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Microscopical image analysis: problems and approaches.

This article reviews some of the problems which have been encountered in the application of automatic image analysis to problems in biology. Some of the questions involved in the actual formulation of such a problem for this approach are considered as well as the difficulties in the analysis due to lack of specific constrast in the image and to its complexity. Various practical methods which have been successful in overcoming these problems are outlined, and the question of the desirability of an opto-manual or semi-automatic system as opposed to a fully automatic version is considered.

Computers↗

Object-oriented developmental environment for image-analysis applications: implementation for 2D gel electrophoretogram analysis.

MOTIVATION: The principal motivation was to design an environment for the development of image-analysis applications which would allow the integration of independent modules into one frame and make available tools for their build-up, running, management and mutual communication. RESULTS: The system was designed as modular, consisting of the core and work modules. The system core focuses on overall management and provides a library of classes for build-up of the work modules, their user interface and data communication. The work modules carry practical implementation of algorithms and data structures for the solution of a particular problem, and were implemented as dynamic-link libraries. They are mutually independent and run as individual threads, communicating with each other via a unified mechanism. The environment was designed to simplify the development and testing of new algorithms or applications. An example of implementation for the particular problem of the analysis of two-dimensional (2D) gel electrophoretograms is presented. The environment was designed for the Windows NT operating system with the use of Microsoft Foundation Class Library employing the possibilities of C++ programming language. AVAILABILITY: Available on request from the authors.

Computer Graphics↗

Clinical and biological applications of image analysis in non-Hodgkin's lymphomas.

The technical procedures and the main applications of image analysis in non-Hodgkin's lymphoma pathology have been described by reviewing the pertinent literature. With the advent of sophisticated computer-based technology, the possibility of objectively evaluating cell morphology, in situ marker expression, DNA indexes, and complex structural features may become a relevant part of the pathologist's expertise.

Algorithms↗

[Image analysis in contrast echocardiography].

The visualization of contrast agents in echocardiography is obtained with complex acquisition sequences and their analysis is dependent on operator experience. In order to be more accessible, images analysis is essential and the main objective is to integrate qualitative and quantitative data in one single image.

Contrast Media↗

A computer-aided image analysis system for area measurement of tooth root surfaces.

The characteristics of a computer-aided Image Analysis System (IAS) to conduct linear and area measurements on tooth root surfaces were investigated. The borders of the features to be measured were extracted from a digitized video image by histogram-based thresholding and binarization. The lengths and square areas were expressed in the number of pixels. The reference-length and area of one pixel were obtained through measurement of the diameter of a metal ball of known diameter in the same digital image. The validity of the IAS was tested using radiographs of teeth containing round metal restorations of known diameter and area. The accuracy of the length measurements ranged from -2.0 to 2.1%, whereas the accuracy of area measurement was between -5.3 and 0.8%. The precision of the IAS to identify the borders of a tooth root surface and to reproducibly calculate its area, expressed in a coefficient of variation, ranges from 0.49 to 4.11%. The results indicate that imaging techniques can be applied to obtain very accurate measurements of tooth root surfaces.

Dental Enamel↗

A landscape parametric profile approach for rat brain image analysis.

This paper develops the use of the landscape parametric profile as a descriptive and interpretative method of analyzing metabolic activity data obtained by neuroimaging techniques such as 2-deoxyglucose autoradiography. The method is suggested as an initial survey-type approach that can be used to map regional activity throughout the coronal planes of a rat brain and to compare relative differences in the patterns of activity between different brains. Images are acquired using preexisting image analysis software and then transformed to landscape parametric profiles derived from each matrix array. The image profile analysis tasks are done using commercially available software for matrix computations such as MATLAB. An effective way of interpreting activity differences between brains is presented by plotting the results from a randomized blocks ANOVA significance test combined with the average profiles of different subjects within the same experimental group. The method greatly reduces the computational burden associated with image averaging and statistical comparison of brains from different subjects while preserving the richness in topographic metabolic information. One significant advantage of the method is that it results in figures that are readily depicted and interpretable in black and white.

Analysis of Variance↗

A comparison of subjective and objective measures of reduction of psoriasis with the use of ultrasound, reflectance colorimetry, computerized video image analysis, and nitric oxide production.

