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Max-Planck-Gesellschaft
Max-Planck-Institut für Experimentelle Medizin
Forschung
Electron Microscopy Core Unit



The Electron Microscopy Core Unit is performing ultrastructural research in collaboration with scientists within the Max-Planck-Institute of Experimental Medicine and the Göttingen research campus and since 2012 also within the Cluster of Excellence and DFG Research Center Nanoscale Microscopy and Molecular Physiology of the Brain.
Since 2004 the Unit is headed by Dr. Wiebke Möbius. Current members of the team are the two technicians Torben Ruhwedel and Boguslawa Sadowski, the Postdoc Dr. Anna Steyer, the PhD student Martin Meschkat and the IT specialist Christos Nardis.

Within the Electron Microscopy Core Unit, we routinely apply transmission electron microscopy (TEM) to analyse ultrastructure at subcellular resolution. Depending on the question under investigation, we use methods for morphological analysis on plastic embedded samples, or we perform immunoelectron microscopy on ultrathin cryosections to localize molecules within the context of the cell or tissue (Möbius, 2016). For ultrastructural analysis, samples are either fixed chemically and processed by conventional embedding or prepared by high-pressure-freezing (HPF) and embedded in resin after freeze substitution (FS). While conventional preparations are more suited for quantitative comparison of phenotypes, HPF-frozen samples often reveal fine structural details that are often lost in conventional preparations due to chemical fixation and embedding. This is especially relevant in ultrastructural myelin research (exemplified in Möbius et al., 2016). We continuously develop and modify protocols of sample preparation such as microwave-assisted embedding for specialized applications like large area or volume imaging using focussed ion beam scanning electron microscopy.

We are equipped with two transmission electron microscopes for routine applications: a Zeiss EM 900 and a LEO 912AB Omega. Since 2011 a Leica HPM100 high pressure freezing machine is available for fast cryo-immobilization of samples which are prone to fixation artefacts. For the possibility to visualize fast events such as synaptic vesicle fusion with a time resolution of milliseconds, the Department of Molecular Neurobiology (Prof. Nils Brose) acquired a Leica HPM100 with light stimulation and a Leica HPM ICE with the option of light- or electrical stimulation in the EM Core Unit. These instruments are operated by Dr. Ben Cooper and Dr. Cordelia Imig from the Department of Molecular Neurobiology.
In 2012 the EM Core Unit joined the Cluster of Excellence and DFG Research Center Nanoscale Microscopy and Molecular Physiology of the Brain (CNMPB) as EM Platform. With the support of the CNMPB in 2014 a Zeiss Crossbeam 540 was installed. This instrument is a focussed ion beam scanning electron microscope (FIB-SEM) in which the ion beam is used for in-situ removal of material by “milling”. With this approach a defined volume of the sample becomes accessible for imaging and 3D reconstruction. The surface area that can be milled is limited, but the milling can be targeted to multiple areas in the same sample. Removal of material as thin as 5 nm is possible, thereby increasing the resolution in Z and creating data stacks with almost isotropic voxel size.
Using FIB milling as a micromachining tool, TEM lamellae of almost any material can be manufactured. Applying this under cryo-conditions (-150°C), vitrified frozen TEM lamellae can be prepared for cryo-TEM tomography of biological samples.

References:
Möbius W (2016) Immunoelectron Microscopy: High-Resolution Immunocytochemistry. In: Ralph A Bradshaw and Philip D Stahl (Editors-in-Chief), Encyclopedia of Cell Biology, Vol 2, Waltham, MA: Academic Press, 2016, pp. 32-43

Möbius W, Nave KA, Werner HB (2016) Electron microscopy of myelin: Structure
preservation by high-pressure freezing. Brain Res. 2016 Jun 15;1641(Pt A):92-100.
doi: 10.1016/j.brainres.2016.02.027. Review. PubMed PMID: 26920467.


