Expansion-assisted selective plane illumination microscopy for nanoscale imaging of centimeter-scale tissues
Recent advances in tissue processing, labeling, and fluorescence microscopy are providing unprecedented views of the structure of cells and tissues at sub-diffraction resolutions and near single molecule sensitivity, driving discoveries in diverse fields of biology, including neuroscience. Biological tissue is organized over scales of nanometers to centimeters. Harnessing molecular imaging across intact, three-dimensional samples on this scale requires new types of microscopes with larger fields of view and working distance, as well as higher throughput. We present a new expansion-assisted selective plane illumination microscope (ExA-SPIM) with aberration-free 1.5 µm×1.5 µm×3 µm optical resolution over a large field of view (10.6×8.0 mm 2 ) and working distance (35 mm) at speeds up to 946 megavoxels/s. Combined with new tissue clearing and expansion methods, the microscope allows imaging centimeter-scale samples with 375 nm lateral and 750 nm axial resolution (4× expansion), including entire mouse brains, with high contrast and without sectioning. We illustrate ExA-SPIM by reconstructing individual neurons across the mouse brain, imaging cortico-spinal neurons in the macaque motor cortex, and visualizing axons in human white matter.
Authors
- David Feng (ORCID: https://orcid.org/0000-0002-4920-8123)
- Steve I. Perlmutter (ORCID: https://orcid.org/0000-0002-1149-4152)
- R. Clay Reid (ORCID: https://orcid.org/0000-0002-8697-6797)
- Brian Kalmbach (ORCID: https://orcid.org/0000-0003-3136-8097)
- Kevin Takasaki (ORCID: https://orcid.org/0009-0000-1531-6022)
- Bosiljka Tasic (ORCID: https://orcid.org/0000-0002-6861-4506)
- Jonathan T. Ting (ORCID: https://orcid.org/0000-0001-8266-0392)
- Kevin Cao (ORCID: https://orcid.org/0000-0002-0226-7687)
- Andrew Recknagel
- Naveen Ouellette (ORCID: https://orcid.org/0009-0005-4472-0880)
- Marcus Hooper (ORCID: https://orcid.org/0000-0003-1228-5958)
- Jayaram Chandrashekar (ORCID: https://orcid.org/0000-0001-6412-0114)
- Sonya Vasquez (ORCID: https://orcid.org/0009-0004-4202-0461)
- Nikolai Dembrow (ORCID: https://orcid.org/0000-0002-7699-6591)
- Ayana Hellevik
- Emily Turschak (ORCID: https://orcid.org/0000-0001-6096-5486)
- Chris Chronopoulos (ORCID: https://orcid.org/0009-0000-1989-1876)
- Saskia de Vries (ORCID: https://orcid.org/0000-0002-3704-3499)
- Pooja Balaram (ORCID: https://orcid.org/0009-0000-0500-9635)
- Gábor Kovács (ORCID: https://orcid.org/0000-0003-1779-775X)
- Cameron Arshadi (ORCID: https://orcid.org/0000-0002-1885-080X)
- Adam K. Glaser (ORCID: https://orcid.org/0000-0003-3558-8994)
- Nathan Clack (ORCID: https://orcid.org/0000-0001-6236-9282)
- Lindsey A. Barner (ORCID: https://orcid.org/0000-0002-9156-0024)
- Karel Svoboda (ORCID: https://orcid.org/0000-0002-6670-7362)
- Anna Grim
- Alan Liddell
- Judith Baka (ORCID: https://orcid.org/0000-0002-8803-0217)
- Xiaoyun Jiang (ORCID: https://orcid.org/0000-0002-4733-9357)
- Shamishtaa Seshamani
- John R. Rohde
- Micah Woodard
- Molly Logsdon
- Rajvi Javeri
Institutions
- Allen Institute for Brain Science (US)
- University of Washington (US)
- Chan Zuckerberg Initiative (United States) (US)
- Allen Institute (US)
- Allen Institute for Neural Dynamics (US)
Publication Details
- Journal
- eLife
- Published
- 2025-06-30
- DOI
- https://doi.org/10.7554/elife.91979.4
- Citations
- 4
- Primary Topic
- Advanced Fluorescence Microscopy Techniques
- Type
- article
- Field-Weighted Citation Impact
- 6.54
Funders
- National Institute of Mental Health
- National Institute of Neurological Disorders and Stroke