Simultaneous Real-Time Three-Dimensional Localization and FRET Measurement of Two Distinct Particles,Nano Letters

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Simultaneous Real-Time Three-Dimensional Localization and FRET Measurement of Two Distinct Particles
Nano Letters ( IF 9.6 ) Pub Date : 2021-09-07 , DOI: 10.1021/acs.nanolett.1c01328
Xingxiang Chen ₁ , Teng Liu ₂ , Xianan Qin ₂ , Quang Quan Nguyen ₂ , Sang Kwon Lee ₃ , Chanwoo Lee ₄ , Yaguang Ren ₅ , Jun Chu ₆ , Guang Zhu _{1,

5} , Tae-Young Yoon ₄ , Chan Young Park ₃ , Hyokeun Park _{1,

2,

7}

Affiliation

Many biological processes employ mechanisms involving the locations and interactions of multiple components. Given that most biological processes occur in three dimensions, the simultaneous measurement of three-dimensional locations and interactions is necessary. However, the simultaneous three-dimensional precise localization and measurement of interactions in real time remains challenging. Here, we report a new microscopy technique to localize two spectrally distinct particles in three dimensions with an accuracy (2.35σ) of tens of nanometers with an exposure time of 100 ms and to measure their real-time interactions using fluorescence resonance energy transfer (FRET) simultaneously. Using this microscope, we tracked two distinct vesicles containing t-SNAREs or v-SNARE in three dimensions and observed FRET simultaneously during single-vesicle fusion in real time, revealing the nanoscale motion and interactions of single vesicles in vesicle fusion. Thus, this study demonstrates that our microscope can provide detailed information about real-time three-dimensional nanoscale locations, motion, and interactions in biological processes.

中文翻译：

两个不同粒子的同时实时三维定位和 FRET 测量

许多生物过程采用涉及多个组件的位置和相互作用的机制。鉴于大多数生物过程发生在三个维度，因此需要同时测量三个维度的位置和相互作用。然而，实时同步三维精确定位和测量交互仍然具有挑战性。在这里，我们报告了一种新的显微技术，以数十纳米的精度 (2.35σ) 在三个维度上定位两个光谱不同的粒子，曝光时间为 100 毫秒，并使用荧光共振能量转移 (FRET) 测量它们的实时相互作用）同时。使用这个显微镜，我们在三个维度上跟踪了两个不同的包含 t-SNARE 或 v-SNARE 的囊泡，并在单囊泡融合过程中实时观察了 FRET，揭示了囊泡融合中单个囊泡的纳米级运动和相互作用。因此，这项研究表明，我们的显微镜可以提供有关生物过程中实时三维纳米级位置、运动和相互作用的详细信息。

更新日期：2021-09-22

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