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PubMed1995

Image analysis quantification in hormone receptor assay of mucinous carcinoma of the breast. Comparison with biochemical analysis.

Mucinous carcinoma (MC) of the breast is characterized by abundant extracellular mucin and by variable epithelial cellularity. Since some MCs are extremely hypocellular, we questioned the validity of biochemical (BIO) assays in these tumors. We analyzed paraffin-embedded tissue from 34 cases of MC of the breast for quantitative estrogen receptor (ER) and progesterone receptor (PR) using immunohistochemistry (IHC) on the Cell Analysis System CAS-200. Of the 34 cases, 31 (91%) were positive for ER, whereas 18 (53%) were positive for PR. In 21 cases the quantitative ER and PR were assayed biochemically by a dextran-coated charcoal method. Using the BIO results as the "true" values, the sensitivity of IHC for ER and PR was 100% and 78%, and the specificity was 13% and 64%, respectively. The low specificity of the values obtained by IHC was attributed to the fact that eight cases were "falsely" false positive (negative by BIO and positive by IHC) for ER and/or PR. Review of the histologic patterns of all 21 cases showed that 7 of the 8 falsely false positive cases were significantly hypocellular (epithelial cellularity 5-20%) as compared to the remaining cases (epithelial cellularity 20-75%). We conclude that immunohistochemical analysis of ER/PR status using image analysis in MC is a sensitive method, with the ability to detect receptor content when their concentration might be too low to be demonstrated by the conventional method as a result of sparse cellularity.

Adenocarcinoma, Mucinous↗
PubMedDate not supplied

Computerized image analysis of nails affected by fungal infection: evaluation using digital photographs and manually defined areas.

Despite the relevant increase in clinical trials on the efficacy of various systemic and/or topical antifungal agents in onychomycosis therapy, the evaluation of the results is largely subjective. The aim of this study was to set up and ensure an objective, reproducible and reliable method to measure nail plate involvement in onychomycosis. In order to validate a specifically designed software for the computerized image analysis of affected areas of the nail, standardized clinical pictures of onychomycosis were prepared by six different clinicians using a sample of 11 affected nails. Diseased areas and total nail plates were measured both on the clinical pictures and on their drawings traced by the different clinicians on transparent tapes adhering to sample nails. The computerized procedure was undertaken by a trained operator who was not a dermatologist. The variation coefficients of measurements on clinical pictures (automatically detected) and on drawings were compared. In addition, the agreement between automatic evaluation and drawing was evaluated by means of Bland-Altman analysis. To consider the effect of possible variations linked to different operators using the computerized method, 11 clinical pictures (one for each clinical case considered) were selected and submitted to computerized image analysis by six different trained operators. The computerized detection of affected nail areas showed a coefficient of variation (vc=8.5%) lower than that observed on drawings (vc=14.7%). The two methods showed appreciable agreement, as demonstrated by Bland-Altman plot. The coefficient of variation of image analysis conducted by six different operators was very low for the total area calculation (vc=0.9%) and acceptable for pathological area detection (vc=4.8%). Based on the results obtained, we conclude that automatic evaluation is a reliable and helpful method for the measurement of the clinical involvement of the nail plate in onychomycosis and for the evaluation of therapies, since it can increase the objectivity and reproducibility of data. However, in a minority of difficult cases, expert dermatological evaluation is needed.

Humans↗
PubMed2005

Comparison of image analysis software packages in the assessment of adhesion of microorganisms to mucosal epithelium using confocal laser scanning microscopy.

We have compared current image analysis software packages in order to find the most useful one for assessing microbial adhesion and inhibition of adhesion to tissue sections. We have used organisms of different sizes, the bacterium Helicobacter pylori and the yeast Candida albicans. Adhesion of FITC-labelled H. pylori and C. albicans was assessed by confocal microscopy. Four different Image analysis software packages, NIH-Image, IP Lab, Image Pro+, and Metamorph, were compared for their ability to quantify adhesion of the two organisms and several quantification methods were devised for each package. For both organisms, the dynamic range that could be detected by the software packages was 1x10(6)-1x10(9) cells/ml. Of the four software packages tested, our results showed that Metamorph software, using our 'Region of Interest' method, with the software's 'Standard Area Method' of counting, was the most suitable for quantifying adhesion of both organisms because of its unique ability to separate clumps of microbial cells. Moreover, fewer steps were required. By pre-incubating H. pylori with the glycoconjugate Lewis b-HSA, an inhibition of binding of 48.8% was achieved using 250 mug/ml Lewis b-HSA. The method we have devised using Metamorph software, provides a simple, quick and accurate way of quantifying adhesion and inhibition of adhesion of microbial cells to the epithelial surface of tissue sections. The method can be applied to organisms ranging in size from small bacteria to larger yeast cells.

