Weidtkamp-Peters Stefanie, Weisshart Klaus, Schmiedeberg Lars, Hemmerich Peter
Leibniz Institute for Age Research, Fritz Lipmann Institute, Jena, Germany.
Methods Mol Biol. 2009;464:321-41. doi: 10.1007/978-1-60327-461-6_18.
Recent developments in cell biology and microscopy techniques enable us to observe macromolecular assemblies in their natural setting: the living cell. These emerging technologies have revealed novel concepts in nuclear cell biology. In order to further elucidate the biochemistry of gene expression, replication, and genome maintenance, the major challenge is now to precisely determine the dynamics of nuclear proteins in the context of the structural organization of the nucleus. Fluorescence correlation spectroscopy (FCS) is an attractive alternative to photobleaching and photoactivation techniques for the analysis of protein dynamics at single-molecule resolution. Here we describe how FCS can be applied to retrieve biophysical parameters of nuclear proteins in living cells.
细胞生物学和显微镜技术的最新进展使我们能够在自然环境（即活细胞）中观察大分子组装体。这些新兴技术揭示了细胞核细胞生物学中的新概念。为了进一步阐明基因表达、复制和基因组维持的生物化学过程，目前的主要挑战是在细胞核结构组织的背景下精确确定核蛋白的动力学。荧光相关光谱法（FCS）是一种有吸引力的替代光漂白和光激活技术的方法，可用于在单分子分辨率下分析蛋白质动力学。在这里，我们描述了如何应用FCS来获取活细胞中核蛋白的生物物理参数。
