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Scattering of an Airy light-sheet by a non-spherical particle using discrete dipole approximation
摘要： The scattering of an Airy light-sheet by a non-spherical particle is investigated using the discrete dipole approximation (DDA). The internal and near-surface fields of the cube placed in an Airy light-sheet are calculated, and the effects of the transverse dimension factor w0, the attenuation factor a, and beam center z0 are mainly discussed. w0 can increase the number of side lobes incident on the target, thereby increase the wave jet energy. Increasing the attenuation coefficient a leads to less wave jet energy. By moving the light-sheet, the side lobe of the incident light-sheet increases, and the wave jet energy increases. Potential applications include optical manipulation of particles, particle sizing, optical imaging, etc.
关键词： Scattering,Airy light-sheet,Angular spectrum decomposition method,Discrete dipole approximation (DDA)
更新于2025-09-23 15:23:52
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Probing Cell Mechanics with Bead-Free Optical Tweezers in the <em>Drosophila</em> Embryo
摘要： Morphogenesis requires coordination between genetic patterning and mechanical forces to robustly shape the cells and tissues. Hence, a challenge to understand morphogenetic processes is to directly measure cellular forces and mechanical properties in vivo during embryogenesis. Here, we present a setup of optical tweezers coupled to a light sheet microscope, which allows to directly apply forces on cell-cell contacts of the early Drosophila embryo, while imaging at a speed of several frames per second. This technique has the advantage that it does not require the injection of beads into the embryo, usually used as intermediate probes on which optical forces are exerted. We detail step by step the implementation of the setup, and propose tools to extract mechanical information from the experiments. By monitoring the displacements of cell-cell contacts in real time, one can perform tension measurements and investigate cell contacts' rheology.
关键词： Drosophila embryo,Developmental Biology,in vivo imaging,optical tweezers,Light sheet microscopy,force measurements,Issue 141,cell mechanics
更新于2025-09-23 15:21:01
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Analysis of aberrations and performance evaluation of adaptive optics in two-photon light-sheet microscopy
摘要： Two-photon light-sheet microscopy (TP-LSM) system performance is greatly degraded by specimen-induced aberrations in illumination path, which limit the field of view, axial resolution and excitation efficiency of the system. Adaptive optics (AO) is an effective method for attenuating these effects. For the design and evaluation of an AO system, a comprehensive analysis of the effects of aberrations is needed. In this paper, a TP-LSM system is simulated, and new indexes based on integral intensity are introduced for the evaluation of an aberrated light-sheet. Then, the influences of each Zernike mode and random aberrations on the illumination path of the TP-LSM system are investigated with a numerical simulation method. Results show that high-order aberrations have little effect on the axial resolution and excitation efficiency of the system and only low-order components require correction. The random aberrations varied in strength with the depth of the specimens, so the number of corrected Zernike modes is variable. A general formula is generated for the estimation of the number of modes that should be detected and corrected under different aberrations and different numerical aperture of the objective. The results can provide important guidance in the design and evaluation of AO units for TP-LSM systems.
关键词： Two-photon Light-sheet Microscopy,Aberration correction,Adaptive optics
更新于2025-09-23 15:21:01
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Restoration of Light Sheet Multi-View Data with the Huygens Fusion and Deconvolution Wizard
摘要： Light sheet fluorescence microscopy (LSFM) allows for high-resolution three-dimensional imaging with minimal photo-damage. By viewing the sample from different directions, different regions of large specimens can be imaged optimally. Moreover, owing to their good spatial resolution and high signal-to-noise ratio, LSFM data are well suited for image deconvolution. Here we present the Huygens Fusion and Deconvolution Wizard, a unique integrated solution for restoring LSFM images, and show that improvements in signal and resolution of 1.5 times and higher are feasible.
