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

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Titel: Volumetric imaging of the 3D orientation of cellular structures with a polarized fluorescence light-sheet microscope
Autoren: Chandler, Talon, Guo, Min, Su, Yijun, Chen, Jiji, Wu, Yicong, Liu, Junyu, Agashe, Atharva, Fischer, Robert, Mehta, Shalin B., Kumar, Abhishek, Baskin, Tobias, Jaumouillé, Valentin, Liu, Huafeng, Swaminathan, Vinay, Nain, Amrinder S., Oldenbourg, Rudolf, La Riviere, Patrick J., Shroff, Hari
Weitere Verfasser: Lund University, Profile areas and other strong research environments, Lund University Profile areas, LU Profile Area: Light and Materials, Lunds universitet, Profilområden och andra starka forskningsmiljöer, Lunds universitets profilområden, LU profilområde: Ljus och material, Originator, Lund University, Faculty of Engineering, LTH, LTH Profile areas, LTH Profile Area: Photon Science and Technology, Lunds universitet, Lunds Tekniska Högskola, LTH profilområden, LTH profilområde: Avancerade ljuskällor, Originator, Lund University, Faculty of Engineering, LTH, LTH Profile areas, LTH Profile Area: Nanoscience and Semiconductor Technology, Lunds universitet, Lunds Tekniska Högskola, LTH profilområden, LTH profilområde: Nanovetenskap och halvledarteknologi, Originator, Lund University, Profile areas and other strong research environments, Strategic research areas (SRA), NanoLund: Centre for Nanoscience, Lunds universitet, Profilområden och andra starka forskningsmiljöer, Strategiska forskningsområden (SFO), NanoLund: Centre for Nanoscience, Originator, Lund University, Profile areas and other strong research environments, Other Strong Research Environments, LUCC: Lund University Cancer Centre, Lunds universitet, Profilområden och andra starka forskningsmiljöer, Övriga starka forskningsmiljöer, LUCC: Lunds universitets cancercentrum, Originator, Lund University, Faculty of Medicine, Department of Clinical Sciences, Lund, Section I, Cell mechanobiology, Lunds universitet, Medicinska fakulteten, Institutionen för kliniska vetenskaper, Lund, Sektion I, Cellmekanobiologi, Originator, Lund University, Faculty of Medicine, WCMM-Wallenberg Centre for Molecular Medicine, Lunds universitet, Medicinska fakulteten, WCMM- Wallenberg center för molekylär medicinsk forskning, Originator
Quelle: Proceedings of the National Academy of Sciences. 122(8):1-12
Schlagwörter: Natural Sciences, Physical Sciences, Biophysics, Naturvetenskap, Fysik, Biofysik, Computer and Information Sciences, Computer graphics and computer vision, Data- och informationsvetenskap (Datateknik), Datorgrafik och datorseende
Beschreibung: 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 allthree-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.
Zugangs-URL: https://doi.org/10.1073/pnas.2406679122
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  Data: 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 allthree-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.
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