Vertical nanopillars for highly localized fluorescence imaging
Vertical nanopillars for highly localized fluorescence imaging
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DOI：
10.1073/pnas.1015589108
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发表时间：
2011-03-08
影响因子：
11.1
通讯作者：
Cui, Bianxiao
中科院分区：
文献类型：
--
作者：
Xie, Chong;Hanson, Lindsey;Cui, Bianxiao
在生物和医学研究中，通过荧光显微镜观察复杂环境中的单个分子变得越来越重要，这需要减少观察体积。在这里，我们展示了使用垂直排列的二氧化硅纳米柱在体外和活细胞内实现低于衍射极限的观察体积。由于直径远小于可见光的波长，嵌入在不透明衬底中的透明二氧化硅纳米柱限制了光的传播，并沿着其垂直表面提供倏逝波激发。这种效应产生高度受限的照明体积，其选择性地激发纳米柱附近的荧光分子。我们表明，这种纳米柱照明可用于在体外高荧光团浓度的单分子检测。此外，我们还证明了垂直纳米柱与活细胞紧密接触，并在细胞内作为高度局部化的光源。此外，纳米柱表面的特定化学修饰使得可以局部招募感兴趣的蛋白质，同时观察它们在细胞复杂拥挤环境中的行为。
Observing individual molecules in a complex environment by fluorescence microscopy is becoming increasingly important in biological and medical research, for which critical reduction of observation volume is required. Here, we demonstrate the use of vertically aligned silicon dioxide nanopillars to achieve below-the-diffraction-limit observation volume in vitro and inside live cells. With a diameter much smaller than the wavelength of visible light, a transparent silicon dioxide nanopillar embedded in a non-transparent substrate restricts the propagation of light and affords evanescence wave excitation along its vertical surface. This effect creates highly confined illumination volume that selectively excites fluorescence molecules in the vicinity of the nanopillar. We show that this nanopillar illumination can be used for in vitro single-molecule detection at high fluorophore concentrations. In addition, we demonstrate that vertical nanopillars interface tightly with live cells and function as highly localized light sources inside the cell. Furthermore, specific chemical modification of the nanopillar surface makes it possible to locally recruit proteins of interest and simultaneously observe their behavior within the complex, crowded environment of the cell.