论文标题
使用小金纳米颗粒作为单探针的相关光电子显微镜
Correlative light electron microscopy using small gold nanoparticles as single probes
论文作者
论文摘要
相关光学显微镜(CLEM)需要在光和电子显微镜中可见的可见稳定探针。在这里,我们演示了使用小金纳米颗粒作为单个探针的Clem方法。使用谐振四波混合(FWM),通过光显微镜在人类癌细胞中与表皮生长因子蛋白结合的单个金纳米颗粒在人类癌细胞中与纳米精度背景无关,并且与相应的透射电子显微镜图像具有高度的相关性。我们使用了10 nm和5 nm半径的纳米颗粒,并在大于10 um的区域上显示出低于60 nm的相关精度,而无需额外的基准标记。通过减少系统误差,将相关精度提高到40 nm以下,而定位精度低于10 nm。极化分辨的FWM与纳米颗粒形状相关,有望在未来的应用中通过形状识别多路复用。由于金纳米颗粒的光稳定性以及FWM显微镜对活细胞的适用性,FWM-Clem为基于荧光的方法提供了强大的替代方案。
Correlative light electron microscopy (CLEM) requires the availability of robust probes which are visible both in light and electron microscopy. Here we demonstrate a CLEM approach using small gold nanoparticles as a single probe. Individual gold nanoparticles bound to the epidermal growth factor protein were located with nanometric precision background-free in human cancer cells by light microscopy using resonant four-wave-mixing (FWM), and were correlatively mapped with high accuracy to the corresponding transmission electron microscopy images. We used nanoparticles of 10 nm and 5 nm radius, and show a correlation accuracy below 60 nm over an area larger than 10 um size, without the need for additional fiducial markers. Correlation accuracy was improved to below 40 nm by reducing systematic errors, while the localisation precision is below 10 nm. Polarisation-resolved FWM correlates with nanoparticle shapes, promising for multiplexing by shape recognition in future applications. Owing to the photostability of gold nanoparticles and the applicability of FWM microscopy to living cells, FWM-CLEM opens up a powerful alternative to fluorescence-based methods.