Lens-free scanning

Boden, Andreas; Volpato, Andrea; Hake, Kayley; York, A.; Testa, Ilaria

doi:10.5281/zenodo.3653386

Cited by 3 publications

(2 citation statements)

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“…Therefore, regardless of the final primary objective position, the light-sheet will maintain its alignment with the focal plane of the tertiary objective during scanning to enable optimal 3D volumetric imaging. This tunability is not supported in the most traditional scan-lens/mirror configurations or other scanning geometries with only one movable mirror [7,14]. Furthermore, compared to the more traditional scan-lens/mirror combination, the two-mirror scanning geometry proposed here offers higher light-efficiency and a more compact footprint, which could be beneficial to all OPM designs regardless of whether they use a commercial base or not.…”

Section: Introductionmentioning

confidence: 91%

User-friendly Oblique Plane Microscopy on a fully functional commercially available microscope base

Sirinakis,

Allgeyer,

Nashchekin

et al. 2024

Preprint

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In this work we present an Oblique Plane Microscope designed to work seamlessly with a commercially available microscope base. To support all the functionality offered by the microscope base, where the position of the objective lens is not fixed, we adopted a two-mirror scanning geometry that can compensate for changes to the position of the objective lens during routine microscope operation. We showed that within the expected displacement range of the 100X, 1.35 NA objective lens away from its designed position, and for most practical applications, there is no significant effect on the resolving power, or the fidelity of the 3D data produced by the microscope. Compared to the more traditional scan-lens/galvo-mirror combination, the two-mirror scanning geometry offers higher light-efficiency and a more compact footprint, which could be beneficial to all OPM designs regardless of the use of a commercial base or not.

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Section: Introductionmentioning

confidence: 91%

User-friendly Oblique Plane Microscopy on a fully functional commercially available microscope base

Sirinakis,

Allgeyer,

Nashchekin

et al. 2024

Preprint

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“…2c and Supplementary Fig. 2) that achieves a pure beam translation in remote object space by reflecting twice off a large galvanometric mirror 19 . This scan system simplifies the alignment procedure and significantly increases detection efficiency.…”

Section: Multi-sheet Resolft Imaging Schemementioning

confidence: 99%

Super-sectioning with multi-sheet reversible saturable optical fluorescence transitions (RESOLFT) microscopy

Bodén,

Ollech,

York

et al. 2024

Nat Methods

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Light-sheet fluorescence microscopy is an invaluable tool for four-dimensional biological imaging of multicellular systems due to the rapid volumetric imaging and minimal illumination dosage. However, it is challenging to retrieve fine subcellular information, especially in living cells, due to the width of the sheet of light (>1 μm). Here, using reversibly switchable fluorescent proteins (RSFPs) and a periodic light pattern for photoswitching, we demonstrate a super-resolution imaging method for rapid volumetric imaging of subcellular structures called multi-sheet RESOLFT. Multiple emission-sheets with a width that is far below the diffraction limit are created in parallel increasing recording speed (1–2 Hz) to provide super-sectioning ability (<100 nm). Our technology is compatible with various RSFPs due to its minimal requirement in the number of switching cycles and can be used to study a plethora of cellular structures. We track cellular processes such as cell division, actin motion and the dynamics of virus-like particles in three dimensions.

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A Versatile Oblique Plane Microscope for Large-Scale and High-Resolution Imaging of Subcellular Dynamics

Sapoznik

Chang

Huh

et al. 2020

Preprint

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We present a single-objective light-sheet microscope, also known as an oblique-plane microscope, that uses a bespoke glass-tipped tertiary objective and improves the resolution, field of view, usability, and stability over previous variants. Owing to its high numerical aperture optics, this microscope achieves the highest lateral resolution in light-sheet fluorescence microscopy, and its axial resolution is similar to that of Lattice Light-Sheet Microscopy. Given this performance, we demonstrate high-resolution imaging of clathrin-mediated endocytosis, vimentin, the endoplasmic reticulum, membrane dynamics, and natural killer cell-mediated cell death. Furthermore, we image biological phenomena that would be otherwise challenging or impossible to perform in a traditional light-sheet microscope geometry, including cell migration through a confined space within a microfluidic device, photoactivation of PI3K, and diffusion of cytoplasmic rheological tracers at a volumetric rate of 14 Hz.

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Lens-free scanning

Cited by 3 publications

References 0 publications

User-friendly Oblique Plane Microscopy on a fully functional commercially available microscope base

User-friendly Oblique Plane Microscopy on a fully functional commercially available microscope base

Super-sectioning with multi-sheet reversible saturable optical fluorescence transitions (RESOLFT) microscopy

A Versatile Oblique Plane Microscope for Large-Scale and High-Resolution Imaging of Subcellular Dynamics

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