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检索条件"任意字段=Three-Dimensional and Multidimensional Microscopy: Image Acquisition and Processing XIX"

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Hyperspectral microscopy as a particle exposure assessment tool 31

Hyperspectral microscopy as a particle exposure assessment t...

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three-dimensional and multidimensional microscopy: image acquisition and processing XXXI 2024

作者： Blanch-Perez Del Notario, Carolina Bertier, Pieter Jayapala, Murali Lambrechts, Andy Imec Kapeldreef 75 Leuven3001 Belgium Belgian Center for Occupational Hygiene BeCOH Brusselsesteenweg 46 Leuven3000 Belgium

ISBN: (纸本)9781510669550

Hyperspectral imaging combines the characteristics of computer vision and point spectroscopy by obtaining an image with both spatial and spectral information. Therefore, in combination with microscopy, it can increase material discrimination possibilities with respect to regular microscopy imaging. We explore this increased discrimination potential to assess exposure to particle contamination, since workplace exposure to specific particle materials poses well-known health hazards. In this respect, we are focusing on discriminating more health relevant particles such as silica in the respirable size fraction. For this purpose, a particle sampling protocol has been proposed and hyperspectral imaging in combination with transmission microscopy is used for particle material identification. We use a Snapscan visual near-infrared (VNIR) camera providing high spectral and spatial resolution in the 460-900 nm range, 150 spectral bands and up to 7 Mpixels of spatial resolution and high acquisition speed. The hyperspectral microscopy system has been tested for discrimination of fifteen different particle materials, such as silica, coal, dolomite, barite, or rutile, among others. The combined analysis of spatial and spectral information shows potential to accurately discriminate the 15 tested particle materials so far by means of a random forest classifier. In addition, a band relevance analysis is performed showing that only a few specific bands are needed to provide accurate discrimination of the tested materials. The hyperspectral hardware and method presented could lead to a faster exposure assessment than traditional techniques used for occupational exposure estimation. © COPYRIGHT SPIE. Downloading of the abstract is permitted for personal use only.

关键词： Health hazards

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Comparison of Optical Quadrature microscopy and Shack-Hartmann Wavefront Sensor

Comparison of Optical Quadrature Microscopy and Shack-Hartma...

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Conference on three-dimensional and multidimensional microscopy - image acquisition and processing XVI

作者： Patel, Yogesh G. Warger, William C., II Ballesta, Jerome DiMarzio, Charles A. Northeastern Univ Dept Elect & Comp Engn 440 Dana Bldg360 Huntington Ave Boston MA 02115 USA Imagine Optic SA F-914000 Orsay France

ISBN: (纸本)9780819474308

Several quantitative phase imaging techniques, such as digital holography, Hilbert-phase microscopy, and phase-shifting interferometry have applications in biological and medical imaging. Quantitative phase imaging measures the changes in the wavefront of the incident light due to refractive index variations throughout a 3-D specimen. We have developed a multimodal microscope which combines optical quadrature microscopy (OQM) and a Shack-Hartmann wavefront sensor for applications in biological imaging. OQM is an interferometric imaging modality that noninvasively measures the amplitude and phase of a signal beam that travels through a transparent specimen. The phase is obtained from interferograms with four different delayed reference wavefronts. The phase is then transformed into a quantitative image of optical path length difference. The Shack-Hartmann wavefront sensor measures the gradient of the wavefront at various points across a beam. A microlens array focuses the local wavefront onto a specific region of the CCD camera. The intensity is given by the maximum amplitude in the region and the phase is determined based on the exact pixel position within the region. We compare the amplitude and quantitative phase information of poly-methyl-meth-acrylate (PMMA) beads in oil and one-cell and two-cell mouse embryos with micrometer resolution using OQM and the Shack-Hartmann. Each pixel in OQM provides a phase measurement, whereas multiple pixels are used in Shack-Hartmann to determine the tilt. Therefore, the simple Shack-Hartmann system is limited by its resolution and field-of-view. Real-time imaging in Shack-Hartmann requires spatial averaging which smoothes the edges of the PMMA beads. The OQM has a greater field-of-view with good resolution;however, it is a complex system requiring multiple optical components and four cameras which may introduce additional artifacts in processing quantitative images. The OQM and Shack-Hartmann has certain advantages depending on

关键词： optical quadrature microscopy quantitative phase imaging shack-hartmann wavefront sensor

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Fluorescence confocal microscopy with spectrally-tuned, cryogenically-cooled diode lasers

Fluorescence confocal microscopy with spectrally-tuned, cryo...

