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Sample records for histopathology tissue samples

  1. A low dimensional entropy-based descriptor of several tissues in skin cancer histopathology samples

    NASA Astrophysics Data System (ADS)

    Álvarez, Pablo; Corredor, Germán.; García-Arteaga, Juan D.; Romero, Eduardo

    2015-12-01

    The use of low-level visual features to assign high level labels in datasets of histopathology images is a possible solution to the problems derived from manual labeling by experts. However, in many cases, the visual cues are not enough. In this article we propose the use of features derived exclusively from the spatial distribution of the cell nuclei. These features are calculated using the weight of k-nn graphs constructed from the distances between cells. Results show that there are k values with enhanced discriminatory power, especially when comparing cancerous and non-cancerous tissue.

  2. Spectral topography of histopathological samples

    NASA Astrophysics Data System (ADS)

    Lerner, Jeremy M.; Lu, Thomas T.; Vari, Sandor G.

    1998-06-01

    The goal of imaging spectroscopy is to obtain independent spectra from individual objects in a field-of-view. In the case of biological materials, such as histopathology samples, it has been well established that spectral characteristic can be indicative of specific diseases including cancer. Diagnosis can be enhanced by the use of probes and stains to indicate the presence of individual genome or other biologically active cell components or substances. To assess a specimen through a microscope is directly analogous to serving the Earth from space to assess natural features. This paper describes a simple and inexpensive imaging spectrometer, with an origin in remote sensing, that demonstrates that it is possible to rapidly identify evidence of disease in histopathology samples using spatially resolved spectral data. The PARISS imaging spectrometer enables a researcher to acquire multi-spectral images that yield functional maps, showing what and where biological molecules are located within a structure. It is the powerful combination of imaging and spectroscopy that provides the tools not readily available to the Life Sciences. The PARISS system incorporates a powerful hybrid neural network analysis to break the data logjam that is often associated with the acquisition and processing of multiple spectra.

  3. Histopathological evaluation of tissue undergoing thermal insult

    PubMed Central

    Chaudhary, Minal; Bonde, Dushyant; Patil, Swati; Gawande, Madhuri; Hande, Alka; Jain, Deepali

    2016-01-01

    Context: Thermal insult is the major cause of thermal injury or death and in case of death due to thermal injury the body often has to be recovered from the site. Histologically, one can predict whether the victim was alive or dead when the fire was on going. However, determination of probable cause of thermal insult to which victim subjected to be difficult when the victim's body is found somewhere else from the crime scene or accident site or found alone. Hence, histopathological evaluation of the tissue which has undergone thermal insult in such conditions could help to place evidence in front of law officials, regarding probable condition, or scenario at time of burn of victim. Aims: Keeping this as a criteria in this study we aim to evaluate burnt tissue histopathologically, that undergone various degree of thermal insult, which simulates various real life scenario for mortality in burn cases. Settings and Design: We evaluate the changes in hematoxylin and eosin staining pattern of tissue which has undergone thermal insult compared to normal tissue and also the progressive changes in staining pattern, architectural, and cellular details. Materials and Methods: Samples were taken from the patients, in various surgical procedures. Each sample was cut into five parts with close margins so that each burnt tissue is evaluated for same field or region. The tissue that obtained was immediately subjected to varying degree of temperature over a specific period so as to simulate the various real-life condition. Then the tissues were fixed, processed, and stained with routine H and E staining. The processed slides of tissue were examined under the microscope, and the staining, and architectural changes were evaluated and described. Results: Results show that there was a progressive changes in the architectural pattern of the epithelium and connective tissue showing cleft formation and vacuolization, staining pattern also shows mixing of stains progressively as the

  4. Critical steps in tissue processing in histopathology.

    PubMed

    Comanescu, Maria; Annaratone, Laura; D'Armento, Giuseppe; Cardos, Georgeta; Sapino, Anna; Bussolati, Gianni

    2012-04-01

    Histopathological diagnosis using Formalin-Fixed Paraffin Embedded (FFPE) tissues is essential for the prognostic and therapeutic management of cancer patients. Pathologists are being confronted with increasing demands, from both clinicians and patients, to provide immunophenotypic and gene expression data from FFPE tissues to allow the planning of personalized therapeutic regimens. Recent improvements in the protocols for pre-analysis processing of pathological tissues aim to better preserve cellular details and to conserve antigens and nucleic acid sequences. These developments have been recently patented. The international protocol for the transporting of surgical specimens from the surgical theatre to the pathology department is to immerse the specimen in formalin. The alternative method of sealing the specimens into bags under a vacuum and then cooling is a well-accepted and environmentally safe procedure that overcomes the many drawbacks linked to transfer in formalin. Importantly, RNA is notoriously poorly preserved in FFPE tissue. Due to this, successful procedures for the extraction of genetic information from archival tissues have been the object of several studies and patents. Novel molecular approaches for RT-qPCR and gene array analysis on FFPE tissues are presented here. Moreover, a major advance is reported in this study, the observation that tissue fixation in cold conditions allows a much better preservation of nucleic acid sequences.

  5. Effect of substrate choice and tissue type on tissue preparation for spectral histopathology by Raman microspectroscopy.

    PubMed

    Fullwood, Leanne M; Griffiths, Dave; Ashton, Katherine; Dawson, Timothy; Lea, Robert W; Davis, Charles; Bonnier, Franck; Byrne, Hugh J; Baker, Matthew J

    2014-01-21

    Raman spectroscopy is a non-destructive, non-invasive, rapid and economical technique which has the potential to be an excellent method for the diagnosis of cancer and understanding disease progression through retrospective studies of archived tissue samples. Historically, biobanks are generally comprised of formalin fixed paraffin preserved tissue and as a result these specimens are often used in spectroscopic research. Tissue in this state has to be dewaxed prior to Raman analysis to reduce paraffin contributions in the spectra. However, although the procedures are derived from histopathological clinical practice, the efficacy of the dewaxing procedures that are currently employed is questionable. Ineffective removal of paraffin results in corruption of the spectra and previous experiments have shown that the efficacy can depend on the dewaxing medium and processing time. The aim of this study was to investigate the influence of commonly used spectroscopic substrates (CaF2, Spectrosil quartz and low-E slides) and the influence of different histological tissue types (normal, cancerous and metastatic) on tissue preparation and to assess their use for spectral histopathology. Results show that CaF2 followed by Spectrosil contribute the least to the spectral background. However, both substrates retain paraffin after dewaxing. Low-E substrates, which exhibit the most intense spectral background, do not retain wax and resulting spectra are not affected by paraffin peaks. We also show a disparity in paraffin retention depending upon the histological identity of the tissue with abnormal tissue retaining more paraffin than normal.

  6. Histopathological findings, phenotyping of inflammatory cells, and expression of markers of nitritative injury in joint tissue samples from calves after vaccination and intraarticular challenge with Mycoplasma bovis strain 1067

    PubMed Central

    2014-01-01

    Background The pathogenesis of caseonecrotic lesions developing in lungs and joints of calves infected with Mycoplasma bovis is not clear and attempts to prevent M. bovis-induced disease by vaccines have been largely unsuccessful. In this investigation, joint samples from 4 calves, i.e. 2 vaccinated and 2 non-vaccinated, of a vaccination experiment with intraarticular challenge were examined. The aim was to characterize the histopathological findings, the phenotypes of inflammatory cells, the expression of class II major histocompatibility complex (MHC class II) molecules, and the expression of markers for nitritative stress, i.e. inducible nitric oxide synthase (iNOS) and nitrotyrosine (NT), in synovial membrane samples from these calves. Furthermore, the samples were examined for M. bovis antigens including variable surface protein (Vsp) antigens and M. bovis organisms by cultivation techniques. Results The inoculated joints of all 4 calves had caseonecrotic and inflammatory lesions. Necrotic foci were demarcated by phagocytic cells, i.e. macrophages and neutrophilic granulocytes, and by T and B lymphocytes. The presence of M. bovis antigens in necrotic tissue lesions was associated with expression of iNOS and NT by macrophages. Only single macrophages demarcating the necrotic foci were positive for MHC class II. Microbiological results revealed that M. bovis had spread to approximately 27% of the non-inoculated joints. Differences in extent or severity between the lesions in samples from vaccinated and non-vaccinated animals were not seen. Conclusions The results suggest that nitritative injury, as in pneumonic lung tissue of M. bovis-infected calves, is involved in the development of caseonecrotic joint lesions. Only single macrophages were positive for MHC class II indicating down-regulation of antigen-presenting mechanisms possibly caused by local production of iNOS and NO by infiltrating macrophages. PMID:25162202

  7. DNA qualification workflow for next generation sequencing of histopathological samples.

    PubMed

    Simbolo, Michele; Gottardi, Marisa; Corbo, Vincenzo; Fassan, Matteo; Mafficini, Andrea; Malpeli, Giorgio; Lawlor, Rita T; Scarpa, Aldo

    2013-01-01

    Histopathological samples are a treasure-trove of DNA for clinical research. However, the quality of DNA can vary depending on the source or extraction method applied. Thus a standardized and cost-effective workflow for the qualification of DNA preparations is essential to guarantee interlaboratory reproducible results. The qualification process consists of the quantification of double strand DNA (dsDNA) and the assessment of its suitability for downstream applications, such as high-throughput next-generation sequencing. We tested the two most frequently used instrumentations to define their role in this process: NanoDrop, based on UV spectroscopy, and Qubit 2.0, which uses fluorochromes specifically binding dsDNA. Quantitative PCR (qPCR) was used as the reference technique as it simultaneously assesses DNA concentration and suitability for PCR amplification. We used 17 genomic DNAs from 6 fresh-frozen (FF) tissues, 6 formalin-fixed paraffin-embedded (FFPE) tissues, 3 cell lines, and 2 commercial preparations. Intra- and inter-operator variability was negligible, and intra-methodology variability was minimal, while consistent inter-methodology divergences were observed. In fact, NanoDrop measured DNA concentrations higher than Qubit and its consistency with dsDNA quantification by qPCR was limited to high molecular weight DNA from FF samples and cell lines, where total DNA and dsDNA quantity virtually coincide. In partially degraded DNA from FFPE samples, only Qubit proved highly reproducible and consistent with qPCR measurements. Multiplex PCR amplifying 191 regions of 46 cancer-related genes was designated the downstream application, using 40 ng dsDNA from FFPE samples calculated by Qubit. All but one sample produced amplicon libraries suitable for next-generation sequencing. NanoDrop UV-spectrum verified contamination of the unsuccessful sample. In conclusion, as qPCR has high costs and is labor intensive, an alternative effective standard workflow for

  8. DNA qualification workflow for next generation sequencing of histopathological samples.

    PubMed

    Simbolo, Michele; Gottardi, Marisa; Corbo, Vincenzo; Fassan, Matteo; Mafficini, Andrea; Malpeli, Giorgio; Lawlor, Rita T; Scarpa, Aldo

    2013-01-01

    Histopathological samples are a treasure-trove of DNA for clinical research. However, the quality of DNA can vary depending on the source or extraction method applied. Thus a standardized and cost-effective workflow for the qualification of DNA preparations is essential to guarantee interlaboratory reproducible results. The qualification process consists of the quantification of double strand DNA (dsDNA) and the assessment of its suitability for downstream applications, such as high-throughput next-generation sequencing. We tested the two most frequently used instrumentations to define their role in this process: NanoDrop, based on UV spectroscopy, and Qubit 2.0, which uses fluorochromes specifically binding dsDNA. Quantitative PCR (qPCR) was used as the reference technique as it simultaneously assesses DNA concentration and suitability for PCR amplification. We used 17 genomic DNAs from 6 fresh-frozen (FF) tissues, 6 formalin-fixed paraffin-embedded (FFPE) tissues, 3 cell lines, and 2 commercial preparations. Intra- and inter-operator variability was negligible, and intra-methodology variability was minimal, while consistent inter-methodology divergences were observed. In fact, NanoDrop measured DNA concentrations higher than Qubit and its consistency with dsDNA quantification by qPCR was limited to high molecular weight DNA from FF samples and cell lines, where total DNA and dsDNA quantity virtually coincide. In partially degraded DNA from FFPE samples, only Qubit proved highly reproducible and consistent with qPCR measurements. Multiplex PCR amplifying 191 regions of 46 cancer-related genes was designated the downstream application, using 40 ng dsDNA from FFPE samples calculated by Qubit. All but one sample produced amplicon libraries suitable for next-generation sequencing. NanoDrop UV-spectrum verified contamination of the unsuccessful sample. In conclusion, as qPCR has high costs and is labor intensive, an alternative effective standard workflow for

  9. Establishing multi-modality datasets with the incorporation of 3D histopathology for soft tissue classification

    NASA Astrophysics Data System (ADS)

    de Ryk, Jessica; Thiesse, Jacqueline; Reinhardt, Joseph M.; Hoffman, Eric A.; McLennan, Geoffrey

    2006-03-01

    The development of multi-modality image analysis has gained increasing popularity over recent years. Multi-modality image databases are being developed to benefit patient clinical care, research and education. The incorporation of histopathology in these multi-modality datasets is complicated by the large differences in image quality, content and spatial association. We have developed a novel system, the large-scale image microtome array (LIMA), to bridge the gap between non-structurally destructive and destructive imaging such that reliable registration and incorporation of three-dimensional (3D) histopathology can be achieved. We have developed registration algorithms to align the micro-CT, LIMA and histopathology data to a common coordinate system. Using this multi-modality image dataset we have developed a classification algorithm to identify on a pixel basis, the tissue types present. The output from the classification processing is a 3D color coded map of tissue distributions. The resulting complete dataset provides an abundance of valuable information relating to the tissue sample including density, anatomical structure, color, texture and cellular information in three dimensions. In this study we have chosen to use normal and diseased lung tissue, however the flexibility of the image acquisition and subsequent processing algorithms makes it applicable to any soft organ tissue.

  10. Tissue Sampling Guides for Porcine Biomedical Models.

    PubMed

    Albl, Barbara; Haesner, Serena; Braun-Reichhart, Christina; Streckel, Elisabeth; Renner, Simone; Seeliger, Frank; Wolf, Eckhard; Wanke, Rüdiger; Blutke, Andreas

    2016-04-01

    This article provides guidelines for organ and tissue sampling adapted to porcine animal models in translational medical research. Detailed protocols for the determination of sampling locations and numbers as well as recommendations on the orientation, size, and trimming direction of samples from ∼50 different porcine organs and tissues are provided in the Supplementary Material. The proposed sampling protocols include the generation of samples suitable for subsequent qualitative and quantitative analyses, including cryohistology, paraffin, and plastic histology; immunohistochemistry;in situhybridization; electron microscopy; and quantitative stereology as well as molecular analyses of DNA, RNA, proteins, metabolites, and electrolytes. With regard to the planned extent of sampling efforts, time, and personnel expenses, and dependent upon the scheduled analyses, different protocols are provided. These protocols are adjusted for (I) routine screenings, as used in general toxicity studies or in analyses of gene expression patterns or histopathological organ alterations, (II) advanced analyses of single organs/tissues, and (III) large-scale sampling procedures to be applied in biobank projects. Providing a robust reference for studies of porcine models, the described protocols will ensure the efficiency of sampling, the systematic recovery of high-quality samples representing the entire organ or tissue as well as the intra-/interstudy comparability and reproducibility of results.

  11. Transillumination hyperspectral imaging for histopathological examination of excised tissue

    NASA Astrophysics Data System (ADS)

    Vasefi, Fartash; Najiminaini, Mohamadreza; Ng, Eldon; Chamson-Reig, Astrid; Kaminska, Bozena; Brackstone, Muriel; Carson, Jeffrey

    2011-08-01

    Angular domain spectroscopic imaging (ADSI) is a novel technique for the detection and characterization of optical contrast in turbid media based on spectral characteristics. The imaging system employs a silicon micromachined angular filter array to reject scattered light traversing a specimen and an imaging spectrometer to capture and discriminate the largely remaining quasiballistic light based on spatial position and wavelength. The imaging modality results in hyperspectral shadowgrams containing two-dimensional (2D) spatial maps of spectral information. An ADSI system was constructed and its performance was evaluated in the near-infrared region on tissue-mimicking phantoms. Image-based spectral correlation analysis using transmission spectra and first order derivatives revealed that embedded optical targets could be resolved. The hyperspectral images obtained with ADSI were observed to depend on target concentration, target depth, and scattering level of the background medium. A similar analysis on a muscle and tumor sample dissected from a mouse resulted in spatially dependent optical transmission spectra that were distinct, which suggested that ADSI may find utility in classifying tissues in biomedical applications.

  12. Revisiting peri-implant soft tissuehistopathological study of the peri-implant soft tissue

    PubMed Central

    Silva, Eduarda; Félix, Sérgio; Rodriguez-Archilla, Alberto; Oliveira, Pedro; Martins dos Santos, José

    2014-01-01

    Peri-implant soft tissues are essential for osseointegration. The peri-implant mucosa may lack vascular supply, and histological observation, even without plaque, shows the presence of inflammatory cells. The objectives of this study were to assess the histopathological changes of the epithelium and connective tissue around the implant. Twenty patients of both genders were studied. Twelve weeks after implant placement, fragments of peri-implant gingival sulcus were harvested and processed for light microscopy. Group I (10): without clinical inflammatory signs (control); Group II (10): with clinical inflammatory signs. Histopathological parameters were analyzed and classified in 3 grades: mild, moderate or severe (grade 1, 2 or 3). Control group showed only slight changes, grade 1. In group II we found edema with moderate to severe cellular and nuclear changes. There are more women than men with all grades of inflammation. All patients with moderate edema are male and all patients with severe edema are female. A significant association (p=0.007) exists between these two variables. Significant differences were found when comparing the degree of inflammation with nuclear alterations (p=0.001) and the same results when comparing the degree of edema and nuclear changes (p<0.001). This study demonstrates that clinical examination can be used, with a small margin of error, to monitor and control the state of the peri-implant mucosa. In clinics the predisposition of female patients to greater degree of edema and inflammation should be accounted for. PMID:24551281

  13. Quality Measures in Pre-Analytical Phase of Tissue Processing: Understanding Its Value in Histopathology

    PubMed Central

    Masilamani, Suresh; Sundaram, Sandhya; Duvuru, Prathiba; Swaminathan, Rajendiran

    2016-01-01

    Introduction Quality monitoring in histopathology unit is categorized into three phases, pre-analytical, analytical and post-analytical, to cover various steps in the entire test cycle. Review of literature on quality evaluation studies pertaining to histopathology revealed that earlier reports were mainly focused on analytical aspects with limited studies on assessment of pre-analytical phase. Pre-analytical phase encompasses several processing steps and handling of specimen/sample by multiple individuals, thus allowing enough scope for errors. Due to its critical nature and limited studies in the past to assess quality in pre-analytical phase, it deserves more attention. Aim This study was undertaken to analyse and assess the quality parameters in pre-analytical phase in a histopathology laboratory. Materials and Methods This was a retrospective study done on pre-analytical parameters in histopathology laboratory of a tertiary care centre on 18,626 tissue specimens received in 34 months. Registers and records were checked for efficiency and errors for pre-analytical quality variables: specimen identification, specimen in appropriate fixatives, lost specimens, daily internal quality control performance on staining, performance in inter-laboratory quality assessment program {External quality assurance program (EQAS)} and evaluation of internal non-conformities (NC) for other errors. Results The study revealed incorrect specimen labelling in 0.04%, 0.01% and 0.01% in 2007, 2008 and 2009 respectively. About 0.04%, 0.07% and 0.18% specimens were not sent in fixatives in 2007, 2008 and 2009 respectively. There was no incidence of specimen lost. A total of 113 non-conformities were identified out of which 92.9% belonged to the pre-analytical phase. The predominant NC (any deviation from normal standard which may generate an error and result in compromising with quality standards) identified was wrong labelling of slides. Performance in EQAS for pre-analytical phase was

  14. Histopathologic changes in liver and kidney tissues induced by carbaryl in Bufotes variabilis (Anura: Bufonidae).

    PubMed

    Çakıcı, Özlem

    2015-03-01

    The purpose of this work was to investigate for the first time histopathologic effects of carbaryl in liver and kidney tissues of Bufotes variabilis. After 96h following exposure to carbaryl (low dose: 0.05, medium dose: 0.1 and high dose: 0.2mg/g), the toads were euthanized and dissected. In liver tissue, vacuolization in hepatocytes, necrosis, mononuclear cell infiltration, an increase in melanomacrophage number, enlargement of sinusoids, hemorrhage and congestion were determined in exposed toads. In kidney tissue, mononuclear cell infiltration, hypertrophied Bowman's capsule cells, deformation, vacuolization, karyolysis and necrosis of renal tubule epithelium, brush border destruction, glomerular shrinkage, hemorrhage and fibrosis were observed in carbaryl-treated groups. According to this investigation, carbaryl caused histopathologic damages in liver and kidney tissues of B. variabilis. PMID:25573057

  15. Histopathologic changes in liver and kidney tissues induced by carbaryl in Bufotes variabilis (Anura: Bufonidae).

    PubMed

    Çakıcı, Özlem

    2015-03-01

    The purpose of this work was to investigate for the first time histopathologic effects of carbaryl in liver and kidney tissues of Bufotes variabilis. After 96h following exposure to carbaryl (low dose: 0.05, medium dose: 0.1 and high dose: 0.2mg/g), the toads were euthanized and dissected. In liver tissue, vacuolization in hepatocytes, necrosis, mononuclear cell infiltration, an increase in melanomacrophage number, enlargement of sinusoids, hemorrhage and congestion were determined in exposed toads. In kidney tissue, mononuclear cell infiltration, hypertrophied Bowman's capsule cells, deformation, vacuolization, karyolysis and necrosis of renal tubule epithelium, brush border destruction, glomerular shrinkage, hemorrhage and fibrosis were observed in carbaryl-treated groups. According to this investigation, carbaryl caused histopathologic damages in liver and kidney tissues of B. variabilis.

  16. Soy consumption and histopathologic markers in breast tissue using tissue microarrays.

    PubMed

    Maskarinec, Gertraud; Erber, Eva; Verheus, Martijn; Hernandez, Brenda Y; Killeen, Jeffrey; Cashin, Suzanne; Cline, J Mark

    2009-01-01

    This study examined the relation of soy intake with hormonal and proliferation markers in benign and malignant breast tissue using tissue microarrays (TMAs). TMAs with up to 4 malignant and 4 benign tissue samples for 268 breast cancer cases were constructed. Soy intake in early life and in adulthood was assessed by questionnaire. The TMAs were stained for estrogen receptor (ER) alpha, ERbeta, progesterone receptor (PR), human epidermal growth factor receptor 2 (HER2/neu), proliferating cell nuclear antigen (PCNA), and Ki-67 using standard immunohistochemical methods. Logistic regression was applied for statistical analysis. A higher percentage of women showed positive marker expression in malignant than in benign tissue. With one exception, HER2/neu, no significant associations between soy intake and pathologic markers were observed. Early life soy intake was associated with lower HER2/neu and PCNA staining of malignant tissue. In benign tissue, early life soy intake showed higher ER and PR expression, but no difference in proliferation markers. The results of this investigation provide some assurance that soy intake does not adversely affect markers of proliferation. TMAs were shown to be a useful tool for epidemiologic research.

  17. The post-analytical phase of histopathology practice: Storage, retention and use of human tissue specimens

    PubMed Central

    Kapila, Supriya Nikita; Boaz, Karen; Natarajan, Srikant

    2016-01-01

    There are several aspects to a histopathology practice besides the acquisition of biopsy specimens and histopathological diagnosis. Pathology Departments are home to an abundant source of knowledge in the form of stored specimens and slides. We attempt to highlight the importance of regulation of storage, retention, and appropriate use of human tissue material in research and ownership rights to the same. We also discuss requirement and waiver of informed consent for scientific work involving the use of such tissues, which in the absence of defined laws come under the purview of Institution Review Boards. Pathology Departments, under the binding of the parent institution, are conceded the responsibility of maintenance and retention of pathology specimens. This communication highlights some of the important aspects in human tissue material handling and research, underscoring the necessity for established regulations regarding the same. PMID:26958513

  18. The post-analytical phase of histopathology practice: Storage, retention and use of human tissue specimens.

    PubMed

    Kapila, Supriya Nikita; Boaz, Karen; Natarajan, Srikant

    2016-01-01

    There are several aspects to a histopathology practice besides the acquisition of biopsy specimens and histopathological diagnosis. Pathology Departments are home to an abundant source of knowledge in the form of stored specimens and slides. We attempt to highlight the importance of regulation of storage, retention, and appropriate use of human tissue material in research and ownership rights to the same. We also discuss requirement and waiver of informed consent for scientific work involving the use of such tissues, which in the absence of defined laws come under the purview of Institution Review Boards. Pathology Departments, under the binding of the parent institution, are conceded the responsibility of maintenance and retention of pathology specimens. This communication highlights some of the important aspects in human tissue material handling and research, underscoring the necessity for established regulations regarding the same.

  19. Histopathological Effects of Tissue Adhesives on Experimental Peripheral Nerve Transection Model in Rats

    PubMed Central

    Çıralık, Harun

    2015-01-01

    Objective Our aim was to evaluate the histopathological effects of tissue adhesives on peripheral nerve regeneration after experimental sciatic nerve transection in rats and to search whether these tissue adhesives may possess a therapeutic potential in peripheral nerve injuries. Methods This experimental study was performed using 42 female Wistar-Albino rats distributed in 6 groups subsequent to transection of right sciatic nerves. Group I underwent external circumferential neurolysis; Group II received suture repair; Group III had local polymeric hydrogel based tissue adhesive administration; Group IV received suture repair and polymeric hydrogel based tissue adhesive application together; Group V had gelatin based tissue adhesive application and Group VI had suture repair and gelatin based tissue adhesive together. After a 6-week follow-up period, biopsies were obtained from site of neural injury and groups were compared with respect to histopathological scoring based on inflammatory, degenerative, necrotic and fibrotic changes. Results There were remarkable differences between control group and study groups with respect to inflammation (p=0.001), degeneration (p=0.002), necrosis (p=0.007), fibrosis (p<0.001) and vascularity (p=0.001). Histopathological scores were similar between study groups and the only noteworthy difference was that Group V displayed a lower score for necrosis and higher score in terms of vascularization. Conclusion Our results imply that tissue adhesives can be useful in repair of peripheral nerve injuries by decreasing the surgical trauma and shortening the duration of intervention. Results with gelatin based tissue adhesive are especially promising since more intense vascularity was observed in tissue after application. However, trials on larger series with longer durations of follow-up are essential for reaching more reliable conclusions. PMID:26819683

  20. Microfluidic sampling system for tissue analytics

    PubMed Central

    Hokkanen, A.; Stuns, I.; Schmid, P.; Kokkonen, A.; Gao, F.; Steinecker, A.; Budczies, J.; Heimala, P.; Hakalahti, L.

    2015-01-01

    We have developed a microfluidics based sampling system for tissue analytics. The proof-of-concept of the sampling system was demonstrated by extracting lipid samples from tissue biopsies. The sample collection system consists of a disposable silicon based multiport microneedle integrated with polymer microfluidics. The polymethyl methacrylate polymer microfluidic chip has a 10 μl sample reservoir and actuation membranes for liquid pumping. A special automated robotic system was developed to control the positioning of the needle and the sampling procedure on preselected spots on the tissue. Real breast cancer tissue samples were used to test the feasibility of the sampling system. We successfully measured indicative cancer biomarkers from the tissue surface. Phosphatidylcholine and phosphoethanolamine were extracted from the tissue membrane with methyl tert-butyl ether solvent and detected by mass spectrometry. In the future, this tool could be used in characterization of preoperative biopsies and tumour tissues removed during surgery. PMID:26421088

  1. Hyperthermic tissue sealing devices: a proposed histopathologic protocol for standardizing the evaluation of thermally sealed vessels

    NASA Astrophysics Data System (ADS)

    Livengood, Ryan H.; Vos, Jeffrey A.; Coad, James E.

    2011-03-01

    Hyperthermic tissue sealing devices are advancing modern laparoscopy and other minimally invasive surgical approaches. Histopathologic evaluation of thermally sealed vessels can provide important information on their associated tissue effects and reactions. However, a standardized systematic approach has not been historically used in the literature. This paper proposes a histologic approach for the analysis of thermally sealed vessels and their basis of hemostasis, including thermal tissue changes, healing, and thrombosis. Histologic evaluation during the first week (Days 3-7) can assess the seal's primary tissue properties. These parameters include the thermal seal's length, architecture, tissue layers involved, adventitial collagen denaturation length, entrapped vapor or blood pockets, tissue homogenization and thermal tissue injury zones. While the architectural features can be assessed in Day 0-3 specimens, the latter thermal injury zones are essentially not assessable in Day 0-3 specimens. Day 14 specimens can provide information on the early healing response to the sealed vessel. Day 30 and longer specimens can be used to evaluate the seal's healing reactions. Assessment of the healing response should include seal site inflammation, granulation tissue, necrosis resorption, fibroproliferative scar healing, and thrombus organization. In order to accurately evaluate these parameters, careful specimen orientation, embedding and multiple histologic sections across the entire seal width are required. When appropriate in vivo post-treatment times are used, thermal vessel seals can be evaluated with routine light microscopy and common histologic staining methods.

  2. Histopathology effects of nickel nanoparticles on lungs, liver, and spleen tissues in male mice

    NASA Astrophysics Data System (ADS)

    Ajdari, Marziyeh; Ziaee Ghahnavieh, Marziyeh

    2014-09-01

    Because of the classification of the nickel compounds as carcinogenic substances, there is a need for in vivo tests to nickel nanoparticles (NiNPs) for observing their effects on health experimentally. Spherical NiNPs with 10 nm in diameter and 75 ppm concentration were applied for investigating their toxicities within male albino mice as an in vivo model. We randomly made sham group, control group, and 75 ppm group (with five animals in each group). Then, the nanoparticles were injected into mice intraperitonealy for 7 days and after that their lungs, liver, and spleen were removed for histopathological observations. At the end of the test, section microscopic observations of liver, spleen, and lung in sham and control groups showed normal tissues but these tissues underwent significant abnormal effects in 75 ppm group. NiNPs can cause undesirable effects in lungs, liver, and spleen tissues with same condition of this study.

  3. Detection of Candida albicans mRNA in Archival Histopathology Samples by Reverse Transcription-PCR

    PubMed Central

    Beggs, Kyle T.; Holmes, Ann R.; Cannon, Richard D.; Rich, Alison M.

    2004-01-01

    The feasibility of detecting Candida albicans mRNA in formalin-fixed paraffin-embedded archival human histopathology specimens by reverse transcription-PCR (RT-PCR) was investigated. RT with gene-specific primers was used to detect five single-copy C. albicans gene transcripts, including those of two housekeeping genes, in oral candidiasis samples up to 8 years of age. PMID:15131211

  4. Is tissue an issue? Current practice and opinion in Western Australia for routine histopathology on products of conception.

    PubMed

    Yap, Shui-Jean; Watts, Jared C; Faithfull, Tiffany J; Wong, Sabrina Z; Wylde, Kate L; McGurgan, Paul M

    2014-10-01

    An anonymous questionnaire-based survey was used to determine current practices and opinions of senior health professionals working in Western Australian (WA) hospitals performing gynaecological procedures, regarding the routine use of histopathology for products of conception (POC) obtained either from the surgical management of miscarriage or termination of pregnancy. Sixty-one senior health professionals completed the survey. Tissue histopathology on POC was routinely requested for miscarriage and termination of pregnancy (TOP) by 87 and 59% of respondents, respectively. Respondents listed the main reasons for requesting routine histopathology as avoidance of misdiagnosis, medico-legal and quality assurance. There were inconsistent practices among WA health professionals regarding sending POC for histopathology; 63% of gynaecology head of departments recommend the introduction of state or national guidelines for the use of histopathology in the surgical management of miscarriages or terminations of pregnancy.

  5. Prevalence of Toxoplasma gondii in Chicken samples from delta of Egypt using ELISA, histopathology and immunohistochemistry.

    PubMed

    Ibrahim, Hany M; Abdel-Ghaffar, Fathy; Osman, Gamalat Y; El-Shourbagy, Safinaz H; Nishikawa, Yoshifumi; Khattab, Reham A

    2016-06-01

    Estimates of the zoonotic diseases are helpful for monitoring and improving public health. Laboratory-based surveillance provides crucial information for assessing zoonotic disease trends and developments. Toxoplasmosis is considered as a zoonotic disease and has both medical and veterinary importance since it leads to abortion in humans and several animal species. In view of the worldwide importance of T. gondii, this study aimed to estimate the prevalence of T. gondii in chickens from the Delta of Egypt. A total of 304 blood and brain samples were collected from Egyptian chickens from Gharbiya, Qalyoubiya, Minufiya, Beheira, Kafr EL-Shaykh and Dakahlia Provinces. In order to determine the serological and histopathological prevalence of T. gondii, the samples were examined by ELISA, histopathology and immunohistochemistry (IHC). The prevalence of T. gondii was 11.18, 6.91, 6.91 % by ELISA, histopathology and IHC, respectively. Statistically significant differences in the prevalence of T. gondii were observed on the basis of season, sex and habitat. These data provide valuable information regarding the epidemiology of T. gondii infections in Egyptian chickens, which can be employed in developing efficient strategies for disease management and control. PMID:27413325

  6. Preparation of tissue samples for X-ray fluorescence microscopy

    NASA Astrophysics Data System (ADS)

    Chwiej, Joanna; Szczerbowska-Boruchowska, Magdalena; Lankosz, Marek; Wojcik, Slawomir; Falkenberg, Gerald; Stegowski, Zdzislaw; Setkowicz, Zuzanna

    2005-12-01

    As is well-known, trace elements, especially metals, play an important role in the pathogenesis of many disorders. The topographic and quantitative elemental analysis of pathologically changed tissues may shed some new light on processes leading to the degeneration of cells in the case of selected diseases. An ideal and powerful tool for such purpose is the Synchrotron Microbeam X-ray Fluorescence technique. It enables the carrying out of investigations of the elemental composition of tissues even at the single cell level. The tissue samples for histopathological investigations are routinely fixed and embedded in paraffin. The authors try to verify the usefulness of such prepared tissue sections for elemental analysis with the use of X-ray fluorescence microscopy. Studies were performed on rat brain samples. Changes in elemental composition caused by fixation in formalin or paraformaldehyde and embedding in paraffin were examined. Measurements were carried out at the bending magnet beamline L of the Hamburger Synchrotronstrahlungslabor HASYLAB in Hamburg. The decrease in mass per unit area of K, Br and the increase in P, S, Fe, Cu and Zn in the tissue were observed as a result of the fixation. For the samples embedded in paraffin, a lower level of most elements was observed. Additionally, for these samples, changes in the composition of some elements were not uniform for different analyzed areas of rat brain.

  7. Molecular and Histopathological Changes in Mouse Intestinal Tissue After Proton Exposure

    NASA Technical Reports Server (NTRS)

    Purgason, A.; Zhang, Y.; Wu, H.

    2010-01-01

    Radiation in space, including types from solar particle events (SPE's), poses serious health risks to astronauts and is especially dangerous for long duration missions. Protons are the most abundant particles in deep space and to date there is little known about the details of the negative consequences crew members will face upon exposure to them. This ongoing project involves a mouse model subjected to several minutes of proton radiation at an energy of 250 MeV and doses of 0 Gy, 0.1 Gy, 1 Gy, and 2 Gy. The gastrointestinal tract of each animal was dissected four hours post-irradiation and the small intestine was isolated and flash-frozen. Three specimens per dose were studied. Tissue was homogenized and RNA was isolated in order for cDNA synthesis and real-time PCR to be performed. Gene expression changes are currently being analyzed specific to mouse apoptosis. Immunohistochemistry will be used to confirm any significant changes found in the analyses. Immunohistochemistry is also being used to observe gamma H2AX staining to learn of any DNA damage that occurred as a result of proton exposure. We expect to see increased DNA damage due to proton exposure. Finally, histopathologic observation of the tissue will be completed using standard H&E staining methods to screen for morphologic changes. Increased apoptosis is expected to be seen in the tissues which is typical of radiation damage. Observations will be confirmed by a pathologist.

  8. [General principles of handling tissues and organs intended for examination in histopathology - pathologists requirements for surgeons].

    PubMed

    Trnková, M

    2014-03-01

    Histopathology has continued to develop into a complex discipline of laboratory medicine in the last 30 years. Implementation of new techniques such as immunohistochemistry, in situ hybridization, molecular pathology and gene profiling yields a large amount of information which is used not only to establish diagnosis, but also for prediction and prognosis. The basic conditions for precision and correctness of this information which directly influence the choice of therapy and the outcome for the patient, and eventually the health and the life of the patient, are obtaining a suitable specimen and the best laboratory processing. The first step in this procedure is collection of samples, fixation and submission to the pathology laboratory - the pre-analytical phase. Knowledge of the principles of this phase and their implementation in daily practice in surgery is of main importance for the quality and quantity of information obtained. We introduce the basic rules for proper handling of different specimens depending on the use of conventional and "new" histopathology methods.

  9. Molecular and Histopathological Changes in Mouse Intestinal Tissue after Proton Exposure

    NASA Astrophysics Data System (ADS)

    Purgason, Ashley; Zhang, Ye; Hamilton, Stanley; Wu, Honglu

    Radiation in space, especially energetic protons emitted from solar particle events (SPEs), poses serious health risks to astronauts and is especially dangerous for long duration missions. Protons are the most abundant particles in space and to date there is little known about the details of the negative consequences crew members will face upon exposure to them. To elucidate some of the possible health effects induced by protons, BALB/C mice were subjected to 250 MeV of proton radiation at doses of 0 Gy, 0.1 Gy, 1 Gy, and 2 Gy. Three specimens per dose were studied. The gastrointestinal tract of each animal was dissected four hours post-irradiation and the isolated small intestinal tissue was fixed in formalin for histopathological examination or snap-frozen in liquid nitrogen for RNA isolation. Histopathologic observation of the tissue using standard HE staining methods to screen for morphologic changes showed a marked increase in apoptotic lesions for even the lowest dose of 0.1 Gy, and the dose response showed possible hyper sensitivities at low dose. Tissue of the gastrointestinal tract was also homogenized and RNA was isolated for cDNA synthesis and real-time PCR analysis for genes involved in apoptosis. Results of gene expression changes revealed consistent up or down regulation of a number of genes for all of the exposure doses that may play a role in proton-induced apoptosis (e.g. Hsp90ab1). In addition, several genes were found to have significant changes in the RNA level after only the low dose (0.1 Gy), but not the high dose (1 and 2 Gy), proton exposures (e.g. Bok and Casp1), whereas some genes had expression changes only after high dose proton exposures (e.g. Tsc22d3). These findings demonstrated that apoptosis may occur in gastrointestinal tracts after even low dose proton exposures, and the different gene expression patterns between low and high dose proton irradiated mice may offer insight into the molecular mechanisms of the possible hyper

  10. A survey of spontaneous occurrence of ochratoxin A residues in chicken tissues and concurrence with histopathological changes in liver and kidneys

    USGS Publications Warehouse

    Milicevic, Dragan; Jovanovic, Milijan; Matekalosverak, Vesna; Radicevic, Tatjana; Petrovic, Milan M.; Lilic, Slobodan

    2011-01-01

    Toxicological and histopathological investigations of tissues of commercially slaughtered chickens were carried out to provide a preliminary evaluation of the incidence of occurrence of ochratoxin A (OTA) in chicken sold in Serbian retail market. In addition, the etiology of nephropathies of these chickens was elucidated. The majority of these tissue samples were not found to contain measurable amounts of OTA. Moreover, the OTA levels found in analyzed tissues were generally low and there was no positive correlation between the presence of OTA and the frequency of histopathological changes. Histopathological changes such as degenerative changes in the kidneys and liver differed from the classical description of the mycotoxic nephropathy, indicating that the chicken nephropathy observed in Serbia may have a multitoxic etiology with possible synergistic effect between microorganisms and natural toxins, usually present in low concentrations. The low OTA results also suggested that chicken meat available in the retail market in Serbia are unlikely to pose any significant adverse health risk to the consumers with respect to OTA toxicity.

  11. Histopathological Analysis of PEEK Wear Particle Effects on the Synovial Tissue of Patients

    PubMed Central

    Jansson, V.; Giurea, A.

    2016-01-01

    Introduction. Increasing interest developed in the use of carbon-fiber-reinforced-poly-ether-ether-ketones (CFR-PEEK) as an alternative bearing material in knee arthroplasty. The effects of CFR-PEEK wear in in vitro and animal studies are controversially discussed, as there are no data available concerning human tissue. The aim of this study was to analyze human tissue containing CFR-PEEK as well as UHMWPE wear debris. The authors hypothesized no difference between the used biomaterials. Methods and Materials. In 10 patients during knee revision surgery of a rotating-hinge-knee-implant-design, synovial tissue samples were achieved (tibial inserts: UHMWPE; bushings and flanges: CFR-PEEK). One additional patient received revision surgery without any PEEK components as a control. The tissue was paraffin-embedded, sliced into 2 μm thick sections, and stained with hematoxylin and eosin in a standard process. A modified panoptical staining was also done. Results. A “wear-type” reaction was seen in the testing and the control group. In all samples, the UHMWPE particles were scattered in the tissue or incorporated in giant cells. CFR-PEEK particles were seen as conglomerates and only could be found next to vessels. CFR-PEEK particles showed no giant-cell reactions. In conclusion, the hypothesis has to be rejected. UHMWPE and PEEK showed a different scatter-behavior in human synovial tissue. PMID:27766256

  12. Tissue distribution and histopathological effects of dietary methylmercury in benthic grubby Myoxocephalus aenaeus

    SciTech Connect

    Pelletier, E.; Audet, C.

    1995-05-01

    There is a need to test deterministic models predicting the behavior and effects of chemicals on aquatic systems by conducting experiments with more than one trophic step at a time. This approach requires the set-up of an experimental food chain in pounds or mesocosm facilities which can be used for dietary uptake studies and assessment of sublethal stress induced by contaminated food. In the course of our current research program at the INRS marine mesocosms facilities, a model benthic food chain including inter- and sub-tidal species such as the mussel (Mytilus edulis), clam (Mya arenaria), starfish (Leptasterias polaris), polychaete (Nereis virens), amphipod (Gammarus sp.), gastropod (Buccinum undatum), and fishes (Pleuronectec americanus, Myoxocephalus aenaeus), is used for testing food uptake models and for the development of sublethal toxicity tests which could be used in the environmental assessment of coastal and estuarine waters. Among these test organisms, the grubby (M. aenaeus) is a small coastal fish (12-15 cm) characterized by a broad head. The grubby is tolerant of water temperature and salinity variations and lives on a wide variety of bottom strata at low depths. The grubby is carnivorous and consumes a wide variety of molluscs and the young of many species of fish. Because of its size, its estuarine and coastal distribution and its large spectrum of prey, this species was seen as an ideal fish to fit into our experimental food chain model. This paper reports a preliminary experiment designed to measure distribution of mercury in tissues and to test the response of various histopathological and biochemical stress indicators in grubby exposed to dietary contamination by methylmercury (MeHg) for a 20-day exposure period. MeHg was chosen because it is rapidly bioaccumulated by most living organisms and its toxicity has been studied for decades in numerous aquatic ecosystems. 17 refs., 2 tabs.

  13. Effect of dietary selenomethionine on growth performance, tissue burden, and histopathology in green and white sturgeon.

    PubMed

    De Riu, Nicola; Lee, Jang-Won; Huang, Susie S Y; Moniello, Giuseppe; Hung, Silas S O

    2014-03-01

    A comparative examination of potential differences in selenium (Se) sensitivity was conducted on two sturgeon species indigenous to the San Francisco Bay-Delta. Juvenile green (Acipenser medirostris), recently given a federally threatened status, and white sturgeon (Acipenser transmontanus) were exposed to one of four nominal concentrations of dietary l-selenomethionine (SeMet) (0 (control), 50, 100, or 200 mg SeMet/kg diet) for 8 weeks. Mortality, growth performance, whole body composition, histopathology, and Se burdens of the whole body, liver, kidneys, gills, heart, and white muscle were determined every 2 to 4 weeks. Significant (p<0.05) mortality was observed in green sturgeon fed the highest SeMet diet after 2 weeks, whereas no mortality was observed in white sturgeon. Growth rates were significantly reduced in both species; however, green sturgeon was more adversely affected by the treatment. Dietary SeMet significantly affected whole body composition and most noticeably, in the decline of lipid contents in green sturgeon. Selenium accumulated significantly in all tissues relative to the control groups. After 4 and 8 weeks of exposure, marked abnormalities were observed in the kidneys and liver of both sturgeon species; however, green sturgeon was more susceptible to SeMet than white sturgeon at all dietary SeMet levels. Our results showed that a dietary Se concentration at 19.7 ± 0.6 mg Se/kg, which is in range with the reported Se concentrations of the benthic macro-vertebrate community of the San Francisco Bay, had adverse effects on both sturgeon species. However, the exposure had a more severe pathological effect on green sturgeon, suggesting that when implementing conservation measures, this federally listed threatened species should be monitored and managed independently from white sturgeon.

  14. Calibration of redox scanning for tissue samples

    NASA Astrophysics Data System (ADS)

    Xu, He N.; Wu, Baohua; Nioka, Shoko; Chance, Britton; Li, Lin Z.

    2009-02-01

    The fluorescence properties of reduced nicotinamide adenine dinucleotide (NADH) and oxidized flavoproteins (Fp) such as flavin adenine dinucleotide (FAD) in the respiratory chain are sensitive indicators of intracellular redox states and have been applied to the studies of mitochondrial function with energy-linked processes. The redox scanner, a threedimensional (3D) redox cryo-imager previously developed by Chance et al., can quantitatively determine the metabolic properties of tissue samples by acquiring the fluorescence images of NADH and Fp. The redox ratios, i.e., Fp/(Fp+NADH) and NADH/(Fp+NADH), obtained on the basis of relative signal intensity ratios, provide a sensitive index of steady-state of the mitochondrial metabolism that has been determined for a variety of biological tissues. This paper presents the further development of the instrument by establishing a calibration method to quantify the concentration of the fluorophores and facilitate the comparison of redox images obtained at different time or with different instrument functions. Calibration curves of both NADH and Fp have been obtained using snap-frozen standard references with NADH concentration ranging from 150-1400 μM and Fp from 80-720 μM. Snap-freeze tissue samples such as human breast tumors xenografted in mice, normal mouse pancreases and spleens were imaged. The NADH and Fp concentrations as well as the redox ratios in the tissue samples were quantified based on the adjacent solution standards of NADH and Fp. The obtained multi-slice redox images revealed high heterogeneity of the tissue samples which can be quantitatively interpreted.

  15. Antioxidant Attenuation of Atrazine Induced Histopathological Changes in Testicular Tissue of Goat In Vitro

    PubMed Central

    Sharma, R. K.; Fulia, Anju; Chauhan, P. K.

    2012-01-01

    During the present investigation the effect of α-tocopherol (100 μmolL-1) in prevention of testicular toxicity induced by atrazine in goat Capra hircus have been analyzed. Vitamin E (α-tocopherol) at dose level 100 μmolL-1 provides attenuation over the histopathological changes generated by pesticide atrazine (100 nmolml-1). Small pieces (approximately 1mm3) of testicular tissue were divided into three groups (one control group + two experimental groups). Experimental group (A) was supplemented with 100 nmolml-1 concentration of atrazine and experimental group (B) was supplemented with 100 nmolml-1 atrazine and 100 μmolL-1 concentrations of vitamin E (α-Tocopherol) and harvesting was carried out after 1, 4 and 8 hrs of exposure. Control was run along with all the experimental groups. In the experimental group (A) treated with atrazine at dose level 100 nmolml-1, revealed histomorphological alterations in the seminiferous tubule. After one hour of exposure duration small vacuoles in cytoplasm of the Sertoli cells and spermatogonia were observed. Chromolysis at pycnosis were also noticed in the spermatogonia and spermatids. In the experimental group (B) exposed with atrazine and simultaneously supplemented with Vitamin E also showed degeneration but it was milder as compared with experimental group treated with atrazine without antioxidant. Atrazine exposure induced a decline in diameter of spermatocytes from 10.51 ± 0.2052 μm in control to 7.915 ± 0.2972, 7.5 ± 0.211 and 7.14 ± 0.225 μm after exposure of 1, 4 and 8 hrs respectively but in case of atrazine supplemented with vitamin E [experimental group (B)], there was less decline in cell diameter that was 8.5 ± 0.1865, 8.1 ± 0.1201 and 7.8 ± 0.2066μm after exposure of 1, 4 and 8 hrs respectively. The result demonstrated that vitamin E delays the degenerative changes induced by atrazine. PMID:23293464

  16. Histopathological Changes in Tissues of Bithynia siamensis goniomphalos Incubated in Crude Extracts of Camellia Seed and Mangosteen Pericarp

    PubMed Central

    Aukkanimart, Ratchadawan; Pinlaor, Somchai; Tesana, Smarn; Aunpromma, Surasit; Booyarat, Chantana; Sriraj, Pranee; Laummaunwai, Porntip; Punjaruk, Wiyada

    2013-01-01

    The present study was performed to observe histopathological changes in tissues of Bithynia siamensis goniomphalos (Gastropoda, Bithyniidae) incubated in crude extract solutions of camellia (Camellia oleifera) seed and mangosteen (Garcinia mangostana) pericarp, and furthermore to estimate the molluscicidal effects of 2 plant substances. Substantial numbers of bithyniid snails were incubated in various concentrations of 2 plant solution for 24 hr. As the positive control, snails incubated in various concentrations of niclosamide, a chemical molluscicide, were used. The histopathological findings were observed in sectioned snail specimens of each experimental and control groups. The results showed that both camellia and mangosteen extracts had molluscicidal effects at 24 hr with 50% lethal concentration (LC50) at concentrations of 0.003 and 0.002 g/ml, respectively, while niclosamide had LC50 at concentrations 0.599 ppm. B. siamensis goniomphalos snail tissues (foot, gill, and digestive system) showed disruption of columnar muscle fibers of the foot, reduction of the length and number of gill cilia, numerous mucous vacuoles, and irregularly shaped of epithelial cells. Irregular apical and calciferous cells, dilatation of the digestive gland tubule, and large hemolymphatic spaces, and irregular apical surfaces, detachment of cilia, and enlargement of lysosomal vacuoles of epidermis were also shown in all groups. By the present study, it is confirmed that 2 plants, camellia and mangosteen, are keeping some substance having molluscicidal effects, and histopathological findings obtained in this study will provide some clues in further studies on their action mechanisms to use them as natural molluscicides. PMID:24327779

  17. A melanin-bleaching methodology for molecular and histopathological analysis of formalin-fixed paraffin-embedded tissue.

    PubMed

    Chung, Joon-Yong; Choi, Jiyeon; Sears, John D; Ylaya, Kris; Perry, Candice; Choi, Chel H; Hong, Seung-Mo; Cho, Hanbyoul; Brown, Kevin M; Hewitt, Stephen M

    2016-10-01

    Removal of excessive melanin from heavily pigmented formalin-fixed paraffin-embedded (FFPE) melanoma tissues is essential for histomorphological and molecular diagnostic assessments. Although there have been efforts to address this issue, current methodologies remain complex and time-consuming, and are not suitable for multiple molecular applications. Herein, we have developed a robust and rapid melanin-bleaching methodology for FFPE tissue specimens. Our approach is based on quick bleaching (15 min) at high temperature (80 °C) with 0.5% diluted hydrogen peroxide (H2O2) in Tris-HCl, PBS, or Tris/Tricine/SDS buffer. Immunostaining for Ki-67 and HMB45 was enhanced by bleaching with 0.5% H2O2 in Tris/Tricine/SDS and Tris-HCl, respectively. In addition to histopathological applications, our approach also facilitates recovery of protein and nucleic acid from archival melanin-rich FFPE tissue sections. Protein extracted from bleached FFPE tissues was compatible with western blotting using anti-human GAPDH and AKT antibodies. Our bleaching condition significantly improved RNA quality compared with unbleached tissues without compromising the yield. Notably, the RNA/DNA obtained from bleached tissues was suitable for end point PCR and real-time quantitative RT-PCR. In conclusion, this improved melanin-bleaching method enhances and simplifies immunostaining procedures, and facilitates the use of melanin-rich FFPE tissues for histomorphological and PCR amplification-based molecular assays.

  18. A melanin-bleaching methodology for molecular and histopathological analysis of formalin-fixed paraffin-embedded tissue.

    PubMed

    Chung, Joon-Yong; Choi, Jiyeon; Sears, John D; Ylaya, Kris; Perry, Candice; Choi, Chel H; Hong, Seung-Mo; Cho, Hanbyoul; Brown, Kevin M; Hewitt, Stephen M

    2016-10-01

    Removal of excessive melanin from heavily pigmented formalin-fixed paraffin-embedded (FFPE) melanoma tissues is essential for histomorphological and molecular diagnostic assessments. Although there have been efforts to address this issue, current methodologies remain complex and time-consuming, and are not suitable for multiple molecular applications. Herein, we have developed a robust and rapid melanin-bleaching methodology for FFPE tissue specimens. Our approach is based on quick bleaching (15 min) at high temperature (80 °C) with 0.5% diluted hydrogen peroxide (H2O2) in Tris-HCl, PBS, or Tris/Tricine/SDS buffer. Immunostaining for Ki-67 and HMB45 was enhanced by bleaching with 0.5% H2O2 in Tris/Tricine/SDS and Tris-HCl, respectively. In addition to histopathological applications, our approach also facilitates recovery of protein and nucleic acid from archival melanin-rich FFPE tissue sections. Protein extracted from bleached FFPE tissues was compatible with western blotting using anti-human GAPDH and AKT antibodies. Our bleaching condition significantly improved RNA quality compared with unbleached tissues without compromising the yield. Notably, the RNA/DNA obtained from bleached tissues was suitable for end point PCR and real-time quantitative RT-PCR. In conclusion, this improved melanin-bleaching method enhances and simplifies immunostaining procedures, and facilitates the use of melanin-rich FFPE tissues for histomorphological and PCR amplification-based molecular assays. PMID:27548802

  19. Retrospective review of diagnostic histopathology samples in research projects: transparency should be the practice with respect to diagnostic discordance.

    PubMed

    Bjugn, Roger; Casati, Bettina

    2012-11-01

    Retrospective review of diagnostic histopathology samples in secondary research projects may generate cases with discordance in diagnosis between the pathologist originally signing out the sample and the reviewer. In this article the authors discuss ethical and legal issues involved in secondary review and propose how diagnostic discrepancies in research projects can be handled. Research participants have the right to know about and control the handling of personal data. Researchers have a duty to inform research participants about research findings of relevance to the current or future health or quality of life. Such feedback is to be provided within a healthcare framework. The authors recommend that pathology laboratories should require researchers to provide feedback on diagnostic discrepancies of potentially clinical significance in secondary research projects using retrospective review of diagnostic histopathology material. The pathology laboratory in charge of the archival material should be responsible for the follow-up on such cases.

  20. A Systematic Approach to the Application of Soft Tissue Histopathology in Paleopathology

    PubMed Central

    Grove, Christina; Peschel, Oliver; Nerlich, Andreas G.

    2015-01-01

    The application of histology to soft tissue remains offers an important technique to obtain diagnostically important information on various physiological and pathological conditions in paleopathology. In a series of 29 cases with mummified tissue ranging between 16 months and c. 5.200 years of postmortem time interval, we systematically investigated paleohistology and the preservation of various tissues. We established a reproducible histological ranking system for the evaluation of mummified tissue preservation. The application of this scheme to the series showed good tissue preservation of tissues with high connective tissue content but also fat tissue and connective tissue rich organs, such as lung tissue, while most other internal organs were less well preserved despite highly different postmortem time intervals. There are some organs with only poor conservation even in short term periods such as the kidneys and CNS. Artificial mummification does not provide better conservation than naturally mummified tissues; “cold” mummies may be much better conserved than those from desert areas. The identification of specific pathologies underlines the potential power of paleohistology. PMID:26346981

  1. Genomic and Histopathological Tissue Biomarkers That Predict Radiotherapy Response in Localised Prostate Cancer

    PubMed Central

    Wilkins, Anna; Dearnaley, David; Somaiah, Navita

    2015-01-01

    Localised prostate cancer, in particular, intermediate risk disease, has varied survival outcomes that cannot be predicted accurately using current clinical risk factors. External beam radiotherapy (EBRT) is one of the standard curative treatment options for localised disease and its efficacy is related to wide ranging aspects of tumour biology. Histopathological techniques including immunohistochemistry and a variety of genomic assays have been used to identify biomarkers of tumour proliferation, cell cycle checkpoints, hypoxia, DNA repair, apoptosis, and androgen synthesis, which predict response to radiotherapy. Global measures of genomic instability also show exciting capacity to predict survival outcomes following EBRT. There is also an urgent clinical need for biomarkers to predict the radiotherapy fraction sensitivity of different prostate tumours and preclinical studies point to possible candidates. Finally, the increased resolution of next generation sequencing (NGS) is likely to enable yet more precise molecular predictions of radiotherapy response and fraction sensitivity. PMID:26504789

  2. Histopathological Changes in the Chorionic Villi and Endometrial Decidual Tissues in the Product of Conception of Spontaneous Abortion Cases.

    PubMed

    Makaju, R; Shrestha, S; Sharma, S; Dhakal, R; Bhandari, S; Shrestha, A; Tamrakar, S

    2015-01-01

    Background Spontaneous abortion refers to a pregnancy that ends spontaneously before the fetus has reached a viable gestational age or expulsion or extraction of an embryo or fetus weighing 500 g or less from its mother. The Maternal Mortality Morbidity Survey of Nepal 2008/09 reported that 7% of maternal deaths in Nepal were due to complications related to abortion. Objective The main objective of this study was to examine the histopathological changes in the chorionic villi and endometrial decidual tissue in products of conception obtained from women with spontaneous abortion. Method This is a retrospective study of 111 patients admitted in the Department of Obstetrics and Gynecology at Dhulikhel Hospital, Kathmandu University Hospital (DH-KUH) with the diagnosis of spontaneous abortion during the period of January 2013 to January 2014. Result Among 111 cases of spontaneous abortions, products of conception was seen in 73 (65.77%) and with only one cases of choriocarcinoma. Majority of cases belongs to age group 21-30 years. The most common decidual changes were inflammation (41.4%) followed by fibrin deposition 29.7%. Majority of the cases shows hydropic changes as histopathological changes in chorionic villi. In the present study, minimum age of lady was 15 years and the maximum age was 45 years and the mean age was 25.09±5.58 years at the time of abortion. Among the cases, maximum 69 (62.2%) of them belonged to age group 21-30 years. Correlating the age group with number of abortions was found to be significantly different (Chi-square= 92.35, df= 3, p < 0.001) among four different age groups. Conclusion The histopathological diagnosis of spontaneous abortion will help in further management of the patient. Further study is required to know the cause of different histopathlogical changes in villi as well as in the decidua. PMID:27423288

  3. Histopathological Changes in the Chorionic Villi and Endometrial Decidual Tissues in the Product of Conception of Spontaneous Abortion Cases.

    PubMed

    Makaju, R; Shrestha, S; Sharma, S; Dhakal, R; Bhandari, S; Shrestha, A; Tamrakar, S

    2015-01-01

    Background Spontaneous abortion refers to a pregnancy that ends spontaneously before the fetus has reached a viable gestational age or expulsion or extraction of an embryo or fetus weighing 500 g or less from its mother. The Maternal Mortality Morbidity Survey of Nepal 2008/09 reported that 7% of maternal deaths in Nepal were due to complications related to abortion. Objective The main objective of this study was to examine the histopathological changes in the chorionic villi and endometrial decidual tissue in products of conception obtained from women with spontaneous abortion. Method This is a retrospective study of 111 patients admitted in the Department of Obstetrics and Gynecology at Dhulikhel Hospital, Kathmandu University Hospital (DH-KUH) with the diagnosis of spontaneous abortion during the period of January 2013 to January 2014. Result Among 111 cases of spontaneous abortions, products of conception was seen in 73 (65.77%) and with only one cases of choriocarcinoma. Majority of cases belongs to age group 21-30 years. The most common decidual changes were inflammation (41.4%) followed by fibrin deposition 29.7%. Majority of the cases shows hydropic changes as histopathological changes in chorionic villi. In the present study, minimum age of lady was 15 years and the maximum age was 45 years and the mean age was 25.09±5.58 years at the time of abortion. Among the cases, maximum 69 (62.2%) of them belonged to age group 21-30 years. Correlating the age group with number of abortions was found to be significantly different (Chi-square= 92.35, df= 3, p < 0.001) among four different age groups. Conclusion The histopathological diagnosis of spontaneous abortion will help in further management of the patient. Further study is required to know the cause of different histopathlogical changes in villi as well as in the decidua.

  4. Radioprotective effects of hesperidin on oxidative damages and histopathological changes induced by X-irradiation in rats heart tissue

    PubMed Central

    Rezaeyan, Abolhasan; Haddadi, Gholam Hassan; Hosseinzadeh, Massood; Moradi, Maryam; Najafi, Masoud

    2016-01-01

    This study was carried out to evaluate radioprotective effects of hesperidin (HES) administration before the irradiation on the cardiac oxidative stress and histopathological changes in an experimental rat model. The cardiovascular complications of radiation exposure cause morbidity and mortality in patients who received radiotherapy. HES, an antioxidant flavonoid found in citrus fruits, suggests the protection against the tissue damage. Fifty-eight rats were divided into four groups: Group 1 received phosphate buffered saline (PBS) and sham radiation; Group 2, HES and sham radiation; Group 3, PBS and radiation; and Group 4, HES and radiation. The rats were exposed to single dose of 18 Gy of 6 MV X-ray. One hundred milligrams per kilogram doses of HES was administered for 7 days before irradiation. The estimation of superoxide dismutase (SOD), malondialdehyde (MDA), and histopathological analyses was performed at 24 h and 8 weeks after radiation exposure. The irradiation of chest area resulted in an elevated MDA level and decreased SOD activity. Moreover, long-term pathological lesions of radiation were inflammation, fibrosis, the increased number of mast cells and macrophages, and development of plaque, vascular leakage, myocardial degeneration, and myocyte necrosis. Although the administration of HES decreases inflammation, fibrosis, mast cell and macrophage numbers, and myocyte necrosis, it did not result in reduced thrombus, myocardium degeneration, and vascular leakage. In conclusion, these results suggest that HES can perform a radioprotection action. The protective effect of HES may be attributable to its immunomodulatory effects and free radical-scavenging properties. PMID:27651565

  5. Radioprotective effects of hesperidin on oxidative damages and histopathological changes induced by X-irradiation in rats heart tissue.

    PubMed

    Rezaeyan, Abolhasan; Haddadi, Gholam Hassan; Hosseinzadeh, Massood; Moradi, Maryam; Najafi, Masoud

    2016-01-01

    This study was carried out to evaluate radioprotective effects of hesperidin (HES) administration before the irradiation on the cardiac oxidative stress and histopathological changes in an experimental rat model. The cardiovascular complications of radiation exposure cause morbidity and mortality in patients who received radiotherapy. HES, an antioxidant flavonoid found in citrus fruits, suggests the protection against the tissue damage. Fifty-eight rats were divided into four groups: Group 1 received phosphate buffered saline (PBS) and sham radiation; Group 2, HES and sham radiation; Group 3, PBS and radiation; and Group 4, HES and radiation. The rats were exposed to single dose of 18 Gy of 6 MV X-ray. One hundred milligrams per kilogram doses of HES was administered for 7 days before irradiation. The estimation of superoxide dismutase (SOD), malondialdehyde (MDA), and histopathological analyses was performed at 24 h and 8 weeks after radiation exposure. The irradiation of chest area resulted in an elevated MDA level and decreased SOD activity. Moreover, long-term pathological lesions of radiation were inflammation, fibrosis, the increased number of mast cells and macrophages, and development of plaque, vascular leakage, myocardial degeneration, and myocyte necrosis. Although the administration of HES decreases inflammation, fibrosis, mast cell and macrophage numbers, and myocyte necrosis, it did not result in reduced thrombus, myocardium degeneration, and vascular leakage. In conclusion, these results suggest that HES can perform a radioprotection action. The protective effect of HES may be attributable to its immunomodulatory effects and free radical-scavenging properties. PMID:27651565

  6. Radioprotective effects of hesperidin on oxidative damages and histopathological changes induced by X-irradiation in rats heart tissue

    PubMed Central

    Rezaeyan, Abolhasan; Haddadi, Gholam Hassan; Hosseinzadeh, Massood; Moradi, Maryam; Najafi, Masoud

    2016-01-01

    This study was carried out to evaluate radioprotective effects of hesperidin (HES) administration before the irradiation on the cardiac oxidative stress and histopathological changes in an experimental rat model. The cardiovascular complications of radiation exposure cause morbidity and mortality in patients who received radiotherapy. HES, an antioxidant flavonoid found in citrus fruits, suggests the protection against the tissue damage. Fifty-eight rats were divided into four groups: Group 1 received phosphate buffered saline (PBS) and sham radiation; Group 2, HES and sham radiation; Group 3, PBS and radiation; and Group 4, HES and radiation. The rats were exposed to single dose of 18 Gy of 6 MV X-ray. One hundred milligrams per kilogram doses of HES was administered for 7 days before irradiation. The estimation of superoxide dismutase (SOD), malondialdehyde (MDA), and histopathological analyses was performed at 24 h and 8 weeks after radiation exposure. The irradiation of chest area resulted in an elevated MDA level and decreased SOD activity. Moreover, long-term pathological lesions of radiation were inflammation, fibrosis, the increased number of mast cells and macrophages, and development of plaque, vascular leakage, myocardial degeneration, and myocyte necrosis. Although the administration of HES decreases inflammation, fibrosis, mast cell and macrophage numbers, and myocyte necrosis, it did not result in reduced thrombus, myocardium degeneration, and vascular leakage. In conclusion, these results suggest that HES can perform a radioprotection action. The protective effect of HES may be attributable to its immunomodulatory effects and free radical-scavenging properties.

  7. Tissue distribution of ochratoxin A as determined by HPLC and ELISA and histopathological effects in chickens.

    PubMed

    Biró, Krisztina; Solti, László; Barna-Vetró, Ildikó; Bagó, György; Glávits, Róbert; Szabó, Erzsébet; Fink-Gremmels, Johanna

    2002-04-01

    Ochratoxin A is a common feed contaminant, which may impair animal health and may lead to residues in edible tissues of slaughter animals. To simulate field conditions, broiler chicks were exposed to a total of 0.5 mg ochratoxin A per week for each of 4 weeks. Plasma toxin levels and tissue residues were measured by high-performance liquid chromatography (HPLC) and microplate enzyme-linked immunosorbent assay (ELISA). Results indicate an accumulation in plasma and wide distribution into all organs, with high levels in the liver and the kidney. Microscopical changes that could primarily be associated with toxin exposure were glomerulonephrosis, tubulonephrosis, focal tubular epithelial cell proliferation and multiple, adenoma-like structures in the renal parenchyma. The HPLC and ELISA methods gave similar results for both tissue distribution and depletion. Differences in absolute tissue toxin concentrations obtained by the two methods might be attributed to the different extraction and clean-up procedures, along with antibody specificity. The findings indicate that the dose applied causes subclinical tissue lesions and measurable tissue residues. PMID:12396358

  8. Histopathological examination of nerve samples from pure neural leprosy patients: obtaining maximum information to improve diagnostic efficiency.

    PubMed

    Antunes, Sérgio Luiz Gomes; Chimelli, Leila; Jardim, Márcia Rodrigues; Vital, Robson Teixeira; Nery, José Augusto da Costa; Corte-Real, Suzana; Hacker, Mariana Andréa Vilas Boas; Sarno, Euzenir Nunes

    2012-03-01

    Nerve biopsy examination is an important auxiliary procedure for diagnosing pure neural leprosy (PNL). When acid-fast bacilli (AFB) are not detected in the nerve sample, the value of other nonspecific histological alterations should be considered along with pertinent clinical, electroneuromyographical and laboratory data (the detection of Mycobacterium leprae DNA with polymerase chain reaction and the detection of serum anti-phenolic glycolipid 1 antibodies) to support a possible or probable PNL diagnosis. Three hundred forty nerve samples [144 from PNL patients and 196 from patients with non-leprosy peripheral neuropathies (NLN)] were examined. Both AFB-negative and AFB-positive PNL samples had more frequent histopathological alterations (epithelioid granulomas, mononuclear infiltrates, fibrosis, perineurial and subperineurial oedema and decreased numbers of myelinated fibres) than the NLN group. Multivariate analysis revealed that independently, mononuclear infiltrate and perineurial fibrosis were more common in the PNL group and were able to correctly classify AFB-negative PNL samples. These results indicate that even in the absence of AFB, these histopathological nerve alterations may justify a PNL diagnosis when observed in conjunction with pertinent clinical, epidemiological and laboratory data.

  9. A histopathological study of the role of periodontal ligament tissue in root resorption in the rat.

    PubMed

    Shiraishi, C; Hara, Y; Abe, Y; Ukai, T; Kato, I

    2001-02-01

    Whether periodontal ligament (PDL) tissue is capable of inducing root resorption was examined. The distal root of the rat molar was sectioned at the furcation and the PDL tissue removed from the root (non-PDL group, n=40). The distal root with the PDL intact was also prepared (PDL-intact group, n=40). The roots were transplanted into the dorsal skin of the rat. On the 1st, 3rd, 5th, 7th, 10th, 14th, 21st or 28th day after transplantation, the roots were removed together with surrounding dorsal subcutaneous tissue and were fixed, demineralized and embedded in paraffin. Serial sections from each block were stained with haematoxylin and eosin or by the tartrate-resistant acid phosphatase (TRAP) method to observe root-resorbing cell formation. Cyclo-oxygenase-2 (COX2) was also detected immunohistologically to examine prostaglandin E(2) production. On the 7th day after transplantation, multinucleated root-resorbing cells with TRAP were observed in the PDL-intact group. The number of TRAP-positive cells peaked on the 10th day after transplantation. COX2-positive cells were observed in PDL during the early experimental stages. No root resorption was seen in the non-PDL group. These results suggest that PDL tissue is involved in the formation of root-resorbing cells and root resorption. PMID:11163317

  10. Ex vivo confocal microscopy imaging to identify tumor tissue on freshly removed brain sample.

    PubMed

    Forest, Fabien; Cinotti, Elisa; Yvorel, Violaine; Habougit, Cyril; Vassal, François; Nuti, Christophe; Perrot, Jean-Luc; Labeille, Bruno; Péoc'h, Michel

    2015-09-01

    Confocal microscopy is a technique able to realize "optic sections" of a tissue with increasing applications. We wondered if we could apply an ex vivo confocal microscope designed for dermatological purpose in a routine use for the most frequent brain tumors. The aim of this work was to identify tumor tissue and its histopathological hallmarks, and to assess grading criteria used in neuropathological practice without tissue loss on freshly removed brain tissue. Seven infiltrating gliomas, nine meningiomas and three metastases of carcinomas were included. We compared imaging results obtained with the confocal microscope to frozen sections, smears and tissue sections of formalin-fixed tissue. Our results show that ex vivo confocal microscopy imaging can be applied to brain tumors in order to quickly identify tumor tissue without tissue loss. It can differentiate tumors and can assess most of grading criteria. Confocal microscopy could represent a new tool to identify tumor tissue on freshly removed sample and could help in selecting areas for biobanking of tumor tissue.

  11. Histopathological changes induced by paraquat on some tissues of gourami fish (Trichogaster trichopterus)

    PubMed Central

    Banaee, M.; Davoodi, M.H.; Zoheiri, F.

    2013-01-01

    Paraquat is a contact and non-selective herbicide which is used for controlling a wide range of terrestrial weeds and aquatic plants. A long-term contact with this xenobiotic can potentially lead to injuries in fishes as live non-target organisms. Therefore, the current study aimed to investigate the effect of sub-lethal toxicity of paraquat on the pathology of gill, liver, and spleen tissues in gourami fish (Trichogaster trichopterus). In this study, sub-lethal concentration is determined based on lethal concentration (LC50 : 7.16±0.69, 4.46±0.43, 2.19±0.27 and 1.41±0.17 mg/l of paraquat within 24, 48, 72 and 96 hours, respectively). The experiment was done with four varied concentrations of paraquat (0.0, 0.07, 0.15, and 0.3 mg/l equal 0.0%, 5%, 10% and 20% of nominal value of 96 h LC50) during 3 weeks. The exposed fish displayed erratic swimming and became lethargic. The changes in gills were characterized by hypertrophy, epithelial, epithelium increase of gill filament, edema and secondary gill lamella. The liver showed hypotrophy of liver cells, cloudy swelling and formation of cytoplasmic vacuoles in the liver tissue of fish treated with 0.15 and 0.3 mg/l concentrations of paraquat. Disorder in the ellipsoid cell and hemosiderin accumulation in melano-macrophage centers was observed in the spleen tissue of fish exposed to 0.15 and 0.3 mg/l of paraquat. PMID:26623309

  12. Multimodal Raman-fluorescence spectroscopy of formalin fixed samples is able to discriminate brain tumors from dysplastic tissue

    NASA Astrophysics Data System (ADS)

    Anand, Suresh; Cicchi, Riccardo; Giordano, Flavio; Buccoliero, Anna Maria; Pavone, Francesco Saverio

    2014-05-01

    In the recent years, there has been a considerable surge in the application of spectroscopy for disease diagnosis. Raman and fluorescence spectra provide characteristic spectral profile related to biochemical and morphological changes when tissues progress from normal state towards malignancy. Spectroscopic techniques offer the advantage of being minimally invasive compared to traditional histopathology, real time and quantitative. In biomedical optical diagnostics, freshly excised specimens are preferred for making ex-vivo spectroscopic measurements. With regard to fresh tissues, if the lab is located far away from the clinic it could pose a problem as spectral measurements have to be performed immediately after dissection. Tissue samples are usually placed in a fixative agent such as 4% formaldehyde to preserve the samples before processing them for routine histopathological studies. Fixation prevents the tissues from decomposition by arresting autolysis. In the present study, we intend to investigate the possibility of using formalin fixed samples for discrimination of brain tumours from dysplastic tissue using Raman and fluorescence spectroscopy. Formalin fixed samples were washed with phosphate buffered saline for about 5 minutes in order to remove the effects of formalin during spectroscopic measurements. In case of fluorescence spectroscopy, changes in spectral profile have been observed in the region between 550-670 nm between dysplastic and tumor samples. For Raman measurements, we found significant differences in the spectral profiles between dysplasia and tumor. In conclusion, formalin fixed samples can be potentially used for the spectroscopic discrimination of tumor against dysplastic tissue in brain samples.

  13. Investigation of biochemical and histopathological effects of Mentha piperita Labiatae and Mentha spicata Labiatae on liver tissue in rats.

    PubMed

    Akdogan, M; Ozguner, M; Aydin, G; Gokalp, O

    2004-01-01

    The plant Mentha piperita, or peppermint, is commonly used in the treatment of loss of appetite, common cold, bronchitis, sinusitis, fever, nausea and vomiting, and indigestion as a herbal agent. In this study, we aimed to investigate biochemical and histological effects of M. piperita Labiatae, growing in the Yenisar Bademli town of Isparta city, and Mentha spicata Labiatae, growing in the Anamas high plateau of the Yenisar Bademli town, on the rat liver tissue. Forty-eight male Wistar albino rats weighing 200-250 g were used for this study. Rats were divided into four groups of 12 animals: Group I received no herbal tea (control group); Group II received 20 g/L M. piperita tea; Group III received 20 g/L M. spicata tea; and Group IV received 40 g/L M. spicata tea. Herbal teas were prepared daily and provided at all times to the rats during 30 days as drinking water. Liver function tests, including aspartate aminotransferase (AST/GOT) and alanine aminotransferase (ALT/GPT) activities were measured. To evaluate liver antioxidant defences, superoxide dismutase (SOD), glutathione peroxidase (GSH-Px), catalase (CAT) and thiobarbituric acid reactive substance (TBARS) activities were determined in the homogenates of liver tissue. In addition, liver tissues were submitted for histopathologic examination. AST and ALT activities were increased in Group II, Group III and Group IV gradually when compared with the control group. The difference between Group II and the control group was not statistically significant (P > 0.016). Increases in AST and ALT activities of Group III and Group IV were statistically significant when compared with the control group. SOD, GSH-Px and CAT activities were increased in Group II when compared with the control group but the difference was not statistically significant (P > 0.016). However, SOD, GSH-Px activities and the TBARS level were significantly increased, and CAT activity was significantly decreased in Group III when compared with the

  14. Antioxidant response and histopathological changes in brain tissue of pigeon exposed to avermectin.

    PubMed

    Li, Ming; You, Tian-Zi; Zhu, Wen-Jun; Qu, Jian-Ping; Liu, Ci; Zhao, Bing; Xu, Shi-Wen; Li, Shu

    2013-10-01

    Avermectins (AVMs) are the active components of some insecticidal and nematicidal products used in agriculture and veterinary medicine for the prevention of parasitic diseases. Residues of AVM drugs or their metabolites in livestock feces have toxic effects on non-target aquatic and terrestrial organisms. In this study, oxidative stress responses and pathological changes on pigeon brain tissues and serum after subchronic exposure to AVM for 30, 60 and 90 days were investigated. The decrease in antioxidant enzyme (superoxide dismutase, SOD and glutathione peroxidase, GSH-Px) activities and increase in methane dicarboxylic aldehyde content in a dose-time-dependent manner in the brain and serum of pigeon were observed. The protein carbonyl content, an indicator of protein oxidation, and DNA-protein crosslink coefficient were significantly augmented with dose-time-dependent properties. The microscopic structures of the cerebrum, cerebellum and optic lobe altered obviously, the severity of which increased with the concentration of AVM and exposure time. The results imply that AVM could induce oxidative damage to the brain tissue and serum of pigeon. The information presented in this study is helpful to understand the mechanism of AVM-induced oxidative stress in birds.

  15. Performance assessment of automated tissue characterization for prostate H and E stained histopathology

    NASA Astrophysics Data System (ADS)

    DiFranco, Matthew D.; Reynolds, Hayley M.; Mitchell, Catherine; Williams, Scott; Allan, Prue; Haworth, Annette

    2015-03-01

    Reliable automated prostate tumor detection and characterization in whole-mount histology images is sought in many applications, including post-resection tumor staging and as ground-truth data for multi-parametric MRI interpretation. In this study, an ensemble-based supervised classification algorithm for high-resolution histology images was trained on tile-based image features including histogram and gray-level co-occurrence statistics. The algorithm was assessed using different combinations of H and E prostate slides from two separate medical centers and at two different magnifications (400x and 200x), with the aim of applying tumor classification models to new data. Slides from both datasets were annotated by expert pathologists in order to identify homogeneous cancerous and non-cancerous tissue regions of interest, which were then categorized as (1) low-grade tumor (LG-PCa), including Gleason 3 and high-grade prostatic intraepithelial neoplasia (HG-PIN), (2) high-grade tumor (HG-PCa), including various Gleason 4 and 5 patterns, or (3) non-cancerous, including benign stroma and benign prostatic hyperplasia (BPH). Classification models for both LG-PCa and HG-PCa were separately trained using a support vector machine (SVM) approach, and per-tile tumor prediction maps were generated from the resulting ensembles. Results showed high sensitivity for predicting HG-PCa with an AUC up to 0.822 using training data from both medical centres, while LG-PCa showed a lower sensitivity of 0.763 with the same training data. Visual inspection of cancer probability heatmaps from 9 patients showed that 17/19 tumors were detected, and HG-PCa generally reported less false positives than LG-PCa.

