Fast super-resolution microscopy by rotating, coherently scattered laser light

通过旋转、相干散射激光实现快速超分辨率显微镜

• 批准号: 413220392
• 负责人: Professor Dr. Alexander Rohrbach
• 金额: --
• 依托单位: Institut für Mikrosystemtechnik (IMTEK)
• 依托单位国家: 德国
• 项目类别: Research Grants
• 财政年份: 2019
• 资助国家: 德国
• 起止时间: 2018-12-31 至 2022-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/413220392?language=en
• 关键词: Fast; super; resolution; microscopy; rotating

The exploration of ever smaller and thus more dynamic structures represents one of the biggest technical challenges of the presence and the future. Long known physical laws seem to set immovable metrological barriers, such as the optical resolution limit in microscopy. It has turned out that many of these barriers can be bypassed with resourceful tricks, without moving the barriers. The smaller the structures to be spatially resolved in light microscopy, the more photons and measurement time are required. However, the smaller the structures in living systems, the faster they move and the less measurement time is available.In this initiative we work on a novel optical microscopy concept, where objects are illuminated coherently and images are generated coherently as well. By exploiting defined multiple interferences under oblique illumination and angular integration of many coherent images, a spatial resolution of nearly 100nm can be achieved in principle, at a temporal resolution of typically 100 Hz and with excellent image contrast. Within the preliminary work of our group, we could achieve a spatial resolution of 150 nm through rotating coherently scattered (ROCS) laser light in TIR-dark-field mode. With this technique we could acquire thousands of images without loss in image quality (e.g. fluorophore bleaching in fluorescence microscopy) and without image reconstruction (as required e.g. in super-resolution microscopy with structured illumination).In the present research proposal, we want to reach two new goals in addition to the expected 120 nm spatial and 100 Hz temporal resolution of this technique. On the one hand, we want to distinguish specifically marked structures in the image through specific absorption and phase retardation of the scattered laser light. On the other hand, we will develop a novel kind of time-correlated microscopy shall be developed - largely independently of the first goal. Here coherent ROCS images of label-free structures with high temporal resolution will be correlated to fluorescence based images of fluorophore distributions with low temporal resolution.

探索更小、更动态的结构是当前和未来最大的技术挑战之一。众所周知的物理定律似乎设置了不可移动的障碍，例如显微镜的光学分辨率极限。事实证明，这些障碍中的许多都可以通过足智多谋的技巧绕过，而不需要移动障碍。在光学显微镜中空间分辨的结构越小，需要的光子和测量时间就越多。然而，生命系统中的结构越小，它们移动的速度就越快，可用的测量时间就越少。在这项计划中，我们致力于一种新的光学显微镜概念，其中物体被相干地照亮，图像也被相干地生成。通过利用倾斜照明下的定义的多个干涉和许多相干图像的角积分，原则上可以实现接近100 nm的空间分辨率，时间分辨率通常为100 Hz，并具有优异的图像对比度。在我们小组的初步工作中，我们可以通过在TIR暗场模式下旋转相干散射（ROCS）激光来实现150 nm的空间分辨率。有了这项技术，我们可以获得数千个图像，而不会损失图像质量（如荧光显微镜中的荧光团漂白）和图像重建（如需要，例如在超分辨率显微镜与结构照明）。在本研究的建议，我们希望达到两个新的目标，除了预期的120 nm的空间和100 Hz的时间分辨率的这项技术。一方面，我们希望通过散射激光的特定吸收和相位延迟来区分图像中的特定标记结构。另一方面，我们将开发一种新的时间相关显微镜应开发-在很大程度上独立的第一个目标。这里，具有高时间分辨率的无标记结构的相干ROCS图像将与具有低时间分辨率的荧光团分布的基于荧光的图像相关。