Animals↗
PubMed1989

Central values and variation of measured nuclear DNA content in imprints of normal tissues determined by image analysis.

A total of 109 slides derived from 37 normal tissues were analyzed for nuclear DNA content using the Cell Analysis Systems (CAS) model 100 image analysis system with the Quantitative DNA Analysis (QDA) software module in order to determine the central values of DNA content and define normal limits. Analysis of the 109 slides revealed an overall mean measured DNA index of 0.997 with a standard deviation of 0.04. Analysis of replicate samples was essentially constant. There were no differences between samples obtained from autopsy and surgical specimens. Small variations were detected among stain batches, individual patients, and tissue types. These results indicate a high degree of accuracy and reproducibility of DNA content determinations using this system.

Cell Nucleus↗
PubMed2001

Quantitative immunohistochemical evaluation of HER2/neu expression with HercepTestTM in breast carcinoma by image analysis.

HercepTestTM (DAKO A/S, Glostrup, Denmark) is an immunohistochemical assay that detects HER2/neu gene products, and evaluates the overexpression status of the HER2/neu protein in determining eligibility for the Trastuzumab (HerceptinR, Genentech, San Francisco, CA, USA) therapy. However, practically, interobserver variability of the HER2/neu interpretation of the immunostained results has caused marked disagreement with regard to the intensity of tumor staining. In this study, we quantitated HER2/neu expression by image analysis, and applied this analyzing system to help to minimize interobserver variability of the interpretation of the HercepTestTM. All the immunostained results were scored semiquantitatively on a range of 0 to 3+ in accordance with the criteria described as per the manufacturer's instructions, and quantitatively evaluated using an image analyzing system with image processing software. Among the 92 cases, 15 were scored as 3+, six were 2+, and 32 were 1+ under intraobservers agreement. When the cases were quantitated, a high correlation was shown between the signal area extracted by image analysis and the corresponding score of staining intensity with the HercepTestTM. By converting the quantitatively extracted data into a scoring system based upon the criteria, the outcome demonstrated a strong concordance with the scoring data obtained from immunostaining. The results indicated that a quantitative scoring system performed by simple image analysis may provide to improve interobserver agreement of the interpretation of the HercepTest TM in clinical practice.

Adult↗
PubMed2006

Digital image analysis of cytoskeletal F-actin disintegration in renal microvascular endothelium following ischemia/reperfusion.

BACKGROUND: Damaged and/or dysfunctional microvascular endothelium has been implicated in the pathogenesis of ischemic acute renal failure (ARF). Rapidly occurring changes in the endothelial F-actin cytoskeleton as observed in vitro might be responsible, but have been proven difficult to measure accurately in situ. Therefore, the purpose of this study was to examine several methods of digital image analysis in order to quantify the alterations of endothelial F-actin after renal ischemia and reperfusion (I/R), and to relate these to deterioration of renal function. METHODS: Frozen sections of Sham and I/R rat kidneys were fixed in 4% formaldehyde and stained with rhodamine-phalloïdin. Microvascular structures were captured using a 3i Marianastrade mark digital imaging fluorescence microscope workstation. Images were analyzed using 3i SlideBooktrade mark software, employing several masking techniques and line-scans. RESULTS: Digital image analysis demonstrated a decrease in the mean intensity of rhodamine-phalloïdin fluorescence after I/R from 1030 +/- 187 to 735 +/- 121 a.u. (arbitrary units, mean +/- SD, n = 7). The number of F-actin fragments per pixel increased from (15.8 +/- 4.9) x 10(-5) to (20.7 +/- 3.5) x 10(-5) (n = 7), indicating cytoskeletal fragmentation. In addition, line-scan analysis demonstrated a disturbed spatial orientation of the F-actin cytoskeleton after I/R. Finally, the loss of F-actin correlated with a rise in plasma creatinine. CONCLUSIONS: The methods of digital image analysis described in the present study demonstrate that renal I/R induces profound changes in the F-actin cytoskeletal structure of microvascular endothelial cells, implicating an injured and dysfunctional microvascular endothelium, which may contribute to acute renal failure (ARF).

