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检索条件"任意字段=Conference on 3-Dimensional and Multidimensional Microscopy: Image Acquisition and Processing IX"

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Programmable beam-splitter for confocal laser scanning microscopy

Programmable beam-splitter for confocal laser scanning micro...

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conference on 3-dimensional and multidimensional microscopy: image acquisition and processing ix

作者： Birk, H Engelhardt, J Storz, R Hartmann, N Bradl, J Ulrich, H Leica Microsyst Heidelberg GMBH D-68165 Mannheim Germany

ISBN: (纸本)0819443603

In confocal microscopy manifold combinations of dyes are used. The entire wavelength range from near-ultraviolet to near-infrared is used to excite these dyes. Their emission is then detected primarily in the visible range. In addition to the number of dyes, the number of possible excitation laser lines is already large and constantly increasing. Due to the almost unlimited number of possible combinations, programmable devices are required. This is true for excitation-modules, beam-splitters and detection modules. Systems using filters with fixed spectral properties can practically fulfill the requirements for only a very small subset of applications. Programmable devices have been realized for excitation using Acousto Optical Tunable Filters (AOTF's). Freely definable spectral detectors are available for a perfect adaptation to the desired emission bands. The missing link for a completely filter-free design is now introduced using an Acousto Optical Beam Splitter (AOBS(TM)).

关键词： multi-spectral fluorescence detection Acousto-Optical Tunable Filter (AOTF) Acousto Optical Beam Splitter (AOBS((TM))) confocal laser scanning

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2-D and 3-D deconvolution of confocal fluorescence images by maximum likelihood estimation

2-D and 3-D deconvolution of confocal fluorescence images by...

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Three-dimensional and multidimensional microscopy: image acquisition and processing ix

作者： Larson, Jeffrey M. Nikon Instruments Inc. Melville NY 11747 United States

Deconvolution of confocal fluorescence images by maximum likelihood estimation (MLE) was investigated for its ability to increase the information content in the images. 3-D MLE algorithms, applied to confocal image stacks, increase lateral and axial resolution and result in a finer optical section. Contrast, especially at edges, is enhanced, improving the documentation quality of high magnification images beyond that possible by confocal microscopy alone. Axial smear associated with spherical aberration was not removed by deconvolution and a lateral thinning artifact was introduced. Single confocal images can be rapidly deconvolved by 2-D MLE by applying a two-dimensional point spread function and treating them as images of planar objects. The 2-D algorithm can also deconvolve a maximum projection of a stack. The method works best when there is a minimal overlap of fluorescent structures. The projection is treated as a two dimensional object. Intensity information excluded by the projection operation cannot be recovered. Deconvolution of images acquired with the pinhole opened to increase sensitivity closely matches images acquired through an optimal opening, although in 2-D MLE, colocalization cannot be distinguished from overlap and the integrity of quantitative data cannot be guaranteed. Properly applied, MLE deconvolution increases the useful information content of confocal images.

关键词： image processing

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Numerical simulation of image formation in near-field optical microscopy

Numerical simulation of image formation in near-field optica...

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Three-dimensional and multidimensional microscopy: image acquisition and processing ix

作者： Baibyrin, Vil B. Volkov, Uryi P. Avetisyan, Uryi A. Krasnikova, Irina V. Bespalova, Natalija V. Saratov State Technical University Politechnitcheskaja 77 Saratov 410007 Russia

Numerical simulation of image formation in near field optical microscopy is needed to understand a relationship between near field images and actual structure of sample since the image can be differs strongly from real structure. In order to estimate the near field image formation, two different approaches are used, namely numerical solution of integral equations with boundary element method or moment method and analytical solution of light scattering problem obtained by assumption of some hypotheses. The first approach results in reasonably complicated algorithm of calculation and often causes a bed conditioned system of linear equations to be formed. The approach based on an analytical solution of scattering problem is more preferable since allow to reduce the algorithm and increase the calculations speed. A new program of simulation of near field image formation for arbitrary sample shape has been developed. The program is based on the results of analytical calculation of near field light diffraction on sinusoidal diffraction grating of arbitrary corrugation amplitude. The sample shape is presented as a number of sinusoidal gratings of various amplitudes by Fourier transformation, for each of the gratings the light scattering amplitude is calculated and the amplitudes are summarized.

关键词： Near field scanning optical microscopy

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Standards in digital microscopy

Standards in digital microscopy

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

作者： Kriete, A Shah, S Inst Anat & Cell Biol IPL D-35385 Giessen Germany

This paper discusses some aspects of standardization in digital microscopy. Included are image acquisition, use of image and graphics formats and data protocols in multidimensions, particular image analysis procedures... 详细信息

ISBN: (纸本)0819439398

关键词： Standards microscopy image acquisition image formats image processing

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Point spread functions for particle imaging using inhomogeneous polarization in scanning optical microscopy

Point spread functions for particle imaging using inhomogene...

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

作者： Youngworth, KS Biss, DP Brown, TG Univ Rochester Inst Opt Rochester NY 14627 USA

ISBN: (纸本)0819439398

Polarization control and analysis continue to be a useful tools for microscopy. We have previously addressed the use of spatially inhomogeneous polarization, specifically azimuthal polarization, in scanning optical microscopy. In this paper we describe new techniques for generating both azimuthally and radially polarized cylindrical-vector beams, investigate high numerical aperture focusing, compare the theoretical and experimental point spread functions for small particle imaging, and explore the use of these beams with microscopy.

关键词： radial polarization azimuthal polarization inhomogeneous polarization cylindrical-vector beams axial electric field high numerical aperture focusing scanning optical microscopy point spread function small particle imaging

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Inverse scattering with subwavelength resolution in the near-zone

Inverse scattering with subwavelength resolution in the near...

