Semi-automated Image Analysis Method Research Articles

The impact of dry eye disease on corneal nerve parameters: A systematic review and meta-analysis.

Dry eye disease (DED) is a growing global health problem with a significant impact on the quality of life of patients. While neurosensory abnormalities have been recognised as a contributor to DED pathophysiology, the potential role of in vivo corneal confocal microscopy in detecting nerve loss or damage remains unclear. This systematic review with meta-analysis (PROSPERO registered CRD42022381861) investigated whether DED has an impact on sub-basal corneal nerve parameters. PubMed, Embase and Web of Science Core Collection databases were searched from inception to 9 December 2022. Studies using laser scanning confocal microscopy to compare corneal nerve parameters of DED with healthy eyes were included. Study selection process and data extraction were performed by two independent members of the review team. Twenty-two studies with 916 participants with DED and 491 healthy controls were included, with 21 of these studies included in subsequent meta-analyses. There was a decrease in total corneal nerve length (-3.85 mm/mm2 ; 95% CI -5.16, -2.55), corneal main nerve trunk density (-4.81 number/mm2 ; 95% CI -7.94, -1.68) and corneal nerve branch density (-15.52 number/mm2 ; 95% CI -27.20, -3.84) in DED eyes compared with healthy eyes, with subgroup analysis demonstrating that these differences were more evident in studies using NeuronJ software, a semi-automated procedure. While this review found evidence of loss of corneal nerve parameters in eyes with DED compared with healthy controls, particularly with the use of a semi-automated image analysis method, it is evident that there is substantial heterogeneity between studies in terms of corneal nerve imaging methodology. Standardisation is required in terms of terminology and analysis, with more research needed to potentially improve the clinical applicability and practicality of corneal nerve imaging. Further investigation is also required to confirm the diagnostic accuracy of this imaging modality and its potential for monitoring DED treatment efficacy.

Ultrasound‐based measures in carpal tunnel syndrome: What to measure?

Carpal tunnel syndrome (CTS) is the most common peripheral entrapment neuropathy and is caused by compression of the median nerve (MN) at the level of transverse carpal ligament of the volar wrist. Radiomics is an advanced semi-automated image analysis method that is utilized to identify characteristics in the MN that can detect CTS with considerable reproducibility.

Reliability of a New Semi-automatic Image Analysis Method for Evaluating the Doppler Signal and Intratendinous Vascular Resistance in Patellar Tendinopathy.

The aim of this study was to determine the intra- and inter-rater reliability of a new semi-automatic image analysis method for quantification of the shape of the Doppler signal and the intratendinous vascular resistance in patellar tendinopathy. Thirty athletes (27.4 y, standard deviation=8.57 y) with patellar intratendinous vascularity were included in a cross-sectional study (42 tendons analyzed). The intratendinous blood flow was assessed with power Doppler and ImageJ (Version 1.50b, National Institutes of Health, Bethesda, MD, USA) quantification software over a manually selected region of interest. Two blinded observers performed the analysis of the Doppler signal (vascular resistance) and shape descriptors (number of signals, pixel intensity, area, perimeter, major diameter, minor diameter, circularity and solidity). The intraclass correlation coefficient (ICC) was calculated, and the Bland-Altman mean of differences (MoD) and 95% limits of agreement (LoA) were determined. Also, small real differences (SRDs) and the standard error of measurement (SEM) were calculated. Intra-rater reliability was at a maximum for area (ICC=0.999, 95% confidence interval [CI]=0.998-0.999) and at a minimum for solidity (ICC=0.782, 95% CI: 0.682-0.853). The MoD and 95% LoA were very low, and the relative SRD and SEM were below 5.3% and 2%, respectively. The inter-rater reliability was the maximum for area (ICC=0.993, 95% CI=0.989-0.996) and the minimum for circularity (ICC=0.73; 95% CI=0.611-0.817). The MoD and 95% LoA were low, with the SRD and SEM below 6% and 2.2%. The proposed quantitative method for studying the intratendinous Doppler signal in the patellar tendon is reliable and reproducible.