关键词： selective plane illumination microscopy (SPIM),Light sheet fluorescence microscopy (LSFM),deconvolution,Huygens,fusion
更新于2025-09-23 15:21:01
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Digital scanned laser lighta??sheet fluorescence lifetime microscopy with widea??field timea??gated imaging
摘要： We develop a multidimensional fluorescence imaging technique by implementing a wide-field time-gated fluorescence lifetime imaging into digital scanned laser light-sheet microscopy (FLIM-DSLM) to measure 3D fluorescence lifetime distribution in mesoscopic specimens with high resolution. This is achieved by acquiring a series of time-gated images at different relative time delays with respect of excitation pulses at different depths. The lifetime is determined for each voxel by iteratively fitting to single exponential decay. The performance of the developed system is evaluated with the measurements of a lifetime reference Rhodamine 6G solution and a sub-resolution fluorescent bead phantom. We also demonstrate the application performances of this system to ex vivo and in vivo imaging of Tg(kdrl:EGFP) transgenic zebrafish embryos, illustrating the lifetime differences between the GFP signal and the autofluorescence signal. The results show that FLIM-DSLM can be used for sample size up to a few millimeters and can be utilized as a powerful and robust method for biomedical research, for example as a readout of protein-protein interactions via F?rster resonance energy transfer.
关键词： Fluorescence lifetime imaging,light-sheet fluorescence microscopy,time-resolved fluorescence imaging
更新于2025-09-23 15:19:57
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Fast objective coupled planar illumination microscopy
摘要： Among optical imaging techniques light sheet fluorescence microscopy is one of the most attractive for capturing high-speed biological dynamics unfolding in three dimensions. The technique is potentially millions of times faster than point-scanning techniques such as two-photon microscopy. However light sheet microscopes are limited by volume scanning rate and/or camera speed. We present speed-optimized Objective Coupled Planar Illumination (OCPI) microscopy, a fast light sheet technique that avoids compromising image quality or photon efficiency. Our fast scan system supports 40 Hz imaging of 700 μm-thick volumes if camera speed is sufficient. We also address the camera speed limitation by introducing Distributed Planar Imaging (DPI), a scaleable technique that parallelizes image acquisition across cameras. Finally, we demonstrate fast calcium imaging of the larval zebrafish brain and find a heartbeat-induced artifact, removable when the imaging rate exceeds 15 Hz. These advances extend the reach of fluorescence microscopy for monitoring fast processes in large volumes.
关键词： Distributed Planar Imaging,calcium imaging,light sheet fluorescence microscopy,zebrafish brain,OCPI microscopy
更新于2025-09-16 10:30:52
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An azimuthally-modified linear phase grating: Generation of varied radial carpet beams over different diffraction orders with controlled intensity sharing among the generated beams
摘要： Diffraction gratings are important optical components and are used in many areas of optics such as in spectroscopy. A diffraction grating is a periodic structure that splits and diffracts the impinging light beam into several beams travelling in different directions. The diffracted beams from a grating are commonly called diffraction orders. The directions of the diffraction orders depend on the grating period and the wavelength of the impinging light beam so that a grating can be used as a dispersive element. In the diffraction of a plane wave from a conventional grating, the intensities of diffracted beams decrease with increasing order of diffraction. Here, we introduce a new type of grating where in the diffraction of a plane wave, the intensity of a given higher order diffracted beam can be higher than the intensity of the lower orders. We construct these gratings by adding an azimuthal periodic dependency to the argument of the transmission function of a linear phase grating that has a sinusoidal profile and we call them azimuthally-modified linear phase gratings (AMLPGs). In this work, in addition to introducing AMLPGs, we present the generation of varied radial carpet beams over different diffraction orders of an AMLPG with controlled intensity sharing among the generated beams. A radial carpet beam is generated in the diffraction of a plane wave from a radial phase grating. We show that for a given value of the phase amplitude over the host linear phase grating, one of the diffraction orders is predominant and by increasing the value of the phase amplitude, the intensity sharing changes in favor of the higher orders. The theory of the work and experimental results are presented. In comparison with the diffraction of a plane wave from radial phase gratings, the use of AMLPGs provides high contrast diffraction patterns and presents varied radial carpet beams over the different diffraction orders of the host linear phase grating. The resulting patterns over different diffraction orders are specified and their differences are determined. The diffraction grating introduced with controlled intensity sharing among different diffraction orders might find wide applications in many areas of optics such as optical switches. We show that AMLPG-based radial carpet beams can be engineered in which they acquire sheet-like spokes. This feature nominates them for potential applications in light sheet microscopy. In addition, a detailed analysis of the multiplication of the diffraction pattern of an AMLPG by the 2D structure of a spatial light modulator is presented. The presented theory is confirmed by respective experiments.