  16. Characteristics of electrically injured skin from human hand tissue samples using Fourier transform infrared microspectroscopy.

    PubMed

    Li, Shi-Ying; Zou, Dong-Hua; Luo, Yi-Wen; Sun, Qi-Ran; Deng, Kai-Fei; Chen, Yi-Jiu; Huang, Ping

    2014-01-01

    This technical note describes a method for distinguishing normal skin tissue samples from those electrically injured by Fourier transform infrared microspectroscopy (FTIR MSP). Furthermore, the infrared spectral features of electrically injured cells and tissues were evaluated to identify molecular changes in epidermal cells. In the present study, 20 human hand tissue samples were evaluated macroscopically and histopathologically. The electrically injured skin samples were subdivided into 2 regions [normal cell regions (NCRs) and polarized cell regions (PCRs)] and 14 major spectral absorption bands were selected. The spectral results showed that the band absorbance at 1080, 1126, 1172, 1242, 1307, 1403, 1456, 1541, 2852, 2925, 2957, 3075, and 3300cm(-1) increased significantly both in the stratum and non-stratum corneum of the PCRs in electrically injured skin tissues samples. No significant difference was found between normal skin and the NCR of the electrically injured skin samples. The band absorbance ratios of A1172/A1126, A1456/A1403, and A2925/A2957 were significantly increased, whereas the A1652/A1541 ratio was decreased in the PCR of the stratum corneum and non-stratum corneum. Baseline changes from 4000 to near 1737cm(-1) were observed in the spectra of the electrically injured skin samples, which were interpreted in terms of the pathological process involved in electrical injury. FTIR-MSP presents a useful method to provide objective spectral markers for the assisted diagnosis of electrical marks.

  17. Diode laser (808 nm) applied to oral soft tissue lesions: a retrospective study to assess histopathological diagnosis and evaluate physical damage.

    PubMed

    Angiero, Francesca; Parma, Luisa; Crippa, Rolando; Benedicenti, Stefano

    2012-03-01

    The diode laser is today widely used in oral pathology to excise lesions; however, some controversy surrounds laser surgery, specifically the accuracy of pathological diagnosis and the control over thermal tissue damage. This study aimed to establish if physical damage induced by the diode laser could affect the histopathological diagnosis and to evaluate the damage caused to the resection margins. Between 2005 and 2010, at S. Gerardo Hospital, Milan, 608 cases of soft tissue lesions localized in the oral cavity (cheek, gingiva, buccal mucosa, tongue, and lips) were examined. Specimens were excised with an 808-nm diode laser, output 1.6-2.7 W, in continuous-wave mode with fibers of 320 μm. Specimens were fixed in 10% buffered formalin solution and examined separately under an optical microscope by two pathologists. In all of the specimens, changes to the epithelium, connective tissue and blood vessels, shape of incision damage, and overall width of modified tissues were evaluated. The data for specimens larger than 3 mm excised with the diode laser were not significant in terms of stromal changes or vascular stasis, while epithelial and stromal changes were significantly more frequent in specimens with a mean size below 3 mm; the diagnosis was not achievable in 46.15%. Our data show that the diode laser is a valid therapeutic instrument for excising oral lesions larger than 3 mm in diameter, but induces serious thermal effects in small lesions (mean size below 3 mm). However, from a clinical standpoint, it is suggested necessary that the specimens taken have in vivo a diameter of at least 5 mm in order to have a reliable reading of the histological sample. PMID:21387158

  18. Molecular and Histopathological Changes in Mouse Intestinal Tissue After Proton Exposure

    NASA Technical Reports Server (NTRS)

    Purgason, Ashley; Wu, Honglu

    2010-01-01

    Whole body exposure to protons in mice causes significant apoptosis in the crypts of the small intestine. Increasing numbers of crypts contained more apoptotic lesions as the dose of exposure increased. 16 genes associated with apoptotic pathways were shown to have significantly altered expression as compared to control samples for at least one of the doses of proton exposure 1 gene, Trp53inp1, was significantly up-regulated across all three doses. Those animals exposed to 0.1 Gy of proton irradiation showed greater amounts of significant alterations in gene expression as compared to 1 Gy and 2 Gy exposures. The differences in gene expression changes of low and high dose proton irradiated mice may offer insight into the molecular mechanisms of the possible high sensitivity at low proton doses. RAIDD (CRADD) may be responsible for the hypersensitivity observed in the duodenum of mice exposed to low doses of protons. Caspase-1 may also play a role in the hypersensitivity seen following proton irradiation at a dose of 0.1 Gy. FOXO3A may be involved in the down-regulation of GILZ observed at high doses of proton exposure.

  19. Organochlorine-induced histopathology in kidney and liver tissue from Arctic fox (Vulpes lagopus).

    PubMed

    Sonne, Christian; Wolkers, Hans; Leifsson, Pall S; Jenssen, Bjørn Munro; Fuglei, Eva; Ahlstrøm, Oystein; Dietz, Rune; Kirkegaard, Maja; Muir, Derek C G; Jørgensen, Even

    2008-04-01

    The effects of persistent organic pollutants on renal and liver morphology in farmed arctic fox (Vulpes lagopus) were studied under experimental conditions. Control animals received a diet containing pork (Sus scrofa) fat with low amounts of persistent organic pollutants, while the diet of the exposed animals contained whale blubber, 'naturally' contaminated with persistent organic pollutants. Polychlorinated biphenyls (PCB) and organochlorine pesticide (OCP) concentrations in the whale blubber were 488 and 395 ng/g wet weight, respectively. Animals were sacrificed and sampled when they were at their fattest (winter) as well as their lowest body weight (summer). The results show that PCB and OCP exposure causes renal (and probably also liver) lesions in arctic foxes. The prevalence of glomerular, tubular and interstitial lesions was significantly highest in the exposed group (chi-square: all p<0.05). The frequency of liver lesions (steatosis, intravascular granulocyte accumulations, interstitial cell infiltrations, lipid granulomas, portal fibrosis and bile duct hyperplasia) were also highest in the exposed group, although not significantly (chi-square: all p>0.05). The prevalence of lesions was not significantly different between lean (winter) and fat (summer) foxes for any of the lesions (chi-square: all p>0.05). We suggest that wild arctic foxes exposed to an environmental cocktail of persistent organic pollutants, such as PCBs and OCPs, in their natural diet are at risk for developing chronic kidney and liver damage. Whether such lesions may have an impact on age and health of the animals remains uncertain.

  20. Heavy metals and hydrocarbon concentrations in water, sediments and tissue of Cyclope neritea from two sites in Suez Canal, Egypt and histopathological effects.

    PubMed

    Sharaf, Hesham M; Shehata, Abdalla M

    2015-01-01

    Heavy metals and hydrocarbons are of the most common marine pollutants around the world. The present study aimed to assess the concentration of petroleum hydrocarbons and heavy metals in tissues of the snail cyclope neritea, water and sediments from two sites of the study area (Temsah lake and Suez canal) represent polluted and unpolluted sites respectively. The results showed that, the levels of the heavy metals (Pb, Cd, Co, Mg and Zn) in the polluted area have reached harmful limits recorded globally. Lead in water, sediment and tissue of the snail reached to 0.95 ppm, 4.54 ppm and 7.93 ppm respectively. Cadmium reached 0.31 ppm, 1.15 ppm and 3.08 ppm in the corresponding samples. Cobalt was not detected in water, but it reached 1.42 ppm and 10.36 ppm in the sediment and snails tissue respectively. Magnesium in water, sediment and tissue of the snail reached 3.73 ppm, 9.44 ppm and12.6 ppm respectively. Zinc reached 0.11 ppm, 3.89 ppm and 12.60ppm in the corresponding samples. Meanwhile, hydrocarbons in the polluted area (site1) reached 110.10 μg/L, 980.15 μg/g and 228.00 μg/g in water sediment and digestive gland tissues of the snails respectively. Whereas, hydrocarbons in the unpolluted area (site2) were estimated as 14.20 μg/L, 55.60 μg/g and 22.66 μg/g in water, sediment and tissue of the snails respectively. The combination of histopathological image with monitoring of the metal level in the digestive gland of the present snail provides an important tool for early detection of impending environmental problems and potential public health issues. Petroleum hydrocarbons are toxic to the marine fauna when present above certain limit in the marine water. The major detoxification organ in molluscs is the digestive gland, which has been used as a bioindicator organ for toxicity assessment. The effect of high crude oil on the digestive gland tubules of exposed snails when examined microscopically reveals a series of histological changes which indicates that the

  1. A Comparison of Molecular and Histopathological Changes in Mouse Intestinal Tissue Following Whole-Body Proton- or Gamma-Irradiation

    NASA Technical Reports Server (NTRS)

    Purgason, Ashley; Mangala, Lingegowda; Zhang, Ye; Hamilton, Stanley; Wu, Honglu

    2010-01-01

    There are many consequences following exposure to the space radiation environment which can adversely affect the health of a crew member. Acute radiation syndrome (ARS) involving nausea and vomiting, damage to radio-sensitive tissue such as the blood forming organs and gastrointestinal tract, and cancer are some of these negative effects. The space radiation environment is ample with protons and contains gamma rays as well. Little knowledge exists to this point, however, regarding the effects of protons on mammalian systems; conversely several studies have been performed observing the effects of gamma rays on different animal models. For the research presented here, we wish to compare our previous work looking at whole-body exposure to protons using a mouse model to our studies of mice experiencing whole-body exposure to gamma rays as part of the radio-adaptive response. Radio-adaptation is a well-documented phenomenon in which cells exposed to a priming low dose of radiation prior to a higher dose display a reduction in endpoints like chromosomal aberrations, cell death, micronucleus formation, and more when compared to their counterparts receiving high dose-irradiation only. Our group has recently completed a radio-adaptive experiment with C57BL/6 mice. For both this study and the preceding proton research, the gastrointestinal tract of each animal was dissected four hours post-irradiation and the isolated small intestinal tissue was fixed in formalin for histopathological examination or snap-frozen in liquid nitrogen for RNA isolation. Histopathologic observation of the tissue using standard H&E staining methods to screen for morphologic changes showed an increase in apoptotic lesions for even the lowest doses of 0.1 Gy of protons and 0.05 Gy of gamma rays, and the percentage of apoptotic cells increased with increasing dose. A smaller percentage of crypts showed 3 or more apoptotic lesions in animals that received 6 Gy of gamma-irradiation compared to mice

  2. Investigation of biochemical and histopathological effects of Mentha piperita L. and Mentha spicata L. on kidney tissue in rats.

    PubMed

    Akdogan, Mehmet; Kilinç, Ibrahim; Oncu, Meral; Karaoz, Erdal; Delibas, Namik

    2003-04-01

    Peppermint plants have been used as a herbal medicine for many conditions, including loss of appetite, common cold, bronchitis, sinusitis, fever, nausea, vomiting and indigestion. This study is aimed at investigating the biochemical and histological effects of Mentha piperita L., growing in the Yenisar Bademli town of Isparta City, and Mentha spicata L., growing on the Anamas high plateau of Isparta City, on rat kidney tissue. Forty-eight male Wistar albino rats weighing 200-250 g were used for this study. Animals were divided into four experimental groups, each with 12 rats, as follows: control group (group I); 20 g/L M. piperita tea (group II); 20 g/L M. spicata tea (group III); 40 g/L M. spicata tea (group IV). The control group rats were given commercial drinking water (Hayat DANONESA water). The tea for the other groups was prepared daily and provided at all times to the rats during 30 days as drinking water. Plasma urea and creatinine levels were determined, and the levels of thiobarbituric acid reactive substance (TBARS) and the activities of glutathione peroxidase (GSH-Px), catalase (CAT) and superoxide dismutase (SOD) were studied in the homogenates of kidney tissue. The levels of plasma urea and creatinine were increased significantly (P < 0.0033) in groups III and IV when compared with group I. The activities of SOD and GSH-Px were decreased significantly (P < 0.0033) in group IV when compared with group I. The activities of CAT were decreased significantly in groups III and IV (P < 0.033, P < 0.0033, respectively) when compared with group I. TBARS levels were increased significantly (P < 0.0033) in groups III and IV when compared with group I. In groups II, III and IV, hydropic degeneration of tubular epithelial cells, the epithelial cells with picnotic nuclei and eosinophilic cytoplasm, tubular dilatation and enlargements in Bowman capsules were observed histologically. However, in group II histopathological changes were more slight than in groups III

  3. Histopathology reconstruction on digital imagery

    NASA Astrophysics Data System (ADS)

    Li, Wenjing; Lieberman, Rich W.; Nie, Sixiang; Xie, Yihua; Eldred, Michael; Oyama, Jody

    2009-02-01

    Diagnosing cervical cancer in a woman is a multi-step procedure involving examination of the cervix, possible biopsy and follow-up. It is open to subjective interpretation and highly dependent upon the skills of cytologists, colposcopists, and pathologists. In an effort to reduce the subjectiveness of the colposcopist-directed biopsy and to improve the diagnostic accuracy of colposcopy, we have developed new colposcopic imaging systems with accompanying computer aided diagnostic (CAD) techniques to guide a colposcopist in deciding if and where to biopsy. If the biopsy's histopathology, the identification of the disease state at the cellular and near-cellular level, is to be used as the gold standard for CAD, then the location of the histopathologic analysis must match exactly to the location of the biopsy tissue in the digital image. Otherwise, no matter how perfect the histopathology and the quality of the digital imagery, the two data sets cannot be matched and the true sensitivity and specificity of the CAD cannot be ascertained. We report here on new approaches to preserving, continuously, the location and orientation of a biopsy sample with respect to its location in the digital image of the cervix so as to preserve the exact spatial relationship throughout the mechanical aspects of the histopathologic analysis. This new approach will allow CAD to produce a linear diagnosis and pinpoint the location of the tissue under examination.

  4. Tabletop magnetic resonance elastography for the measurement of viscoelastic parameters of small tissue samples

    NASA Astrophysics Data System (ADS)

    Ipek-Ugay, Selcan; Drießle, Toni; Ledwig, Michael; Guo, Jing; Hirsch, Sebastian; Sack, Ingolf; Braun, Jürgen

    2015-02-01

    We demonstrate the feasibility of low-cost tabletop MR elastography (MRE) for quantifying the complex shear modulus G∗ of small soft biological tissue samples as provided by pathologists. The MRE system was developed based on a tabletop MRI scanner equipped with a 0.5 T permanent magnet and a tissue sample holder mounted to a loudspeaker. A spin echo sequence was enhanced with motion-encoding gradients of 250 mT/m amplitude synchronized to acoustic vibration frequencies. Shear wave images suitable for elastography were acquired between vibration frequencies of 0.5 and 1 kHz in agarose, ultrasound gel, porcine liver, porcine skeletal muscle, and bovine heart with a spatial resolution of 234 μm pixel edge length. The measured frequency dependence of G∗ agreed well with previous work based on high-field MR systems. The ratio between loss and storage moduli was highest in liver and ultrasound gel, followed by muscle tissue and agarose gel while ultrasound gel and liver showed similarly low storage moduli compared to the other samples. The shear wave to noise ratio is an important imaging criteria for MRE and was about 4.2 times lower for the preliminary setup of the 0.5 T tabletop system compared to a 7 T animal scanner. In the future, the new tabletop MRE system may serve as a low cost device for preclinical research on the correlation of viscoelastic parameters with histopathology of biological samples.

  5. Technical note: Alternatives to reduce adipose tissue sampling bias.

    PubMed

    Cruz, G D; Wang, Y; Fadel, J G

    2014-10-01

    Understanding the mechanisms by which nutritional and pharmaceutical factors can manipulate adipose tissue growth and development in production animals has direct and indirect effects in the profitability of an enterprise. Adipocyte cellularity (number and size) is a key biological response that is commonly measured in animal science research. The variability and sampling of adipocyte cellularity within a muscle has been addressed in previous studies, but no attempt to critically investigate these issues has been proposed in the literature. The present study evaluated 2 sampling techniques (random and systematic) in an attempt to minimize sampling bias and to determine the minimum number of samples from 1 to 15 needed to represent the overall adipose tissue in the muscle. Both sampling procedures were applied on adipose tissue samples dissected from 30 longissimus muscles from cattle finished either on grass or grain. Briefly, adipose tissue samples were fixed with osmium tetroxide, and size and number of adipocytes were determined by a Coulter Counter. These results were then fit in a finite mixture model to obtain distribution parameters of each sample. To evaluate the benefits of increasing number of samples and the advantage of the new sampling technique, the concept of acceptance ratio was used; simply stated, the higher the acceptance ratio, the better the representation of the overall population. As expected, a great improvement on the estimation of the overall adipocyte cellularity parameters was observed using both sampling techniques when sample size number increased from 1 to 15 samples, considering both techniques' acceptance ratio increased from approximately 3 to 25%. When comparing sampling techniques, the systematic procedure slightly improved parameters estimation. The results suggest that more detailed research using other sampling techniques may provide better estimates for minimum sampling.

  6. Specimen Sample Preservation for Cell and Tissue Cultures

    NASA Technical Reports Server (NTRS)

    Meeker, Gabrielle; Ronzana, Karolyn; Schibner, Karen; Evans, Robert

    1996-01-01

    The era of the International Space Station with its longer duration missions will pose unique challenges to microgravity life sciences research. The Space Station Biological Research Project (SSBRP) is responsible for addressing these challenges and defining the science requirements necessary to conduct life science research on-board the International Space Station. Space Station will support a wide range of cell and tissue culture experiments for durations of 1 to 30 days. Space Shuttle flights to bring experimental samples back to Earth for analyses will only occur every 90 days. Therefore, samples may have to be retained for periods up to 60 days. This presents a new challenge in fresh specimen sample storage for cell biology. Fresh specimen samples are defined as samples that are preserved by means other than fixation and cryopreservation. The challenge of long-term storage of fresh specimen samples includes the need to suspend or inhibit proliferation and metabolism pending return to Earth-based laboratories. With this challenge being unique to space research, there have not been any ground based studies performed to address this issue. It was decided hy SSBRP that experiment support studies to address the following issues were needed: Fixative Solution Management; Media Storage Conditions; Fresh Specimen Sample Storage of Mammalian Cell/Tissue Cultures; Fresh Specimen Sample Storage of Plant Cell/Tissue Cultures; Fresh Specimen Sample Storage of Aquatic Cell/Tissue Cultures; and Fresh Specimen Sample Storage of Microbial Cell/Tissue Cultures. The objective of these studies was to derive a set of conditions and recommendations that can be used in a long duration microgravity environment such as Space Station that will permit extended storage of cell and tissue culture specimens in a state consistent with zero or minimal growth, while at the same time maintaining their stability and viability.

  7. Analysis of chemical components from plant tissue samples

    NASA Technical Reports Server (NTRS)

    Laseter, J. L.

    1972-01-01

    Information is given on the type and concentration of sterols, free fatty acids, and total fatty acids in plant tissue samples. All samples were analyzed by gas chromatography and then by gas chromatography-mass spectrometry combination. In each case the mass spectral data was accumulated as a computer printout and plot. Typical gas chromatograms are included as well as tables describing test results.

  8. Multivariate classification of infrared spectra of cell and tissue samples

    DOEpatents

    Haaland, David M.; Jones, Howland D. T.; Thomas, Edward V.

    1997-01-01

    Multivariate classification techniques are applied to spectra from cell and tissue samples irradiated with infrared radiation to determine if the samples are normal or abnormal (cancerous). Mid and near infrared radiation can be used for in vivo and in vitro classifications using at least different wavelengths.

  9. Solid phase microextraction fills the gap in tissue sampling protocols.

    PubMed

    Bojko, Barbara; Gorynski, Krzysztof; Gomez-Rios, German Augusto; Knaak, Jan Matthias; Machuca, Tiago; Spetzler, Vinzent Nikolaus; Cudjoe, Erasmus; Hsin, Michael; Cypel, Marcelo; Selzner, Markus; Liu, Mingyao; Keshavjee, Shaf; Pawliszyn, Janusz

    2013-11-25

    Metabolomics and biomarkers discovery are an integral part of bioanalysis. However, untargeted tissue analysis remains as the bottleneck of such studies due to the invasiveness of sample collection, as well as the laborious and time-consuming sample preparation protocols. In the current study, technology integrating in vivo sampling, sample preparation and global extraction of metabolites--solid phase microextraction was presented and evaluated during liver and lung transplantation in pig model. Sampling approaches, including selection of the probe, transportation, storage conditions and analyte coverage were discussed. The applicability of the method for metabolomics studies was demonstrated during lung transplantation experiments. PMID:24216199

  10. Heavy metals in selected tissues and histopathological changes in liver and kidney of common moorhen (Gallinula chloropus) from Anzali Wetland, the south Caspian Sea, Iran.

    PubMed

    Salamat, Negin; Etemadi-Deylami, Eelia; Movahedinia, Abdolali; Mohammadi, Yaghoob

    2014-12-01

    The present study aimed to measure the concentrations of Sn, Pb, Zn, Hg, Cu, Ni and Cd in the muscle and liver of 40 Common Moorhens (Gallinula chloropus) hunted from four stations in Anzali Wetland (Pirbazar, Ghalam-Koudeh, Selkeh and Abkenar). The histopathologic alteration index (HAI) of liver and kidney was also assessed in these birds. The highest concentrations of selected metals were measured in the liver of birds collected from Ghalam-Koudeh (Pb: 4.59±0.21, Sn: 6.663±0.282, Zn: 29.867±2.011, Cu: 24.07±1.84, Hg: 7.5±0.257, Ni: 6.85±0.52, Cd: 1.879±0.4mg kg(-1) dw). The lowest concentrations of metals were measured in the muscle of birds caught from Abkenar (Pb: 0.799±0.207, Sn: 1.873±0.066, Zn: 18.533±1.582, Hg: 0.86±0.08, Ni: 0.53±0.117, Cu: 6.63±1.114, Cd: 0.08±0.002mg kg(-1) dw). Also the highest and lowest concentrations of metals were recorded in sediment of Ghalam-Koudeh and Abkenar stations, respectively. These stations were located next to multi-industry Anzali Port. However, the concentration of Sn and Zn in sediment and tissues of Common Moorhens collected from different stations was lower than the permissible limit suggested by WHO and Canadian Council of Ministers of the Environment (CCME). But, Pb, Hg and Ni concentration in sediment and birds caught from all stations was higher than the permissible limit defined by WHO and CCME. Cu and Cd concentration in tissue samples and sediment of Ghalam-Koudeh and Pirbazar was also higher than the permissible limit defined by WHO and CCME. Hemorrhage, melanomacrophage aggregations, sinusoidal congestion and hepatocyte vacuolation were the most pathological changes found in the liver. Reduction of the Bowman space, melanomacrophage aggregations and hemorrhage also were observed in the kidney. The HAI means of G. chloropus collected from Ghalam-Koudeh and Pirbazar were significantly higher than other sites. Based on the HAI values and metal bioaccumulation in the tissues of G. chloropus

  11. Heavy metals in selected tissues and histopathological changes in liver and kidney of common moorhen (Gallinula chloropus) from Anzali Wetland, the south Caspian Sea, Iran.

    PubMed

    Salamat, Negin; Etemadi-Deylami, Eelia; Movahedinia, Abdolali; Mohammadi, Yaghoob

    2014-12-01

    The present study aimed to measure the concentrations of Sn, Pb, Zn, Hg, Cu, Ni and Cd in the muscle and liver of 40 Common Moorhens (Gallinula chloropus) hunted from four stations in Anzali Wetland (Pirbazar, Ghalam-Koudeh, Selkeh and Abkenar). The histopathologic alteration index (HAI) of liver and kidney was also assessed in these birds. The highest concentrations of selected metals were measured in the liver of birds collected from Ghalam-Koudeh (Pb: 4.59±0.21, Sn: 6.663±0.282, Zn: 29.867±2.011, Cu: 24.07±1.84, Hg: 7.5±0.257, Ni: 6.85±0.52, Cd: 1.879±0.4mg kg(-1) dw). The lowest concentrations of metals were measured in the muscle of birds caught from Abkenar (Pb: 0.799±0.207, Sn: 1.873±0.066, Zn: 18.533±1.582, Hg: 0.86±0.08, Ni: 0.53±0.117, Cu: 6.63±1.114, Cd: 0.08±0.002mg kg(-1) dw). Also the highest and lowest concentrations of metals were recorded in sediment of Ghalam-Koudeh and Abkenar stations, respectively. These stations were located next to multi-industry Anzali Port. However, the concentration of Sn and Zn in sediment and tissues of Common Moorhens collected from different stations was lower than the permissible limit suggested by WHO and Canadian Council of Ministers of the Environment (CCME). But, Pb, Hg and Ni concentration in sediment and birds caught from all stations was higher than the permissible limit defined by WHO and CCME. Cu and Cd concentration in tissue samples and sediment of Ghalam-Koudeh and Pirbazar was also higher than the permissible limit defined by WHO and CCME. Hemorrhage, melanomacrophage aggregations, sinusoidal congestion and hepatocyte vacuolation were the most pathological changes found in the liver. Reduction of the Bowman space, melanomacrophage aggregations and hemorrhage also were observed in the kidney. The HAI means of G. chloropus collected from Ghalam-Koudeh and Pirbazar were significantly higher than other sites. Based on the HAI values and metal bioaccumulation in the tissues of G. chloropus

  12. Using tissue samples for proteomic studies-critical considerations.

    PubMed

    Becker, Karl-Friedrich

    2015-04-01

    From my experience of 22 years working in a pathology research laboratory and overseeing dozens of collaborations with research groups from basic sciences and industry, I have the impression that researchers are rarely aware of the special issues related to acquisition and processing of frozen or formalin-fixed tissue samples for proteomic analysis. While challenges are expected for formalin-fixed tissues because of the cross-linking activities of formaldehyde, researchers believe when using frozen tissue samples they are safe and always have excellent material to analyze-but this is not always the case. It is alarming that many researchers do not question the quality of the tissue samples they are analyzing and focus only on their analytical technique. Standardization of the entire workflow from test ordering to the report of the proteomic assay, with special emphasis on the preanalytical phase, is crucial for successful integration of proteomic studies in the clinic as protein profiles may change due to sample processing before the proteomic analysis is performed. The aim of this review is to discuss the progress of proteomic studies with human tissues and to highlight the challenges that must be understood and addressed for successful translation of proteomic methods to clinical practice.

  13. Tissue sampling methods and standards for vertebrate genomics

    PubMed Central

    2012-01-01

    The recent rise in speed and efficiency of new sequencing technologies have facilitated high-throughput sequencing, assembly and analyses of genomes, advancing ongoing efforts to analyze genetic sequences across major vertebrate groups. Standardized procedures in acquiring high quality DNA and RNA and establishing cell lines from target species will facilitate these initiatives. We provide a legal and methodological guide according to four standards of acquiring and storing tissue for the Genome 10K Project and similar initiatives as follows: four-star (banked tissue/cell cultures, RNA from multiple types of tissue for transcriptomes, and sufficient flash-frozen tissue for 1 mg of DNA, all from a single individual); three-star (RNA as above and frozen tissue for 1 mg of DNA); two-star (frozen tissue for at least 700 μg of DNA); and one-star (ethanol-preserved tissue for 700 μg of DNA or less of mixed quality). At a minimum, all tissues collected for the Genome 10K and other genomic projects should consider each species’ natural history and follow institutional and legal requirements. Associated documentation should detail as much information as possible about provenance to ensure representative sampling and subsequent sequencing. Hopefully, the procedures outlined here will not only encourage success in the Genome 10K Project but also inspire the adaptation of standards by other genomic projects, including those involving other biota. PMID:23587255

  14. Histopathological Osteomyelitis Evaluation Score (HOES) – an innovative approach to histopathological diagnostics and scoring of osteomyelitis

    PubMed Central

    Tiemann, A.; Hofmann, G. O.; Krukemeyer, M. G.; Krenn, V.; Langwald, S.

    2014-01-01

    Background: Treatment and diagnosis of osteomyelitis are still a challenging problem for surgeons, microbiologists and histopathologists. A direct microbiological detection of bacteria in tissues is still gold standard, but it is not always successful for example in chronic osteomyelitis and/or when an antibiotic treatment has already been started or in cases of low virulent bacteria. The goal of this study was to define diagnostic criteria of osteomyelitis, the inflammatory regression of osteomyelitis (“osteomyelitis score”) under specific therapy by the correlation of histopathological and microbiological and clinical standard tests. Methods: In this retrospective analysis patients with medical history and clinically clear signs of bacterial infection and osteomyelitis underwent surgery between 01.01.2013 and 31.12.2012. Their formal consent was given. Tissue samples were taken during surgery according to defined criteria including surgical interventions. Histopathological diagnosis was carried out by conventional techniques based on defined criteria of bacterial infection in connective tissue, peri-implant membrane and bone. These results were carried out in tables by numbers representing the histopathological criteria of acute osteomyelitis (A1 to A3) as well as the chronic criteria (C1 and C2) in a semiquantitative way (scale 0 to 3). On the other hand a notational, graduated histopathological report was performed. Preoperative clinical diagnosis, perioperative macroscopic diagnosis, histopathological and microbiological findings were correlated. Results: Histopathological samples of 52 surgical interventions based on the preoperative diagnosis “osteomyelitis” (AOM, ECOM or COM) were included. 37 times preoperatively signs of a chronic osteomyelitis (COM), 10 times preoperatively acute osteomyelitis (AOM) was diagnosed. Another 5 patients were preoperatively diagnosed as acute exacerbated osteomyelitis (ECOM). The correlation of the histopathological

  15. Translational research in pediatrics: tissue sampling and biobanking.

    PubMed

    Brisson, Alayne R; Matsui, Doreen; Rieder, Michael J; Fraser, Douglas D

    2012-01-01

    Translational research is expanding and has become a focus of National Research funding agencies, touted as the primary avenue to improve health care practice. The use of human tissues for research on disease etiology is a pillar of translational research, particularly with innovations in research technologies to investigate the building blocks of disease. In pediatrics, translational research using human tissues has been hindered by the many practical and ethical considerations associated with tissue procurement from children and also by a limited population base for study, by the increasing complexities in conducting clinical research, and by a lack of dedicated child-health research funding. Given these obstacles, pediatric translational research can be enhanced by developing strategic and efficient biobanks that will provide scientists with quality tissue specimens to render accurate and reproducible research results. Indeed, tissue sampling and biobanking within pediatric academic settings has potential to impact child health by promoting bidirectional interaction between clinicians and scientists, helping to maximize research productivity, and providing a competitive edge for attracting and maintaining high-quality personnel. The authors of this review outline key issues and practical solutions to optimize pediatric tissue sampling and biobanking for translational research, activities that will ultimately reduce the burden of childhood disease. PMID:22144705

  16. Histopathological follow-up by tissue micro-array in a survival study after melanoma treated by nanosecond pulsed electric fields (nsPEF).

    PubMed

    Chen, Xinhua; Chen, Xinmei; Swanson, R James; Schoenbach, Karl H; Yin, Shengyong; Zheng, Shusen

    2011-06-01

    A recent study has shown that nanosecond pulsed electric fields (nsPEF) can affect the intracellular structures of melanoma within weeks. nsPEF is a non-drug, non-thermal treatment using ultrashort, intense pulsed electric fields with nanosecond durations. In the current study we followed up melanoma histopathology and metastasis with tissue micro-array 5 months post-nsPEF. After nsPEF treatment, tumor growth, tumor histology, metastasis, peri-tumor vessel and micro-vessel density were examined for the effect of nsPEF treatment on melanoma in vivo. The 17 nsPEF-treated mice were tumor-free for 169 days, significantly longer than those 19 control mice bearing melanoma without nsPEF. Histopathology follow-up showed that melanoma did not recur to the primary injection place after complete elimination. Compared with the control tumor, nsPEF-treated tumors present decreased micro-vessel density in a time-course manner in this survival study. Treatment with nsPEF caused continuous histopathological changes in melanomas, eliminated melanoma without recurrence at the primary site and prolonged animal survival time by inhibiting tumor blood supply and leading to tumor infarction. Thus, nsPEF could be applied in a non-ionizing therapeutic approach, without other agents, to locally treat tumors within a defined boundary.

  17. Histopathology of Incontinence-Associated Skin Lesions: Inner Tissue Damage Due to Invasion of Proteolytic Enzymes and Bacteria in Macerated Rat Skin

    PubMed Central

    Mugita, Yuko; Minematsu, Takeo; Huang, Lijuan; Nakagami, Gojiro; Kishi, Chihiro; Ichikawa, Yoshie; Nagase, Takashi; Oe, Makoto; Noguchi, Hiroshi; Mori, Taketoshi; Abe, Masatoshi; Sugama, Junko; Sanada, Hiromi

    2015-01-01

    A common complication in patients with incontinence is perineal skin lesions, which are recognized as a form of dermatitis. In these patients, perineal skin is exposed to digestive enzymes and intestinal bacterial flora, as well as excessive water. The relative contributions of digestive enzymes and intestinal bacterial flora to skin lesion formation have not been fully shown. This study was conducted to reveal the process of histopathological changes caused by proteases and bacterial inoculation in skin maceration. For skin maceration, agarose gel containing proteases was applied to the dorsal skin of male Sprague-Dawley rats for 4 h, followed by Pseudomonas aeruginosa inoculation for 30 min. Macroscopic changes, histological changes, bacterial distribution, inflammatory response, and keratinocyte proliferation and differentiation were examined. Proteases induced digestion in the prickle cell layer of the epidermis, and slight bleeding in the papillary dermis and around hair follicles in the macerated skin without macroscopic evidence of erosion. Bacterial inoculation of the skin macerated by proteolytic solution resulted in the formation of bacteria-rich clusters comprising numerous microorganisms and inflammatory cells within the papillary dermis, with remarkable tissue damage around the clusters. Tissue damage expanded by day 2. On day 3, the proliferative keratinocyte layer was elongated from the bulge region of the hair follicles. Application of proteases and P. aeruginosa induced skin lesion formation internally without macroscopic erosion of the overhydrated area, suggesting that the histopathology might be different from regular dermatitis. The healing process of this lesion is similar to transepidermal elimination. PMID:26407180

  18. Histopathology of Incontinence-Associated Skin Lesions: Inner Tissue Damage Due to Invasion of Proteolytic Enzymes and Bacteria in Macerated Rat Skin.

    PubMed

    Mugita, Yuko; Minematsu, Takeo; Huang, Lijuan; Nakagami, Gojiro; Kishi, Chihiro; Ichikawa, Yoshie; Nagase, Takashi; Oe, Makoto; Noguchi, Hiroshi; Mori, Taketoshi; Abe, Masatoshi; Sugama, Junko; Sanada, Hiromi

    2015-01-01

    A common complication in patients with incontinence is perineal skin lesions, which are recognized as a form of dermatitis. In these patients, perineal skin is exposed to digestive enzymes and intestinal bacterial flora, as well as excessive water. The relative contributions of digestive enzymes and intestinal bacterial flora to skin lesion formation have not been fully shown. This study was conducted to reveal the process of histopathological changes caused by proteases and bacterial inoculation in skin maceration. For skin maceration, agarose gel containing proteases was applied to the dorsal skin of male Sprague-Dawley rats for 4 h, followed by Pseudomonas aeruginosa inoculation for 30 min. Macroscopic changes, histological changes, bacterial distribution, inflammatory response, and keratinocyte proliferation and differentiation were examined. Proteases induced digestion in the prickle cell layer of the epidermis, and slight bleeding in the papillary dermis and around hair follicles in the macerated skin without macroscopic evidence of erosion. Bacterial inoculation of the skin macerated by proteolytic solution resulted in the formation of bacteria-rich clusters comprising numerous microorganisms and inflammatory cells within the papillary dermis, with remarkable tissue damage around the clusters. Tissue damage expanded by day 2. On day 3, the proliferative keratinocyte layer was elongated from the bulge region of the hair follicles. Application of proteases and P. aeruginosa induced skin lesion formation internally without macroscopic erosion of the overhydrated area, suggesting that the histopathology might be different from regular dermatitis. The healing process of this lesion is similar to transepidermal elimination. PMID:26407180

  19. Histopathology of Incontinence-Associated Skin Lesions: Inner Tissue Damage Due to Invasion of Proteolytic Enzymes and Bacteria in Macerated Rat Skin.

    PubMed

    Mugita, Yuko; Minematsu, Takeo; Huang, Lijuan; Nakagami, Gojiro; Kishi, Chihiro; Ichikawa, Yoshie; Nagase, Takashi; Oe, Makoto; Noguchi, Hiroshi; Mori, Taketoshi; Abe, Masatoshi; Sugama, Junko; Sanada, Hiromi

    2015-01-01

    A common complication in patients with incontinence is perineal skin lesions, which are recognized as a form of dermatitis. In these patients, perineal skin is exposed to digestive enzymes and intestinal bacterial flora, as well as excessive water. The relative contributions of digestive enzymes and intestinal bacterial flora to skin lesion formation have not been fully shown. This study was conducted to reveal the process of histopathological changes caused by proteases and bacterial inoculation in skin maceration. For skin maceration, agarose gel containing proteases was applied to the dorsal skin of male Sprague-Dawley rats for 4 h, followed by Pseudomonas aeruginosa inoculation for 30 min. Macroscopic changes, histological changes, bacterial distribution, inflammatory response, and keratinocyte proliferation and differentiation were examined. Proteases induced digestion in the prickle cell layer of the epidermis, and slight bleeding in the papillary dermis and around hair follicles in the macerated skin without macroscopic evidence of erosion. Bacterial inoculation of the skin macerated by proteolytic solution resulted in the formation of bacteria-rich clusters comprising numerous microorganisms and inflammatory cells within the papillary dermis, with remarkable tissue damage around the clusters. Tissue damage expanded by day 2. On day 3, the proliferative keratinocyte layer was elongated from the bulge region of the hair follicles. Application of proteases and P. aeruginosa induced skin lesion formation internally without macroscopic erosion of the overhydrated area, suggesting that the histopathology might be different from regular dermatitis. The healing process of this lesion is similar to transepidermal elimination.

  20. Identification of fungal pathogens in Formalin-fixed, Paraffin-embedded tissue samples by molecular methods.

    PubMed

    Rickerts, Volker

    2016-02-01

    The etiology of invasive fungal infections (IFI) is incompletely understood due to diagnostic limitations including insensitivity of cultures and failure of histopathology to discriminate between different species. This diagnostic gap precludes the optimal use of antifungals, leading to adverse patient outcomes. The identification of fungal pathogens from Formalin-fixed, Paraffin-embedded tissue (FFPE) blocks by molecular methods is emerging as an alternative approach to study the etiology of IFI. PCR assays, including species specific- and broadrange fungal tests are used with FFPE samples from patients with proven IFI. Fungal species identification is achieved in 15-90% of the samples. This heterogeneity may be explained by the samples studied. However, comparison of different studies is impaired, as controls ruling out false positive-, false negative test results or PCR inhibition are frequently not reported. Studies using in situ hybridization also vary in the clinical samples included and the targeted fungi. In addition, target sequences, the probe chemistry and the detection of hybridization signals also account for the differences in diagnostic sensitivity. Using both approaches in parallel yields additive insights, potentially leading to a superior identification of fungal etiology and awareness of the limitations of both molecular diagnostic approaches.

  1. Semiautomated Device for Batch Extraction of Metabolites from Tissue Samples

    PubMed Central

    2012-01-01

    Metabolomics has become a mainstream analytical strategy for investigating metabolism. The quality of data derived from these studies is proportional to the consistency of the sample preparation. Although considerable research has been devoted to finding optimal extraction protocols, most of the established methods require extensive sample handling. Manual sample preparation can be highly effective in the hands of skilled technicians, but an automated tool for purifying metabolites from complex biological tissues would be of obvious utility to the field. Here, we introduce the semiautomated metabolite batch extraction device (SAMBED), a new tool designed to simplify metabolomics sample preparation. We discuss SAMBED’s design and show that SAMBED-based extractions are of comparable quality to extracts produced through traditional methods (13% mean coefficient of variation from SAMBED versus 16% from manual extractions). Moreover, we show that aqueous SAMBED-based methods can be completed in less than a quarter of the time required for manual extractions. PMID:22292466

  2. Characterization of different tissue changes in normal, betel chewers, potentially malignant lesions, conditions and oral squamous cell carcinoma using reflectance confocal microscopy: correlation with routine histopathology.

    PubMed

    Anuthama, Krishnamurthy; Sherlin, Herald J; Anuja, N; Ramani, Pratibha; Premkumar, Priya; Chandrasekar, T

    2010-04-01

    The goal of this study was to characterize the features of normal mucosa, mucosa in betel chewers and smokers, potentially malignant lesions and squamous cell carcinoma of oral mucosa using reflectance confocal microscopy. Oral cavity biopsies were acquired from 25 patients from College of Dental Surgery, Saveetha University who underwent screening for suspected lesions of Oral precancer and Oral cancer along with normal patients who underwent impaction. Biopsies were acquired from the clinically suspicious area and immediately placed in Dulbecco modified eagles growth medium (DMEM). Reflectance confocal images were obtained at multiple image plane depths from biopsies within 6h of excision. After imaging, biopsies were fixed in 10% formalin and submitted for routine histopathological examination by an experienced oral and maxillofacial pathologist. Reflectance confocal images were compared with histological images from the same sample to determine the tissue features which contribute to early cellular changes, image contrast and early diagnosis. The confocal images were obtained to a depth of up to 150 microns on intact biopsy specimens and subsequent 3-dimensional images, keratin thickness measurements, cell measurements, cell density analysis and graphical representations were performed using Leica image analysis software. In normal mucosa keratin deposition were seen as alternating dark and bright stacks and in different cell layers the nuclei were seen as disks of varying intensities. In pre-cancerous lesions the keratin thickness and cell nuclear density were found to be increased when compared to normal controls. In OSMF cases confocal images of fibrosis show scattering from individual fibres as hyperdense areas. Oral squamous cell carcinoma cases demonstrated extensive variations in cell size, nuclear size and nuclear morphology. At cellular level, dysplastic features like increased nuclear density, increased nuclear cytoplasmic ratio, nuclear and cellular

  3. Effects of acrylamide exposure on serum hormones, gene expression, cell proliferation, and histopathology in male reproductive tissues of Fischer 344 rats.

    PubMed

    Camacho, L; Latendresse, J R; Muskhelishvili, L; Patton, R; Bowyer, J F; Thomas, M; Doerge, D R

    2012-06-01

    Acrylamide (AA) is a reactive monomer used in many technological applications, but it is the incidental formation during cooking of common starchy foods that leads to pervasive human exposure, typically in the range of 1 μg/kg body weight (bw)/day (d). AA is carcinogenic in multiple organs from both sexes of several rodent models and a consistent body of evidence points to a genotoxic mechanism based on metabolism to a DNA-reactive epoxide, glycidamide (GA). In F344 rats, tumorigenesis occurs in several hormonally regulated tissues (thyroid, mammary gland, and peri-testicular mesothelium), which has prompted speculation about endocrine dysregulation as a possible mechanism. The present study evaluated the effects of a 14 d exposure to AA administered through the drinking water on reproductive tissues and the hypothalamic-pituitary-testes (HPG) axis in male F344 rats. The doses selected encompass a range from approximately 2.5 mg/kg bw/d, which is carcinogenic after lifetime exposure, to 50 mg/kg bw/d, a maximally tolerable dose that causes hind limb paralysis. AA caused significant changes in serum hormones, histopathology, testicular gene expression, and cell proliferation, especially at the highest dose. Despite strong evidence for activation of the HPG axis subsequent to decreases in testosterone levels, and histopathological changes associated with significant effects on Leydig and germ cells, with concomitant mRNA expression changes, the precise mechanism(s) for AA-induced testicular toxicity remains unclear; however, the absence of evidence for increased proliferation of the peri-testicular mesothelium (Ki-67 immunoreactivity) does not support hormonal dysregulation as a contributing factor to the predisposition of this tissue to the carcinogenic effects of AA.

  4. An adapted tissue microarray for the development of a matrix arrangement of tissue samples.

    PubMed

    Gurgel, Daniel C; Dornelas, Conceição A; Lima-Júnior, Roberto C P; Ribeiro, Ronaldo A; Almeida, Paulo R C

    2012-03-15

    The arrangement of tissue samples in a matrix, known as the tissue microarray (TMA) method, is a well-recognized method worldwide. This technique makes it possible to assess the expression of molecular markers on a large scale with high yields in terms of time, costs, and archived material. Some researchers are trying to adapt the technique to expand the research possibilities. This study proposes an adaptive simplification of low-cost instruments for obtaining samples that will be used in the construction of the TMA. The use of a manual leather puncher, which has a very low cost and a long expected life and eliminates the need to use a press machine, is a simple and effective alternative to building blocks of tissue microarrays.

  5. Laser microdissection of small tissue samples--application to chronic pancreatitis tissues.

    PubMed

    Heinmöller, Ernst; Bockholt, Anke; Werther, Meike; Ziemer, Maria; Müller, Annegret; Ghadimi, B Michael; Rüschoff, Josef

    2003-01-01

    Laser microdissection is considered to be the gold standard of tissue sampling, especially if a defined small tissue area consisting of single or few cells within a heterogeneous tissue compartment is of interest. This sophisticated technique offers the opportunity of rapid and contamination-free tissue sampling for RNA- or DNA-based molecular genetic studies. We have applied laser microdissection to a molecular genetic study of pancreatic intraductal lesions (PanINs) in tissues of chronic pancreatitis, where an exact microdissection of small ducts within a dense fibrous tissue is of paramount importance for following analysis. From nine patients suffering from chronic pancreatitis, formalin-fixed, paraffin-embedded tissue specimens were laser microdissected, and a total of 202 normal ducts and PanINs of grade PanIN-1A to grade PanIN-2 were harvested. After whole genome amplification by improved primer extension and preamplification PCR (I-PEP-PCR), microsatellite-PCR based loss of heterozygosity analysis (LOH) of the tumor suppressor gene loci TP53, p16INK4, and DPC4 was performed. One of 85 informative duct lesions (1.2%) had LOH of TP53, 1 of 76 duct lesions (1.3%) had LOH of DPC4, and 2/29 duct lesions (6.9%) showed LOH of p16INK4. Microsatellite instability (MSI) was seen in 2 of 178 duct lesions (1.1%). Immunohistochemical staining of p53 protein and DPC4 protein revealed no aberrant expression. These preliminary data indicate that LOH of tumor suppressor genes, important in pancreatic cancer genesis or MSI, can be found in chronic pancreatitis tissues, but their incidence is low. PMID:12924436

  6. Mercury and histopathology of the vulnerable goliath grouper, Epinephelus itajara, in U.S. waters: a multi-tissue approach.

    PubMed

    Adams, Douglas H; Sonne, Christian

    2013-10-01

    Goliath grouper have undergone significant global population declines with potential biological extinction for some subpopulations. Although overfishing and habitat loss are important drivers of these declines, the negative effects of contaminants may also play a role. The life history patterns of goliath grouper may make this species especially prone to exposure to contaminants and may exacerbate bioaccumulation of toxic substances, including mercury, which has documented detrimental health effects. Therefore, we analyzed mercury (in muscle, liver, kidney, gonad, and brain tissue) and the histology of key organs (liver, kidney and gill tissue) in 56 goliath groupers from U.S. waters. Total mercury concentration was greatest in liver tissue, followed by kidney, muscle, gonad, and brain. Maximum mercury concentration ranged from 22.68 μg/g in liver tissue to 0.89 μg/g in brain tissue. Mean mercury concentration ranged from 2.87 μg/g in liver tissue to 0.37 μg/g in brain tissue with a mean of 0.63 μg/g in muscle. Mean mercury concentrations observed in goliath grouper from U.S. waters were within the range known to cause direct health effects in fish after long-term exposure. The lesions and histological changes observed in the liver, kidney, and gills of goliath groupers were similar to those found in other fish following laboratory mercury-exposure trials and to those found in mercury-contaminated fish in wild populations carrying similar or even lower concentrations. We suggest that exposure to mercury and other environmental influences such as pathogens and reduced temperatures could be co-factors in the histological effects or anomalies observed in the present study, and resulting stresses may be involved in the observed population declines in the species. PMID:23830062

  7. Mercury and histopathology of the vulnerable goliath grouper, Epinephelus itajara, in U.S. waters: a multi-tissue approach.

    PubMed

    Adams, Douglas H; Sonne, Christian

    2013-10-01

    Goliath grouper have undergone significant global population declines with potential biological extinction for some subpopulations. Although overfishing and habitat loss are important drivers of these declines, the negative effects of contaminants may also play a role. The life history patterns of goliath grouper may make this species especially prone to exposure to contaminants and may exacerbate bioaccumulation of toxic substances, including mercury, which has documented detrimental health effects. Therefore, we analyzed mercury (in muscle, liver, kidney, gonad, and brain tissue) and the histology of key organs (liver, kidney and gill tissue) in 56 goliath groupers from U.S. waters. Total mercury concentration was greatest in liver tissue, followed by kidney, muscle, gonad, and brain. Maximum mercury concentration ranged from 22.68 μg/g in liver tissue to 0.89 μg/g in brain tissue. Mean mercury concentration ranged from 2.87 μg/g in liver tissue to 0.37 μg/g in brain tissue with a mean of 0.63 μg/g in muscle. Mean mercury concentrations observed in goliath grouper from U.S. waters were within the range known to cause direct health effects in fish after long-term exposure. The lesions and histological changes observed in the liver, kidney, and gills of goliath groupers were similar to those found in other fish following laboratory mercury-exposure trials and to those found in mercury-contaminated fish in wild populations carrying similar or even lower concentrations. We suggest that exposure to mercury and other environmental influences such as pathogens and reduced temperatures could be co-factors in the histological effects or anomalies observed in the present study, and resulting stresses may be involved in the observed population declines in the species.

  8. Immunoelectron microscopic localization of elastic tissue components in archival tissue samples.

    PubMed

    Fanning, J C; White, J F; Polewski, R; Cleary, E G

    1991-06-01

    Tissue samples that have been stored for many years, in different media and under a variety of conditions, have been examined by modern techniques of immunoelectron microscopy, using antibodies against elastic tissue components. A range of postembedding restorative procedures has been identified, which will allow reliable immunolocalization of antibodies against the elastic tissue component of such specimens. These methods have been applied successfully to autopsy-derived material, fixed in buffered formaldehyde, to archival material stored frozen at -70 or -20 degrees C, to specimens fixed for electron microscopy and stored for many years in buffer, and even to archival material from formaldehyde-fixed, paraffin-embedded blocks, reprocessed for electron microscopic examination. The successful restorative methods included pre-treatment of the sections with 6 M guanidine hydrochloride, or 1 M Tris/saline, each containing 100 mM dithiothreitol (a reducing agent) followed by alkylation with 220 mM iodoacetamide. The application of these techniques allowed reliable study of elastic tissue antibody distributions in archival tissues that could not be obtained again, as well as comparative studies with tissues processed many years previously.

  9. Postoperative morbidity and histopathologic characteristics of tonsillar tissue following coblation tonsillectomy in children: a prospective randomized single-blind study.

    PubMed

    Roje, Zeljka; Racić, Goran; Dogas, Zoran; Pisac, Valdi Pesutić; Timms, Michael

    2009-03-01

    The aim of this prospective randomized single blind study was to determine the depth of thermal damage to tonsillar tissue due to coblation, and to compare it with thermal damage to tonsillar tissue following conventional tonsillectomy; to correlate the depth of thermal damage to tonsillar tissue with the parameters of postoperative morbidity, to compare intraoperative blood loss, postoperative pain severity, time to resuming normal physical activity, and incidence of postoperative bleeding between two groups of tonsillectomized children aged up to 16 years. 72 children aged 3-16 years scheduled for tonsillectomy randomly assigned into two groups submitted either to conventional tonsillectomy with bipolar diathermy coagulation or to coblation tonsillectomy, with a 14-day follow up. Statistically significant differences were observed in the depth of thermal damage to tonsillar tissue (p < 0.001), intraoperative blood loss (p < 0.004), in postoperative pain severity (p < 0.05) and in time to resuming normal physical activity between the two groups (p < 0.001). There was no case of reactionary or secondary bleeding in either group. In this paper for the first time we have correlated postoperative morbidity and thermal tissue damage: less thermal damage is associated with less postoperative morbidity.

  10. Effective attenuation of atrazine-induced histopathological changes in testicular tissue by antioxidant N-phenyl-4-aryl-polyhydroquinolines.

    PubMed

    Chandak, Navneet; Bhardwaj, Jitender K; Zheleva-Dimitrova, Dimitrina; Kitanov, Gerassim; Sharma, Rajnesh K; Sharma, Pawan K; Saso, Luciano

    2015-01-01

    Some of the environmental toxicants acting as endocrine disruptors have been associated with health hazards in human and wildlife by modulating hormonal actions. Atrazine, a strong endocrine disruptor, induces detrimental effects on gonads in male and female, and causes impairment of fertility and developmental problems as well as sex alterations. Atrazine decreases the activities of antioxidant enzymes and thus responsible for oxidative stress. Natural antioxidants have shown ability to reduce/slow down the apoptotic effect of atrazine on testicular tissue. In the present study, some N-phenyl-4-aryl-polyhydroquinolines bearing phenolic or/and alkoxy group(s) (6a-6g) were synthesized and evaluated for antioxidant activity in four different assays. Three best compounds (6e-6g) were studied for their ameliorative effect on testicular tissue supplemented with atrazine in vitro.

  11. Histopathological changes of renal tissue following sodium fluoride administration in two consecutive generations of mice. Correlation with the urinary elimination of fluoride.

    PubMed

    Dimcevici Poesina, Nicoleta; Bălălău, Cristian; Nimigean, Vanda Roxana; Nimigean, Victor; Ion, Ion; Baconi, Daniela; Bârcă, Maria; Băran Poesina, Violeta

    2014-01-01

    The present study was designed to investigate the toxic effects (evaluated as histopathological changes) of sodium fluoride on the kidney in two consecutive generations of NMRI mice. An attempt to correlate the toxicity with the urinary elimination of fluoride has been made, as urinary fluoride excretion has been widely used as an indicator of fluoride intake and exposure. Six mixed (males and females) animal groups have been constituted by dividing the populations of mice derived from pregnant females (named "mothers" 0.5 mg sodium fluoride) treated with 0.5 mg sodium fluoride by daily gavage and pregnant females (named "mothers" 0.25 mg sodium fluoride) treated with 0.25 mg sodium fluoride by daily gavage; three types of sodium fluoride treatments were administrated: homeopathic, allopathic-homeopathic and allopathic. When the animals reached the adulthood, by randomization, they were selected in pairs for giving birth to the second generation of mice. No treatments were administrated to the second generation of mice; thus, the urinary elimination of fluoride in the second generation is attributed to exposure at sodium fluoride before birth. The administration of sodium fluoride to the first generation (F1) is realized until the mice reached the adulthood. For the first generation, the urine was collected at three times, every three weeks: at the age of four weeks, seven weeks and 11 weeks; single sampling urine, at the age of four weeks, has been conducted for the second generation. The urine samples have been analyzed using the ion selective electrode method for fluoride. For the histopathological examination, the animals were killed by cervical dislocation; the kidneys were collected in a 10% formalin solution. The preparation of samples for optical microscopy was realized with Hematoxylin-Eosin staining. The results indicate that the elimination of fluoride was similar (at the second evaluation, at 7-week-old of the first generation) for the both generations

  12. Histopathology of a mesoparasitic hatschekiid copepod in hospite: does Mihbaicola sakamakii (Copepoda: Siphonostomatoida: Hatschekiidae) fast within the host fish tissue?

    PubMed

    Hirose, Euichi; Uyeno, Daisuke

    2014-08-01

    Mihbaicola sakamakii is a mesoparasitic copepod that infests the branchiostegal membranes of groupers (Perciformes: Serranidae). In this study, we observed M. sakamakii within host tissue. Histologically, copepods were found enclosed inside a pouch composed of the thickened epidermis of the host, tightly encased on all sides by the host epidermal pouch wall. There were no host blood cells or other food resources in the pouch lumen. Since the host epidermis was intact and continuous, even in the vicinity of the oral region of the parasite, the copepod would not have access to the host blood in this state. However, the stomach (ampullary part of the mid gut) was filled with granular components, the majority of which were crystalloids that likely originated from fish erythrocyte hemoglobin. We supposed that the parasite drinks blood exuded from the lesion in the fish caused by copepod entry into the host tissue. Invasion of the parasite may elicit immune responses in the host, but there were no traces on the copepod of any cellular immune reactions, such as encapsulation. The array of minute protuberances on the copepod cuticle surface may be involved in avoidance of cell adhesion. After the lesion has healed, the copepod is enclosed in a tough epidermal pouch, in which it gradually digests the contents of its stomach and continues egg production. PMID:25088597

  13. Measurement of phthalates in small samples of mammalian tissue

    SciTech Connect

    Acott, P.D.; Murphy, M.G.; Ogborn, M.R.; Crocker, J.F.S.

    1987-03-01

    Di-(2-ethylhexyl)-phthalate (DEHP) is a phthalic acid ester that is used as a plasticizer in polyvinyl chloride products, many of which have widespread medical application. DEHP has been shown to be leached from products used for storage and delivery of blood transfusions during procedures such as plasmaphoresis, hemodialysis and open heart surgery. Results of studies in this laboratory have suggested that there is an association between the absorption and deposition of DEHP (and/or related chemicals) in the kidney and the acquired renal cystic disease (ACD) frequently seen in patients who have undergone prolonged dialysis treatment. In order to determine the relationship between the two, it has been necessary to establish a method for extracting and accurately quantitating minute amounts of these chemicals in small tissue samples. The authors have now established such a method using kidneys from normal rats and from a rat model for ACD.

  14. Artefacts in histopathology.

    PubMed

    Chatterjee, Shailja

    2014-09-01

    Histopathology is the science of slide analysis for the diagnostic and research purposes. However, sometimes the presence of certain artefacts in a microscopic section can result in misinterpretations leading to diagnostic pitfalls that can result in increased patient morbidity. This article reviews the common artefacts encountered during slide examination alongside the remedial measures which can be undertaken to differentiate between an artefact and tissue constituent.

  15. Immunohistochemical and histopathologic correlates of Alzheimer's disease-associated Alz-50 immunoreactivity quantified in homogenates of cerebral tissue.

    PubMed Central

    de la Monte, S. M.; Spratt, R. A.; Chong, J.; Ghanbari, H. A.; Wands, J. R.

    1992-01-01

    Alz-50 is a monoclonal antibody that immunoreacts with neurofibrillary tangles and neurites in brains with Alzheimer's disease (AD). In addition, the levels of Alz-50 immunoreactivity in brain, measured by either enzyme-linked immunosorbent assay or ALZ-enzyme-linked immunosorbant assay (EIA), are increased in AD relative to age-matched controls. The current study compares the distribution and extent of Alz-50 immunostaining with quantified levels of Alz-50 immunoreactivity measured in adjacent frozen blocks of tissue by ALZ-EIA. The brain tissue studied was obtained from individuals with AD, AD + Down's syndrome (AD + DN), Parkinson's disease with dementia (PD), or AD + PD, and from nondemented aged controls. In AD, AD + DN, and AD + PD, there were significantly higher densities of Alz-50-immunoreactive (AFI) neurons, more abundant diffuse AFI neurites, and higher ALZ-EIA values than in aged controls. In PD, the overall mean density of AFI neurons was significantly lower than in AD and AD + DN, but AFI neurites were as abundant as they were in brains with an AD diagnosis. However, PD was readily distinguished from AD and AD + DN by significantly lower mean ALZ-EIA values, and significantly lower densities of neurofilament-immuno-reactive AD lesions. Multiple-regression analysis demonstrated significant correlations between ALZ-EIA levels and the severity of AD lesions, and the density of AFI neurites, but not with the density of AFI neurons. Therefore, ALZ-EIA levels may represent only a portion of the Alz-50 immunoreactivity detectable by immunohistochemical staining. Images Figure 1 PMID:1466403

  16. Optimization of Evans blue quantitation in limited rat tissue samples

    NASA Astrophysics Data System (ADS)

    Wang, Hwai-Lee; Lai, Ted Weita

    2014-10-01

    Evans blue dye (EBD) is an inert tracer that measures plasma volume in human subjects and vascular permeability in animal models. Quantitation of EBD can be difficult when dye concentration in the sample is limited, such as when extravasated dye is measured in the blood-brain barrier (BBB) intact brain. The procedure described here used a very small volume (30 µl) per sample replicate, which enabled high-throughput measurements of the EBD concentration based on a standard 96-well plate reader. First, ethanol ensured a consistent optic path length in each well and substantially enhanced the sensitivity of EBD fluorescence spectroscopy. Second, trichloroacetic acid (TCA) removed false-positive EBD measurements as a result of biological solutes and partially extracted EBD into the supernatant. Moreover, a 1:2 volume ratio of 50% TCA ([TCA final] = 33.3%) optimally extracted EBD from the rat plasma protein-EBD complex in vitro and in vivo, and 1:2 and 1:3 weight-volume ratios of 50% TCA optimally extracted extravasated EBD from the rat brain and liver, respectively, in vivo. This procedure is particularly useful in the detection of EBD extravasation into the BBB-intact brain, but it can also be applied to detect dye extravasation into tissues where vascular permeability is less limiting.

  17. Incidence of mycobacterial infections in cats in Great Britain: estimate from feline tissue samples submitted to diagnostic laboratories.

    PubMed

    Gunn-Moore, D A; Gaunt, C; Shaw, D J

    2013-08-01

    The aim of this study was to estimate the incidence of mycobacterial infections in cats in Great Britain (GB). This was performed using the proxy measure of feline tissue samples submitted to diagnostic laboratories in GB that were found to have histopathological changes typical of mycobacterial infection ('MYC'). Sixteen primary diagnostic laboratories were asked for information on the number of feline samples submitted in 2009, the number with MYC, the number undergoing Ziehl-Neelsen (ZN) staining and, for comparison, the number diagnosed with lymphoma. Eight laboratories provided full data for the whole year: 11,782 samples; lymphoma 3.2% (mean, 95% CI: 2.89, 3.5), MYC 1.16% (0.98; 1.37) and ZN-positive 0.31% (0.22; 0.43). Data on 1569 samples from seven laboratories that provided partial data on samples for the whole year revealed similar results, although all changes were more frequent: lymphoma 5.42% (4.35; 6.66), MYC 2.36% (1.66; 3.23) and ZN-positive 0.77% (0.40; 1.33). One laboratory only provided data for part of the year (4.5 months), reporting all three types of histopathology less frequently: 18,232 samples; lymphoma 0.2% (0.18; 0.32), MYC 0.07% (0.04; 0.12) and ZN-positive 0.05% (0.02; 0.09). The reasons for low reporting rates in this high-throughput laboratory are unclear. In total, 187 samples were reported as having MYC. Five Reference laboratories were also contacted, reporting 174 feline tissue submissions in 2009, with mycobacteria being cultured from 90. The study shows that MYC are frequently reported in tissue samples from cats in GB, being reported in ~1% of samples, with confirmation as ZN-positive in ~0.3%. Lymphoma is recognized as a common disease in cats, being seen in ~3% of samples in this study. When compared against MYC, lymphoma was reported only twice as frequently. This confirms that far from being rare, clinically significant mycobacterial infections occur commonly in cats in GB.

  18. Histopathological Image Analysis: A Review

    PubMed Central

    Gurcan, Metin N.; Boucheron, Laura; Can, Ali; Madabhushi, Anant; Rajpoot, Nasir; Yener, Bulent

    2010-01-01

    Over the past decade, dramatic increases in computational power and improvement in image analysis algorithms have allowed the development of powerful computer-assisted analytical approaches to radiological data. With the recent advent of whole slide digital scanners, tissue histopathology slides can now be digitized and stored in digital image form. Consequently, digitized tissue histopathology has now become amenable to the application of computerized image analysis and machine learning techniques. Analogous to the role of computer-assisted diagnosis (CAD) algorithms in medical imaging to complement the opinion of a radiologist, CAD algorithms have begun to be developed for disease detection, diagnosis, and prognosis prediction to complement to the opinion of the pathologist. In this paper, we review the recent state of the art CAD technology for digitized histopathology. This paper also briefly describes the development and application of novel image analysis technology for a few specific histopathology related problems being pursued in the United States and Europe. PMID:20671804

  19. Clinical and Histopathological Investigation of Seborrheic Keratosis

    PubMed Central

    Roh, Nam Kyung; Hahn, Hyung Jin; Lee, Yang Won; Choe, Yong Beom

    2016-01-01

    Background Seborrheic keratosis (SK) is one of the most common epidermal tumors of the skin. However, only a few large-scale clinicohistopathological investigations have been conducted on SK or on the possible correlation between histopathological SK subtype and location. Objective The aim of this study was to analyze the clinical and histopathological features of a relatively large number of cases of diagnosed SK. Methods Two hundred and seventy-one pathology slides of skin tissue from patients with clinically diagnosed SK and 206 cases of biopsy-proven SK were analyzed. The biopsy-proven cases of SK were assessed for histopathological subclassification. The demographic, clinical, and histopathological data of the patients were collected for analysis of associated factors. Results The most frequent histopathological subtype was the acanthotic type, followed by mixed, hyperkeratotic, melanoacanthoma, clonal, irritated, and adenoid types; an unexpectedly high percentage (9.2%) of the melanoacanthoma variant was observed. The adenoid type was more common in sun-exposed sites than in sun-protected sites (p=0.028). Premalignant and malignant entities together represented almost one-quarter (24.2%) of the clinicopathological mismatch cases (i.e., mismatch between the clinical and histopathological diagnoses). Regarding the location of SK development, the frequency of mismatch for the sun-exposed areas was significantly higher than that for sun-protected areas (p=0.043). Conclusion The adenoid type was more common in sun-exposed sites. Biopsy sampling should be performed for lesions situated in sun-exposed areas to exclude other premalignant or malignant diseases. PMID:27081260

  20. Long-term room temperature preservation of corpse soft tissue: an approach for tissue sample storage

    PubMed Central

    2011-01-01

    Background Disaster victim identification (DVI) represents one of the most difficult challenges in forensic sciences, and subsequent DNA typing is essential. Collected samples for DNA-based human identification are usually stored at low temperature to halt the degradation processes of human remains. We have developed a simple and reliable procedure for soft tissue storage and preservation for DNA extraction. It ensures high quality DNA suitable for PCR-based DNA typing after at least 1 year of room temperature storage. Methods Fragments of human psoas muscle were exposed to three different environmental conditions for diverse time periods at room temperature. Storage conditions included: (a) a preserving medium consisting of solid sodium chloride (salt), (b) no additional substances and (c) garden soil. DNA was extracted with proteinase K/SDS followed by organic solvent treatment and concentration by centrifugal filter devices. Quantification was carried out by real-time PCR using commercial kits. Short tandem repeat (STR) typing profiles were analysed with 'expert software'. Results DNA quantities recovered from samples stored in salt were similar up to the complete storage time and underscored the effectiveness of the preservation method. It was possible to reliably and accurately type different genetic systems including autosomal STRs and mitochondrial and Y-chromosome haplogroups. Autosomal STR typing quality was evaluated by expert software, denoting high quality profiles from DNA samples obtained from corpse tissue stored in salt for up to 365 days. Conclusions The procedure proposed herein is a cost efficient alternative for storage of human remains in challenging environmental areas, such as mass disaster locations, mass graves and exhumations. This technique should be considered as an additional method for sample storage when preservation of DNA integrity is required for PCR-based DNA typing. PMID:21846338

  1. Validation of a postfixation tissue storage and transport medium to preserve histopathology and molecular pathology analyses (total and phosphoactivated proteins, and FISH).

    PubMed

    Stumm, Michael M; Walker, Maja R; Stork, Caroline; Hanoteau, Noelle; Wagner, Urs; O'Reilly, Terence M

    2012-03-01

    Tumor biomarker studies are integral to oncology clinical trials but may yield artifactual results owing to variation in sample procurement and processing. Ethanol, 70% vol/vol, was validated as a sample transport medium using markers of the PI3K/Akt/mTOR pathway. BT474 tumor xenografts were excised and slices were immediately placed into formaldehyde and fixed for 24 hours. Fixed tissue slices were immediately processed into paraffin or transferred to 70% vol/vol ethanol and stored at room temperature for 1, 2, and 4 weeks before further processing. Freshly cut tissue sections were evaluated for pAKT(S473), HER2, pHER-2(Y1248), pS6(S235/236), and pS6(S240/244), Ki-67, and HER2 by fluorescence in situ hybridization and stained with H&E and Masson trichrome. No significant changes were observed when comparing samples stored in 70% ethanol for up to 4 weeks with immediately processed tissue. Ethanol, 70% vol/vol, provides a safe storage medium for formaldehyde-fixed tumor tissue, facilitating sample transport during multicenter clinical trials.

  2. Histopathology and bioaccumulation of heavy metals (Cr, Ni and Pb) in fish (Channa striatus and Heteropneustes fossilis) tissue: a study for toxicity and ecological impacts.

    PubMed

    Fatima, Mahino; Usmani, Nazura

    2013-05-01

    Abstract: The water samples were collected from the 22 km segment III of Yamuna River from Okhla barrage. This segment receives water from 17 sewage drains of Delhi, Western Yamuna Canal (WYC), upper Ganga canal via Najafgarh drain and Hindon cut canal. Hence, the water samples collected were used to determine the presence of Chromium, Nickel and Lead through Atomic Absorption Spectrophotometry. The concentration of these heavy metals were much above the maximum permissible limits set by WHO. This was bound to have its influence on the riverine flora and fauna. To evaluate this, two popularly consumed fish species such as Channa striatus and Heteropneustes fossilis were caught and the bioaccumulation of these heavy metals were estimated in different organs (liver, kidney, gill and muscle). It was found that Cr accumulated the most in these organs (gill being most influenced) in both the species. The accumulation of all these heavy metals were above MPL set by World Health Organisation (WHO) and Food and Drug Administration (FDA). Histopathology was also conducted where heavy damages were observed in both liver and kidney of both the species. PMID:24498804

  3. Histopathology and bioaccumulation of heavy metals (Cr, Ni and Pb) in fish (Channa striatus and Heteropneustes fossilis) tissue: a study for toxicity and ecological impacts.

    PubMed

    Fatima, Mahino; Usmani, Nazura

    2013-05-01

    Abstract: The water samples were collected from the 22 km segment III of Yamuna River from Okhla barrage. This segment receives water from 17 sewage drains of Delhi, Western Yamuna Canal (WYC), upper Ganga canal via Najafgarh drain and Hindon cut canal. Hence, the water samples collected were used to determine the presence of Chromium, Nickel and Lead through Atomic Absorption Spectrophotometry. The concentration of these heavy metals were much above the maximum permissible limits set by WHO. This was bound to have its influence on the riverine flora and fauna. To evaluate this, two popularly consumed fish species such as Channa striatus and Heteropneustes fossilis were caught and the bioaccumulation of these heavy metals were estimated in different organs (liver, kidney, gill and muscle). It was found that Cr accumulated the most in these organs (gill being most influenced) in both the species. The accumulation of all these heavy metals were above MPL set by World Health Organisation (WHO) and Food and Drug Administration (FDA). Histopathology was also conducted where heavy damages were observed in both liver and kidney of both the species.

  4. Evidence of presence of Mycobacterium tuberculosis in bovine tissue samples by multiplex PCR: possible relevance to reverse zoonosis.

    PubMed

    Mittal, M; Chakravarti, S; Sharma, V; Sanjeeth, B S; Churamani, C P; Kanwar, N S

    2014-04-01

    Bovine tuberculosis, caused by Mycobacterium bovis, remains one of the most important zoonotic health concerns worldwide. The transmission of Mycobacterium tuberculosis from humans to animals also occurs especially in countries where there is close interaction of humans with the animals. In the present study, thirty bovine lung tissue autopsy samples from an organized dairy farm located in North India were screened for the presence of Mycobacterium tuberculosis complex by smear microscopy, histopathological findings and PCR. Differential diagnosis of M. tuberculosis and M. bovis was made based on the deletion of mce-3 operon in M. bovis. The present study found eight of these samples positive for M. tuberculosis by multiplex PCR. Sequencing was performed on two PCR-positive representative samples and on annotation, and BLAST analysis confirmed the presence of gene fragment specific to Mycobacterium tuberculosis. The presence of M. tuberculosis in all the positive samples raises the possibility of human-to-cattle transmission and possible adaptation of this organism in bovine tissues. This study accentuates the importance of screening and differential diagnosis of Mycobacterium tuberculosis complex in humans and livestock for adopting effective TB control and eradication programmes.

  5. Simulating an investigative study of clinical cancer samples: use of tissue slides and PCR-based promoter-hypermethylation analysis.

    PubMed

    Chew, Yuan Yuan; Chin, Cheen Fei; Yeong, Foong May

    2015-01-01

    Topics on the molecular basis underlying cancer are quite popular among students. Also, excellent textbooks abound that provide interesting materials for discussion during lectures and tutorials about major events leading to cancer formation and progression. However, much less is available for students to conduct experiments for the analysis of cancer samples in undergraduate modules where there is a limited time-frame. Given the difficulty of working with cancer samples and the scarcity of good samples even in the clinical laboratories, it is impossible to run large-class practicals using patients' samples. Here, we describe the use of tissue slides in combination with polymerase-chain reaction (PCR) as a means of simulating an investigative approach to supplement students' learning of clinical research. By using tissue slides for histo-pathological examinations and specific budding yeast genomic DNA and primers adapted to demonstrate methylation-specific PCR, we designed an inquiry-based lab session to simulate the clinical investigation of a cohort of biopsies that students could analyze in a one-session practical.

  6. Variation in glycogen concentrations within mantle and foot tissue in Amblema plicata plicata: Implications for tissue biopsy sampling

    USGS Publications Warehouse

    Naimo, T.J.; Monroe, E.M.

    1999-01-01

    With the development of techniques to non-lethally biopsy tissue from unionids, a new method is available to measure changes in biochemical, contaminant, and genetic constituents in this imperiled faunal group. However, before its widespread application, information on the variability of biochemical components within and among tissues needs to be evaluated. We measured glycogen concentrations in foot and mantle tissue in Amblema plicata plicata (Say, 1817) to determine if glycogen was evenly distributed within and between tissues and to determine which tissue might be more responsive to the stress associated with relocating mussels. Glycogen was measured in two groups of mussels: those sampled from their native environment (undisturbed mussels) and quickly frozen for analysis and those relocated into an artificial pond (relocated mussels) for 24 months before analysis. In both undisturbed and relocated mussels, glycogen concentrations were evenly distributed within foot, but not within mantle tissue. In mantle tissue, concentrations of glycogen varied about 2-fold among sections. In addition, glycogen varied significantly between tissues in undisturbed mussels, but not in relocated mussels. Twenty-four months after relocation, glycogen concentrations had declined by 80% in mantle tissue and by 56% in foot tissue relative to the undisturbed mussels. These data indicate that representative biopsy samples can be obtained from foot tissue, but not mantle tissue. We hypothesize that mantle tissue could be more responsive to the stress of relocation due to its high metabolic activity associated with shell formation.

  7. Automated Classification of Glandular Tissue by Statistical Proximity Sampling

    PubMed Central

    Azar, Jimmy C.; Simonsson, Martin

    2015-01-01

    Due to the complexity of biological tissue and variations in staining procedures, features that are based on the explicit extraction of properties from subglandular structures in tissue images may have difficulty generalizing well over an unrestricted set of images and staining variations. We circumvent this problem by an implicit representation that is both robust and highly descriptive, especially when combined with a multiple instance learning approach to image classification. The new feature method is able to describe tissue architecture based on glandular structure. It is based on statistically representing the relative distribution of tissue components around lumen regions, while preserving spatial and quantitative information, as a basis for diagnosing and analyzing different areas within an image. We demonstrate the efficacy of the method in extracting discriminative features for obtaining high classification rates for tubular formation in both healthy and cancerous tissue, which is an important component in Gleason and tubule-based Elston grading. The proposed method may be used for glandular classification, also in other tissue types, in addition to general applicability as a region-based feature descriptor in image analysis where the image represents a bag with a certain label (or grade) and the region-based feature vectors represent instances. PMID:25685143

  8. Studying Genes in Tissue Samples From Younger and Adolescent Patients With Soft Tissue Sarcomas

    ClinicalTrials.gov

    2016-05-13

    Childhood Alveolar Soft-part Sarcoma; Childhood Angiosarcoma; Childhood Desmoplastic Small Round Cell Tumor; Childhood Epithelioid Sarcoma; Childhood Fibrosarcoma; Childhood Leiomyosarcoma; Childhood Liposarcoma; Childhood Malignant Mesenchymoma; Childhood Neurofibrosarcoma; Childhood Synovial Sarcoma; Chordoma; Desmoid Tumor; Metastatic Childhood Soft Tissue Sarcoma; Nonmetastatic Childhood Soft Tissue Sarcoma; Recurrent Childhood Soft Tissue Sarcoma

  9. Robust and artifact-free mounting of tissue samples for atomic force microscopy.

    PubMed

    Morgan, Joshua T; Raghunathan, Vijay Krishna; Thomasy, Sara M; Murphy, Christopher J; Russell, Paul

    2014-01-01

    Immobilization of tissue-samples for atomic for microscopy (AFM) is typically done using either semi-dry tissue or by gluing the tissue sample down, both of which can introduce artifacts. Here, we describe the design of a Soft- Clamping Immobilizing Retainer of Tissue (SCIRT) for consistent and nondestructive immobilization of tissues for AFM analysis. We compare the performance of our SCIRT method with glue-immobilization for two difficult to handle tissue types: human trabecular meshwork (HTM) and rabbit cornea (RC). Our results demonstrate that the SCIRT method has several advantages, including: (i) allowing for small sample sizes, (ii) enabling continuous hydration, (iii) eliminating contact with glue or associated solvents, (iv) permitting sample recovery following measurement, and (v) ease of use. In conclusion, the SCIRT method is a simple and effective means of immobilizing soft, hydrated tissue samples consistently and without artifacts.

  10. Robust and artifact-free mounting of tissue samples for atomic force microscopy

    PubMed Central

    Morgan, Joshua T.; Raghunathan, Vijay Krishna; Thomasy, Sara M.; Murphy, Christopher J.; Russell, Paul

    2015-01-01

    Immobilization of tissue-samples for atomic for microscopy (AFM) is typically done using either semi-dry tissue or by gluing the tissue sample down, both of which can introduce artifacts. Here, we describe the design of a Soft-Clamping Immobilizing Retainer of Tissue (SCIRT) for consistent and nondestructive immobilization of tissues for AFM analysis. We compare the performance of our SCIRT method with glue-immobilization for two difficult to handle tissue types: human trabecular meshwork (HTM) and rabbit cornea (RC). Our results demonstrate that the SCIRT method has several advantages, including: (i) allowing for small sample sizes, (ii) enabling continuous hydration, (iii) eliminating contact with glue or associated solvents, (iv) permitting sample recovery following measurement, and (v) ease of use. In conclusion, the SCIRT method is a simple and effective means of immobilizing soft, hydrated tissue samples consistently and without artifacts. PMID:24447138

  11. Histopathological characteristics of human non-tumor thyroid tissues in a long-term model of adenomatous goiter xenografts in the NOD/Shi-scid, IL-2Rγ(null) mouse.