Impact of Coseismic Frictional Melting on Particle Size, Shape Distribution and Chemistry of Experimentally-Generated Pseudotachylite

In natural friction melts, or pseudotachylites, clast textures and glass compositions can influence the frictional behavior of faults hosting pseudotachylites, and are, in turn, sensitive to the processes involved in pseudotachylite formation. Quantification of these parameters in situations where the host rock composition and formation conditions are well-constrained, such as analogue experiments, may yield calibrations that can be employed in analysis of natural pseudotachylites. In this paper, we experimentally-generated pseudotachylites in granitoid rocks (tonalite and Westerly granite) at Pconf= 40 MPa and slip rates of ∼0.1 m s−1, comparable to the conditions under which natural pseudotachylite is known to form in Earth’s upper crust. We find variations in both clast textures and glass compositions that reflect formation processes, and probably influence the frictional behavior of similar natural faults hosting pseudotachylite. Quantification of particle size and shape distribution with a semi-automatic image analysis method, combined with analysis of glass and host-rock composition of these experimentally generated pseudotachylites, reveals that the textures of pseudotachylite material evolved by combinations of 1) comminution, 2) heterogeneous frictional flash melting, and 3) homogeneous (diffusive) clast melting and/or marginal decrepitation. Fractal dimensions of pseudotachylite-hosted clasts (D ∼ 3) that are greater than those of marginal fragmented host rock particles (gouge, D ∼ 2.4), reflect an increase of the intensity of comminution by slip localisation during a pre-melting phase. Chemical analyses demonstrate that these pseudotachylite glasses were generated by frictional flash melting, where host rock phases melt individually. Biotite is the least resistant to melting, feldspar intermediate, and quartz is the most resistant. The peudotachylite glass generated in these experiments has an alkaline composition, is depleted in SiO2compared to the bulk host-rock, and shows heterogeneous compositions in a single sample related to proximity to host-rock minerals. The percentage contributions of host rock phases to the melt, calculated by a mixing model, shows that glass compositions are dominated by plagioclase and biotite. Within the melt, margins of clasts were dissolved uniformly by diffusion and/or affected by marginal decrepitation, resulting in convex and round shapes with convexities averaging ∼0.8 and circularities averaging ∼0.65.

Measuring the dynamic response of the thylakoid architecture in plant leaves by electron microscopy.

The performance of the photosynthesis machinery in plants, including light harvesting, electron transport, and protein repair, is controlled by structural changes in the thylakoid membrane system inside the chloroplasts. In particular, the structure of the stacked grana area of thylakoid membranes is highly dynamic, changing in response to different environmental cues such as light intensity. For example, the aqueous thylakoid lumen enclosed by thylakoid membranes in grana has been documented to swell in the presence of light. However, light‐induced alteration of the stromal gap in the stacked grana (partition gap) and of the unstacked stroma lamellae has not been well characterized. Light‐induced changes in the entire thylakoid membrane system, including the lumen in both stacked and unstacked domains as well as the partition gap, are presented here, and the functional implications are discussed. This structural analysis was made possible by development of a robust semi‐automated image analysis method combined with optimized plant tissue fixation techniques for transmission electron microscopy generating quantitative structural results for the analysis of thylakoid ultrastructure.Significance StatementA methodical pipeline ranging from optimized leaf tissue preparation for electron microscopy to quantitative image analysis was established. This methodical development was employed to study details of light‐induced changes in the plant thylakoid ultrastructure. It was found that the lumen of the entire thylakoid system (stacked and unstacked domains) undergoes light‐induced swelling, whereas adjacent membranes on the stroma side in stacked grana thylakoid approach each other.

