sm43 =e 77, (1 ASEOL, Application Note Scanning Microscope ES Specimen Preparation Methods for Scanning Electron Microscopes _JEOL |I. Introduction Il, Specimen Preparation Methods for Scanning Electron Microscopes IL-1, Chemical Specimen Preparation Method 11-2. Details of Chemical Preparation Method 112-1, Cut off a slice and cleaning of biological specimen 1-2-2. Fixation 1-2-3. Wash after fixation 1-2-4. Dehydration 1-2-5. Drying IL-3. How to Prepare Chemicals for Chemical Specimen Preparation Method Preparation of buffer solution How to prepare fixatives How to prepare dehydrating agents 14, Examples of Specimen Preparation 14-1, Plant leaves 1-4-2, Soft tissue such as small intestines 11-4-3, Red blood cells (free cells) 1-4-4. Yeast (microorganism) 114-5. General conductive staining pH and Osmotic Pressure of Fixative IL-6, Physical Specimen Preparation Method I6-1 Preeze-drying method 1-6-2. Freezing method (Cryo-SEM method) 1-7. Combination of Chemical and Physical Preparation Methods I1-7-1, Alcohol fracturing method II, Conclusion 20I. Introduction In the observation of biological specimens using an electron micros ‘ope, it is necessary that specimens be kept in a state as close to their living state as possible. However, 70 to 80% of the components of al specimen is water. Therefore, if a specimen is inserted into an el tron microscope as it is after cut off a slice, the water in it will evaporate, often causing the specimen to be contracted. In addition, unless a biological specimen is treated immediately after cut off a slice, it cannot maintain a state close to its living state, as a result of decomposition. In this connection, to observe a biological specimen with an electron microscope, the following method is normally used Firstly, the protein and lipid that constitute a specimen are chemically or physically fixed to keep them in a state as close to their living state as possible. Secondly, it is further dehydrated and dried, before being inserted into the electron microscope. The method of preparing biological specimens, originally developed for the transmission electron microscope, has now been almost established. ‘Therefore, specimens for scan ng elect on microscopy are often prepared on the basis of that method. For transmission electron microscopy, specimens are prepared into ultrathin sections to observe transmission images, whereas for scanning electron microscopy, specimens in a bulk state are used to observe three-dimensional images. Although there is some difference in specimen preparation between these two observation techniques, it has been proved that the specimen preparation method originally developed for transmission electron microscopy is also bringing about an almost satisfactory result for scanning electron microscopy. Described hereunder are the general procedures for chemical preparation needed to observe biological specimens using the scanning electron microscope. We therefore ommend that you consult individual reference books for the specific aspects of specimen preparation, which differ among research fields.