A digital light-sheet microscope for the analysis of biological processes under almost natural conditions

多年来，生物成像一直在向使用越来越多的生理相关系统的实验迈进，新技术正在使这种实验方法逐渐变得可行。这样的系统，包括整个模型生物体、组织外植体和三维(3D)细胞培养，必须以一种最大限度地减少扰动的方式进行成像，以保持其作为实验模型的生理完整性。长期以来，光诱导的光损伤和光毒性在生物成像领域一直是一个具有挑战性的问题，并且可以在所有生物组织水平上对健康和功能产生相当大的影响。因此，对如此大而敏感的样品进行成像需要一种有效的3D成像方法，以减少样品在光下的暴露。光片显微镜代表一种平面照明技术，它彻底改变了生物样品的光学成像方式。基本上，通过分离照明和检测光路，使得光片技术成为可能，从而允许优化光曝光和信号收集效率的新颖照明策略。使用光片显微镜，较低的照明强度可以与平面照明相结合，以最小的样本光曝光提供更好的信号，从而实现高帧速率和长时间的成像。已经从这种成像方式中受益颇多的领域包括发育生物学、神经生物学、药物发现、植物生物学等。然而，定制的薄片显微镜可能非常难以使用，通常需要两个或更多目标、非标准的样品制备方案、困难的对准程序以及处理由这些技术产生的大量数据的精心规划的工作流程。我们的目标是购买一种商用光片显微镜，这是目前可用的最简单、最直观和最可靠的系统，与简单的数据处理和分析平台相结合，可以使研究人员能够在几乎自然的条件下研究生物过程。

Biological imaging has been moving consistently towards experimentation employing increasingly physiologically relevant systems for many years, and new technologies are making such experimental approaches progressively feasible. Such systems, comprising whole model organisms, tissue explants, and three-dimensional (3D) cell cultures, must be imaged in a way that minimises perturbations in order to maintain their physiological integrity as experimental models. Light-induced photodamage and phototoxicity have long been challenging in the field of biological imaging and can have quite a dramatic effect on health and function at all biological levels of organisation. Imaging such large and sensitive specimens thus demands an efficient approach to 3D imaging that lessens exposure of the sample to light.The light-sheet microscope stands for a planar illumination technique that has revolutionised how optical imaging of biological specimens can be performed. Basically, the light-sheet technique is made possible by decoupling the illumination and detection optical pathways, allowing for novel illumination strategies that optimise the light exposure and signal collection efficiency. With light-sheet microscopy, lower illumination intensities can be used in combination with planar illumination to provide improved signal with minimal sample light exposure, enabling imaging with high frame rates and over long periods of time. Fields that have already benefited considerably from such an imaging modality include developmental biology, neurobiology, drug discovery, plant biology, and beyond.However, custom-made light-sheet microscopes can be very difficult to use, usually requiring two or more objectives, non-standard sample preparation protocols, difficult alignment procedures, and a well-planned workflow for dealing with the massive amounts of data generated by such techniques. Our aim is to purchase a commercial light-sheet microscope that is the simplest, most intuitive and most reliable system available that in combination with a straightforward data handling and analysis platform can empower researchers to investigate biological processes under almost natural conditions.