Cluster-based milling method for large-field-of-view volume electron microscopy

Kormacheva, Maria (2023): Cluster-based milling method for large-field-of-view volume electron microscopy. Dissertation, LMU München: Fakultät für Physik

Abstract

Current methods for automated volume image acquisition that allow resolving the structure of neural circuits have a limited field-of-view. This work investigates using a gas cluster ion beam (GCIB) to overcome the field-of-view limitation and explores the combination with a multi-beam scanning electron microscope (mSEM) to create a system for the acquisition of the whole mouse brain. To this end, a staining protocol for 500 micrometre thick whole-coronal cross-sections is established, and a GCIB is incorporated with a scanning electron microscope (SEM) to identify optimal system parameters. In addition, an electron beam irradiation system is built and automated to induce conductivity in collected sections for SEM imaging. The results verify that ion milling can keep up with the imaging rate in the mSEM while maintaining adequate quality. In addition, software is implemented for targeted image acquisition in the mSEM. Finally, calculations show that acquiring the whole mouse brain is feasible but heavily dependent on the imaging rate and the number of parallel GCIB-mSEM systems.

• Dokumententyp: Dissertationen (Dissertation, LMU München)
• Keywords: connectomics, electron microscopy
• Themengebiete: 500 Naturwissenschaften und Mathematik ; 500 Naturwissenschaften und Mathematik > 530 Physik
• Fakultäten: Fakultät für Physik
• Sprache der Hochschulschrift: Englisch
• Datum der mündlichen Prüfung: 19. Juni 2023
• 1. Bericht­erstatter:in: Denk, Winfried