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000276116 0247_ $$2doi$$a10.6019/s-biad1274
000276116 0247_ $$2doi$$a10.6019/S-BIAD1274
000276116 0247_ $$2doi$$a10.6019/s-biad1273
000276116 037__ $$aDZNE-2025-00197
000276116 1001_ $$0P:(DE-2719)2812260$$aSchifferer, Martina$$b0$$udzne
000276116 245__ $$aDataset: ATUM-Tomo: A multi-scale approach to cellular ultrastructure by combined volume scanning electron microscopy and electron tomography
000276116 260__ $$bBioImage Archive$$c2024
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000276116 500__ $$aThis work was supported by the DFG under Germany’s Excellence Strategy within the framework of the Munich Cluster for Systems Neurology (SyNergy; EXC 2145 – ID 390857198), TRR 274/1 2020 (projects Z01 and B03 – ID 408885537) and FOR Immunostroke (Mi 694/9-1 A03 – ID 428663564). Research in the Müller-Reichert lab was funded by the DFG (grant MU 1423/8-2 and 8-3 to TMR). All animal experiments were supported by DFG grant 457586042.
000276116 520__ $$aLike other volume electron microscopy approaches, Automated Tape Collecting Ultramicrotomy (ATUM) enables imaging of serial sections deposited on thick plastic tapes by scanning electron microscopy (SEM). ATUM is unique in enabling hierarchical imaging and thus efficient screening for target structures, as needed for correlative light and electron microscopy. However, SEM of sections on tape can only access the section surface, thereby limiting the axial resolution to the typical size of cellular vesicles with an order of magnitude lower than the acquired xy resolution. In contrast, serial-section electron tomography (ET), a transmission electron microscopy-based approach, yields isotropic voxels at full EM resolution, but requires deposition of sections on electron-stable thin and fragile films, thus making screening of large section libraries difficult and prone to section loss. To combine the strength of both approaches, we developed ‘ATUM-Tomo’, a hybrid method, where sections are first reversibly attached to plastic tape via a dissolvable coating, and after screening detached and transferred to the ET-compatible thin films. ATUM-SEM of serial semi-thick sections and consecutive ET of selected re-mounted sections combines SEM’s fast target recognition and coarse rendering capability with high-resolution volume visualizations, respectively, thus enabling multi-scale interrogation of cellular ultrastructure. As a proof-of-principle, we applied correlative ATUM-Tomo to study ultrastructural features of blood brain barrier (BBB) leakiness around microthrombi in a mouse model of traumatic brain injury. Microthrombi and associated sites of BBB leakiness were identified by confocal imaging of injected fluorescent and electron-dense nanoparticles, then relocalized by ATUM-SEM, and finally interrogated by correlative ATUM-Tomo. This workflow is a seamless zoom-in on structural BBB pathology from the micro- to the nanometer scale. Overall, our new ATUM-Tomo approach will substantially advance ultrastructural analysis of biological phenomena that require cell- and tissue-level contextualization of the finest subcellular textures.
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000276116 536__ $$0G:(GEPRIS)390857198$$aDFG project G:(GEPRIS)390857198 - EXC 2145: Munich Cluster for Systems Neurology (SyNergy) (390857198)$$c390857198$$x1
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000276116 7001_ $$0P:(DE-2719)9001702$$aRodriguez Martinez, Lucia$$b1$$udzne
000276116 7001_ $$0P:(DE-2719)9000614$$aKislinger, Georg$$b2$$udzne
000276116 7001_ $$0P:(DE-HGF)0$$aFabig, Gunar$$b3
000276116 7001_ $$0P:(DE-HGF)0$$aWehn, Antonia$$b4
000276116 7001_ $$0P:(DE-2719)9001710$$aJiang, Hanyi$$b5$$udzne
000276116 7001_ $$0P:(DE-2719)9001883$$aNiemann, Cornelia$$b6$$udzne
000276116 7001_ $$0P:(DE-HGF)0$$aKlymchenko, Andrey S.$$b7
000276116 7001_ $$0P:(DE-2719)9000853$$aPlesnila, Nikolaus$$b8
000276116 7001_ $$0P:(DE-2719)2810727$$aMisgeld, Thomas$$b9$$udzne
000276116 7001_ $$0P:(DE-HGF)0$$aMüller-Reichert, Thomas$$b10
000276116 7001_ $$0P:(DE-HGF)0$$aKhalin, Igor$$b11
000276116 773__ $$a10.6019/s-biad1274
000276116 7870_ $$0DZNE-2024-00953$$aKislinger, Georg et.al.$$dCambridge : eLife Sciences Publications, 2024$$iRelatedTo$$tCombining array tomography with electron tomography provides insights into leakiness of the blood-brain barrier in mouse cortex.
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