    PubMed

    Fujii, Etsuko; Kato, Atsuhiko; Chen, Yu Jau; Matsubara, Koichi; Ohnishi, Yasuyuki; Suzuki, Masami

    2014-07-01

    There is a growing need for modeling the human thyroid to link data obtained from animals to humans because of its sensitivity to radiation exposure and endocrine disruption chemicals. In a scid mouse model produced by transplanting human thyroid tissues, leakiness and thymic lymphoma that occurs spontaneously in the scid mouse can complicate the interpretation of experimental results. Considering that the NOD.Cg-Prkdc(scid)Il2rg(tm1Sug)/Jic mouse (NOD/Shi-scid, IL-2Rγ(null) or NOG mouse) may be a better host because this strain has low incidence of leakiness and thymic lymphoma, we have evaluated the potential of a model that allows long-term observation of non-tumor human thyroid tissues in this mouse. We transplanted tissues of human adenomatous goiter into NOG mice and examined the tissues histopathologically. The morphology of human adenomatous goiter tissues was maintained from 24 to 44 weeks after transplantation in NOG mice with no noted differences between donor-matched tissues or the weeks after transplantation. The tissues expressed thyroglobulin protein and mRNA as well as thyroperoxidase. Endothelial cells originating from human were found in the transplanted tissues and were thought to be a characteristic of this model. The intactness of the tissues before transplantation was found to affect the rate of tissue engraftment. From the present results we have concluded that transplanted thyroid tissues in NOG mice maintain the histopathological characteristics of their origin for long terms. Therefore this model was thought feasible for toxicity evaluation.

  12. Functional relationships among selenium concentrations in the diet, target tissues, and nondestructive tissue samples of two species of snakes.

    PubMed

    Hopkins, William A; Snodgrass, Joel W; Baionno, Jennifer A; Roe, John H; Staub, Brandon P; Jackson, Brian P

    2005-02-01

    Nondestructive sampling methods, such as removal of feathers for contaminant analysis, are desirable in ecological monitoring programs that seek to minimize the impacts of harvesting organisms. Although many reptiles are declining worldwide, nondestructive sampling techniques seldom have been employed for assessing contaminant exposure in these organisms. In this study, we examined the utility of nondestructive tissue sampling for assessing Se exposure in reptiles. We describe the functional relationships among dietary Se concentrations, target tissue Se concentrations, and Se concentrations in nondestructive tissue samples (blood and tail tissue biopsy) in two species of snakes that had been exposed to Se under very different experimental protocols. Using nonlinear regression, we found strong positive correlations (r2 > 0.92) in all comparisons among Se concentrations in nondestructive tissues, diet, and target tissues. Moreover, equations describing these relationships can be used to estimate concentrations of Se in diet and target organs, from known concentrations of Se in nondestructive tissue samples. Although the current paucity of toxicity data on reptiles precludes tests of our models, we demonstrate how the equations describing these relationships might be used to make predictions about Se accumulation in target organs for risk assessment. Future studies on reptiles that examine these relationships under different Se exposure conditions, and those that document physiological responses of reptiles to various concentrations of Se, will help to refine our models and test their efficacy for predicting health risk.

  13. Out-of-Sample Extrapolation utilizing Semi-Supervised Manifold Learning (OSE-SSL): Content Based Image Retrieval for Histopathology Images

    PubMed Central

    Sparks, Rachel; Madabhushi, Anant

    2016-01-01

    Content-based image retrieval (CBIR) retrieves database images most similar to the query image by (1) extracting quantitative image descriptors and (2) calculating similarity between database and query image descriptors. Recently, manifold learning (ML) has been used to perform CBIR in a low dimensional representation of the high dimensional image descriptor space to avoid the curse of dimensionality. ML schemes are computationally expensive, requiring an eigenvalue decomposition (EVD) for every new query image to learn its low dimensional representation. We present out-of-sample extrapolation utilizing semi-supervised ML (OSE-SSL) to learn the low dimensional representation without recomputing the EVD for each query image. OSE-SSL incorporates semantic information, partial class label, into a ML scheme such that the low dimensional representation co-localizes semantically similar images. In the context of prostate histopathology, gland morphology is an integral component of the Gleason score which enables discrimination between prostate cancer aggressiveness. Images are represented by shape features extracted from the prostate gland. CBIR with OSE-SSL for prostate histology obtained from 58 patient studies, yielded an area under the precision recall curve (AUPRC) of 0.53 ± 0.03 comparatively a CBIR with Principal Component Analysis (PCA) to learn a low dimensional space yielded an AUPRC of 0.44 ± 0.01. PMID:27264985

  14. IHC Profiler: An Open Source Plugin for the Quantitative Evaluation and Automated Scoring of Immunohistochemistry Images of Human Tissue Samples

    PubMed Central

    Malhotra, Renu; De, Abhijit

    2014-01-01

    In anatomic pathology, immunohistochemistry (IHC) serves as a diagnostic and prognostic method for identification of disease markers in tissue samples that directly influences classification and grading the disease, influencing patient management. However, till today over most of the world, pathological analysis of tissue samples remained a time-consuming and subjective procedure, wherein the intensity of antibody staining is manually judged and thus scoring decision is directly influenced by visual bias. This instigated us to design a simple method of automated digital IHC image analysis algorithm for an unbiased, quantitative assessment of antibody staining intensity in tissue sections. As a first step, we adopted the spectral deconvolution method of DAB/hematoxylin color spectra by using optimized optical density vectors of the color deconvolution plugin for proper separation of the DAB color spectra. Then the DAB stained image is displayed in a new window wherein it undergoes pixel-by-pixel analysis, and displays the full profile along with its scoring decision. Based on the mathematical formula conceptualized, the algorithm is thoroughly tested by analyzing scores assigned to thousands (n = 1703) of DAB stained IHC images including sample images taken from human protein atlas web resource. The IHC Profiler plugin developed is compatible with the open resource digital image analysis software, ImageJ, which creates a pixel-by-pixel analysis profile of a digital IHC image and further assigns a score in a four tier system. A comparison study between manual pathological analysis and IHC Profiler resolved in a match of 88.6% (P<0.0001, CI = 95%). This new tool developed for clinical histopathological sample analysis can be adopted globally for scoring most protein targets where the marker protein expression is of cytoplasmic and/or nuclear type. We foresee that this method will minimize the problem of inter-observer variations across labs and further help in

  15. Impact of metals on histopathology and expression of HSP 70 in different tissues of Milk fish (Chanos chanos) of Kaattuppalli Island, South East Coast, India.

    PubMed

    Rajeshkumar, Sivakumar; Munuswamy, Natesan

    2011-04-01

    Histological and Immunohistochemical studies were carried out to document the possible impact of heavy metal contamination in different tissues of Chanos chanos. Heavy metals such as Cu, Pb, Zn, Cd, Mn and Fe were predominant in water, sediment and biota of Kaattuppalli Island and varied significantly between two different sites. Histological changes such as swelling of muscle fiber and break down of muscle bundles were noted in the muscle. Similarly gill filament cell proliferation, increase in intercellular spaces and primary and secondary lamellar epithelium were evident in gills. The hepatocytes showed damage of central vein and rupture of irregular hepatic plate with more number of vacuoles in the fish collected from polluted site compared to that from the less polluted site. The impact of pollution was also assessed in different tissues by immunohistochemistry using primary antibody (mouse monoclonal HSP70 antibody 1:2000) and secondary antibody (HRP conjugated antibody) for expression of stress protein. Immunostaining analysis showed expression of HSP70 with high intensity in the tissues of fish collected from polluted site compared to less polluted sites. Further, HSP70 positive cells were analyzed from six locations per fish tissue section. One-way analysis of variance (ANOVA) followed by least significant difference (LSD) was used to check if the expression was significant. Results indicate that the values are statistically significant at the two different sampling sites (P<0.05).

  16. Ethical use of tissue samples in genetic research.

    PubMed

    Azarow, Kenneth S; Olmstead, Francis L; Hume, Roderick F; Myers, Jerome; Calhoun, Bryon C; Martin, Laura S

    2003-06-01

    Many centrally based cancer protocols have begun to address the ethical issues concerning tissue banking for genetic research. A multidisciplinary subcommittee of the Madigan Army Medical Center Institutional Review Board was established to determine the scope of the problem and offer a concise, user-friendly policy with guidelines on how to control and monitor the use of stored tissue for future genetic and molecular research. Our institution participates in 69 Southern Oncology Group or National Surgical Adjuvant Breast and Bowel Project protocols and 47 Children's Oncology Group protocols. Of these protocols, 22 of 69 and 36 of 47, respectively, asked for tissue to be stored for future biologic study. Only 4 of 69 and 3 of 47, respectively, deal with specific consent for future genetic/biologic research. The multidisciplinary committee developed a policy that dealt with the following areas: exempt status, waived consent, informed consent, deceased status, family studies, and information flow. An algorithm was created to establish a system of checks and balances concerning privacy, protection and an appeals process.

  17. Final LDRD report : development of sample preparation methods for ChIPMA-based imaging mass spectrometry of tissue samples.

    SciTech Connect

    Maharrey, Sean P.; Highley, Aaron M.; Behrens, Richard, Jr.; Wiese-Smith, Deneille

    2007-12-01

    The objective of this short-term LDRD project was to acquire the tools needed to use our chemical imaging precision mass analyzer (ChIPMA) instrument to analyze tissue samples. This effort was an outgrowth of discussions with oncologists on the need to find the cellular origin of signals in mass spectra of serum samples, which provide biomarkers for ovarian cancer. The ultimate goal would be to collect chemical images of biopsy samples allowing the chemical images of diseased and nondiseased sections of a sample to be compared. The equipment needed to prepare tissue samples have been acquired and built. This equipment includes an cyro-ultramicrotome for preparing thin sections of samples and a coating unit. The coating unit uses an electrospray system to deposit small droplets of a UV-photo absorbing compound on the surface of the tissue samples. Both units are operational. The tissue sample must be coated with the organic compound to enable matrix assisted laser desorption/ionization (MALDI) and matrix enhanced secondary ion mass spectrometry (ME-SIMS) measurements with the ChIPMA instrument Initial plans to test the sample preparation using human tissue samples required development of administrative procedures beyond the scope of this LDRD. Hence, it was decided to make two types of measurements: (1) Testing the spatial resolution of ME-SIMS by preparing a substrate coated with a mixture of an organic matrix and a bio standard and etching a defined pattern in the coating using a liquid metal ion beam, and (2) preparing and imaging C. elegans worms. Difficulties arose in sectioning the C. elegans for analysis and funds and time to overcome these difficulties were not available in this project. The facilities are now available for preparing biological samples for analysis with the ChIPMA instrument. Some further investment of time and resources in sample preparation should make this a useful tool for chemical imaging applications.

  18. Micro-organisms isolated from cadaveric samples of allograft musculoskeletal tissue.

    PubMed

    Varettas, Kerry

    2013-12-01

    Allograft musculoskeletal tissue is commonly used in orthopaedic surgical procedures. Cadaveric donors of musculoskeletal tissue supply multiple allografts such as tendons, ligaments and bone. The microbiology laboratory of the South Eastern Area Laboratory Services (SEALS, Australia) has cultured cadaveric allograft musculoskeletal tissue samples for bacterial and fungal isolates since 2006. This study will retrospectively review the micro-organisms isolated over a 6-year period, 2006-2011. Swab and tissue samples were received for bioburden testing and were inoculated onto agar and/or broth culture media. Growth was obtained from 25.1 % of cadaveric allograft musculoskeletal tissue samples received. The predominant organisms isolated were coagulase-negative staphylococci and coliforms, with the heaviest bioburden recovered from the hemipelvis. The rate of bacterial and fungal isolates from cadaveric allograft musculoskeletal tissue samples is higher than that from living donors. The type of organism isolated may influence the suitability of the allograft for transplant.

  19. Elemental distribution and sample integrity comparison of freeze-dried and frozen-hydrated biological tissue samples with nuclear microprobe

    NASA Astrophysics Data System (ADS)

    Vavpetič, P.; Vogel-Mikuš, K.; Jeromel, L.; Ogrinc Potočnik, N.; Pongrac, P.; Drobne, D.; Pipan Tkalec, Ž.; Novak, S.; Kos, M.; Koren, Š.; Regvar, M.; Pelicon, P.

    2015-04-01

    The analysis of biological samples in frozen-hydrated state with micro-PIXE technique at Jožef Stefan Institute (JSI) nuclear microprobe has matured to a point that enables us to measure and examine frozen tissue samples routinely as a standard research method. Cryotome-cut slice of frozen-hydrated biological sample is mounted between two thin foils and positioned on the sample holder. The temperature of the cold stage in the measuring chamber is kept below 130 K throughout the insertion of the samples and the proton beam exposure. Matrix composition of frozen-hydrated tissue is consisted mostly of ice. Sample deterioration during proton beam exposure is monitored during the experiment, as both Elastic Backscattering Spectrometry (EBS) and Scanning Transmission Ion Microscopy (STIM) in on-off axis geometry are recorded together with the events in two PIXE detectors and backscattered ions from the chopper in a single list-mode file. The aim of this experiment was to determine differences and similarities between two kinds of biological sample preparation techniques for micro-PIXE analysis, namely freeze-drying and frozen-hydrated sample preparation in order to evaluate the improvements in the elemental localisation of the latter technique if any. In the presented work, a standard micro-PIXE configuration for tissue mapping at JSI was used with five detection systems operating in parallel, with proton beam cross section of 1.0 × 1.0 μm2 and a beam current of 100 pA. The comparison of the resulting elemental distributions measured at the biological tissue prepared in the frozen-hydrated and in the freeze-dried state revealed differences in elemental distribution of particular elements at the cellular level due to the morphology alteration in particular tissue compartments induced either by water removal in the lyophilisation process or by unsatisfactory preparation of samples for cutting and mounting during the shock-freezing phase of sample preparation.

  20. Monitoring the marine environment using marine mammal tissue samples

    SciTech Connect

    Jones, P.D.; Hannah, D.J.; Day, P.J.

    1995-12-31

    Marine environments, both inshore and open ocean, receive numerous inputs of anthropogenic chemicals. Cetaceans provide a valuable resource for monitoring the low level contamination of marine environments with persistent organic contaminants. Comparative studies using inshore and offshore southern ocean cetaceans have revealed significant differences in the types of contamination in these two environments. The polychlorinated biphenyls (PCBs) deposited in the southern oceans are characterized by an abundance of lower chlorinated congeners. Polychlorinated dibenzo-p-dioxins and dibenzofurans (PCDD/F) are not present at significant concentrations in cetaceans from the open southern ocean. In contrast significant concentrations of PCDD/F congeners are detected in the blubber of the inshore living Hector`s dolphin. This species lives close to the shore and has a very small home range (approximately 30 km) for a cetacean. Analysis of tissue PCDD/F and PCB profiles from different populations and their food sources will be presented. The data are being used to determine if there are local variations in the contamination of the New Zealand inshore marine environment.

  1. Tissue Microarray Technology for Molecular Applications: Investigation of Cross-Contamination between Tissue Samples Obtained from the Same Punching Device

    PubMed Central

    Vassella, Erik; Galván, José A.; Zlobec, Inti

    2015-01-01

    Background: Tissue microarray (TMA) technology allows rapid visualization of molecular markers by immunohistochemistry and in situ hybridization. In addition, TMA instrumentation has the potential to assist in other applications: punches taken from donor blocks can be placed directly into tubes and used for nucleic acid analysis by PCR approaches. However, the question of possible cross-contamination between samples punched with the same device has frequently been raised but never addressed. Methods: Two experiments were performed. (1) A block from mycobacterium tuberculosis (TB) positivetissue and a second from an uninfected patient were aligned side-by-side in an automated tissue microarrayer. Four 0.6 mm punches were cored from each sample and placed inside their corresponding tube. Between coring of each donor block, a mechanical cleaning step was performed by insertion of the puncher into a paraffin block. This sequence of coring and cleaning was repeated three times, alternating between positive and negative blocks. A fragment from the 6110 insertion sequence specific for mycobacterium tuberculosis was analyzed; (2) Four 0.6 mm punches were cored from three KRAS mutated colorectal cancer blocks, alternating with three different wild-type tissues using the same TMA instrument (sequence of coring: G12D, WT, G12V, WT, G13D and WT). Mechanical cleaning of the device between each donor block was made. Mutation analysis by pyrosequencing was carried out. This sequence of coring was repeated manually without any cleaning step between blocks. Results/Discussion: In both analyses, all alternating samples showed the expected result (samples 1, 3 and 5: positive or mutated, samples 2, 4 and 6: negative or wild-type). Similar results were obtained without cleaning step. These findings suggest that no cross-contamination of tissue samples occurs when donor blocks are punched using the same device, however a cleaning step is nonetheless recommended. Our result supports

  2. Experimental implementation of coded aperture coherent scatter spectral imaging of cancerous and healthy breast tissue samples

    NASA Astrophysics Data System (ADS)

    Lakshmanan, Manu N.; Greenberg, Joel A.; Samei, Ehsan; Kapadia, Anuj J.

    2015-03-01

    A fast and accurate scatter imaging technique to differentiate cancerous and healthy breast tissue is introduced in this work. Such a technique would have wide-ranging clinical applications from intra-operative margin assessment to breast cancer screening. Coherent Scatter Computed Tomography (CSCT) has been shown to differentiate cancerous from healthy tissue, but the need to raster scan a pencil beam at a series of angles and slices in order to reconstruct 3D images makes it prohibitively time consuming. In this work we apply the coded aperture coherent scatter spectral imaging technique to reconstruct 3D images of breast tissue samples from experimental data taken without the rotation usually required in CSCT. We present our experimental implementation of coded aperture scatter imaging, the reconstructed images of the breast tissue samples and segmentations of the 3D images in order to identify the cancerous and healthy tissue inside of the samples. We find that coded aperture scatter imaging is able to reconstruct images of the samples and identify the distribution of cancerous and healthy tissues (i.e., fibroglandular, adipose, or a mix of the two) inside of them. Coded aperture scatter imaging has the potential to provide scatter images that automatically differentiate cancerous and healthy tissue inside of ex vivo samples within a time on the order of a minute.

  3. Mitochondrial Respiration Chain Enzymatic Activities in the Human Brain: Methodological Implications for Tissue Sampling and Storage.

    PubMed

    Ronsoni, Marcelo Fernando; Remor, Aline Pertile; Lopes, Mark William; Hohl, Alexandre; Troncoso, Iris H Z; Leal, Rodrigo Bainy; Boos, Gustavo Luchi; Kondageski, Charles; Nunes, Jean Costa; Linhares, Marcelo Neves; Lin, Kátia; Latini, Alexandra Susana; Walz, Roger

    2016-04-01

    Mitochondrial respiratory chain complexes enzymatic (MRCCE) activities were successfully evaluated in frozen brain samples. Epilepsy surgery offers an ethical opportunity to study human brain tissue surgically removed to treat drug resistant epilepsies. Epilepsy surgeries are done with hemodynamic and laboratory parameters to maintain physiology, but there are no studies analyzing the association among these parameters and MRCCE activities in the human brain tissue. We determined the intra-operative parameters independently associated with MRCCE activities in middle temporal neocortex (Cx), amygdala (AMY) and head of hippocampus (HIP) samples of patients (n = 23) who underwent temporal lobectomy using multiple linear regressions. MRCCE activities in Cx, AMY and HIP are differentially associated to trans-operative mean arterial blood pressure, O2 saturation, hemoglobin, and anesthesia duration to time of tissue sampling. The time-course between the last seizure occurrence and tissue sampling as well as the sample storage to biochemical assessments were also associated with enzyme activities. Linear regression models including these variables explain 13-17 % of MRCCE activities and show a moderate to strong effect (r = 0.37-0.82). Intraoperative hemodynamic and laboratory parameters as well as the time from last seizure to tissue sampling and storage time are associated with MRCCE activities in human samples from the Cx, AMYG and HIP. Careful control of these parameters is required to minimize confounding biases in studies using human brain samples collected from elective neurosurgery. PMID:26586405

  4. Culture methods of allograft musculoskeletal tissue samples in Australian bacteriology laboratories.

    PubMed

    Varettas, Kerry

    2013-12-01

    Samples of allograft musculoskeletal tissue are cultured by bacteriology laboratories to determine the presence of bacteria and fungi. In Australia, this testing is performed by 6 TGA-licensed clinical bacteriology laboratories with samples received from 10 tissue banks. Culture methods of swab and tissue samples employ a combination of solid agar and/or broth media to enhance micro-organism growth and maximise recovery. All six Australian laboratories receive Amies transport swabs and, except for one laboratory, a corresponding biopsy sample for testing. Three of the 6 laboratories culture at least one allograft sample directly onto solid agar. Only one laboratory did not use a broth culture for any sample received. An international literature review found that a similar combination of musculoskeletal tissue samples were cultured onto solid agar and/or broth media. Although variations of allograft musculoskeletal tissue samples, culture media and methods are used in Australian and international bacteriology laboratories, validation studies and method evaluations have challenged and supported their use in recovering fungi and aerobic and anaerobic bacteria.

  5. Heavy Metal Content in Thoracic Tissue Samples from Patients with and without NSCLC.

    PubMed

    Tran, Jessica Q; Dranikov, Alexandra; Iannucci, Anita; Wagner, Walter P; LoBello, Janine; Allen, Jeffrey; Weiss, Glen J

    2014-01-01

    Objectives. Environmental factors expose an individual to heavy metals that may stimulate cancer growth preclinically including non-small cell lung cancer (NSCLC) cells. Here, we examine the prevalence of four heavy metals present in postsurgical tissues from individuals with and without NSCLC. Materials and Methods. Thoracic tissue samples from two separate sample sets were analyzed for cadmium (Cd), arsenic (As), mercury (Hg), and lead (Pb) content. Results. In the first sample set, there was no significant measurable amount of Pb and Hg found in either NSCLC tissue or nonmalignant lung tissue samples. Cd was the most prevalent heavy metal and As was present in moderate amounts. In the second sample set, Cd was measurable across all tissue types taken from 28 NSCLC patients and significantly higher Cd was measurable in noncancer benign lung (n = 9). In the NSCLC samples, As was measurable in moderate amounts, while Hg and Pb amounts were negligible. Conclusion. Cd and As are present in lung tissues for patients with NSCLC. With existing preclinical evidence of their tumorigenecity, it is plausible that Cd and/or As may have an impact on NSCLC development. Additional studies examining the prevalence and association between smokers and nonsmokers are suggested. PMID:26316947

  6. Investigation of false positives associated with loop-mediated isothermal amplification assays for detection of Toxoplasma gondii in archived tissue samples of captive felids.

    PubMed

    Suleman, Essa; Mtshali, Moses Sibusiso; Lane, Emily

    2016-09-01

    Toxoplasma gondii is a ubiquitous protozoan parasite that infects humans and many different animals, including felids. Many molecular and serologic tests have been developed for detection of T. gondii in a wide range of hosts. Loop-mediated isothermal amplification (LAMP) is a field-friendly technique that lacks the practical drawbacks of other molecular and serologic tests, and LAMP assays have been successfully developed for detection of T. gondii in fresh tissue samples. In the current study, both a previously published and a de-novo designed LAMP assay were compared to a quantitative real-time (q)PCR assay, for the detection of T. gondii in archived formalin-fixed, paraffin-embedded (FFPE) tissue samples from captive wildlife. The LAMP assays produced conflicting results, generating both false positives and false negatives. Furthermore, the LAMP assays were unable to positively identify samples with low levels of parasites as determined by qPCR and histopathology. Therefore, these LAMP assays may not be the most suitable assays for detection of T. gondii in archived FFPE and frozen tissue samples. PMID:27449130

  7. Efficient and scalable serial extraction of DNA and RNA from frozen tissue samples.

    PubMed

    Mathot, Lucy; Lindman, Monica; Sjöblom, Tobias

    2011-01-01

    Advances in cancer genomics have created a demand for scalable sample processing. We here present a process for serial extraction of nucleic acids from the same frozen tissue sample based on magnetic silica particles. The process is automation friendly with high recoveries of pure DNA and RNA suitable for analysis.

  8. Terahertz spectroscopy and detection of brain tumor in rat fresh-tissue samples

    NASA Astrophysics Data System (ADS)

    Yamaguchi, S.; Fukushi, Y.; Kubota, O.; Itsuji, T.; Yamamoto, S.; Ouchi, T.

    2015-03-01

    Terahertz (THz) spectroscopy and imaging of biomedical samples is expected to be an important application of THz analysis techniques. Identification and localization of tumor tissue, imaging of biological samples, and analysis of DNA by THz spectroscopy have been reported. THz time-domain spectroscopy (TDS) is useful for obtaining the refractive index over a broad frequency range. However, THz-TDS spectra of fresh tissue samples are sensitive to procedures such as sample preparation, and a standardized measurement protocol is required. Therefore, in this work, we establish a protocol for measurements of THz spectra of fresh tissue and demonstrate reliable detection of rat brain tumor tissue. We use a reflection THz-TDS system to measure the refractive index spectra of the samples mounted on a quartz plate. The tissue samples were measured immediately after sectioning to avoid sample denaturalization during storage. Special care was taken in THz data processing to eliminate parasitic reflections and reduce noise. The error level in our refractive index measurements was as low as 0.02 in the frequency range 0.8-1.5 THz. With increasing frequency, the refractive index in the tumor and normal regions monotonically decreased, similarly to water, and it was 0.02 higher in the tumor regions. The spectral data suggest that the tumor regions have higher water content. Hematoxylin-eosin stained images showed that increased cell density was also responsible for the observed spectral features. A set of samples from 10 rats showed consistent results. Our results suggest that reliable tumor detection in fresh tissue without pretreatment is possible with THz spectroscopy measurements. THz spectroscopy has the potential to become a real-time in vivo diagnostic method.

  9. Sample Preparation for Mass Spectrometry Imaging of Plant Tissues: A Review.

    PubMed

    Dong, Yonghui; Li, Bin; Malitsky, Sergey; Rogachev, Ilana; Aharoni, Asaph; Kaftan, Filip; Svatoš, Aleš; Franceschi, Pietro

    2016-01-01

    Mass spectrometry imaging (MSI) is a mass spectrometry based molecular ion imaging technique. It provides the means for ascertaining the spatial distribution of a large variety of analytes directly on tissue sample surfaces without any labeling or staining agents. These advantages make it an attractive molecular histology tool in medical, pharmaceutical, and biological research. Likewise, MSI has started gaining popularity in plant sciences; yet, information regarding sample preparation methods for plant tissues is still limited. Sample preparation is a crucial step that is directly associated with the quality and authenticity of the imaging results, it therefore demands in-depth studies based on the characteristics of plant samples. In this review, a sample preparation pipeline is discussed in detail and illustrated through selected practical examples. In particular, special concerns regarding sample preparation for plant imaging are critically evaluated. Finally, the applications of MSI techniques in plants are reviewed according to different classes of plant metabolites. PMID:26904042

  10. Sample Preparation for Mass Spectrometry Imaging of Plant Tissues: A Review

    PubMed Central

    Dong, Yonghui; Li, Bin; Malitsky, Sergey; Rogachev, Ilana; Aharoni, Asaph; Kaftan, Filip; Svatoš, Aleš; Franceschi, Pietro

    2016-01-01

    Mass spectrometry imaging (MSI) is a mass spectrometry based molecular ion imaging technique. It provides the means for ascertaining the spatial distribution of a large variety of analytes directly on tissue sample surfaces without any labeling or staining agents. These advantages make it an attractive molecular histology tool in medical, pharmaceutical, and biological research. Likewise, MSI has started gaining popularity in plant sciences; yet, information regarding sample preparation methods for plant tissues is still limited. Sample preparation is a crucial step that is directly associated with the quality and authenticity of the imaging results, it therefore demands in-depth studies based on the characteristics of plant samples. In this review, a sample preparation pipeline is discussed in detail and illustrated through selected practical examples. In particular, special concerns regarding sample preparation for plant imaging are critically evaluated. Finally, the applications of MSI techniques in plants are reviewed according to different classes of plant metabolites. PMID:26904042

  11. A tissue fixative that protects macromolecules (DNA, RNA, and protein) and histomorphology in clinical samples.

    PubMed

    Vincek, Vladimir; Nassiri, Mehdi; Nadji, Mehrdad; Morales, Azorides R

    2003-10-01

    Preservation of macromolecules (DNA, RNA, and proteins) in tissue is traditionally achieved by immediate freezing of the sample. Although isolation of PCR-able RNA has been reported from formalin-fixed, paraffin-embedded tissues, the process has not been shown to be reproducible because high molecular weight RNA is usually degraded. We investigated the potential value of a new universal molecular fixative (UMFIX, Sakura Finetek USA, Inc., Torrance, California) in preservation of macromolecules in paraffin-embedded tissue. Mouse and human tissues were fixed in UMFIX from 1 hour to 8 weeks. They were then processed by a rapid tissue processing (RTP) system, embedded in paraffin, and evaluated for routine histology as well as for the quality and quantity of DNA, RNA, and proteins. Formalin-fixed tissues were processed by RTP and evaluated in a similar manner. Fresh-frozen samples were used as controls. The morphology of UMFIX-exposed tissue was comparable to that fixed in formalin. High molecular weight RNA was preserved in tissue that was immediately fixed in UMFIX and stored from 1 hour to 8 weeks at room temperature. There were no significant differences between UMFIX-exposed and frozen tissues on PCR, RT-PCR, real-time PCR, and expression microarrays. Similarly, physical and antigenic preservation of proteins in UMFIX tissue was similar to fresh state. Both RNA and proteins were substantially degraded in formalin-fixed and similarly processed specimens. We concluded that it is now possible to preserve histomorphology and intact macromolecules in the same archival paraffin-embedded tissue through the use of a novel fixative and a rapid processing system.

  12. MALDI direct analysis and imaging of frozen versus FFPE tissues: what strategy for which sample?

    PubMed

    Wisztorski, Maxence; Franck, Julien; Salzet, Michel; Fournier, Isabelle

    2010-01-01

    Significant advances have been made in the past decade in the field of mass spectrometry imaging with MALDI ion sources (MALDI-MSI). While MALDI-MSI has high potential in the field of biology and in the clinic, a challenge for MALDI-MSI has been to adapt itself to a greater range of sample types. In particular, much of the biological archived materials for pathology studies are tissue biopsies fixed with paraformaldehyde and embedded in paraffin (FFPE tissues) because of the high stability of such samples. Thus, there has been a need to develop strategies for analyzing FFPE samples as this would allow retrospective studies of past clinical cases on large cohorts of existing samples. Obviously, PAF fixation, by inducing protein cross-linking, causes problems for molecular analysis by MS. We developed on tissue digestion strategies for overcoming these difficulties and allowing molecular data to be retrieved from FFPE samples no matter how long they have been stored. These digestion strategies preserve localization from digested proteins making MALDI-MSI of proteins possible by monitoring the resulting peptides. We present methods and protocols for FFPE samples. These strategies have proven to be valuable for all tested FFPE samples and have opened archived tissues from hospital banks to MALDI-MSI.

  13. Broth versus solid agar culture of swab samples of cadaveric allograft musculoskeletal tissue.

    PubMed

    Varettas, Kerry

    2013-12-01

    As part of the donor assessment protocol, bioburden assessment must be performed on allograft musculoskeletal tissue samples collected at the time of tissue retrieval. Swab samples of musculoskeletal tissue allografts from cadaveric donors are received at the microbiology department of the South Eastern Area Laboratory Services (Australia) to determine the presence of bacteria and fungi. This study will review the isolation rate of organisms from solid agar and broth culture of swab samples of cadaveric allograft musculoskeletal tissue over a 6-year period, 2006-2011. Swabs were inoculated onto horse blood agar (anaerobic, 35 °C) and chocolate agar (CO2, 35 °C) and then placed into a cooked meat broth (aerobic, 35 °C). A total of 1,912 swabs from 389 donors were received during the study period. 557 (29.1 %) swabs were culture positive with the isolation of 713 organisms, 249 (34.9 %) from solid agar culture and an additional 464 (65.1 %) from broth culture only. This study has shown that the broth culture of cadaveric allograft musculoskeletal swab samples recovered a greater amount of organisms than solid agar culture. Isolates such as Clostridium species and Staphylococcus aureus would not have been isolated from solid agar culture alone. Broth culture is an essential part of the bioburden assessment protocol of swab samples of cadaveric allograft musculoskeletal tissue in this laboratory.

  14. An antigen trapping ELISA for the detection of capripoxvirus in tissue culture supernatant and biopsy samples.

    PubMed

    Carn, V M

    1995-01-01

    A trapping ELISA for the detection of capripoxvirus antigen in tissue culture supernatant and biopsy material was developed, using a guinea-pig polyclonal detector antiserum raised against a recombinant capripoxvirus specific antigen, expressed in Escherichia coli using the plasmid vector pGEX-2T. The ELISA detected antigen in tissue culture samples that on virus titration contained equal to or in excess of 10(2.8) TCID50/ml. Virus isolation and ELISA were compared for the detection of capripoxvirus in skin biopsy samples from sheep, goats and cattle. The ELISA compared well with virus isolation, and has applications as a diagnostic test. This assay reduces the reliance of diagnostic laboratories on tissue culture facilities, and can be used to confirm the presence of capripoxvirus in tissue culture.

  15. Detection of Slit2 promoter hypermethylation in tissue and serum samples from breast cancer patients.

    PubMed

    Kim, Ga-Eon; Lee, Kyung Hwa; Choi, Yoo Duk; Lee, Ji Shin; Lee, Jae Hyuk; Nam, Jong Hee; Choi, Chan; Park, Min Ho; Yoon, Jung Han

    2011-10-01

    Promoter hypermethylation has been shown to be a common mechanism for inactivation of tumor suppressor genes in breast cancer. The aim of this study was to investigate the prevalence of Slit2 promoter hypermethylation in both the tumor and serum samples of breast cancer patients with ductal carcinoma in situ (DCIS) or invasive breast carcinoma (IBC). The methylation status of Slit2 was investigated in 210 tissue samples (15 breast with no pathological findings, 26 DCIS, and 169 IBC samples) and 123 corresponding serum samples (15 breast with no pathological findings, 26 DCIS, and 82 IBC samples) using methylation-specific polymerase chain reaction. Immunohistochemical staining for Slit2 was also performed using tissue microarray blocks to determine whether Slit2 promoter hypermethylation correlated with loss of Slit2 expression. Slit2 promoter hypermethylation was not detected in breast tissue and serum samples from patients with no pathological findings. DCIS or IBC showed a statistically higher frequency of Slit2 promoter hypermethylation compared to breast with no pathological findings in both the tissue and serum samples; however, there were no statistically significant differences between DCIS and IBC samples. Similar Slit2 promoter hypermethylation patterns were seen in the tissue samples and corresponding serum specimens (p < 0.001). Slit2 promoter hypermethylation was associated with loss of Slit2 expression. These results suggest that Slit2 promoter hypermethylation appears to be responsible for functionally silencing Slit2 expression. Slit2 promoter hypermethylation may be considered as a possible serum marker for early detection of breast cancer.

  16. X-ray scattering for the characterization of lyophilized breast tissue samples

    NASA Astrophysics Data System (ADS)

    Elshemey, Wael M.; Mohamed, Fayrouz S.; Khater, Ibrahim M.

    2013-09-01

    This work investigates the possibility of characterizing breast cancer by measuring the X-ray scattering profiles of lyophilized excised breast tissue samples. Since X-ray scattering from water-rich tissue is dominated by scattering from water, the removal of water by lyophilization would enhance the characterization process. In the present study, X-ray scattering profiles of 22 normal, 22 malignant and 10 benign breast tissue samples are measured. The cut-offs of scatter diagrams, sensitivity, specificity and diagnostic accuracy of three characterization parameters (full width at half maximum (FWHM) for the peak at 1.1 nm-1, area under curve (AUC), and ratio of 1st to 2nd scattering peak intensities (I1/I2%)) are calculated and compared to the data from non-lyophilized samples. Results show increased sensitivity (up to 100%) of the present data on lyophilized breast tissue samples compared to previously reported data for non-lyophilized samples while the specificity (up to 95.4%), diagnostic accuracy (up to 95.4%) and receiver operating characteristic (ROC) curve values (up to 0.9979) for both sets of data are comparable. The present study shows significant differences between normal samples and each of malignant and benign samples. Only subtle differences exist between malignant and benign lyophilized breast tissue samples where FWHM=0.7±0.1 and 0.8±0.3, AUC=1.3±0.2 and 1.4±0.2 and I1/I2%=44.9±11.0 and 52.4±7.6 for malignant and benign samples respectively.

  17. Effects of formalin fixation on tissue optical properties of in-vitro brain samples

    NASA Astrophysics Data System (ADS)

    Anand, Suresh; Cicchi, Riccardo; Martelli, Fabrizio; Giordano, Flavio; Buccoliero, Anna Maria; Guerrini, Renzo; Pavone, Francesco S.

    2015-03-01

    Application of light spectroscopy based techniques for the detection of cancers have emerged as a promising approach for tumor diagnostics. In-vivo or freshly excised samples are normally used for point spectroscopic studies. However, ethical issues related to in-vivo studies, rapid decay of surgically excised tissues and sample availability puts a limitation on in-vivo and in-vitro studies. There has been a few studies reported on the application of formalin fixed samples with good discrimination capability. Usually formalin fixation is performed to prevent degradation of tissues after surgical resection. Fixing tissues in formalin prevents cell death by forming cross-linkages with proteins. Previous investigations have revealed that washing tissues fixed in formalin using phosphate buffered saline is known to reduce the effects of formalin during spectroscopic measurements. But this could not be the case with reflectance measurements. Hemoglobin is a principal absorbing medium in biological tissues in the visible range. Formalin fixation causes hemoglobin to seep out from red blood cells. Also, there could be alterations in the refractive index of tissues when fixed in formalin. In this study, we propose to investigate the changes in tissue optical properties between freshly excised and formalin fixed brain tissues. The results indicate a complete change in the spectral profile in the visible range where hemoglobin has its maximum absorption peaks. The characteristic bands of oxy-hemoglobin at 540, 580 nm and deoxy-hemoglobin at 555 nm disappear in the case of samples fixed in formalin. In addition, an increased spectral intensity was observed for the wavelengths greater than 650 nm where scattering phenomena are presumed to dominate.

  18. Comparison of organochlorine residues in human adipose tissue autopsy samples from two Ontario municipalities

    SciTech Connect

    Williams, D.T.; LeBel, G.L.; Junkins, E.

    1984-01-01

    Human adipose tissue samples obtained during autopsies in a Canadian Great Lakes community, Kingston, Ontario, and a second community, Ottawa, Ontario, were analyzed for organochlorine pesticides, polychlorobiphenyls, chlorobenzenes, and chlorophenols. Significantly different levels of Dichlorodiphenyl-dichlorethane, mirex, hexachlorobenzene, and 2,3,4,6-tetrachlorophenol were found in Kingston adipose tissues compared to Ottawa tissues. Residue levels of oxychlordane, mirex, and polychlorinated biphenyls were significantly different in Kingston males versus Kingston females. The means and ranges of residue levels were contrasted with those reported in previous Canadian surveys.

  19. Quantitative mapping of collagen fiber alignment in thick tissue samples using transmission polarized-light microscopy.

    PubMed

    Yakovlev, Dmitry D; Shvachkina, Marina E; Sherman, Maria M; Spivak, Andrey V; Pravdin, Alexander B; Yakovlev, Dmitry A

    2016-07-01

    Immersion optical clearing makes it possible to use transmission polarized-light microscopy for characterization of thick (200 to 2000  μm) layers of biological tissues. We discuss polarization properties of thick samples in the context of the problem of characterization of collagen fiber alignment in connective tissues such as sclera and dermis. Optical chirality caused by azimuthal variations of the macroscopic (effective) optic axis of the medium across the sample thickness should be considered in polarization mapping of thick samples of these tissues. We experimentally evaluate to what extent the optical chirality affects the measurement results in typical situations and show under what conditions it can be easily taken into account and does not hinder, but rather helps, in characterization of collagen fiber alignment.

  20. Quantitative mapping of collagen fiber alignment in thick tissue samples using transmission polarized-light microscopy

    NASA Astrophysics Data System (ADS)

    Yakovlev, Dmitry D.; Shvachkina, Marina E.; Sherman, Maria M.; Spivak, Andrey V.; Pravdin, Alexander B.; Yakovlev, Dmitry A.

    2016-07-01

    Immersion optical clearing makes it possible to use transmission polarized-light microscopy for characterization of thick (200 to 2000 μm) layers of biological tissues. We discuss polarization properties of thick samples in the context of the problem of characterization of collagen fiber alignment in connective tissues such as sclera and dermis. Optical chirality caused by azimuthal variations of the macroscopic (effective) optic axis of the medium across the sample thickness should be considered in polarization mapping of thick samples of these tissues. We experimentally evaluate to what extent the optical chirality affects the measurement results in typical situations and show under what conditions it can be easily taken into account and does not hinder, but rather helps, in characterization of collagen fiber alignment.

  1. Isolation of high quality protein samples from punches of formalin fixed and paraffin embedded tissue blocks.

    PubMed

    Kroll, J; Becker, K F; Kuphal, S; Hein, R; Hofstädter, F; Bosserhoff, A K

    2008-04-01

    In general, it is believed that the extraction of proteins from formalin-fixed paraffin embedded samples is not feasible. However, recently a new technique was developed, presenting the extraction of non-degraded, full length proteins from formalin fixed tissues, usable for western blotting and protein arrays. In the study presented here, we applied this technique to punch biopsies of formalin fixed tissues embedded in paraffin to reduce heterogeneity of the tissue represented in sections, and to ensure analysing mainly defined cellular material. Successful extraction was achieved even from very small samples (0.7 mm(3)). Additionally, we were able to detect highly glycosylated proteins and protein modification, such as phosphorylation. Interestingly, with this technique it is feasible to extract high quality proteins from 14 year old samples. In summary, the new technique makes a great pool of material now usable for molecular analysis with high throughput tools. PMID:18228195

  2. Histopathological lesions associated with equine periodontal disease.

    PubMed

    Cox, Alistair; Dixon, Padraic; Smith, Sionagh

    2012-12-01

    Equine periodontal disease (EPD) is a common and painful condition, the aetiology and pathology of which are poorly understood. To characterise the histopathological lesions associated with EPD, the skulls of 22 horses were assessed grossly for the presence of periodontal disease, and a standard set of interdental tissues taken from each for histopathological examination. Histological features of EPD included ulceration and neutrophilic inflammation of the gingival epithelium. Mononuclear and eosinophilic inflammation of the gingival lamina propria and submucosa was commonly present irrespective of the presence or degree of periodontal disease. Gingival hyperplasia was present to some degree in all horses, and was only weakly associated with the degree of periodontal disease. In all horses dental plaque was present at the majority of sites examined and was often associated with histological evidence of peripheral cemental erosion. Bacteria (including spirochaetes in four horses) were identified in gingival samples by Gram and silver impregnation techniques and were significantly associated with the presence of periodontal disease. This is the first study to describe histological features of EPD, and the first to identify associated spirochaetes in some cases. Histological features were variable, and there was considerable overlap of some features between the normal and diseased gingiva. Further investigation into the potential role of bacteria in the pathogenesis and progression of EPD is warranted.

  3. Method for the detection of desmethylbromethalin in animal tissue samples for the determination of bromethalin exposure.

    PubMed

    Filigenzi, Michael S; Bautista, Adrienne C; Aston, Linda S; Poppenga, Robert H

    2015-06-01

    Bromethalin, a potent neurotoxin, is widely available for use as a rodenticide. As access to other rodenticides is reduced due to regulatory pressure, the use of bromethalin is likely to increase with a concomitant increase in poisonings in nontarget animals. Analytical methods for the detection of bromethalin residues in animals suspected to have been exposed to this rodenticide are needed to support post-mortem diagnosis of toxicosis. This paper describes a novel method for the analysis of desmethylbromethalin (DMB), bromethalin's toxic metabolite, in tissue samples such as liver, brain, and adipose. Samples were extracted with 5% ethanol in ethyl acetate, and an aliquot of the extract was evaporated dry, reconstituted, and analyzed by reverse phase ultrahigh-performance liquid chromatography-mass spectrometry. The mass spectrometer utilized electrospray ionization in negative ion mode with multiple reaction monitoring. This method was qualitatively validated at a level of 1.0 ng/g in liver tissue. The quantitative potential of the method was also evaluated, and a method detection limit of 0.35 ng/g wet weight was determined in fat tissue. DMB was detected in tissue samples from animals suspected to have been poisoned by this compound. To the authors' knowledge, there have been no other methods reported for analysis of DMB in tissue samples using LC-MS/MS. PMID:25688571

  4. Sample processing considerations for detecting copy number changes in formalin-fixed, paraffin-embedded tissues.

    PubMed

    Jacobs, Sharoni

    2012-11-01

    The Whole Genome Sampling Analysis (WGSA) assay in combination with Affymetrix GeneChip Mapping Arrays is used for copy number analysis of high-quality DNA samples (i.e., samples that have been collected from blood, fresh or frozen tissue, or cell lines). Formalin-fixed, paraffin-embedded (FFPE) samples, however, represent the most prevalent form of archived clinical samples, but they provide additional challenges for molecular assays. FFPE processing usually results in the degradation of FFPE DNA and in the contamination and chemical modification of these DNA samples. Because of these issues, FFPE DNA is not suitable for all molecular assays designed for high-quality DNA samples. Strategies recommended for processing FFPE DNA samples through WGSA and to the Mapping arrays are described here. PMID:23118355

  5. Sample processing considerations for detecting copy number changes in formalin-fixed, paraffin-embedded tissues.

    PubMed

    Jacobs, Sharoni

    2012-11-01

    The Whole Genome Sampling Analysis (WGSA) assay in combination with Affymetrix GeneChip Mapping Arrays is used for copy number analysis of high-quality DNA samples (i.e., samples that have been collected from blood, fresh or frozen tissue, or cell lines). Formalin-fixed, paraffin-embedded (FFPE) samples, however, represent the most prevalent form of archived clinical samples, but they provide additional challenges for molecular assays. FFPE processing usually results in the degradation of FFPE DNA and in the contamination and chemical modification of these DNA samples. Because of these issues, FFPE DNA is not suitable for all molecular assays designed for high-quality DNA samples. Strategies recommended for processing FFPE DNA samples through WGSA and to the Mapping arrays are described here.

  6. Citrulline as a Biomarker in the Murine Total-Body Irradiation Model: Correlation of Circulating and Tissue Citrulline to Small Intestine Epithelial Histopathology.

    PubMed

    Jones, Jace W; Tudor, Gregory; Li, Fei; Tong, Yan; Katz, Barry; Farese, Ann M; MacVittie, Thomas J; Booth, Catherine; Kane, Maureen A

    2015-11-01

    The use of plasma citrulline as a biomarker for gastrointestinal acute radiation syndrome via exposure to total-body irradiation in a murine model was investigated. The radiation exposure covered lethal, mid-lethal, and sub-lethal gastrointestinal acute radiation syndrome. Plasma citrulline profiles were generated over the first 6 d following total-body irradiation exposure of 6-15 Gy. In addition, plasma citrulline was comprehensively evaluated in the context of matching small intestine citrulline and histopathology. Higher plasma citrulline was significantly associated with lower irradiation doses over the first 6 d following the irradiation insult. Furthermore, higher plasma citrulline was significantly associated with higher crypt survival. The correlation of the plasma citrulline to crypt survival was more robust for higher irradiation doses and for later time points. The data suggested plasma citrulline was most informative for reflecting gastrointestinal injury resulting from exposure to 9-15 Gy total-body irradiation covering time-points 2-5 d post the irradiation insult. PMID:26425905

  7. Citrulline as a Biomarker in the Murine Total-Body Irradiation Model: Correlation of Circulating and Tissue Citrulline to Small Intestine Epithelial Histopathology.

    PubMed

    Jones, Jace W; Tudor, Gregory; Li, Fei; Tong, Yan; Katz, Barry; Farese, Ann M; MacVittie, Thomas J; Booth, Catherine; Kane, Maureen A

    2015-11-01

    The use of plasma citrulline as a biomarker for gastrointestinal acute radiation syndrome via exposure to total-body irradiation in a murine model was investigated. The radiation exposure covered lethal, mid-lethal, and sub-lethal gastrointestinal acute radiation syndrome. Plasma citrulline profiles were generated over the first 6 d following total-body irradiation exposure of 6-15 Gy. In addition, plasma citrulline was comprehensively evaluated in the context of matching small intestine citrulline and histopathology. Higher plasma citrulline was significantly associated with lower irradiation doses over the first 6 d following the irradiation insult. Furthermore, higher plasma citrulline was significantly associated with higher crypt survival. The correlation of the plasma citrulline to crypt survival was more robust for higher irradiation doses and for later time points. The data suggested plasma citrulline was most informative for reflecting gastrointestinal injury resulting from exposure to 9-15 Gy total-body irradiation covering time-points 2-5 d post the irradiation insult.

  8. Collecting and Storing Tissue, Blood, and Bone Marrow Samples From Patients With Rhabdomyosarcoma or Other Soft Tissue Sarcoma

    ClinicalTrials.gov

    2016-09-23

    Adult Rhabdomyosarcoma; Childhood Desmoplastic Small Round Cell Tumor; Chordoma; Desmoid Tumor; Metastatic Childhood Soft Tissue Sarcoma; Nonmetastatic Childhood Soft Tissue Sarcoma; Previously Treated Childhood Rhabdomyosarcoma; Previously Untreated Childhood Rhabdomyosarcoma; Recurrent Adult Soft Tissue Sarcoma; Recurrent Childhood Rhabdomyosarcoma; Recurrent Childhood Soft Tissue Sarcoma; Stage I Adult Soft Tissue Sarcoma; Stage II Adult Soft Tissue Sarcoma; Stage III Adult Soft Tissue Sarcoma; Stage IV Adult Soft Tissue Sarcoma

  9. Automated MALDI matrix coating system for multiple tissue samples for imaging mass spectrometry.

    PubMed

    Mounfield, William P; Garrett, Timothy J

    2012-03-01

    Uniform matrix deposition on tissue samples for matrix-assisted laser desorption/ionization (MALDI) is key for reproducible analyte ion signals. Current methods often result in nonhomogenous matrix deposition, and take time and effort to produce acceptable ion signals. Here we describe a fully-automated method for matrix deposition using an enclosed spray chamber and spray nozzle for matrix solution delivery. A commercial air-atomizing spray nozzle was modified and combined with solenoid controlled valves and a Programmable Logic Controller (PLC) to control and deliver the matrix solution. A spray chamber was employed to contain the nozzle, sample, and atomized matrix solution stream, and to prevent any interference from outside conditions as well as allow complete control of the sample environment. A gravity cup was filled with MALDI matrix solutions, including DHB in chloroform/methanol (50:50) at concentrations up to 60 mg/mL. Various samples (including rat brain tissue sections) were prepared using two deposition methods (spray chamber, inkjet). A linear ion trap equipped with an intermediate-pressure MALDI source was used for analyses. Optical microscopic examination showed a uniform coating of matrix crystals across the sample. Overall, the mass spectral images gathered from tissues coated using the spray chamber system were of better quality and more reproducible than from tissue specimens prepared by the inkjet deposition method.

  10. Rapid mass spectrometric conversion of tissue biopsy samples into permanent quantitative digital proteome maps

    PubMed Central

    Guo, Tiannan; Kouvonen, Petri; Koh, Ching Chiek; Gillet, Ludovic C; Wolski, Witold E; Röst, Hannes L; Rosenberger, George; Collins, Ben C; Blum, Lorenz C; Gillessen, Silke; Joerger, Markus; Jochum, Wolfram; Aebersold, Ruedi

    2015-01-01

    Clinical specimens are each inherently unique, limited and non-renewable. As such, small samples such as tissue biopsies are often completely consumed after a limited number of analyses. Here we present a method that enables fast and reproducible conversion of a small amount of tissue (approximating the quantity obtained by a biopsy) into a single, permanent digital file representing the mass spectrometry-measurable proteome of the sample. The method combines pressure cycling technology (PCT) and SWATH mass spectrometry (MS), and the resulting proteome maps can be analyzed, re-analyzed, compared and mined in silico to detect and quantify specific proteins across multiple samples. We used this method to process and convert 18 biopsy samples from 9 renal cell carcinoma patients into SWATH-MS fragment ion maps. From these proteome maps we detected and quantified more than 2,000 proteins with a high degree of reproducibility across all samples. The identified proteins clearly separated tumorous kidney tissues from healthy tissue, and differentiated distinct histomorphological kidney cancer subtypes. PMID:25730263

  11. Automated MALDI Matrix Coating System for Multiple Tissue Samples for Imaging Mass Spectrometry

    NASA Astrophysics Data System (ADS)

    Mounfield, William P.; Garrett, Timothy J.

    2012-03-01

    Uniform matrix deposition on tissue samples for matrix-assisted laser desorption/ionization (MALDI) is key for reproducible analyte ion signals. Current methods often result in nonhomogenous matrix deposition, and take time and effort to produce acceptable ion signals. Here we describe a fully-automated method for matrix deposition using an enclosed spray chamber and spray nozzle for matrix solution delivery. A commercial air-atomizing spray nozzle was modified and combined with solenoid controlled valves and a Programmable Logic Controller (PLC) to control and deliver the matrix solution. A spray chamber was employed to contain the nozzle, sample, and atomized matrix solution stream, and to prevent any interference from outside conditions as well as allow complete control of the sample environment. A gravity cup was filled with MALDI matrix solutions, including DHB in chloroform/methanol (50:50) at concentrations up to 60 mg/mL. Various samples (including rat brain tissue sections) were prepared using two deposition methods (spray chamber, inkjet). A linear ion trap equipped with an intermediate-pressure MALDI source was used for analyses. Optical microscopic examination showed a uniform coating of matrix crystals across the sample. Overall, the mass spectral images gathered from tissues coated using the spray chamber system were of better quality and more reproducible than from tissue specimens prepared by the inkjet deposition method.

  12. Rapid quantification of inflammation in tissue samples using perfluorocarbon emulsion and fluorine-19 nuclear magnetic resonance

    PubMed Central

    Ahrens, Eric T.; Young, Won-Bin; Xu, Hongyan; Pusateri, Lisa K.

    2016-01-01

    Quantification of inflammation in tissue samples can be a time-intensive bottleneck in therapeutic discovery and preclinical endeavors. We describe a versatile and rapid approach to quantitatively assay macrophage burden in intact tissue samples. Perfluorocarbon (PFC) emulsion is injected intravenously, and the emulsion droplets are effectively taken up by monocytes and macrophages. These ‘in situ’ labeled cells participate in inflammatory events in vivo resulting in PFC accumulation at inflammatory loci. Necropsied tissues or intact organs are subjected to conventional fluorine-19 (19F) NMR spectroscopy to quantify the total fluorine content per sample, proportional to the macrophage burden. We applied these methods to a rat model of experimental allergic encephalomyelitis (EAE) exhibiting extensive inflammation and demyelination in the central nervous system (CNS), particularly in the spinal cord. In a cohort of EAE rats, we used 19F NMR to derive an inflammation index (IFI) in intact CNS tissues. Immunohistochemistry was used to confirm intracellular colocalization of the PFC droplets within CNS CD68+ cells having macrophage morphology. The IFI linearly correlated to mRNA levels of CD68 via real-time PCR analysis. This 19F NMR approach can accelerate tissue analysis by at least an order of magnitude compared with histological approaches. PMID:21548906

  13. Semiquantitative determination of polychlorinated biphenyls in tissue samples by thin layer chromatography

    USGS Publications Warehouse

    Mulhern, B.M.; Cromartie, E.; Reichel, W.L.; Belisle, A.A.

    1971-01-01

    A method is described for the analysis of polychlorinated biphenyl (PCB) compounds in tissue samples. Cleanup by hexane-aceto-nitrile partitioning and Florisil column chromatography are performed on samples before oxidative treatment to convert DDE to DCBP. PCB components are then determined semi-quantitatively by TLC. No prior separation of PCB from chlorinated pesticides is required. The lower limit of sensitivity is 0.2 ?g.

  14. Three Dimensional Imaging of Paraffin Embedded Human Lung Tissue Samples by Micro-Computed Tomography

    PubMed Central

    Scott, Anna E.; Vasilescu, Dragos M.; Seal, Katherine A. D.; Keyes, Samuel D.; Mavrogordato, Mark N.; Hogg, James C.; Sinclair, Ian; Warner, Jane A.; Hackett, Tillie-Louise; Lackie, Peter M.

    2015-01-01

    Background Understanding the three-dimensional (3-D) micro-architecture of lung tissue can provide insights into the pathology of lung disease. Micro computed tomography (µCT) has previously been used to elucidate lung 3D histology and morphometry in fixed samples that have been stained with contrast agents or air inflated and dried. However, non-destructive microstructural 3D imaging of formalin-fixed paraffin embedded (FFPE) tissues would facilitate retrospective analysis of extensive tissue archives of lung FFPE lung samples with linked clinical data. Methods FFPE human lung tissue samples (n = 4) were scanned using a Nikon metrology µCT scanner. Semi-automatic techniques were used to segment the 3D structure of airways and blood vessels. Airspace size (mean linear intercept, Lm) was measured on µCT images and on matched histological sections from the same FFPE samples imaged by light microscopy to validate µCT imaging. Results The µCT imaging protocol provided contrast between tissue and paraffin in FFPE samples (15mm x 7mm). Resolution (voxel size 6.7 µm) in the reconstructed images was sufficient for semi-automatic image segmentation of airways and blood vessels as well as quantitative airspace analysis. The scans were also used to scout for regions of interest, enabling time-efficient preparation of conventional histological sections. The Lm measurements from µCT images were not significantly different to those from matched histological sections. Conclusion We demonstrated how non-destructive imaging of routinely prepared FFPE samples by laboratory µCT can be used to visualize and assess the 3D morphology of the lung including by morphometric analysis. PMID:26030902

  15. Phase-contrast Hounsfield units of fixated and non-fixated soft-tissue samples

    SciTech Connect

    Willner, Marian; Fior, Gabriel; Marschner, Mathias; Birnbacher, Lorenz; Schock, Jonathan; Braun, Christian; Fingerle, Alexander A.; Noël, Peter B.; Rummeny, Ernst J.; Pfeiffer, Franz; Herzen, Julia; Rozhkova, Elena A.

    2015-08-31

    X-ray phase-contrast imaging is a novel technology that achieves high soft-tissue contrast. Although its clinical impact is still under investigation, the technique may potentially improve clinical diagnostics. In conventional attenuation-based X-ray computed tomography, radiological diagnostics are quantified by Hounsfield units. Corresponding Hounsfield units for phase-contrast imaging have been recently introduced, enabling a setup-independent comparison and standardized interpretation of imaging results. Thus far, the experimental values of few tissue types have been reported; these values have been determined from fixated tissue samples. This study presents phase-contrast Hounsfield units for various types of non-fixated human soft tissues. A large variety of tissue specimens ranging from adipose, muscle and connective tissues to liver, kidney and pancreas tissues were imaged by a grating interferometer with a rotating-anode X-ray tube and a photon-counting detector. In addition, we investigated the effects of formalin fixation on the quantitative phase-contrast imaging results.

  16. Evaluation of frozen tissue-derived prognostic gene expression signatures in FFPE colorectal cancer samples

    PubMed Central

    Zhu, Jing; Deane, Natasha G.; Lewis, Keeli B.; Padmanabhan, Chandrasekhar; Washington, M. Kay; Ciombor, Kristen K.; Timmers, Cynthia; Goldberg, Richard M.; Beauchamp, R. Daniel; Chen, Xi

    2016-01-01

    Defining molecular features that can predict the recurrence of colorectal cancer (CRC) for stage II-III patients remains challenging in cancer research. Most available clinical samples are Formalin-Fixed, Paraffin-Embedded (FFPE). NanoString nCounter® and Affymetrix GeneChip® Human Transcriptome Array 2.0 (HTA) are the two platforms marketed for high-throughput gene expression profiling for FFPE samples. In this study, to evaluate the gene expression of frozen tissue-derived prognostic signatures in FFPE CRC samples, we evaluated the expression of 516 genes from published frozen tissue-derived prognostic signatures in 42 FFPE CRC samples measured by both platforms. Based on HTA platform-derived data, we identified both gene (99 individual genes, FDR < 0.05) and gene set (four of the six reported multi-gene signatures with sufficient information for evaluation, P < 0.05) expression differences associated with survival outcomes. Using nCounter platform-derived data, one of the six multi-gene signatures (P < 0.05) but no individual gene was associated with survival outcomes. Our study indicated that sufficiently high quality RNA could be obtained from FFPE tumor tissues to detect frozen tissue-derived prognostic gene expression signatures for CRC patients. PMID:27623752

  17. Comparison of Microdialysis Sampling Perfusion Fluid Components on the Foreign Body Reaction in Rat Subcutaneous Tissue

    PubMed Central

    Keeler, Geoffrey D.; Durdik, Jeannine M.; Stenken, Julie A.

    2013-01-01

    Microdialysis sampling is a commonly used technique for collecting solutes from the extracellular space of tissues in laboratory animals and humans. Large molecular weight solutes can be collected using high molecular weight cutoff (MWCO) membranes (100 kDa or greater). High MWCO membranes require addition of high molecular weight dextrans or albumin to the perfusion fluid to prevent fluid loss via ultrafiltration. While these perfusion fluid additives are commonly used during microdialysis sampling, the tissue response to the loss of these compounds across the membrane is poorly understood. Tissue reactions to implanted microdialysis sampling probes containing different microdialysis perfusion fluids were compared over a 7-day time period in rats. The base perfusion fluid was Ringer’s solution supplemented with either bovine serum albumin (BSA), rat serum albumin (RSA), Dextran-70, or Dextran-500. A significant inflammatory response to Dextran-70 was observed. No differences in the tissue response between BSA and RSA were observed. Among these agents, the BSA, RSA, and Dextran-500 produced a significantly reduced inflammatory response compared to the Dextran-70. This work demonstrates that use of Dextran-70 in microdialysis sampling perfusion fluids should be eliminated and replaced with Dextran-500 or other alternatives. PMID:24239995

  18. Evaluation of frozen tissue-derived prognostic gene expression signatures in FFPE colorectal cancer samples.

    PubMed

    Zhu, Jing; Deane, Natasha G; Lewis, Keeli B; Padmanabhan, Chandrasekhar; Washington, M Kay; Ciombor, Kristen K; Timmers, Cynthia; Goldberg, Richard M; Beauchamp, R Daniel; Chen, Xi

    2016-01-01

    Defining molecular features that can predict the recurrence of colorectal cancer (CRC) for stage II-III patients remains challenging in cancer research. Most available clinical samples are Formalin-Fixed, Paraffin-Embedded (FFPE). NanoString nCounter® and Affymetrix GeneChip® Human Transcriptome Array 2.0 (HTA) are the two platforms marketed for high-throughput gene expression profiling for FFPE samples. In this study, to evaluate the gene expression of frozen tissue-derived prognostic signatures in FFPE CRC samples, we evaluated the expression of 516 genes from published frozen tissue-derived prognostic signatures in 42 FFPE CRC samples measured by both platforms. Based on HTA platform-derived data, we identified both gene (99 individual genes, FDR < 0.05) and gene set (four of the six reported multi-gene signatures with sufficient information for evaluation, P < 0.05) expression differences associated with survival outcomes. Using nCounter platform-derived data, one of the six multi-gene signatures (P < 0.05) but no individual gene was associated with survival outcomes. Our study indicated that sufficiently high quality RNA could be obtained from FFPE tumor tissues to detect frozen tissue-derived prognostic gene expression signatures for CRC patients. PMID:27623752

  19. Tissue sampling from live blue mussels, Mytilus edulis. A field study from the Swedish west coast

    NASA Astrophysics Data System (ADS)

    Svärdh, Lillemor

    2003-05-01

    Histological techniques are often used to study environmental effects on mussels, but since these techniques include killing of the individuals, rare or endangered populations cannot be studied using conventional tissue sampling. This study is an attempt to find a method that can be used repeatedly with the same mussel individual and which does not affect growth and survival. From 200 mussels, Mytilus edulis, tissue was sampled in different ways, such as drilling a hole in the shell or prising apart the shell valves. Two kind of instruments were used, an injection needle and surgery forceps. Some of the drilled mussels had their holes sealed again with cement. Drilling a hole in the shell, removing tissue sample with surgical forceps and then leaving the holes open did not seriously harm the mussels during the two months the experiment lasted. But if the holes were sealed with cement, both length and weight growth were negatively affected (35% lower length growth and 36% lower weight growth compared to the control mussels). Mortality was highest among the drilled and sealed mussels (80% higher than among the other treatments). The vulnerability of the population, the aim of the study and the duration of the experiment should decide what method to use for tissue sampling. For long-term experiments and repeated sampling, opening the mussels by prizing apart the valves is a better alternative than drilling holes in the shells, but depending on the morphology of the species it could be difficult to sample the anterior part of the mussel body. For a short experiment and to sample anterior parts, drilling the shells, leaving the holes open and using surgical forceps, seems to be an acceptable compromise between the different treatments used.

  20. The Novel Application of Non-Lethal Citizen Science Tissue Sampling in Recreational Fisheries.

    PubMed

    Williams, Samuel M; Holmes, Bonnie J; Pepperell, Julian G

    2015-01-01

    Increasing fishing pressure and uncertainty surrounding recreational fishing catch and effort data promoted the development of alternative methods for conducting fisheries research. A pilot investigation was undertaken to engage the Australian game fishing community and promote the non-lethal collection of tissue samples from the black marlin Istiompax indica, a valuable recreational-only species in Australian waters, for the purpose of future genetic research. Recruitment of recreational anglers was achieved by publicizing the project in magazines, local newspapers, social media, blogs, websites and direct communication workshops at game fishing tournaments. The Game Fishing Association of Australia and the Queensland Game Fishing Association were also engaged to advertise the project and recruit participants with a focus on those anglers already involved in the tag-and-release of marlin. Participants of the program took small tissue samples using non-lethal methods which were stored for future genetic analysis. The program resulted in 165 samples from 49 participants across the known distribution of I. indica within Australian waters which was a sufficient number to facilitate a downstream population genetic analysis. The project demonstrated the potential for the development of citizen science sampling programs to collect tissue samples using non-lethal methods in order to achieve targeted research objects in recreationally caught species. PMID:26376487

  1. The Novel Application of Non-Lethal Citizen Science Tissue Sampling in Recreational Fisheries.

    PubMed

    Williams, Samuel M; Holmes, Bonnie J; Pepperell, Julian G

    2015-01-01

    Increasing fishing pressure and uncertainty surrounding recreational fishing catch and effort data promoted the development of alternative methods for conducting fisheries research. A pilot investigation was undertaken to engage the Australian game fishing community and promote the non-lethal collection of tissue samples from the black marlin Istiompax indica, a valuable recreational-only species in Australian waters, for the purpose of future genetic research. Recruitment of recreational anglers was achieved by publicizing the project in magazines, local newspapers, social media, blogs, websites and direct communication workshops at game fishing tournaments. The Game Fishing Association of Australia and the Queensland Game Fishing Association were also engaged to advertise the project and recruit participants with a focus on those anglers already involved in the tag-and-release of marlin. Participants of the program took small tissue samples using non-lethal methods which were stored for future genetic analysis. The program resulted in 165 samples from 49 participants across the known distribution of I. indica within Australian waters which was a sufficient number to facilitate a downstream population genetic analysis. The project demonstrated the potential for the development of citizen science sampling programs to collect tissue samples using non-lethal methods in order to achieve targeted research objects in recreationally caught species.

  2. The Novel Application of Non-Lethal Citizen Science Tissue Sampling in Recreational Fisheries

    PubMed Central

    Williams, Samuel M.; Holmes, Bonnie J.; Pepperell, Julian G.

    2015-01-01

    Increasing fishing pressure and uncertainty surrounding recreational fishing catch and effort data promoted the development of alternative methods for conducting fisheries research. A pilot investigation was undertaken to engage the Australian game fishing community and promote the non-lethal collection of tissue samples from the black marlin Istiompax indica, a valuable recreational-only species in Australian waters, for the purpose of future genetic research. Recruitment of recreational anglers was achieved by publicizing the project in magazines, local newspapers, social media, blogs, websites and direct communication workshops at game fishing tournaments. The Game Fishing Association of Australia and the Queensland Game Fishing Association were also engaged to advertise the project and recruit participants with a focus on those anglers already involved in the tag-and-release of marlin. Participants of the program took small tissue samples using non-lethal methods which were stored for future genetic analysis. The program resulted in 165 samples from 49 participants across the known distribution of I. indica within Australian waters which was a sufficient number to facilitate a downstream population genetic analysis. The project demonstrated the potential for the development of citizen science sampling programs to collect tissue samples using non-lethal methods in order to achieve targeted research objects in recreationally caught species. PMID:26376487

  3. Non-lethal sampling of walleye for stable isotope analysis: a comparison of three tissues

    USGS Publications Warehouse

    Chipps, Steven R.; VanDeHey, J.A.; Fincel, M.J.

    2012-01-01

    Stable isotope analysis of fishes is often performed using muscle or organ tissues that require sacrificing animals. Non-lethal sampling provides an alternative for evaluating isotopic composition for species of concern or individuals of exceptional value. Stable isotope values of white muscle (lethal) were compared with those from fins and scales (non-lethal) in walleye, Sander vitreus (Mitchill), from multiple systems, size classes and across a range of isotopic values. Isotopic variability was also compared among populations to determine the potential of non-lethal tissues for diet-variability analyses. Muscle-derived isotope values were enriched compared with fins and depleted relative to scales. A split-sample validation technique and linear regression found that isotopic composition of walleye fins and scales was significantly related to that in muscle tissue for both δ13C and δ15N (r2 = 0.79–0.93). However, isotopic variability was significantly different between tissue types in two of six populations for δ15N and three of six populations for δ13C. Although species and population specific, these findings indicate that isotopic measures obtained from non-lethal tissues are indicative of those obtained from muscle.

  4. Imaging of Proteins in Tissue Samples Using Nanospray Desorption Electrospray Ionization Mass Spectrometry.

    PubMed

    Hsu, Cheng-Chih; Chou, Pi-Tai; Zare, Richard N

    2015-11-17

    Chemical maps of tissue samples provide important information on biological processes therein. Recently, advances in tissue imaging have been achieved using ambient ionization techniques, such as desorption electrospray ionization mass spectrometry (DESI-MS), but such techniques have been almost exclusively confined to the mapping of lipids and metabolites. We report here the use of nanospray desorption electrospray ionization (nanoDESI) that allows us to image proteins in tissue samples in a label-free manner at atmospheric pressure with only minimum sample preparation. Multiply charged proteins with masses up to 15 kDa were successfully detected by nanoDESI using an LTQ Orbitrap mass spectrometer. In an adult mice brain section, expression of proteins including ubiquitin, β-thymosin, myelin basic protein, and hemoglobin were spatially mapped and characterized. We also determined the location of methylation on myelin basic protein. This imaging modality was further implemented to MYC-induced lymphomas. We observed an array of truncated proteins in the region where normal thymus cells were infiltrated by tumor cells, in contrast to healthy tissue.

  5. Phase-contrast Hounsfield units of fixated and non-fixated soft-tissue samples

    DOE PAGES

    Willner, Marian; Fior, Gabriel; Marschner, Mathias; Birnbacher, Lorenz; Schock, Jonathan; Braun, Christian; Fingerle, Alexander A.; Noël, Peter B.; Rummeny, Ernst J.; Pfeiffer, Franz; et al

    2015-08-31

    X-ray phase-contrast imaging is a novel technology that achieves high soft-tissue contrast. Although its clinical impact is still under investigation, the technique may potentially improve clinical diagnostics. In conventional attenuation-based X-ray computed tomography, radiological diagnostics are quantified by Hounsfield units. Corresponding Hounsfield units for phase-contrast imaging have been recently introduced, enabling a setup-independent comparison and standardized interpretation of imaging results. Thus far, the experimental values of few tissue types have been reported; these values have been determined from fixated tissue samples. This study presents phase-contrast Hounsfield units for various types of non-fixated human soft tissues. A large variety of tissuemore » specimens ranging from adipose, muscle and connective tissues to liver, kidney and pancreas tissues were imaged by a grating interferometer with a rotating-anode X-ray tube and a photon-counting detector. In addition, we investigated the effects of formalin fixation on the quantitative phase-contrast imaging results.« less

  6. MicroRNA Stability in FFPE Tissue Samples: Dependence on GC Content

    PubMed Central

    Kakimoto, Yu; Tanaka, Masayuki; Kamiguchi, Hiroshi; Ochiai, Eriko; Osawa, Motoki

    2016-01-01

    MicroRNAs (miRNAs) are small non-coding RNAs responsible for fine-tuning of gene expression at post-transcriptional level. The alterations in miRNA expression levels profoundly affect human health and often lead to the development of severe diseases. Currently, high throughput analyses, such as microarray and deep sequencing, are performed in order to identify miRNA biomarkers, using archival patient tissue samples. MiRNAs are more robust than longer RNAs, and resistant to extreme temperatures, pH, and formalin-fixed paraffin-embedding (FFPE) process. Here, we have compared the stability of miRNAs in FFPE cardiac tissues using next-generation sequencing. The mode read length in FFPE samples was 11 nucleotides (nt), while that in the matched frozen samples was 22 nt. Although the read counts were increased 1.7-fold in FFPE samples, compared with those in the frozen samples, the average miRNA mapping rate decreased from 32.0% to 9.4%. These results indicate that, in addition to the fragmentation of longer RNAs, miRNAs are to some extent degraded in FFPE tissues as well. The expression profiles of total miRNAs in two groups were highly correlated (0.88 tissues instead of miR-1, which was predominant before fixation. Subsequent quantitative PCR (qPCR) analyses revealed that miRNAs with GC content of less than 40% are more degraded than GC-rich miRNAs (p<0.0001). We showed that deep sequencing data obtained using FFPE samples cannot be directly compared with that of fresh frozen samples. The combination of miRNA deep sequencing and other quantitative analyses, such as qPCR, may improve the utility of archival FFPE tissue samples. PMID:27649415

  7. MicroRNA Stability in FFPE Tissue Samples: Dependence on GC Content.

    PubMed

    Kakimoto, Yu; Tanaka, Masayuki; Kamiguchi, Hiroshi; Ochiai, Eriko; Osawa, Motoki

    2016-01-01

    MicroRNAs (miRNAs) are small non-coding RNAs responsible for fine-tuning of gene expression at post-transcriptional level. The alterations in miRNA expression levels profoundly affect human health and often lead to the development of severe diseases. Currently, high throughput analyses, such as microarray and deep sequencing, are performed in order to identify miRNA biomarkers, using archival patient tissue samples. MiRNAs are more robust than longer RNAs, and resistant to extreme temperatures, pH, and formalin-fixed paraffin-embedding (FFPE) process. Here, we have compared the stability of miRNAs in FFPE cardiac tissues using next-generation sequencing. The mode read length in FFPE samples was 11 nucleotides (nt), while that in the matched frozen samples was 22 nt. Although the read counts were increased 1.7-fold in FFPE samples, compared with those in the frozen samples, the average miRNA mapping rate decreased from 32.0% to 9.4%. These results indicate that, in addition to the fragmentation of longer RNAs, miRNAs are to some extent degraded in FFPE tissues as well. The expression profiles of total miRNAs in two groups were highly correlated (0.88 tissues instead of miR-1, which was predominant before fixation. Subsequent quantitative PCR (qPCR) analyses revealed that miRNAs with GC content of less than 40% are more degraded than GC-rich miRNAs (p<0.0001). We showed that deep sequencing data obtained using FFPE samples cannot be directly compared with that of fresh frozen samples. The combination of miRNA deep sequencing and other quantitative analyses, such as qPCR, may improve the utility of archival FFPE tissue samples. PMID:27649415

  8. CT-guided Irreversible Electroporation in an Acute Porcine Liver Model: Effect of Previous Transarterial Iodized Oil Tissue Marking on Technical Parameters, 3D Computed Tomographic Rendering of the Electroporation Zone, and Histopathology

    SciTech Connect

    Sommer, C. M.; Fritz, S.; Vollherbst, D.; Zelzer, S.; Wachter, M. F. Bellemann, N. Gockner, T. Mokry, T. Schmitz, A.; Aulmann, S.; Stampfl, U.; Pereira, P.; Kauczor, H. U.; Werner, J.; Radeleff, B. A.

    2015-02-15

    PurposeTo evaluate the effect of previous transarterial iodized oil tissue marking (ITM) on technical parameters, three-dimensional (3D) computed tomographic (CT) rendering of the electroporation zone, and histopathology after CT-guided irreversible electroporation (IRE) in an acute porcine liver model as a potential strategy to improve IRE performance.MethodsAfter Ethics Committee approval was obtained, in five landrace pigs, two IREs of the right and left liver (RL and LL) were performed under CT guidance with identical electroporation parameters. Before IRE, transarterial marking of the LL was performed with iodized oil. Nonenhanced and contrast-enhanced CT examinations followed. One hour after IRE, animals were killed and livers collected. Mean resulting voltage and amperage during IRE were assessed. For 3D CT rendering of the electroporation zone, parameters for size and shape were analyzed. Quantitative data were compared by the Mann–Whitney test. Histopathological differences were assessed.ResultsMean resulting voltage and amperage were 2,545.3 ± 66.0 V and 26.1 ± 1.8 A for RL, and 2,537.3 ± 69.0 V and 27.7 ± 1.8 A for LL without significant differences. Short axis, volume, and sphericity index were 16.5 ± 4.4 mm, 8.6 ± 3.2 cm{sup 3}, and 1.7 ± 0.3 for RL, and 18.2 ± 3.4 mm, 9.8 ± 3.8 cm{sup 3}, and 1.7 ± 0.3 for LL without significant differences. For RL and LL, the electroporation zone consisted of severely widened hepatic sinusoids containing erythrocytes and showed homogeneous apoptosis. For LL, iodized oil could be detected in the center and at the rim of the electroporation zone.ConclusionThere is no adverse effect of previous ITM on technical parameters, 3D CT rendering of the electroporation zone, and histopathology after CT-guided IRE of the liver.

  9. Screening of Viral Pathogens from Pediatric Ileal Tissue Samples after Vaccination

    DOE PAGES

    Hewitson, Laura; Thissen, James B.; Gardner, Shea N.; McLoughlin, Kevin S.; Glausser, Margaret K.; Jaing, Crystal J.

