Department of Biological Sciences, Columbia University, New York, NY, USA.
Department of Neuroscience, Karolinska Institutet, Stockholm,, Sweden.
BMC Biol. 2018 May 29;16(1):57. doi: 10.1186/s12915-018-0521-8.
Advances in tissue clearing and molecular labeling methods are enabling unprecedented optical access to large intact biological systems. These developments fuel the need for high-speed microscopy approaches to image large samples quantitatively and at high resolution. While light sheet microscopy (LSM), with its high planar imaging speed and low photo-bleaching, can be effective, scaling up to larger imaging volumes has been hindered by the use of orthogonal light sheet illumination.
To address this fundamental limitation, we have developed light sheet theta microscopy (LSTM), which uniformly illuminates samples from the same side as the detection objective, thereby eliminating limits on lateral dimensions without sacrificing the imaging resolution, depth, and speed. We present a detailed characterization of LSTM, and demonstrate its complementary advantages over LSM for rapid high-resolution quantitative imaging of large intact samples with high uniform quality.
The reported LSTM approach is a significant step for the rapid high-resolution quantitative mapping of the structure and function of very large biological systems, such as a clarified thick coronal slab of human brain and uniformly expanded tissues, and also for rapid volumetric calcium imaging of highly motile animals, such as Hydra, undergoing non-isomorphic body shape changes.
组织透明化和分子标记方法的进步使得对大型完整生物系统进行前所未有的光学访问成为可能。这些发展激发了对高速显微镜方法的需求，以定量和高分辨率对大型样本进行成像。虽然光片显微镜 (LSM) 具有高平面成像速度和低光漂白的特点，但由于使用正交光片照明，其扩展到更大的成像体积受到了限制。
为了解决这个基本限制，我们开发了光片 theta 显微镜 (LSTM)，它从与检测物镜相同的一侧均匀地照亮样品，从而在不牺牲成像分辨率、深度和速度的情况下消除了对横向尺寸的限制。我们对 LSTM 进行了详细的表征，并展示了它相对于 LSM 的互补优势，可用于快速高分辨率定量成像大型完整样品，具有高均匀质量。
所报道的 LSTM 方法是快速高分辨率定量映射大型生物系统结构和功能的重要步骤，例如人类大脑的厚冠状切片和均匀扩展的组织，以及快速对高度运动的动物（如 Hydra）进行体积钙成像，这些动物正在经历非同形的身体形状变化。