The role of imaging in early detection of patients at risk for neuropathy during primary chemotherapy for ovarian carcinoma.

e18071 Background: Neuropathy is a common adverse event requiring dose reduction or regimen change in patients receiving platinum-taxane chemotherapy for ovarian cancer. Our aim was to determine whether specific biomarkers, such as skeletal muscle, visceral and subcutaneous fat cross-sectional areas, measured using computerized tomography (CT), were associated with the development of neuropathy. Analytic Morphomics, a semi-automated image analysis method, has been used to predict oncologic outcomes with immunotherapy and to identify patients at higher risk for postoperative complications from major cancer surgeries. We hypothesized that patients who required dose reductions or regimen changes would have CT imaging findings that distinguished them from their counterparts who did not require dose reduction or regimen change. Methods: All patients at our institution diagnosed with high-grade ovarian carcinoma between January 1, 2014 and December 31, 2018 were identified retrospectively. Analytic Morphomics was used to extract measures of body composition from CT images at the level of the third lumbar vertebra. Demographic and clinical characteristics were abstracted from the electronic medical record. We analyzed several binary outcomes (neuropathy, delay in chemotherapy, or dose reduction by cycle 1, 3 or 6) and used unpaired, two-tailed t-tests to compare means between each outcome group at each timepoint. Results: Of 156 patients screened for the study, 138 patients were included and had pre-treatment CT scans available and completed six cycles of platinum-taxane chemotherapy. Patients who developed neuropathy in any of the six cycles, or had a change in their chemotherapy regimen by cycle six, had higher mean visceral fat area than those who did not develop neuropathy. This finding was statistically significant throughout three cycles (p = 0.023) and six cycles (p = 0.030). Conclusions: Increased visceral fat on CT imaging was significantly associated with a higher incidence of neuropathy and change in chemotherapy regimen in patients undergoing primary platinum-taxane chemotherapy for ovarian carcinoma. Weight and BMI did not have a significant association with development of neuropathy, dose reduction, or treatment delay. Further study is needed to determine if dose reductions or regimen changes due to neuropathy can be better predicted using morphomics.

A method for semi-automated image analysis of HLA class I tumour epithelium expression in rectal cancer.

Biomarkers may hold the key towards development and improvement of personalized cancer treatment. For instance, tumour expression of immune system-related proteins may reveal the tumour immune status and, accordingly, determine choice for type of immunotherapy. Therefore, objective evaluation of tumour biomarker expression is needed but often challenging. For instance, human leukocyte antigen (HLA) class I tumour epithelium expression is cumbersome to quantify by eye due to its presence on both tumour epithelial cells and tumour stromal cells, as well as tumourinfiltrating immune cells. In this study, we solved this problem by setting up an immunohistochemical (IHC) double staining using a tissue microarray (TMA) of rectal tumours wherein HLA class I expression was coloured with a blue chromogen, whereas non-epithelial tissue was visualized with a brown chromogen. We subsequently developed a semi-automated image analysis method that identified tumour epithelium as well as the percentage of HLA class I-positive tumour epithelium. Using this technique, we compared HCA2/HC10 and EMR8-5 antibodies for the assessment of HLA class I tumour expression and concluded that EMR8-5 is the superior antibody for this purpose. This IHC double staining can in principle be used for scoring of any biomarker expressed by tumour epithelium.

Abstract P5-07-01: Computational scoring of tumor infiltrating lymphocytes in triple-negative breast cancer

