Department of Applied Chemistry, Graduate School of Science and Engineering, Saitama University, 255 Shimo-Okubo, Sakura, Saitama 338-8570, Japan.
J Chem Phys. 2017 Jul 28;147(4):041101. doi: 10.1063/1.4985871.
We present standing evanescent-wave fluorescence correlation spectroscopy (SEW-FCS). This technique utilizes the interference of two evanescent waves which generates a standing evanescent-wave. Fringe-pattern illumination created by a standing evanescent-wave enables us to measure the diffusion coefficients of molecules with a super-resolution corresponding to one fringe width. Because the fringe width can be reliably estimated by a simple procedure, utilization of fringes is beneficial to quantitatively analyze the slow diffusion of molecules in a supported lipid bilayer (SLB), a model biomembrane formed on a solid substrate, with the timescale relevant for reliable FCS analysis. Furthermore, comparison of the data between SEW-FCS and conventional total-internal reflection FCS, which can also be performed by the SEW-FCS instrument, effectively eliminates the artifact due to afterpulsing of the photodiode detector. The versatility of SEW-FCS is demonstrated by its application to various SLBs.
我们提出了静态消逝波荧光相关光谱（SEW-FCS）技术。该技术利用两个消逝波的干涉产生静态消逝波。静态消逝波产生的条纹图案照明使我们能够测量扩散系数，其分辨率超精细对应于一个条纹宽度。由于条纹宽度可以通过简单的程序可靠地估计，因此利用条纹有利于定量分析在固体基底上形成的模型生物膜支撑脂质双层（SLB）中分子的缓慢扩散，该时间尺度与可靠的 FCS 分析相关。此外，通过静态消逝波荧光相关光谱仪进行的静态消逝波荧光相关光谱和传统的全内反射荧光相关光谱之间的数据比较，可以有效地消除光电二极管探测器的后脉冲伪影。静态消逝波荧光相关光谱的多功能性通过其在各种 SLB 中的应用得到了证明。
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