCS imaging will become a useful method for the precise characterization of cellular reaction-diffusion processes.
扩散过程和细胞内的局部动力平衡导致分子的非均匀空间分布，这对于核组织和细胞分裂、分化和迁移中的信号转导等过程至关重要。为了理解这些机制，需要对蛋白质丰度、迁移率和相互作用进行空间分辨的定量测量，但目前的方法在研究动态参数方面能力有限。在这里，我们描述了一种基于光片照明的显微镜，它允许大规模并行荧光相关光谱（FCS）测量，并利用它来可视化哺乳动物细胞和分离的蝇组织中蛋白质的扩散和相互作用。通过对核内异染色质蛋白 HP1α（参考文献 4）的流动性进行成像，我们可以提供高分辨率的扩散图谱，揭示具有异染色质样 HP1α-染色质相互作用的常染色质区域。我们预计 FCS 成像将成为精确描述细胞反应-扩散过程的有用方法。