Abstract Background: Stromal Tumor Infiltrating Lymphocytes (sTIL) are an established prognostic feature in triple-negative breast cancer, yet manual assessment or visual estimation of sTILs with conventional light microscopy may be subject to inter-pathologist variability. Recently published guidelines by the International TIL Working Group help address inter-pathologist variability, yet there remains a need for more objective and quantitative computational sTIL scoring. Methods: Our study used 120 triple-negative breast cancer slides (one slide per patient). A deep-learning based image analysis workflow is used to perform segmentation and classification of tissue regions and cells on the digital whole slide image. We used 14 annotated slides to train and validate the deep learning model, and to obtain image segmentation and classification accuracy statistics. Non-training slides were used to evaluate the concordance of manual (m-sTIL) and computationally derived (c-sTIL) scores. To generate data to create the model we manually annotated tissue regions in FFPE H&E stained digital slides, including: tumor, stroma, and necrosis. Initial classification of cell nuclei was performed using a semi-automated image analysis method, and then manually corrected to generate ground truth for tumor, stroma (fibroblasts), and lymphocytes. All annotations were performed by a trained research fellow and reviewed by a board-certified pathologist. Corresponding region and nucleus-level annotations were combined to train and validate a fully-convolutional neural network that jointly classifies tissue regions and cell nuclei. Tissue region segmentation accuracy was assessed by the Dice coefficient to measure degree of overlap between predicted tissue regions and ground truth annotations. Cell classification accuracy was assessed using area under curve (AUC). Two board-certified pathologists independently generated an m-sTIL score for all slides according to clinical guidelines, and discrepancies between pathologists were resolved by consensus. c-sTIL scores were calculated as the percentage of classified stromal areas occupied by nuclei classified as lymphocytic infiltrates. Results: Tissue region segmentation was accurate for both stroma (0.77 Dice) and tumor (0.83 Dice) regions, and accurate overall (0.78 Dice). Cell classification was highly accurate for lymphocytes (0.89 AUC), tumor cells (0.90 AUC), stromal cells (0.78 AUC), and overall (0.89 AUC, micro average). Inter observer spearman correlation between the m-sTIL scores of our two pathologists was 0.66 (p < 0.001). By comparison, the correlation between c-sTIL and consensus m-sTIL was higher at 0.73 (p < 0.001). Dichotomizing at a threshold sTIL score of 10%, c-sTIL scoring identifies low-sTIL patients with an accuracy of 85%. High- and Low- sTIL score patient groups show clear separation on a Kaplan-Meier curve for both c-sTIL and m-sTIL scoring approaches. Conclusions: Our pipeline quantifies stromal TILs with high concordance with manual pathologist scores, and sheds light on the ability of computational approaches in standardizing diagnostic pathology workflows. Future work will investigate how other computationally driven histology biomarkers can predict outcomes and help prognosticate breast cancer patients. Citation Format: Amgad M, Sarkar A, Srinivas C, Redman R, Ratra S, Bechert CJ, Calhoun BC, Mrazeck K, Kurkure U, Cooper LA, Barnes M. Computational scoring of tumor infiltrating lymphocytes in triple-negative breast cancer [abstract]. In: Proceedings of the 2018 San Antonio Breast Cancer Symposium; 2018 Dec 4-8; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2019;79(4 Suppl):Abstract nr P5-07-01.

An innovative method to quantitate tissue integration of hyaluronic acid-based dermal fillers.

Following intradermal injection, hyaluronic acid (HA)-based fillers tend to spread within the reticular dermis and to distribute between the dermal fibers. This biointegration is commonly measured qualitatively using histological methods. We developed a "toolbox" consisting of a visual scoring and a semi-automatic image analysis method using internal developed algorithm to quantitate the biointegration of Restylane® in histological sections. Restylane® was injected intradermally in the abdominal skin of 10 healthy human subjects scheduled for abdominoplasty. The injections were performed either in vivo before surgery or ex vivo on samples taken post-surgery at different time points. The samples were processed for histology by visual scoring and image analysis using algorithms developed in Definiens to assess biointegration. The image analysis segmentation was accurate with <5% manual changes. Furthermore, the results calculated with the semi-automatic method were consistent with the visual scores obtained on injected human skin samples by means of a 5-grade photographic scale. A modified hematoxylin-eosin staining was found adequate to visualize both, the filler and the general morphology, on the same section. An excellent correlation was observed between the integration results obtained with PAS/Alcian Blue and HE-stained slides, allowing for a single staining in future studies. We developed a modified HE staining histological method and a new histomorphometric image analysis tool to quantitate biointegration of HA-based fillers in human skin. The results obtained in this study confirmed the known intermediate biointegration properties of Restylane®, thus validating these innovative methods.

