Spatial cluster analysis of nanoscopically mapped serotonin receptors for classification of fixed brain tissue

NASA Astrophysics Data System (ADS)

Sams, Michael; Silye, Rene; Göhring, Janett; Muresan, Leila; Schilcher, Kurt; Jacak, Jaroslaw

2014-01-01

We present a cluster spatial analysis method using nanoscopic dSTORM images to determine changes in protein cluster distributions within brain tissue. Such methods are suitable to investigate human brain tissue and will help to achieve a deeper understanding of brain disease along with aiding drug development. Human brain tissue samples are usually treated postmortem via standard fixation protocols, which are established in clinical laboratories. Therefore, our localization microscopy-based method was adapted to characterize protein density and protein cluster localization in samples fixed using different protocols followed by common fluorescent immunohistochemistry techniques. The localization microscopy allows nanoscopic mapping of serotonin 5-HT1A receptor groups within a two-dimensional image of a brain tissue slice. These nanoscopically mapped proteins can be confined to clusters by applying the proposed statistical spatial analysis. Selected features of such clusters were subsequently used to characterize and classify the tissue. Samples were obtained from different types of patients, fixed with different preparation methods, and finally stored in a human tissue bank. To verify the proposed method, samples of a cryopreserved healthy brain have been compared with epitope-retrieved and paraffin-fixed tissues. Furthermore, samples of healthy brain tissues were compared with data obtained from patients suffering from mental illnesses (e.g., major depressive disorder). Our work demonstrates the applicability of localization microscopy and image analysis methods for comparison and classification of human brain tissues at a nanoscopic level. Furthermore, the presented workflow marks a unique technological advance in the characterization of protein distributions in brain tissue sections.

Evaporation process in histological tissue sections for neutron autoradiography.

PubMed

Espector, Natalia M; Portu, Agustina; Santa Cruz, Gustavo A; Saint Martin, Gisela

2018-05-01

The analysis of the distribution and density of nuclear tracks forming an autoradiography in a nuclear track detector (NTD) allows the determination of 10 B atoms concentration and location in tissue samples from Boron Neutron Capture Therapy (BNCT) protocols. This knowledge is of great importance for BNCT dosimetry and treatment planning. Tissue sections studied with this technique are obtained by cryosectioning frozen tissue specimens. After the slicing procedure, the tissue section is put on the NTD and the sample starts drying. The thickness varies from its original value allowing more particles to reach the detector and, as the mass of the sample decreases, the boron concentration in the sample increases. So in order to determine the concentration present in the hydrated tissue, the application of corrective coefficients is required. Evaporation mechanisms as well as various factors that could affect the process of mass variation are outlined in this work. Mass evolution for tissue samples coming from BDIX rats was registered with a semimicro analytical scale and measurements were analyzed with software developed to that end. Ambient conditions were simultaneously recorded, obtaining reproducible evaporation curves. Mathematical models found in the literature were applied for the first time to this type of samples and the best fit of the experimental data was determined. The correlation coefficients and the variability of the parameters were evaluated, pointing to Page's model as the one that best represented the evaporation curves. These studies will contribute to a more precise assessment of boron concentration in tissue samples by the Neutron Autoradiography technique.

Characterization of the Tissue and Stromal Cell Components of Micro-Superficial Enhanced Fluid Fat Injection (Micro-SEFFI) for Facial Aging Treatment.

PubMed

Rossi, Martina; Roda, Barbara; Zia, Silvia; Vigliotta, Ilaria; Zannini, Chiara; Alviano, Francesco; Bonsi, Laura; Zattoni, Andrea; Reschiglian, Pierluigi; Gennai, Alessandro

2018-06-14

New microfat preparations provide material suitable for use as a regenerative filler for different facial areas. To support the development of new robust techniques for regenerative purposes, the cellular content of the sample should be considered. To evaluate the stromal vascular fraction (SVF) cell components of micro-superficial enhanced fluid fat injection (SEFFI) samples via a technique to harvest re-injectable tissue with minimum manipulation. The results were compared to those obtained from SEFFI samples. Microscopy analysis was performed to visualize the tissue structure. Micro-SEFFI samples were also fractionated using Celector ®, an innovative non-invasive separation technique, to provide an initial evaluation of sample fluidity and composition. SVFs obtained from SEFFI and micro-SEFFI were studied. Adipose stromal cells (ASCs) were isolated and characterized by proliferation and differentiation capacity assays. Microscopic and quality analyses of micro-SEFFI samples by Celector® confirmed the high fluidity and sample cellular composition in terms of red blood cell contamination, the presence of cell aggregates and extracellular matrix fragments. ASCs were isolated from adipose tissue harvested using SEFFI and micro-SEFFI systems. These cells were demonstrated to have a good proliferation rate and differentiation potential towards mesenchymal lineages. Despite the small sizes and low cellularity observed in micro-SEFFI-derived tissue, we were able to isolate stem cells. This result partially explains the regenerative potential of autologous micro-SEFFI tissue grafts. In addition, using this novel Celector® technology, tissues used for aging treatment were characterized analytically, and the adipose tissue composition was evaluated with no need for extra sample processing.

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.

Bruised Poultry Tissue as a Possible Source of Staphylococcal Infection

PubMed Central

Roskey, C. T.; Hamdy, M. K.

1972-01-01

Bacteriological analyses were made on 45 swab samples secured from hands of poultry workers on processing line, on 31 bruised and 15 normal poultry tissue samples, and on 15 swabs obtained from infected lacerations and exudates of abcesses on hands, arms, chest, and abdomen of poultry workers. A total of 170 Staphylococcus cultures were isolated from samples examined. These cultures were characterized morphologically and biochemically and then grouped into six distinct patterns. S. aureus was found in 86.6% of swab samples obtained from infected workers, in 40% of swabs from hands of workers who handle bruised birds, and in 38.7% of bruised tissues, and was absent from all samples obtained from hands of workers who do not handle bruised birds. All the coagulase-positive staphylococcal isolates were bacteriophage-typed, and the results showed that the same phage-type S. aureus was found in many poultry bruises and in infected lesions of poultry workers as well as on hands of workers who handle bruised birds. These results indicate that poultry bruises are a source of staphylococcal infection encountered among poultry workers. PMID:4553136

Rapid Molecular Microbiologic Diagnosis of Prosthetic Joint Infection

PubMed Central

Cazanave, Charles; Greenwood-Quaintance, Kerryl E.; Hanssen, Arlen D.; Karau, Melissa J.; Schmidt, Suzannah M.; Gomez Urena, Eric O.; Mandrekar, Jayawant N.; Osmon, Douglas R.; Lough, Lindsay E.; Pritt, Bobbi S.; Steckelberg, James M.

2013-01-01

We previously showed that culture of samples obtained by prosthesis vortexing and sonication was more sensitive than tissue culture for prosthetic joint infection (PJI) diagnosis. Despite improved sensitivity, culture-negative cases remained; furthermore, culture has a long turnaround time. We designed a genus-/group-specific rapid PCR assay panel targeting PJI bacteria and applied it to samples obtained by vortexing and sonicating explanted hip and knee prostheses, and we compared the results to those with sonicate fluid and periprosthetic tissue culture obtained at revision or resection arthroplasty. We studied 434 subjects with knee (n = 272) or hip (n = 162) prostheses; using a standardized definition, 144 had PJI. Sensitivities of tissue culture, of sonicate fluid culture, and of PCR were 70.1, 72.9, and 77.1%, respectively. Specificities were 97.9, 98.3, and 97.9%, respectively. Sonicate fluid PCR was more sensitive than tissue culture (P = 0.04). PCR of prosthesis sonication samples is more sensitive than tissue culture for the microbiologic diagnosis of prosthetic hip and knee infection and provides same-day PJI diagnosis with definition of microbiology. The high assay specificity suggests that typical PJI bacteria may not cause aseptic implant failure. PMID:23658273

Scanning ion conductance microscopy for visualizing the three-dimensional surface topography of cells and tissues.

PubMed

Nakajima, Masato; Mizutani, Yusuke; Iwata, Futoshi; Ushiki, Tatsuo

2018-01-01

Scanning ion conductance microscopy (SICM), which belongs to the family of scanning probe microscopy, regulates the tip-sample distance by monitoring the ion current through the use of an electrolyte-filled nanopipette as the probing tip. Thus, SICM enables "contact-free" imaging of cell surface topography in liquid conditions. In this paper, we applied hopping mode SICM for obtaining topographical images of convoluted tissue samples such as trachea and kidney in phosphate buffered saline. Some of the SICM images were compared with the images obtained by scanning electron microscopy (SEM) after drying the same samples. We showed that the imaging quality of hopping mode SICM was excellent enough for investigating the three-dimensional surface structure of the soft tissue samples. Thus, SICM is expected to be used for imaging a wide variety of cells and tissues - either fixed or alive- at high resolution under physiologically relevant liquid conditions. Copyright © 2017 Elsevier Ltd. All rights reserved.

Tumor Acquisition for Biomarker Research in Lung Cancer

PubMed Central

Stevenson, Marvaretta; Christensen, Jared; Shoemaker, Debra; Foster, Traci; Barry, William T.; Tong, Betty C.; Wahidi, Momen; Shofer, Scott; Datto, Michael; Ginsburg, Geoffrey; Crawford, Jeffrey; D’Amico, Thomas; Ready, Neal

2015-01-01

The biopsy collection data from two lung cancer trials that required fresh tumor samples be obtained for microarray analysis were reviewed. In the trial for advanced disease, microarray data were obtained on 50 patient samples, giving an overall success rate of 60.2%. The majority of the specimens were obtained through CT-guided lung biopsies (N=30). In the trial for early-stage patients, 28 tissue specimens were collected from excess tumor after surgical resection with a success rate of 85.7%. This tissue procurement program documents the feasibility in obtaining fresh tumor specimens prospectively that could be used for molecular testing. PMID:24810245

System and method for controlling depth of imaging in tissues using fluorescence microscopy under ultraviolet excitation following staining with fluorescing agents

DOE Office of Scientific and Technical Information (OSTI.GOV)

Demos, Stavros; Levenson, Richard

The present disclosure relates to a method for analyzing tissue specimens. In one implementation the method involves obtaining a tissue sample and exposing the sample to one or more fluorophores as contrast agents to enhance contrast of subcellular compartments of the tissue sample. The tissue sample is illuminated by an ultraviolet (UV) light having a wavelength between about 200 nm to about 400 nm, with the wavelength being selected to result in penetration to only a specified depth below a surface of the tissue sample. Inter-image operations between images acquired under different imaging parameters allow for improvement of the imagemore » quality via removal of unwanted image components. A microscope may be used to image the tissue sample and provide the image to an image acquisition system that makes use of a camera. The image acquisition system may create a corresponding image that is transmitted to a display system for processing and display.« less

Relationship of In Vivo MR Parameters to Histopathological and Molecular Characteristics of Newly Diagnosed, Nonenhancing Lower-Grade Gliomas.

PubMed

Luks, Tracy L; McKnight, Tracy Richmond; Jalbert, Llewellyn E; Williams, Aurelia; Neill, Evan; Lobo, Khadjia A; Persson, Anders I; Perry, Arie; Phillips, Joanna J; Molinaro, Annette M; Chang, Susan M; Nelson, Sarah J

2018-06-05

The goal of this research was to elucidate the relationship between WHO 2016 molecular classifications of newly diagnosed, nonenhancing lower grade gliomas (LrGG), tissue sample histopathology, and magnetic resonance (MR) parameters derived from diffusion, perfusion, and 1 H spectroscopic imaging from the tissue sample locations and the entire tumor. A total of 135 patients were scanned prior to initial surgery, with tumor cellularity scores obtained from 88 image-guided tissue samples. MR parameters were obtained from corresponding sample locations, and histograms of normalized MR parameters within the T2 fluid-attenuated inversion recovery lesion were analyzed in order to evaluate differences between subgroups. For tissue samples, higher tumor scores were related to increased normalized apparent diffusion coefficient (nADC), lower fractional anisotropy (nFA), lower cerebral blood volume (nCBV), higher choline (nCho), and lower N-acetylaspartate (nNAA). Within the T2 lesion, higher tumor grade was associated with higher nADC, lower nFA, and higher Cho to NAA index. Pathological analysis confirmed that diffusion and metabolic parameters increased and perfusion decreased with tumor cellularity. This information can be used to select targets for tissue sampling and to aid in making decisions about treating residual disease. Copyright © 2018 The Authors. Published by Elsevier Inc. All rights reserved.

A high-resolution optical imaging system for obtaining the serial transverse section images of biologic tissue

NASA Astrophysics Data System (ADS)

Wu, Li; Zhang, Bin; Wu, Ping; Liu, Qian; Gong, Hui

2007-05-01

A high-resolution optical imaging system was designed and developed to obtain the serial transverse section images of the biologic tissue, such as the mouse brain, in which new knife-edge imaging technology, high-speed and high-sensitive line-scan CCD and linear air bearing stages were adopted and incorporated with an OLYMPUS microscope. The section images on the tip of the knife-edge were synchronously captured by the reflection imaging in the microscope while cutting the biologic tissue. The biologic tissue can be sectioned at interval of 250 nm with the same resolution of the transverse section images obtained in x and y plane. And the cutting job can be automatically finished based on the control program wrote specially in advance, so we save the mass labor of the registration of the vast images data. In addition, by using this system a larger sample can be cut than conventional ultramicrotome so as to avoid the loss of the tissue structure information because of splitting the tissue sample to meet the size request of the ultramicrotome.

Equilibrium sampling of environmental pollutants in fish: comparison with lipid-normalized concentrations and homogenization effects on chemical activity.

PubMed

Jahnke, Annika; Mayer, Philipp; Adolfsson-Erici, Margaretha; McLachlan, Michael S

2011-07-01

Equilibrium sampling of organic pollutants into the silicone polydimethylsiloxane (PDMS) has recently been applied in biological tissues including fish. Pollutant concentrations in PDMS can then be multiplied with lipid/PDMS distribution coefficients (D(Lipid,PDMS) ) to obtain concentrations in fish lipids. In the present study, PDMS thin films were used for equilibrium sampling of polychlorinated biphenyls (PCBs) in intact tissue of two eels and one salmon. A classical exhaustive extraction technique to determine lipid-normalized PCB concentrations, which assigns the body burden of the chemical to the lipid fraction of the fish, was additionally applied. Lipid-based PCB concentrations obtained by equilibrium sampling were 85 to 106% (Norwegian Atlantic salmon), 108 to 128% (Baltic Sea eel), and 51 to 83% (Finnish lake eel) of those determined using total extraction. This supports the validity of the equilibrium sampling technique, while at the same time confirming that the fugacity capacity of these lipid-rich tissues for PCBs was dominated by the lipid fraction. Equilibrium sampling was also applied to homogenates of the same fish tissues. The PCB concentrations in the PDMS were 1.2 to 2.0 times higher in the homogenates (statistically significant in 18 of 21 cases, p < 0.05), indicating that homogenization increased the chemical activity of the PCBs and decreased the fugacity capacity of the tissue. This observation has implications for equilibrium sampling and partition coefficients determined using tissue homogenates. Copyright © 2011 SETAC.

Quantitation of repaglinide and metabolites in mouse whole-body thin tissue sections using droplet-based liquid microjunction surface sampling-high-performance liquid chromatography-electrospray ionization tandem mass spectrometry.

PubMed

Chen, Weiqi; Wang, Lifei; Van Berkel, Gary J; Kertesz, Vilmos; Gan, Jinping

2016-03-25

Herein, quantitation aspects of a fully automated autosampler/HPLC-MS/MS system applied for unattended droplet-based surface sampling of repaglinide dosed thin tissue sections with subsequent HPLC separation and mass spectrometric analysis of parent drug and various drug metabolites were studied. Major organs (brain, lung, liver, kidney and muscle) from whole-body thin tissue sections and corresponding organ homogenates prepared from repaglinide dosed mice were sampled by surface sampling and by bulk extraction, respectively, and analyzed by HPLC-MS/MS. A semi-quantitative agreement between data obtained by surface sampling and that by employing organ homogenate extraction was observed. Drug concentrations obtained by the two methods followed the same patterns for post-dose time points (0.25, 0.5, 1 and 2 h). Drug amounts determined in the specific tissues was typically higher when analyzing extracts from the organ homogenates. In addition, relative comparison of the levels of individual metabolites between the two analytical methods also revealed good semi-quantitative agreement. Copyright © 2015 Elsevier B.V. All rights reserved.

Quantitation of repaglinide and metabolites in mouse whole-body thin tissue sections using droplet-based liquid microjunction surface sampling-high-performance liquid chromatography-electrospray ionization tandem mass spectrometry

DOE PAGES

Chen, Weiqi; Wang, Lifei; Van Berkel, Gary J.; ...

2015-11-03

Herein, quantitation aspects of a fully automated autosampler/HPLC-MS/MS system applied for unattended droplet-based surface sampling of repaglinide dosed thin tissue sections with subsequent HPLC separation and mass spectrometric analysis of parent drug and various drug metabolites was studied. Major organs (brain, lung, liver, kidney, muscle) from whole-body thin tissue sections and corresponding organ homogenates prepared from repaglinide dosed mice were sampled by surface sampling and by bulk extraction, respectively, and analyzed by HPLC-MS/MS. A semi-quantitative agreement between data obtained by surface sampling and that by employing organ homogenate extraction was observed. Drug concentrations obtained by the two methods followed themore » same patterns for post-dose time points (0.25, 0.5, 1 and 2 h). Drug amounts determined in the specific tissues was typically higher when analyzing extracts from the organ homogenates. Furthermore, relative comparison of the levels of individual metabolites between the two analytical methods also revealed good semi-quantitative agreement.« less

Quantitation of repaglinide and metabolites in mouse whole-body thin tissue sections using droplet-based liquid microjunction surface sampling-high-performance liquid chromatography-electrospray ionization tandem mass spectrometry

DOE Office of Scientific and Technical Information (OSTI.GOV)

Chen, Weiqi; Wang, Lifei; Van Berkel, Gary J.

Herein, quantitation aspects of a fully automated autosampler/HPLC-MS/MS system applied for unattended droplet-based surface sampling of repaglinide dosed thin tissue sections with subsequent HPLC separation and mass spectrometric analysis of parent drug and various drug metabolites was studied. Major organs (brain, lung, liver, kidney, muscle) from whole-body thin tissue sections and corresponding organ homogenates prepared from repaglinide dosed mice were sampled by surface sampling and by bulk extraction, respectively, and analyzed by HPLC-MS/MS. A semi-quantitative agreement between data obtained by surface sampling and that by employing organ homogenate extraction was observed. Drug concentrations obtained by the two methods followed themore » same patterns for post-dose time points (0.25, 0.5, 1 and 2 h). Drug amounts determined in the specific tissues was typically higher when analyzing extracts from the organ homogenates. Furthermore, relative comparison of the levels of individual metabolites between the two analytical methods also revealed good semi-quantitative agreement.« less

Immunohistochemical identification of Propionibacterium acnes in granuloma and inflammatory cells of myocardial tissues obtained from cardiac sarcoidosis patients.

PubMed

Asakawa, Naoya; Uchida, Keisuke; Sakakibara, Mamoru; Omote, Kazunori; Noguchi, Keiji; Tokuda, Yusuke; Kamiya, Kiwamu; Hatanaka, Kanako C; Matsuno, Yoshihiro; Yamada, Shiro; Asakawa, Kyoko; Fukasawa, Yuichiro; Nagai, Toshiyuki; Anzai, Toshihisa; Ikeda, Yoshihiko; Ishibashi-Ueda, Hatsue; Hirota, Masanori; Orii, Makoto; Akasaka, Takashi; Uto, Kenta; Shingu, Yasushige; Matsui, Yoshiro; Morimoto, Shin-Ichiro; Tsutsui, Hiroyuki; Eishi, Yoshinobu

2017-01-01

Although rare, cardiac sarcoidosis (CS) is potentially fatal. Early diagnosis and intervention are essential, but histopathologic diagnosis is limited. We aimed to detect Propionibacterium acnes, a commonly implicated etiologic agent of sarcoidosis, in myocardial tissues obtained from CS patients. We examined formalin-fixed paraffin-embedded myocardial tissues obtained by surgery or autopsy and endomyocardial biopsy from patients with CS (n = 26; CS-group), myocarditis (n = 15; M-group), or other cardiomyopathies (n = 39; CM-group) using immunohistochemistry (IHC) with a P. acnes-specific monoclonal antibody. We found granulomas in 16 (62%) CS-group samples. Massive (≥14 inflammatory cells) and minimal (<14 inflammatory cells) inflammatory foci, respectively, were detected in 16 (62%) and 11 (42%) of the CS-group samples, 10 (67%) and 10 (67%) of the M-group samples, and 1 (3%) and 18 (46%) of the CM-group samples. P. acnes-positive reactivity in granulomas, massive inflammatory foci, and minimal inflammatory foci were detected in 10 (63%), 10 (63%), and 8 (73%) of the CS-group samples, respectively, and in none of the M-group and CM-group samples. Frequent identification of P. acnes in sarcoid granulomas of originally aseptic myocardial tissues suggests that this indigenous bacterium causes granuloma in many CS patients. IHC detection of P. acnes in massive or minimal inflammatory foci of myocardial biopsy samples without granulomas may be useful for differentiating sarcoidosis from myocarditis or other cardiomyopathies.

Extraction efficiency and implications for absolute quantitation of propranolol in mouse brain, liver and kidney thin tissue sections using droplet-based liquid microjunction surface sampling-HPLC ESI-MS/MS

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kertesz, Vilmos; Weiskittel, Taylor M.; Vavek, Marissa

Currently, absolute quantitation aspects of droplet-based surface sampling for thin tissue analysis using a fully automated autosampler/HPLC-ESI-MS/MS system are not fully evaluated. Knowledge of extraction efficiency and its reproducibility is required to judge the potential of the method for absolute quantitation of analytes from thin tissue sections. Methods: Adjacent thin tissue sections of propranolol dosed mouse brain (10- μm-thick), kidney (10- μm-thick) and liver (8-, 10-, 16- and 24- μm-thick) were obtained. Absolute concentration of propranolol was determined in tissue punches from serial sections using standard bulk tissue extraction protocols and subsequent HPLC separations and tandem mass spectrometric analysis. Thesemore » values were used to determine propranolol extraction efficiency from the tissues with the droplet-based surface sampling approach. Results: Extraction efficiency of propranolol using 10- μm-thick brain, kidney and liver thin tissues using droplet-based surface sampling varied between ~45-63%. Extraction efficiency decreased from ~65% to ~36% with liver thickness increasing from 8 μm to 24 μm. Randomly selecting half of the samples as standards, precision and accuracy of propranolol concentrations obtained for the other half of samples as quality control metrics were determined. Resulting precision ( ±15%) and accuracy ( ±3%) values, respectively, were within acceptable limits. In conclusion, comparative quantitation of adjacent mouse thin tissue sections of different organs and of various thicknesses by droplet-based surface sampling and by bulk extraction of tissue punches showed that extraction efficiency was incomplete using the former method, and that it depended on the organ and tissue thickness. However, once extraction efficiency was determined and applied, the droplet-based approach provided the required quantitation accuracy and precision for assay validations. Furthermore, this means that once the extraction efficiency was calibrated for a given tissue type and drug, the droplet-based approach provides a non-labor intensive and high-throughput means to acquire spatially resolved quantitative analysis of multiple samples of the same type.« less

Extraction efficiency and implications for absolute quantitation of propranolol in mouse brain, liver and kidney thin tissue sections using droplet-based liquid microjunction surface sampling-HPLC ESI-MS/MS

DOE PAGES

Kertesz, Vilmos; Weiskittel, Taylor M.; Vavek, Marissa; ...

2016-06-22

Currently, absolute quantitation aspects of droplet-based surface sampling for thin tissue analysis using a fully automated autosampler/HPLC-ESI-MS/MS system are not fully evaluated. Knowledge of extraction efficiency and its reproducibility is required to judge the potential of the method for absolute quantitation of analytes from thin tissue sections. Methods: Adjacent thin tissue sections of propranolol dosed mouse brain (10- μm-thick), kidney (10- μm-thick) and liver (8-, 10-, 16- and 24- μm-thick) were obtained. Absolute concentration of propranolol was determined in tissue punches from serial sections using standard bulk tissue extraction protocols and subsequent HPLC separations and tandem mass spectrometric analysis. Thesemore » values were used to determine propranolol extraction efficiency from the tissues with the droplet-based surface sampling approach. Results: Extraction efficiency of propranolol using 10- μm-thick brain, kidney and liver thin tissues using droplet-based surface sampling varied between ~45-63%. Extraction efficiency decreased from ~65% to ~36% with liver thickness increasing from 8 μm to 24 μm. Randomly selecting half of the samples as standards, precision and accuracy of propranolol concentrations obtained for the other half of samples as quality control metrics were determined. Resulting precision ( ±15%) and accuracy ( ±3%) values, respectively, were within acceptable limits. In conclusion, comparative quantitation of adjacent mouse thin tissue sections of different organs and of various thicknesses by droplet-based surface sampling and by bulk extraction of tissue punches showed that extraction efficiency was incomplete using the former method, and that it depended on the organ and tissue thickness. However, once extraction efficiency was determined and applied, the droplet-based approach provided the required quantitation accuracy and precision for assay validations. Furthermore, this means that once the extraction efficiency was calibrated for a given tissue type and drug, the droplet-based approach provides a non-labor intensive and high-throughput means to acquire spatially resolved quantitative analysis of multiple samples of the same type.« less

Comparison of two modalities: a novel technique, 'chromohysteroscopy', and blind endometrial sampling for the evaluation of abnormal uterine bleeding.

PubMed

Alay, Asli; Usta, Taner A; Ozay, Pinar; Karadugan, Ozgur; Ates, Ugur

2014-05-01

The objective of this study was to compare classical blind endometrial tissue sampling with hysteroscopic biopsy sampling following methylene blue dyeing in premenopausal and postmenopausal patients with abnormal uterine bleeding. A prospective case-control study was carried out in the Office Hysteroscopy Unit. Fifty-four patients with complaints of abnormal uterine bleeding were evaluated. Data of 38 patients were included in the statistical analysis. Three groups were compared by examining samples obtained through hysteroscopic biopsy before and after methylene blue dyeing, and classical blind endometrial tissue sampling. First, uterine cavity was evaluated with office hysteroscopy. Methylene blue dye was administered through the hysteroscopic inlet. Tissue samples were obtained from stained and non-stained areas. Blind endometrial sampling was performed in the same patients immediately after the hysteroscopy procedure. The results of hysteroscopic biopsy from methylene blue stained and non-stained areas and blind biopsy were compared. No statistically significant differences were determined in the comparison of biopsy samples obtained from methylene-blue stained, non-stained areas and blind biopsy (P > 0.05). We suggest that chromohysteroscopy is not superior to endometrial sampling in cases of abnormal uterine bleeding. Further studies with greater sample sizes should be performed to assess the validity of routine use of endometrial dyeing. © 2014 The Authors. Journal of Obstetrics and Gynaecology Research © 2014 Japan Society of Obstetrics and Gynecology.

2-hydroxyethyl metahcrylate/gelatin based superporous hydrogels for tissue regeneration

NASA Astrophysics Data System (ADS)

Tomić, Simonida Lj.; Babić, Marija M.; Vuković, Jovana S.; Perišić, Marija D.; Filipović, Vuk V.; Davidović, Sladjana Z.; Filipović, Jovanka M.

2016-05-01

In this study, superporous hydrogels were synthesized by free radical polymerization of 2-hydroxyethyl methacrylate without and in the presence of gelatin. Highly porous hydrogel structures were obtained by two different techniques: using a gas blowing agent, sodium bicarbonate, and a cryogenic treatment followed by freeze-drying. After the gel synthesis, gelatin molecules were covalently immobilised onto PHEMA via glytaraldehyde activation. All samples were characterized for morphological, mechanical, swelling and antibacterial properties. The results obtained show that samples with gelatin show better properties in comparison with PHEMA samples, which make these materials highly attractive for developing hydrogel scaffolds for tissue regeneration.

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.

An efficient, reliable and inexpensive device for the rapid homogenization of multiple tissue samples by centrifugation.

PubMed

Ilyin, S E; Plata-Salamán, C R

2000-02-15

Homogenization of tissue samples is a common first step in the majority of current protocols for RNA, DNA, and protein isolation. This report describes a simple device for centrifugation-mediated homogenization of tissue samples. The method presented is applicable to RNA, DNA, and protein isolation, and we show examples where high quality total cell RNA, DNA, and protein were obtained from brain and other tissue samples. The advantages of the approach presented include: (1) a significant reduction in time investment relative to hand-driven or individual motorized-driven pestle homogenization; (2) easy construction of the device from inexpensive parts available in any laboratory; (3) high replicability in the processing; and (4) the capacity for the parallel processing of multiple tissue samples, thus allowing higher efficiency, reliability, and standardization.

Selenium levels in human breast carcinoma tissue are associated with a common polymorphism in the gene for SELENOP (Selenoprotein P).

PubMed

Ekoue, Dede N; Zaichick, Sofia; Valyi-Nagy, Klara; Picklo, Matthew; Lacher, Craig; Hoskins, Kent; Warso, Michael A; Bonini, Marcelo G; Diamond, Alan M

2017-01-01

Selenium supplementation of the diets of rodents has consistently been shown to suppress mammary carcinogenesis and some, albeit not all, human epidemiological studies have indicated an inverse association between selenium and breast cancer risk. In order to better understand the role selenium plays in breast cancer, 30 samples of tumor tissue were obtained from women with breast cancer and analyzed for selenium concentration, the levels of several selenium-containing proteins and the levels of the MnSOD anti-oxidant protein. Polymorphisms within the genes for these same proteins were determined from DNA isolated from the tissue samples. There was a wide range of selenium in these tissues, ranging from 24 to 854ng/gm. The selenium levels in the tissues were correlated to the genotype of the SELENOP selenium carrier protein, but not to other proteins whose levels have been reported to be responsive to selenium availability, including GPX1, SELENOF and SBP1. There was an association between a polymorphism in the gene for MnSOD and the levels of the encoded protein. These studies were the first to examine the relationship between selenium levels, genotypes and protein levels in human tissues. Furthermore, the obtained data provide evidence for the need to obtain data about the effects of selenium in breast cancer by examining samples from that particular tissue type. Copyright © 2016 The Authors. Published by Elsevier GmbH.. All rights reserved.

Bladder cancer diagnosis during cystoscopy using Raman spectroscopy

NASA Astrophysics Data System (ADS)

Grimbergen, M. C. M.; van Swol, C. F. P.; Draga, R. O. P.; van Diest, P.; Verdaasdonk, R. M.; Stone, N.; Bosch, J. H. L. R.

2009-02-01

Raman spectroscopy is an optical technique that can be used to obtain specific molecular information of biological tissues. It has been used successfully to differentiate normal and pre-malignant tissue in many organs. The goal of this study is to determine the possibility to distinguish normal tissue from bladder cancer using this system. The endoscopic Raman system consists of a 6 Fr endoscopic probe connected to a 785nm diode laser and a spectral recording system. A total of 107 tissue samples were obtained from 54 patients with known bladder cancer during transurethral tumor resection. Immediately after surgical removal the samples were placed under the Raman probe and spectra were collected and stored for further analysis. The collected spectra were analyzed using multivariate statistical methods. In total 2949 Raman spectra were recorded ex vivo from cold cup biopsy samples with 2 seconds integration time. A multivariate algorithm allowed differentiation of normal and malignant tissue with a sensitivity and specificity of 78,5% and 78,9% respectively. The results show the possibility of discerning normal from malignant bladder tissue by means of Raman spectroscopy using a small fiber based system. Despite the low number of samples the results indicate that it might be possible to use this technique to grade identified bladder wall lesions during endoscopy.

A large-scale measurement of dielectric properties of normal and malignant colorectal tissues obtained from cancer surgeries at Larmor frequencies.

PubMed

Li, Zhou; Deng, Guanhua; Li, Zhe; Xin, Sherman Xuegang; Duan, Song; Lan, Maoying; Zhang, Sa; Gao, Yixin; He, Jun; Zhang, Songtao; Tang, Hongming; Wang, Weiwei; Han, Shuai; Yang, Qing X; Zhuang, Ling; Hu, Jiani; Liu, Feng

2016-11-01

Knowledge of dielectric properties of malignant human tissues is necessary for the recently developed magnetic resonance (MR) technique called MR electrical property tomography. This technique may be used in early tumor detection based on the obvious differentiation of the dielectric properties between normal and malignant tissues. However, the dielectric properties of malignant human tissues in the scale of the Larmor frequencies are not completely available in the literature. In this study, the authors focused only on the dielectric properties of colorectal tumor tissue. The dielectric properties of 504 colorectal malignant samples excised from 85 patients in the scale of the Larmor frequencies were measured using the precision open-ended coaxial probe method. The obtained complex-permittivity data were fitted to the single-pole Cole-Cole model. The median permittivity and conductivity for the malignant tissue sample were 79.3 and 0.881 S/m at 128 MHz, which were 14.6% and 17.0% higher, respectively, than those of normal tissue samples. Significant differences between normal and malignant tissues were found for the dielectric properties (p < 0.05). Experimental results indicated that the dielectric properties were significantly different between normal and malignant tissues for colorectal tissue. This large-scale clinical measurement provides more subtle base data to validate the technique of MR electrical property tomography.

Tissue recommendations for precision cancer therapy using next generation sequencing: a comprehensive single cancer center’s experiences

PubMed Central

Hong, Mineui; Bang, Heejin; Van Vrancken, Michael; Kim, Seungtae; Lee, Jeeyun; Park, Se Hoon; Park, Joon Oh; Park, Young Suk; Lim, Ho Yeong; Kang, Won Ki; Sun, Jong-Mu; Lee, Se Hoon; Ahn, Myung-Ju; Park, Keunchil; Kim, Duk Hwan; Lee, Seunggwan; Park, Woongyang; Kim, Kyoung-Mee

2017-01-01

To generate accurate next-generation sequencing (NGS) data, the amount and quality of DNA extracted is critical. We analyzed 1564 tissue samples from patients with metastatic or recurrent solid tumor submitted for NGS according to their sample size, acquisition method, organ, and fixation to propose appropriate tissue requirements. Of the 1564 tissue samples, 481 (30.8%) consisted of fresh-frozen (FF) tissue, and 1,083 (69.2%) consisted of formalin-fixed paraffin-embedded (FFPE) tissue. We obtained successful NGS results in 95.9% of cases. Out of 481 FF biopsies, 262 tissue samples were from lung, and the mean fragment size was 2.4 mm. Compared to lung, GI tract tumor fragments showed a significantly lower DNA extraction failure rate (2.1 % versus 6.1%, p = 0.04). For FFPE biopsy samples, the size of biopsy tissue was similar regardless of tumor type with a mean of 0.8 × 0.3 cm, and the mean DNA yield per one unstained slide was 114 ng. We obtained highest amount of DNA from the colorectum (2353 ng) and the lowest amount from the hepatobiliary tract (760.3 ng) likely due to a relatively smaller biopsy size, extensive hemorrhage and necrosis, and lower tumor volume. On one unstained slide from FFPE operation specimens, the mean size of the specimen was 2.0 × 1.0 cm, and the mean DNA yield per one unstained slide was 1800 ng. In conclusions, we present our experiences on tissue requirements for appropriate NGS workflow: > 1 mm2 for FF biopsy, > 5 unstained slides for FFPE biopsy, and > 1 unstained slide for FFPE operation specimens for successful test results in 95.9% of cases. PMID:28477007

Penetration of Clindamycin into Decubitus Ulcers

PubMed Central

Berger, Stephen A.; Barza, Michael; Haher, Jane; McFarland, James J.; Louie, Samuel; Kane, Anne

1978-01-01

Forty tissue samples, primarily of skin and bone, were obtained from 29 patients undergoing excision of decubitus ulcers after intravenous injection of 600 mg of clindamycin. Antibiotic concentrations exceeded 2.5 μg/g in 80% of the samples. In 50% of the instances, tissue levels were greater than those simultaneously present in the serum. PMID:708028

Multiview boosting digital pathology analysis of prostate cancer.

PubMed

Kwak, Jin Tae; Hewitt, Stephen M

2017-04-01

Various digital pathology tools have been developed to aid in analyzing tissues and improving cancer pathology. The multi-resolution nature of cancer pathology, however, has not been fully analyzed and utilized. Here, we develop an automated, cooperative, and multi-resolution method for improving prostate cancer diagnosis. Digitized tissue specimen images are obtained from 5 tissue microarrays (TMAs). The TMAs include 70 benign and 135 cancer samples (TMA1), 74 benign and 89 cancer samples (TMA2), 70 benign and 115 cancer samples (TMA3), 79 benign and 82 cancer samples (TMA4), and 72 benign and 86 cancer samples (TMA5). The tissue specimen images are segmented using intensity- and texture-based features. Using the segmentation results, a number of morphological features from lumens and epithelial nuclei are computed to characterize tissues at different resolutions. Applying a multiview boosting algorithm, tissue characteristics, obtained from differing resolutions, are cooperatively combined to achieve accurate cancer detection. In segmenting prostate tissues, the multiview boosting method achieved≥ 0.97 AUC using TMA1. For detecting cancers, the multiview boosting method achieved an AUC of 0.98 (95% CI: 0.97-0.99) as trained on TMA2 and tested on TMA3, TMA4, and TMA5. The proposed method was superior to single-view approaches, utilizing features from a single resolution or merging features from all the resolutions. Moreover, the performance of the proposed method was insensitive to the choice of the training dataset. Trained on TMA3, TMA4, and TMA5, the proposed method obtained an AUC of 0.97 (95% CI: 0.96-0.98), 0.98 (95% CI: 0.96-0.99), and 0.97 (95% CI: 0.96-0.98), respectively. The multiview boosting method is capable of integrating information from multiple resolutions in an effective and efficient fashion and identifying cancers with high accuracy. The multiview boosting method holds a great potential for improving digital pathology tools and research. Copyright © 2017 Elsevier B.V. All rights reserved.

Triple Gene Analysis Using Samples Obtained by Endobronchial Ultrasound-guided Transbronchial Needle Aspiration

PubMed Central

Lee, Kyungjong; Um, Sang-Won; Jeong, Byeong-Ho; Yang, Jung Wook; Choi, Yoon-La; Han, Joungho; Kim, Hojoong; Kwon, O Jung

2016-01-01

Objective A mutational analysis of tumor tissue samples is an important part of advanced lung cancer treatment strategies. This study evaluated the efficacy of a triple gene analysis using samples obtained via endobronchial ultrasound-guided transbronchial needle aspiration (EBUS-TBNA). Methods Either metastatic lymph nodes or primary lung mass samples obtained by EBUS-TBNA were collected between May 2011 and May 2013. We consecutively analyzed epidermal growth factor receptor (EGFR), V-Ki-ras2 Kirsten rat sarcoma viral oncogene homolog (KRAS), and anaplastic lymphoma kinase (ALK) fusion genes using remnant tissue samples. Results A total of 109 patients were diagnosed with non-small cell lung cancer (NSCLC). Of these, 70% were adenocarcinoma, 27% squamous cell carcinoma with NSCLC, and 3% were related to other types of lung cancer. EGFR mutations were detected in 23 cases (21.1%), KRAS mutations in 13 cases (11.9%), and ALK fusion genes in 5 cases (4.9%). The ALK fusion genes could not be analyzed in four cases because of insufficient tissue samples remaining after routine histochemistry and an EGFR/KRAS mutation analysis. We found that small biopsy samples from EBUS-TBNA were adequate for performing a triple gene analysis in 97 patients (96%). ALK fusion protein immunohistochemistry (IHC) was 100% consistent with fluorescence in situ hybridization (FISH). Conclusion Small samples obtained by EBUS-TBNA were found to be sufficient for performing a triple gene analysis following routine histology and IHC. ALK IHC showed a very good concordance with FISH for detecting ALK fusion genes. PMID:27803402

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.

Colon Cancer Associated Transcript-1 (CCAT1) Expression in Adenocarcinoma of the Stomach.

PubMed

Mizrahi, Ido; Mazeh, Haggi; Grinbaum, Ronit; Beglaibter, Nahum; Wilschanski, Michael; Pavlov, Vera; Adileh, Muchamad; Stojadinovic, Alexander; Avital, Itzhak; Gure, Ali Osmay; Halle, David; Nissan, Aviram

2015-01-01

Long non-coding RNAs (lncRNAs) have been shown to have functional roles in cancer biology and are dys-regulated in many tumors. Colon Cancer Associated Transcript -1 (CCAT1) is a lncRNA, previously shown to be significantly up-regulated in colon cancer. The aim of this study is to determine expression levels of CCAT1 in gastric carcinoma (GC). Tissue samples were obtained from patients undergoing resection for gastric carcinoma (n=19). For each patient, tumor tissue and normal appearing gastric mucosa were taken. Normal gastric tissues obtained from morbidly obese patients, undergoing laparoscopic sleeve gastrectomy served as normal controls (n=19). A human gastric carcinoma cell line (AGS) served as positive control. RNA was extracted from all tissue samples and CCAT1 expression was analyzed using quantitative real time-PCR (qRT-PCR). Low expression of CCAT1 was identified in normal gastric mucosa samples obtained from morbidly obese patients [mean Relative Quantity (RQ) = 1.95±0.4]. AGS human gastric carcinoma cell line showed an elevated level of CCAT1 expression (RQ=8.02). Expression levels of CCAT1 were approximately 10.8 fold higher in GC samples than in samples taken from the negative control group (RQ=21.1±5 vs. RQ=1.95±0.4, respectively, p<0.001). Interestingly, CCAT1 expression was significantly overexpressed in adjacent normal tissues when compared to the negative control group (RQ = 15.25±2 vs. RQ=1.95±0.4, respectively, p<0.001). Tissues obtained from recurrent GC cases showed the highest expression levels (RQ = 88.8±31; p<0.001). Expression levels increased with tumor stage (T4- 36.4±15, T3- 16.1±6, T2- 4.7±1), however this did not reach statistical significance (p=0.2). There was no difference in CCAT1 expression between intestinal and diffuse type GC (RQ=22.4±7 vs. 22.4±16, respectively, p=0.9). Within the normal gastric tissue samples, no significant difference in CCAT1 expression was observed in helicobacter pylori negative and positive patients (RQ= 2.4±0.9 vs. 0.93±0.2, respectively, p=0.13). CCAT1 is up-regulated in gastric cancer, and may serve as a potential bio-marker for early detection and surveillance.

Development of the technique of terahertz pulse spectroscopy for diagnostic malignant tumors during gastrointestinal surgeries

NASA Astrophysics Data System (ADS)

Goryachuk, A. A.; Khodzitsky, M. K.; Borovkova, M. A.; Khamid, A. K.; Dutkinskii, P. S.; Shishlo, D. A.

2016-08-01

Samples of fresh excised tissues obtained from patients who had undergone gastric cancer have been investigated. Samples were consisted of cancer zone, normal zone and zone mixed of normal and cancer tissues. Their optical properties and spectral features were investigated by terahertz time-domain spectroscopy (TDS) in reflection mode. It was found that waveforms of reflected signals from normal and cancer tissues were well distinguished so it can be concluded that it is easy to discriminate gastric cancer tissue from normal by using THz TDS.

Successful Application of Microarray Technology to Microdissected Formalin-Fixed, Paraffin-Embedded Tissue

PubMed Central

Coudry, Renata A.; Meireles, Sibele I.; Stoyanova, Radka; Cooper, Harry S.; Carpino, Alan; Wang, Xianqun; Engstrom, Paul F.; Clapper, Margie L.

2007-01-01

The establishment of a reliable method for using RNA from formalin-fixed, paraffin-embedded (FFPE) tissue would provide an opportunity to obtain novel gene expression data from the vast amounts of archived tissue. A custom-designed 22,000 oligonucleotide array was used in the present study to compare the gene expression profile of colonic epithelial cells isolated by laser capture microdissection from FFPE-archived samples with that of the same cell population from matched frozen samples, the preferred source of RNA. Total RNA was extracted from FFPE tissues, amplified, and labeled using the Paradise Reagent System. The quality of the input RNA was assessed by the Bioanalyzer profile, reverse transcriptase-polymerase chain reaction, and agarose gel electrophoresis. The results demonstrate that it is possible to obtain reliable microarray data from FFPE samples using RNA acquired by laser capture microdissection. The concordance between matched FFPE and frozen samples was evaluated and expressed as a Pearson’s correlation coefficient, with values ranging from 0.80 to 0.97. The presence of ribosomal RNA peaks in FFPE-derived RNA was reflected by a high correlation with paired frozen samples. A set of practical recommendations for evaluating the RNA integrity and quality in FFPE samples is reported. PMID:17251338

Cell and tissue microarray technologies for protein and nucleic acid expression profiling.

PubMed

Cardano, Marina; Diaferia, Giuseppe R; Falavigna, Maurizio; Spinelli, Chiara C; Sessa, Fausto; DeBlasio, Pasquale; Biunno, Ida

2013-02-01

Tissue microarray (TMA) and cell microarray (CMA) are two powerful techniques that allow for the immunophenotypical characterization of hundreds of samples simultaneously. In particular, the CMA approach is particularly useful for immunophenotyping new stem cell lines (e.g., cardiac, neural, mesenchymal) using conventional markers, as well as for testing the specificity and the efficacy of newly developed antibodies. We propose the use of a tissue arrayer not only to perform protein expression profiling by immunohistochemistry but also to carry out molecular genetics studies. In fact, starting with several tissues or cell lines, it is possible to obtain the complete signature of each sample, describing the protein, mRNA and microRNA expression, and DNA mutations, or eventually to analyze the epigenetic processes that control protein regulation. Here we show the results obtained using the Galileo CK4500 TMA platform.

[Transciptome among Mexicans: a large scale methodology to analyze the genetics expression profile of simultaneous samples in muscle, adipose tissue and lymphocytes obtained from the same individual].

PubMed

Bastarrachea, Raúl A; López-Alvarenga, Juan Carlos; Kent, Jack W; Laviada-Molina, Hugo A; Cerda-Flores, Ricardo M; Calderón-Garcidueñas, Ana Laura; Torres-Salazar, Amada; Torres-Salazar, Amanda; Nava-González, Edna J; Solis-Pérez, Elizabeth; Gallegos-Cabrales, Esther C; Cole, Shelley A; Comuzzie, Anthony G

2008-01-01

We describe the methodology used to analyze multiple transcripts using microarray techniques in simultaneous biopsies of muscle, adipose tissue and lymphocytes obtained from the same individual as part of the standard protocol of the Genetics of Metabolic Diseases in Mexico: GEMM Family Study. We recruited 4 healthy male subjects with BM1 20-41, who signed an informed consent letter. Subjects participated in a clinical examination that included anthropometric and body composition measurements, muscle biopsies (vastus lateralis) subcutaneous fat biopsies anda blood draw. All samples provided sufficient amplified RNA for microarray analysis. Total RNA was extracted from the biopsy samples and amplified for analysis. Of the 48,687 transcript targets queried, 39.4% were detectable in a least one of the studied tissues. Leptin was not detectable in lymphocytes, weakly expressed in muscle, but overexpressed and highly correlated with BMI in subcutaneous fat. Another example was GLUT4, which was detectable only in muscle and not correlated with BMI. Expression level concordance was 0.7 (p< 0.001) for the three tissues studied. We demonstrated the feasibility of carrying out simultaneous analysis of gene expression in multiple tissues, concordance of genetic expression in different tissues, and obtained confidence that this method corroborates the expected biological relationships among LEPand GLUT4. TheGEMM study will provide a broad and valuable overview on metabolic diseases, including obesity and type 2 diabetes.

Value of amniocentesis versus fetal tissue for cytogenetic analysis in cases of fetal demise.

PubMed

Bryant Borders, Ann E; Greenberg, Jessica; Plaga, Stacey; Shepard-Hinton, Megan; Yates, Carin; Elias, Sherman; Shulman, Lee P

2009-01-01

Use of fetal tissue for cytogenetic analysis in cases of second- and third-trimester fetal demise frequently results in unacceptably high failure rates. We reviewed our ongoing use of amniocentesis prior to uterine evacuation to determine if this provided a better source of cells for cytogenetic analysis. We compared cytogenetic results using fetal tissues obtained following uterine evacuation to our ongoing use of amniotic fluid cell obtained by transabdominal amniocentesis prior to uterine evacuation from 2003 to 2008. In 49 of the 63 cases evaluated by fetal tissue biopsies performed after uterine evacuation, a karyotypic analysis was obtained (77.8%). Among the 38 cases evaluated by amniocentesis, an amniotic fluid sample and fetal cytogenetic results were obtained in all 38 (100%) cases. Our findings indicate that amniocentesis is a more reliable source of cytogenetic information than fetal tissue in cases of second- and third-trimester fetal demise.

An HR-MAS MR Metabolomics Study on Breast Tissues Obtained with Core Needle Biopsy

PubMed Central

Cho, Nariya; Chang, Jung Min; Koo, Hye Ryoung; Yi, Ann; Kim, Hyeonjin; Park, Sunghyouk; Moon, Woo Kyung

2011-01-01

Background Much research has been devoted to the development of new breast cancer diagnostic measures, including those involving high-resolution magic angle spinning (HR-MAS) magnetic resonance (MR) spectroscopic techniques. Previous HR-MAS MR results have been obtained from post-surgery samples, which limits their direct clinical applicability. Methodology/Principal Findings In the present study, we performed HR-MAS MR spectroscopic studies on 31 breast tissue samples (13 cancer and 18 non-cancer) obtained by percutaneous core needle biopsy. We showed that cancer and non-cancer samples can be discriminated very well with Orthogonal Projections to Latent Structure-Discriminant Analysis (OPLS-DA) multivariate model on the MR spectra. A subsequent blind test showed 69% sensitivity and 94% specificity in the prediction of the cancer status. A spectral analysis showed that in cancer cells, taurine- and choline-containing compounds are elevated. Our approach, additionally, could predict the progesterone receptor statuses of the cancer patients. Conclusions/Significance HR-MAS MR metabolomics on intact breast tissues obtained by core needle biopsy may have a potential to be used as a complement to the current diagnostic and prognostic measures for breast cancers. PMID:22028780

Assessment of adenosine deaminase (ADA) activity and oxidative stress in patients with chronic tonsillitis.

PubMed

Garca, Mehmet Fatih; Demir, Halit; Turan, Mahfuz; Bozan, Nazım; Kozan, Ahmet; Belli, Şeyda Bayel; Arslan, Ayşe; Cankaya, Hakan

2014-06-01

To emphasize the effectiveness of adenosine deaminase (ADA) enzyme, which has important roles in the differentiation of lymphoid cells, and oxidative stress in patients with chronic tonsillitis. Serum and tissue samples were obtained from 25 patients who underwent tonsillectomy due to recurrent episodes of acute tonsillitis. In the control group, which also had 25 subjects, only serum samples were taken as obtaining tissue samples would not have been ethically appropriate. ADA enzyme activity, catalase (CAT), carbonic anhydrase (CA), nitric oxide (NO) and malondialdehyde (MDA) were measured in the serum and tissue samples of patients and control group subjects. The serum values of both groups were compared. In addition, the tissue and serum values of patients were compared. Serum ADA activity and the oxidant enzymes MDA and NO values of the patient group were significantly higher than those of the control group (p < 0.001), the antioxidant enzymes CA and CAT values of the patient group were significantly lower than those of the control group (p < 0.001). In addition, while CA, CAT and NO enzyme levels were found to be significantly higher in the tonsil tissue of the patient group when compared to serum levels (p < 0.05), there was no difference between tissue and serum MDA and ADA activity (p > 0.05). Elevated ADA activity may be effective in the pathogenesis of chronic tonsillitis both by impairing tissue structure and contributing to SOR formation.

In Vitro Tissue Differentiation using Dynamics of Tissue Mechanical Properties

NASA Astrophysics Data System (ADS)

Lin, Wei-Chiang; Phillips, Paul J.

2002-03-01

Dynamics of tissue mechanical properties of various human tissue types were studied at macroscopic as well as microscopic level in vitro. This study was conducted to enable the development of a feedback system based on dynamics of tissue mechanical properties for intraoperative guidance for tumor treatment (e.g., RF ablation of liver tumor) and noninvasive tumor localization. Human liver tissues, including normal, cancerous, and cirrhotic tissues, were obtained from patients receiving liver transplant or tumor resection at Vanderbilt University Medical Center with the approval of the Vanderbilt Institutional Review Board. Tissue samples, once resected from the patients, were snap-frozen using liquid nitrogen and stored at -70 oC. Measurements of the mechanical properties of these tissue samples were conducted at the University of Tennessee at Knoxville. Dynamics of tissue mechanical properties were measured from both native and thermally coagulated tissue samples at macroscopic and microscopic level. Preliminary results suggest the dynamics of mechanical properties of normal liver tissues are very different from those of cancerous liver tissues. The correlation between the dynamics of mechanical properties at macroscopic level and those at microscopic level is currently under investigation.

Use of a new jumbo forceps improves tissue acquisition of Barrett's esophagus surveillance biopsies.

PubMed

Komanduri, Sri; Swanson, Garth; Keefer, Laurie; Jakate, Shriram

2009-12-01

The major risk factors for the development of esophageal adenocarcinoma remain long-standing GERD and resultant Barrett's esophagus (BE). Finding the exact method of adequate tissue sampling for surveillance of dysplasia in BE remains a dilemma. We prospectively compared standard large-capacity biopsy forceps with a new jumbo biopsy forceps for dysplasia detection in BE. Prospective, single-center investigation. We prospectively enrolled 32 patients undergoing surveillance endoscopy for BE. Biopsy samples were obtained in paired fashion alternating between the experimental (jumbo) and control (large-capacity) forceps. Each sample was assessed for histopathology, specimen size, and adequacy. A total of 712 specimens were available for analysis for this investigation. Six patients were found to have dysplasia, and in 5 of those patients, the dysplasia was only detected with the jumbo forceps. The mean width was significantly greater in the Radial Jaw 4 jumbo group (3.3 mm vs 1.9 mm [P < .005]) as was the mean depth (2.0 mm vs 1.1 mm [P < .005]). Sixteen percent of samples obtained with the standard forceps provided an adequate sample, whereas the jumbo forceps provided an adequate sample 79% of the time (P < .05). A lack of a validated index for assessment of tissue adequacy in BE. The Radial Jaw 4 jumbo biopsy forceps significantly improves dysplasia detection and adequate tissue sampling in patients undergoing endoscopy for BE.

The lung tissue microbiota of mild and moderate chronic obstructive pulmonary disease.

PubMed

Pragman, Alexa A; Lyu, Tianmeng; Baller, Joshua A; Gould, Trevor J; Kelly, Rosemary F; Reilly, Cavan S; Isaacson, Richard E; Wendt, Chris H

2018-01-09

Oral taxa are often found in the chronic obstructive pulmonary disease (COPD) lung microbiota, but it is not clear if this is due to a physiologic process such as aspiration or experimental contamination at the time of specimen collection. Microbiota samples were obtained from nine subjects with mild or moderate COPD by swabbing lung tissue and upper airway sites during lung lobectomy. Lung specimens were not contaminated with upper airway taxa since they were obtained surgically. The microbiota were analyzed with 16S rRNA gene qPCR and 16S rRNA gene hypervariable region 3 (V3) sequencing. Data analyses were performed using QIIME, SourceTracker, and R. Streptococcus was the most common genus in the oral, bronchial, and lung tissue samples, and multiple other taxa were present in both the upper and lower airways. Each subject's own bronchial and lung tissue microbiota were more similar to each other than were the bronchial and lung tissue microbiota of two different subjects (permutation test, p = 0.0139), indicating more within-subject similarity than between-subject similarity at these two lung sites. Principal coordinate analysis of all subject samples revealed clustering by anatomic sampling site (PERMANOVA, p = 0.001), but not by subject. SourceTracker analysis found that the sources of the lung tissue microbiota were 21.1% (mean) oral microbiota, 8.7% nasal microbiota, and 70.1% unknown. An analysis using the neutral theory of community ecology revealed that the lung tissue microbiota closely reflects the bronchial, oral, and nasal microbiota (immigration parameter estimates 0.69, 0.62, and 0.74, respectively), with some evidence of ecologic drift occurring in the lung tissue. This is the first study to evaluate the mild-moderate COPD lung tissue microbiota without potential for upper airway contamination of the lung samples. In our small study of subjects with COPD, we found oral and nasal bacteria in the lung tissue microbiota, confirming that aspiration is a source of the COPD lung microbiota.

Analysis of EML4-ALK rearrangement in non-small cell lung cancer in Argentina.