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Conference on 3-dimensional and multidimensional microscopy - image acquisition and processing XII

作者： Sivers, N de Workeen, BV Lee, SA SUNY Albany MMM Lab Colonie NY 12205 USA

ISBN: (纸本)0819456756

In confocal microscopy, the laser excitation must be near the fluorochrome absorption peak to efficiently excite the fluorochrome. However, most fluorochromes emitting in the red to infrared have absorption and emission spectra with overlapping wavelength ranges, i.e. small Stokes shifts. As a result, the laser excitation extends into the fluorescence wavelengths hindering separation of the reflected laser signal from the information-containing fluorescence signal. Therefore, compromises are necessary: (1) the entire reflected laser spectrum is filtered, eliminating some of the fluorescence signal;or (2) the entire fluorescence signal is recorded, including unwanted reflected laser light, increasing the background noise. These compromises must be addressed, even with fluorochromes like Cy5 - having a relatively large Stokes shift. Cryogenically cooled diode lasers eliminate the need for these compromises by tuning the output wavelength away from the fluorochrome emission. By separating the excitation and emission spectra, the confocal fluorescence signal-to-noise ratio can be increased by filtering more of the reflected laser emission, without losing valuable fluorescence information. However, this results in slightly less efficient excitation of the fluorochrome. We will present spectrophotometric analyses of fluorochrome absorption, fluorescence, and diode laser emission as a function of diode operating temperature. We will show that as the diode lasers are cooled their output power increases and more than compensates for the lower fluorochrome excitation, resulting in significantly more intense fluorescence. Thus, by tuning the diode laser, more fluorescence information and less reflected laser light reach the detector, creating images with greater intensity and less background noise.

关键词： confocal microscopy laser diodes cryogenic cooling Cy5 fluorochrome

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Hybrid of two-photon microscopy and optical multimodality imaging for multi-scale imaging of small animals 25

Hybrid of two-photon microscopy and optical multimodality im...

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Conference on three-dimensional and multidimensional microscopy: image acquisition and processing XXV

作者： Li, Tianmeng Hui, Hui Ma, He Yang, Xin Tian, Jie Northeastern Univ Sinodutch Biomed & Informat Engn Sch Shenyang Liaoning Peoples R China Chinese Acad Sci Inst Automat Key Lab Mol Imaging Beijing 100190 Peoples R China Beijing Key Lab Mol Imaging Inst Automat Beijing 100190 Peoples R China

ISBN: (数字)9781510614840

ISBN: (纸本)9781510614840;9781510614833

Non-invasive imaging technologies, such as magnetic resonance imaging (MRI) and optical multimodality imaging methods, are commonly used for diagnosing and supervising the development of inflammatory bowel disease (IBD). These in vivo imaging methods can provide morphology changes information of IBD in macro-scale. However, it is difficult to investigate the intestinal wall in molecular and cellular level. State-of-art light-sheet and two-photon microscopy have the ability to acquire the changes for IBD in micro-scale. The aim of this work is to evaluate the size of the enterocoel and the thickness of colon wall using both MRI for in vivo imaging, and light-sheet and two-photon microscope for in vitro imaging. C57BL/6 mice were received 3.5% Dextran sodium sulfate (DSS) in the drinking water for 5 days to build IBD model. Mice were imaged with MRI on days 0, 6 to observe colitis progression. After MRI imaging, the mice were sacrificed to take colons for tissue clearing. Then, light-sheet and two-photon microscopies are used for in vitro imaging of the cleared samples. The experimental group showed symptoms of bloody stools, sluggishness and weight loss. It showed that the colon wall was thicker while the enterocoel was narrower compare to control group. The more details are observed using light-sheet and two-photon microscope. It is demonstrated that hybrid of MRI in macro-scale and light-sheet and two-photon microscopy in micro-scale imaging is feasible for colon inflammation diagnosing and supervising.

关键词： Multi-scale molecular imaging Inflammatory bowel disease Magnetic resonance imaging Two-photon microscopy Light-sheet microscopy

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Data compression in microscopy: a comparative study

Data compression in microscopy: a comparative study

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Proceedings of the 1999 three-dimensional and multidimensional microscopy: image acquisition and processing VI

作者： Kriete, Andres Klein, Norbert Berger, Lars C. Inst of Anatomy and Cell Biology Giessen Germany

ISBN: (纸本)0819430757

image compression in microscopy is a valuable technique, in particular if applied to multidimensional data. Information-preserving and competitive information-losing compression is applied to microscopical data and the resulting image quality is evaluated on a quantitative basis both in the spatial and frequency domain. Included are image data featuring different signal-to-noise ratios, but also voxel data for volume representations as well as graphical data for surface representations. The effect of compression on 3-D visualization and image management with data bases is included.

关键词： image compression

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Validation of Excitation-Scan Hyperspectral Multi-faceted Mirror Array Prototype System Advancements to Hyperspectral Imaging Applications 29

Validation of Excitation-Scan Hyperspectral Multi-faceted Mi...

