Imaging Fluorescence-Correlation Spectroscopy for Measuring Fast Surface Diffusion at Liquid/Solid Interfaces
Imaging Fluorescence-Correlation Spectroscopy for Measuring Fast Surface Diffusion at Liquid/Solid Interfaces
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DOI：
10.1021/ac5014354
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发表时间：
2014-08-05
影响因子：
7.4
通讯作者：
Harris, Joel M.
中科院分区：
文献类型：
--
作者：
Cooper, Justin T.;Harris, Joel M.
开发探测界面分子传输的技术对于理解和优化基于表面的分析方法（包括表面增强光谱、生物测定和化学分离）是重要的。单分子荧光成像和跟踪已被用于测量荧光分子在表面的横向扩散速率，但该技术仅限于研究较慢的扩散，其中分子必须在图像采集期间保持相对静止，以便在斑点中建立足够的强度来检测和定位分子。虽然更快的时间分辨率可以通过荧光相关光谱（FCS），其中在一个小斑点的强度波动与表面上的分子的运动有关，长寿命的吸附事件所产生的表面不均匀性可以压倒相关测量和掩模的表面扩散的移动人口。在这里，我们利用这两种技术的组合，成像-FCS，用于测量快速界面传输在模型色谱表面。这是通过使用电子倍增电荷耦合器件（CCD）相机对表面进行快速成像来实现的，同时将采集限制在相机上的小区域以允许快速成帧速率。从采样区域的总强度是自相关的，以确定分子的表面扩散速率与毫秒时间分辨率。该技术允许电子控制的采集区域，这可以用来避免强吸附网站，从而最大限度地减少其对测量的自相关衰减的贡献，并改变采集面积，以解决表面扩散的吸附和解吸动力学。作为概念证明，使用成像-FCS测量1，1 '-双十八烷基-3，3，3' 3 '-四甲基吲哚碳菁（DiI）在平面C-18-和C-1-改性表面上的表面扩散速率、界面群体和吸附解吸速率。
The development of techniques to probe interfacial molecular transport is important for understanding and optimizing surface-based analytical methods including surface-enhanced spectroscopies, biological assays, and chemical separations. Single-molecule-fluorescence imaging and tracking has been used to measure lateral diffusion rates of fluorescent molecules at surfaces, but the technique is limited to the study of slower diffusion, where molecules must remain relatively stationary during acquisition of an image in order to build up sufficient intensity in a spot to detect and localize the molecule. Although faster time resolution can be achieved by fluorescence-correlation spectroscopy (FCS), where intensity fluctuations in a small spot are related to the motions of molecules on the surface, long-lived adsorption events arising from surface inhomogeneity can overwhelm the correlation measurement and mask the surface diffusion of the moving population. Here, we exploit a combination of these two techniques, imaging-FCS, for measurement of fast interfacial transport at a model chromatographic surface. This is accomplished by rapid imaging of the surface using an electron-multiplied-charged-coupled-device (CCD) camera, while limiting the acquisition to a small area on the camera to allow fast framing rates. The total intensity from the sampled region is autocorrelated to determine surface diffusion rates of molecules with millisecond time resolution. The technique allows electronic control over the acquisition region, which can be used to avoid strong adsorption sites and thus minimize their contribution to the measured autocorrelation decay and to vary the acquisition area to resolve surface diffusion from adsorption and desorption kinetics. As proof of concept, imaging-FCS was used to measure surface diffusion rates, interfacial populations, and adsorption desorption rates of 1,1'-dioctadecyl-3,3,3'3'-tetramethylindocarbocyanine (DiI) on planar C-18- and C-1-modified surfaces.