Oral Presentation 51st Lorne Proteins Conference 2026

High-resolution cryo-EM using a common 120-keV LaB6 electron microscope equipped with a sub–200-keV direct electron detector. (130828)

Hariprasad Venugopal 1 , Sergey Gorelick 1 , Jesse Mobbs 2 3 , Cyntia Taveneau 4 , Daniel R Fox 5 6 7 , Ziva Vuckovic 8 , Sahil Gulati 9 , Gavin Knott 4 , Rhys Grinter 2 6 7 , David Thal 2 3 , Stephen Mick 9 , Cory Czarnik 9 , Georg Ramm 1 4
  1. Ramaciotti Center for Cryo-Electron Microscopy, Monash University, Clayton, MELBOURNE, VICTORIA, Australia
  2. Australian Research Council Centre for Cryo-Electron Microscopy of Membrane Proteins, Monash Institute of Pharmaceutical Sciences, Monash University, Parkville, MELBOURNE, Victoria, Australia
  3. Drug Discovery Biology, Monash Institute of Pharmaceutical Sciences, Monash University, Parkville, MELBOURNE, Victoria, Australia
  4. Department of Biochemistry and Molecular Biology, Monash Biomedicine Discovery Institute, Monash University, Clayton, MELBOURNE, Victoria, Australia
  5. Australian Research Council Centre for Cryo-Electron Microscopy of Membrane Proteins, Monash Institute of Pharmaceutical Sciences, Monash University, Parkville, MELBOURNE, Victoria, Australia
  6. Department of Biochemistry and Pharmacology, Bio21 Molecular Science and Biotechnology Institute, The University of Melbourne, Parkville, MELBOURNE, Victoria, Australia
  7. Department of Microbiology, Biomedicine Discovery Institute, Monash University, Clayton, MELBOURNE, Victoria, Australia
  8. GlycoEra AG, Einsiedlerstrasse 34, 8820 Wädenswil, Switzerland
  9. GATAN, AMETEK, Pleasanton, CA, USA

Cryo-electron microscopy (Cryo-EM) single particle analysis (SPA) has become a major structural biology technique in recent years. High-resolution cryo-EM typically requires 200-300-keV cryo-capable transmission electron microscopes (cryo-TEMs) with coherent field emission gun (FEG) as source, stable columns, autoloader system and direct electron detector (DED). These setups are specialised for Cryo-EM work and are expensive to establish and maintain. More recently, the concept of using 100-keV cryo-TEMs has been introduced as a way to make cryo-EM more affordable and hence accessible to a larger group of researchers (1, 2). So far, the implementation of these 100-keV cryo-TEMs have relied on specialised microscopes with FEG sources as well as more stable optics.

Recently, we explored, whether a standard 120-keV LaB6 source TEM, commonly available at many laboratories worldwide, can be upgraded with a sub 200-keV optimised DED (GATAN, Alpine) and its suitability for high-resolution cryo-EM using a standard side entry cryo-holder (3). Using this imaging configuration, we successfully achieved a 2.65Å resolution reconstruction for apoferritin. We were able to solve a challenging, asymmetric, 153 kDa membrane protein GPCR (M4 muscarinic acid receptor) to a resolution of 4.4Å. Additionally, we even managed to resolve a small protein target like haemoglobin (64 kDa) to 4.33Å. Importantly, all these results were achieved using an automated data collection routine implemented through SerialEM, making it feasible to collect large cryo-EM data sets with a side entry cryo-holder. Building on this work, we demonstrate results at 80-keV and implications for imaging sub 100 kDa molecules. Further, we also showcase preliminary results into modifying the existing LaB6 source to improve its coherence.

Overall, these findings point to a promising, widely accessible path for obtaining high-quality cryo-EM structures. We believe this imaging configuration offers many laboratories the opportunity to establish effective cryo-EM SPA screening capabilities without investing in costly specialised cryo-TEMs. This could help to considerably lower the economic entry barrier for cryo-EM SPA and contribute to the “democratisation” of cryo-EM.

  1. 1. K. Naydenova, G. McMullan, M. Peet, Y. Lee, P. Edwards, S. Chen, E. Leahy, S. Scotcher, R. Henderson, C. Russo. IUCrJ 6, 1086-1098 (2019).
  2. 2. G. McMullan, K. Naydenova, D. Mihaylov, K. Yamashita, M. J. Peet, H. Wilson, J. L. Dickerson, S. Chen, G. Cannone, Y. Lee. Proceedings of the National Academy of Sciences 120, e2312905120 (2023).
  3. 3. H. Venugopal, J. Mobbs, C. Taveneau, D. R. Fox, Z. Vuckovic, S. Gulati, G. Knott, R. Grinter, D. Thal, S. Mick, C. Czarnik, G. Ramm. Science Advances 11, eadr0438 (2025).