A Dragonfly multimodal fast imaging platform with SRRF-stream (Super-Resolution Radial Fluctuation) in the Liverpool Centre for Cell Imaging (CCI)

植物或动物的单个细胞暴露于环境的变化中（生化信号、温度、机械力、光线变化……）。细胞必须解释这些信息以适应并做出适当的反应。然而，生物体中的细胞并不是孤立地起作用的，而是一个复杂的三维环境的一部分。因此，通过实验重建这样的环境，并在完整器官或3D培养物中可视化单个细胞的能力，对于研究生物过程至关重要。随着时间的推移，成像技术正在迅速发展，以研究多细胞环境中的单个细胞。对从纳米（阐明细胞内部发生了什么）到毫米（阐明细胞在组织中是如何组织的）的细胞过程可视化的需求，以最小的侵入方式和实时的方式，引发了新的显微镜和成像技术的发展。我们建议购买一种显微镜（蜻蜓，由Andor商业化），它可以：1)实时快速成像生物过程，2)高质量的3D样品成像，固定或生活，不需要特定的样品制备，3)成像在非常小的尺度，超过传统显微镜的光学分辨率，不损坏样品，因为高水平的光4)成像在近红外光谱的光，这是更常用的在体内成像。这将使在小动物模型中使用相同的荧光探针从细胞到组织的测量整合起来。5)生理条件下成像（控制温度、湿度、CO2、O2）。蜻蜓是一种多功能显微镜，由于它有3种不同的照明模式，是多用户设施的理想仪器。我们将把它安装在利物浦细胞成像中心（CCI），这是一个拥有来自学术界和工业界约100名注册用户的开放访问和共享设施。该显微镜将服务于BBSRC职权范围内的广泛科学研究。我们简要介绍以下两个将受益于它的研究课题：1。更好地了解细菌和藻类的光合作用，以进一步改造作物并提高其生产力光合作用是一个必不可少的生物过程。在光合作用过程中，蓝藻、藻类和高等植物等光养生物将太阳光转化为化学能，并产生动物生命所必需的氧气。我们需要更好地了解光合作用，以推动未来的作物工程以提高产量。这是通过阐明细菌和藻类中非常有效的光合作用机制的组织来实现的。利物浦和约克的研究小组已经开发出分子工具，可以在活细菌或藻类中观察到它。他们现在需要能够非常精确地（在纳米范围内）成像这些组件，并测量它们的快速运动。他们将使用“蜻蜓”对必要组件进行快速、高通量和高分辨率成像。氧感蛋白在细胞分裂中的意外作用氧对多细胞生物的生命至关重要，动物已经进化出了应对氧浓度降低的机制。有趣的是，最近发现的氧感应蛋白也能调节细胞分裂过程中的基本过程。连接氧感应酶和细胞分裂的分子机制需要进一步研究，使用人体组织模型。由于细胞分裂过程快速且发生在细胞的特定区域，因此需要能够高速成像和高分辨率的显微镜。此外，本研究在体外重建的假人体组织是脆弱的，需要温和的成像条件，低光水平，以避免光引起的损伤。蜻蜓将为这项研究提供所需的成像条件，并将使发现控制人体组织细胞分裂的重要机制成为可能。

Individual cells in a plant or an animal are exposed to changes in their environment (biochemical signals, temperature, mechanical forces, light variations...). Cells have to interpret this information to adapt and respond appropriately. However, cells in an organism do not function in isolation, but are part of a complex 3D environment. The ability to experimentally recreate such environments and visualise individual cells in intact organs or 3D cultures is therefore essential to study biological processes. Imaging technologies are developing fast to study individual cells in a multicellular environment over-time. The need to visualise cellular processes from the nanometre (to elucidate what happens inside cells) to the millimetre (to elucidate how cells are organised in a tissue) scale in the least invasive manner and in real time has triggered the development of new microscopes and imaging technologies. We propose to purchase a microscope (Dragonfly, commercialised by Andor) which allows:1) Fast imaging of the biological processes in real-time, 2) High quality imaging of 3D samples, fixed or living, without the need of specific sample preparation,3) Imaging at very small scales, beyond the optical resolution of traditional microscopes, without damaging the sample due to high levels of light4) Imaging in the near-infrared spectrum of light, which is more commonly used for in vivo imaging. This will enable to integrate measurements done with the same fluorescent probes from cells to tissues in small animal models.5) Imaging in physiological conditions (controlled temperature, humidity, CO2, O2).Dragonfly is a versatile microscope, thanks to its 3 different modes of illumination and is an ideal instrument for a multi-user facility. We will install it in the Liverpool Centre for Cell Imaging (CCI), an open access and shared facility with ~100 registered users from academia and industry. The microscope will serve a breadth of science across the BBSRC remit. We briefly present below two research topics, which will benefit from it:1. A better understanding of photosynthesis in bacteria and algae to further engineer crops and boost their productivityPhotosynthesis is an essential biological process. During photosynthesis, phototrophs such as cyanobacteria, algae and higher plants convert solar light into chemical energy and generate oxygen necessary for animal life. A better understanding of photosynthesis is required to drive the future engineering of crop plants to increase yields. This is achieved by the elucidation of the organisation of the very efficient photosynthetic machinery in bacteria and in algae. Groups in Liverpool and York have developed molecular tools to visualise it in living bacteria or algae. They now need to be able to image the components very precisely (at nanometre range) and to measure their fast movements. They will use the Dragonfly for fast, high-throughput and high-resolution imaging of the necessary components.2. The unexpected role of oxygen sensing proteins in cell divisionOxygen is essential for life in multicellular organisms and animals have evolved mechanisms to cope with decreased oxygen concentration. Interestingly, oxygen-sensing proteins have recently been discovered, to also regulate essential processes during cell division. The molecular mechanisms that connect oxygen-sensing enzymes and cell division needs to be further investigated, using models of human tissues. Because cell division processes are fast and occur in defined areas of the cells, a microscope allowing high speed imaging and high resolution is required. Moreover, the pseudo-human tissues recreated in vitro for this study are fragile and require the gentle imaging conditions, with low light levels, to avoid light-induced damages. The Dragonfly will provide the required imaging conditions for this study and will enable the discovery of important mechanisms that control cell division in human tissues.