Research


The main focus of this EM Core Unit is myelin, a membranous multi-layered structure which builds the so-called white matter in the brain. Myelin is made by oligodendrocytes in the central nervous system (CNS) and by Schwann cells in the peripheral nervous system (PNS). The myelin sheath is a key feature of long axons because it speeds up conduction velocity as much as 100 times that of non-myelinated axons due to saltatory action potential propagation. Apart from this, the myelin sheath has protecting and supporting functions for the myelinated axons.
By using mouse genetics we investigate the biology and function of the myelinating glia cells in the CNS and PNS. We are focusing on the mechanisms of myelin biogenesis, maintenance and turnover. By electron microscopy, one astonishing feature of myelin is the appearance as a solid and rather non-dynamic structure. Moreover, myelin proteins are known to turn over very slowly. To study mechanisms of myelin maintenance, we apply inducible knock-out strategies in the adult mouse and observe spatial and temporal changes in the myelin ultrastructure.
Due to the intimate relationship between the myelin sheath and the wrapped axon, also called the axo-glial unit, pathological changes in the myelinating glia cell affect axonal integrity. We study morphological changes induced by myelin mutants to better understand the functions of the intact myelin sheath as well as processes in demyelinating disease and ageing. Impairment of the myelin sheath and its consequences are studied in demyelinating disease models of multiple sclerosis and neuromyelitis optica (Weil et al., 2016 and 2017).
Apart from mouse models we are also interested in evolutionary aspects of myelin development. To address questions in myelin evolution we selected an agnathan model species, the marine lamprey Petromyzon marinus, which does not possess myelin and represents a putative ancestral stage in vertebrate evolution before the emergence of myelin. To test the hypothesis that evolutionary progenitors of myelinating cells are conserved in this recent species we assessed the axo-glial units in the lateral line nerve (representing the PNS) and the spinal cord in the larval and adult stage of the marine lamprey with transmission EM and volume imaging by FIB-SEM as well as fluorescent in-situ hybridization (Weil et al, 2018).

References:
Weil M-T, Möbius W, Winkler A, Ruhwedel T, Wrzos C, Romanelli E, Bennett JL, Enz L, Goebels N, Nave KA, Kerschensteiner M, Schaeren-Wiemers N, Stadelmann C, Simons M (2016) Loss of Myelin Basic Protein Function Triggers Myelin Breakdown in Models of Demyelinating Diseases. Cell Rep. Jul 12;16(2):314-22.

Weil, M-T, Ruhwedel, T, Möbius W, and Simons, M. (2017) Intracerebral injections and ultrastructural analysis of high-pressure frozen brain tissue. Curr. Protoc. Neurosci. 78:2.27.1-2.27.18. doi: 10.1002/cpns.22

Weil M-T, Heibeck S, Töpperwien M, tom Dieck S, Ruhwedel T, Salditt T, Rodicio MC, Morgan JR, Nave K-A, Möbius W and Werner HB. (2018) Axonal Ensheathment in the Nervous System of Lamprey: Implications for the Evolution of Myelinating Glia. Journal of Neuroscience 18 July 2018, 38 (29) 6586-6596


Göttingen Graduate School of Neurosciences and Molecular Biosciences (GGNB) and the International Max-Planck Research School (IMPRS) Neuroscience


Since 2009 we offer an advanced methods course: A044 “Subcellular localization of proteins by immunoelectron microscopy” applying immunolabeling of cryosections according to Tokuyasu for the undergraduate students of this school. Starting in 2018 we provide another course A228 “A practical introduction into electron microscopy” to provide insight into the basic methodology of biomedical electron microscopy.
Since 2015 we are involved in the International Max-Planck Research School (IMPRS) Neuroscience Study Program by providing a lecture (“Introduction into Electron Microscopy”), a practical EM-Course and by offering lab rotations.


Exchange of information within the EM community


“Submicron”-mailing list
Everybody working in the field of electron microscopy is often confronted with problems or questions which could be solved by discussion with experienced colleagues. For this purpose we launched an international mailing list called “submicron-list” for a quick exchange of information. This platform is also ideal to announce conferences, workshops or send out job offers. If you like to become a participant, please contact moebius@em.mpg.de or send a request to submicron-l@gwdg.de


DGE (German Society of Electron Microscopy) and the Workgroup PANOS


The workgroup PANOS (Präparation und Abbildung Nativer Organischer Systeme) of the German Society of Electron Microscopy (DGE, Deutsche Gesellschaft für Elektronenmikroskopie) is organized by members of the DGE working in the field of biomedical electron microscopy. We organize regular annual spring meetings to promote exchange and discuss interesting topics presented by researchers or students. This meeting is open for non-members of the DGE.
https://www.dge-homepage.de/
Information is exchanged by a mailing list: panos-list@gwdg.de



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