Actins↗
PubMed1996

Sendai virus-induced IFN-alpha production analysed by immunocytochemistry and computerized image analysis.

IFN-alpha production in Sendai virus-stimulated human buffy coat cultures could readily be demonstrated in individual cells at a protein level by the use of a novel immuno-enzymatic staining procedure. A distinctive rounded, juxtanuclear staining pattern was generated in producer cells by the accumulation of the intracellularly synthesized IFN-alpha in the Golgi stacks. The technology is based upon acquiring a video image of stained monolayers of cells, viewed in a microscope by a colour camera, which then transfers binary images directly into a computer-controlled operating system. The characteristic appearance of the immunocytochemical staining enabled a computerized image-analysis system to measure IFN-alpha producing cells based on defined criteria set for morphology, intensity, colour and size. The automated system could accurately and reproducibly register a range of 0.1-7.0% of the total cell population as IFN-alpha producing cells during the kinetic studies of the response. Congruent results were obtained with manual microscopy and image analysis concerning the assessment of the incidence of IFN-alpha producing cells in the total cell populations. All IFN-alpha producing cells expressed surface HLA-DR molecules and 95% of these cells belonged to the myelomonocytic lineage. The image analysis system provided, in contrast to conventional microscopy, an opportunity to assess and document differences of signal intensity and cell size of individual IFN-alpha producing as well as non-producing cells.

Cell Size↗
PubMed1997

[The establishment of the digital imaging analysis system on the lower maxillofacial soft tissue]

Soft tissue shape is an important parameter to evaluate the postprosthetic effect of facial profile.In this study,the technique of digital imaging analysis was used to transferring from imaging informations to digital information,and then these information were imputed to computer for capturing,measuring and analyzing.So far,a set of digital imaging analysis system relating to the speciality of prosthodontics has been established preliminarily,with the advantage of quantitative analysis,non invasion and accuracy.It will provide a new way for reconstructing vertical dimension of occlusion.

Journal Article↗
PubMedDate not supplied

Flow visualization tools for image analysis of capillary networks.

OBJECTIVE: Video recordings of red blood cell (RBC) flow through capillary networks contain a considerable amount of information pertaining to oxygen transport through the microcirculation. Image analysis of these video recordings has been widely used to determine RBC dynamics (velocity, lineal density and supply rate) and oxygenation (Brunner et al., 2000; Ellis et al., 1990, 1992; Ellsworth et al., 1987; Klyscz et al., 1997; Pries 1988). However, not all capillaries in a given field of view are suitable for image analysis. Typically, capillary segments that are relatively straight and in sharp focus, and exhibit flow of individual RBCs that are well separated by plasma gaps, are good candidates for analysis. We have developed several image processing tools to aid in the selection of such capillaries for analysis and to obtain quick overviews of RBC flow through the microcirculation. METHODS: Burgess et al. (Microcirc. 2:75, 1995) and Burkell et al. (Annals Biomed. Eng. 24:1, 1996; J. Vasc. Res. 35:2, 1998) have previously introduced mean and variance images to aid in the selection of capillaries for analysis. We have extended their concept and developed similar two dimensional visualization techniques for studies of RBC flow through capillary networks. RESULTS: Five new methods of processing video data were developed. The minimum image highlights all capillaries containing RBCs in a given field of view. The maximum image identifies capillaries that exhibit high lineal density or stopped flow. The range image represents the difference between the maximum and minimum light intensity values that occur at a given pixel over a given time period, and helps to identify capillary segments that are in good focus and are perfused by RBCs and plasma. The difference image represents the cumulative sum of the square of differences in intensity values between consecutive frames and gives an indication of the frequency of passage of RBCs separated by plasma gaps. The transition image represents the number of times the intensity at a given pixel crosses a predefined threshold and indicates the number of RBCs (or trains of RBCs) that passes a given location during the observation period. CONCLUSIONS: The above flow visualization techniques are valuable tools to aid in the study of image focus, network geometry, RBC flow paths and dynamics, that can then be used in identifying capillaries for subsequent (separate) detailed analysis to provide quantitative information about RBC flow.