PubMed

Verzura, María; Batagelj, Emilio; Bagnes, Claudia; Martin, Claudio; Enrico, Diego; Richardet, Eduardo; De la Iglesia, Paola

2018-02-26

Identification of EML4-ALK rearrangement by FISH test has become standard in advanced NSCLC patients. There is limited information about the prevalence and clinical characteristics of ALK translocation in Latin America. The aim of our study was to evaluate this lung cancer subtype features in Argentinian patients and the factibility of FISH test with different methods used for obtaining tissue samples. Between August 2014 and February 2017, 183 non-squamous NSCLC patients were prospectively enrolled from five Argentinian institutions. Different techniques and procedures were used to obtained tissue samples material. ALK determination was performed by FISH and immunohistochemistry (IHC). Correlation with clinico-pathological information and different biopsy procedures was assessed. From 183 non-squamous NSCLC samples, 131 could perform FISH test, finding 123 (93.9%) negative and 8 (6.1%) positive patients. Fifty-one samples were not evaluable by FISH, 35 because of technical problems and 16 due to not/weak signal. The difficulties in obtaining adequate FISH tests were observed significantly more frequently for fine-needle aspiration (FNA) and core-needle biopsy than for excisional and incisional biopsy (p = 0.009). Regarding the procedures, surgery was the most efficient, obtaining only 12.7% (10/79) of not evaluable samples for FISH, while CT guided biopsy and transbronchial biopsy (TBB) failed in 43.8% (21/48) and 41.3% (19/46) of patients respectively (p < 0.001). We observed a significant association between ALK translocation and never smoking habit (p = 0.004). Our ALK rearrangements frequency (6.1%) was similar to the reports worldwide. One of the major determinants for the ALK FISH test success is the quality of the tissue sample obtained. Copyright © 2018 Elsevier Inc. All rights reserved.

Sensitivity of bhk-21 cells supplemented with diethylaminoethyl-dextran for detection of street rabies virus in saliva samples.

PubMed Central

Larghi, O P; Nebel, A E; Lazaro, L; Savy, V L

1975-01-01

A tissue culture system for detecting rabies virus from saliva samples of suspected animals was developed and compared to suckling mouse inoculation. Swab samples were obtained from the mouth of the animal heads received for rabies diagnosis; these swabs were submerged in maintenance medium. The maintenance medium was inoculated intracerebrally into suckling mice and onto BHK-21 cells with diethylaminoethyl (DEAE)-dextran (BHK/DEAE) and without (BHK). Rabies immunofluorescence was performed on the brain of the mice dying during the observation period and also on both tissue culture systems every day after infection. The BHK-DEAE system detected 28 positive samples obtained from 48 rabid animals and the BHK system detected 18. By suckling mouse inoculation only 11 of the same positive samples were detected. A total of 90 samples was studied by the three methods. Rabies virus was detected by the tissue culture methods earlier than by suckling mouse inoculation. The BHK-DEAE method was an economic and fast method for rabies virus detection in saliva samples, which could be used for ecological and pathogenesis studies, as well for rabies diagnosis before the death of the suspected animal. PMID:1100655

Fluorescence spectroscopy using excitation and emission matrix for quantification of tissue native fluorophores and cancer diagnosis

NASA Astrophysics Data System (ADS)

Wu, Binlin; Gayen, S. K.; Xu, M.

2014-03-01

Native fluorescence spectrum of normal and cancerous human prostate tissues is studied to distinguish between normal and cancerous tissues, and cancerous tissues at different cancer grade. The tissue samples were obtained from Cooperative Human Tissue Network (CHTN) and National Disease Research Interchange(NDRI). An excitation and emission matrix (EEM) was generated for each tissue sample by acquiring native fluorescence spectrum of the sample using multiple excitation wavelengths. The non-negative matrix factorization algorithm was used to generate fluorescence EEMs that correspond to the fluorophores in biological tissues, including tryptophan, collagen, elastin, nicotinamide adenine dinucleotide (NADH), flavin adenine dinucleotide (FAD) and the background paraffin. We hypothesize that, as a consequence of metabolic changes associated with the development of cancer, the concentrations of NADH and FAD are different in normal and cancerous tissues, and also different for different cancer grades. We used the ratio of the abundances of FAD and NADH to distinguish between normal and cancerous tissues, and the tissue cancer grade. The FAD-to-NADH ratio was found to be the highest for normal tissue and decreased as the cancer grade increased.

Characterization of human breast cancer tissues by infrared imaging.

PubMed

Verdonck, M; Denayer, A; Delvaux, B; Garaud, S; De Wind, R; Desmedt, C; Sotiriou, C; Willard-Gallo, K; Goormaghtigh, E

2016-01-21

Fourier Transform InfraRed (FTIR) spectroscopy coupled to microscopy (IR imaging) has shown unique advantages in detecting morphological and molecular pathologic alterations in biological tissues. The aim of this study was to evaluate the potential of IR imaging as a diagnostic tool to identify characteristics of breast epithelial cells and the stroma. In this study a total of 19 breast tissue samples were obtained from 13 patients. For 6 of the patients, we also obtained Non-Adjacent Non-Tumor tissue samples. Infrared images were recorded on the main cell/tissue types identified in all breast tissue samples. Unsupervised Principal Component Analyses and supervised Partial Least Square Discriminant Analyses (PLS-DA) were used to discriminate spectra. Leave-one-out cross-validation was used to evaluate the performance of PLS-DA models. Our results show that IR imaging coupled with PLS-DA can efficiently identify the main cell types present in FFPE breast tissue sections, i.e. epithelial cells, lymphocytes, connective tissue, vascular tissue and erythrocytes. A second PLS-DA model could distinguish normal and tumor breast epithelial cells in the breast tissue sections. A patient-specific model reached particularly high sensitivity, specificity and MCC rates. Finally, we showed that the stroma located close or at distance from the tumor exhibits distinct spectral characteristics. In conclusion FTIR imaging combined with computational algorithms could be an accurate, rapid and objective tool to identify/quantify breast epithelial cells and differentiate tumor from normal breast tissue as well as normal from tumor-associated stroma, paving the way to the establishment of a potential complementary tool to ensure safe tumor margins.

Cell and Tissue Microarray Technologies for Protein and Nucleic Acid Expression Profiling

PubMed Central

Cardano, Marina; Diaferia, Giuseppe R.; Falavigna, Maurizio; Spinelli, Chiara C.; Sessa, Fausto; DeBlasio, Pasquale

2013-01-01

Tissue microarray (TMA) and cell microarray (CMA) are two powerful techniques that allow for the immunophenotypical characterization of hundreds of samples simultaneously. In particular, the CMA approach is particularly useful for immunophenotyping new stem cell lines (e.g., cardiac, neural, mesenchymal) using conventional markers, as well as for testing the specificity and the efficacy of newly developed antibodies. We propose the use of a tissue arrayer not only to perform protein expression profiling by immunohistochemistry but also to carry out molecular genetics studies. In fact, starting with several tissues or cell lines, it is possible to obtain the complete signature of each sample, describing the protein, mRNA and microRNA expression, and DNA mutations, or eventually to analyze the epigenetic processes that control protein regulation. Here we show the results obtained using the Galileo CK4500 TMA platform. PMID:23172795

Optical coefficient measurements using bulk living tissue by an optical fiber puncture with FOV change

NASA Astrophysics Data System (ADS)

Nakazawa, Haruna; Doi, Marika; Ogawa, Emiyu; Arai, Tsunenori

2018-02-01

To avoid an instability of the optical coefficient measurement using sliced tissue preparation, we proposed the combination of light intensity measurement through an optical fiber puncturing into a bulk tissue varying field of view (FOV) and ray tracing calculation using Monte-Carlo method. The optical coefficients of myocardium such as absorption coefficient μa, scattering coefficient μs, and anisotropic parameter g are used in the myocardium optical propagation. Since optical coefficients obtained using thin sliced tissue could be instable because they are affected by dehydration and intracellular fluid effusion on the sample surface, variety of coefficients have been reported over individual optical differences of living samples. The proposed method which combined the experiment using the bulk tissue with ray tracing calculation were performed. In this method, a 200 μmΦ high-NA silica fiber installed in a 21G needle was punctured up to the bottom of the myocardial bulk tissue over 3 cm in thickness to measure light intensity changing the fiber-tip depth and FOV. We found that the measured attenuation coefficients decreased as the FOV increased. The ray trace calculation represented the same FOV dependence in above mentioned experimental result. We think our particular fiber punctured measurement using bulk tissue varying FOV with Inverse Monte-Carlo method might be useful to obtain the optical coefficients to avoid sample preparation instabilities.

Investigation of formalin influence over hard and soft biological tissues fluorescent spectra in vitro

NASA Astrophysics Data System (ADS)

Borisova, E.; Uzunov, Tz.; Vladimirov, B.; Avramov, L.

2007-05-01

In order to investigate the formalin influence over fluorescence properties of hard and soft biological tissues during conservation, emission spectra have been registered. Nitrogen laser at 337 nm and light-emitting diode with maximum at 405 nm have been used as excitation sources. For investigation of formalin influence over hard tissues, an experiment was made on teeth samples. Sound teeth were demineralized with a phosphoric acid for 10 seconds to obtain enamel structure near to the tooth lesion, and were fixed in formalin. Before and after teeth treatment spectra from the areas of interest were detected. There were not observed changes in the shape of the teeth spectra, related to the introduction of formalin fluorescence. Samples from mucosa of esophagus and stomach, where initially an ALA/Protoporphyrin IX diagnosis was applied, were used as soft tissue specimens. After fluorescent diagnosis in vivo biopsy samples were obtained from normal and cancerous areas and were conserved in formalin. Initially, spectrum observed has one autofluorescence maximum from the mucous tissue at 500-600 nm and secondary maxima from the protoporphyrin fluorescence at 635 nm and 720 nm, as well as pronounced minima at 540 and 575 nm related to hemoglobin absorption. After formalin conservation hemoglobin absorption was strongly reduced that increases mucous emission signal in green-yellow spectral region. Simultaneously the maxima at 635 nm and 720 nm were reduced. As conclusion we could say that formalin has negligible influence over fluorescence spectra of conserved hard tissues and has more pronounced influence over fluorescence spectra obtained in the case of soft tissue conservation, which has to be taking into account in measurements in vitro.

Colon Cancer Associated Transcript-1 (CCAT1) Expression in Adenocarcinoma of the Stomach

PubMed Central

Mizrahi, Ido; Mazeh, Haggi; Grinbaum, Ronit; Beglaibter, Nahum; Wilschanski, Michael; Pavlov, Vera; Adileh, Muchamad; Stojadinovic, Alexander; Avital, Itzhak; Gure, Ali Osmay; Halle, David; Nissan, Aviram

2015-01-01

Background: Long non-coding RNAs (lncRNAs) have been shown to have functional roles in cancer biology and are dys-regulated in many tumors. Colon Cancer Associated Transcript -1 (CCAT1) is a lncRNA, previously shown to be significantly up-regulated in colon cancer. The aim of this study is to determine expression levels of CCAT1 in gastric carcinoma (GC). Methods: Tissue samples were obtained from patients undergoing resection for gastric carcinoma (n=19). For each patient, tumor tissue and normal appearing gastric mucosa were taken. Normal gastric tissues obtained from morbidly obese patients, undergoing laparoscopic sleeve gastrectomy served as normal controls (n=19). A human gastric carcinoma cell line (AGS) served as positive control. RNA was extracted from all tissue samples and CCAT1 expression was analyzed using quantitative real time-PCR (qRT-PCR). Results: Low expression of CCAT1 was identified in normal gastric mucosa samples obtained from morbidly obese patients [mean Relative Quantity (RQ) = 1.95±0.4]. AGS human gastric carcinoma cell line showed an elevated level of CCAT1 expression (RQ=8.02). Expression levels of CCAT1 were approximately 10.8 fold higher in GC samples than in samples taken from the negative control group (RQ=21.1±5 vs. RQ=1.95±0.4, respectively, p<0.001). Interestingly, CCAT1 expression was significantly overexpressed in adjacent normal tissues when compared to the negative control group (RQ = 15.25±2 vs. RQ=1.95±0.4, respectively, p<0.001). Tissues obtained from recurrent GC cases showed the highest expression levels (RQ = 88.8±31; p<0.001). Expression levels increased with tumor stage (T4- 36.4±15, T3- 16.1±6, T2- 4.7±1), however this did not reach statistical significance (p=0.2). There was no difference in CCAT1 expression between intestinal and diffuse type GC (RQ=22.4±7 vs. 22.4±16, respectively, p=0.9). Within the normal gastric tissue samples, no significant difference in CCAT1 expression was observed in helicobacter pylori negative and positive patients (RQ= 2.4±0.9 vs. 0.93±0.2, respectively, p=0.13). Conclusion: CCAT1 is up-regulated in gastric cancer, and may serve as a potential bio-marker for early detection and surveillance. PMID:25561974

Histology of periapical lesions obtained during apical surgery.

PubMed

Schulz, Malte; von Arx, Thomas; Altermatt, Hans Jörg; Bosshardt, Dieter

2009-05-01

The aim of this was to evaluate the histology of periapical lesions in teeth treated with periapical surgery. After root-end resection, the root tip was removed together with the periapical pathological tissue. Histologic sectioning was performed on calcified specimens embedded in methylmethacrylate (MMA) and on demineralized specimens embedded in LR White (Fluka, Buchs, Switzerland). The samples were evaluated with light and transmission electron microscopy (TEM). The histologic findings were classified into periapical abscesses, granulomas, or cystic lesions (true or pocket cysts). The final material comprised 70% granulomas, 23% cysts and 5% abscesses, 1% scar tissues, and 1% keratocysts. Six of 125 samples could not be used. The cystic lesions could not be subdivided into pocket or true cysts. All cysts had an epithelium-lined cavity, two of them with cilia-lined epithelium. These results show the high incidence of periapical granulomas among periapical lesions obtained during apical surgery. Periapical abscesses were a rare occasion. The histologic findings from samples obtained during apical surgery may differ from findings obtained by teeth extractions. A determination between pocket and true apical cysts is hardly possible when collecting samples by apical surgery.

Expression profiling of microRNAs in human bone tissue from postmenopausal women.

PubMed

De-Ugarte, Laura; Serra-Vinardell, Jenny; Nonell, Lara; Balcells, Susana; Arnal, Magdalena; Nogues, Xavier; Mellibovsky, Leonardo; Grinberg, Daniel; Diez-Perez, Adolfo; Garcia-Giralt, Natalia

2018-01-01

Bone tissue is composed of several cell types, which express their own microRNAs (miRNAs) that will play a role in cell function. The set of total miRNAs expressed in all cell types configures the specific signature of the bone tissue in one physiological condition. The aim of this study was to explore the miRNA expression profile of bone tissue from postmenopausal women. Tissue was obtained from trabecular bone and was analyzed in fresh conditions (n = 6). Primary osteoblasts were also obtained from trabecular bone (n = 4) and human osteoclasts were obtained from monocyte precursors after in vitro differentiation (n = 5). MicroRNA expression profiling was obtained for each sample by microarray and a global miRNA analysis was performed combining the data acquired in all the microarray experiments. From the 641 miRNAs detected in bone tissue samples, 346 (54%) were present in osteoblasts and/or osteoclasts. The other 46% were not identified in any of the bone cells analyzed. Intersection of osteoblast and osteoclast arrays identified 101 miRNAs shared by both cell types, which accounts for 30-40% of miRNAs detected in these cells. In osteoblasts, 266 miRNAs were detected, of which 243 (91%) were also present in the total bone array, representing 38% of all bone miRNAs. In osteoclasts, 340 miRNAs were detected, of which 196 (58%) were also present in the bone tissue array, representing 31% of all miRNAs detected in total bone. These analyses provide an overview of miRNAs expressed in bone tissue, broadening our knowledge in the microRNA field.

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

DOE Office of Scientific and Technical Information (OSTI.GOV)

Van Berkel, Gary J; Kertesz, Vilmos; Koeplinger, Kenneth A.

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 bodymore » 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.« less

Characterisation of signal enhancements achieved when utilizing a photon diode in deep Raman spectroscopy of tissue

PubMed Central

Vardaki, Martha Z.; Matousek, Pavel; Stone, Nicholas

2016-01-01

We characterise the performance of a beam enhancing element (‘photon diode’) for use in deep Raman spectroscopy (DRS) of biological tissues. The optical component enhances the number of laser photons coupled into a tissue sample by returning escaping photons back into it at the illumination zone. The method is compatible with transmission Raman spectroscopy, a deep Raman spectroscopy concept, and its implementation leads to considerable enhancement of detected Raman photon rates. In the past, the enhancement concept was demonstrated with a variety of samples (pharmaceutical tablets, tissue, etc) but it was not systematically characterized with biological tissues. In this study, we investigate the enhancing properties of the photon diode in the transmission Raman geometry as a function of: a) the depth and b) the optical properties of tissue samples. Liquid tissue phantoms were employed to facilitate systematic variation of optical properties. These were chosen to mimic optical properties of human tissues, including breast and prostate. The obtained results evidence that a photon diode can enhance Raman signals of tissues by a maximum of × 2.4, although it can also decrease the signals created towards the back of samples that exhibit high scattering or absorption properties. PMID:27375932

Wavelet analysis in two-dimensional tomography

NASA Astrophysics Data System (ADS)

Burkovets, Dimitry N.

2002-02-01

The diagnostic possibilities of wavelet-analysis of coherent images of connective tissue in its pathological changes diagnostics. The effectiveness of polarization selection in obtaining wavelet-coefficients' images is also shown. The wavelet structures, characterizing the process of skin psoriasis, bone-tissue osteoporosis have been analyzed. The histological sections of physiological normal and pathologically changed samples of connective tissue of human skin and spongy bone tissue have been analyzed.

SPRUCE Pretreatment Plant Tissue Analyses, 2009 through 2013

DOE Data Explorer

Phillips, J. R. [Oak Ridge National Laboratory, U.S. Department of Energy, Oak Ridge, Tennessee, U.S.A; Brice, D. J. [Oak Ridge National Laboratory, U.S. Department of Energy, Oak Ridge, Tennessee, U.S.A; Hanson, P. J. [Oak Ridge National Laboratory, U.S. Department of Energy, Oak Ridge, Tennessee, U.S.A; Childs, J. [Oak Ridge National Laboratory, U.S. Department of Energy, Oak Ridge, Tennessee, U.S.A; Iversen, C. M. [Oak Ridge National Laboratory, U.S. Department of Energy, Oak Ridge, Tennessee, U.S.A; Norby, R. J. [Oak Ridge National Laboratory, U.S. Department of Energy, Oak Ridge, Tennessee, U.S.A; Warren, J. M. [Oak Ridge National Laboratory, U.S. Department of Energy, Oak Ridge, Tennessee, U.S.A

2009-12-01

This data set reports the results of elemental analyses of foliar and stem/woody twig plant tissues collected at the SPRUCE site in 2009, 2012, and 2013. Samples were obtained at various locations around the S1 Bog and from within the developing experimental treatment plots. These are pretreatment vegetation samples, collected prior to initiation of the SPRUCE experiment heating and elevated CO2 treatments.

Pre-operative intra-articular deep tissue sampling with novel retrograde forceps improves the diagnostics in periprosthetic joint infection.

PubMed

Wimmer, Matthias D; Ploeger, Milena M; Friedrich, Max J; Hügle, Thomas; Gravius, Sascha; Randau, Thomas M

2017-07-01

Histopathological tissue analysis is a key parameter within the diagnostic algorithm for suspected periprosthetic joint infections (PJIs), conventionally acquired in open surgery. In 2014, Hügle and co-workers introduced novel retrograde forceps for retrograde synovial biopsy with simultaneous fluid aspiration of the knee joint. We hypothesised that tissue samples acquired by retrograde synovial biopsy are equal to intra-operatively acquired deep representative tissue samples regarding bacterial detection and differentiation of periprosthetic infectious membranes. Thirty patients (male n = 15, 50%; female n = 15, 50%) with 30 suspected PJIs in painful total hip arthroplasties (THAs) were included in this prospective, controlled, non-blinded trial. The results were compared with intra-operatively obtained representative deep tissue samples. In summary, 27 out of 30 patients were diagnosed correctly as infected (17/17) or non-infected (10/13). The sensitivity to predict a PJI using the Retroforce® sampling forceps in addition to standard diagnostics was 85%, the specificity 100%. Retrograde synovial biopsy is a new and rapid diagnostic procedure under local anaesthesia in patients with painful THAs with similar histological results compared to deep tissue sampling.

Fabrication and characterization of biological tissue phantoms with embedded nanoparticles

NASA Astrophysics Data System (ADS)

Skaptsov, A. A.; Ustalkov, S. O.; Mohammed, A. H. M.; Savenko, O. A.; Novikova, A. S.; Kozlova, E. A.; Kochubey, V. I.

2017-11-01

Phantoms are imitations of biological tissue, which are used for modelling of the light propagation in biological tissues. Carrying out any biophysical experiments requires an indispensable constancy of the initial experiment conditions. The use of solid undegradable phantoms is the basis to obtain reliable reproducible experimental results. The fabrication of biological tissues phantoms containing high absorbance or fluorescence nanoparticles and corresponding to specific mechanical, optical properties is an actual task. This work describes development, fabrication and characterization of such solid tissue phantoms with embedded CdSe/ZnS quantum dots, gold and upconversion nanoparticles. Luminescence of samples with CdSe/ZnS quantum dots and upconversion nanoparticles were recorded. A sample of gold nanorods was analyzed using thermal gravimetric analysis. It can be concluded that the samples are well suited for experiments on laser thermolysis.

[GSTP1, APC and RASSF1 gene methylation in prostate cancer samples: comparative analysis of MS-HRM method and Infinium HumanMethylation450 BeadChip beadchiparray diagnostic value].

PubMed

Skorodumova, L O; Babalyan, K A; Sultanov, R; Vasiliev, A O; Govorov, A V; Pushkar, D Y; Prilepskaya, E A; Danilenko, S A; Generozov, E V; Larin, A K; Kostryukova, E S; Sharova, E I

2016-11-01

There is a clear need in molecular markers for prostate cancer (PC) risk stratification. Alteration of DNA methylation is one of processes that occur during ÐÑ progression. Methylation-sensitive PCR with high resolution melting curve analysis (MS-HRM) can be used for gene methylation analysis in routine laboratory practice. This method requires very small amounts of DNA for analysis. Numerous results have been accumulated on DNA methylation in PC samples analyzed by the Infinium HumanMethylation450 BeadChip (HM450). However, the consistency of MS-HRM results with chip hybridization results has not been examined yet. The aim of this study was to assess the consistency of results of GSTP1, APC and RASSF1 gene methylation analysis in ÐÑ biopsy samples obtained by MS-HRM and chip hybridization. The methylation levels of each gene determined by MS-HRM were statistically different in the group of PC tissue samples and the samples without signs of tumor growth. Chip hybridization data analysis confirmed the results obtained with the MS-HRM. Differences in methylation levels between tumor tissue and histologically intact tissue of each sample determined by MS-HRM and chip hybridization, were consistent with each other. Thus, we showed that the assessment of GSTP1, APC and RASSF1 gene methylation analysis using MS-HRM is suitable for the design of laboratory assays that will differentiate the PC tissue from the tissue without signs of tumor growth.

Cell block samples from malignant pleural effusion might be valid alternative samples for anaplastic lymphoma kinase detection in patients with advanced non-small-cell lung cancer.

PubMed

Zhou, Jianya; Yao, Hongtian; Zhao, Jing; Zhang, Shumeng; You, Qihan; Sun, Ke; Zou, Yinying; Zhou, Caicun; Zhou, Jianying

2015-06-01

To evaluate the clinical value of cell block samples from malignant pleural effusion (MPE) as alternative samples to tumour tissue for anaplastic lymphoma kinase (ALK) detection in patients with advanced non-small-cell lung cancer (NSCLC). Fifty-two matched samples were eligible for analysis. ALK status was detected by Ventana immunohistochemistry (IHC) (with the D5F3 clone), reverse transcription polymerase chain reaction (RT-PCR) and fluorescence in-situ hybridization (FISH) in MPE cell block samples, and by FISH in tumour tissue block samples. In total, ALK FISH results were obtained for 52 tumour tissue samples and 41 MPE cell block samples. Eight cases (15.4%) were ALK-positive in tumour tissue samples by FISH, and among matched MPE cell block samples, five were ALK-positive by FISH, seven were ALK-positive by RT-PCR, and eight were ALK-positive by Ventana IHC. The ALK status concordance rates between tumour tissue and MPE cell block samples were 78.9% by FISH, 98.1% by RT-PCR, and 100% by Ventana IHC. In MPE cell block samples, the sensitivity and specificity of Ventana IHC (100% and 100%) and RT-PCR (87.5% and 100%) were higher than those of FISH (62.5% and 100%). Malignant pleural effusion cell block samples had a diagnostic performance for ALK detection in advanced NSCLC that was comparable to that of tumour tissue samples. MPE cell block samples might be valid alternative samples for ALK detection when tissue is not available. Ventana IHC could be the most suitable method for ALK detection in MPE cell block samples. © 2014 John Wiley & Sons Ltd.

A novel method of genomic DNA extraction for Cactaceae1

PubMed Central

Fehlberg, Shannon D.; Allen, Jessica M.; Church, Kathleen

2013-01-01

• Premise of the study: Genetic studies of Cactaceae can at times be impeded by difficult sampling logistics and/or high mucilage content in tissues. Simplifying sampling and DNA isolation through the use of cactus spines has not previously been investigated. • Methods and Results: Several protocols for extracting DNA from spines were tested and modified to maximize yield, amplification, and sequencing. Sampling of and extraction from spines resulted in a simplified protocol overall and complete avoidance of mucilage as compared to typical tissue extractions. Sequences from one nuclear and three plastid regions were obtained across eight genera and 20 species of cacti using DNA extracted from spines. • Conclusions: Genomic DNA useful for amplification and sequencing can be obtained from cactus spines. The protocols described here are valuable for any cactus species, but are particularly useful for investigators interested in sampling living collections, extensive field sampling, and/or conservation genetic studies. PMID:25202521

Flexible cystoscopic bladder biopsies: a technique for outpatient evaluation of the lower urinary tract urothelium.

PubMed

Beaghler, M; Grasso, M

1994-11-01

Routine urothelial biopsies of the lower urinary tract are obtained using the cold cup biopsy technique. This procedure is most often performed in the surgical suite and requires rigid endoscopic access and the use of biopsy forceps and Bugbee electrodes to obtain tissue for histologic examination. A new single-step biopsy forceps has been used through the flexible cystoscope. Using a 16 F actively deflectable, flexible cystoscope and the 5.4 F Therma Jaw Hot Urologic Forceps, bladder biopsies were obtained in 27 patients for a variety of indications. This biopsy forceps allows simultaneous tissue sampling and electrocoagulation of the biopsy site, thus eliminating the need for exchange of instruments through the flexible cystoscope. Tissue samples are somewhat protected from thermal changes during coagulation through the use of a Faraday cage. Biopsies were frequently obtained in an outpatient setting, requiring only local topical anesthesia (2% lidocaine jelly). Carcinoma in situ, transitional cell carcinoma, acute and chronic inflammation, and normal bladder mucosa were differentiated histologically. Using this technique, lower urinary tract urothelial mapping can be performed safely in the office with minimal patient discomfort.

Optimal molecular profiling of tissue and tissue components: defining the best processing and microdissection methods for biomedical applications.

PubMed

Bova, G Steven; Eltoum, Isam A; Kiernan, John A; Siegal, Gene P; Frost, Andra R; Best, Carolyn J M; Gillespie, John W; Emmert-Buck, Michael R

2005-01-01

Isolation of well-preserved pure cell populations is a prerequisite for sound studies of the molecular basis of pancreatic malignancy and other biological phenomena. This chapter reviews current methods for obtaining anatomically specific signals from molecules isolated from tissues, a basic requirement for productive linking of phenotype and genotype. The quality of samples isolated from tissue and used for molecular analysis is often glossed-over or omitted from publications, making interpretation and replication of data difficult or impossible. Fortunately, recently developed techniques allow life scientists to better document and control the quality of samples used for a given assay, creating a foundation for improvement in this area. Tissue processing for molecular studies usually involves some or all of the following steps: tissue collection, gross dissection/identification, fixation, processing/embedding, storage/archiving, sectioning, staining, microdissection/annotation, and pure analyte labeling/identification. High-quality tissue microdissection does not necessarily mean high-quality samples to analyze. The quality of biomaterials obtained for analysis is highly dependent on steps upstream and downstream from tissue microdissection. We provide protocols for each of these steps, and encourage you to improve upon these. It is worth the effort of every laboratory to optimize and document its technique at each stage of the process, and we provide a starting point for those willing to spend the time to optimize. In our view, poor documentation of tissue and cell type of origin and the use of nonoptimized protocols is a source of inefficiency in current life science research. Even incremental improvement in this area will increase productivity significantly.

Comparison of inferred fractions of n-3 and n-6 polyunsaturated fatty acids in feral domestic cat diets with those in commercial feline extruded diets.

PubMed

Backus, Robert C; Thomas, David G; Fritsche, Kevin L

2013-04-01

To compare presumed fatty acid content in natural diets of feral domestic cats (inferred from body fat polyunsatrated fatty acids content) with polyunsaturated fatty acid content of commercial feline extruded diets. Subcutaneous and intra-abdominal adipose tissue samples (approx 1 g) from previously frozen cadavers of 7 adult feral domestic cats trapped in habitats remote from human activity and triplicate samples (200 g each) of 7 commercial extruded diets representing 68% of market share obtained from retail stores. Lipid, triacylglycerol, and phospholipid fractions in adipose tissue samples and ether extracts of diet samples were determined by gas chromatography of methyl esters. Triacylglycerol and phospholipid fractions in the adipose tissue were isolated by thin-layer chromatography. Diet samples were also analyzed for proximate contents. For the adipose tissue samples, with few exceptions, fatty acids fractions varied only moderately with lipid fraction and site from which tissue samples were obtained. Linoleic, α-linolenic, arachidonic, eicosapentaenoic, and docosahexaenoic acid fractions were 15.0% to 28.2%, 4.5% to 18.7%, 0.9% to 5.0%, < 0.1% to 0.2%, and 0.6% to 1.7%, respectively. As inferred from the adipose findings, dietary fractions of docosahexaenoic and α-linolenic acid were significantly greater than those in the commercial feline diets, but those for linoleic and eicosapentaenoic acids were not significantly different. The fatty acid content of commercial extruded feline diets differed from the inferred content of natural feral cat diets, in which dietary n-3 and possibly n-6 polyunsaturated fatty acids were more abundant. The impact of this difference on the health of pet cats is not known.

Ex vivo validation of photo-magnetic imaging.

PubMed

Luk, Alex; Nouizi, Farouk; Erkol, Hakan; Unlu, Mehmet B; Gulsen, Gultekin

2017-10-15

We recently introduced a new high-resolution diffuse optical imaging technique termed photo-magnetic imaging (PMI), which utilizes magnetic resonance thermometry (MRT) to monitor the 3D temperature distribution induced in a medium illuminated with a near-infrared light. The spatiotemporal temperature distribution due to light absorption can be accurately estimated using a combined photon propagation and heat diffusion model. High-resolution optical absorption images are then obtained by iteratively minimizing the error between the measured and modeled temperature distributions. We have previously demonstrated the feasibility of PMI with experimental studies using tissue simulating agarose phantoms. In this Letter, we present the preliminary ex vivo PMI results obtained with a chicken breast sample. Similarly to the results obtained on phantoms, the reconstructed images reveal that PMI can quantitatively resolve an inclusion with a 3 mm diameter embedded deep in a biological tissue sample with only 10% error. These encouraging results demonstrate the high performance of PMI in ex vivo biological tissue and its potential for in vivo imaging.

Comparison of premortem and postmortem estimates of plutonium deposited in the skeleton and liver of six individuals

DOE Office of Scientific and Technical Information (OSTI.GOV)

Sula, M.J.; Bihl, D.E.; Carbaugh, E.H.

1988-04-01

Assessment of organ burdens after internal exposures to radionuclides is often necessary to evaluate the health and regulatory implications of the exposure. The assessment of plutonium activity in skeleton and liver is usually estimated from measurements of plutonium excreted via urine. As part of the overall evaluation of internal dose assessment techniques, it is useful to compare the results of organ burden estimates made from evaluation of urinary excretion data with those made at death from tissue samples collected posthumously from the individual. Estimates of plutonium in the skeleton and liver, based on postmortem analysis of tissue samples for sixmore » individuals, were obtained from the US Transuranium Registry (USTR). Bioassay data and other radiation exposure information obtained from the individuals' files were used to estimate their skeleton and liver burdens at the times of their deaths, and these estimates were compared to those obtained through tissue analysis. 6 refs., 2 tabs.« less

Direct-to-PCR tissue preservation for DNA profiling.

PubMed

Sorensen, Amy; Berry, Clare; Bruce, David; Gahan, Michelle Elizabeth; Hughes-Stamm, Sheree; McNevin, Dennis

2016-05-01

Disaster victim identification (DVI) often occurs in remote locations with extremes of temperatures and humidities. Access to mortuary facilities and refrigeration are not always available. An effective and robust DNA sampling and preservation procedure would increase the probability of successful DNA profiling and allow faster repatriation of bodies and body parts. If the act of tissue preservation also released DNA into solution, ready for polymerase chain reaction (PCR), the DVI process could be further streamlined. In this study, we explored the possibility of obtaining DNA profiles without DNA extraction, by adding aliquots of preservative solutions surrounding fresh human muscle and decomposing human muscle and skin tissue samples directly to PCR. The preservatives consisted of two custom preparations and two proprietary solutions. The custom preparations were a salt-saturated solution of dimethyl sulfoxide (DMSO) with ethylenediaminetetraacetic (EDTA) and TENT buffer (Tris, EDTA, NaCl, Tween 20). The proprietary preservatives were DNAgard (Biomatrica(®)) and Tissue Stabilising Kit (DNA Genotek). We obtained full PowerPlex(®) 21 (Promega) and GlobalFiler(®) (Life Technologies) DNA profiles from fresh and decomposed tissue preserved at 35 °C for up to 28 days for all four preservatives. The preservative aliquots removed from the fresh muscle tissue samples had been stored at -80 °C for 4 years, indicating that long-term archival does not diminish the probability of successful DNA typing. Rather, storage at -80 °C seems to reduce PCR inhibition.

Determination of the complex refractive index segments of turbid sample with multispectral spatially modulated structured light and models approximation

NASA Astrophysics Data System (ADS)

Meitav, Omri; Shaul, Oren; Abookasis, David

2017-09-01

Spectral data enabling the derivation of a biological tissue sample's complex refractive index (CRI) can provide a range of valuable information in the clinical and research contexts. Specifically, changes in the CRI reflect alterations in tissue morphology and chemical composition, enabling its use as an optical marker during diagnosis and treatment. In the present work, we report a method for estimating the real and imaginary parts of the CRI of a biological sample using Kramers-Kronig (KK) relations in the spatial frequency domain. In this method, phase-shifted sinusoidal patterns at single high spatial frequency are serially projected onto the sample surface at different near-infrared wavelengths while a camera mounted normal to the sample surface acquires the reflected diffuse light. In the offline analysis pipeline, recorded images at each wavelength are converted to spatial phase maps using KK analysis and are then calibrated against phase-models derived from diffusion approximation. The amplitude of the reflected light, together with phase data, is then introduced into Fresnel equations to resolve both real and imaginary segments of the CRI at each wavelength. The technique was validated in tissue-mimicking phantoms with known optical parameters and in mouse models of ischemic injury and heat stress. Experimental data obtained indicate variations in the CRI among brain tissue suffering from injury. CRI fluctuations correlated with alterations in the scattering and absorption coefficients of the injured tissue are demonstrated. This technique for deriving dynamic changes in the CRI of tissue may be further developed as a clinical diagnostic tool and for biomedical research applications. To the best of our knowledge, this is the first report of the estimation of the spectral CRI of a mouse head following injury obtained in the spatial frequency domain.

Scanless nonlinear optical microscope for image reconstruction and space-time correlation analysis

NASA Astrophysics Data System (ADS)

Ceffa, N. G.; Radaelli, F.; Pozzi, P.; Collini, M.; Sironi, L.; D'alfonso, L.; Chirico, G.

2017-06-01

Optical Microscopy has been applied to life science from its birth and reached widespread application due to its major advantages: limited perturbation of the biological tissue and the easy accessibility of the light sources. However, as the spatial and time resolution requirements and the time stability of the microscopes increase, researchers are struggling against some of its limitations: limited transparency and the refractivity of the living tissue to light and the field perturbations induced by the path in the tissue. We have developed a compact stand-alone, completely scan-less, optical setup that allows to acquire non-linear excitation images and to measure the sample dynamics simultaneously on an ensemble of arbitrary chosen regions of interests. The image is obtained by shining a square array of spots on the sample obtained by a spatial light modulator and by shifting it (10 ms refresh time) on the sample. The final image is computed from the superposition of (100-1000) images. Filtering procedures can be applied to the raw images of the excitation array before building the image. We discuss results that show how this setup can be used for the correction of wave front aberrations induced by turbid samples (such as living tissues) and for the computation of space-time cross-correlations in complex networks.

Preventing disease transmission by deceased tissue donors by testing blood for viral nucleic acid.

PubMed

Strong, D Michael; Nelson, Karen; Pierce, Marge; Stramer, Susan L

2005-01-01

Nucleic acid testing (NAT) has reduced the risk of transmitting infectious disease through blood transfusion. Currently NAT for HIV-1 and HCV are FDA licensed and performed by nearly all blood collection facilities, but HBV NAT is performed under an investigational study protocol. Residual risk estimates indicate that NAT could potentially reduce disease transmission through transplanted tissue. However, tissue donor samples obtained post-mortem have the potential to produce an invalid NAT result due to inhibition of amplification reactions by hemolysis and other factors. The studies reported here summarize the development of protocols to allow NAT of deceased donor samples with reduced rates of invalid results. Using these protocols, inventories from two tissue centers were tested with greater than 99% of samples producing a valid test result.

Pulsed terahertz imaging of breast cancer in freshly excised murine tumors

NASA Astrophysics Data System (ADS)

Bowman, Tyler; Chavez, Tanny; Khan, Kamrul; Wu, Jingxian; Chakraborty, Avishek; Rajaram, Narasimhan; Bailey, Keith; El-Shenawee, Magda

2018-02-01

This paper investigates terahertz (THz) imaging and classification of freshly excised murine xenograft breast cancer tumors. These tumors are grown via injection of E0771 breast adenocarcinoma cells into the flank of mice maintained on high-fat diet. Within 1 h of excision, the tumor and adjacent tissues are imaged using a pulsed THz system in the reflection mode. The THz images are classified using a statistical Bayesian mixture model with unsupervised and supervised approaches. Correlation with digitized pathology images is conducted using classification images assigned by a modal class decision rule. The corresponding receiver operating characteristic curves are obtained based on the classification results. A total of 13 tumor samples obtained from 9 tumors are investigated. The results show good correlation of THz images with pathology results in all samples of cancer and fat tissues. For tumor samples of cancer, fat, and muscle tissues, THz images show reasonable correlation with pathology where the primary challenge lies in the overlapping dielectric properties of cancer and muscle tissues. The use of a supervised regression approach shows improvement in the classification images although not consistently in all tissue regions. Advancing THz imaging of breast tumors from mice and the development of accurate statistical models will ultimately progress the technique for the assessment of human breast tumor margins.

Direct microCT imaging of non-mineralized connective tissues at high resolution.

PubMed

Naveh, Gili R S; Brumfeld, Vlad; Dean, Mason; Shahar, Ron; Weiner, Steve

2014-01-01

The 3D imaging of soft tissues in their native state is challenging, especially when high resolution is required. An X-ray-based microCT is, to date, the best choice for high resolution 3D imaging of soft tissues. However, since X-ray attenuation of soft tissues is very low, contrasting enhancement using different staining materials is needed. The staining procedure, which also usually involves tissue fixation, causes unwanted and to some extent unknown tissue alterations. Here, we demonstrate that a method that enables 3D imaging of soft tissues without fixing and staining using an X-ray-based bench-top microCT can be applied to a variety of different tissues. With the sample mounted in a custom-made loading device inside a humidity chamber, we obtained soft tissue contrast and generated 3D images of fresh, soft tissues with a resolution of 1 micron voxel size. We identified three critical conditions which make it possible to image soft tissues: humidified environment, mechanical stabilization of the sample and phase enhancement. We demonstrate the capability of the technique using different specimens: an intervertebral disc, the non-mineralized growth plate, stingray tessellated radials (calcified cartilage) and the collagenous network of the periodontal ligament. Since the scanned specimen is fresh an interesting advantage of this technique is the ability to scan a specimen under load and track the changes of the different structures. This method offers a unique opportunity for obtaining valuable insights into 3D structure-function relationships of soft tissues.

Classification of normal and malignant human gastric mucosa tissue with confocal Raman microspectroscopy and wavelet analysis

NASA Astrophysics Data System (ADS)

Hu, Yaogai; Shen, Aiguo; Jiang, Tao; Ai, Yong; Hu, Jiming

2008-02-01

Thirty-two samples from the human gastric mucosa tissue, including 13 normal and 19 malignant tissue samples were measured by confocal Raman microspectroscopy. The low signal-to-background ratio spectra from human gastric mucosa tissues were obtained by this technique without any sample preparation. Raman spectral interferences include a broad featureless sloping background due to fluorescence and noise. They mask most Raman spectral feature and lead to problems with precision and quantitation of the original spectral information. A preprocessed algorithm based on wavelet analysis was used to reduce noise and eliminate background/baseline of Raman spectra. Comparing preprocessed spectra of malignant gastric mucosa tissues with those of counterpart normal ones, there were obvious spectral changes, including intensity increase at ˜1156 cm -1 and intensity decrease at ˜1587 cm -1. The quantitative criterion based upon the intensity ratio of the ˜1156 and ˜1587 cm -1 was extracted for classification of the normal and malignant gastric mucosa tissue samples. This could result in a new diagnostic method, which would assist the early diagnosis of gastric cancer.

Evaluation of specimen preparation techniques for micro-PIXE localisation of elements in hyperaccumulating plants

NASA Astrophysics Data System (ADS)

Kachenko, Anthony G.; Siegele, Rainer; Bhatia, Naveen P.; Singh, Balwant; Ionescu, Mihail

2008-04-01

Hybanthus floribundus subsp. floribundus, a rare Australian Ni-hyperaccumulating shrub and Pityrogramma calomelanos var. austroamericana, an Australian naturalized As-hyperaccumulating fern are promising species for use in phytoremediation of contaminated sites. Micro-proton-induced X-ray emission (μ-PIXE) spectroscopy was used to map the elemental distribution of the accumulated metal(loid)s, Ca and K in leaf or pinnule tissues of the two plant species. Samples were prepared by two contrasting specimen preparation techniques: freeze-substitution in tetrahydrofuran (THF) and freeze-drying. The specimens were analysed to compare the suitability of each technique in preserving (i) the spatial elemental distribution and (ii) the tissue structure of the specimens. Further, the μ-PIXE results were compared with concentration of elements in the bulk tissue obtained by ICP-AES analysis. In H. floribundus subsp. floribundus, μ-PIXE analysis revealed Ni, Ca and K concentrations in freeze-dried leaf tissues were at par with bulk tissue concentrations. Elemental distribution maps illustrated that Ni was preferentially localised in the adaxial epidermal tissues (1% DW) and least concentration was found in spongy mesophyll tissues (0.53% DW). Conversely, elemental distribution maps of THF freeze-substituted tissues indicated significantly lower Ni, Ca and K concentrations than freeze-dried specimens and bulk tissue concentrations. Moreover, Ni concentrations were uniform across the whole specimen and no localisation was observed. In P. calomelanos var. austroamericana freeze-dried pinnule tissues, μ-PIXE revealed statistically similar As, Ca and K concentrations as compared to bulk tissue concentrations. Elemental distribution maps showed that As localisation was relatively uniform across the whole specimen. Once again, THF freeze-substituted tissues revealed a significant loss of As compared to freeze-dried specimens and the concentrations obtained by bulk tissue analysis. The results demonstrate that freeze-drying is a suitable sample preparation technique to study elemental distribution of ions in H. floribundus and P. calomelanos plant tissues using μ-PIXE spectroscopy. Furthermore, cellular structure was preserved in samples prepared using this technique.

Evaluation of a multiplex real-time PCR assay for detecting pathogens in cardiac valve tissue in patients with endocarditis.

PubMed

Fernández, Angel L; Varela, Eduardo; Martínez, Lucía; Martínez, Amparo; Sierra, Juan; González-Juanatey, José R; Regueiro, Benito

2010-10-01

With a novel real-time multiplex polymerase chain reaction test, the LightCycler SeptiFast® test, 25 bacterial and fungal species can be identified directly in blood. The SeptiFast® test has been used for rapid etiologic diagnosis in infectious endocarditis using blood samples but has not been evaluated directly on cardiac vegetations in patients being treated for infectious endocarditis. We prospectively analyzed 15 samples of heart valve tissue with active infectious endocarditis using the SeptiFast® test and compared the test's sensitivity with that of blood culture, valve tissue culture, and the SeptiFast® test in blood. The sensitivity of the SeptiFast test in heart valve tissue was 100%. The test results confirmed the diagnosis obtained using blood culture in 13 cases and identified the pathogen in 2 cases where blood culture tested negative. The sensitivity of the SeptiFast® test in heart valve tissue was greater than that obtained with conventional culture of vegetations or with the SeptiFast test in blood.

Multiple-wavelength spectroscopic quantitation of light-absorbing species in scattering media

DOEpatents

Nathel, Howard; Cartland, Harry E.; Colston, Jr., Billy W.; Everett, Matthew J.; Roe, Jeffery N.

2000-01-01

An oxygen concentration measurement system for blood hemoglobin comprises a multiple-wavelength low-coherence optical light source that is coupled by single mode fibers through a splitter and combiner and focused on both a target tissue sample and a reference mirror. Reflections from both the reference mirror and from the depths of the target tissue sample are carried back and mixed to produce interference fringes in the splitter and combiner. The reference mirror is set such that the distance traversed in the reference path is the same as the distance traversed into and back from the target tissue sample at some depth in the sample that will provide light attenuation information that is dependent on the oxygen in blood hemoglobin in the target tissue sample. Two wavelengths of light are used to obtain concentrations. The method can be used to measure total hemoglobin concentration [Hb.sub.deoxy +Hb.sub.oxy ] or total blood volume in tissue and in conjunction with oxygen saturation measurements from pulse oximetry can be used to absolutely quantify oxyhemoglobin [HbO.sub.2 ] in tissue. The apparatus and method provide a general means for absolute quantitation of an absorber dispersed in a highly scattering medium.

Multimodal-3D imaging based on μMRI and μCT techniques bridges the gap with histology in visualization of the bone regeneration process.

PubMed

Sinibaldi, R; Conti, A; Sinjari, B; Spadone, S; Pecci, R; Palombo, M; Komlev, V S; Ortore, M G; Tromba, G; Capuani, S; Guidotti, R; De Luca, F; Caputi, S; Traini, T; Della Penna, S

2018-03-01

Bone repair/regeneration is usually investigated through X-ray computed microtomography (μCT) supported by histology of extracted samples, to analyse biomaterial structure and new bone formation processes. Magnetic resonance imaging (μMRI) shows a richer tissue contrast than μCT, despite at lower resolution, and could be combined with μCT in the perspective of conducting non-destructive 3D investigations of bone. A pipeline designed to combine μMRI and μCT images of bone samples is here described and applied on samples of extracted human jawbone core following bone graft. We optimized the coregistration procedure between μCT and μMRI images to avoid bias due to the different resolutions and contrasts. Furthermore, we used an Adaptive Multivariate Clustering, grouping homologous voxels in the coregistered images, to visualize different tissue types within a fused 3D metastructure. The tissue grouping matched the 2D histology applied only on 1 slice, thus extending the histology labelling in 3D. Specifically, in all samples, we could separate and map 2 types of regenerated bone, calcified tissue, soft tissues, and/or fat and marrow space. Remarkably, μMRI and μCT alone were not able to separate the 2 types of regenerated bone. Finally, we computed volumes of each tissue in the 3D metastructures, which might be exploited by quantitative simulation. The 3D metastructure obtained through our pipeline represents a first step to bridge the gap between the quality of information obtained from 2D optical microscopy and the 3D mapping of the bone tissue heterogeneity and could allow researchers and clinicians to non-destructively characterize and follow-up bone regeneration. Copyright © 2017 John Wiley & Sons, Ltd.

Equilibrium Passive Sampling of POP in Lipid-Rich and Lean Fish Tissue: Quality Control Using Performance Reference Compounds.

PubMed

Rusina, Tatsiana P; Carlsson, Pernilla; Vrana, Branislav; Smedes, Foppe

2017-10-03

Passive sampling is widely used to measure levels of contaminants in various environmental matrices, including fish tissue. Equilibrium passive sampling (EPS) of persistent organic pollutants (POP) in fish tissue has been hitherto limited to application in lipid-rich tissue. We tested several exposure methods to extend EPS applicability to lean tissue. Thin-film polydimethylsiloxane (PDMS) passive samplers were exposed statically to intact fillet and fish homogenate and dynamically by rolling with cut fillet cubes. The release of performance reference compounds (PRC) dosed to passive samplers prior to exposure was used to monitor the exchange process. The sampler-tissue exchange was isotropic, and PRC were shown to be good indicators of sampler-tissue equilibration status. The dynamic exposures demonstrated equilibrium attainment in less than 2 days for all three tested fish species, including lean fish containing 1% lipid. Lipid-based concentrations derived from EPS were in good agreement with lipid-normalized concentrations obtained using conventional solvent extraction. The developed in-tissue EPS method is robust and has potential for application in chemical monitoring of biota and bioaccumulation studies.

Laser-induced autofluorescence of oral cavity hard tissues

NASA Astrophysics Data System (ADS)

Borisova, E. G.; Uzunov, Tz. T.; Avramov, L. A.

2007-03-01

In current study oral cavity hard tissues autofluorescence was investigated to obtain more complete picture of their optical properties. As an excitation source nitrogen laser with parameters - 337,1 nm, 14 μJ, 10 Hz (ILGI-503, Russia) was used. In vitro spectra from enamel, dentine, cartilage, spongiosa and cortical part of the periodontal bones were registered using a fiber-optic microspectrometer (PC2000, "Ocean Optics" Inc., USA). Gingival fluorescence was also obtained for comparison of its spectral properties with that of hard oral tissues. Samples are characterized with significant differences of fluorescence properties one to another. It is clearly observed signal from different collagen types and collagen-cross links with maxima at 385, 430 and 480-490 nm. In dentine are observed only two maxima at 440 and 480 nm, related also to collagen structures. In samples of gingival and spongiosa were observed traces of hemoglobin - by its re-absorption at 545 and 575 nm, which distort the fluorescence spectra detected from these anatomic sites. Results, obtained in this study are foreseen to be used for development of algorithms for diagnosis and differentiation of teeth lesions and other problems of oral cavity hard tissues as periodontitis and gingivitis.

Gastric Cancer-Specific Protein Profile Identified Using Endoscopic Biopsy Samples via MALDI Mass Spectrometry

PubMed Central

Kim, Hark Kyun; Reyzer, Michelle L.; Choi, Il Ju; Kim, Chan Gyoo; Kim, Hee Sung; Oshima, Akira; Chertov, Oleg; Colantonio, Simona; Fisher, Robert J.; Allen, Jamie L.; Caprioli, Richard M.; Green, Jeffrey E.

2012-01-01

To date, proteomic analyses on gastrointestinal cancer tissue samples have been performed using surgical specimens only, which are obtained after a diagnosis is made. To determine if a proteomic signature obtained from endoscopic biopsy samples could be found to assist with diagnosis, frozen endoscopic biopsy samples collected from 63 gastric cancer patients and 43 healthy volunteers were analyzed using matrix-assisted laser desorption/ionization (MALDI) mass spectrometry. A statistical classification model was developed to distinguish tumor from normal tissues using half the samples and validated with the other half. A protein profile was discovered consisting of 73 signals that could classify 32 cancer and 22 normal samples in the validation set with high predictive values (positive and negative predictive values for cancer, 96.8% and 91.3%; sensitivity, 93.8%; specificity, 95.5%). Signals overexpressed in tumors were identified as α-defensin-1, α-defensin-2, calgranulin A, and calgranulin B. A protein profile was also found to distinguish pathologic stage Ia (pT1N0M0) samples (n = 10) from more advanced stage (Ib or higher) tumors (n = 48). Thus, protein profiles obtained from endoscopic biopsy samples may be useful in assisting with the diagnosis of gastric cancer and, possibly, in identifying early stage disease. PMID:20557134

DNA Yield From Tissue Samples in Surgical Pathology and Minimum Tissue Requirements for Molecular Testing.

PubMed

Austin, Melissa C; Smith, Christina; Pritchard, Colin C; Tait, Jonathan F

2016-02-01

Complex molecular assays are increasingly used to direct therapy and provide diagnostic and prognostic information but can require relatively large amounts of DNA. To provide data to pathologists to help them assess tissue adequacy and provide prospective guidance on the amount of tissue that should be procured. We used slide-based measurements to establish a relationship between processed tissue volume and DNA yield by A260 from 366 formalin-fixed, paraffin-embedded tissue samples submitted for the 3 most common molecular assays performed in our laboratory (EGFR, KRAS, and BRAF). We determined the average DNA yield per unit of tissue volume, and we used the distribution of DNA yields to calculate the minimum volume of tissue that should yield sufficient DNA 99% of the time. All samples with a volume greater than 8 mm(3) yielded at least 1 μg of DNA, and more than 80% of samples producing less than 1 μg were extracted from less than 4 mm(3) of tissue. Nine square millimeters of tissue should produce more than 1 μg of DNA 99% of the time. We conclude that 2 tissue cores, each 1 cm long and obtained with an 18-gauge needle, will almost always provide enough DNA for complex multigene assays, and our methodology may be readily extrapolated to individual institutional practice.

Expression of interleukins, neuropeptides, and growth hormone receptor and leptin receptor genes in adipose tissue from growing broiler chickens

USDA-ARS?s Scientific Manuscript database

In this study, total RNA was collected from abdominal adipose tissue samples obtained from ten broiler chickens at 3, 4, 5, and 6 weeks of age and prepared for quantitative real-time PCR analysis. Studies of the gene expression of cytokines and associated genes in chicken adipose tissue were initia...

Molecular profiles of pre- and postoperative breast cancer tumours reveal differentially expressed genes.

PubMed

Riis, Margit L H; Lüders, Torben; Markert, Elke K; Haakensen, Vilde D; Nesbakken, Anne-Jorun; Kristensen, Vessela N; Bukholm, Ida R K

2012-01-01

Gene expression studies on breast cancer have generally been performed on tissue obtained at the time of surgery. In this study, we have compared the gene expression profiles in preoperative tissue (core needle biopsies) while tumor is still in its normal milieu to postoperative tissue from the same tumor obtained during surgery. Thirteen patients were included of which eleven had undergone sentinel node diagnosis procedure before operation. Microarray gene expression analysis was performed using total RNA from all the samples. Paired significance analysis of microarrays revealed 228 differently expressed genes, including several early response stress-related genes such as members of the fos and jun families as well as genes of which the expression has previously been associated with cancer. The expression profiles found in the analyses of breast cancer tissue must be evaluated with caution. Different profiles may simply be the result of differences in the surgical trauma and timing of when samples are taken and not necessarily associated with tumor biology.

Molecular Profiles of Pre- and Postoperative Breast Cancer Tumours Reveal Differentially Expressed Genes

PubMed Central

Riis, Margit L. H.; Lüders, Torben; Markert, Elke K.; Haakensen, Vilde D.; Nesbakken, Anne-Jorun; Kristensen, Vessela N.; Bukholm, Ida R. K.

2012-01-01

Gene expression studies on breast cancer have generally been performed on tissue obtained at the time of surgery. In this study, we have compared the gene expression profiles in preoperative tissue (core needle biopsies) while tumor is still in its normal milieu to postoperative tissue from the same tumor obtained during surgery. Thirteen patients were included of which eleven had undergone sentinel node diagnosis procedure before operation. Microarray gene expression analysis was performed using total RNA from all the samples. Paired significance analysis of microarrays revealed 228 differently expressed genes, including several early response stress-related genes such as members of the fos and jun families as well as genes of which the expression has previously been associated with cancer. The expression profiles found in the analyses of breast cancer tissue must be evaluated with caution. Different profiles may simply be the result of differences in the surgical trauma and timing of when samples are taken and not necessarily associated with tumor biology. PMID:23227362

Exome copy number variation detection: Use of a pool of unrelated healthy tissue as reference sample.