    2014-01-01

    In 2010, researchers reported that the two US-licensed rotavirus vaccines contained DNA or DNA fragments from porcine circovirus (PCV). Although PCV, a common virus among pigs, is not thought to cause illness in humans, these findings raised several safety concerns. In this study, we sought to determine whether viruses, including PCV, could be detected in ileal tissue samples of children vaccinated with one of the two rotavirus vaccines. A broad spectrum, novel DNA detection technology, the Lawrence Livermore Microbial Detection Array (LLMDA), was utilized, and confirmation of viral pathogens using the polymerase chain reaction (PCR) was conducted. The LLMDAmore » technology was recently used to identify PCV from one rotavirus vaccine. Ileal tissue samples were analyzed from 21 subjects, aged 15–62 months. PCV was not detected in any ileal tissue samples by the LLMDA or PCR. LLMDA identified a human rotavirus A from one of the vaccinated subjects, which is likely due to a recent infection from a wild type rotavirus. LLMDA also identified human parechovirus, a common gastroenteritis viral infection, from two subjects. Additionally, LLMDA detected common gastrointestinal bacterial organisms from the Enterobacteriaceae , Bacteroidaceae , and Streptococcaceae families from several subjects. This study provides a survey of viral and bacterial pathogens from pediatric ileal samples, and may shed light on future studies to identify pathogen associations with pediatric vaccinations.« less

  10. Swine infectious agents in Tayassu pecari and Pecari tajacu tissue samples from Brazil.

    PubMed

    de Castro, Alessandra Marnie Martins Gomes; Brombila, Talita; Bersano, Josete Garcia; Soares, Herbert Sousa; Silva, Sheila Oliveira de Souza; Minervino, Antonio Humberto Hamad; Ogata, Renato Akio; Gennari, Solange Maria; Richtzenhain, Leonardo Jose

    2014-04-01

    Peccaries and pigs, Tayassuidae and Suidae respectively, diverged approximately one million years ago from a common ancestor. Because these families share some pathogens, peccaries can act as reservoirs of infectious pathogens for domestic and wild swine. We evaluated the presence of swine infectious agents in the spleen and lung tissues of white-lipped peccaries (WLP; Tayassu pecari) and collared peccaries (CP; Pecari tajacu) in Brazil. Samples from 10 adult CP and three WLP, which had been hunted by locals or hit by motor vehicles, were obtained from two free-ranging Brazilian populations. The samples were tested by PCR for Mycoplasma hyopneumoniae, Bordetella bronchiseptica, Pasteurella multocida, porcine circovirus 2 (PCV2), Suid herpesvirus 1 (SuHV-1), and porcine parvovirus (PPV). Positive samples were sequenced. Both species were negative for PPV and B. bronchiseptica and positive for PCV2 and SuHV-1. The lungs of two animals were positive for M. hyopneumoniae and P. multocida. This report is the first demonstration of PCV2 and SuHV-1 swine viruses and of M. hyopneumoniae and P. multocida bacteria in peccaries. One factor contributing to this detection was access to tissue samples, which is uncommon. The role of these infectious agents in peccaries is unknown and further epidemiologic studies should be performed. This study identified several infectious agents in peccaries and highlighted the importance of the tissue type used to detect pathogens.

  11. In situ hybridization method for studies of cell wall deficient M. paratuberculosis in tissue samples.

    PubMed

    Hulten, K; Karttunen, T J; El-Zimaity, H M; Naser, S A; Almashhrawi, A; Graham, D Y; El-Zaatari, F A

    2000-12-20

    Cell wall deficient forms of mycobacteria may be important in the pathogenesis of Crohn's disease and sarcoidosis. However, no method has been available to localize this type of organisms in tissue sections. We developed an in situ hybridization method for the demonstration of Mycobacterium paratuberculosis spheroplasts (cell wall deficient forms) in paraffin embedded tissue sections.M. paratuberculosis spheroplasts were prepared by treatment with glycine and lysozyme. Pieces of beef were injected with the prepared spheroplasts. The samples were fixed in buffered formalin and paraffin embedded. A M. paratuberculosis-specific probe derived from the IS900 gene was used. Specificity was controlled by using an irrelevant probe and by hybridizing sections with spheroplasts from other bacteria. Beef samples injected with M. paratuberculosis spheroplasts were the only samples that hybridized with the probe. Beef samples containing acid-fast or spheroplast forms of M. smegmatis and M. tuberculosis as well as the acid-fast forms of M. paratuberculosis did not hybridize with the probe. Unrelated bacterial controls, i.e. Helicobacter pylori and Escherichia coli were also negative in the assay. In situ hybridization with the IS900 probe provides a specific way to localize M. paratuberculosis spheroplasts in tissue sections and may be useful for studies of the connection between M. paratuberculosis and Crohn's disease and sarcoidosis. The assay may also be valuable for studies on Johne's diseased animals. PMID:11118736

  12. Targeted or whole genome sequencing of formalin fixed tissue samples: potential applications in cancer genomics

    PubMed Central

    Zhao, Yue; Cottrell, Joseph; Klotzle, Brandy; Godwin, Andrew K.; Koestler, Devin; Beyerlein, Peter; Fan, Jian-Bing; Bibikova, Marina; Chien, Jeremy

    2015-01-01

    Current genomic studies are limited by the poor availability of fresh-frozen tissue samples. Although formalin-fixed diagnostic samples are in abundance, they are seldom used in current genomic studies because of the concern of formalin-fixation artifacts. Better characterization of these artifacts will allow the use of archived clinical specimens in translational and clinical research studies. To provide a systematic analysis of formalin-fixation artifacts on Illumina sequencing, we generated 26 DNA sequencing data sets from 13 pairs of matched formalin-fixed paraffin-embedded (FFPE) and fresh-frozen (FF) tissue samples. The results indicate high rate of concordant calls between matched FF/FFPE pairs at reference and variant positions in three commonly used sequencing approaches (whole genome, whole exome, and targeted exon sequencing). Global mismatch rates and C·G > T·A substitutions were comparable between matched FF/FFPE samples, and discordant rates were low (<0.26%) in all samples. Finally, low-pass whole genome sequencing produces similar pattern of copy number alterations between FF/FFPE pairs. The results from our studies suggest the potential use of diagnostic FFPE samples for cancer genomic studies to characterize and catalog variations in cancer genomes. PMID:26305677

  13. Targeted or whole genome sequencing of formalin fixed tissue samples: potential applications in cancer genomics.

    PubMed

    Munchel, Sarah; Hoang, Yen; Zhao, Yue; Cottrell, Joseph; Klotzle, Brandy; Godwin, Andrew K; Koestler, Devin; Beyerlein, Peter; Fan, Jian-Bing; Bibikova, Marina; Chien, Jeremy

    2015-09-22

    Current genomic studies are limited by the poor availability of fresh-frozen tissue samples. Although formalin-fixed diagnostic samples are in abundance, they are seldom used in current genomic studies because of the concern of formalin-fixation artifacts. Better characterization of these artifacts will allow the use of archived clinical specimens in translational and clinical research studies. To provide a systematic analysis of formalin-fixation artifacts on Illumina sequencing, we generated 26 DNA sequencing data sets from 13 pairs of matched formalin-fixed paraffin-embedded (FFPE) and fresh-frozen (FF) tissue samples. The results indicate high rate of concordant calls between matched FF/FFPE pairs at reference and variant positions in three commonly used sequencing approaches (whole genome, whole exome, and targeted exon sequencing). Global mismatch rates and C · G > T · A substitutions were comparable between matched FF/FFPE samples, and discordant rates were low (<0.26%) in all samples. Finally, low-pass whole genome sequencing produces similar pattern of copy number alterations between FF/FFPE pairs. The results from our studies suggest the potential use of diagnostic FFPE samples for cancer genomic studies to characterize and catalog variations in cancer genomes.

  14. Targeted or whole genome sequencing of formalin fixed tissue samples: potential applications in cancer genomics.

    PubMed

    Munchel, Sarah; Hoang, Yen; Zhao, Yue; Cottrell, Joseph; Klotzle, Brandy; Godwin, Andrew K; Koestler, Devin; Beyerlein, Peter; Fan, Jian-Bing; Bibikova, Marina; Chien, Jeremy

    2015-09-22

    Current genomic studies are limited by the poor availability of fresh-frozen tissue samples. Although formalin-fixed diagnostic samples are in abundance, they are seldom used in current genomic studies because of the concern of formalin-fixation artifacts. Better characterization of these artifacts will allow the use of archived clinical specimens in translational and clinical research studies. To provide a systematic analysis of formalin-fixation artifacts on Illumina sequencing, we generated 26 DNA sequencing data sets from 13 pairs of matched formalin-fixed paraffin-embedded (FFPE) and fresh-frozen (FF) tissue samples. The results indicate high rate of concordant calls between matched FF/FFPE pairs at reference and variant positions in three commonly used sequencing approaches (whole genome, whole exome, and targeted exon sequencing). Global mismatch rates and C · G > T · A substitutions were comparable between matched FF/FFPE samples, and discordant rates were low (<0.26%) in all samples. Finally, low-pass whole genome sequencing produces similar pattern of copy number alterations between FF/FFPE pairs. The results from our studies suggest the potential use of diagnostic FFPE samples for cancer genomic studies to characterize and catalog variations in cancer genomes. PMID:26305677

  15. Simultaneous sampling of tissue oxygenation and oxygen consumption in skeletal muscle.

    PubMed

    Nugent, William H; Song, Bjorn K; Pittman, Roland N; Golub, Aleksander S

    2016-05-01

    Under physiologic conditions, microvascular oxygen delivery appears to be well matched to oxygen consumption in respiring tissues. We present a technique to measure interstitial oxygen tension (PISFO2) and oxygen consumption (VO2) under steady-state conditions, as well as during the transitions from rest to activity and back. Phosphorescence Quenching Microscopy (PQM) was employed with pneumatic compression cycling to achieve 1 to 10 Hz sampling rates of interstitial PO2 and simultaneous recurrent sampling of VO2 (3/min) in the exteriorized rat spinotrapezius muscle. The compression pressure was optimized to 120-130 mmHg without adverse effect on the tissue preparation. A cycle of 5s compression followed by 15s recovery yielded a resting VO2 of 0.98 ± 0.03 ml O2/100 cm(3)min while preserving microvascular oxygen delivery. The measurement system was then used to assess VO2 dependence on PISFO2 at rest and further tested under conditions of isometric muscle contraction to demonstrate a robust ability to monitor the on-kinetics of tissue respiration and the compensatory changes in PISFO2 during contraction and recovery. The temporal and spatial resolution of this approach is well suited to studies seeking to characterize microvascular oxygen supply and demand in thin tissues. PMID:26683232

  16. Analysis of dissected tissues with digital holographic microscopy: quantification of inflammation mediated tissue alteration, influence of sample preparation, and reliability of numerical autofocusing

    NASA Astrophysics Data System (ADS)

    Kemper, Björn; Lenz, Philipp; Bettenworth, Dominik; Krausewitz, Philipp; Domagk, Dirk; Ketelhut, Steffi

    2015-03-01

    Quantitative phase imaging with digital holographic microscopy (DHM) allows label-free imaging of tissue sections and quantification of the spatial refractive index distribution, which is of interest for applications in digital pathology. We show that DHM allows quantitative imaging of different layers in unstained tissue samples by detection of refractive index changes. In addition, we evaluate the automated refocussing feature of DHM for application on dissected tissues and could achieve highly reproducible holographic autofocusing for unstained and moderately stained samples. Finally, it is demonstrated that in human ulcerative colitis patients the average tissue refractive index is reduced significantly in all parts of the inflamed colonic wall in comparison to patients in remission.

  17. A pilot study of sampling subcutaneous adipose tissue to examine biomarkers of cancer risk.

    PubMed

    Campbell, Kristin L; Makar, Karen W; Kratz, Mario; Foster-Schubert, Karen E; McTiernan, Anne; Ulrich, Cornelia M

    2009-01-01

    Examination of adipose tissue biology may provide important insight into mechanistic links for the observed association between higher body fat and risk of several types of cancer, in particular colorectal and breast cancer. We tested two different methods of obtaining adipose tissue from healthy individuals. Ten overweight or obese (body mass index, 25-40 kg/m(2)), postmenopausal women were recruited. Two subcutaneous abdominal adipose tissue samples were obtained per individual (i.e., right and left lower abdominal regions) using two distinct methods (method A: 14-gauge needle with incision, versus method B: 16-gauge needle without incision). Gene expression was examined at the mRNA level for leptin, adiponectin, aromatase, interleukin 6 (IL-6), and tumor necrosis factor-alpha (TNF-alpha) in flash-frozen tissue, and at the protein level for leptin, adiponectin, IL-6, and TNF-alpha following short-term culture. Participants preferred biopsy method A and few participants reported any of the usual minor side effects. Gene expression was detectable for leptin, adiponectin, and aromatase, but was below detectable limits for IL-6 and TNF-alpha. For detectable genes, relative gene expression in adipose tissue obtained by methods A and B was similar for adiponectin (r = 0.64, P = 0.06) and leptin (r = 0.80, P = 0.01), but not for aromatase (r = 0.37,P = 0.34). Protein levels in tissue culture supernatant exhibited good intra-assay agreement [coefficient of variation (CV), 1-10%], with less agreement for intraindividual agreement (CV, 17-29%) and reproducibility, following one freeze-thaw cycle (CV, >14%). Subcutaneous adipose tissue biopsies from healthy, overweight individuals provide adequate amounts for RNA extraction, gene expression, and other assays of relevance to cancer prevention research. PMID:19139016

  18. Effects of sample preparation on the optical properties of breast tissue

    NASA Astrophysics Data System (ADS)

    Marks, Fay A.

    1996-04-01

    The optical properties of biological tissue should be determined in vivo whenever possible. However, for those instances when in vivo studies are impractical, too expensive or inappropriate, and when blood flow is not an issue, the ability to perform in vitro studies then becomes invaluable. Optical absorption spectroscopy shows that it may be possible to obtain meaningful information about the optical properties of human breast tissue from in vitro samples if strict preparation and measuring protocols are used. That a strict protocol for storing and handling tissue is critical can be seen from our observations of changes in the optical absorption spectra that occur in response to formalin fixation, the passage of time, application of stains and dyes, and storage in growth medium of the excised tissue. In vivo optical absorption spectroscopy measurements have been made on human breast cancer xenografts and compared with in vitro measurements on breast biopsies prepared according to precise collection and treatment protocols. There is a 'window of opportunity' before time dependent changes in the UV optical absorption spectra of the excised tissue specimens occur. This time window of opportunity widens at longer wavelengths with the least changes occurring in the optical spectra in the NIR.

  19. Transgenic zebrafish reveal tissue-specific differences in estrogen signaling in response to environmental water samples

    USGS Publications Warehouse

    Gorelick, Daniel A.; Iwanowicz, Luke R.; Hung, Alice L.; Blazer, Vicki; Halpern, Marnie E.

    2014-01-01

    Background: Environmental endocrine disruptors (EED) are exogenous chemicals that mimic endogenous hormones, such as estrogens. Previous studies using a zebrafish transgenic reporter demonstrated that the EEDs bisphenol A and genistein preferentially activate estrogen receptors (ER) in the larval heart compared to the liver. However, it was not known whether the transgenic zebrafish reporter was sensitive enough to detect estrogens from environmental samples, whether environmental estrogens would exhibit similar tissue-specific effects as BPA and genistein or why some compounds preferentially target receptors in the heart. Methods: We tested surface water samples using a transgenic zebrafish reporter with tandem estrogen response elements driving green fluorescent protein expression (5xERE:GFP). Reporter activation was colocalized with tissue-specific expression of estrogen receptor genes by RNA in situ hybridization. Results: Selective patterns of ER activation were observed in transgenic fish exposed to river water samples from the Mid-Atlantic United States, with several samples preferentially activating receptors in embryonic and larval heart valves. We discovered that tissue-specificity in ER activation is due to differences in the expression of estrogen receptor subtypes. ERα is expressed in developing heart valves but not in the liver, whereas ERβ2 has the opposite profile. Accordingly, subtype-specific ER agonists activate the reporter in either the heart valves or the liver. Conclusion: The use of 5xERE:GFP transgenic zebrafish has revealed an unexpected tissue-specific difference in the response to environmentally relevant estrogenic compounds. Exposure to estrogenic EEDs in utero is associated with adverse health effects, with the potentially unanticipated consequence of targeting developing heart valves.

  20. Molecular classification of cancer with the 92-gene assay in cytology and limited tissue samples

    PubMed Central

    Brachtel, Elena F.; Operaña, Theresa N.; Sullivan, Peggy S.; Kerr, Sarah E.; Cherkis, Karen A.; Schroeder, Brock E.; Dry, Sarah M.; Schnabel, Catherine A.

    2016-01-01

    Background Detailed molecular evaluation of cytology and limited tissue samples is increasingly becoming the standard for cancer care. Reproducible and accurate diagnostic approaches with reduced demands on cellularity are an ongoing unmet need. This study evaluated the performance of a 92-gene assay for molecular diagnosis of tumor type/subtype in cytology and limited tissue samples. Methods Clinical validation of accuracy for the 92-gene assay in limited tissue samples such as cytology cell blocks, core biopsies and small excisions was conducted in a blinded multi-institutional study (N = 109, 48% metastatic, 53% grade II and III). Analytical success rate and diagnostic utility were evaluated in a consecutive series of 644 cytology cases submitted for clinical testing. Results The 92-gene assay demonstrated 91% sensitivity (95% CI [0.84, 0.95]) for tumor classification, with high accuracy maintained irrespective of specimen type (100%, 92%, and 86% in FNA/cytology cell blocks, core biopsies, and small excisions, respectively; p = 0.26). The assay performed equally well for metastatic versus primary tumors (90% vs 93%, p = 0.73), and across histologic grades (100%, 90%, 89%, in grades I, II, and III, respectively; p = 0.75). In the clinical case series, a molecular diagnosis was reported in 87% of the 644 samples, identifying 23 different tumor types and allowing for additional mutational analysis in selected cases. Conclusions These findings demonstrate high accuracy and analytical success rate of the 92-gene assay, supporting its utility in the molecular diagnosis of cancer for specimens with limited tissue. PMID:27034010

  1. Quantitative laser-induced breakdown spectroscopy analysis of calcified tissue samples

    NASA Astrophysics Data System (ADS)

    Samek, O.; Beddows, D. C. S.; Telle, H. H.; Kaiser, J.; Liška, M.; Cáceres, J. O.; Gonzáles Ureña, A.

    2001-06-01

    We report on the application of laser-induced breakdown spectroscopy (LIBS) to the analysis of important minerals and the accumulation of potentially toxic elements in calcified tissue, to trace e.g. the influence of environmental exposure, and other medical or biological factors. This theme was exemplified for quantitative detection and mapping of Al, Pb and Sr in representative samples, including teeth (first teeth of infants, second teeth of children and teeth of adults) and bones (tibia and femur). In addition to identifying and quantifying major and trace elements in the tissues, one- and two-dimensional profiles and maps were generated. Such maps (a) provide time/concentration relations, (b) allow to follow mineralisation of the hydroxyapatite matrix and the migration of the elements within it and (c) enable to identify disease states, such as caries in teeth. In order to obtain quantitative calibration, reference samples in the form of pressed pellets with calcified tissue-equivalent material (majority compound of pellets is CaCO 3) were used whose physical properties closely resembled hydroxyapatite. Compounds of Al, Sr and Pb were added to the pellets, containing atomic concentrations in the range 100-10 000 ppm relative to the Ca content of the matrix. Analytical results based on this calibration against artificial samples for the trace elements under investigation agree with literature values, and with our atomic absorption spectroscopy (AAS) cross-validation measurements.

  2. Distribution of polybrominated diphenyl ethers in Japanese autopsy tissue and body fluid samples.

    PubMed

    Hirai, Tetsuya; Fujimine, Yoshinori; Watanabe, Shaw; Nakano, Takeshi

    2012-09-01

    Brominated flame retardants are components of many plastics and are used in products such as cars, textiles, televisions, and personal computers. Human exposure to polybrominated diphenyl ether (PBDE) flame retardants has increased exponentially during the last three decades. Our objective was to measure the body burden and distribution of PBDEs and to determine the concentrations of the predominant PBDE congeners in samples of liver, bile, adipose tissue, and blood obtained from Japanese autopsy cases. Tissues and body fluids obtained from 20 autopsy cases were analyzed. The levels of 25 PBDE congeners, ranging from tri- to hexa-BDEs, were assessed. The geometric means of the sum of the concentrations of PBDE congeners having detection frequencies >50 % (ΣPBDE) in the blood, liver, bile, and adipose tissue were 2.4, 2.6, 1.4, and 4.3 ng/g lipid, respectively. The most abundant congeners were BDE-47 and BDE-153, followed by BDE-100, BDE-99, and BDE-28+33. These concentrations of PBDE congeners were similar to other reports of human exposure in Japan but were notably lower than concentrations than those reported in the USA. Significant positive correlations were observed between the concentrations of predominant congeners and ΣPBDE among the samples analyzed. The ΣPBDE concentration was highest in the adipose tissue, but PBDEs were distributed widely among the tissues and body fluids analyzed. The PBDE levels observed in the present study are similar to those reported in previous studies in Japan and significantly lower than those reported in the USA. PMID:22544599

  3. Optimal transport time and conditions for cartilage tissue samples and expanded chondrocyte suspensions.

    PubMed

    Yilmaz, Banu Coskun; Yilmaz, Cengiz; Yilmaz, Necat S; Balli, Ebru; Tasdelen, Bahar

    2010-01-01

    For autologous chondrocyte implantation, the chondral tissue obtained is transferred from the operating room to the laboratory using specialized carrier systems within 24 hours. Similar expenses are used for the transport of cultured chondrocytes. The purpose of this study was to find the optimal temperature, size of tissue, and time that the chondrocytes can stand without losing viability and proliferative capacity. Fresh calf cartilage was harvested and divided into 24 groups. Half of the samples were diced into 1- to 2-mm(3) pieces. All 12 groups were kept at either 4 degrees C, 25 degrees C, or 37 degrees C for 1, 3, 5, or 7 days and were seeded for cell culture. Times to reach confluence values were compared. Produced cell suspensions were grouped similarly and tested similarly. Neither the temperature nor the waiting days caused any difference in the proliferative capacity of the cells. Diced tissues yielded a shorter time to reach confluence values. Chondral tissue obtained from the patient can be transferred to the laboratory at temperatures ranging from 4 degrees C to 37 degrees C in up to 7 days. These conditions did not affect the proliferative capacity or the viability of the chondrocytes. Dicing the tissue prior to transport will shorten total culturing time. The expanded cell suspensions should be transferred at temperatures from 4 degrees C to 25 degrees C within 3 days. Specialized carrier systems to get the chondral tissue from the operating room to the laboratory and to take the expanded chondrocytes back to the operating room within hours may not be necessary.

  4. Threshold-dependent sample sizes for selenium assessment with stream fish tissue

    USGS Publications Warehouse

    Hitt, Nathaniel P.; Smith, David

    2013-01-01

    Natural resource managers are developing assessments of selenium (Se) contamination in freshwater ecosystems based on fish tissue concentrations. We evaluated the effects of sample size (i.e., number of fish per site) on the probability of correctly detecting mean whole-body Se values above a range of potential management thresholds. We modeled Se concentrations as gamma distributions with shape and scale parameters fitting an empirical mean-to-variance relationship in data from southwestern West Virginia, USA (63 collections, 382 individuals). We used parametric bootstrapping techniques to calculate statistical power as the probability of detecting true mean concentrations up to 3 mg Se/kg above management thresholds ranging from 4-8 mg Se/kg. Sample sizes required to achieve 80% power varied as a function of management thresholds and type-I error tolerance (α). Higher thresholds required more samples than lower thresholds because populations were more heterogeneous at higher mean Se levels. For instance, to assess a management threshold of 4 mg Se/kg, a sample of 8 fish could detect an increase of ∼ 1 mg Se/kg with 80% power (given α = 0.05), but this sample size would be unable to detect such an increase from a management threshold of 8 mg Se/kg with more than a coin-flip probability. Increasing α decreased sample size requirements to detect above-threshold mean Se concentrations with 80% power. For instance, at an α-level of 0.05, an 8-fish sample could detect an increase of ∼ 2 units above a threshold of 8 mg Se/kg with 80% power, but when α was relaxed to 0.2 this sample size was more sensitive to increasing mean Se concentrations, allowing detection of an increase of ∼ 1.2 units with equivalent power. Combining individuals into 2- and 4-fish composite samples for laboratory analysis did not decrease power because the reduced number of laboratory samples was compensated by increased precision of composites for estimating mean

  5. Threshold-dependent sample sizes for selenium assessment with stream fish tissue.

    PubMed

    Hitt, Nathaniel P; Smith, David R

    2015-01-01

    Natural resource managers are developing assessments of selenium (Se) contamination in freshwater ecosystems based on fish tissue concentrations. We evaluated the effects of sample size (i.e., number of fish per site) on the probability of correctly detecting mean whole-body Se values above a range of potential management thresholds. We modeled Se concentrations as gamma distributions with shape and scale parameters fitting an empirical mean-to-variance relationship in data from southwestern West Virginia, USA (63 collections, 382 individuals). We used parametric bootstrapping techniques to calculate statistical power as the probability of detecting true mean concentrations up to 3 mg Se/kg above management thresholds ranging from 4 to 8 mg Se/kg. Sample sizes required to achieve 80% power varied as a function of management thresholds and Type I error tolerance (α). Higher thresholds required more samples than lower thresholds because populations were more heterogeneous at higher mean Se levels. For instance, to assess a management threshold of 4 mg Se/kg, a sample of eight fish could detect an increase of approximately 1 mg Se/kg with 80% power (given α=0.05), but this sample size would be unable to detect such an increase from a management threshold of 8 mg Se/kg with more than a coin-flip probability. Increasing α decreased sample size requirements to detect above-threshold mean Se concentrations with 80% power. For instance, at an α-level of 0.05, an 8-fish sample could detect an increase of approximately 2 units above a threshold of 8 mg Se/kg with 80% power, but when α was relaxed to 0.2, this sample size was more sensitive to increasing mean Se concentrations, allowing detection of an increase of approximately 1.2 units with equivalent power. Combining individuals into 2- and 4-fish composite samples for laboratory analysis did not decrease power because the reduced number of laboratory samples was compensated for by increased precision of composites

  6. Infrared microspectroscopy of benign and neoplastic prostate: correlation of spectral patterns with histopathology

    NASA Astrophysics Data System (ADS)

    Chiriboga, Luis; Diem, Max; Yee, Herman T.

    2000-05-01

    The diagnosis of prostate cancer is based on the visible microscopic evaluation of both cytological and architectural features of the prostate tissue sections. In order to determine whether IR spectral 'mapping' can be used to objectively distinguish between normal and neoplastic prostate tissue, a comparison between 'visual, point-by- point' and 'automated, point-by-point' IR measurements was performed. Automated, point-by-point analysis was performed without any prior diagnostic information. Visual, point-by- point measurements were based on histopathology, histochemistry and immunohistochemical analysis of the tissue samples. The spectra obtained from these measurements were compared to the spectra obtained from automated point- by-point analysis. Our results indicate that the spectra obtained from histopathologically directed measurements compares well with those of automated mapping methods. Therefore, we believe that current mapping methodology can be directly correlated with pathological diagnoses.

  7. Determination of benzophenones in human placental tissue samples by liquid chromatography-tandem mass spectrometry.

    PubMed

    Vela-Soria, F; Jiménez-Díaz, I; Rodríguez-Gómez, R; Zafra-Gómez, A; Ballesteros, O; Navalón, A; Vílchez, J L; Fernández, M F; Olea, N

    2011-09-30

    Benzophenones (BPs) are a family of compounds widely used to protect the skin and hair from UV irradiation. Despite human exposure to BPs through dermal application of products containing sunscreen agents and the increasing evidence that BPs are able to interfere with endocrine systems, few studies have examined the occurrence of BPs in humans. In this work, we propose a new liquid chromatography-tandem mass spectrometry (LC-MS/MS) method to determine six BPs, namely, benzophenone-1 (BP-1), benzophenone-2 (BP-2), benzophenone-3 (BP-3), benzophenone-6 (BP-6), benzophenone-8 (BP-8) and 4-hydroxybenzophenone (4-OH-BP) in human placental tissue samples. The method involves an extraction step of the analytes from the samples using ethyl acetate, followed by a clean-up step using centrifugation prior to their quantification by LC-MS/MS using an atmospheric pressure chemical ionization (APCI) interface in the positive mode. Benzophenone-d(10) (BP-d(10)) was used as surrogate. Found detection limits (LOD) ranged from 0.07 to 0.3 ng g(-1) and quantification limits (LOQ) from 0.3 to 1.0 ng g(-1), while inter- and intra-day variability was under 5%. The method was validated using standard addition calibration and a recovery assay. Recovery rates for spiked samples ranged from 98 to 104%. This method was satisfactorily applied for the determination of BPs in 16 placental tissue samples collected from women who live in Granada (Spain).

  8. The relationship between decorrelation time and sample thickness in acute rat brain tissue slices (Conference Presentation)

    NASA Astrophysics Data System (ADS)

    Brake, Joshua; Jang, Mooseok; Yang, Changhuei

    2016-03-01

    The optical opacity of biological tissue has long been a challenge in biomedical optics due to the strong scattering nature of tissue in the optical regime. While most conventional optical techniques attempt to gate out multiply scattered light and use only unscattered light, new approaches in the field of wavefront shaping exploit the time reversible symmetry of optical scattering in order to focus light inside or through scattering media. While these approaches have been demonstrated effectively on static samples, it has proven difficult to apply them to dynamic biological samples since even small changes in the relative positions of the scatterers within will cause the time symmetry that wavefront shaping relies upon to decorrelate. In this paper we investigate the decorrelation curves of acute rat brain slices for thicknesses in the range 1-3 mm (1/e decorrelation time on the order of seconds) using multi-speckle diffusing wave spectroscopy (MSDWS) and compare the results with theoretical predictions. The results of this study demonstrate that the 1/L^2 relationship between decorrelation time and thickness predicted by diffusing wave spectroscopy provides a good rule of thumb for estimating how the decorrelation of a sample will change with increasing thickness. Understanding this relationship will provide insight to guide the future development of biophotonic wavefront shaping tools by giving an estimate of how fast wavefront shaping systems need to operate to overcome the dynamic nature of biological samples.

  9. A single lysis solution for the analysis of tissue samples by different proteomic technologies.

    PubMed

    Gromov, Pavel; Celis, Julio E; Gromova, Irina; Rank, Fritz; Timmermans-Wielenga, Vera; Moreira, José M A

    2008-12-01

    Cancer, being a major healthcare concern worldwide, is one of the main targets for the application of emerging proteomic technologies and these tools promise to revolutionize the way cancer will be diagnosed and treated in the near future. Today, as a result of the unprecedented advances that have taken place in molecular biology, cell biology and genomics there is a pressing need to accelerate the translation of basic discoveries into clinical applications. This need, compounded by mounting evidence that cellular model systems are unable to fully recapitulate all biological aspects of human dissease, is driving scientists to increasingly use clinically relevant samples for biomarker and target discovery. Tissues are heterogeneous and as a result optimization of sample preparation is critical for generating accurate, representative, and highly reproducible quantitative data. Although a large number of protocols for preparation of tissue lysates has been published, so far no single recipe is able to provide a "one-size fits all" solubilization procedure that can be used to analyse the same lysate using different proteomics technologies. Here we present evidence showing that cell lysis buffer 1 (CLB1), a lysis solution commercialized by Zeptosens [a division of Bayer (Schweiz) AG], provides excellent sample solubilization and very high 2D PAGE protein resolution both when using carrier ampholytes and immobilized pH gradient strips. Moreover, this buffer can also be used for array-based proteomics (reverse-phase lysate arrays or direct antibody arrays), allowing the direct comparison of qualitative and quantitative data yielded by these technologies when applied to the same samples. The usefulness of the CLB1 solution for gel-based proteomics was further established by 2D PAGE analysis of a number of technically demanding specimens such as breast carcinoma core needle biopsies and problematic tissues such as brain cortex, cerebellum, skeletal muscle, kidney cortex and

  10. Determination of optical properties of oxidative bleaching human dental tissue samples using optical coherence tomography

    NASA Astrophysics Data System (ADS)

    Ni, Y. R.; Guo, Z. Y.; Shu, S. Y.; Zeng, C. C.; Zhong, H. Q.; Chen, B. L.; Liu, Z. M.; Bao, Y.

    2011-10-01

    Oxidative bleaching changes of human teeth induced changes in the optical properties of dental tissue. We introduced 1310 nm wavelengths of optical coherence tomography (OCT) attenuation coefficient method which is a relatively novel and rarely reported methodology to measure the correlation coefficient during the teeth oxidative bleaching procedure. And the quantitative parameters of enamel optical thickness and disruption of the entrance signal (DES) were extracted from the OCT images. The attenuation coefficient of the bleached tissue is 6.2 mm-1 which is significant (p < 0.001) higher than that unbleached sample is 1.4 mm-1. But attenuation coefficient varied significantly (p < 0.001) between 5.9 and 1.5 mm-1 in dentine which is downtrend. Furthermore, the persistence of bleaching oxidation in 35% hydrogen peroxide-induced optical thickness of enamel is similar with unbleached tissue which may indicate the refractive index of enamel is unchanged. Moreover, disruption of the entrance signal (DES) analysis showed that remarkable difference was appeared at enamel surface. The results indicate that optical properties of oxidative bleaching human dental tissue can be determined by attenuation coefficient using OCT system.

  11. Quantified Histopathology of the Keratoconic Cornea

    PubMed Central

    Mathew, Jessica H.; Goosey, John D.; Bergmanson, Jan P. G.

    2011-01-01

    Purpose The present study systematically investigated and quantified histopathological changes in a series of keratoconic (Kc) corneas utilizing a physiologically formulated fixative to not further distort the already distorted diseased corneas. Methods Twelve surgically removed Kc corneal buttons were immediately preserved and processed for light and transmission electron microscopy using an established corneal protocol. Measurements were taken from the central cone and peripheral regions of the host button. The sample size examined ranged in length from 390–2608um centrally and 439–2242um peripherally. Results The average corneal thickness was 437um centrally and 559um peripherally. Epithelial thickness varied centrally from 14–92um and peripherally from 30–91um. A marked thickening of the epithelial basement membrane was noted in 58% of corneas. Centrally, anterior limiting lamina (ALL) was thinned or lost over 60% of the area examined, while peripheral cornea was also affected, but to a lesser extent. Histopathologically, posterior cornea remained undisturbed by the disease. Anteriorly in the stroma, an increased number of cells and tissue debris were encountered and some of these cells were clearly not keratocytes. Conclusions It is concluded that Kc pathology, at least initially, has a distinct anterior focus involving the epithelium, ALL and anterior stroma. The epithelium had lost its cellular uniformity and was compromised by the loss or damage to the ALL. The activity of the hitherto unreported recruited stromal cells may be to break down and remove ALL and anterior stromal lamellae leading to the overall thinning that accompanies this disease. PMID:21623252

  12. Molecular strain identification of the Mycobacterium tuberculosis complex in archival tissue samples

    PubMed Central

    Zink, A R; Nerlich, A G

    2004-01-01

    Aims: To investigate the use of different molecular analyses that can identify distinct strains of human pathogenic mycobacteria in formalin fixed and paraffin wax embedded archival tissue samples to see whether it is possible to differentiate between the members of the Mycobacterium tuberculosis complex (M tuberculosis, M bovis, M africanum, M microti, or M canettii) and/or substrains in a high number of samples. This would be of interest for identifying individual infection traits and superinfection by different mycobacterial strains. Methods: Forty nine archival tissue samples with clinically and/or histologically suspected tuberculosis infection were subjected to molecular DNA analysis. Results: The molecular analysis revealed the presence of M tuberculosis complex DNA in 20 samples, whereas acid fast bacilli could be detected by Ziehl-Neelsen staining in only eight samples. All IS6110 positive samples were further characterised by spoligotyping and seven cases provided M tuberculosis specific signatures, whereas M bovis specific signatures were obtained in four cases. The analysis of mtp40, oxyR, and pncA partial gene sequences confirmed the presence of M tuberculosis in six cases and M bovis in one case. The amplification and sequencing of four further genetic regions (katG, gyrA, TbD1, RD9) characterised six “modern” M tuberculosis strains belonging to genetic groups 2 or 3. Conclusion: This study provides clear evidence that archival paraffin wax embedded material can be used for further studies on the strain identification of M tuberculosis complex strains and can therefore unequivocally be used for the study of the epidemiology and evolution of tuberculosis pathogens. PMID:15509681

  13. A workflow to preserve genome-quality tissue samples from plants in botanical gardens and arboreta1

    PubMed Central

    Gostel, Morgan R.; Kelloff, Carol; Wallick, Kyle; Funk, Vicki A.

    2016-01-01

    Premise of the study: Internationally, gardens hold diverse living collections that can be preserved for genomic research. Workflows have been developed for genomic tissue sampling in other taxa (e.g., vertebrates), but are inadequate for plants. We outline a workflow for tissue sampling intended for two audiences: botanists interested in genomics research and garden staff who plan to voucher living collections. Methods and Results: Standard herbarium methods are used to collect vouchers, label information and images are entered into a publicly accessible database, and leaf tissue is preserved in silica and liquid nitrogen. A five-step approach for genomic tissue sampling is presented for sampling from living collections according to current best practices. Conclusions: Collecting genome-quality samples from gardens is an economical and rapid way to make available for scientific research tissue from the diversity of plants on Earth. The Global Genome Initiative will facilitate and lead this endeavor through international partnerships. PMID:27672517

  14. A workflow to preserve genome-quality tissue samples from plants in botanical gardens and arboreta1

    PubMed Central

    Gostel, Morgan R.; Kelloff, Carol; Wallick, Kyle; Funk, Vicki A.

    2016-01-01

    Premise of the study: Internationally, gardens hold diverse living collections that can be preserved for genomic research. Workflows have been developed for genomic tissue sampling in other taxa (e.g., vertebrates), but are inadequate for plants. We outline a workflow for tissue sampling intended for two audiences: botanists interested in genomics research and garden staff who plan to voucher living collections. Methods and Results: Standard herbarium methods are used to collect vouchers, label information and images are entered into a publicly accessible database, and leaf tissue is preserved in silica and liquid nitrogen. A five-step approach for genomic tissue sampling is presented for sampling from living collections according to current best practices. Conclusions: Collecting genome-quality samples from gardens is an economical and rapid way to make available for scientific research tissue from the diversity of plants on Earth. The Global Genome Initiative will facilitate and lead this endeavor through international partnerships.

  15. Liquid microjunction surface sampling of acetaminophen, terfenadine and their metabolites in thin tissue sections

    SciTech Connect

    Kertesz, Vilmos; Paranthaman, Nithya; Moench, Paul; Catoire, Alexandre; Flarakos, Jimmy; Van Berkel, Gary J.

    2014-10-01

    The aim of this paper was to evaluate the analytical performance of a fully automated droplet-based surface-sampling system for determining the distribution of the drugs acetaminophen and terfenadine, and their metabolites, in rat thin tissue sections. The following are the results: The rank order of acetaminophen concentration observed in tissues was stomach > small intestine > liver, while the concentrations of its glucuronide and sulfate metabolites were greatest in the liver and small intestine. Terfenadine was most concentrated in the liver and kidney, while its major metabolite, fexofenadine, was found in the liver and small intestine. In conclusion, the spatial distributions of both drugs and their respective metabolites observed in this work were consistent with previous studies using radiolabeled drugs.

  16. Liquid microjunction surface sampling of acetaminophen, terfenadine and their metabolites in thin tissue sections

    DOE PAGES

    Kertesz, Vilmos; Paranthaman, Nithya; Moench, Paul; Catoire, Alexandre; Flarakos, Jimmy; Van Berkel, Gary J.

    2014-10-01

    The aim of this paper was to evaluate the analytical performance of a fully automated droplet-based surface-sampling system for determining the distribution of the drugs acetaminophen and terfenadine, and their metabolites, in rat thin tissue sections. The following are the results: The rank order of acetaminophen concentration observed in tissues was stomach > small intestine > liver, while the concentrations of its glucuronide and sulfate metabolites were greatest in the liver and small intestine. Terfenadine was most concentrated in the liver and kidney, while its major metabolite, fexofenadine, was found in the liver and small intestine. In conclusion, the spatialmore » distributions of both drugs and their respective metabolites observed in this work were consistent with previous studies using radiolabeled drugs.« less

  17. Determination of /sup 35/S-aminoacyl-transfer ribonucleic acid specific radioactivity in small tissue samples

    SciTech Connect

    Samarel, A.M.; Ogunro, E.A.; Ferguson, A.G.; Lesch, M.

    1981-11-15

    Rate determination of protein synthesis utilizing tracer amino acid incorporation requires accurate assessment of the specific radioactivity of the labeled precursor aminoacyl-tRNA pool. Previously published methods presumably useful for the measurement of any aminoacyl-tRNA were unsuccessful when applied to (/sup 35/S)methionine, due to the unique chemical properties of this amino acid. Herein we describe modifications of these methods necessary for the measurement of /sup 35/S-aminoacyl-tRNA specific radioactivity from small tissue samples incubated in the presence of (/sup 35/S)methionine. The use of (/sup 35/S)methionine of high specific radioactivity enables analysis of the methionyl-tRNA from less than 100 mg of tissue. Conditions for optimal recovery of /sup 35/S-labeled dansyl-amino acid derivatives are presented and possible applications of this method are discussed.

  18. Report filing in histopathology.

    PubMed Central

    Blenkinsopp, W K

    1977-01-01

    An assessment of alternative methods of filing histopathology report forms in alphabetical order showed that orthodox card index filing is satisfactory up to about 100000 reports but, because of the need for long-term retrieval, when the reports filed exceed this number they should be copied on jacketed microfilm and a new card index file begun. PMID:591645

  19. Evaluation of sample holders designed for long-lasting X-ray micro-tomographic scans of ex-vivo soft tissue samples

    NASA Astrophysics Data System (ADS)

    Dudak, J.; Zemlicka, J.; Krejci, F.; Karch, J.; Patzelt, M.; Zach, P.; Sykora, V.; Mrzilkova, J.

    2016-03-01

    X-ray microradiography and microtomography are imaging techniques with increasing applicability in the field of biomedical and preclinical research. Application of hybrid pixel detector Timepix enables to obtain very high contrast of low attenuating materials such as soft biological tissue. However X-ray imaging of ex-vivo soft tissue samples is a difficult task due to its structural instability. Ex-vivo biological tissue is prone to fast drying-out which is connected with undesired changes of sample size and shape producing later on artefacts within the tomographic reconstruction. In this work we present the optimization of our Timepix equipped micro-CT system aiming to maintain soft tissue sample in stable condition. Thanks to the suggested approach higher contrast of tomographic reconstructions can be achieved while also large samples that require detector scanning can be easily measured.

  20. Postmortem interval alters the water relaxation and diffusion properties of rat nervous tissue--implications for MRI studies of human autopsy samples.

    PubMed

    Shepherd, Timothy M; Flint, Jeremy J; Thelwall, Peter E; Stanisz, Greg J; Mareci, Thomas H; Yachnis, Anthony T; Blackband, Stephen J

    2009-02-01

    High-resolution imaging of human autopsy tissues may improve our understanding of in vivo MRI findings, but interpretation is complicated because samples are obtained by immersion fixation following a postmortem interval (PMI). This study tested the hypotheses that immersion fixation and PMI's from 0-24 h would alter the water relaxation and diffusion properties in rat cortical slice and spinal cord models of human nervous tissue. Diffusion data collected from rat cortical slices at multiple diffusion times (10-60 ms) and b-values (7-15,000 s/mm(2)) were analyzed using a two-compartment model with exchange. Rat spinal cords were characterized with standard diffusion tensor imaging (21 directions, b=1250 s/mm(2)). Switching from perfusion- to immersion-fixation at 0 h PMI altered most MRI properties of rat cortical slices and spinal cords, including a 22% decrease in fractional anisotropy (P<0.001). After 4 h PMI, cortical slice T(1) and T(2) increased 22% and 65% respectively (P<0.001), transmembrane water exchange decreased 23% (P<0.001) and intracellular proton fraction increased 25% (P=0.002). After 6 h PMI, spinal cord white matter fractional anisotropy had decreased 38% (P<0.001). MRI property changes were observed for PMIs up to 24 h. The MRI changes correlated with protease activity and histopathological signs of autolysis. Thus, immersion fixation and/or even short PMIs (4-6 h) altered the MRI properties of rat nervous tissue. This suggests comparisons between in vivo clinical MRI and MRI data from human autopsy tissues should be interpreted with caution. PMID:18996206

  1. Microscopy and elemental analysis in tissue samples using computed microtomography with synchrotron x-rays

    SciTech Connect

    Spanne, P.; Rivers, M.L.

    1988-01-01

    The initial development shows that CMT using synchrotron x-rays can be developed to ..mu..m spatial resolution and perhaps even better. This creates a new microscopy technique which is of special interest in morphological studies of tissues, since no chemical preparation or slicing of the sample is necessary. The combination of CMT with spatial resolution in the ..mu..m range and elemental mapping with sensitivity in the ppM range results in a new tool for elemental mapping at the cellular level. 7 refs., 1 fig.

  2. Histopathological features of tungiasis in Peru.

    PubMed

    Maco, Vicente; Maco, Vicente P; Tantalean, Manuel E; Gotuzzo, Eduardo

    2013-06-01

    Tungiasis is an ectoparasitic skin disease caused by Tunga penetrans and Tunga trimamillata. There is a lack of histopathological studies that evaluate the recognition of this flea in tissues. We describe the ex vivo dermoscopic and the histopathological patterns of six cases and relate the findings to the developmental stage of the parasite as defined by the Fortaleza classification: two were classified as Fortaleza 3b, 3 as 4a, and 1 as 4b. Two dermoscopic patterns were observed: a brown pigmented ring and a radial crown with a central pore. The most common histopathological findings were an eosinophilic cuticle, eggs in different stages of development, tracheal rings (parasite), and basal hyperplasia (host). The eosinophilic cuticle, eggs in different stages of evolution, and tracheal rings can help to establish the diagnosis when other parts of the parasite are lacking. The Fortaleza staging may represent a tool for pathology reporting purposes. PMID:23478579

  3. Histopathological Features of Tungiasis in Peru

    PubMed Central

    Maco, Vicente; Maco, Vicente P.; Tantalean, Manuel E.; Gotuzzo, Eduardo

    2013-01-01

    Tungiasis is an ectoparasitic skin disease caused by Tunga penetrans and Tunga trimamillata. There is a lack of histopathological studies that evaluate the recognition of this flea in tissues. We describe the ex vivo dermoscopic and the histopathological patterns of six cases and relate the findings to the developmental stage of the parasite as defined by the Fortaleza classification: two were classified as Fortaleza 3b, 3 as 4a, and 1 as 4b. Two dermoscopic patterns were observed: a brown pigmented ring and a radial crown with a central pore. The most common histopathological findings were an eosinophilic cuticle, eggs in different stages of development, tracheal rings (parasite), and basal hyperplasia (host). The eosinophilic cuticle, eggs in different stages of evolution, and tracheal rings can help to establish the diagnosis when other parts of the parasite are lacking. The Fortaleza staging may represent a tool for pathology reporting purposes. PMID:23478579

  4. Histopathology of Lyme arthritis in LSH hamsters

    SciTech Connect

    Hejka, A.; Schmitz, J.L.; England, D.M.; Callister, S.M.; Schell, R.F.

    1989-05-01

    The authors studied the histopathologic evolution of arthritis in nonirradiated and irradiated hamsters infected with Borrelia burgdorferi. Nonirradiated hamsters injected in the hind paws with B. burgdorferi developed an acute inflammatory reaction involving the synovium, periarticular soft tissues, and dermis. This acute inflammatory reaction was short-lived and was replaced by a mild chronic synovitis as the number of detectable spirochetes in the synovium, periarticular soft tissues, and perineurovascular areas diminished. Exposing hamsters to radiation before inoculation with B. burgdorferi exacerbated and prolonged the acute inflammatory phase. Spirochetes also persisted longer in the periarticular soft tissues. A major histopathologic finding was destructive and erosive bone changes of the hind paws, which resulted in deformation of the joints. These studies should be helpful in defining the immune mechanism participating in the onset, progression, and resolution of Lyme arthritis.

  5. Assessment of bioburden on human and animal tissues: part 2--results of testing of human tissue and qualification of a composite sample for routine bioburden determination.

    PubMed

    Kowalski, John B; Merritt, Karen; Gocke, David; Osborne, Joel

    2012-08-01

    A quantitative method was developed and validated to assess bioburden on tissue from human donors and to compare bioburden determination results to swab culture results from the same donor. An initial study with allograft tissue from 101 donors showed a wide range of bioburden levels; values from no colony-forming units (CFU) detected to >28,000 CFU were observed. Tissues from donors that had swab cultures negative for objectionable microorganisms generally had lower bioburden than tissues from donors where objectionable microorganisms were recovered by swab culturing. In a follow-up study with 1,445 donors, a wide range of bioburden levels was again observed on tissues from donors that were swab culture negative for objectionable microorganisms. Tissues from 885 (61%) of these donors had no recoverable bioburden (<2 CFU). Importantly, tissues from 560 (39%) of the donors had recoverable bioburden which ranged from 1 to >24,000 CFU. Identification of bioburden isolates showed a diversity of genera and species. In compliance with the recent revision of the American Association of Tissue Banks K2.210 Standard, the quantitative bioburden determination method was validated with a composite tissue sample that contains bone and soft tissue sections tested together in one extraction vessel. A recovery efficiency of 68% was validated and the composite sample was shown to be representative of all of the tissues recovered from a donor. The use of the composite sample in conjunction with the quantitative bioburden determination method will facilitate an accurate assessment of the numbers and types of contaminating microorganisms on allografts prior to disinfection/sterilization. This information will ensure that disinfection/sterilization processes are properly validated and the capability of the overall allograft process is understood on a donor by donor basis.

  6. METHODS FOR USING 3-D ULTRASOUND SPECKLE TRACKING IN BIAXIAL MECHANICAL TESTING OF BIOLOGICAL TISSUE SAMPLES

    PubMed Central

    Yap, Choon Hwai; Park, Dae Woo; Dutta, Debaditya; Simon, Marc; Kim, Kang

    2014-01-01

    Being multilayered and anisotropic, biological tissues such as cardiac and arterial walls are structurally complex, making full assessment and understanding of their mechanical behavior challenging. Current standard mechanical testing uses surface markers to track tissue deformations and does not provide deformation data below the surface. In the study described here, we found that combining mechanical testing with 3-D ultrasound speckle tracking could overcome this limitation. Rat myocardium was tested with a biaxial tester and was concurrently scanned with high-frequency ultrasound in three dimensions. The strain energy function was computed from stresses and strains using an iterative non-linear curve-fitting algorithm. Because the strain energy function consists of terms for the base matrix and for embedded fibers, spatially varying fiber orientation was also computed by curve fitting. Using finite-element simulations, we first validated the accuracy of the non-linear curve-fitting algorithm. Next, we compared experimentally measured rat myocardium strain energy function values with those in the literature and found a matching order of magnitude. Finally, we retained samples after the experiments for fiber orientation quantification using histology and found that the results satisfactorily matched those computed in the experiments. We conclude that 3-D ultrasound speckle tracking can be a useful addition to traditional mechanical testing of biological tissues and may provide the benefit of enabling fiber orientation computation. PMID:25616585

  7. Persistent synthetic chlorinated hydrocarbons in albatross tissue samples from Midway Atoll

    SciTech Connect

    Jones, P.D.; Hannah, D.J.; Buckland, S.J.

    1996-10-01

    Anthropogenic organic contaminants have been found in even the most remote locations. To assess the global distribution and possible effects of such contaminants, the authors examined the tissues of two species of albatross collected from Midway Atoll in the central North Pacific Ocean. These birds have an extensive feeding range covering much of the subtropical and northern Pacific Ocean. Anthropogenic contaminants were found at relatively great concentrations in these birds. The sum of 19 polychlorinated biphenyl (PCB) congeners ranged from 177 ng/g wet weight in eggs to 2,750 ng/g wet weight in adult fat. Total toxic equivalents (TEQs) derived from polychlorinated dibenzo-p-dioxins (PCDDs) and dibenzofurans (PCDFs) ranged from 17.2 to 297 pg/g wet weight in the same tissues, while the inclusion of TEQs from PCBs increased these values to 48.4 and 769 pg/g wet weight, respectively. While contaminant concentrations varied between species and tissues, the contaminant profile was relatively uniform. The profile of contaminants detected was unusual in that much of the TEQs was contributed by two pentachlorinated congeners (2,3,4,7,8-pentachlorinated dibenzo-p-dioxin), and the profiles of PCB congeners did not match known sources. When compared to other studies the concentrations detected in the Midway Atoll samples were near or above the thresholds known to cause adverse effects in other fish-eating bird species.

  8. MicroRNA Profiling from RSV-Infected Biofluids, Whole Blood, and Tissue Samples.

    PubMed

    Anderson, Lydia; Jorquera, Patricia A; Tripp, Ralph A

    2016-01-01

    Several studies have shown that respiratory syncytial virus (RSV) can modulate the host innate immune response by dysregulation of host microRNAs (miRNAs) related to the antiviral response, a feature that also affects the memory immune response to RSV (Thornburg et al. MBio 3(6), 2012). miRNAs are small, endogenous, noncoding RNAs that function in posttranscriptional gene regulation. Here, we explain a compilation of methods for the purification, quantification, and characterization of miRNA expression profiles in biofluids, whole blood samples, and tissue samples obtained from in vivo studies. In addition, this chapter describes methods for the isolation of exosomal miRNA populations. Understanding alterations in miRNA expression profiles and identifying miRNA targets genes, and their contribution to the pathogenesis of RSV, may help elucidate novel mechanism of host-virus interaction (Rossi et al., Pediatr Pulmonol, 2015). PMID:27464696

  9. Variable ploidy of ovarian clear cell carcinomas. Implications for adequacy of tissue sampling.

    PubMed

    Listinsky, C M; Bonfiglio, T A; Leary, J

    1988-02-01

    Ten cases of clear cell (mesonephroid) adenocarcinoma of the ovary were examined for (1) variations of morphology within each tumor and its metastases, (2) ploidy of each morphologic region and (3) clinical behavior. Correlations were sought among these factors. Analysis of the ploidy in up to six morphologic regions per tumor showed variations in the ploidy in seven of the ten cases, with all seven having both diploid and nondiploid regions. The presence or absence of abnormal ploidy was not predictable based on the histomorphologic appearance of a given section. These results suggest that (1) the evaluation of a single random tissue sample may not discover aneuploidy that is present and (2) future ploidy studies on malignant tumors may require extensive tumor sampling in order to definitively exclude the presence of aneuploid populations.

  10. Rapid microarray-based genotyping of Chlamydia spp. strains from clinical tissue samples.

    PubMed

    Sachse, Konrad; Ruettger, Anke

    2015-01-01

    Pathogenic Chlamydia (C.) psittaci and C. trachomatis strains can be genotyped based on variations in the ompA genomic locus. In the present chapter, we describe rapid genotyping assays for both chlamydial agents using the ArrayStrip™ (AS) microarray platform. The test is targeting multiple discriminatory sites in the variable domains of the ompA gene by using 35 (C. psittaci) and 61 (C. trachomatis) oligonucleotide probes representing genotype-specific polymorphisms. In addition to discrimination among the established genotypes, this approach allows identification of atypical strains that were not accessible to typing using previously established techniques, such as PCR-RFLP or serotyping. The present DNA microarray assay can be conducted directly on clinical tissue samples and is suitable for tracing epidemiological chains and exploring the dissemination of particular genotypes. The procedure is easy to handle and economically affordable, and it allows genotyping of up to 32 clinical samples per day, thus lending itself for routine diagnosis as well.

  11. Quality and safety aspects in histopathology laboratory

    PubMed Central

    Adyanthaya, Soniya; Jose, Maji

    2013-01-01

    Histopathology is an art of analyzing and interpreting the shapes, sizes and architectural patterns of cells and tissues within a given specific clinical background and a science by which the image is placed in the context of knowledge of pathobiology, to arrive at an accurate diagnosis. To function effectively and safely, all the procedures and activities of histopathology laboratory should be evaluated and monitored accurately. In histopathology laboratory, the concept of quality control is applicable to pre-analytical, analytical and post-analytical activities. Ensuring safety of working personnel as well as environment is also highly important. Safety issues that may come up in a histopathology lab are primarily those related to potentially hazardous chemicals, biohazardous materials, accidents linked to the equipment and instrumentation employed and general risks from electrical and fire hazards. This article discusses quality management system which can ensure quality performance in histopathology laboratory. The hazards in pathology laboratories and practical safety measures aimed at controlling the dangers are also discussed with the objective of promoting safety consciousness and the practice of laboratory safety. PMID:24574660

  12. Quality and safety aspects in histopathology laboratory.

    PubMed

    Adyanthaya, Soniya; Jose, Maji

    2013-09-01

    Histopathology is an art of analyzing and interpreting the shapes, sizes and architectural patterns of cells and tissues within a given specific clinical background and a science by which the image is placed in the context of knowledge of pathobiology, to arrive at an accurate diagnosis. To function effectively and safely, all the procedures and activities of histopathology laboratory should be evaluated and monitored accurately. In histopathology laboratory, the concept of quality control is applicable to pre-analytical, analytical and post-analytical activities. Ensuring safety of working personnel as well as environment is also highly important. Safety issues that may come up in a histopathology lab are primarily those related to potentially hazardous chemicals, biohazardous materials, accidents linked to the equipment and instrumentation employed and general risks from electrical and fire hazards. This article discusses quality management system which can ensure quality performance in histopathology laboratory. The hazards in pathology laboratories and practical safety measures aimed at controlling the dangers are also discussed with the objective of promoting safety consciousness and the practice of laboratory safety.

  13. Feasibility of direct digital sampling for diffuse optical frequency domain spectroscopy in tissue

    NASA Astrophysics Data System (ADS)

    Roblyer, Darren; O'Sullivan, Thomas D.; Warren, Robert V.; Tromberg, Bruce J.

    2013-04-01

    Frequency domain optical spectroscopy in the diffusive regime is currently being investigated for biomedical applications including tumor detection, therapy monitoring, exercise metabolism and others. Analog homodyne or heterodyne detection of sinusoidally modulated signals has been the predominant method for measuring phase and amplitude of photon density waves that have traversed through tissue. Here we demonstrate the feasibility of utilizing direct digital sampling of modulated signals using a 3.6 gigasample/second 12 bit analog to digital converter. Digitally synthesized modulated signals between 50 MHz and 400 MHz were measured on tissue-simulating phantoms at six near-infrared wavelengths. An amplitude and phase precision of 1% and 0.6° were achieved during drift tests. Amplitude, phase, scattering and absorption values were compared with a well-characterized network analyzer-based diffuse optical device. Optical properties measured with both systems were within 3.6% for absorption and 2.8% for scattering over a range of biologically relevant values. Direct digital sampling represents a viable method for frequency domain diffuse optical spectroscopy and has the potential to reduce system complexity, size and cost.

  14. Evaluation of convenient pretreatment protocols for RNA virus metagenomics in serum and tissue samples.

    PubMed

    Rosseel, Toon; Ozhelvaci, Orkun; Freimanis, Graham; Van Borm, Steven

    2015-09-15

    Viral metagenomic approaches are increasingly being used for viral discovery. Various strategies are applied to enrich viral sequences, but there is often a lack of knowledge about their effective influence on the viral discovery sensitivity. We evaluate some convenient and widely used approaches for RNA virus discovery in clinical samples in order to reveal their sensitivity and potential bias introduced by the enrichment or amplifications steps. An RNA virus was artificially spiked at a fixed titer in serum and lung tissue, respectively, low and high nucleic acid content matrices. For serum, a simple DNase treatment on the RNA extract gave the maximum gain in proportion of viral sequences (83×), and a subsequent ribosomal RNA removal nearly doubled once more the proportion of viral sequences. For lung tissue, a ribosomal RNA depletion step on the RNA extract had the biggest gain in proportion of viral sequences (32×). We show also that direct sequencing of cDNA is recommended above an extra random PCR amplification step, and a that the virion enrichment strategy (filtration and nuclease treatment) has a beneficial effect for sequencing-based virus discovery. Our findings provide sample-dependent guidelines for targeted virus discovery strategies.

  15. Protocol: High-throughput and quantitative assays of auxin and auxin precursors from minute tissue samples

    PubMed Central

    2012-01-01

    Background The plant hormone auxin, indole-3-acetic acid (IAA), plays important roles in plant growth and development. The signaling response to IAA is largely dependent on the local concentration of IAA, and this concentration is regulated by multiple mechanisms in plants. Therefore, the precise quantification of local IAA concentration provides insights into the regulation of IAA and its biological roles. Meanwhile, pathways and genes involved in IAA biosynthesis are not fully understood, so it is necessary to analyze the production of IAA at the metabolite level for unbiased studies of IAA biosynthesis. Results We have developed high-throughput methods to quantify plant endogenous IAA and its biosynthetic precursors including indole, tryptophan, indole-3-pyruvic acid (IPyA), and indole-3-butyric acid (IBA). The protocol starts with homogenizing plant tissues with stable-labeled internal standards added, followed by analyte purification using solid phase extraction (SPE) tips and analyte derivatization. The derivatized analytes are finally analyzed by selected reaction monitoring on a gas chromatograph-mass spectrometer (GC-MS/MS) to determine the precise abundance of analytes. The amount of plant tissue required for the assay is small (typically 2–10 mg fresh weight), and the use of SPE tips is simple and convenient, which allows preparation of large sets of samples within reasonable time periods. Conclusions The SPE tips and GC-MS/MS based method enables high-throughput and accurate quantification of IAA and its biosynthetic precursors from minute plant tissue samples. The protocol can be used for measurement of these endogenous compounds using isotope dilution, and it can also be applied to analyze IAA biosynthesis and biosynthetic pathways using stable isotope labeling. The method will potentially advance knowledge of the role and regulation of IAA. PMID:22883136

  16. Liquid Microjunction Surface Sampling Probe Electrospray Mass Spectrometry for Detection of Drugs and Metabolites in Thin Tissue Sections

    SciTech Connect

    Van Berkel, Gary J; Kertesz, Vilmos; Koeplinger, Kenneth A.; Vavek, Marissa; Kong, Ah-Ng Tony

    2008-01-01

    A self-aspirating, liquid micro-junction surface sampling probe/electrospray emitter mass spectrometry system was demonstrated for use in the direct analysis of spotted and dosed drugs and their metabolites in thin tissue sections. Proof-of-principle sampling and analysis directly from tissue without the need for sample preparation was demonstrated first by raster scanning a region on a section of rat liver onto which reserpine was spotted. The mass spectral signal from selected reaction monitoring was used to develop a chemical image of the spotted drug on the tissue. The probe was also used to selectively spot sample areas of sagittal whole mouse body tissue sections that had been dosed orally (90 mg/kg) with R,S-sulforaphane 3 hrs prior to sacrifice. Sulforaphane and its glutathione and N-acetyl cysteine conjugates were monitored with selected reaction monitoring and detected in the stomach and various other tissues from the dosed mouse. No signal for these species was observed in the tissue from a control mouse. The same dosed tissue section was used to illustrate the possibility of obtaining a line scan across the whole body section. In total these results illustrate the potential for rapid screening of the distribution of drugs and metabolites in tissue sections with the micro-liquid junction surface sampling probe/electrospray mass spectrometry approach.

  17. Reporting Tumor Molecular Heterogeneity in Histopathological Diagnosis

    PubMed Central

    Mafficini, Andrea; Amato, Eliana; Fassan, Matteo; Simbolo, Michele; Antonello, Davide; Vicentini, Caterina; Scardoni, Maria; Bersani, Samantha; Gottardi, Marisa; Rusev, Borislav; Malpeli, Giorgio; Corbo, Vincenzo; Barbi, Stefano; Sikora, Katarzyna O.; Lawlor, Rita T.; Tortora, Giampaolo; Scarpa, Aldo

    2014-01-01

    Background Detection of molecular tumor heterogeneity has become of paramount importance with the advent of targeted therapies. Analysis for detection should be comprehensive, timely and based on routinely available tumor samples. Aim To evaluate the diagnostic potential of targeted multigene next-generation sequencing (TM-NGS) in characterizing gastrointestinal cancer molecular heterogeneity. Methods 35 gastrointestinal tract tumors, five of each intestinal type gastric carcinomas, pancreatic ductal adenocarcinomas, pancreatic intraductal papillary mucinous neoplasms, ampulla of Vater carcinomas, hepatocellular carcinomas, cholangiocarcinomas, pancreatic solid pseudopapillary tumors were assessed for mutations in 46 cancer-associated genes, using Ion Torrent semiconductor-based TM-NGS. One ampulla of Vater carcinoma cell line and one hepatic carcinosarcoma served to assess assay sensitivity. TP53, PIK3CA, KRAS, and BRAF mutations were validated by conventional Sanger sequencing. Results TM-NGS yielded overlapping results on matched fresh-frozen and formalin-fixed paraffin-embedded (FFPE) tissues, with a mutation detection limit of 1% for fresh-frozen high molecular weight DNA and 2% for FFPE partially degraded DNA. At least one somatic mutation was observed in all tumors tested; multiple alterations were detected in 20/35 (57%) tumors. Seven cancers displayed significant differences in allelic frequencies for distinct mutations, indicating the presence of intratumor molecular heterogeneity; this was confirmed on selected samples by immunohistochemistry of p53 and Smad4, showing concordance with mutational analysis. Conclusions TM-NGS is able to detect and quantitate multiple gene alterations from limited amounts of DNA, moving one step closer to a next-generation histopathologic diagnosis that integrates morphologic, immunophenotypic, and multigene mutational analysis on routinely processed tissues, essential for personalized cancer therapy. PMID:25127237

  18. Mapping international practice patterns in EUS-guided tissue sampling: outcome of a global survey

    PubMed Central

    van Riet, Priscilla A.; Cahen, Djuna L.; Poley, Jan-Werner; Bruno, Marco J.

    2016-01-01

    Background and study aims: Although Endoscopic Ultrasound (EUS)-guided tissue sampling is widely used, the optimal sampling strategy remains subject of debate. We evaluated practice patterns within the international endosonographic community. Patients and methods: An online questionnaire was sent to 400 endosonographers from the United States, Europe, and Asia. Results: A total of 186 (47 %) endosonographers participated: United States 54 (29 %), Europe 85 (46 %), and Asia 47 (25 %). European (75 %) and Asian (84 %) respondents routinely check coagulation status, whereas US respondents only check on indication (64 %, P = 0.007). While propofol sedation is standard in the United States (83 %), conscious sedation is still widely used in Europe (52 %) and Asia (84 %, P < 0.001). Overall, the 22-gauge needle is most commonly used (52 %). For fine-needle aspiration (FNA) of solid pancreatic lesions, 22-gauge (45 %) and 25-gauge (49 %) needles are used equally. For fine-needle biopsy (FNB) of solid masses, the 25-gauge device is less favored than the 22-gauge FNA device (49 % versus 21 %). The 19-gauge needle is generally used for FNB of submucosal masses (62 %). Rapid on-site pathological evaluation (ROSE) is utilized more often by US (98 %) than by European and Asian respondents (51 %, P < 0.001). Cytolyt (52 %), formalin (15 %) and alcohol (15 %) are used for FNA specimen preservation in the United States and Europe, while saline (27 %) and alcohol (38 %) are widely used in Asia (P < 0.001). Conclusions: EUS-guided tissue sampling practices vary substantially within the international endosonographic community and differ considerably from recommendations expressed in guidelines. Because the clinical relevance of these variations is largely unknown, the outcome of this survey suggests a need for further studies. PMID:27227103

  19. One-step labeling of degenerative neurons in unfixed brain tissue samples using Fluoro-Jade C.

    PubMed

    Gu, Qiang; Schmued, Larry C; Sarkar, Sumit; Paule, Merle G; Raymick, Bryan

    2012-06-30

    Neurodegeneration is the underlying cause of a vast majority of neurological disorders and often a result of brain trauma, stroke, or neurotoxic insult. Here we describe a simple method for labeling degenerating neurons in unfixed brain tissue samples. This method could provide a new avenue for identifying and harvesting degenerative neurons from unfixed brain tissues for subsequent molecular analyses.

  20. The effects of endosulfan on the great ramshorn snail Planorbarius corneus (Gastropoda, Pulmonata): a histopathological study.

    PubMed

    Otludil, Birgül; Cengiz, Elif Ipek; Yildirim, M Zeki; Unver, Ozkan; Unlü, Erhan

    2004-08-01

    In this study the great ramshorn snail (Planorbarius corneus), one of the most abundant gastropod of Turkish limnic systems, was investigated to determine the histopathological effects of endosulfan on the digestive gland, foot and mantle under laboratory conditions. Samples were collected from small artificial pools in Karaot at Gelendost-Isparta (southwest of Turkey), where agricultural activities are widespread. The snails were exposed to two sublethal concentrations of endosulfan (0.4 and 0.8 mg/l) for periods of 10, 20 and 30 days. Fifteen snails were kept in 2.5 l glass jars containing dechlorinated tap water and exposed under semi-static test (daily exchange of test water). All the testing was carried out on adult specimens, and snails were maintained on a photothermal period with 16 light hours at 22 +/- 2 degrees C. The histopathological examinations revealed the following changes: amoebocytes infiltration, dilatation in hemolymphatic spaces between the tubules, degeneration of cells, abnormal lumen, necrosis of cells and atrophy in the connective tissue of digestive gland; desquamation of the epithelium cells, changes in the number of mucocytes and protein gland cells, lipid vacuolus and atrophy of the columnar muscle fibers of the foot and mantle tissues. Pycnotic state of cells was also seen in the mantle tissues. Endosulfan caused significant histopathological alterations in the digestive gland, foot and mantle tissues of the snail, irrespective of concentrations of the pesticide and its exposure periods. The results are discussed, particularly in comparison to those of other aquatic organisms.

  1. Sources of Technical Variability in Quantitative LC-MS Proteomics: Human Brain Tissue Sample Analysis.

    SciTech Connect

    Piehowski, Paul D.; Petyuk, Vladislav A.; Orton, Daniel J.; Xie, Fang; Moore, Ronald J.; Ramirez Restrepo, Manuel; Engel, Anzhelika; Lieberman, Andrew P.; Albin, Roger L.; Camp, David G.; Smith, Richard D.; Myers, Amanda J.

    2013-05-03

    To design a robust quantitative proteomics study, an understanding of both the inherent heterogeneity of the biological samples being studied as well as the technical variability of the proteomics methods and platform is needed. Additionally, accurately identifying the technical steps associated with the largest variability would provide valuable information for the improvement and design of future processing pipelines. We present an experimental strategy that allows for a detailed examination of the variability of the quantitative LC-MS proteomics measurements. By replicating analyses at different stages of processing, various technical components can be estimated and their individual contribution to technical variability can be dissected. This design can be easily adapted to other quantitative proteomics pipelines. Herein, we applied this methodology to our label-free workflow for the processing of human brain tissue. For this application, the pipeline was divided into four critical components: Tissue dissection and homogenization (extraction), protein denaturation followed by trypsin digestion and SPE clean-up (digestion), short-term run-to-run instrumental response fluctuation (instrumental variance), and long-term drift of the quantitative response of the LC-MS/MS platform over the 2 week period of continuous analysis (instrumental stability). From this analysis, we found the following contributions to variability: extraction (72%) >> instrumental variance (16%) > instrumental stability (8.4%) > digestion (3.1%). Furthermore, the stability of the platform and its’ suitability for discovery proteomics studies is demonstrated.

  2. Cellulase activity screening using pure carboxymethylcellulose: application to soluble cellulolytic samples and to plant tissue prints.

    PubMed

    Johnsen, Hanne R; Krause, Kirsten

    2014-01-01

    Reliable, rapid and inexpensive detection of cellulolytic enzymes that can be used for a wide variety of biological and environmental samples are currently in high demand. Here, a new cellulase detection protocol is described that circumvents problems observed with popular agar-based methods by exploiting the ability of carboxymethylcellulose (CMC) to form gel-like surfaces on its own. These pure CMC-layers are sensitive to cellulolytic degradation and stainable by Gram's iodine without showing unwelcome reactions with other enzymes. The staining intensity negatively correlates with the enzyme activity and can be used for quantification. Cellulase activities are not obstructed by high sugar contents (e.g., in plant material) which limit the applicability of other quantification methods, making our new method particularly attractive for screening of plant extracts. A useful variant of this new method is its applicability to plant tissue prints for spatial mapping of the cellulolytic activity in a zymogram-like fashion. PMID:24413752

  3. In vivo optical detection of pH in microscopic tissue samples of Arabidopsis thaliana.

    PubMed

    Kašík, Ivan; Podrazký, Ondřej; Mrázek, Jan; Martan, Tomáš; Matějec, Vlastimil; Hoyerová, Klára; Kamínek, Miroslav

    2013-12-01

    Minimally invasive in vivo measurement of pH in microscopic biological samples of μm or μl size, e.g. plant cells, tissues and saps, may help to explain complex biological processes. Consequently, techniques to achieve such measurements are a focus of interest for botanists. This paper describes a technique for the in vivo measurement of pH in the range pH5.0 to pH7.8 in microscopic plant tissue samples of Arabidopsis thaliana based on a ratiometric fluorescence method using low-loss robust tapered fiber probes. For this purpose tapered fiber probes were prepared and coated with a detection layer containing ion-paired fluorescent pH-transducer 8-hydroxypyrene-1,3,6-trisulfonic acid trisodium salt (c-HPTS). A fluorescence ratiometric approach was employed based on excitation at 415 nm and 450 nm and on the comparison of the fluorescence response at 515 nm. The suitability of tapered fiber probes for local detection of pH between 5.0 and 7.8 was demonstrated. A pH sensitivity of 0.15 pH units was achieved within the pH ranges 5.0-5.9 and 7.1-7.8, and this was improved to 0.04 pH units within the pH range 5.9-7.1. Spatial resolution of the probes was better than 20 μm and a time response within 15-20s was achieved. Despite the minute dimensions of the tapered fiber probes the setup developed was relatively robust and compact in construction and performed reliably. It has been successfully employed for the in vivo local determination of pH of mechanically resistant plant tissues of A. thaliana of microscopic scale. The detection of momentary pH gradients across the intact plant seems to be a good tool for the determination of changes in pH in response to experimental treatments affecting for example enzyme activities, availability of mineral nutrients, hormonal control of plant development and plant responses to environmental cues.

  4. Elastic moduli of normal and pathological human breast tissues: an inversion-technique-based investigation of 169 samples

    NASA Astrophysics Data System (ADS)

    Samani, Abbas; Zubovits, Judit; Plewes, Donald

    2007-03-01

    Understanding and quantifying the mechanical properties of breast tissues has been a subject of interest for the past two decades. This has been motivated in part by interest in modelling soft tissue response for surgery planning and virtual-reality-based surgical training. Interpreting elastography images for diagnostic purposes also requires a sound understanding of normal and pathological tissue mechanical properties. Reliable data on tissue elastic properties are very limited and those which are available tend to be inconsistent, in part as a result of measurement methodology. We have developed specialized techniques to measure tissue elasticity of breast normal tissues and tumour specimens and applied them to 169 fresh ex vivo breast tissue samples including fat and fibroglandular tissue as well as a range of benign and malignant breast tumour types. Results show that, under small deformation conditions, the elastic modulus of normal breast fat and fibroglandular tissues are similar while fibroadenomas were approximately twice the stiffness. Fibrocystic disease and malignant tumours exhibited a 3-6-fold increased stiffness with high-grade invasive ductal carcinoma exhibiting up to a 13-fold increase in stiffness compared to fibrogalndular tissue. A statistical analysis showed that differences between the elastic modulus of the majority of those tissues were statistically significant. Implications for the specificity advantages of elastography are reviewed.

  5. Alaska Marine Mammal Tissue Archival Project: Sample inventory and results of analyses of selected samples for organic compounds and trace elements

    SciTech Connect

    Becker, P.R.; Wise, S.A.; Schantz, M.M.; Koster, B.J.; Zeisler, R.

    1992-02-01

    In 1987, the Alaska Marine Mammal Tissue Archival Project (AMMTAP) was established as part of the National Biomonitoring Specimen Bank (NBSB) program at the National Institute of Standards and Technology (NIST).The purpose of the AMMTAP was to establish a representative collection of Alaska marine mammal tissues for future contaminant analyses and documentation of long-term trends in environmental quality. Since 1987, specimens have been collected from 65 animals (seven species) from six different sites. The report contains the current sample inventory and the results of the analysis of selected samples for the measurement of inorganic and organic compounds.

  6. Analysis of protein biomarkers in human clinical tumor samples: critical aspects to success from tissue acquisition to analysis.

    PubMed

    Warren, Madhuri V; Chan, W Y Iris; Ridley, John M

    2011-04-01

    There has been increased interest in the analysis of protein biomarkers in clinical tumor tissues in recent years. Tissue-based biomarker assays can add value and aid decision-making at all stages of drug development, as well as being developed for use as predictive biomarkers and for patient stratification and prognostication in the clinic. However, there must be an awareness of the legal and ethical issues related to the sourcing of human tissue samples. This article also discusses the limits of scope and critical aspects on the successful use of the following tissue-based methods: immunohistochemistry, tissue microarrays and automated image analysis. Future advances in standardization of tissue biobanking methods, immunohistochemistry and quantitative image analysis techniques are also discussed. PMID:21473728

  7. Low invasive in vivo tissue sampling for monitoring biomarkers and drugs during surgery.

    PubMed

    Bojko, Barbara; Gorynski, Krzysztof; Gomez-Rios, German A; Knaak, Jan M; Machuca, Tiago; Cudjoe, Erasmus; Spetzler, Vinzent N; Hsin, Michael; Cypel, Marcelo; Selzner, Markus; Liu, Mingyao; Keshjavee, Shaf; Pawliszyn, Janusz

    2014-05-01

    The techniques currently used for drug, metabolite, and biomarker determination are based on sample collection, and therefore they are not suitable for repeated analysis because of the high invasiveness. Here, we present a novel method of biochemical analysis directly in organ during operation without need of a separate sample collection step: solid-phase microextraction (SPME). The approach is based on flexible microprobe coated with biocompatible extraction phase that is inserted to the tissue with no damage or disturbance of the organ. The method was evaluated during lung and liver transplantations using normothermic ex vivo liver perfusion (NEVLP) and ex vivo lung perfusion (EVLP). The study demonstrated feasibility of the method to extract wide range of endogenous compounds and drugs. Statistical analysis allowed observing metabolic changes of lung during cold ischemic time, perfusion, and reperfusion. It was also demonstrated that the level of drugs and their metabolites can be monitored over time. Based on the methylprednisolone as a selected example, the impairment of enzymatic properties of liver was detected in the injured organs but not in healthy control. This finding was supported by changes in pathways of endogenous metabolites. The SPME probe was also used for analysis of perfusion fluid using stopcock connection. The evaluation of biochemical profile of perfusates demonstrated potential of the approach for monitoring organ function during ex vivo perfusion. The simplicity of the device makes it convenient to use by medical personnel. With the microprobe, different areas of the organ or various organs can be sampled simultaneously. The technology allows assessment of organ function by biochemical profiling, determination of potential biomarkers, and drug monitoring. The use of this method for preintervention analysis could enhance the decision-making process for the best possible personalized approach, whereas post-transplantation monitoring would be

  8. Effects of MRTI sampling characteristics on estimation of HIFU SAR and tissue thermal diffusivity

    NASA Astrophysics Data System (ADS)

    Dillon, C. R.; Todd, N.; Payne, A.; Parker, D. L.; Christensen, D. A.; Roemer, R. B.

