Gerlich D, Beaudouin J, Gebhard M, Ellenberg J, Eils R
Intelligent Bioinformatics Systems Department, German Cancer Research Centre, 69120 Heidelberg, Germany.
Nat Cell Biol. 2001 Sep;3(9):852-5. doi: 10.1038/ncb0901-852.
Live-cell imaging technology using fluorescent proteins (green fluorescent protein and its homologues) has revolutionized the study of cellular dynamics. But tools that can quantitatively analyse complex spatiotemporal processes in live cells remain lacking. Here we describe a new technique--fast multi-colour four-dimensional imaging combined with automated and quantitative time-space reconstruction--to fill this gap. As a proof of principle, we apply this method to study the re-formation of the nuclear envelope in live cells. Four-dimensional imaging of three spectrally distinct fluorescent proteins is used to simultaneously visualize three different cellular compartments at high speed and with high spatial resolution. The highly complex data, comprising several thousand images from a single cell, were quantitatively reconstructed in time-space by software developed in-house. This analysis reveals quantitative and qualitative insights into the highly ordered topology of nuclear envelope formation, in correlation with chromatin expansion - results that would have been impossible to achieve by manual inspection alone. Our new technique will greatly facilitate study of the highly ordered dynamic architecture of eukaryotic cells.
利用荧光蛋白（绿色荧光蛋白及其同源物）的活细胞成像技术彻底改变了细胞动力学的研究。但仍缺乏能够定量分析活细胞中复杂时空过程的工具。在此，我们描述了一种新技术——快速多色四维成像结合自动定量时空重建——以填补这一空白。作为原理验证，我们应用此方法研究活细胞中核膜的重新形成。对三种光谱不同的荧光蛋白进行四维成像，以高速和高空间分辨率同时可视化三个不同的细胞区室。由内部开发的软件对来自单个细胞的数千张图像组成的高度复杂数据进行了时空定量重建。该分析揭示了与染色质扩张相关的核膜形成高度有序拓扑结构的定量和定性见解——这些结果仅凭人工检查是不可能实现的。我们的新技术将极大地促进对真核细胞高度有序动态结构的研究。
