Whole-cell multi-target single-molecule super-resolution imaging in 3D with microfluidics and a single-objective tilted light sheet

dc.citation.articleNumber10187en_US
dc.citation.journalTitleNature Communicationsen_US
dc.citation.volumeNumber15en_US
dc.contributor.authorSaliba, Nahimaen_US
dc.contributor.authorGagliano, Gabriellaen_US
dc.contributor.authorGustavsson, Anna-Karinen_US
dc.contributor.orgSmalley-Curl Institute;Center for Nanoscale Imaging Sciencesen_US
dc.date.accessioned2025-01-09T20:17:04Zen_US
dc.date.available2025-01-09T20:17:04Zen_US
dc.date.issued2024en_US
dc.description.abstractMulti-target single-molecule super-resolution fluorescence microscopy offers a powerful means of understanding the distributions and interplay between multiple subcellular structures at the nanoscale. However, single-molecule super-resolution imaging of whole mammalian cells is often hampered by high fluorescence background and slow acquisition speeds, especially when imaging multiple targets in 3D. In this work, we have mitigated these issues by developing a steerable, dithered, single-objective tilted light sheet for optical sectioning to reduce fluorescence background and a pipeline for 3D nanoprinting microfluidic systems for reflection of the light sheet into the sample. This easily adaptable microfluidic fabrication pipeline allows for the incorporation of reflective optics into microfluidic channels without disrupting efficient and automated solution exchange. We combine these innovations with point spread function engineering for nanoscale localization of individual molecules in 3D, deep learning for analysis of overlapping emitters, active 3D stabilization for drift correction and long-term imaging, and Exchange-PAINT for sequential multi-target imaging without chromatic offsets. We then demonstrate that this platform, termed soTILT3D, enables whole-cell multi-target 3D single-molecule super-resolution imaging with improved precision and imaging speed.en_US
dc.identifier.citationSaliba, N., Gagliano, G., & Gustavsson, A.-K. (2024). Whole-cell multi-target single-molecule super-resolution imaging in 3D with microfluidics and a single-objective tilted light sheet. Nature Communications, 15(1), 10187. https://doi.org/10.1038/s41467-024-54609-zen_US
dc.identifier.digitals41467-024-54609-zen_US
dc.identifier.doihttps://doi.org/10.1038/s41467-024-54609-zen_US
dc.identifier.urihttps://hdl.handle.net/1911/118144en_US
dc.language.isoengen_US
dc.publisherSpringer Natureen_US
dc.rightsExcept where otherwise noted, this work is licensed under a Creative Commons Attribution-NonCommercial-NoDerivatives (CC BY-NC-ND) license. Permission to reuse, publish, or reproduce the work beyond the terms of the license or beyond the bounds of fair use or other exemptions to copyright law must be obtained from the copyright holder.en_US
dc.rights.urihttps://creativecommons.org/licenses/by-nc-nd/4.0/en_US
dc.titleWhole-cell multi-target single-molecule super-resolution imaging in 3D with microfluidics and a single-objective tilted light sheeten_US
dc.typeJournal articleen_US
dc.type.dcmiTexten_US
dc.type.publicationpublisher versionen_US
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