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Conference on three-dimensional and multidimensional microscopy - image acquisition and processing Xxix

作者： Parker, Marina Mayes, Sam A. Browning, Craig M. Deal, Joshua Gunn-Mayes, Samantha Rich, Thomas C. Leavesley, Silas J. Univ S Alabama Chem & Biomol Engn Mobile AL 36688 USA Univ S Alabama Pharmacol Mobile AL 36688 USA Univ S Alabama Ctr Lung Biol Mobile AL 36688 USA Univ S Alabama Dept Syst Engn Mobile AL 36688 USA

ISBN: (纸本)9781510648043;9781510648036

Hyperspectral imaging technologies (HSI) have undergone rapid development since their beginning stages. While original applications were in remote sensing, other uses include agriculture, food safety and medicine. HSI has shown great utility in fluorescence microscopy for detecting signatures from many fluorescent molecules;however, acquisitions speeds have been slow due to light losses associated with spectral filtering. Therefore, we designed a novel light emitting diode (LED)-based rapid excitation scanning hyperspectral imaging platform allowing users to obtain simultaneous measurements of fluorescent labels without compromising acquisition speeds. Previously, we reported our results of the optical ray trace simulations and the geometrical capability of designing a multifaceted mirror imaging system as an initial approach to combine light at many wavelengths. The design utilized LEDs and a multifaceted mirror array to combine light sources into a liquid light guide. The computational model was constructed using Monte Carlo optical ray software (TracePro, Lambda Research Corp.). Recent prototype validation results show that when compared to a commercial emission scanning spectral confocal microscope (Zeiss-LSM-980), the novel LED-based excitation scanning HSI prototype successfully detected and separated six fluorescent labels from a custom 6-label African green monkey kidney epithelial cells. We report on the prototype's ability to overcome limitations of acquisition speeds, sensitivity, and specificity present in conventional systems. Future work will evaluate prototype's light losses to determine latent design modifications needed to demonstrate the system's feasibility as a promising solution for overcoming HSI acquisition speeds. This work was supported by NSF award MRI1725937.

关键词： Systems Design LED Tissue imaging HSI Spectroscopy microscopy Transmission Spectral Fluorescence Molecules Cells

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Normalized level set model for segmentation of low-contrast objects in 2- and 3-dimensional images 31

Normalized level set model for segmentation of low-contrast ...

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three-dimensional and multidimensional microscopy: image acquisition and processing XXXI 2024

作者： Baig, Mirza M. Junaid Wang, Yao L. Chung, Samuel H. Stepanyants, Armen Department of Physics Northeastern University 360 Huntington Ave BostonMA02115 United States Department of Bioengineering Northeastern University 360 Huntington Ave BostonMA02115 United States

ISBN: (纸本)9781510669550

Analyses of biomedical images often rely on accurate segmentation of structures of interest. Traditional segmentation methods based on thresholding, watershed, fast marching, and level set perform well in high-contrast images containing structures of similar intensities. However, such methods can under-segment or miss entirely low-intensity objects on noisy backgrounds. Machine learning segmentation methods promise superior performance but require large training datasets of labeled images which are difficult to create, particularly in 3D. Here, we propose an algorithm based on the Local Binary Fitting level set method and its application specifically designed to improve the segmentation accuracy for low-contrast structures even with significant noise levels present. The proposed algorithm, the Normalized Local Binary Fitting level set method, shows promise in enhancing the segmentation of low-contrast structures in biomedical images, addressing the limitations of traditional segmentation methods, and offering an alternative to machine learning approaches that require extensive training datasets. © COPYRIGHT SPIE. Downloading of the abstract is permitted for personal use only.

关键词： Machine learning

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Implementation of PSF Engineering in High-Resolution 3D microscopy Imaging with a LCoS (Reflective) SLM

Implementation of PSF Engineering in High-Resolution 3D Micr...

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Conference on three-dimensional and multidimensional microscopy - image acquisition and processing XXI

作者： King, Sharon V. Doblas, Ana Patwary, Nurmohammed Saavedra, Genaro Mart-Nez-Corral, Manuel Preza, Chrysanthe Univ Memphis Dept Elect & Comp Engn Memphis TN 38152 USA Univ Valencia Dept Opt 3D Imaging & Display Lab E-46100 Burjassot Spain