BACKGROUND: Studies of antipsoriatic therapy often rely on subjective scoring. Objective measures have been developed but have not previously been compared with subjective scoring. OBJECTIVE: Our purpose was to compare subjective and objective measures of reduction of psoriasis with topical therapy. METHODS: A 2-week, double-blind, left-to-right comparative trial of betamethasone valerate against white soft paraffin was performed in 12 patients. The subjective scores were erythema, elevation, scale, and a composite total. Objective measures were nitric oxide production measured by chemiluminescence; erythema reflectance; ultrasound scan for thickness, scale, and echo-poor zone; and computerized image analysis of video images. RESULTS: Subjective and objective measures had similar power to detect therapeutic effect. The subjective measures showed greater variation and relatively overestimated improvement. There was correlation between measures and estimates for area, redness, and thickness. Nitric oxide production was the most powerful objective measure. CONCLUSION: Thickness determined by ultrasound scan and nitric oxide production are useful measures of reduction of psoriasis, which lend themselves to more powerful statistical tests than subjective interval data.

Administration, Topical↗

[Image analysis of optic disk swelling in diabetic retinopathy].

Optic disc swelling was measured by computerized image analysis in diabetic patients. The subjects were divided into four groups. Group 1 contained 13 eyes without diabetic retinopathy, group 2, 14 eyes with a mild stage of simple diabetic retinopathy, group 3, 14 eyes with a severe stage of simple diabetic retinopathy and group 4, 14 eyes with preproliferative diabetic retinopathy. There were statistically significant differences in the optic disc swelling between group 1 and group 4, and between group 2 and group 4 (p < 0.05). The optic disc swelling increased as the stage of diabetic retinopathy progressed.

Diabetic Retinopathy↗

Image analysis: a consumer's guide.

The last years have seen an explosion of systems in image analysis. It is hard for the pathologist or the cytologist to make the right choice of equipment. All machines are stupid, and the only valuable thing is the human work put into it. So make your benefit of the work other people have done for you. Chose a method largely used on many systems and which has proved fertile in many domains and not only for your specific to day's application: Mathematical Morphology, to which are to be added the linear convolutions present on all machines is a strong candidate for becoming such a method. The paper illustrates a working day of an ideal system: research and diagnostic directed work during the working hours, automatic screening of cervical (or other) smears during night.

Animals↗

An image analysis system: an objective and accurate alternative for reading the agar diffusion test.

A computerized image analysis system (IAS) has been used to develop a new method for reading the agar diffusion test automatically. In four experiments a total of 88 porcine plasma and 95 urine samples were screened for oxytetracycline by the agar diffusion test. The inhibition zones were measured by hand and by the IAS directly from the bioassay plate and by the IAS from the photo-negative taken from the plate. Both methods were positively correlated with the hand method for plasma (0.9716, 0.9669) and urine (0.9878, 0.9731) in the range tested for 0.1 to 2.0 micrograms/ml. Moreover, the coefficient of variation and the day-to-day-variation amounted to 1.72% and 1.47% respectively, for the method by hand and 1.10, 1.54% and 0.27, 0.38% respectively, for the IAS methods. It is concluded that the IAS method is an objective and accurate alternative for reading the agar diffusion test.

Agar↗

Image analysis of gunshot residue on entry wounds. I--The technique and preliminary study.

An automated image analysis (IA) technique has been developed to obtain a measure of the amount (i.e. number and area) of gunshot residue (GSR) particles within and around a gunshot wound. Sample preparation and IA procedures were standardised to improve the reproducibility of the IA measurements of GSR. Measurements of GSR from test firings into goat hide were enhanced by staining the barium and lead components present on the skin sections with Alizarin Red S. The amount of GSR detected on the stained skin sections was compared with backscatter electron micrographs of the same sections. The differences were deemed to be insignificant when the variability in the repeated test firings were taken into consideration. Preliminary results indicated that the decreasing relationship between firing range and the amount of GSR deposited was non-linear, and that for firing ranges of up to 20 cm the amount of GSR deposited from repeated shots fired from the same distance was highly variable.

Animals↗

Assessment of the nucleolar organizer regions by automated image analysis in benign and malignant colonic tumours and adjacent tissues in rats.

