Our knowledge about the organisation of cells, biological specimen structures, composition and properties in submicroscopic detail of biological samples is associated to a large extent to transmission electron microscopy (TEM). The Electron Microscopy Core (EM Core) Facility provides advice, technical services, training, equipment and facilities to NHLBI intramural scientists if their research requires electron microscopy (EM) to clarify questions. Using an electron microscope offers the advantage of increasing both the magnification of an object and the resolution over other imaging tools. These could be issues involving subcellular, supramolecular or macromolecular structure at a level of resolution below that obtained by a light microscope. The NHLBI EM Core Facility has supported projects using the following techniques in the past year: 1. Chemical fixation, embedding, ultra-thin sectioning and transmission EM digital imaging of tissues and cell culture. 2. EM immunocytochemistry, including immunogold, nanogold with silver enhancement and immunoperoxidase localisation of proteins and other antigens within and on the surface of tissues and cells by pre-embedding techniques. 3. Negative staining of large proteins, polymers and supramolecular structures as well as lipid and membrane vesicles for transmission EM digital imaging. 4. Rotary shadowing of large protein molecules, DNA and other macromolecules. 5. Preparation of platinum replicas of cytoskeletons, partially lysed cells and freeze-fractured/freeze- dried tissues and cells. 6. Chemical fixation, critical point drying, sputter-coating and scanning EM digital imaging of small organisms, organs, tissues and cells, as well as other materials such as artificial matrices. 7. Thawed cryosection immuno labelling technique (also called Tokuyasu method). 8. Immuno correlative light and electron microscopy (CLEM) on Tokuyasu cryosections. 9. Cryo-electron microscopy of vitreous sections (CEMOVIS) to observe biological samples in their most native, fully hydrated state. 10. High-pressure freezing (HPF) and freeze substitution (FS) allows improved morphological preservation compared to the conventional preparation. These techniques are available as a service to be performed by the EM Core Staff for the customer or they are available to be learned by users who will be trained to be independent users of the EM Core. In the past year, the NHLBI EM Core supported a total of 81 recorded projects for 44 investigators within the NIH. 54.5% of the services were requested by NHLBI research groups. The remaining 45.5% are divided as follows: CC (2.3%), NCATS (2.3%), NCI (9.1%), NEI (2.3%), NHGRI (2.3%), NIAMS (2.3%), NIDCR (3%), NIDDK (9.1%), NINDS (4.5%), NINR (2.3%) and OD (2.3%). We have also trained several postdoctoral fellows, summer students, contractors, staff scientists and investigators in the use of the scanning and transmission electron microscopes and sample preparation techniques.
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