    2013-10-01

    While the non-invasive and three-dimensional nature of magnetic-resonance temperature imaging (MRTI) makes it a valuable tool for high-intensity focused ultrasound (HIFU) treatments, random and systematic errors in MRTI measurements may propagate into temperature-based parameter estimates used for pretreatment planning. This study assesses the MRTI effects of zero-mean Gaussian noise (SD = 0.0-2.0 °C), temporal sampling (tacq = 1.0-8.0 s), and spatial averaging (Res = 0.5-2.0 mm isotropic) on HIFU temperature measurements and temperature-based estimates of the amplitude and full width half maximum (FWHM) of the HIFU specific absorption rate and of tissue thermal diffusivity. The ultrasound beam used in simulations and ex vivo pork loin experiments has lateral and axial FWHM dimensions of 1.4 mm and 7.9 mm respectively. For spatial averaging simulations, beams with lateral FWHM varying from 1.2-2.2 mm are also assessed. Under noisy conditions, parameter estimates are improved by fitting to data from larger voxel regions. Varying the temporal sampling results in minimal changes in measured temperatures (<2% change) and parameter estimates (<5% change). For the HIFU beams studied, a spatial resolution of 1 × 1 × 3 mm3 or smaller is required to keep errors in temperature and all estimated parameters less than 10%. By quantifying the errors associated with these sampling characteristics, this work provides researchers with appropriate MRTI conditions for obtaining estimates of parameters essential to pretreatment modeling of HIFU thermal therapies.

  9. Freezing skeletal muscle tissue does not affect its decomposition in soil: evidence from temporal changes in tissue mass, microbial activity and soil chemistry based on excised samples.

    PubMed

    Stokes, Kathryn L; Forbes, Shari L; Tibbett, Mark

    2009-01-10

    The study of decaying organisms and death assemblages is referred to as forensic taphonomy, or more simply the study of graves. This field is dominated by the fields of entomology, anthropology and archaeology. Forensic taphonomy also includes the study of the ecology and chemistry of the burial environment. Studies in forensic taphonomy often require the use of analogues for human cadavers or their component parts. These might include animal cadavers or skeletal muscle tissue. However, sufficient supplies of cadavers or analogues may require periodic freezing of test material prior to experimental inhumation in the soil. This study was carried out to ascertain the effect of freezing on skeletal muscle tissue prior to inhumation and decomposition in a soil environment under controlled laboratory conditions. Changes in soil chemistry were also measured. In order to test the impact of freezing, skeletal muscle tissue (Sus scrofa) was frozen (-20 degrees C) or refrigerated (4 degrees C). Portions of skeletal muscle tissue (approximately 1.5 g) were interred in microcosms (72 mm diameter x 120 mm height) containing sieved (2mm) soil (sand) adjusted to 50% water holding capacity. The experiment had three treatments: control with no skeletal muscle tissue, microcosms containing frozen skeletal muscle tissue and those containing refrigerated tissue. The microcosms were destructively harvested at sequential periods of 2, 4, 6, 8, 12, 16, 23, 30 and 37 days after interment of skeletal muscle tissue. These harvests were replicated 6 times for each treatment. Microbial activity (carbon dioxide respiration) was monitored throughout the experiment. At harvest the skeletal muscle tissue was removed and the detritosphere soil was sampled for chemical analysis. Freezing was found to have no significant impact on decomposition or soil chemistry compared to unfrozen samples in the current study using skeletal muscle tissue. However, the interment of skeletal muscle tissue had a

  10. Detection of Cryptococcus neoformans DNA in Tissue Samples by Nested and Real-Time PCR Assays

    PubMed Central

    Bialek, Ralf; Weiss, Michael; Bekure-Nemariam, Kubrom; Najvar, Laura K.; Alberdi, Maria B.; Graybill, John R.; Reischl, Udo

    2002-01-01

    Two PCR protocols targeting the 18S rRNA gene of Cryptococcus neoformans were established, compared, and evaluated in murine cryptococcal meningitis. One protocol was designed as a nested PCR to be performed in conventional block thermal cyclers. The other protocol was designed as a quantitative single-round PCR adapted to LightCycler technology. One hundred brain homogenates and dilutions originating from 20 ICR mice treated with different azoles were examined. A fungal burden of 3 × 101 to 2.9 × 104 CFU per mg of brain tissue was determined by quantitative culture. Specific PCR products were amplified by the conventional and the LightCycler methods in 86 and 87 samples, respectively, with products identified by DNA sequencing and real-time fluorescence detection. An analytical sensitivity of 1 CFU of C. neoformans per mg of brain tissue and less than 10 CFU per volume used for extraction was observed for both PCR protocols, while homogenates of 70 organs from mice infected with other fungi were PCR negative. Specificity testing was performed with genomic DNA from 31 hymenomycetous fungal species and from the ustilaginomycetous yeast Malassezia furfur, which are phylogenetically related to C. neoformans. Twenty-four strains, including species of human skin flora like M. furfur and Trichosporon spp., were PCR negative. Amplification was observed with Cryptococcus amylolentus, Filobasidiella depauperata, Cryptococcus laurentii, and five species unrelated to clinical specimens. LightCycler PCR products from F. depauperata and Trichosporon faecale could be clearly discriminated by melting curve analysis. The sensitive and specific nested PCR assay as well as the rapid and quantitative LightCycler PCR assay might be useful for the diagnosis and monitoring of human cryptococcal infections. PMID:11874894

  11. Preservation of nucleic acids and tissue morphology in paraffin-embedded clinical samples: comparison of five molecular fixatives.

    PubMed

    Staff, Synnöve; Kujala, Paula; Karhu, Ritva; Rökman, Annika; Ilvesaro, Joanna; Kares, Saara; Isola, Jorma

    2013-09-01

    Formalin fixation preserves tissue morphology at the expense of macromolecule integrity. Freshly frozen samples are the golden standard for DNA and RNA analyses but require laborious deep-freezing and frozen sectioning for morphological studies. Alternative tissue stabilisation methods are therefore needed. We analysed the preservation of nucleic acids, immunohistochemical staining properties and tissue morphology in paraffin-embedded clinical tissue samples fixed with Z7, RCL2, PAXgene, Allprotect and RNAlater. Formalin-fixed and deep-frozen samples were used as controls. Immunohistochemical analyses showed good preservation of antigenicity in all except Allprotect and RNAlater-fixed samples. RNA quality, based on RNA integrity number value by Bioanalyzer, was comparable with freshly frozen samples only in PAXgene-fixed samples. According to quantitative reverse transcription-polymerase chain reaction (qRT-PCR) analyses, RNA from PAXgene samples yielded results similar to freshly frozen samples. No difference between fixatives was seen in DNA analyses (PCR and real-time PCR). In conclusion, PAXgene seems to be superior to other molecular fixatives and formaldehyde.

  12. Design and implementation of coded aperture coherent scatter spectral imaging of cancerous and healthy breast tissue samples.

    PubMed

    Lakshmanan, Manu N; Greenberg, Joel A; Samei, Ehsan; Kapadia, Anuj J

    2016-01-01

    A scatter imaging technique for the differentiation of cancerous and healthy breast tissue in a heterogeneous sample is introduced in this work. Such a technique has potential utility in intraoperative margin assessment during lumpectomy procedures. In this work, we investigate the feasibility of the imaging method for tumor classification using Monte Carlo simulations and physical experiments. The coded aperture coherent scatter spectral imaging technique was used to reconstruct three-dimensional (3-D) images of breast tissue samples acquired through a single-position snapshot acquisition, without rotation as is required in coherent scatter computed tomography. We perform a quantitative assessment of the accuracy of the cancerous voxel classification using Monte Carlo simulations of the imaging system; describe our experimental implementation of coded aperture scatter imaging; show the reconstructed images of the breast tissue samples; and present segmentations of the 3-D images in order to identify the cancerous and healthy tissue in the samples. From the Monte Carlo simulations, we find that coded aperture scatter imaging is able to reconstruct images of the samples and identify the distribution of cancerous and healthy tissues (i.e., fibroglandular, adipose, or a mix of the two) inside them with a cancerous voxel identification sensitivity, specificity, and accuracy of 92.4%, 91.9%, and 92.0%, respectively. From the experimental results, we find that the technique is able to identify cancerous and healthy tissue samples and reconstruct differential coherent scatter cross sections that are highly correlated with those measured by other groups using x-ray diffraction. Coded aperture scatter imaging has the potential to provide scatter images that automatically differentiate cancerous and healthy tissue inside samples within a time on the order of a minute per slice. PMID:26962543

  13. Vibrational spectroscopy studies of formalin-fixed cervix tissues.

    PubMed

    Krishna, C M; Sockalingum, G D; Vadhiraja, B M; Maheedhar, K; Rao, A C K; Rao, L; Venteo, L; Pluot, M; Fernandes, D J; Vidyasagar, M S; Kartha, V B; Manfait, M

    2007-02-15

    Optical histopathology is fast emerging as a potential tool in cancer diagnosis. Fresh tissues in saline are ideal samples for optical histopathology. However, evaluation of suitability of ex vivo handled tissues is necessitated because of severe constraints in sample procurement, handling, and other associated problems with fresh tissues. Among these methods, formalin-fixed samples are shown to be suitable for optical histopathology. However, it is necessary to further evaluate this method from the point of view discriminating tissues with minute biochemical variations. A pilot Raman and Fourier transform infrared (FTIR) microspectroscopic studies of formalin-fixed tissues normal, malignant, and after-2-fractions of radiotherapy from the same malignant cervix subjects were carried out, with an aim to explore the feasibility of discriminating these tissues, especially the tissues after-2-fractions of radiotherapy from other two groups. Raman and FTIR spectra exhibit large differences for normal and malignant tissues and subtle differences are seen between malignant and after-2-fractions of radiotherapy tissues. Spectral data were analyzed by principal component analysis (PCA) and it provided good discrimination of normal and malignant tissues. PCA of data of three tissues, normal, malignant, and 2-fractions after radiotherapy, gave two clusters corresponding to normal and malignant + after-2-fractions of radiotherapy tissues. A second step of PCA was required to achieve discrimination between malignant and after-2-fractions of radiotherapy tissues. Hence, this study not only further supports the use of formalin-fixed tissues in optical histopathology, especially from Raman spectroscopy point of view, it also indicates feasibility of discriminating tissues with minute biochemical differences such as malignant and after-2-fractions of radiotherapy.

  14. Critical comparison of sample preparation strategies for shotgun proteomic analysis of formalin-fixed, paraffin-embedded samples: insights from liver tissue

    PubMed Central

    2014-01-01

    Background The growing field of formalin-fixed paraffin-embedded (FFPE) tissue proteomics holds promise for improving translational research. Direct tissue trypsinization (DT) and protein extraction followed by in solution digestion (ISD) or filter-aided sample preparation (FASP) are the most common workflows for shotgun analysis of FFPE samples, but a critical comparison of the different methods is currently lacking. Experimental design DT, FASP and ISD workflows were compared by subjecting to the same label-free quantitative approach three independent technical replicates of each method applied to FFPE liver tissue. Data were evaluated in terms of method reproducibility and protein/peptide distribution according to localization, MW, pI and hydrophobicity. Results DT showed lower reproducibility, good preservation of high-MW proteins, a general bias towards hydrophilic and acidic proteins, much lower keratin contamination, as well as higher abundance of non-tryptic peptides. Conversely, FASP and ISD proteomes were depleted in high-MW proteins and enriched in hydrophobic and membrane proteins; FASP provided higher identification yields, while ISD exhibited higher reproducibility. Conclusions These results highlight that diverse sample preparation strategies provide significantly different proteomic information, and present typical biases that should be taken into account when dealing with FFPE samples. When a sufficient amount of tissue is available, the complementary use of different methods is suggested to increase proteome coverage and depth. PMID:25097466

  15. The use of histopathology in the practice of necropsy.

    PubMed Central

    Williams, J O; Goddard, M J; Gresham, G A; Wyatt, B A

    1997-01-01

    AIM: To examine current practice and to establish criteria for the use of histopathology in necropsy practice. METHODS: During an audit of necropsy reporting, consensus could not be reached about the use of routine histopathology. Therefore local guidelines were formulated and current practice was compared with these guidelines. Fifteen consecutive necropsies undertaken by each consultant were reviewed and the use of histopathology noted. RESULTS: In general, the standard of necropsy reporting was reasonably high. Tissue was retained for histopathology in 25% of necropsies and 72% of these necropsy reports included a histopathology report. Using the guidelines, the assessors judged that histopathology might have been valuable in a further 19%. It was felt that routine histopathology would not have been helpful in determining the cause of death in the remaining 56%. The importance of the pathologist's clinical judgement in individual cases was stressed. At reaudit, nearly two years later, there was no significant change in practice, reflecting the lack of consensus. CONCLUSIONS: Even when histopathology might contribute to finding the cause of death, it was not always done. However, the assumption that histology is invariably helpful in determining the cause of death is challenged. PMID:9301557

  16. Integrated genome-wide analysis of genomic changes and gene regulation in human adrenocortical tissue samples

    PubMed Central

    Gara, Sudheer Kumar; Wang, Yonghong; Patel, Dhaval; Liu-Chittenden, Yi; Jain, Meenu; Boufraqech, Myriem; Zhang, Lisa; Meltzer, Paul S.; Kebebew, Electron

    2015-01-01

    To gain insight into the pathogenesis of adrenocortical carcinoma (ACC) and whether there is progression from normal-to-adenoma-to-carcinoma, we performed genome-wide gene expression, gene methylation, microRNA expression and comparative genomic hybridization (CGH) analysis in human adrenocortical tissue (normal, adrenocortical adenomas and ACC) samples. A pairwise comparison of normal, adrenocortical adenomas and ACC gene expression profiles with more than four-fold expression differences and an adjusted P-value < 0.05 revealed no major differences in normal versus adrenocortical adenoma whereas there are 808 and 1085, respectively, dysregulated genes between ACC versus adrenocortical adenoma and ACC versus normal. The majority of the dysregulated genes in ACC were downregulated. By integrating the CGH, gene methylation and expression profiles of potential miRNAs with the gene expression of dysregulated genes, we found that there are higher alterations in ACC versus normal compared to ACC versus adrenocortical adenoma. Importantly, we identified several novel molecular pathways that are associated with dysregulated genes and further experimentally validated that oncostatin m signaling induces caspase 3 dependent apoptosis and suppresses cell proliferation. Finally, we propose that there is higher number of genomic changes from normal-to-adenoma-to-carcinoma and identified oncostatin m signaling as a plausible druggable pathway for therapeutics. PMID:26446994

  17. Integrated genome-wide analysis of genomic changes and gene regulation in human adrenocortical tissue samples.

    PubMed

    Gara, Sudheer Kumar; Wang, Yonghong; Patel, Dhaval; Liu-Chittenden, Yi; Jain, Meenu; Boufraqech, Myriem; Zhang, Lisa; Meltzer, Paul S; Kebebew, Electron

    2015-10-30

    To gain insight into the pathogenesis of adrenocortical carcinoma (ACC) and whether there is progression from normal-to-adenoma-to-carcinoma, we performed genome-wide gene expression, gene methylation, microRNA expression and comparative genomic hybridization (CGH) analysis in human adrenocortical tissue (normal, adrenocortical adenomas and ACC) samples. A pairwise comparison of normal, adrenocortical adenomas and ACC gene expression profiles with more than four-fold expression differences and an adjusted P-value < 0.05 revealed no major differences in normal versus adrenocortical adenoma whereas there are 808 and 1085, respectively, dysregulated genes between ACC versus adrenocortical adenoma and ACC versus normal. The majority of the dysregulated genes in ACC were downregulated. By integrating the CGH, gene methylation and expression profiles of potential miRNAs with the gene expression of dysregulated genes, we found that there are higher alterations in ACC versus normal compared to ACC versus adrenocortical adenoma. Importantly, we identified several novel molecular pathways that are associated with dysregulated genes and further experimentally validated that oncostatin m signaling induces caspase 3 dependent apoptosis and suppresses cell proliferation. Finally, we propose that there is higher number of genomic changes from normal-to-adenoma-to-carcinoma and identified oncostatin m signaling as a plausible druggable pathway for therapeutics.

  18. Stable isotope analysis of 1987-1991 zooplankton samples and bowhead whale tissues. Final report

    SciTech Connect

    Schell, D.M.

    1992-06-01

    Stable isotope analyses of bowhead whale tissue samples and bowhead whale prey organisms collected over the years 1987 to 1991 were used to provide detail on the isotope ratio gradients evident in the arctic Alaskan zooplankton and to verify previous findings regarding the growth rates and age determination techniques developed for bowhead whales. Zooplankton of the Bering and Chukchi seas are enriched in (13)C relative to the eastern Beaufort Sea. The analysis of baleen from bowhead whales taken between 1987 to 1990 indicate that the whales are slow-growing and the young animals between year one and about six to seven years of age, undergo a period of little or no linear growth. The authors estimate that bowheads require 16-18 years to reach the length of sexual maturity, i.e., 13-14 m. From baleen Delta(13C) cycles, a 20 year record of the isotope ratios in the phytoplankton of the northern Bering and Chukchi seas was constructed. The long-term record has been compared with the temperature anomalies in surface waters of the Bering Sea. The Delta(13C) of the zooplankton is inversely correlated with temperature and refutes current models attempting to relate ocean temperature, and atmospheric carbon dioxide levels with the Delta(13C) of ocean sediment organic matter.

  19. Gene expression profiling of human breast tissue samples using SAGE-Seq.

    PubMed

    Wu, Zhenhua Jeremy; Meyer, Clifford A; Choudhury, Sibgat; Shipitsin, Michail; Maruyama, Reo; Bessarabova, Marina; Nikolskaya, Tatiana; Sukumar, Saraswati; Schwartzman, Armin; Liu, Jun S; Polyak, Kornelia; Liu, X Shirley

    2010-12-01

    We present a powerful application of ultra high-throughput sequencing, SAGE-Seq, for the accurate quantification of normal and neoplastic mammary epithelial cell transcriptomes. We develop data analysis pipelines that allow the mapping of sense and antisense strands of mitochondrial and RefSeq genes, the normalization between libraries, and the identification of differentially expressed genes. We find that the diversity of cancer transcriptomes is significantly higher than that of normal cells. Our analysis indicates that transcript discovery plateaus at 10 million reads/sample, and suggests a minimum desired sequencing depth around five million reads. Comparison of SAGE-Seq and traditional SAGE on normal and cancerous breast tissues reveals higher sensitivity of SAGE-Seq to detect less-abundant genes, including those encoding for known breast cancer-related transcription factors and G protein-coupled receptors (GPCRs). SAGE-Seq is able to identify genes and pathways abnormally activated in breast cancer that traditional SAGE failed to call. SAGE-Seq is a powerful method for the identification of biomarkers and therapeutic targets in human disease.

  20. Apicomplexa primers amplify Proteromonas (Stramenopiles, Slopalinida, Proteromonadidae) in tissue and blood samples from lizards.

    PubMed

    Maia, João P M C; Gómez-Díaz, Elena; Harris, D James

    2012-12-01

    Microscopy has traditionally been the most common method in parasitological studies, but in recent years molecular screening has become increasingly frequent to detect protozoan parasites in a wide range of vertebrate hosts and vectors. During routine molecular screening of apicomplexan parasites in reptiles using the 18S rRNA gene, we have amplified and sequenced Proteromonas parasites from three lizard hosts (less than 1% prevalence). We conducted phylogenetic analysis to confirm the taxonomic position and infer their relationships with other stramenopiles. Although our phylogeny is limited due to scarcity of molecular data on these protists, our results confirm they are closely related to Proteromonas lacertae. Our findings show that unexpected parasites can be amplified from host samples (blood and tissue) using general procedures to detect hemoparasites, and stress that positive PCR amplifications alone should not be considered as definitive proof of infection by particular parasites. Further validation by sequence confirmation and thorough phylogenetic assessment will not only avoid false positives and biased prevalence estimates but also provide valuable information on the biodiversity and phylogenetic relationships of other parasitic organisms. More generally, our results illustrate the perils of general diagnosis protocols in parasitological studies and the need of cross-validation procedures.

  1. Grinding and polishing instead of sectioning for the tissue samples with a graft: Implications for light and electron microscopy.

    PubMed

    Mukhamadiyarov, Rinat A; Sevostyanova, Victoria V; Shishkova, Daria K; Nokhrin, Andrey V; Sidorova, Olga D; Kutikhin, Anton G

    2016-06-01

    A broad use of the graft replacement requires a detailed investigation of the host-graft interaction, including both histological examination and electron microscopy. A high quality sectioning of the host tissue with a graft seems to be complicated; in addition, it is difficult to examine the same tissue area by both of the mentioned microscopy techniques. To solve these problems, we developed a new technique of epoxy resin embedding with the further grinding, polishing, and staining. Graft-containing tissues prepared by grinding and polishing preserved their structure; however, sectioning frequently required the explantation of the graft and led to tissue disintegration. Moreover, stained samples prepared by grinding and polishing may then be assessed by both light microscopy and backscattered scanning electron microscopy. Therefore, grinding and polishing outperform sectioning when applied to the tissues with a graft. PMID:27023831

  2. Discordant assessment of tumor biomarkers by histopathological and molecular assays in the EORTC randomized controlled 10041/BIG 03-04 MINDACT trial breast cancer : Intratumoral heterogeneity and DCIS or normal tissue components are unlikely to be the cause of discordance.

    PubMed

    Viale, Giuseppe; Slaets, Leen; de Snoo, Femke A; Bogaerts, Jan; Russo, Leila; van't Veer, Laura; Rutgers, Emiel J T; Piccart-Gebhart, Martine J; Stork-Sloots, Lisette; Dell'Orto, Patrizia; Glas, Annuska M; Cardoso, Fatima

    2016-02-01

    Accurate identification of breast cancer patients most likely to benefit from adjuvant systemic therapies is crucial. Better understanding of differences between methods can lead to an improved ER, PgR, and HER-2 assessment. The purpose of this preplanned translational research is to investigate the correlation of central IHC/FISH assessments with microarray mRNA readouts of ER, PgR, and HER-2 status in the MINDACT trial and to determine if any discordance could be attributed to intratumoral heterogeneity or the DCIS and normal tissue components in the specimens. MINDACT is an international, prospective, randomized, phase III trial investigating the clinical utility of MammaPrint in selecting patients with early breast cancer for adjuvant chemotherapy (n = 6694 patients). Gene-expression data were obtained by TargetPrint; IHC and/or FISH were assessed centrally (n = 5788; 86 %). Macroscopic and microscopic evaluation of centrally submitted FFPE blocks identified 1427 cases for which the very same sample was submitted for gene-expression analysis. TargetPrint ER had a positive agreement of 98 %, and a negative agreement of 95 % with central pathology. Corresponding figures for PgR were 85 and 94 % and for HER-2 72 and 99 %. Agreement of mRNA versus central protein was not different when the same or a different portion of the tumor tissue was analyzed or when DCIS and/or normal tissue was included in the sample subjected to mRNA assays. This is the first large analysis to assess the discordance rate between protein and mRNA analysis of breast cancer markers, and to look into intratumoral heterogeneity, DCIS, or normal tissue components as a potential cause of discordance. The observed difference between mRNA and protein assessment for PgR and HER-2 needs further research; the present analysis does not support intratumoral heterogeneity or the DCIS and normal tissue components being likely causes of the discordance.

  3. Molecular histopathology by nonlinear interferometric vibrational imaging

    NASA Astrophysics Data System (ADS)

    Boppart, Stephen A.

    2011-07-01

    A rapid label-free approach for molecular histopathology is presented and reviewed. Broadband vibrational spectra are generated by nonlinear interferometric vibrational imaging (NIVI), a coherent anti-Stokes Raman scattering (CARS)- based technique that uses interferometry and signal processing approaches to acquire Raman-like profiles with suppression of the non-resonant background. This allows for the generation of images that provide contrast based on quantitative chemical composition with high spatial and spectral resolution. Algorithms are demonstrated for reducing the diagnostic spectral information into color-coded composite images for the rapid identification of chemical constituents in skin, as well as differentiating normal from abnormal tissue in a pre-clinical tumor model for human breast cancer. This technology and methodology could result in an alternative method to the traditional histological staining and subjective interpretation procedure currently used in the diagnosis of disease, and has the potential for future in vivo molecular histopathology.

  4. [Bacteria isolated from urine and renal tissue samples and their relation to renal histology].

    PubMed

    Gökalp, A; Gültekin, E Y; Bakici, M Z; Ozdeşlik, B

    1988-01-01

    The bacteria from the urine and renal biopsy specimens of 40 patients undergoing renal surgery were isolated and their relations with renal histology investigated. The urine cultures were positive in 14 patients, the same organisms being isolated from the renal tissue in 7 cases. In 6 patients with negative urine cultures, bacteria were isolated from renal tissues. Of the 28 cases pathologically diagnosed as chronic pyelonephritis, bacteria were isolated from the renal tissue in 13 cases, the urine cultures being positive in only 11 cases. E. coli was the most commonly encountered bacteria in both the urine and renal tissues.

  5. Comparison between whole mount tissue preparations and virtual tissue microarray samples for measuring Ki-67 and apoptosis indices in human bladder cancer: A cross-sectional study.

    PubMed

    Oshiro, Hisashi; Czerniak, Bogdan A; Sakamaki, Kentaro; Tsuta, Koji; Bondaruk, Jolanta; Keyhani, Afsaneh; Dinney, Colin P; Nagai, Takeshi; Kamat, Ashish M

    2016-08-01

    Recent tissue microarray (TMA)-based studies have shown that cell proliferation- and apoptosis-related biomarkers are associated with clinical outcomes in patients with bladder urothelial carcinoma. However, little is known about the differences in these biomarker measurements between whole mount tissue preparations and TMAs. This study aimed to elucidate the discrepancy in the measurements of Ki-67 indices (KIs) and apoptosis indices (AIs) between whole mount tissue preparations and TMAs of bladder urothelial carcinoma samples.Whole mount tissue preparations for Ki-67 immunohistochemistry and terminal deoxynucleotidyl transferase dUTP nick end labeling were made from 30 patients who underwent transurethral resection of bladder urothelial carcinoma. Digital microscopy-assisted virtual TMAs, consisting of 3 small round areas (1 or 0.6 mm in diameter), were generated from the same whole mount tissue preparations. The measurement results in highly reactive areas of biomarkers were compared between the whole mount tissue preparation- and the TMA-based methods. Bland-Altman plot analysis, regression analysis, and Kendall τ were performed to investigate differences in the measurement results, systematic biases, and correlations between biomarkers.Although the Bland-Altman plot analysis demonstrated that almost all the plots were within the limits of agreement, fixed biases were detected in the 1- and 0.6-mm TMAs for the KI (0.181 and 0.222, respectively) and the AI (0.055 and 0.063, respectively). Proportional biases were also detected in the 1- and 0.6-mm TMAs for the AI (P < 0.001 and P < 0.001, respectively). Furthermore, positive correlations between KIs and AIs were observed in whole mount tissue preparations (r = 0.260, P = 0.044) and in the 1 mm TMAs (r = 0.375, P = 0.004); however, no such correlation was observed in the 0.6 mm TMAs.Our study suggests that the measurement results for certain biomarkers of bladder urothelial

  6. Comparison between whole mount tissue preparations and virtual tissue microarray samples for measuring Ki-67 and apoptosis indices in human bladder cancer

    PubMed Central

    Oshiro, Hisashi; Czerniak, Bogdan A.; Sakamaki, Kentaro; Tsuta, Koji; Bondaruk, Jolanta; Keyhani, Afsaneh; Dinney, Colin P.; Nagai, Takeshi; Kamat, Ashish M.

    2016-01-01

    Abstract Recent tissue microarray (TMA)-based studies have shown that cell proliferation- and apoptosis-related biomarkers are associated with clinical outcomes in patients with bladder urothelial carcinoma. However, little is known about the differences in these biomarker measurements between whole mount tissue preparations and TMAs. This study aimed to elucidate the discrepancy in the measurements of Ki-67 indices (KIs) and apoptosis indices (AIs) between whole mount tissue preparations and TMAs of bladder urothelial carcinoma samples. Whole mount tissue preparations for Ki-67 immunohistochemistry and terminal deoxynucleotidyl transferase dUTP nick end labeling were made from 30 patients who underwent transurethral resection of bladder urothelial carcinoma. Digital microscopy-assisted virtual TMAs, consisting of 3 small round areas (1 or 0.6 mm in diameter), were generated from the same whole mount tissue preparations. The measurement results in highly reactive areas of biomarkers were compared between the whole mount tissue preparation- and the TMA-based methods. Bland–Altman plot analysis, regression analysis, and Kendall τ were performed to investigate differences in the measurement results, systematic biases, and correlations between biomarkers. Although the Bland–Altman plot analysis demonstrated that almost all the plots were within the limits of agreement, fixed biases were detected in the 1- and 0.6-mm TMAs for the KI (0.181 and 0.222, respectively) and the AI (0.055 and 0.063, respectively). Proportional biases were also detected in the 1- and 0.6-mm TMAs for the AI (P < 0.001 and P < 0.001, respectively). Furthermore, positive correlations between KIs and AIs were observed in whole mount tissue preparations (r = 0.260, P = 0.044) and in the 1 mm TMAs (r = 0.375, P = 0.004); however, no such correlation was observed in the 0.6 mm TMAs. Our study suggests that the measurement results for certain biomarkers of bladder

  7. An Unsupervised MVA Method to Compare Specific Regions in Human Breast Tumor Tissue Samples Using ToF-SIMS

    PubMed Central

    Bluestein, Blake M.; Morrish, Fionnuala; Graham, Daniel J.; Guenthoer, Jamie; Hockenbery, David; Porter, Peggy; Gamble, Lara J.

    2016-01-01

    Imaging time-of-flight secondary ion mass spectrometry (ToF-SIMS) and principal component analysis (PCA) were used to investigate two sets of pre- and post-chemotherapy human breast tumor tissue sections to characterize lipids associated with tumor metabolic flexibility and response to treatment. The micron spatial resolution imaging capability of ToF-SIMS provides a powerful approach to attain spatially-resolved molecular and cellular data from cancerous tissues not available with conventional imaging techniques. Three ca. 1 mm2 areas per tissue section were analyzed by stitching together 200 μm × 200 μm raster area scans. A method to isolate and analyze specific tissue regions of interest by utilizing PCA of ToF-SIMS images is presented, which allowed separation of cellularized areas from stromal areas. These PCA-generated regions of interest were then used as masks to reconstruct representative spectra from specifically stromal or cellular regions. The advantage of this unsupervised selection method is a reduction in scatter in the spectral PCA results when compared to analyzing all tissue areas or analyzing areas highlighted by a pathologist. Utilizing this method, stromal and cellular regions of breast tissue biopsies taken pre- versus post-chemotherapy demonstrate chemical separation using negatively-charged ion species. In this sample set, the cellular regions were predominantly all cancer cells. Fatty acids (i.e. palmitic, oleic, and stearic), monoacylglycerols, diacylglycerols and vitamin E profiles were distinctively different between the pre- and post-therapy tissues. These results validate a new unsupervised method to isolate and interpret biochemically distinct regions in cancer tissues using imaging ToF-SIMS data. In addition, the method developed here can provide a framework to compare a variety of tissue samples using imaging ToF-SIMS, especially where there is section-to-section variability that makes it difficult to use a serial hematoxylin and

  8. Optical coherence tomography detection of shear wave propagation in inhomogeneous tissue equivalent phantoms and ex-vivo carotid artery samples

    PubMed Central

    Razani, Marjan; Luk, Timothy W.H.; Mariampillai, Adrian; Siegler, Peter; Kiehl, Tim-Rasmus; Kolios, Michael C.; Yang, Victor X.D.

    2014-01-01

    In this work, we explored the potential of measuring shear wave propagation using optical coherence elastography (OCE) in an inhomogeneous phantom and carotid artery samples based on a swept-source optical coherence tomography (OCT) system. Shear waves were generated using a piezoelectric transducer transmitting sine-wave bursts of 400 μs duration, applying acoustic radiation force (ARF) to inhomogeneous phantoms and carotid artery samples, synchronized with a swept-source OCT (SS-OCT) imaging system. The phantoms were composed of gelatin and titanium dioxide whereas the carotid artery samples were embedded in gel. Differential OCT phase maps, measured with and without the ARF, detected the microscopic displacement generated by shear wave propagation in these phantoms and samples of different stiffness. We present the technique for calculating tissue mechanical properties by propagating shear waves in inhomogeneous tissue equivalent phantoms and carotid artery samples using the ARF of an ultrasound transducer, and measuring the shear wave speed and its associated properties in the different layers with OCT phase maps. This method lays the foundation for future in-vitro and in-vivo studies of mechanical property measurements of biological tissues such as vascular tissues, where normal and pathological structures may exhibit significant contrast in the shear modulus. PMID:24688822

  9. Profiling of adrenocorticotropic hormone and arginine vasopressin in human pituitary gland and tumor thin tissue sections using droplet-based liquid-microjunction surface-sampling-HPLC–ESI-MS–MS

    SciTech Connect

    Kertesz, Vilmos; Calligaris, David; Feldman, Daniel R.; Changelian, Armen; Laws, Edward R.; Santagata, Sandro; Agar, Nathalie Y. R.; Van Berkel, Gary J.

    2015-06-18

    Described here are the results from the profiling of the proteins arginine vasopressin (AVP) and adrenocorticotropic hormone (ACTH) from normal human pituitary gland and pituitary adenoma tissue sections using a fully automated droplet-based liquid microjunction surface sampling-HPLC-ESI-MS/MS system for spatially resolved sampling, HPLC separation, and mass spectral detection. Excellent correlation was found between the protein distribution data obtained with this droplet-based liquid microjunction surface sampling-HPLC-ESI-MS/MS system and those data obtained with matrix assisted laser desorption ionization (MALDI) chemical imaging analyses of serial sections of the same tissue. The protein distributions correlated with the visible anatomic pattern of the pituitary gland. AVP was most abundant in the posterior pituitary gland region (neurohypophysis) and ATCH was dominant in the anterior pituitary gland region (adenohypophysis). The relative amounts of AVP and ACTH sampled from a series of ACTH secreting and non-secreting pituitary adenomas correlated with histopathological evaluation. ACTH was readily detected at significantly higher levels in regions of ACTH secreting adenomas and in normal anterior adenohypophysis compared to non-secreting adenoma and neurohypophysis. AVP was mostly detected in normal neurohypophysis as anticipated. This work demonstrates that a fully automated droplet-based liquid microjunction surface sampling system coupled to HPLC-ESI-MS/MS can be readily used for spatially resolved sampling, separation, detection, and semi-quantitation of physiologically-relevant peptide and protein hormones, such as AVP and ACTH, directly from human tissue. In addition, the relative simplicity, rapidity and specificity of the current methodology support the potential of this basic technology with further advancement for assisting surgical decision-making.

  10. Profiling of adrenocorticotropic hormone and arginine vasopressin in human pituitary gland and tumor thin tissue sections using droplet-based liquid-microjunction surface-sampling-HPLC–ESI-MS–MS

    DOE PAGES

    Kertesz, Vilmos; Calligaris, David; Feldman, Daniel R.; Changelian, Armen; Laws, Edward R.; Santagata, Sandro; Agar, Nathalie Y. R.; Van Berkel, Gary J.

    2015-06-18

    Described here are the results from the profiling of the proteins arginine vasopressin (AVP) and adrenocorticotropic hormone (ACTH) from normal human pituitary gland and pituitary adenoma tissue sections using a fully automated droplet-based liquid microjunction surface sampling-HPLC-ESI-MS/MS system for spatially resolved sampling, HPLC separation, and mass spectral detection. Excellent correlation was found between the protein distribution data obtained with this droplet-based liquid microjunction surface sampling-HPLC-ESI-MS/MS system and those data obtained with matrix assisted laser desorption ionization (MALDI) chemical imaging analyses of serial sections of the same tissue. The protein distributions correlated with the visible anatomic pattern of the pituitary gland.more » AVP was most abundant in the posterior pituitary gland region (neurohypophysis) and ATCH was dominant in the anterior pituitary gland region (adenohypophysis). The relative amounts of AVP and ACTH sampled from a series of ACTH secreting and non-secreting pituitary adenomas correlated with histopathological evaluation. ACTH was readily detected at significantly higher levels in regions of ACTH secreting adenomas and in normal anterior adenohypophysis compared to non-secreting adenoma and neurohypophysis. AVP was mostly detected in normal neurohypophysis as anticipated. This work demonstrates that a fully automated droplet-based liquid microjunction surface sampling system coupled to HPLC-ESI-MS/MS can be readily used for spatially resolved sampling, separation, detection, and semi-quantitation of physiologically-relevant peptide and protein hormones, such as AVP and ACTH, directly from human tissue. In addition, the relative simplicity, rapidity and specificity of the current methodology support the potential of this basic technology with further advancement for assisting surgical decision-making.« less

  11. Gross signs and histopathology of Ostrea puelchana infected by a Bonamia exitiosa-like parasite (Haplosporidia).

    PubMed

    Kroeck, Marina A

    2010-04-01

    Haplosporidian microcells belonging to the genus Bonamia parasitise various species of oysters around the world. In Argentina, Bonamia sp. was the causative agent of mass mortality among Ostrea puelchana cultured in San Antonio Bay (San Matías Gulf), and it was detected in natural beds inside San Matías Gulf. In order to describe the gross and histopathological signs caused by Bonamia sp. in O. puelchana, cultured and wild oysters were sampled and analysed by traditional techniques including heart imprints and histology. Cells of Bonamia sp. were observed in connective tissue, free or within haemocytes, in gills and around the digestive gland, stomach, intestine and gonad. Gross signs, histopathological alterations in O. puelchana, and Bonamia sp. cytological morphology resemble those reported for B. exitiosa. However, I propose to treat the Argentinean species as B. exitiosa-like until more molecular and ultrastructural studies are conducted to determine the correct taxonomy.

  12. Single-pixel hyperspectral imaging for real-time cancer detection: detecting damage in ex vivo porcine tissue samples

    NASA Astrophysics Data System (ADS)

    Peller, Joseph; Farahi, Faramarz; Trammell, Susan R.

    2016-03-01

    We are developing a single-pixel hyperspectral imaging system based on compressive sensing that acquires spatial and spectral information simultaneously. Our spectral imaging system uses autofluorescencent emission from collagen (400 nm) and NAD(P)H (475 nm), as well as, differences in the optical reflectance spectra as diagnostics for differentiating between healthy and diseased tissue. In this study, we demonstrate the ability of our imaging system to discriminate between healthy and damaged porcine epidermal tissue. Healthy porcine epidermal tissue samples (n=11) were imaged ex vivo using our hyperspectral system. The amount of NAD(P)H emission and the reflectance properties were approximately constant across the surface of healthy tissue samples. The tissue samples were then thermally damaged using an 1850 nm thulium fiber laser and re-imaged after laser irradiation. The damaged regions were clearly visible in the hyperspectral images as the thermal damage altered the fluorescent emission of NAD(P)H and changed the scattering properties of the tissue. The extent of the damaged regions was determined based on the hyperspectral images and these estimates were compared to damage extents measured in white light images acquired with a traditional camera. The extent of damage determined via hyperspectral imaging was in good agreement with estimates based on white light imaging indicating that our system is capable of differentiating between healthy and damaged tissue. Possible applications of our single pixel hyperspectral imaging system range from real-time determination of tumor margins during surgery to the use of this technique in the pathology lab to aid with cancer diagnosis and staging.

  13. Characterization and antimicrobial resistance of Salmonella isolated from internal tissues, ceca and rinse samples from commercial broiler chickens

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The presence, serotype, and antimicrobial resistance profile of Salmonella from internal tissues (spleen, liver/gall bladder, thymus, Meckel’s diverticulum, and free floating yolk), ceca and carcass rinse samples were determined from six-week-old (n=30) and eight-week-old (n=40) commercial broilers ...

  14. Comparative evaluation of specific methods for labeling of Encephalitozoon cuniculi in paraffin wax-embedded tissue samples.

    PubMed

    Habenbacher, Bettina; Klang, Andrea; Fragner, Karin; Dinhopl, Nora; Künzel, Frank; Weissenböck, Herbert

    2012-03-01

    Detection of the microsporidian Encephalitozoon cuniculi in tissue samples is considered difficult. The aim of the current study was to determine whether immunohistochemistry (IHC) and in situ hybridization (ISH) represent reliable methods for the detection of E. cuniculi in postmortem tissue samples of rabbits. Paraffin-embedded tissue sections of brain and kidneys of 48 naturally infected pet rabbits, 10 negative controls, and the eyes of 3 further rabbits were used for all investigations. By IHC in 19 animals (37.3%), spores could be clearly detected and were all equally stained. By ISH using a digoxigenin-labeled oligonucleotide probe, only 6 animals (11.8%) proved undoubtedly positive. In these cases, many parasite-like objects revealed strong typical purple-black positive signals. However, several of the examined samples showed only partial staining of the pathogen or unclear results. Thus, in order to find an explanation for these inconsistent ISH results and to take a more detailed look at the different developmental stages of the organism, electron microscopy was applied. Empty spores, which had already discharged their polar filaments, prevailed in total number. Taken together, both techniques are rather insensitive, but under the condition that sufficient numbers of microsporidia are present, IHC can be recommended for specific identification of E. cuniculi in tissue samples. In contrast, ISH failed to detect some developmental stages of the organism, and, as such, ISH is therefore considered an inappropriate diagnostic method.

  15. Development and application of specific cytokine assays in tissue samples from a bottlenose dolphin with hyperinsulinemia

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Chronic inflammation has been associated with insulin resistance and type 2 diabetes in humans. Postmortem hepatic and splenic tissue from a 46-year old geriatric male bottlenose dolphin (Tursiops truncatus) with insulin resistance (chronic hyperinsulinemia with hyperglycemia) , chronic = inflamma...

  16. Histopathologic criteria to confirm white-nose syndrome in bats

    USGS Publications Warehouse

    Meteyer, C.U.; Buckles, E.L.; Blehert, D.S.; Hicks, A.C.; Green, D.E.; Shearn-Bochsler, V.; Thomas, N.J.; Gargas, A.; Behr, M.J.

    2009-01-01

    White-nose syndrome (WNS) is a cutaneous fungal disease of hibernating bats associated with a novel Geomyces sp. fungus. Currently, confirmation of WNS requires histopathologic examination. Invasion of living tissue distinguishes this fungal infection from those caused by conventional transmissible dermatophytes. Although fungal hyphae penetrate the connective tissue of glabrous skin and muzzle, there is typically no cellular inflammatory response in hibernating bats. Preferred tissue samples to diagnose this fungal infection are rostral muzzle with nose and wing membrane fixed in 10% neutral buffered formalin. To optimize detection, the muzzle is trimmed longitudinally, the wing membrane is rolled, and multiple cross-sections are embedded to increase the surface area examined. Periodic acid-Schiff stain is essential to discriminate the nonpigmented fungal hyphae and conidia. Fungal hyphae form cup-like epidermal erosions and ulcers in the wing membrane and pinna with involvement of underlying connective tissue. In addition, fungal hyphae are present in hair follicles and in sebaceous and apocrine glands of the muzzle with invasion of tissue surrounding adnexa. Fungal hyphae in tissues are branching and septate, but the diameter and shape of the hyphae may vary from parallel walls measuring 2 ??m in diameter to irregular walls measuring 3-5 ??m in diameter. When present on short aerial hyphae, curved conidia are approximately 2.5 ??m wide and 7.5 ??m in curved length. Conidia have a more deeply basophilic center, and one or both ends are usually blunt. Although WNS is a disease of hibernating bats, severe wing damage due to fungal hyphae may be seen in bats that have recently emerged from hibernation. These recently emerged bats also have a robust suppurative inflammatory response.

  17. An exploratory survey of professionals on the use of stored tissue samples from minors for genetic research.

    PubMed

    Hens, K; Snoeck, J; Nys, H; Cassiman, J-J; Dierickx, K

    2010-01-01

    he ethical aspects of the use of stored tissue samples collected from minors are of topical interest. However, the views of professionals working in the field of genetics have not been investigated in depth anywhere. We conducted a survey among 194 such professionals in Belgium. This list was composed of the members of the High Council for Anthropogenetics, supplemented with all professionals working in the field of genetics that we found on the websites of the eight Belgian centers of human genetics and of the associated university registries. We achieved a response rate of 35.5%. The vast majority (92%) think that research on stored tissue samples is useful. Most respondents stated that parental consent is valid (82.5%), and 76.5% thought that children should also be given the right to assent when they are able to comprehend the implications of the storage of biological samples and of genetic research. Slightly more than half put the age at which young people can understand storage or research rather high: 16-18 years (51 and 53.1%, respectively). Although there is some consensus in the literature that donors should be allowed to give broad consent for future research on their biological samples, only 47.6% in our survey thought that parents should be allowed to consent to any future research on their children's samples. The aim of our study was to give some basis for future ethical reflections and policies on the subject of stored tissue samples from minors for genetic research. We concluded that a large majority of Belgian researchers and clinicians in the field of genetic research think research on stored tissue samples from minors is useful. They also think that parental consent for such research is valid, but that children should be allowed to assent as they grow older.

  18. Simultaneous determination of perfluorinated compounds and their potential precursors in mussel tissue and fish muscle tissue and liver samples by liquid chromatography-electrospray-tandem mass spectrometry.

    PubMed

    Zabaleta, I; Bizkarguenaga, E; Prieto, A; Ortiz-Zarragoitia, M; Fernández, L A; Zuloaga, O

    2015-03-27

    An analytical method for the simultaneous determination in fish liver and muscle tissue and mussel samples of 14 perfluorinated compounds (PFCs), including three perfluoroalkylsulfonates (PFSAs), seven perfluorocarboxylic acids (PFCAs), three perfluorophosphonic acids (PFPAs) and perfluorooctanesulfonamide (PFOSA), and 10 potential precursors, including four polyfluoroalkyl phosphates (PAPs), four fluorotelomer saturated acids (FTCAs) and two fluorotelomer unsaturated acids (FTUCAs), was developed in the present work. Different clean-up strategies by means of solid-phase extraction (SPE) using a mix-mode weak anion exchanger (WAX), reverse phase Envi-Carb or a combination of them was optimized and evaluated for the clean-up of focused ultrasonic solid-liquid (FUSLE) extracts before the analysis by liquid chromatography-triple quadrupole tandem mass spectrometry (LC-MS/MS). Mix-mode WAX coupled in-line to Envi-Carb was finally selected since it rendered the cleanest extracts and minimum matrix effect. The FUSLE-SPE-LC-MS/MS methodology was validated in terms of recovery, precision and method detection limits (MDLs). Apparent recovery values in the 65-116%, 59-119% and 67-126% range and MDLs in the 0.1-2.7 ng/g, 0.1-3.8 ng/g and 0.2-3.1ng/g range were obtained for liver, mussel and fish muscle tissue samples, respectively. The method developed was applied to the analysis of grey mullet liver (Chelon labrosus) and mussel (Mytilus galloprovincialis) samples from the Basque Coast (North of Spain) and Yellowfin tuna muscle tissue (Thunnus albacares) samples from the Indian Ocean. To the best of our knowledge this is the first method that describes the simultaneous determination of 14 PFCs and 10 potential precursors in fish liver, fish muscle tissue and mussel samples. Besides, this is the first time that 8:2 monosubstituted polyfluorodecyl phosphate (8:2 monoPAP) and 8:2 disubstituted polyfluorodecyl phosphate (8:2 diPAP) were detected in mussel and tuna samples

  19. Measurement of characteristic prompt gamma rays emitted from oxygen and carbon in tissue-equivalent samples during proton beam irradiation

    NASA Astrophysics Data System (ADS)

    Polf, Jerimy C.; Panthi, Rajesh; Mackin, Dennis S.; McCleskey, Matt; Saastamoinen, Antti; Roeder, Brian T.; Beddar, Sam

    2013-09-01

    The purpose of this work was to characterize how prompt gamma (PG) emission from tissue changes as a function of carbon and oxygen concentration, and to assess the feasibility of determining elemental concentration in tissues irradiated with proton beams. For this study, four tissue-equivalent water-sucrose samples with differing densities and concentrations of carbon, hydrogen, and oxygen were irradiated with a 48 MeV proton pencil beam. The PG spectrum emitted from each sample was measured using a high-purity germanium detector, and the absolute detection efficiency of the detector, average beam current, and delivered dose distribution were also measured. Changes to the total PG emission from 12C (4.44 MeV) and 16O (6.13 MeV) per incident proton and per Gray of absorbed dose were characterized as a function of carbon and oxygen concentration in the sample. The intensity of the 4.44 MeV PG emission per incident proton was found to be nearly constant for all samples regardless of their carbon concentration. However, we found that the 6.13 MeV PG emission increased linearly with the total amount (in grams) of oxygen irradiated in the sample. From the measured PG data, we determined that 1.64 × 107 oxygen PGs were emitted per gram of oxygen irradiated per Gray of absorbed dose delivered with a 48 MeV proton beam. These results indicate that the 6.13 MeV PG emission from 16O is proportional to the concentration of oxygen in tissue irradiated with proton beams, showing that it is possible to determine the concentration of oxygen within tissues irradiated with proton beams by measuring 16O PG emission.

  20. Measurement of characteristic prompt gamma rays emitted from oxygen and carbon in tissue-equivalent samples during proton beam irradiation

    PubMed Central

    Polf, Jerimy C; Panthi, Rajesh; Mackin, Dennis S; McCleskey, Matt; Saastamoinen, Antti; Roeder, Brian T; Beddar, Sam

    2013-01-01

    The purpose of this work was to characterize how prompt gamma (PG) emission from tissue changes as a function of carbon and oxygen concentration, and to assess the feasibility of determining elemental concentration in tissues irradiated with proton beams. For this study, four tissue-equivalent water-sucrose samples with differing densities and concentrations of carbon, hydrogen, and oxygen were irradiated with a 48 MeV proton pencil beam. The PG spectrum emitted from each sample was measured using a high-purity germanium detector, and the absolute detection efficiency of the detector, average beam current, and delivered dose distribution were also measured. Changes to the total PG emission from 12C (4.44 MeV) and 16O (6.13 MeV) per incident proton and per Gray of absorbed dose were characterized as a function of carbon and oxygen concentration in the sample. The intensity of the 4.44 MeV PG emission per incident proton was found to be nearly constant for all samples regardless of their carbon concentration. However, we found that the 6.13 MeV PG emission increased linearly with the total amount (in grams) of oxygen irradiated in the sample. From the measured PG data, we determined that 1.64 × 107 oxygen PGs were emitted per gram of oxygen irradiated per Gray of absorbed dose delivered with a 48 MeV proton beam. These results indicate that the 6.13 MeV PG emission from 16O is proportional to the concentration of oxygen in tissue irradiated with proton beams, showing that it is possible to determine the concentration of oxygen within tissues irradiated with proton beams by measuring 16O PG emission. PMID:23920051

  1. Stain-less staining for computed histopathology

    PubMed Central

    Mayerich, David; Walsh, Michael J.; Kadjacsy-Balla, Andre; Ray, Partha S.; Hewitt, Stephen M.; Bhargava, Rohit

    2015-01-01

    Dyes such as hematoxylin and eosin (H&E) and immunohistochemical stains have been increasingly used to visualize tissue composition in research and clinical practice. We present an alternative approach to obtain the same information using stain-free chemical imaging. Relying on Fourier transform infrared (FT-IR) spectroscopic imaging and computation, stainless computed histopathology can enable a rapid, digital, quantitative and non-perturbing visualization of morphology and multiple molecular epitopes simultaneously in a variety of research and clinical pathology applications. PMID:26029735

  2. Pathology Tissue-quantitative Mass Spectrometry Analysis to Profile Histone Post-translational Modification Patterns in Patient Samples*

    PubMed Central

    Noberini, Roberta; Uggetti, Andrea; Pruneri, Giancarlo; Minucci, Saverio

    2016-01-01

    Histone post-translational modifications (hPTMs) generate a complex combinatorial code that has been implicated with various pathologies, including cancer. Dissecting such a code in physiological and diseased states may be exploited for epigenetic biomarker discovery, but hPTM analysis in clinical samples has been hindered by technical limitations. Here, we developed a method (PAThology tissue analysis of Histones by Mass Spectrometry - PAT-H-MS) that allows to perform a comprehensive, unbiased and quantitative MS-analysis of hPTM patterns on formalin-fixed paraffin-embedded (FFPE) samples. In pairwise comparisons, histone extracted from formalin-fixed paraffin-embedded tissues showed patterns similar to fresh frozen samples for 24 differentially modified peptides from histone H3. In addition, when coupled with a histone-focused version of the super-SILAC approach, this method allows the accurate quantification of modification changes among breast cancer patient samples. As an initial application of the PAThology tissue analysis of Histones by Mass Spectrometry method, we analyzed breast cancer samples, revealing significant changes in histone H3 methylation patterns among Luminal A-like and Triple Negative disease subtypes. These results pave the way for retrospective epigenetic studies that combine the power of MS-based hPTM analysis with the extensive clinical information associated with formalin-fixed paraffin-embedded archives. PMID:26463340

  3. Probing focal cortical dysplasia in formalin fixed samples using tissue optical spectroscopy

    NASA Astrophysics Data System (ADS)

    Anand, Suresh; Cicchi, Riccardo; Giordano, Flavio; Buccoliero, Anna Maria; Conti, Valerio; Guerrini, Renzo; Pavone, Francesco Saverio

    2016-03-01

    Focal cortical dysplasia (FCD) is one of most common causes of intractable epilepsy in pediatric population and these are often insensitive to anti-epileptic drugs. FCD is characterized by a disarray in localized regions of the cerebral cortex and abnormal neurons which results them to misfire with incorrect signals. Resective neurosurgery to remove or disconnect the affected parts from the rest of the brain seems to be a viable option to treat FCD. Before neurosurgery the subject could undergo imaging studies including magnetic resonance imaging (MRI) or computed tomography (CT) scans. On the downside FCD could be elusive in MRI images and may be practically invisible in CT scans. Furthermore, unnecessary removal of normal tissues is to be taken into consideration as this could lead to neurological defects. In this context, optical spectroscopy have been widely investigated as an alternative technique for the detection of abnormal tissues in different organ sites. Disease progression is accompanied by a number of architectural, biochemical and morphological changes. These variations are reflected in the spectral intensity and line shape. Here, in this proof of concept study we propose to investigate the application of tissue optical spectroscopy based on fluorescence excitation at two wavelength 378 and 445 nm coupled along with Raman spectroscopy for the detection of FCD on formalin fixed tissue specimens from pediatric subjects. For fluorescence at both the excitation wavelengths FCD showed a decreased intensity at longer wavelength when compared to normal tissues. Also, differences exist in the Raman spectral profiles of normal and FCD.

  4. How You Can Help Medical Research: Donating Your Blood, Tissue, and Other Samples

    MedlinePlus

    ... sample to make sure the proposed research is ethical, useful, and based on good science. After the ... discuss further? • Am I comfortable with experts making decisions about how my samples will be used in ...

  5. Optical parameter measurement of highly diffusive tissue body phantoms with specifically designed sample holder for photo diagnostic and PDT applications

    NASA Astrophysics Data System (ADS)

    Rehman, A.; Rehman, K.; Anwar, S.; Firdous, S.; Nawaz, M.

    2015-12-01

    Knowledge of optical properties (absorption coefficients, scattering Coefficients, and anisotropy) is necessary for understanding light tissue interactions. Optical parameters define the behavior of light in the tissues. Intralipid and Indian ink are well-established tissue body phantoms. Quantitative characterization of biological tissues in terms of optical properties is achieved with integrating sphere. However, samples having significantly higher scattering and absorption coefficients such as malignant tissues potentially reduce the signal to noise ratio (SNR) and accuracy of integrating sphere. We have measured the diffuse reflection and transmission of these phantoms by placing them in integrating sphere at 632.8 nm and then applied IAD method to determine the optical properties tissue phantoms composed of Indian ink (1.0%) and Intralipid (20%). We have fabricated a special sample holder with thin microscopic cover slips which can be used to measure signal from highly concentrated intralipid and Indian ink solutions. Experiments conducted with various phantoms reveal significant improvement of SNR for a wide range of optical properties. This approach opens up a field for potential applications in measurement of optical properties of highly diffusive biological tissues. For 20% intralipid μa =0.112+/-0.046 cm-1 and μs =392.299+/-10.090 cm-1 at 632.8 nm and for 1.0% Indian ink μa =9.808+/-0.490 cm-1 and μs =1.258+/-0.063 cm-1 at same wavelength. System shows good repeatability and reproducibility within 4.9% error. Work may have important biomedical applications in photo-diagnosis and Photodynamic therapy.

  6. Towards a minimally invasive sampling tool for high resolution tissue analytical mapping

    NASA Astrophysics Data System (ADS)

    Gottardi, R.

    2015-09-01

    Multiple spatial mapping techniques of biological tissues have been proposed over the years, but all present limitations either in terms of resolution, analytical capacity or invasiveness. Ren et al (2015 Nanotechnology 26 284001) propose in their most recent work the use of a picosecond infrared laser (PIRL) under conditions of ultrafast desorption by impulsive vibrational excitation (DIVE) to extract small amounts of cellular and molecular components, conserving their viability, structure and activity. The PIRL DIVE technique would then work as a nanobiopsy with minimal damage to the surrounding tissues, which could potentially be applied for high resolution local structural characterization of tissues in health and disease with the spatial limit determined by the laser focus.

  7. Lack of mutation-histopathology correlation in a patient with Proteus syndrome.

    PubMed

    Doucet, Meggie E; Bloomhardt, Hadley M; Moroz, Krzysztof; Lindhurst, Marjorie J; Biesecker, Leslie G

    2016-06-01

    Proteus syndrome (PS) is characterized by progressive, disproportionate, segmental overgrowth, and tumor susceptibility caused by a somatic mosaic AKT1 activating mutation. Each individual has unique manifestations making this disorder extremely heterogeneous. We correlated three variables in 38 tissue samples from a patient who died with PS: the gross affection status, the microscopic affection status, and the mutation level. The AKT1 mutation was measured using a PCR-based RFLP assay. Thirteen samples were grossly normal; six had detectable mutation (2-29%) although four of these six were histopathologically normal. Of the seven grossly normal samples that had no mutation, only four were histologically normal. The mutation level in the grossly abnormal samples was 3-35% and all but the right and left kidneys, skull, and left knee bone, with mutation levels of 19%, 15%, 26%, and 17%, respectively, had abnormal histopathology. The highest mutation level was in a toe bone sample whereas the lowest levels were in the soft tissue surrounding that toe, and an omental fat nodule. We also show here that PS overgrowth can be caused by cellular proliferation or by extracellular matrix expansion. Additionally, papillary thyroid carcinoma was identified, a tumor not previously associated with PS. We conclude that gross pathology and histopathology correlate poorly with mutation levels in PS, that overgrowth can be mediated by cellular proliferation or extracellular matrix expansion, and that papillary thyroid carcinoma is part of the tumor susceptibility of PS. New methods need to be developed to facilitate genotype-phenotype correlation in mosaic disorders. © 2016 Wiley Periodicals, Inc.

  8. Measurement of intravenously administered γ-Fe2O3 particle amount in mice tissues using vibrating sample magnetometer.

    PubMed

    Kishimoto, Mikio; Miyamoto, Ryoichi; Oda, Tatsuya; Ohara, Yusuke; Yanagihara, Hideto; Ohkohchi, Nobuhiro; Kita, Eiji

    2014-12-01

    Dispersions of platelet γ-Fe2O3 particles 30-50nm in size were intravenously administered to mice and the amount of particles accumulated in each tissue was obtained by magnetization measurement using a vibrating sample magnetometer. Background noise was greatly reduced by measuring dried tissues under a magnetic field of 500 Oe so that the effect of diamagnetism was slight. Remarkable particle accumulation was observed in the liver and spleen. Considerable particle accumulation was observed in the lung when a large quantity of γ-Fe2 O3 particles was administered. There was no significant particle accumulation in the kidney and heart.

  9. Revised histopathological consensus classification of joint implant related pathology.

    PubMed

    Krenn, V; Morawietz, L; Perino, G; Kienapfel, H; Ascherl, R; Hassenpflug, G J; Thomsen, M; Thomas, P; Huber, M; Kendoff, D; Baumhoer, D; Krukemeyer, M G; Natu, S; Boettner, F; Zustin, J; Kölbel, B; Rüther, W; Kretzer, J P; Tiemann, A; Trampuz, A; Frommelt, L; Tichilow, R; Söder, S; Müller, S; Parvizi, J; Illgner, U; Gehrke, T

    2014-12-01

    This extended classification of joint implant related pathology is a practical histopathologic classification based on defined morphological criteria covering the complete spectrum of pathohistologic changes in periprosthetic tissues. These changes may occur as a consequence of endoprosthetic replacement of large joints and may lead to a reduction in the prosthesis survival rate. We describe the established consensus classification of the periprosthetic membrane, in which aseptic and septic prosthetic loosening can be subdivided into four histological types, as well as histopathological criteria for additional significant pathologies including endoprosthetic-associated arthrofibrosis, particle-induced immunological, inflammatory and toxic mechanisms (adverse reactions), and bone tissue pathologies. These characteristic tissue alterations and their relationships are summarized in the extended classification. Since particle heterogeneity in periprosthetic tissue is high and particle identification is a necessary part of diagnosis, the identification of different types of particles is described in the histopathological particle algorithm. The morphological qualities of prosthetic material particles and the demarcation between abrasion and non-abrasion endogenous particles are also summarized. This feasible classification which is based on low cost standard tissue processing and examination and on well-defined diagnostic criteria is a solid platform for the histological diagnosis of implant associated pathologies providing a stable and reproducible tool for the surgical pathologist. Since this classification is suitable for standardized histopathological diagnostics, it might also provide a useful data set for joint arthroplasty registers, particularly for registers based on so-called routine data.

  10. Comparison of Accuracy of Whole-Exome Sequencing with Formalin-Fixed Paraffin-Embedded and Fresh Frozen Tissue Samples

    PubMed Central

    Kwon, Mi Jeong; Kim, Ryong Nam; Kim, Yu Jin; Song, Ji-Young; Jung, Kyung Soo; Shin, Young Kee

    2015-01-01

    Formalin fixing with paraffin embedding (FFPE) has been a standard sample preparation method for decades, and archival FFPE samples are still very useful resources. Nonetheless, the use of FFPE samples in cancer genome analysis using next-generation sequencing, which is a powerful technique for the identification of genomic alterations at the nucleotide level, has been challenging due to poor DNA quality and artificial sequence alterations. In this study, we performed whole-exome sequencing of matched frozen samples and FFPE samples of tissues from 4 cancer patients and compared the next-generation sequencing data obtained from these samples. The major differences between data obtained from the 2 types of sample were the shorter insert size and artificial base alterations in the FFPE samples. A high proportion of short inserts in the FFPE samples resulted in overlapping paired reads, which could lead to overestimation of certain variants; >20% of the inserts in the FFPE samples were double sequenced. A large number of soft clipped reads was found in the sequencing data of the FFPE samples, and about 30% of total bases were soft clipped. The artificial base alterations, C>T and G>A, were observed in FFPE samples only, and the alteration rate ranged from 200 to 1,200 per 1M bases when sequencing errors were removed. Although high-confidence mutation calls in the FFPE samples were compatible to that in the frozen samples, caution should be exercised in terms of the artifacts, especially for low-confidence calls. Despite the clearly observed artifacts, archival FFPE samples can be a good resource for discovery or validation of biomarkers in cancer research based on whole-exome sequencing. PMID:26641479

  11. Comparison of Accuracy of Whole-Exome Sequencing with Formalin-Fixed Paraffin-Embedded and Fresh Frozen Tissue Samples.

    PubMed

    Oh, Ensel; Choi, Yoon-La; Kwon, Mi Jeong; Kim, Ryong Nam; Kim, Yu Jin; Song, Ji-Young; Jung, Kyung Soo; Shin, Young Kee

    2015-01-01

    Formalin fixing with paraffin embedding (FFPE) has been a standard sample preparation method for decades, and archival FFPE samples are still very useful resources. Nonetheless, the use of FFPE samples in cancer genome analysis using next-generation sequencing, which is a powerful technique for the identification of genomic alterations at the nucleotide level, has been challenging due to poor DNA quality and artificial sequence alterations. In this study, we performed whole-exome sequencing of matched frozen samples and FFPE samples of tissues from 4 cancer patients and compared the next-generation sequencing data obtained from these samples. The major differences between data obtained from the 2 types of sample were the shorter insert size and artificial base alterations in the FFPE samples. A high proportion of short inserts in the FFPE samples resulted in overlapping paired reads, which could lead to overestimation of certain variants; >20% of the inserts in the FFPE samples were double sequenced. A large number of soft clipped reads was found in the sequencing data of the FFPE samples, and about 30% of total bases were soft clipped. The artificial base alterations, C>T and G>A, were observed in FFPE samples only, and the alteration rate ranged from 200 to 1,200 per 1M bases when sequencing errors were removed. Although high-confidence mutation calls in the FFPE samples were compatible to that in the frozen samples, caution should be exercised in terms of the artifacts, especially for low-confidence calls. Despite the clearly observed artifacts, archival FFPE samples can be a good resource for discovery or validation of biomarkers in cancer research based on whole-exome sequencing.

  12. Comparison of Accuracy of Whole-Exome Sequencing with Formalin-Fixed Paraffin-Embedded and Fresh Frozen Tissue Samples.

    PubMed

    Oh, Ensel; Choi, Yoon-La; Kwon, Mi Jeong; Kim, Ryong Nam; Kim, Yu Jin; Song, Ji-Young; Jung, Kyung Soo; Shin, Young Kee

    2015-01-01

    Formalin fixing with paraffin embedding (FFPE) has been a standard sample preparation method for decades, and archival FFPE samples are still very useful resources. Nonetheless, the use of FFPE samples in cancer genome analysis using next-generation sequencing, which is a powerful technique for the identification of genomic alterations at the nucleotide level, has been challenging due to poor DNA quality and artificial sequence alterations. In this study, we performed whole-exome sequencing of matched frozen samples and FFPE samples of tissues from 4 cancer patients and compared the next-generation sequencing data obtained from these samples. The major differences between data obtained from the 2 types of sample were the shorter insert size and artificial base alterations in the FFPE samples. A high proportion of short inserts in the FFPE samples resulted in overlapping paired reads, which could lead to overestimation of certain variants; >20% of the inserts in the FFPE samples were double sequenced. A large number of soft clipped reads was found in the sequencing data of the FFPE samples, and about 30% of total bases were soft clipped. The artificial base alterations, C>T and G>A, were observed in FFPE samples only, and the alteration rate ranged from 200 to 1,200 per 1M bases when sequencing errors were removed. Although high-confidence mutation calls in the FFPE samples were compatible to that in the frozen samples, caution should be exercised in terms of the artifacts, especially for low-confidence calls. Despite the clearly observed artifacts, archival FFPE samples can be a good resource for discovery or validation of biomarkers in cancer research based on whole-exome sequencing. PMID:26641479

  13. Evaluation of a novel tagging and tissue preservation system for potential use in forensic sample collection.

    PubMed

    Grassberger, Martin; Stein, Christina; Hanslik, Stefan; Hochmeister, Manfred

    2005-07-16

    The authors describe a new, easy-to-use barcode-based tissue collection, preservation and body tracking system, which might prove instrumental in the containment of mass fatalities such as aircraft accidents, war related accidents, environmental disasters (e.g. earthquakes, hurricanes, and floods) terrorist bombings or mass murders.

  14. A novel approach in personal identification from tissue samples undergone different processes through STR typing.

    PubMed

    Staiti, N; Di Martino, D; Saravo, L

    2004-12-01

    Short tandem repeats (STRs) or microsatellites have been recognized worldwide as a powerful tool for human identification. They have become widely used in human identification especially in criminal cases and mass disasters. Police departments are often interested in cases where tissues are already decomposed and only do bones remain to let them perform laboratory analyses. Bone is the most resistant tissue in animal body to time depending degradation and putrefaction, but it is often hard to extract DNA from it because of its highly mineralized structure, which makes DNA extraction and/or amplification hard to carry out. We have performed human nuclear DNA extraction and STR typing in three different cases, on bones and bone fragments from long time dead persons found buried, in the sea, almost completely burnt and whose tissues were already decomposed. We report these caseworks as we would like to show how forensic scientists are improving their skill in identifying people whose corps have undergone several kinds of processes, even independently on the time passed and the level of putrefaction of their tissues.

  15. Evaluation of a novel tagging and tissue preservation system for potential use in forensic sample collection.

    PubMed

    Grassberger, Martin; Stein, Christina; Hanslik, Stefan; Hochmeister, Manfred

    2005-07-16

    The authors describe a new, easy-to-use barcode-based tissue collection, preservation and body tracking system, which might prove instrumental in the containment of mass fatalities such as aircraft accidents, war related accidents, environmental disasters (e.g. earthquakes, hurricanes, and floods) terrorist bombings or mass murders. PMID:15939157

  16. The focus on sample quality: Influence of colon tissue collection on reliability of qPCR data

    PubMed Central

    Korenkova, Vlasta; Slyskova, Jana; Novosadova, Vendula; Pizzamiglio, Sara; Langerova, Lucie; Bjorkman, Jens; Vycital, Ondrej; Liska, Vaclav; Levy, Miroslav; Veskrna, Karel; Vodicka, Pavel; Vodickova, Ludmila; Kubista, Mikael; Verderio, Paolo

    2016-01-01

    Successful molecular analyses of human solid tissues require intact biological material with well-preserved nucleic acids, proteins, and other cell structures. Pre-analytical handling, comprising of the collection of material at the operating theatre, is among the first critical steps that influence sample quality. The aim of this study was to compare the experimental outcomes obtained from samples collected and stored by the conventional means of snap freezing and by PAXgene Tissue System (Qiagen). These approaches were evaluated by measuring rRNA and mRNA integrity of the samples (RNA Quality Indicator and Differential Amplification Method) and by gene expression profiling. The collection procedures of the biological material were implemented in two hospitals during colon cancer surgery in order to identify the impact of the collection method on the experimental outcome. Our study shows that the pre-analytical sample handling has a significant effect on the quality of RNA and on the variability of qPCR data. PAXgene collection mode proved to be more easily implemented in the operating room and moreover the quality of RNA obtained from human colon tissues by this method is superior to the one obtained by snap freezing. PMID:27383461

  17. Identification of immune cell infiltration in hematoxylin-eosin stained breast cancer samples: texture-based classification of tissue morphologies

    NASA Astrophysics Data System (ADS)

    Turkki, Riku; Linder, Nina; Kovanen, Panu E.; Pellinen, Teijo; Lundin, Johan

    2016-03-01

    The characteristics of immune cells in the tumor microenvironment of breast cancer capture clinically important information. Despite the heterogeneity of tumor-infiltrating immune cells, it has been shown that the degree of infiltration assessed by visual evaluation of hematoxylin-eosin (H and E) stained samples has prognostic and possibly predictive value. However, quantification of the infiltration in H and E-stained tissue samples is currently dependent on visual scoring by an expert. Computer vision enables automated characterization of the components of the tumor microenvironment, and texture-based methods have successfully been used to discriminate between different tissue morphologies and cell phenotypes. In this study, we evaluate whether local binary pattern texture features with superpixel segmentation and classification with support vector machine can be utilized to identify immune cell infiltration in H and E-stained breast cancer samples. Guided with the pan-leukocyte CD45 marker, we annotated training and test sets from 20 primary breast cancer samples. In the training set of arbitrary sized image regions (n=1,116) a 3-fold cross-validation resulted in 98% accuracy and an area under the receiver-operating characteristic curve (AUC) of 0.98 to discriminate between immune cell -rich and - poor areas. In the test set (n=204), we achieved an accuracy of 96% and AUC of 0.99 to label cropped tissue regions correctly into immune cell -rich and -poor categories. The obtained results demonstrate strong discrimination between immune cell -rich and -poor tissue morphologies. The proposed method can provide a quantitative measurement of the degree of immune cell infiltration and applied to digitally scanned H and E-stained breast cancer samples for diagnostic purposes.

  18. Bisulfite-Based DNA Methylation Analysis from Recent and Archived Formalin-Fixed, Paraffin Embedded Colorectal Tissue Samples.

    PubMed

    Kalmár, Alexandra; Péterfia, Bálint; Hollósi, Péter; Wichmann, Barnabás; Bodor, András; Patai, Árpád V; Schöller, Andrea; Krenács, Tibor; Tulassay, Zsolt; Molnár, Béla

    2015-09-01

    We aimed to test the applicability of formalin-fixed and paraffin-embedded (FFPE) tissue samples for gene specific DNA methylation analysis after using two commercially available DNA isolation kits. Genomic DNA was isolated from 5 colorectal adenocarcinomas and 5 normal adjacent tissues from "recent", collected within 6 months, and "archived", collected more than 5 years ago, FFPE tissues using either High Pure FFPET DNA Isolation kit or QIAamp DNA FFPE Tissue kit. DNA methylation analysis of MAL, SFRP1 and SFRP2 genes, known to be hypermethylated in CRC, was performed using methylation-sensitive high resolution melting (MS-HRM) analysis and sequencing. QIAamp (Q) method resulted in slightly higher recovery in archived (HP: 1.22 ± 3.18 μg DNA; Q: 3.00 ± 4.04 μg DNA) and significantly (p < 0.05) higher recovery in recent samples compared to High Pure method (HP) (HP: 4.10 ± 2.91 μg DNA; Q: 11.51 ± 7.50 μg DNA). Both OD260/280 and OD260/230 ratios were lower, but still high in the High Pure isolated archived and recent samples compared to those isolated with QIAamp. Identical DNA methylation patterns were detected for all 3 genes tested by MS-HRM with both isolation kits in the recent group. However, despite of higher DNA recovery in QIAamp slightly more reproducible methylation results were obtained from High Pure isolated archived samples. Sequencing confirmed DNA hypermethylation in CRCs. In conclusion, reproducible DNA methylation patterns were obtained from recent samples using both isolation kits. However, long term storage may affect the reliability of the results leading to moderate differences between the efficiency of isolation kits.

  19. Bisulfite-Based DNA Methylation Analysis from Recent and Archived Formalin-Fixed, Paraffin Embedded Colorectal Tissue Samples.

    PubMed

    Kalmár, Alexandra; Péterfia, Bálint; Hollósi, Péter; Wichmann, Barnabás; Bodor, András; Patai, Árpád V; Schöller, Andrea; Krenács, Tibor; Tulassay, Zsolt; Molnár, Béla

    2015-09-01

    We aimed to test the applicability of formalin-fixed and paraffin-embedded (FFPE) tissue samples for gene specific DNA methylation analysis after using two commercially available DNA isolation kits. Genomic DNA was isolated from 5 colorectal adenocarcinomas and 5 normal adjacent tissues from "recent", collected within 6 months, and "archived", collected more than 5 years ago, FFPE tissues using either High Pure FFPET DNA Isolation kit or QIAamp DNA FFPE Tissue kit. DNA methylation analysis of MAL, SFRP1 and SFRP2 genes, known to be hypermethylated in CRC, was performed using methylation-sensitive high resolution melting (MS-HRM) analysis and sequencing. QIAamp (Q) method resulted in slightly higher recovery in archived (HP: 1.22 ± 3.18 μg DNA; Q: 3.00 ± 4.04 μg DNA) and significantly (p < 0.05) higher recovery in recent samples compared to High Pure method (HP) (HP: 4.10 ± 2.91 μg DNA; Q: 11.51 ± 7.50 μg DNA). Both OD260/280 and OD260/230 ratios were lower, but still high in the High Pure isolated archived and recent samples compared to those isolated with QIAamp. Identical DNA methylation patterns were detected for all 3 genes tested by MS-HRM with both isolation kits in the recent group. However, despite of higher DNA recovery in QIAamp slightly more reproducible methylation results were obtained from High Pure isolated archived samples. Sequencing confirmed DNA hypermethylation in CRCs. In conclusion, reproducible DNA methylation patterns were obtained from recent samples using both isolation kits. However, long term storage may affect the reliability of the results leading to moderate differences between the efficiency of isolation kits. PMID:25991403

  20. Histopathologic and histomorphometric studies and determination of IL-8 in patients with periodontal disease

    PubMed Central

    Koss, Myriam A.; Castro, Cecilia E.; Carino, Silvia; López, Maria E.

    2014-01-01

    Background: Periodontitis is an inflammatory disease characterized by connective tissue breakdown and alveolar bone resorption. Objective: The aim of this study was to make a quantitative evaluation of the gingival tissue components in biopsies from patients with different clinical states of periodontal disease and to determine the relationship between the presence of interleukin-8 and the tissue destruction. Materials and Methods: The study group consisted of 33 biopsies from adult subjects. A total of 25 periodontal patients were sub-classified on the basis of the clinical and radiographic criteria in mild, moderate and advanced periodontitis. Gingival samples were obtained from patients in the course of basic periodontal surgeries as a muco-periostic collate. Data were analyzed by the SPSS system. Results: Collagen fibres decreased as non-specific chronic states increased. A lymphocyte inflammatory infiltrate changed to a lymphoplasmocytary form. Reactivity to interleukin-8 was detected with the severity of the histopathologic diagnoses; however there was no association with the clinical diagnoses. Conclusions: Histopathology and histomorphometry of gingival tissue changed with the degree of inflammation. No evidence of interleukin-8 as a biomarker for clinical diagnoses of periodontitis was obtained. PMID:24872619

  1. Histopathology and MR image fusion of the prostate

    NASA Astrophysics Data System (ADS)

    Jo, Hyun Hee; Jung, Julip; Jang, Yujin; Hong, Helen; Lee, Hak Jong

    2008-03-01

    We propose a method for combining histopathology image with MR image of the prostate by using image correction and nonrigid registration. Our method consists of four steps. First, two or four tissue sections of the prostate in histopathology image are combined to produce a single prostate image by stitching. Second, the intensity of prostate bleeding area on T2-weighted MR image is substituted for that on T1-weighted MR image. Our intensity correction prevents a mistake which a prostate bleeding is considered as a tumor on T2-weighted MR image. Third, rough and fine registration is performed to find the best match for pixel overlap between histopathology and MR images. Then the result of rigid registration is deformed by the TPS warping. Finally, aligned images are visualized by the intensity intermixing. Experimental results show that the prostate tumor lesion can be properly located and clearly visualized within MR images for tissue characterization comparison.

  2. Three-dimensional digital breast histopathology imaging

    NASA Astrophysics Data System (ADS)

    Clarke, G. M.; Peressotti, C.; Mawdsley, G. E.; Eidt, S.; Ge, M.; Morgan, T.; Zubovits, J. T.; Yaffe, M. J.

    2005-04-01

    We have developed a digital histology imaging system that has the potential to improve the accuracy of surgical margin assessment in the treatment of breast cancer by providing finer sampling and 3D visualization. The system is capable of producing a 3D representation of histopathology from an entire lumpectomy specimen. We acquire digital photomicrographs of a stack of large (120 x 170 mm) histology slides cut serially through the entire specimen. The images are then registered and displayed in 2D and 3D. This approach dramatically improves sampling and can improve visualization of tissue structures compared to current, small-format histology. The system consists of a brightfield microscope, adapted with a freeze-frame digital video camera and a large, motorized translation stage. The image of each slide is acquired as a mosaic of adjacent tiles, each tile representing one field-of-view of the microscope, and the mosaic is assembled into a seamless composite image. The assembly is done by a program developed to build image sets at six different levels within a multiresolution pyramid. A database-linked viewing program has been created to efficiently register and display the animated stack of images, which occupies about 80 GB of disk space per lumpectomy at full resolution, on a high-resolution (3840 x 2400 pixels) colour monitor. The scanning or tiling approach to digitization is inherently susceptible to two artefacts which disrupt the composite image, and which impose more stringent requirements on system performance. Although non-uniform illumination across any one isolated tile may not be discernible, the eye readily detects this non-uniformity when the entire assembly of tiles is viewed. The pattern is caused by deficiencies in optical alignment, spectrum of the light source, or camera corrections. The imaging task requires that features as small as 3.2 &mum in extent be seamlessly preserved. However, inadequate accuracy in positioning of the translation

  3. Histopathological baseline levels and confounding factors in common sole (Solea solea) for marine environmental risk assessment.