Algorithms↗
PubMed1992

DNA quantitation by image analysis of paraffin-embedded colorectal adenocarcinomas and its prognostic value.

DNA content was measured by image analysis in a retrospective study of formalin-fixed paraffin-embedded colorectal carcinomas from 213 patients who were followed up for at least 5 yr. DNA histograms were classified as diploid, aneuploid, or tetraploid. Diploid tumors comprised 29% of all cases, aneuploid 50%, and tetraploid 21%. Aneuploid tumors were found more often in patients with advanced disease and in carcinomas arising in the rectum. Pathologic stage, histologic grade, and ploidy were individually related to survival and recurrence. However, after stage stratification, histologic grade was no longer a significant prognostic factor. In patients without regional or distant metastases (Dukes' Stage A and Stage B), patients with aneuploid tumors had a statistically worse prognosis than patients with diploid or tetraploid tumors (P less than 0.01). The prognostic value of ploidy in this group of patients was maintained only in tumors arising in the distal colon and rectum (P less than 0.04). In patients with regional or distant metastases, DNA content did not provide additional prognostic information. In conclusion, DNA quantitation can be evaluated reliably by image analysis of archival material and can provide valuable prognostic information, especially in patients with Dukes' Stage A and Stage B disease. It may prove useful in guiding adjuvant therapy in these patients.

Adenocarcinoma↗
PubMed1997

Tumor angiogenesis as a prognostic factor in ovarian carcinoma: quantification of endothelial immunoreactivity by image analysis.

BACKGROUND: The growth of a malignant tumor requires the formation of new capillaries. Quantification of these microvessels is difficult. The purpose of this study was to establish an objective technique for quantifying angiogenesis and to evaluate whether microvessel quantity may predict tumor aggressiveness in patients with ovarian carcinoma. METHODS: Endothelial area was used to quantify microvessel density in immunohistochemically stained sections of 28 International Federation of Gynecology and Obstetrics Stage IIIC ovarian carcinomas. The endothelial area was measured with a computer-aided image analysis system in the subepithelial stroma of highest vascularization. The endothelial area in the specimens of 14 patients who survived for > or =6 years was compared with that of 14 patients matched for stage and treatment who died of the disease. RESULTS: The mean tumor area analyzed was 5.04 +/- 0.23 mm2. The mean endothelial area per mm2 of stroma from survivors and dead patients was 0.038 +/- 0.026 mm2 and 0.110 +/- 0.034 mm2, respectively (P < 0.0001). No significant differences were found in histology, tumor grade, status of lymph nodes, and amount of residual tumor. CONCLUSIONS: Image analysis was used to overcome the potential subjectivity of manual counts. Computer-assisted image analysis can evaluate accurately the angiogenic potential in ovarian carcinomas. Tumor angiogenesis may prove to be a prognostic factor in patients with ovarian carcinoma. This study suggests that the measurement of the endothelial area would be clinically useful in determining microvessel density [See editorial on pages 2219-21, this issue.]

Antigens, CD34↗
PubMed1999

Digital image analysis: improving accuracy and reproducibility of radiographic measurement.

