Rapid lightsheet fluorescence imaging of whole Drosophila brains at nanoscale resolution by potassium acrylate-based expansion microscopy

Taking advantage of the good mechanical strength of expanded Drosophila brains and to tackle their relatively large size that can complicate imaging, we apply potassium (poly)acrylate-based hydrogels for expansion microscopy (ExM), resulting in a 40x plus increased resolution of transgenic fluorescent proteins preserved by glutaraldehyde fixation in the nervous system. Large-volume ExM is realized by using an axicon-based Bessel lightsheet microscope, featuring gentle multi-color fluorophore excitation and intrinsic optical sectioning capability, enabling visualization of Tm5a neurites and L3 lamina neurons with photoreceptors in the optic lobe. We also image nanometer-sized dopaminergic neurons across the same intact iteratively expanded Drosophila brain, enabling us to measure the 3D expansion ratio. Here we show that at a tile scanning speed of ~1 min/mm<sup>3</sup> with 10<sup>12</sup> pixels over 14 hours, we image the centimeter-sized fly brain at an effective resolution comparable to electron microscopy, allowing us to visualize mitochondria within presynaptic compartments and Bruchpilot (Brp) scaffold proteins distributed in the central complex, enabling robust analyses of neurobiological topics.

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Publication Details

Journal
Nature Communications
Published
2024-12-30
DOI
https://doi.org/10.1038/s41467-024-55305-8
Citations
9
Primary Topic
Advanced Fluorescence Microscopy Techniques
Type
article
Field-Weighted Citation Impact
5.96

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article

Rapid lightsheet fluorescence imaging of whole Drosophila brains at nanoscale resolution by potassium acrylate-based expansion microscopy

Suewei Lin, Bi‐Chang Chen, Chi‐Hon Lee, Li‐An Chu et al.
9 citations
Nature Communications
Advanced Fluorescence Microscopy Techniques
5.96
article

Rapid lightsheet fluorescence imaging of whole Drosophila brains at nanoscale resolution by potassium acrylate-based expansion microscopy

Suewei Lin, Bi‐Chang Chen, Chi‐Hon Lee, Li‐An Chu, Jui-Cheng Hsu, Chia-Ming Lee, Po Ting Lin, Chien‐Kai Wang, Yi‐Chung Tung, Yi‐Ping Hsueh, Chiao-Hui Tu, Fan‐Gang Tseng, Tzu‐Yang Lin, Tung‐Han Hsieh, Xuejiao Tian, Wenjie Chen, Peilin Chen, Min-Ju Tsai, Hsin Chen
article en
9 citations

Abstract

Taking advantage of the good mechanical strength of expanded Drosophila brains and to tackle their relatively large size that can complicate imaging, we apply potassium (poly)acrylate-based hydrogels for expansion microscopy (ExM), resulting in a 40x plus increased resolution of transgenic fluorescent proteins preserved by glutaraldehyde fixation in the nervous system. Large-volume ExM is realized by using an axicon-based Bessel lightsheet microscope, featuring gentle multi-color fluorophore excitation and intrinsic optical sectioning capability, enabling visualization of Tm5a neurites and L3 lamina neurons with photoreceptors in the optic lobe. We also image nanometer-sized dopaminergic neurons across the same intact iteratively expanded Drosophila brain, enabling us to measure the 3D expansion ratio. Here we show that at a tile scanning speed of ~1 min/mm<sup>3</sup> with 10<sup>12</sup> pixels over 14 hours, we image the centimeter-sized fly brain at an effective resolution comparable to electron microscopy, allowing us to visualize mitochondria within presynaptic compartments and Bruchpilot (Brp) scaffold proteins distributed in the central complex, enabling robust analyses of neurobiological topics.

Nature CommunicationsVol. 15(1)
National Taiwan University (TW), National Tsing Hua University (TW), Research Center for Applied Science, Academia Sinica (TW), Institute of Cellular and Organismic Biology, Academia Sinica (TW), Institute of Molecular Biology, Academia Sinica (TW), Academia Sinica (TW), National Cheng Kung University (TW)
Academia Sinica, Ministry of Science and Technology, Taiwan
Openalex Percentile: Top 4%
Advanced Fluorescence Microscopy Techniques
5.96
DOIOpen Access PDFOpenAlex
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