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bioRxiv · 10.64898/2026.09.27.753993

Whole cryo-FIB lamella acquisition at high magnification with rectangular beam montage cryo-electron tomography

Brown, H. G.·Valimehr, S.·Styczynski, D.·Mudaliyar, M.·Kundapura, S. S.·Johnson, M. D.·Ghosal, D.·Hanssen, E.
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Abstract

The visualisation of large biological samples in cryo-electron tomography requires both few-nanometre resolution and a wide, ideally >10 micrometrre, field of view. For a single image, increasing magnification trades increased resolution for a smaller field of view. Montaging, where high-magnification images are stitched together to increase field of view, addresses the trade-off but is dose-inefficient with round beams that are standard in transmission electron microscopy (TEM), since this illumination shape does not tessellate. Our workflow greatly simplifies montaging schemes for dose-sensitive specimens, allowing large montages of any shape - from individual cells or other features of interest up to whole lamellae - to be collected at high resolution without compromising electron dose efficiency. We demonstrate the workflow by acquiring montage tomograms of whole yeast (Saccharomyces cerevisiae) cells in cryo-FIB lamellae and of human neurons.

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Brown, H. G., Valimehr, S., Styczynski, D., Mudaliyar, M., Kundapura, S. S., Johnson, M. D., Ghosal, D., Hanssen, E.. 2026-09-27. Whole cryo-FIB lamella acquisition at high magnification with rectangular beam montage cryo-electron tomography. https://doi.org/10.64898/2026.09.27.753993

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