摘要:

SUMMARYIn a three‐dimensional (3‐D) image data set obtained through optical sectioning, each two‐dimensional (2‐D) segment is blurred by out‐of‐focus information from neighbouring focal planes superimposed on the in‐focus segments from that plane. Instead of attempting to remove this redundant information over the full 3‐D data set, we have developed a technique for restoring stereoscopic views. In this paper we describe the implementation of a Wiener‐type inverse filtering method for generating stereo pairs of bright‐field micrographs. A theoretical optical transfer function valid under certain simplifying approximations has been used in implementing this filtering technique. In developing this method the slice theorem of computed tomography is used. In this way the image reconstruction problem is reduced to one of processing 2‐D arrays rather than 3‐D arrays and the problem of restoring missing Fourier components within the missing‐cone region is circumvented. Limited experimentation with real micrographs shows that the approach provides images that display an effective increased depth of field and 3‐D attributes of the specimen, even though some of the underlying assumptions on which this method is based are difficult to verify explicitly. The method can be implemented with a relatively fast executi

ISSN:0022-2720    

DOI:10.1111/j.1365-2818.1992.tb03256.x

出版商:Blackwell Publishing Ltd    

年代:1992

数据来源: WILEY

摘要:

SUMMARYThe optical transfer function of several scanning microscope systems is derived, using a physically intuitive approach. The technique allows a wide range of systems to be modelled with only minor modifications to the basic formulation.The results are then used to determine the response of various scanning microscopes for objects both in and out of the focal plane. The possibility of performing extended‐focus phase imaging in heterodyne microscopes by scanning the sample along the optical axis is also examined. This mode of operation should allow measurements of minute topographical and phase variations on tilted or warped samples with the same lateral resolution as would be obtained when the sample is in focus throughout the entire sca

ISSN:0022-2720    

DOI:10.1111/j.1365-2818.1992.tb03257.x

出版商:Blackwell Publishing Ltd    

年代:1992

数据来源: WILEY

摘要:

SUMMARYFast secondary electrons (FSE), which result from inelastic scattering of incident electrons, are known to generate a significant number of X‐rays for light elements, and also to degrade the spatial resolution of X‐ray microanalysis in thin foils. A Monte Carlo program simulating the generation and diffusion of FSE in binary systems has been developed to study the effect of composition on κABfactors and spatial resolution. The effect of accelerating voltage and thickness is also prese

ISSN:0022-2720    

DOI:10.1111/j.1365-2818.1992.tb03258.x

出版商:Blackwell Publishing Ltd    

年代:1992

数据来源: WILEY

摘要:

SUMMARYCryo‐fixed yeastParameciaand sea urchin embryos were investigated with an in‐lens type field‐emission SEM using a cold stage. The goal was to further develop and investigate the processing of frozen samples for the low‐temperature scanning electron microscope (LTSEM).Uncoated frozen‐hydrated samples were imaged with the low‐voltage backscattered electron signal (BSE). Resolution and contrast were sufficient to visualize cross‐fractured membranes, nuclear pores and small vesicles in the cytoplasm. It is assumed that the resolution of this approach is limited by the extraction depth of the BSE which depends upon the accelerating voltage of the primary beam (V0). In this study, the lowest possible (V0) was 2.6 kV because below this value the sensitivity of the BSE detector is insufficient. It is concluded that the resolution of the uncoated specimen could be improved if equipment were available for high‐resolution BSE imaging at 0.5–2kV.Higher resolution was obtained with platinum cryo‐coated samples, on which intramembranous particles were easily imaged. These images even show the ring‐like appearance of the hexagonally arranged intramembranous particles known from high‐resolution replica studies. On fully hydrated samples at high magnification, the observation time for a particular area is limited by mass loss caused by electron irradiation. Other potential sources of artefacts are the deposition of water vapour contamination and shrinkage caused by the sublimation of ice.Imaging of partially dehydrated (partially freeze‐dried) samples, e.g. high‐pressure frozenParameciumand sea urchin embryos, will probably become the main application in cell biology. In spite of possible shrinkage problems, this approach has a number of advantages compared with any other electron microscopy preparation method: no chemical fixation is necessary, eliminating this source of artefacts; due to partial removal of the water additional structures in the cytoplasm can be investigated; and finally, the mass loss due to electron beam irradiation is greatly reduced compared to full

ISSN:0022-2720    

DOI:10.1111/j.1365-2818.1992.tb03259.x

出版商:Blackwell Publishing Ltd    

年代:1992

数据来源: WILEY

摘要:

SUMMARYThe normally ‘condensed’ (darkly stained) chromosomes of dinofiagellates decondense by swelling. This occurs in an increasing number of cells when the concentration of added OsO4is decreased. With different fixatives other types of disintegration can be observed, which vary with the concentration. With cryofixation and freeze‐substitution the chromosomes are most ‘condensed’.Escherichia coltinfected with bacteriophage T4, with or without active lysozyme production, were studied by optical densitometry for partial lysis and by light and electron microscopy for observing swelling. When active lysozyme is present some of the acrolein (2.5%) ‐ glutaraldehyde (2%)‐fixed cells swell at 0°C, but do not in the absence of lysozyme nor when fixed at room temperature. If OsO4is added at concentrations ≤0.5%, partial lysis occurs when lysozyme is present. The optical density decreases, the cells lose some matter and swell slightly. The corresponding electron micrographs show gap formation by curdling and/or a decreased concentration of the cytoplasm which reveals certain phage

ISSN:0022-2720    

DOI:10.1111/j.1365-2818.1992.tb03260.x

出版商:Blackwell Publishing Ltd    

年代:1992

数据来源: WILEY

摘要:

SUMMARYA variation of the technique of encapsulating samples in agar cubes is presented. The samples are enclosed by bubbles in the agar. The hollow space is occupied by the specimen and a drop of melted agar is used to fill the remainder of the cavity and to cover it. The solidified agar is cut into a small cube containing the bubble, and is processed for TEM in the usual way.

ISSN:0022-2720    

DOI:10.1111/j.1365-2818.1992.tb03261.x

出版商:Blackwell Publishing Ltd    

年代:1992

数据来源: WILEY

ISSN:0022-2720    

DOI:10.1111/j.1365-2818.1992.tb03262.x

出版商:Blackwell Publishing Ltd    

年代:1992

数据来源: WILEY