scholarly journals Three-photon light-sheet fluorescence microscopy

2018 ◽  
Author(s):  
Adriá Escobet-Montalbán ◽  
Federico M. Gasparoli ◽  
Jonathan Nylk ◽  
Pengfei Liu ◽  
Zhengyi Yang ◽  
...  
Keyword(s):  
Fluorescence Microscopy ◽  
Imaging Modality ◽  
Light Sheet ◽  
Wide Field ◽  
Excitation Light ◽  
Volumetric Imaging ◽  
Cellular Spheroids ◽  
Light Sheet Microscopy ◽  
Photon Excitation ◽  
Light Sheet Fluorescence Microscopy

We present the first demonstration of three-photon excitation light-sheet fluorescence microscopy. Light-sheet fluorescence microscopy in single- and two-photon modes has emerged as a powerful wide-field, low photo-damage technique for fast volumetric imaging of biological samples. We extend this imaging modality to the three-photon regime enhancing its penetration depth. Our present study uses a standard conventional femtosecond pulsed laser at 1000 nm wavelength for the imaging of 450 µm diameter cellular spheroids. In addition, we show, experimentally and through numerical simulations, the potential advantages in three-photon light-sheet microscopy of using propagation-invariant Bessel beams in preference to Gaussian beams.

scholarly journals Light sheet fluorescence microscopy of optically cleared brains for studying the glymphatic system

2020 ◽  
Vol 40 (10) ◽  
pp. 1975-1986
Author(s):  
Nicholas B Bèchet ◽  
Tekla M Kylkilahti ◽  
Bengt Mattsson ◽  
Martina Petrasova ◽  
Nagesh C Shanbhag ◽  
...  
Keyword(s):  
Fluorescence Microscopy ◽  
Fluid Transport ◽  
Light Sheet ◽  
Brain Parenchyma ◽  
Sleep State ◽  
Glymphatic System ◽  
Light Sheet Microscopy ◽  
Highly Active ◽  
Quantitative Analyses ◽  
Light Sheet Fluorescence Microscopy

Fluid transport in the perivascular space by the glia-lymphatic (glymphatic) system is important for the removal of solutes from the brain parenchyma, including peptides such as amyloid-beta which are implicated in the pathogenesis of Alzheimer’s disease. The glymphatic system is highly active in the sleep state and under the influence of certain of anaesthetics, while it is suppressed in the awake state and by other anaesthetics. Here we investigated whether light sheet fluorescence microscopy of whole optically cleared murine brains was capable of detecting glymphatic differences in sleep- and awake-mimicking anaesthesia, respectively. Using light-sheet imaging of whole brains, we found anaesthetic-dependent cerebrospinal fluid (CSF) influx differences, including reduced tracer influx along tertiary branches of the middle cerebral artery and reduced influx along dorsal and anterior penetrating arterioles, in the awake-mimicking anaesthesia. This study establishes that light sheet microscopy of optically cleared brains is feasible for quantitative analyses and can provide images of the entire glymphatic system in whole brains.

scholarly journals Characterization of Scattering Effects in Phantom Samples using Single and Two-Photon Excitation Light Sheet Microscopy

2012 ◽  
Vol 102 (3) ◽  
pp. 195a-196a
Author(s):  
Zeno Lavagnino ◽  
Francesca Cella Zanacchi ◽  
Emiliano Ronzitti ◽  
Ivan Coto Hernandez ◽  
Alberto Diaspro
Keyword(s):  
Light Sheet ◽  
Excitation Light ◽  
Two Photon ◽  
Light Sheet Microscopy ◽  
Photon Excitation ◽  
Scattering Effects ◽  
Two Photon Excitation

scholarly journals Wide field intravital imaging by two-photon-excitation digital-scanned light-sheet microscopy (2p-DSLM) with a high-pulse energy laser

2014 ◽  
Vol 5 (10) ◽  
pp. 3311 ◽  
Author(s):  
Atsushi Maruyama ◽  
Yusuke Oshima ◽  
Hiroko Kajiura-Kobayashi ◽  
Shigenori Nonaka ◽  
Takeshi Imamura ◽  
...  
Keyword(s):  
Pulse Energy ◽  
Light Sheet ◽  
Intravital Imaging ◽  
Wide Field ◽  
High Pulse Energy ◽  
Two Photon ◽  
Light Sheet Microscopy ◽  
Photon Excitation ◽  
High Pulse ◽  
Energy Laser

scholarly journals Light-Sheet Fluorescence Microscopy with Scanning Non-diffracting Beams

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Hosein Kafian ◽  
Meelad Lalenejad ◽  
Sahar Moradi-Mehr ◽  
Shiva Akbari Birgani ◽  
Daryoush Abdollahpour
Keyword(s):  
Fluorescence Microscopy ◽  
Spatial Coherence ◽  
Three Dimensional ◽  
Light Sheet ◽  
Tissue Imaging ◽  
Wide Field ◽  
Self Healing ◽  
Main Challenge ◽  
Cancer Tumor ◽  
Light Sheet Fluorescence Microscopy

