Quantitative Light Imaging Laboratory, Department of Mechanical Science and Engineering, Beckman Institute for Advanced Science and Technology, University of Illinois at Urbana-Champaign, Urbana, Illinois 61801, USA.
Phys Rev Lett. 2012 Nov 2;109(18):188104. doi: 10.1103/PhysRevLett.109.188104.
Because of its ability to study specifically labeled structures, fluorescence microscopy is the most widely used technique for investigating live cell dynamics and function. Fluorescence correlation spectroscopy is an established method for studying molecular transport and diffusion coefficients at a fixed spatial scale. We propose a new approach, dispersion-relation fluorescence spectroscopy (DFS), to study the transport dynamics over a broad range of spatial and temporal scales. The molecules of interest are labeled with a fluorophore whose motion gives rise to spontaneous fluorescence intensity fluctuations that are analyzed to quantify the governing mass transport dynamics. These data are characterized by the effective dispersion relation. We report on experiments demonstrating that DFS can distinguish diffusive from advection motion in a model system, where we obtain quantitatively accurate values of both diffusivities and advection velocities. Because of its spatially resolved information, DFS can distinguish between directed and diffusive transport in living cells. Our data indicate that the fluorescently labeled actin cytoskeleton exhibits active transport motion along a direction parallel to the fibers and diffusive in the perpendicular direction.
由于能够研究特定标记的结构，荧光显微镜是研究活细胞动力学和功能最广泛使用的技术。荧光相关光谱学是一种用于在固定空间尺度上研究分子输运和扩散系数的成熟方法。我们提出了一种新的方法，即弥散关系荧光光谱学（DFS），以研究广泛的时空尺度上的输运动力学。感兴趣的分子用荧光团标记，其运动导致自发荧光强度波动，对其进行分析以定量地描述控制质量输运动力学。这些数据的特征是有效弥散关系。我们报告了实验结果，表明 DFS 可以区分模型系统中的扩散和对流运动，我们获得了两种扩散率和对流速度的定量准确值。由于其具有空间分辨信息，DFS 可以区分活细胞中的定向和扩散输运。我们的数据表明，荧光标记的肌动蛋白细胞骨架表现出沿纤维平行方向的主动输运运动，而在垂直方向上则表现出扩散运动。