An automated computer image analysis technique was used to study the morphological parameters of argyrophilic nucleolar organizer regions (AgNORs) in chemically induced rat colonic tumours of different grades. Different nuclear parameters were of different diagnostic value. For distinguishing tumorous tissue from normal tissue all the parameters studied were capable of serving as diagnostic markers. Malignant processes could, however, be more reliably detected by means of the area of the nucleus, nuclear shape factor, area of AgNOR and ratio of AgNOR area to nuclear area. In macroscopically normal tissue adjacent to a tumour, the values of all the AgNOR parameters studied were similar to those in tumorous tissues. It can be concluded that the initial stages in tumorigenesis are accompanied by changes in all of the nucleus and AgNOR parameters, but as malignancy develops, only some of these parameters continue to change. Close correlations between nuclear parameters in malignant tissue suggest that for diagnostic purposes only nuclear and AgNOR areas should be used.

Adenocarcinoma↗

Variations with age and serum cholesterol level in the topographic distribution of macroscopic aortic atherosclerotic lesions as assessed by image analysis methods.

The topographic distribution of macroscopic atherosclerotic lesions in the aorta was studied by image analysis of aortas taken at autopsy from 155 males aged 10 to 54 yr. In both the thoracic and abdominal aorta, fatty streaks (FS) appear before raised lesions (RL) affecting preferential areas located longitudinally along the intercostal ostia, at the aorta bifurcation, and surrounding the points of origin of branches of the abdominal aorta. Later RL appear in the same areas of the aortic intima affected by FS at younger ages. As for the effect of serum total cholesterol (TC) in the progression of atherosclerosis, the prevalent involvement areas by FS and RL were similarly distributed as indicated by our analyses. However, areas showing frequent occurrence of FS and RL in each aortic segment were distributed more extensively at identical areas in the higher serum TC groups than in the lower TC level group. Thus, it can be concluded that FS are the precursor lesions of RL in the aorta and that the serum TC promotes the progression of atherosclerosis.

Adolescent↗

Image analysis evaluation of ploidy, S-phase fraction and nuclear area in canine mammary tumours.

Image analysis was used on cytocentrifuge preparations of Feulgen-stained nuclei extracted from formalin-fixed, paraffin wax-embedded tissues to determine ploidy (assessed by integrated optical density), S-phase fraction, and nuclear area of 90 canine mammary tumours (30 benign and 60 malignant). Only two lesions identified histologically as benign were aneuploid, suggesting potential malignancy. Of malignant tumours, 37% were aneuploid. No distinction based on ploidy could be made within benign or malignant groups, except for tubular adenocarcinomas of complex type, all of which were diploid. Mean S-phase fraction of malignant lesions (15.60%) was twice that of benign lesions (7.87%; P < 0.0001). The mean nuclear surface was significantly smaller in malignant lesions (70.62 microns2) than in benign counterparts (77.45 microns2; P < 0.05). No relation was found between a given tumour type and S-phase fraction or nuclear area within histologically malignant or benign tumour groups. Infiltration of lymphatics by neoplastic cells, known to indicate a poor prognosis, was associated with aneuploid lesions (mean DNA index > 1.25) and high S-phase fraction. The presence of necrosis and of a high number of mitoses was significantly associated with high S-phase fraction lesions.

Animals↗

Determination of pseudophakic accommodation with translation lenses using Purkinje image analysis.

PURPOSE: To determine pseudophakic accommodation of an accommodating posterior chamber intraocular lens (translation lens) using Purkinje image analysis and linear matrix methods in the paraxial space. METHODS: A 2 x 2 system matrix was defined for each Purkinje image I to IV using refraction, translation and mirror matrices. Image size (m) and axial image position (z) was determined as an example for an off-axis object (a 0.2 m off-axis object located 0.5 m in front of the cornea.). First, our method was applied to the phakic relaxed (emmetropic) and accommodated (6.96 D) Le Grand eye. Secondly, for demonstration of the applicability of the calculation scheme to the pseudophakic eye, we provide a clinical example where we determine the accommodation amplitude of the translation lens (1 CU, HumanOptics, Erlangen, Germany) from photographed Purkinje images in the relaxed and accommodated state. From the biometric data: axial length 23.7 mm, corneal power 43.5, corneal thickness 550 microns, implanted intraocular lens (IOL) with a refractive power of 20.5 D (shape equi-biconvex, refractive index 1.46), and refractive indices of the cornea, aqueous and vitreous from the Le Grand model eye, we calculated the refractive state and the sizes of Purkinje images I and III initiated from two off-axis light sources. RESULTS: For the Le Grand model eye, Purkinje image II (z/m = 3.5850 mm/0.0064) is slightly smaller than and directly in front of image I (z/m = 3.8698 mm/0.0077). Purkinje image III (z/m = 10.6097 mm/0.0151) is nearly double the size of image I and during accommodation it moves from the vitreous into the crystalline lens. Purkinje IV (z/m = 4.3244 mm/-0.0059) is inverted, three quarters the size of image I, lies in the crystalline lens and moves slightly towards the retina. For the pseudophakic eye, pseudophakic accommodation of 1.10 D was calculated from the proportion of distances between both Purkinje images I and III in the relaxed (3.04) and accommodated (2.75) state, which is in contrast to the total subjective accommodation of 2.875 D evaluated with an accommodometer. CONCLUSIONS: We present a straightforward mathematical strategy for calculation of the Purkinje images I-IV. Results of our model calculation resemble the values provided by Le Grand. In addition, this approach yields a simple en bloc scheme for determination of pseudophakic accommodation in pseudophakic eyes with accommodative lenses (translation lenses) using Purkinje image photography.