ISBN: (纸本)9780819498625

Wavefront coding techniques are currently used to engineer unique point spread functions (PSFs) that enhance existing microscope modalities or create new ones. Previous work in this field demonstrated that simulated intensity PSFs encoded with a generalized cubic phase mask (GCPM) are invariant to spherical aberration or misfocus;dependent on parameter selection. Additional work demonstrated that simulated PSFs encoded with a squared cubic phase mask (SQUBIC) produce a depth invariant focal spot for application in confocal scanning microscopy. Implementation of PSF engineering theory with a liquid crystal on silicon (LCoS) spatial light modulator (SLM) enables validation of WFC phase mask designs and parameters by manipulating optical wavefront properties with a programmable diffractive element. To validate and investigate parameters of the GCPM and SQUBIC WFC masks, we implemented PSF engineering in an upright microscope modified with a dual camera port and a LCoS SLM. We present measured WFC PSFs and compare them to simulated PSFs through analysis of their effect on the microscope imaging system properties. Experimentally acquired PSFs show the same intensity distribution as simulation for the GCPM phase mask, the SQUBIC-mask and the well-known and characterized cubic-phase mask (CPM), first applied to high NA microscopy by Arnison et al.(10), for extending depth of field. These measurements provide experimental validation of new WFC masks and demonstrate the use of the LCoS SLM as a WFC design tool. Although efficiency improvements are needed, this application of LCoS technology renders the microscope capable of switching among multiple WFC modes.

关键词： microscopy wave-front encoding spatial light modulators liquid crystal

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Investigation of second harmonic generation and multispectral imaging as new contrast mechanisms in scanning laser optical tomography 25

Investigation of second harmonic generation and multispectra...

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Conference on three-dimensional and multidimensional microscopy: image acquisition and processing XXV

作者： Nolte, Lena Antonopoulos, Georgios C. Heisterkamp, Alexander Ripken, Tammo Meyer, Heiko Laser Zentrum Hannover eV Ind & Biomed Opt Dept Hollerithallee 8 D-30419 Hannover Germany Leibniz Univ Hannover Inst Quantenopt Hannover Germany

ISBN: (数字)9781510614840

ISBN: (纸本)9781510614840;9781510614833

Scanning laser optical tomography (SLOT) is a 3D imaging technique, based on the principle of computed tomography to visualize samples up to magnitude of several centimeters. Intrinsic contrast mechanisms as absorption, scattering and autofluorescence provide information about the 3D architecture and composition of the sample. Another valuable intrinsic contrast mechanism is second harmonic generation (SHG), which is generated in non-centrosymmetric materials and commonly used to image collagen in biological samples. The angular dependence of the SHG signal, however, produces artifacts in reconstructed optical tomography datasets (OPT, SLOT). Thus, successful use of this intrinsic contrast mechanism is impaired. We investigate these artifacts by simulation and experiment and propose an elimination procedure that enables successful reconstruction of SHG-SLOT data. Nevertheless, in many cases specific labeling of certain structures is necessary to make them visible. Using multiple dyes in one sample can lead to crosstalk between the different channels and reduce contrast of the images. Also autofluorescence of the sample itself can account for that. By using multispectral imaging in combination with spectral unmixing techniques, this loss can be compensated. Therefore either a spectrally resolved detection path, or spectrally resolved excitation is required. Therefore we integrated a white supercontinuum light source in our SLOT-setup that enables a spectral selection of the excitation beam and extended the detection path to a four channel setup. This enables the detection of three fluorescence channels and one absorption channel in parallel, and increases the contrast in the reconstructed 3D images significantly.

关键词： Scanning Laser Optical Tomography 3D microscopy Tomographic imaging Nonlinear Optics Second Harmonic Generation Multispectral imaging Spectral unmixing

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Application of a novel 8×8 PMT-array detector to light microscopy

Application of a novel 8×8 PMT-array detector to light micr...

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Proceedings of three-dimensional and multidimensional microscopy: image acquisition and processing V

作者： Spring, Kenneth R. Kovbasnjuk, Olga N. Gibson, Carter C. Bungay, Peter M. National Institutes of Health 10/6N260 Bethesda MD 20892-1603 United States

A multi-anode photomultiplier tube (PMT) attached to a light microscope was used to measure the low-light-level signals from fluorescent dyes trapped in the extracellular spaces between living, cultured kidney epithelial cells. The detection assembly utilized photon counting of all 64 channels (maximum count rate 2 MHz) to record the fluorescence produced after the photoactivation of caged fluorophores of different molecular weight and charge. Photoactivation was accomplished by a brief (5ns, 0.5 mJ) pulse of light at 355 nm from a frequency-tripled Nd:YAG laser. Fluorescence of the uncaged fluorophores was excited by the output of an argon laser equipped with an acousto-optical tunable filter for control of wavelength and power. Diffusion coefficients for the fluorescent indicators in the extracellular spaces were calculated from the spatial and temporal decay of the fluorescence after uncaging of the dye in a small region (3 μm diameter spot) of the 30-90 μm-diameter microscope field. The system magnification was adjusted so that each 2.5-mm square PMT channel corresponded to a 12.5-μm square region of the microscope field. The spatial decay of the fluorescence was obtained by sampling multiple adjacent PMT channels, while the temporal decay was determined from the PMT channel encompassing the uncaging site.

关键词： Photomultipliers

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