PubMed

Wenric, Stephane; Sticca, Tiberio; Caberg, Jean-Hubert; Josse, Claire; Fasquelle, Corinne; Herens, Christian; Jamar, Mauricette; Max, Stéphanie; Gothot, André; Caers, Jo; Bours, Vincent

2017-01-01

An increasing number of bioinformatic tools designed to detect CNVs (copy number variants) in tumor samples based on paired exome data where a matched healthy tissue constitutes the reference have been published in the recent years. The idea of using a pool of unrelated healthy DNA as reference has previously been formulated but not thoroughly validated. As of today, the gold standard for CNV calling is still aCGH but there is an increasing interest in detecting CNVs by exome sequencing. We propose to design a metric allowing the comparison of two CNV profiles, independently of the technique used and assessed the validity of using a pool of unrelated healthy DNA instead of a matched healthy tissue as reference in exome-based CNV detection. We compared the CNV profiles obtained with three different approaches (aCGH, exome sequencing with a matched healthy tissue as reference, exome sequencing with a pool of eight unrelated healthy tissue as reference) on three multiple myeloma samples. We show that the usual analyses performed to compare CNV profiles (deletion/amplification ratios and CNV size distribution) lack in precision when confronted with low LRR values, as they only consider the binary status of each CNV. We show that the metric-based distance constitutes a more accurate comparison of two CNV profiles. Based on these analyses, we conclude that a reliable picture of CNV alterations in multiple myeloma samples can be obtained from whole-exome sequencing in the absence of a matched healthy sample. © 2016 WILEY PERIODICALS, INC.

Lewis x is highly expressed in normal tissues: a comparative immunohistochemical study and literature revision.

PubMed

Croce, María V; Isla-Larrain, Marina; Rabassa, Martín E; Demichelis, Sandra; Colussi, Andrea G; Crespo, Marina; Lacunza, Ezequiel; Segal-Eiras, Amada

2007-01-01

An immunohistochemical analysis was employed to determine the expression of carbohydrate antigens associated to mucins in normal epithelia. Tissue samples were obtained as biopsies from normal breast (18), colon (35) and oral cavity mucosa (8). The following carbohydrate epitopes were studied: sialyl-Lewis x, Lewis x, Lewis y, Tn hapten, sialyl-Tn and Thomsen-Friedenreich antigen. Mucins were also studied employing antibodies against MUC1, MUC2, MUC4, MUC5AC, MUC6 and also normal colonic glycolipid. Statistical analysis was performed and Kendall correlations were obtained. Lewis x showed an apical pattern mainly at plasma membrane, although cytoplasmic staining was also found in most samples. TF, Tn and sTn haptens were detected in few specimens, while sLewis x was found in oral mucosa and breast tissue. Also, normal breast expressed MUC1 at a high percentage, whereas MUC4 was observed in a small number of samples. Colon specimens mainly expressed MUC2 and MUC1, while most oral mucosa samples expressed MUC4 and MUC1. A positive correlation between MUC1VNTR and TF epitope (r=0.396) was found in breast samples, while in colon specimens MUC2 and colonic glycolipid versus Lewis x were statistically significantly correlated (r=0.28 and r=0.29, respectively). As a conclusion, a defined carbohydrate epitope expression is not exclusive of normal tissue or a determined localization, and it is possible to assume that different glycoproteins and glycolipids may be carriers of carbohydrate antigens depending on the tissue localization considered.

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.

PubMed

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

2015-08-01

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 spectrometric detection. Excellent correlation was found between the protein distribution data obtained with this method and 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 with non-secreting adenoma and neurohypophysis. AVP was mostly detected in normal neurohypophysis, as expected. This work reveals 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, including AVP and ACTH, directly from human tissue. In addition, the relative simplicity, rapidity, and specificity of this method support the potential of this basic technology, with further advancement, for assisting surgical decision-making. Graphical Abstract Mass spectrometry based profiling of hormones in human pituitary gland and tumor thin tissue sections.

Assessment of tissue-specific cortisol activity with regard to degeneration of the suspensory ligaments in horses with pituitary pars intermedia dysfunction.

PubMed

Hofberger, Sina C; Gauff, Felicia; Thaller, Denise; Morgan, Ruth; Keen, John A; Licka, Theresia F

2018-02-01

OBJECTIVE To identify signs of tissue-specific cortisol activity in samples of suspensory ligament (SL) and neck skin tissue from horses with and without pituitary pars intermedia dysfunction (PPID). SAMPLE Suspensory ligament and neck skin tissue samples obtained from 26 euthanized horses with and without PPID. PROCEDURES Tissue samples were collected from 12 horses with and 14 horses without PPID (controls). Two control horses had received treatment with dexamethasone; data from those horses were not used in statistical analyses. The other 12 control horses were classified as old horses (≥ 14 years old) and young horses (≤ 9 years old). Standard histologic staining, staining for proteoglycan accumulation, and immunostaining of SL and neck skin tissue sections for glucocorticoid receptors, insulin, 11β hydroxysteroid dehydrogenase type 1, and 11β hydroxysteroid dehydrogenase type 2 were performed. Findings for horses with PPID were compared with findings for young and old horses without PPID. RESULTS Compared with findings for old and young control horses, there were significantly more cells stained for glucocorticoid receptors in SL samples and for 11 β hydroxysteroid dehydrogenase type 1 in SL and skin tissue samples from horses with PPID. Insulin could not be detected in any of the SL or skin tissue samples. Horses with PPID had evidence of SL degeneration with significantly increased proteoglycan accumulation. Neck skin tissue was found to be significantly thinner in PPID-affected horses than in young control horses. CONCLUSIONS AND CLINICAL RELEVANCE Results suggested that tissue-specific dysregulation of cortisol metabolism may contribute to the SL degeneration associated with PPID in horses.

Impact of Collection and Storage of Lung Tumor Tissue on Whole Genome Expression Profiling

PubMed Central

Freidin, Maxim B.; Bhudia, Neesa; Lim, Eric; Nicholson, Andrew G.; Cookson, William O.; Moffatt, Miriam F.

2012-01-01

Gene expression profiling could assist in revealing biomarkers of lung cancer prognosis and progression. The handling of biological samples may strongly influence global gene expression, a fact that has not been addressed in many studies. We sought to investigate the changes in gene expression that may occur as a result of sample processing time and conditions. Using Illumina Human WG-6 arrays, we quantified gene expression in lung carcinoma samples from six patients obtained at chest opening before and immediately after lung resection with storage in RNAlater [T1a(CO) and T1b(LR)], after receipt of the sample for histopathology, placed in RNAlater [T2a(HP)]; snap frozen [T2b(HP.SF)]; or snap frozen and stored for 1 week [T2c(HP.SFA)], as well as formalin-fixed, paraffin-embedded (FFPE) block samples. Sampling immediately after resection closely represented the tissue obtained in situ, with only 1% of genes differing more than twofold [T1a(CO) versus T1b(LR)]. Delaying tissue harvest for an average of 30 minutes from the operating theater had a significant impact on gene expression, with approximately 25% of genes differing between T1a(CO) and T2a(HP). Many genes previously identified as lung cancer biomarkers were altered during this period. Examination of FFPE specimens showed minimal correlation with fresh samples. This study shows that tissue collection immediately after lung resection with conservation in RNAlater is an optimal strategy for gene expression profiling. PMID:22240448

Core needle biopsy of soft tissue tumors, CEUS vs US guided: a pilot study.

PubMed

Coran, Alessandro; Di Maggio, Antonio; Rastrelli, Marco; Alberioli, Enrico; Attar, Shady; Ortolan, Paolo; Bortolanza, Carlo; Tosi, Annalisa; Montesco, Maria Cristina; Bezzon, Elisabetta; Rossi, Carlo Riccardo; Stramare, Roberto

2015-12-01

The purpose of this study was to evaluate the usefulness of contrast-enhanced ultrasonography (CEUS) in the bioptic sampling of soft tissue tumors (STT) compared with unenhanced ultrasonography alone. This is a prospective longitudinal study of 40 patients subjected to ultrasonography (US)-guided core needle biopsy (CNB) to characterize a suspected STT. Three series of bioptic samplings were carried out on each patient, respectively using unenhanced US alone and CEUS in both the areas of the tumor enhanced or not by the contrast medium. All bioptic samples underwent a histological evaluation and the results were analyzed by comparing the histology of the biopsy with the definitive diagnosis in 15 surgically excised samples. 27 (67.5 %) of the 40 patients completed the entire study procedure; in 19 cases (70.3 %) the three bioptic samplings gave unanimous results, also when compared to the surgical specimen; in seven cases (25.9 %) use of CEUS allowed to obtain additional or more accurate information about the mass in question, compared to simple US guidance without contrast; in one patient (3.7 %) sampling obtained using unenhanced ultrasonography guidance and in the areas enhanced by the contrast agent had precisely the same results of the surgical specimen. CEUS, due to its ability to evaluate microvascular areas, has proven to be a promising method in guiding bioptic sampling of soft tissue tumor, directing the needle to the most significant areas of the tumor. Given the small number of patients evaluated in our study, to achieve statistically significant results, it would be appropriate to obtain a larger sample size, since the very first results seem to be encouraging and to justify the increase of the population.

Brain tumor imaging of rat fresh tissue using terahertz spectroscopy

NASA Astrophysics Data System (ADS)

Yamaguchi, Sayuri; Fukushi, Yasuko; Kubota, Oichi; Itsuji, Takeaki; Ouchi, Toshihiko; Yamamoto, Seiji

2016-07-01

Tumor imaging by terahertz spectroscopy of fresh tissue without dye is demonstrated using samples from a rat glioma model. The complex refractive index spectrum obtained by a reflection terahertz time-domain spectroscopy system can discriminate between normal and tumor tissues. Both the refractive index and absorption coefficient of tumor tissues are higher than those of normal tissues and can be attributed to the higher cell density and water content of the tumor region. The results of this study indicate that terahertz technology is useful for detecting brain tumor tissue.

CODIFI (Concordance in Diabetic Foot Ulcer Infection): a cross-sectional study of wound swab versus tissue sampling in infected diabetic foot ulcers in England.

PubMed

Nelson, Andrea; Wright-Hughes, Alexandra; Backhouse, Michael Ross; Lipsky, Benjamin A; Nixon, Jane; Bhogal, Moninder S; Reynolds, Catherine; Brown, Sarah

2018-01-31

To determine the extent of agreement and patterns of disagreement between wound swab and tissue samples in patients with an infected diabetic foot ulcer (DFU). Multicentre, prospective, cross-sectional study. Primary and secondary care foot ulcer/diabetic outpatient clinics and hospital wards across England. Inclusion criteria: consenting patients aged ≥18 years; diabetes mellitus; suspected infected DFU. clinically inappropriate to take either sample. Wound swab obtained using Levine's technique; tissue samples collected using a sterile dermal curette or scalpel. Coprimary: reported presence, and number, of pathogens per sample; prevalence of resistance to antimicrobials among likely pathogens. Secondary: recommended change in antibiotic therapy based on blinded clinical review; adverse events; sampling costs. 400 consenting patients (79% male) from 25 centres.Most prevalent reported pathogens were Staphylococcus aureus (43.8%), Streptococcus (16.7%) and other aerobic Gram-positive cocci (70.6%). At least one potential pathogen was reported from 70.1% of wound swab and 86.1% of tissue samples. Pathogen results differed between sampling methods in 58% of patients, with more pathogens and fewer contaminants reported from tissue specimens.The majority of pathogens were reported significantly more frequently in tissue than wound swab samples (P<0.01), with equal disagreement for S. aureus and Pseudomonas aeruginosa. Blinded clinicians more often recommended a change in antibiotic regimen based on tissue compared with wound swab results (increase of 8.9%, 95% CI 2.65% to 15.3%). Ulcer pain and bleeding occurred more often after tissue collection versus wound swabs (pain: 9.3%, 1.3%; bleeding: 6.8%, 1.5%, respectively). Reports of tissue samples more frequently identified pathogens, and less frequently identified non-pathogens compared with wound swab samples. Blinded clinicians more often recommended changes in antibiotic therapy based on tissue compared with wound swab specimens. Further research is needed to determine the effect of the additional information provided by tissue samples. ISRCTN52608451. © Article author(s) (or their employer(s) unless otherwise stated in the text of the article) 2018. All rights reserved. No commercial use is permitted unless otherwise expressly granted.

A novel semi-quantitative method for measuring tissue bleeding.

PubMed

Vukcevic, G; Volarevic, V; Raicevic, S; Tanaskovic, I; Milicic, B; Vulovic, T; Arsenijevic, S

2014-03-01

In this study, we describe a new semi-quantitative method for measuring the extent of bleeding in pathohistological tissue samples. To test our novel method, we recruited 120 female patients in their first trimester of pregnancy and divided them into three groups of 40. Group I was the control group, in which no dilation was applied. Group II was an experimental group, in which dilation was performed using classical mechanical dilators. Group III was also an experimental group, in which dilation was performed using a hydraulic dilator. Tissue samples were taken from the patients' cervical canals using a Novak's probe via energetic single-step curettage prior to any dilation in Group I and after dilation in Groups II and III. After the tissue samples were prepared, light microscopy was used to obtain microphotographs at 100x magnification. The surfaces affected by bleeding were measured in the microphotographs using the Autodesk AutoCAD 2009 program and its "polylines" function. The lines were used to mark the area around the entire sample (marked A) and to create "polyline" areas around each bleeding area on the sample (marked B). The percentage of the total area affected by bleeding was calculated using the formula: N = Bt x 100 / At where N is the percentage (%) of the tissue sample surface affected by bleeding, At (A total) is the sum of the surfaces of all of the tissue samples and Bt (B total) is the sum of all the surfaces affected by bleeding in all of the tissue samples. This novel semi-quantitative method utilizes the Autodesk AutoCAD 2009 program, which is simple to use and widely available, thereby offering a new, objective and precise approach to estimate the extent of bleeding in tissue samples.

A validated UPLC-MS/MS method for the analysis of linezolid and a novel oxazolidinone derivative (PH027) in plasma and its application to tissue distribution study in rabbits.

PubMed

Hedaya, Mohsen A; Thomas, Vidhya; Abdel-Hamid, Mohamed E; Kehinde, Elijah O; Phillips, Oludotun A

2017-01-01

Linezolid is the first approved oxazolidinone antibacterial agent, whereas PH027 is a novel compound of the same class that exhibits good in vitro antibacterial activity. The objective of this study was to develop an UPLC-MS/MS assay for the analysis of linezolid and PH027 in plasma and to apply the method for comparative pharmacokinetic and tissue distribution studies of both compounds. Plasma samples and calibrators were extracted with diethyl ether after addition of the internal standard solution. After evaporation of the ether layer, the residue was reconstituted in mobile phase and injected into UPLC-MS/MS. The mobile phase consisted of 2mM ammonium acetate buffer solution and acetonitrile (70:30) at a flow rate of 0.2ml/min. Separation was achieved using UPLC BEH C 18 column, and quantitative determination of the analytes was performed using multiple-reaction monitoring (MRM) scanning mode. The method was validated by analyzing quality control tissue homogenate samples, and was applied to analyze tissue homogenate samples obtained following IV injections of linezolid and PH027 in rabbits. The developed UPLC-MS/MS method was linear in the concentration range of 50-5000ng/ml. Validation of the method proved that the method's precision, selectivity and stability were all within the acceptable limits. Linezolid and PH027 concentrations were accurately determined in the quality control tissue homogenate samples, and analysis of samples obtained following IV administration of the two compounds showed that the tissue to plasma concentration ratio of PH027 was higher than that of linezolid probably due to its higher lipophilicity. The developed UPLC-MS/MS method for the analysis of linezolid and PH027 in rabbit's plasma can accurately determine the concentrations of these compounds in different tissues. Copyright © 2016 Elsevier B.V. All rights reserved.

A random sampling approach for robust estimation of tissue-to-plasma ratio from extremely sparse data.

PubMed

Chu, Hui-May; Ette, Ene I

2005-09-02

his study was performed to develop a new nonparametric approach for the estimation of robust tissue-to-plasma ratio from extremely sparsely sampled paired data (ie, one sample each from plasma and tissue per subject). Tissue-to-plasma ratio was estimated from paired/unpaired experimental data using independent time points approach, area under the curve (AUC) values calculated with the naïve data averaging approach, and AUC values calculated using sampling based approaches (eg, the pseudoprofile-based bootstrap [PpbB] approach and the random sampling approach [our proposed approach]). The random sampling approach involves the use of a 2-phase algorithm. The convergence of the sampling/resampling approaches was investigated, as well as the robustness of the estimates produced by different approaches. To evaluate the latter, new data sets were generated by introducing outlier(s) into the real data set. One to 2 concentration values were inflated by 10% to 40% from their original values to produce the outliers. Tissue-to-plasma ratios computed using the independent time points approach varied between 0 and 50 across time points. The ratio obtained from AUC values acquired using the naive data averaging approach was not associated with any measure of uncertainty or variability. Calculating the ratio without regard to pairing yielded poorer estimates. The random sampling and pseudoprofile-based bootstrap approaches yielded tissue-to-plasma ratios with uncertainty and variability. However, the random sampling approach, because of the 2-phase nature of its algorithm, yielded more robust estimates and required fewer replications. Therefore, a 2-phase random sampling approach is proposed for the robust estimation of tissue-to-plasma ratio from extremely sparsely sampled data.

Glioma tissue obtained by modern ultrasonic aspiration with a simple sterile suction trap for primary cell culture and pathological evaluation.

PubMed

Schroeteler, Juliane; Reeker, Ralf; Suero Molina, Eric; Brokinkel, Benjamin; Holling, Markus; Grauer, Oliver M; Senner, Volker; Stummer, Walter; Ewelt, Christian

2014-01-01

Ultrasonic aspiration is widely used in the resection of brain tumors. Nevertheless, tumor tissue fragments obtained by ultrasonic aspiration are usually discarded. In this study, we demonstrate that these fragments are possible sources of material for histopathological study and tissue culture and compare their microscopic features and viability in tissue culture of cavitron ultrasonic surgical aspirator tissue fragments. Brain tumor tissue collected by ultrasonic aspiration (CUSA EXcel®; Integra Radionics Inc.) in a simple sterile suction trap during resection was processed for primary cell culture. Cell viability and immunohistological markers were measured by the WST-1 test, microscopy and immunofluorescent evaluation. Six gliomas are presented to demonstrate that these tissue fragments show good preservation of histological detail and tissue viability in culture. Utilization of this material may facilitate pathological interpretation by providing a more representative sample of tumor histology as well as an adequate and sterile biosource of material for tissue culture studies.

Aetiology of extrahepatic epithelial iron deposits in siderosis in Bantu

PubMed Central

Buchanan, W. M.

1969-01-01

Twenty-seven specimens of human tissue, obtained by operation, were tested to evaluate the theory that iron uptake by tissues from serum is greater when transferrin is nearly completely saturated than when the degree of saturation is normal. Samples of each tissue were incubated in autologous serum so prepared that in one instance the transferrin was 50% saturated and in the second 90% saturated with iron containing 59Fe. In all samples the uptake of iron was greater from the transferrin which was 90% saturated. The uptake by tissues of epithelial origin was significantly greater than that by non-epithelial tissues. Considerable variation in uptake was noted between samples of the same tissue from different individuals. The role of iron stores in the tissue and folic acid deficiency are discussed. It is concluded that the degree of transferrin saturation is important in determining iron uptake by tissues, especially in those of epithelial origin, and that such uptake may be modified by tissue stores and folic acid deficiency. It is felt that these factors are probably responsible for the extrahepatic parenchymal deposits of iron sometimes found in Bantu subjects with siderosis. PMID:5784982

Proteomic Analysis of Human Tendon and Ligament: Solubilization and Analysis of Insoluble Extracellular Matrix in Connective Tissues.

PubMed

Sato, Nori; Taniguchi, Takako; Goda, Yuichiro; Kosaka, Hirofumi; Higashino, Kosaku; Sakai, Toshinori; Katoh, Shinsuke; Yasui, Natsuo; Sairyo, Koichi; Taniguchi, Hisaaki

2016-12-02

Connective tissues such as tendon, ligament and cartilage are mostly composed of extracellular matrix (ECM). These tissues are insoluble, mainly due to the highly cross-linked ECM proteins such as collagens. Difficulties obtaining suitable samples for mass spectrometric analysis render the application of modern proteomic technologies difficult. Complete solubilization of them would not only elucidate protein composition of normal tissues but also reveal pathophysiology of pathological tissues. Here we report complete solubilization of human Achilles tendon and yellow ligament, which is achieved by chemical digestion combined with successive protease treatment including elastase. The digestion mixture was subjected to liquid chromatography-mass spectrometry. The low specificity of elastase was overcome by accurate mass analysis achieved using FT-ICR-MS. In addition to the detailed proteome of both tissues, we also quantitatively determine the major protein composition of samples, by measuring peak area of some characteristic peptides detected in tissue samples and in purified proteins. As a result, differences between human Achilles tendon and yellow ligament were elucidated at molecular level.

Complete validation of a unique digestion assay to detect Trichinella larvae in horse meat demonstrates the reliability of this assay for meeting food safety and trade requirements.

PubMed

Forbes, L B; Hill, D E; Parker, S; Tessaro, S V; Gamble, H R; Gajadhar, A A

2008-03-01

A tissue digestion assay using a double separatory funnel procedure for the detection of Trichinella larvae in horse meat was validated for application in food safety programs and trade. The assay consisted of a pepsin-HCl digestion step to release larvae from muscle tissue and two sequential sedimentation steps in separatory funnels to recover and concentrate larvae for detection with a stereomicroscope. With defined critical control points, the assay was conducted within a quality assurance system compliant with International Organization for Standardization-International Electrotechnical Commission (ISO/IEC) 17025 guidelines. Samples used in the validation were obtained from horses experimentally infected with Trichinella spiralis to obtain a range of muscle larvae densities. One-, 5-, and 10-g samples of infected tissue were combined with 99, 95, and 90 g, respectively, of known negative horse tissue to create a 100-g sample for testing. Samples of 5 and 10 g were more likely to be positive than were 1-g samples when larval densities were less than three larvae per gram (lpg). This difference is important because ingested meat with 1 lpg is considered the threshold for clinical disease in humans. Using a 5-g sample size, all samples containing 1.3 to 2 lpg were detected, and 60 to 100% of samples with infected horse meat containing 0.1 to 0.7 lpg were detected. In this study, the double separatory funnel digestion assay was efficient and reliable for its intended use in food safety and trade. This procedure is the only digestion assay for Trichinella in horse meat that has been validated as consistent and effective at critical levels of sensitivity.

Detection of Toxoplasma gondii DNA in Brazilian oysters (Crassostrea rhizophorae).

PubMed

Ribeiro, L A; Santos, L K N S S; Brito, P A; Maciel, B M; Da Silva, A V; Albuquerque, G R

2015-05-04

The aim of this study was to detect evidence of Toxoplasma gondii using polymerase chain reaction (PCR)-based techniques in oysters (Crassostrea rhizophorae) obtained from the southern coastal region of Bahia, Brazil. A total of 624 oysters were collected, and the gills and digestive glands were dissected. Each tissue sample was separated into pools containing tissues (of the same type) from three animals, leading to a total of 416 experimental samples for analysis (208 samples each from the gills and digestive glands). Molecular analysis using PCR-based detection of the T. gondii AF 146527 repetitive fragment yielded negative results for all samples. However, when nested-PCR was used for detection of the T. gondii SAG-1 gene, 17 samples were positive, with the gills being the tissue with maximal detection of the parasite. These positive results were confirmed by sample sequencing. It is therefore suggested that C. rhizophorae oysters are capable of filtering and retaining T. gondii oocysts in their tissue. This represents a risk to public health because they are traditionally ingested in natura.

Microtomography evaluation of dental tissue wear surface induced by in vitro simulated chewing cycles on human and composite teeth.

PubMed

Bedini, Rossella; Pecci, Raffaella; Notarangelo, Gianluca; Zuppante, Francesca; Persico, Salvatore; Di Carlo, Fabio

2012-01-01

In this study a 3D microtomography display of tooth surfaces after in vitro dental wear tests has been obtained. Natural teeth have been compared with prosthetic teeth, manufactured by three different polyceramic composite materials. The prosthetic dental element samples, similar to molars, have been placed in opposition to human teeth extracted by paradontology diseases. After microtomography analysis, samples have been subjected to in vitro fatigue test cycles by servo-hydraulic mechanical testing machine. After the fatigue test, each sample has been subjected again to microtomography analysis to obtain volumetric value changes and dental wear surface images. Wear surface images were obtained by 3D reconstruction software and volumetric value changes were measured by CT analyser software. The aim of this work has been to show the potential of microtomography technique to display very clear and reliable wear surface images. Microtomography analysis methods to evaluate volumetric value changes have been used to quantify dental tissue and composite material wear.

Tissue diagnosis by means of endogenous fluorophores

NASA Astrophysics Data System (ADS)

Lohmann, Wolfgang; Dreyer, Thomas; Nilles, M.; Bohle, Rainer M.; Schill, Wolf-Bernhard; Glanz, Hiltrud; Fryen, Andreas; Horn, Gisela; Pollich, Beate

1995-01-01

In vitro or in situ single spot or area excitation of human tissue with either 337 nm or 365 nm results in a fluorescence spectrum at about 470 nm. Since in most tissue samples the spectra obtained look alike and differ in intensity only, the 2D tomographical distribution of the fluorescence intensity was also investigated. It could be shown that the fluorescence images of unfixed and unstained cryosections match the histological images (HE stained cryosections). They exhibit a fluorescence pattern unique for the different types of diseases investigated (basalioma, melanoma, nevi, psoriasis, seborrheic keratosis). Additional information can be obtained by elastica van Gieson stained images and by illuminating the HE stained cryosections with 365 nm. Since the cryosections can be prepared and evaluated in less than 5 minutes, this technique might be used as a fast cut technique for determining e.g. the diagnosis of a biopsy sample, also during surgery.

Method development and subsequent survey analysis of biological tissues for platinum, lead, and manganese content.

PubMed Central

Yoakum, A M; Stewart, P L; Sterrett, J E

1975-01-01

An emission spectrochemical method is described for the determination of trace quantities of platinum, lead, and manganese in biological tissues. Total energy burns in an argon-oxygen atmosphere are employed. Sample preparation, conditions of analysis, and preparation of standards are discussed. The precision of the method is consistently better than +/- 15%, and comparative analyses indicate comparable accuracies. Data obtained for experimental rat tissues and for selected autopsy tissues are presented. PMID:1157798

Correlations of trace elements in breast human tissues: Evaluation of spatial distribution using μ-XRF

NASA Astrophysics Data System (ADS)

Silva, Marina Piacenti da; Silva, Deisy Mara da; Ribeiro-Silva, Alfredo; Poletti, Martin Eduardo

2012-05-01

The aim of this work is to investigate microscopic correlations between trace elements in breast human tissues. A synchrotron X-ray fluorescence microprobe system (μ-XRF) was used to obtain two-dimensional distribution of trace element Ca, Fe, Cu and Zn in normal (6 samples) and malignant (14 samples) breast tissues. The experiment was performed in X-ray Fluorescence beam line at Laboratório Nacional de Luz Síncrotron (LNLS), Campinas, Brazil. The white microbeam was generated with a fine conical capillary with a 20 μm output diameter. The samples were supported on a XYZ table. An optical microscope with motorized zoom was used for sample positioning and choice the area to be scanned. Automatic two-dimensional scans were programmed and performed with steps of 30 μm in each direction (x, y) on the selected area. The fluorescence signals were recorded using a Si(Li) detector, positioned at 90 degrees with respect to the incident beam, with a collection time of 10 s per point. The elemental maps obtained from each sample were overlap to observe correlation between trace elements. Qualitative results showed that the pairs of elements Ca-Zn and Fe-Cu could to be correlated in malignant breast tissues. Quantitative results, achieved by Spearman correlation tests, indicate that there is a spatial correlation between these pairs of elements (p < 0.001) suggesting the importance of these elements in metabolic processes associated with the development of the tumor.

Assessment of a 1H high-resolution magic angle spinning NMR spectroscopy procedure for free sugars quantification in intact plant tissue.

PubMed

Delgado-Goñi, Teresa; Campo, Sonia; Martín-Sitjar, Juana; Cabañas, Miquel E; San Segundo, Blanca; Arús, Carles

2013-08-01

In most plants, sucrose is the primary product of photosynthesis, the transport form of assimilated carbon, and also one of the main factors determining sweetness in fresh fruits. Traditional methods for sugar quantification (mainly sucrose, glucose and fructose) require obtaining crude plant extracts, which sometimes involve substantial sample manipulation, making the process time-consuming and increasing the risk of sample degradation. Here, we describe and validate a fast method to determine sugar content in intact plant tissue by using high-resolution magic angle spinning nuclear magnetic resonance spectroscopy (HR-MAS NMR). The HR-MAS NMR method was used for quantifying sucrose, glucose and fructose in mesocarp tissues from melon fruits (Cucumis melo var. reticulatus and Cucumis melo var. cantalupensis). The resulting sugar content varied among individual melons, ranging from 1.4 to 7.3 g of sucrose, 0.4-2.5 g of glucose; and 0.73-2.83 g of fructose (values per 100 g fw). These values were in agreement with those described in the literature for melon fruit tissue, and no significant differences were found when comparing them with those obtained using the traditional, enzymatic procedure, on melon tissue extracts. The HR-MAS NMR method offers a fast (usually <30 min) and sensitive method for sugar quantification in intact plant tissues, it requires a small amount of tissue (typically 50 mg fw) and avoids the interferences and risks associated with obtaining plant extracts. Furthermore, this method might also allow the quantification of additional metabolites detectable in the plant tissue NMR spectrum.

Qualification of serological infectious disease assays for the screening of samples from deceased tissue donors.

PubMed

Kitchen, A D; Newham, J A

2011-05-01

Whilst some of the assays used for serological screening of post-mortem blood samples from deceased tissue donors in some countries have been specifically validated by the manufacturer for this purpose, a significant number of those currently in use globally have not. Although specificity has previously been considered a problem in the screening of such samples, we believe that ensuring sensitivity is more important. The aim of this study was to validate a broader range of assays for the screening of post-mortem blood samples from deceased tissue donors. Six microplate immunoassays currently in use within National Health Service Blood and Transplant (NHSBT) for the screening of blood, tissue and stem cell donations were included. Representative samples from confirmed positive donors were titrated in screen negative post-mortem samples in parallel with normal pooled negative serum to determine if there was any inhibition with the post-mortem samples. There were no significant differences seen (P < 0.005) between the dilution curves obtained for the positive samples diluted in post-mortem samples and normal pooled sera. Although small numbers of samples were studied, it can be surmised that the post-mortem blood samples from deceased tissue donors, collected according to United Kingdom guidelines, are a suitable substrate for the assays evaluated. No diminution of reactivity was seen when dilution with sera from deceased donors was compared to dilution using pooled serum from live donors. In the absence of genuine low titre positive post-mortem samples, the use of samples spiked with various levels of target material provides a means of qualifying serological screening assays used by NHSBT for the screening of post-mortem blood samples from deceased tissue donors.

Inflammatory cytokine expression following the use of bipolar electrocoagulation, ultracision harmonic scalpel and cold knife biopsy.

PubMed

Litta, Pietro; Saccardi, Carlo; Gizzo, Salvatore; Conte, Lorena; Ambrosi, Giulia; Sissi, Claudia; Palumbo, Manlio

2015-08-01

Electrical surgical devices may determine tissue damage through lateral thermal spread and activation of inflammatory processes. Several tissue effects are associated with the use of different surgical instruments. The aim of the present study was to compare tissue damage following the application of cold knife biopsy, bipolar electrocoagulation and the ultracision harmonic scalpel, through the analysis of inflammatory gene mediator expression. Three fragments of the round ligament (length 0.5 cm) were obtained from 22 females who had undergone total or subtotal laparoscopic hysterectomy using three different modes of resection: Cold knife biopsy, bipolar electrocoagulation and ultracision harmonic scalpel. The tissue fragments were examined by quantitative polymerase chain reaction (qPCR) analysis of selected cytokines. Gene expression analysis demonstrated large standard deviations due to individual variability among patients and indicated variability in the concentrations of cytokines in the three different samples. The quantity of cytokine mRNA in the cold knife biopsy samples was generally greater than those obtained by other techniques. Tumor necrosis factor-α expression was significantly higher in the sample obtained with the ultracision harmonic scalpel and bipolar electrocoagulation (P=0.033) when compared with cold knife biopsy. The inflammatory response was analyzed by the quantification of gene expression through the use of qPCR. The ultracision harmonic scalpel and bipolar electrocoagulation triggered the inflammatory cascade and resulted in an increased production of cytokines compared with cold knife biopsy.

p16 promoter hypermethylation: A useful serum marker for early detection of gastric cancer

PubMed Central

Abbaszadegan, Mohammad Reza; Moaven, Omeed; Sima, Hamid Reza; Ghafarzadegan, Kamran; A'rabi, Azadeh; Forghani, Mohammad Naser; Raziee, Hamid Reza; Mashhadinejad, Ali; Jafarzadeh, Mostafa; Esmaili-Shandiz, Ehsan; Dadkhah, Ezzat

2008-01-01

AIM: To determine p16 promoter hypermethylation in gastric tumoral tissue and serum samples, its impact on p16-protein expression, and correlation with clinical and histological features. METHODS: Samples were obtained from 52 histologically confirmed cases of gastric adenocarcinoma. Gastric tissue and serum of 50 age- and sex-matched individuals with normal gastroscopy and biopsy were obtained as control samples. Methylation-specific polymerase chain reaction (MSP) was used to evaluate methylation status of p16 promoter. p16-protein expression was analyzed by immunohistochemical staining on paraffin-embedded sections. RESULTS: Methylation was detected in 44.2% (23/52) of tumoral tissues. 60.9% of them were also methylated in serum, i.e., 26.9% of all patients (14/52). Methylation was not detected in tissue and sera of control samples. p16-protein expression was decreased in 61.5% of cases (32/52), and was significantly associated with promoter hypermethylation (P < 0.001). Methylation was significantly more frequent in higher pathological grades (P < 0.05). Methylation was not associated with other clinicopathological features and environmental factors including H pylori infection and smoking. CONCLUSION: p16 promoter hypermethylation is an important event in gastric carcinogenesis. It is the principle mechanism of p16 gene silencing. It is related to malignant tumor behavior. Detection of DNA methylation in serum may be a biomarker for early detection of gastric cancer. PMID:18395906

Preparation and Respirometric Assessment of Mitochondria Isolated from Skeletal Muscle Tissue Obtained by Percutaneous Needle Biopsy

PubMed Central

Bharadwaj, Manish S.; Tyrrell, Daniel J.; Lyles, Mary F.; Demons, Jamehl L.; Rogers, George W.; Molina, Anthony J. A.

2015-01-01

Respirometric profiling of isolated mitochondria is commonly used to investigate electron transport chain function. We describe a method for obtaining samples of human Vastus lateralis, isolating mitochondria from minimal amounts of skeletal muscle tissue, and plate based respirometric profiling using an extracellular flux (XF) analyzer. Comparison of respirometric profiles obtained using 1.0, 2.5 and 5.0 μg of mitochondria indicate that 1.0 μg is sufficient to measure respiration and that 5.0 μg provides most consistent results based on comparison of standard errors. Western blot analysis of isolated mitochondria for mitochondrial marker COX IV and non-mitochondrial tissue marker GAPDH indicate that there is limited non-mitochondrial contamination using this protocol. The ability to study mitochondrial respirometry in as little as 20 mg of muscle tissue allows users to utilize individual biopsies for multiple study endpoints in clinical research projects. PMID:25741892

Focused ion beam (FIB)/scanning electron microscopy (SEM) in tissue structural research.

PubMed

Leser, Vladka; Milani, Marziale; Tatti, Francesco; Tkalec, Ziva Pipan; Strus, Jasna; Drobne, Damjana

2010-10-01

The focused ion beam (FIB) and scanning electron microscope (SEM) are commonly used in material sciences for imaging and analysis of materials. Over the last decade, the combined FIB/SEM system has proven to be also applicable in the life sciences. We have examined the potential of the focused ion beam/scanning electron microscope system for the investigation of biological tissues of the model organism Porcellio scaber (Crustacea: Isopoda). Tissue from digestive glands was prepared as for conventional SEM or as for transmission electron microscopy (TEM). The samples were transferred into FIB/SEM for FIB milling and an imaging operation. FIB-milled regions were secondary electron imaged, back-scattered electron imaged, or energy dispersive X-ray (EDX) analyzed. Our results demonstrated that FIB/SEM enables simultaneous investigation of sample gross morphology, cell surface characteristics, and subsurface structures. The same FIB-exposed regions were analyzed by EDX to provide basic compositional data. When samples were prepared as for TEM, the information obtained with FIB/SEM is comparable, though at limited magnification, to that obtained from TEM. A combination of imaging, micro-manipulation, and compositional analysis appears of particular interest in the investigation of epithelial tissues, which are subjected to various endogenous and exogenous conditions affecting their structure and function. The FIB/SEM is a promising tool for an overall examination of epithelial tissue under normal, stressed, or pathological conditions.

Endoscopic ultrasound guided fine needle aspiration and useful ancillary methods

PubMed Central

Tadic, Mario; Stoos-Veic, Tajana; Kusec, Rajko

2014-01-01

The role of endoscopic ultrasound (EUS) in evaluating pancreatic pathology has been well documented from the beginning of its clinical use. High spatial resolution and the close proximity to the evaluated organs within the mediastinum and abdominal cavity allow detection of small focal lesions and precise tissue acquisition from suspected lesions within the reach of this method. Fine needle aspiration (FNA) is considered of additional value to EUS and is performed to obtain tissue diagnosis. Tissue acquisition from suspected lesions for cytological or histological analysis allows, not only the differentiation between malignant and non-malignant lesions, but, in most cases, also the accurate distinction between the various types of malignant lesions. It is well documented that the best results are achieved only if an adequate sample is obtained for further analysis, if the material is processed in an appropriate way, and if adequate ancillary methods are performed. This is a multi-step process and could be quite a challenge in some cases. In this article, we discuss the technical aspects of tissue acquisition by EUS-guided-FNA (EUS-FNA), as well as the role of an on-site cytopathologist, various means of specimen processing, and the selection of the appropriate ancillary method for providing an accurate tissue diagnosis and maximizing the yield of this method. The main goal of this review is to alert endosonographers, not only to the different possibilities of tissue acquisition, namely EUS-FNA, but also to bring to their attention the importance of proper sample processing in the evaluation of various lesions in the gastrointestinal tract and other accessible organs. All aspects of tissue acquisition (needles, suction, use of stylet, complications, etc.) have been well discussed lately. Adequate tissue samples enable comprehensive diagnoses, which answer the main clinical questions, thus enabling targeted therapy. PMID:25339816

Improving tissue preparation for matrix-assisted laser desorption ionization mass spectrometry imaging. Part 1: using microspotting.

PubMed

Franck, Julien; Arafah, Karim; Barnes, Alan; Wisztorski, Maxence; Salzet, Michel; Fournier, Isabelle

2009-10-01

Nowadays, matrix-assisted laser desorption ionization mass spectrometry imaging (MALDI MSI) is a powerful technique to obtain the distribution of endogenous and exogenous molecules within tissue sections. It can, thus, be used to study the evolution of molecules across different physiological stages in order to find out markers or get knowledge on signaling pathways. In order to provide valuable information, we must carefully control the sample preparation to avoid any delocalization of molecules of interest inside the tissue during this step. Currently, two strategies can be used to deposit chemicals, such as the MALDI matrix, onto the tissue both involving generation of microdroplets that will be dropped off onto the surface. First strategy involves microspraying of solutions. Here, we have been interested in the development of a microspotting strategy, where nanodroplets of solvent are ejected by a piezoelectric device to generate microspots at the tissue level. Such systems allow one to precisely control sample preparation by creating an array of spots. In terms of matrix crystallization, a microspotting MALDI matrix is hardly compatible with the results by classical (pipetting) methods. We have thus synthesized and studied new solid ionic matrixes in order to obtain high analytical performance using such a deposition system. These developments have enabled optimization of the preparation time because of the high stability of the printing that is generated in these conditions. We have also studied microspotting for performing on-tissue digestion in order to go for identification of proteins or to work from formalin fixed and paraffin embedded (FFPE) tissue samples. We have shown that microspotting is an interesting approach for on tissue digestion. Peptides, proteins, and lipids were studied under this specific preparation strategy to improve imaging performances for this class of molecules.

Structural locus of transmucosal albumin efflux in canine ileum. A fluorescent study.

PubMed

Granger, D N; Cook, B H; Taylor, A E

1976-12-01

This study demonstrates the effects of elevated intestinal venous pressure on the intestinal tissue spaces and the histological locus of the transmucosal albumin flux under such conditions. The authors were able to localize albumin in the tissues using an Evans blue-albumin fluorescence technique. This technique makes use of the fluorescence properties and albumin affinity of Evans blue dye (T-1824). Evans blue dye has a high affinity for albumin and emits a red-orange fluorescence at a wavelength of 720 nm. Evans blue was mixed with a solution of bovine serum albumin at concentrations that yield negligible amounts of free dye. Control ileal samples were obtained in order to visualize the natural tissue morphology and fluorescence. The Evans blue-albumin solution was injected and tissue samples were obtained 15 and 60 min postinjection, then venous outflow was occluded and after 15 and 60 min the tissues were sampled. Each sample was immediately frozen, freeze dried, embedded in paraffin, and 7-mu sections were made. The Evans blue-albumin was demonstrated histologically with a fluorescence microscope. No leakage sites were apparent at normal venous pressures. However, after elevation of venous pressure, Evans blue-albumin was observed in the interepithelial and/or intraepithelial spaces of villus tips, but no Evans blue-albumin was observed either between or within the epithelial cells of the crypts, or within the tubular crypt lumina. These results indicate that at elevated venous pressures, the transmucosal albumin flux occurs exclusively at the villus tip region, suggesting a great vulnerability of the cells found in this region to elevations in tissue pressure as compared to the crypt epithelial cells.

Spectroscopic analysis of bladder cancer tissues using Fourier transform infrared spectroscopy

NASA Astrophysics Data System (ADS)

Al-Muslet, Nafie A.; Ali, Essam E.

2012-03-01

Bladder cancer is one of the most common cancers in Africa. It takes several days to reach a diagnosis using histological examinations of specimens obtained by endoscope, which increases the medical expense. Recently, spectroscopic analysis of bladder cancer tissues has received considerable attention as a diagnosis technique due to its sensitivity to biochemical variations in the samples. This study investigated the use of Fourier transform infrared (FTIR) spectroscopy to analyze a number of bladder cancer tissues. Twenty-two samples were collected from 11 patients diagnosed with bladder cancer from different hospitals without any pretreatment. From each patient two samples were collected, one normal and another cancerous. FTIR spectrometer was used to differentiate between normal and cancerous bladder tissues via changes in spectra of these samples. The investigations detected obvious changes in the bands of proteins (1650, 1550 cm-1), lipids (2925, 2850 cm-1), and nucleic acid (1080, 1236 cm-1). The results show that FTIR spectroscopy is promising as a rapid, accurate, nondestructive, and easy to use alternative method for identification and diagnosis of bladder cancer tissues.

A survey of hemoparasites and ectoparasites in Nasua nasua Linnaeus, 1766 with a redescription of Hepatozoon procyonis Richards, 1961 based on morphological and molecular data.

PubMed

da Silva, Maria Regina Lucas; Fornazari, Felipe; Martins, Thiago Fernandes; Hippólito, Alícia Giolo; Rolim, Luna Scarpari; Bisca, Jacqueline Muniz; Teixeira, Carlos Roberto; O'Dwyer, Lucia Helena

2018-07-01

Hemoparasites are vector-borne parasites that infect wild carnivores worldwide. Since data on hemoparasite infections in Nasua nasua from Brazil are lacking, the aim of this study was to investigate the occurrence of hemoparasites and ectoparasites in N. nasua from different areas of Brazil. Blood samples and ectoparasites from 83 N. nasua were collected in Botucatu, Palmital, and São Paulo municipalities. Samples were screened via microscopy and molecular methods to detect hemoparasites. Tissues from two N. nasua were obtained for histopathological and molecular analyses. All 83 samples were negative for piroplasms on morphological and molecular examination. Thin blood smears of nine animals were positive for Hepatozoon gamonts. The gamonts shared morphological characteristics of Hepatozoon procyonis. Meronts were detected in the liver and spleen tissue of one animal. Twenty-one blood samples and four tissue samples were PCR positive for Hepatozoon sp. The sequences obtained were 97% identical to those of Hepatozoon felis, Hepatozoon ursi, and Hepatozoon sp. Based on searches for similarity and morphology, we identified the sequences as belonging to H. procyonis. This study provides epidemiological data on hemoparasite infections and redescribe H. procyonis based on morphological, morphometrical, and molecular analyses.

Behavior of optical properties of coagulated blood sample at 633 nm wavelength

NASA Astrophysics Data System (ADS)

Morales Cruzado, Beatriz; Vázquez y Montiel, Sergio; Delgado Atencio, José Alberto

2011-03-01

Determination of tissue optical parameters is fundamental for application of light in either diagnostics or therapeutical procedures. However, in samples of biological tissue in vitro, the optical properties are modified by cellular death or cellular agglomeration that can not be avoided. This phenomena change the propagation of light within the biological sample. Optical properties of human blood tissue were investigated in vitro at 633 nm using an optical setup that includes a double integrating sphere system. We measure the diffuse transmittance and diffuse reflectance of the blood sample and compare these physical properties with those obtained by Monte Carlo Multi-Layered (MCML). The extraction of the optical parameters: absorption coefficient μa, scattering coefficient μs and anisotropic factor g from the measurements were carried out using a Genetic Algorithm, in which the search procedure is based in the evolution of a population due to selection of the best individual, evaluated by a function that compares the diffuse transmittance and diffuse reflectance of those individuals with the experimental ones. The algorithm converges rapidly to the best individual, extracting the optical parameters of the sample. We compare our results with those obtained by using other retrieve procedures. We found that the scattering coefficient and the anisotropic factor change dramatically due to the formation of clusters.

Collagen-chitosan scaffold - Lauric acid plasticizer for skin tissue engineering on burn cases

NASA Astrophysics Data System (ADS)

Widiyanti, Prihartini; Setyadi, Ewing Dian; Rudyardjo, Djony Izak

2017-02-01

The prevalence of burns in the world is more than 800 cases per one million people each year and this is the second highest cause of death due to trauma after traffic accident. Many studies are turning to skin substitute methods of tissue engineering. The purpose of this study is to determine the composition of the collagen, chitosan, and lauric acid scaffold, as well as knowing the results of the characterization of the scaffold. The synthesis of chitosan collagen lauric acid scaffold as a skin tissue was engineered using freeze dried method. Results from making of collagen chitosan lauric acid scaffold was characterized physically, biologically and mechanically by SEM, cytotoxicity, biodegradation, and tensile strength. From the morphology test, the result obtained is that pore diameter size ranges from 94.11 to 140.1 µm for samples A,B,C,D, which are in the range of normal pore size 63-150 µm, while sample E has value below the standard which is about 37.87 to 47.36 µm. From cytotoxicity assay, the result obtained is the percentage value of living cells between 20.11 to 21.51%. This value is below 50% the standard value of living cells. Incompatibility is made possible because of human error mainly the replication of washing process over the standard. Degradation testing obtained values of 19.44% - 40% by weight which are degraded during the 7 days of observation. Tensile test results obtained a range of values of 0.192 - 3.53 MPa. Only sample A (3.53 MPa) and B (1.935 MPa) meet the standard values of skin tissue scaffold that is 1-24 MPa. Based on the results of the characteristics of this study, composite chitosan collagen scaffold with lauric acid plasticizer has a potential candidate for skin tissue engineering for skin burns cases.

Detection of Foot-and-mouth Disease Virus RNA and Capsid Protein in Lymphoid Tissues of Convalescent Pigs Does Not Indicate Existence of a Carrier State.

PubMed

Stenfeldt, C; Pacheco, J M; Smoliga, G R; Bishop, E; Pauszek, S J; Hartwig, E J; Rodriguez, L L; Arzt, J

2016-04-01

A systematic study was performed to investigate the potential of pigs to establish and maintain persistent foot-and-mouth disease virus (FMDV) infection. Infectious virus could not be recovered from sera, oral, nasal or oropharyngeal fluids obtained after resolution of clinical infection with any of five FMDV strains within serotypes A, O and Asia-1. Furthermore, there was no isolation of live virus from tissue samples harvested at 28-100 days post-infection from convalescent pigs recovered from clinical or subclinical FMD. Despite lack of detection of infectious FMDV, there was a high prevalence of FMDV RNA detection in lymph nodes draining lesion sites harvested at 35 days post-infection, with the most frequent detection recorded in popliteal lymph nodes (positive detection in 88% of samples obtained from non-vaccinated pigs). Likewise, at 35 dpi, FMDV capsid antigen was localized within follicles of draining lymph nodes, but without concurrent detection of FMDV non-structural protein. There was a marked decline in the detection of FMDV RNA and antigen in tissue samples by 60 dpi, and no antigen or viral RNA could be detected in samples obtained at 100 dpi. The data presented herein provide the most extensive investigation of FMDV persistence in pigs. The overall conclusion is that domestic pigs are unlikely to be competent long-term carriers of infectious FMDV; however, transient persistence of FMDV protein and RNA in lymphoid tissues is common following clinical or subclinical infection. © Published 2014. This article is a US Government work and is in the public domain in the USA.

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.; ...

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

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 Office of Scientific and Technical Information (OSTI.GOV)

Kertesz, Vilmos; Calligaris, David; Feldman, Daniel R.

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

Modeling and measurement of tissue elastic moduli using optical coherence elastography

NASA Astrophysics Data System (ADS)

Liang, Xing; Oldenburg, Amy L.; Crecea, Vasilica; Kalyanam, Sureshkumar; Insana, Michael F.; Boppart, Stephen A.

2008-02-01

Mechanical forces play crucial roles in tissue growth, patterning and development. To understand the role of mechanical stimuli, biomechanical properties are of great importance, as well as our ability to measure biomechanical properties of developing and engineered tissues. To enable these measurements, a novel non-invasive, micron-scale and high-speed Optical Coherence Elastography (OCE) system has been developed utilizing a titanium:sapphire based spectral-domain Optical Coherence Tomography (OCT) system and a mechanical wave driver. This system provides axial resolution of 3 microns, transverse resolution of 13 microns, and an acquisition rate as high as 25,000 lines per second. External lowfrequency vibrations are applied to the samples in the system. Step and sinusoidal steady-state responses are obtained to first characterize the OCE system and then characterize samples. Experimental results of M-mode OCE on silicone phantoms and human breast tissues are obtained, which correspond to biomechanical models developed for this analysis. Quantified results from the OCE system correspond directly with results from an indentation method from a commercial. With micron-scale resolution and a high-speed acquisition rate, our OCE system also has the potential to rapidly measure dynamic 3-D tissue biomechanical properties.

Phase-resolved acoustic radiation force optical coherence elastography

NASA Astrophysics Data System (ADS)

Qi, Wenjuan; Chen, Ruimin; Chou, Lidek; Liu, Gangjun; Zhang, Jun; Zhou, Qifa; Chen, Zhongping

2012-11-01

Many diseases involve changes in the biomechanical properties of tissue, and there is a close correlation between tissue elasticity and pathology. We report on the development of a phase-resolved acoustic radiation force optical coherence elastography method (ARF-OCE) to evaluate the elastic properties of tissue. This method utilizes chirped acoustic radiation force to produce excitation along the sample's axial direction, and it uses phase-resolved optical coherence tomography (OCT) to measure the vibration of the sample. Under 500-Hz square wave modulated ARF signal excitation, phase change maps of tissue mimicking phantoms are generated by the ARF-OCE method, and the resulting Young's modulus ratio is correlated with a standard compression test. The results verify that this technique could efficiently measure sample elastic properties accurately and quantitatively. Furthermore, a three-dimensional ARF-OCE image of the human atherosclerotic coronary artery is obtained. The result indicates that our dynamic phase-resolved ARF-OCE method can delineate tissues with different mechanical properties.

Using Digital Image Correlation to Characterize Local Strains on Vascular Tissue Specimens.

PubMed

Zhou, Boran; Ravindran, Suraj; Ferdous, Jahid; Kidane, Addis; Sutton, Michael A; Shazly, Tarek

2016-01-24

Characterization of the mechanical behavior of biological and engineered soft tissues is a central component of fundamental biomedical research and product development. Stress-strain relationships are typically obtained from mechanical testing data to enable comparative assessment among samples and in some cases identification of constitutive mechanical properties. However, errors may be introduced through the use of average strain measures, as significant heterogeneity in the strain field may result from geometrical non-uniformity of the sample and stress concentrations induced by mounting/gripping of soft tissues within the test system. When strain field heterogeneity is significant, accurate assessment of the sample mechanical response requires measurement of local strains. This study demonstrates a novel biomechanical testing protocol for calculating local surface strains using a mechanical testing device coupled with a high resolution camera and a digital image correlation technique. A series of sample surface images are acquired and then analyzed to quantify the local surface strain of a vascular tissue specimen subjected to ramped uniaxial loading. This approach can improve accuracy in experimental vascular biomechanics and has potential for broader use among other native soft tissues, engineered soft tissues, and soft hydrogel/polymeric materials. In the video, we demonstrate how to set up the system components and perform a complete experiment on native vascular tissue.

Multimodal fiber-probe spectroscopy as a clinical tool for diagnosing and classifying biological tissues

NASA Astrophysics Data System (ADS)

Cicchi, Riccardo; Anand, Suresh; Fantechi, Riccardo; Giordano, Flavio; Gacci, Mauro; Conti, Valerio; Nesi, Gabriella; Buccoliero, Anna Maria; Carini, Marco; Guerrini, Renzo; Pavone, Francesco Saverio

2017-07-01

An optical fiber probe for multimodal spectroscopy was designed, developed and used for tissue diagnostics. The probe, based on a fiber bundle with optical fibers of various size and properties, allows performing spectroscopic measurements with different techniques, including fluorescence, Raman, and diffuse reflectance, using the same probe. Two visible laser diodes were used for fluorescence spectroscopy, a laser diode emitting in the NIR was used for Raman spectroscopy, and a fiber-coupled halogen lamp for diffuse reflectance. The developed probe was successfully employed for diagnostic purposes on various tissues, including brain and bladder. In particular, the device allowed discriminating healthy tissue from both tumor and dysplastic tissue as well as to perform tumor grading. The diagnostic capabilities of the method, determined using a cross-validation method with a leave-one-out approach, demonstrated high sensitivity and specificity for all the examined samples, as well as a good agreement with histopathological examination performed on the same samples. The obtained results demonstrated that the multimodal approach is crucial for improving diagnostic capabilities with respect to what can be obtained from individual techniques. The experimental setup presented here can improve diagnostic capabilities on a broad range of tissues and has the potential of being used clinically for guiding surgical resection in the near future.

Synchronous fluorescence spectroscopy of colon neoplasia

NASA Astrophysics Data System (ADS)

Borisova, Ekaterina; Semyachkina-Glushkovskaya, Oxana; Genova, Tsanislava; Penkov, Nikolay; Terziev, Ivan; Vladimirov, Borislav; Avramov, Latchezar

2017-03-01

Synchronous fluorescence spectroscopy (SFS) is a steady-state approach that we used for evaluation of specific fluorescence characteristics of cancerous colorectal tissues. SFS allow narrowing of the fluorescence spectra received, which increase the spectral resolution and improve the analysis of the fluorescence origin in such complex objects, such as biological tissues. In our study we investigate the characteristic differences, with diagnostic meaning, in the synchronous fluorescence spectra (SFS) of cancerous and healthy colorectal tissues ex vivo using a spectrofluorimeter FluoroLog3 (HORIBA, JobinYvon, France) for obtaining of the SFS data in a broad spectral range (300-800 nm) using excitation in the range of 280-440 nm with a delta lambda between 0 and 200 nm with a 10 nm step between scanning excitation and emission data. The procedure of obtaining the investigated samples ex vivo includes their excision during surgery for removal of neoplasia lesions. After the surgical removal biological samples are transported in isothermal conditions and safekeeping solution from the hospital to the spectral laboratory, where their spectral properties were investigated. All patients received and signed written informed consent and this research is approved by Ethics committee of University Hospital "Tsaritsa Yoanna", Sofia. Histological analysis was used as "gold standard" for evaluation of tissue samples and comparison of the spectral data received.

