Observation of highly dynamic processes inside living cells at the single molecule level is key for a better understanding of biological systems. However, imaging of single molecules in living cells is usually limited by the spatial and temporal resolution, photobleaching and the signal-to-background ratio. To overcome these limitations, light-sheet microscopes with thin selective plane illumination, for example, in a reflected geometry with a high numerical aperture imaging objective, have been developed. Here, we developed a reflected light-sheet microscope with active optics for fast, high contrast, two-colour acquisition of -stacks. We demonstrate fast volume scanning by imaging a two-colour giant unilamellar vesicle (GUV) hemisphere. In addition, the high contrast enabled the imaging and tracking of single lipids in the GUV cap. The enhanced reflected scanning light-sheet microscope enables fast 3D scanning of artificial membrane systems and potentially live cells with single-molecule sensitivity and thereby could provide quantitative and molecular insight into the operation of cells.

中文翻译：

---

使用具有单分子灵敏度的反射光片显微镜对巨型单层囊泡进行快速 3D 成像

在单分子水平上观察活细胞内部的高度动态过程是更好地理解生物系统的关键。然而，活细胞中单个分子的成像通常受到空间和时间分辨率、光漂白和信背景比的限制。为了克服这些限制，已经开发了具有薄选择性平面照明的光片显微镜，例如，在具有高数值孔径成像物镜的反射几何中。在这里，我们开发了一种带有主动光学元件的反射光片显微镜，用于快速、高对比度、双色采集-堆栈。我们通过对双色巨型单层囊泡 (GUV) 半球进行成像来展示快速体积扫描。此外，高对比度能够对 GUV 帽中的单个脂质进行成像和跟踪。增强型反射扫描光片显微镜能够对人工膜系统和具有单分子敏感性的潜在活细胞进行快速 3D 扫描，从而可以提供对细胞操作的定量和分子洞察。