Protocol for high-throughput electron tomography exemplified on centrioles in primary human plasma cells
Summary: Electron microscopy is the gold standard to characterize cellular ultrastructure. However, production of significant morphometrical data is highly limited by acquisition time. Here, we describe a semi-automated high-throughput strategy using single-axis serial section electron tomography to...
| Main Authors: | , , , , , |
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| Format: | Article |
| Language: | English |
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Elsevier
2023-09-01
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| Series: | STAR Protocols |
| Subjects: | |
| Online Access: | http://www.sciencedirect.com/science/article/pii/S2666166723003404 |
| _version_ | 1827917664287195136 |
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| author | Sebastian Köhrer Tobias Dittrich Martin Schorb Isabella Haberbosch Yannick Schwab Alwin Krämer |
| author_facet | Sebastian Köhrer Tobias Dittrich Martin Schorb Isabella Haberbosch Yannick Schwab Alwin Krämer |
| author_sort | Sebastian Köhrer |
| collection | DOAJ |
| description | Summary: Electron microscopy is the gold standard to characterize cellular ultrastructure. However, production of significant morphometrical data is highly limited by acquisition time. Here, we describe a semi-automated high-throughput strategy using single-axis serial section electron tomography to investigate and analyze centriole ultrastructure in bone-marrow-derived, primary human CD138pos plasma cells. The protocol comprises steps for electron microscopy sample preparation, semi-automated transmission electron microscopy screening, and screening evaluation for cells of interest. Thereafter, we detail tomography acquisition, data reconstruction, and joining.For complete details on the use and execution of this protocol, please refer to Dittrich et al.1 : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics. |
| first_indexed | 2024-03-13T03:31:38Z |
| format | Article |
| id | doaj.art-ab5e2ce23d5b4e5badf5a3dbdf45d638 |
| institution | Directory Open Access Journal |
| issn | 2666-1667 |
| language | English |
| last_indexed | 2024-03-13T03:31:38Z |
| publishDate | 2023-09-01 |
| publisher | Elsevier |
| record_format | Article |
| series | STAR Protocols |
| spelling | doaj.art-ab5e2ce23d5b4e5badf5a3dbdf45d6382023-06-24T05:19:06ZengElsevierSTAR Protocols2666-16672023-09-0143102373Protocol for high-throughput electron tomography exemplified on centrioles in primary human plasma cellsSebastian Köhrer0Tobias Dittrich1Martin Schorb2Isabella Haberbosch3Yannick Schwab4Alwin Krämer5Clinical Cooperation Unit Molecular Hematology/Oncology, German Cancer Research Center (DKFZ), and Department of Internal Medicine V, University of Heidelberg, 69120 Heidelberg, Germany; Cell Biology and Biophysics Unit, European Molecular Biology Laboratory (EMBL), 69117 Heidelberg, Germany; Corresponding authorClinical Cooperation Unit Molecular Hematology/Oncology, German Cancer Research Center (DKFZ), and Department of Internal Medicine V, University of Heidelberg, 69120 Heidelberg, Germany; Cell Biology and Biophysics Unit, European Molecular Biology Laboratory (EMBL), 69117 Heidelberg, Germany; Department of Internal Medicine V, University of Heidelberg, 69120 Heidelberg, Germany; Amyloidosis Center, University of Heidelberg, 69120 Heidelberg, GermanyElectron Microscopy Core Facility, European Molecular Biology Laboratory, 69117 Heidelberg, Germany; Corresponding authorClinical Cooperation Unit Molecular Hematology/Oncology, German Cancer Research Center (DKFZ), and Department of Internal Medicine V, University of Heidelberg, 69120 Heidelberg, Germany; Department of Internal Medicine V, University of Heidelberg, 69120 Heidelberg, GermanyCell Biology and Biophysics Unit, European Molecular Biology Laboratory (EMBL), 69117 Heidelberg, Germany; Electron Microscopy Core Facility, European Molecular Biology Laboratory, 69117 Heidelberg, GermanyClinical Cooperation Unit Molecular Hematology/Oncology, German Cancer Research Center (DKFZ), and Department of Internal Medicine V, University of Heidelberg, 69120 Heidelberg, Germany; Department of Internal Medicine V, University of Heidelberg, 69120 Heidelberg, Germany; Corresponding authorSummary: Electron microscopy is the gold standard to characterize cellular ultrastructure. However, production of significant morphometrical data is highly limited by acquisition time. Here, we describe a semi-automated high-throughput strategy using single-axis serial section electron tomography to investigate and analyze centriole ultrastructure in bone-marrow-derived, primary human CD138pos plasma cells. The protocol comprises steps for electron microscopy sample preparation, semi-automated transmission electron microscopy screening, and screening evaluation for cells of interest. Thereafter, we detail tomography acquisition, data reconstruction, and joining.For complete details on the use and execution of this protocol, please refer to Dittrich et al.1 : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.http://www.sciencedirect.com/science/article/pii/S2666166723003404High Throughput ScreeningMicroscopy |
| spellingShingle | Sebastian Köhrer Tobias Dittrich Martin Schorb Isabella Haberbosch Yannick Schwab Alwin Krämer Protocol for high-throughput electron tomography exemplified on centrioles in primary human plasma cells STAR Protocols High Throughput Screening Microscopy |
| title | Protocol for high-throughput electron tomography exemplified on centrioles in primary human plasma cells |
| title_full | Protocol for high-throughput electron tomography exemplified on centrioles in primary human plasma cells |
| title_fullStr | Protocol for high-throughput electron tomography exemplified on centrioles in primary human plasma cells |
| title_full_unstemmed | Protocol for high-throughput electron tomography exemplified on centrioles in primary human plasma cells |
| title_short | Protocol for high-throughput electron tomography exemplified on centrioles in primary human plasma cells |
| title_sort | protocol for high throughput electron tomography exemplified on centrioles in primary human plasma cells |
| topic | High Throughput Screening Microscopy |
| url | http://www.sciencedirect.com/science/article/pii/S2666166723003404 |
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