Quality control of human tissues--experience from the Indiana University Cancer Center-Lilly Research Labs human tissue bank.

PubMed

Sandusky, George E; Teheny, Katie Heinz; Esterman, Mike; Hanson, Jeff; Williams, Stephen D

2007-01-01

The success of molecular research and its applications in both the clinical and basic research arenas is strongly dependent on the collection, handling, storage, and quality control of fresh human tissue samples. This tissue bank was set up to bank fresh surgically obtained human tissue using a Clinical Annotated Tissue Database (CATD) in order to capture the associated patient clinical data and demographics using a one way patient encryption scheme to protect patient identification. In this study, we determined that high quality of tissue samples is imperative for both genomic and proteomic molecular research. This paper also contains a brief compilation of the literature involved in the patient ethics, patient informed consent, patient de-identification, tissue collection, processing, and storage as well as basic molecular research generated from the tissue bank using good clinical practices. The current applicable rules, regulations, and guidelines for handling human tissues are briefly discussed. More than 6,610 cancer patients have been consented (97% of those that were contacted by the consenter) and 16,800 tissue specimens have been banked from these patients in 9 years. All samples collected in the bank were QC'd by a pathologist. Approximately 1,550 tissue samples have been requested for use in basic, clinical, and/or biomarker cancer research studies. Each tissue aliquot removed from the bank for a research study were evaluated by a second H&E, if the samples passed the QC, they were submitted for genomic and proteomic molecular analysis/study. Approximately 75% of samples evaluated were of high histologic quality and used for research studies. Since 2003, we changed the patient informed consent to allow the tissue bank to gather more patient clinical follow-up information. Ninety two percent of the patients (1,865 patients) signed the new informed consent form and agreed to be re-contacted for follow-up information on their disease state. In addition, eighty five percent of patients (1,584) agreed to be re-contacted to provide a biological fluid sample to be used for biomarker research.

Development of a wearable CMOS-based contact imaging system for real-time skin condition diagnosis

NASA Astrophysics Data System (ADS)

Petitdidier, Nils; Koenig, Anne; Gerbelot, Rémi; Gioux, Sylvain; Dinten, Jean-Marc

2017-07-01

Diffuse reflectance spectroscopy has been widely used in the field of biological tissue characterization with various modalities [1-5,6]. One of these modalities consists in measuring the spatially resolved diffuse reflectance (SRDR). In this technique, light is collected at multiple distances from the excitation point. The obtained reflectance decay curve is used to determine scattering and absorption properties of the tissue [7], which are directly related to tissue content and structure. Existing systems usually use fiber optics to collect light reflected from the tissue and transfer it to an optical sensor [1,6]. Such devices make it possible to perform SRDR measurements directly in contact with the tissue. However, they offer poor spatial sampling of the reflectance and low light collection efficiency. We propose to overcome these limitations by using a CMOS sensor placed in contact with the tissue to achieve light collection with high spatial sampling over several millimeters and with increased fill factor. Our objective in this paper is to demonstrate the potential of our instrument to determine the optical properties of tissues from SRDR measurements. We first describe the instrument and the employed methodology. Then, preliminary results obtained on optical phantoms are presented. Finally, the potential of our system for SRDR measurements is evaluated through comparison with a fiber-optic probe previously developed in our laboratory [6,8].

3D imaging of optically cleared tissue using a simplified CLARITY method and on-chip microscopy

PubMed Central

Zhang, Yibo; Shin, Yoonjung; Sung, Kevin; Yang, Sam; Chen, Harrison; Wang, Hongda; Teng, Da; Rivenson, Yair; Kulkarni, Rajan P.; Ozcan, Aydogan

2017-01-01

High-throughput sectioning and optical imaging of tissue samples using traditional immunohistochemical techniques can be costly and inaccessible in resource-limited areas. We demonstrate three-dimensional (3D) imaging and phenotyping in optically transparent tissue using lens-free holographic on-chip microscopy as a low-cost, simple, and high-throughput alternative to conventional approaches. The tissue sample is passively cleared using a simplified CLARITY method and stained using 3,3′-diaminobenzidine to target cells of interest, enabling bright-field optical imaging and 3D sectioning of thick samples. The lens-free computational microscope uses pixel super-resolution and multi-height phase recovery algorithms to digitally refocus throughout the cleared tissue and obtain a 3D stack of complex-valued images of the sample, containing both phase and amplitude information. We optimized the tissue-clearing and imaging system by finding the optimal illumination wavelength, tissue thickness, sample preparation parameters, and the number of heights of the lens-free image acquisition and implemented a sparsity-based denoising algorithm to maximize the imaging volume and minimize the amount of the acquired data while also preserving the contrast-to-noise ratio of the reconstructed images. As a proof of concept, we achieved 3D imaging of neurons in a 200-μm-thick cleared mouse brain tissue over a wide field of view of 20.5 mm2. The lens-free microscope also achieved more than an order-of-magnitude reduction in raw data compared to a conventional scanning optical microscope imaging the same sample volume. Being low cost, simple, high-throughput, and data-efficient, we believe that this CLARITY-enabled computational tissue imaging technique could find numerous applications in biomedical diagnosis and research in low-resource settings. PMID:28819645

A Multicenter comparative trial of a novel EUS-guided core biopsy needle (SharkCore™) with the 22-gauge needle in patients with solid pancreatic mass lesions

PubMed Central

Naveed, Mariam; Siddiqui, Ali A.; Kowalski, Thomas E.; Loren, David E.; Khalid, Ammara; Soomro, Ayesha; Mazhar, Syed M.; Yoo, Joseph; Hasan, Raza; Yalamanchili, Silpa; Tarangelo, Nicholas; Taylor, Linda J.; Adler, Douglas G.

2018-01-01

Background and Objectives: The ability to obtain adequate tissue of solid pancreatic lesions by EUS-guided remains a challenge. The aim of this study was to compare the performance characteristics and safety of EUS-FNA for evaluating solid pancreatic lesions using the standard 22-gauge needle versus a novel EUS biopsy needle. Methods: This was a multicenter retrospective study of EUS-guided sampling of solid pancreatic lesions between 2009 and 2015. Patients underwent EUS-guided sampling with a 22-gauge SharkCore (SC) needle or a standard 22-gauge FNA needle. Technical success, performance characteristics of EUS-FNA, the number of needle passes required to obtain a diagnosis, diagnostic accuracy, and complications were compared. Results: A total of 1088 patients (mean age = 66 years; 49% female) with pancreatic masses underwent EUS-guided sampling with a 22-gauge SC needle (n = 115) or a standard 22-gauge FNA needle (n = 973). Technical success was 100%. The frequency of obtaining an adequate cytology by EUS-FNA was similar when using the SC and the standard needle (94.1% vs. 92.7%, respectively). The sensitivity, specificity, and diagnostic accuracy of EUS-FNA for tissue diagnosis were not significantly different between two needles. Adequate sample collection leading to a definite diagnosis was achieved by the 1st, 2nd, and 3rd pass in 73%, 92%, and 98% of procedures using the SC needle and 20%, 37%, and 94% procedures using the standard needle (P < 0.001), respectively. The median number of passes to obtain a tissue diagnosis using the SC needle was significantly less as compared to the standard needle (1 and 3, respectively; P < 0.001). Conclusions: The EUS SC biopsy needle is safe and technically feasible for EUS-FNA of solid pancreatic mass lesions. Preliminary results suggest that the SC needle has a diagnostic yield similar to the standard EUS needle and significantly reduces the number of needle passes required to obtain a tissue diagnosis. PMID:29451167

[Radiobiological Human Tissue repository: progress and perspectives for solving the problems of radiation safety and health protection of personnel and population].

PubMed

Kirillova, E N; Romanov, S A; Loffredo, C A; Zakharova, M L; Revina, V S; Sokolova, S N; Goerlitz, D S; Zubkova, O V; Lukianova, T V; Uriadnitzkaia, T I; Pavlova, O S; Slukinova, U V; Kolosova, A V; Muksinova, K N

2014-01-01

Radiobiological Human Tissue repository was established in order to obtain and store biological material from Mayak PA workers occupationally exposed to ionizing (α- and/or γ-) radiation in a wide dose range, from the residents exposed to long term radiation due to radiation accidents and transfer of the samples to scientists for the purpose of studying the effects of radiation for people and their offspring. The accumulated biomaterial is the informational and research potential that form the basis for the work of the scientists in different spheres of biology and medicine. The repository comprises 5 sections: tumor and non-tumor tissues obtained in the course of autopsies, biopsies, surgeries, samples of blood and its components, of DNA, induced sputum, saliva, and other from people exposed or unexposed (control) to radiation. The biomaterial is stored in formalin, in paraffin blocks, slides, as well as in the freezers under low temperatures. All the information on the samples and the registrants (medical, dosimetry, demographic, and occupational data) was obtained and entered into the electronic database. A constantly updated website of the repository was developed in order to provide a possibility to get acquainted with the material and proceed with application for biosamples for scientists from Russia and abroad. Some data obtained in the course of scientific research works on the basis of the biomaterial from the Repository are briefly introduced in the review.

Correcting the effect of refraction and dispersion of light in FT-IR spectroscopic imaging in transmission through thick infrared windows.

PubMed

Chan, K L Andrew; Kazarian, Sergei G

2013-01-15

Transmission mode is one of the most common sampling methods for FT-IR spectroscopic imaging because the spectra obtained generally have a reasonable signal-to-noise ratio. However, dispersion and refraction of infrared light occurs when samples are sandwiched between infrared windows or placed underneath a layer of liquid. Dispersion and refraction cause infrared light to focus with different focal lengths depending on the wavelength (wavenumber) of the light. As a result, images obtained are in focus only at a particular wavenumber while they are defocused at other wavenumber values. In this work, a solution to correct this spread of focus by means of adding a lens on top of the infrared transparent window, such that a pseudo hemisphere is formed, has been investigated. Through this lens (or pseudo hemisphere), refraction of light is removed and the light across the spectral range has the same focal depth. Furthermore, the lens acts as a solid immersion objective and an increase of both magnification and spatial resolution (by 1.4 times) is demonstrated. The spatial resolution was investigated using an USAF resolution target, showing that the Rayleigh criterion can be achieved, as well as a sample with a sharp polymer interface to indicate the spatial resolution that can be expected in real samples. The reported approach was used to obtain chemical images of cross sections of cancer tissue and hair samples sandwiched between infrared windows showing the versatility and applicability of the method. In addition to the improved spatial resolution, the results reported herein also demonstrate that the lens can reduce the effect of scattering near the edges of tissue samples. The advantages of the presented approach, obtaining FT-IR spectroscopic images in transmission mode with the same focus across all wavenumber values and simultaneous improvement in spatial resolution, will have wide implications ranging from studies of live cells to sorption of drugs into tissues.

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.

Coherent X-ray diffraction from collagenous soft tissues

PubMed Central

Berenguer de la Cuesta, Felisa; Wenger, Marco P. E.; Bean, Richard J.; Bozec, Laurent; Horton, Michael A.; Robinson, Ian K.

2009-01-01

Coherent X-ray diffraction has been applied in the imaging of inorganic materials with great success. However, its application to biological specimens has been limited to some notable exceptions, due to the induced radiation damage and the extended nature of biological samples, the last limiting the application of most part of the phasing algorithms. X-ray ptychography, still under development, is a good candidate to overcome such difficulties and become a powerful imaging method for biology. We describe herein the feasibility of applying ptychography to the imaging of biological specimens, in particular collagen rich samples. We report here speckles in diffraction patterns from soft animal tissue, obtained with an optimized small angle X-ray setup that exploits the natural coherence of the beam. By phasing these patterns, dark field images of collagen within tendon, skin, bone, or cornea will eventually be obtained with a resolution of 60–70 nm. We present simulations of the contrast mechanism in collagen based on atomic force microscope images of the samples. Simulations confirmed the ‘speckled’ nature of the obtained diffraction patterns. Once inverted, the patterns will show the disposition and orientation of the fibers within the tissue, by enhancing the phase contrast between protein and no protein regions of the sample. Our work affords the application of the most innovative coherent X-ray diffraction tools to the study of biological specimens, and this approach will have a significant impact in biology and medicine because it overcomes many of the limits of current microscopy techniques. PMID:19706395

Coherent X-ray diffraction from collagenous soft tissues.

PubMed

Berenguer de la Cuesta, Felisa; Wenger, Marco P E; Bean, Richard J; Bozec, Laurent; Horton, Michael A; Robinson, Ian K

2009-09-08

Coherent X-ray diffraction has been applied in the imaging of inorganic materials with great success. However, its application to biological specimens has been limited to some notable exceptions, due to the induced radiation damage and the extended nature of biological samples, the last limiting the application of most part of the phasing algorithms. X-ray ptychography, still under development, is a good candidate to overcome such difficulties and become a powerful imaging method for biology. We describe herein the feasibility of applying ptychography to the imaging of biological specimens, in particular collagen rich samples. We report here speckles in diffraction patterns from soft animal tissue, obtained with an optimized small angle X-ray setup that exploits the natural coherence of the beam. By phasing these patterns, dark field images of collagen within tendon, skin, bone, or cornea will eventually be obtained with a resolution of 60-70 nm. We present simulations of the contrast mechanism in collagen based on atomic force microscope images of the samples. Simulations confirmed the 'speckled' nature of the obtained diffraction patterns. Once inverted, the patterns will show the disposition and orientation of the fibers within the tissue, by enhancing the phase contrast between protein and no protein regions of the sample. Our work affords the application of the most innovative coherent X-ray diffraction tools to the study of biological specimens, and this approach will have a significant impact in biology and medicine because it overcomes many of the limits of current microscopy techniques.

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.

Influence of soft tissue in the assessment of the primary fixation of acetabular cup implants using impact analyses.

PubMed

Bosc, Romain; Tijou, Antoine; Rosi, Giuseppe; Nguyen, Vu-Hieu; Meningaud, Jean-Paul; Hernigou, Philippe; Flouzat-Lachaniette, Charles-Henri; Haiat, Guillaume

2018-06-01

The acetabular cup (AC) implant primary stability is an important determinant for the success of cementless hip surgery but it remains difficult to assess the AC implant fixation in the clinic. A method based on the analysis of the impact produced by an instrumented hammer on the ancillary has been developed by our group (Michel et al., 2016a). However, the soft tissue thickness present around the acetabulum may affect the impact response, which may hamper the robustness of the method. The aim of this study is to evaluate the influence of the soft tissue thickness (STT) on the acetabular cup implant primary fixation evaluation using impact analyses. To do so, different AC implants were inserted in five bovine bone samples. For each sample, different stability conditions were obtained by changing the cavity diameter. For each configuration, the AC implant was impacted 25 times with 10 and 30 mm of soft tissues positioned underneath the sample. The averaged indicator I m was determined based on the amplitude of the signal for each configuration and each STT and the pull-out force was measured. The results show that the resonance frequency of the system increases when the value of the soft tissue thickness decreases. Moreover, an ANOVA analysis shows that there was no significant effect of the value of soft tissue thickness on the values of the indicator I m (F = 2.33; p-value = 0.13). This study shows that soft tissue thickness does not appear to alter the prediction of the acetabular cup implant primary fixation obtained using the impact analysis approach, opening the path towards future clinical trials. Copyright © 2018 Elsevier Ltd. All rights reserved.

Significance of biological resource collection and tumor tissue bank creation.

PubMed

Yu, Ying-Yan; Zhu, Zheng-Gang

2010-01-15

Progress in the molecular oncology of gastrointestinal carcinomas depends on high quality cancer tissues for research. Recent acceleration on new technological platforms as well as the "omics" revolution increases the demands on tissues and peripheral blood for research at the DNA, mRNA and protein levels. Tissue bank creation emerges as a priority. Tumor tissue banks are facilities that are organized to collect, store and distribute samples of tumor and normal tissue for further use in basic and translational cancer research. The samples are generally obtained immediately after excision, prior to fixation, to ensure optimal preservation of proteins and nucleic acids. It is possible for surgeons or pathologists to collect fresh tissue prospectively during their routine dissection procedures. Most tissue banks are "project-driven" tumor banks, which are specialized collections of tumor samples on which their research is based. Systematic collection of all available tumor tissue is much rarer. High quality tissue banks need the collaboration of clinicians and basic scientists, but also the informed consent of patients and ethical approval. Through the standard operation procedure, snap frozen fresh tissue collection, storage and quality control for cryopreserved tissues are the pivotal factors on tissue bank construction and maintaining. The purpose of the tissue bank creation is enhancing the quality and speed on both the basic and translational research on gastrointestinal cancer. The quality assurance and quality control are handled based on reviewing HE staining slides or touch imprint cytology by pathologists.

Microarray expression profiling in adhesion and normal peritoneal tissues.

PubMed

Ambler, Dana R; Golden, Alicia M; Gell, Jennifer S; Saed, Ghassan M; Carey, David J; Diamond, Michael P

2012-05-01

To identify molecular markers associated with adhesion and normal peritoneal tissue using microarray expression profiling. Comparative study. University hospital. Five premenopausal women. Adhesion and normal peritoneal tissue samples were obtained from premenopausal women. Ribonucleic acid was extracted using standard protocols and processed for hybridization to Affymetrix Whole Transcript Human Gene Expression Chips. Microarray data were obtained from five different patients, each with adhesion tissue and normal peritoneal samples. Real-time polymerase chain reaction was performed for confirmation using standard protocols. Gene expression in postoperative adhesion and normal peritoneal tissues. A total of 1,263 genes were differentially expressed between adhesion and normal tissues. One hundred seventy-three genes were found to be up-regulated and 56 genes were down-regulated in the adhesion tissues compared with normal peritoneal tissues. The genes were sorted into functional categories according to Gene Ontology annotations. Twenty-six up-regulated genes and 11 down-regulated genes were identified with functions potentially relevant to the pathophysiology of postoperative adhesions. We evaluated and confirmed expression of 12 of these specific genes via polymerase chain reaction. The pathogenesis, natural history, and optimal treatment of postoperative adhesive disease remains unanswered. Microarray analysis of adhesions identified specific genes with increased and decreased expression when compared with normal peritoneum. Knowledge of these genes and ontologic pathways with altered expression provide targets for new therapies to treat patients who have or are at risk for postoperative adhesions. Copyright © 2012 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.

Near infrared diffuse reflection and laser-induced fluorescence spectroscopy for myocardial tissue characterisation

NASA Astrophysics Data System (ADS)

Nilsson, A. M. K.; Heinrich, D.; Olajos, J.; Andersson-Engels, S.

1997-10-01

In order to evaluate the potential of cardiovascular tissue characterisation using near-infrared (NIR) spectroscopy, spectra in a previously unexplored wavelength region 0.8-2.3 μm were recorded from various pig heart tissue samples in vitro: normal myocardium (with and without endo/epicardium), aorta, fatty and fibrous heart tissue. The spectra were analysed with principal component analysis (PCA), revealing several spectroscopically characteristic features enabling tissue classification. Several of the identified spectral features could be attributed to specific tissue constituents by comparing the tissue signals with spectra obtained from water, elastin, collagen and cholesterol as well as with published data. The results obtained with the NIR spectroscopy technique in terms of its potential to classify different tissue types were compared with those from laser-induced fluorescence (LIF) using 337 nm excitation. LIF and NIR spectroscopy can in combination with PCA be used to discriminate between all previously mentioned tissue groups, apart from fatty versus fibrous tissue (LIF) and aorta versus fibrous tissue (NIR), respectively. The NIR analysis was improved by focusing the PCA to the wavelength segment 2.0-2.3 μm, resulting in successful spectral characterisation of all cardiovascular tissue groups.

Development and characterization of non-resonant multiphoton photoacoustic spectroscopy (NMPPAS) for brain tumor margining

NASA Astrophysics Data System (ADS)

Dahal, Sudhir

During tumor removal surgery, due to the problems associated with obtaining high-resolution, real-time chemical images of where exactly the tumor ends and healthy tissue begins (tumor margining), it is often necessary to remove a much larger volume of tissue than the tumor itself. In the case of brain tumor surgery, however, it is extremely unsafe to remove excess tissue. Therefore, without an accurate image of the tumor margins, some of the tumor's finger-like projections are inevitably left behind in the surrounding parenchyma to grow again. For this reason, the development of techniques capable of providing high-resolution real-time images of tumor margins up to centimeters below the surface of a tissue is ideal for the diagnosis and treatment of tumors, as well as surgical guidance during brain tumor excision. A novel spectroscopic technique, non-resonant multiphoton photoacoustic spectroscopy (NMPPAS), is being developed with the capabilities of obtaining high-resolution subsurface chemical-based images of underlying tumors. This novel technique combines the strengths of multiphoton tissue spectroscopy and photoacoustic spectroscopy into a diagnostic methodology that will, ultimately, provide unparalleled chemical information and images to provide the state of sub-surface tissues. The NMPPAS technique employs near-infrared light (in the diagnostic window) to excite ultraviolet and/or visible light absorbing species deep below the tissue's surface. Once a multiphoton absorption event occurs, non-radiative relaxation processes generates a localized thermal expansion and subsequent acoustic wave that can be detected using a piezoelectric transducer. Since NMPPAS employs an acoustic detection modality, much deeper diagnoses can be performed than that is possible using current state of the art high-resolution chemical imaging techniques such as multiphoton fluorescence spectroscopy. NMPPAS was employed to differentiate between excised brain tumors (astrocytoma III) and healthy tissue with over 99% accuracy. NMPPAS spectral features showed evident differences between tumor and healthy tissues, and ratiometric analysis ensured that only a few wavelengths could be used for excitation instead of using numerous wavelength excitations to create spectra. This process would significantly reduce the analysis time while maintaining the same degree of accuracy. Tissue phantoms were fabricated in order to characterize the properties of NMPPAS. Scattering particles were doped into the phantoms to simulate their light scattering properties to real tissues. This allowed for better control over shape, size, reproducibility and doping in the sample while maintaining the light-tissue interaction properties of real tissue. To make NMPPAS viable for clinical applications, the technique was characterized to determine the spatial (lateral and longitudinal) resolution, depth of penetration and its ability to image in three-dimension through layers of tissue. Both resolutions were determined to be near-cellular level resolution (50-70 microm), obtained initially with the aid of the technique of multiphoton fluorescence, and later verified using NMPPAS imaging. Additionally, the maximum depth of penetration and detection was determined to be about 1.4cm, making the technique extremely suitable to margin tumors from underlying tissues in the brain. The capability of NMPPAS to detect and image layers that lie beneath other structures and blood vessels was also investigated. Three-dimensional images were obtained for the first time using NMPPAS. The images were obtained from different depths and structures were imaged through other layers of existing structures in the sample. This verified that NMPPAS was capable of detecting and imaging structures that lie embedded within the tissues. NMPPAS images of embedded structures were also obtained with the presence of hemoglobin, which is potentially the largest source of background in blood-perfused tissues, thus showing that the technique is capable of detecting and differentiating in blood-perfused samples.

Translational Research in Pediatrics IV: Solid Tissue Collection and Processing.

PubMed

Gillio-Meina, Carolina; Zielke, H Ronald; Fraser, Douglas D

2016-01-01

Solid tissues are critical for child-health research. Specimens are commonly obtained at the time of biopsy/surgery or postmortem. Research tissues can also be obtained at the time of organ retrieval for donation or from tissue that would otherwise have been discarded. Navigating the ethics of solid tissue collection from children is challenging, and optimal handling practices are imperative to maximize tissue quality. Fresh biopsy/surgical specimens can be affected by a variety of factors, including age, gender, BMI, relative humidity, freeze/thaw steps, and tissue fixation solutions. Postmortem tissues are also vulnerable to agonal factors, body storage temperature, and postmortem intervals. Nonoptimal tissue handling practices result in nucleotide degradation, decreased protein stability, artificial posttranslational protein modifications, and altered lipid concentrations. Tissue pH and tryptophan levels are 2 methods to judge the quality of solid tissue collected for research purposes; however, the RNA integrity number, together with analyses of housekeeping genes, is the new standard. A comprehensive clinical data set accompanying all tissue samples is imperative. In this review, we examined: the ethical standards relating to solid tissue procurement from children; potential sources of solid tissues; optimal practices for solid tissue processing, handling, and storage; and reliable markers of solid tissue quality. Copyright © 2016 by the American Academy of Pediatrics.

Determination of selected neurotoxic insecticides in small amounts of animal tissue utilizing a newly constructed mini-extractor.

PubMed

Seifertová, Marta; Čechová, Eliška; Llansola, Marta; Felipo, Vicente; Vykoukalová, Martina; Kočan, Anton

2017-10-01

We developed a simple analytical method for the simultaneous determination of representatives of various groups of neurotoxic insecticides (carbaryl, chlorpyrifos, cypermethrin, and α-endosulfan and β-endosulfan and their metabolite endosulfan sulfate) in limited amounts of animal tissues containing different amounts of lipids. Selected tissues (rodent fat, liver, and brain) were extracted in a special in-house-designed mini-extractor constructed on the basis of the Soxhlet and Twisselmann extractors. A dried tissue sample placed in a small cartridge was extracted, while the nascent extract was simultaneously filtered through a layer of sodium sulfate. The extraction was followed by combined clean-up, including gel permeation chromatography (in case of high lipid content), ultrasonication, and solid-phase extraction chromatography using C 18 on silica and aluminum oxide. Gas chromatography coupled with high-resolution mass spectrometry was used for analyte separation, detection, and quantification. Average recoveries for individual insecticides ranged from 82 to 111%. Expanded measurement uncertainties were generally lower than 35%. The developed method was successfully applied to rat tissue samples obtained from an animal model dealing with insecticide exposure during brain development. This method may also be applied to the analytical treatment of small amounts of various types of animal and human tissue samples. A significant advantage achieved using this method is high sample throughput due to the simultaneous treatment of many samples. Graphical abstract Optimized workflow for the determination of selected insecticides in small amounts of animal tissue including newly developed mini-extractor.

Magnetoacoustic imaging of human liver tumor with magnetic induction

NASA Astrophysics Data System (ADS)

Hu, Gang; Cressman, Erik; He, Bin

2011-01-01

Magnetoacoustic tomography with magnetic induction (MAT-MI) is an imaging technique under development to achieve imaging of electrical impedance contrast in biological tissues with spatial resolution close to ultrasound imaging. However, previously reported MAT-MI experimental results are obtained either from low salinity gel phantoms, or from normal animal tissue samples. In this study, we report the experimental study on the performance of the MAT-MI imaging method for imaging in vitro human liver tumor tissue. The present promising experimental results suggest the feasibility of MAT-MI to image electrical impedance contrast between the cancerous tissue and its surrounding normal tissues.

A new diagnostic real-time PCR method for huanglongbing detection in citrus root tissue

USDA-ARS?s Scientific Manuscript database

Citrus fibrous root tissue was evaluated as an alternative source material for Huanglongbing (HLB) diagnosis by real-time PCR using primer-probe set TXCChlb, developed in the present study based on 16S rDNA of “Candidatus Liberibacter asiaticus” (CLas). Real-time PCR data obtained with DNA samples p...

Investigation of metabolite changes in the transition from pre-invasive to invasive cervical cancer measured using (1)H and (31)P magic angle spinning MRS of intact tissue.

PubMed

De Silva, Sonali S; Payne, Geoffrey S; Thomas, Valerie; Carter, Paul G; Ind, Thomas E J; deSouza, Nandita M

2009-02-01

The aim of this study was to determine the metabolic changes in the transition from pre-invasive to invasive cervical cancer using high-resolution magic angle spinning (HR-MAS) MRS. Biopsy specimens were obtained from women with histologically normal cervix (n = 5), cervical intraepithelial neoplasia (CIN; mild, n = 5; moderate/severe, n = 40), and invasive cancer (n = 23). (1)H HR-MAS MRS data were acquired using a Bruker Avance 11.74 T spectrometer (Carr-Purcell-Meiboom-Gill sequence; TR = 4.8 s; TE = 135 ms; 512 scans; 41 min acquisition). (31)P HR-MAS spectra were obtained from the normal subjects and cancer patients only (as acetic acid applied before tissue sampling in patients with CIN impaired spectral quality) using a (1)H-decoupled pulse-acquire sequence (TR = 2.82 s; 2048 scans; 96 min acquisition). Peak assignments were based on values reported in the literature. Peak areas were measured using the AMARES algorithm. Estimated metabolite concentrations were compared between patient diagnostic categories and tissue histology using independent samples t tests. Comparisons based on patient category at diagnosis showed significantly higher estimated concentrations of choline (P = 0.0001) and phosphocholine (P = 0.002) in tissue from patients with cancer than from patients with high-grade dyskaryosis, but no differences between non-cancer groups. Division by histology of the sample also showed increases in choline (P = 0.002) and phosphocholine (P = 0.002) in cancer compared with high-grade CIN tissue. Phosphoethanolamine was increased in cancer compared with normal tissue (P = 0.0001). Estimated concentrations of alanine (P = 0.01) and creatine (P = 0.008) were significantly reduced in normal tissue from cancer patients compared with normal tissue from non-cancer patients. The estimated concentration of choline was significantly increased in CIN tissue from cancer patients compared with CIN tissue from non-cancer patients (P = 0.0001). Estimated concentrations of choline-containing metabolites increased from pre-invasive to invasive cervical cancer. Concurrent metabolite depletion occurs in normal tissue adjacent to cancer tissue. Copyright (c) 2008 John Wiley & Sons, Ltd.

Validation of proton stopping power ratio estimation based on dual energy CT using fresh tissue samples

NASA Astrophysics Data System (ADS)

Taasti, Vicki T.; Michalak, Gregory J.; Hansen, David C.; Deisher, Amanda J.; Kruse, Jon J.; Krauss, Bernhard; Muren, Ludvig P.; Petersen, Jørgen B. B.; McCollough, Cynthia H.

2018-01-01

Dual energy CT (DECT) has been shown, in theoretical and phantom studies, to improve the stopping power ratio (SPR) determination used for proton treatment planning compared to the use of single energy CT (SECT). However, it has not been shown that this also extends to organic tissues. The purpose of this study was therefore to investigate the accuracy of SPR estimation for fresh pork and beef tissue samples used as surrogates of human tissues. The reference SPRs for fourteen tissue samples, which included fat, muscle and femur bone, were measured using proton pencil beams. The tissue samples were subsequently CT scanned using four different scanners with different dual energy acquisition modes, giving in total six DECT-based SPR estimations for each sample. The SPR was estimated using a proprietary algorithm (syngo.via DE Rho/Z Maps, Siemens Healthcare, Forchheim, Germany) for extracting the electron density and the effective atomic number. SECT images were also acquired and SECT-based SPR estimations were performed using a clinical Hounsfield look-up table. The mean and standard deviation of the SPR over large volume-of-interests were calculated. For the six different DECT acquisition methods, the root-mean-square errors (RMSEs) for the SPR estimates over all tissue samples were between 0.9% and 1.5%. For the SECT-based SPR estimation the RMSE was 2.8%. For one DECT acquisition method, a positive bias was seen in the SPR estimates, having a mean error of 1.3%. The largest errors were found in the very dense cortical bone from a beef femur. This study confirms the advantages of DECT-based SPR estimation although good results were also obtained using SECT for most tissues.

Technique for examining biological materials using diffuse reflectance spectroscopy and the kubelka-munk function

DOEpatents

Alfano, Robert R.; Yang, Yuanlong

2003-09-02

Method and apparatus for examining biological materials using diffuse reflectance spectroscopy and the Kubelka-Munk function. In one aspect, the method is used to determine whether a tissue sample is cancerous or not and comprises the steps of (a) measuring the diffuse reflectance from the tissue sample at a first wavelength and at a second wavelength, wherein the first wavelength is a wavelength selected from the group consisting of 255-265 nm and wherein the second wavelength is a wavelength selected from the group consisting of 275-285 nm; (b) using the Kubelka-Munk function to transform the diffuse reflectance measurement obtained at the first and second wavelengths; and (c) comparing a ratio or a difference of the transformed Kubelka-Munk measurements at the first and second wavelengths to appropriate standards determine whether or not the tissue sample is cancerous. One can use the spectral profile of KMF between 250 nm to 300 nm to determine whether or not the tissue sample is cancerous or precancerous. According to the value at the first and second wavelengths determine whether or not the malignant tissue is invasive or mixed invasive and in situ or carcinoma in situ.

Cancer related gene alterations can be detected with next-generation sequencing analysis of bile in diffusely infiltrating type cholangiocarcinoma.

PubMed

Lee, Chang Hun; Wang, Hong En; Seo, Seung Young; Kim, Seong Hun; Kim, In Hee; Kim, Sang Wook; Lee, Soo Teik; Kim, Dae Ghon; Han, Myung Kwan; Lee, Seung Ok

2016-08-01

Genome-wide association study in diffusely infiltrating type cholangiocarcinoma (CC) can be limited due to the difficulty of obtaining tumor tissue. We aimed to evaluate the genomic alterations of diffusely infiltrating type CC using next-generation sequencing (NGS) of bile and to compare the variations with those of mass-forming type CC. A total of 24 bile samples obtained during endoscopic retrograde cholangiopancreatography (ERCP) and 17 surgically obtained tumor tissue samples were evaluated. Buffy coat and normal tissue samples were used as controls for a somatic mutation analysis. After extraction of genomic DNA, NGS analysis was performed for 48 cancer related genes. There were 27 men and 14 women with a mean age of 65.0±11.8years. The amount of extracted genomic DNA from 3cm(3) of bile was 66.0±84.7μg and revealed a high depth of sequencing coverage. All of the patients had genomic variations, with an average number of 19.4±2.8 and 22.3±3.3 alterations per patient from the bile and tumor tissue, respectively. After filtering process, damaging SNPs (8 sites for each type of CC) were predicted by analyzing tools, and their target genes showed relevant differences between the diffusely infiltrating and mass-forming type CC. Finally, in somatic mutation analysis, tumor-normal paired 14 tissue and 6 bile samples were analyzed, genomic alterations of EGFR, FGFR1, ABL1, PIK3CA, and CDKN2A gene were seen in the diffusely infiltrating type CC, and TP53, KRAS, APC, GNA11, ERBB4, ATM, SMAD4, BRAF, and IDH1 were altered in the mass-forming type CC group. STK11, GNAQ, RB1, KDR, and SMO genes were revealed in both groups. The NGS analysis was feasible with bile sample and diffusely infiltrating type CC revealed genetic differences compared with mass-forming type CC. Genome-wide association study could be performed using bile sample in the patients with CC undergoing ERCP and a different genetic approach for accurate diagnosis, pathogenesis study, and targeted therapy will be needed in diffusely infiltrating type CC. Copyright © 2016 Elsevier Inc. All rights reserved.

The Prostate, Lung, Colorectal and Ovarian Cancer (PLCO) Screening Trial Pathology Tissue Resource.

PubMed

Zhu, Claire S; Huang, Wen-Yi; Pinsky, Paul F; Berg, Christine D; Sherman, Mark; Yu, Kelly J; Carrick, Danielle M; Black, Amanda; Hoover, Robert; Lenz, Petra; Williams, Craig; Hawkins, Laura; Chaloux, Matthew; Yurgalevitch, Susan; Mathew, Sunitha; Miller, Amy; Olivo, Vanessa; Khan, Asia; Pretzel, Shannon M; Multerer, Deborah; Beckmann, Patricia; Broski, Karen G; Freedman, Neal D

2016-12-01

Pathology tissue specimens with associated epidemiologic and clinical data are valuable for cancer research. The Prostate, Lung, Colorectal and Ovarian (PLCO) Cancer Screening Trial undertook a large-scale effort to create a public resource of pathology tissues from PLCO participants who developed a cancer during the trial. Formalin-fixed paraffin-embedded tissue blocks were obtained from pathology laboratories on a loan basis for central processing of tissue microarrays, with additional free-standing tissue cores collected for nucleic acid extraction. Pathology tissue specimens were obtained for prostate cancer (n = 1,052), lung cancer (n = 434), colorectal cancer (n = 675) and adenoma (n = 658), ovarian cancer and borderline tumors (n = 212), breast cancer (n = 870), and bladder cancer (n = 204). The process of creating this resource was complex, involving multidisciplinary teams with expertise in pathology, epidemiology, information technology, project management, and specialized laboratories. Creating the PLCO tissue resource required a multistep process, including obtaining medical records and contacting pathology departments where pathology materials were stored after obtaining necessary patient consent and authorization. The potential to link tissue biomarkers to prospectively collected epidemiologic information, screening and clinical data, and matched blood or buccal samples offers valuable opportunities to study etiologic heterogeneity, mechanisms of carcinogenesis, and biomarkers for early detection and prognosis. The methods and protocols developed for this effort, and the detailed description of this resource provided here, will be useful for those seeking to use PLCO pathology tissue specimens for their research and may also inform future tissue collection efforts in other settings. Cancer Epidemiol Biomarkers Prev; 25(12); 1635-42. ©2016 AACR. ©2016 American Association for Cancer Research.

Investigation of Helicobacter pylori in tonsillary tissue with Pronto Dry test and pathologic examination.

PubMed

Aslan, Sundus; Yilmaz, Ismail; Bal, Nebil; Sener, Mesut; Butros, Reha; Demirhan, Beyhan; Ozluoglu, Levent N

2007-09-01

The objectives of this clinical study were to identify, by means of the Pronto Dry test and pathologic examination, Helicobacter pylori (HP) in tonsillary tissue and to establish the role of HP in tonsillary microbiology by identifying that bacterium in the tonsillary mucosa or within the tonsil core. The subjects consisted of 52 patients (25 men and 27 women; age range, 3-65 years; mean age, 15.1+/-14.5 years) who were scheduled to undergo tonsillectomy for the treatment of chronic tonsillitis and who had not been treated with an antibiotic or a bismuth-containing compound for 6 months before the initiation of the study. In each patient, two specimens (one 4 mm x 4 mm tissue sample from the nonmucosal tonsil core and one 4 mm x 4 mm sample of mucosal tissue) were excised from both tonsils immediately after tonsillectomy. The specimens were placed in the Pronto Dry test kit, and the test results were obtained 1 h later. The remaining tonsillary tissues were submitted for pathologic analysis via hematoxylin-eosin stain, Giemsa stain, Warthin-Starry silver stain, and staining for inducible nitric oxide synthase (iNOS). The results of the Pronto Dry test were positive for HP in 42% (n=22) of the excised mucosal tissue and in 47% (n=24) of the excised core tissue. In 27% (n=14) of the patients, both the core and the mucosal tissues tested positive for HP. There was no significant difference between the positive Pronto Dry test ratios of the biopsies obtained from the mucosa and those obtained from the core (P=0.693). iNOS staining showed that macrophage iNOS activity was significantly higher (P=0.025) in biopsied mucosal tissues with a positive Pronto Dry test result than in those with a negative result. Light microscopy revealed no HP in samples stained with hematoxylin-eosin stain, Giemsa stain, or Warthin-Starry silver stain. Positive Pronto Dry test results and the results of iNOS staining showed that HP contributes to chronic tonsillitis, especially at the mucosal layer. Although HP does not colonize, it contributes to the chronic tonsillary inflammatory process as a triggering agent by affecting macrophages in the tonsil and thus increasing iNOS expression.

Measurement of indicator genes using global complementary DNA (cDNA) amplification, by polyadenylic acid reverse transcriptase polymerase chain reaction (poly A RT-PCR): A feasibility study using paired samples from tissue and ductal juice in patients undergoing pancreatoduodenectomy.

PubMed

Sanyal, Sudip; Siriwardena, Ajith K; Byers, Richard

2018-06-01

The aim of this study is to compare gene expression profiles in RNA isolated from pancreatic ductal juice with the RNA expression profiles of the same genes from matched intra-operative tissue samples from pancreatic tumours. Intra-operative sampling of pancreatic juice and collection of matched tissue samples was undertaken in patients undergoing pancreatoduodenectomy for clinically suspected pancreatic cancer and a precursor lesion, main-duct intraductal papillary mucinous neoplasm. RNA was isolated and Poly A PCR was used to globally amplify the RNA. Real-time polymerase chain reaction (RT-PCR) was used to measure expression levels of 17 genes selected from microarray studies. Spearman's rank correlation test was used to examine the relationship of gene expression between pancreatic juice and tissue. The study was approved by Regional Ethics Committee. Mesothelin (MSLN) showed significant correlation (p < 0.008) in expression levels between paired pancreatic juice and tissue samples in pancreas cancer. In intraductal papillary mucinous neoplasms (IPMN), Matrix Metalloproteinase 7 (MMP7), showed significant correlation (p < 0.01) in the expression levels between paired pancreatic juice and tissue samples. This study confirms that RNA analysis of paired pancreatic juice and tissue samples and establishment of cDNA using poly A PCR is technically feasible. Application of the technique to non-invasively obtained pancreatic juice during endoscopic assessment of tumours and the use of gene arrays of cancer indicator genes are the next steps in development of this technique. Copyright © 2018 IAP and EPC. Published by Elsevier B.V. All rights reserved.

Determination of 240Pu/239Pu isotopic ratios in human tissues collected from areas around the Semipalatinsk Nuclear Test Site by sector-field high resolution ICP-MS.

PubMed

Yamamoto, M; Oikawa, S; Sakaguchi, A; Tomita, J; Hoshi, M; Apsalikov, K N

2008-09-01

Information on the 240Pu/239Pu isotope ratios in human tissues for people living around the Semipalatinsk Nuclear Test Site (SNTS) was deduced from 9 sets of soft tissues and bones, and 23 other bone samples obtained by autopsy. Plutonium was radiochemically separated and purified, and plutonium isotopes (239Pu and 240Pu) were determined by sector-field high resolution inductively coupled plasma-mass spectrometry. For most of the tissue samples from the former nine subjects, low 240Pu/239Pu isotope ratios were determined: bone, 0.125 +/- 0.018 (0.113-0.145, n = 4); lungs, 0.063 +/- 0.010 (0.051-0.078, n = 5); and liver, 0.148 +/- 0.026 (0.104-0.189, n = 9). Only 239Pu was detected in the kidney samples; the amount of 240Pu was too small to be measured, probably due to the small size of samples analyzed. The mean 240Pu/239Pu isotope ratio for bone samples from the latter 23 subjects was 0.152 +/- 0.034, ranging from 0.088 to 0.207. A significant difference (a two-tailed Student's t test; 95% significant level, alpha = 0.05) between mean 240Pu/239Pu isotope ratios for the tissue samples and for the global fallout value (0.178 +/- 0.014) indicated that weapons-grade plutonium from the atomic bombs has been incorporated into the human tissues, especially lungs, in the residents living around the SNTS. The present 239,240Pu concentrations in bone, lung, and liver samples were, however, not much different from ranges found for human tissues from other countries that were due solely to global fallout during the 1970's-1980's.

Mass Spectrometric Rapid Diagnosis of Infectious Diseases.

DTIC Science & Technology

1980-01-01

the analytical procedures to warrant reporting anew the whole analytical procedure. A. Sample Collection and Storage Procedure Urine samples were...positives or false-negatives. Next we have carried out a longitudinal study on urine samples obtained from groups of volunteer subjects vaccinated with...sterilization and storage procedures. 2. Developed new, simpler sample preparation techniques including one to handle tissue culture media. 3. Improved on the

[Measurement of the status of trace elements in cattle using liver biopsy samples].

PubMed

Ouweltjes, W; de Zeeuw, A C; Moen, A; Counotte, G H M

2007-02-01

Serum, plasma, or urine samples are usually used for the measurement of the trace elements copper; zinc, iron, selenium, because these samples are easy to obtain; however; these samples are not always appropriate. For example, it is not possible to measure molybdenum, the major antagonist of copper; in blood or urine. Therefore measurement of trace elements in liver tissue is considered the gold standard. For the assessment of selenium the method of choice remains determination of glutathion peroxidase in erythrocytes and for the assessment of magnesium determination of magnesium in urine. We determined the accuracy and repeatability of measuring trace elements in liver biopsies and whole liver homogenates. The levels of trace elements measured were similar in both preparations (92% agreement). Liver biopsy in live animals is a relatively simple procedure but not common in The Netherlands. Reference levels of trace elements, classified as too low, low, adequate, high, and too high, were established on the basis of our research and information in the literature. In a second study we investigated the practical aspects of obtaining liver tissue samples and their use. Samples were collected from cattle on a commercial dairy farm. Liver biopsy provided additional information to that obtained from serum and urine samples. We prepared a biopsy protocol and a test package, which we tested on 14 farms where an imbalance of trace minerals was suspected. Biopsy samples taken from 4 to 6 animals revealed extreme levels of trace elements.

Usability of Immunohistochemistry in Forensic Samples With Varying Decomposition.

PubMed

Lesnikova, Iana; Schreckenbach, Marc Niclas; Kristensen, Maria Pihlmann; Papanikolaou, Liv Lindegaard; Hamilton-Dutoit, Stephen

2018-05-24

Immunohistochemistry (IHC) is an important diagnostic tool in anatomic and surgical pathology but is used less frequently in forensic pathology. Degradation of tissue because of postmortem decomposition is believed to be a major limiting factor, although it is unclear what impact such degradation actually has on IHC staining validity. This study included 120 forensic autopsy samples of liver, lung, and brain tissues obtained for diagnostic purposes. The time from death to autopsy ranged between 1 and more than 14 days. Samples were prepared using the tissue microarray technique. The antibodies chosen for the study included KL1 (for staining bile duct epithelium), S100 (for staining glial cells and myelin), vimentin (for endothelial cells in cerebral blood vessels), and CD45 (for pulmonary lymphocytes). Slides were evaluated by light microscopy. Immunohistochemistry reactions were scored according to a system based on the extent and intensity of the positive stain. An overall correlation between the postmortem interval and the IHC score for all tissue samples was found. Samples from decedents with a postmortem interval of 1 to 3 days showed positive staining with all antibodies, whereas samples from decedents with a longer postmortem interval showed decreased staining rates. Our results suggest that IHC analysis can be successfully used for postmortem diagnosis in a range of autopsy samples showing lesser degrees of decomposition.

Use of a tissue sectioner to expose internal structures of biological samples for scanning electron microscopy.

PubMed

Brown, M F; Brotzman, H G; Kinden, D A

1976-09-01

A procedure yielding sections of unembedded biological samples for observation by scanning electron microscopy is described. Sections of samples, fixed and hardened in OsO4, were obtained in quantity with a tissue sectioner. Subsequent treatments to osmium-coat cut surfaces were employed prior to critical point drying. The procedure yields cleanly cut surfaces through cells and cytoplasmic organelles which are retained in their normal position. Sections of apple leaf and mouse kidney are illustrated. Sections can be readily cut in a desired plane with less structural damage than is typically encountered by other sectioning or dissection techniques.

Video of Tissue Grown in Space in NASA Bioreactor

NASA Technical Reports Server (NTRS)

2003-01-01

Principal investigator Leland Chung grew prostate cancer and bone stromal cells aboard the Space Shuttle Columbia during the STS-107 mission. Although the experiment samples were lost along with the ill-fated spacecraft and crew, he did obtain downlinked video of the experiment that indicates the enormous potential of growing tissues in microgravity. Cells grown aboard Columbia had grown far larger tissue aggregates at day 5 than did the cells grown in a NASA bioreactor on the ground.

Assessment of hyaline cartilage matrix composition using near infrared spectroscopy.

PubMed

Palukuru, Uday P; McGoverin, Cushla M; Pleshko, Nancy

2014-09-01

Changes in the composition of the extracellular matrix (ECM) are characteristic of injury or disease in cartilage tissue. Various imaging modalities and biochemical techniques have been used to assess the changes in cartilage tissue but lack adequate sensitivity, or in the case of biochemical techniques, result in destruction of the sample. Fourier transform near infrared (FT-NIR) spectroscopy has shown promise for the study of cartilage composition. In the current study NIR spectroscopy was used to identify the contributions of individual components of cartilage in the NIR spectra by assessment of the major cartilage components, collagen and chondroitin sulfate, in pure component mixtures. The NIR spectra were obtained using homogenous pellets made by dilution with potassium bromide. A partial least squares (PLS) model was calculated to predict composition in bovine cartilage samples. Characteristic absorbance peaks between 4000 and 5000 cm(-1) could be attributed to components of cartilage, i.e. collagen and chondroitin sulfate. Prediction of the amount of collagen and chondroitin sulfate in tissues was possible within 8% (w/dw) of values obtained by gold standard biochemical assessment. These results support the use of NIR spectroscopy for in vitro and in vivo applications to assess matrix composition of cartilage tissues, especially when tissue destruction should be avoided. Copyright © 2014. Published by Elsevier B.V.

The direct analysis of drug distribution of rotigotine-loaded microspheres from tissue sections by LESA coupled with tandem mass spectrometry.

PubMed

Xu, Li-Xiao; Wang, Tian-Tian; Geng, Yin-Yin; Wang, Wen-Yan; Li, Yin; Duan, Xiao-Kun; Xu, Bin; Liu, Charles C; Liu, Wan-Hui

2017-09-01

The direct analysis of drug distribution of rotigotine-loaded microspheres (RoMS) from tissue sections by liquid extraction surface analysis (LESA) coupled with tandem mass spectrometry (MS/MS) was demonstrated. The RoMS distribution in rat tissues assessed by the ambient LESA-MS/MS approach without extensive or tedious sample pretreatment was compared with that obtained by a conventional liquid chromatography tandem mass spectrometry (LC-MS/MS) method in which organ excision and subsequent solvent extraction were commonly employed before analysis. Results obtained from the two were well correlated for a majority of the organs, such as muscle, liver, stomach, and hippocampus. The distribution of RoMS in the brain, however, was found to be mainly focused in the hippocampus and striatum regions as shown by the LESA-imaged profiles. The LESA approach we developed is sensitive enough, with an estimated LLOQ at 0.05 ng/mL of rotigotine in brain tissue, and information-rich with minimal sample preparation, suitable, and promising in assisting the development of new drug delivery systems for controlled drug release and protection. Graphical abstract Workflow for the LESA-MS/MS imaging of brain tissue section after intramuscular RoMS administration.

Determination of iodine in bread and fish using the iodide ion-selective electrode

DOE Office of Scientific and Technical Information (OSTI.GOV)

Steiner, J.B.

The purpose of this study was to assess the potential for use of the ion-selective electrode (ISE) as a method for measuring the iodine content in bread and fish. Ashing methods, sample preparation and electrode responses were evaluated. The iodine values obtained using the iodide electrode were compared to iodine values obtained by the arsenic-cerium method (As-Ce). Ashing methods were used in preparing bread and haddock for iodine analysis by the ISE. The values were compared to unashed samples measured by the ISE. Electrode response to iodide was examined by varying the sample pH, measuring electrode equilibrium times, and comparingmore » direct measurement in ppm to iodide values obtained by the method of known addition. Oyster reference tissue with a known iodine concentration was used to determine rates of recovery. For the As-Ce procedure, an alkaline dry ash for two hour followed by colorimetric analysis at 320 nm was recommended. The study showed that the pre-treatment of bread and fish was necessary for ISE measurement. The iodine values obtained by the ISE in the analysis of oyster reference tissue, haddock and bread were not in agreement with their corresponding As-Ce values. Further work needs to be done to determine an ashing procedure that has minimal iodide loss an/or develop sample treatments that will improve the reliability and precision of iodine values obtained using the ion-selective electrode.« less

Multilayer porous UHMWPE scaffolds for bone defects replacement.

PubMed

Maksimkin, A V; Senatov, F S; Anisimova, N Yu; Kiselevskiy, M V; Zalepugin, D Yu; Chernyshova, I V; Tilkunova, N A; Kaloshkin, S D

2017-04-01

Reconstruction of the structural integrity of the damaged bone tissue is an urgent problem. UHMWPE may be potentially used for the manufacture of porous implants simulating as closely as possible the porous cancellous bone tissue. But the extremely high molecular weight of the polymer does not allow using traditional methods of foaming. Porous and multilayer UHMWPE scaffolds with nonporous bulk layer and porous layer that mimics cancellous bone architecture were obtained by solid-state mixing, thermopressing and washing in subcritical water. Structural and mechanical properties of the samples were studied. Porous UHMWPE samples were also studied in vitro and in vivo. The pores of UHMWPE scaffold are open and interconnected. Volume porosity of the obtained samples was 79±2%; the pore size range was 80-700μm. Strong connection of the two layers in multilayer UHMWPE scaffolds was observed with decreased number of fusion defects. Functionality of implants based on multilayer UHMWPE scaffolds is provided by the fixation of scaffolds in the bone defect through ingrowths of the connective tissue into the pores, which ensures the maintenance of the animals' mobility. Copyright © 2016 Elsevier B.V. All rights reserved.

Tuberculosis prevalence in forensic autopsies.

PubMed

Ozsoy, Sait; Demirel, Birol; Albay, Ali; Kisa, Ozgul; Dinc, Ahmet H; Safali, Mukerrem

2010-03-01

According to the 2008 World Health Organization report, in 2006, 9.2 million new cases were determined, and 1.7 million people have lost their life due to tuberculosis (TB) in all around the world. In our country (Turkey), it is estimated that 35,000 to 40,000 people have TB disease annually. The Ministry of Health could just determine 18,500 of these cases, and only 6500 patient could be treated effectively. According to the Tuberculosis Dispensary records, the incidence for TB in Turkey is 28/100,000. It is aimed to determine the infection with Mycobacterium tuberculosis using acidoresistant bacilli microscopy, TB culture, and histopathological methods in tissue samples that were obtained from lungs of forensic cases whose autopsies had been performed in Council of Forensic Medicine Ankara Department Morgue Specialized Committee. A total of 3 tissue samples that were obtained from lungs of randomized 302 cases, were positive for TB in Löwenstein-Jensen medium. Granuloma with caseating necrosis was found in histopathological examination and acidoresistant (+) bacilli (1+, 2+, and 2+, respectively) in microscopically analysis were also demonstrated in this 3 tissue samples. For this reason, we think that autopsy workers have to be careful about tuberculosis during their autopsy working.

Detection of beryllium in digested autopsy tissues by inductively coupled plasma mass spectrometry using a high matrix interface configuration.

PubMed

Larivière, Dominic; Tremblay, Mélodie; Durand-Jézéquel, Myriam; Tolmachev, Sergei

2012-04-01

This article describes a robust methodology using the combination of instrumental design (high matrix interface-HMI), sample dilution and internal standardization for the quantification of beryllium (Be) in various digested autopsy tissues using inductively coupled plasma mass spectrometry. The applicability of rhodium as a proper internal standard for Be was demonstrated in three types of biological matrices (i.e., femur, hair, lung tissues). Using HMI, it was possible to achieve instrumental detection limits and sensitivity of 0.6 ng L(-1) and 157 cps L ng(-1), respectively. Resilience to high salt matrices of the HMI setup was also highlighted using bone mimicking solution ([Ca(2+)] = 26 to 1,400 mg L(-1)), providing a 14-fold increase in tolerance and a 2.7-fold decrease in method detection limit compared to optimized experimental conditions obtained without the HMI configuration. Precision of the methodology to detect low levels of Be in autopsy samples was demonstrated using hair and blood certified reference materials. Be concentration ranging from 0.015 to 255 μg kg(-1) in autopsy samples obtained from the U.S. Transuranium and Uranium Registries were measured using the methodology presented.

The effects of different preservation processes on the total protein and growth factor content in a new biological product developed from human amniotic membrane.

PubMed

Russo, Alessandra; Bonci, Paola; Bonci, Paolo

2012-06-01

The aim of this work is to quantify the total protein and growth factors content in a tissue-suspension obtained from processed human amniotic membrane (hAM). hAM was collected, frozen, freeze dried, powdered and sterilized by γ-irradiation. At each step of the process, samples were characterized for the total protein amounts by a Bradford protein assay and for the growth factor concentrations by ELISA test of the tissue suspensions. Frozen-hAM samples show higher release of total proteins and specific growth factors in the tissue suspension in comparison with freeze-dried hAM. We observed that even if the protein extraction is hindered once the tissue is dried, the powdering process allows a greater release in the tissue suspension of total proteins and growth factors after tissue re-solubilization in comparison with only the freeze-drying process (+91 ± 13% for EGF, +16 ± 4% for HGF, +11 ± 5% for FGF, +16 ± 9% for TGF-β1), and a greater release of EGF (85 ± 10%) in comparison with only the freezing process, because proteins become much readily solubilized in the solution. According with these results, we describe a protocol to obtain a new sterile biological product from hAM tissue, with well-known effects of thermal, mechanical and physical processes on the total protein and grow factors contents.

Identification of Foreign Particles in Human Tissues using Raman Microscopy.