Accommodation, Ocular↗

Cell cycle expression of estrogen receptors determined by image analysis on human breast cancer cells in vitro and in vivo.

We have investigated, by image analysis, the cell cycle expression of estrogen receptors (ER) on MCF-7 cell line and on MCF-7 xenografts. The results demonstrate, in vitro as well as in vivo, an increase of ER concentration during the G0/G1-phase, followed by a decrease during the S-phase until the late S-phase where a rapid increase was noted. These results confirm that estrogens are involved in the DNA synthesis since ER is expressed in vivo at a maximal level in the late G1. In presence of saturating concentrations of 17 beta-estradiol, the mean ER concentration in G0/G1 phase is significantly decreased compared with the control cells cultured in estrogen-deprived medium. This indicates that 17 beta-estradiol down-regulates ER preferentially in the G0/G1 phase. These data suggest that ER in S and G2/M phases is unable to interact with its ligand. Consequently, estrogens may have no effects on the entry of cells in mitosis. Finally, after long-term tamoxifen treatment of MCF-7 xenografts, a tamoxifen-resistant tumor was developed which was characterized by a change in the profile of ER concentration during the G0/G1 phase. In conclusion, it is possible that the differences in cell cycle distribution of ER could be correlated with different phenotypes of breast cancer and also with different clinical phases of tumoral evolution. However, it remains to be known what is the clinical significance of the ER cell cycle expression in relation to tumor aggressiveness and survival.

Animals↗

Morphology of arachnoid granulations and villi in the region of the human sella turcica--light microscopy and three-dimensional image analysis.

The distribution and structure of the arachnoid granulations and villi in the region of the sella turcica in human adult brains were observed under light microscopy. In order to study the interrelationships between the arachnoid projections, hypophysis, cavernous sinus, and sella turcica, we performed a three-dimensional reconstruction and analysis of the region with the use of a digital image-processing method. Arachnoid projections penetrating the dura were villous when they were few in number, but more frequently granular when they were numerous. No relationship was observed between the type of projections and the age or the underlying disease of the subjects. Brains which showed no or few arachnoid granulations or villi had a small and deep hypophyseal fossa with a poorly-developed intercavernous sinus or venous plexus. Three-dimensional image analysis revealed that arachnoid granulations and villi form a maze in the dura or connective tissue between the venous plexuses.

Adult↗

Semiquantitative computerized image analysis of fetal hemoglobin distribution patterns in sickle cell anemia and its variants.

Computerized, semiquantitative image analysis of 2913 erythrocytes from 29 young patients with sickle cell anemia and its variants was conducted utilizing a modified method for determination of fetal hemoglobin distribution by an acid elution procedure. Histograms of eight arbitrary levels of staining intensity, proportional to fetal hemoglobin levels, were analyzed in relationship to both computer-generated, mathematical parameters of erythrocytic shape abnormalities and clinical parameters of disease severity. Shifts in fetal hemoglobin distribution were observed in analysis of sequential specimens. An inverse correlation was observed between angularity of erythrocytes and their individual fetal hemoglobin content (p less than or equal to 0.05). A positive correlation was noted between growth percentiles and percentage erythrocyte ghosts by acid elution (p less than or equal to 0.01).

Anemia, Sickle Cell↗