Technical note: A toy as tool: a low-cost image analysis system for the evaluation of tumor size in experimental small animal models

Image analysis systems are an essential tool in measurements of size of intraparenchymal tumors or lesions in experimental small animal models. Conventional image analysis systems are relatively expensive. We therefore compared the performance of a professional image analysis system with an inexpensive setup by evaluating tumor size in an orthotopic glioma mouse model. The maximum cross-sectional tumor area of H&E stained brain-slides of two groups of mice (treatment and control group) was measured by two independent investigators using a professional image analysis system (Leica DM IRB microscope) with the Leica Quantimet 500c software, and a low-cost-system (Intel QX3 microscope) with a non-commercial image analysis software. Mean tumor volumes were calculated and the results from each of the image analysis systems, investigators, and treatment effects were compared. The tumor volumes as measured with the low-cost and the professional system differed between -3.7 and +7.5% (P = 0.69-0.99). Measurements made by investigator A and B differed between -7.0 and +3.9% (P = 0.69-0.88). Treatment in all cases significantly reduced the tumor volume between 58.4 and 62.7% (P = 0.0002 or 0.0003), regardless of the investigator or the used image analysis system. We therefore conclude that the QX3 low-cost microscope in combination with a non-commercial image-analysis software represents an inexpensive solution to reliably analyze the size of regions of interest, if they provide a sufficient contrast. However, the low-cost setup due to its low resolution definitely limits a detailed analysis of histologic features.     

Mechanical scanning of the specimen in the scanning electron microscope

A scanning electron microscope (SEM) system equipped with a motor drive specimen stage fully controlled with a personal computer (PC) has been utilized for obtaining ultralow magnification SEM images. This modem motor drive stage works as a mechanical scanning device. To produce ultra-low magnification SEM images, we use a successful combination of the mechanical scanning, electronic scanning, and digital image processing techniques. This new method is extremely labor and time saving for ultra-low magnification and wide-area observation. The option of ultra-low magnification observation (while maintaining the original SEM functions and performance) is important during a scanning electron microscopy session.     

Scanning the pressure-induced distortion of fingerprints

Fingerprint recognition technology is an important part of criminal investigations it is the basis of some security systems and an important tool of government operations such as the Immigration and Naturalization Services, registration procedures in the Armed Forces, and so forth. After the tragic events of September 11, 2001, the importance of reliable fingerprint recognition technology became even more obvious. In the current study, pressure-induced changes of distances between ridges of a fingerprint were measured. Using calibrated silicon pressure sensors we scanned the distribution of pressure across a finger pixel by pixel, and also generated maps of an average pressure distribution during fingerprinting. Emulating the fingerprinting procedure employed with widely used optical scanners, we found that on average the distance between ridges decreases by about 20% when a finger is positioned on a scanner. Controlled loading of a finger demonstrated that it is impossible to reproduce the same distribution of pressure across a given finger during repeated fingerprinting procedures.     

Robust incremental compensation of the light attenuation with depth in 3D fluorescence microscopy

Fluorescent signal intensities from confocal laser scanning microscopes (CLSM) suffer from several distortions inherent to the method. Namely, layers which lie deeper within the specimen are relatively dark due to absorption and scattering of both excitation and fluorescent light, photobleaching and/or other factors. Because of these effects, a quantitative analysis of images is not always possible without correction. Under certain assumptions, the decay of intensities can be estimated and used for a partial depth intensity correction. In this paper we propose an original robust incremental method for compensating the attenuation of intensity signals. Most previous correction methods are more or less empirical and based on fitting a decreasing parametric function to the section mean intensity curve computed by summing all pixel values in each section. The fitted curve is then used for the calculation of correction factors for each section and a new compensated sections series is computed. However, these methods do not perfectly correct the images. Hence, the algorithm we propose for the automatic correction of intensities relies on robust estimation, which automatically ignores pixels where measurements deviate from the decay model. It is based on techniques adopted from the computer vision literature for image motion estimation. The resulting algorithm is used to correct volumes acquired in CLSM. An implementation of such a restoration filter is discussed and examples of successful restorations are given.     

Estimation of true dihedral angles and relative grain boundary energies in polycrystals using the disector

Relative grain boundary energies can be simply related to true dihedral angles, which are the angles between grain boundary planes meeting at triple edges in polycrystals. Some limited efforts in the measurement of true dihedral angles have used the technique of serial sectioning, which is usually cumbersome and time consuming. In this study the three‐dimensional probe called the ‘disector’ has been employed to evaluate true dihedral angles. This probe, combined with automated image processing, introduces precision as well as efficiency, overcomes the disadvantages of the two‐dimensional probe and is far less tedious and less complicated than serial sectioning. It is shown that the technique is relatively simple and therefore can be applied to obtain a significantly large and accurate statistical sample of true dihedral angles. The application of this method is demonstrated by evaluating the triple junction geometry and the associated relative grain boundary energies in polycrystalline 316L austenitic stainless steel.     