PubMed

Campion, Alan; Smith, Kenneth J; Fedulov, Alexey V; Gregory, David; Fan, Yuwei; Godleski, John J

2018-06-12

The precise identification of foreign particles in tissue for patient care and research has been studied using polarized light microscopy, electron microscopy with X-ray analysis, and electron diffraction. The goal of this study was to unambiguously identify particles in tissues using a combina-tion of polarized light microscopy and Raman microscopy, which provides chemical composition and microstructural characterization of complex materials with submicron spatial resolution. We designed a model system of stained and unstained cells that contained birefringent talc particles, and systematically investigated the influence of slide and coverslip materials, laser wavelengths, and mounting media on the Raman spectra ob-tained. Hematoxylin and eosin stained slides did not produce useful results because of fluorescence interference from the stains. Unstained cell samples prepared with standard slides and coverslips produce high quality Raman spectra when excited at 532 nm; the spectra are uniquely as-signed to talc. We also obtain high quality Raman spectra specific for talc in unstained tissue samples (pleural tissue following talc pleurodesis and ovarian tissue following long-term perineal talc exposure). Raman microscopy is sufficiently sensitive and compositionally selective to identify particles as small as one micron in diameter. Among commonly used coverslip mounting media, Cytoseal 60 is recommended; Permount was unacceptable due to intense background interference. Raman spectra have been catalogued for thousands of substances, which suggests that this approach is likely to be successful in identifying other particles of interest in tissues, potentially making Raman microscopy a powerful new tool in pathology.

Detecting quinoxaline-2-carboxylic acid in animal tissues by using sensitive rapid enzyme-linked immunosorbent assay and time-resolved fluoroimmunoassay.

PubMed

Le, Tao; Yu, Huan; Niu, Xiaodong

2015-05-15

An indirect competitive enzyme-linked immunosorbent assay (ic-ELISA) and time-resolved fluoroimmunoassay (TR-FIA) based on an anti-N-butylquinoxaline-2-carboxamide (BQCA) monoclonal antibody were standardized and validated for quinoxaline-2-carboxylic acid (QCA) screening in animal tissues and its performance were compared to HPLC. The sensitivities obtained for edible tissue extracts were 1.62 and 1.12 ng ml(-1) for ic-ELISA and TR-FIA detection, respectively. Two samples were spiked with QCA and analyzed by both methods. The recovery values ranged from 92.6% to 112.2% and the coefficients of variation were less than 15% for QCA spiking into swine tissue samples at concentrations of 2.5-50.0 μg kg(-1). Excellent correlations (r(2)=0.987-0.996) of the ic-ELISA/HPLC and TR-FIA/HPLC data were observed for processed samples. The results demonstrated that the ic-ELISA and TR-FIA methods were rapid and accurate for the residue detection of QCA in animal tissues. Copyright © 2014 Elsevier Ltd. All rights reserved.

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, in order to attain more detailed results that could be crucial in understanding the process of carcinogenesis.

An evaluation of the histological effects of intra-articular methadone in the canine model.

PubMed

Jones, Timothy A; Hand, Walter R; Ports, Michael D; Unger, Daniel V; Herbert, Daniel; Pellegrini, Joseph E

2003-02-01

Methadone hydrochloride is an opiate that has pharmacodynamic and pharmacokinetic properties that suggest it may provide longer analgesia than morphine when administered via the intra-articular route. However, no studies to date have been conducted examining the effects of intra-articular methadone hydrochloride on local tissues. Therefore, the purpose of this study was to determine the histopathologic effects of intra-articular methadone hydrochloride on local tissues in the canine knee. Nine canines, 1 to 4 years old, weighing between 20 kg and 23 kg were used. All canines had their knees randomized to receive either bupivacaine, 0.5% with epinephrine 1:200,000 (4.5 mL), and 5 mg methadone hydrochloride (0.5 mL) for the study knee, or bupivacaine, 0.5% with epinephrine 1:200,000 (4.5 mL), and 0.5 mL normal saline for the control knee. Serum methadone hydrochloride levels were obtained on all canines at 6 and 24 hours. Canines were randomly assigned to 1 of 3 groups to be euthanized at either 24 hours, 14 days, or 28 days. Following euthanization and necropsy, synovial fluid levels and tissue samples were obtained and examined for histopathologic changes. Synovial fluid samples noted a few white blood cells at 24 hours and none at 14 and 28 days. Tissue samples showed no histopathologic changes, and serum concentration levels of methadone hydrochloride were negligible.

A High-Throughput Method for Direct Detection of Therapeutic Oligonucleotide-Induced Gene Silencing In Vivo

PubMed Central

Coles, Andrew H.; Osborn, Maire F.; Alterman, Julia F.; Turanov, Anton A.; Godinho, Bruno M.D.C.; Kennington, Lori; Chase, Kathryn; Aronin, Neil

2016-01-01

Preclinical development of RNA interference (RNAi)-based therapeutics requires a rapid, accurate, and robust method of simultaneously quantifying mRNA knockdown in hundreds of samples. The most well-established method to achieve this is quantitative real-time polymerase chain reaction (qRT-PCR), a labor-intensive methodology that requires sample purification, which increases the potential to introduce additional bias. Here, we describe that the QuantiGene® branched DNA (bDNA) assay linked to a 96-well Qiagen TissueLyser II is a quick and reproducible alternative to qRT-PCR for quantitative analysis of mRNA expression in vivo directly from tissue biopsies. The bDNA assay is a high-throughput, plate-based, luminescence technique, capable of directly measuring mRNA levels from tissue lysates derived from various biological samples. We have performed a systematic evaluation of this technique for in vivo detection of RNAi-based silencing. We show that similar quality data is obtained from purified RNA and tissue lysates. In general, we observe low intra- and inter-animal variability (around 10% for control samples), and high intermediate precision. This allows minimization of sample size for evaluation of oligonucleotide efficacy in vivo. PMID:26595721

Various clinical application of phase contrast X-ray

NASA Astrophysics Data System (ADS)

Oh, Chilhwan; Park, Sangyong; Ha, Seunghan; Park, Gyuman; Lee, Gunwoo; Lee, Onseok; Je, Jungho

2008-02-01

In biomedical application study using phase contrast X-ray, both sample thickness or density and absorption difference are very important factors in aspects of contrast enhancement. We present experimental evidence that synchrotron hard X-ray are suitable for radiological imaging of biological samples down to the cellular level. We investigated the potential of refractive index radiology using un-monochromatized synchrotron hard X-rays for the imaging of cell and tissue in various diseases. Material had been adopted various medical field, such as apoE knockout mouse in cardiologic field, specimen from renal and prostatic carcinoma patient in urology, basal cell epithelioma in dermatology, brain tissue from autosy sample of pakinson's disease, artificially induced artilrtis tissue from rabbits and extracted tooth from patients of crack tooth syndrome. Formalin and paraffin fixed tissue blocks were cut in 3 mm thickness for the X-ray radiographic imaging. From adjacent areas, 4 μm thickness sections were also prepared for hematoxylin-eosin staining. Radiographic images of dissected tissues were obtained using the hard X-rays from the 7B2 beamline of the Pohang Light Source (PLS). The technique used for the study was the phase contrast images were compared with the optical microscopic images of corresponding histological slides. Radiographic images of various diseased tissues showed clear histological details of organelles in normal tissues. Most of cancerous lesions were well differentiated from adjacent normal tissues and detailed histological features of each tumor were clearly identified. Also normal microstructures were identifiable by the phase contrast imaging. Tissue in cancer or other disease showed clearly different findings from those of surrounding normal tissue. For the first time we successfully demonstrated that synchrotron hard X-rays can be used for radiological imaging of relatively thick tissue samples with great histological details.

Kinetic Modeling of PET Data Without Blood Sampling

NASA Astrophysics Data System (ADS)

Bentourkia, M.

2005-06-01

In positron emission tomography (PET) imaging, application of kinetic modeling always requires an input curve (IC) together with the PET data. The IC can be obtained by means of external blood sampling or, in the case of cardiac studies, by means of a region-of-interest (ROI) drawn on the blood pool. It is, however, very unsuitable to withdraw and to analyze blood samples, and in small animals, these operations become difficult, while ICs determined from ROIs are generally contaminated by emissions from neighboring sites, or they are underestimated because of partial volume effect. In this paper, we report a new method to extract kinetic parameters from dynamic PET studies without a priori knowledge of the IC. The method is applied in human brain data measured with fluorodeoxyglucose (FDG) human-brain and in cardiac-rat perfusion studies with /sup 13/N-ammonia and /sup 11/C-acetate. The tissue blood volume (TBV), usually fitted together with the rate constants, is extracted simultaneously with the tissue time activity curves for cardiac studies, while for brain gray matter, TBV is known to be about 4% to 7%. The shape of IC is obtained by means of factor analysis from an ROI drawn around a cardiac tissue or a brain artery. The results show a good correlation (p<0.05) between the cerebral metabolic rate of glucose, myocardial blood flow, and oxygen consumption obtained with the new method in comparison to the usual method. In conclusion, it is possible to apply kinetic modeling without any blood sampling, which significantly simplifies PET acquisition and data analysis.

Validation of an LC-MS/MS method for the quantification of choline-related compounds and phospholipids in foods and tissues.

PubMed

Xiong, Yeping; Zhao, Yuan-Yuan; Goruk, Sue; Oilund, Kirsten; Field, Catherine J; Jacobs, René L; Curtis, Jonathan M

2012-12-12

A hydrophilic interaction liquid chromatography-tandem mass spectrometry (HILIC LC-MS/MS) method was developed and validated to simultaneously quantify six aqueous choline-related compounds and eight major phospholipids classes in a single run. HILIC chromatography was coupled to positive ion electrospray mass spectrometry. A combination of multiple scan modes including precursor ion scan, neutral loss scan and multiple reaction monitoring was optimized for the determination of each compound or class in a single LC/MS run. This work developed a simplified extraction scheme in which both free choline and related compounds along with phospholipids were extracted into a homogenized phase using chloroform/methanol/water (1:2:0.8) and diluted into methanol for the analysis of target compounds in a variety of sample matrices. The analyte recoveries were evaluated by spiking tissues and food samples with two isotope-labeled internal standards, PC-d(3) and Cho-d(3). Recoveries of between 90% and 115% were obtained by spiking a range of sample matrices with authentic standards containing all 14 of the target analytes. The precision of the analysis ranged from 1.6% to 13%. Accuracy and precision was comparable to that obtained by quantification of selected phospholipid classes using (31)P NMR. A variety of sample matrices including egg yolks, human diets and animal tissues were analyzed using the validated method. The measurements of total choline in selected foods were found to be in good agreement with values obtained from the USDA choline database. Copyright © 2012 Elsevier B.V. All rights reserved.

Spatial mapping of metals in tissue-sections using combination of mass-spectrometry and histology through image registration

NASA Astrophysics Data System (ADS)

Anyz, Jiri; Vyslouzilova, Lenka; Vaculovic, Tomas; Tvrdonova, Michaela; Kanicky, Viktor; Haase, Hajo; Horak, Vratislav; Stepankova, Olga; Heger, Zbynek; Adam, Vojtech

2017-01-01

We describe a new procedure for the parallel mapping of selected metals in histologically characterized tissue samples. Mapping is achieved via image registration of digital data obtained from two neighbouring cryosections by scanning the first as a histological sample and subjecting the second to laser ablation inductively coupled plasma mass spectrometry. This computer supported procedure enables determination of the distribution and content of metals of interest directly in the chosen histological zones and represents a substantial improvement over the standard approach, which determines these values in tissue homogenates or whole tissue sections. The potential of the described procedure was demonstrated in a pilot study that analysed Zn and Cu levels in successive development stages of pig melanoma tissue using MeLiM (Melanoma-bearing-Libechov-Minipig) model. We anticipate that the procedure could be useful for a complex understanding of the role that the spatial distribution of metals plays within tissues affected by pathological states including cancer.

A polarization sensitive hyperspectral imaging system for detection of differences in tissue properties

NASA Astrophysics Data System (ADS)

Peller, Joseph A.; Ceja, Nancy K.; Wawak, Amanda J.; Trammell, Susan R.

2018-02-01

Polarized light imaging and optical spectroscopy can be used to distinguish between healthy and diseased tissue. In this study, the design and testing of a single-pixel hyperspectral imaging system that uses differences in the polarization of light reflected from tissue to differentiate between healthy and thermally damaged tissue is discussed. Thermal lesions were created in porcine skin (n = 8) samples using an IR laser. The damaged regions were clearly visible in the polarized light hyperspectral images. Reflectance hyperspectral and white light imaging was also obtained for all tissue samples. Sizes of the thermally damaged regions as measured via polarized light hyperspectral imaging are compared to sizes of these regions as measured in the reflectance hyperspectral images and white light images. Good agreement between the sizes measured by all three imaging modalities was found. Hyperspectral polarized light imaging can differentiate between healthy and damaged tissue. Possible applications of this imaging system include determination of tumor margins during cancer surgery or pre-surgical biopsy.

Spatial mapping of metals in tissue-sections using combination of mass-spectrometry and histology through image registration

PubMed Central

Anyz, Jiri; Vyslouzilova, Lenka; Vaculovic, Tomas; Tvrdonova, Michaela; Kanicky, Viktor; Haase, Hajo; Horak, Vratislav; Stepankova, Olga; Heger, Zbynek; Adam, Vojtech

2017-01-01

We describe a new procedure for the parallel mapping of selected metals in histologically characterized tissue samples. Mapping is achieved via image registration of digital data obtained from two neighbouring cryosections by scanning the first as a histological sample and subjecting the second to laser ablation inductively coupled plasma mass spectrometry. This computer supported procedure enables determination of the distribution and content of metals of interest directly in the chosen histological zones and represents a substantial improvement over the standard approach, which determines these values in tissue homogenates or whole tissue sections. The potential of the described procedure was demonstrated in a pilot study that analysed Zn and Cu levels in successive development stages of pig melanoma tissue using MeLiM (Melanoma-bearing-Libechov-Minipig) model. We anticipate that the procedure could be useful for a complex understanding of the role that the spatial distribution of metals plays within tissues affected by pathological states including cancer. PMID:28071735

Application of SEC-ICP-MS for comparative analyses of metal-containing species in cancerous and healthy human thyroid samples.

PubMed

Boulyga, Sergei F; Loreti, Valeria; Bettmer, Jörg; Heumann, Klaus G

2004-09-01

Size exclusion chromatography (SEC) was coupled on-line to inductively coupled plasma mass spectrometry (ICP-MS) for speciation study of trace metals in cancerous thyroid tissues in comparison to healthy thyroids aimed to estimation of changes in metalloprotein speciation in pathological tissue. The study showed a presence of species binding Cu, Zn, Cd and Pb in healthy thyroid tissue with a good reproducibility of chromatographic results, whereas the same species could not be detected in cancerous tissues. Thus, remarkable differences with respect to metal-binding species were revealed between healthy and pathological thyroid samples, pointing out a completely different distribution of trace metals in cancerous tissues. The metal-binding species could not be identified in the frame of this work because of a lack of appropriate standards. Nevertheless, the results obtained confirm the suitability of SEC-ICP-MS for monitoring of changes in trace metal distribution in cancerous tissue and will help to better understand the role of metal-containing species in thyroid pathology.

Microscopic fluorescence spectral analysis of basal cell carcinomas

NASA Astrophysics Data System (ADS)

He, Qingli; Lui, Harvey; Zloty, David; Cowan, Bryce; Warshawski, Larry; McLean, David I.; Zeng, Haishan

2007-05-01

Background and Objectives. Laser-induced autofluorescence (LIAF) is a promising tool for cancer diagnosis. This method is based on the differences in autofluorescence spectra between normal and cancerous tissues, but the underlined mechanisms are not well understood. The objective of this research is to study the microscopic origins and intrinsic fluorescence properties of basal cell carcinoma (BCC) for better understanding of the mechanism of in vivo fluorescence detection and margin delineation of BCCs on skin patients. A home-made micro- spectrophotometer (MSP) system was used to image the fluorophore distribution and to measure the fluorescence spectra of various microscopic structures and regions on frozen tissue sections. Materials and Methods. BCC tissue samples were obtained from 14 patients undergoing surgical resections. After surgical removal, each tissue sample was immediately embedded in OCT medium and snap-frozen in liquid nitrogen. The frozen tissue block was then cut into 16-μm thickness sections using a cryostat microtome and placed on microscopic glass slides. The sections for fluorescence study were kept unstained and unfixed, and then analyzed by the MSP system. The adjacent tissue sections were H&E stained for histopathological examination and also served to help identify various microstructures on the adjacent unstained sections. The MSP system has all the functions of a conventional microscope, plus the ability of performing spectral analysis on selected micro-areas of a microscopic sample. For tissue fluorescence analysis, 442nm He-Cd laser light is used to illuminate and excite the unstained tissue sections. A 473-nm long pass filter was inserted behind the microscope objective to block the transmitted laser light while passing longer wavelength fluorescence signal. The fluorescence image of the sample can be viewed through the eyepieces and also recorded by a CCD camera. An optical fiber is mounted onto the image plane of the photograph port of the microscope to collect light from a specific micro area of the sample. The collected light is transmitted via the fiber to a disperserve type CCD spectrometer for spectral analysis. Results. The measurement results showed significant spectral differences between normal and cancerous tissues. For normal tissue regions, the spectral results agreed with our previous findings on autofluorescence of normal skin sections. For the cancerous regions, the epidermis showed very weak fluorescence signal, while the stratum corneum exhibited fluorescence emissions peaking at about 510 nm. In the dermis, the basal cell island and a band of surrounding areas showed very weak fluorescence signal, while distal dermis above and below the basal cell island showed greater fluorescence signal but with different spectral shapes. The very weak autofluorescence from the basal cell island and its surrounding area may be attributed to their degenerative properties that limited the production of collagens. Conclusions. The obtained microscopic results very well explain the in vivo fluorescence properties of BCC lesions in that they have decreased fluorescence intensity compared to the surrounding normal skin. The intrinsic spectra of various microstructures and the microscopic fluorescence images (corresponding fluorophore distribution in tissue) obtained in this study will be used for further theoretical modeling of in vivo fluorescence spectroscopy and imaging of skin cancers.

In vivo microsampling to capture the elusive exposome

NASA Astrophysics Data System (ADS)

Bessonneau, Vincent; Ings, Jennifer; McMaster, Mark; Smith, Richard; Bragg, Leslie; Servos, Mark; Pawliszyn, Janusz

2017-03-01

Loss and/or degradation of small molecules during sampling, sample transportation and storage can adversely impact biological interpretation of metabolomics data. In this study, we performed in vivo sampling using solid-phase microextraction (SPME) in combination with non-targeted liquid chromatography and high-resolution tandem mass spectrometry (LC-MS/MS) to capture the fish tissue exposome using molecular networking analysis, and the results were contrasted with molecular differences obtained with ex vivo SPME sampling. Based on 494 MS/MS spectra comparisons, we demonstrated that in vivo SPME sampling provided better extraction and stabilization of highly reactive molecules, such as 1-oleoyl-sn-glycero-3-phosphocholine and 1-palmitoleoyl-glycero-3-phosphocholine, from fish tissue samples. This sampling approach, that minimizes sample handling and preparation, offers the opportunity to perform longitudinal monitoring of the exposome in biological systems and improve the reliability of exposure-measurement in exposome-wide association studies.

In vivo microsampling to capture the elusive exposome

PubMed Central

Bessonneau, Vincent; Ings, Jennifer; McMaster, Mark; Smith, Richard; Bragg, Leslie; Servos, Mark; Pawliszyn, Janusz

2017-01-01

Loss and/or degradation of small molecules during sampling, sample transportation and storage can adversely impact biological interpretation of metabolomics data. In this study, we performed in vivo sampling using solid-phase microextraction (SPME) in combination with non-targeted liquid chromatography and high-resolution tandem mass spectrometry (LC-MS/MS) to capture the fish tissue exposome using molecular networking analysis, and the results were contrasted with molecular differences obtained with ex vivo SPME sampling. Based on 494 MS/MS spectra comparisons, we demonstrated that in vivo SPME sampling provided better extraction and stabilization of highly reactive molecules, such as 1-oleoyl-sn-glycero-3-phosphocholine and 1-palmitoleoyl-glycero-3-phosphocholine, from fish tissue samples. This sampling approach, that minimizes sample handling and preparation, offers the opportunity to perform longitudinal monitoring of the exposome in biological systems and improve the reliability of exposure-measurement in exposome-wide association studies. PMID:28266605

Synthesis and characterization of poly (lactic acid)/chitosan nanocomposites based on renewable resources as biobased-material

NASA Astrophysics Data System (ADS)

Suryani; Agusnar, H.; Wirjosentono, B.; Rihayat, T.; Salisah, Z.

2018-01-01

Biobased becomes one of the new breakthrough in the smart engineering, especially in biomedical applications, such as tissue engineering that serves as a supporting physical structure to trigger the growth of skin tissue. From various studies which had been done, it was known that the optimal Biobased healed wounds or injuries in a relatively short time. In this study, a Biobased natural polymer based e.g Poly(Lactic Acid) (PLA)/Chitosan Nanocomposites was made. PLA was synthesized from saba banana (Musa acuminata) as raw material using Ring-Opening Polymerization (ROP) method. PLA was mixed with Chitosan with Chitosan concentration variations of 1%, 3%, and 5% to form a nanocomposites. The analysis result showed that Chitosan concentration in PLA/Chitosan Nanocomposites sample affected the value of tensile strength. The highest value of tensile strength was obtained on a sample of 100 ml volume with a concentration of 3%, which was 120.396 MPa. The highest percentage of elongation was obtained in 100 ml volume sample with 5% concentration, which was 26.3686%. In the hydrophilicity test, the highest percentage of water absorption was obtained in a 200 ml volume sample with 5% concentration, which was 44.615%. The addition of Chitosan to the sample affected the functional group bonding, where there was a functional group of NH2 at the wave number of 2923.92 cm-1. The sample characteristics based on water absorption indicated that the sample was potentially to be used as Biobased construction material.

Vibrational Profiling of Brain Tumors and Cells

PubMed Central

Nelson, Sultan L; Proctor, Dustin T; Ghasemloonia, Ahmad; Lama, Sanju; Zareinia, Kourosh; Ahn, Younghee; Al-Saiedy, Mustafa R; Green, Francis HY; Amrein, Matthias W; Sutherland, Garnette R

2017-01-01

This study reports vibration profiles of neuronal cells and tissues as well as brain tumor and neocortical specimens. A contact-free method and analysis protocol was designed to convert an atomic force microscope into an ultra-sensitive microphone with capacity to record and listen to live biological samples. A frequency of 3.4 Hz was observed for both cultured rat hippocampal neurons and tissues and vibration could be modulated pharmacologically. Malignant astrocytoma tissue samples obtained from operating room, transported in artificial cerebrospinal fluid, and tested within an hour, vibrated with a much different frequency profile and amplitude, compared to meningioma or lateral temporal cortex providing a quantifiable measurement to accurately distinguish the three tissues in real-time. Vibration signals were converted to audible sound waves by frequency modulation, thus demonstrating, acoustic patterns unique to meningioma, malignant astrocytoma and neocortex. PMID:28744324

Real time monitoring of pulsatile change in hemoglobin concentrations of cerebral tissue by a portable tissue oximeter with a 10-Hz sampling rate

NASA Astrophysics Data System (ADS)

Shiga, Toshikazu; Chihara, Eiichi; Tanabe, Kazuhisa; Tanaka, Yoshifumi; Yamamoto, Katsuyuki

1998-01-01

A portable CW tissue oximeter of a 10-Hz sampling rate was developed for examination of pulsatile components of the output signals as a mean of checking the signal reliability during long-term monitoring. Feasible studies were performed on a healthy subject. Changes in Hb and HbO2 signals of cerebral tissue were continuously measured by placing a photoprobe on the forehead during 6-hour sleep. Pulsatile changes in Hb and HbO2 were steadily observed over a whole period of the recording. The phase relation of pulsation in Hb and HbO2 was almost inverse. Not only information for reliable monitoring but also physiological parameters with respect to cerebral circulation and metabolism could be obtained by measuring the pulsatile components.

Real time monitoring of pulsatile change in hemoglobin concentrations of cerebral tissue by a portable tissue oximeter with a 10-Hz sampling rate

NASA Astrophysics Data System (ADS)

Shiga, Toshikazu; Chihara, Eiichi; Tanabe, Kazuhisa; Tanaka, Yoshifumi; Yamamoto, Katsuyuki

1997-12-01

A portable CW tissue oximeter of a 10-Hz sampling rate was developed for examination of pulsatile components of the output signals as a mean of checking the signal reliability during long-term monitoring. Feasible studies were performed on a healthy subject. Changes in Hb and HbO2 signals of cerebral tissue were continuously measured by placing a photoprobe on the forehead during 6-hour sleep. Pulsatile changes in Hb and HbO2 were steadily observed over a whole period of the recording. The phase relation of pulsation in Hb and HbO2 was almost inverse. Not only information for reliable monitoring but also physiological parameters with respect to cerebral circulation and metabolism could be obtained by measuring the pulsatile components.

Comparison of Follicular and Luteal Phase Mucosal Markers of HIV Susceptibility in Healthy Women

PubMed Central

Chandra, Neelima; Yousefieh, Nazita; Zalenskaya, Irina; Kimble, Thomas; Asin, Susana; Rollenhagen, Christiane; Anderson, Sharon M.; Herold, Betsy; Mesquita, Pedro M.M.; Richardson-Harman, Nicola; Cunningham, Tina; Schwartz, Jill L.; Doncel, Gustavo F.

2016-01-01

Abstract The purpose of this study was to evaluate differences in vaginal immune cell populations, vaginal tissue gene expression, antimicrobial activity of the cervicovaginal (CV) lavage (CVL), vaginal flora, and p24 antigen production from CV tissues after ex vivo human immunodeficiency virus (HIV) infection between follicular (FOL) and luteal (LUT) phases of the menstrual cycle. CV tissue biopsies, CV secretions, and blood samples were obtained as part of two longitudinal clinical trials of healthy women (CONRAD D11-119 and A12-124 studies). Participants (n = 39) were HIV-seronegative women not using exogenous hormone supplementation, with normal menstrual cycles, who were screened to exclude sexually transmitted and reproductive tract infections. Serum levels of estradiol and progesterone were significantly higher in the LUT versus the FOL phase of the menstrual cycle. Controlling for race, reported contraceptive use/sexual practices, and clinical trial, we found no differences in vaginal tissue immune cell populations and activation status, transcriptomes, inhibition of HIV, herpes simplex virus type 2 and Escherichia coli by the CVL, vaginal pH or Nugent score, or production of p24 antigen after ex vivo infection by HIV-1BaL between CV samples obtained in the FOL phase versus the LUT phase of the menstrual cycle. There were no significant correlations between serum estradiol and progesterone levels and CV endpoints. The hypothesis that the LUT phase of the menstrual cycle represents a more vulnerable stage for mucosal infection with HIV was not supported by data from samples obtained from the lower genital tract (ectocervix and vagina) from these two clinical trials. PMID:26750085

Characterizing viscoelastic mechanical properties of highly compliant polymers and biological tissues using impact indentation.

PubMed

Mijailovic, Aleksandar S; Qing, Bo; Fortunato, Daniel; Van Vliet, Krystyn J

2018-04-15

Precise and accurate measurement of viscoelastic mechanical properties becomes increasingly challenging as sample stiffness decreases to elastic moduli <1 kPa, largely due to difficulties detecting initial contact with the compliant sample surface. This limitation is particularly relevant to characterization of biological soft tissues and compliant gels. Here, we employ impact indentation which, in contrast to shear rheology and conventional indentation, does not require contact detection a priori, and present a novel method to extract viscoelastic moduli and relaxation time constants directly from the impact response. We first validate our approach by using both impact indentation and shear rheology to characterize polydimethylsiloxane (PDMS) elastomers of stiffness ranging from 100 s of Pa to nearly 10 kPa. Assuming a linear viscoelastic constitutive model for the material, we find that the moduli and relaxation times obtained from fitting the impact response agree well with those obtained from fitting the rheological response. Next, we demonstrate our validated method on hydrated, biological soft tissues obtained from porcine brain, murine liver, and murine heart, and report the equilibrium shear moduli, instantaneous shear moduli, and relaxation time constants for each tissue. Together, our findings provide a new and straightforward approach capable of probing local mechanical properties of highly compliant viscoelastic materials with millimeter scale spatial resolution, mitigating complications involving contact detection or sample geometric constraints. Characterization and optimization of mechanical properties can be essential for the proper function of biomaterials in diverse applications. However, precise and accurate measurement of viscoelastic mechanical properties becomes increasingly difficult with increased compliance (particularly for elastic moduli <1 kPa), largely due to challenges detecting initial contact with the compliant sample surface and measuring response at short timescale or high frequency. By contrast, impact indentation has highly accurate contact detection and can be used to measure short timescale (glassy) response. Here, we demonstrate an experimental and analytical method that confers significant advantages over existing approaches to extract spatially resolved viscoelastic moduli and characteristic time constants of biological tissues (e.g., brain and heart) and engineered biomaterials. Copyright © 2018 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Increasing the speed of tumour diagnosis during surgery with selective scanning Raman microscopy

NASA Astrophysics Data System (ADS)

Kong, Kenny; Rowlands, Christopher J.; Varma, Sandeep; Perkins, William; Leach, Iain H.; Koloydenko, Alexey A.; Pitiot, Alain; Williams, Hywel C.; Notingher, Ioan

2014-09-01

One of the main challenges in cancer surgery is ensuring that all tumour cells are removed during surgery, while sparing as much healthy tissue as possible. Histopathology, the gold-standard technique for cancer diagnosis, is often impractical for intra-operative use because of the time-consuming tissue preparation procedures (sectioning and staining). Raman micro-spectroscopy is a powerful technique that can discriminate between tumours and healthy tissues with high accuracy, based entirely on intrinsic chemical differences. However, raster-scanning Raman micro-spectroscopy is a slow imaging technique that typically requires data acquisition times as long as several days for typical tissue samples obtained during surgery (1 × 1 cm2) - in particular when high signal-to-noise ratio spectra are required to ensure accurate diagnosis. In this paper we present two techniques based on selective sampling Raman micro-spectroscopy that can overcome these limitations. In selective sampling, information regarding the spatial features of the tissue, either measured by an alternative optical technique or estimated in real-time from the Raman spectra, can be used to drastically reduce the number of Raman spectra required for diagnosis. These sampling strategies allowed diagnosis of basal cell carcinoma in skin tissue samples excised during Mohs micrographic surgery faster than frozen section histopathology, and two orders of magnitude faster than previous techniques based on raster-scanning Raman microscopy. Further development of these techniques may help during cancer surgery by providing a fast and objective way for surgeons to ensure the complete removal of tumour cells while sparing as much healthy tissue as possible.

Introducing Human Population Biology through an Easy Laboratory Exercise on Mitochondrial DNA

ERIC Educational Resources Information Center

Pardinas, Antonio F.; Dopico, Eduardo; Roca, Agustin; Garcia-Vazquez, Eva; Lopez, Belen

2010-01-01

This article describes an easy and cheap laboratory exercise for students to discover their own mitochondrial haplogroup. Students use buccal swabs to obtain mucosa cells as noninvasive tissue samples, extract DNA, and with a simple polymerase chain reaction-restriction fragment length polymorphism analysis they can obtain DNA fragments of…

Microultrasound characterisation of ex vivo porcine tissue for ultrasound capsule endoscopy

NASA Astrophysics Data System (ADS)

Lay, H. S.; Cox, B. F.; Sunoqrot, M.; Démoré, C. E. M.; Näthke, I.; Gomez, T.; Cochran, S.

2017-01-01

Gastrointestinal (GI) disease development and progression is often characterised by cellular and tissue architectural changes within the mucosa and sub-mucosa layers. Current clinical capsule endoscopy and other approaches are heavily reliant on optical techniques which cannot detect disease progression below the surface layer of the tissue. To enhance the ability of clinicians to detect cellular changes earlier and more confidently, both quantitative and qualitative microultrasound (μUS) techniques are investigated in healthy ex vivo porcine GI tissue. This work is based on the use of single-element, focussed μUS transducers made with micromoulded piezocomposite operating at around 48 MHz. To explore the possibility that μUS can detect Crohn’s disease and other inflammatory bowel diseases, ex vivo porcine small bowel tissue samples were cannulised and perfused with phosphate-buffered saline followed by various dilutions of polystyrene microspheres. Comparison with fluorescent imaging showed that the microspheres had infiltrated the microvasculature of the samples and that μUS was able to successfully detect this as a mimic of inflammation. Samples without microspheres were analysed using quantitative ultrasound to assess mechanical properties. Attenuation coefficients of 1.78 ± 0.66 dB/mm and 1.92 ± 0.77 dB/mm were obtained from reference samples which were surgically separated from the muscle layer. Six intact samples were segmented using a software algorithm and the acoustic impedance, Z, for varying tissue thicknesses, and backscattering coefficient, BSC, were calculated using the reference attenuation values and tabulated.

Class I and II histone deacetylase expression in human chronic periodontitis gingival tissue.

PubMed

Cantley, M D; Dharmapatni, A A S S K; Algate, K; Crotti, T N; Bartold, P M; Haynes, D R

2016-04-01

Histone deacetylase inhibitors (HDACi) are being considered to treat chronic inflammatory diseases at low doses. Currently HDACi that are more specific are being developed to target particular HDACs; therefore, this study aimed to determine levels and distribution of class I and II HDAC in human gingival samples obtained from patients with chronic periodontitis. Gingival biopsies were obtained from patients with and without (mild inflammation, no bone loss) periodontitis. Total RNA was isolated for real-time quantitative polymerase chain reaction to determine expression of HDACs 1-10. Immunohistochemistry was used to determine protein distribution of HDACs 1, 5, 8 and 9. Factor VIII, CD3 and tartrate resistant acid phosphatase (TRAP) were detected in serial sections to identify blood vessels, lymphocytes, pre-osteoclasts and osteoclasts cells respectively. Tumour necrosis factor α (TNF-α) expression was also assessed. mRNA for HDAC 1, 5, 8 and 9 were significantly upregulated in chronic periodontitis gingival tissues compared to non-periodontitis samples (p < 0.05). Significantly higher HDAC 1 protein expression was observed in chronic periodontitis samples (p < 0.05), and was associated with CD3, TRAP and TNF-α-positive cells. HDAC 1, 5, 8 and 9 were expressed strongly by the factor VIII-positive microvasculature in the chronic periodontitis gingival tissues. HDAC 1, 5, 8 and 9 expression was higher in gingival tissues from patients with chronic periodontitis compared to non-periodontitis samples. Results suggest that these HDACs could therefore be targeted with specific acting HDACi. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Short Communication An efficient method for simultaneous extraction of high-quality RNA and DNA from various plant tissues.

PubMed

Oliveira, R R; Viana, A J C; Reátegui, A C E; Vincentz, M G A

2015-12-29

Determination of gene expression is an important tool to study biological processes and relies on the quality of the extracted RNA. Changes in gene expression profiles may be directly related to mutations in regulatory DNA sequences or alterations in DNA cytosine methylation, which is an epigenetic mark. Correlation of gene expression with DNA sequence or epigenetic mark polymorphism is often desirable; for this, a robust protocol to isolate high-quality RNA and DNA simultaneously from the same sample is required. Although commercial kits and protocols are available, they are mainly optimized for animal tissues and, in general, restricted to RNA or DNA extraction, not both. In the present study, we describe an efficient and accessible method to extract both RNA and DNA simultaneously from the same sample of various plant tissues, using small amounts of starting material. The protocol was efficient in the extraction of high-quality nucleic acids from several Arabidopsis thaliana tissues (e.g., leaf, inflorescence stem, flower, fruit, cotyledon, seedlings, root, and embryo) and from other tissues of non-model plants, such as Avicennia schaueriana (Acanthaceae), Theobroma cacao (Malvaceae), Paspalum notatum (Poaceae), and Sorghum bicolor (Poaceae). The obtained nucleic acids were used as templates for downstream analyses, such as mRNA sequencing, quantitative real time-polymerase chain reaction, bisulfite treatment, and others; the results were comparable to those obtained with commercial kits. We believe that this protocol could be applied to a broad range of plant species, help avoid technical and sampling biases, and facilitate several RNA- and DNA-dependent analyses.

Preparation and Application of an Innovative Thrombocyte/Leukocyte-Enriched Plasma to Promote Tissue Repair in Chelonians

PubMed Central

Di Ianni, Francesco; Merli, Elisa; Burtini, Francesca; Conti, Virna; Pelizzone, Igor; Di Lecce, Rosanna; Parmigiani, Enrico; Squassino, Gian Paolo; Del Bue, Maurizio; Lucarelli, Enrico; Ramoni, Roberto; Grolli, Stefano

2015-01-01

Platelet concentrates are widely used in mammalian regenerative medicine to improve tissue healing. Chelonians (Testudines) would benefit from the application of thrombocyte preparations to regenerate damaged tissues, since traumatic injuries are leading causes of morbidity and mortality for both wild-living and domesticated animals. The aim of this study was to establish a protocol that optimized the recovery of the thrombocytes from blood samples and to show the efficacy of thrombocyte-enriched plasma in chelonians. Peripheral blood samples were obtained from Testudo spp. (n = 12) and Trachemys scripta elegans (n = 10). Blood cells were fractionated by sodium diatrizoate-sodium polysucrose density gradient using a two-step centrifugation protocol. Thrombocytes and leukocytes were isolated and resuspended to obtain thrombocyte-leucocyte rich plasma (TLRP). The mean recovery of leukocytes and thrombocytes was 48.9% (±4.0 SEM, n = 22) of the whole blood cell content. No statistically significant difference was observed between blood samples collected from different turtle species. The ability of TLRP to form a gel was evaluated by adding variable concentrations of calcium gluconate at room temperature and at 37°C. A reliable and consistent clotting of the TLRP was obtained in glass tubes and dishes by adding 5-20% v/v of a 100 mg/ml solution of calcium gluconate. Furthermore, in order to test the clinical efficacy of TLRP, a preliminary evaluation was performed on four turtles (Testudo spp.) with traumatic injuries. In all the four animals, a successful clinical outcome was observed. The results demonstrated that a thrombocyte-enriched plasma, comparable to mammalian platelet rich plasma, can be prepared from chelonian blood samples. Furthermore, although the low number of cases presented does not allow definitive conclusions from a clinical point of view, their outcome suggests that TLRP application could be further investigated to improve the healing process of both soft and hard tissue injuries in chelonians. PMID:25901960

Near-infrared confocal micro-Raman spectroscopy combined with PCA-LDA multivariate analysis for detection of esophageal cancer

NASA Astrophysics Data System (ADS)

Chen, Long; Wang, Yue; Liu, Nenrong; Lin, Duo; Weng, Cuncheng; Zhang, Jixue; Zhu, Lihuan; Chen, Weisheng; Chen, Rong; Feng, Shangyuan

2013-06-01

The diagnostic capability of using tissue intrinsic micro-Raman signals to obtain biochemical information from human esophageal tissue is presented in this paper. Near-infrared micro-Raman spectroscopy combined with multivariate analysis was applied for discrimination of esophageal cancer tissue from normal tissue samples. Micro-Raman spectroscopy measurements were performed on 54 esophageal cancer tissues and 55 normal tissues in the 400-1750 cm-1 range. The mean Raman spectra showed significant differences between the two groups. Tentative assignments of the Raman bands in the measured tissue spectra suggested some changes in protein structure, a decrease in the relative amount of lactose, and increases in the percentages of tryptophan, collagen and phenylalanine content in esophageal cancer tissue as compared to those of a normal subject. The diagnostic algorithms based on principal component analysis (PCA) and linear discriminate analysis (LDA) achieved a diagnostic sensitivity of 87.0% and specificity of 70.9% for separating cancer from normal esophageal tissue samples. The result demonstrated that near-infrared micro-Raman spectroscopy combined with PCA-LDA analysis could be an effective and sensitive tool for identification of esophageal cancer.

Photoacoustic imaging in both soft and hard biological tissue

NASA Astrophysics Data System (ADS)

Li, T.; Dewhurst, R. J.

2010-03-01

To date, most Photoacoustic (PA) imaging results have been from soft biotissues. In this study, a PA imaging system with a near-infrared pulsed laser source has been applied to obtain 2-D and 3-D images from both soft tissue and post-mortem dental samples. Imaging results showed that the PA technique has the potential to image human oral disease, such as early-stage teeth decay. For non-invasive photoacoustic imaging, the induced temperature and pressure rises within biotissues should not cause physical damage to the tissue. Several simulations based on the thermoelastic effect have been applied to predict initial temperature and pressure fields within a tooth sample. Predicted initial temperature and pressure rises are below corresponding safety limits.

PIXE analysis of elements in gastric cancer and adjacent mucosa

NASA Astrophysics Data System (ADS)

Liu, Qixin; Zhong, Ming; Zhang, Xiaofeng; Yan, Lingnuo; Xu, Yongling; Ye, Simao

1990-04-01

The elemental regional distributions in 20 resected human stomach tissues were obtained using PIXE analysis. The samples were pathologically divided into four types: normal, adjacent mucosa A, adjacent mucosa B and cancer. The targets for PIXE analysis were prepared by wet digestion with a pressure bomb system. P, K, Fe, Cu, Zn and Se were measured and statistically analysed. We found significantly higher concentrations of P, K, Cu, Zn and a higher ratio of Cu compared to Zn in cancer tissue as compared with normal tissue, but statistically no significant difference between adjacent mucosa and cancer tissue was found.

A Method to Evaluate Genome-Wide Methylation in Archival Formalin-Fixed, Paraffin-Embedded Ovarian Epithelial Cells

PubMed Central

Li, Qiling; Li, Min; Ma, Li; Li, Wenzhi; Wu, Xuehong; Richards, Jendai; Fu, Guoxing; Xu, Wei; Bythwood, Tameka; Li, Xu; Wang, Jianxin; Song, Qing

2014-01-01

Background The use of DNA from archival formalin and paraffin embedded (FFPE) tissue for genetic and epigenetic analyses may be problematic, since the DNA is often degraded and only limited amounts may be available. Thus, it is currently not known whether genome-wide methylation can be reliably assessed in DNA from archival FFPE tissue. Methodology/Principal Findings Ovarian tissues, which were obtained and formalin-fixed and paraffin-embedded in either 1999 or 2011, were sectioned and stained with hematoxylin-eosin (H&E).Epithelial cells were captured by laser micro dissection, and their DNA subjected to whole genomic bisulfite conversion, whole genomic polymerase chain reaction (PCR) amplification, and purification. Sequencing and software analyses were performed to identify the extent of genomic methylation. We observed that 31.7% of sequence reads from the DNA in the 1999 archival FFPE tissue, and 70.6% of the reads from the 2011 sample, could be matched with the genome. Methylation rates of CpG on the Watson and Crick strands were 32.2% and 45.5%, respectively, in the 1999 sample, and 65.1% and 42.7% in the 2011 sample. Conclusions/Significance We have developed an efficient method that allows DNA methylation to be assessed in archival FFPE tissue samples. PMID:25133528

A New Paradigm for Tissue Diagnostics: Tools and Techniques to Standardize Tissue Collection, Transport, and Fixation.

PubMed

Bauer, Daniel R; Otter, Michael; Chafin, David R

2018-01-01

Studying and developing preanalytical tools and technologies for the purpose of obtaining high-quality samples for histological assays is a growing field. Currently, there does not exist a standard practice for collecting, fixing, and monitoring these precious samples. There has been some advancement in standardizing collection for the highest profile tumor types, such as breast, where HER2 testing drives therapeutic decisions. This review examines the area of tissue collection, transport, and monitoring of formalin diffusion and details a prototype system that could be used to help standardize tissue collection efforts. We have surveyed recent primary literature sources and conducted several site visits to understand the most error-prone processes in histology laboratories. This effort identified errors that resulted from sample collection techniques and subsequent transport delays from the operating room (OR) to the histology laboratories. We have therefore devised a prototype sample collection and transport concept. The system consists of a custom data logger and cold transport box and takes advantage of a novel cold + warm (named 2 + 2) fixation method. This review highlights the beneficial aspects of standardizing tissue collection, fixation, and monitoring. In addition, a prototype system is introduced that could help standardize these processes and is compatible with use directly in the OR and from remote sites.

IgG and IgM anti-snRNP reactivity in sequentially obtained serum samples from patients with connective tissue diseases.

PubMed Central

Nyman, U; Lundberg, I; Hedfors, E; Wahren, M; Pettersson, I

1992-01-01

Sequentially obtained serum samples from 30 patients with connective tissue disease positive for antibody to ribonucleoprotein (RNP) were examined to determine the specificities of IgG and IgM antibodies to snRNP during the disease course using immunoblotting of nuclear extracts. The antibody patterns were correlated with disease activity. The patterns of antibody to snRNP of individual patients were mainly stable during the study but changes in levels of antibody to snRNP were seen corresponding to changes in clinical activity. These results indicate that increased reactivity of serum IgM antibodies against the B/B' proteins seems to precede a clinically evident exacerbation of disease whereas IgG antibody reactivity to the 70 K protein peaks at the time of a disease flare. Images PMID:1485812

Histological study of subcutaneous fat at NIR laser treatment of the rat skin in vivo

NASA Astrophysics Data System (ADS)

Yanina, I. Y.; Svenskaya, Yu. I.; Navolokin, N. A.; Matveeva, O. V.; Bucharskaya, A. B.; Maslyakova, G. N.; Gorin, D. A.; Sukhorukov, G. B.; Tuchin, V. V.

2015-07-01

The goal of this work is to quantify impact of in vivo photochemical treatment using indocyanine green (ICG) or encapsulated ICG and NIR laser irradiation through skin of rat with obesity by the follow up tissue sampling and histochemistry. After 1 hour elapsed since 1-min light exposure samples of rat skin with subcutaneous tissue of thickness of 1.5-2.5 mm were taken by surgery from rats within marked 4-zones of the skin site. For hematoxylin-eosin histological examination of excised tissue samples, fixation was carried out by 10%-formaldehyde solution. For ICG and encapsulated ICG subcutaneous injection and subsequent 1-min diode laser irradiation with power density of 8 W/cm2, different necrotic regions with lipolysis of subcutaneous fat were observed. The obtained data can be used for safe layer-by-layer laser treatment of obesity and cellulite.

Oblique incidence reflectometry: optical models and measurements using a side-viewing gradient index lens-based endoscopic imaging system

NASA Astrophysics Data System (ADS)

Wall, R. Andrew; Barton, Jennifer K.

2014-06-01

A side-viewing, 2.3-mm diameter oblique incidence reflectometry endoscope has been designed to obtain optical property measurements of turbid samples. Light from a single-mode fiber is relayed obliquely onto the tissue with a gradient index lens-based distal optics assembly and the resulting diffuse reflectance profile is imaged and collected with a 30,000 element, 0.72 mm clear aperture fiber bundle. Sampling the diffuse reflectance in two-dimensions allows for fitting of the reflected intensity profile to a well-known theoretical model, permitting the extraction of both absorption and reduced scattering coefficients of the tissue sample. Models and measurements of the endoscopic imaging system are presented in tissue phantoms and in vivo mouse colon, verifying the endoscope's capabilities to accurately measure effective attenuation coefficient and differentiate diseased from normal colon.

Thermographic study of laser on arteries.

PubMed

Mnitentag, J; Marques, E F; Ribeiro, M P; Braga, G A; Navarro, M R; Veratti, A B; Armelin, E; Macruz, R; Jatene, A D

1987-01-01

We analyzed the different effects of CO2, Nd-YAG, and argon lasers on aorta by using a Thermovision infrared system that registered the laser interaction with mongrel dog aorta. The images (thermograms) obtained were processed by a computer, which associated each area of the same temperature with a determined color. These thermograms were compared to histological analysis of the respective samples and the following results were obtained: (1) After the application of each laser there is very little propagation of heat in tissue. (2) The CO2 laser makes tissue reach 100 degrees C in less than 0.05 seconds. (3) The heat dissipation time was higher with the Nd-YAG laser due to higher scattering on tissue. Based on this research we conclude that the CO2 laser was best absorbed, the Nd-YAG laser penetrated human tissue with the best results, and the argon laser had the most significant backscattering.

ABO blood grouping from hard and soft tissues of teeth by modified absorption-elution technique.

PubMed

Ramnarayan, Bk; Manjunath, M; Joshi, Anagha Ananth

2013-01-01

Teeth have always been known as stable tissue that can be preserved both physically and chemically for long periods of time. Blood group substances have been known to be present in both the hard and soft tissues of the teeth. This study aimed at detection of ABO blood group substances from soft and hard tissues of teeth and also to evaluate the reliability of teeth stored for a relatively long period as a source of blood group substances by absorption-elution technique with some modifications. Blood group obtained from the teeth was compared with those obtained from the blood sample. Pulp showed a very large correlation in both fresh and long-standing teeth though it decreased slightly in the latter. Hard tissue showed a large correlation in both the groups indicating that hard tissue is quite reliable to detect blood group and that there is no much difference in the reliability in both the groups. However, combining pulp and hard tissue, correlation is moderate. Correlation of blood grouping with the age, sex, and jaw distribution was carried out. Blood group identification from hard and soft tissues of teeth aids in the identification of an individual.

Comparative proteomic analysis using samples obtained with laser microdissection and saturation dye labelling.

PubMed

Wilson, Kate E; Marouga, Rita; Prime, John E; Pashby, D Paul; Orange, Paul R; Crosier, Steven; Keith, Alexander B; Lathe, Richard; Mullins, John; Estibeiro, Peter; Bergling, Helene; Hawkins, Edward; Morris, Christopher M

2005-10-01

Comparative proteomic methods are rapidly being applied to many different biological systems including complex tissues. One pitfall of these methods is that in some cases, such as oncology and neuroscience, tissue complexity requires isolation of specific cell types and sample is limited. Laser microdissection (LMD) is commonly used for obtaining such samples for proteomic studies. We have combined LMD with sensitive thiol-reactive saturation dye labelling of protein samples and 2-D DIGE to identify protein changes in a test system, the isolated CA1 pyramidal neurone layer of a transgenic (Tg) rat carrying a human amyloid precursor protein transgene. Saturation dye labelling proved to be extremely sensitive with a spot map of over 5,000 proteins being readily produced from 5 mug total protein, with over 100 proteins being significantly altered at p < 0.0005. Of the proteins identified, all showed coherent changes associated with transgene expression. It was, however, difficult to identify significantly different proteins using PMF and MALDI-TOF on gels containing less than 500 mug total protein. The use of saturation dye labelling of limiting samples will therefore require the use of highly sensitive MS techniques to identify the significantly altered proteins isolated using methods such as LMD.

Extending neutron autoradiography technique for boron concentration measurements in hard tissues.

PubMed

Provenzano, Lucas; Olivera, María Silvina; Saint Martin, Gisela; Rodríguez, Luis Miguel; Fregenal, Daniel; Thorp, Silvia I; Pozzi, Emiliano C C; Curotto, Paula; Postuma, Ian; Altieri, Saverio; González, Sara J; Bortolussi, Silva; Portu, Agustina

2018-07-01

The neutron autoradiography technique using polycarbonate nuclear track detectors (NTD) has been extended to quantify the boron concentration in hard tissues, an application of special interest in Boron Neutron Capture Therapy (BNCT). Chemical and mechanical processing methods to prepare thin tissue sections as required by this technique have been explored. Four different decalcification methods governed by slow and fast kinetics were tested in boron-loaded bones. Due to the significant loss of the boron content, this technique was discarded. On the contrary, mechanical manipulation to obtain bone powder and tissue sections of tens of microns thick proved reproducible and suitable, ensuring a proper conservation of the boron content in the samples. A calibration curve that relates the 10 B concentration of a bone sample and the track density in a Lexan NTD is presented. Bone powder embedded in boric acid solution with known boron concentrations between 0 and 100 ppm was used as a standard material. The samples, contained in slim Lexan cases, were exposed to a neutron fluence of 10 12 cm -2 at the thermal column central facility of the RA-3 reactor (Argentina). The revealed tracks in the NTD were counted with an image processing software. The effect of track overlapping was studied and corresponding corrections were implemented in the presented calibration curve. Stochastic simulations of the track densities produced by the products of the 10 B thermal neutron capture reaction for different boron concentrations in bone were performed and compared with the experimental results. The remarkable agreement between the two curves suggested the suitability of the obtained experimental calibration curve. This neutron autoradiography technique was finally applied to determine the boron concentration in pulverized and compact bone samples coming from a sheep experimental model. The obtained results for both type of samples agreed with boron measurements carried out by ICP-OES within experimental uncertainties. The fact that the histological structure of bone sections remains preserved allows for future boron microdistribution analysis. Copyright © 2018 Elsevier Ltd. All rights reserved.

Untargeted UPLC-MS Profiling Pipeline to Expand Tissue Metabolome Coverage: Application to Cardiovascular Disease

PubMed Central

2015-01-01

Metabolic profiling studies aim to achieve broad metabolome coverage in specific biological samples. However, wide metabolome coverage has proven difficult to achieve, mostly because of the diverse physicochemical properties of small molecules, obligating analysts to seek multiplatform and multimethod approaches. Challenges are even greater when it comes to applications to tissue samples, where tissue lysis and metabolite extraction can induce significant systematic variation in composition. We have developed a pipeline for obtaining the aqueous and organic compounds from diseased arterial tissue using two consecutive extractions, followed by a different untargeted UPLC-MS analysis method for each extract. Methods were rationally chosen and optimized to address the different physicochemical properties of each extract: hydrophilic interaction liquid chromatography (HILIC) for the aqueous extract and reversed-phase chromatography for the organic. This pipeline can be generic for tissue analysis as demonstrated by applications to different tissue types. The experimental setup and fast turnaround time of the two methods contributed toward obtaining highly reproducible features with exceptional chromatographic performance (CV % < 0.5%), making this pipeline suitable for metabolic profiling applications. We structurally assigned 226 metabolites from a range of chemical classes (e.g., carnitines, α-amino acids, purines, pyrimidines, phospholipids, sphingolipids, free fatty acids, and glycerolipids) which were mapped to their corresponding pathways, biological functions and known disease mechanisms. The combination of the two untargeted UPLC-MS methods showed high metabolite complementarity. We demonstrate the application of this pipeline to cardiovascular disease, where we show that the analyzed diseased groups (n = 120) of arterial tissue could be distinguished based on their metabolic profiles. PMID:25664760

Small angle scattering polarization biopsy: a comparative analysis of various skin diseases

NASA Astrophysics Data System (ADS)

Zimnyakov, D. A.; Alonova, M. V.; Yermolenko, S. B.; Ivashko, P. V.; Reshetnikova, E. M.; Galkina, E. M.; Utz, S. R.

2013-12-01

An approach to differentiation of the morphological features of normal and pathological human epidermis on the base of statistical analysis of the local polarization states of laser light forward scattered by in-vitro tissue samples is discussed. The eccentricity and the azimuth angle of local polarization ellipses retrieved for various positions of the focused laser beam on the tissue surface, and the coefficient of collimated transmittance are considered as the diagnostic parameters for differentiation. The experimental data obtained with the psoriasis, discoid lupus erythematosus, alopecia, lichen planus, scabies, demodex, and normal skin samples are presented.

Immunohistochemical evaluation of neuroreceptors in healthy and pathological temporo-mandibular joint.

PubMed

Favia, Gianfranco; Corsalini, Massimo; Di Venere, Daniela; Pettini, Francesco; Favia, Giorgio; Capodiferro, Saverio; Maiorano, Eugenio

2013-01-01

A study was performed on the articular disk and periarticular tissues of the temporo-mandibular joint (TMJ) with immunohistochemical techniques to give evidence to the presence of neuroreceptors (NRec) in these sites. The study was carried out on tissue samples obtained from 10 subjects without TMJ disease and from 7 patients with severe TMJ arthritis and arthrosis. We use antibodies directed against following antigens: Gliofibrillary Acidic Protein (GFAP), Leu-7, Myelin Basic Protein (MBP), Neurofilaments 68 kD (NF), Neuron Specific Enolase (NSE), S-100 protein (S-100) and Synaptophysin (SYN). This study revealed that Ruffini's-like, Pacini's-like and Golgi's-like receptors can be demonstrated in TMJ periarticular tissues and that free nervous endings are present in the subsynovial tissues but not within the articular disk. We observed elongated cytoplamic processes of chondrocytes that demonstrated strong S-100 immunoreactivity but they were unreactive with all other antibodies. These cytoplamic processes were more abundant and thicker in the samples obtained from patients with disease TMJ. The results of this study confirm that different Nrec are detectable in TMJ periarticular tissues but they are absent within the articular disk. In the latter site, only condrocytic processes are evident, especially in diseased TMJ, and they might have been confused with nervous endings in previous morphological studies. Nevertheless the absence of immunoreactivity for NF, NSE and SYN proves that they are not of neural origin.