    PubMed

    Cuevas, N; Zorita, I; Costa, P M; Larreta, J; Franco, J

    2015-09-01

    Liver and gonad histopathology, biometric parameters and hepatic metal bioaccumulation were assessed monthly over a one-year period in common soles from the Basque continental shelf, in order to determine baseline levels and confounding factors within biomonitoring studies. Biometric parameters and hepatic metal bioaccumulation varied according to season and gender. Accordingly, hepatic histopathological traits presented seasonal variations related to the reproductive cycle. However, the hepatic histopathological index showed that seasonality and gender were not significant confounding factors. Conversely, the gonad histopathological index was modulated by season and gender. As for organ comparison, the liver endured more severe histopathological damage than the gonad. In brief, the sampling period and gender may not affect the estimation of hepatic histopathological indices for biomonitoring purposes. Nonetheless, due to different sensitivities to environmental 'noise' variables, the sampling period and gender differentiation should be thoroughly considered for the assessment of gonad histopathology, biometrics and metal bioaccumulation. PMID:26364682

  4. Limit of Detection in X-ray Diffraction Measurements of Tissue Equivalent Samples

    NASA Astrophysics Data System (ADS)

    Zheng, Y.; Vassiljev, N.; Konstantinidis, A.; Griffiths, J.; Speller, R.

    2015-09-01

    There is a suggestion of a new approach to mammography whereby following a conventional mammogram, the radiologist could interrogate suspicious regions using X-ray diffraction whilst the patient is still present and to establish the true extent of disease. A starting point for this work is to quantify the minimum detectable amount of breast cancer within a realistic thickness phantom. Perspex has a similar diffraction pattern to healthy breast tissue whilst water is similar to breast tumour, hence these two materials are used as tissue equivalent test objects for X-ray diffraction measurements. The preliminary results show linear agreement between the ratio of Perspex to water and the ratio of the diffraction peak intensities at 0.7 nm-1 and 1.5 nm-1. The minimum detectable limit for a component of the two ‘tissue’ mix was found to be 4.1%. This suggests that X-ray diffraction can be used to quantify tissue like mixtures down to the 4.1% / 95.9% mix level and hence has a strong potential for delineating the extent of infiltration disease.

  5. Histopathological characterization of a Cameron lesion.

    PubMed

    Katz, Jordan; Brar, Sonia; Sidhu, Jagmohan S

    2012-10-01

    Cameron lesions are linear erosions located at the neck of a hiatal hernia (HH) in patients with a large HH. The prevalence has been seen in up to 5% of patients with HH who undergo esophagogastroduodenoscopy, and they can be associated with overt gastrointestinal bleeding or anemia. These lesions occur due to vascular compression by the diaphragm in a large sliding HH. Histopathologic changes seen in the biopsy tissue of a Cameron lesion are due to ischemia, but this ischemia is reversible with treatment of HH. The existence of this entity and the histopathologic picture of a Cameron lesion is not well known to pathologists, and therefore, a microscopic picture of a Cameron lesion can be easily confused with ischemic gastritis. Ischemic gastritis is the result of atherosclerosis, usually seen in older people, unrelated to HH, and is not easily reversible. The authors received a gastric biopsy of a hiatal hernia without any associated clinical diagnosis of a Cameron lesion conveyed to the pathologist. This biopsy tissue showed ischemic changes in the gastric mucosa on microscopic examination. Diagnosis of ischemic gastritis was considered but ruled out after the case was discussed with the gastroenterologist. The correct diagnosis was made once the clinical diagnosis of HH with Cameron lesion (ie, a vertical red erosion) was made known to the pathologist. By reporting this case, the authors aim to increase awareness of Cameron lesion among pathologists so that they ask about the presence of a Cameron lesion before making the diagnosis of ischemic gastritis.

  6. Aspergillus PCR-Based Investigation of Fresh Tissue and Effusion Samples in Patients with Suspected Invasive Aspergillosis Enhances Diagnostic Capabilities

    PubMed Central

    Reinwald, M.; Spiess, B.; Heinz, W. J.; Heussel, C. P.; Bertz, H.; Cornely, O. A.; Hahn, J.; Lehrnbecher, T.; Kiehl, M.; Laws, H. J.; Wolf, H. H.; Schwerdtfeger, R.; Schultheis, B.; Burchardt, A.; Klein, M.; Dürken, M.; Claus, B.; Schlegel, F.; Hummel, M.; Hofmann, W.-K.

    2013-01-01

    Although it is a severe complication in immunocompromised patients, diagnosing invasive fungal disease (IFD), especially invasive aspergillosis (IA), remains difficult. In certain clinical scenarios, examining tissue samples for identification of the infectious organism becomes important. As culture-based methods rarely yield results, the performance of an Aspergillus-specific nested PCR in fresh tissue or pleural effusion samples was evaluated. Fresh tissue (n = 59) and effusion (n = 47) specimens from 79 immunocompromised patients were subjected to an Aspergillus-specific PCR assay. Twenty-six patients had proven (n = 20) or probable (n = 6) IFD, according to the European Organization for Research and Treatment of Cancer/Invasive Fungal Infections Cooperative Group and National Institute of Allergy and Infectious Diseases Mycoses Study Group (EORTC/MSG) criteria, while the remaining patients were classified as having either possible IFD (n = 30) or no IFD (n = 23). IA was identified as the underlying IFD in 21/26 proven/probable cases. PCR positivity was observed for 18/21 proven/probable and 6 possible IA cases; cases classified as no IA did not show positive signals. Patients with proven IFD (n = 5) with cultures positive for non-Aspergillus molds also had negative Aspergillus PCR results. Aspergillus PCR performance analysis yielded sensitivity and specificity values of 86% (95% confidence interval [CI], 65% to 95%) and 100% (95% CI, 86% to 100%), respectively, thus leading to a diagnostic odds ratio of >200. In this analysis, good diagnostic performance of the PCR assay for detection of IA was observed for tissue samples, while effusion samples showed lower sensitivity rates. PCR testing represents a complementary tool; a positive PCR result strengthens the likelihood of IA, whereas IA seems unlikely in cases with negative results but findings could indicate non-Aspergillus IFD. Thus, PCR testing of these specimens enhances the diagnostic capabilities. PMID

  7. Storage and shipping of tissue samples for DNA analyses: A case study on earthworms☆

    PubMed Central

    Straube, Daniela; Juen, Anita

    2013-01-01

    Nowadays, molecular analyses play an important role in studies of soil dwelling animals, for example in taxonomy, phylogeography or food web analyses. The quality of the DNA, used for later molecular analyses, is an important factor and depends on collection and preservation of samples prior to DNA extraction. Ideally, DNA samples are frozen immediately upon collection, but if samples are collected in the field, suitable preservation methods might be limited due to unavailability of resources or remote field sites. Moreover, shipping samples over long distances can cause loss of DNA quality e.g. by thawing or leaking of preservation liquid. In this study we use earthworms, a key organism in soil research, to compare three different DNA preservation methods – freezing at −20 °C, storing in 75% ethanol, and freeze drying. Samples were shipped from the United States of America to Austria. The DNA of the samples was extracted using two different extraction methods, peqGOLD™ and Chelex® 100. The DNA amplification success was determined by amplifying four DNA fragments of different length. The PCR amplification success is significantly influenced by preservation method and extraction method and differed significantly depending on the length of the DNA fragment. Freeze drying samples was the best preservation method when samples were extracted using the silica based extraction method peqGOLD™. For samples that were extracted with Chelex® 100, storage in ethanol was the best preservation method. However, the overall amplification success was significantly lower for the extraction procedure based on Chelex® 100. The detection of the small DNA fragments was higher and independent from the extraction method, while the amplification success was significantly reduced for the longer DNA fragments. We recommend freeze drying of DNA samples, especially when they have to be shipped for longer distances. No special packaging or declaration is needed for freeze dried

  8. Polychlorinated dibenzo-p-dioxins, dibenzofurans, and polychlorinated biphenyls in human tissues, meat, fish, and wildlife samples from India.

    PubMed

    Kumar, K S; Kannan, K; Paramasivan, O N; Shanmuga Sundaram, V P; Nakanishi, J; Masunaga, S

    2001-09-01

    Concentrations of polychlorinated dibenzo-p-dioxins (PCDDs), dibenzofurans (PCDFs), and non- and mono-ortho-substituted polychlorinated biphenyls (dioxin-like PCBs) were measured in tissues of humans, fishes, chicken, lamb, goat, predatory birds, and Ganges River dolphins collected from various locations in India. PCDDs/DFs were found in most of the samples analyzed with the liver of spotted owlet containing the highest concentration of 3,300 pg/g, fat wt. 2,3,7,8-Substituted PCDDs and PCDFs were found in human fattissues at concentrations ranging from 170 to 1,300 pg/g, fat wt. Concentrations of PCDDs were generally greaterthan those of PCDFs in human tissues, fishes, animal fat, and dolphin. Among fishes, meat, and wildlife samples analyzed, concentrations of PCDDs/DFs were found in the following order: country chicken < goat/lamb fat < fishes < river dolphins < predatory birds. Hepta-CDDs and OCDD were the major PCDD homologues found in humans, fishes, meat products, and dolphins. 2,3,7,8-Tetrachlorodibenzo-p-dioxin equivalents of PCDDs/DFs were greater than those of PCBs in selected fish, dolphin, and human samples. To our knowledge, this is the first report of PCDDs and PCDFs in human tissues, fishes, meat, and wildlife collected from India. PMID:11563645

  9. Histopathological changes in the gills and liver of Prochilodus lineatus from the Salado River basin (Santa Fe, Argentina).

    PubMed

    Troncoso, Ileana C; Cazenave, Jimena; Bacchetta, Carla; Bistoni, María de Los Angeles

    2012-06-01

    This study evaluated the histopathological changes in gills and liver of Prochilodus lineatus inhabiting the Salado River basin. Fish were collected in four different sampling stations. The histological lesions in the tissues were examined under light microscopy and evaluated with quantitative analyses. The morphometric analysis of the gills showed a significant shortening of secondary lamellae and a lower percentage of area for gas exchange in fish from station 1 (an urban area, located near the mouth of the Salado River) in comparison with fish gills from the reference site (station 4, a relatively pristine area). Moreover, a significantly higher area occupied with necrotic foci and the occurrence of an important inflammatory response were observed in fish liver of station 1 than the samples caught from other stations. Thus, histopathological evidences showed differences among sites, which could be related to different environmental conditions. PMID:21850399

  10. NOTE: Image analysis can be used to detect spatial changes in the histopathology of pancreatic tumours

    NASA Astrophysics Data System (ADS)

    Sims, A. J.; Bennett, M. K.; Murray, A.

    2003-07-01

    Pancreatic cancer is frequently associated with intense growth of fibrous tissue at the periphery of tumours, but the histopathological quantification of this stromal reaction has not yet been used as a prognostic factor because of the difficulty of obtaining quantitative measures using manual methods. Manual histological grading is a poor indicator of outcome in this type of cancer and there is a clinical need to establish a more sensitive indicator. Recent pancreatic tumour biology research has focused upon the stromal reaction and there is an indication that its histopathological quantification may lead to a new prognostic indicator. Histological samples from 21 cases of pancreatic carcinoma were stained using the sirius red, light-green method. Multiple images from the centre and periphery of each tumour were automatically segmented using colour cluster analysis to subdivide each image into representative colours. These were classified manually as stroma, cell cytoplasm or lumen in order to measure the area of each component in each image. Measured areas were analysed to determine whether the technique could detect spatial differences in the area of each tissue component over all samples, and within individual samples. Over all 21 cases, the area of stromal tissue at the periphery of the tumours exceeded that at the centre by an average of 10.0 percentage points (P < 0.001). Within individual tumours, the algorithm was able to detect significantly more stroma (P < 0.05) at the periphery than the centre in 11 cases, whilst none of the remaining cases had significantly more stromal tissue at the centre than the periphery. The results demonstrate that semi-automated analysis can be used to detect spatial differences in the area of fibrous tissue in routinely stained sections of pancreatic cancer.

  11. X-Ray Fluorescence Microscopy for Investigation of Archival Tissues

    PubMed Central

    Paunesku, T.; Wanzer, M. B.; Kirillova, E. N.; Muksinova, K. N.; Revina, V. S.; Romanov, S. A.; Lyubchansky, E. R.; Grosche, B.; Birschwilks, M.; Vogt, S.; Finney, L.; Woloschak, G. E.

    2013-01-01

    Several recent efforts in radiation biology community worldwide have amassed records and archival tissues from animals exposed to different radionuclides and external beam irradiation. In most cases, these samples come from life-long studies on large animal populations conducted in national laboratories and equivalent institutions throughout Europe, North America, and Japan. While many of these tissues were used for histopathological analyses, much more information may still be obtained from these samples. A new technique suitable for imaging of these tissues is X-Ray Fluorescence Microscopy (XFM). Following development of third generation synchrotrons, XFM has emerged as an ideal technique for study of metal content, speciation, and localization in cells, tissues and organs. Here we review some of the recent XFM literature pertinent to tissue sample studies and present examples of XFM data obtained from tissue sections of beagle dog samples which show that the quality of archival tissues allows XFM investigation. PMID:22951477

  12. Comparing paraffined and deparaffinized breast cancer tissue samples and an analysis of Raman spectroscopy and infrared methods

    NASA Astrophysics Data System (ADS)

    Depciuch, J.; Kaznowska, E.; Szmuc, K.; Zawlik, I.; Cholewa, M.; Heraud, P.; Cebulski, J.

    2016-05-01

    Breast cancer makes up a quarter of all cancer in women, which is why research into new diagnostic methods and sample preparations need to be developed at an accelerated pace. Researchers are looking for diagnostic tools to detect when an individual has cancer cells and use that information to see what measurements and approaches can be used to take further diagnostic steps. The most common method of sample preparation is the imbibing of tumor tissue in paraffin, which can produce a background for spectroscopic measurements in the range of 500-3500 cm-1. In this study we demonstrated that proper preparation of paraffin-embedded specimens and the measurement methodology can eliminate paraffin vibration, as was done in the work Depciuch et al. 2015. Thanks to this spectroscopic technique there may become a reliable and accurate method of diagnosing breast cancer based on the evidence found from the prepared samples. The study compared the results obtained through Raman spectroscopy and FTIR (Fourier Transform Infrared) measurements of healthy and cancerous breast tissues that were either embedded in paraffin or deparaffinized. The resulting spectrum and accurate analysis led to the conclusion that the appropriate measurement of the background and the elimination of peaks from the paraffin had the greatest impact on the reliability of results. Furthermore, after the accurate, detailed studies FTIR and Raman spectroscopy on samples of breast tissue that were deparaffinized or embedded in paraffin, including a complete analysis of the peak after transformation Kramers-Kröning (KK), it was found that sample preparation did not affect the result obtained by measuring the reflectance in the mid-infrared range, and that this only had a minimal effect relating to the intensity obtained by the measurement of the Raman peak. Only in special cases, when Raman spectroscopic methods are used for research to find the peculiarities of the spectra, are deparaffinization recommended

  13. Pathologic evaluation of a new endoscopic ultrasound needle designed to obtain core tissue samples: A pilot study

    PubMed Central

    Adler, Douglas G.; Witt, Benjamin; Chadwick, Barbara; Wells, Jason; Taylor, Linda Jo; Dimaio, Christopher; Schmidt, Robert

    2016-01-01

    Background and Objectives: Standard endoscopic ultrasound-fine-needle aspiration (EUS-FNA) needles are in widespread use. Meaningful differences between the available needles have been difficult to identify. Recently, a new EUS needle (Shark Core®, Covidien, Dublin, Leinster, Ireland), has been introduced in an attempt to improve diagnostic accuracy, tissue yield, and to potentially obtain a core tissue sample. We performed a pilot study prospectively to evaluate this new needle when compared to a standard EUS-FNA needle. Materials and Methods: Analysis of the first 15 patients undergoing EUS-FNA with the Shark Core needle was performed and it was compared to EUS-FNA in 15 patients who underwent EUS-FNA with a standard needle. Results: The Shark Core needle required fewer needle passes to obtain diagnostic adequacy than the standard needle [(χ2(1) = 11.3, P < 0.001]. The Shark Core needle required 1.5 passes to reach adequacy, whereas the standard needle required three passes. For cases with cell blocks, the Shark Core needle produced diagnostic material in 85% of cases [95% confidence interval (CI): 54–98], whereas the standard needle produced diagnostic material in 38% of the cases (95% CI: 9-76). The Shark Core needle produced actual tissue cores 82% of the time (95% CI: 48–98) and the standard needle produced no tissue cores (95% CI: 0-71) (P = 0.03). Conclusion: This pilot study found that the Shark Core needle had a high rate of producing adequate cytologic material for the diagnosis of pancreatic and peri-pancreatic lesions sampled by EUS with fewer passes required to obtain a definitive diagnosis and with a high rate of tissue cores being obtained when compared to a standard FNA needle. PMID:27386475

  14. Equine Odontoclastic Tooth Resorption and Hypercementosis: Histopathologic Features.

    PubMed

    Smedley, R C; Earley, E T; Galloway, S S; Baratt, R M; Rawlinson, J E

    2015-09-01

    Equine odontoclastic tooth resorption and hypercementosis (EOTRH) is a painful progressive condition of older horses that involves multiple teeth, including canines and incisors. EOTRH is uncommonly recognized by veterinary pathologists and in some cases may be misdiagnosed as cementoblastoma. The cause is unknown. The goals of this study were to describe the histopathologic features of EOTRH in 17 affected horses from the United States and to increase awareness of this condition. Samples ranged from affected tooth to the entire rostral mandible and maxilla. Affected teeth exhibited cemental hyperplasia and lysis. The marked proliferation of cementum in severe cases caused bulbous enlargement of the intra-alveolar portions of affected teeth. Several teeth contained necrotic debris, bacteria, and plant material in the regions of cemental lysis. All horses exhibited dentinal lysis in at least affected tooth, and several contained necrotic debris in these regions. Endodontic disease was often present with inflammation, lysis, necrotic debris, fibrosis, and/or a thin rim of atubular mineralized tissue in the pulp cavity. Periodontal disease was a common feature that was primarily characterized by moderate lymphoplasmacytic inflammation. Resorption with secondary hypercementosis appears to begin on the external surface of the teeth rather than within the pulp cavity. Distinguishing EOTRH from other diseases requires a complete history that includes the number and location of affected teeth, a gross description of regional hard/soft tissue health, and radiographic findings.

  15. Redescription of Onchocerca lupi (Spirurida: Onchocercidae) with histopathological observations

    PubMed Central

    2013-01-01

    Background Onchocerca lupi is a dog parasite of increasing zoonotic concern, with new human cases diagnosed in Turkey, Tunisia, Iran, and the United States. Information about the morphology of this nematode is scant and a detailed re-description of this species is overdue. In addition, histopathological data of potential usefulness for the identification of O. lupi infections are provided. Methods Male and female nematodes, collected from the connective tissue of a dog, were examined using light microscopy and scanning electron microscopy (SEM), and an histological evaluation was performed on biopsy samples from periocular tissues. Results The morphological identification was confirmed by molecular amplification and partial sequencing of cytochrome oxidase subunit 1 gene. This study provides the first comprehensive morphological and morphometric description of O. lupi from a dog based on light microscopy, SEM, molecular characterization, and histological observations. Conclusions Data herein presented contribute to a better understanding of this little known parasitic zoonosis, whose impact on human and animal health is still underestimated. The presence of granulomatous reactions only around the female adult suggests that the release of microfilariae from the uterus might be the cause of the inflammatory reaction observed. PMID:24499611

  16. Histopathology of ultrashort-laser-pulse retinal damage

    NASA Astrophysics Data System (ADS)

    Toth, Cynthia A.; Narayan, Drew G.; Osborne, Catherine; Rockwell, Benjamin A.; Stein, Cindy D.; Amnotte, Rodney E.; DiCarlo, Cheryl D.; Roach, William P.; Noojin, Gary D.; Cain, Clarence P.

    1996-05-01

    Recent studies of retinal damage due to ultrashort laser pulses have shown interesting behavior. Laser induced retinal damage for ultrashort (i.e. less than 1 ns) laser pulses is produced at lower energies than in the nanosecond to microsecond laser pulse regime and the energy required for hemorrhagic lesions is much greater times greater for the nanosecond regime. We investigated the tissue effects exhibited in histopathology of retinal tissues exposed to ultrashort laser pulses.

  17. Determination of protein carbonyls in plasma, cell extracts, tissue homogenates, isolated proteins: Focus on sample preparation and derivatization conditions

    PubMed Central

    Weber, Daniela; Davies, Michael J.; Grune, Tilman

    2015-01-01

    Protein oxidation is involved in regulatory physiological events as well as in damage to tissues and is thought to play a key role in the pathophysiology of diseases and in the aging process. Protein-bound carbonyls represent a marker of global protein oxidation, as they are generated by multiple different reactive oxygen species in blood, tissues and cells. Sample preparation and stabilization are key steps in the accurate quantification of oxidation-related products and examination of physiological/pathological processes. This review therefore focuses on the sample preparation processes used in the most relevant methods to detect protein carbonyls after derivatization with 2,4-dinitrophenylhydrazine with an emphasis on measurement in plasma, cells, organ homogenates, isolated proteins and organelles. Sample preparation, derivatization conditions and protein handling are presented for the spectrophotometric and HPLC method as well as for immunoblotting and ELISA. An extensive overview covering these methods in previously published articles is given for researchers who plan to measure protein carbonyls in different samples. PMID:26141921

  18. MicroRNA Stability in Postmortem FFPE Tissues: Quantitative Analysis Using Autoptic Samples from Acute Myocardial Infarction Patients.

    PubMed

    Kakimoto, Yu; Kamiguchi, Hiroshi; Ochiai, Eriko; Satoh, Fumiko; Osawa, Motoki

    2015-01-01

    MicroRNAs (miRNAs) are very short (18-24 nucleotides) nucleic acids that are expressed in a number of biological tissues and have been shown to be more resistant to extreme temperatures and pH compared to longer RNA molecules, like mRNAs. As miRNAs contribute to diverse biological process and respond to various kinds of cellular stress, their utility as diagnostic biomarkers and/or therapeutic targets has recently been explored. Here, we have evaluated the usefulness of miRNA quantification during postmortem examination of cardiac tissue from acute myocardial infarction (AMI) patients. Cardiac tissue was collected within one week of the patient's death and either frozen (19 samples) or fixed in formalin for up to three years (36 samples). RNA integrity was evaluated with an electropherogram, and it appears that longer RNAs are fragmented after death in the long-term fixed samples. Quantitative PCR was also performed for seven miRNAs and three other small RNAs in order to determine the appropriate controls for our postmortem analysis. Our data indicate that miR-191 and miR-26b are more suitable than the other types of small RNA molecules as they are stably detected after death and long-term fixation. Further, we also applied our quantitation method, using these endogenous controls, to evaluate the expression of three previously identified miRNA biomarkers, miR-1, miR-208b, and miR-499a, in formalin-fixed tissues from AMI patients. Although miR-1 and miR-208b decreased (1.4-fold) and increased (1.2-fold), respectively, in the AMI samples compared to the controls, the significance of these changes was limited by our sample size. In contrast, the relative level of miR-499a was significantly decreased in the AMI samples (2.1-fold). This study highlights the stability of miRNAs after death and long-term fixation, validating their use as reliable biomarkers for AMI during postmortem examination. PMID:26046358

  19. Determination of five nitroimidazole residues in artificial porcine muscle tissue samples by capillary electrophoresis.

    PubMed

    Lin, Yingyun; Su, Yan; Liao, Xiulin; Yang, Na; Yang, Xiupei; Choi, Martin M F

    2012-01-15

    A capillary electrophoresis (CE) method with ultraviolet detection has been developed for simultaneous detection and quantification of five nitroimidazoles including benzoylmetronidazole, dimetridazole, metronidazole, ronidazole, and secnidazole in porcine muscles. Nitroimidazoles in samples were extracted by ethyl acetate with subsequent clean-up by a strong cation exchange solid phase extraction column. The clean extracts were subjected to CE separation with optimal experimental conditions: pH 3.0 running buffer containing 25mM sodium phosphate and 0.10mM tetrabutylammonium bromide, 5s hydrodynamic injection at 0.5psi and 28kV separation voltage. The nitroimidazoles could be monitored and detected at 320nm within 18min. The limits of detection were below 1.0μg/kg and limits of quantification were lower than 3.2μg/kg for all nitroimidazoles in the muscle samples. The recoveries and relative standard deviations were 85.4-96.0, 83.5-92.5, 1.3-3.9, and 1.1-4.2%, respectively for the intra-day and inter-day analyses. The proposed CE method has been successfully applied to determine nitroimidazoles in artificial porcine muscle samples with good accuracy and recovery, demonstrating that it has potential for detection and quantification of multi-nitroimidazole residue in real muscle samples. PMID:22265553

  20. Biodistribution Analysis of Oncolytic Adenoviruses in Patient Autopsy Samples Reveals Vascular Transduction of Noninjected Tumors and Tissues

    PubMed Central

    Koski, Anniina; Bramante, Simona; Kipar, Anja; Oksanen, Minna; Juhila, Juuso; Vassilev, Lotta; Joensuu, Timo; Kanerva, Anna; Hemminki, Akseli

    2015-01-01

    In clinical trials with oncolytic adenoviruses, there has been no mortality associated with treatment vectors. Likewise, in the Advanced Therapy Access Program (ATAP), where 290 patients were treated with 10 different viruses, no vector-related mortality was observed. However, as the patient population who received adenovirus treatments in ATAP represented heavily pretreated patients, often with very advanced disease, some patients died relatively soon after receiving their virus treatment mandating autopsy to investigate cause of death. Eleven such autopsies were performed and confirmed disease progression as the cause of death in each case. The regulatory requirement for investigating the safety of advanced therapy medical products presented a unique opportunity to study tissue samples collected as a routine part of the autopsies. Oncolytic adenoviral DNA was recovered in a wide range of tissues, including injected and noninjected tumors and various normal tissues, demonstrating the ability of the vector to disseminate through the vascular route. Furthermore, we recovered and cultured viable virus from samples of noninjected brain metastases of an intravenously treated patient, confirming that oncolytic adenovirus can reach tumors through the intravascular route. Data presented here give mechanistic insight into mode of action and biodistribution of oncolytic adenoviruses in cancer patients. PMID:26156245

  1. Digital simulation of staining in histopathology multispectral images: enhancement and linear transformation of spectral transmittance.

    PubMed

    Bautista, Pinky A; Yagi, Yukako

    2012-05-01

    Hematoxylin and eosin (H&E) stain is currently the most popular for routine histopathology staining. Special and/or immuno-histochemical (IHC) staining is often requested to further corroborate the initial diagnosis on H&E stained tissue sections. Digital simulation of staining (or digital staining) can be a very valuable tool to produce the desired stained images from the H&E stained tissue sections instantaneously. We present an approach to digital staining of histopathology multispectral images by combining the effects of spectral enhancement and spectral transformation. Spectral enhancement is accomplished by shifting the N-band original spectrum of the multispectral pixel with the weighted difference between the pixel's original and estimated spectrum; the spectrum is estimated using M < N principal component (PC) vectors. The pixel's enhanced spectrum is transformed to the spectral configuration associated to its reaction to a specific stain by utilizing an N × N transformation matrix, which is derived through application of least mean squares method to the enhanced and target spectral transmittance samples of the different tissue components found in the image. Results of our experiments on the digital conversion of an H&E stained multispectral image to its Masson's trichrome stained equivalent show the viability of the method.

  2. Digital simulation of staining in histopathology multispectral images: enhancement and linear transformation of spectral transmittance

    NASA Astrophysics Data System (ADS)

    Bautista, Pinky A.; Yagi, Yukako

    2012-05-01

    Hematoxylin and eosin (H&E) stain is currently the most popular for routine histopathology staining. Special and/or immuno-histochemical (IHC) staining is often requested to further corroborate the initial diagnosis on H&E stained tissue sections. Digital simulation of staining (or digital staining) can be a very valuable tool to produce the desired stained images from the H&E stained tissue sections instantaneously. We present an approach to digital staining of histopathology multispectral images by combining the effects of spectral enhancement and spectral transformation. Spectral enhancement is accomplished by shifting the N-band original spectrum of the multispectral pixel with the weighted difference between the pixel's original and estimated spectrum; the spectrum is estimated using M < N principal component (PC) vectors. The pixel's enhanced spectrum is transformed to the spectral configuration associated to its reaction to a specific stain by utilizing an N × N transformation matrix, which is derived through application of least mean squares method to the enhanced and target spectral transmittance samples of the different tissue components found in the image. Results of our experiments on the digital conversion of an H&E stained multispectral image to its Masson's trichrome stained equivalent show the viability of the method.

  3. Digital simulation of staining in histopathology multispectral images: enhancement and linear transformation of spectral transmittance.

    PubMed

    Bautista, Pinky A; Yagi, Yukako

    2012-05-01

    Hematoxylin and eosin (H&E) stain is currently the most popular for routine histopathology staining. Special and/or immuno-histochemical (IHC) staining is often requested to further corroborate the initial diagnosis on H&E stained tissue sections. Digital simulation of staining (or digital staining) can be a very valuable tool to produce the desired stained images from the H&E stained tissue sections instantaneously. We present an approach to digital staining of histopathology multispectral images by combining the effects of spectral enhancement and spectral transformation. Spectral enhancement is accomplished by shifting the N-band original spectrum of the multispectral pixel with the weighted difference between the pixel's original and estimated spectrum; the spectrum is estimated using M < N principal component (PC) vectors. The pixel's enhanced spectrum is transformed to the spectral configuration associated to its reaction to a specific stain by utilizing an N × N transformation matrix, which is derived through application of least mean squares method to the enhanced and target spectral transmittance samples of the different tissue components found in the image. Results of our experiments on the digital conversion of an H&E stained multispectral image to its Masson's trichrome stained equivalent show the viability of the method. PMID:22612136

  4. Scene segmentation in a machine vision system for histopathology

    NASA Astrophysics Data System (ADS)

    Thompson, Deborah B.; Bartels, H. G.; Haddad, J. W.; Bartels, Peter H.

    1990-07-01

    Algorithms and procedures employed to attain reliable and exhaustive segmentation in histopathologic imagery of colon and prostate sections are detailed. The algorithms are controlled and selectively called by a scene segmentation expert system as part of a machine vision system for the diagnostic interpretation of histopathologic sections. At this time, effective segmentation of scenes of glandular tissues is produced, with the system being conservative in the identification of glands; for the segmentation of overlapping glandular nuclei an overall success rate of approximately 90% has been achieved.

  5. Micro-scaled high-throughput digestion of plant tissue samples for multi-elemental analysis

    PubMed Central

    Hansen, Thomas H; Laursen, Kristian H; Persson, Daniel P; Pedas, Pai; Husted, Søren; Schjoerring, Jan K

    2009-01-01

    Background Quantitative multi-elemental analysis by inductively coupled plasma (ICP) spectrometry depends on a complete digestion of solid samples. However, fast and thorough sample digestion is a challenging analytical task which constitutes a bottleneck in modern multi-elemental analysis. Additional obstacles may be that sample quantities are limited and elemental concentrations low. In such cases, digestion in small volumes with minimum dilution and contamination is required in order to obtain high accuracy data. Results We have developed a micro-scaled microwave digestion procedure and optimized it for accurate elemental profiling of plant materials (1-20 mg dry weight). A commercially available 64-position rotor with 5 ml disposable glass vials, originally designed for microwave-based parallel organic synthesis, was used as a platform for the digestion. The novel micro-scaled method was successfully validated by the use of various certified reference materials (CRM) with matrices rich in starch, lipid or protein. When the micro-scaled digestion procedure was applied on single rice grains or small batches of Arabidopsis seeds (1 mg, corresponding to approximately 50 seeds), the obtained elemental profiles closely matched those obtained by conventional analysis using digestion in large volume vessels. Accumulated elemental contents derived from separate analyses of rice grain fractions (aleurone, embryo and endosperm) closely matched the total content obtained by analysis of the whole rice grain. Conclusion A high-throughput micro-scaled method has been developed which enables digestion of small quantities of plant samples for subsequent elemental profiling by ICP-spectrometry. The method constitutes a valuable tool for screening of mutants and transformants. In addition, the method facilitates studies of the distribution of essential trace elements between and within plant organs which is relevant for, e.g., breeding programmes aiming at improvement of the

  6. Use of Curcuma longa L. extract to stain various tissue samples for histological studies

    PubMed Central

    Kumar, Sachin; Singh, Narendra Nath; Singh, Arun; Singh, Neelakshi; Sinha, Raman Kant

    2014-01-01

    Background: Curcuma longa L. is a perennial herb and a member of the Zingiberaceae (ginger) family, which is used extensively in foods as well as in Ayurvedic and Chinese systems of medicine. Current researches have focused on its antioxidant, hepatoprotective, anti-inflammatory, anticarcinogenic and antimicrobial properties. Until now, very few studies suggested its role as a histological stain. Aim: To ascertain its efficacy to be used as a counterstain after hematoxylin, to compare it's staining ability with that of routinely used eosin dye and also to ascertain its role in various collagen diseases. Materials and Methods: Turmeric rhizomes were cut into small pieces and were dried. These dried turmeric rhizomes were milled to form fine powder, which was then processed to form dye for staining tissue structures. Results: It revealed that turmeric can be used as a counterstain after hematoxylin, its staining ability was also good and comparable to that of eosin dye with a special affinity for collagen and muscle fibers. Conclusion: Turmeric dye can be used as a histological stain, which stains similar to eosin dye and its specific affinity for collagen and muscle fibers authenticates its role in the treatment of collagen and muscle disorders. PMID:26195911

  7. Gross and histopathological observations of long-term catheterized vessels in experimental sheep.

    PubMed

    Barraza, M L; Strickland, J R; Zepeda, H; Taylor, J B; Krehbiel, C R; Bell, G S; Klotz, J L

    2006-06-01

    Chronic indwelling central vessel catheters provide vascular access for compartmental infusion or sampling. However, complications with catheter patency during the postoperative and/or experimental period often arise. In order to identify physiological occurrences common with such complications, 10 multicatheterized sheep (61.8 +/- 7.8 kg BW), obtained from a previous nutrient flux study were used for gross and histopathological investigation. Catheters had been surgically placed in a hepatic portal vein (PVC), a hepatic vein (HVC), a distal mesenteric vein (MVC) and a mesenteric artery (MAC). In the previous study, catheters (PVC, HVC and MAC) were used to collect blood samples or infuse (MVC) p-aminohippurate. Catheters were maintained for a total of 58 days prior to necropsy. Histopathological findings indicated that catheter failures were associated with the following tissue responses: (i) thromboses with frequent focal vasculitis; (ii) euplastic tissues associated with extensive fibrosis; (iii) granulomas; (iv) neo-vascularization of the media; (v) calcification processes; and (vi) micro-abscesses. Additional studies are needed that address and incorporate improvement of catheter design and placement to minimize irritation of endothelium, improvement of catheter treatments and therapeutic regimes, and development and use of alternative anti-coagulants. A greater understanding of the mechanisms leading to failure will help researchers improve catheter performance and patency.

  8. Metals bioaccumulation and histopathological biomarkers in Carcinus maenas crab from Bizerta lagoon, Tunisia.

    PubMed

    Ben-Khedher, Sana; Jebali, Jamel; Houas, Zohra; Nawéli, Hmida; Jrad, Amel; Banni, Mohamed; Boussetta, Hamadi

    2014-03-01

    Metals concentrations and histolopathological lesions of gills and digestive gland were investigated in Carcinus maenas crabs sampled from Bizerta Lagoon and Kuriat Island (Tunisia) as control site. The concentrations of trace metals varied between tissues, sites and sampling time. The highest levels of the analysed metals in gills and digestive gland were noted in Menzel Bourguiba and Cimentery sites at both sampling times (February and July). The higher metals loads were associated with severe and various tissues alterations in contaminated crabs. We particularly noted in the gills a haemocytic infiltration, distension and enlargement of the lamellae, lifting of lamellar epithelium, necrotic lesions and fusion of lamellae in the most polluted sites (Menzel Bourguiba and Cimentery). Moreover, others pathological alterations were observed in digestive gland of crabs collected from polluted sites and with a severity site dependent. We observed necrotic tubules containing tissue debris in the lumen with more intensity in crabs collected from Cimentery site in both sampling times. The thickened basal laminae and the walling off of the tubules by haemocytes around the thickened basal laminae were more abundant at Menzel Bourguiba than at others sites. The coagulation in the thickened basal laminae was observed only at Cimentery in February. Tissues histopathological lesions were sensitive to discriminate crabs of different sites and demonstrated its usefulness in this biomonitoring study. We recommend the association of histopatholocial lesions to biochemical biomarkers in future biomonitoring studies. PMID:24323326

  9. Non-lethal sampling of liver tissue for toxicologic evaluation of Florida cottonmouths snakes, Agkistrodon piscivorus conanti.

    PubMed

    Quesada, Rolando J; McCleary, Ryan J R; Heard, Darryl J; Lillywhite, Harvey B

    2014-01-01

    Due to their longevity, strong site tenure, poikilothermic metabolism, and low-energy specializations, reptiles might serve as excellent environmental sentinels. Cottonmouth snakes are generalist predators and scavengers, and as such, may have higher exposure to persistent environmental contaminants as a result of bioaccumulation. Traditionally, assessment and monitoring of contaminant exposure in reptiles have involved lethal sampling techniques. In this paper, we describe a non-destructive technique for sampling liver tissue in live anesthetized Florida cottonmouths. Wild-caught snakes (n = 21) were anesthetized with propofol, and a liver wedge biopsy was obtained by clamping the edge of the organ with two small hemostatic mosquito forceps via right-sided coeliotomy incision. A minimum required tissue sample weighing >100 mg was harvested from all except one of the animals. No mortalities occurred during the procedures or recovery from anesthesia, and all snakes were released back into the field after the animal had consumed prey and defecated, usually within 2 weeks following surgery. Hemorrhage was a minor complication in most snakes, especially those with friable discolored livers. The procedure appeared to have no short-term deleterious effects, and two biopsied individuals were captured after being released into the field and appeared to be normal and healthy. However, follow-up studies and recapture of more snakes are needed to assess long-term survivability. Our non-destructive liver sampling technique might be implemented in toxicological studies of other squamates and could help to minimize the lethal sampling of threatened species. PMID:24197420

  10. Museums and disease: using tissue archive and museum samples to study pathogens.

    PubMed

    Tsangaras, Kyriakos; Greenwood, Alex D

    2012-01-20

    Molecular studies of archival and fossil samples have traditionally focused on the nucleic acids derived from the host species. However, there has recently been an increase in ancient DNA research on the identification and characterization of infectious agents within the hosts. The study of pathogens from the past provides great opportunities for discovering the causes of historical infection events, characterizing host-microorganism co-evolution and directly investigating the evolution of specific pathogens. Several research teams have been able to isolate and characterize a variety of different bacterial, parasite and viral microorganisms. However, this emerging field is not without obstacles. The diagenetic processes that make ancient DNA research generally difficult are also impediments to ancient pathogen research and perhaps more so given that their DNA may represent an even rarer proportion of the remaining nucleic acids in a fossil sample than host DNA. However, studies performed under controlled conditions and following stringent ancient DNA protocols can and have yielded reliable and often surprising results. This article reviews the advantages, problems, and failures of ancient microbiological research.

  11. Genome-wide detection of DNase I hypersensitive sites in single cells and FFPE tissue samples.

    PubMed

    Jin, Wenfei; Tang, Qingsong; Wan, Mimi; Cui, Kairong; Zhang, Yi; Ren, Gang; Ni, Bing; Sklar, Jeffrey; Przytycka, Teresa M; Childs, Richard; Levens, David; Zhao, Keji

    2015-12-01

    DNase I hypersensitive sites (DHSs) provide important information on the presence of transcriptional regulatory elements and the state of chromatin in mammalian cells. Conventional DNase sequencing (DNase-seq) for genome-wide DHSs profiling is limited by the requirement of millions of cells. Here we report an ultrasensitive strategy, called single-cell DNase sequencing (scDNase-seq) for detection of genome-wide DHSs in single cells. We show that DHS patterns at the single-cell level are highly reproducible among individual cells. Among different single cells, highly expressed gene promoters and enhancers associated with multiple active histone modifications display constitutive DHS whereas chromatin regions with fewer histone modifications exhibit high variation of DHS. Furthermore, the single-cell DHSs predict enhancers that regulate cell-specific gene expression programs and the cell-to-cell variations of DHS are predictive of gene expression. Finally, we apply scDNase-seq to pools of tumour cells and pools of normal cells, dissected from formalin-fixed paraffin-embedded tissue slides from patients with thyroid cancer, and detect thousands of tumour-specific DHSs. Many of these DHSs are associated with promoters and enhancers critically involved in cancer development. Analysis of the DHS sequences uncovers one mutation (chr18: 52417839G>C) in the tumour cells of a patient with follicular thyroid carcinoma, which affects the binding of the tumour suppressor protein p53 and correlates with decreased expression of its target gene TXNL1. In conclusion, scDNase-seq can reliably detect DHSs in single cells, greatly extending the range of applications of DHS analysis both for basic and for translational research, and may provide critical information for personalized medicine. PMID:26605532

  12. Feasibility of digitally stained multimodal confocal mosaics to simulate histopathology

    NASA Astrophysics Data System (ADS)

    Gareau, Daniel S.

    2009-05-01

    Fluorescence confocal mosaicing microscopy of tissue biopsies stained with acridine orange has been shown to accurately identify tumors and with an overall sensitivity of 96.6% and specificity of 89.2%. However, fluorescence shows only nuclear detail similar to hematoxylin in histopathology and does not show collagen or cytoplasm, which may provide necessary negative contrast information similar to eosin used in histopathology. Reflectance mode contrast is sensitive to collagen and cytoplasm without staining. To further improve sensitivity and specificity, digitally stained confocal mosaics combine confocal fluorescence and reflectance images in a multimodal pseudo-color image to mimic the appearance of histopathology with hematoxylin and eosin and facilitate the introduction of confocal microscopy into the clinical realm.

  13. Graph run-length matrices for histopathological image segmentation.

    PubMed

    Tosun, Akif Burak; Gunduz-Demir, Cigdem

    2011-03-01

    The histopathological examination of tissue specimens is essential for cancer diagnosis and grading. However, this examination is subject to a considerable amount of observer variability as it mainly relies on visual interpretation of pathologists. To alleviate this problem, it is very important to develop computational quantitative tools, for which image segmentation constitutes the core step. In this paper, we introduce an effective and robust algorithm for the segmentation of histopathological tissue images. This algorithm incorporates the background knowledge of the tissue organization into segmentation. For this purpose, it quantifies spatial relations of cytological tissue components by constructing a graph and uses this graph to define new texture features for image segmentation. This new texture definition makes use of the idea of gray-level run-length matrices. However, it considers the runs of cytological components on a graph to form a matrix, instead of considering the runs of pixel intensities. Working with colon tissue images, our experiments demonstrate that the texture features extracted from "graph run-length matrices" lead to high segmentation accuracies, also providing a reasonable number of segmented regions. Compared with four other segmentation algorithms, the results show that the proposed algorithm is more effective in histopathological image segmentation.

  14. In situ assessment of mRNA accessibility in heterogeneous tissue samples using elongation factor-1 alpha (EF-1 alpha).

    PubMed

    Gruber, A D; Levine, R A

    1997-05-01

    Elongation factor-1 alpha (EF-1 alpha) is an evolutionarily highly conserved universal cofactor of protein synthesis in all living cells. In this study, its use as a positive control in situ hybridization assays for specific detection of mRNA sequences was evaluated. Northern blot analysis of various non-neoplastic and neoplastic cultured cells of different stages of confluence, cell shape, and cell cycle status revealed that EF-1 alpha had a lower and more homogeneous expression than did beta-actin. In situ hybridization assays using digoxigenin-labeled riboprobes for the detection of EF-1 alpha mRNA in routinely formalin-fixed, paraffin-embedded tissue sections showed that EF-1 alpha is a suitable positive control in all types of cells. However, variation of protease pretreatments demonstrated distinct and sometimes mutually exclusive digestion conditions for different cell types within the same tissue sample. Our results indicate that detection of EF-1 alpha mRNA is an appropriate internal standard for in situ hybridization assays and that it is useful to control artifacts such as false negatives caused by inappropriate protease pretreatments. The observed variability of optimal protease pretreatments for different cell types within the same tissue section strengthens the importance of a positive control in in situ hybridization assays.

  15. Development of a remote laser-induced breakdown spectroscopy system for investigation of calcified tissue samples

    SciTech Connect

    Hrdlicka, Ales; Prokes, Lubomir; Stankova, Alice; Novotny, Karel; Vitesnikova, Anna; Kanicky, Viktor; Otruba, Vitezslav; Kaiser, Jozef; Novotny, Jan; Malina, Radomir; Palenikova, Katerina

    2010-05-01

    The development of a remote laser-induced breakdown spectroscopy (LIBS) setup with an off-axis Newtonian collection optics, Galilean-based focusing telescope, and a 532 nm flattop laser beam source is presented. The device was tested at a 6 m distance on a slice of bone to simulate its possible use in the field, e.g., during archaeological excavations. It is shown that this setup is sufficiently sensitive to both major (P, Mg) and minor elements (Na, Zn, Sr). The measured quantities of Mg, Zn, and Sr correspond to the values obtained by reference laser ablation-inductively coupled plasma-mass spectrometry (LA-ICP-MS) measurements within an approximately 20% range of uncertainty. A single point calibration was performed by use of a bone meal standard . The radial element distribution is almost invariable by use of LA-ICP-MS, whereas the LIBS measurement showed a strong dependence on the sample porosity. Based on these results, this remote LIBS setup with a relatively large (350 mm) collecting mirror is capable of semiquantitative analysis at the level of units of mg kg{sup -1}.

  16. The problem with medical research on tissue and organ samples taken in connection with forensic autopsies in France.

    PubMed

    Rougé-Maillart, C; Dupont, V; Jousset, N

    2016-02-01

    Currently, in France, it is legally impossible to conduct scientific research on tissue and organ samples taken from forensic autopsies. In fact, the law schedules the destruction of such samples at the end of the judicial investigation, and the common law rules governing cadaver research cannot be applied to the forensic context. However, nothing seems in itself to stand in the way of such research since, despite their specific nature, these samples from forensic autopsies could be subject, following legislative amendments, to common law relating to medical research on samples taken from deceased persons. But an essential legislative amendment, firstly to allow the Biomedicine Agency to become authorized to issue a research permit and secondly, to change the research conditions in terms of the non-opposition of the deceased to said research. Such an amendment would be a true breakthrough because it would allow teams to continue to move forward calmly in research, and allow this research to be placed within a legal framework, which would promote international exchanges.

  17. The problem with medical research on tissue and organ samples taken in connection with forensic autopsies in France.

    PubMed

    Rougé-Maillart, C; Dupont, V; Jousset, N

    2016-02-01

    Currently, in France, it is legally impossible to conduct scientific research on tissue and organ samples taken from forensic autopsies. In fact, the law schedules the destruction of such samples at the end of the judicial investigation, and the common law rules governing cadaver research cannot be applied to the forensic context. However, nothing seems in itself to stand in the way of such research since, despite their specific nature, these samples from forensic autopsies could be subject, following legislative amendments, to common law relating to medical research on samples taken from deceased persons. But an essential legislative amendment, firstly to allow the Biomedicine Agency to become authorized to issue a research permit and secondly, to change the research conditions in terms of the non-opposition of the deceased to said research. Such an amendment would be a true breakthrough because it would allow teams to continue to move forward calmly in research, and allow this research to be placed within a legal framework, which would promote international exchanges. PMID:26694871

  18. Monitoring impacts of air pollution: PIXE analysis and histopathological modalities in evaluating relative risks of elemental contamination.

    PubMed

    Ejaz, Sohail; Camer, Gerry Amor; Anwar, Khaleeq; Ashraf, Muhammad

    2014-04-01

    Environmental toxicants invariably affect all biological organisms resulting to sufferings ranging from subclinical to debilitating clinical conditions. This novel research aimed to determine the toxic burdens of increased environmental elements in some vital organs/tissues of the wild animals (starling, owl, crow and pigeon), exposed to air polluted environment were assessed using particle induced X-ray emission and histopathological approaches. The presence of significantly elevated amounts of elemental toxicants namely: Aluminum (Al), Chlorine (Cl), Iron (Fe), Potassium (K), Magnesium (Mg), Manganese (Mn), Silicon (Si) and Vanadium (V) from the skin, muscle, lungs, liver and kidney of sampled animals were in concurrence with the observed histopathological changes. The skin of sampled starling, owl, pigeon and crow spotlighted highly significant increase (P < 0.001) in Al, Cl, Mg and Si. Muscle samples with myodegenerative lesions and mineral depositions highlighted substantial augmentation (P < 0.001) in the amount of Al, Fe, Mn, Si and V. The lungs of starling, owl, and pigeon were severely intoxicated (P < 0.001) with increased amount of Al, Fe, K, Mn and Si producing pulmonary lesions of congestion, edema, pneumonitis and mineral debris depositions. Liver samples revealed that the sampled animals were laden with Cl, Fe, Mg, Mn and V with histopathological profound degenerative changes and hepatic necrosis. Kidney sections presented severe tubular degenerative and necrotic changes that may be attributed to increased amounts of Cl and Fe. These current findings implied that the environmental/elemental toxicants and the accompanying lesions that were discerned in the organs/tissues of sampled birds may as well be afflicting people living within the polluted area. Further assessment to more conclusively demonstrate correlations of current findings to those of the populace within the area is encouraged.

  19. Evaluation of two-dimensional electrophoresis and liquid chromatography – tandem mass spectrometry for tissue-specific protein profiling of laser-microdissected plant samples

    SciTech Connect

    Schad, Martina; Lipton, Mary S.; Giavalisco, Patrick; Smith, Richard D.; Kehr, Julia

    2005-07-14

    Laser microdissection (LM) allows the collection of homogeneous tissue- and cell specific plant samples. The employment of this technique with subsequent protein analysis has thus far not been reported for plant tissues, probably due to the difficulties associated with defining a reasonable cellular morphology and, in parallel, allowing efficient protein extraction from tissue samples. The relatively large sample amount needed for successful proteome analysis is an additional issue that complicates protein profiling on a tissue- or even cell-specific level. In contrast to transcript profiling that can be performed from very small sample amounts due to efficient amplification strategies, there is as yet no amplification procedure for proteins available. In the current study, we compared different tissue preparation techniques prior to LM/laser pressure catapulting (LMPC) with respect to their suitability for protein retrieval. Cryosectioning was identified as the best compromise between tissue morphology and effective protein extraction. After collection of vascular bundles from Arabidopsis thaliana stem tissue by LMPC, proteins were extracted and subjected to protein analysis, either by classical two-dimensional gel electrophoresis (2-DE), or by high-efficiency liquid chromatography (LC) in conjunction with tandem mass spectrometry (MS/MS). Our results demonstrate that both methods can be used with LMPC collected plant material. But because of the significantly lower sample amount required for LC-MS/MS than for 2-DE, the combination of LMPC and LC-MS/MS has a higher potential to promote comprehensive proteome analysis of specific plant tissues.

  20. Brief Report: S6 Ribosomal Protein Phosphorylation in Autistic Frontal Cortex and Cerebellum: A Tissue Array Analysis

    ERIC Educational Resources Information Center

    Eberhart, Charles G.; Copeland, Joshua; Abel, Ty W.

    2006-01-01

    Few autistic brain samples are available for study, limiting investigations into molecular and histopathological abnormalities associated with this common disease. To facilitate distribution of samples, we have constructed a tissue array containing cerebral and cerebellar cores from 5 autistic children, 1 girl with Rett syndrome, and 5 age-matched…

  1. 42 CFR 493.1219 - Condition: Histopathology.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ... 42 Public Health 5 2010-10-01 2010-10-01 false Condition: Histopathology. 493.1219 Section 493....1219 Condition: Histopathology. If the laboratory provides services in the subspecialty of Histopathology, the laboratory must meet the requirements specified in §§ 493.1230 through 493.1256, §...

  2. Mediastinitis after cardiac surgery: improvement of bacteriological diagnosis by use of multiple tissue samples and strain typing.

    PubMed

    Tammelin, Ann; Hambraeus, Anna; Ståhle, Elisabeth

    2002-08-01

    The diagnosis of postsurgical mediastinitis (PSM) among patients with sternal wound complication (SWC) after cardiac surgery is sometimes difficult, as fever, elevated C-reactive protein levels, and chest pain can be caused by a general inflammatory reaction to the operative trauma and/or sternal dehiscence without infection. The definitions of PSM usually used emphasize clinical signs and symptoms easily observed by the surgeon. The aim of the study was to investigate whether the use of standardized multiple tissue sampling, optimal culturing methods, and strain typing, together with a microbiological criterion for infection, could identify more infected patients than clinical assessment alone. Patients reexplored due to SWC after cardiac artery bypass grafting (CABG) or heart valve replacement (HVR) with or without CABG performed at the Department for Cardio-Thoracic Surgery at the Uppsala University Hospital between 10 March 1998 and 9 September 2000 were investigated prospectively. Tissue samples were taken from the sternum or adjacent mediastinal tissue, preferably before the administration of antibiotics. Culturing was performed both directly (on agar plates) and using enrichment broth. Species identification was performed by standard methods, and strain typing was performed by pulsed-field gel electrophoresis. A total of 41 cases with at least five tissue samples each were included in the study group. Of these patients, 32 were infected according to the microbiological criterion (i.e., the same strain was found in >/=50% of the samples). Staphylococcus epidermidis was the primary pathogen in 38% of the cases (12/32), S. aureus was the primary pathogen in 31% (10/32), P. acnes was the primary pathogen in 25% (8/32), and S. simulans and S. haemolyticus were the primary pathogens in 3% (1/32) each. All cases of S. aureus infection and 86% (12/14) of coagulase-negative staphylococcus (CoNS) infections were identified from primary cultures. All cases fulfilling

  3. Porcine proliferative enteropathy in Estonian pig herds: histopathology and detection of Lawsonia intracellularis by PCR.

    PubMed

    Järveots, Tönu; Saar, Tiiu; Lepp, Elbi; Suuroja, Toivo; Lindjärv, Raivo; Nathues, Heiko; Sütt, Silva; Põdersoo, Diivi

    2011-01-01

    Lawsonia intracellularis is the causative agent of proliferative enteritis in pigs (PPE). This bacterium is difficult to culture from clinical samples and antemortem demonstration is therefore usually performed by PCR on faecal samples. The aim of this study was to elucidate the frequency of L. intracellularis infection in pig herds in Estonia using PCR, histopathological methods and electronmicroscopical studies. The frequency of demonstration of L. intracellularis was highest in 9-12 weeks old pigs (68.1%). It was more frequent in growing pigs with enteritis on small farms where the system of "all-in all-out" was not practiced and where standards of hygiene were poor. Gross and histopathological studies demonstrated that characteristic macroscopic changes associated with PPE were localised to the distal jejunum and ileum.Thickened longitudinal and circumferential folds occurred in the mucosa of the affected regions of the bowel. Samples from pigs aged 4 to 20 weeks exhibited the most intensive inflammatory changes. The distal part of the jejunum, ileum and the upper third of proximal colon and cecum wall were visibly thickened with reduced luminal diameter. Hyperplasia of lymphoid tissue and, in many cases, pseudomembranous or fibrinous inflammation was found. L. intracellularis was detected in 56 young pigs using histopathological methods. Additionally, in 8 of these pigs intracellular bacteria were demonstrated in ilial epithelial cells by transmission electronmicroscopical (TEM) investigation. On the basis of these TEM investigations it was concluded that L. intracellularis causes disturbances of normal growth, differentiation and apoptosis of the epithelial cells of ileum. PMID:21306056

  4. A New Sample Substrate for Imaging and Correlating Organic and Trace Metal Composition in Biological Cells and Tissues

    SciTech Connect

    Miller,L.; Wang, Q.; Smith, R.; Zhong, H.; Elliott, D.; Warren, J.

    2007-01-01

    Many disease processes involve alterations in the chemical makeup of tissue. Synchrotron-based infrared (IR) and X-ray fluorescence (XRF) microscopes are becoming increasingly popular tools for imaging the organic and trace metal compositions of biological materials, respectively, without the need for extrinsic labels or stains. Fourier transform infrared microspectroscopy (FTIRM) provides chemical information on the organic components of a material at a diffraction-limited spatial resolution of 2-10 {mu}m in the mid-infrared region. The synchrotron X-ray fluorescence (SXRF) microprobe is a complementary technique used to probe trace element content in the same systems with a similar spatial resolution. However to be most beneficial, it is important to combine the results from both imaging techniques on a single sample, which requires precise overlap of the IR and X-ray images. In this work, we have developed a sample substrate containing a gold grid pattern on its surface, which can be imaged with both the IR and X-ray microscopes. The substrate consists of a low trace element glass slide that has a gold grid patterned on its surface, where the major and minor parts of the grid contain 25 and 12 nm gold, respectively. This grid pattern can be imaged with the IR microscope because the reflectivity of gold differs as a function of thickness. The pattern can also be imaged with the SXRF microprobe because the Au fluorescence intensity changes with gold thickness. The tissue sample is placed on top of the patterned substrate. The grid pattern's IR reflectivity image and the gold SXRF image are used as fiducial markers for spatially overlapping the IR and SXRF images from the tissue. Results show that IR and X-ray images can be correlated precisely, with a spatial resolution of less than one pixel (i.e., 2-3 microns). The development of this new tool will be presented along with applications to paraffin-embedded metalloprotein crystals, Alzheimer's disease, and hair

  5. Paralytic shellfish poisoning: post-mortem analysis of tissue and body fluid samples from human victims in the Patagonia fjords.

    PubMed

    García, Carlos; del Carmen Bravo, María; Lagos, Marcelo; Lagos, Néstor

    2004-02-01

    In July 5, 2002 fishermen working in harvesting sea urchin (Loxechinus albus) in the Patagonia Chilean fjords were intoxicated by consumption of filter-feeder bivalve Aulacomya ater. After the ingestion of 7-9 ribbed mussel, two fishermen died 3-4 h after shellfish consumption. The forensic examination in both victims did not show pathological abnormalities with the exception of the lungs conditions, crackling to the touch, pulmonary congestion and edema. The toxic mussel sample showed a toxicity measured by mouse bioassay of 8575 microg of STX (saxitoxin) equivalent by 100 g of shellfish meat. Using post-column derivatization HPLC method with fluorescent on line detection was possible to measure mass amount of each paralytic shellfish poisoning (PSP) toxin yielding individual toxin concentrations. These PSP toxins were identified in the gastric content, body fluids (urine, bile and cerebrospinal fluid) and tissue samples (liver, kidney, lung, stomach, spleen, heart, brain, adrenal glands, pancreas and thyroids glands). The toxin profiles of each body fluid and tissue samples and the amount of each PSP toxin detected are reported. The PSP toxins found in the gastric content, were STX and the gonyautoxins (GTX4, GTX1, GTX5, GTX3 and GTX2) which showed to be the major amount of PSP toxins found in the victims biological samples. The PSP toxin composition in urine and bile showed as major PSP toxins neoSaxitoxin (neoSTX) and GTX4/GTX1 epimers, both STX analogues with an hydroxyl group (-OH) in the N(1) of the tetrahydropurine nucleus. The neoSTX was not present in the gastric content sample, indicating that the oxidation of N(1) in the STX tetrahydropurine nucleus resulted neoSTX, in a similar way that GTX3/GTX2 epimers were transformed in GTX4/GTX1 epimers. Beside this metabolic transformation, also the hydrolysis of carbamoyl group from STX to form its decarbomoyl analogue decarbamoylsaxitoxin was detected in liver, kidney and lung. These two findings show that PSP

  6. Radiation-Induced Liver Damage: Correlation of Histopathology with Hepatobiliary Magnetic Resonance Imaging, a Feasibility Study

    SciTech Connect

    Seidensticker, Max; Burak, Miroslaw; Kalinski, Thomas; Garlipp, Benjamin; Koelble, Konrad; Wust, Peter; Antweiler, Kai; Seidensticker, Ricarda; Mohnike, Konrad; Pech, Maciej; Ricke, Jens

    2015-02-15

    PurposeRadiotherapy of liver malignancies shows promising results (radioembolization, stereotactic irradiation, interstitial brachytherapy). Regardless of the route of application, a certain amount of nontumorous liver parenchyma will be collaterally damaged by radiation. The functional reserve may be significantly reduced with an impact on further treatment planning. Monitoring of radiation-induced liver damage by imaging is neither established nor validated. We performed an analysis to correlate the histopathological presence of radiation-induced liver damage with functional magnetic resonance imaging (MRI) utilizing hepatobiliary contrast media (Gd-BOPTA).MethodsPatients undergoing local high-dose-rate brachytherapy for whom a follow-up hepatobiliary MRI within 120 days after radiotherapy as well as an evaluable liver biopsy from radiation-exposed liver tissue within 7 days before MRI were retrospectively identified. Planning computed tomography (CT)/dosimetry was merged to the CT-documentation of the liver biopsy and to the MRI. Presence/absence of radiation-induced liver damage (histopathology) and Gd-BOPTA uptake (MRI) as well as the dose applied during brachytherapy at the site of tissue sampling was determined.ResultsFourteen biopsies from eight patients were evaluated. In all cases with histopathological evidence of radiation-induced liver damage (n = 11), no uptake of Gd-BOPTA was seen. In the remaining three, cases no radiation-induced liver damage but Gd-BOPTA uptake was seen. Presence of radiation-induced liver damage and absence of Gd-BOPTA uptake was correlated with a former high-dose exposition.ConclusionsAbsence of hepatobiliary MRI contrast media uptake in radiation-exposed liver parenchyma may indicate radiation-induced liver damage. Confirmatory studies are warranted.

  7. Routine sample preparation and HPLC analysis for ascorbic acid (vitamin C) determination in wheat plants and Arabidopsis leaf tissues.

    PubMed

    Szalai, Gabriella; Janda, T; Pál, Magda

    2014-06-01

    Plants have developed various mechanisms to protect themselves against oxidative stress. One of the most important non-enzymatic antioxidants is ascorbic acid. There is thus a need for a rapid, sensitive method for the analysis of the reduced and oxidised forms of ascorbic acid in crop plants. In this paper a simple, economic, selective, precise and stable HPLC method is presented for the detection of ascorbate in plant tissue. The sensitivity, the short retention time and the simple isocratic elution mean that the method is suitable for the routine quantification of ascorbate in a high daily sample number. The method has been found to be better than previously reported methods, because of the use of an economical, readily available mobile phase, UV detection and the lack of complicated extraction procedures. The method has been tested on Arabidopsis plants with different ascorbate levels and on wheat plants during Cd stress.

  8. Gas-liquid chromatographic and gas-liquid-mass spectometric determination of fenvalerate and permethrin residues in grasshoppers and duck tissue samples

    USGS Publications Warehouse

    Reichel, W.L.; Kolbe, E.J.; Stafford, C.J.

    1981-01-01

    A procedure is described for determining fenvalerate and permethrin residues in grasshoppers and duck tissues. Samples are Soxhlet-extracted with hexane and cleaned up by gel permeation chromatography with an in-line alumina column. Samples are analyzed by gas-liquid chromatography with electron capture detection, and confirmed by gas-liquid chromatography-mass spectrometry. The average recovery from fortified tissues was 97%.

  9. Basic Histopathological Methods and Breast Lesion Types for Research.

    PubMed

    Ouyang, Nengtai; Wang, Lin

    2016-01-01

    The in situ observation on the tissues, such as histopathology, immunohistochemistry (IHC), immunofluorescence (IF), and in situ hybridization (ISH), is one of the most important methods in the biomedical scientific research. In this chapter we introduce the most often used methods-hematoxylin and eosin (H&E) and double IF staining. H&E staining is used for general morphology by which the different pathological types of breast lesions are identified. The double IF staining is often used to study the protein-protein interaction on tissues for signaling mechanisms. This chapter also includes the histopathology of primary or simplified breast lesion types that is essential for applying the above methods and the reclassification of breast cancers by molecular markers.

  10. Digital staining for histopathology multispectral images by the combined application of spectral enhancement and spectral transformation.

    PubMed

    Bautista, Pinky A; Yagi, Yukako

    2011-01-01

    In this paper we introduced a digital staining method for histopathology images captured with an n-band multispectral camera. The method consisted of two major processes: enhancement of the original spectral transmittance and the transformation of the enhanced transmittance to its target spectral configuration. Enhancement is accomplished by shifting the original transmittance with the scaled difference between the original transmittance and the transmittance estimated with m dominant principal component (PC) vectors;the m-PC vectors were determined from the transmittance samples of the background image. Transformation of the enhanced transmittance to the target spectral configuration was done using an nxn transformation matrix, which was derived by applying a least square method to the enhanced and target spectral training data samples of the different tissue components. Experimental results on the digital conversion of a hematoxylin and eosin (H&E) stained multispectral image to its Masson's trichrome stained (MT) equivalent shows the viability of the method.

  11. Sensitivity and specificity of serological tests, histopathology and immunohistochemistry for detection of Toxoplasma gondii infection in domestic chickens.

    PubMed

    Casartelli-Alves, L; Boechat, V C; Macedo-Couto, R; Ferreira, L C; Nicolau, J L; Neves, L B; Millar, P R; Vicente, R T; Oliveira, R V C; Muniz, A G; Bonna, I C F; Amendoeira, M R R; Silva, R C; Langoni, H; Schubach, T M P; Menezes, R C

    2014-08-29

    Since free-range chickens are important for the epidemiology of toxoplasmosis, this study evaluated the sensitivity and specificity of different laboratory techniques for the diagnosis of Toxoplasma gondii in these animals. Serum samples from 135 adult domestic chickens were tested for anti-T. gondii antibodies by the indirect fluorescent antibody test (IFAT), enzyme-linked immunosorbent assay (ELISA), modified agglutination test (MAT), and indirect hemagglutination test (IHAT). Tissue samples from all animals were analyzed by histopathology, immunohistochemistry and mouse bioassay (gold standard). Fifty-four chickens were positive for T. gondii in the bioassay. The sensitivity and specificity of the different tests were, respectively, 85% and 56% for ELISA; 80% and 52% for IFAT; 76% and 68% for MAT; 61% and 80% for IHAT; 7% and 98% for immunohistochemistry, and 6% and 98% for histopathology. The MAT was the most effective method for the diagnosis of T. gondii infection in chickens, followed by ELISA. Histopathology and immunohistochemistry are useful tools for the diagnosis of T. gondii infection in chickens due to their specificity.

  12. Histopathological and biochemical alterations of the earthworm (Lumbricus Terrestris) as biomarker of soil pollution along Porsuk River Basin (Turkey).

    PubMed

    Kılıç, Gözde Aydoğan

    2011-05-01

    This study investigated biomarker responses of the earthworm Lumbricus terrestris in order to evaluate the soil pollution along Porsuk River Basin. Samples consisted of animals from six sites that are agricultural regions and a forested control. Histopathological and biochemical alterations were examined. Significant histopathological alterations were observed in animals from three of the sampling sites. There was an enlargement of epithelial cell lining, mucus cell hyperplasia and increase in mucus secretion. Circular and longitudinal muscles lost their structural integrity. Chloragogenous tissue was dilated and vocuolized. Necrosis was observed in the cells and tissues of some affected worms. A load of heavy metals in tissues of animals was determined. Heavy metals were found to be accumulated particularly in longitudinal muscles of animals. CAT activity was found to be increased in animals from three of the experimental sites. GST activity was also increased in five sites while it was stable in one site. The results have shown that animals from locations particularly that are close to urbanized and industrialized regions were seriously affected from the soil pollution around the basin. These results are reflecting the biological effects of soil pollution around Porsuk River Basin on the indicator organism L. terrestris and constitute an early warning of ecological change in relation to human health. PMID:21251693

  13. Histopathologic studies of ischemic optic neuropathy.

    PubMed Central

    Knox, D L; Kerrison, J B; Green, W R

    2000-01-01

    PURPOSE: To define the histopathologic features of eyes in which a pathologic diagnosis of ischemic optic neuropathy had been made in the years 1951 through 1998. METHODS: The following data were documented: age of patient, race, sex, source of tissue, cause of death, clinical history, interval from loss of vision to death, enucleation, exenteration, and biopsy. The histopathologic criteria for diagnosis of ischemic optic neuropathy were the presence of localized ischemic edema, cavernous degeneration, or an area of atrophy located superior or inferior in the optic nerve. Cases with history of abrupt loss of vision were combined with reports from the literature to construct a time table of histopathologic features and associated conditions. RESULTS: Ischemic optic neuropathy was present in 193 eyes. There were 88 females and 65 males. The average age was 71.6 years. Ischemic edema without (early) and with (later) gitter macrophages was present in 26 (13.5%). Cavernous degeneration was present in 69 nerves (36%). Mucopolysaccharide (MPS) was present in 37 cavernous lesions 1 month or longer after loss of vision. Cavernous lesions were seen in 3 eyes in which peripapillary retinal nerve fiber layer hemorrhage had been observed prior to death. Atrophic lesions, the most common pattern, were observed in 133 optic nerves (66.8%). More than 1 ischemic lesion was seen in 38 optic nerves (19.7%). Bilateral ischemic lesions were seen in 50 (35.2%) of 142 paired eyes. CONCLUSIONS: Ischemic optic nerve lesions are initially acellular and later show macrophage infiltration. Cavernous lesions with MPS are present 4 weeks or longer after vision loss. The location of MPS posteriorly and along the internal margin suggests that MPS is produced at the edges of lesions. Progressive vision loss in ischemic optic neuropathy may be secondary to compression of intact nerve from ischemic edema and cavernous swelling, or a second ischemic lesion. Images FIGURE 2 FIGURE 3 FIGURE 4 FIGURE 5

  14. Balanced-PCR amplification allows unbiased identification of genomic copy changes in minute cell and tissue samples.

    PubMed

    Wang, Gang; Brennan, Cameron; Rook, Martha; Wolfe, Jia Liu; Leo, Christopher; Chin, Lynda; Pan, Hongjie; Liu, Wei-Hua; Price, Brendan; Makrigiorgos, G Mike

    2004-05-21

    Analysis of genomic DNA derived from cells and fresh or fixed tissues often requires whole genome amplification prior to microarray screening. Technical hurdles to this process are the introduction of amplification bias and/or the inhibitory effects of formalin fixation on DNA amplification. Here we demonstrate a balanced-PCR procedure that allows unbiased amplification of genomic DNA from fresh or modestly degraded paraffin-embedded DNA samples. Following digestion and ligation of a target and a control genome with distinct linkers, the two are mixed and amplified in a single PCR, thereby avoiding biases associated with PCR saturation and impurities. We demonstrate genome-wide retention of allelic differences following balanced-PCR amplification of DNA from breast cancer and normal human cells and genomic profiling by array-CGH (cDNA arrays, 100 kb resolution) and by real-time PCR (single gene resolution). Comparison of balanced-PCR with multiple displacement amplification (MDA) demonstrates equivalent performance between the two when intact genomic DNA is used. When DNA from paraffin-embedded samples is used, balanced PCR overcomes problems associated with modest DNA degradation and produces unbiased amplification whereas MDA does not. Balanced-PCR allows amplification and recovery of modestly degraded genomic DNA for subsequent retrospective analysis of human tumors with known outcomes.

  15. Activity assays of mammalian thioredoxin and thioredoxin reductase: fluorescent disulfide substrates, mechanisms, and use with tissue samples.

    PubMed

    Montano, Sergio J; Lu, Jun; Gustafsson, Tomas N; Holmgren, Arne

    2014-03-15

    Thioredoxin (Trx) is a protein disulfide reductase that, together with nicotinamide adenine dinucleotide phosphate (NADPH) and thioredoxin reductase (TrxR), controls oxidative stress or redox signaling via thiol redox control. Human cytosolic Trx1 has Cys32 and Cys35 as the active site and three additional cysteine residues (Cys62, Cys69, and Cys73), which by oxidation generates inactive Cys62 to Cys69 two-disulfide Trx. This, combined with TrxR with a broad substrate specificity, complicates assays of mammalian Trx and TrxR. We sought to understand the autoregulation of Trx and TrxR and to generate new methods for quantification of Trx and TrxR. We optimized the synthesis of two fluorescent substrates, di-eosin-glutathione disulfide (Di-E-GSSG) and fluorescein isothiocyanate-labeled insulin (FiTC-insulin), which displayed higher fluorescence on disulfide reduction. Di-E-GSSG showed a very large increase in fluorescence quantum yield but had a relatively low affinity for Trx and was also a weak direct substrate for TrxR, in contrast to GSSG. FiTC-insulin was used to develop highly sensitive assays for TrxR and Trx. Reproducible conditions were developed for reactivation of modified Trx, commonly present in frozen or oxidized samples. Trx in cell extracts and tissue samples, including plasma and serum, were subsequently analyzed, showing highly reproducible results and allowing measurement of trace amounts of Trx.

  16. Histopathologic Diagnosis of Fungal Infections in the 21st Century

    PubMed Central

    Guarner, Jeannette; Brandt, Mary E.

    2011-01-01

    Summary: Fungal infections are becoming more frequent because of expansion of at-risk populations and the use of treatment modalities that permit longer survival of these patients. Because histopathologic examination of tissues detects fungal invasion of tissues and vessels as well as the host reaction to the fungus, it is and will remain an important tool to define the diagnostic significance of positive culture isolates or results from PCR testing. However, there are very few instances where the morphological characteristics of fungi are specific. Therefore, histopathologic diagnosis should be primarily descriptive of the fungus and should include the presence or absence of tissue invasion and the host reaction to the infection. The pathology report should also include a comment stating the most frequent fungi associated with that morphology as well as other possible fungi and parasites that should be considered in the differential diagnosis. Alternate techniques have been used to determine the specific agent present in the histopathologic specimen, including immunohistochemistry, in situ hybridization, and PCR. In addition, techniques such as laser microdissection will be useful to detect the now more frequently recognized dual fungal infections and the local environment in which this phenomenon occurs. PMID:21482725

  17. [Update on protein analysis of fixed tissues].

    PubMed

    Becker, K-F; Berg, D; Malinowsky, K; Wolff, C; Ergin, B; Meding, S; Walch, A; Höfler, H

    2010-10-01

    Tissue samples have been routinely used for decades to distinguish healthy from diseased tissue in histopathological characterization. While nucleic acid-based methodologies have been successfully in use for many years, protein-based techniques, in contrast, are at a very early stage (with the exception of immunohistochemistry). One reason for this delay may be that the scientific community has long thought that formalin-fixed and paraffin embedded (FFPE) tissues are unfit for protein analysis. However, recent reports demonstrate that many protein methods that are routinely used for frozen tissues can also be applied for FFPE tissues, including Western blot, protein microarray, matrix-assisted laser desorption/ionization (MALDI) imaging and 2D gel electrophoresis. The present article provides an overview of recent developments in this field, focussing particular attention on quantitative analysis and high throughput technologies that have the potential to be integrated into the routine workflow of clinical pathology laboratories.

  18. Comparison between cytology and histopathology to evaluate subclinical endometritis in dairy cows.

    PubMed

    Bogado Pascottini, Osvaldo; Hostens, Miel; Dini, Pouya; Vandepitte, Jan; Ducatelle, Richard; Opsomer, Geert

    2016-10-01

    The aim of the present study was to compare endometrial cytology with histopathology to diagnose subclinical endometritis (SCE) in dairy cows. Endometrial cytology samples were collected from Holstein-Friesian cows (n = 32) just before slaughtering. Half of them were obtained by in vivo cytobrush (IV-CB), whereas the other half by in vivo low-volume lavage (IV-LVL). After slaughtering, reproductive tracts were collected, and the endometrium was sampled at eight locations. At each location, both a ex vivo cytobrush sample (EV-CB) and a tissue sample for histopathologic examination were taken. In the histopathology slides, polymorphonuclear (PMN) cell counts were differentiated as PMN cells in direct contact with the epithelial cells of the endometrium (PMN-EP), and PMN cells present in the deeper stratum compactum (PMN-SC). Summation of both countings was referred to as PMN-total. Pearson's correlation and Cohen's kappa coefficient were used to assess the correlation and agreement between both sampling methods (in vivo cytology [IV-CB and IV-LVL] with EV-CB and PMN-total). A Poisson mixed effect model was used to analyze the PMN cells' distribution. The prevalence of SCE was 18.75% (n = 6/32) for in vivo cytology. The SCE prevalence based on EV-CB analyses and on the assessment of PMN-total was determined both at the sample (n = 256) as well as at the cow level (n = 32): EV-CB 25% (n = 64/256) and 35.5% (n = 12/32), and PMN-total 37.11% (n = 95/256) and 59.38% (n = 19/32). Correlation and agreement between IV-CB and EV-CB were r = 0.81 and k = 0.97, whereas between IV-CB and PMN-total r = 0.15 and k = 0.23, respectively. In vivo low-volume lavage correlation and agreement were r = 0.52 and k = 0.66 with EV-CB, and r = 0.45 and k = 0.44 with PMN-total. Moreover, correlation and agreement between EV-CB and PMN-total were r = 0.60 and k = 0.50, respectively. More PMN cells (P < 0.05) were detected in PMN-SC when compared to PMN

  19. Comparison between cytology and histopathology to evaluate subclinical endometritis in dairy cows.

    PubMed

    Bogado Pascottini, Osvaldo; Hostens, Miel; Dini, Pouya; Vandepitte, Jan; Ducatelle, Richard; Opsomer, Geert

    2016-10-01

    The aim of the present study was to compare endometrial cytology with histopathology to diagnose subclinical endometritis (SCE) in dairy cows. Endometrial cytology samples were collected from Holstein-Friesian cows (n = 32) just before slaughtering. Half of them were obtained by in vivo cytobrush (IV-CB), whereas the other half by in vivo low-volume lavage (IV-LVL). After slaughtering, reproductive tracts were collected, and the endometrium was sampled at eight locations. At each location, both a ex vivo cytobrush sample (EV-CB) and a tissue sample for histopathologic examination were taken. In the histopathology slides, polymorphonuclear (PMN) cell counts were differentiated as PMN cells in direct contact with the epithelial cells of the endometrium (PMN-EP), and PMN cells present in the deeper stratum compactum (PMN-SC). Summation of both countings was referred to as PMN-total. Pearson's correlation and Cohen's kappa coefficient were used to assess the correlation and agreement between both sampling methods (in vivo cytology [IV-CB and IV-LVL] with EV-CB and PMN-total). A Poisson mixed effect model was used to analyze the PMN cells' distribution. The prevalence of SCE was 18.75% (n = 6/32) for in vivo cytology. The SCE prevalence based on EV-CB analyses and on the assessment of PMN-total was determined both at the sample (n = 256) as well as at the cow level (n = 32): EV-CB 25% (n = 64/256) and 35.5% (n = 12/32), and PMN-total 37.11% (n = 95/256) and 59.38% (n = 19/32). Correlation and agreement between IV-CB and EV-CB were r = 0.81 and k = 0.97, whereas between IV-CB and PMN-total r = 0.15 and k = 0.23, respectively. In vivo low-volume lavage correlation and agreement were r = 0.52 and k = 0.66 with EV-CB, and r = 0.45 and k = 0.44 with PMN-total. Moreover, correlation and agreement between EV-CB and PMN-total were r = 0.60 and k = 0.50, respectively. More PMN cells (P < 0.05) were detected in PMN-SC when compared to PMN

  20. A Simple Protocol for High Efficiency Protein Isolation After RNA Isolation from Mouse Thyroid and Other Very Small Tissue Samples.

