Refined cryo-imaging quality of extracellular vesicles through TEM-adaptions in our newly implemented cryo-TEM facility

Andreas Marl; Astrid Obermayer; Jakub Zalesak; Gregor Zickler; Cristian-Tudor Matea; Nicole Meisner-Kober; Simone Pokrant

Refined cryo-imaging quality of extracellular vesicles through TEM-adaptions in our newly implemented cryo-TEM facility

Andreas Marl, Astrid Obermayer^* (Corresponding author), Jakub Zalesak^* (Corresponding author), Gregor Zickler^* (Corresponding author), Cristian-Tudor Matea^* (Corresponding author), Nicole Meisner-Kober^* (Corresponding author), Simone Pokrant^* (Corresponding author)

^*Corresponding author for this work

Research output: Contribution to conference › Poster › peer-review

Abstract

Extracellular vesicles (EVs) are nanosized phospholipid membrane-enclosed particles, released by almost every cell type, playing key roles in intracellular communication. The interest in EVs has grown exponentially with the discovery of their importance in various physiological and pathophysiological processes along with their potential for diagnostic and therapeutic applications. The understanding of structure-function relationships requires imaging of these vesicles in their native environment under close-to-live conditions at the nanoscale.

In the initial phase of our FFG infrastructure call project (BioMat-TEM) together with additional funding of the Land Salzburg we established a Transmission Electron Microscopy lab including a plunge freezing system for sample vitrification and implemented a cryo holder with a transfer unit. Additionally, we developed operating procedures for sample preparation of EVs and conducted TEM analysis of these samples under cryo conditions. In the second phase we acquired an energy filter and a new prefilter camera capable of capturing higher contrast images. This enables us now to greatly improve imaging quality and visualize more structural details such as the membrane bilayer.

The next phase of the project will focus on optimizing sample preparation with the aim of developing standard operation procedures for each EV source. This will enable us to increase the throughput of EV cryo-TEM imaging and to obtain data that allow comparisons across different variants of EVs.

Original language	English
Publication status	Published - 5 Apr 2024

Fields of Science and Technology Classification 2012

205 Materials Engineering
106 Biology
103 Physics, Astronomy

Cite this

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title = "Refined cryo-imaging quality of extracellular vesicles through TEM-adaptions in our newly implemented cryo-TEM facility",

abstract = "Extracellular vesicles (EVs) are nanosized phospholipid membrane-enclosed particles, released by almost every cell type, playing key roles in intracellular communication. The interest in EVs has grown exponentially with the discovery of their importance in various physiological and pathophysiological processes along with their potential for diagnostic and therapeutic applications. The understanding of structure-function relationships requires imaging of these vesicles in their native environment under close-to-live conditions at the nanoscale.In the initial phase of our FFG infrastructure call project (BioMat-TEM) together with additional funding of the Land Salzburg we established a Transmission Electron Microscopy lab including a plunge freezing system for sample vitrification and implemented a cryo holder with a transfer unit. Additionally, we developed operating procedures for sample preparation of EVs and conducted TEM analysis of these samples under cryo conditions. In the second phase we acquired an energy filter and a new prefilter camera capable of capturing higher contrast images. This enables us now to greatly improve imaging quality and visualize more structural details such as the membrane bilayer.The next phase of the project will focus on optimizing sample preparation with the aim of developing standard operation procedures for each EV source. This will enable us to increase the throughput of EV cryo-TEM imaging and to obtain data that allow comparisons across different variants of EVs. ",

author = "Andreas Marl and Astrid Obermayer and Jakub Zalesak and Gregor Zickler and Cristian-Tudor Matea and Nicole Meisner-Kober and Simone Pokrant",

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T1 - Refined cryo-imaging quality of extracellular vesicles through TEM-adaptions in our newly implemented cryo-TEM facility

AU - Marl, Andreas

AU - Obermayer, Astrid

AU - Zalesak, Jakub

AU - Zickler, Gregor

AU - Matea, Cristian-Tudor

AU - Meisner-Kober, Nicole

AU - Pokrant, Simone

PY - 2024/4/5

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N2 - Extracellular vesicles (EVs) are nanosized phospholipid membrane-enclosed particles, released by almost every cell type, playing key roles in intracellular communication. The interest in EVs has grown exponentially with the discovery of their importance in various physiological and pathophysiological processes along with their potential for diagnostic and therapeutic applications. The understanding of structure-function relationships requires imaging of these vesicles in their native environment under close-to-live conditions at the nanoscale.In the initial phase of our FFG infrastructure call project (BioMat-TEM) together with additional funding of the Land Salzburg we established a Transmission Electron Microscopy lab including a plunge freezing system for sample vitrification and implemented a cryo holder with a transfer unit. Additionally, we developed operating procedures for sample preparation of EVs and conducted TEM analysis of these samples under cryo conditions. In the second phase we acquired an energy filter and a new prefilter camera capable of capturing higher contrast images. This enables us now to greatly improve imaging quality and visualize more structural details such as the membrane bilayer.The next phase of the project will focus on optimizing sample preparation with the aim of developing standard operation procedures for each EV source. This will enable us to increase the throughput of EV cryo-TEM imaging and to obtain data that allow comparisons across different variants of EVs.

AB - Extracellular vesicles (EVs) are nanosized phospholipid membrane-enclosed particles, released by almost every cell type, playing key roles in intracellular communication. The interest in EVs has grown exponentially with the discovery of their importance in various physiological and pathophysiological processes along with their potential for diagnostic and therapeutic applications. The understanding of structure-function relationships requires imaging of these vesicles in their native environment under close-to-live conditions at the nanoscale.In the initial phase of our FFG infrastructure call project (BioMat-TEM) together with additional funding of the Land Salzburg we established a Transmission Electron Microscopy lab including a plunge freezing system for sample vitrification and implemented a cryo holder with a transfer unit. Additionally, we developed operating procedures for sample preparation of EVs and conducted TEM analysis of these samples under cryo conditions. In the second phase we acquired an energy filter and a new prefilter camera capable of capturing higher contrast images. This enables us now to greatly improve imaging quality and visualize more structural details such as the membrane bilayer.The next phase of the project will focus on optimizing sample preparation with the aim of developing standard operation procedures for each EV source. This will enable us to increase the throughput of EV cryo-TEM imaging and to obtain data that allow comparisons across different variants of EVs.

M3 - Poster

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