Assessment of nematode biomass in marine sediments: A semi-automated image analysis method

Assessment of nematode biomass in marine sediments: A semi-automated image analysis method

3D characterization of rolling contact fatigue crack networks

Rolling contact fatigue (RCF) damage is becoming more frequent with increased traffic, accelerations, and loading conditions in the railway industry. Defects which are characterized by a two-lobe darkened surface and a V-shaped surface-breaking crack are defined as squats. The origination and propagation of squats in railway rails is the topic of many recent studies; the associated crack networks develop with complicated geometry near the surface of rails, but can be difficult to detect and distinguish from normally existing head checks in their early stages, using in-field non-destructive detection techniques. After cutting out damaged sections of rail, there are a number of options to characterize the damage. The aim of this study was to evaluate different methods to geometrically describe squat crack networks; through X-ray radiography complemented with geometrical reconstruction, metallography, X-ray tomography, and topography measurements. The experiments were performed on squats from rail sections taken from the field. In the first method, high-resolution and high-energy X-ray images exposed through the entire rail head from a range of angles were combined using a semi-automated image analysis method for geometrical reconstruction, and a 3D representation of the complex crack network was achieved. This was compared with measurements on cross-sections after repeated metallographic sectioning to determine the accuracy of prediction of the geometrical reconstruction. A second squat was investigated by X-ray tomography after extraction of a section of the rail head. A third squat was opened by careful cutting, which gave full access to the crack faces, and the topography was measured by stylus profilometry. The high-energy X-ray, 3D reconstruction method showed accurate main crack geometry at medium depths; the advantage of the method being that it potentially could be developed for non-destructive testing in field. However significant drawbacks exist due to limitations in radiography in terms of detecting tightly closed cracks in very thick components. This includes the inability to detect the crack tips which is an important factor in determining the risks associated to a specific crack. Metallographic investigation of the cracks gave good interpretation of crack geometry along the sections examined, and gave the possibility to study microstructure and plastic deformation adjacent to the crack face. However this time-consuming method requires destruction of the specimen investigated. The X-ray tomography revealed the 3D crack network including side branches in a 10×10×30mm3 sample, and provided topographic information without completely opening the squat. Topography measurements acquired by stylus profilometry provided an accurate description of the entire main crack surface texture, including features such as surface ridges and beach marks.

Evaluation and Validation of a Semiautomatic Image Analysis Method: Quantification of Nares Cell Proliferation of Rhinella arenarum (Anura) Detected with the Avidin-Biotin Complex/Peroxidase, DAB

The current report presents the development and application of a novel methodological approach for computer-based methods of processing and analysis of proliferative tissues labeled by ABC-peroxidase method using 3, 3′-diaminobenzidine (DAB) as chromogen. This semiautomatic method is proposed to replace the classical manual approach, widely accepted as gold standard. Our method is based on a visual analysis of the microscopy image features from which a computational model is built to generate synthetic images which are used to evaluate and validate the methods of image processing and analysis. The evaluation allows knowing whether the computational methods applied are affected by the change of the image characteristics. Validation allows determining the method’s reliability and analyzing the concordance between the proposed method and a gold standard one. Additional strongness of this new approach is that it may be a framework adaptable to other studies made on any kind of microscopy.

A sensitive, reproducible and objective immunofluorescence analysis method of dystrophin in individual fibers in samples from patients with duchenne muscular dystrophy.