OBJECTIVE: To assess the accuracy and reproducibility of a digital image analyser and the human eye, in measuring radiographic dimensions. DESIGN: We experimentally compared radiographic measurement using either an image analyser system or the human eye with digital caliper. BACKGROUND: The assessment of total hip arthroplasty wear from radiographs relies on both the accuracy of radiographic images and the accuracy of radiographic measurement. METHODS: Radiographs were taken of a slip gauge (30+/-0.00036 mm) and slip gauge with a femoral stem. The projected dimensions of the radiographic images were calculated by trigonometry. The radiographic dimensions were then measured by blinded observers using both techniques. RESULTS: For a single radiograph, the human eye was accurate to 0.26 mm and reproducible to +/-0.1 mm. In comparison the digital image analyser system was accurate to 0.01 mm with a reproducibility of +/-0.08 mm. In an arthroplasty model, where the dimensions of an object were corrected for magnification by the known dimensions of a femoral head, the human eye was accurate to 0.19 mm, whereas the image analyser system was accurate to 0.04 mm. CONCLUSIONS: The digital image analysis system is up to 20 times more accurate than the human eye, and in an arthroplasty model the accuracy of measurement increases four-fold. We believe such image analysis may allow more accurate and reproducible measurement of wear from standard follow-up radiographs.

Hip Prosthesis↗
PubMed1992

[Visualized three-dimensional reconstruction and image analysis in orthopedics and trauma surgery].

Computer tomography is a commonly used technique for detecting pathological alterations in soft tissue and the skeleton. The remote access to image informations as well as the allocation of display and processing tools via networks enables improved diagnostic and therapeutic practice in orthopaedic and traumatologic surgery. The realization of a user friendly image analysis system displays and processes the acquired images in a modality oriented manner. Our method is based on a unique file format and specific evaluation procedures to produce the input data for three-dimensional display and algorithms for the individual design of implants. Our image analysis system can process the data of conventional computer tomographies with three or more mm distance. In contrast to the available systems there is a low significance of radiation effects.

Computer Graphics↗
PubMed2006

Biological parametric mapping: A statistical toolbox for multimodality brain image analysis.

In recent years, multiple brain MR imaging modalities have emerged; however, analysis methodologies have mainly remained modality-specific. In addition, when comparing across imaging modalities, most researchers have been forced to rely on simple region-of-interest type analyses, which do not allow the voxel-by-voxel comparisons necessary to answer more sophisticated neuroscience questions. To overcome these limitations, we developed a toolbox for multimodal image analysis called biological parametric mapping (BPM), based on a voxel-wise use of the general linear model. The BPM toolbox incorporates information obtained from other modalities as regressors in a voxel-wise analysis, thereby permitting investigation of more sophisticated hypotheses. The BPM toolbox has been developed in Matlab with a user-friendly interface for performing analyses, including voxel-wise multimodal correlation, ANCOVA, and multiple regression. It has a high degree of integration with the SPM (statistical parametric mapping) software relying on it for visualization and statistical inference. Furthermore, statistical inference for a correlation field, rather than a widely used T-field, has been implemented in the correlation analysis for more accurate results. An example with in vivo data is presented, demonstrating the potential of the BPM methodology as a tool for multimodal image analysis.

Brain↗
PubMed1994

Image analysis of proliferating cells in tumors of the human nervous system: an immunohistological study with the monoclonal antibody Ki-67.

Obtaining growth fractions from immunohistological preparations by the commonly used cell count calculation method is time consuming. For the first time, we investigated and compared the detection of proliferating cells in immunohistologically labeled tissue from tumors of the nervous system using the monoclonal antibody Ki-67 by a new computerized image analysis system and by cell count calculation. The two methods showed a high correlation (correlation index, 0.98) in 37 gliomas (2 pilocytic astrocytomas, 10 Grade II astrocytomas, 5 Grade III astrocytomas, 20 Grade IV astrocytomas and glioblastoma multiforme) and a heterogenous group of 10 additional tumors of the nervous system, including oligodendroglioma, pineoblastoma, primary central nervous system lymphoma, and neurofibroma. Advantages of and indications for image analysis are as follows: 1) time-saving evaluation of immunohistological preparations enables their use in neuro-oncological routine diagnostics and examination of larger cell populations, thus leading to more precisely reproducible results, especially in heterogenous tumors; 2) image analysis, calculating the area of cell nuclei rather than their number, avoids difficulties with fragmented or overlapping nuclei; 3) analysis of different antibodies (for example, Ki-67 and anti-proliferating cell nuclear antigen) may be performed with the same program; 4) investigation of a larger patient group may lead, in combination with the histopathological diagnosis and clinical parameters, to better adapted therapeutic concepts.

