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Volumetric imaging of the 3D orientation of cellular structures with a polarized fluorescence light-sheet microscope

Chandler, Talon ; Guo, Min ; Su, Yijun ; Chen, Jiji ; Wu, Yicong ; Liu, Junyu ; Agashe, Atharva ; Fischer, Robert ; Mehta, Shalin B. and Kumar, Abhishek , et al. (2025) In Proceedings of the National Academy of Sciences 122(8). p.1-12
Abstract
Polarized fluorescence microscopy is a valuable tool for measuring molecular orientations in biological samples, but techniques for recovering three-dimensional orientations and positions of fluorescent ensembles are limited. We report a polarized dual-view light-sheet system for determining the diffraction-limited three-dimensional distribution of the orientations and positions of ensembles of fluorescent dipoles that label biological structures. We share a set of visualization, histogram, and profiling tools for interpreting these positions and orientations. We model the distributions based on the polarization-dependent efficiency of excitation and detection of emitted fluorescence, using coarse-grained representations we call... (More)
Polarized fluorescence microscopy is a valuable tool for measuring molecular orientations in biological samples, but techniques for recovering three-dimensional orientations and positions of fluorescent ensembles are limited. We report a polarized dual-view light-sheet system for determining the diffraction-limited three-dimensional distribution of the orientations and positions of ensembles of fluorescent dipoles that label biological structures. We share a set of visualization, histogram, and profiling tools for interpreting these positions and orientations. We model the distributions based on the polarization-dependent efficiency of excitation and detection of emitted fluorescence, using coarse-grained representations we call orientation distribution functions (ODFs). We apply ODFs to create physics-informed models of image formation with spatio-angular point-spread and transfer functions. We use theory and experiment to conclude that light-sheet tilting is a necessary part of our design for recovering all three-dimensional orientations. We use our system to extend known two-dimensional results to three dimensions in FM1-43-labeled giant unilamellar vesicles, fast-scarlet-labeled cellulose in xylem cells, and phalloidin-labeled actin in U2OS cells. Additionally, we observe phalloidin-labeled actin in mouse fibroblasts grown on grids of labeled nanowires and identify correlations between local actin alignment and global cell-scale orientation, indicating cellular coordination across length scales. (Less)
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organization
publishing date
type
Contribution to journal
publication status
published
subject
in
Proceedings of the National Academy of Sciences
volume
122
issue
8
article number
e2406679122
pages
1 - 12
publisher
National Academy of Sciences
external identifiers
  • scopus:85219101879
  • pmid:39982748
ISSN
0027-8424
DOI
10.1073/pnas.2406679122
language
English
LU publication?
yes
id
3c22d0da-262e-494c-b0c0-dc444779a9b8
date added to LUP
2025-05-30 11:01:06
date last changed
2025-06-03 03:00:05
@article{3c22d0da-262e-494c-b0c0-dc444779a9b8,
  abstract     = {{Polarized fluorescence microscopy is a valuable tool for measuring molecular orientations in biological samples, but techniques for recovering three-dimensional orientations and positions of fluorescent ensembles are limited. We report a polarized dual-view light-sheet system for determining the diffraction-limited three-dimensional distribution of the orientations and positions of ensembles of fluorescent dipoles that label biological structures. We share a set of visualization, histogram, and profiling tools for interpreting these positions and orientations. We model the distributions based on the polarization-dependent efficiency of excitation and detection of emitted fluorescence, using coarse-grained representations we call orientation distribution functions (ODFs). We apply ODFs to create physics-informed models of image formation with spatio-angular point-spread and transfer functions. We use theory and experiment to conclude that light-sheet tilting is a necessary part of our design for recovering all three-dimensional orientations. We use our system to extend known two-dimensional results to three dimensions in FM1-43-labeled giant unilamellar vesicles, fast-scarlet-labeled cellulose in xylem cells, and phalloidin-labeled actin in U2OS cells. Additionally, we observe phalloidin-labeled actin in mouse fibroblasts grown on grids of labeled nanowires and identify correlations between local actin alignment and global cell-scale orientation, indicating cellular coordination across length scales.}},
  author       = {{Chandler, Talon and Guo, Min and Su, Yijun and Chen, Jiji and Wu, Yicong and Liu, Junyu and Agashe, Atharva and Fischer, Robert and Mehta, Shalin B. and Kumar, Abhishek and Baskin, Tobias and Jaumouillé, Valentin and Liu, Huafeng and Swaminathan, Vinay and Nain, Amrinder S. and Oldenbourg, Rudolf and La Riviere, Patrick J. and Shroff, Hari}},
  issn         = {{0027-8424}},
  language     = {{eng}},
  month        = {{02}},
  number       = {{8}},
  pages        = {{1--12}},
  publisher    = {{National Academy of Sciences}},
  series       = {{Proceedings of the National Academy of Sciences}},
  title        = {{Volumetric imaging of the 3D orientation of cellular structures with a polarized fluorescence light-sheet microscope}},
  url          = {{http://dx.doi.org/10.1073/pnas.2406679122}},
  doi          = {{10.1073/pnas.2406679122}},
  volume       = {{122}},
  year         = {{2025}},
}