Duchenne muscular dystrophy (DMD) is characterized by the absence or reduced levels of dystrophin expression on the inner surface of the sarcolemmal membrane of muscle fibers. Clinical development of therapeutic approaches aiming to increase dystrophin levels requires sensitive and reproducible measurement of differences in dystrophin expression in muscle biopsies of treated patients with DMD. This, however, poses a technical challenge due to intra- and inter-donor variance in the occurrence of revertant fibers and low trace dystrophin expression throughout the biopsies. We have developed an immunofluorescence and semi-automated image analysis method that measures the sarcolemmal dystrophin intensity per individual fiber for the entire fiber population in a muscle biopsy. Cross-sections of muscle co-stained for dystrophin and spectrin have been imaged by confocal microscopy, and image analysis was performed using Definiens software. Dystrophin intensity has been measured in the sarcolemmal mask of spectrin for each individual muscle fiber and multiple membrane intensity parameters (mean, maximum, quantiles per fiber) were calculated. A histogram can depict the distribution of dystrophin intensities for the fiber population in the biopsy. This method was tested by measuring dystrophin in DMD, Becker muscular dystrophy, and healthy muscle samples. Analysis of duplicate or quadruplicate sections of DMD biopsies on the same or multiple days, by different operators, or using different antibodies, was shown to be objective and reproducible (inter-assay precision, CV 2–17% and intra-assay precision, CV 2–10%). Moreover, the method was sufficiently sensitive to detect consistently small differences in dystrophin between two biopsies from a patient with DMD before and after treatment with an investigational compound.

Retinal Vessel Diameter Assessment in Papilledema by Semi-Automated Analysis of SLO Images: Feasibility and Reliability

To report feasibility and reliability of a semi-automated image analysis method for retinal vessel diameter measurements in subjects with papilledema before and after treatment. Scanning laser ophthalmoscopy (SLO) was performed in seven normal, five pseudopapilledema, and seven papilledema subjects. In four papilledema subjects, SLO was performed both before and following treatment. Two observers measured diameters of superior and inferior retinal arteries and veins from SLO images using two methods: manual analysis and semi-automated customized analysis. Vessel measurements were compared between observers and between image analysis methods. Retinal vein and artery diameters for each subject were compared between papilledema, pseudopapilledema, and normal subjects, and before and following treatment for papilledema subjects. Interobserver reliability was 0.97 (Pearson's correlation, r) and 0.90 for semi-automated and manual measurements, respectively. Correlation coefficient of manual and semi-automated measurements was 0.85. Retinal vein diameter in papilledema subjects was larger than in pseudopapilledema and normal subjects (P = 0.03, 0.04, Mann-Whitney). Papilledema subjects had a decrease in retinal vein diameter following treatment for and resolution of papilledema (P = 0.04, Wilcoxon signed rank). Retinal artery diameters were not significantly different between papilledema and pseudopapilledema or normal groups, and did not significantly change following papilledema treatment. A feasible and reliable semi-automated image analysis method for measurement of retinal artery and vein diameters from SLO images of elevated optic nerves is reported. Further studies are needed to determine the clinical utility of retinal vein diameter measurements as a marker for diagnosis and treatment of papilledema.

Semi-Automated Image Analysis of Xenopus Laevis Behavioral Response to Visual Stimuli

High-throughput screens to measure the behavioural effects of various genetic or pharmacological manipulations will require the development of assays that do not rely on subjective human observer input. The Xenopus laevis tadpole is a low-cost, easily accessible vertebrate animal model for studying brain development. We present a new semi-automated image analysis method to quantify avoidance swimming behaviour by multiple tadpoles in parallel. In our setup, Xenopus tadpoles swim in a water-filled Petri dish placed on a computer screen which displays moving dots as escape-evoking visual stimuli on a grey background. Each experiment is recorded by a camera as an image time series. An algorithm was developed to precisely track tadpole movements in space and time. In the control experiments, presenting only the background grey field, we measured the speed distribution of tadpole displacements per image interval. This distribution was used as a reference to assess the tadpole behavioural responses to contrast varying visual stimuli. In experiments where stimuli are brighter or darker with respect to the grey background intensity, we mapped the spatial distribution of responses as a function of the closest dot position with respect to tadpole orientation. We observed a greater response concentration within an area of 5 mm radius around the tadpole. By defining potential reaction events within this threshold minimal distance, we could evaluate the response probability for each tadpole. Our results show a higher tadpole response probability to darker dots than to brighter dots. Furthermore, we show an angular dependence of the response probability that varies with dot contrast. We conclude that this tracking and analysis technique can be applied to a variety of behavioural studies, enabling a rapid and more objective, automated evaluation of visuomotor responses under a wide range of experimental manipulations.