    PubMed

    Ziros, Panos G; Chartoumpekis, Dionysios V; Sykiotis, Gerasimos P

    2016-01-01

    As a dedicated hormone-secreting organ, the thyroid gland possesses a complement of proteostatic systems, including antioxidant, unfolded protein, and autophagic responses. The vast majority of animal investigations of thyroid physiology and, more recently, proteostasis, have utilized as model the rat, rather than the mouse. This is due to the very small size of the thyroid gland in the latter, with a total weight of ~2 mg (~1 mg per thyroid lobe). However, this strategy has limited the utilization of genetic approaches, such as taking advantage of the various transgenic and knockout mouse models. Here, we describe a simple and highly efficient protocol for the simultaneous isolation of mRNA, micro-RNA and 150-200 μg of protein from as little as 1 mg of mouse thyroid tissue, the average weight of one of the two thyroid lobes, thus preserving the other lobe for immunohistochemical or other analyses. While our workflow is similar to other protocols published in the literature and/or proposed by commercial reagent providers, we have introduced a key modification that addresses efficiently the most challenging step of the protein isolation process: the solubilization of the protein pellet after RNA extraction and protein precipitation. We demonstrate the feasibility of our approach and its utility for downstream analyses (including Western blotting) that facilitate the comparative study of proteostatic pathways in the mouse thyroid. We have also successfully applied this protocol on samples from mouse liver, brown and white adipose tissue, as well as from rodent cell lines. PMID:27613051

  1. Exploring the cellular and tissue uptake of nanomaterials in a range of biological samples using multimodal nonlinear optical microscopy.

    PubMed

    Johnston, Helinor J; Mouras, Rabah; Brown, David M; Elfick, Alistair; Stone, Vicki

    2015-12-18

    The uptake of nanomaterials (NMs) by cells is critical in determining their potential biological impact, whether beneficial or detrimental. Thus, investigation of NM internalization by cells is a common consideration in hazard and efficacy studies. There are currently a number of approaches that are routinely used to investigate NM-cell interactions, each of which have their own advantages and limitations. Ideally, imaging modalities used to investigate NM uptake by cells should not require the NM to be labelled (e.g. with fluorophores) to facilitate its detection. We present a multimodal imaging approach employing a combination of label-free microscopies that can be used to investigate NM-cell interactions. Coherent anti-Stokes Raman scattering microscopy was used in combination with either two-photon photoluminescence or four-wave mixing (FWM) to visualize the uptake of gold or titanium dioxide NMs respectively. Live and fixed cell imaging revealed that NMs were internalized by J774 macrophage and C3A hepatocyte cell lines (15-31 μg ml(-1)). Sprague Dawley rats were exposed to NMs (intratracheal instillation, 62 μg) and NMs were detected in blood and lung leucocytes, lung and liver tissue, demonstrating that NMs could translocate from the exposure site. Obtained data illustrate that multimodal nonlinear optical microscopy may help overcome current challenges in the assessment of NM cellular uptake and biodistribution. It is therefore a powerful tool that can be used to investigate unlabelled NM cellular and tissue uptake in three dimensions, requires minimal sample preparation, and is applicable to live and fixed cells. PMID:26584818

  2. Exploring the cellular and tissue uptake of nanomaterials in a range of biological samples using multimodal nonlinear optical microscopy

    NASA Astrophysics Data System (ADS)

    Johnston, Helinor J.; Mouras, Rabah; Brown, David M.; Elfick, Alistair; Stone, Vicki

    2015-12-01

    The uptake of nanomaterials (NMs) by cells is critical in determining their potential biological impact, whether beneficial or detrimental. Thus, investigation of NM internalization by cells is a common consideration in hazard and efficacy studies. There are currently a number of approaches that are routinely used to investigate NM-cell interactions, each of which have their own advantages and limitations. Ideally, imaging modalities used to investigate NM uptake by cells should not require the NM to be labelled (e.g. with fluorophores) to facilitate its detection. We present a multimodal imaging approach employing a combination of label-free microscopies that can be used to investigate NM-cell interactions. Coherent anti-Stokes Raman scattering microscopy was used in combination with either two-photon photoluminescence or four-wave mixing (FWM) to visualize the uptake of gold or titanium dioxide NMs respectively. Live and fixed cell imaging revealed that NMs were internalized by J774 macrophage and C3A hepatocyte cell lines (15-31 μg ml-1). Sprague Dawley rats were exposed to NMs (intratracheal instillation, 62 μg) and NMs were detected in blood and lung leucocytes, lung and liver tissue, demonstrating that NMs could translocate from the exposure site. Obtained data illustrate that multimodal nonlinear optical microscopy may help overcome current challenges in the assessment of NM cellular uptake and biodistribution. It is therefore a powerful tool that can be used to investigate unlabelled NM cellular and tissue uptake in three dimensions, requires minimal sample preparation, and is applicable to live and fixed cells.

  3. A novel method for single sample multi-axial nanoindentation of hydrated heterogeneous tissues based on testing great white shark jaws.

    PubMed

    Ferrara, Toni L; Boughton, Philip; Slavich, Eve; Wroe, Stephen

    2013-01-01

    Nanomechanical testing methods that are suitable for a range of hydrated tissues are crucial for understanding biological systems. Nanoindentation of tissues can provide valuable insights into biology, tissue engineering and biomimetic design. However, testing hydrated biological samples still remains a significant challenge. Shark jaw cartilage is an ideal substrate for developing a method to test hydrated tissues because it is a unique heterogeneous composite of both mineralized (hard) and non-mineralized (soft) layers and possesses a jaw geometry that is challenging to test mechanically. The aim of this study is to develop a novel method for obtaining multidirectional nanomechanical properties for both layers of jaw cartilage from a single sample, taken from the great white shark (Carcharodon carcharias). A method for obtaining multidirectional data from a single sample is necessary for examining tissue mechanics in this shark because it is a protected species and hence samples may be difficult to obtain. Results show that this method maintains hydration of samples that would otherwise rapidly dehydrate. Our study is the first analysis of nanomechanical properties of great white shark jaw cartilage. Variation in nanomechanical properties were detected in different orthogonal directions for both layers of jaw cartilage in this species. The data further suggest that the mineralized layer of shark jaw cartilage is less stiff than previously posited. Our method allows multidirectional nanomechanical properties to be obtained from a single, small, hydrated heterogeneous sample. Our technique is therefore suitable for use when specimens are rare, valuable or limited in quantity, such as samples obtained from endangered species or pathological tissues. We also outline a method for tip-to-optic calibration that facilitates nanoindentation of soft biological tissues. Our technique may help address the critical need for a nanomechanical testing method that is applicable

  4. A Novel Method for Single Sample Multi-Axial Nanoindentation of Hydrated Heterogeneous Tissues Based on Testing Great White Shark Jaws

    PubMed Central

    Ferrara, Toni L.; Boughton, Philip; Slavich, Eve; Wroe, Stephen

    2013-01-01

    Nanomechanical testing methods that are suitable for a range of hydrated tissues are crucial for understanding biological systems. Nanoindentation of tissues can provide valuable insights into biology, tissue engineering and biomimetic design. However, testing hydrated biological samples still remains a significant challenge. Shark jaw cartilage is an ideal substrate for developing a method to test hydrated tissues because it is a unique heterogeneous composite of both mineralized (hard) and non-mineralized (soft) layers and possesses a jaw geometry that is challenging to test mechanically. The aim of this study is to develop a novel method for obtaining multidirectional nanomechanical properties for both layers of jaw cartilage from a single sample, taken from the great white shark (Carcharodon carcharias). A method for obtaining multidirectional data from a single sample is necessary for examining tissue mechanics in this shark because it is a protected species and hence samples may be difficult to obtain. Results show that this method maintains hydration of samples that would otherwise rapidly dehydrate. Our study is the first analysis of nanomechanical properties of great white shark jaw cartilage. Variation in nanomechanical properties were detected in different orthogonal directions for both layers of jaw cartilage in this species. The data further suggest that the mineralized layer of shark jaw cartilage is less stiff than previously posited. Our method allows multidirectional nanomechanical properties to be obtained from a single, small, hydrated heterogeneous sample. Our technique is therefore suitable for use when specimens are rare, valuable or limited in quantity, such as samples obtained from endangered species or pathological tissues. We also outline a method for tip-to-optic calibration that facilitates nanoindentation of soft biological tissues. Our technique may help address the critical need for a nanomechanical testing method that is applicable

  5. Histological observations in the Hawaiian reef coral, Porites compressa, affected by Porites bleaching with tissue loss

    USGS Publications Warehouse

    Sudek, M.; Work, T.M.; Aeby, G.S.; Davy, S.K.

    2012-01-01

    The scleractinian finger coral Porites compressa is affected by the coral disease Porites bleaching with tissue loss (PBTL). This disease initially manifests as bleaching of the coenenchyme (tissue between polyps) while the polyps remain brown with eventual tissue loss and subsequent algal overgrowth of the bare skeleton. Histopathological investigation showed a loss of symbiont and melanin-containing granular cells which was more pronounced in the coenenchyme than the polyps. Cell counts confirmed a 65% reduction in symbiont density. Tissue loss was due to tissue fragmentation and necrosis in affected areas. In addition, a reduction in putative bacterial aggregate densities was found in diseased samples but no potential pathogens were observed.

  6. Histological observations in the Hawaiian reef coral, Porites compressa, affected by Porites bleaching with tissue loss.

    PubMed

    Sudek, M; Work, T M; Aeby, G S; Davy, S K

    2012-10-01

    The scleractinian finger coral Porites compressa is affected by the coral disease Porites bleaching with tissue loss (PBTL). This disease initially manifests as bleaching of the coenenchyme (tissue between polyps) while the polyps remain brown with eventual tissue loss and subsequent algal overgrowth of the bare skeleton. Histopathological investigation showed a loss of symbiont and melanin-containing granular cells which was more pronounced in the coenenchyme than the polyps. Cell counts confirmed a 65% reduction in symbiont density. Tissue loss was due to tissue fragmentation and necrosis in affected areas. In addition, a reduction in putative bacterial aggregate densities was found in diseased samples but no potential pathogens were observed.

  7. Histopathological review of 667 cases of oral mucoceles with emphasis on uncommon histopathological variations.

    PubMed

    de Brito Monteiro, Bárbara Vanessa; Bezerra, Thâmara Manoela Marinho; da Silveira, Éricka Janine Dantas; Nonaka, Cassiano Francisco Weege; da Costa Miguel, Márcia Cristina

    2016-04-01

    Mucoceles can occur in the oral cavity, appendix, bladder, paranasal sinuses, and lacrimal sac. In the oral cavity, mucoceles arise from pathological alterations in the minor salivary gland ducts. In this study, we aimed to histologically reevaluate cases of oral mucoceles to identify possible variants. A total of 667 slides containing tissue sections stained with hematoxylin and eosin diagnosed as a phenomenon of mucus extravasation were analyzed under light microscopy by 4 previously trained examiners. In 128 cases (19.1%), 1 or more histopathological changes were identified. Twenty cases (2.9%) exhibited collagenous globular structures compatible with myxoglobulosis. In 30 cases (4.49%), dissociation of collagen fibers after mucin extravasation was observed. Fifty-four cases (8.09%) exhibited papillary synovial metaplasia-like change, and 32 (4.79%) showed a significant reduction in the lumen of the cavity due to large papillae. Twenty cases (2.9%) were compatible with superficial mucoceles, and in 11 cases (1.64%), the foamy macrophages showed an unusual solid arrangement, known as clear cell change. It is essential to recognize the possible histopathological changes in oral mucoceles to avoid diagnostic pitfalls. PMID:27040930

  8. Histopathology of meibomian gland dysfunction.

    PubMed

    Gutgesell, V J; Stern, G A; Hood, C I

    1982-09-01

    We conducted a histopathologic study of he meibomian glands of seven patients (all men, ranging in age from 58 to 83 years) who had severe or moderately severe meibomian dysfunction and who were undergoing ectropion or entropion repair. Abnormal features included signs of obstruction and dilatation of ducts, enlargement of acini with cystic degeneration and squamous metaplasia, foreign-body reaction and granuloma formation, a mild increase in inflammatory cells, and abnormal keratinization. Demodex organisms were found in both acini and ducts of one patient. These findings were similar to those reported in other entities involving meibomian duct obstruction, probably related to abnormalities of keratinization, plays an important role in the pathogenesis of meibomian gland dysfunction.

  9. Measurement of Inflammatory Chemokines in Micro-dissected Tissue Biopsy Samples by Chip-Based Immunoaffinity Capillary Electrophoresis.

    PubMed

    Phillips, Terry M; Wellner, Edward; McMohan, Shane; Kalish, Heather

    2016-01-01

    To aid in the biochemical analysis of human skin biopsies, a chip-based immunoaffinity capillary electrophoresis (ICE) system has been developed for measuring inflammatory chemokines in micro-dissected areas of the biopsy. Following isolation of the areas of interest, the tissue was solubilized and the analytes of interest were isolated by the immunoaffinity disk within the chip. The captured analytes were labeled in situ with a 635 nm light-emitting laser dye and electro-eluted into the chip separation channel. Electrophoretic separation of all of the analytes was achieved in 2.5 min with quantification of each peak being performed by online LIF detection and integration of each peak area. The degree of accuracy and precision achieved by the chip-based system is comparable to conventional immunoassays and the system is robust enough to be applied to the analysis of clinical samples. Further, with the expanding array of antibodies that are commercially available, this chip-based system can be applied to a wide variety of different biomedical and clinical analyses. PMID:27473486

  10. Confidentiality, informed consent, and children's participation in research involving stored tissue samples: interviews with medical professionals from the Middle East.

    PubMed

    Alahmad, Ghiath; Al Jumah, Mohammed; Dierickx, Kris

    2015-01-01

    Ethical issues regarding research biobanks continue to be a topic of intense debate, especially issues of confidentiality, informed consent, and child participation. Although considerable empirical literature concerning research biobank ethics exists, very little information is available regarding the opinions of medical professionals doing genetics research from the Middle East, especially Arabic speaking countries. Ethical guidelines for research biobanks are critically needed as some countries in the Middle East are starting to establish national research biobanks. Islam is the dominant religion in these countries, and it affects people's behavior and influences their positions. Moreover, communities in these countries enjoy a set of customs, traditions and social norms, and have social and familial structures that must be taken into account when developing research policies. We interviewed 12 medical professionals from the Middle East currently working with stored tissue samples to document their opinions. We found general agreement. Participants' primary concerns were similar to the views of researchers internationally. Since children tend to represent a high percentage of Middle Eastern populations, and because children's bodies are not just small adult bodies, the interviewed professionals strongly believed that it is imperative to include children in biobank research. Participants generally believed that protecting confidentiality is socially very important and that informed consent/assent must be obtained from both adult and child participants. This study provides a starting point for additional studies.

  11. [Detection of B-lymphocyte clonality in samples of salivary gland tissue in patients with primary Sjogren syndrome].

    PubMed

    Vojvodic, D; Magic, Z; Stefanovic, D; Cikota, B; Ilic, V; Jovic, M; Tatic, V; Colic, M

    2001-01-01

    Intensive lymphoplasmocytic infiltration with atrophy of glandular tissue structures is the dominant patohistological feature found in exocrine glands of patients with Sjögren syndrome (SS). The infiltrates consist of T and B lymphocyte clusters that make the structures resembling germinal centers, and numerous plasmocytes that are secreting imunoglobulines locally, including autoantibodies. By applying the polymerase chain reaction (PCR) in our study we have shown the existence of dominant B cell clone in salivary glands samples of 4 out of 6 patients with SS, in the absence of clinical, routine laboratory, and patohistological signs of the lymphoma. B lymphocyte clones were detected upon the amplification of gene segment that encoded variable heavy chain immunoglobulin CDR3 region. Finding of single, dominant B lymphocyte clone could be of predictive significance, because these patients are predisposed to non-Hodgkin lymphoma (NHL) for which there is an assumption that it originates out of salivary glands from one of the clusters of proliferating B lymphocytes. PMID:11769416

  12. Application of a Liquid Extraction Based Sealing Surface Sampling Probe for Mass Spectrometric Analysis of Dried Blood Spots and Mouse Whole-Body Thin Tissue Sections

    SciTech Connect

    Van Berkel, Gary J; Kertesz, Vilmos

    2009-01-01

    The utility of a liquid extraction based sealing surface sampling probe (SSSP) for the direct mass spectrometric analysis of targeted drugs and metabolites in dried blood spots (DBSs) and whole mouse thin tissue sections was demonstrated. The accuracy and precision for the quantitative analysis of a minimum of 50 ng/mL sitamaquine or acetaminophen in DBSs on paper were well within the required 15% dictated by internationally recognized acceptance criteria for assay validations. Analysis of whole-body mouse thin tissue sections from animals dosed with propranolol, adhered to an adhesive tape substrate, provided semi-quantitative information for propranolol and its hydroxyproranolol glucuronide metabolite within specific organs of the tissue. The relative abundances recorded for the two compounds in the brain, lung, kidney and liver were in nominal agreement with previously reported amounts based on analysis using a liquid microjunction surface sampling probe (LMJ-SSP), and whole-body autoradiography (WBA) and HPLC-MS analysis. The ability to sample and analyze from tape-adhered tissue samples, which are generally employed in WBA analysis, presents the possibility of consecutive WBA and SSSP-MS analysis of the same tissue section. This would facilitate assignment, and possibly quantitation, of the different molecular forms of total drug related material detected in the WBA analysis. The flexibility to sample larger or smaller spot sizes, alternative probe sealing mechanisms, and a reduction in internal volumes and associated sample carryover issues will be among the first simple improvements necessary to make the SSSP-MS method a practical DBS and/or thin tissue section analysis tool or to expand its use to other surface sampling applications.

  13. Chondroid syringoma of the scalp: case report and discussion of clinical features, histopathology, and treatment.

    PubMed

    Paik, Young S; Liess, Benjamin D

    2011-04-01

    Chondroid syringoma is a generally benign, adnexal tumor most commonly found in the head and neck. It is typically diagnosed retrospectively from histopathology, and careful evaluation must differentiate it from tumors of salivary gland origin. It presents as a slowly growing, subcutaneous or intradermal nodule. Treatment consists of wide local excision with a margin of normal tissue. We report a case involving a 51-year-old man with a chondroid syringoma arising from the scalp. Clinical features, histopathology, and treatment are discussed.

  14. The Histopathological Spectrum of Pyogenic Granuloma: A Case Series.

    PubMed

    Marla, Vinay; Shrestha, Ashish; Goel, Khushboo; Shrestha, Sajeev

    2016-01-01

    Background. Pyogenic granuloma is a reactive tumor-like lesion commonly affecting the oral cavity. These lesions usually appear as localized solitary nodule with a sessile or pedunculated base and colour varying from red, purplish, or pink, depending on the vascularity of the lesion. Pyogenic granuloma shows predilection for gingiva and is usually slow growing, but at times it shows rapid growth. The natural course of this lesion can be categorized into three distinct phases, namely, (i) cellular phase, (ii) capillary phase/vascular phase, and (iii) involutionary phase. Histopathologically, pyogenic granuloma is classified into lobular capillary hemangioma (LCH) and non-lobular capillary hemangioma (non-LCH). Case Presentation. In this series, four cases (varied age groups and both genders) of pyogenic granuloma showing varying histopathological presentation in relation to its clinical course have been described. The lesion in its early phase reveals diffuse endothelial cells, with few budding into capillaries. Among the capillary phase, the LCH type shows numerous blood vessels organized into lobular aggregates whereas the non-LCH type does not show any such organization and resembles granulation tissue. The involutionary phase shows healing of the lesion and is characterized by extensive fibrosis in the connective tissue. Conclusion. In conclusion, knowledge of the various histopathological presentation of this lesion is necessary for proper identification. PMID:27382492

  15. Weakly supervised histopathology cancer image segmentation and classification.

    PubMed

    Xu, Yan; Zhu, Jun-Yan; Chang, Eric I-Chao; Lai, Maode; Tu, Zhuowen

    2014-04-01

    Labeling a histopathology image as having cancerous regions or not is a critical task in cancer diagnosis; it is also clinically important to segment the cancer tissues and cluster them into various classes. Existing supervised approaches for image classification and segmentation require detailed manual annotations for the cancer pixels, which are time-consuming to obtain. In this paper, we propose a new learning method, multiple clustered instance learning (MCIL) (along the line of weakly supervised learning) for histopathology image segmentation. The proposed MCIL method simultaneously performs image-level classification (cancer vs. non-cancer image), medical image segmentation (cancer vs. non-cancer tissue), and patch-level clustering (different classes). We embed the clustering concept into the multiple instance learning (MIL) setting and derive a principled solution to performing the above three tasks in an integrated framework. In addition, we introduce contextual constraints as a prior for MCIL, which further reduces the ambiguity in MIL. Experimental results on histopathology colon cancer images and cytology images demonstrate the great advantage of MCIL over the competing methods.

  16. The Histopathological Spectrum of Pyogenic Granuloma: A Case Series

    PubMed Central

    Shrestha, Sajeev

    2016-01-01

    Background. Pyogenic granuloma is a reactive tumor-like lesion commonly affecting the oral cavity. These lesions usually appear as localized solitary nodule with a sessile or pedunculated base and colour varying from red, purplish, or pink, depending on the vascularity of the lesion. Pyogenic granuloma shows predilection for gingiva and is usually slow growing, but at times it shows rapid growth. The natural course of this lesion can be categorized into three distinct phases, namely, (i) cellular phase, (ii) capillary phase/vascular phase, and (iii) involutionary phase. Histopathologically, pyogenic granuloma is classified into lobular capillary hemangioma (LCH) and non-lobular capillary hemangioma (non-LCH). Case Presentation. In this series, four cases (varied age groups and both genders) of pyogenic granuloma showing varying histopathological presentation in relation to its clinical course have been described. The lesion in its early phase reveals diffuse endothelial cells, with few budding into capillaries. Among the capillary phase, the LCH type shows numerous blood vessels organized into lobular aggregates whereas the non-LCH type does not show any such organization and resembles granulation tissue. The involutionary phase shows healing of the lesion and is characterized by extensive fibrosis in the connective tissue. Conclusion. In conclusion, knowledge of the various histopathological presentation of this lesion is necessary for proper identification. PMID:27382492

  17. Assessment of the histopathological lesions and chemical analysis of feral cats to the smoke from the Kuwait oil fires.

    PubMed

    Moeller, R B; Kalasinsky, V F; Razzaque, M; Centeno, J A; Dick, E J; Abdal, M; Petrov, I I; DeWitt, T W; al-Attar, M; Pletcher, J M

    1994-01-01

    Twenty-six adult or subadult feral cats were collected from Kuwait approximately 8 months after the ignition of the Kuwait oil wells. These animals were obtained from two sources: 12 animals from Kuwait City, a relatively smoke-free area, and 14 from the city of Ahmadi, an area with heavy smoke. Animals were euthanized and a complete set of tissues consisting of all major organs was taken for histopathology. Samples of lung, liver, kidney, urine, and blood were also taken for toxicology. Histopathological lesions observed in the lung were mild accumulations of anthracotic pigment in the lungs of 17 cats. Hyperplasia of the bronchial and bronchiolar gland in 8 cats, and smooth muscle hyperplasia of bronchioles in 14 cats. Tracheal gland hyperplasia was observed in 7 cats, and minimal squamous metaplasia of the tracheal mucosa in 17 cats, Laryngeal lesions consisted of submucosal gland hyperplasia in 2 cats and squamous metaplasia of the mucosa in 5 cats. Hyperplasia of the nasal submucosal glands was observed in 6 animals. The pharyngeal mucosa as well as other organs and organ systems were normal in all cats. Atomic absorption analysis for 11 metals was performed; vanadium and nickel levels (two metals that were present in the smoke from the oil fires) are not indicative of substantial exposure to the oil fires. Based on the histopathological findings and toxicological analysis, it is felt that inhalation of air contaminated with smoke from the oil fires had little or no long-term effect on the animals examined.

  18. Clinical, histopathological and genetic studies in a case of fatal familial insomnia with review of the literature.

    PubMed

    Peng, Bin; Zhang, Shenqi; Dong, Hongjuan; Lu, Zuneng

    2015-01-01

    To explore clinical, histopathological and genetic features of a case with fatal familial insomnia (FFI) and review the related literatures. A middle-aged woman who complained of "insomnia for 9 months and psychosis for 3 months" was suspicious of FFI. The clinical features of the patient were analyzed, and the dead patient was examined by autopsy and the brain tissues were obtained for histopathological studies, and the blood samples from the patient and some of her familial members were collected for the sequencing of prion protein gene (PRNP). The main clinical features included intractable insomnia, psychiatric symptoms and abnormal night sleep behavior, unsteady gait, difficulty swallowing, sudden death, and positive family history. The pathological studies showed neuronal loss and gliosis of multiple brain tissues in the proband, predominated with thalamus; and analysis of PRNP revealed gene D178N mutation, and linkage with 129 methionine (Met) allele in the proband and a relative. FFI patients may manifest as sudden death, and may have prominent psychiatric symptoms; the corresponding gene mutation could occur in the asymptomatic carriers; the data of autopsy and brain tissue pathology is helpful for further understanding of this disease.

  19. Value of histopathologic examination of uterine products after first-trimester miscarriage.

    PubMed

    Alsibiani, Sharifa Ali

    2014-01-01

    The main rationale of routine histopathologic examination of products after first-trimester miscarriages is to detect an ectopic pregnancy or a molar pregnancy, which require further management. An alternative approach is to examine the products only when there is a definite indication. As there is no agreement, we aimed to study whether routine histopathological examination of tissues obtained after first-trimester miscarriage is of any clinical value in our populations. Medical records of all (558) patients with a diagnosis of first-trimester miscarriage over 4 years (2007-2010) at King Abdulaziz University Hospital, Jeddah, Saudi Arabia, were reviewed. Histopathologic examination confirmed products of conception in 537 (96.2%) patients, no products of conception in 17 (3%) patients, molar pregnancy in 2 (0.4%) patients, and decidual tissues without chorionic villi (Arias-Stella reaction) in 2 (0.4%) patients. After clinical correlation, only one unsuspected partial molar pregnancy was diagnosed by histopathology examination. Conclusion is that it does not appear reasonable to perform histopathological examination routinely after all first-trimester miscarriages in our studied population. We recommend that histopathological examination be performed in select instances: when the diagnosis is uncertain, when fewer tissues have been obtained during surgery, when unexpected pathology was seen, when ultrasound suggests a molar pregnancy, or when patients are considered at high risk for trophoblastic disease.

  20. Imaging and histopathological aspects in aseptic osteonecrosis of the femoral head.

    PubMed

    Trăistaru, Magdalena Rodica; Kamal, Diana; Kamal, Kamal Constantin; Rogoveanu, Otilia Constantina; Popescu, Mihai; Bondari, Simona; Alexandru, Dragoş Ovidiu; Ionovici, Nina; Grecu, Dan Cristian

    2015-01-01

    Aseptic osteonecrosis causes various clinical manifestations, depending on its location, but has in common a histopathological and radiological substrate. Aseptic osteonecrosis of the femoral head is a condition whose pathogenesis remains unclear despite many theories developed so far, and the discovery of numerous risk factors. The objective of this study is to emphasize the role of imaging techniques and correlating histology and immunohistochemistry methods in order to more accurately stage the disease. This retrospective study was performed on a total of 103 patients with clinical and radiological suspicion of unilateral or bilateral osteonecrosis. For the diagnosis criteria, we used clinical information, pelvic X-ray images, magnetic resonance imaging (MRI) or computed tomography (CT). For the inclusion of patients in a disease stage, we used the Association Research Circulation Osseous (ARCO) classification system. For patients diagnosed at an advanced stage, who underwent hip arthroplasty, we harvested biological material necessary for the histopathological study. There were differences in the appearance and extent of the lesion on the histological samples compared to macroscopic examination and even those obtained through imaging means, particularly for patients in evolutionary stage III. Aspects such as the extension of the area of fibrosis, bone tissue remodeling, the density of the newly formed vascular network and degree of impairment of the cartilage, are determined more accurately using histology and immunohistochemistry techniques. Before classifying patients in a certain stage, after correlating clinical and imaging data, histopathological aspects have to be considered, particularly in patients in stages III and IV, in which total hip arthroplasty could be delayed.

  1. Comparison of two histopathologic methods for evaluating subcutaneous reaction to mineral trioxide aggregate

    PubMed Central

    Lotfi, Mehrdad; Moradzadeh, Monir; Aghbali, Amirala; Rahimi, Saeed; Saghiri, Mohammadali; Zand, Vahid; Mehdipour, Masoumeh; Ranjkesh, Bahram; Doosti, Sirvan

    2012-01-01

    Objectives: One of the most important factors for suitable materials for pulp therapy is biocompatibility. Two histopathologic methods of Cox and Federation Dentaire International (FDI) were used to evaluate inflammation. In Cox method, density of inflammatory cells, tissue reactions like fibrosis, vascular responses like congestion and fibrin extravasation have been used to evaluate inflammatory reactions. The aim of this study was to compare the accuracy of pathologists’ interpretations using two different methods. Study design: Three pathologists observed the degree of inflammation in 225 histopathologic sections. These sections showed inflammation in subcutaneous connective tissue of rats adjacent to polyethylene tubes, filled with white or gray mineral trioxide aggregate. Empty tubes served as controls. Samples were harvested after 7-, 15-, 30-, 60-, and 90-days. All pathologists examined the sections under a light microscope (Carl Zeiss, Oberkochen, Germany) at ×400 magnifications. Chi-Square test was used to evaluate the difference between inflammation grades when one pathologist used two methods. Cohen’s Kappa value was used to measure agreement of three pathologists to recognize the degrees of inflammations when using one of the methods. Results: There were no significant differences between the two methods when one of the pathologist used these methods to report the degree of inflammation (p=0.054). However, two other pathologists reported significant differences between two methods (p=0.005, p=0.001). In the FDI method, there was an acceptable agreement between first and second, and first and third pathologist in terms of the degree of inflammation, and intermediate agreement existed between the second and third pathologist. With the Cox method, no agreement among the pathologists could be found. Conclusion: The results of three pathologists in terms of rating inflammation with the FDI method showed better agreement than with the Cox method

  2. Primary adenocarcinoma of ureter: A rare histopathological variant

    PubMed Central

    Chaudhary, Prekshi; Agarwal, Rashi; Srinivasan, Shashank; Singh, Dinesh

    2016-01-01

    Primary carcinoma of ureter is an uncommon malignancy. Of which, mostly are transitional cell carcinomas followed by squamous cell carcinomas and adenocarcinomas being the rarest histopathology encountered. We report a case of adenocarcinoma ureter in a middle-aged male along with its clinical scenario. A 62-year-old male, presented with complaints of lower urinary tract symptoms. Computerized tomography urogram showed a soft tissue lesion at the right ureterovesical junction. Cystoscopic biopsy reported villous adenoma. Diethylene triamine pentaacetic acid scan reported nonfunctioning right kidney. He underwent laparoscopic right nephroureterectomy, and histopathology reported adenocarcinoma of the right lower third of ureter, with positive distal and close radial margins. The patient received external beam radiation to the postoperative bed and lymph nodes, and he is disease-free till date. PMID:27453661

  3. Scalable histopathological image analysis via active learning.

    PubMed

    Zhu, Yan; Zhang, Shaoting; Liu, Wei; Metaxas, Dimitris N

    2014-01-01

    Training an effective and scalable system for medical image analysis usually requires a large amount of labeled data, which incurs a tremendous annotation burden for pathologists. Recent progress in active learning can alleviate this issue, leading to a great reduction on the labeling cost without sacrificing the predicting accuracy too much. However, most existing active learning methods disregard the "structured information" that may exist in medical images (e.g., data from individual patients), and make a simplifying assumption that unlabeled data is independently and identically distributed. Both may not be suitable for real-world medical images. In this paper, we propose a novel batch-mode active learning method which explores and leverages such structured information in annotations of medical images to enforce diversity among the selected data, therefore maximizing the information gain. We formulate the active learning problem as an adaptive submodular function maximization problem subject to a partition matroid constraint, and further present an efficient greedy algorithm to achieve a good solution with a theoretically proven bound. We demonstrate the efficacy of our algorithm on thousands of histopathological images of breast microscopic tissues. PMID:25320821

  4. Fish biomarkers for environmental monitoring: An integrated model supporting enzyme activity and histopathological lesions

    NASA Astrophysics Data System (ADS)

    Neta, Raimunda Nonata Fortes Carvalho; Torres Junior, Audalio Rebelo

    2014-10-01

    We present a mathematical model describing the association between glutathione-S-transferase activity and brachial lesions in the catfish, Sciades herzbergii (Ariidae) from a polluted port. The catfish were sampled from a port known to be contaminated with heavy metals and organic compounds and from a natural reserve in São Marcos Bay, Brazil. Two biomarkers, hepatic glutathione S-transferase (GST) activity and histopathological lesions, in gills tissue were measured. The values for GST activity were modeled with the occurrence of branchial lesions by fitting a third order polynomial. Results from the mathematical model indicate that GST activity has a strong polynomial relationship with the occurrence of branchial lesions in both the wet and the dry seasons, but only at the polluted port site. The model developed in this study indicates that branchial and hepatic lesions are initiated when GST activity reaches 2.15 μmol min-1 mg protein-1. Beyond this limit, GST activity decreased to very low levels and irreversible histopathological lesions occurred. This mathematical model provides a realistic approach to analyze predictive biomarkers of environmental health status.

  5. 42 CFR 493.1273 - Standard: Histopathology.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ... 42 Public Health 5 2010-10-01 2010-10-01 false Standard: Histopathology. 493.1273 Section 493.1273 Public Health CENTERS FOR MEDICARE & MEDICAID SERVICES, DEPARTMENT OF HEALTH AND HUMAN SERVICES... Systems § 493.1273 Standard: Histopathology. (a) As specified in § 493.1256(e)(3), fluorescent...

  6. Evaluation of cytokeratin-19 in breast cancer tissue samples: a comparison of automatic and manual evaluations of scanned tissue microarray cylinders

    PubMed Central

    2015-01-01

    Background Digital image (DI) analysis avoids visual subjectivity in interpreting immunohistochemical stains and provides more reproducible results. An automated procedure consisting of two variant methods for quantifying the cytokeratin-19 (CK19) marker in breast cancer tissues is presented. Methods The first method (A) excludes the holes inside selected CK19 stained areas, and the second (B) includes them. 93 DIs scanned from complete cylinders of tissue microarrays were evaluated visually by two pathologists and by the automated procedures. Results and conclusions There was good concordance between the two automated methods, both of which tended to identify a smaller CK19-positive area than did the pathologists. The results obtained with method B were more similar to those of the pathologists; probably because it takes into account the entire positive tumoural area, including the holes. However, the pathologists overestimated the positive area of CK19. Further studies are needed to confirm the utility of this automated procedure in prognostic studies. PMID:26329009

  7. Influence of sample preparation and reliability of automated numerical refocusing in stain-free analysis of dissected tissues with quantitative phase digital holographic microscopy

    NASA Astrophysics Data System (ADS)

    Kemper, Björn; Lenz, Philipp; Bettenworth, Dominik; Krausewitz, Philipp; Domagk, Dirk; Ketelhut, Steffi

    2015-05-01

    Digital holographic microscopy (DHM) has been demonstrated to be a versatile tool for high resolution non-destructive quantitative phase imaging of surfaces and multi-modal minimally-invasive monitoring of living cell cultures in-vitro. DHM provides quantitative monitoring of physiological processes through functional imaging and structural analysis which, for example, gives new insight into signalling of cellular water permeability and cell morphology changes due to toxins and infections. Also the analysis of dissected tissues quantitative DHM phase contrast prospects application fields by stain-free imaging and the quantification of tissue density changes. We show that DHM allows imaging of different tissue layers with high contrast in unstained tissue sections. As the investigation of fixed samples represents a very important application field in pathology, we also analyzed the influence of the sample preparation. The retrieved data demonstrate that the quality of quantitative DHM phase images of dissected tissues depends strongly on the fixing method and common staining agents. As in DHM the reconstruction is performed numerically, multi-focus imaging is achieved from a single digital hologram. Thus, we evaluated the automated refocussing feature of DHM for application on different types of dissected tissues and revealed that on moderately stained samples highly reproducible holographic autofocussing can be achieved. Finally, it is demonstrated that alterations of the spatial refractive index distribution in murine and human tissue samples represent a reliable absolute parameter that is related of different degrees of inflammation in experimental colitis and Crohn's disease. This paves the way towards the usage of DHM in digital pathology for automated histological examinations and further studies to elucidate the translational potential of quantitative phase microscopy for the clinical management of patients, e.g., with inflammatory bowel disease.

  8. A statistical model and national data set for partioning fish-tissue mercury concentration variation between spatiotemporal and sample characteristic effects

    USGS Publications Warehouse

    Wente, Stephen P.

    2004-01-01

    Many Federal, Tribal, State, and local agencies monitor mercury in fish-tissue samples to identify sites with elevated fish-tissue mercury (fish-mercury) concentrations, track changes in fish-mercury concentrations over time, and produce fish-consumption advisories. Interpretation of such monitoring data commonly is impeded by difficulties in separating the effects of sample characteristics (species, tissues sampled, and sizes of fish) from the effects of spatial and temporal trends on fish-mercury concentrations. Without such a separation, variation in fish-mercury concentrations due to differences in the characteristics of samples collected over time or across space can be misattributed to temporal or spatial trends; and/or actual trends in fish-mercury concentration can be misattributed to differences in sample characteristics. This report describes a statistical model and national data set (31,813 samples) for calibrating the aforementioned statistical model that can separate spatiotemporal and sample characteristic effects in fish-mercury concentration data. This model could be useful for evaluating spatial and temporal trends in fishmercury concentrations and developing fish-consumption advisories. The observed fish-mercury concentration data and model predictions can be accessed, displayed geospatially, and downloaded via the World Wide Web (http://emmma.usgs.gov). This report and the associated web site may assist in the interpretation of large amounts of data from widespread fishmercury monitoring efforts.

  9. Nonlesions, misdiagnoses, missed diagnoses, and other interpretive challenges in fish histopathology studies: a guide for investigators, authors, reviewers, and readers

    USGS Publications Warehouse

    Wolf, Jeffrey C.; Baumgartner, Wes A.; Blazer, Vicki; Camus, Alvin C.; Engelhardt, Jeffrey A.; Fournie, John W.; Frasca, Salvatore; Groman, David B.; Kent, Michael L.; Khoo, Lester H.; Law, Jerry M.; Lombardini, Eric D.; Ruehl-Fehlert, Christine; Segner, Helmut E.; Smith, Stephen A.; Spitsbergen, Jan M.; Weber, Klaus; Wolfe, Marilyn J.

    2015-01-01

    Differentiating salient histopathologic changes from normal anatomic features or tissue artifacts can be decidedly challenging, especially for the novice fish pathologist. As a consequence, findings of questionable accuracy may be reported inadvertently, and the potential negative impacts of publishing inaccurate histopathologic interpretations are not always fully appreciated. The objectives of this article are to illustrate a number of specific morphologic findings in commonly examined fish tissues (e.g., gills, liver, kidney, and gonads) that are frequently either misdiagnosed or underdiagnosed, and to address related issues involving the interpretation of histopathologic data. To enhance the utility of this article as a guide, photomicrographs of normal and abnormal specimens are presented. General recommendations for generating and publishing results from histopathology studies are additionally provided. It is hoped that the furnished information will be a useful resource for manuscript generation, by helping authors, reviewers, and readers to critically assess fish histopathologic data.

  10. Nonlesions, misdiagnoses, missed diagnoses, and other interpretive challenges in fish histopathology studies: a guide for investigators, authors, reviewers, and readers.

    PubMed

    Wolf, Jeffrey C; Baumgartner, Wes A; Blazer, Vicki S; Camus, Alvin C; Engelhardt, Jeffery A; Fournie, John W; Frasca, Salvatore; Groman, David B; Kent, Michael L; Khoo, Lester H; Law, Jerry M; Lombardini, Eric D; Ruehl-Fehlert, Christine; Segner, Helmut E; Smith, Stephen A; Spitsbergen, Jan M; Weber, Klaus; Wolfe, Marilyn J

    2015-04-01

    Differentiating salient histopathologic changes from normal anatomic features or tissue artifacts can be decidedly challenging, especially for the novice fish pathologist. As a consequence, findings of questionable accuracy may be reported inadvertently, and the potential negative impacts of publishing inaccurate histopathologic interpretations are not always fully appreciated. The objectives of this article are to illustrate a number of specific morphologic findings in commonly examined fish tissues (e.g., gills, liver, kidney, and gonads) that are frequently either misdiagnosed or underdiagnosed, and to address related issues involving the interpretation of histopathologic data. To enhance the utility of this article as a guide, photomicrographs of normal and abnormal specimens are presented. General recommendations for generating and publishing results from histopathology studies are additionally provided. It is hoped that the furnished information will be a useful resource for manuscript generation, by helping authors, reviewers, and readers to critically assess fish histopathologic data.

  11. Histopathology of laser skin resurfacing

    NASA Astrophysics Data System (ADS)

    Thomsen, Sharon L.; Baldwin, Bonnie; Chi, Eric; Ellard, Jeff; Schwartz, Jon A.

    1997-05-01

    Pulsed carbon-dioxide laser skin resurfacing is a purportedly 'non-thermal' procedure enjoying wide application as a cosmetic treatment for skin wrinkles. Treatment success has been based on clinical assessments of skin smoothness. Skin lesions (1 cm2) created by one, two or three superimposed carbon-dioxide laser passes were placed on the backs of 28 'fuzzy' Harlan Sprague Dawley rats. The variable laser irradiation parameters included measured energies ranging from 112 to 387/pulse with pulse widths of 65 and 125 microseconds and a repetition rate of 8 Hz. The square, flat laser beam measured 3 mm2 at the focal point. The lesions were collected from 0 to 10 days after treatment for qualitative and quantitative histopathology. Thermal damage and treatment effect tended to increase in severity and, to a lesser extent, depth with increased delivery parameters. In acute lesions, the vacuolated and fragmented, desiccated and thermally coagulated epidermis was partially removed exposing the underlying thermally coagulated dermal collagen and cells. Epidermal and dermal necrosis and slough occurred between 24 to 72 hours after treatment. Epithelial regeneration originated from the adnexa and the lesion edges. Dermal fibrous scar formation began at 5 days below the regenerated epidermis and became more prominent at 7 and 10 days.

  12. Histopathology of gastrointestinal neuroendocrine neoplasms

    PubMed Central

    Hirabayashi, Kenichi; Zamboni, Giuseppe; Nishi, Takayuki; Tanaka, Akira; Kajiwara, Hiroshi; Nakamura, Naoya

    2013-01-01

    Gastrointestinal neuroendocrine neoplasms (GI-NENs) arise from neuroendocrine cells distributed mainly in the mucosa and submucosa of the gastrointestinal tract. In 2010, the World Health Organization (WHO) classification of NENs of the digestive system was changed, categorizing these tumors as grade 1 neuroendocrine tumor (NET), grade-2NET, neuroendocrine carcinoma (large- or small-cell type), or mixed adenoneuroendocrine carcinoma (MANEC). Such a classification is based on the Ki-67 index and mitotic count in histological material. For the accurate pathological diagnosis and grading of NENs, it is important to clearly recognize the characteristic histological features of GI-NENs and to understand the correct method of counting Ki-67 and mitoses. In this review, we focus on the histopathological features of GI-NENs, particularly regarding biopsy and cytological diagnoses, neuroendocrine markers, genetic and molecular features, and the evaluation of the Ki-67 index and mitotic count. In addition, we will address the histological features of GI-NEN in specific organs. PMID:23346552

  13. Comparison of Nanostring nCounter® Data on FFPE Colon Cancer Samples and Affymetrix Microarray Data on Matched Frozen Tissues.

    PubMed

    Chen, Xi; Deane, Natasha G; Lewis, Keeli B; Li, Jiang; Zhu, Jing; Washington, M Kay; Beauchamp, R Daniel

    2016-01-01

    The prognosis of colorectal cancer (CRC) stage II and III patients remains a challenge due to the difficulties of finding robust biomarkers suitable for testing clinical samples. The majority of published gene signatures of CRC have been generated on fresh frozen colorectal tissues. Because collection of frozen tissue is not practical for routine surgical pathology practice, a clinical test that improves prognostic capabilities beyond standard pathological staging of colon cancer will need to be designed for formalin-fixed paraffin-embedded (FFPE) tissues. The NanoString nCounter® platform is a gene expression analysis tool developed for use with FFPE-derived samples. We designed a custom nCounter® codeset based on elements from multiple published fresh frozen tissue microarray-based prognostic gene signatures for colon cancer, and we used this platform to systematically compare gene expression data from FFPE with matched microarray array data from frozen tissues. Our results show moderate correlation of gene expression between two platforms and discovery of a small subset of genes as candidate biomarkers for colon cancer prognosis that are detectable and quantifiable in FFPE tissue sections. PMID:27176004

  14. Histopathological effects of intramuscular metamizole sodium on rat sciatic nerve

    PubMed Central

    Emir, Abdurrahman; Kalkan, Yıldıray; Bostan, Habib

    2016-01-01

    Objective(s): We investigated the histopathological effects of metamizole sodium (MS) on the sciatic nerve. Materials and Methods: This study was performed using 48 adult male Wistar albino rats. Ten groups were constituted with 6 rats in each group. MS injection into the sciatic nerve (group 1), MS injection into the muscle [group 3 (50 mg/kg, 0.4 ml) and group 5 (50 mg/kg, 0.8 ml)], MS injection into the muscle cavity in the vicinity of the sciatic nerve [group 2 (50 mg/kg, 0.4 ml) and group 4 (50 mg/kg, 0.8 ml)], normal saline injection into the muscle in the vicinity of the sciatic nerve [group 6A (0.4 ml) and 6B (0.8 ml)], subjected to injury by drilling the entire layer of nerve without injecting any drug, normal saline injection in the sciatic nerve, and control group. Nerve and muscle samples were taken 7 days after administrations. Tissue sections were stained using a hematoxylin and eosin-Luxol® fast blue stain, assessed by a histologist. Results: The levels of axonal degeneration of the rats in groups 1, 2, 3, 4, 5, 6A, and 8 were found to be significantly higher compared to the levels of the rats in the control group (P<0.05). Myelin degeneration of the rats in all groups was found to be significantly higher compared to myelin degeneration of the rats in the control group (P<0.05). Conclusion: It was observed that MS could lead to injury in the sciatic nerve with a toxic effect due to diffusion. PMID:27746863

  15. Assessment of Epidermal Growth Factor Receptor (EGFR) signaling in paired colorectal cancer and normal colon tissue samples using computer-aided immunohistochemical analysis.

    PubMed

    Messersmith, Wells; Oppenheimer, Darin; Peralba, Josep; Sebastiani, Valeria; Amador, Maria; Jimeno, Antonio; Embuscado, Erlinda; Hidalgo, Manuel; Iacobuzio-Donahue, Christine

    2005-12-01

    The Epidermal Growth Factor Receptor (EGFR) plays a role in multiple tumor cell processes and is targeted by several anticancer therapies. Although EGFR mutations may determine tumor susceptibility in a small proportion of patients, knowledge of the EGFR signaling pathway status in tumors may help guide further drug development and hypothesis-driven combination studies. We aimed to validate and apply a novel computer-aided immunohistochemical (IHC) technique to characterize the status of EGFR signaling in matched colorectal tumor and normal colon tissue samples. Tissue Microarrays (TMA)were made from both cancerous and normal colorectal tissue in 18 patients and stained with antibodies against EGFR, phospho-EGFR (pEGFR), Akt, pAkt, MAPK, and pMAPK. TMA's were quantitatively scored using the Automated Cellular Imaging System (ACIS II, Chromavision, Inc). ACIS was compared against cell line Western blotting, ELISA, and visual scoring (0-3+) by a pathologist. We found that ACIS analysis was highly reproducible and results were well correlated with other techniques. A post-scan "image microdissection" technique of analyzing heterogeneous human samples showed good correlation between paired human samples [Pearson correlation for tumors, 0.922 (p < .001)]. Cancer samples had markedly higher staining of pEGFR, Akt, pAkt, MAPK, and pMAPK. We conclude that ACIS IHC of human tissue samples is quantitative, reproducible, and correlates with Western blots and ELISA in cell line pellets as well as pathologist's scores of human samples. Colorectal tumors show higher staining of pEGFR and downstream effectors compared to matched normal colorectal tissues.

  16. Breaking the stalemate: a prospective regulatory framework for unforseen research uses of human tissue samples and health information.

    PubMed

    Greely, H T

    1999-01-01

    The legal and ethical issues raised by new research uses of previously collected human tissues and health information are increasingly important to genetics research. This Article discusses and criticizes current positions on such uses, including the recent report of the National Bioethics Advisory Commission, Research Involving Human Biological Materials. It then proposes a new regulatory framework for tissue and information collected in the future that would better protect the interests of the people who provide them. It ends by suggesting a resolution for the problems of previously collected tissue and information.

  17. High-resolution MALDI-FT-ICR MS imaging for the analysis of metabolites from formalin-fixed, paraffin-embedded clinical tissue samples.

    PubMed

    Buck, Achim; Ly, Alice; Balluff, Benjamin; Sun, Na; Gorzolka, Karin; Feuchtinger, Annette; Janssen, Klaus-Peter; Kuppen, Peter J K; van de Velde, Cornelis J H; Weirich, Gregor; Erlmeier, Franziska; Langer, Rupert; Aubele, Michaela; Zitzelsberger, Horst; Aichler, Michaela; Walch, Axel

    2015-09-01

    We present the first analytical approach to demonstrate the in situ imaging of metabolites from formalin-fixed, paraffin-embedded (FFPE) human tissue samples. Using high-resolution matrix-assisted laser desorption/ionization Fourier-transform ion cyclotron resonance mass spectrometry imaging (MALDI-FT-ICR MSI), we conducted a proof-of-principle experiment comparing metabolite measurements from FFPE and fresh frozen tissue sections, and found an overlap of 72% amongst 1700 m/z species. In particular, we observed conservation of biomedically relevant information at the metabolite level in FFPE tissues. In biomedical applications, we analysed tissues from 350 different cancer patients and were able to discriminate between normal and tumour tissues, and different tumours from the same organ, and found an independent prognostic factor for patient survival. This study demonstrates the ability to measure metabolites in FFPE tissues using MALDI-FT-ICR MSI, which can then be assigned to histology and clinical parameters. Our approach is a major technical, histochemical, and clinicopathological advance that highlights the potential for investigating diseases in archived FFPE tissues. PMID:25965788

  18. Histopathology mapping of biochemical changes in myocardial infarction by Fourier transform infrared spectral imaging.

    PubMed

    Yang, Tian T; Weng, Shi F; Zheng, Na; Pan, Qing H; Cao, Hong L; Liu, Liang; Zhang, Hai D; Mu, Da W

    2011-04-15

    Fourier transform infrared (FTIR) imaging and microspectroscopy have been extensively applied in the identification and investigation of both healthy and diseased tissues. FTIR imaging can be used to determine the biodistribution of several molecules of interest (carbohydrates, lipids, proteins) for tissue analysis, without the need for prior staining of these tissues. Molecular structure data, such as protein secondary structure and collagen triple helix exhibits, can also be obtained from the same analysis. Thus, several histopathological lesions, for example myocardial infarction, can be identified from FTIR-analyzed tissue images, the latter which can allow for more accurate discrimination between healthy tissues and pathological lesions. Accordingly, we propose FTIR imaging as a new tool integrating both molecular and histopathological assessment to investigate the degree of pathological changes in tissues. In this study, myocardial infarction is presented as an illustrative example of the wide potential of FTIR imaging for biomedical applications.

  19. Analytical procedure for mapping the distribution of 10B and 99Tc markers in cryo-sections of animal tissue samples by secondary ion mass spectrometry

    NASA Astrophysics Data System (ADS)

    Marchetti, Ilaria; Menichetti, Luca; Kusmic, Claudia; de las Heras, Laura Aldave; Salvadori, Piero; Fuoco, Roger; Belloni, Fabio; L'Abbate, Antonio; Betti, Maria

    2009-09-01

    The development of a complete, standard analytical procedure for a quantitative use of secondary ion mass spectrometry to map the distribution in animal tissues of exogenous isotopes presents difficulties inherently related to sample preparation and preservation, as well as to the specific application being considered. We have tested in two very different cases a procedure based on the cryo-preparation of samples and calibration standards. The applications under investigation were the mapping of 10B in mouse brain tissue, with relevance to the boron neutron capture therapy, and of the perfusion tracer 99Tc in mouse heart tissue, with relevance to the study of microcirculation and cardiovascular pathologies. Scanning electron microscopy and inductively coupled mass spectrometry analysis were used as reference techniques for secondary ion mass spectrometry images and analyte measurements, respectively. Cryo-preparation of tissue sections for ion microscopy proved to be simple and efficient (in terms of structural and chemical integrity) for both brain and heart samples derived from fresh organs. This technique, however, turned out to be reliable only on the brain tissue when applied to the preparation of standards, which required chemical fixation of portions of organs. Brain and heart tissues showed a totally different response to chemical fixation, from both a structural and an analytical point of view. On the one hand, we were able to estimate a relative sensitivity factor for 10B in the cryo-sectioned brain matrix; on the other hand, even without the possibility of an absolute quantification of the 99Tc signal and notwithstanding the presence of an isobaric interference, secondary ion mass spectrometry mapping however proved to be capable to resolve the specific response of the cardiac tissue to the perfusion mechanism.

  20. Histopathological Image Classification Using Discriminative Feature-Oriented Dictionary Learning.

    PubMed

    Vu, Tiep Huu; Mousavi, Hojjat Seyed; Monga, Vishal; Rao, Ganesh; Rao, U K Arvind

    2016-03-01

    In histopathological image analysis, feature extraction for classification is a challenging task due to the diversity of histology features suitable for each problem as well as presence of rich geometrical structures. In this paper, we propose an automatic feature discovery framework via learning class-specific dictionaries and present a low-complexity method for classification and disease grading in histopathology. Essentially, our Discriminative Feature-oriented Dictionary Learning (DFDL) method learns class-specific dictionaries such that under a sparsity constraint, the learned dictionaries allow representing a new image sample parsimoniously via the dictionary corresponding to the class identity of the sample. At the same time, the dictionary is designed to be poorly capable of representing samples from other classes. Experiments on three challenging real-world image databases: 1) histopathological images of intraductal breast lesions, 2) mammalian kidney, lung and spleen images provided by the Animal Diagnostics Lab (ADL) at Pennsylvania State University, and 3) brain tumor images from The Cancer Genome Atlas (TCGA) database, reveal the merits of our proposal over state-of-the-art alternatives. Moreover, we demonstrate that DFDL exhibits a more graceful decay in classification accuracy against the number of training images which is highly desirable in practice where generous training is often not available.

  1. Histopathological effects of copper on selected epithelial tissues of snails

    SciTech Connect

    Wolmarans, C.T.; van Aardt, W.J.; Coetzee, J.

    1986-06-01

    The exposure of freshwater snails to copper sulfate has several harmful effects on the snails. These effects include several physiological disturbances as well as certain behavioral and histological changes. On the strength of these findings it was decided to investigate, by means of transmission electron microscopy, whether histological changes occur in the surface epithelium of the snail Bulinus tropicus after exposure to copper sulfate.

  2. Influence of parasite density and sample storage time on the reliability of Entamoeba histolytica-specific PCR from formalin-fixed and paraffin-embedded tissues.

    PubMed

    Frickmann, Hagen; Tenner-Racz, Klara; Eggert, Petra; Schwarz, Norbert G; Poppert, Sven; Tannich, Egbert; Hagen, Ralf M

    2013-12-01

    We report on the reliability of polymerase chain reaction (PCR) for the detection of Entamoeba histolytica from formalin-fixed, paraffin-embedded tissue in comparison with microscopy and have determined predictors that may influence PCR results. E. histolytica-specific and Entamoeba dispar-specific real-time PCR and microscopy from adjacent histologic sections were performed using a collection of formalin-fixed, paraffin-embedded tissue specimens obtained from patients with invasive amebiasis. Specimens had been collected during the previous 4 decades. Association of sample age, parasite density, and reliability of PCR was analyzed. E. histolytica PCR was positive in 20 of 34 biopsies (58.8%); 2 of these 20 were microscopically negative for amebae in neighboring tissue sections. PCR was negative in 9 samples with visible amebae in neighboring sections and in 5 samples without visible parasites in neighboring sections. PCR was negative in all specimens that were older than 3 decades. Low parasite counts and sample ages older than 20 years were predictors for false-negative PCR results. All samples were negative for E. dispar DNA. PCR is suitable for the detection of E. histolytica in formalin-fixed, paraffin-embedded tissue samples that are younger than 2 decades and that contain intermediate to high parasite numbers. Negative results in older samples were due to progressive degradation of DNA over time as indicated by control PCRs targeting the human 18S rRNA gene. Moreover, our findings support previous suggestions that only E. histolytica but not E. dispar is responsible for invasive amebiasis.

  3. Stain-free histopathology by programmable supercontinuum pulses

    NASA Astrophysics Data System (ADS)

    Tu, Haohua; Liu, Yuan; Turchinovich, Dmitry; Marjanovic, Marina; Lyngsø, Jens K.; Lægsgaard, Jesper; Chaney, Eric J.; Zhao, Youbo; You, Sixian; Wilson, William L.; Xu, Bingwei; Dantus, Marcos; Boppart, Stephen A.

    2016-08-01

    The preparation, staining, visualization and interpretation of histological images of tissue is well accepted as the gold standard process for the diagnosis of disease. These methods have a long history of development, and are used ubiquitously in pathology, despite being highly time- and labour-intensive. Here, we introduce a unique optical imaging platform and methodology for label-free multimodal multiphoton microscopy that uses a novel photonic-crystal fibre source to generate tailored chemical contrast based on programmable supercontinuum pulses. We demonstrate the collection of optical signatures of the tumour microenvironment, including evidence of mesoscopic biological organization, tumour cell migration and (lymph-) angiogenesis collected directly from fresh ex vivo mammary tissue. Acquisition of these optical signatures and other cellular or extracellular features, which are largely absent from histologically processed and stained tissue, combined with an adaptable platform for optical alignment-free programmable-contrast imaging, offers the potential to translate stain-free molecular histopathology into routine clinical use.

  4. Histopathologic lesions in sea otters exposed to crude oil

    SciTech Connect

    Lipscomb, T.P.; Harris, R.K.; Moeller, R.B.; Pletcher, J.M.; Haebler, R.J.

    1993-01-01

    On 24 March 1989, the oil tanker Exxon Valdez ran aground on Bligh Reef in Prince William Sound, Alaska. In the months following the spill, over 1,000 sea otters (Enhydra lutris) from oil spill-affected areas are known to have died. The purpose of the study is to identify and describe histopathologic lesions associated with crude oil exposure in sea otters and to discuss possible pathogeneses of the lesions. Materials available included tissues from oil-contaminated and uncontaminated otters that died in rehabilitation centers following the oil spill and tissues from otters that were found dead in the oil spill-affected area with external oil present. Tissues from apparently normal sea otters from an area not contaminated by crude oil, were also examined.

  5. Cellulite histopathology and related mechanobiology.

    PubMed

    Quatresooz, P; Xhauflaire-Uhoda, E; Piérard-Franchimont, C; Piérard, G E

    2006-06-01

    Cellulite, sometimes called gynoid lipodystrophy, is much more prevalent in women than in men. There are glaring discrepancies regarding the microanatomical descriptions of this condition in the literature. A lumpy aspect of the dermo-hypodermal interface is often cited, but it appears to represent a gender-linked characteristic of the thighs and buttocks without being a specific sign of cellulite. Incipient cellulite recognized by a discrete padded look or 'orange peel' aspect appears correlated with the presence of a network of focally enlarged fibrosclerotic strands partitioning the hypodermis and serving as a physiological buttress limiting the outpouching of fat lobules on pinching the skin. These connective tissue structures might represent a hormonal-dependent reactive process to sustained mechanical tensions caused by the adipocyte lobules. Full blown cellulite is recognized by a lumpy-bumpy and dimpled skin surface. It likely represents subjugation of the hypertrophic response of the hypodermal connective tissue strands when their resistance is overcome by progressive fat accumulation. In these cases, histological aspects reminiscent of striae distensae are identified within the hypodermal connective tissue strands. The mechanical properties of skin involved by cellulite process are altered, but may tend to resume to normal under treatment. These functional changes influence the mechanobiology of connective tissue cells, in particular the Factor XIIIa-positive dermal dendrocytes.

  6. Automated segmentation of the melanocytes in skin histopathological images.

    PubMed

    Lu, Cheng; Mahmood, Muhammad; Jha, Naresh; Mandal, Mrinal

    2013-03-01

    In the diagnosis of skin melanoma by analyzing histopathological images, the detection of the melanocytes in the epidermis area is an important step. However, the detection of melanocytes in the epidermis area is dicult because other keratinocytes that are very similar to the melanocytes are also present. This paper proposes a novel computer-aided technique for segmentation of the melanocytes in the skin histopathological images. In order to reduce the local intensity variant, a mean-shift algorithm is applied for the initial segmentation of the image. A local region recursive segmentation algorithm is then proposed to filter out the candidate nuclei regions based on the domain prior knowledge. To distinguish the melanocytes from other keratinocytes in the epidermis area, a novel descriptor, named local double ellipse descriptor (LDED), is proposed to measure the local features of the candidate regions. The LDED uses two parameters: region ellipticity and local pattern characteristics to distinguish the melanocytes from the candidate nuclei regions. Experimental results on 28 dierent histopathological images of skin tissue with dierent zooming factors show that the proposed technique provides a superior performance.

  7. Evaluation of a new immunodiagnostic assay for Helicobacter pylori antibody detection: correlation with histopathological and microbiological results.

    PubMed Central

    Pronovost, A D; Rose, S L; Pawlak, J W; Robin, H; Schneider, R

    1994-01-01

    Infection with Helicobacter pylori has been associated with the pathogenesis of chronic active gastritis and gastric and duodenal ulcer disease. Detection of immunoglobulin G antibodies to H. pylori offers a simple alternative to direct detection of the organism in biopsied tissue by culture or histopathological methods. A rapid flow-through membrane-based enzyme immunoassay for the detection of human immunoglobulin G antibodies to H. pylori has been developed and evaluated. Clinical evaluations were performed with 256 patient serum samples obtained from four clinical sites. Biopsy samples were obtained by endoscopic procedures at the same time as the serum samples, and were histopathologically and microbiologically categorized for the presence or absence of H. pylori. Sensitivity and specificity for this rapid enzyme immunoassay were 92 and 88%, respectively, compared directly with endoscopy results. After discordant results were resolved by a quantitative microwell enzyme-linked immunosorbent assay, the resulting sensitivity and specificity were 94 and > 99%, respectively. These results indicate that this rapid enzyme immunoassay is a useful technique to determine H. pylori infection status and is a viable alternative to invasive endoscopic procedures. PMID:8126203

  8. Some Physical, Chemical, and Biological Parameters of Samples of Scleractinium Coral Aquaculture Skeleton Used for Reconstruction/Engineering of the Bone Tissue.

    PubMed

    Popov, A A; Sergeeva, N S; Britaev, T A; Komlev, V S; Sviridova, I K; Kirsanova, V A; Akhmedova, S A; Dgebuadze, P Yu; Teterina, A Yu; Kuvshinova, E A; Schanskii, Ya D

    2015-08-01

    Physical and chemical (phase and chemical composition, dynamics of resorption, and strength properties), and biological (cytological compatibility and scaffold properties of the surface) properties of samples of scleractinium coral skeletons from aquacultures of three types and corresponding samples of natural coral skeletons (Pocillopora verrucosa, Acropora formosa, and Acropora nobilis) were studied. Samples of scleractinium coral aquaculture skeleton of A. nobilis, A. formosa, and P. verrucosa met the requirements (all study parameters) to materials for osteoplasty and 3D-scaffolds for engineering of bone tissue.

  9. Vitamin-K-Dependent Protection of the Renal Microvasculature: Histopathological Studies in Normal and Diseased Kidneys

    PubMed Central

    Wei, Fang-Fei; Drummen, Nadja E.A.; Thijs, Lutgarde; Jacobs, Lotte; Herfs, Marjolein; van't Hoofd, Cynthia; Vermeer, Cees; Staessen, Jan A.

    2016-01-01

    Vitamin-K-dependent carboxylation of matrix Gla protein (MGP) protects the macrocirculation against calcification. We recently reported in a multiethnic population study that the estimated glomerular filtration rate, a microvascular trait, decreased and the risk of chronic kidney disease increased with higher circulating levels of inactive dephospho-uncarboxylated MGP, a marker of vitamin K deficiency. These findings highlighted the possibility that vitamin K might have a beneficial effect on the renal microcirculation. To substantiate these epidemiological findings, we undertook a pilot study, in which we stained renal tissue samples obtained by biopsy from 2 healthy kidney donors and 4 patients with nephropathy for carboxylated and uncarboxylated MGP and calcium deposits. Three patients had renal calcifications, which were consistently associated with carboxylated and uncarboxylated MGP. Normal renal tissue was devoid of microcalcifications and staining for carboxylated and uncarboxylated MGP. Pending confirmation in a larger study covering a wider range of renal pathologies, these histopathological findings suggest that MGP might inhibit calcification not only in large arteries, as was known before, but in renal tissue as well, thereby highlighting potentially new avenues for promoting renal health, for instance by vitamin K supplementation. PMID:27752480

  10. Happenings in histopathology - a post-World War II perspective.

    PubMed

    Shanmugaratnam, K

    2007-08-01

    There have been several important developments in the practice of histopathology since World War II; those reviewed in this lecture are grouped under 4 headings: new techniques (cytopathology, electron microscopy, immunohistochemistry and molecular pathology), organisational issues (recruitment, training and certification, subspecialties, quality control and consultations), ethical and legal issues (service costs, and the ownership and uses of biopsy tissues) and globalisation (international associations, standardised classification and nomenclature, and telepathology). Advances in the fields of molecular pathology and telepathology are expected to have the greatest impact on the practice of pathology in the next decade. PMID:17767342

  11. Happenings in histopathology - a post-World War II perspective.

    PubMed

    Shanmugaratnam, K

    2007-08-01

    There have been several important developments in the practice of histopathology since World War II; those reviewed in this lecture are grouped under 4 headings: new techniques (cytopathology, electron microscopy, immunohistochemistry and molecular pathology), organisational issues (recruitment, training and certification, subspecialties, quality control and consultations), ethical and legal issues (service costs, and the ownership and uses of biopsy tissues) and globalisation (international associations, standardised classification and nomenclature, and telepathology). Advances in the fields of molecular pathology and telepathology are expected to have the greatest impact on the practice of pathology in the next decade.

  12. Histopathology of kidney of albino rat poisoned with uranyl nitrate

    SciTech Connect

    Goel, K.A.; Garg, V.K.; Garg, V.

    1980-01-01

    Heavy metals input into the media either terrestrial or aquatic is an important aspect of environmental pollution. Heavy metals are known to produce toxic effects on the different tissues of various terrestrial and aquatic animals. Some of these are highly toxic at even very low concentrations and they alter the cellular architecture of many organs including the kidney. Little has been done on the effect of rare earth metals, particularly that of uranium on the kidney of animals. In the present paper histopathological changes produced by uranium on the kidney of albino rats are discussed.

  13. Evaluation of sample preparation and chromatographic separation for the parallel determination of taurine and edaravone in rat tissues using HILIC-MS/MS.

    PubMed

    Li, Yin-jie; Li, Zheng; Zheng, Xiao-xiao; Wu, Xiao-wen; Wang, Shi-rui; Guo, Hao; Yu, Yan-yan; Guo, Meng-zhe; Yan, Dong-zhi; Tang, Dao-quan

    2015-05-01

    The quantitative analysis of taurine and edaravone in biological sample is critical in pharmaceutical studies. Although each of them can be individually analyzed by different approaches, concurrent quantification is still a highly challenging task with respect to their great polarity variation and the complex composition of tissue sample. In the present study, to simultaneously determine taurine and edaravone in rat tissue, the sample preparation and chromatographic separation conditions were evaluated and discussed in detail. As for the sample preparation, four kinds of solvent and the volume ratio of the optimal solvent to biological sample were both tested and evaluated based on the chromatographic profile, extraction recovery, and matrix effect (ME). The chromatographic separation was performed in a reverse phase (RP) and two hydrophilic interaction liquid chromatography (HILIC) modes, and the corresponding separation efficiencies were assessed using chromatographic parameters like half-width (W 1/2 ), tailing factor (f t), theoretical plates number (N), and ME. Furthermore, adopted composition of two mobile phase systems and the concentrations of the additives in the optimum buffer system were also investigated on an Atlantis HILIC silica column according to the resultant chromatographic profiles and peak areas of the analytes. The optimal results were obtained when the biological samples were deproteined by 4-fold volume of methanol/acetonitrile (1:3, v/v) and separated on a HILIC column with a gradient elution of acetonitrile/water containing 0.2 % formic acid and 10 mM ammonium formate. The proposed approach was validated and successfully applied to the parallel determination of the tissue distribution of edaravone and taurine in rat tissues.

  14. Fine mapping of tissue properties on excised samples of melanoma and skin without the need for histological staining.

    PubMed

    Tittmann, Bernhard R; Miyasaka, Chiaki; Maeva, Elena; Shum, David

    2013-02-01

    This paper develops a novel two-frequency approach for noninvasive evaluation of cancerous tissue with optimum depth and resolution. Frequencies of about 50 MHz are used in thickly sliced tissue to detect differences of the relative attenuation (C-scan mode scanning) with relatively limited resolution. Thus, suspect zones can be identified according to a quantitative criterion. These suspect zones are then selected for preparation of thin, transversal slices from within the original thick slices. Very-high-resolution (1-μm) visualization of cells is obtained at around 600 MHz on these transversal sections and adjacent sections are prepared for histological study in parallel. The technique's feasibility and potential are demonstrated on both normal and cancerous (melanoma) skin tissue. Isotropy of the specimens is experimentally verified to ensure that conditions were coherent for use of a 5-layer, angular spectrum model made to simulate longitudinal velocity, allowing estimation of longitudinal velocity from semiquantitative V(z) data.

  15. Virus isolation vs RT-PCR: which method is more successful in detecting VHSV and IHNV in fish tissue sampled under field conditions?

    PubMed

    Knüsel, R; Bergmann, S M; Einer-Jensen, K; Casey, J; Segner, H; Wahli, T

    2007-09-01

    This study compared the results of reverse transcription-polymerase chain reaction (RT-PCR) and traditional virus isolation on cell culture in detection of viral haemorrhagic septicaemia virus (VHSV) and infectious haematopoietic necrosis virus (IHNV). RT-PCR was used for 172 tissue sample pools (total of 859 fish) originating from a field survey on the occurrence of VHSV and IHNV in farmed and wild salmonids in Switzerland. These samples represented all sites with fish that were either identified as virus-positive by means of virus isolation (three sites, four positive tissue sample pools) and/or demonstrated positive anti-VHSV-antibody titres (83 sites, 121 positive blood samples) in a serum plaque neutralization test (SPNT). The RT-PCR technique confirmed the four VHSV-positive tissue sample pools detected by virus isolation and additionally identified one VHSV-positive sample that showed positive anti-VHSV-AB titres, but was negative in virus isolation. With IHNV, RT-PCR detected two positive samples not identified by virus isolation while in these fish the SPNT result had been questionable. One of the IHNV-positive samples represents the first detection of IHNV-RNA in wild brown trout in Switzerland. Compared to SPNT, the RT-PCR method detected, as with virus isolation, a much lower number of positive cases; reasons for this discrepancy are discussed. Our results indicate that RT-PCR can not only be successfully applied in field surveys, but may also be slightly more sensitive than virus isolation. However, in a titration experiment under laboratory conditions, the sensitivity of RT-PCR was not significantly higher when compared with virus isolation.

  16. Mercury concentrations in water and hybrid striped bass (Morone saxatilis × M. chrysops) muscle tissue samples collected from the Ohio River, USA.

    PubMed

    Emery, Erich B; Spaeth, John P

    2011-04-01

    We report on long-term aqueous mercury (Hg) measurements collected at fixed locations along the Ohio River, offer insights into patterns of water and fish tissue Hg levels, and calculate site-specific bioaccumulation factors (BAFs) along an extensive longitudinal basis. We examined the relationship between total recoverable Hg concentrations in water and fish samples collected from 12 locations on the mainstem Ohio River. Water samples were collected on a bimonthly basis from each location over a 6-year period preceding the collection of fish tissue samples. This abundance of data enabled us to calculate the long-term average aqueous Hg concentrations and approximate the lifetime aqueous Hg exposure experienced by fish, enabling the calculation of appropriate BAFs. Hybrid striped bass (HSB; Morone saxatilis × M. chrysops) were collected from the Ohio River, composited (three fish), and analyzed for Hg in muscle tissue from each location. Concentrations ranged from 0.2 to 0.4 mg/kg and 41.7% of all samples collected were higher than the US Environmental Protection Agency regulatory threshold of 0.3 mg Hg/kg wet weight. Hg levels generally increased with fish weight, length, and age. However, Hg concentration in the water was the strongest predictor of tissue concentrations. We found that both water and tissue concentrations increased with drainage area, albeit at different rates. This discrepancy in spatial patterns revealed that the bioaccumulation rate of methylmercury might not be consistent throughout the Ohio River mainstem. BAFs calculated at each location supported this finding, as values decreased with increasing drainage area. Our study serves to fill critical, previously identified data gaps and provides decision-makers with the information necessary to develop more appropriate BAF development and risk-management strategies.

  17. High-throughput quantum cascade laser (QCL) spectral histopathology: a practical approach towards clinical translation.

    PubMed

    Pilling, Michael J; Henderson, Alex; Bird, Benjamin; Brown, Mick D; Clarke, Noel W; Gardner, Peter

    2016-06-23

    Infrared microscopy has become one of the key techniques in the biomedical research field for interrogating tissue. In partnership with multivariate analysis and machine learning techniques, it has become widely accepted as a method that can distinguish between normal and cancerous tissue with both high sensitivity and high specificity. While spectral histopathology (SHP) is highly promising for improved clinical diagnosis, several practical barriers currently exist, which need to be addressed before successful implementation in the clinic. Sample throughput and speed of acquisition are key barriers and have been driven by the high volume of samples awaiting histopathological examination. FTIR chemical imaging utilising FPA technology is currently state-of-the-art for infrared chemical imaging, and recent advances in its technology have dramatically reduced acquisition times. Despite this, infrared microscopy measurements on a tissue microarray (TMA), often encompassing several million spectra, takes several hours to acquire. The problem lies with the vast quantities of data that FTIR collects; each pixel in a chemical image is derived from a full infrared spectrum, itself composed of thousands of individual data points. Furthermore, data management is quickly becoming a barrier to clinical translation and poses the question of how to store these incessantly growing data sets. Recently, doubts have been raised as to whether the full spectral range is actually required for accurate disease diagnosis using SHP. These studies suggest that once spectral biomarkers have been predetermined it may be possible to diagnose disease based on a limited number of discrete spectral features. In this current study, we explore the possibility of utilising discrete frequency chemical imaging for acquiring high-throughput, high-resolution chemical images. Utilising a quantum cascade laser imaging microscope with discrete frequency collection at key diagnostic wavelengths, we

  18. Distribution of 3-hydroxy iC17:0 in subgingival plaque and gingival tissue samples: relationship to adult periodontitis.

    PubMed

    Nichols, F C

    1994-09-01

    Gram-negative organisms incorporate hydroxy fatty acids into the lipid A moiety of lipopolysaccharide (LPS), and in the case of some members of the family Enterobacteriaceae, hydroxy fatty acids are incorporated exclusively into lipid A. However, a limited number of Bacteroides species have been shown to incorporate several classes of 3-hydroxy fatty acids, particularly 3-hydroxy iC17:0, into constitutive lipids as well as LPS. The present study examined the distribution of hydroxy fatty acids in two periodontal pathogens, Prevotella intermedia and Porphyromonas gingivalis, by employing a phospholipid extraction procedure (E. G. Bligh and W. J. Dyer, Can. J. Biochem. Physiol. 37:911-917, 1959) which partitioned constitutive lipids into the organic solvent phase and LPS into the aqueous phase. The distribution of hydroxy fatty acids within organic solvent and aqueous extracts of these bacterial species was then compared with the distribution in subgingival plaque samples isolated from either gingivitis or severe periodontitis sites as well as the distribution in gingival tissue samples. The organic solvent and aqueous extracts were hydrolyzed under strong alkaline conditions, and the free fatty acids were treated to form pentafluorobenzyl-ester, trimethylsilyl-ether derivatives. Hydroxy fatty acid levels were quantified by using gas chromatography-negative-ion chemical ionization-mass spectrometry. By using this approach, the mean values of the 3-hydroxy iC17:0 recovered within organic solvent extracts of P. gingivalis strains ranged from 56 to 63% of total 3-hydroxy iC17:0. Substantially less 3-hydroxy iC17:0 (< 5%) was recovered in organic solvent extracts of P. intermedia. By comparison, 75% of the 3-hydroxy iC17:0 in periodontitis subgingival plaque samples was recovered in organic solvent extracts, while only 43% of the 3-hydroxy iC17:0 in gingivitis plaque samples from the same patients was recovered in organic solvent extracts. However, 3-hydroxy iC17:0 was

  19. Collecting and Studying Blood and Tissue Samples From Patients With Locally Recurrent or Metastatic Prostate or Bladder/Urothelial Cancer

    ClinicalTrials.gov

    2016-06-06

    Healthy Control; Localized Urothelial Carcinoma of the Renal Pelvis and Ureter; Metastatic Malignant Neoplasm in the Bone; Metastatic Malignant Neoplasm in the Soft Tissues; Metastatic Urothelial Carcinoma of the Renal Pelvis and Ureter; Recurrent Bladder Carcinoma; Recurrent Prostate Carcinoma; Recurrent Urothelial Carcinoma of the Renal Pelvis and Ureter; Stage IV Bladder Cancer; Stage IV Bladder Urothelial Carcinoma; Stage IV Prostate Cancer

  20. Detection of Ki-ras mutations in tissue and plasma samples of patients with pancreatic cancer using PNA-mediated PCR clamping and hybridisation probes

    PubMed Central

    Däbritz, J; Hänfler, J; Preston, R; Stieler, J; Oettle, H

    2005-01-01

    In the present study, we combined the PCR-clamping approach with melting curve analysis using mutant specific hybridisation probes and wild-type specific peptide nucleic acids (PNAs) to determine the genotypes of the most frequent point mutation in codon 12 of the proto-oncogene Ki-ras in tissue and plasma samples of patients with pancreatic cancer. The sensitivity of our assay was 1–5 × 10−5. The melting curve analysis of tissue samples of four patients revealed two valine mutations, one none-valine mutation and one wild-type sequence. Ki-ras alterations were found in 28% of DNAs (18 out of 64) of nonrelated plasma samples of 10 patients with ductal adenocarcinoma of the pancreas. The valine mutation was the predominantly detected gene alteration (83%). Out of ten patients investigated, four patients (40%) became positive during clinical observation with respect to Ki-ras mutation. All four patients exhibited progressive disease and high levels of tumour marker CA 19-9. In conclusion, the one-step procedure discribed may be a useful clinical tool for analysing Ki-ras point mutations in tissue and plasmas samples. In addition, this method can be adapted for simultanous detection of multiple mutations and quantitation. PMID:15655549

  1. A strategy for extending the applicability of a validated plasma calibration curve to quantitative measurements in multiple tissue homogenate samples: a case study from a rat tissue distribution study of JI-101, a triple kinase inhibitor.

