随着科学的进步，生命科学的研究对象由单个器官向组织体、离体组织切片及发育过程中的活体胚胎转变。荧光特异性标记的出现，为追踪物质在单细胞、组织体、器官甚至整个胚胎内的转移过程提供了手段。为了实现整个追踪过程，需要对活体胚胎进行无损、非侵入式的亚细胞级别成像，这就对荧光显微技术提出了更高的要求。在传统荧光显微技术基础上发展了光片照明和超分辨荧光显微技术。前者通过选择平面照明方式，只激发探测物镜焦平面附近的样品，因其具有高穿透深度、低漂白和高成像速度而广泛应用于三维活体组织成像；后者利用特殊的光调控手段将显微镜的分辨率提升至纳米水平，成为研究亚细胞水平生命活动的有力**。通过介绍2大技术的发展、融合以及目前所遇到的问题，探究新型的、适宜观察三维厚组织样品亚细胞结构和生命过程的成像方法。

With the progress of science, the research objects of life science change from single organ to tissue, in vitro tissue slice, and live embryonic during development. The appearance of fluorescent-specific markers provides a means for tracking the process of the substance transfer within a single cell, tissue, organ, and even the entire embryo. In order to track the whole process, it is necessary to make nondestructive, non-invasive imaging of living embryos with sub-cellular level resolution, which poses higher requirements for fluorescence microscopy. We develop light-sheet fluorescence microscope and super-resolution fluorescence microscopy on the basis of traditional fluorescence microscopy. Light-sheet fluorescence microscope only excites the sample near the objective′s focal plane. Because of its high penetration depth, low photo-toxicity and photobleaching, and high imaging speed, light-sheet fluorescence microscope can be widely used in three-dimensional in vivo large scale biological imaging. Super-resolution fluorescence microscopy uses special light control means to improve the resolution of the microscope to nanometer level, which becomes a powerful weapon for exploring sub-cellular life activities. We introduce the development, integration, and the current problems encountered in the two technologies, and try to explore an imaging technique suitable for observing the subcellular structure and life process of three-dimensional large scale′s samples.