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 cells (65 cases-2 mutations; 3.1%). PMID:25683726

Development of a biaxial compression device for biological samples: preliminary experimental results for a closed cell foam.

PubMed

Little, J P; Tevelen, G; Adam, C J; Evans, J H; Pearcy, M J

2009-07-01

Biological tissues are subjected to complex loading states in vivo and in order to define constitutive equations that effectively simulate their mechanical behaviour under these loads, it is necessary to obtain data on the tissue's response to multiaxial loading. Single axis and shear testing of biological tissues is often carried out, but biaxial testing is less common. We sought to design and commission a biaxial compression testing device, capable of obtaining repeatable data for biological samples. The apparatus comprised a sealed stainless steel pressure vessel specifically designed such that a state of hydrostatic compression could be created on the test specimen while simultaneously unloading the sample along one axis with an equilibrating tensile pressure. Thus a state of equibiaxial compression was created perpendicular to the long axis of a rectangular sample. For the purpose of calibration and commissioning of the vessel, rectangular samples of closed cell ethylene vinyl acetate (EVA) foam were tested. Each sample was subjected to repeated loading, and nine separate biaxial experiments were carried out to a maximum pressure of 204 kPa (30 psi), with a relaxation time of two hours between them. Calibration testing demonstrated the force applied to the samples had a maximum error of 0.026 N (0.423% of maximum applied force). Under repeated loading, the foam sample demonstrated lower stiffness during the first load cycle. Following this cycle, an increased stiffness, repeatable response was observed with successive loading. While the experimental protocol was developed for EVA foam, preliminary results on this material suggest that this device may be capable of providing test data for biological tissue samples. The load response of the foam was characteristic of closed cell foams, with consolidation during the early loading cycles, then a repeatable load-displacement response upon repeated loading. The repeatability of the test results demonstrated the ability of the test device to provide reproducible test data and the low experimental error in the force demonstrated the reliability of the test data.

SU-F-T-220: Validation of Hounsfield Unit-To-Stopping Power Ratio Calibration Used for Dose Calculation in Proton Radiotherapy

DOE Office of Scientific and Technical Information (OSTI.GOV)

Polf, J; Chung, H; Langen, K

Purpose: To validate the stoichiometric calibration of the Hounsfield Unit (HU) to Stopping Power Ratio (SPR) calibration used to commission a commercial treatment planning system (TPS) for proton radiotherapy dose calculation. Methods and Materials: The water equivalent thickness (WET) of several individual pig tissues (lung, fat, muscle, liver, intestine, rib, femur), mixed tissue samples (muscle/rib, ice/femur, rib/air cavity/muscle), and an intact pig head were measured with a multi-layer ionization chamber (MLIC). A CT scan of each sample was obtained and imported into a commercial TPS. The WET calculated by the TPS for each tissue sample was compared to the measuredmore » WET value to determine the accuracy of the HU-to-SPR calibration curve used by the TPS to calculate dose. Results: The WET values calculated by the TPS showed good agreement (< 2.0%) with the measured values for bone and all soft tissues except fat (3.1% difference). For the mixed tissue samples and the intact pig head measurements, the difference in the TPS and measured WET values all agreed to within 3.5%. In addition, SPR values were calculated from the measured WET of each tissue, and compared to SPR values of reference tissues from ICRU 46 used to generate the HU-to-SPR calibration for the TPS. Conclusion: For clinical scenarios where the beam passes through multiple tissue types and its path is dominated by soft tissues, we believe using an uncertainty of 3.5% of the planned beam range is acceptable to account for uncertainties in the TPS WET determination.« less

Polarimetric imaging of biological tissues based on the indices of polarimetric purity.

PubMed

Van Eeckhout, Albert; Lizana, Angel; Garcia-Caurel, Enric; Gil, José J; Sansa, Adrià; Rodríguez, Carla; Estévez, Irene; González, Emilio; Escalera, Juan C; Moreno, Ignacio; Campos, Juan

2018-04-01

We highlight the interest of using the indices of polarimetric purity (IPPs) to the inspection of biological tissues. The IPPs were recently proposed in the literature and they result in a further synthetization of the depolarizing properties of samples. Compared with standard polarimetric images of biological samples, IPP-based images lead to larger image contrast of some biological structures and to a further physical interpretation of the depolarizing mechanisms inherent to the samples. In addition, unlike other methods, their calculation do not require advanced algebraic operations (as is the case of polar decompositions), and they result in 3 indicators of easy implementation. We also propose a pseudo-colored encoding of the IPP information that leads to an improved visualization of samples. This last technique opens the possibility of tailored adjustment of tissues contrast by using customized pseudo-colored images. The potential of the IPP approach is experimentally highlighted along the manuscript by studying 3 different ex-vivo samples. A significant image contrast enhancement is obtained by using the IPP-based methods, compared to standard polarimetric images. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

A quantum dot-based immunoassay for screening of tylosin and tilmicosin in edible animal tissues.

PubMed

Le, Tao; Zhu, Liqian; Yang, Xian

2015-01-01

A rapid, indirect competitive fluorescence-linked immunosorbent assay (ic-FLISA) based on quantum dots (QDs) as the fluorescent marker was developed for the detection of tylosin and tilmicosin in edible animal tissues. The end point fluorescent detection system was carried out using QDs conjugated with goat anti-mouse secondary antibody. The limits of detection (LODs) for the determination of tylosin and tilmicosin were 0.02 and 0.04 μg kg(-1), respectively. This detection method was used to analyse spiked samples and the recoveries ranged from 83.5% to 98.7% for tylosin and from 81.8% to 98.2% for tilmicosin. In real porcine tissue sample analysis, the results of ic-FLISA were similar to those obtained from an indirect competitive enzyme-linked immunosorbent assay (ic-ELISA) to an HPLC method indicating its potential for tylosin and tilmicosin screening in edible animal tissues.

Device for high spatial resolution chemical analysis of a sample and method of high spatial resolution chemical analysis

DOEpatents

Van Berkel, Gary J.

2015-10-06

A system and method for analyzing a chemical composition of a specimen are described. The system can include at least one pin; a sampling device configured to contact a liquid with a specimen on the at least one pin to form a testing solution; and a stepper mechanism configured to move the at least one pin and the sampling device relative to one another. The system can also include an analytical instrument for determining a chemical composition of the specimen from the testing solution. In particular, the systems and methods described herein enable chemical analysis of specimens, such as tissue, to be evaluated in a manner that the spatial-resolution is limited by the size of the pins used to obtain tissue samples, not the size of the sampling device used to solubilize the samples coupled to the pins.

Detection of Leptospira interrogans DNA and antigen in fixed equine eyes affected with end-stage equine recurrent uveitis.

PubMed

Pearce, Jacqueline W; Galle, Laurence E; Kleiboeker, Steve B; Turk, James R; Schommer, Susan K; Dubielizig, Richard R; Mitchell, William J; Moore, Cecil P; Giuliano, Elizabeth A

2007-11-01

Equine recurrent uveitis (ERU) is the most frequent cause of blindness in horses worldwide. Leptospira has been implicated as an etiologic agent in some cases of ERU and has been detected in fresh ocular tissues of affected horses. The objective of this study was to determine the presence of Leptospira antigen and DNA in fixed equine ocular tissues affected with end-stage ERU. Sections of eyes from 30 horses were obtained. Controls included 1) 10 normal equine eyes and 2) 10 equine eyes with a nonrecurrent form of uveitis. The experimental group consisted of 10 eyes diagnosed with ERU based on clinical signs and histologic lesions. Sections were subjected to immunohistochemical staining with an array of rabbit anti-Leptospira polyclonal antibodies. DNA extractions were performed by using a commercial kit designed for fixed tissue. Real-time PCR analysis was completed on extracted DNA. The target sequence for PCR was designed from alignments of available Leptospira 16S rDNA partial sequences obtained from GenBank. Two of 10 test samples were positive for Leptospira antigen by immunohistochemical assay. Zero of 20 controls were positive for Leptospira antigen. All test samples and controls were negative for Leptospira DNA by real-time PCR analysis. Leptospira was detected at a lower frequency than that previously reported for fresh ERU-affected aqueous humor and vitreous samples. Leptospira is not frequently detectable in fixed ocular tissues of horses affected with ERU when using traditional immunohistochemical and real-time PCR techniques.

In vitro chronic hepatic disease characterization with a multiparametric ultrasonic approach.

PubMed

Meziri, M; Pereira, W C A; Abdelwahab, A; Degott, C; Laugier, P

2005-03-01

Although, high resolution, real-time ultrasonic (US) imaging is routinely available, image interpretation is based on grey-level and texture and quantitative evaluation is limited. Other potentially useful diagnostic information from US echoes may include modifications in tissue acoustic parameters (speed, attenuation and backscattering) resulting from disease development. Changes in acoustical parameters can be detected using time-of-flight and spectral analysis techniques. The objective of this study is to explore the potential of three parameters together (attenuation coefficient, US speed and integrated backscatter coefficient-IBC) to discriminate healthy and fibrosis subgroups in liver tissue. Echoes from 21 fresh in vitro samples of human liver and from a plane reflector were obtained using a 20-MHz central frequency transducer (6-30 MHz bandpass). The scan plane was parallel to the reflector placed beneath the liver. A 30 x 20 matrix of A-scans was obtained, with a 200-microm step. The samples were classified according to the Metavir scale in five different degrees of fibrosis. US speed, attenuation and IBC were estimated from standard methods described in the literature. Statistical tests were applied to the results of each parameter individually and indicated that it was not possible to identify all the fibrosis groups. Then a discriminant analysis was performed for the three parameters together resulting in a reasonable separation of fibrotic groups. Although the number of tissue samples is limited, this study opens the possibility of enhancing the discriminant capability of ultrasonic parameters of liver tissue disease when they are combined together.

Climatic signals registered as Carbon isotopic values in Metasequoia leaf tissues: A statistical analysis

NASA Astrophysics Data System (ADS)

Yang, H.; Blais, B.; Perez, G.; Pagani, M.

2006-12-01

To examine climatic signals registered as carbon isotopic values in leaf tissues of C3 plants, we collected mature leaf tissues from sun and shade leaves of Metasequoia trees germinated from the 1947 batch of seeds from China and planted along a latitudinal gradient of the United States. Samples from 40 individual trees, along with fossilized material from the early Tertiary of the Canadian Arctic, were analyzed for C and concentration and isotopic values using EA-IRMS after the removal of free lipids. The generated datasets were then merged with climate data compiled from each tree site recorded as average values over the past thirty years (1971-2002, NOAA database). When the isotope data were cross plotted against each geographic and climatic indicator, Latitude, Mean Annual Temperature (MAT), Average Summer Mean Temperature (ASMT)(June-August), Mean Annual Precipitation (MAP), and Average Summer Mean Precipitation (ASMP) respectively correlation patterns were revealed. The best correlating trend was obtained between temperature parameters and C isotopic values, and this correlation is stronger in the northern leaf samples than the southern samples. We discovered a strong positive correlation between latitude and the offset of C isotopic values between shade and sun leaves. This investigation represents a comprehensive examination on climatic signals registered as C isotopic values on a single species that is marked by single genetic source. The results bear implications on paleoclimatic interpretations of C isotopic signals obtained from fossil plant tissues.

Ultra-high performance liquid chromatographic determination of levofloxacin in human plasma and prostate tissue with use of experimental design optimization procedures.

PubMed

Szerkus, O; Jacyna, J; Wiczling, P; Gibas, A; Sieczkowski, M; Siluk, D; Matuszewski, M; Kaliszan, R; Markuszewski, M J

2016-09-01

Fluoroquinolones are considered as gold standard for the prevention of bacterial infections after transrectal ultrasound guided prostate biopsy. However, recent studies reported that fluoroquinolone- resistant bacterial strains are responsible for gradually increasing number of infections after transrectal prostate biopsy. In daily clinical practice, antibacterial efficacy is evaluated only in vitro, by measuring the reaction of bacteria with an antimicrobial agent in culture media (i.e. calculation of minimal inhibitory concentration). Such approach, however, has no relation to the treated tissue characteristics and might be highly misleading. Thus, the objective of this study was to develop, with the use of Design of Experiments approach, a reliable, specific and sensitive ultra-high performance liquid chromatography- diode array detection method for the quantitative analysis of levofloxacin in plasma and prostate tissue samples obtained from patients undergoing prostate biopsy. Moreover, correlation study between concentrations observed in plasma samples vs prostatic tissue samples was performed, resulting in better understanding, evaluation and optimization of the fluoroquinolone-based antimicrobial prophylaxis during transrectal ultrasound guided prostate biopsy. Box-Behnken design was employed to optimize chromatographic conditions of the isocratic elution program in order to obtain desirable retention time, peak symmetry and resolution of levofloxacine and ciprofloxacine (internal standard) peaks. Fractional Factorial design 2(4-1) with four center points was used for screening of significant factors affecting levofloxacin extraction from the prostatic tissue. Due to the limited number of tissue samples the prostatic sample preparation procedure was further optimized using Central Composite design. Design of Experiments approach was also utilized for evaluation of parameter robustness. The method was found linear over the range of 0.030-10μg/mL for human plasma and 0.300-30μg/g for human prostate tissue samples. The intra-day and inter-day variability for levofloxacine from both plasma and prostate samples were less than 10%, with accuracies between 93 and 108% of the nominal values. The limit of detection and the limit of quantification for human plasma were 0.01μg/mL and 0.03μg/mL, respectively. For the prostate tissue, the limit of detection and the limit of quantification were 0.1μg/g and 0.3μg/g, respectively. The average recoveries of levofloxacin were in the range from 99 to 106%. Also, the method fulfills requirements of robustness what was determined and proved by Design of Experiments. The developed method was successfully applied to examine prostate tissue and plasma samples from 140 hospitalized patients enrolled into the clinical study, 12h after oral administration of LVF at a dose of 500mg. The mean (±SD) LVF concentration in prostate was 6.22±3.52μg/g and in plasma 2.54±1.14μg/mL. Due to simplicity of the method and relative small amount of sample needed for the assay, the method can be applied in clinical practice for monitoring of LVF concentrations in plasma and prostate gland. Copyright © 2016 Elsevier B.V. All rights reserved.

Correlation of virus load in plasma and lymph node tissue in human immunodeficiency virus infection. INCAS Study Group. Italy, Netherlands, Canada, Australia, and (United) States.

PubMed

Harris, M; Patenaude, P; Cooperberg, P; Filipenko, D; Thorne, A; Raboud, J; Rae, S; Dailey, P; Chernoff, D; Todd, J; Conway, B; Montaner, J S

1997-11-01

The impact of long-term changes in plasma viremia, produced by effective combination antiretroviral therapy, on human immunodeficiency virus (HIV) burden within tissue reservoirs is unknown. Fifteen patients who had received at least 1 year of therapy with two or three drug combinations of zidovudine, didanosine, and nevirapine had suitable samples of lymph node tissue obtained by ultrasound-guided core needle biopsy. HIV RNA was extracted from homogenized tissue samples and quantitated using a modified branched DNA assay. Results were correlated with antiretroviral treatment effect on the basis of plasma virus load measurements over the preceding 12-18 months. A statistically significant negative correlation was observed between magnitude of treatment effect on plasma viremia and lymph node virus load. These data suggest that combinations of antiretroviral drugs that produce sustained suppression of plasma HIV RNA may also be able to reduce the virus burden in lymphoid tissues.

Nonnegative constraint analysis of key fluorophores within human breast cancer using native fluorescence spectroscopy excited by selective wavelength of 300 nm

NASA Astrophysics Data System (ADS)

Pu, Yang; Sordillo, Laura A.; Alfano, Robert R.

2015-03-01

Native fluorescence spectroscopy offers an important role in cancer discrimination. It is widely acknowledged that the emission spectrum of tissue is a superposition of spectra of various salient fluorophores. In this study, the native fluorescence spectra of human cancerous and normal breast tissues excited by selected wavelength of 300 nm are used to investigate the key building block fluorophores: tryptophan and reduced nicotinamide adenine dinucleotide (NADH). The basis spectra of these key fluorophores' contribution to the tissue emission spectra are obtained by nonnegative constraint analysis. The emission spectra of human cancerous and normal tissue samples are projected onto the fluorophore spectral subspace. Since previous studies indicate that tryptophan and NADH are key fluorophores related with tumor evolution, it is essential to obtain their information from tissue fluorescence but discard the redundancy. To evaluate the efficacy of for cancer detection, linear discriminant analysis (LDA) classifier is used to evaluate the sensitivity, and specificity. This research demonstrates that the native fluorescence spectroscopy measurements are effective to detect changes of fluorophores' compositions in tissues due to the development of cancer.

Reducing the orientation influence of Mueller matrix measurements for anisotropic scattering media

NASA Astrophysics Data System (ADS)

Sun, Minghao; He, Honghui; Zeng, Nan; Du, E.; He, Yonghong; Ma, Hui

2014-09-01

Mueller matrix polarimetry techniques contain rich micro-structural information of samples, such as the sizes and refractive indices of scatterers. Recently, Mueller matrix imaging methods have shown great potentials as powerful tools for biomedical diagnosis. However, the orientations of anisotropic fibrous structures in tissues have prominent influence on Mueller matrix measurements, resulting in difficulties for extracting micro-structural information effectively. In this paper, we apply the backscattering Mueller matrix imaging technique to biological samples with different microstructures, such as chicken heart muscle, bovine skeletal muscle, porcine liver and fat tissues. Experimental results show that the directions of the muscle fibers have prominent influence on the Mueller matrix elements. In order to reduce the orientation influence, we adopt the rotation-independent MMT and RLPI parameters, which were proposed in our previous studies, to the tissue samples. Preliminary results in this paper show that the orientation-independent parameters and their statistic features are helpful for analyzing the tissues to obtain their micro-structural properties. Since the micro-structure variations are often related to the pathological changes, the method can be applied to microscope imaging techniques and used to detect abnormal tissues such as cancer and other lesions for diagnosis purposes.

Chemical Clearing and Dehydration of GFP Expressing Mouse Brains

PubMed Central

Saghafi, Saiedeh; Weiler, Reto; Dodt, Hans-Ulrich

2012-01-01

Generally, chemical tissue clearing is performed by a solution consisting of two parts benzyl benzoate and one part benzyl alcohol. However, prolonged exposure to this mixture markedly reduces the fluorescence of GFP expressing specimens, so that one has to compromise between clearing quality and fluorescence preservation. This can be a severe drawback when working with specimens exhibiting low GFP expression rates. Thus, we screened for a substitute and found that dibenzyl ether (phenylmethoxymethylbenzene, CAS 103-50-4) can be applied as a more GFP-friendly clearing medium. Clearing with dibenzyl ether provides improved tissue transparency and strikingly improved fluorescence intensity in GFP expressing mouse brains and other samples as mouse spinal cords, or embryos. Chemical clearing, staining, and embedding of biological samples mostly requires careful foregoing tissue dehydration. The commonly applied tissue dehydration medium is ethanol, which also can markedly impair GFP fluorescence. Screening for a substitute also for ethanol we found that tetrahydrofuran (CAS 109-99-9) is a more GFP-friendly dehydration medium than ethanol, providing better tissue transparency obtained by successive clearing. Combined, tetrahydrofuran and dibenzyl ether allow dehydration and chemical clearing of even delicate samples for UM, confocal microscopy, and other microscopy techniques. PMID:22479475

Chemical clearing and dehydration of GFP expressing mouse brains.

PubMed

Becker, Klaus; Jährling, Nina; Saghafi, Saiedeh; Weiler, Reto; Dodt, Hans-Ulrich

2012-01-01

Generally, chemical tissue clearing is performed by a solution consisting of two parts benzyl benzoate and one part benzyl alcohol. However, prolonged exposure to this mixture markedly reduces the fluorescence of GFP expressing specimens, so that one has to compromise between clearing quality and fluorescence preservation. This can be a severe drawback when working with specimens exhibiting low GFP expression rates. Thus, we screened for a substitute and found that dibenzyl ether (phenylmethoxymethylbenzene, CAS 103-50-4) can be applied as a more GFP-friendly clearing medium. Clearing with dibenzyl ether provides improved tissue transparency and strikingly improved fluorescence intensity in GFP expressing mouse brains and other samples as mouse spinal cords, or embryos. Chemical clearing, staining, and embedding of biological samples mostly requires careful foregoing tissue dehydration. The commonly applied tissue dehydration medium is ethanol, which also can markedly impair GFP fluorescence. Screening for a substitute also for ethanol we found that tetrahydrofuran (CAS 109-99-9) is a more GFP-friendly dehydration medium than ethanol, providing better tissue transparency obtained by successive clearing. Combined, tetrahydrofuran and dibenzyl ether allow dehydration and chemical clearing of even delicate samples for UM, confocal microscopy, and other microscopy techniques.

[Criteria of the molecular pathology testing of lung cancer].

PubMed

Tímár, József

2014-06-01

From the aspect of the contemporary pathologic diagnostics of lung cancer the tissue obtained is a key issue since small biopsies and cytology still play a major role. In the non-small cell lung cancer era cytology is considered equal to biopsy however, in recent years it is unable to provide quality diagnosis and must be replaced by biopsy. Various molecular techniques can handle various different tissue samples which must be considered during molecular pathology diagnosis. Moreover, tumor cell-normal cell ratio in the obtained tissue, as well as the absolute tumor cell number have great significance, which information must be provided in the primary lung cancer diagnosis. Last but not least, for continuous sustainable molecular diagnostics of lung cancer rational algorithms, affordable technology and appropriate reimbursement are equally necessary.

Minimal requirements for the molecular testing of lung cancer.

PubMed

Popper, Helmut H; Tímár, József; Ryska, Ales; Olszewski, Wlodzimierz

2014-10-01

From the aspect of the contemporary pathologic diagnostics of lung cancer, it is a key issue of the tissue obtained since small biopsies and cytology still play a major role. In the non-small cell lung cancer era, cytology considered equal to biopsy. However, in recent years it is unable to provide quality diagnosis and must be replaced by biopsy. Various molecular techniques can handle various different tissue samples which must be considered during molecular pathology diagnosis. Besides, tumor cell-normal cell ratio in the obtained tissue as well as the absolute tumor cell number have great significance whose information must be provided in the primary lung cancer diagnosis. Last but not least, for continuous sustainable molecular diagnostics of lung cancer rational algorythms, affordable technology and appropriate reimbursement are equally necessary.

Bdellovibrios in Callinectus sapidus, the Blue Crab

PubMed Central

Kelley, Jacqueline I.; Williams, Henry N.

1992-01-01

Bdellovibrios were recovered from the gill tissue of all of 31 crabs sampled and from all samples of epibiota obtained from the ventral shell surface of 15 crabs. The results suggest that the blue crab is a reservoir for bdellovibrios. The association with crabs may be an important factor in the ecology of the bdellovibrios. PMID:16348706

Fetal Tissue Procurement for Karyotype Analysis: Clinician or Pathologist - Which is Better?

PubMed

Conant, Joanna L; Tang, Mary E; Waters, Brenda L

2016-01-01

Chromosomal abnormalities are detected in up to 13% of stillbirths and over 20% of those with developmental anomalies. These estimates may be low since up to 50% of samples fail to achieve a result due to microbial overgrowth or nonviability. Tissue for cytogenetics can be procured at bedside by the clinician or by the pathologist in the laboratory. With clinical collection, tissue is placed into culture media immediately, increasing chances of growth. However, collection competes for attention with other activities, which may result in microbial overgrowth or selection of maternal rather than fetal tissue. Laboratory procurement occurs in a controlled environment using sterile technique, but delay in collection may decrease viability. Our goal was to determine which collection method yields better results. We reviewed cases from 2007-2013 that had two samples submitted for cytogenetics, one from the clinician and one from the pathologist. Specimen source, delivery, collection, and culture setup times, harvest date, cell growth, microbial overgrowth, maternal contamination and final result were obtained from medical records and cytogenetic culture sheets. There was no difference in growth rate, maternal cell contamination, or reporting time between clinician- and pathologist-procured samples despite delay in collection time for laboratory samples. Clinical samples had more microbial overgrowth. Compared to samples collected at bedside, samples collected in the laboratory had a lower rate of microbial contamination with similar growth and maternal cell contamination rates, despite prolonged time to collection. Collecting samples both at bedside and in the laboratory is unnecessary.

Liquid microjunction surface sampling coupled with high-pressure liquid chromatography-electrospray ionization-mass spectrometry for analysis of drugs and metabolites in whole-body thin tissue sections.

PubMed

Kertesz, Vilmos; Van Berkel, Gary J

2010-07-15

In this work, a commercially available autosampler was adapted to perform direct liquid microjunction (LMJ) surface sampling followed by a high-pressure liquid chromatography (HPLC) separation of the extract components and detection with electrospray ionization mass spectrometry (ESI-MS). To illustrate the utility of coupling a separation with this direct liquid extraction based surface sampling approach, four different organs (brain, lung, kidney, and liver) from whole-body thin tissue sections of propranolol dosed and control mice were examined. The parent drug was observed in the chromatograms of the surface sampling extracts from all the organs of the dosed mouse examined. In addition, two isomeric phase II metabolites of propranolol (an aliphatic and an aromatic hydroxypropranolol glucuronide) were observed in the chromatograms of the extracts from lung, kidney, and liver. Confirming the presence of one or the other or both of these glucuronides in the extract from the various organs was not possible without the separation. These drug and metabolite data obtained using the LMJ surface sampling/HPLC-MS method and the results achieved by analyzing similar samples by conventional extraction of the tissues and subsequent HPLC-MS analysis were consistent. The ability to directly and efficiently sample from thin tissue sections via a liquid extraction and then perform a subsequent liquid phase separation increases the utility of this liquid extraction surface sampling approach.

Comparison of blood chemistry values for samples collected from juvenile chinook salmon by three methods

USGS Publications Warehouse

Congleton, J.L.; LaVoie, W.J.

2001-01-01

Thirteen blood chemistry indices were compared for samples collected by three commonly used methods: caudal transection, heart puncture, and caudal vessel puncture. Apparent biases in blood chemistry values for samples obtained by caudal transection were consistent with dilution with tissue fluids: alanine aminotransferase (ALT), aspartate aminotransferase (AST), lactate dehydrogenase (LDH), creatine kinase (CK), triglyceride, and K+ were increased and Na+ and Cl- were decreased relative to values for samples obtained by caudal vessel puncture. Some enzyme activities (ALT, AST, LDH) and K+ concentrations were also greater in samples taken by heart puncture than in samples taken by caudal vessel puncture. Of the methods tested, caudal vessel puncture had the least effect on blood chemistry values and should be preferred for blood chemistry studies on juvenile salmonids.

Towards intra-operative diagnosis of tumours during breast conserving surgery by selective-sampling Raman micro-spectroscopy

NASA Astrophysics Data System (ADS)

Kong, Kenny; Zaabar, Fazliyana; Rakha, Emad; Ellis, Ian; Koloydenko, Alexey; Notingher, Ioan

2014-10-01

Breast-conserving surgery (BCS) is increasingly employed for the treatment of early stage breast cancer. One of the key challenges in BCS is to ensure complete removal of the tumour while conserving as much healthy tissue as possible. In this study we have investigated the potential of Raman micro-spectroscopy (RMS) for automated intra-operative evaluation of tumour excision. First, a multivariate classification model based on Raman spectra of normal and malignant breast tissue samples was built and achieved diagnosis of mammary ductal carcinoma (DC) with 95.6% sensitivity and 96.2% specificity (5-fold cross-validation). The tumour regions were discriminated from the healthy tissue structures based on increased concentration of nucleic acids and reduced concentration of collagen and fat. The multivariate classification model was then applied to sections from fresh tissue of new patients to produce diagnosis images for DC. The diagnosis images obtained by raster scanning RMS were in agreement with the conventional histopathology diagnosis but were limited to long data acquisition times (typically 10 000 spectra mm-2, which is equivalent to ~5 h mm-2). Selective-sampling based on integrated auto-fluorescence imaging and Raman spectroscopy was used to reduce the number of Raman spectra to ~20 spectra mm-2, which is equivalent to an acquisition time of ~15 min for 5 × 5 mm2 tissue samples. This study suggests that selective-sampling Raman microscopy has the potential to provide a rapid and objective intra-operative method to detect mammary carcinoma in tissue and assess resection margins.

Sinonasal microbiome sampling: a comparison of techniques.

PubMed

Bassiouni, Ahmed; Cleland, Edward John; Psaltis, Alkis James; Vreugde, Sarah; Wormald, Peter-John

2015-01-01

The role of the sino-nasal microbiome in CRS remains unclear. We hypothesized that the bacteria within mucosal-associated biofilms may be different from the more superficial-lying, free-floating bacteria in the sinuses and that this may impact on the microbiome results obtained. This study investigates whether there is a significant difference in the microbiota of a sinonasal mucosal tissue sample versus a swab sample. Cross-sectional study with paired design. Mucosal biopsy and swab samples were obtained intra-operatively from the ethmoid sinuses of 6 patients with CRS. Extracted DNA was sequenced on a Roche-454 sequencer using 16S-rRNA gene targeted primers. Data were analyzed using QIIME 1.8 software package. At a maximum subsampling depth of 1,100 reads, the mean observed species richness was 33.3 species (30.6 for swab, versus 36 for mucosa; p > 0.05). There was no significant difference in phylogenetic and non-phylogenetic alpha diversity metrics (Faith's PD_Whole_Tree and Shannon's index) between the two sampling methods (p > 0.05). The type of sample also had no significant effect on phylogenetic and non-phylogenetic beta diversity metrics (Unifrac and Bray-Curtis; p > 0.05). We observed no significant difference between the microbiota of mucosal tissue and swab samples. This suggests that less invasive swab samples are representative of the sinonasal mucosa microbiome and can be used for future sinonasal microbiome studies.

Observation of human tissue with phase-contrast x-ray computed tomography

NASA Astrophysics Data System (ADS)

Momose, Atsushi; Takeda, Tohoru; Itai, Yuji; Tu, Jinhong; Hirano, Keiichi

1999-05-01

Human tissues obtained from cancerous kidneys fixed in formalin were observed with phase-contrast X-ray computed tomography (CT) using 17.7-keV synchrotron X-rays. By measuring the distributions of the X-ray phase shift caused by samples using an X-ray interferometer, sectional images that map the distribution of the refractive index were reconstructed. Because of the high sensitivity of phase- contrast X-ray CT, a cancerous lesion was differentiated from normal tissue and a variety of other structures were revealed without the need for staining.

Numerical examination of like-honeycomb structures

NASA Astrophysics Data System (ADS)

John, Małgorzata; John, Antoni; Skarka, Wojciech

2018-01-01

In the paper based on the analogy with the biological tissue of bones, it was decided to examine more homogenous structure and also a heterogeneous structure too. Here, a new approach is proposed based on results from literature obtained using topology optimization 2D and 3D structures like beam, girder and cantilever. Proposed model of structure is similar to spatial trusses with honeycomb-shape porous. Parameters varied not only uniformly throughout the volume of the sample, but also be modified depending on various factors. They underwent a change in cell dimensions, among other things, the thickness of the wall. The obtained results were compared with those obtained previously for homogeneous samples.

Tissue Banking, Bioinformatics, and Electronic Medical Records: The Front-End Requirements for Personalized Medicine

PubMed Central

Suh, K. Stephen; Sarojini, Sreeja; Youssif, Maher; Nalley, Kip; Milinovikj, Natasha; Elloumi, Fathi; Russell, Steven; Pecora, Andrew; Schecter, Elyssa; Goy, Andre

2013-01-01

Personalized medicine promises patient-tailored treatments that enhance patient care and decrease overall treatment costs by focusing on genetics and “-omics” data obtained from patient biospecimens and records to guide therapy choices that generate good clinical outcomes. The approach relies on diagnostic and prognostic use of novel biomarkers discovered through combinations of tissue banking, bioinformatics, and electronic medical records (EMRs). The analytical power of bioinformatic platforms combined with patient clinical data from EMRs can reveal potential biomarkers and clinical phenotypes that allow researchers to develop experimental strategies using selected patient biospecimens stored in tissue banks. For cancer, high-quality biospecimens collected at diagnosis, first relapse, and various treatment stages provide crucial resources for study designs. To enlarge biospecimen collections, patient education regarding the value of specimen donation is vital. One approach for increasing consent is to offer publically available illustrations and game-like engagements demonstrating how wider sample availability facilitates development of novel therapies. The critical value of tissue bank samples, bioinformatics, and EMR in the early stages of the biomarker discovery process for personalized medicine is often overlooked. The data obtained also require cross-disciplinary collaborations to translate experimental results into clinical practice and diagnostic and prognostic use in personalized medicine. PMID:23818899

Evaluation of different pulverisation methods for RNA extraction in squash fruit: lyophilisation, cryogenic mill and mortar grinding.

PubMed

Román, Belén; González-Verdejo, Clara I; Peña, Francisco; Nadal, Salvador; Gómez, Pedro

2012-01-01

Quality and integrity of RNA are critical for transcription studies in plant molecular biology. In squash fruit and other high water content crops, the grinding of tissue with mortar and pestle in liquid nitrogen fails to produce a homogeneous and fine powered sample desirable to ensure a good penetration of the extraction reagent. To develop an improved pulverisation method to facilitate the homogenisation process of squash fruit tissue prior to RNA extraction without reducing quality and yield of the extracted RNA. Three methods of pulverisation, each followed by the same extraction protocol, were compared. The first approach consisted of the lyophilisation of the sample in order to remove the excess of water before grinding, the second one used a cryogenic mill and the control one a mortar grinding of frozen tissue. The quality of the isolated RNA was tested by carrying out a quantitative real time downstream amplification. In the three situations considered, mean values for A(260) /A(280) indicated minimal interference by proteins and RNA quality indicator (RQI) values were considered appropriate for quantitative real-time polymerase chain reaction (qRT-PCR) amplification. Successful qRT-PCR amplifications were obtained with cDNA isolated with the three protocols. Both apparatus can improve and facilitate the grinding step in the RNA extraction process in zucchini, resulting in isolated RNA of high quality and integrity as revealed by qRT-PCR downstream application. This is apparently the first time that a cryogenic mill has been used to prepare fruit samples for RNA extraction, thereby improving the sampling strategy because the fine powder obtained represents a homogeneous mix of the organ tissue. Copyright © 2012 John Wiley & Sons, Ltd.

The serological screening of deceased tissue donors within the English Blood Service for infectious agents--a review of current outcomes and a more effective strategy for the future.

PubMed

Kitchen, A D; Gillan, H L

2010-04-01

The overall effectiveness of the NHSBT screening programme for infectious agents in deceased tissue donors is examined and evaluated in terms of current outcomes and how to improve upon these outcomes. The screening results and any subsequent confirmatory results from a total of 1659 samples from NHSBT deceased donors referred to NTMRL for screening for infectious agents were included in the analysis. Overall 1566/1659 (94.4%) of the samples were screen negative. A total of 93 were repeat reactive on screening for one or more of the mandatory markers screened for, of which only 12 (13%) were subsequently confirmed to be positive on confirmatory testing. The majority of the repeat reactive samples were demonstrating non-specific reactivity with the screening assays in use. Overall, the NHSBT screening programme for infectious agents in deceased tissue donors is very effective with a relatively low overall loss of donors because of non-specific reactivity. However, unnecessary loss of tissue products is not acceptable, and although this programme compares favourably with the outcomes of other such programmes, the confirmatory results obtained demonstrate both the need and the potential for improving the outcomes. This is particularly important as one donor may donate more than one product, and can be achieved very easily with a change to the screening algorithm followed, using the confirmatory data obtained to support and validate this change. CONTENTS SUMMARY: Critical analysis of the NHSBT screening programme for infectious agents in deceased tissue donors and a strategy involving the design and use of a different screening algorithm to improve these outcomes.

The effect of electronically steering a phased array ultrasound transducer on near-field tissue heating.

PubMed

Payne, Allison; Vyas, Urvi; Todd, Nick; de Bever, Joshua; Christensen, Douglas A; Parker, Dennis L

2011-09-01

This study presents the results obtained from both simulation and experimental techniques that show the effect of mechanically or electronically steering a phased array transducer on proximal tissue heating. The thermal response of a nine-position raster and a 16-mm diameter circle scanning trajectory executed through both electronic and mechanical scanning was evaluated in computer simulations and experimentally in a homogeneous tissue-mimicking phantom. Simulations were performed using power deposition maps obtained from the hybrid angular spectrum (HAS) method and applying a finite-difference approximation of the Pennes' bioheat transfer equation for the experimentally used transducer and also for a fully sampled transducer to demonstrate the effect of acoustic window, ultrasound beam overlap and grating lobe clutter on near-field heating. Both simulation and experimental results show that electronically steering the ultrasound beam for the two trajectories using the 256-element phased array significantly increases the thermal dose deposited in the near-field tissues when compared with the same treatment executed through mechanical steering only. In addition, the individual contributions of both beam overlap and grating lobe clutter to the near-field thermal effects were determined through comparing the simulated ultrasound beam patterns and resulting temperature fields from mechanically and electronically steered trajectories using the 256-randomized element phased array transducer to an electronically steered trajectory using a fully sampled transducer with 40 401 phase-adjusted sample points. Three distinctly different three distinctly different transducers were simulated to analyze the tradeoffs of selected transducer design parameters on near-field heating. Careful consideration of design tradeoffs and accurate patient treatment planning combined with thorough monitoring of the near-field tissue temperature will help to ensure patient safety during an MRgHIFU treatment.

Pathology Reports

MedlinePlus

... pathology report will include the results of these tests. For example, the pathology report may include information obtained from ... markers or indicators of a specific cancer. For example, the Philadelphia chromosome ... ( 3 ). Some tests that might be performed on a tissue sample ...

Controlling for anthropogenically induced atmospheric variation in stable carbon isotope studies

USGS Publications Warehouse

Long, E.S.; Sweitzer, R.A.; Diefenbach, D.R.; Ben-David, M.

2005-01-01

Increased use of stable isotope analysis to examine food-web dynamics, migration, transfer of nutrients, and behavior will likely result in expansion of stable isotope studies investigating human-induced global changes. Recent elevation of atmospheric CO2 concentration, related primarily to fossil fuel combustion, has reduced atmospheric CO2 ??13C (13C/12C), and this change in isotopic baseline has, in turn, reduced plant and animal tissue ??13C of terrestrial and aquatic organisms. Such depletion in CO2 ??13C and its effects on tissue ??13C may introduce bias into ??13C investigations, and if this variation is not controlled, may confound interpretation of results obtained from tissue samples collected over a temporal span. To control for this source of variation, we used a high-precision record of atmospheric CO2 ??13C from ice cores and direct atmospheric measurements to model modern change in CO2 ??13C. From this model, we estimated a correction factor that controls for atmospheric change; this correction reduces bias associated with changes in atmospheric isotopic baseline and facilitates comparison of tissue ??13C collected over multiple years. To exemplify the importance of accounting for atmospheric CO2 ??13C depletion, we applied the correction to a dataset of collagen ??13C obtained from mountain lion (Puma concolor) bone samples collected in California between 1893 and 1995. Before correction, in three of four ecoregions collagen ??13C decreased significantly concurrent with depletion of atmospheric CO2 ??13C (n ??? 32, P ??? 0.01). Application of the correction to collagen ??13C data removed trends from regions demonstrating significant declines, and measurement error associated with the correction did not add substantial variation to adjusted estimates. Controlling for long-term atmospheric variation and correcting tissue samples for changes in isotopic baseline facilitate analysis of samples that span a large temporal range. ?? Springer-Verlag 2005.

Use of focused ultrasonication in activity-based profiling of deubiquitinating enzymes in tissue.

PubMed

Nanduri, Bindu; Shack, Leslie A; Rai, Aswathy N; Epperson, William B; Baumgartner, Wes; Schmidt, Ty B; Edelmann, Mariola J

2016-12-15

To develop a reproducible tissue lysis method that retains enzyme function for activity-based protein profiling, we compared four different methods to obtain protein extracts from bovine lung tissue: focused ultrasonication, standard sonication, mortar & pestle method, and homogenization combined with standard sonication. Focused ultrasonication and mortar & pestle methods were sufficiently effective for activity-based profiling of deubiquitinases in tissue, and focused ultrasonication also had the fastest processing time. We used focused-ultrasonicator for subsequent activity-based proteomic analysis of deubiquitinases to test the compatibility of this method in sample preparation for activity-based chemical proteomics. Copyright © 2016 Elsevier Inc. All rights reserved.

Nonlocal correlations of polarization-entangled photons through brain tissue (Conference Presentation)

NASA Astrophysics Data System (ADS)

Galvez, Enrique J.; Shi, Lingyan; Alfano, Robert R.

2017-02-01

We investigated the preservation of non-local correlations between polarization-entangled photons when one of them traveled through brain tissue slices of different thicknesses. Using down-converted photons at a wavelength of 802 nm minimized the absorption by the tissue. After the light passed through the tissue samples, we performed quantum state tomography to obtain quantitative measures of the entanglement. We found that entanglement is preserved to a surprising degree, and when it degrades, it does so following a particular path in a tangle versus linear-entropy graph. Such a trajectory reveals direct transfer of probability from entangled to mixed state.

Understanding the effect of pulsed electric fields on thermostability of connective tissue isolated from beef pectoralis muscle using a model system.

PubMed

Alahakoon, A U; Oey, I; Silcock, P; Bremer, P

2017-10-01

Brisket is a low value/tough meat cut that contains a large amount of connective tissue. Conversion of collagen into gelatin during heating reduces the toughness of the connective tissue however this conversion is slow at low cooking temperatures (around 60°C). The objective of this project was to determine the ability of pulsed electric field (PEF) processing to reduce the thermal stability of connective tissue. To achieve this, a novel model system was designed in which connective tissue obtained from beef deep pectotalis muscle (brisket) was exposed to PEF at combinations of electric field strength (1.0 and 1.5kV/cm) and specific energy (50 and 100kJ/kg) within an agar matrix at electrical conductivities representing the electrical conductivity found in brisket. Differential scanning calorimetry showed that PEF treatment significantly (p<0.05) decreased the denaturation temperature of connective tissue compared to untreated samples. Increasing electric field strength and the specific energy increased the Ringer soluble collagen fraction. PEF treated samples showed higher solubilization compared to the untreated samples at both 60°C and 70°C in heat solubility test. SEM examination of PEF treated (at 1.5kV/cm and 100kJ/kg) and untreated samples revealed that PEF appeared to increase the porosity of the connective tissue structure. These finding suggest that PEF processing is a technology that could be used to improve the tenderness and decrease the cooking time of collagen rich, meat cuts. Copyright © 2017 Elsevier Ltd. All rights reserved.

Partition of environmental chemicals between maternal and fetal blood and tissues.

PubMed

Needham, Larry L; Grandjean, Philippe; Heinzow, Birger; Jørgensen, Poul J; Nielsen, Flemming; Patterson, Donald G; Sjödin, Andreas; Turner, Wayman E; Weihe, Pal

2011-02-01

Passage of environmental chemicals across the placenta has important toxicological consequences, as well as for choosing samples for analysis and for interpreting the results. To obtain systematic data, we collected in 2000 maternal and cord blood, cord tissue, placenta, and milk in connection with births in the Faroe Islands, where exposures to marine contaminants is increased. In 15 sample sets, we measured a total of 87 environmental chemicals, almost all of which were detected both in maternal and fetal tissues. The maternal serum lipid-based concentrations of organohalogen compounds averaged 1.7 times those of cord serum, 2.8 times those of cord tissue and placenta, and 0.7 those of milk. For organohalogen compounds detectable in all matrices, a high degree of correlation between concentrations in maternal serum and the other tissues investigated was generally observed (r(2) > 0.5). Greater degree of chlorination resulted in lower transfer from maternal serum into milk. Concentrations of pentachlorbenzene, γ-hexachlorocyclohexane, and several polychlorinated biphenyl congeners with low chlorination were higher in fetal samples and showed poor correlation with maternal levels. Perfluorinated compounds occurred in lower concentrations in cord serum than in maternal serum. Cadmium, lead, mercury, and selenium were all detected in fetal samples, but only mercury showed close correlations among concentrations in different matrices. Although the environmental chemicals examined pass through the placenta and are excreted into milk, partitions between maternal and fetal samples are not uniform.

Comparison of different biopsy forceps models for tissue sampling in eosinophilic esophagitis.

PubMed

Bussmann, Christian; Schoepfer, Alain M; Safroneeva, Ekaterina; Haas, Nadine; Godat, Sébastien; Sempoux, Christine; Simon, Hans-Uwe; Straumann, Alex

2016-12-01

Background and aims: Eosinophilic esophagitis (EoE) is a mixed inflammatory and fibrostenotic disease. Unlike superficial inflammatory changes, subepithelial fibrosis is not routinely sampled in esophageal biopsies. This study aimed to evaluate the efficacy and safety of deep esophageal sampling with four different types of biopsy forceps. Patients and methods: In this cross-sectional study, esophageal biopsies were taken in 30 adult patients by one expert endoscopist. Biopsies sampled from distal esophagus using a static jaw forceps (Olympus, FB-11K-1) were compared with proximal biopsies sampled with static jaw (Olympus, FB-45Q-1), alligator jaw (Olympus, FB-210K), and large-capacity forceps (Boston Scientific, Radial Jaw 4). One pathologist calculated the surface area of epithelial and subepithelial layers in hematoxylin and eosin (H&E)-stained biopsies. Results: Subepithelial tissue was acquired in 97 % (static jaw FB-11K-1), 93 % (static jaw FB-45Q-1), 80 % (alligator jaw), and 55 % (large-capacity) of samples. Median (interquartile [IQR]) ratios of surface area of epithelial to subepithelial tissue were: static jaw FB-45Q-1, 1.07 (0.65 - 4.465); static jaw FB-11K-1, 1.184 (0.608 - 2.545); alligator jaw, 2.353 (1.312 - 4.465); and large-capacity, 2.71 (1.611 - 4.858). The static jaw models obtained a larger surface area of subepithelial tissue compared with the alligator jaw ( P  < 0.001 and P  = 0.037, for FB-11K-1 and FB-45Q-1, respectively) and the large-capacity forceps ( P  < 0.001, for both static jaw models). No esophageal perforations occurred. Conclusions: The static jaw forceps models allowed sampling of subepithelial tissue in > 90 % of biopsies and appear to be superior to alligator or large-capacity forceps in sampling larger amounts of subepithelial tissue. © Georg Thieme Verlag KG Stuttgart · New York.

Evaluation of blood and muscle tissues for molecular detection and characterization of hematozoa infections in northern pintails (Anas acuta) wintering in California

USGS Publications Warehouse

Ramey, Andy M.; Schmutz, Joel A.; Fleskes, Joseph P.; Yabsley, Michael J.

2013-01-01

Information on the molecular detection of hematozoa from different tissue types and multiple years would be useful to inform sample collection efforts and interpret results of meta-analyses or investigations spanning multiple seasons. In this study, we tested blood and muscle tissue collected from northern pintails (Anas acuta) during autumn and winter of different years to evaluate prevalence and genetic diversity ofLeucocytozoon, Haemoproteus, and Plasmodium infections in this abundant waterfowl species of the Central Valley of California. We first compared results for paired blood and wing muscle samples to assess the utility of different tissue types for molecular investigations of haemosporidian parasites. Second, we explored inter-annual variability of hematozoa infection in Central Valley northern pintails and investigated possible effects of age, sex, and sub-region of sample collection on estimated parasite detection probability and prevalence. We found limited evidence for differences between tissue types in detection probability and prevalence ofLeucocytozoon, Haemoproteus, and Plasmodium parasites, which supports the utility of both sample types for obtaining information on hematozoan infections. However, we detected 11 haemosporidian mtDNA cyt bhaplotypes in blood samples vs. six in wing muscle tissue collected during the same sample year suggesting an advantage to using blood samples for investigations of genetic diversity. Estimated prevalence ofLeucocytozoon parasites was greater during 2006–2007 as compared to 2011–2012 and four unique haemosporidian mtDNA cyt b haplotypes were detected in the former sample year but not in the latter. Seven of 15 mtDNA cyt b haplotypes detected in northern pintails had 100% identity with previously reported hematozoa lineages detected in waterfowl (Haemoproteus and Leucocytozoon) or other avian taxa (Plasmodium) providing support for lack of host specificity for some parasite lineages.

Association between infection of virulence cagA gene Helicobacter pylori and laryngeal squamous cell carcinoma

PubMed Central

Burduk, Paweł Krzysztof

2013-01-01

Background The aim of the study was to evaluate the presence of cagA gene Helicobacter pylori in etiopathogenesis of initiation and development of larynx squamous cell carcinoma (LSCC) and its predictable role as a prognostic factor. Material/Methods The prospective, controlled study involved a series of 75 patients (65 male, 10 female, mean age 59.1 years, range 43 to 79 years) with larynx cancer. Samples of larynx cancerous tissue, each of 10–15 mg, were obtained from fresh tissues and were used for nucleic acid purification. DNA was extracted from 225 samples (larynx tumor – I (75), margin of tumor and normal tissue – II (75) and normal larynx tissue from opposite side to the tumor – III). All samples were subjected to H. pylori ureA detection by the PCR H. pylori diagnostic test. Samples that were positive for ureA H. pylori gene were evaluated for cagA H. pylori gene. Results Presence of H. pylori cagA gene was identified in 46,7% to 49,3% of 75 H. pylori ureA gene-positive larynx cancer depending of tissue location. There was a correlation of high incidence of positive cagA gene in larynx cancer tissue in supraglottic versus subglottic and glottic location. We observed a predominance of cagA gene in LSCC in patients with positive cervical lymph nodes and clinical stage T3 and T4. Conclusions H. pylori is present in larynx tissue and may be a possible carcinogen or co-carcinogen in LSCC development, but that must be addressed by future investigations. The presence of cagA gene in larynx cancer tissues significantly decreases survival rate and increases the disease recurrence possibilities. PMID:23860397

Association between infection of virulence cagA gene Helicobacter pylori and laryngeal squamous cell carcinoma.

PubMed

Burduk, Paweł Krzysztof

2013-07-17

The aim of the study was to evaluate the presence of cagA gene Helicobacter pylori in etiopathogenesis of initiation and development of larynx squamous cell carcinoma (LSCC) and its predictable role as a prognostic factor. The prospective, controlled study involved a series of 75 patients (65 male, 10 female, mean age 59.1 years, range 43 to 79 years) with larynx cancer. Samples of larynx cancerous tissue, each of 10-15 mg, were obtained from fresh tissues and were used for nucleic acid purification. DNA was extracted from 225 samples (larynx tumor - I (75), margin of tumor and normal tissue - II (75) and normal larynx tissue from opposite side to the tumor - III). All samples were subjected to H. pylori ureA detection by the PCR H. pylori diagnostic test. Samples that were positive for ureA H. pylori gene were evaluated for cagA H. pylori gene. Presence of H. pylori cagA gene was identified in 46,7% to 49,3% of 75 H. pylori ureA gene-positive larynx cancer depending of tissue location. There was a correlation of high incidence of positive cagA gene in larynx cancer tissue in supraglottic versus subglottic and glottic location. We observed a predominance of cagA gene in LSCC in patients with positive cervical lymph nodes and clinical stage T3 and T4. H. pylori is present in larynx tissue and may be a possible carcinogen or co-carcinogen in LSCC development, but that must be addressed by future investigations. The presence of cagA gene in larynx cancer tissues significantly decreases survival rate and increases the disease recurrence possibilities.

A DNA methylation fingerprint of 1628 human samples

PubMed Central

Fernandez, Agustin F.; Assenov, Yassen; Martin-Subero, Jose Ignacio; Balint, Balazs; Siebert, Reiner; Taniguchi, Hiroaki; Yamamoto, Hiroyuki; Hidalgo, Manuel; Tan, Aik-Choon; Galm, Oliver; Ferrer, Isidre; Sanchez-Cespedes, Montse; Villanueva, Alberto; Carmona, Javier; Sanchez-Mut, Jose V.; Berdasco, Maria; Moreno, Victor; Capella, Gabriel; Monk, David; Ballestar, Esteban; Ropero, Santiago; Martinez, Ramon; Sanchez-Carbayo, Marta; Prosper, Felipe; Agirre, Xabier; Fraga, Mario F.; Graña, Osvaldo; Perez-Jurado, Luis; Mora, Jaume; Puig, Susana; Prat, Jaime; Badimon, Lina; Puca, Annibale A.; Meltzer, Stephen J.; Lengauer, Thomas; Bridgewater, John; Bock, Christoph; Esteller, Manel

2012-01-01

Most of the studies characterizing DNA methylation patterns have been restricted to particular genomic loci in a limited number of human samples and pathological conditions. Herein, we present a compromise between an extremely comprehensive study of a human sample population with an intermediate level of resolution of CpGs at the genomic level. We obtained a DNA methylation fingerprint of 1628 human samples in which we interrogated 1505 CpG sites. The DNA methylation patterns revealed show this epigenetic mark to be critical in tissue-type definition and stemness, particularly around transcription start sites that are not within a CpG island. For disease, the generated DNA methylation fingerprints show that, during tumorigenesis, human cancer cells underwent a progressive gain of promoter CpG-island hypermethylation and a loss of CpG methylation in non-CpG-island promoters. Although transformed cells are those in which DNA methylation disruption is more obvious, we observed that other common human diseases, such as neurological and autoimmune disorders, had their own distinct DNA methylation profiles. Most importantly, we provide proof of principle that the DNA methylation fingerprints obtained might be useful for translational purposes by showing that we are able to identify the tumor type origin of cancers of unknown primary origin (CUPs). Thus, the DNA methylation patterns identified across the largest spectrum of samples, tissues, and diseases reported to date constitute a baseline for developing higher-resolution DNA methylation maps and provide important clues concerning the contribution of CpG methylation to tissue identity and its changes in the most prevalent human diseases. PMID:21613409

Comparison of osmotic swelling influences on meniscal fibrocartilage and articular cartilage tissue mechanics in compression and shear.

PubMed

Nguyen, An M; Levenston, Marc E

2012-01-01

Although the contribution of the circumferential collagen bundles to the anisotropic tensile stiffness of meniscal tissue has been well described, the implications of interactions between tissue components for other mechanical properties have not been as widely examined. This study compared the effects of the proteoglycan-associated osmotic swelling stress on meniscal fibrocartilage and articular cartilage (AC) mechanics by manipulating the osmotic environment and tissue compressive offset. Cylindrical samples were obtained from the menisci and AC of bovine stifles, equilibrated in phosphate-buffered saline solutions ranging from 0.1× to 10×, and tested in oscillatory torsional shear and unconfined compression. Biochemical analysis indicated that treatments and testing did not substantially alter tissue composition. Mechanical testing revealed tissue-specific responses to both increasing compressive offset and decreasing bath salinity. Most notably, reduced salinity dramatically increased the shear modulus of both axially and circumferentially oriented meniscal tissue explants to a much greater extent than for cartilage samples. Combined with previous studies, these findings suggest that meniscal proteoglycans have a distinct structural role, stabilizing, and stiffening the matrix surrounding the primary circumferential collagen bundles. Copyright © 2011 Orthopaedic Research Society.

Acoustic properties of a short-finned pilot whale head with insight into temperature influence on tissues' sound velocity.

PubMed

Dong, Jianchen; Song, Zhongchang; Li, Songhai; Gong, Zining; Li, Kuan; Zhang, Peijun; Zhang, Yu; Zhang, Meng

2017-10-01

Acoustic properties of odontocete head tissues, including sound velocity, density, and acoustic impedance, are important parameters to understand dynamics of its echolocation. In this paper, acoustic properties of head tissues from a freshly dead short-finned pilot whale (Globicephala macrorhynchus) were reconstructed using computed tomography (CT) and ultrasound. The animal's forehead soft tissues were cut into 188 ordered samples. Sound velocity, density, and acoustic impedance of each sample were either directly measured or calculated by formula, and Hounsfield Unit values (HUs) were obtained from CT scanning. According to relationships between HUs and sound velocity, HUs and density, as well as HUs and acoustic impedance, distributions of acoustic properties in the head were reconstructed. The inner core in the melon with low-sound velocity and low-density is an evidence for its potential function of sound focusing. The increase in acoustic impedance of forehead tissues from inner core to outer layer may be important for the acoustic impedance matching between the outer layer tissue and seawater. In addition, temperature dependence of sound velocity in soft tissues was also examined. The results provide a guide to the simulation of the sound emission of the short-finned pilot whale.

An Automated Platform for High-Resolution Tissue Imaging Using Nanospray Desorption Electrospray Ionization Mass Spectrometry

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lanekoff, Ingela T.; Heath, Brandi S.; Liyu, Andrey V.