Minimizing photobleaching during confocal microscopy of fluorescent probes bound to chromatin: role of anoxia and photon flux

Exposure to light can destroy the ability of a molecule to fluoresce. Such photobleaching limits the use of fluorescence and confocal microscopy in biological studies. Loss of fluorescence decreases the signal‐to‐noise ratio and so image resolution; it also prevents the acquisition of meaningful data late during repeated scanning (e.g. when collecting three‐dimensional images). The aim of this work was to investigate the role of oxygen in the photobleaching of fluorophores bound to DNA in fixed cells, and to explore whether anoxia could minimize such bleaching. Anoxia significantly reduced bleaching rates and changed the order of reaction of both propidium iodide (an intercalator) and chromomycin A₃ (a minor‐groove binder) bound to DNA; it afforded the greatest protection at low photon fluxes. However, it had no effect on the bleaching of the green fluorescent protein (GFP) covalently attached to a histone and so bound to DNA, probably because the protein shielded the chromophore from oxygen. Bleaching of all three fluorophores depended on photon flux. Practical ways of minimizing bleaching were examined, and examples of three‐dimensional images of DNA marked by propidium and GFP (collected under standard and optimized conditions) are presented.     

Relationship between cavitation structures and cavitation damage

A study of visual and erosion effects of cavitation on simple single hydrofoil configurations in a cavitation tunnel was made. A two-dimensional hydrofoil with circular leading edge was used for the experiments. In addition, the hydrofoil geometry was modified to obtain some three-dimensional cavitation effects. A thin copper foil, applied to the surface of the hydrofoil, was used as an erosion sensor. Cavitation phenomenon above hydrofoils at different flow conditions (system pressure, water gas content) was observed. Images of vapour cavities from above and from side view were taken. A statistical evaluation of cavitation structures was made. Images of damaged copper coated hydrofoil surface were taken under sufficient magnification. A pit-count method, based on computer-aided image processing, was used for direct measurement of the cavitation erosion by evaluating the damage of the surface of the hydrofoil. A relation between characteristics of cavitation structures and cavitation damage was established.     

基于FPGA的电子内窥镜CCD彩色图像采集与显示系统

本文介绍了电子内窥镜ＣＣＤ图像采集和显示系统的研究与实现,并主要论述了以现场可编程门阵列（ＦＰＧＡ）为系统控制核心的ＣＸＣＤ图像放视频显示缓存管理及窗口控制,以及基于锁相原理的系统时钟的包括同步信号发生电路和高频主时钟电路。     

Methodological principles for fractal analysis of trabecular bone

A standardized methodology for the fractal analysis of histological sections of trabecular bone has been established.
A modified box counting method has been developed for use on a PC-based image analyser. The effect of image analyser settings, magnification, image orientation and threshold levels was determined. Also, the range of scale over which trabecular bone is effectively fractal was determined and a method formulated to calculate objectively more than one fractal dimension from the modified Richardson plot.
The results show that magnification, image orientation and threshold settings have little effect on the estimate of fractal dimension. Trabecular bone has a lower limit below which it is not fractal (λ < 25 μm) and the upper limit is 4250 μm. There are three distinct fractal dimensions for trabecular bone (sectional fractals), with magnitudes greater than 1.0 and less than 2.0.
It has been shown that trabecular bone is effectively fractal over a defined range of scale. Also, within this range, there is more than one fractal dimension, describing spatial structural entities. Fractal analysis is a model-independent method for describing a complex multifaceted structure, which can be adapted for the study of other biological systems. This may be at the cell, tissue or organ level and complements conventional histomorphometric and stereological techniques.     

Automatic image acquisition, calibration and montage assembly for biological X-ray microscopy

We describe a system for the automatic acquisition and processing of digital images in a high-resolution X-ray microscope, including the formation of large-field high-resolution image montages. A computer-controlled sample positioning stage provides approximate coordinates for each high-resolution subimage. Individual subimages are corrected to compensate for time-varying, non-uniform illumination and CCD-related artefacts. They are then automatically assembled into a montage. The montage assembly algorithm is designed to use the overlap between each subimage and multiple neighbours to improve the performance of the registration step and the fidelity of the result. This is accomplished by explicit use of recorded stage positions, optimized ordering of subimage insertion, and registration of subimages to the developing montage. Using this procedure registration errors are below the resolution limit of the microscope (43 nm). The image produced is a seamless, large-field montage at full resolution, assembled automatically without human intervention. Beyond this, it is also an accurate X-ray transmission map that allows the quantitative measurement of anatomical and chemical features of the sample. Applying these tools to a biological problem, we have conducted the largest X-ray microscopical study to date.