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  4. Overcoming Preferred Orientation in Cryo-EM With Ultrasonic Excitation During Vitrification
 
preprint

Overcoming Preferred Orientation in Cryo-EM With Ultrasonic Excitation During Vitrification

Williams, Harry M.  
•
Curtis, Wyatt A.  
•
Haubner, Michal  
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September 17, 2025

Preferred particle orientation remains a frequently encountered issue in cryo-electron microscopy that arises when proteins adsorb to the air-water interface in only a limited number of orientations. This can significantly increase the data acquisition time required to reach a desired resolution or even make it impossible to obtain a reconstruction altogether. Here, we show that preferred orientation can be overcome by continuously exciting the sample with ultrasonic waves during vitrification. Our experiments suggest that mechanical oscillations induced in the sample support continuously shake proteins loose form the air-water interface, thereby scrambling their orientations. The simple, physical nature of this mechanism should make it applicable to a wide range of proteins. Since our method can be easily implemented in existing vitrification devices, it should find widespread adoption.

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Type
preprint
DOI
10.1101/2025.09.14.676144
Author(s)
Williams, Harry M.  

École Polytechnique Fédérale de Lausanne

Curtis, Wyatt A.  

École Polytechnique Fédérale de Lausanne

Haubner, Michal  

École Polytechnique Fédérale de Lausanne

Hruby, J

École Polytechnique Fédérale de Lausanne

Drabbels, Marcel  

École Polytechnique Fédérale de Lausanne

Lorenz, Ulrich J.  

École Polytechnique Fédérale de Lausanne

Date Issued

2025-09-17

Publisher

bioRxiv

Written at

EPFL

EPFL units
LND  
LUXS  
Available on Infoscience
February 18, 2026
Use this identifier to reference this record
https://infoscience.epfl.ch/handle/20.500.14299/259693
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