Learning and developing scanning electron microscopy (SEM) techniques for biological research

Abstract

Electron microscopy (EM) is a sophisticated imaging technique invented in the 1930s that can capture images of extremely small structures. Instead of using light as in standard microscopy, EM uses a beam of electrons. Electrons have a shorter wavelength than photons and therefore can report smaller structures than light can. Dr Daisuke Koga of Asahikawa Medical University, Japan, is a world-leading expert in electron microscopy. He explains that electron microscopes can now focus on examining everything from relatively large structures such as tissues and whole cells down to individual protein structures. This makes the technique extraordinarily flexible and therefore useful to a wide range of biological fields, and experts in the technique are continually developing novel methods to examine an even wider range of biological features. Through his extensive research efforts, Koga has developed a novel EM technique capable of constructing 3D images of the Golgi apparatus. This organelle has attracted a lot of morphologists for a long time because of its mysterious and beautiful structure. The morphological studies of the Golgi have been mainly performed by TEM. Although TEM of a single ultrathin section provides the ultrastructure of the Golgi, the 3D shape of this organelle remains uncleared. "The Golgi apparatus is responsible for protein modification and trafficking," he explains. The Golgi takes freshly synthesised proteins from the endoplasmic reticulum and adds additional features such as phosphoryl groups and sugars that confer on additional functions on the protein." He says that it also packages proteins that need to be secreted out or even for storage. These functions are diverse and extremely important for normal cell operations. Koga has now established serial section scanning electron microscopy (SEM) that has the power to accurately reconstruct 3D images of various subcellular structures. His success arose from improvements in signal detection systems of SEM, which allowed for the imaging of ultrastructure information from tissue sections embedded in resins. He has combined these instrumental improvements with a unique sample preparation technique. By slicing hundreds of extremely thin sections of samples embedded in resin and imaging the regions of interest, the 3D structure of subcellular components can be revealed by 3D reconstruction of consecutive SEM images. "I have revealed the entire 3D shape of the Golgi apparatus in different cells such as the epithelial principal cells in epididymis, pancreatic acinar cells, parotid acinar cells and cerebellar Purkinje cells using the serial section SEM and 3D reconstruction method," outlines Koga. He has printed 3D printed models of the Golgi apparatus to clarify the 3D configuration of this organelle. Now, he is trying to reveal the 3D morphological diversity of the Golgi in the anterior pituitary tissue which comprises several kinds of endocrine cells (somatotrope, mammotrope, gonadotrope, thyrotrope and corticotrope).