    PubMed

    Gurav, Sandip Dhondiram; Jeniffer, Sherine; Punde, Ravindra; Gilibili, Ravindranath Reddy; Giri, Sanjeev; Srinivas, Nuggehally R; Mullangi, Ramesh

    2012-04-01

    A general practice in bioanalysis is that, whatever the biological matrix the analyte is being quantified in, the validation is performed in the same matrix as per regulatory guidelines. In this paper, we are presenting the applicability of a validated LC-MS/MS method in rat plasma for JI-101, to estimate the concentrations of JI-101 in various tissues that were harvested in a rat tissue distribution study. A simple protein precipitation technique was used to extract JI-101 and internal standard from the tissue homogenates. The recovery of JI-101 in all the matrices was found to be >70%. Chromatographic separation was achieved using a binary gradient using mobile phase A (acetonitrile) and B (0.2% formic acid in water) at a flow rate of 0.30 mL/min on a Prodigy ODS column with a total run time of 4.0 min. The MS/MS ion transitions monitored were 466.1 → 265 for JI-101 and 180.1 → 110.1 for internal standard. The linearity range was 5.02-4017 ng/mL. The JI-101 levels were quantifiable in the various tissue samples harvested in this study. Therefore, the use of a previously validated JI-101 assay in plasma circumvented the tedious process of method development/validation in various tissue matrices.

  2. Microbiological and histopathological aspects of canine pyometra

    PubMed Central

    Coggan, Jennifer Anne; Melville, Priscilla Anne; de Oliveira, Clair Motos; Faustino, Marcelo; Moreno, Andréa Micke; Benites, Nilson Roberti

    2008-01-01

    As pyometra is recognized as one of the main causes of disease and death in the bitch the purposes of this study were to evaluate microbiological and histopathological aspects of canine pyometra and to research the virulence factors of the E. coli isolates identifying possible risks to human health. The microbiological isolation from the intrauterine contents of 100 dogs with pyometra was carried out and the virulence factors in the E. coli strains were identified using PCR method. This study also consisted of the counting of microorganisms colonies forming units in samples of intrauterine content, tests of antimicrobial susceptibility of the E. coli isolates and the histological examination of the uterus. E. coli was the most prevalent microorganism isolated (76.6%) and 120 strains (79.5%) were positive for sfa, 86 (56.9%) were positive for cnf, 87 (57.6%) were positive for pap, 52 (34.4%) were positive for hly, 51 (33.8%) were positive for iuc and 5 (3.3%) were positive for afa genes. One observed more sensitivity of E. coli to norfloxacin, polimixin B, sulphazotrin, chloranfenicol and enrofloxacin. In 42% of the samples of uterine walls where microorganisms were isolated, the sizes of the areas of the inflammatory responses corresponded to 39–56%. Virulence factors were identified in 98.0% of the strains evaluated, demonstrating a high frequency of potentially pathogenic E. coli. It must be considered that dogs are animals that are living in close proximity to man for thousands of years and have an important role in the transmission of E. coli to other animals and to man. PMID:24031249

  3. Plant tissue-based chemiluminescence flow biosensor for determination of unbound dopamine in rabbit blood with on-line microdialysis sampling.

    PubMed

    Li, Baoxin; Zhang, Zhujun; Jin, Yan

    2002-06-01

    A novel plant tissue-based chemiluminescence (CL) biosensor for dopamine combined with flow injection analysis is presented in this paper. The potato roots act as molecular recognition elements. Dopamine is oxidized by oxygen under the catalysis of polyphenol oxidase in the tissue column to produce hydrogen peroxide, which can react with luminol in the presence of peroxidase of potato tissue to generate CL signal. The CL emission intensity was linear with dopamine concentration in the range of 1x10(-5)-1x10(-7) g/ml and the detection limit was 5.3x10(-8) g/ml (3sigma) with a relative standard deviation of 1.7%. Combined with microdialysis sampling, the biosensor was applied to monitor the variation of dopamine level in the blood of rabbit after the administration of dopamine to demonstrate the favorable resolution and reliability of the system for in vivo on-line monitoring. PMID:11959481

  4. Application of non-lethal stable isotope analysis to assess feeding patterns of juvenile pallid sturgeon Scaphirhynchus albus: a comparison of tissue types and sample preservation methods

    USGS Publications Warehouse

    Andvik, R.T.; VanDeHey, J.A.; Fincel, M.J.; French, William E.; Bertrand, K.N.; Chipps, Steven R.; Klumb, R.A.; Graeb, B.D.S.

    2010-01-01

    Traditional techniques for stable isotope analysis (SIA) generally require sacrificing animals to collect tissue samples; this can be problematic when studying diets of endangered species such as the pallid sturgeon Scaphirhynchus albus. Our objectives were to (i) determine if pectoral fin tissue (non-lethal) could be a substitute for muscle tissue (lethal) in SIA of juvenile pallid sturgeon, and (ii) evaluate the influence of preservation techniques on stable isotope values. In the laboratory, individual juvenile pallid sturgeon were held for up to 186 day and fed chironomids, fish, or a commercially available pellet diet. Significant, positive relationships (r² ≥ 0.8) were observed between fin and muscle tissues for both δ15N and δ13C; in all samples isotopes were enriched in fins compared to muscle tissue. Chironomid and fish based diets of juvenile pallid sturgeon were distinguishable for fast growing fish (0.3 mm day−1) using stable δ15N and δ13C isotopes. Frozen and preserved fin tissue δ15N isotopes were strongly related (r2 = 0.89) but δ13C isotopes were weakly related (r2 = 0.16). Therefore, freezing is recommended for preservation of fin clips to avoid the confounding effect of enrichment by ethanol. This study demonstrates the utility of a non-lethal technique to assess time integrated food habits of juvenile pallid sturgeon and should be applicable to other threatened or endangered species.

  5. Needle optical coherence elastography for the measurement of microscale mechanical contrast deep within human breast tissues

    NASA Astrophysics Data System (ADS)

    Kennedy, Kelsey M.; McLaughlin, Robert A.; Kennedy, Brendan F.; Tien, Alan; Latham, Bruce; Saunders, Christobel M.; Sampson, David D.

    2013-12-01

    Optical coherence elastography (OCE) is an emerging imaging technique that probes microscale mechanical contrast in tissues with the potential to differentiate healthy and malignant tissues. However, conventional OCE techniques are limited to imaging the first 1 to 2 mm of tissue in depth. We demonstrate, for the first time, OCE measurements deep within human tissues using needle OCE, extending the potential of OCE as a surgical guidance tool. We use needle OCE to detect tissue interfaces based on mechanical contrast in both normal and malignant breast tissues in freshly excised human mastectomy samples, as validated against histopathology. Further, we demonstrate the feasibility of in situ measurements >4 cm from the tissue surface using ultrasound guidance of the OCE needle probe. With further refinement, our method may potentially aid in accurate detection of the boundary of the tumor to help ensure full removal of all malignant tissues, which is critical to the success of breast-conserving surgery.

  6. MALDI mass spectrometry imaging: A cutting-edge tool for fundamental and clinical histopathology.

    PubMed

    Longuespée, Rémi; Casadonte, Rita; Kriegsmann, Mark; Pottier, Charles; Picard de Muller, Gaël; Delvenne, Philippe; Kriegsmann, Jörg; De Pauw, Edwin

    2016-07-01

    Histopathological diagnoses have been done in the last century based on hematoxylin and eosin staining. These methods were complemented by histochemistry, electron microscopy, immunohistochemistry (IHC), and molecular techniques. Mass spectrometry (MS) methods allow the thorough examination of various biocompounds in extracts and tissue sections. Today, mass spectrometry imaging (MSI), and especially matrix-assisted laser desorption ionization (MALDI) imaging links classical histology and molecular analyses. Direct mapping is a major advantage of the combination of molecular profiling and imaging. MSI can be considered as a cutting edge approach for molecular detection of proteins, peptides, carbohydrates, lipids, and small molecules in tissues. This review covers the detection of various biomolecules in histopathological sections by MSI. Proteomic methods will be introduced into clinical histopathology within the next few years.

  7. Proteomics analysis of tissue samples from patients with squamous cell carcinoma of the penis and positive to human papillomavirus

    PubMed Central

    Koifman, Leandro; Ornellas, Paulo; Ornellas, Antonio Augusto; Pereira, Denise de Abreu; Zingali, Benedeta Russolina; Cavalcanti, Silvia Maria Baeta; Afonso, Larissa Alves; Sandim, Vanessa; Alves, Gilda

    2015-01-01

    ABSTRACT Purpose: The aim of this study was to identify possible protein biomarkers and/or candidates for therapeutic targets in tissues of patients with SCCP, infected by HPV, applying one dimensional electrophoresis (1DE), followed by direct mass spectrometry (MS) analysis. Materials and Methods: Tissues from 10 HPV positive patients with SCCP and from 10 patients with HPV negative non-tumorous penile foreskins were analyzed applying 1D electrophoresis, followed by analysis with direct mass spectrometry (MS). Results: Sixty-three different proteins were identified in the first group and 50 in the second group. Recognition was possible for 28 proteins exclusively detected in Group 1 and 21 proteins presented only in Group 2. Conclusion: Some proteins in the first group are directly involved in the development of other types of cancer, and therefore, suitable for analysis. Complement C3 protein is a strong candidate for evaluating SCCP patients. PMID:26401855

  8. Maintaining Breast Cancer Specimen Integrity and Individual or Simultaneous Extraction of Quality DNA, RNA, and Proteins from Allprotect-Stabilized and Nonstabilized Tissue Samples

    PubMed Central

    Carroll, Paul; Donatello, Simona; Connolly, Elizabeth; Griffin, Mairead; Dunne, Barbara; Burke, Louise; Flavin, Richard; Rizkalla, Hala; Ryan, Ciara; Hayes, Brian; D'Adhemar, Charles; Banville, Niamh; Faheem, Nazia; Muldoon, Cian; Gaffney, Eoin F.

    2011-01-01

    The Saint James's Hospital Biobank was established in 2008, to develop a high-quality breast tissue BioResource, as a part of the breast cancer clinical care pathway. The aims of this work were: (1) to ascertain the quality of RNA, DNA, and protein in biobanked carcinomas and normal breast tissues, (2) to assess the efficacy of AllPrep® (Qiagen) in isolating RNA, DNA, and protein simultaneously, (3) to compare AllPrep with RNEasy® and QIAamp® (both Qiagen), and (4) to examine the effectiveness of Allprotect® (Qiagen), a new tissue stabilization medium in preserving DNA, RNA, and proteins. One hundred eleven frozen samples of carcinoma and normal breast tissue were analyzed. Tumor and normal tissue morphology were confirmed by frozen sections. Tissue type, tissue treatment (Allprotect vs. no Allprotect), extraction kit, and nucleic acid quantification were analyzed by utilizing a 4 factorial design (SPSS PASW 18 Statistics Software®). QIAamp (DNA isolation), AllPrep (DNA, RNA, and Protein isolation), and RNeasy (RNA isolation) kits were assessed and compared. Mean DNA yield and A260/280 values using QIAamp were 33.2 ng/μL and 1.86, respectively, and using AllPrep were 23.2 ng/μL and 1.94. Mean RNA yield and RNA Integrity Number (RIN) values with RNeasy were 73.4 ng/μL and 8.16, respectively, and with AllPrep were 74.8 ng/μL and 7.92. Allprotect-treated tissues produced higher RIN values of borderline significance (P=0.055). No discernible loss of RNA stability was detected after 6 h incubation of stabilized or nonstabilized tissues at room temperature or 4°C or in 9 freeze-thaw cycles. Allprotect requires further detailed evaluation, but we consider AllPrep to be an excellent option for the simultaneous extraction of RNA, DNA, and protein from tumor and normal breast tissues. The essential presampling procedures that maintain the diagnostic integrity of pathology specimens do not appear to compromise the quality of molecular isolates. PMID

  9. Maintaining Breast Cancer Specimen Integrity and Individual or Simultaneous Extraction of Quality DNA, RNA, and Proteins from Allprotect-Stabilized and Nonstabilized Tissue Samples.

    PubMed

    Mee, Blanaid C; Carroll, Paul; Donatello, Simona; Connolly, Elizabeth; Griffin, Mairead; Dunne, Barbara; Burke, Louise; Flavin, Richard; Rizkalla, Hala; Ryan, Ciara; Hayes, Brian; D'Adhemar, Charles; Banville, Niamh; Faheem, Nazia; Muldoon, Cian; Gaffney, Eoin F

    2011-12-01

    The Saint James's Hospital Biobank was established in 2008, to develop a high-quality breast tissue BioResource, as a part of the breast cancer clinical care pathway. The aims of this work were: (1) to ascertain the quality of RNA, DNA, and protein in biobanked carcinomas and normal breast tissues, (2) to assess the efficacy of AllPrep(®) (Qiagen) in isolating RNA, DNA, and protein simultaneously, (3) to compare AllPrep with RNEasy(®) and QIAamp(®) (both Qiagen), and (4) to examine the effectiveness of Allprotect(®) (Qiagen), a new tissue stabilization medium in preserving DNA, RNA, and proteins. One hundred eleven frozen samples of carcinoma and normal breast tissue were analyzed. Tumor and normal tissue morphology were confirmed by frozen sections. Tissue type, tissue treatment (Allprotect vs. no Allprotect), extraction kit, and nucleic acid quantification were analyzed by utilizing a 4 factorial design (SPSS PASW 18 Statistics Software(®)). QIAamp (DNA isolation), AllPrep (DNA, RNA, and Protein isolation), and RNeasy (RNA isolation) kits were assessed and compared. Mean DNA yield and A(260/280) values using QIAamp were 33.2 ng/μL and 1.86, respectively, and using AllPrep were 23.2 ng/μL and 1.94. Mean RNA yield and RNA Integrity Number (RIN) values with RNeasy were 73.4 ng/μL and 8.16, respectively, and with AllPrep were 74.8 ng/μL and 7.92. Allprotect-treated tissues produced higher RIN values of borderline significance (P=0.055). No discernible loss of RNA stability was detected after 6 h incubation of stabilized or nonstabilized tissues at room temperature or 4°C or in 9 freeze-thaw cycles. Allprotect requires further detailed evaluation, but we consider AllPrep to be an excellent option for the simultaneous extraction of RNA, DNA, and protein from tumor and normal breast tissues. The essential presampling procedures that maintain the diagnostic integrity of pathology specimens do not appear to compromise the quality of molecular isolates.

  10. Histopathological Study of Cyclosporine Pulmonary Toxicity in Rats

    PubMed Central

    Elshama, Said Said; EL-Kenawy, Ayman El-Meghawry; Osman, Hosam-Eldin Hussein

    2016-01-01

    Cyclosporine is considered one of the common worldwide immunosuppressive drugs that are used for allograft rejection prevention. However, articles that address adverse effects of cyclosporine use on the vital organs such as lung are still few. This study aims to investigate pulmonary toxic effect of cyclosporine in rats by assessment of pulmonary histopathological changes using light and electron microscope examination. Sixty male adult albino rats were divided into three groups; each group consists of twenty rats. The first received physiological saline while the second and third groups received 25 and 40 mg/kg/day of cyclosporine, respectively, by gastric gavage for forty-five days. Cyclosporine reduced the lung and body weight with shrinkage or pyknotic nucleus of pneumocyte type II, degeneration of alveoli and interalveolar septum beside microvilli on the alveolar surface, emphysema, inflammatory cellular infiltration, pulmonary blood vessels congestion, and increase of fibrous tissues in the interstitial tissues and around alveoli with negative Periodic Acid-Schiff staining. Prolonged use of cyclosporine induced pulmonary ultrastructural and histopathological changes with the lung and body weight reduction depending on its dose. PMID:26941796

  11. Histopathological Study of Cyclosporine Pulmonary Toxicity in Rats.

    PubMed

    Elshama, Said Said; El-Kenawy, Ayman El-Meghawry; Osman, Hosam-Eldin Hussein

    2016-01-01

    Cyclosporine is considered one of the common worldwide immunosuppressive drugs that are used for allograft rejection prevention. However, articles that address adverse effects of cyclosporine use on the vital organs such as lung are still few. This study aims to investigate pulmonary toxic effect of cyclosporine in rats by assessment of pulmonary histopathological changes using light and electron microscope examination. Sixty male adult albino rats were divided into three groups; each group consists of twenty rats. The first received physiological saline while the second and third groups received 25 and 40 mg/kg/day of cyclosporine, respectively, by gastric gavage for forty-five days. Cyclosporine reduced the lung and body weight with shrinkage or pyknotic nucleus of pneumocyte type II, degeneration of alveoli and interalveolar septum beside microvilli on the alveolar surface, emphysema, inflammatory cellular infiltration, pulmonary blood vessels congestion, and increase of fibrous tissues in the interstitial tissues and around alveoli with negative Periodic Acid-Schiff staining. Prolonged use of cyclosporine induced pulmonary ultrastructural and histopathological changes with the lung and body weight reduction depending on its dose. PMID:26941796

  12. Numerical and structural genomic aberrations are reliably detectable in tissue microarrays of formalin-fixed paraffin-embedded tumor samples by fluorescence in-situ hybridization.

    PubMed

    Horn, Heike; Bausinger, Julia; Staiger, Annette M; Sohn, Maximilian; Schmelter, Christopher; Gruber, Kim; Kalla, Claudia; Ott, M Michaela; Rosenwald, Andreas; Ott, German

    2014-01-01

    Few data are available regarding the reliability of fluorescence in-situ hybridization (FISH), especially for chromosomal deletions, in high-throughput settings using tissue microarrays (TMAs). We performed a comprehensive FISH study for the detection of chromosomal translocations and deletions in formalin-fixed and paraffin-embedded (FFPE) tumor specimens arranged in TMA format. We analyzed 46 B-cell lymphoma (B-NHL) specimens with known karyotypes for translocations of IGH-, BCL2-, BCL6- and MYC-genes. Locus-specific DNA probes were used for the detection of deletions in chromosome bands 6q21 and 9p21 in 62 follicular lymphomas (FL) and six malignant mesothelioma (MM) samples, respectively. To test for aberrant signals generated by truncation of nuclei following sectioning of FFPE tissue samples, cell line dilutions with 9p21-deletions were embedded into paraffin blocks. The overall TMA hybridization efficiency was 94%. FISH results regarding translocations matched karyotyping data in 93%. As for chromosomal deletions, sectioning artefacts occurred in 17% to 25% of cells, suggesting that the proportion of cells showing deletions should exceed 25% to be reliably detectable. In conclusion, FISH represents a robust tool for the detection of structural as well as numerical aberrations in FFPE tissue samples in a TMA-based high-throughput setting, when rigorous cut-off values and appropriate controls are maintained, and, of note, was superior to quantitative PCR approaches.

  13. Pre-Analytical Considerations for Successful Next-Generation Sequencing (NGS): Challenges and Opportunities for Formalin-Fixed and Paraffin-Embedded Tumor Tissue (FFPE) Samples

    PubMed Central

    Arreaza, Gladys; Qiu, Ping; Pang, Ling; Albright, Andrew; Hong, Lewis Z.; Marton, Matthew J.; Levitan, Diane

    2016-01-01

    In cancer drug discovery, it is important to investigate the genetic determinants of response or resistance to cancer therapy as well as factors that contribute to adverse events in the course of clinical trials. Despite the emergence of new technologies and the ability to measure more diverse analytes (e.g., circulating tumor cell (CTC), circulating tumor DNA (ctDNA), etc.), tumor tissue is still the most common and reliable source for biomarker investigation. Because of its worldwide use and ability to preserve samples for many decades at ambient temperature, formalin-fixed, paraffin-embedded tumor tissue (FFPE) is likely to be the preferred choice for tissue preservation in clinical practice for the foreseeable future. Multiple analyses are routinely performed on the same FFPE samples (such as Immunohistochemistry (IHC), in situ hybridization, RNAseq, DNAseq, TILseq, Methyl-Seq, etc.). Thus, specimen prioritization and optimization of the isolation of analytes is critical to ensure successful completion of each assay. FFPE is notorious for producing suboptimal DNA quality and low DNA yield. However, commercial vendors tend to request higher DNA sample mass than what is actually required for downstream assays, which restricts the breadth of biomarker work that can be performed. We evaluated multiple genomics service laboratories to assess the current state of NGS pre-analytical processing of FFPE. Significant differences in pre-analytical capabilities were observed. Key aspects are highlighted and recommendations are made to improve the current practice in translational research. PMID:27657050

  14. Development of an indirect immunofluorescence assay for diagnosis of bovine viral diarrhoea virus on ear notch tissue samples in cattle infected persistently.

    PubMed

    Bedeković, Tomislav; Lemo, Nina; Lojkić, Ivana; Cvetnić, Zeljko; Cač, Zeljko; Madić, Josip

    2011-12-01

    Bovine viral diarrhoea virus (BVDV) causes a disease that has a wide range of clinical symptoms in domestic and wild ruminants. It is a major problem in cattle and causes significant economic losses in the cattle industry. The virus infects bovines of all ages and causes both immunosuppression and reproductive, respiratory and digestive disorders. Cattle infected persistently, as a continuing source of the virus and the main factor in transmission of the disease between and among herds, are the main source of BVDV and a primary factor in the epidemiology of the disease. To determine whether a BVDV infection is persistent, two samples should be taken at 3-4 week intervals and tested for the virus antigen. Animal sera, whole blood, organ and ear notch tissue samples can be used for BVDV diagnosis. In ear notch tissue, viral antigen can be detected by an antigen enzyme-linked immunosorbent assay (antigen ELISA), immunohistochemistry (IHC) and reverse-transcription polymerase chain reaction (RT-PCR). This paper describes the development and implementation of an indirect immunofluorescence (IF) method using ear notch tissue samples for diagnosis of cattle infected persistently. Results obtained by this method show that IF is a good alternative to RT-PCR and antigen ELISA and can be a quick and accurate method in diagnosis of BVDV in cattle infected persistently with this virus.

  15. Histopathologic ear findings of syphilis: a temporal bone study.

    PubMed

    Hızlı, Ömer; Hızlı, Pelin; Kaya, Serdar; Monsanto, Rafael da Costa; Paparella, Michael M; Cureoglu, Sebahattin

    2016-09-01

    To the best of our knowledge, histopathologic studies of syphilitic ears have generally focused on hydropic changes; so far, no such studies have investigated peripheral vestibular otopathology using differential interference contrast microscopy, in patients with syphilis. For this study, we examined 13 human temporal bone samples from 8 patients with a history of syphilis. Using conventional light microscopy, we performed qualitative histopathologic assessment. In addition, using differential interference contrast microscopy, we performed type I and type II vestibular hair cell counts on each vestibular sense organ with minimal autolysis; in which the neuroepithelium was oriented perpendicular to the plane of section. We then compared vestibular hair cell densities (cells per 0.01 mm² surface area) in the syphilis group vs. the control group. In the syphilis group, we observed precipitate in the endolymphatic or perilymphatic spaces in 1 (7.7 %) of the samples and endolymphatic hydrops in eight (61.5 %) of the samples. Hydrops involved the cochlea (four samples) and/or saccule (four samples). In addition, the syphilis group experienced a significant loss of type II vestibular hair cells in the maculae of the utricle and saccule, and in the cristae of the lateral and posterior semicircular canals, as compared with the control group (P < 0.05). PMID:26573155

  16. Histopathologic ear findings of syphilis: a temporal bone study.

    PubMed

    Hızlı, Ömer; Hızlı, Pelin; Kaya, Serdar; Monsanto, Rafael da Costa; Paparella, Michael M; Cureoglu, Sebahattin

    2016-09-01

    To the best of our knowledge, histopathologic studies of syphilitic ears have generally focused on hydropic changes; so far, no such studies have investigated peripheral vestibular otopathology using differential interference contrast microscopy, in patients with syphilis. For this study, we examined 13 human temporal bone samples from 8 patients with a history of syphilis. Using conventional light microscopy, we performed qualitative histopathologic assessment. In addition, using differential interference contrast microscopy, we performed type I and type II vestibular hair cell counts on each vestibular sense organ with minimal autolysis; in which the neuroepithelium was oriented perpendicular to the plane of section. We then compared vestibular hair cell densities (cells per 0.01 mm² surface area) in the syphilis group vs. the control group. In the syphilis group, we observed precipitate in the endolymphatic or perilymphatic spaces in 1 (7.7 %) of the samples and endolymphatic hydrops in eight (61.5 %) of the samples. Hydrops involved the cochlea (four samples) and/or saccule (four samples). In addition, the syphilis group experienced a significant loss of type II vestibular hair cells in the maculae of the utricle and saccule, and in the cristae of the lateral and posterior semicircular canals, as compared with the control group (P < 0.05).

  17. Histopathology reveals correlative and unique phenotypes in a high-throughput mouse phenotyping screen.

    PubMed

    Adissu, Hibret A; Estabel, Jeanne; Sunter, David; Tuck, Elizabeth; Hooks, Yvette; Carragher, Damian M; Clarke, Kay; Karp, Natasha A; Newbigging, Susan; Jones, Nora; Morikawa, Lily; White, Jacqueline K; McKerlie, Colin

    2014-05-01

    The Mouse Genetics Project (MGP) at the Wellcome Trust Sanger Institute aims to generate and phenotype over 800 genetically modified mouse lines over the next 5 years to gain a better understanding of mammalian gene function and provide an invaluable resource to the scientific community for follow-up studies. Phenotyping includes the generation of a standardized biobank of paraffin-embedded tissues for each mouse line, but histopathology is not routinely performed. In collaboration with the Pathology Core of the Centre for Modeling Human Disease (CMHD) we report the utility of histopathology in a high-throughput primary phenotyping screen. Histopathology was assessed in an unbiased selection of 50 mouse lines with (n=30) or without (n=20) clinical phenotypes detected by the standard MGP primary phenotyping screen. Our findings revealed that histopathology added correlating morphological data in 19 of 30 lines (63.3%) in which the primary screen detected a phenotype. In addition, seven of the 50 lines (14%) presented significant histopathology findings that were not associated with or predicted by the standard primary screen. Three of these seven lines had no clinical phenotype detected by the standard primary screen. Incidental and strain-associated background lesions were present in all mutant lines with good concordance to wild-type controls. These findings demonstrate the complementary and unique contribution of histopathology to high-throughput primary phenotyping of mutant mice.

  18. Multi-scale occupancy approach to estimate Toxoplasma gondii prevalence and detection probability in tissues: an application and guide for field sampling.

    PubMed

    Elmore, Stacey A; Huyvaert, Kathryn P; Bailey, Larissa L; Iqbal, Asma; Su, Chunlei; Dixon, Brent R; Alisauskas, Ray T; Gajadhar, Alvin A; Jenkins, Emily J

    2016-08-01

    Increasingly, birds are recognised as important hosts for the ubiquitous parasite Toxoplasma gondii, although little experimental evidence exists to determine which tissues should be tested to maximise the detection probability of T. gondii. Also, Arctic-nesting geese are suspected to be important sources of T. gondii in terrestrial Arctic ecosystems, but the parasite has not previously been reported in the tissues of these geese. Using a domestic goose model, we applied a multi-scale occupancy framework to demonstrate that the probability of detection of T. gondii was highest in the brain (0.689, 95% confidence interval=0.486, 0.839) and the heart (0.809, 95% confidence interval=0.693, 0.888). Inoculated geese had an estimated T. gondii infection probability of 0.849, (95% confidence interval=0.643, 0.946), highlighting uncertainty in the system, even under experimental conditions. Guided by these results, we tested the brains and hearts of wild Ross's Geese (Chen rossii, n=50) and Lesser Snow Geese (Chen caerulescens, n=50) from Karrak Lake, Nunavut, Canada. We detected 51 suspected positive tissue samples from 33 wild geese using real-time PCR with melt-curve analysis. The wild goose prevalence estimates generated by our multi-scale occupancy analysis were higher than the naïve estimates of prevalence, indicating that multiple PCR repetitions on the same organs and testing more than one organ could improve T. gondii detection. Genetic characterisation revealed Type III T. gondii alleles in six wild geese and Sarcocystis spp. in 25 samples. Our study demonstrates that Arctic nesting geese are capable of harbouring T. gondii in their tissues and could transport the parasite from their southern overwintering grounds into the Arctic region. We demonstrate how a multi-scale occupancy framework can be used in a domestic animal model to guide resource-limited sample collection and tissue analysis in wildlife. Secondly, we confirm the value of traditional occupancy in

  19. Multi-scale occupancy approach to estimate Toxoplasma gondii prevalence and detection probability in tissues: an application and guide for field sampling.

    PubMed

    Elmore, Stacey A; Huyvaert, Kathryn P; Bailey, Larissa L; Iqbal, Asma; Su, Chunlei; Dixon, Brent R; Alisauskas, Ray T; Gajadhar, Alvin A; Jenkins, Emily J

    2016-08-01

    Increasingly, birds are recognised as important hosts for the ubiquitous parasite Toxoplasma gondii, although little experimental evidence exists to determine which tissues should be tested to maximise the detection probability of T. gondii. Also, Arctic-nesting geese are suspected to be important sources of T. gondii in terrestrial Arctic ecosystems, but the parasite has not previously been reported in the tissues of these geese. Using a domestic goose model, we applied a multi-scale occupancy framework to demonstrate that the probability of detection of T. gondii was highest in the brain (0.689, 95% confidence interval=0.486, 0.839) and the heart (0.809, 95% confidence interval=0.693, 0.888). Inoculated geese had an estimated T. gondii infection probability of 0.849, (95% confidence interval=0.643, 0.946), highlighting uncertainty in the system, even under experimental conditions. Guided by these results, we tested the brains and hearts of wild Ross's Geese (Chen rossii, n=50) and Lesser Snow Geese (Chen caerulescens, n=50) from Karrak Lake, Nunavut, Canada. We detected 51 suspected positive tissue samples from 33 wild geese using real-time PCR with melt-curve analysis. The wild goose prevalence estimates generated by our multi-scale occupancy analysis were higher than the naïve estimates of prevalence, indicating that multiple PCR repetitions on the same organs and testing more than one organ could improve T. gondii detection. Genetic characterisation revealed Type III T. gondii alleles in six wild geese and Sarcocystis spp. in 25 samples. Our study demonstrates that Arctic nesting geese are capable of harbouring T. gondii in their tissues and could transport the parasite from their southern overwintering grounds into the Arctic region. We demonstrate how a multi-scale occupancy framework can be used in a domestic animal model to guide resource-limited sample collection and tissue analysis in wildlife. Secondly, we confirm the value of traditional occupancy in

  20. Genotyping of human papillomavirus in paraffin embedded cervical tissue samples from women in Ethiopia and the Sudan.

    PubMed

    Abate, Ebba; Aseffa, Abraham; El-Tayeb, Muntasir; El-Hassan, Ibrahim; Yamuah, Lawrence; Mihret, Wude; Bekele, Liku; Ashenafi, Senait; El-Dawi, Nadia; Belayneh, Meseret; El-Hassan, Ahmed; Engers, Howard

    2013-02-01

    Cervical cancer is the most frequent female malignancy in most developing countries. Previous studies have demonstrated a strong association of human papillomavirus (HPV) infection with dysplasia and carcinoma of the uterine cervix. The objective of this study was to identify the prevailing HPV genotypes responsible for the development of cervical cancer among women in Ethiopia and the Sudan. A molecular characterization of HPV was done on 245 paraffin embedded cervical biopsy samples collected from the two countries. Amplification of HPV and subsequent genotyping was done using SPF10 primers and Line probe assay. Of samples collected from Ethiopian patients, 93% (149/160) and 13% (21/160) had high risk and low risk HPV genotypes, respectively. Among samples collected from the Sudan, 94% (80/85) harbored high risk and 11.7% (10/85) low risk HPV genotypes. Human papillomavirus 16 was the most frequent genotype identified in samples from Ethiopia (91%, 136/149) and the Sudan (82.5%, 66/80). HPV 52, 58, and 18 were the second, third and fourth common genotypes identified in Ethiopia, whereas HPV 18, 45, and 52 were the second, third, and fourth genotypes identified in samples collected from the Sudan. Thus, individuals living in different geographical localities should receive vaccines based on the specific genotypes circulating in the area and a vaccine targeting HPV 16, 18, 45, 52, and 58 may be optimal for the control of cervical cancer in the two countries.

  1. Interpreting histopathology in the epididymis

    PubMed Central

    De Grava Kempinas, Wilma; Klinefelter, Gary Robert

    2014-01-01

    While most of this Special Issue is devoted to the testis (which is where most drug and chemically induced toxicity of the male reproductive tract is identified), being able to recognize and understand the potential effects of toxicants on the epididymis is immensely important and an area that is often overlooked. The epididymis is the organ where the post-testicular sperm differentiation occurs, through a complex and still not completely understood sperm maturation process, allowing them to fertilize the oocyte. Also in the epididymis, sperm are stored until ejaculation, while being protected from immunogenic reaction by a blood-epididymis barrier. From a toxicologic perspective the epididymis is inherently complicated as its structure and function can be altered both indirectly and directly. In this review we will discuss the factors that must be considered when attempting to distinguish between indirect and direct epididymal toxicity and highlight what is currently known about mechanisms of epididymal toxicants, using the rat as a reference model. We identify 2 distinguishable signature lesions – one representing androgen deprivation (secondary to Leydig cell toxicity in the testis) and another representing a direct acting toxicant. Other commonly observed alterations will also be shown and discussed. Finally, we point out that many of the key functions of the epididymis can be altered in the absence of a detectable change in tissue structure. Collectively, we hope this will provide pathologists with increased confidence in identification of epididymal toxicity and enable more informed guidance as mechanism of action is considered. PMID:26413396

  2. High-throughput RNA sequencing of a formalin-fixed, paraffin-embedded autopsy lung tissue sample from the 1918 influenza pandemic

    PubMed Central

    Xiao, Yong-Li; Kash, John C; Beres, Stephen B; Sheng, Zong-Mei; Musser, James M; Taubenberger, Jeffery K

    2013-01-01

    Most biopsy and autopsy tissues are formalin-fixed and paraffin-embedded (FFPE), but this process leads to RNA degradation that limits gene expression analysis. The RNA genome of the 1918 pandemic influenza virus was previously determined in a 9-year effort by overlapping RT-PCR from post-mortem samples. Here, the full genome of the 1918 virus at 3000× coverage was determined in one high-throughput sequencing run of a library derived from total RNA of a 1918 FFPE sample after duplex-specific nuclease treatments. Bacterial sequences associated with secondary bacterial pneumonias were also detected. Host transcripts were well represented in the library. Compared to a 2009 pandemic influenza virus FFPE post-mortem library, the 1918 sample showed significant enrichment for host defence and cell death response genes, concordant with prior animal studies. This methodological approach should assist in the analysis of FFPE tissue samples isolated over the past century from a variety of diseases. PMID:23180419

  3. Evaluation of a digestion assay and determination of sample size and tissue for the reliable detection of Trichinella larvae in walrus meat.

    PubMed

    Leclair, Daniel; Forbes, Lorry B; Suppa, Sandy; Gajadhar, Alvin A

    2003-03-01

    A digestion assay was validated for the detection of Trichinella larvae in walrus (Odobenus rosmarus) meat, and appropriate samples for testing were determined using tissues from infected walruses harvested for food. Examination of muscles from 3 walruses showed that the tongue consistently contained approximately 2-6 times more larvae than the pectoral and intercostal muscles. Comparison of numbers of larvae in the root, body, and apex of the tongue from 3 walruses failed to identify a predilection site within the tongue, but the apex was considered an optimal tissue because of the high larval density within the tongue and the ease of collection. All 31 spiked samples weighing 50 g each and containing between 0.1 and 0.4 larvae per gram (lpg) were correctly identified as infected, indicating that the sensitivity of this procedure is adequate for diagnostic use. A sample size of 10 g consistently detected larvae in 2 walrus tongues containing > or = 0.3 lpg (n = 40), and until additional data are available, sample sizes from individual walrus tongues should be a minimum of 10 g. This study provides the preliminary data that were used for the development of a food safety analytical protocol for the detection of Trichinella in walrus meat in arctic communities.

  4. Genetic Characterization of Echinococcus granulosus from a Large Number of Formalin-Fixed, Paraffin-Embedded Tissue Samples of Human Isolates in Iran

    PubMed Central

    Rostami, Sima; Torbaghan, Shams Shariat; Dabiri, Shahriar; Babaei, Zahra; Mohammadi, Mohammad Ali; Sharbatkhori, Mitra; Harandi, Majid Fasihi

    2015-01-01

    Cystic echinococcosis (CE), caused by the larval stage of Echinococcus granulosus, presents an important medical and veterinary problem globally, including that in Iran. Different genotypes of E. granulosus have been reported from human isolates worldwide. This study identifies the genotype of the parasite responsible for human hydatidosis in three provinces of Iran using formalin-fixed paraffin-embedded tissue samples. In this study, 200 formalin-fixed paraffin-embedded tissue samples from human CE cases were collected from Alborz, Tehran, and Kerman provinces. Polymerase chain reaction amplification and sequencing of the partial mitochondrial cytochrome c oxidase subunit 1 gene were performed for genetic characterization of the samples. Phylogenetic analysis of the isolates from this study and reference sequences of different genotypes was done using a maximum likelihood method. In total, 54.4%, 0.8%, 1%, and 40.8% of the samples were identified as the G1, G2, G3, and G6 genotypes, respectively. The findings of the current study confirm the G1 genotype (sheep strain) to be the most prevalent genotype involved in human CE cases in Iran and indicates the high prevalence of the G6 genotype with a high infectivity for humans. Furthermore, this study illustrates the first documented human CE case in Iran infected with the G2 genotype. PMID:25535316

  5. High resolution Physio-chemical Tissue Analysis: Towards Non-invasive In Vivo Biopsy

    PubMed Central

    Xu, Guan; Meng, Zhuo-xian; Lin, Jian-die; Deng, Cheri X.; Carson, Paul L.; Fowlkes, J. Brian; Tao, Chao; Liu, Xiaojun; Wang, Xueding

    2016-01-01

    Conventional gold standard histopathologic diagnosis requires information of both high resolution structural and chemical changes in tissue. Providing optical information at ultrasonic resolution, photoacoustic (PA) technique could provide highly sensitive and highly accurate tissue characterization noninvasively in the authentic in vivo environment, offering a replacement for histopathology. A two-dimensional (2D) physio-chemical spectrogram (PCS) combining micrometer to centimeter morphology and chemical composition simultaneously can be generated for each biological sample with PA measurements at multiple optical wavelengths. This spectrogram presents a unique 2D “physio-chemical signature” for any specific type of tissue. Comprehensive analysis of PCS, termed PA physio-chemical analysis (PAPCA), can lead to very rich diagnostic information, including the contents of all relevant molecular and chemical components along with their corresponding histological microfeatures, comparable to those accessible by conventional histology. PAPCA could contribute to the diagnosis of many diseases involving diffusive patterns such as fatty liver. PMID:26842459

  6. High resolution Physio-chemical Tissue Analysis: Towards Non-invasive In Vivo Biopsy

    NASA Astrophysics Data System (ADS)

    Xu, Guan; Meng, Zhuo-Xian; Lin, Jian-Die; Deng, Cheri X.; Carson, Paul L.; Fowlkes, J. Brian; Tao, Chao; Liu, Xiaojun; Wang, Xueding

    2016-02-01

    Conventional gold standard histopathologic diagnosis requires information of both high resolution structural and chemical changes in tissue. Providing optical information at ultrasonic resolution, photoacoustic (PA) technique could provide highly sensitive and highly accurate tissue characterization noninvasively in the authentic in vivo environment, offering a replacement for histopathology. A two-dimensional (2D) physio-chemical spectrogram (PCS) combining micrometer to centimeter morphology and chemical composition simultaneously can be generated for each biological sample with PA measurements at multiple optical wavelengths. This spectrogram presents a unique 2D “physio-chemical signature” for any specific type of tissue. Comprehensive analysis of PCS, termed PA physio-chemical analysis (PAPCA), can lead to very rich diagnostic information, including the contents of all relevant molecular and chemical components along with their corresponding histological microfeatures, comparable to those accessible by conventional histology. PAPCA could contribute to the diagnosis of many diseases involving diffusive patterns such as fatty liver.

  7. Automated Liquid Microjunction Surface Sampling-HPLC-MS/MS Analysis of Drugs and Metabolites in Whole-Body Thin Tissue Sections

    SciTech Connect

    Kertesz, Vilmos; Van Berkel, Gary J

    2013-01-01

    A fully automated liquid extraction-based surface sampling system utilizing a commercially available autosampler coupled to high performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) detection is reported. Discrete spots selected for droplet-based sampling and automated sample queue generation for both the autosampler and MS were enabled by using in-house developed software. In addition, co-registration of spatially resolved sampling position and HPLC-MS information to generate heatmaps of compounds monitored for subsequent data analysis was also available in the software. The system was evaluated with whole-body thin tissue sections from propranolol dosed rat. The hands-free operation of the system was demonstrated by creating heatmaps of the parent drug and its hydroxypropranolol glucuronide metabolites with 1 mm resolution in the areas of interest. The sample throughput was approximately 5 min/sample defined by the time needed for chromatographic separation. The spatial distributions of both the drug and its metabolites were consistent with previous studies employing other liquid extraction-based surface sampling methodologies.

  8. A hardware approach for histological and histopathological digital image stain normalization.

    PubMed

    Şerbănescu, Mircea Sebastian; Pleşea, Iancu Emil

    2015-01-01

    Advances in technology made the migration of pathological diagnosis to digital slides possible. As the need for objectivity and automation emerged, new computer software algorithms were proposed. Computer algorithms demand accurate color and intensity values in order to provide reliable results. The tissue samples undergo several processing steps from histological preparation to digitalization, which cannot be completely standardized. Thus, non-standardized input data generates unreliable output data. In this article, we discuss a new computational normalization algorithm for histopathological stained slides that uses a hardware color marker. The marker is added to the glass slide together with the tissue section, exposed to all the processing steps and altered in the same manner as the biological material of interest, thus becoming a solid color marker for image normalization. The results of the proposed method are numerically and perceptually tested in order to prove the advantages of the method. We conclude that our combined hardware-software technique for staining normalization of digital slides is superior to the existing methods based on only software normalization, and that its implementation will tackle not only the acquisition errors but also the technical errors that may occur during the staining process.

  9. Histopathological retrospective study of canine renal disease in Korea, 2003~2008

    PubMed Central

    Yhee, Ji-Young; Yu, Chi-Ho; Kim, Jong-Hyuk; Im, Keum-Soon; Chon, Seung-Ki

    2010-01-01

    Renal disease includes conditions affecting the glomeruli, tubules, interstitium, pelvis, and vasculature. Diseases of the kidney include glomerular diseases, diseases of the tubules and interstitium, diseases of renal pelvis, and developmental abnormalities. Renal tissue samples (n = 70) submitted to the Department of Veterinary Pathology of Konkuk University from 2003 to 2008 were included in this study. Tissue histopathology was performed using light microscopy with hematoxylin and eosin stains. Masson's trichrome, Congo Red, and Warthin starry silver staining were applied in several individual cases. Glomerular diseases (22.9%), tubulointerstitial diseases (8.6%), neoplastic diseases (8.6%), conditions secondary to urinary obstruction (24.3%), and other diseases (35.7%) were identified. Glomerulonephritis (GN) cases were classified as acute proliferative GN (5.7%), membranous GN (4.3%), membranoproliferative GN (4.3%), focal segmental GN (2.9%), and other GN (4.2%). The proportion of canine GN cases presently identified was not as high as the proportions identified in human studies. Conversely, urinary obstruction and end-stage renal disease cases were relatively higher in dogs than in human populations. PMID:21113095

  10. Histopathological analysis of unilateral condylar hyperplasia: difficulties in diagnosis and characterization of the disease.

    PubMed

    Vásquez, B; Olate, S; Cantín, M; Sandoval, C; Fariña, R; Del Sol, M

    2016-05-01

    The aim of this study was to perform a histological analysis of the conditions in patients undergoing surgery for unilateral condylar hyperplasia (UCH) using different histopathological analytical techniques and to describe the complications and existing controversy in order to better define the disease. A partial condylectomy was performed in five patients who had been diagnosed with UCH using clinical and imaging methods. The samples obtained were analyzed using routine histological techniques including haematoxylin-eosin, Van Gieson, picrosirius, alcian blue/haematoxylin-eosin, and AgNOR staining. The analyses were performed by an observer who was blinded to the clinical parameters of the disease. The cellularity, tissue layers, size of the anatomical structures, and the relationships between them were assessed. The analysis of these patients was complemented by a review of the scientific literature. Variability was observed in the analysis of the cases. The presence of connective tissue was detected at the bone level, with cartilage formation at different levels. Each island presented levels of involvement that could indicate various degrees of aggressiveness. Type I collagen was observed in most cases, although type III was also identified. The development of histological diagnostic methods to determine the aggressiveness or level of involvement in UCH is not currently possible. Further studies are needed to establish new histological classifications. PMID:26723497

  11. A hardware approach for histological and histopathological digital image stain normalization.

    PubMed

    Şerbănescu, Mircea Sebastian; Pleşea, Iancu Emil

    2015-01-01

    Advances in technology made the migration of pathological diagnosis to digital slides possible. As the need for objectivity and automation emerged, new computer software algorithms were proposed. Computer algorithms demand accurate color and intensity values in order to provide reliable results. The tissue samples undergo several processing steps from histological preparation to digitalization, which cannot be completely standardized. Thus, non-standardized input data generates unreliable output data. In this article, we discuss a new computational normalization algorithm for histopathological stained slides that uses a hardware color marker. The marker is added to the glass slide together with the tissue section, exposed to all the processing steps and altered in the same manner as the biological material of interest, thus becoming a solid color marker for image normalization. The results of the proposed method are numerically and perceptually tested in order to prove the advantages of the method. We conclude that our combined hardware-software technique for staining normalization of digital slides is superior to the existing methods based on only software normalization, and that its implementation will tackle not only the acquisition errors but also the technical errors that may occur during the staining process. PMID:26429166

  12. Histopathological analysis of unilateral condylar hyperplasia: difficulties in diagnosis and characterization of the disease.

    PubMed

    Vásquez, B; Olate, S; Cantín, M; Sandoval, C; Fariña, R; Del Sol, M

    2016-05-01

    The aim of this study was to perform a histological analysis of the conditions in patients undergoing surgery for unilateral condylar hyperplasia (UCH) using different histopathological analytical techniques and to describe the complications and existing controversy in order to better define the disease. A partial condylectomy was performed in five patients who had been diagnosed with UCH using clinical and imaging methods. The samples obtained were analyzed using routine histological techniques including haematoxylin-eosin, Van Gieson, picrosirius, alcian blue/haematoxylin-eosin, and AgNOR staining. The analyses were performed by an observer who was blinded to the clinical parameters of the disease. The cellularity, tissue layers, size of the anatomical structures, and the relationships between them were assessed. The analysis of these patients was complemented by a review of the scientific literature. Variability was observed in the analysis of the cases. The presence of connective tissue was detected at the bone level, with cartilage formation at different levels. Each island presented levels of involvement that could indicate various degrees of aggressiveness. Type I collagen was observed in most cases, although type III was also identified. The development of histological diagnostic methods to determine the aggressiveness or level of involvement in UCH is not currently possible. Further studies are needed to establish new histological classifications.

  13. Epizootiology and histopathology of Parvicapsula sp. in coho salmon Oncorhynchus kisutch

    USGS Publications Warehouse

    Yasutake, William T.; Elliott, Diane G.

    2003-01-01

    The epizootiology and histopathology of the myxosporean Parvicapsula sp. was studied during monthly health surveys of 4 groups of coho salmon Oncorhynchus kisutch at a commercial farm in Puget Sound, Washington, USA, from 1984 to 1986. No Parvicapsula sp. was detected in histological samples taken from juvenile fish in fresh water, but the parasite was detected in fish from all groups 2 to 8 mo after transfer to seawater net pens. Groups placed in seawater net pens in November and December had a higher prevalence of infection, and a longer period of continuous detected infection, than those introduced into net pens in May. For the groups transferred to seawater in November and December, the highest infection prevalence (45 to 90%) was detected during the following March and April. Among 13 tissues examined histologically, only the pseudobranch and kidney were positive for Parvicapsula sp., with 26 (62%) of 42 positive fish showing infections only in the pseudobranch, 5 (12%) showing infections only in the kidney, and 11 (26%) showing infections in both organs. Both the pseudobranch and kidney were apparent primary infection sites, but pseudobranch infections appeared to persist longer in a population. Pseudobranch infections were frequently heavy and associated with extensive inflammation and necrosis of filament and lamellar tissues. The kidney had been the only infection site reported for Parvicapsula sp. in previous studies of coho salmon.

  14. A correlative method for imaging identical regions of samples by micro-CT, light microscopy, and electron microscopy: imaging adipose tissue in a model system.

    PubMed

    Sengle, Gerhard; Tufa, Sara F; Sakai, Lynn Y; Zulliger, Martin A; Keene, Douglas R

    2013-04-01

    We present a method in which a precise region of interest within an intact organism is spatially mapped in three dimensions by non-invasive micro-computed X-ray tomography (micro-CT), then further evaluated by light microscopy (LM) and transmission electron microscopy (TEM). Tissues are prepared as if for TEM including osmium fixation, which imparts soft tissue contrast in the micro-CT due to its strong X-ray attenuation. This method may therefore be applied to embedded, archived TEM samples. Upon selection of a two-dimensional (2-D) projection from a region of interest (ROI) within the three-dimensional volume, the epoxy-embedded sample is oriented for microtomy so that the sectioning plane is aligned with the micro-CT projection. Registration is verified by overlaying LM images with 2-D micro-CT projections. Structures that are poorly resolved in the micro-CT may be evaluated at TEM resolution by observing the next serial ultrathin section, thereby accessing the same ROI by all three imaging techniques. We compare white adipose tissue within the forelimbs of mice harboring a lipid-altering mutation with their littermate controls. We demonstrate that individual osmium-stained lipid droplets as small as 15 µm and separated by as little as 35 µm may be discerned as separate entities in the micro-CT, validating this to be a high-resolution, non-destructive technique for evaluation of fat content.

  15. Meniere's disease: histopathology, cytochemistry, and imaging.

    PubMed

    Ishiyama, Gail; Lopez, Ivan A; Sepahdari, Ali R; Ishiyama, Akira

    2015-04-01

    Meniere's disease is a poorly understood, disabling syndrome causing spells of vertigo, hearing fluctuation, tinnitus, and aural fullness. In this paper, we present a review of the histopathology, cytochemistry, and imaging of Meniere's disease. Histopathology is significant for neuroepithelial damage with hair cell loss, basement membrane thickening, and perivascular microvascular damage. Cytochemical alterations are significant for altered AQP4 and AQP6 expression in the supporting cell, and altered cochlin and mitochondrial protein expression. Current developments include imaging techniques to determine the degree and presence of endolymphatic hydrops, and future studies will endeavor to correlate the observance of hydrops with clinical findings. PMID:25766597

  16. Surgery on the Trabecular Meshwork: Histopathological Evidence

    PubMed Central

    Bhartiya, Shibal; Ichhpujani, Parul

    2015-01-01

    ABSTRACT Juxtacanalicular (JXT) trabecular meshwork and endothelial lining of Schlemm’s canal have been cited as the loci of aqueous outflow resistance, both in a normal as well as a glaucomatous eye. In this review, we attempt to understand the currently available surgical modalities in light of the available histopathological evidence, regarding localization of outflow resistance. How to cite this article: Bhartiya S, Ichhpujani P, Shaarawy T. Surgery on the Trabecular Meshwork: Histopathological Evidence. J Curr Glaucoma Pract 2015;9(2):51-61. PMID:26997835

  17. The determination of 22 natural brassinosteroids in a minute sample of plant tissue by UHPLC-ESI-MS/MS.

    PubMed

    Tarkowská, Danuše; Novák, Ondřej; Oklestkova, Jana; Strnad, Miroslav

    2016-09-01

    The triterpenoid plant hormones brassinosteroids (BRs) are believed to influence almost every aspect of plant growth and development. We have developed a sensitive mass spectrometry-based method for the simultaneous profiling of twenty-two naturally occurring brassinosteroids including biosynthetic precursors and the majority of biologically active metabolites. Using ultra-high performance liquid chromatographic (UHPLC) analysis, the run time was reduced up to three times (to 9 min) in comparison to standard HPLC BRs analyses, the retention time stability was improved to 0.1-0.2 % RSD and the injection accuracy was increased to 1.1-4.9 % RSD. The procedures for extraction and for two-step purification based on solid-phase extraction (SPE) were optimised in combination with subsequent UHPLC analysis coupled to electrospray ionisation tandem mass spectrometry (ESI-MS/MS) using Brassica flowers and Arabidopsis plant tissue extracts. In multiple reaction monitoring (MRM) mode, the average detection limit for BRs analysed was close to 7 pg, and the linear range covered up to 3 orders of magnitude. The low detection limits for this broad range of BR metabolites enabled as little as 50 mg of plant tissue to be used for quantitative analyses. The results of determinations exploiting internal standards showed that this approach provides a high level of practicality, reproducibility and recovery. The method we have established will enable researchers to gain a better understanding of the dynamics of the biosynthesis and metabolism of brassinosteroids and their modes of action in plant growth and development. PMID:27531032

  18. The determination of 22 natural brassinosteroids in a minute sample of plant tissue by UHPLC-ESI-MS/MS.

    PubMed

    Tarkowská, Danuše; Novák, Ondřej; Oklestkova, Jana; Strnad, Miroslav

    2016-09-01

    The triterpenoid plant hormones brassinosteroids (BRs) are believed to influence almost every aspect of plant growth and development. We have developed a sensitive mass spectrometry-based method for the simultaneous profiling of twenty-two naturally occurring brassinosteroids including biosynthetic precursors and the majority of biologically active metabolites. Using ultra-high performance liquid chromatographic (UHPLC) analysis, the run time was reduced up to three times (to 9 min) in comparison to standard HPLC BRs analyses, the retention time stability was improved to 0.1-0.2 % RSD and the injection accuracy was increased to 1.1-4.9 % RSD. The procedures for extraction and for two-step purification based on solid-phase extraction (SPE) were optimised in combination with subsequent UHPLC analysis coupled to electrospray ionisation tandem mass spectrometry (ESI-MS/MS) using Brassica flowers and Arabidopsis plant tissue extracts. In multiple reaction monitoring (MRM) mode, the average detection limit for BRs analysed was close to 7 pg, and the linear range covered up to 3 orders of magnitude. The low detection limits for this broad range of BR metabolites enabled as little as 50 mg of plant tissue to be used for quantitative analyses. The results of determinations exploiting internal standards showed that this approach provides a high level of practicality, reproducibility and recovery. The method we have established will enable researchers to gain a better understanding of the dynamics of the biosynthesis and metabolism of brassinosteroids and their modes of action in plant growth and development.

  19. [Histopathological diagnostic work-up of joint endoprosthesis-associated pathologies].

    PubMed

    Krenn, V; Perino, G; Krenn, V T; Wienert, S; Saberi, D; Hügle, T; Hopf, F; Huber, M

    2016-05-01

    Increasing classes of joint implants and the combination of materials results in increased and wear-associated pathologies. According to the revised consensus classification, the following types can be recognized at conventional histological examination: Type I, particle-induced type; Type II, infection type; Type III, combination type; Type IV, indifferent type; Type V arthrofibrotic type; Type VI, allergic/immunological/toxic adverse reactions and Type VII, bone pathologies. Wear particles are histopathologically characterized according to the Krenn particle algorithm which focuses on a descriptive identification of wear particles and the differentiation of other nonwear-related particles. Type VII is considered histologically when there is evidence of a perivascular/interstitial lymphocytic CD20- and CD3-positive infiltrate, presence of mast cells and eosinophils, and tissue necrosis/infarction associated with implant wear material. Since wear particle-induced toxicity cannot be differentiated with certainty from hypersensitivity/allergic reaction on histological examination, immunological-allergological and clinical data should be used as supplementary criteria for the differential diagnosis. Tissue sampling should be performed from periprosthetic soft tissue with location mapping and when feasible also from bone tissue. Additional information regarding the type of implant and clinical, radiological, immunological, and microbiology data should be available to the pathologist. Further immunohistochemical studies are recommended in the following settings: infection (CD15, CD20, CD68); prosthesis-associated arthrofibrosis (β‑catenin); allergic/immunologic/toxic adverse reactions (CD20, CD3, CD4, CD8, CD117 and for T‑cell characterization T‑bet, GATA-3, and FOXP3).

  20. Results of quality-control sampling of water, bed sediment, and tissue in the Western Lake Michigan Drainages study unit of the National Water-Quality Assessment Program

    USGS Publications Warehouse

    Fitzgerald, S.A.

    1997-01-01

    This report contains the quality control results of the Western Lake Michigan Drainages study unit of the National Water Quality Assessment Program. Quality control samples were collected in the same manner and contemporaneously with environmental samples during the first highintensity study phase in the unit (1992 through 1995) and amounted to approximately 15 percent of all samples collected. The accuracy and precision of hundreds of chemical analyses of surface and ground-water, bed sediment, and tissue was determined through the collection and analysis of field blanks, field replicates and splits, matrix spikes, and surrogates. Despite the several detections of analytes in the field blanks, the concentrations of most constituents in the environmental samples will likely be an order of magnitude or higher than those in the blanks. However, frequent detections, and high concentrations, of dissolved organic carbon (DOC) in several surface and ground-water blanks are probably significant with respect to commonly measured environmental concentrations, and the environmental data will have to be qualified accordingly. The precision of sampling of water on a percent basis, as determined from replicates and splits, was generally proportional to the concentration of the constituents, with constituents present in relatively high concentrations generally having less sampling variability than those with relatively low concentrations. In general, analytes with relatively high variability between replicates were present at concentrations near the reporting limit or were associated with relatively small absolute concentration differences, or both. Precision of replicates compared to that for splits in bed sediment samples was similar, thus eliminating sampling as a major source of variability in analyte concentrations. In the case the phthalates in bed sediment, contamination in either the field or laboratory could have caused the relatively large variability between replicate

  1. Impact of tissue type and content of neoplastic cells of samples on the quality of epidermal growth factor receptor mutation analysis among patients with lung adenocarcinoma

    PubMed Central

    PALIOGIANNIS, PANAGIOTIS; ATTENE, FEDERICO; COSSU, ANTONIO; DEFRAIA, EFISIO; PORCU, GIUSEPPE; CARTA, ANNAMARIA; SOTGIU, MARIA IGNAZIA; PAZZOLA, ANTONIO; CORDERO, LORENZO; CAPELLI, FRANCESCA; FADDA, GIOVANNI MARIA; ORTU, SALVATORE; SOTGIU, GIOVANNI; PALOMBA, GRAZIA; SINI, MARIA CRISTINA; PALMIERI, GIUSEPPE; COLOMBINO, MARIA

    2015-01-01

    Assessment of the epidermal growth factor receptor (EGFR) mutational status has become crucial in recent years in the molecular classification of patients with lung cancer. The impact of the type and quantity of malignant cells of the neoplastic specimen on the quality of mutation analysis remains to be elucidated, and only empirical and sporadic data are available. The aim of the present study was to investigate the impact of tissue type and content of neoplastic cells in the specimen on the quality of EGFR mutation analysis among patients with lung adenocarcinoma. A total of 515 patients with histologically-confirmed disease were included in the present study. Formalin-fixed paraffin embedded tissue samples were used for the mutation analysis and the content of the neoplastic cells was evaluated using light microscopy. Genomic DNA was isolated using a standard protocol. The coding sequences and splice junctions of exons 18, 19 and 21 in the EGFR gene were then screened for mutations by direct automated sequencing. The mean age of the patients examined was 64.9 years and 357 (69.3%) were male. A total of 429 tissue samples (83.3%) were obtained by biopsy and the remaining samples were obtained by surgery. A total of 456 samples (88.5%) were observed from primary lung adenocarcinomas, while 59 (11.5%) were from metastatic lesions. EGFR mutations occurred in 59 cases (11.5%); exon 18 mutations were detected in one case (1.7%), whereas exon 19 and 21 mutations were detected in 30 (51%) and 28 (47.3%) cases, respectively. EGFR mutations were more frequent in females and patients that had never smoked. The distribution of the mutations among primary and metastatic tissues exhibited no significant differences in the proportions of EGFR mutations detected. However, a statistically significant difference in the number of mutations detected was found between samples with at least 50% of neoplastic cells (450 cases-57 mutations; 12.7%) and those with <50% of neoplastic

  2. A generic nuclei detection method for histopathological breast images

    NASA Astrophysics Data System (ADS)

    Kost, Henning; Homeyer, André; Bult, Peter; Balkenhol, Maschenka C. A.; van der Laak, Jeroen A. W. M.; Hahn, Horst K.

    2016-03-01

    The detection of cell nuclei plays a key role in various histopathological image analysis problems. Considering the high variability of its applications, we propose a novel generic and trainable detection approach. Adaption to specific nuclei detection tasks is done by providing training samples. A trainable deconvolution and classification algorithm is used to generate a probability map indicating the presence of a nucleus. The map is processed by an extended watershed segmentation step to identify the nuclei positions. We have tested our method on data sets with different stains and target nuclear types. We obtained F1-measures between 0.83 and 0.93.

  3. Sample Preparation for Mass Spectrometry Analysis of Protein-Protein Interactions in Cancer Cell Lines and Tissues.

    PubMed

    Beigbeder, Alice; Vélot, Lauriane; James, D Andrew; Bisson, Nicolas

    2016-01-01

    A precisely controlled network of protein-protein interactions constitutes the basis for functional signaling pathways. This equilibrium is more often than not disrupted in cancer cells, by the aberrant expression or activation of oncogenic proteins. Therefore, the analysis of protein interaction networks in cancer cells has become crucial to expand our comprehension of the molecular underpinnings of tumor formation and progression. This protocol describes a sample preparation method for the analysis of signaling complexes by mass spectrometry (MS), following the affinity purification of a protein of interest from a cancer cell line or a solid tumor. In particular, we provide a spin tip-based protease digestion procedure that offers a more rapid and controlled alternative to other gel-based and gel-free methods. This sample preparation protocol represents a useful strategy to identify protein interactions and to gain insight into the molecular mechanisms that contribute to a given cancer phenotype. PMID:27581032

  4. Selective Extraction and Effective Separation of Galactosylsphingosine (Psychosine) and Glucosylsphingosine from Other Glycosphingolipids in Pathological Tissue Samples

    PubMed Central

    Li, Su-Chen; Buck, Wayne R.; Haskins, Mark E.; Wu, Sz-Wei; Khoo, Kay-Hooi; Sidransky, Ellen; Bunnell, Bruce A.

    2012-01-01

    To facilitate the study of the chemical pathology of galactosylsphingosine (psychosine, GalSph) in Krabbe disease and glucosylsphingosine (GlcSph) in Gaucher disease, we have devised a facile method for the effective separation of these two glycosylsphingosines from other glycosphingolipids (GSLs) in Krabbe brain and Gaucher spleen samples. The procedure involves the use of acetone to selectively extract GalSph and GlcSph, respectively, from Krabbe brain and Gaucher spleen samples. Since acetone does not extract other GSLs except modest amounts of galactosylceramide, sulfatide, and glucosylceramide, the positively charged GalSph or GlcSph in the acetone extract can be readily separated from other GSLs by batchwise cation-exchange chromatography using a Waters Accell Plus CM Cartridge. GalSph or GlcSph enriched by this simple procedure can be readily analyzed by thin-layer chromatography or high-performance liquid chromatography. PMID:21136152

  5. Direct online HPLC-CV-AFS method for traces of methylmercury without derivatisation: a matrix-independent method for urine, sediment and biological tissue samples.

    PubMed

    Brombach, Christoph-Cornelius; Gajdosechova, Zuzana; Chen, Bin; Brownlow, Andrew; Corns, Warren T; Feldmann, Jörg; Krupp, Eva M

    2015-01-01

    Mercury (Hg) is a global pollutant which occurs in different species, with methylmercury (MeHg) being the critical compound due to its neurotoxicity and bioaccumulation through the food chain. Methods for trace speciation of MeHg are therefore needed for a vast range of sample matrices, such as biological tissues, fluids, soils or sediments. We have previously developed an ultra-trace speciation method for methylmercury in water, based on a preconcentration HPLC cold vapour atomic fluorescence spectrometry (HPLC-CV-AFS) method. The focus of this work is mercury speciation in a variety of sample matrices to assess the versatility of the method. Certified reference materials were used where possible, and samples were spiked where reference materials were not available, e.g. human urine. Solid samples were submitted for commonly used digestion or extraction processes to obtain a liquid sample for injection into the analytical system. For MeHg in sediment samples, an extraction procedure was adapted to accommodate MeHg separation from high amounts of Hg(2+) to avoid an overload of the column. The recovery for MeHg determination was found to be in the range of 88-104% in fish reference materials (DOLT-2, DOLT-4, DORM-3), lobster (TORT-2), seaweed (IAEA-140/TM), sediments (ERM(®)-CC580) and spiked urine and has been proven to be robust, reliable, virtually matrix-independent and relatively cost-effective. Applications in the ultra-trace concentration range are possible using the preconcentration up to 200 mL, while for higher MeHg-containing samples, lower volumes can be applied. A comparison was carried out between species-specific isotope dilution gas chromatography inductively coupled plasma mass spectrometry (SSID-GC-ICP-MS) as the gold standard and HPLC-CV-AFS for biological tissues (liver, kidney and muscle of pilot whales), showing a slope of 1.008 and R (2) = 0.97, which indicates that the HPLC-CV-AFS method achieves well-correlated results for MeHg in

  6. Direct online HPLC-CV-AFS method for traces of methylmercury without derivatisation: a matrix-independent method for urine, sediment and biological tissue samples.

    PubMed

    Brombach, Christoph-Cornelius; Gajdosechova, Zuzana; Chen, Bin; Brownlow, Andrew; Corns, Warren T; Feldmann, Jörg; Krupp, Eva M

    2015-01-01

    Mercury (Hg) is a global pollutant which occurs in different species, with methylmercury (MeHg) being the critical compound due to its neurotoxicity and bioaccumulation through the food chain. Methods for trace speciation of MeHg are therefore needed for a vast range of sample matrices, such as biological tissues, fluids, soils or sediments. We have previously developed an ultra-trace speciation method for methylmercury in water, based on a preconcentration HPLC cold vapour atomic fluorescence spectrometry (HPLC-CV-AFS) method. The focus of this work is mercury speciation in a variety of sample matrices to assess the versatility of the method. Certified reference materials were used where possible, and samples were spiked where reference materials were not available, e.g. human urine. Solid samples were submitted for commonly used digestion or extraction processes to obtain a liquid sample for injection into the analytical system. For MeHg in sediment samples, an extraction procedure was adapted to accommodate MeHg separation from high amounts of Hg(2+) to avoid an overload of the column. The recovery for MeHg determination was found to be in the range of 88-104% in fish reference materials (DOLT-2, DOLT-4, DORM-3), lobster (TORT-2), seaweed (IAEA-140/TM), sediments (ERM(®)-CC580) and spiked urine and has been proven to be robust, reliable, virtually matrix-independent and relatively cost-effective. Applications in the ultra-trace concentration range are possible using the preconcentration up to 200 mL, while for higher MeHg-containing samples, lower volumes can be applied. A comparison was carried out between species-specific isotope dilution gas chromatography inductively coupled plasma mass spectrometry (SSID-GC-ICP-MS) as the gold standard and HPLC-CV-AFS for biological tissues (liver, kidney and muscle of pilot whales), showing a slope of 1.008 and R (2) = 0.97, which indicates that the HPLC-CV-AFS method achieves well-correlated results for MeHg in

  7. Persistent organochlorine pesticides detected in blood and tissue samples of vultures from different localities in South Africa.

    PubMed

    van Wyk, E; Bouwman, H; van der Bank, H; Verdoorn, G H; Hofmann, D

    2001-07-01

    Gas chromatography was used to establish the presence of quantifiable residues of 14 persistent chlorinated hydrocarbon pollutants in whole blood, clotted blood, heart, kidney, liver and muscle samples obtained from individual African whitebacked (Pseudogyps africanus), Cape griffon (Gyps coprotheres) and Lappetfaced (Torgos tracheliotos) vultures from different localities in South Africa. The levels of pesticides measured in whole blood samples of live specimens were compared between nestlings from two natural breeding colonies, adults from a wildlife area and birds held in captivity. Statistically significant (P<0.05) differences between populations were detected in geometric means calculated for gamma-BHC (lindane), alpha(cis)-chlordane and alpha-endosulfan. Five of the organochlorine contaminants displayed significant variations between concentrations detected in the clotted blood, organs and muscles excised from vulture carcasses. This includes residues of gamma-BHC, alpha-chlordane, dieldrin, beta-endosulfan and heptachlor epoxide. Values of the respective biocides measured in vulture samples were generally low in comparison to results documented for a number of avian species. Although no threat is posed by any of the organochloride pesticides, continual monitoring of especially breeding colonies is recommended. Furthermore, the suitability of African whitebacked vulture nestlings as basic bioindicators is highly advocated. PMID:11461840

  8. Imaging diagnosis--computed tomographic, surgical, and histopathologic characteristics of an infiltrative angiolipoma in a dog.

    PubMed

    Kraun, Michael B; Nelson, Nathan C; Hollinger, Charlotte

    2015-01-01

    A 6-year-old female spayed Shetland Sheepdog presented for evaluation of a subcutaneous mass over the right prescapular region. The mass had been cytologically diagnosed as a lipoma by the referring veterinarian 20 months prior, but had grown significantly and was very firm. CT scan of the mass was suggestive of neoplasia; however, the tissue of origin could not be determined. Histopathologic evaluation diagnosed infiltrative angiolipoma, and marginal resection of the tumor was performed. Infiltrative angiolipomas are benign but locally aggressive neoplasms uncommonly reported in veterinary medicine. This report correlates the clinical, CT, and histopathologic characteristics of an infiltrative angiolipoma.

  9. [Imaging Mass Spectrometry in Histopathologic Analysis].

    PubMed

    Yamazaki, Fumiyoshi; Seto, Mitsutoshi

    2015-04-01

    Matrix-assisted laser desorption/ionization (MALDI)-imaging mass spectrometry (IMS) enables visualization of the distribution of a range of biomolecules by integrating biochemical information from mass spectrometry with positional information from microscopy. IMS identifies a target molecule. In addition, IMS enables global analysis of biomolecules containing unknown molecules by detecting the ratio of the molecular weight to electric charge without any target, which makes it possible to identify novel molecules. IMS generates data on the distribution of lipids and small molecules in tissues, which is difficult to visualize with either conventional counter-staining or immunohistochemistry. In this review, we firstly introduce the principle of imaging mass spectrometry and recent advances in the sample preparation method. Secondly, we present findings regarding biological samples, especially pathological ones. Finally, we discuss the limitations and problems of the IMS technique and clinical application, such as in drug development. PMID:26536781

  10. Histopathological features of hypertrophic bone mass of temporomandibular joint ankylosis (TMJA): An explanation of pathogenesis of TMJA.

    PubMed

    Duan, Denghui; Li, Jiangming; Xiao, E; He, Linhai; Yan, Yingbin; Chen, Yan; Zhang, Yi

    2015-07-01

    Temporomandibular joint ankylosis (TMJA) is a severe organic disease with progressive limitation of the mouth opening. Histopathologically, a residual joint space is reported to consist of fibrous tissue and/or cartilage, indicating two types of interface (osteo-fibrous and osteo-chondral) of residual joint space. It is well known that adverse mechanical stress results in pathological changes of osteoarthritis and enthesopathy in these interfaces. What would happen pathologically in these interfaces of TMJA under repeated mandible movement has not been elucidated. Fourteen tissue samples of residual joint space and temporal and condylar bone were stained with hematoxylin and eosin and evaluated by collagen I and II immunohistochemistry. A pathological study of 14 TMJA patients showed that the residual joint space presented a fibrocartilage entheses structure and an articular cartilage structure. Moreover, these two structures were associated with pathological alterations of both osteoarthritis and enthesopathy, including degenerated and necrotized tissue, chondrocyte cloning, crack and fissure, various bone scleroses, and inflammatory granulation tissue. It is suggested that the pathological alterations of both osteoarthritis and enthesopathy occurred in TMJA, which hints at mechanical stress on TMJA development. PMID:26026887

  11. Tissue and serum samples of patients with papillary thyroid cancer with and without benign background demonstrate different altered expression of proteins

    PubMed Central

    Abdullah, Mardiaty Iryani; Lee, Ching Chin; Mat Junit, Sarni; Ng, Khoon Leong

    2016-01-01

    Background Papillary thyroid cancer (PTC) is mainly diagnosed using fine-needle aspiration biopsy. This most common form of well-differentiated thyroid cancer occurs with or without a background of benign thyroid goiter (BTG). Methods In the present study, a gel-based proteomics analysis was performed to analyse the expression of proteins in tissue and serum samples of PTC patients with (PTCb; n = 6) and without a history of BTG (PTCa; n = 8) relative to patients with BTG (n = 20). This was followed by confirmation of the levels of proteins which showed significant altered abundances of more than two-fold difference (p < 0.01) in the tissue and serum samples of the same subjects using ELISA. Results The data of our study showed that PTCa and PTCb distinguish themselves from BTG in the types of tissue and serum proteins of altered abundance. While higher levels of alpha-1 antitrypsin (A1AT) and heat shock 70 kDa protein were associated with PTCa, lower levels of A1AT, protein disulfide isomerase and ubiquitin-conjugating enzyme E2 N seemed apparent in the PTCb. In case of the serum proteins, higher abundances of A1AT and alpha 1-beta glycoprotein were detected in PTCa, while PTCb was associated with enhanced apolipoprotein A-IV and alpha 2-HS glycoprotein (AHSG). The different altered expression of tissue and serum A1AT as well as serum AHSG between PTCa and PTCb patients were also validated by ELISA. Discussion The distinctive altered abundances of the tissue and serum proteins form preliminary indications that PTCa and PTCb are two distinct cancers of the thyroid that are etiologically and mechanistically different although it is currently not possible to rule out that they may also be due other reasons such as the different stages of the malignant disease. These proteins stand to have a potential use as tissue or serum biomarkers to discriminate the three different thyroid neoplasms although this requires further validation in clinically representative

  12. Tissue and serum samples of patients with papillary thyroid cancer with and without benign background demonstrate different altered expression of proteins

    PubMed Central

    Abdullah, Mardiaty Iryani; Lee, Ching Chin; Mat Junit, Sarni; Ng, Khoon Leong

    2016-01-01

    Background Papillary thyroid cancer (PTC) is mainly diagnosed using fine-needle aspiration biopsy. This most common form of well-differentiated thyroid cancer occurs with or without a background of benign thyroid goiter (BTG). Methods In the present study, a gel-based proteomics analysis was performed to analyse the expression of proteins in tissue and serum samples of PTC patients with (PTCb; n = 6) and without a history of BTG (PTCa; n = 8) relative to patients with BTG (n = 20). This was followed by confirmation of the levels of proteins which showed significant altered abundances of more than two-fold difference (p < 0.01) in the tissue and serum samples of the same subjects using ELISA. Results The data of our study showed that PTCa and PTCb distinguish themselves from BTG in the types of tissue and serum proteins of altered abundance. While higher levels of alpha-1 antitrypsin (A1AT) and heat shock 70 kDa protein were associated with PTCa, lower levels of A1AT, protein disulfide isomerase and ubiquitin-conjugating enzyme E2 N seemed apparent in the PTCb. In case of the serum proteins, higher abundances of A1AT and alpha 1-beta glycoprotein were detected in PTCa, while PTCb was associated with enhanced apolipoprotein A-IV and alpha 2-HS glycoprotein (AHSG). The different altered expression of tissue and serum A1AT as well as serum AHSG between PTCa and PTCb patients were also validated by ELISA. Discussion The distinctive altered abundances of the tissue and serum proteins form preliminary indications that PTCa and PTCb are two distinct cancers of the thyroid that are etiologically and mechanistically different although it is currently not possible to rule out that they may also be due other reasons such as the different stages of the malignant disease. These proteins stand to have a potential use as tissue or serum biomarkers to discriminate the three different thyroid neoplasms although this requires further validation in clinically representative

  13. Assessment of the histopathological lesions and chemical analysis of feral cats to the smoke from Kuwait oil fires

    SciTech Connect

    Moeller, R.B.; Kalasinsky, V.F.; Razzaque, M.; Centeno, J.A.; Dick, E.J.

    1994-12-31

    Twenty-six adult or subadult feral cats were collected from Kuwait approximately 8 months after the ignition of the Kuwait oil wells. These animals were obtained from two sources: 12 animals from Kuwait City, a relatively Co smoke-free area, and 14 from the city of Alimadi, an area with heavy smoke. Animals were euthanized and a complete set of tissues consisting of all 0 major organs was taken for histopathology. Samples of lung, liver, kidney, urine, and blood were also taken for toxicology. Histopathological lesions observed in the lung were mild accumulations of anthracotic pigment in the lungs of 17 cats. Hyperplasia of the bronchial and bronchiolar gland in 8 cats, and smooth muscle hyperplasia of bronchioles in 14 cats. Iracheal gland hyperplasia was observed in 7 cats, and minimal squamous metaplasia of the tracheal mucosa in 17 cats, Laryngeal lesions consisted of submucosal gland hyperplasia in 2 cats and squamous metaplasia of the mucosa in 5 cats. Hyperplasia of the nasal submucosal glands was observed in 6 animals. The pharyngeal mucosa as well as other organs and organ systems (a) were normal in all cats. Atomic absorption analysis for 11 metals was performed; vanadium and nickel levels (two metals that were present in the smoke from the oil fires) are not indicative of substantial exposure to the oil fires. Based on the histopathological findings and toxicological analysis, it is felt that inhalation of air contaminated with smoke from the oil fires had little or no long-term effect on the animals examined.

  14. Assessment of the histopathological lesions and chemical analysis of feral cats to the smoke from the Kuwait oil fires

    SciTech Connect

    Moeller, R.B. Jr.; Dick, E.J.; Pletcher, J.M.

    1994-12-3