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Three-dimensional image study on the vascular structure after angiopoietin-1 transduction in isolated mouse pancreatic islets

Three-dimensional image study on the vascular structure afte...

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conference on Three-dimensional and multidimensional microscopy - image acquisition and processing XV

作者： He, Jing Su, Dongming Trucco, Massimo Univ Pittsburgh Sch Med Dept Pediat Div ImmunogenetRangos Res Ctr 5th Ave 3460 Pittsburgh PA 15213 USA

ISBN: (纸本)9780819470362

Angiopoletin-1 (Ang-1) is essential for remodeling the primitive vascular plexus during embryonic development and for reducing plasma leakage in inflammation of adult vasculature. However, the role for Ang-1 in maintenance of vascular stability in isolated pancreatic islets is not fully understood. In this study, we compared the difference of vascular morphology between Ang-1 treated (n=5) and control mouse islets (n=5) using both two- and three-dimensional optical image analysis. Isolated mouse islets were transduced with Ang-1 or Lac Z (control) vector at 37 degrees C for 16 hours. Islets were incubated with both rat anti-CD31 antibody and rabbit anti-insulin antibody followed by incubation with Rhodamine-conjugated goat anti-rat IgG and Alexa-488 conjugated goat anti-rabbit IgG. Islets were viewed under a Nikon confocal microscope. Serial optical section images were captured and reconstructed using Nikon EZ-C1 software. Individual two-D and reconstructed three-D images were analyzed using MetaMorph image Analysis software. Islet vascular density was determined. In two-D images, there was no significant difference of vascular density between the two groups. The vascular morphology didn't show any obvious differences in two-D images either. However, in the three-D images, we found higher vascular density and more vascular branches in the Ang-1 transducted islets and vascular dilation in control group. In conclusion, using three-D image analysis, Ang-1 displayed functions in maintenance of vascular stability and in stimulating growth of vascular branches in isolated mouse pancreatic islets. In order to study further the regeneration of different cell contents in the spherical pancreatic islet, three-D image analysis is an effective method to approach this goal.

关键词： pancreatic islet three-D image two-D image 3D-reconstruction Ang-1 vascular structure confocal microscope mouse pancreas diabetics transplantation

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Confocal photoluminescence microscopy in II-VI materials: annealing and degradation dynamics

Confocal photoluminescence microscopy in II-VI materials: an...

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conference on 3-dimensional and multidimensional microscopy - image acquisition and processing VI

作者： McCabe, EM Jordan, C Fewer, DT Donegan, JF Taniguchi, S Hino, T Nakano, K Ishibashi, A Uusimaa, P Pessa, M Univ Dublin Trinity Coll Dublin 2 Ireland

ISBN: (纸本)0819430757

Confocal photoluminescence imaging is an important tool in the investigation of recombination in semiconductors and in the characterisation of material growth. This characterisation is particularly important for II-VI wide band-gap semiconductors where the potential for blue-green lasers is being explored currently. To achieve room-temperature cw operation of these lasers over the multi-thousand hours necessary for commercialisation, extremely low defect densities are required. The confocal microscope is used in this work to image photoluminescence from II-VI materials to characterise the defect formation and propagation within the quantum well region of the material. This imaging approach permits the degradation to be monitored in real time and over a large area in samples with low defect densities. The additional advantages of this set-up over a conventional microscope are, of course, the higher lateral resolution and narrow depth of field associated with a confocal microscope. While considerable effort has been focused on the degradation in these II-VI semiconductors, we have recently observed that annealing can occur simultaneously in the same sample when the material is exposed to intense optical excitation. images of annealing and degradation of a range of II-VI samples will be presented to highlight these observations.

关键词： confocal microscopy photoluminescence imaging semiconductors II-VI materials blue green lasers

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Efficient Confocal microscopy with a Dual-Wedge Scanner

Efficient Confocal Microscopy with a Dual-Wedge Scanner

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conference on Three-dimensional and multidimensional microscopy - image acquisition and processing XVI

作者： Warger, William C., II Guerrera, Stephen A. Eastman, Zachary DiMarzio, Charles A. Northeastern Univ Dept Elect & Comp Engn 360 Huntington Ave Boston MA 02115 USA Lucid Inc Rochester NY 14623 USA Massachusetts Gen Hosp Boston MA 02114 USA MIT Cambridge MA 02139 USA

ISBN: (纸本)9780819474308

Confocal microscopes achieve high spatial resolution by focusing both a light source and a detector to a single point with an objective having a high numerical aperture. In order to produce an image, it is then necessary to scan this common focal spot through the specimen, and it is often important to image the full field of view in a short time. In order to avoid vignetting, the scanning must be done in or near the pupil of the optical system. For some fast scanners, this requires the use of multiple relay telescopes to create multiple pupils. Galvanometric scanners impose a practical limit on the scan speed because of the angular accelerations involved in reversing direction. Rotating polygons are often used to achieve greater speed. For a scanner consisting of a rotating polygon and a galvanometric scanner, two relay telescopes are normally used. We have developed a dual-wedge scanner which has the potential to perform the scan in a configuration which is very short in the axial direction, thereby eliminating much of the complexity of current high-speed scanners. We have demonstrated a prototype of the scanner in a reflectance confocal microscope. Transverse and axial resolution are comparable to those of other scanning systems. The selection of rotation speeds for the wedges is important to ensuring full coverage of the field of view in a reasonable time. Various tradeoffs on these parameters will be discussed. The beam behavior in the pupil will be discussed. Resolution limits and aberrations will be shown from ray-tracing analysis, and compared to experimental results.