Quantitative analysis of patch patterns in mosaic tissues with ClonalTools software.

Quantitative analysis of mosaic tissues is a powerful method for following developmental lineages; however, analytical techniques are often subjective and repetitious. Here a flexible, semi-automated image analysis method for mosaic patterns is described. ClonalTools is a free customizable tool-set designed for the open-source image analysis package ImageJ. Circular, polygonal or linear one-dimensional mosaic arrays can be interrogated to provide measurements of the total number and width of positive and negative patches in a region of interest. These results are adjusted for the effects of random clumping using a previously described method to correct for differences in the contribution of the positive and negative cell type. The applicability of ClonalTools to different systems is discussed with reference to the analysis of mosaic patterns in the mouse corneal epithelium and adrenal cortex and in the outgrowth of neurites from explant cultures of mouse retina as example systems. To validate ClonalTools quantitatively, a recently published manual clonal analysis of the corneal epithelium of X-inactivation beta-Gal-mosaic mice was re-analysed. The semi-automated results did not differ significantly from the published data. Rapid quantification of such patterns to produce biologically relevant results represents a welcome improvement in terms of ease and speed of use over previous methods.

Changes of Dextrose Particles with Pneumatic Conveying: Analysis of Size and Shape

When conveying particulate materials, changes in size and shape of individual particles can be observed. These changes can have a great impact on the bulk powder and affect its flow properties in the pipeline. Changes can be wanted or unwanted depending on the whole process chain and final use of the powder. In this investigation, dextrose monohydrate particles were pneumatically conveyed repetitively in a pilot plant–scale rig, and the size and shape of the particles were characterized by a semiautomatic image analysis method. This characterization was done qualitatively by observing micrographs and quantitatively for each individual particle (a total of 16,120 dextrose crystals) by using two statistical diameters and two shape factors. The effect of the changes in shape and size of the particles on the mass flow rate in the system was studied. It was concluded that the mass flow rate of the pneumatic conveying tests was not affected considerably by the changes of the dextrose particles.

Relationship between PCNA Proliferating Marker and Angiogenesis

The goal of the study was to evaluate the prognosis significance of intratumor angiogenesis that was compared with PCNA proliferating marker, histology grade, mitotic index, tumor size, and histology type. The study had been realized using different histology types of bitch mammary tumors according to WHO classifications. There were evaluated some potential angiogenesis markers, such as intratumor microvessel density, total microvascular area and perimeter according to microscopic tumor area, and average vessel area and perimeter in different benign and malign mammary tumors. The study was realized in 8 bitch mammary tumors represented by two benign lesions (simple adenoma and fibroadenoma) and 6 carcinomas (grade I, II and III tumors). From all tumors had been harvested fragments represented by 5 mm slices that were fixed in 10% buffered formalin and proceeded by paraffin technique. The slides were stained by usual methods (HE and TM). To realize immunohistochemistry reaction paraffin embedded slices were attached onto silanized glass slides (Dako). PCNA proliferating marker (clone PC10, izotype IgG2a kappa - Dako) was evaluated by counting by 3 evaluators 8-10 high power magnified fields (400x), about 1000 cells (immunomarked nuclei). There was established average PCNA percentage for each tumor type. Intratumor angiogenesis had been established using CD31 marker (clone JC70A, izotype IgG1 kappa - Dako) and evaluated using semiautomatic image analysis method. Both immunohistochemical reactions utilized LSAB technique. To evaluate new methods necessary to establish bitch mammary tumors malignancy, which is important for mammary cancer prognosis, we utilized several malignancy markers. Also, because intratumor angiogenesis quantification provided contradictory results obtained by many authors, this parameter was related and compared with classic and reliable markers (PCNA antigen, mitotic index, histology grade, tumor size, and tumors’ characteristics).