Abstract Light-sheet fluorescence microscopy (LSFM) has now become a unique tool in different fields ranging from three-dimensional (3D) tissue imaging to real-time functional imaging of neuronal activities. Nevertheless, obtaining high-quality artifact-free images from large, dense and inhomogeneous samples is the main challenge of the method that still needs to be adequately addressed. Here, we demonstrate significant enhancement of LSFM image qualities by using scanning non-diffracting illuminating beams, both through experimental and numerical investigations. The effect of static and scanning illumination with several beams are analyzed and compared, and it is shown that scanning 2D Airy light-sheet is minimally affected by the inhomogeneities in the samples, and provides higher contrasts and uniform resolution over a wide field-of-view, due to its reduced spatial coherence, self-healing feature and longer penetration depth. Further, the capabilities of the illumination scheme is utilized for both single-and double-wavelength 3D imaging of large and dense mammospheres of cancer tumor cells as complex inhomogeneous biological samples.

Light sheet fluorescence microscopy (LSFM): past, present and future

The Analyst ◽  
2014 ◽  
Vol 139 (19) ◽  
pp. 4758-4768 ◽  
Author(s):  
John Lim ◽  
Hwee Kuan Lee ◽  
Weimiao Yu ◽  
Sohail Ahmed
Keyword(s):  
Fluorescence Microscopy ◽  
Imaging Modality ◽  
Light Sheet ◽  
Light Sheet Fluorescence Microscopy ◽  
Over Time

Light sheet fluorescence microscopy (LSFM) has emerged as an important imaging modality to follow biology in live 3D samples over time with reduced phototoxicity and photobleaching.

scholarly journals Wide field light-sheet microscopy with lens-axicon controlled two-photon Bessel beam illumination

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Sota Takanezawa ◽  
Takashi Saitou ◽  
Takeshi Imamura
Keyword(s):  
High Speed ◽  
Bessel Beam ◽  
Light Sheet ◽  
Time Lapse ◽  
Whole Body ◽  
Wide Field ◽  
Two Photon ◽  
Light Sheet Microscopy ◽  
Photon Excitation ◽  
Two Photon Excitation

AbstractTwo-photon excitation can lower phototoxicity and improve penetration depth, but its narrow excitation range restricts its applications in light-sheet microscopy. Here, we propose simple illumination optics, a lens-axicon triplet composed of an axicon and two convex lenses, to generate longer extent Bessel beams. This unit can stretch the beam full width at half maximum of 600–1000 μm with less than a 4-μm waist when using a 10× illumination lens. A two-photon excitation digital scanned light-sheet microscope possessing this range of field of view and ~2–3-μm axial resolution is constructed and used to analyze the cellular dynamics over the whole body of medaka fish. We demonstrate long-term time-lapse observations over several days and high-speed recording with ~3 mm3 volume per 4 s of the embryos. Our system is minimal and suppresses laser power loss, which can broaden applications of two-photon excitation in light-sheet microscopy.

scholarly journals Light-Sheet Fluorescence Microscopy with Scanning Non-diffracting Beams

2019 ◽  
Author(s):  
Hosein Kafian ◽  
Meelad Lalenejad ◽  
Sahar Moradi-Mehr ◽  
Shiva Akbari Birgani ◽  
Daryoush Abdollahpour
Keyword(s):  
Fluorescence Microscopy ◽  
Spatial Coherence ◽  
Three Dimensional ◽  
Light Sheet ◽  
Tissue Imaging ◽  
Wide Field ◽  
Self Healing ◽  
Main Challenge ◽  
Cancer Tumor ◽  
Light Sheet Fluorescence Microscopy

AbstractLight-sheet fluorescence microscopy (LSFM) has now become a unique technique in different fields ranging from three-dimensional (3D) tissue imaging to real-time functional imaging of neuronal activities. Nevertheless, obtaining high-quality artifact-free images from large, dense and inhomogeneous samples is the main challenge of the method that still needs to be adequately addressed. Here, we demonstrate significant enhancement of LSFM image qualities by using scanning non-diffracting illuminating beams, both through experimental and numerical investigations. The effect of static and scanning illumination with several beams are analyzed and compared, and it is shown that scanning 2D Airy light sheet is minimally affected by the artifacts, and provide higher contrasts and uniform resolution over a wide field-of-view, due to its reduced spatial coherence, self-healing feature and higher penetration depth. Further, the capabilities of the illumination scheme is utilized for both single and double wavelength 3D imaging of a large and dense mammospheres of cancer tumor cells as complex inhomogeneous biological samples.

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