A super-resolution microscopy platform for imaging cells at multiple spatial scales

荧光成像使研究人员能够探索细胞内结构的定位、动态和可塑性。虽然在当时是革命性的，但经典荧光成像显微镜的分辨率受到光的衍射极限的限制。这意味着细胞中分子的精确位置和组织不能被精确地可视化。相反，只有对它们的位置的模糊描述是可能的，没有任何可能揭示单个分子如何在空间中排列。超分辨率显微镜的出现改变了这一切——现在有可能利用荧光成像来解决单个分子的位置，并开始探索发生在纳米尺度上的分子排列的全新世界。事实上，这些新方法有助于揭示细胞内新的亚细胞结构，这将有助于更好地理解它们的功能。为了深入研究这个极小的新世界，重要的是要有合适的设备来进行单分子成像。特别是，它需要能够对标记有不同荧光化合物的分子这样做，以揭示多分子结构，在3D和大视场中具有高精度，以包含尽可能多的信息。最后，重要的是要有机会获得成像工具，提供低分辨率的感兴趣的分子图。通过这种方式，我们的目标是提供细胞和神经元中分子分布的大规模描述，并随后进入其中一些域，以非常高的空间分辨率表征它们。在这里，我们建议使用这种方法来回答不同系统（包括完整组织切片）中的基本细胞生物学问题，以建立大脑中神经元形成的突触的特性，并了解体外培养细胞中细胞骨架的局部结构。

Fluorescence imaging has allowed researchers to explore the localisation, dynamics and plasticity of structures within cells. Although revolutionary at the time, the resolution of classical fluorescence imaging microscopes is constrained by the diffraction limit of light. This means that the precise location and organisation of molecules in a cell cannot be accurately visualised. Instead, only a blurred description of their whereabouts was possible, without any possibility of uncovering how single molecules were arranged in space. The advent super-resolution microscopy has changed all this - it is now possible to resolve the location of individual molecules using fluorescence imaging and begin to explore a whole new world of molecular arrangements that takes place at the nanoscale. In fact, these new approaches have been instrumental in uncovering novel sub-cellular structures within cells, which will help better understand their function. To delve into this new world of the very small, it is important to have the right equipment that allows single-molecule imaging. In particular, it needs to be able to do so for molecules tagged with different fluorescent compounds to uncover multi-molecular structures, with high precision in 3D and over large fields of view to encompass as much information as possible. Finally, it is important to also have access to imaging tools that give low resolution maps of the molecules of interest. In this way, we aim to provide large-scale descriptions of the distribution of molecules in cells and neurons and subsequently home into some of these domains to characterise them at very high spatial resolution. Here, we propose to use this approach to answer basic cell biology questions in different systems, including in intact tissue slices, to establish the properties of synapses formed by neurons in the brain and understand the local structure of the cytoskeleton in cells grown in vitro.