Adolescent↗
PubMed2006

HER2/neu expression and gene alterations in pancreatic ductal adenocarcinoma: a comparative immunohistochemistry and chromogenic in situ hybridization study based on tissue microarrays and computerized image analysis.

CONTEXT: HER2/neu overexpression is observed in many cancers including pancreatic ductal adenocarcinoma. Although immunohistochemistry remains the basic method for evaluating HER2/neu protein expression, significant information regarding gene status cannot be assessed. DESIGN: Using tissue microarray technology, fifty histologically confirmed pancreatic ductal adenocarcinomas were cored twice and re-embedded in one paraffin block. Immunohistochemistry (clone TAB 250) and chromogenic (HER2/neu amplification Spot Light kit) in situ hybridization protocols were performed. The immunostained slides were evaluated by conventional eye microscopy and digital image analysis. The chi square test and the kappa statistic were applied by running the SPSS package. MAIN OUTCOME MEASURES: The levels of staining intensity were estimated by the performance of a semi automated image analysis system. RESULTS: HER2/neu gene amplification was detected in 8/50 cases (16%). Chromosome 17 aneuploidy was detected in 19 cases (38%). Significant improvement in interobserver agreement (kappa=0.76 vs. 0.94) was achieved correlating the immunohistochemical results obtained by conventional eye and digital microscopy, especially in the cases of overexpression (2+, 3+). Finally, 29 (58%), 11 (22%), 6 (12%) and 4 (8%) cases were characterized as 0, 1+, 2+ and 3+, respectively. HER2/neu protein expression was significantly associated with grade (P=0.019), but not with stage (P=0.466). in addition, chromosome 17 and gene status were not correlated with stage and grade. CONCLUSION: Our results indicate that a subset of pancreatic ductal adenocarcinomas is characterized by HER2/neu gene amplification. In contrast to breast cancer, protein overexpression does not predict this specific gene deregulation mechanism. This event may reflect the different biological role of the molecule in those two solid tumours, affecting the response to novel targeted agents, such as monoclonal anti-HER2/neu antibodies. Furthermore, evaluation of HER2/neu protein expression based on digital image analysis and not only on conventional eye microscopy improves the accuracy and reliability of immunohistochemical estimation, although that does not demonstrate clinical significance and prognostic value in pancreatic ductal adenocarcinoma.

Aged↗
PubMed1996

Quality control of proliferation marker (MIB-1) in image analysis systems utilizing cell culture-based control materials.

Standard controls for quality control of cell growth (proliferation) assays in image analysis systems are not currently available. The authors have developed a system of controls, based on cultured and harvested human cell lines, that can mimic tissue sections. These controls help ensure quality control for the entire cell proliferation analysis process, from the initial cutting of the paraffin block through fixation, immunohistochemical staining, and interactive image analysis. The use of cell line controls is advantageous because of the greater cell population homogeneity and the volume of uniform slides that are obtainable, as opposed to the use of a heterogeneous tissue sample. This system provides an excellent means of evaluating the day-to-day performance of cell proliferation analysis and may also be adapted for use as a method of multi-institutional proficiency testing.

Biomarkers, Tumor↗
PubMed1981

Structural transitions of chromatin in isolated Xenopus erythrocyte nuclei. II. Computer-based image analysis.

The morphology of isolated Xenopus erythrocyte nuclei is largely determined by the ionic constitution of the isolation medium. Electron microscopy of thin sections of nuclei isolated under varying ionic conditions has revealed four different morphological types with characteristic chromatin configurations. These nuclei have been used to study the condensation of chromatin within the confines of the nucleus. Electron micrographs of thin nuclear sections have been analysed by computer on the basis of their intensity of staining with lead citrate/uranyl acetate. The staining pattern (image analysis scan) is characteristic for each nuclear morphology and can be explained by a model which assumes that there are only four higher-order conformations that chromatin can adopt within the nucleus. These are present in varying amounts in the different nuclear types and an attempt has been made to equate the peaks identified by image analysis with previously-described chromatin structures. The results of this study serve to emphasize the potential of computer-based image analysis in the study of chromatin structure.

Animals↗