关键词： radial carpet beams,diffraction gratings,optical switches,light sheet microscopy,azimuthally-modified linear phase gratings
更新于2025-09-11 14:15:04
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Automated high-throughput light-sheet fluorescence microscopy of larval zebrafish
摘要： Light sheet fluorescence microscopy enables fast, minimally phototoxic, three-dimensional imaging of live specimens, but is currently limited by low throughput and tedious sample preparation. Here, we describe an automated high-throughput light sheet fluorescence microscope in which specimens are positioned by and imaged within a fluidic system integrated with the sheet excitation and detection optics. We demonstrate the ability of the instrument to rapidly examine live specimens with minimal manual intervention by imaging fluorescent neutrophils over a nearly 0.3 mm3 volume in dozens of larval zebrafish. In addition to revealing considerable inter-individual variability in neutrophil number, known previously from labor-intensive methods, three-dimensional imaging allows assessment of the correlation between the bulk measure of total cellular fluorescence and the spatially resolved measure of actual neutrophil number per animal. We suggest that our simple experimental design should considerably expand the scope and impact of light sheet imaging in the life sciences.
关键词： neutrophils,light sheet fluorescence microscopy,automated imaging,high-throughput imaging,larval zebrafish
更新于2025-09-10 09:29:36
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K-domain transform based three-dimensional microscopy
摘要： A K-domain transform based three-dimensional microscopy technique is proposed. By illuminating an object with a light sheet along the optical axis and recording the complex amplitude, including the modulus and the phase (or wave-front) of the re?ected light in the epi-direction, the structure of the illuminated slice of the specimen can be clearly reconstructed by transforming the re?ected light from the vertical plane to the axial plane. While the principle of this proposed technique is theoretically illustrated, its feasibility is veri?ed both numerically and experimentally. Because the illuminating and collecting optics comprise a coaxial imaging system, the proposed technique can achieve high-speed and high-resolution three-dimensional imaging with a simple optical setup, which can be realized using a common commercial microscope with only slight modi?cation.
关键词： complex amplitude,three-dimensional microscopy,high-resolution imaging,K-domain transform,light sheet
更新于2025-09-10 09:29:36
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[IEEE 2018 40th Annual International Conference of the IEEE Engineering in Medicine and Biology Society (EMBC) - Honolulu, HI, USA (2018.7.18-2018.7.21)] 2018 40th Annual International Conference of the IEEE Engineering in Medicine and Biology Society (EMBC) - Tracking Gene Expression via Light Sheet Microscopy and Computer Vision in Living Organisms
摘要： Automated tracking of spatiotemporal gene expression using in vivo microscopy images have given great insight into understanding developmental processes in multicellular organisms. Many existing analysis tools rely on the fluorescent tagging of cell wall or cell nuclei localized proteins to assess position, orientation, and overall shape of an organism; information necessary for determining locations of gene expression activity. Particularly in plants, organism lines that have fluorescent tags can take months to develop, which can be time consuming and costly. We propose an automated solution for analyzing spatial characteristics of gene expression without the necessity of fluorescent tagged cell walls or cell nuclei. Our solution indicates, segments, and tracks gene expression using a fluorescent imaging channel of a light sheet microscope while determining gene expression location within an organism from a Brightfield (non-fluorescent) imaging channel. We use the images obtained from the Arabidopsis thaliana root as a proof of concept for our solution by studying the effects of heat shock stress on CYCLIN B1 protein production.
关键词： computer vision,CYCLIN B1,light sheet microscopy,gene expression,Arabidopsis thaliana
更新于2025-09-09 09:28:46