Cytological analyses by advanced electron microscopy

Abstract

Recently several microscopy techniques have revealed the dynamics and structure of intracellular material such as organelles and supramolecular proteins for a better understanding of the fundamental properties of a cell. Electron microscopy provides information on extremely small biological specimens, providing images of several target molecules and organelles, including mitochondria and plastids, and their surrounding intracellular environments. Technical development using several advanced electron microscopes has been advanced to obtain 3D structural modeling. Among them, cryo-TEM and tomography techniques are useful for visualization of fine structures, but the observation area is limited. Conversely, "FIB-SEM" theoretically is not affected by the thickness of the sample, and FIB-SEM and reconstruction are effective for 3D structural modeling of C. merolae cells (2-5 microns) during cell division at the whole cell level. Another technique, UHVEM tomography, visualizes a detailed 3D structure of the light-harvesting complex on the surface of the thylakoid membrane in the plastid and the micro-compartments (cristae) in mitochondria. Furthermore, fine whole cell 3D structural model using cryo-STEM tomography, which is obtained under physical fixation and without staining, instead of chemical fixation, is compared with the models of FIB-SEM and 3D reconstruction.