EM facility of the institute of neuroscience (previously the brain research institute) was established in 1980. At present, the facility is equipped with three precision electron microscope instruments, including a 80 kV TEM (Jeol, JEM-1230), a LaB6 120 kV TEM (Thermo Scientific, Talos L120C)) and a field-emission scanning electron microscopy (Zeiss,Gemin300). Around these EM devices, the facility has a complete sample preparation system for the study of neurobiology ultrastructure, including Cryo-Transfer Holder (Gatan, Elsa 698), automated tape-collecting ultramicrotome (ATUM, RMC), high-pressure freezer-freeze substitution system (Leica EM HPM100, Leica EM AFS2 & FSP), ultra-thin microtome (Leica UC7/UC6), low temperature sectioning system (Leica EM FC6),versatile sputter coater/turbo evaporator(Quorum, Q150T), Vitrobot glow discharge cleaning system (PELCO easiGlowTM 91000) and some related histology equipments. With all these setups, the facility has been exploring applications of EM imaging techniques inneuronal systems for all researchers to elucidatethe relationship between ultrastructure and function.
In recent years, the technology team has focused on high quality sample preparation and observation solutions:
(i) IEM (immune-electron microscope) technique is used to localize molecules at the ultrastructural level by labeling them with specific antibodies, which are visualized by electron-opaque markers (colloidal gold particles orenzyme reaction) attached to them. The effect is to produce an electron-dense label at the site of the antigen-antibody reaction.
(ii) ATUM-SEM (Automatic tape collecting ultra microtome-SEM) technique is a type of serial-section electron microscopy, and usually used in analyzing the connectivity in volumetric samples of brain tissue. The technology is collecting ultrathin sections (30-70 nm) serially on a kapton tape by AUTOME, and then imaging these sections in sequence by SEM to obtain a large number of sequence images and form a “Ultrathin section libraries”. These images of serial sections are automatic recognition, alignment, and finally reconstruct three-dimensional reconstruction at nanometer resolutions. Imaging and tissue handling of the ATUM-SEM is more complicated technology.
(iii) HPF-FS (high pressure freezing-freezing substitution) technique in ultrastructural study of nervous tissue is one of the methods of cryofixation. It is fast freezing the living tissues or cells to 77 K, under hydrostatic pressure of 2100 bars. Compared with conventional chemical fixation methods, this method has the following advantages: 1.has faster fixed speed, can capture the instantaneous physiological state; 2. can obtain more native structure; 3. has higher resolution. After high pressure freezing, samples usually need pass the process of freeze-substitution.
The facility is open 9 hours per day and 5 days per week for services on the electron microscopes and 14 hours per day and 6 days per week for use of the cryostats. We provide service to investigators of CEBSIT as well as other institutions on an hourly-fee basis or a fee-for-service basis. Outside users from surrounding universities and local biotechnology industries are accommodated schedule permitting.
