TY - CHAP
T1 - Neurons on tape
T2 - Automated Tape Collecting Ultramicrotomy-mediated volume EM for targeting neuropathology
AU - Kislinger, Georg
AU - Niemann, Cornelia
AU - Rodriguez, Lucia
AU - Jiang, Hanyi
AU - Fard, Maryam K.
AU - Snaidero, Nicolas
AU - Schumacher, Adrian Minh
AU - Kerschensteiner, Martin
AU - Misgeld, Thomas
AU - Schifferer, Martina
N1 - Publisher Copyright:
© 2023 Elsevier Inc.
PY - 2023/1
Y1 - 2023/1
N2 - In this chapter, we review Automated Tape Collecting Ultramicrotomy (ATUM), which, among other array tomography methods, substantially simplified large-scale volume electron microscopy (vEM) projects. vEM reveals biological structures at nanometer resolution in three dimensions and resolves ambiguities of two-dimensional representations. However, as the structures of interest—like disease hallmarks emerging from neuropathology—are often rare but the field of view is small, this can easily turn a vEM project into a needle in a haystack problem. One solution for this is correlated light and electron microscopy (CLEM), providing tissue context, dynamic and molecular features before switching to targeted vEM to hone in on the object's ultrastructure. This requires precise coordinate transfer between the two imaging modalities (e.g., by micro computed tomography), especially for block face vEM which relies on physical destruction of sections. With array tomography methods, serial ultrathin sections are collected into a tissue library, thus allowing storage of precious samples like human biopsies and enabling repetitive imaging at different resolution levels for an SEM-based search strategy. For this, ATUM has been developed to reliably collect serial ultrathin sections via a conveyor belt onto a plastic tape that is later mounted onto silicon wafers for serial scanning EM (SEM). The ATUM-SEM procedure is highly modular and can be divided into sample preparation, serial ultramicrotomy onto tape, mounting, serial image acquisition—after which the acquired image stacks can be used for analysis. Here, we describe the steps of this workflow and how ATUM-SEM enables targeting and high resolution imaging of specific structures. ATUM-SEM is widely applicable. To illustrate this, we exemplify the approach by reconstructions of focal pathology in an Alzheimer mouse model and CLEM of a specific cortical synapse.
AB - In this chapter, we review Automated Tape Collecting Ultramicrotomy (ATUM), which, among other array tomography methods, substantially simplified large-scale volume electron microscopy (vEM) projects. vEM reveals biological structures at nanometer resolution in three dimensions and resolves ambiguities of two-dimensional representations. However, as the structures of interest—like disease hallmarks emerging from neuropathology—are often rare but the field of view is small, this can easily turn a vEM project into a needle in a haystack problem. One solution for this is correlated light and electron microscopy (CLEM), providing tissue context, dynamic and molecular features before switching to targeted vEM to hone in on the object's ultrastructure. This requires precise coordinate transfer between the two imaging modalities (e.g., by micro computed tomography), especially for block face vEM which relies on physical destruction of sections. With array tomography methods, serial ultrathin sections are collected into a tissue library, thus allowing storage of precious samples like human biopsies and enabling repetitive imaging at different resolution levels for an SEM-based search strategy. For this, ATUM has been developed to reliably collect serial ultrathin sections via a conveyor belt onto a plastic tape that is later mounted onto silicon wafers for serial scanning EM (SEM). The ATUM-SEM procedure is highly modular and can be divided into sample preparation, serial ultramicrotomy onto tape, mounting, serial image acquisition—after which the acquired image stacks can be used for analysis. Here, we describe the steps of this workflow and how ATUM-SEM enables targeting and high resolution imaging of specific structures. ATUM-SEM is widely applicable. To illustrate this, we exemplify the approach by reconstructions of focal pathology in an Alzheimer mouse model and CLEM of a specific cortical synapse.
KW - ATUM
KW - Alzheimer ‘s disease
KW - Array tomography
KW - CLEM
KW - Correlation
KW - Neuroscience
KW - Plaques
KW - SEM
KW - Targeting
UR - http://www.scopus.com/inward/record.url?scp=85149833902&partnerID=8YFLogxK
U2 - 10.1016/bs.mcb.2023.01.012
DO - 10.1016/bs.mcb.2023.01.012
M3 - Chapter
C2 - 37451765
AN - SCOPUS:85149833902
SN - 9780323916073
T3 - Methods in Cell Biology
SP - 125
EP - 170
BT - Volume Electron Microscopy
A2 - Narayan, Kedar
A2 - Narayan, Kedar
A2 - Collinson, Lucy
A2 - Verkade, Paul
PB - Academic Press Inc.
ER -