Light-sheet mesoscopy with the Mesolens provides fast sub-cellular resolution imaging throughout large tissue volumes

Battistella, Eliana; Schniete, Jan; Wesencraft, Katrina; Quintana, Juan F.; McConnell, Gail

doi:10.1016/j.isci.2022.104797

Cited by 12 publications

(9 citation statements)

References 42 publications

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“…Ideally in a LS microscope, there would be an infinitesimally thin sheet of infinite extent. This is the reason that some more complex LS microscopes employ Bessel beams 17 or Airy beams, 18 which are nondiffractive but carry extra fringes whose effects must be mitigated via other means. Using standard optics, a tightly focused beam axially ( Z , perpendicular to the imaging plane) means a tightly focused beam laterally (in X ).…”

Section: Resultsmentioning

confidence: 99%

Practical strategies for robust and inexpensive imaging of aqueous‐cleared tissues

Williams

Bloom

Robertus

et al. 2023

Journal of Microscopy

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Lightsheet microscopy offers an ideal method for imaging of large (mm–cm scale) biological tissues rendered transparent via optical clearing protocols. However the diversity of clearing technologies and tissue types, and how these are adapted to the microscope can make tissue mounting complicated and somewhat irreproducible. Tissue preparation for imaging can involve glues and or equilibration in a variety of expensive and/or proprietary formulations. Here we present practical advice for mounting and capping cleared tissues in optical cuvettes for macroscopic imaging, providing a standardised 3D cell that can be imaged routinely and relatively inexpensively. We show that acrylic cuvettes cause minimal spherical aberration with objective numerical apertures less than 0.65. Furthermore, we describe methods for aligning and assessing the light sheets, discriminating fluorescence from autofluorescence, identifying chromatic artefacts due to differential scattering and removing streak artefacts such that they do not confound downstream 3D object segmentation analyses, with mouse embryo, liver and heart imaging as demonstrated examples.

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Section: Resultsmentioning

confidence: 99%

Practical strategies for robust and inexpensive imaging of aqueous‐cleared tissues

Williams

Bloom

Robertus

et al. 2023

Journal of Microscopy

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“…For optical sectioning, various options exist for achieving 3D fluorescence imaging in the visible wavelength region. These include scanning confocal microscopy (4,14), light-sheet microscopy (15), light-field microscopy (16), and spatiotemporal focusing (17). Among these options, we selected the confocal imaging method for this study owing to the limitations of light-sheet method in uniformly illuminating areas larger than 1 cm (15,18), and the trade-off between spatial resolution and the ability to resolve three dimensions in a single shot for the light-field method (19).…”

Section: Resultsmentioning

confidence: 99%

Volumetric trans-scale imaging of massive quantity of heterogeneous cell populations in centimeter-wide tissue and embryo

Ichimura,

Kakizuka,

Taniguchi

et al. 2023

Preprint

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We established a volumetric trans-scale imaging system with an ultra-large field-of-view (FOV) that enables simultaneous observation of millions of cellular dynamics in centimeter-wide three-dimensional (3D) tissues and embryos. Using a custom-made giant lens system with a magnification of 2× and a numerical aperture (NA) of 0.25, and a CMOS camera with more than 100 megapixels, we built a trans-scale scope AMATERAS-2, and realized fluorescence imaging with a lateral spatial resolution of approximately 1.2 µm across an FOV of approximately 1.5 × 1.0 cm2. The 3D resolving capability was realized through a combination of optical and computational sectioning techniques tailored for our low-power imaging system. We applied the imaging technique to 1.2 cm-wide section of mouse brain, and successfully observed various regions of the brain with sub-cellular resolution in a single FOV. We also performed time-lapse imaging of a 1-cm-wide vascular network during quail embryo development for over 24 hours, visualizing the movement of over 4.0 × 105vascular endothelial cells and quantitatively analyzing their dynamics. Our results demonstrate the potential of this technique in accelerating production of comprehensive reference maps of all cells in organisms and tissues, which contributes to understanding developmental processes, brain functions, and pathogenesis of disease, as well as high-throughput quality check of tissues used for transplantation medicine.

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“…To visualise and quantify data in three-dimensions, OME.TIFF files from the two channels were sequentially opened with Imaris and these data were subsequently converted to the proprietary Imaris file format (Imaris 9.8, Oxford Instruments). The 'Surfaces' tool was used to segment objects in each channel from the background to facilitate measurements [19]. Next, the 'Statistics' function was used to extract volumetric information from the image data.…”

Section: Image Processing and Analysismentioning

confidence: 99%

Multimodal optical mesoscopy reveals the quantity and spatial distribution of gram-positive biofilms inex vivotonsils

Clapperton,

Kunanandam,

Florea

et al. 2023

Preprint

Self Cite

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Biofilms are known to be present in tonsils, but little is known about their spatial location and size distribution throughout the tonsil. Studies of the location and distribution of biofilms in tonsil specimens have thus far been limited to either high-magnification methods such as electron microscopy, which enables high resolution imaging but only from a tiny tissue volume, or lower magnification techniques such as light microscopy, which allow imaging of larger specimens but with poor spatial resolution. To overcome these limitations, we report the use of multimodal optical mesoscopy to visualize and quantify the number and spatial distribution of gram-positive biofilms in fresh, excised paediatric tonsils. This methodology supports simultaneous imaging of both the tonsil host and biofilms in whole mounts of tissue up to 5 mm x 5 mm x 3 mm with subcellular resolution throughout. A quantitative assessment of thirty-six tonsil specimens revealed no statistically significant difference between biofilm presence on the tonsil surface and the interior of the tonsil. This new quantitative mesoscale imaging approach may prove useful in understanding the role of biofilms in tonsillar diseases and other infections.

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Light-sheet mesoscopy with the Mesolens provides fast sub-cellular resolution imaging throughout large tissue volumes

Cited by 12 publications

References 42 publications

Practical strategies for robust and inexpensive imaging of aqueous‐cleared tissues

Practical strategies for robust and inexpensive imaging of aqueous‐cleared tissues

Volumetric trans-scale imaging of massive quantity of heterogeneous cell populations in centimeter-wide tissue and embryo

Multimodal optical mesoscopy reveals the quantity and spatial distribution of gram-positive biofilms inex vivotonsils

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