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

作者： Carney, PS Schotland, JC Washington Univ Dept Elect Engn St Louis MO 63130 USA

ISBN: (纸本)0819439398

In classical microscopy and inverse scattering, images are made from far field measurements. It will be shown that the standard Abbe or Rayleigh resolution limits arise as a result of assumptions which do not hold in the near field. Existing technologies such as TIRM and PSTM that exploit these properties of the near field produce two dimensional images which exhibit structure on a subwavelength scale. These images have proven difficult to interpret when the sample presents variations in structure and optical properties simultaneously in three dimensions. We present the analytic singular value decomposition of the linearized scattering operator. This representation leads to a robust inversion algorithm for the inverse scattering problem in the near zone. Numerical implementation of the algorithm is demonstrated using simulated noisy data to produce tomographic images of a three dimensional scatterer.

关键词： near-field optics inverse scattering microscopy tomography

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Automated 3D dendritic spine detection and analysis from two-photon microscopy

Automated 3D dendritic spine detection and analysis from two...

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

作者： Koh, IYY Lindquist, WB SUNY Stony Brook Dept Appl Math & Stat Stony Brook NY 11794 USA

ISBN: (纸本)0819439398

The functional significance of dendritic spines and their plasticity to a wide spectrum of developmental and pathological conditions has led to extensive studies based on spine morphology. The advances in image acquisition techniques and the associated generation of large 3D data sets of optical micrographs have not been accompanied by comparable advances in data analysis techniques. We present an automated 3D spine detection and quantification procedure suitable for images obtained by laser scanning microscopy. The image is first processed by deconvolution and the dendritic phase consisting of the neuronal cytoplasm is extracted by segmentation. Spines are detected as geometrical protrusions relative to the dendritic backbone, ks very thin necks may not be imaged, some spine 'heads' may be detached from the dendrite and are detected as detached components. These detected heads are merged with spine 'bases' where appropriate. Morphological characterizations on spine length, volume, density and shape classifications are obtained. For time-lapse data, images are registered and individual spines are traced through the image sequence. Successful comparison results on spine lengths and densities with manual analysis are obtained. This method is highly automatic and allows detailed and objective quantification of the structure and dynamics of dendritic spines, which can be important predictors for the function of neural networks.

关键词： image processing dendrite spine two-photon medial axis laser scanning microscopy

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Phase retrieval of widefield microscopy point spread functions

Phase retrieval of widefield microscopy point spread functio...

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

作者： Hanser, BM Gustafsson, MGL Agard, DA Sedat, JW Univ Calif San Francisco Grad Grp Biophys San Francisco CA 94143 USA

ISBN: (纸本)0819439398

The three-dimensional imaging properties of a light microscope are traditionally described through an intensity point spread function (PSF) or its Fourier transform, the optical transfer function (OTF). However, the imaging properties can be more compactly described by a two-dimensional generalized pupil function. Use of the pupil function allows easy modification of an observed PSF to introduce known aberrations, a task that is much more difficult when using a PSF or OTF. Unfortunately, it is not straightforward to determine the complex-valued pupil function from the measured intensity PSF, because of the lack of phase information. This is the problem of phase retrieval. Several phase retrieval algorithms have been developed for two-dimensional imaging in astronomy. We have modified one such algorithm to be appropriate for the high-aperture, non-paraxial case of high resolution light microscopy. PSFs reconstructed from phase-retrieved pupil functions, modified by calculated aberrations closely reproduce the features of measured, aberrated PSFs.

关键词： phase retrieval Point Spread Function (PSF) wide-field microscopy fluorescence microscopy Optical Transfer Function (OTF) phase diversity optical microscopy pupil function

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Three-dimensional imaging with a quadrature tomographic microscope

Three-dimensional imaging with a quadrature tomographic micr...

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

作者： Stott, JJ Bennett, RE Warner, CM DiMarzio, CA Northeastern Univ Ctr Subsurface Sensing & Imaging Syst Boston MA 02115 USA

ISBN: (纸本)0819439398

The quadrature tomographic microscope combines conventional microscopy with interferometric quadrature detection techniques to yield a simultaneous measurement of both the amplitude and phase of the electric field. This is of tremendous utility in imaging nearly-transparent objects since changes in phase can be directly attributed to changes in the object's index of refraction while the small amplitude changes provide additional structural information. In this paper we will discuss the progress we have made towards reconstructing three dimensional images from the measured two-dimensional complex electric field. BS applying an artificial curvature to the measured field and back-propagating the Gaussian beams, we are able to digitally manipulate the imaging kernel and image different focal planes.

关键词： microscopy optical quadrature tomography quantitative biology

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Programmable array microsope employing two ferroelectric liquid crystal spatial light modulators

Programmable array microsope employing two ferroelectric liq...

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

作者： Taylor, CM McCabe, EM Univ Dublin Trinity Coll Dept Phys Dublin 2 Ireland

ISBN: (纸本)0819439398

We present a programmable array microscope (PAM) which uses a pair of ferroelectric liquid crystal spatial light modulators (FLC SLMs). The system is similar to a confocal imaging system called a direct-view microscope (DVM), in which scanned aperture arrays are used to obtain real-time confocal images. The PAM, unlike the DVM, allows arrays to be scanned electronically rather than mechanically. In our system, one SLM is placed in the source plane of a conventional microscope system;the other is placed in the detector plane. Confocal aperture arrays are displayed and scanned synchronously on the SLMs and confocal imaging results. The resolution is improved when compared to a similar previously presented system which employed a single SLR I. We present axial resolution measurements for a variety of array dimensions and investigate the use of aperture correlation techniques to improve the light throughput in the devices.

关键词： programmable array microscopy ferroelectric liquid crystal spatial light modulator confocal microscope direct-view microscopy

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