Microcompounding of organoclay–ABS/PA6 blend‐based nanocomposites

AbstractIn this study, acrylonitrile–butadiene–styrene (ABS) and polyamide‐6 (PA6) were blended in the presence of an olefin‐based compatibilizer and organoclays. The effects of ABS to PA6 ratio, clay content, and screw speed of the microcompounder were examined by performing morphological (i.e., XRD, SEM, and TEM) and tensile tests. The average aspect ratio of the clay platelets after processing was obtained by applying semiautomatic image analysis method. SEM analysis showed that addition of the compatibilizers to the ABS/PA6 blend system resulted in a decrease in diameter of dispersed phase when one of the phases was continuous. The addition of 5 wt% compatibilizer altered the dispersed morphology to cocontinuous morphology when the weight percentage of ABS was equal to that of PA6. The results of XRD analysis implied that clays were exfoliated in the presence of PA6. It was observed in TEM micrographs that clays were selectively dispersed in PA6 phase. Aspect ratio of the platelets increased as the PA6 content increased. Moduli of the nanocomposites were improved by enriching blend with PA6 and increasing screw speed. POLYM. COMPOS., 2008. © 2008 Society of Plastics Engineers

Clinical Measurement of Prognostic Immune Signature in Follicular Lymphoma by RT-PCR Based Gene Expression Profiling and Immunohistochemistry Demonstrates Favourable T-Cell and Unfavorable Macrophage Infiltration.

Gene expression profiling studies have demonstrated immune response gene signatures predictive of outcome in follicular lymphoma (FL) and there is a need for validation of these signatures and for their translation to clinical use. However, measurement of these genes in routine practice remains difficult and to date there have been very few studies validating the hypothesis. We have previously demonstrated the utility of real-time PCR measurement of gene expression levels in globally amplified polyA cDNA as a clinically practical method for translation of gene signatures to clinical use. In this project we extended the method to analysis of immune response signatures in FL. We used real-time PCR to measure expression levels (normalised to the mean of 4 housekeeping genes) of 36 candidate Indicator genes, selected from microarray studies, in polyA cDNAs prepared using polyA PCR (method detailed in Sakhinia et al 2007) from 58 archived human frozen lymph nodes, together with immunohistochemistry for CD3, CD4, CD7, CD8, CD10, CD20, CD21 and CD68 in parallel formalin fixed paraffin embedded tissue samples to measure immune response in FL. Immunohistochemical positivity was measured by a semi-automated image analysis method using spectral unmixing to identify areas of immunopositivity. Kaplan-Mier survival analysis was performed against the normalised real-time PCR expression levels of each of the genes and against the percentage immunohistochemical postivity for CD3, CD4, CD7, CD8, CD10, CD20, CD21; for CD68 survival analysis was performed for cases with either 15 or less or more than 15 CD68 positive cells per high power field (hpf). High levels of CCR1, a marker of monocyte actication, were associated with a shorter survival interval (p<0.02) (figure 1a), whilst immunohistochemistry demonstrated association of high numbers of CD7 positive T-cells with longer survival interval (figure 1b) (p<0.032) and of high numbers of CD68 positive macrophages with a shorter survival interval (figure 1c) (p<0.02). The results confirm the role of the host immune response in outcome in FL and identify CCR1 as a prognostic indicator and marker of immune switch between macrophage and T-cell dominant response. The methods used are clinically applicable, whilst the clinical utility of polyA DNA and real-time PCR for measurement of gene signatures and the strength of this approach as a “molecular block” are confirmed. [Display omitted]