2012-10-02

An automated platform has been developed for acquisition and visualization of mass spectrometry imaging (MSI) data using nanospray desorption electrospray ionization (nano-DESI). The new system enables robust operation of the nano-DESI imaging source over many hours. This is achieved by controlling the distance between the sample and the probe by mounting the sample holder onto an automated XYZ stage and defining the tilt of the sample plane. This approach is useful for imaging of relatively flat samples such as thin tissue sections. Custom software called MSI QuickView was developed for visualization of large data sets generated in imaging experiments. MSImore » QuickView enables fast visualization of the imaging data during data acquisition and detailed processing after the entire image is acquired. The performance of the system is demonstrated by imaging rat brain tissue sections. High resolution mass analysis combined with MS/MS experiments enabled identification of lipids and metabolites in the tissue section. In addition, high dynamic range and sensitivity of the technique allowed us to generate ion images of low-abundance isobaric lipids. High-spatial resolution image acquired over a small region of the tissue section revealed the spatial distribution of an abundant brain metabolite, creatine, in the white and gray matter that is consistent with the literature data obtained using magnetic resonance spectroscopy.« less

In vitro double-integrating-sphere optical properties of tissues between 630 and 1064 nm

NASA Astrophysics Data System (ADS)

Beek, J. F.; Blokland, P.; Posthumus, P.; Aalders, M.; Pickering, J. W.; Sterenborg, H. J. C. M.; van Gemert, M. J. C.

1997-11-01

The optical properties (absorption and scattering coefficients and the scattering anisotropy factor) were measured in vitro for cartilage, liver, lung, muscle, myocardium, skin, and tumour (colon adenocarcinoma CC 531) at 630, 632.8, 790, 850 and 1064 nm. Rabbits, rats, piglets, goats, and dogs were used to obtain the tissues. A double-integrating-sphere setup with an intervening sample was used to determine the reflectance, and the diffuse and collimated transmittances of the sample. The inverse adding - doubling algorithm was used to determine the optical properties from the measurements. The overall results were comparable to those available in the literature, although only limited data are available at 790 - 850 nm. The results were reproducible for a specific sample at a specific wavelength. However, when comparing the results of different samples of the same tissue or different lasers with approximately the same wavelength (e.g. argon dye laser at 630 nm and HeNe laser at 632.8 nm) variations are large. We believe these variations in optical properties should be explained by biological variations of the tissues. In conclusion, we report on an extensive set of in vitro absorption and scattering properties of tissues measured with the same equipment and software, and by the same group. Although the accuracy of the method requires further improvement, it is highly likely that the other existing data in the literature have a similar level of accuracy.

Viscoelastic Property Measurement in Thin Tissue Constructs Using Ultrasound

PubMed Central

Liu, Dalong; Ebbini, Emad S.

2010-01-01

We present a dual-element concave ultrasound transducer system for generating and tracking of localized tissue displacements in thin tissue constructs on rigid substrates. The system is comprised of a highly focused PZT-4 5-MHz acoustic radiation force (ARF) transducer and a confocal 25-MHz polyvinylidene fluoride imaging transducer. This allows for the generation of measurable displacements in tissue samples on rigid substrates with thickness values down to 500 µm. Impulse-like and longer duration sine-modulated ARF pulses are possible with intermittent M-mode data acquisition for displacement tracking. The operations of the ARF and imaging transducers are strictly synchronized using an integrated system for arbitrary waveform generation and data capture with a shared timebase. This allows for virtually jitter-free pulse-echo data well suited for correlation-based speckle tracking. With this technique we could faithfully capture the entire dynamics of the tissue axial deformation at pulse-repetition frequency values up to 10 kHz. Spatio-temporal maps of tissue displacements in response to a variety of modulated ARF beams were produced in tissue-mimicking elastography phantoms on rigid substrates. The frequency response was measured for phantoms with different modulus and thickness values. The frequency response exhibited resonant behavior with the resonance frequency being inversely proportional to the sample thickness. This resonant behavior can be used in obtaining high-contrast imaging using magnitude and phase response to sinusoidally modulated ARF beams. Furthermore, a second order forced harmonic oscillator (FHO) model was shown to capture this resonant behavior. Based on the FHO model, we used the extended Kalman filter (EKF) for tracking the apparent modulus and viscosity of samples subjected to dc and sinusoidally modulated ARF. The results show that the stiffness (apparent modulus) term in the FHO is largely time-invariant and can be estimated robustly using the EKF. On the other hand, the damping (apparent viscosity) is time varying. These findings were confirmed by comparing the magnitude response of the FHO (with parameters obtained using the EKF) with the measured ones for different thin tissue constructs. PMID:18334343

Preservation and rapid purification of DNA from decomposing human tissue samples.

PubMed

Sorensen, Amy; Rahman, Elizabeth; Canela, Cassandra; Gangitano, David; Hughes-Stamm, Sheree

2016-11-01

One of the key features to be considered in a mass disaster is victim identification. However, the recovery and identification of human remains are sometimes complicated by harsh environmental conditions, limited facilities, loss of electricity and lack of refrigeration. If human remains cannot be collected, stored, or identified immediately, bodies decompose and DNA degrades making genotyping more difficult and ultimately decreasing DNA profiling success. In order to prevent further DNA damage and degradation after collection, tissue preservatives may be used. The goal of this study was to evaluate three customized (modified TENT, DESS, LST) and two commercial DNA preservatives (RNAlater and DNAgard ® ) on fresh and decomposed human skin and muscle samples stored in hot (35°C) and humid (60-70% relative humidity) conditions for up to three months. Skin and muscle samples were harvested from the thigh of three human cadavers placed outdoors for up to two weeks. In addition, the possibility of purifying DNA directly from the preservative solutions ("free DNA") was investigated in order to eliminate lengthy tissue digestion processes and increase throughput. The efficiency of each preservative was evaluated based on the quantity of DNA recovered from both the "free DNA" in solution and the tissue sample itself in conjunction with the quality and completeness of downstream STR profiles. As expected, DNA quantity and STR success decreased with time of decomposition. However, a marked decrease in DNA quantity and STR quality was observed in all samples after the bodies entered the bloat stage (approximately six days of decomposition in this study). Similar amounts of DNA were retrieved from skin and muscle samples over time, but slightly more complete STR profiles were obtained from muscle tissue. Although higher amounts of DNA were recovered from tissue samples than from the surrounding preservative, the average number of reportable alleles from the "free DNA" was comparable. Overall, DNAgard ® and the modified TENT buffer were the most successful tissue preservatives tested in this study based on STR profile success from "free DNA" in solution when decomposing tissues were stored for up to three months in hot, humid conditions. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

Identification of a Novel Human Papillomavirus, Type HPV199, Isolated from a Nasopharynx and Anal Canal, and Complete Genomic Characterization of Papillomavirus Species Gamma-12

PubMed Central

Oštrbenk, Anja; Kocjan, Boštjan J.; Hošnjak, Lea; Li, Jingjing; Deng, Qiuju; Šterbenc, Anja; Poljak, Mario

2015-01-01

The novel human papillomavirus type 199 (HPV199) was initially identified in a nasopharyngeal swab sample obtained from a 25 year-old immunocompetent male. The complete genome of HPV199 is 7,184 bp in length with a GC content of 36.5%. Comparative genomic characterization of HPV199 and its closest relatives showed the classical genomic organization of Gammapapillomaviruses (Gamma-PVs). HPV199 has seven major open reading frames (ORFs), encoding five early (E1, E2, E4, E6, and E7) and two late (L1 and L2) proteins, while lacking the E5 ORF. The long control region (LCR) of 513 bp is located between the L1 and E6 ORFs. Phylogenetic analysis additionally confirmed that HPV-199 clusters into the Gamma-PV genus, species Gamma-12, additionally containing HPV127, HV132, HPV148, HPV165, and three putative HPV types: KC5, CG2 and CG3. HPV199 is most closely related to HPV127 (nucleotide identity 77%). The complete viral genome sequence of additional HPV199 isolate was determined from anal canal swab sample. Two HPV199 complete viral sequences exhibit 99.4% nucleotide identity. To the best of our knowledge, this is the first member of Gamma-PV with complete nucleotide sequences determined from two independent clinical samples. To evaluate the tissue tropism of the novel HPV type, 916 clinical samples were tested using HPV199 type-specific real-time PCR: HPV199 was detected in 2/76 tissue samples of histologically confirmed common warts, 2/108 samples of eyebrow hair follicles, 2/137 anal canal swabs obtained from individuals with clinically evident anal pathology, 4/184 nasopharyngeal swabs and 3/411 cervical swabs obtained from women with normal cervical cytology. Although HPV199 was found in 1.4% of cutaneous and mucosal samples only, it exhibits dual tissue tropism. According to the results of our study and literature data, dual tropism of all Gamma-12 members is highly possible. PMID:26375679

Automated cell disruption is a reliable and effective method of isolating RNA from fresh snap-frozen normal and malignant oral mucosa samples.

PubMed

Van der Vorst, Sébastien; Dekairelle, Anne-France; Irenge, Léonid; Hamoir, Marc; Robert, Annie; Gala, Jean-Luc

2009-01-01

This study compared automated vs. manual tissue grinding in terms of RNA yield obtained from oral mucosa biopsies. A total of 20 patients undergoing uvulectomy for sleep-related disorders and 10 patients undergoing biopsy for head and neck squamous cell carcinoma were enrolled in the study. Samples were collected, snap-frozen in liquid nitrogen, and divided into two parts of similar weight. Sample grinding was performed on one sample from each pair, either manually or using an automated cell disruptor. The performance and efficacy of each homogenization approach was compared in terms of total RNA yield (spectrophotometry, fluorometry), mRNA quantity [densitometry of specific TP53 amplicons and TP53 quantitative reverse-transcribed real-time PCR (qRT-PCR)], and mRNA quality (functional analysis of separated alleles in yeast). Although spectrophotometry and fluorometry results were comparable for both homogenization methods, TP53 expression values obtained by amplicon densitometry and qRT-PCR were significantly and consistently better after automated homogenization (p<0.005) for both uvula and tumor samples. Functional analysis of separated alleles in yeast results was better with the automated technique for tumor samples. Automated tissue homogenization appears to be a versatile, quick, and reliable method of cell disruption and is especially useful in the case of small malignant samples, which show unreliable results when processed by manual homogenization.

Using Stable Isotopes of Carbon and Nitrogen to Evaluate Trophic Interactions in Aquatic Environments

ERIC Educational Resources Information Center

Christensen, David R.; LaRoche, Andrew

2012-01-01

This paper describes a series of laboratory exercises for upper level biology courses, independent research and/or honors programs. Students sampled fish from a local water body with the assistance of a local fish and wildlife agency. Tissue samples from collected fish were utilized to obtain estimates of the stable isotopes delta[superscript 13]C…

Sampling and Analyzing Air Pollution: An Apparatus Suitable for Use in Schools.

ERIC Educational Resources Information Center

Rockwell, Dean M.; Hansen, Tony

1994-01-01

Describes two variations of an air sampler and analyzer that are inexpensive to construct, easy to operate, and designed to be used in an educational program. Variations use vacuum cleaners and aquarium pumps, and white facial tissues serve as filters. Samples of air pollution obtained by this method may be used from early grade school to advanced…

Determination of organic acids in tissues and exudates of maize, lupin, and chickpea by high-performance liquid chromatography-tandem mass spectrometry.

PubMed

Erro, Javier; Zamarreño, Angel M; Yvin, Jean-Claude; Garcia-Mina, Jose M

2009-05-27

This article describes a fast and simple methodology for the extraction and determination of organic acids in tissues and root exudates of maize, lupin, and chickpea by LC/MS/MS. Its main advantage is that it does not require sample prepurification before HPLC analysis or sample derivatization to improve sensibility. The results obtained showed good precision and accuracy, a recovery close to 100%, and no significant matrix effect. Moreover, the sensibility of the method is in general better than that of previously described methodologies, with detection limits between 15 and 900 pg injected.

A family of hyperelastic models for human brain tissue

NASA Astrophysics Data System (ADS)

Mihai, L. Angela; Budday, Silvia; Holzapfel, Gerhard A.; Kuhl, Ellen; Goriely, Alain

2017-09-01

Experiments on brain samples under multiaxial loading have shown that human brain tissue is both extremely soft when compared to other biological tissues and characterized by a peculiar elastic response under combined shear and compression/tension: there is a significant increase in shear stress with increasing axial compression compared to a moderate increase with increasing axial tension. Recent studies have revealed that many widely used constitutive models for soft biological tissues fail to capture this characteristic response. Here, guided by experiments of human brain tissue, we develop a family of modeling approaches that capture the elasticity of brain tissue under varying simple shear superposed on varying axial stretch by exploiting key observations about the behavior of the nonlinear shear modulus, which can be obtained directly from the experimental data.

Improved Butanol-Methanol (BUME) Method by Replacing Acetic Acid for Lipid Extraction of Biological Samples.

PubMed

Cruz, Mutya; Wang, Miao; Frisch-Daiello, Jessica; Han, Xianlin

2016-07-01

Extraction of lipids from biological samples is a critical step in lipidomics, especially for shotgun lipidomics where lipid extracts are directly infused into a mass spectrometer. The butanol-methanol (BUME) extraction method was originally developed to extract lipids from plasma samples with 1 % acetic acid. Considering some lipids are sensitive to acidic environments, we modified this protocol by replacing acetic acid with lithium chloride solution and extended the modified extraction to tissue samples. Although no significant reduction of plasmalogen levels in the acidic BUME extracts of rat heart samples was found, the modified method was established to extract various tissue samples, including rat liver, heart, and plasma. Essentially identical profiles of the majority of lipid classes were obtained from the extracts of the modified BUME and traditional Bligh-Dyer methods. However, it was found that neither the original, nor the modified BUME method was suitable for 4-hydroxyalkenal species measurement in biological samples.

Improved Butanol-Methanol (BUME) Method by Replacing Acetic Acid for Lipid Extraction of Biological Samples

PubMed Central

Cruz, Mutya; Wang, Miao; Frisch-Daiello, Jessica; Han, Xianlin

2016-01-01

Extraction of lipids from biological samples is a critical step in lipidomics, especially for shotgun lipidomics where lipid extracts are directly infused into a mass spectrometer. The butanol-methanol (BUME) extraction method was originally developed to extract lipids from plasma samples with 1% acetic acid. Considering some lipids are sensitive to acidic environments, we modified this protocol by replacing acetic acid with lithium chloride solution and extended the modified extraction to tissue samples. Although no significant reduction of plasmalogen levels in the acidic BUME extracts of rat heart samples was found, the modified method was established to extract various tissue samples, including rat liver, heart, and plasma. Essentially identical profiles of the majority of lipid classes were obtained from the extracts of the modified BUME and traditional Bligh-Dyer methods. However, it was found that neither the original, nor the modified BUME method was suitable for 4-hydroxyalkenal species measurement in biological samples. PMID:27245345

Detection of virus in shrimp using digital color correlation

NASA Astrophysics Data System (ADS)

Alvarez-Borrego, Josue; Chavez-Sanchez, Cristina; Bueno-Ibarra, Mario A.

1999-07-01

Detection of virus in shrimp tissue using digital color correlation is presented. Phase filters in three channels (red, green and blue) were used in order to detect HPV virus like target. These first results obtained showed that is possible to detect virus in shrimp tissue. More research must be made with color correlation in order to consider natural morphology of the virus, color, scale and rotation and noise in the samples.

RT-PCR analysis of RNA extracted from Bouin-fixed and paraffin-embedded lymphoid tissues.

PubMed

Gloghini, Annunziata; Canal, Barbara; Klein, Ulf; Dal Maso, Luigino; Perin, Tiziana; Dalla-Favera, Riccardo; Carbone, Antonino

2004-11-01

In the present study, we have investigated whether RNA can be efficiently isolated from Bouin-fixed or formalin-fixed, paraffin-embedded lymphoid tissue specimens. To this aim, we applied a new and simple method that includes the combination of proteinase K digestion and column purification. By this method, we demonstrated that the amplification of long fragments could be accomplished after a pre-heating step before cDNA synthesis associated with the use of enzymes that work at high temperature. By means of PCR using different primers for two examined genes (glyceraldehyde-3-phosphate dehydrogenase [GAPDH]- and CD40), we amplified segments of cDNA obtained by reverse transcription of the isolated RNA extracted from Bouin-fixed or formalin-fixed paraffin-embedded tissues. Amplified fragments of the expected sizes were obtained for both genes tested indicating that this method is suitable for the isolation of high-quality RNA. To explore the possibility for giving accurate real time quantitative RT-PCR results, cDNA obtained from matched frozen, Bouin-fixed and formalin-fixed neoplastic samples (two diffuse large cell lymphomas, one plasmacytoma) was tested for the following target genes: CD40, Aquaporin-3, BLIMP1, IRF4, Syndecan-1. Delta threshold cycle (DeltaC(T)) values for Bouin-fixed and formalin-fixed paraffin-embedded tissues and their correlation with those for frozen samples showed an extremely high correlation (r > 0.90) for all of the tested genes. These results show that the method of RNA extraction we propose is suitable for giving accurate real time quantitative RT-PCR results.

Classification of superficial lesions of the eye with an optical biopsy system: First trials with the Los Alamos instrument

DOE Office of Scientific and Technical Information (OSTI.GOV)

Glickman, R.D.; Gritz, D.C.; Held, K.S.

the clinical diagnosis of a lesion often requires that a histological analysis be made of a physical specimen of the suspect tissue. In the present work, the authors have utilized an optical biopsy system (OBS) developed at Los Alamos National Laboratory which is safe for patient use and provides a large amount of optical data from the sampled tissue. An earlier version of this system has been used to study age-related changes in the ocular lens (10). The purpose of the present study is to establish the potential clinical utility of the OBS by determining if characteristic features in themore » optical signatures, obtained from a variety of ophthalmic lesions, are correlated with the histological features of tissue biopsies obtained from these patients.« less

Absolute quantification methods in tissue near-infrared spectroscopy

NASA Astrophysics Data System (ADS)

Matcher, Steven J.; Kirkpatrick, Peter J.; Nahid, K.; Cope, Mark; Delpy, David T.

1995-05-01

Recent work aimed at providing an absolute measurement of tissue haemoglobin saturation and a new instrument development, the spatially resolved spectrometer (SRS), are discussed. The theoretical basis of operation of this device and its hardware implementation are described and the results of validation studies on tissue simulating phantoms are presented as are preliminary measurements on human volunteers and observations on patients undergoing neurosurgery. In its present form the instrument appears to produce absolute haemoglobin saturation values for resting human skeletal muscle and the normally perfused human head which are rather low based on physiological expectations. However, we obtained a tight correlation between the saturation values measured by the SRS instrument and those obtained from blood-gas analysis of samples drawn from a jugular bulb catheter in one neurosurgery subject during clamping of the right carotid arteries.

Monochromatic computed microtomography using laboratory and synchrotron sources and X-ray fluorescence analysis for comprehensive analysis of structural changes in bones.

PubMed

Buzmakov, Alexey; Chukalina, Marina; Nikolaev, Dmitry; Gulimova, Victoriya; Saveliev, Sergey; Tereschenko, Elena; Seregin, Alexey; Senin, Roman; Zolotov, Denis; Prun, Victor; Shaefer, Gerald; Asadchikov, Victor

2015-06-01

A combination of X-ray tomography at different wavelengths and X-ray fluorescence analysis was applied in the study of two types of bone tissue changes: prolonged presence in microgravity conditions and age-related bone growth. The proximal tail vertebrae of geckos were selected for investigation because they do not bear the supporting load in locomotion, which allows them to be considered as an independent indicator of gravitational influence. For the vertebrae of geckos no significant differences were revealed in the elemental composition of the flight samples and the synchronous control samples. In addition, the gecko bone tissue samples from the jaw apparatus, spine and shoulder girdle were measured. The dynamics of structural changes in the bone tissue growth was studied using samples of a human fetal hand. The hands of human fetuses of 11-15 weeks were studied. Autonomous zones of calcium accumulation were found not only in individual fingers but in each of the investigated phalanges. The results obtained are discussed.

Monochromatic computed microtomography using laboratory and synchrotron sources and X-ray fluorescence analysis for comprehensive analysis of structural changes in bones1

PubMed Central

Buzmakov, Alexey; Chukalina, Marina; Nikolaev, Dmitry; Gulimova, Victoriya; Saveliev, Sergey; Tereschenko, Elena; Seregin, Alexey; Senin, Roman; Zolotov, Denis; Prun, Victor; Shaefer, Gerald; Asadchikov, Victor

2015-01-01

A combination of X-ray tomography at different wavelengths and X-ray fluorescence analysis was applied in the study of two types of bone tissue changes: prolonged presence in microgravity conditions and age-related bone growth. The proximal tail vertebrae of geckos were selected for investigation because they do not bear the supporting load in locomotion, which allows them to be considered as an independent indicator of gravitational influence. For the vertebrae of geckos no significant differences were revealed in the elemental composition of the flight samples and the synchronous control samples. In addition, the gecko bone tissue samples from the jaw apparatus, spine and shoulder girdle were measured. The dynamics of structural changes in the bone tissue growth was studied using samples of a human fetal hand. The hands of human fetuses of 11–15 weeks were studied. Autonomous zones of calcium accumulation were found not only in individual fingers but in each of the investigated phalanges. The results obtained are discussed. PMID:26089762

Design and optimization of reverse-transcription quantitative PCR experiments.

PubMed

Tichopad, Ales; Kitchen, Rob; Riedmaier, Irmgard; Becker, Christiane; Ståhlberg, Anders; Kubista, Mikael

2009-10-01

Quantitative PCR (qPCR) is a valuable technique for accurately and reliably profiling and quantifying gene expression. Typically, samples obtained from the organism of study have to be processed via several preparative steps before qPCR. We estimated the errors of sample withdrawal and extraction, reverse transcription (RT), and qPCR that are introduced into measurements of mRNA concentrations. We performed hierarchically arranged experiments with 3 animals, 3 samples, 3 RT reactions, and 3 qPCRs and quantified the expression of several genes in solid tissue, blood, cell culture, and single cells. A nested ANOVA design was used to model the experiments, and relative and absolute errors were calculated with this model for each processing level in the hierarchical design. We found that intersubject differences became easily confounded by sample heterogeneity for single cells and solid tissue. In cell cultures and blood, the noise from the RT and qPCR steps contributed substantially to the overall error because the sampling noise was less pronounced. We recommend the use of sample replicates preferentially to any other replicates when working with solid tissue, cell cultures, and single cells, and we recommend the use of RT replicates when working with blood. We show how an optimal sampling plan can be calculated for a limited budget. .

The effects of CD147 on the cell proliferation, apoptosis, invasion, and angiogenesis in glioma.

PubMed

Yin, Haoyuan; Shao, Ying; Chen, Xuan

2017-01-01

To analyze the effects of extracellular matrix metalloproteinase inducer (CD147) on glioma proliferation, apoptosis, invasion, and angiogenesis. Tissue samples were obtained from 101 glioma cases while normal brain tissues were obtained from 30 brain injury cases. Immunohistochemical assay was performed to detect the expressions of CD147, CD34, and VEGF in tissue samples. QRT-PCR was performed to detect the relative expression of CD147 mRNA in human glioma cell lines. CD147 siRNA was transfected into glioma cell line U251. Cell proliferation, apoptosis, invasion, and angiogenesis were tested by MTT, flow cytometry, Transwell assay, and vasculogenic mimicry assay, respectively. Expressions of relative proteins were analyzed with western blot. CD147 was positively expressed with the percentage of 0, 37.5, 44.8, 67.9, and 85.7 % in normal tissues and glioma tissues with WHO grades I-IV, respectively, and the scores of MVDand VEGF were associated with the expression of CD147. CD147 was significantly upregulated in the human glioma cell lines (P < 0.05). Downregulated the expression of CD147 suppressed cell proliferation, blocked cell cycle, induced apoptosis, inhibited cell invasion and angiogenesis in glioma cells in vitro. The expression of CD147 was significantly associated with WHO tumor grade and angiogenesis; silencing of CD147 contributed to inhibition of glioma proliferation, invasion, and angiogenesis. Our study provided firm evidence that CD 147 is a potential glioma target for anti-angiogenic therapies.

Immunohistochemical Evaluation of Neuroreceptors in Healthy and Pathological Temporo-Mandibular Joint

PubMed Central

Favia, Gianfranco; Corsalini, Massimo; Di Venere, Daniela; Pettini, Francesco; Favia, Giorgio; Capodiferro, Saverio; Maiorano, Eugenio

2013-01-01

Aim: A study was performed on the articular disk and periarticular tissues of the temporo-mandibular joint (TMJ) with immunohistochemical techniques to give evidence to the presence of neuroreceptors (NRec) in these sites. Methods: The study was carried out on tissue samples obtained from 10 subjects without TMJ disease and from 7 patients with severe TMJ arthritis and arthrosis. We use antibodies directed against following antigens: Gliofibrillary Acidic Protein (GFAP), Leu-7, Myelin Basic Protein (MBP), Neurofilaments 68 kD (NF), Neuron Specific Enolase (NSE), S-100 protein (S-100) and Synaptophysin (SYN). Results: This study revealed that Ruffini's-like, Pacini's-like and Golgi's-like receptors can be demonstrated in TMJ periarticular tissues and that free nervous endings are present in the subsynovial tissues but not within the articular disk. We observed elongated cytoplamic processes of chondrocytes that demonstrated strong S-100 immunoreactivity but they were unreactive with all other antibodies. These cytoplamic processes were more abundant and thicker in the samples obtained from patients with disease TMJ. Conclusion: The results of this study confirm that different Nrec are detectable in TMJ periarticular tissues but they are absent within the articular disk. In the latter site, only condrocytic processes are evident, especially in diseased TMJ, and they might have been confused with nervous endings in previous morphological studies. Nevertheless the absence of immunoreactivity for NF, NSE and SYN proves that they are not of neural origin. PMID:24151441

Automated acoustic matrix deposition for MALDI sample preparation.

PubMed

Aerni, Hans-Rudolf; Cornett, Dale S; Caprioli, Richard M

2006-02-01

Novel high-throughput sample preparation strategies for MALDI imaging mass spectrometry (IMS) and profiling are presented. An acoustic reagent multispotter was developed to provide improved reproducibility for depositing matrix onto a sample surface, for example, such as a tissue section. The unique design of the acoustic droplet ejector and its optimization for depositing matrix solution are discussed. Since it does not contain a capillary or nozzle for fluid ejection, issues with clogging of these orifices are avoided. Automated matrix deposition provides better control of conditions affecting protein extraction and matrix crystallization with the ability to deposit matrix accurately onto small surface features. For tissue sections, matrix spots of 180-200 microm in diameter were obtained and a procedure is described for generating coordinate files readable by a mass spectrometer to permit automated profile acquisition. Mass spectral quality and reproducibility was found to be better than that obtained with manual pipet spotting. The instrument can also deposit matrix spots in a dense array pattern so that, after analysis in a mass spectrometer, two-dimensional ion images may be constructed. Example ion images from a mouse brain are presented.

Bi-PROF

PubMed Central

Gries, Jasmin; Schumacher, Dirk; Arand, Julia; Lutsik, Pavlo; Markelova, Maria Rivera; Fichtner, Iduna; Walter, Jörn; Sers, Christine; Tierling, Sascha

2013-01-01

The use of next generation sequencing has expanded our view on whole mammalian methylome patterns. In particular, it provides a genome-wide insight of local DNA methylation diversity at single nucleotide level and enables the examination of single chromosome sequence sections at a sufficient statistical power. We describe a bisulfite-based sequence profiling pipeline, Bi-PROF, which is based on the 454 GS-FLX Titanium technology that allows to obtain up to one million sequence stretches at single base pair resolution without laborious subcloning. To illustrate the performance of the experimental workflow connected to a bioinformatics program pipeline (BiQ Analyzer HT) we present a test analysis set of 68 different epigenetic marker regions (amplicons) in five individual patient-derived xenograft tissue samples of colorectal cancer and one healthy colon epithelium sample as a control. After the 454 GS-FLX Titanium run, sequence read processing and sample decoding, the obtained alignments are quality controlled and statistically evaluated. Comprehensive methylation pattern interpretation (profiling) assessed by analyzing 102-104 sequence reads per amplicon allows an unprecedented deep view on pattern formation and methylation marker heterogeneity in tissues concerned by complex diseases like cancer. PMID:23803588

Dispersion analysis of collagen fiber networks in cervical tissue using optical coherence tomography (Conference Presentation)

NASA Astrophysics Data System (ADS)

Gan, Yu; Yao, Wang; Myers, Kristin M.; Vink, Joy Y.; Wapner, Ronald J.; Hendon, Christine P.

2016-02-01

Understanding the human cervical collagen fiber network is critical to delineating the physiology of cervical remodeling during pregnancy. Previously, we presented our methodology to study the ultrastructure of collagen fibers over an entire field of transverse slices of human cervix tissue using optical coherence tomography. Here, we present a pixel-wise fiber orientation method to enable dispersion analysis on entire slices of human cervical tissues. We obtained en face images that were parallel to the surface. In each en face image, we masked the collagen fiber region based on signal noise ratio. Then, we extracted fiber orientations in each pixel using a weighted summation scheme and generated a pixel-wise directionality map within the entire region. The weight was determined by intensity variations between a pixel of interest and its neighboring pixels and their corresponding distances. We divided the directionality map into regions of 400 μm × 400 μm along radial direction in all four quadrants. In each region, we fit von-Mises distribution to fiber orientations of pixels with mode θ and dispersion b. We compared dispersions among regions and samples. Using IRB approved protocols, we obtained whole transverse slices of cervical tissue from pregnant (n = 2) and non-pregnant (n = 13) women. We observed higher dispersion in pregnant samples compared to non-pregnant samples and higher dispersions in patient's right/left zones than posterior/anterior zones within an axial slice. Future studies will analyze how collagen fiber dispersion patterns change from the internal to the external os.

Experimental and constitutive modeling approaches for a study of biomechanical properties of human coronary arteries.

PubMed

Jankowska, Malgorzata A; Bartkowiak-Jowsa, Magdalena; Bedzinski, Romuald

2015-10-01

The study concerns the determination of mechanical properties of human coronary arterial walls with both experimental and constitutive modeling approaches. The research material was harvested from 18 patients (range 50-84 years). On the basis of hospital records and visual observation, each tissue sample was classified according to the stage (0, I, II, III) of atherosclerosis development (SAD). Then, strip samples considered as a membrane with the shape of rectangular parallelepiped were preconditioned and subjected to uniaxial tensile tests in longitudinal (n=27) and circumferential (n=4) direction. With experimental data obtained, the stress-strain characteristics were prepared. Furthermore, tensile strengths and related strains, stiffness coefficients and tangent modules of elasticity were computed. For a constitutive model of passive mechanical behavior of coronary arteries, values of material parameters were computed. The studies led to the following conclusions. Most importantly, the atherosclerotic changes affect all the mechanical properties of arterial walls. A progress of arteriosclerosis contributes to an increase of vascular stiffness. The highest values of the stiffness coefficients are obtained for the tissues in the advanced stage of the disease. We were also able to observe that gradual calcification, progression of atherosclerosis and degradation of collagen in the tissue caused a decrease of tensile strengths and related strains. Finally, a comparison made for the tissues with the advanced SAD showed that the tensile strengths and strains were much higher in the case of the samples with the circumferential orientation rather than those with the longitudinal one. Copyright © 2015 Elsevier Ltd. All rights reserved.

Re-generation of tissue about an animal-based scaffold: AMS studies of the fate of the scaffold

NASA Astrophysics Data System (ADS)

Rickey, Frank A.; Elmore, David; Hillegonds, Darren; Badylak, Stephen; Record, Rae; Simmons-Byrd, Abby

2000-10-01

Small intestinal submucosa (SIS) is an unusual tissue, which shows great promise for the repair of damaged tissues in humans. When the SIS is used as a surgical implant, the porcine-derived material is not rejected by the host immune system, and in fact stimulates the constructive re-modeling of damaged tissue. In dogs, these SIS scaffolds have been used to grow new arteries, tendons, and urinary bladders. Moreover, the SIS scaffold tissue seems to disappear from the implant region after a few months. The fate of this SIS tissue is of considerable importance if it is to be used in human tissue repair. SIS is obtained from pigs. We have labeled the SIS in several pigs by intraveneous administration of 14C enriched proline from the age of three weeks until they reach market weight. The prepared SIS was then implanted in dogs as scaffolds for urinary bladder patches. During the remaining life of each dog, blood, urine and feces samples were collected on a regular schedule. AMS analyses of these specimens were performed to measure the elimination rate of the SIS. At different intervals, the dogs were sacrificed. Tissue samples were analyzed by AMS to determine the whole-body distribution of the labeled SIS.

High-quality RNA extracted from biopsied samples dehydrated and stored dried at room temperature without chemical preservation for up to 3 months as evidenced by RT-PCR results.

PubMed

Sadler, Theodore R; Khodavirdi, Ani C

2015-07-01

Handling and maintenance of biological tissues for nucleic acid and/or protein analysis has long been a challenge because of the perceived instability of these molecules at room temperature if not preserved or processed. Structural damage and compromised integrity of aforementioned biomolecules subsequent to preservation have also posed difficulties in their use in research. The development of technologies employing nonfixative methods with the capability to store at room temperature have been of growing interest. Our previous publication exploring preservation of proteins by desiccation challenged the convention of their unstable nature. Herein, we report the results of quantitative and qualitative analyses of RNA from tissue samples that were desiccated and stored at room temperature for up to 3 months. Our results indicate that viable RNA can be obtained from dehydrated ex vivo tissue samples that have been stored at room temperature.

SPRUCE Shrub-Layer Growth Assessments in S1-Bog Plots and SPRUCE Experimental Plots beginning in 2010

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hanson, P.J.; Phillips, J.R.; Brice, D.J.

This data set reports shrub layer growth assessments for the S1-Bog on the Marcell Experimental Forest in Minnesota from 2010 through 2017. Data were obtained by destructively harvesting two 0.25 m2 plots within defined plot areas of the S1-Bog or SPRUCE experimental plots. In 2015, SPRUCE plots 4, 6, 8, 10, 11, 13, 16, 17, 19 and 20 were enclosed in the SPRUCE enclosures. Prior to 2015 all data are for open ambient conditions. In early years a distinct hummock and a hollow sampling square were both collected, but in later years unsampled hollow areas became unavailable due to priormore » sampling or instrument installations. All vegetation material above the Sphagnum surface of the bog was clipped and transferred to plastic storage bags which were then frozen until the samples could be sorted. Sorting was done by species, tissue type (leaves vs. stems) and tissue age (current-year vs. older tissues).« less

A rabbit model of non-typhoidal Salmonella bacteremia.

PubMed

Panda, Aruna; Tatarov, Ivan; Masek, Billie Jo; Hardick, Justin; Crusan, Annabelle; Wakefield, Teresa; Carroll, Karen; Yang, Samuel; Hsieh, Yu-Hsiang; Lipsky, Michael M; McLeod, Charles G; Levine, Myron M; Rothman, Richard E; Gaydos, Charlotte A; DeTolla, Louis J

2014-09-01

Bacteremia is an important cause of morbidity and mortality in humans. In this study, we focused on the development of an animal model of bacteremia induced by non-typhoidal Salmonella. New Zealand White rabbits were inoculated with a human isolate of non-typhoidal Salmonella strain CVD J73 via the intra-peritoneal route. Blood samples were collected at specific time points and at euthanasia from infected rabbits. Additionally, tissue samples from the heart, lungs, spleen, gastrointestinal tract, liver and kidneys were obtained at euthanasia. All experimentally infected rabbits displayed clinical signs of disease (fever, dehydration, weight loss and lethargy). Tissues collected at necropsy from the animals exhibited histopathological changes indicative of bacteremia. Non-typhoidal Salmonella bacteria were detected in the blood and tissue samples of infected rabbits by microbiological culture and real-time PCR assays. The development of this animal model of bacteremia could prove to be a useful tool for studying how non-typhoidal Salmonella infections disseminate and spread in humans. Copyright © 2014 Elsevier Ltd. All rights reserved.

Determination of mercury in fish tissue using a minianalyzer based on cold vapor atomic absorption spectrometry at the 184.9 nm line.

PubMed

Rizea, Maria-Cristina; Bratu, Maria-Cristina; Danet, Andrei Florin; Bratu, Adrian

2007-09-01

A sensitive method was proposed and optimized for the determination of total mercury in fish tissue by using wet digestion, followed by cold vapor atomic absorption spectrometry (CVAAS) at the main resonance line of mercury (184.9 nm). The measurements were made using a new type of a non-dispersive mercury minianalyzer. This instrument was initially designed and built for atmospheric mercury-vapor detection. For determining mercury in aqueous samples, the minianalyzer was linked with a mercury/hydride system, Perkin Elmer Model MHS-10. To check the method, the analyzed samples were spiked with a standard solution of mercury. The recoveries of mercury spiked to wet fish tissue were >90% for 0.5 - 0.8 g samples. The results showed a better sensitivity (about 2.5 times higher) when using the mercury absorption line at 184.9 nm compared with the sensitivity obtained by conventional CVAAS at 253.7 nm.

The first liquid biopsy test approved. Is it a new era of mutation testing for non-small cell lung cancer?

PubMed Central

2017-01-01

Specific mutations in epidermal growth factor receptor (EGFR) gene are predictive for response to the EGFR tyrosine kinase inhibitors (TKIs) in non-small cell lung cancer patients (NSCLC). According to international guidelines, the molecular testing in patients with advanced NSCLC of a non-squamous subtype is recommended. However, obtain a tissue sample could be challenging. Liquid biopsy allows to determine patients suitable for EGFR-targeted therapy by analysis of circulating-free tumor DNA (cfDNA) in peripheral blood samples and might replace tissue biopsy. It allows to acquire a material in convenient minimally invasive manner, is easily repeatable, could be used for molecular identification and molecular changes monitoring. Many studies show a high concordance rate between tissue and plasma samples testing. When U.S. Food and Drug Administration (FDA) approved the first liquid biopsy test, analysis of driver gene mutation from cfDNA becomes a reality in clinical practice for patients with NSCLC. PMID:28251125

Usage of CT data in biomechanical research

NASA Astrophysics Data System (ADS)

Safonov, Roman A.; Golyadkina, Anastasiya A.; Kirillova, Irina V.; Kossovich, Leonid Y.

2017-02-01

Object of study: The investigation is focused on development of personalized medicine. The determination of mechanical properties of bone tissues based on in vivo data was considered. Methods: CT, MRI, natural experiments on versatile test machine Instron 5944, numerical experiments using Python programs. Results: The medical diagnostics methods, which allows determination of mechanical properties of bone tissues based on in vivo data. The series of experiments to define the values of mechanical parameters of bone tissues. For one and the same sample, computed tomography (CT), magnetic resonance imaging (MRI), ultrasonic investigations and mechanical experiments on single-column test machine Instron 5944 were carried out. The computer program for comparison of CT and MRI images was created. The grayscale values in the same points of the samples were determined on both CT and MRI images. The Haunsfield grayscale values were used to determine rigidity (Young module) and tensile strength of the samples. The obtained data was compared to natural experiments results for verification.

The utility of electron microscopy in detecting asbestos fibers and particles in BALF in diffuse lung diseases.

PubMed

Kido, Takashi; Morimoto, Yasuo; Yatera, Kazuhiro; Ishimoto, Hiroshi; Ogoshi, Takaaki; Oda, Keishi; Yamasaki, Kei; Kawanami, Toshinori; Shimajiri, Shohei; Mukae, Hiroshi

2017-04-21

In patients with diffuse lung diseases, differentiating occupational lung diseases from other diseases is clinically important. However, the value of assessing asbestos and particles in bronchoalveolar lavage fluid (BALF) in diffuse lung diseases by electron microscopy (EM) remains unclear. We evaluated the utility of EM in detecting asbestos fibers and particles in patients with diffuse lung diseases. The BALF specimens of 107 patients with diffuse lung diseases were evaluated. First, detection of asbestos by EM and light microscopy (LM) were compared. Second, the detection of asbestos using surgically obtained lung tissues of 8 of 107 patients were compared with the results of EM and LM in BALF. Third, we compared the results of mineralogical components of particles in patients with (n = 48) and without (n = 59) a history of occupational exposure to inorganic dust. BALF asbestos were detected in 11 of 48 patients with a history of occupational exposure by EM; whereas asbestos as asbestos bodies (ABs) were detected in BALF in 4 of these 11 patients by LM. Eight of 107 patients in whom lung tissue samples were surgically obtained, EM detected BALF asbestos at a level of >1,000 fibers/ml in all three patients who had ABs in lung tissue samples by LM at a level of >1,000 fibers/g. The BALF asbestos concentration by EM and in lung tissue by LM were positively correlated. The particle fractions of iron and phosphorus were increased in patients with a history of occupational exposure and both correlated with a history of occupational exposure by a multiple regression analysis. EM using BALF seemed to be superior to LM using BALF and displayed a similar sensitivity to LM using surgically-obtained lung tissue samples in the detection of asbestos. Our results also suggest that detection of elements, such as iron and phosphorus in particles, is useful for evaluating occupational exposure. We conclude that the detection of asbestos and iron and phosphorus in particles in BALF by EM is very useful for the evaluation of occupational exposure.

Multispectral detection of cutaneous lesions using spectroscopy and microscopy approaches

NASA Astrophysics Data System (ADS)

Borisova, E.; Genova-Hristova, Ts.; Troyanova, P.; Pavlova, E.; Terziev, I.; Semyachkina-Glushkovskaya, O.; Lomova, M.; Genina, E.; Stanciu, G.; Tranca, D.; Avramov, L.

2018-02-01

Autofluorescence, diffuse-reflectance and transmission spectral, and microscopic measurements were made on different cutaneous neoplastic lesions, namely basal cell carcinoma, squamous cell carcinoma, malignant melanoma, and dysplastic and benign lesions related. Spectroscopic measurements were made on ex vivo tissue samples, and confocal microscopy investigations were made on thin tissue slices. Fluorescence spectra obtained reveal statistically significant differences between the different benign, dysplastic and malignant lesions by the level of emission intensity, as well by spectral shape, which are fingerprints applicable for differentiation algorithms. In reflectance mode the most significant differences are related to the influence of skin pigments - melanin and hemoglobin. Transmission spectroscopy mode gave complementary optical properties information about the tissue samples investigated to that one of reflectance and absorption spectroscopy. Using autofluorescence detection of skin lesions we obtain very good diagnostic performance for distinguishing of nonmelanoma lesions. Using diffuse reflectance and transmission spectroscopy we obtain significant tool for pigmented pathologies differentiation, but it is a tool with moderate sensitivity for non-melanoma lesions detection. One could rapidly increase the diagnostic accuracy of the received combined "optical biopsy" method when several spectral detection techniques are applied in common algorithm for lesions' differentiation. Specific spectral features observed in each type of lesion investigated on micro and macro level would be presented and discussed. Correlation between the spectral data received and the microscopic features observed would be discussed in the report.

Tissue-type plasminogen activator-induced fibrinolysis is enhanced in patients with breast, lung, pancreas and colon cancer.

PubMed

Nielsen, Vance G; Matika, Ryan W; Ley, Michele L B; Waer, Amy L; Gharagozloo, Farid; Kim, Samuel; Nfonsam, Valentine N; Ong, Evan S; Jie, Tun; Warneke, James A; Steinbrenner, Evangelina B

2014-04-01

Although cancer-mediated changes in hemostatic proteins unquestionably promote hypercoagulation, the effects of neoplasia on fibrinolysis in the circulation are less well defined. The goals of the present investigation were to determine if plasma obtained from patients with breast, lung, pancreas and colon cancer was less or more susceptible to lysis by tissue-type plasminogen activator (tPA) compared to plasma obtained from normal individuals. Archived plasma obtained from patients with breast (n = 18), colon/pancreas (n = 27) or lung (n = 19) was compared to normal individual plasma (n = 30) using a thrombelastographic assay that assessed fibrinolytic vulnerability to exogenously added tPA. Plasma samples were activated with tissue factor/celite, had tPA added, and had data collected until clot lysis occurred. Additional, similar samples had potato carboxypeptidase inhibitor added to assess the role played by thrombin-activatable fibrinolysis inhibitor in cancer-modulated fibrinolysis. Rather than inflicting a hypofibrinolytic state, the three groups of cancers demonstrated increased vulnerability to tPA (e.g. decreased time to lysis, increased speed of lysis, decreased clot lysis time). However, hypercoagulation manifested as increased speed of clot formation and strength compensated for enhanced fibrinolytic vulnerability, resulting in a clot residence time that was not different from normal individual thrombi. In sum, enhanced hypercoagulability associated with cancer was in part diminished by enhanced fibrinolytic vulnerability to tPA.

Method for procuring specific populations of viable human prostate cells for research.

PubMed

Fischer, A H; Philips, A; Taysavang, P; McKenney, J K; Amin, M B

2001-04-01

A wider range of research can be conducted on viable tissue samples than on fixed or frozen samples. A major obstacle to studying viable prostate tissue samples is the inability to accurately identify cancer on direct examination of unembedded tissue. We used a dissecting microscope to identify cancer in unfixed prostate tissue samples stained on the cut surface with 0.5% aqueous toluidine blue. We measured the diagnostic accuracy of this technique in 25 consecutive prostatectomies, determined the viability of procured samples, and estimated the effect on final pathologic assessment. Both surfaces of a 3- to 5-mm thick cross-section taken midway between base and apex of the prostate were examined. A 4-mm punch biopsy was directed to one benign and one malignant area when clearly present. The dissecting microscope allowed clearcut recognition of carcinoma in 17 of the 25 cross-sections, and carcinoma was confirmed in all 17 (100%). In 8 of 25 cases, no procurement was attempted because no carcinoma was evident in the one cross-section studied. Twenty of 25 cross-sections were adequate for benign tissue procurement; five of the cross-sections were not suitable for procurement because of the presence of extensive carcinoma or atrophy. Seventeen of the 20 were accurately diagnosed as benign (85%); one showed pseudohyperplastic adenocarcinoma, one showed focal high-grade prostatic intraepithelial neoplasia, and one showed urothelial carcinoma in situ. Prostatic epithelium obtained with the technique remains viable and can be separated from stroma. The dissecting microscope technique appears to facilitate rather than interfere with accurate pathologic assessment: extraprostatic extension or positive margins were correctly identified during tissue procurement in three cases. The procedure takes only about 30 minutes.

Comparative trace elemental analysis of cancerous and non-cancerous tissues of rectal cancer patients using PIXE

NASA Astrophysics Data System (ADS)

Naga Raju, G. J.; Sarita, P.; Murthy, K. S. R.

2017-08-01

Particle Induced X-ray Emission (PIXE), an accelerator based analytical technique has been employed in this work for the analysis of trace elements in the cancerous and non-cancerous tissues of rectal cancer patients. A beam of 3 MeV protons generated from 3 MV Pelletron accelerator at the Ion Beam Laboratory of Institute of Physics, Bhubaneswar, India was used as projectile to excite the atoms present in the tissues samples. PIXE technique, with its capability to detect simultaneously several elements present at very low concentrations, offers an excellent tool for trace element analysis. The characteristic X-rays emitted by the samples were recorded by a high resolution Si (Li) detector. On the basis of the PIXE spectrum obtained for each sample, the elements Cl, K, Ca, Ti, V, Cr, Mn, Fe, Co, Ni, Cu, Zn, As, and Br were identified and their relative concentrations were estimated in the cancerous and non-cancerous tissues of rectum. The levels of Mn, Fe, Co, Cu, Zn, and As were higher (p < 0.005) while the levels of Ca, Cr and Ni were lower (p < 0.005) in the cancer tissues relative to the normal tissues. The alterations in the levels of the trace elements observed in the present work are discussed in this paper with respect to their potential role in the initiation, promotion and inhibition of cancer of the rectum.

Vibro-acoustography with 1.75D ultrasound array transducer for detection and localization of permanent prostate brachytherapy seeds: ex vivo study

NASA Astrophysics Data System (ADS)

Mehrmohammadi, Mohammad; Alizad, Azra; Kinnick, Randall R.; Davis, Brian J.; Fatemi, Mostafa

2013-03-01

Effective brachytherapy procedures require precise placement of radioactive seeds in the prostate. Currently, transrectal ultrasound (TRUS) imaging is one of the main intraoperative imaging modalities to assist physicians in placement of brachytherapy seeds. However, the seed detection rate with TRUS is poor mainly because ultrasound imaging is highly sensitive to variations in seed orientation. The purpose of this study is to investigate the abilities of a new acoustic radiation force imaging modality, vibro-acoustography (VA), equipped with a 1.75D array transducer and implemented on a customized clinical ultrasound scanner, to image and localize brachytherapy seeds in prostatic tissue. To perform experiments, excised cadaver prostate specimens were implanted with dummy brachytherapy seeds, and embedded in tissue mimicking gel to simulate the properties of the surrounding soft tissues. The samples were scanned using the VA system and the resulting VA signals were used to reconstruct VA images at several depths inside the tissue. To further evaluate the performance of VA in detecting seeds, X-ray computed tomography (CT) images of the same tissue sample, were obtained and used as a gold-standard to compare the number of seeds detected by the two methods. Our results indicate that VA is capable of imaging of brachytherapy seeds with accuracy and high contrast, and can detect a large percentage of the seeds implanted within the tissue samples.

Contrast-enhanced 3D micro-CT of plant tissues using different impregnation techniques.

PubMed

Wang, Zi; Verboven, Pieter; Nicolai, Bart

2017-01-01

X-ray micro-CT has increasingly been used for 3D imaging of plant structures. At the micrometer resolution however, limitations in X-ray contrast often lead to datasets with poor qualitative and quantitative measures, especially within dense cell clusters of plant tissue specimens. The current study developed protocols for delivering a cesium based contrast enhancing solution to varying plant tissue specimens for the purpose of improving 3D tissue structure characterization within plant specimens, accompanied by new image processing workflows to extract the additional data generated by the contrast enhanced scans. Following passive delivery of a 10% cesium iodide contrast solution, significant increases of 85.4 and 38.0% in analyzable cell volumes were observed in pear fruit hypanthium and tomato fruit outer mesocarp samples. A significant increase of 139.6% in the number of analyzable cells was observed in the pear fruit samples along the added ability to locate and isolate better brachysclereids and vasculature in the sample volume. Furthermore, contrast enhancement resulted in significant improvement in the definition of collenchyma and parenchyma in the petiolule of tomato leaflets, from which both qualitative and quantitative data can be extracted with respect to cell measures. However, contrast enhancement was not achieved in leaf vasculature and mesophyll tissue due to fundamental limitations. Active contrast delivery to apple fruit hypanthium samples did yield a small but insignificant increase in analyzable volume and cells, but data on vasculature can now be extracted better in correspondence to the pear hypanthium samples. Contrast delivery thus improved visualization and analysis the most in dense tissue types. The cesium based contrast enhancing protocols and workflows can be utilized to obtain detailed 3D data on the internal microstructure of plant samples, and can be adapted to additional samples of interest with minimal effort. The resulting datasets can therefore be utilized for more accurate downstream studies that requires 3D data.

Brain Slice Staining and Preparation for Three-Dimensional Super-Resolution Microscopy

PubMed Central

German, Christopher L.; Gudheti, Manasa V.; Fleckenstein, Annette E.; Jorgensen, Erik M.

2018-01-01

Localization microscopy techniques – such as photoactivation localization microscopy (PALM), fluorescent PALM (FPALM), ground state depletion (GSD), and stochastic optical reconstruction microscopy (STORM) – provide the highest precision for single molecule localization currently available. However, localization microscopy has been largely limited to cell cultures due to the difficulties that arise in imaging thicker tissue sections. Sample fixation and antibody staining, background fluorescence, fluorophore photoinstability, light scattering in thick sections, and sample movement create significant challenges for imaging intact tissue. We have developed a sample preparation and image acquisition protocol to address these challenges in rat brain slices. The sample preparation combined multiple fixation steps, saponin permeabilization, and tissue clarification. Together, these preserve intracellular structures, promote antibody penetration, reduce background fluorescence and light scattering, and allow acquisition of images deep in a 30 μm thick slice. Image acquisition challenges were resolved by overlaying samples with a permeable agarose pad and custom-built stainless steel imaging adapter, and sealing the imaging chamber. This approach kept slices flat, immobile, bathed in imaging buffer, and prevented buffer oxidation during imaging. Using this protocol, we consistently obtained single molecule localizations of synaptic vesicle and active zone proteins in three-dimensions within individual synaptic terminals of the striatum in rat brain slices. These techniques may be easily adapted to the preparation and imaging of other tissues, substantially broadening the application of super-resolution imaging. PMID:28924666

Investigation of enrofloxacin residues in broiler tissues using ELISA and LC-MS/MS.

PubMed

Panzenhagen, Pedro Henrique N; Aguiar, Waldemir S; Gouvêa, Raquel; de Oliveira, Andréa M G; Barreto, Fabiano; Pereira, Virgínia L A; Aquino, Maria Helena C

2016-01-01

This study investigated the efficiency of an enrofloxacin ELISA test kit to detect the presence of enrofloxacin residues in broiler tissues compared with LC-MS/MS. Broiler tissues from 72 samples consisting of 60 breast muscle, six pools of livers (500 g each) and six pools of kidneys (500 g each) were obtained from six different slaughterhouses. Breast muscle from 10 carcasses and pools of livers and kidneys from approximately 200 carcasses of the same flock were collected from each slaughterhouse. ELISA and HPLC were used to identify and quantify the contamination of the samples with enrofloxacin. A total of 72% of the analysed samples contained enrofloxacin residues detected by the ELISA and 22.2% were detected by LC-MS/MS. The mean values of enrofloxacin contamination found in chicken breast by ELISA and HPLC were 8.63 and 12.25 μg kg(-1), respectively. None of the samples exceeded the maximum limit of 100 μg kg(-1) by both methods set by the European Union as well as the Brazilian Agriculture Ministry. All positive samples for enrofloxacin residues detected by LC-MS/MS were also positive by ELISA. These data confirm the efficiency of the ELISA test, and suggest its use as a screening method for enrofloxacin residues in poultry tissues due to its quick results, low price and ease of applicability.

Biophotonics in diagnosis and modeling of tissue pathologies

NASA Astrophysics Data System (ADS)

Serafetinides, A. A.; Makropoulou, M.; Drakaki, E.

2008-12-01

Biophotonics techniques are applied to several fields in medicine and biology. The laser based techniques, such as the laser induced fluorescence (LIF) spectroscopy and the optical coherence tomography (OCT), are of particular importance in dermatology, where the laser radiation could be directly applied to the tissue target (e.g. skin). In addition, OCT resolves architectural tissue properties that might be useful as tumour discrimination parameters for skin as well as for ocular non-invasive visualization. Skin and ocular tissues are complex multilayered and inhomogeneous organs with spatially varying optical properties. This fact complicates the quantitative analysis of the fluorescence and/or light scattering spectra, even from the same tissue sample. To overcome this problem, mathematical simulation is applied for the investigation of the human tissue optical properties, in the visible/infrared range of the spectrum, resulting in a better discrimination of several tissue pathologies. In this work, we present i) a general view on biophotonics applications in diagnosis of human diseases, ii) some specific results on laser spectroscopy techniques, as LIF measurements, applied in arterial and skin pathologies and iii) some experimental and theoretical results on ocular OCT measurements. Regarding the LIF spectroscopy, we examined the autofluorescence properties of several human skin samples, excised from humans undergoing biopsy examination. A nitrogen laser was used as an excitation source, emitting at 337 nm (ultraviolet excitation). Histopathology examination of the samples was also performed, after the laser spectroscopy measurements and the results from the spectroscopic and medical analysis were compared, to differentiate malignancies, e.g. basal cell carcinoma tissue (BCC), from normal skin tissue. Regarding the OCT technique, we correlated human data, obtained from patients undergoing OCT examination, with Monte Carlo simulated cornea and retina tissues for diagnosis of ocular diseases.