关键词： Prisms Scanners Confocal microscopy Microscopes Objectives Polygon scanners Raster graphics Skin Telescopes Reflectivity

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Feature extraction and 3D reconstruction of Nomarski DIC microscope images using local energy and neural net techniques

Feature extraction and 3D reconstruction of Nomarski DIC mic...

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conference on 3-dimensional and multidimensional microscopy - image acquisition and processing VI

作者： Kyan, M Guan, L Arison, M Cogswell, C Univ Sydney Dept Elect Engn Sydney NSW 2006 Australia

ISBN: (纸本)0819430757

The Nomarski differential interference contrast (DIC) mode is commonly used for imaging translucent biological specimens and it exhibits several major advantages over other phase contrast techniques, including a boost in high spatial frequencies in the region of focus. However, DIC images (unlike confocal) are limited by the presence of low spatial frequency blur and also by a differential shading gradient at feature boundaries, which make normal confocal visualisation techniques unsuitable for feature extraction or for 3D volume rendering of focus-series datasets. To remedy this problem, we employ a neural network technique based on competitive learning, known as Kohonen's self-organising feature map (SOFM), to perform segmentation, using a collection of statistics (know as features) defining the image. Our past investigation showed that standard features such as the localised mean and variance of pixel intensities provided reasonable extraction of objects such asmitotic chromosomes, but surface detail was only moderately resolved. In this current work, local energy is investigated as an alternative image statistic based on phase congruency in the image. This, along with different combinations of other image statistics, is applied in a SOFM, producing 3D images exhibiting vast improvement in the level of detail and clearly isolating the chromosomes from the background.

关键词： differential interference contrast DIC feature extraction feature space image segmentation self organising feature map SOFM phase congruency local energy Morlet wavelet

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Simple and fast spectral domain algorithm for quantitative phase imaging of living cells with digital holographic microscopy 24

Simple and fast spectral domain algorithm for quantitative p...

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conference on Three-dimensional and multidimensional microscopy - image acquisition and processing XXIV

作者： Min, Junwei Yao, Baoli Ketelhut, Steffi Kemper, Bjoern Univ Munster Biomed Technol Ctr Mendelstr 17 D-48149 Munster Germany Chinese Acad Sci Xian Inst Opt & Precis Mech State Key Lab Transient Opt & Photon Xian 710119 Shaanxi Peoples R China

ISBN: (数字)9781510605824

ISBN: (纸本)9781510605817;9781510605824

The modular combination of optical microscopes with digital holographic microscopy (DHM) has been proven to be a powerful tool for quantitative live cell imaging. The introduction of condenser and different microscope objectives (MO) simplifies the usage of the technique and makes it easier to measure different kinds of specimens with different magnifications. However, the high flexibility of illumination and imaging also causes variable phase aberrations that need to be eliminated for high resolution quantitative phase imaging. The existent phase aberrations compensation methods either require add additional elements into the reference arm or need specimen free reference areas or separate reference holograms to build up suitable digital phase masks. These inherent requirements make them unpractical for usage with highly variable illumination and imaging systems and prevent on-line monitoring of living cells. In this paper, we present a simple numerical method for phase aberration compensation based on the analysis of holograms in spatial frequency domain with capabilities for on-line quantitative phase imaging. From a single shot off-axis hologram, the whole phase aberration can be eliminated automatically without numerical fitting or pre-knowledge of the setup. The capabilities and robustness for quantitative phase imaging of living cancer cells are demonstrated.

关键词： Digital holographic microscopy aberration compensation frequency spectrum analysis quantitative phase imaging quantitative live cell analysis

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image processing in 3-D standing-wave fluorescence microscopy

Image processing in 3-D standing-wave fluorescence microscop...

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conference on Three-dimensional microscopy - image acquisition and processing III

作者： Krishnamurthi, V Bailey, B Lanni, F CARNEGIE MELLON UNIV CTR LIGHT MICROSCOPE IMAGING & BIOTECHNOLDEPT PHYSBIOMED ENGN PROGRAMPITTSBURGHPA 15213

ISBN: (纸本)0819420298

Standing-wave fluorescence microscopy, a method which utilizes interference to create a periodic excitation pattern along the optical axis, has been shown to provide improved axial resolution in thin, fluorescently labeled specimens. In each plane of focus, a complete standing wave data set is obtained by acquiring an image at each of three distinct positions of the interference fringes. Thicker specimens require through-focus data consisting of three images per plane. In this report we describe the recovery of information from this data using 3D image processing. The effective optical transfer function (OTF) of the standing wave microscope consists of the conventional OTF and two sidebands which are copies of the conventional OTF shifted axially by the spatial frequency of the interference fringes. The large gaps between the central band and the sidebands lead to significant ringing in the 3D reconstruction if linear deconvolution methods are employed. The use of non-linear, constrained image processing techniques has been shown to allow accurate extrapolation outside the OTF band limit. We demonstrate the extent to which the sidebands enhance recovery of information in the gaps, and provide a comparison between deconvolution using inverse-filtering and maximum-likelihood estimation.

