摘要:

SUMMARYFibrous structures like polymers, glass fibres, muscle fibres and capillaries are important components of materials and tissues. A spatial fibre process is the union of smoothly curved or linear one‐dimensional features of finite length, arranged in an unbounded three‐dimensional reference space according to some random mechanism. Design‐based stereology was combined with confocal scanning laser microscopy to study samples of fibre‐reinforced composites, which were considered as realizations of not necessarily isotropic fibre processes. The methods enable the unbiased estimation of the intensity and of the directional distribution of spatial fibre processes from arbitrarily directed pairs of registered parallel optical sections a known distance apart. The directions of fibres sampled by a frame of observation on the reference plane are estimated from the coordinates of the intersection points of the fibres with both optical planes using confocal scanning laser microscopy. The true directional distribution of the fibre process is estimated by weighting each measured direction by the reciprocal of its chance of being sampled, which can be inferred from the data. The concept of complete directional randomness for uniformly and independently distributed spatial directions is introduced. The cumulative distribution function of the angular distances between different directions and other exact relations are derived for complete randomness of vectorial and axial directions. A Monte Carlo method is constructed to test spatial fibre processes, whose fibres have negligibly small curvature, for complete directional randomness. Confocal scanning laser microscopy was used to study the angular distribution of glass fibres in a polymer composite which was subjected to increasing hydrostatic extrusion. The hypothesis of complete directional randomness had to be rejected for all samples with 1% probability of error. The directional distribution was of the bipolar type, with the principal axis directed parallel to the axis of extrusion. Progressive stretching of the material increased the degree of anisotropy of the glass fibres. Although presented for an application in polymer physics, the methods are general and may also be applied in biological investi

ISSN:0022-2720    

DOI:10.1111/j.1365-2818.1994.tb03432.x

出版商:Blackwell Publishing Ltd    

年代:1994

数据来源: WILEY

摘要:

SUMMARYThree‐dimensional (3‐D) image analysis algorithms and experimental results that demonstrate the feasibility of fully automated tracing of neurons from fluorescence confocal microscopy data are presented. The input to the automated analysis is a set of successive optical slices that have been acquired using a confocal scanning laser microscope. The output of the system is a labelled graph representation of the neuronal topology that is spatially aligned with the 3‐D image data. A variety of topological and metric analyses can be carried out using this representation. For instance, precise measurements of volumes, lengths, diameters and tortuosities can be made over specific portions of the neuron that are specified in terms of the graph representation. The effectiveness of the method is demonstrated for a set of sample fields featuring selectively stained neurons. Additional work will be needed to refine the method for unsupervised use with complex data involving multiple intertwined neurons and extremely fine dendritic struc

ISSN:0022-2720    

DOI:10.1111/j.1365-2818.1994.tb03433.x

出版商:Blackwell Publishing Ltd    

年代:1994

数据来源: WILEY

摘要:

SUMMARYMethods are presented for the automated, quantitative and three‐dimensional (3‐D) analysis of cell populations in thick, essentially intact tissue sections while maintaining intercell spatial relationships. This analysis replaces current manual methods which are tedious and subjective.The thick sample is imaged in three dimensions using a confocal scanning laser microscope. The stack of optical slices is processed by a 3‐D segmentation algorithm that separates touching and overlapping structures using localization constraints. Adaptive data reduction is used to achieve computational efficiency. A hierarchical cluster analysis algorithm is used automatically to characterize the cell population by a variety of cell features. It allows automatic detection and characterization of patterns such as the 3‐D spatial clustering of cells, and the relative distributions of cells of various sizes. It also permits the detection of structures that are much smaller, larger, brighter, darker, or differently shaped than the rest of the population. The overall method is demonstrated for a set of rat brain tissue sections that were labelled for tyrosine hydroxylase using fluorescein‐conjugated antibodies. The automated system was verified by comparison with computer‐assisted manual counts from the same i

ISSN:0022-2720    

DOI:10.1111/j.1365-2818.1994.tb03434.x

出版商:Blackwell Publishing Ltd    

年代:1994

数据来源: WILEY

摘要:

SUMMARYDigital imaging fluorescence microscopy (DIFM) of tissue sections was used to quantify uptake of labelled plasma proteins by the arterial wall. Several aspects of the measuring system were investigated so that absolute tracer concentrations and their local variation could be derived from digitized images. These investigations may be relevant to other studies employing DIFM.Nonlinearities were found to arise from offsets in the video digitizers, from background fluorescence and stray light within the microscope and from the transfer characteristics of the intensified CCD camera. Camera gain controls showed complex behaviour. Camera output fell substantially for several hours after switching on and was affected by room temperature. Large spatial variations in response were caused by the geometry of the microscope optics and by the image intensifier. However, the ratios between areas were not affected by light intensity or camera gain settings. Measured intensities were independent of the depthwise location of fluorophores within tissue sections but they were affected by the emission from objects outside the measuring area. Photobleaching of tracer varied significantly over the range of excitation intensities and durations used but was not concentration dependent. Methods used to correct these effects and obtain a uniform, linear and constant relationship between concentration and grey level are described.Using the system and appropriate corrections,in vivouptake of sulphorhodamine‐B‐labelled serum albumin by the rabbit aortic wall was investigated. Results obtained for the mean uptake of tracer and its local variation were quantitatively similar to those previously obtained with nonmicroscopic meth

ISSN:0022-2720    

DOI:10.1111/j.1365-2818.1994.tb03435.x

出版商:Blackwell Publishing Ltd    

年代:1994

数据来源: WILEY

摘要:

SUMMARYMixtures of ethanol, 2‐propanol and 2‐butanol can be used as a cryoglue to mount vitrified biological specimens for ultrathin cryosectioning. Brought directly from room temperature to a cutting support at 140K in the cold chamber of a cryoultramicrotome, these alcohols stiffen to a viscous and gluey consistency allowing the attachment or embedding of a vitreous biological sample. The mass hardens at lower temperatures fixing the sample well for microtomy. With ethanol: 2‐propanol (2:3), samples are applied at 140 K and ultrathin cutting can be done at

ISSN:0022-2720    

DOI:10.1111/j.1365-2818.1994.tb03436.x

出版商:Blackwell Publishing Ltd    

年代:1994

数据来源: WILEY

摘要:

SUMMARYThe incorporation into rat incisor dentin of two calcium isotopes, the stable44Ca and the radioactive45Ca, was studied using secondary ion mass spectrometry (SIMS) stepscanning and imaging, and autoradiography, respectively. The results demonstrated a time‐dependent incorporation of the calcium isotopes into the mineral phase of dentin. With the SIMS step‐scanning, detecting44Ca, the ion yield was high in the odontoblasts 2 min after intravenous injection. After 10 min a marked increase in signal intensity was found at the dentin mineralization front. This result was consistent with those obtained by45Ca autoradiography; a peak of incorporation occurred 10 min after injection of the isotope. Likewise, localization of44Ca to the mineralization front could be demonstrated 10 min after injection by SIMS imaging. In images obtained at earlier intervals, no such increase in ion yield could be detected. The results show that the nonradioactive, stable isotope44Ca can be used as a marker for biomineralization in a similar way to radioactive4

ISSN:0022-2720    

DOI:10.1111/j.1365-2818.1994.tb03437.x

出版商:Blackwell Publishing Ltd    

年代:1994

数据来源: WILEY

摘要:

SUMMARYEvidence from high‐resolution images of primary cell walls suggests that the cell wall is constructed from at least two independent yet coextensive fibrous networks, one based on cellulose/hemicellulose and the other on pectin. The ability to analyse the structure of each of these networks in isolation has been hampered by a lack of suitable biological material such as mutants. However, the recent use of the cellulose‐synthesis inhibitor 2,6‐dichlorobenzonitrile (DCB) that prevents the formation of the cellulose‐xyloglucan network while allowing the pectin network to form a functional wall offers the unique opportunity of studying at least the pectin network independently. A range of electron microscopy techniques and a novel spectroscopy method are used to study the walls from tomato suspension cells adapted to growth on DCB. Measurements of the minimum cell wall thickness derived from thin sections of dehydrated walls show that the marked reduction in level of the cellulose/hemicellulose network affects neither the thickness of the wall formed, nor the apparent spacing of pectin molecules. However, images obtained by the fast‐freeze, deep‐etch, rotary‐shadowed (FDR) replica technique show that the three‐dimensional architecture of these pectin‐rich walls is very different from that of nonadapted walls. Fourier transform infrared (FTIR) microspectroscopy data and immunogold‐labelling studies provide additional evidence that supports the previo

ISSN:0022-2720    

DOI:10.1111/j.1365-2818.1994.tb03438.x

出版商:Blackwell Publishing Ltd    

年代:1994

数据来源: WILEY

摘要:

SUMMARYA new algorithm for image segmentation is proposed, which is capable of extracting particles in the presence of noise and background fluctuations. It begins with the detection of small regions belonging to the background, called background nuclei, and then lets these nuclei grow to become the entire background. Edge information and region information of the image are used simultaneously.

ISSN:0022-2720    

DOI:10.1111/j.1365-2818.1994.tb03439.x

出版商:Blackwell Publishing Ltd    

年代:1994

数据来源: WILEY