1)State Key Laboratory of Primate Biomedical Research, Institute of Primate Translational Medicine, Kunming University of Science and Technology, Kunming650500, China;2)Yunnan Key Laboratory of Primate Biomedical Research, Kunming 650500, China;3)Faculty of Life Science and Technology, Kunming University of Science and Technology, Kunming 650500, China
This work was supported by grants from The Natural Science Foundation of Yunnan Province (202001BC070001, 202102AA100053).
Expansion microscopy (ExM) is a new super-resolution imaging technique. With the aid of expandable hydrogel, biological samples are uniformly physically amplified and can be imaged in super resolution by using conventional optical imaging microscopes. In ExM, after immunofluorescence staining, gel embedding, protease digestion and water swelling, the relative distance of fluorescent labeled molecules inside the biological samples was increased, so the sample can bypass the optical diffraction limit in conventional fluorescence microscope to achieve the super-resolution imaging. ExM is widely suitable for many types of biological samples such as cell and tissue sections. Proteins, nucleic acids, lipids and other biological macromolecules can also be imaged by ExM. ExM can be combined with confocal microscopy, light-sheet microscopy and super-resolution microscopy to further improve imaging resolution. In recent years, a variety of derivative technologies have been developed from base ExM, which further promotes the practical application of this technology. Protein retention expansion microscopy (proExM) can avoid complicated sample preparation process and directly image endogenous fluorescent proteins. Magnified analysis of the proteome (MAP) was suitable for super-resolution imaging in large biological samples. Iterative expansion microscopy (iExM) can increase the final expansion factor of biological samples to 16-22 times by changing the gel embedding steps. Cryo-expansion microscopy (Cryo-ExM) can provide better image fidelity. Expansion fluorescent in situ hybridization (ExFISH) and Click-ExM can achieve super-resolution imaging in nonprotein biomolecules, such as RNA, lipids, and polysaccharides. Expansion pathology (ExPath) can be used for clinicopathologic specimens imaging. The combination of ExM and light-sheet microscope can improve the image resolution to super-resolution level in the deep imaging depth. The application of ExM in super-resolution microscopy can further increase the resolution of images to 10-30 nm. In this paper, we reviewed the basic principles of ExM and its derivative technology, the research progress of combining ExM with different imaging technologies, the application progress of ExM in observing different types of biological samples, and the prospective of spreading ExM technology in the future.
YANG Zhen-Yu, GUAN Miao, SUN Zheng-Long. The Basic Principles and Application of Expansion Microscopy[J]. Progress in Biochemistry and Biophysics,2023,50(3):505-512
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