关键词： Composites Fermium Frequency modulation Optical transfer functions image processing Luminescence Microscopes microscopy 3D image processing Point spread functions

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Excitation-scan Mirror Array System Advancements to Hyperspectral Imaging Applications 27

Excitation-scan Mirror Array System Advancements to Hyperspe...

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conference on Three-dimensional and multidimensional microscopy - image acquisition and processing XXVII

作者： Browning, Craig M. Mayes, Sam A. Deal, Joshua Gunn-Mayes, Samantha Rich, Thomas C. Leavesley, Silas J. Parker, Marina 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)9781510632547

Hyperspectral imaging (HSI) technology has been applied in a range of fields for target detection and mixture analysis. While its original applications were in remote sensing, modern uses include agriculture, historical document authentications and medicine. HSI has shown great utility in fluorescence microscopy;however, acquisition speeds have been slow due to light losses associated with spectral filtering. We are currently developing a rapid hyperspectral imaging platform for 5-dimensional imaging (RHIP-5D), a confocal imaging system that will allow users to obtain simultaneous measurements of many fluorescent labels. We have previously reported on optical modeling performance of the system. This previous model investigated geometrical capability of designing a multifaceted mirror imaging system as an initial approach to sample light at many wavelengths. The design utilized light-emitting diodes (LEDs) and a multifaceted mirror array to combine light sources into a liquid light guide (LLG). The computational model was constructed using Monte Carlo optical ray software (TracePro, Lambda Research Corp.). Recent results presented here show transmission has increased up to 9% through parametric optimization of each component. Future work will involve system validation using a prototype engineered based on our optimized model. System requirements will be evaluated to determine if potential design changes are needed to improve the system. We will report on-spectral resolution to demonstrate feasibility of the RHIP-5D as a promising solution for overcoming current HSI acquisition speed and sensitivity limitations.

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

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Principles for modeling and functional simulation of biological microstructures

Principles for modeling and functional simulation of biologi...

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conference on Three-dimensional microscopy - image acquisition and processing IV

作者： Kriete, A Univ. of Giessen (Germany)

ISBN: (纸本)0819423955

This paper discusses some aspects of computer based modeling of biological microstructures. The workflow to model and simulate a biological structure is described as a feedback - loop. Beside the system definition by structural and dynamical properties, the simulation is discussed as a mathematical representation coupled with a computer visualization. As an example, the investigation of the functional behavior of lung structures is described with special emphasis to the modeling of respiratory units.

关键词： Mathematical modeling Computer simulations Lung 3D modeling Visualization 3D acquisition Data modeling Process modeling Systems modeling Convection

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A novel method for image denoising of fluorescence molecular imaging based on fuzzy C-Means clustering 22

A novel method for image denoising of fluorescence molecular...

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conference on Three-dimensional and multidimensional microscopy - image acquisition and processing XXII

作者： An, Yu Liu, Jie Ye, Jinzuo Mao, Yamin Yang, Xin Jiang, Shixin Chi, Chongwei Tian, Jie Beijing Jiaotong Univ Sch Comp & Informat Technol Dept Biomed Engn Beijing 100044 Peoples R China Inst Chinese Acad Sci Key Lab Mol Imaging Beijing 100190 Peoples R China

ISBN: (纸本)9781628414202

As an important molecular imaging modality, fluorescence molecular imaging (FMI) has the advantages of high sensitivity, low cost and ease of use. By labeling the regions of interest with fluorophore, FMI can noninvasively obtain the distribution of fluorophore in-vivo. However, due to the fact that the spectrum of fluorescence is in the section of the visible light range, there are mass of autofluorescence on the surface of the bio-tissues, which is a major disturbing factor in FMI. Meanwhile, the high-level of dark current for charge-coupled device (CCD) camera and other influencing factor can also produce a lot of background noise. In this paper, a novel method for image denoising of FMI based on fuzzy C-Means clustering (FCM) is proposed, because the fluorescent signal is the major component of the fluorescence images, and the intensity of autofluorescence and other background signals is relatively lower than the fluorescence signal. First, the fluorescence image is smoothed by sliding-neighborhood operations to initially eliminate the noise. Then, the wavelet transform (WLT) is performed on the fluorescence images to obtain the major component of the fluorescent signals. After that, the FCM method is adopt to separate the major component and background of the fluorescence images. Finally, the proposed method was validated using the original data obtained by in vivo implanted fluorophore experiment, and the results show that our proposed method can effectively obtain the fluorescence signal while eliminate the background noise, which could increase the quality of fluorescence images.

关键词： image denoising fluorescence molecular imaging fuzzy C-Means clustering

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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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