Comparative evaluation of human pulp tissue dissolution by different concentrations of chlorine dioxide, calcium hypochlorite and sodium hypochlorite: An in vitro study

PubMed Central

Taneja, Sonali; Mishra, Neha; Malik, Shubhra

2014-01-01

Introduction: Irrigation plays an indispensable role in removal of tissue remnants and debris from the complicated root canal system. This study compared the human pulp tissue dissolution by different concentrations of chlorine dioxide, calcium hypochlorite and sodium hypochlorite. Materials and Methods: Pulp tissue was standardized to a weight of 9 mg for each sample. In all,60 samples obtained were divided into 6 groups according to the irrigating solution used- 2.5% sodium hypochlorite (NaOCl), 5.25% NaOCl, 5% calcium hypochlorite (Ca(OCl)2), 10% Ca(OCl)2, 5%chlorine dioxide (ClO2) and 13% ClO2. Pulp tissue was placed in each test tube carrying irrigants of measured volume (5ml) according to their specified subgroup time interval: 30 minutes (Subgroup A) and 60 minutes (Subgroup B). The solution from each sample test tube was filtered and was left for drying overnight. The residual weight was calculated by filtration method. Results: Mean tissue dissolution increases with increase in time period. Results showed 5.25% NaOCl to be most effective at both time intervals followed by 2.5% NaOCl at 60 minutes, 10%Ca(OCl)2 and 13% ClO2 at 60 minutes. Least amount of tissue dissolving ability was demonstrated by 5% Ca(OCl)2 and 5% ClO2 at 30 minutes. Distilled water showed no pulp tissue dissolution. Conclusion: Withinthe limitations of the study, NaOCl most efficiently dissolved the pulp tissue at both concentrations and at both time intervals. Mean tissue dissolution by Ca(OCl)2 and ClO2 gradually increased with time and with their increase in concentration. PMID:25506141

Composite technique for regional neurochemical studies: measurement of energy and neurotransmitter metabolites in single tissue sample.

PubMed

Djuricic, B M; Ueki, Y; Spatz, M

1985-06-01

A combined method is described for the determination of various metabolites from a single tissue sample of the brain. It comprises a quick inactivation of cerebral enzymes by microwave irradiation, easy separation of the desired brain regions, and perchloric acid extraction of tissue substances, which are assayed either by specific enzymatic techniques or by HPLC with electrochemical detection. The obtained values of most energy and neurotransmitter metabolites in the brain are in agreement with those reported using other methods. However, this technique, in contrast to the brain freezing in vitro or freeze-blowing, provides a more efficient procedure for rapid arrest of cerebral metabolism even in the deep brain structures and is therefore suitable for detection of early changes particularly those occurring in experimental pathological conditions such as ischemia.

Imaging Fibrosis and Separating Collagens using Second Harmonic Generation and Phasor Approach to Fluorescence Lifetime Imaging

PubMed Central

Ranjit, Suman; Dvornikov, Alexander; Stakic, Milka; Hong, Suk-Hyun; Levi, Moshe; Evans, Ronald M.; Gratton, Enrico

2015-01-01

In this paper we have used second harmonic generation (SHG) and phasor approach to auto fluorescence lifetime imaging (FLIM) to obtain fingerprints of different collagens and then used these fingerprints to observe bone marrow fibrosis in the mouse femur. This is a label free approach towards fast automatable detection of fibrosis in tissue samples. FLIM has previously been used as a method of contrast in different tissues and in this paper phasor approach to FLIM is used to separate collagen I from collagen III, the markers of fibrosis, the largest groups of disorders that are often without any effective therapy. Often characterized by an increase in collagen content of the corresponding tissue, the samples are usually visualized by histochemical staining, which is pathologist dependent and cannot be automated. PMID:26293987

Biochemical surface modification of Co-Cr-Mo.

PubMed

Puleo, D A

1996-01-01

Because of the limited mechanical properties of tissue substitutes formed by culturing cells on polymeric scaffolds, other approaches to tissue engineering must be explored for applications that require complete and immediate ability to bear weight, e.g. total joint replacements. Biochemical surface modification offers a way to partially regulate events at the bone-implant interface to obtain preferred tissue responses. Tresyl chloride, gamma-aminopropyltriethoxysilane (APS) and p-nitrophenyl chloroformate (p-NPC) immobilization schemes were used to couple a model enzyme, trypsin, on bulk samples of Co-Cr-Mo. For comparison, samples were simply adsorbed with protein. The three derivatization schemes resulted in different patterns and levels of activity. Tresyl chloride was not effective in immobilizing active enzyme on Co-Cr-Mo. Aqueous silanization with 12.5% APS resulted in optimal immobilized activity. Activity on samples derivatized with 0.65 mg p-NPC cm-2 was four to five times greater than that on samples simple adsorbed with enzyme or optimally derivatized with APS and was about eight times that on tresylated samples. This work demonstrates that, although different methods have different effectiveness, chemical derivatization can be used to alter the amount and/or stability of biomolecules immobilized on the surface of Co-Cr-Mo.

Testing the use of juvenile Salmo trutta L. as biomonitors of heavy metal pollution in freshwater.

PubMed

Lamas, S; Fernández, J A; Aboal, J R; Carballeira, A

2007-02-01

Individual specimens of Salmo trutta were captured, from four sampling sites in Galician rivers (NW Spain) affected by different types of contamination: diffuse urban waste, run-off from an unrestored dump at a copper mine and waste from a fish farm. The ages of the captured trouts were established and only those belonging to the 1+ age class were selected for study. The liver and kidney were removed from each fish and analysed to determine the tissue concentrations of Cu, Fe and Zn. The results obtained showed that: (i) the use of 1+ individuals allowed differentiation of contamination scenarios on the basis of the tissue concentrations of metal; (ii) the use of 1+ individuals allowed standardization of the time of exposure, which was sufficiently long for differential uptake to have taken place; (iii) liver tissue provided the best results as, less effort was required than for processing kidney tissue, and significant differences between sampling sites were detected because the intrapopulational variability in metal levels was lower than for kidney, and (iv) the levels of elements detected were not affected by basal tissue concentrations or residual concentrations due to past contamination, which older trouts may have been exposed to. In addition, the use of 1+ trout may provide better results in annual environmental sampling surveys.

Characterization of the Optical Properties of Turbid Media by Supervised Learning of Scattering Patterns.

PubMed

Hassaninia, Iman; Bostanabad, Ramin; Chen, Wei; Mohseni, Hooman

2017-11-10

Fabricated tissue phantoms are instrumental in optical in-vitro investigations concerning cancer diagnosis, therapeutic applications, and drug efficacy tests. We present a simple non-invasive computational technique that, when coupled with experiments, has the potential for characterization of a wide range of biological tissues. The fundamental idea of our approach is to find a supervised learner that links the scattering pattern of a turbid sample to its thickness and scattering parameters. Once found, this supervised learner is employed in an inverse optimization problem for estimating the scattering parameters of a sample given its thickness and scattering pattern. Multi-response Gaussian processes are used for the supervised learning task and a simple setup is introduced to obtain the scattering pattern of a tissue sample. To increase the predictive power of the supervised learner, the scattering patterns are filtered, enriched by a regressor, and finally characterized with two parameters, namely, transmitted power and scaled Gaussian width. We computationally illustrate that our approach achieves errors of roughly 5% in predicting the scattering properties of many biological tissues. Our method has the potential to facilitate the characterization of tissues and fabrication of phantoms used for diagnostic and therapeutic purposes over a wide range of optical spectrum.

Light-induced autofluorescence of animal skin used in tissue optical modeling

NASA Astrophysics Data System (ADS)

Borisova, E.; Bliznakova, I.; Troyanova, P.; Avramov, L.

2007-07-01

Light-induced autofluorescence spectroscopy provides many possibilities for medical diagnostics needs for differentiation of tissue pathologies including cancer. For the needs of clinical practice scientists collect spectral data from patients in vivo or they study different tumor models to obtain objective information for fluorescent properties of every kind of normal and diseased tissue. Therefore it is very important to find the most appropriate and close to the human skin samples from the point of view of laser-induced fluorescence spectroscopy, which will give the possibility for easier transfer of data obtained in animal models to spectroscopic medical diagnostics in humans. In this study are presented some results for in vitro detection of the autofluorescence signals of the animal skin (pig and chicken) with using of LEDs as excitation sources (maximum emission at 365, 375, 385 and 400 nm). The autofluorescence signals from in vivo human skin were also detected for comparison with the models' results. Specific features of the spectra measured are discussed and there are proposed some of the origins of the fluorescence signals obtained. Fluorescence maxima detected are addressed to the typical fluorophores existing in the cutaneous tissues. Influence of main skin absorbers, namely melanin and hemoglobin, is also discussed.

Laser capture microdissection to study flower morphogenesis

NASA Astrophysics Data System (ADS)

Pawełkowicz, Magdalena Ewa; Skarzyńska, Agnieszka; Kowalczuk, Cezary; PlÄ der, Wojciech; Przybecki, Zbigniew

2017-08-01

Laser Capture Microdissection (LCM) is a sample preparation microscopic method that enables isolation of an interesting cell or cells population from human, animal or plant tissue. This technique allows for obtaining pure sample from heterogeneous mixture. From isolated cells, it is possible to obtain the appropriate quality material used for genomic research in transcriptomics, proteomics and metabolomics. We used LCM method to study flower morphogenesis and specific bud's organ organization and development. The genes expression level in developing flower buds of male (B10) and female (2gg) lines were analyzed with qPCR. The expression was checked for stamen and carpel primordia obtained with LCM and for whole flower buds at successive stages of growth.

A fast and mild decellularization protocol for obtaining extracellular matrix.

PubMed

Mirzarafie, Ariana; Grainger, Rhian K; Thomas, Ben; Bains, William; Ustok, Fatma I; Lowe, Chris R

2014-04-01

Degradation of extracellular matrix (ECM) function with age is a major cause of loss of tissue function with age that we would wish to reverse. Tissue engineering to provide replacement tissue requires an ECM-mimicking scaffold for cell organization. The standard protocols for achieving this take 10 days and include steps that may change the protein structure of the ECM. Here we describe a much shorter protocol for decellularizing chicken muscle, skin, and tendon samples that achieves the same efficiency as the original protocol without protein cross-link interference. Our protocol can be completed in 72 hr.

Optical pathology of human brain metastasis of lung cancer using combined resonance Raman and spatial frequency spectroscopies

NASA Astrophysics Data System (ADS)

Zhou, Yan; Liu, Cheng-hui; Pu, Yang; Cheng, Gangge; Zhou, Lixin; Chen, Jun; Zhu, Ke; Alfano, Robert R.

2016-03-01

Raman spectroscopy has become widely used for diagnostic purpose of breast, lung and brain cancers. This report introduced a new approach based on spatial frequency spectra analysis of the underlying tissue structure at different stages of brain tumor. Combined spatial frequency spectroscopy (SFS), Resonance Raman (RR) spectroscopic method is used to discriminate human brain metastasis of lung cancer from normal tissues for the first time. A total number of thirty-one label-free micrographic images of normal and metastatic brain cancer tissues obtained from a confocal micro- Raman spectroscopic system synchronously with examined RR spectra of the corresponding samples were collected from the identical site of tissue. The difference of the randomness of tissue structures between the micrograph images of metastatic brain tumor tissues and normal tissues can be recognized by analyzing spatial frequency. By fitting the distribution of the spatial frequency spectra of human brain tissues as a Gaussian function, the standard deviation, σ, can be obtained, which was used to generate a criterion to differentiate human brain cancerous tissues from the normal ones using Support Vector Machine (SVM) classifier. This SFS-SVM analysis on micrograph images presents good results with sensitivity (85%), specificity (75%) in comparison with gold standard reports of pathology and immunology. The dual-modal advantages of SFS combined with RR spectroscopy method may open a new way in the neuropathology applications.

In vivo classification of human skin burns using machine learning and quantitative features captured by optical coherence tomography

NASA Astrophysics Data System (ADS)

Singla, Neeru; Srivastava, Vishal; Singh Mehta, Dalip

2018-02-01

We report the first fully automated detection of human skin burn injuries in vivo, with the goal of automatic surgical margin assessment based on optical coherence tomography (OCT) images. Our proposed automated procedure entails building a machine-learning-based classifier by extracting quantitative features from normal and burn tissue images recorded by OCT. In this study, 56 samples (28 normal, 28 burned) were imaged by OCT and eight features were extracted. A linear model classifier was trained using 34 samples and 22 samples were used to test the model. Sensitivity of 91.6% and specificity of 90% were obtained. Our results demonstrate the capability of a computer-aided technique for accurately and automatically identifying burn tissue resection margins during surgical treatment.

Multivariate classification of the infrared spectra of cell and tissue samples

DOE Office of Scientific and Technical Information (OSTI.GOV)

Haaland, D.M.; Jones, H.D.; Thomas, E.V.

1997-03-01

Infrared microspectroscopy of biopsied canine lymph cells and tissue was performed to investigate the possibility of using IR spectra coupled with multivariate classification methods to classify the samples as normal, hyperplastic, or neoplastic (malignant). IR spectra were obtained in transmission mode through BaF{sub 2} windows and in reflection mode from samples prepared on gold-coated microscope slides. Cytology and histopathology samples were prepared by a variety of methods to identify the optimal methods of sample preparation. Cytospinning procedures that yielded a monolayer of cells on the BaF{sub 2} windows produced a limited set of IR transmission spectra. These transmission spectra weremore » converted to absorbance and formed the basis for a classification rule that yielded 100{percent} correct classification in a cross-validated context. Classifications of normal, hyperplastic, and neoplastic cell sample spectra were achieved by using both partial least-squares (PLS) and principal component regression (PCR) classification methods. Linear discriminant analysis applied to principal components obtained from the spectral data yielded a small number of misclassifications. PLS weight loading vectors yield valuable qualitative insight into the molecular changes that are responsible for the success of the infrared classification. These successful classification results show promise for assisting pathologists in the diagnosis of cell types and offer future potential for {ital in vivo} IR detection of some types of cancer. {copyright} {ital 1997} {ital Society for Applied Spectroscopy}« less

Intra-operative on-line discrimination of kidney cancer from normal tissue by IR ATR spectroscopy of extracellular fluid

NASA Astrophysics Data System (ADS)

Urboniene, V.; Velicka, M.; Ceponkus, J.; Pucetaite, M.; Jankevicius, F.; Sablinskas, V.; Steiner, G.

2016-03-01

Determination of cancerous and normal kidney tissues during partial, simple or radical nephrectomy surgery was performed by using differences in the IR absorption spectra of extracellular fluid taken from the corresponding tissue areas. The samples were prepared by stamping of the kidney tissue on ATR diamond crystal. The spectral measurements were performed directly in the OR during surgery for 58 patients. It was found that intensities of characteristic spectral bands of glycogen (880-1200 cm-1) in extracellular fluid are sensitive to the type of the tissue and can be used as spectral markers of tumours. Characteristic spectral band of lactic acid (1730 cm-1) - product of the anaerobic glycolysis, taking place in the cancer cells is not suitable for use as a spectral marker of cancerous tissue, since it overlaps with the band of carbonyl stretch in phospholipids and fatty acids. Results of hierarchical cluster analysis of the spectra show that the spectra of healthy and tumour tissue films can be reliably separated into two groups. On the other hand, possibility to differentiate between tumours of different types and grades remains in question. While the fluid from highly malignant G3 tumour tissue contains highly pronounced glycogen spectral bands and can be well separated from benign and G1 tumours by principal component analysis, the variations between spectra from sample to sample prevent from obtaining conclusive results about the grouping between different tumour types and grades. The proposed method is instant and can be used in situ and even in vivo.

Quantifying the ultrastructure of carotid arteries using high-resolution micro-diffusion tensor imaging—comparison of intact versus open cut tissue

NASA Astrophysics Data System (ADS)

Salman Shahid, Syed; Gaul, Robert T.; Kerskens, Christian; Flamini, Vittoria; Lally, Caitríona

2017-12-01

Diffusion magnetic resonance imaging (dMRI) can provide insights into the microstructure of intact arterial tissue. The current study employed high magnetic field MRI to obtain ultra-high resolution dMRI at an isotropic voxel resolution of 117 µm3 in less than 2 h of scan time. A parameter selective single shell (128 directions) diffusion-encoding scheme based on Stejskel-Tanner sequence with echo-planar imaging (EPI) readout was used. EPI segmentation was used to reduce the echo time (TE) and to minimise the susceptibility-induced artefacts. The study utilised the dMRI analysis with diffusion tensor imaging (DTI) framework to investigate structural heterogeneity in intact arterial tissue and to quantify variations in tissue composition when the tissue is cut open and flattened. For intact arterial samples, the region of interest base comparison showed significant differences in fractional anisotropy and mean diffusivity across the media layer (p  <  0.05). For open cut flat samples, DTI based directionally invariant indices did not show significant differences across the media layer. For intact samples, fibre tractography based indices such as calculated helical angle and fibre dispersion showed near circumferential alignment and a high degree of fibre dispersion, respectively. This study demonstrates the feasibility of fast dMRI acquisition with ultra-high spatial and angular resolution at 7 T. Using the optimised sequence parameters, this study shows that DTI based markers are sensitive to local structural changes in intact arterial tissue samples and these markers may have clinical relevance in the diagnosis of atherosclerosis and aneurysm.

Antigen recovery and preservation using the microwave irradiation of biological samples for transmission electron microscopy analysis.

PubMed

Aïoun, Josiane; Chat, Sophie; Bordat, Christian; Péchoux, Christine

2013-01-01

Most studies using microwave irradiation (MWI) for the preparation of tissue samples have reported an improvement in structural integrity. However, there have been few studies on the effect of microwave (MW) on antigen preservation during sample preparation prior to immunolocalization. This report documents our experience of specimen preparation using an automatic microwave apparatus to obtain antigen preservation and retrieval. We tested the effects of MW processing vs. conventional procedures on the morphology and antigenicity of two different tissues: the brain and mammary gland, whose chemical composition and anatomical organization are quite different. We chose to locate the transcription factor PPARβ/δ using immunocytochemistry on brain tissue sections from hamsters. Antigen retrieval protocols involving MWI were used to restore immunoreactivity. We also studied the efficiency of the ultrastructural immunolocalization of both PPARγ and caveolin-1 following MWI vs. conventional treatment, on mammary gland tissue from mice at 10 days of lactation. Our findings showed that the treatment of tissue samples with MWI, in the context of a process lasting just a few hours from fixation to immunolocalization, enabled similar, or even better, results than conventional protocols. The quantification of immunolabeling for cav-1 indicated an increase in density of up to three-fold in tissues processed in the microwave oven. Furthermore, MW treatment permitted the localization of PPARβ/δ in glutaraldehyde-fixed specimens, which was impossible in the absence of MWI. This study thus showed that techniques involving the use of microwaves could largely improve both ultrastructure and immunodetection. Copyright © 2013 Elsevier Ltd. All rights reserved.

Silver halide fiber optic radiometry for temperature monitoring and control of tissues heated by microwave

NASA Astrophysics Data System (ADS)

Shenfeld, Ofer; Belotserkovsky, Edward; Goldwasser, Benad; Zur, Albert; Katzir, Abraham

1993-02-01

The heating of tissue by microwave radiation has attained a place of importance in various medical fields, such as the treatment of malignancies, urinary retention, and hypothermia. Accurate temperature measurements in these treated tissues is important for treatment planning and for the control of the heating process. It is also important to be able to measure spacial temperature distribution in the tissues because they are heated in a nonuniform way by the microwave radiation. Conventional temperature sensors used today are inaccurate in the presence of microwave radiation and require contact with the heated tissue. Fiber optic radiometry makes it possible to measure temperatures accurately in the presence of microwave radiation and does not require contact with the tissue. Accurate temperature measurements of tissues heated by microwave was obtained using a silver halide optic radiometer, enabling control of the heating process in other regions of the tissue samples. Temperature mappings of the heated tissues were performed and the nonuniform temperature distributions in these tissues was demonstrated.

Interventional Vitamin C A Strategy for Attenuation of Coagulopathy and Inflammation in Hemorrhagic Trauma and Shock

DTIC Science & Technology

2016-10-01

evident in liver and kidney sections. Treatment with VitC (200mg/kg) was associated with a lower degree of histological tissue injury and a...significantly reduced ALI score. Treatment with VitC also reduced the expression of pro-inflammatory mediators in lungs, liver and kidneys . In addition, VitC...Euthasol IV  Tissue samples (lung, liver and kidney ) obtained for histology and molecular characterization Definitive trial conducted: Eight (8

Plasma and white adipose tissue lipid composition in marmots.

PubMed

Florant, G L; Nuttle, L C; Mullinex, D E; Rintoul, D A

1990-05-01

White adipose tissue biopsies and plasma samples were obtained from hibernating yellow-bellied marmots (Marmota flaviventris) maintained in the laboratory. In addition, biopsies and plasma samples were obtained from normothermic animals in the field and laboratory. Measurement of plasma free fatty acid (FA) levels indicated that winter laboratory animals exhibited increased lipolysis. Additionally, analysis of white adipose tissue triacylglycerol revealed that the FA composition of the storage fat in animals maintained on the standard laboratory diet is remarkably simple and uniform between different adipose depots in the same animal. Three FAs (palmitic, oleic, and linoleic acids) made up greater than 95% of the total. Triene (alpha-linolenate) was found in newly captured animals, but the percentage of this FA decreased rapidly when the animals were maintained on the standard laboratory diet. Throughout the hibernation season (October to April), white adipose tissue-saturated FA percentage decreased, monoene percentage remained constant, and diene percentage increased. Analysis of plasma FA composition suggested that these animals tended to metabolize saturated FAs from stored lipid during hibernation and that dienes were mobilized briefly after the last arousal from hibernation in spring. From these observations, we hypothesize that marmots preferentially metabolize saturated fats during the hibernation period and that essential FAs of the omega 6 series tend to be metabolized more slowly than other FAs. These characteristics suggest that marmots are a valuable animal model in which to study lipid metabolism.

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.

Identical mitochondrial somatic mutations unique to chronic periodontitis and coronary artery disease

PubMed Central

Pallavi, Tokala; Chandra, Rampalli Viswa; Reddy, Aileni Amarender; Reddy, Bavigadda Harish; Naveen, Anumala

2016-01-01

Context: The inflammatory processes involved in chronic periodontitis and coronary artery diseases (CADs) are similar and produce reactive oxygen species that may result in similar somatic mutations in mitochondrial deoxyribonucleic acid (mtDNA). Aims: The aims of the present study were to identify somatic mtDNA mutations in periodontal and cardiac tissues from subjects undergoing coronary artery bypass surgery and determine what fraction was identical and unique to these tissues. Settings and Design: The study population consisted of 30 chronic periodontitis subjects who underwent coronary artery surgery after an angiogram had indicated CAD. Materials and Methods: Gingival tissue samples were taken from the site with deepest probing depth; coronary artery tissue samples were taken during the coronary artery bypass grafting procedures, and blood samples were drawn during this surgical procedure. These samples were stored under aseptic conditions and later transported for mtDNA analysis. Statistical Analysis Used: Complete mtDNA sequences were obtained and aligned with the revised Cambridge reference sequence (NC_012920) using sequence analysis and auto assembler tools. Results: Among the complete mtDNA sequences, a total of 162 variations were spread across the whole mitochondrial genome and present only in the coronary artery and the gingival tissue samples but not in the blood samples. Among the 162 variations, 12 were novel and four of the 12 novel variations were found in mitochondrial NADH dehydrogenase subunit 5 complex I gene (33.3%). Conclusions: Analysis of mtDNA mutations indicated 162 variants unique to periodontitis and CAD. Of these, 12 were novel and may have resulted from destructive oxidative forces common to these two diseases. PMID:27041832

Implication of Gastric Cancer Molecular Genetic Markers in Surgical Practice.

PubMed

Nemtsova, Marina V; Strelnikov, Vladimir V; Tanas, Alexander S; Bykov, Igor I; Zaletaev, Dmitry V; Rudenko, Viktoria V; Glukhov, Alexander I; Kchorobrich, Tatiana V; Li, Yi; Tarasov, Vadim V; Barreto, George E; Aliev, Gjumrakch

2017-10-01

We have investigated aberrant methylation of genes CDH1, RASSF1A, MLH1, N33, DAPK, expression of genes hTERT, MMP7, MMP9, BIRC5 (survivin), PTGS2, and activity of telomerase of 106 gastric tumor samples obtained intra-operatively and 53 gastric tumor samples from the same group of patients obtained endoscopically before surgery. Biopsy specimens obtained from 50 patients with chronic calculous cholecystitis were used as a control group. Together with tissue samples obtained from different sites remote to tumors, a total of 727 samples have been studied. The selected parameters comprise a system of molecular markers that can be used in both diagnostics of gastric cancer and in dynamic monitoring of patients after surgery. Special attention was paid to the use of molecular markers for the diagnostics of malignant process in the material obtained endoscopically since the efficacy of morphological diagnostics in biopsies is compromised by intratumoral heterogeneity, which may prevent reliable identification of tumor cells in the sampling. Our data indicated that certain molecular genetic events provided more sensitive yet specific markers of the tumor. We demonstrated that molecular profiles detected in preoperative biopsies were confirmed by the material obtained intra-operatively. The use of endoscopic material facilitates gastric tumors pre-operative diagnostics, improving early detection of gastric cancer and potential effective treatment strategies.

Electromagnetic Spectroscopy of Normal Breast Tissue Specimens Obtained From Reduction Surgeries: Comparison of Optical and Microwave Properties

PubMed Central

Lazebnik, Mariya; Zhu, Changfang; Palmer, Gregory M.; Harter, Josephine; Sewall, Sarah; Ramanujam, Nirmala; Hagness, Susan C.

2009-01-01

Techniques utilizing electromagnetic energy at microwave and optical frequencies have been shown to be promising for breast cancer detection and diagnosis. Since different biophysical mechanisms are exploited at these frequencies to discriminate between healthy and diseased tissue, combining these two modalities may result in a more powerful approach for breast cancer detection and diagnosis. Toward this end, we performed microwave dielectric spectroscopy and optical diffuse reflectance spectroscopy measurements at the same sites on freshly-excised normal breast tissues obtained from reduction surgeries at the University of Wisconsin Hospital, using microwave and optical probes with very similar sensing volumes. We found that the microwave dielectric constant and effective conductivity are correlated with tissue composition across the entire measurement frequency range (|r|~0.5–0.6, p<0.01), and that the optical absorption coefficient at 460 nm and optical scattering coefficient are correlated with tissue composition (|r|~ 0.4–0.6, p<0.02). Finally, we found that the optical absorption coefficient at 460 nm is correlated with the microwave dielectric constant and effective conductivity (r=−0.55, p<0.01). Our results suggest that combining optical and microwave modalities for analyzing breast tissue samples may serve as a crosscheck and provide complementary information about tissue composition. PMID:18838370

Electromagnetic spectroscopy of normal breast tissue specimens obtained from reduction surgeries: comparison of optical and microwave properties.

PubMed

Lazebnik, Mariya; Zhu, Changfang; Palmer, Gregory M; Harter, Josephine; Sewall, Sarah; Ramanujam, Nirmala; Hagness, Susan C

2008-10-01

Techniques utilizing electromagnetic energy at microwave and optical frequencies have been shown to be promising for breast cancer detection and diagnosis. Since different biophysical mechanisms are exploited at these frequencies to discriminate between healthy and diseased tissue, combining these two modalities may result in a more powerful approach for breast cancer detection and diagnosis. Toward this end, we performed microwave dielectric spectroscopy and optical diffuse reflectance spectroscopy measurements at the same sites on freshly excised normal breast tissues obtained from reduction surgeries at the University of Wisconsin Hospital, using microwave and optical probes with very similar sensing volumes. We found that the microwave dielectric constant and effective conductivity are correlated with tissue composition across the entire measurement frequency range (|r| approximately 0.5-0.6, p<0.01) and that the optical absorption coefficient at 460 nm and optical scattering coefficient are correlated with tissue composition (|r| approximately 0.4-0.6, p<0.02). Finally, we found that the optical absorption coefficient at 460 nm is correlated with the microwave dielectric constant and effective conductivity (r=-0.55, p<0.01). Our results suggest that combining optical and microwave modalities for analyzing breast tissue samples may serve as a crosscheck and provide complementary information about tissue composition.

Imaging using a supercontinuum laser to assess tumors in patients with breast carcinoma

NASA Astrophysics Data System (ADS)

Sordillo, Laura A.; Sordillo, Peter P.; Alfano, R. R.

2016-03-01

The supercontinuum laser light source has many advantages over other light sources, including broad spectral range. Transmission images of paired normal and malignant breast tissue samples from two patients were obtained using a Leukos supercontinuum (SC) laser light source with wavelengths in the second and third NIR optical windows and an IR- CCD InGaAs camera detector (Goodrich Sensors Inc. high response camera SU320KTSW-1.7RT with spectral response between 900 nm and 1,700 nm). Optical attenuation measurements at the four NIR optical windows were obtained from the samples.

[Induced pluripotent stem cells: a new paradigm to study human tissues].

PubMed

Sansac, Caroline; Assou, Said; Bouckenheimer, Julien; Lemaître, Jean-Marc; De Vos, John

2016-01-01

Induced pluripotent stem cells (iPSCs) are obtained by reprogramming differentiated cells through forced expression of four embryonic transcription factors. The discovery of this technology, able to transform a differentiated cell into a pluripotent cell, has profoundly shifted the paradigm of the concept of cell identity, since it is now possible to obtain in vitro any cell type from an initial sample of skin or blood cells from a healthy volunteer or patient. Applications of iPSCs are exceedingly large, and comprise the in vitro modeling of normal or pathological tissues, including for massive drug screening. They also open new therapeutic avenues in the field of regenerative medicine. © Société de Biologie, 2016.

Liquid Microjunction Surface Sampling Coupled with High-Pressure Liquid Chromatography-Electrospray Ionization-Mass Spectrometry for Analysis of Drugs and Metabolites in Whole-Body Thin Tissue Sections

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kertesz, Vilmos; Van Berkel, Gary J

2010-01-01

In this work, a commercially available autosampler was adapted to perform direct liquid microjunction (LMJ) surface sampling followed by a high-pressure liquid chromatography (HPLC) separation of the extract components and detection with electrospray ionization mass spectrometry (ESI-MS). To illustrate the utility of coupling a separation with this direct liquid extraction based surface sampling approach, four different organs (brain, lung, kidney, and liver) from whole-body thin tissue sections of propranolol dosed and control mice were examined. The parent drug was observed in the chromatograms of the surface sampling extracts from all the organs of the dosed mouse examined. In addition, twomore » isomeric phase II metabolites of propranolol (an aliphatic and an aromatic hydroxypropranolol glucuronide) were observed in the chromatograms of the extracts from lung, kidney, and liver. Confirming the presence of one or the other or both of these glucuronides in the extract from the various organs was not possible without the separation. These drug and metabolite data obtained using the LMJ surface sampling/HPLC-MS method and the results achieved by analyzing similar samples by conventional extraction of the tissues and subsequent HPLC-MS analysis were consistent.« less

LIBS analysis of artificial calcified tissues matrices.

PubMed

Kasem, M A; Gonzalez, J J; Russo, R E; Harith, M A

2013-04-15

In most laser-based analytical methods, the reproducibility of quantitative measurements strongly depends on maintaining uniform and stable experimental conditions. For LIBS analysis this means that for accurate estimation of elemental concentration, using the calibration curves obtained from reference samples, the plasma parameters have to be kept as constant as possible. In addition, calcified tissues such as bone are normally less "tough" in their texture than many samples, especially metals. Thus, the ablation process could change the sample morphological features rapidly, and result in poor reproducibility statistics. In the present work, three artificial reference sample sets have been fabricated. These samples represent three different calcium based matrices, CaCO3 matrix, bone ash matrix and Ca hydroxyapatite matrix. A comparative study of UV (266 nm) and IR (1064 nm) LIBS for these three sets of samples has been performed under similar experimental conditions for the two systems (laser energy, spot size, repetition rate, irradiance, etc.) to examine the wavelength effect. The analytical results demonstrated that UV-LIBS has improved reproducibility, precision, stable plasma conditions, better linear fitting, and the reduction of matrix effects. Bone ash could be used as a suitable standard reference material for calcified tissue calibration using LIBS with a 266 nm excitation wavelength. Copyright © 2013 Elsevier B.V. All rights reserved.

Assessment of Sample Preparation Bias in Mass Spectrometry-Based Proteomics.

PubMed

Klont, Frank; Bras, Linda; Wolters, Justina C; Ongay, Sara; Bischoff, Rainer; Halmos, Gyorgy B; Horvatovich, Péter

2018-04-17

For mass spectrometry-based proteomics, the selected sample preparation strategy is a key determinant for information that will be obtained. However, the corresponding selection is often not based on a fit-for-purpose evaluation. Here we report a comparison of in-gel (IGD), in-solution (ISD), on-filter (OFD), and on-pellet digestion (OPD) workflows on the basis of targeted (QconCAT-multiple reaction monitoring (MRM) method for mitochondrial proteins) and discovery proteomics (data-dependent acquisition, DDA) analyses using three different human head and neck tissues (i.e., nasal polyps, parotid gland, and palatine tonsils). Our study reveals differences between the sample preparation methods, for example, with respect to protein and peptide losses, quantification variability, protocol-induced methionine oxidation, and asparagine/glutamine deamidation as well as identification of cysteine-containing peptides. However, none of the methods performed best for all types of tissues, which argues against the existence of a universal sample preparation method for proteome analysis.

High quality copy number and genotype data from FFPE samples using Molecular Inversion Probe (MIP) microarrays

PubMed Central

Wang, Yuker; Carlton, Victoria EH; Karlin-Neumann, George; Sapolsky, Ronald; Zhang, Li; Moorhead, Martin; Wang, Zhigang C; Richardson, Andrea L; Warren, Robert; Walther, Axel; Bondy, Melissa; Sahin, Aysegul; Krahe, Ralf; Tuna, Musaffe; Thompson, Patricia A; Spellman, Paul T; Gray, Joe W; Mills, Gordon B; Faham, Malek

2009-01-01

Background A major challenge facing DNA copy number (CN) studies of tumors is that most banked samples with extensive clinical follow-up information are Formalin-Fixed Paraffin Embedded (FFPE). DNA from FFPE samples generally underperforms or suffers high failure rates compared to fresh frozen samples because of DNA degradation and cross-linking during FFPE fixation and processing. As FFPE protocols may vary widely between labs and samples may be stored for decades at room temperature, an ideal FFPE CN technology should work on diverse sample sets. Molecular Inversion Probe (MIP) technology has been applied successfully to obtain high quality CN and genotype data from cell line and frozen tumor DNA. Since the MIP probes require only a small (~40 bp) target binding site, we reasoned they may be well suited to assess degraded FFPE DNA. We assessed CN with a MIP panel of 50,000 markers in 93 FFPE tumor samples from 7 diverse collections. For 38 FFPE samples from three collections we were also able to asses CN in matched fresh frozen tumor tissue. Results Using an input of 37 ng genomic DNA, we generated high quality CN data with MIP technology in 88% of FFPE samples from seven diverse collections. When matched fresh frozen tissue was available, the performance of FFPE DNA was comparable to that of DNA obtained from matched frozen tumor (genotype concordance averaged 99.9%), with only a modest loss in performance in FFPE. Conclusion MIP technology can be used to generate high quality CN and genotype data in FFPE as well as fresh frozen samples. PMID:19228381

A SELDI mass spectrometry study of experimental autoimmune encephalomyelitis: sample preparation, reproducibility, and differential protein expression patterns.

PubMed

Azzam, Sausan; Broadwater, Laurie; Li, Shuo; Freeman, Ernest J; McDonough, Jennifer; Gregory, Roger B

2013-05-01

Experimental autoimmune encephalomyelitis (EAE) is an autoimmune, inflammatory disease of the central nervous system that is widely used as a model of multiple sclerosis (MS). Mitochondrial dysfunction appears to play a role in the development of neuropathology in MS and may also play a role in disease pathology in EAE. Here, surface enhanced laser desorption ionization mass spectrometry (SELDI-MS) has been employed to obtain protein expression profiles from mitochondrially enriched fractions derived from EAE and control mouse brain. To gain insight into experimental variation, the reproducibility of sub-cellular fractionation, anion exchange fractionation as well as spot-to-spot and chip-to-chip variation using pooled samples from brain tissue was examined. Variability of SELDI mass spectral peak intensities indicates a coefficient of variation (CV) of 15.6% and 17.6% between spots on a given chip and between different chips, respectively. Thinly slicing tissue prior to homogenization with a rotor homogenizer showed better reproducibility (CV = 17.0%) than homogenization of blocks of brain tissue with a Teflon® pestle (CV = 27.0%). Fractionation of proteins with anion exchange beads prior to SELDI-MS analysis gave overall CV values from 16.1% to 18.6%. SELDI mass spectra of mitochondrial fractions obtained from brain tissue from EAE mice and controls displayed 39 differentially expressed proteins (p≤ 0.05) out of a total of 241 protein peaks observed in anion exchange fractions. Hierarchical clustering analysis showed that protein fractions from EAE animals with severe disability clearly segregated from controls. Several components of electron transport chain complexes (cytochrome c oxidase subunit 6b1, subunit 6C, and subunit 4; NADH dehydrogenase flavoprotein 3, alpha subcomplex subunit 2, Fe-S protein 4, and Fe-S protein 6; and ATP synthase subunit e) were identified as possible differentially expressed proteins. Myelin Basic Protein isoform 8 (MBP8) (14.2 kDa) levels were lower in EAE samples with advanced disease relative to controls, while an MBP fragment (12. 4kDa), likely due to calpain digestion, was increased in EAE relative to controls. The appearance of MBP in mitochondrially enriched fractions is due to tissue freezing and storage, as MBP was not found associated with mitochondria obtained from fresh tissue. SELDI mass spectrometry can be employed to explore the proteome of a complex tissue (brain) and obtain protein profiles of differentially expressed proteins from protein fractions. Appropriate homogenization protocols and protein fractionation using anion exchange beads can be employed to reduce sample complexity without introducing significant additional variation into the SELDI mass spectra beyond that inherent in the SELDI- MS method itself. SELDI-MS coupled with principal component analysis and hierarchical cluster analysis provides protein patterns that can clearly distinguish the disease state from controls. However, identification of individual differentially expressed proteins requires a separate purification of the proteins of interest by polyacrylamide electrophoresis prior to trypsin digestion and peptide mass fingerprint analysis, and unambiguous identification of differentially expressed proteins can be difficult if protein bands consist of several proteins with similar molecular weights.

A SELDI mass spectrometry study of experimental autoimmune encephalomyelitis: sample preparation, reproducibility, and differential protein expression patterns

PubMed Central

2013-01-01

Background Experimental autoimmune encephalomyelitis (EAE) is an autoimmune, inflammatory disease of the central nervous system that is widely used as a model of multiple sclerosis (MS). Mitochondrial dysfunction appears to play a role in the development of neuropathology in MS and may also play a role in disease pathology in EAE. Here, surface enhanced laser desorption ionization mass spectrometry (SELDI-MS) has been employed to obtain protein expression profiles from mitochondrially enriched fractions derived from EAE and control mouse brain. To gain insight into experimental variation, the reproducibility of sub-cellular fractionation, anion exchange fractionation as well as spot-to-spot and chip-to-chip variation using pooled samples from brain tissue was examined. Results Variability of SELDI mass spectral peak intensities indicates a coefficient of variation (CV) of 15.6% and 17.6% between spots on a given chip and between different chips, respectively. Thinly slicing tissue prior to homogenization with a rotor homogenizer showed better reproducibility (CV = 17.0%) than homogenization of blocks of brain tissue with a Teflon® pestle (CV = 27.0%). Fractionation of proteins with anion exchange beads prior to SELDI-MS analysis gave overall CV values from 16.1% to 18.6%. SELDI mass spectra of mitochondrial fractions obtained from brain tissue from EAE mice and controls displayed 39 differentially expressed proteins (p≤ 0.05) out of a total of 241 protein peaks observed in anion exchange fractions. Hierarchical clustering analysis showed that protein fractions from EAE animals with severe disability clearly segregated from controls. Several components of electron transport chain complexes (cytochrome c oxidase subunit 6b1, subunit 6C, and subunit 4; NADH dehydrogenase flavoprotein 3, alpha subcomplex subunit 2, Fe-S protein 4, and Fe-S protein 6; and ATP synthase subunit e) were identified as possible differentially expressed proteins. Myelin Basic Protein isoform 8 (MBP8) (14.2 kDa) levels were lower in EAE samples with advanced disease relative to controls, while an MBP fragment (12. 4kDa), likely due to calpain digestion, was increased in EAE relative to controls. The appearance of MBP in mitochondrially enriched fractions is due to tissue freezing and storage, as MBP was not found associated with mitochondria obtained from fresh tissue. Conclusions SELDI mass spectrometry can be employed to explore the proteome of a complex tissue (brain) and obtain protein profiles of differentially expressed proteins from protein fractions. Appropriate homogenization protocols and protein fractionation using anion exchange beads can be employed to reduce sample complexity without introducing significant additional variation into the SELDI mass spectra beyond that inherent in the SELDI- MS method itself. SELDI-MS coupled with principal component analysis and hierarchical cluster analysis provides protein patterns that can clearly distinguish the disease state from controls. However, identification of individual differentially expressed proteins requires a separate purification of the proteins of interest by polyacrylamide electrophoresis prior to trypsin digestion and peptide mass fingerprint analysis, and unambiguous identification of differentially expressed proteins can be difficult if protein bands consist of several proteins with similar molecular weights. PMID:23635033

Buccal swab as a reliable predictor for X inactivation ratio in inaccessible tissues.

PubMed

de Hoon, Bas; Monkhorst, Kim; Riegman, Peter; Laven, Joop S E; Gribnau, Joost

2015-11-01

As a result of the epigenetic phenomenon of X chromosome inactivation (XCI) every woman is a mosaic of cells with either an inactive paternal X chromosome or an inactive maternal X chromosome. The ratio between inactive paternal and maternal X chromosomes is different for every female individual, and can influence an X-encoded trait or disease. A multitude of X linked conditions is known, and for many of them it is recognised that the phenotype in affected female carriers of the causative mutation is modulated by the XCI ratio. To predict disease severity an XCI ratio is usually determined in peripheral blood samples. However, the correlation between XCI ratios in peripheral blood and disease affected tissues, that are often inaccessible, is poorly understood. Here, we tested several tissues obtained from autopsies of 12 female individuals for patch size and XCI ratio. XCI ratios were analysed using methyl-sensitive PCR-based assays for the AR, PCSK1N and SLITRK4 loci. XCI patch size was analysed by testing the XCI ratio of tissue samples with decreasing size. XCI patch size was analysed for liver, muscle, ovary and brain samples and was found too small to confound testing for XCI ratio in these tissues. XCI ratios were determined in the easily accessible tissues, blood, buccal epithelium and hair follicle, and compared with ratios in several inaccessible tissues. Buccal epithelium is preferable over peripheral blood for predicting XCI ratios of inaccessible tissues. Ovary is the only inaccessible tissue showing a poor correlation to blood and buccal epithelium, but has a good correlation to hair follicle instead. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://group.bmj.com/group/rights-licensing/permissions.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Berenguer de la Cuesta, Felisa; Wenger, Marco P.E.; Bean, Richard J.

Coherent X-ray diffraction has been applied in the imaging of inorganic materials with great success. However, its application to biological specimens has been limited to some notable exceptions, due to the induced radiation damage and the extended nature of biological samples, the last limiting the application of most part of the phasing algorithms. X-ray ptychography, still under development, is a good candidate to overcome such difficulties and become a powerful imaging method for biology. We describe herein the feasibility of applying ptychography to the imaging of biological specimens, in particular collagen rich samples. We report here speckles in diffraction patternsmore » from soft animal tissue, obtained with an optimized small angle X-ray setup that exploits the natural coherence of the beam. By phasing these patterns, dark field images of collagen within tendon, skin, bone, or cornea will eventually be obtained with a resolution of 60-70 nm. We present simulations of the contrast mechanism in collagen based on atomic force microscope images of the samples. Simulations confirmed the 'speckled' nature of the obtained diffraction patterns. Once inverted, the patterns will show the disposition and orientation of the fibers within the tissue, by enhancing the phase contrast between protein and no protein regions of the sample. Our work affords the application of the most innovative coherent X-ray diffraction tools to the study of biological specimens, and this approach will have a significant impact in biology and medicine because it overcomes many of the limits of current microscopy techniques.« less

Decalcification using ethylenediaminetetraacetic acid for clear microstructure imaging of cochlea through optical coherence tomography

NASA Astrophysics Data System (ADS)

Lee, Jaeyul; Kim, Kanghae; Wijesinghe, Ruchire Eranga; Jeon, Doekmin; Lee, Sang Heun; Jeon, Mansik; Jang, Jeong Hun

2016-08-01

The aim of this study was to analyze the effectiveness of decalcification using ethylenediaminetetraacetic acid (EDTA) as an optical clearing method to enhance the depth visibility of internal soft tissues of cochlea. Ex vivo mouse and guinea pig cochlea samples were soaked in EDTA solutions for decalcification, and swept source optical coherence tomography (OCT) was used as imaging modality to monitor the decalcified samples consecutively. The monitored noninvasive cross-sectional images showed that the mouse and guinea pig cochlea samples had to be decalcified for subsequent 7 and 14 days, respectively, to obtain the optimal optical clearing results. Using this method, difficulties in imaging of internal cochlea microstructures of mice could be evaded. The obtained results verified that the depth visibility of the decalcified ex vivo samples was enhanced.

An electrical impedance tomography (EIT) multi-electrode needle-probe device for local assessment of heterogeneous tissue impeditivity.

PubMed

Meroni, Davide; Maglioli, Camilla Carpano; Bovio, Dario; Greco, Francesco G; Aliverti, Andrea

2017-07-01

Electrical Impedance Tomography (EIT) is an image reconstruction technique applied in medicine for the electrical imaging of living tissues. In literature there is the evidence that a large resistivity variation related to the differences of the human tissues exists. As a result of this interest for the electrical characterization of the biological samples, recently the attention is also focused on the identification and characterization of the human tissue, by studying the homogeneity of its structure. An 8 electrodes needle-probe device has been developed with the intent of identifying the structural inhomogeneities under the surface layers. Ex-vivo impeditivity measurements, by placing the needle-probe in 5 different patterns of fat and lean porcine tissue, were performed, and impeditivity maps were obtained by EIDORS open source software for image reconstruction in electrical impedance. The values composing the maps have been analyzed, pointing out a good tissue discrimination, and the conformity with the real images. We conclude that this device is able to perform impeditivity maps matching to reality for position and orientation. In all the five patterns presented is possible to identify and replicate correctly the heterogeneous tissue under test. This new procedure can be helpful to the medical staff to completely characterize the biological sample, in different unclear situations.

Pharmacokinetics and brain penetration of carbapenems in mice.

PubMed

Matsumoto, Kazuaki; Kurihara, Yuji; Kuroda, Yuko; Hori, Seiji; Kizu, Junko

2016-05-01

An adverse effect associated with the administration of carbapenems is central nervous system (CNS) toxicity, with higher brain concentrations of carbapenems being linked to an increased risk of seizures. However, the pharmacokinetics and brain penetration of carbapenems have not yet been examined. Thus, the aim of this in vivo investigation was to determine the pharmacokinetics and brain penetration of carbapenems in mice. Blood samples and brain tissue samples were obtained 10, 20, 30, 60, and 120 min after the subcutaneous administration of carbapenems (91 mg/kg). We obtained the following values for the pharmacokinetic parameters of carbapenems in mice: 1.20-1.71 L/h/kg for CLtotal/F, 1.41-2.03 h(-1) for Ke, 0.34-0.51 h for T1/2, 0.66-0.95 L/kg for Vss/F, 0.49-0.73 h for MRT, 83.46-110.58 μg/mL for Cmax, plasma, and 0.28-0.83 μg/g for Cmax, brain tissue. The AUC0-∞ of the carbapenems tested in plasma were in the following order: doripenem > meropenem > biapenem > imipenem, and in brain tissue were: imipenem > doripenem > meropenem > biapenem. The degrees of brain tissue penetration, defined as the AUC0-∞, brain tissue/fAUC0-∞, plasma ratio, were 0.016 for imipenem, 0.004 for meropenem, 0.002 for biapenem, and 0.008 for doripenem. The results of the present study demonstrated that, of the carbapenems examined, imipenem penetrated brain tissue to the greatest extent. Copyright © 2015 Japanese Society of Chemotherapy and The Japanese Association for Infectious Diseases. Published by Elsevier Ltd. All rights reserved.

Time-resolved fluorescence (TRF) and diffuse reflectance spectroscopy (DRS) for margin analysis in breast cancer.

PubMed

Shalaby, Nourhan; Al-Ebraheem, Alia; Le, Du; Cornacchi, Sylvie; Fang, Qiyin; Farrell, Thomas; Lovrics, Peter; Gohla, Gabriela; Reid, Susan; Hodgson, Nicole; Farquharson, Michael

2018-03-01

One of the major problems in breast cancer surgery is defining surgical margins and establishing complete tumor excision within a single surgical procedure. The goal of this work is to establish instrumentation that can differentiate between tumor and normal breast tissue with the potential to be implemented in vivo during a surgical procedure. A time-resolved fluorescence and reflectance spectroscopy (tr-FRS) system is used to measure fluorescence intensity and lifetime as well as collect diffuse reflectance (DR) of breast tissue, which can subsequently be used to extract optical properties (absorption and reduced scatter coefficient) of the tissue. The tr-FRS data obtained from patients with Invasive Ductal Carcinoma (IDC) whom have undergone lumpectomy and mastectomy surgeries is presented. A preliminary study was conducted to determine the validity of using banked pre-frozen breast tissue samples to study the fluorescence response and optical properties. Once the validity was established, the tr-FRS system was used on a data-set of 40 pre-frozen matched pair cases to differentiate between tumor and normal breast tissue. All measurements have been conducted on excised normal and tumor breast samples post surgery. Our results showed the process of freezing and thawing did not cause any significant differences between fresh and pre-frozen normal or tumor breast tissue. The tr-FRS optical data obtained from 40 banked matched pairs showed significant differences between normal and tumor breast tissue. The work detailed in the main study showed the tr-FRS system has the potential to differentiate malignant from normal breast tissue in women undergoing surgery for known invasive ductal carcinoma. With further work, this successful outcome may result in the development of an accurate intraoperative real-time margin assessment system. Lasers Surg. Med. 50:236-245, 2018. © 2018 Wiley Periodicals, Inc. © 2018 Wiley Periodicals, Inc.

QUANTITATIVE PLUTONIUM MICRODISTRIBUTION IN BONE TISSUE OF VERTEBRA FROM A MAYAK WORKER

PubMed Central

Lyovkina, Yekaterina V.; Miller, Scott C.; Romanov, Sergey A.; Krahenbuhl, Melinda P.; Belosokhov, Maxim V.

2010-01-01

The purpose was to obtain quantitative data on plutonium microdistribution in different structural elements of human bone tissue for local dose assessment and dosimetric models validation. A sample of the thoracic vertebra was obtained from a former Mayak worker with a rather high plutonium burden. Additional information was obtained on occupational and exposure history, medical history, and measured plutonium content in organs. Plutonium was detected in bone sections from its fission tracks in polycarbonate film using neutron-induced autoradiography. Quantitative analysis of randomly selected microscopic fields on one of the autoradiographs was performed. Data included fission fragment tracks in different bone tissue and surface areas. Quantitative information on plutonium microdistribution in human bone tissue was obtained for the first time. From these data, quantitative relationship of plutonium decays in bone volume to decays on bone surface in cortical and trabecular fractions were defined as 2.0 and 0.4, correspondingly. The measured quantitative relationship of decays in bone volume to decays on bone surface does not coincide with recommended models for the cortical bone fraction by the International Commission on Radiological Protection. Biokinetic model parameters of extrapulmonary compartments might need to be adjusted after expansion of the data set on quantitative plutonium microdistribution in other bone types in human as well as other cases with different exposure patterns and types of plutonium. PMID:20838087

First-time detection of Mycobacterium bovis in livestock tissues and milk in the West Bank, Palestinian Territories.

PubMed

Ereqat, Suheir; Nasereddin, Abedelmajeed; Levine, Hagai; Azmi, Kifaya; Al-Jawabreh, Amer; Greenblatt, Charles L; Abdeen, Ziad; Bar-Gal, Gila Kahila

2013-01-01

Bovine tuberculosis, bTB, is classified by the WHO as one of the seven neglected zoonontic diseases that cause animal health problems and has high potential to infect humans. In the West Bank, bTB was not studied among animals and the prevalence of human tuberculosis caused by M. bovis is unknown. Therefore, the aim of this study was to estimate the prevalence of bTB among cattle and goats and identify the molecular characteristics of bTB in our area. A total of 208 tissue samples, representing 104 animals, and 150 raw milk samples, obtained from cows and goats were examined for the presence of mycobacteria. The tissue samples were collected during routine meat inspection from the Jericho abattoir. DNA was extracted from all samples, milk and tissue biopsies (n = 358), and screened for presence of TB DNA by amplifying a 123-bp segment of the insertion sequence IS6110. Eight out of 254 animals (3.1%) were found to be TB positive based on the IS6110-PCR. Identification of M. bovis among the positive TB samples was carried out via real time PCR followed by high resolution melt curve analysis, targeting the A/G transition along the oxyR gene. Spoligotyping analysis revealed a new genotype of M. bovis that was revealed from one tissue sample. Detection of M. bovis in tissue and milk of livestock suggests that apparently healthy cattle and goats are a potential source of infection of bTB and may pose a risk to public health. Hence, appropriate measures including meat inspection at abattoirs in the region are required together with promotion of a health campaign emphasizing the importance of drinking pasteurized milk. In addition, further studies are essential at the farm level to determine the exact prevalence of bTB in goats and cattle herds in the West Bank and Israel.

First-Time Detection of Mycobacterium bovis in Livestock Tissues and Milk in the West Bank, Palestinian Territories

PubMed Central

Ereqat, Suheir; Nasereddin, Abedelmajeed; Levine, Hagai; Azmi, Kifaya; Al-Jawabreh, Amer; Greenblatt, Charles L.; Abdeen, Ziad; Bar-Gal, Gila Kahila

2013-01-01

Background Bovine tuberculosis, bTB, is classified by the WHO as one of the seven neglected zoonontic diseases that cause animal health problems and has high potential to infect humans. In the West Bank, bTB was not studied among animals and the prevalence of human tuberculosis caused by M. bovis is unknown. Therefore, the aim of this study was to estimate the prevalence of bTB among cattle and goats and identify the molecular characteristics of bTB in our area. Methodology/principal findings A total of 208 tissue samples, representing 104 animals, and 150 raw milk samples, obtained from cows and goats were examined for the presence of mycobacteria. The tissue samples were collected during routine meat inspection from the Jericho abattoir. DNA was extracted from all samples, milk and tissue biopsies (n = 358), and screened for presence of TB DNA by amplifying a 123-bp segment of the insertion sequence IS6110. Eight out of 254 animals (3.1%) were found to be TB positive based on the IS6110-PCR. Identification of M. bovis among the positive TB samples was carried out via real time PCR followed by high resolution melt curve analysis, targeting the A/G transition along the oxyR gene. Spoligotyping analysis revealed a new genotype of M. bovis that was revealed from one tissue sample. Significance Detection of M. bovis in tissue and milk of livestock suggests that apparently healthy cattle and goats are a potential source of infection of bTB and may pose a risk to public health. Hence, appropriate measures including meat inspection at abattoirs in the region are required together with promotion of a health campaign emphasizing the importance of drinking pasteurized milk. In addition, further studies are essential at the farm level to determine the exact prevalence of bTB in goats and cattle herds in the West Bank and Israel. PMID:24069475

Differentiated embryonic chondrocytes 1 expression of periodontal ligament tissue and gingival tissue in the patients with chronic periodontitis.

PubMed

Hu, Shenlin; Shang, Wei; Yue, Haitao; Chen, Ruini; Dong, Zheng; Hu, Jinhua; Mao, Zhao; Yang, Jian

2015-04-01

To evaluate the DEC1 expression of periodontal ligament tissue and gingival tissue in the patients with chronic periodontitis. 20 non-smoking patients with chronic periodontitis and 20 healthy individuals were enrolled. Periodontal ligament tissue and gingival tissue samples from healthy subjects were collected during teeth extraction for orthodontic reason or the third molar extraction. The parallel samples from patients with chronic periodontitis were obtained during periodontal flap operations or teeth extraction as part of periodontal treatment. The DEC1 expression and the alkaline phosphatase (ALP) activity of both the periodontal ligament tissue and gingival tissue were determined by Western blot, Immunohistochemistry and ALP Detection Kit. The DEC1 expression of periodontal ligament tissue in the patients with chronic periodontitis decreased significantly along with the decreased ALP activity. On the contrary, the DEC1 expression of gingival tissue in the patients with chronic periodontitis increased significantly. Further study found that the DEC1 expression of gingival tissue increased mainly in the suprabasal layer of gingival epithelial cells but decreased in the gingival connective tissue of the patients with chronic periodontitis. The DEC1 expression decreases in the periodontal ligament tissue which is related to the osteogenic capacity, whereas the DEC1 expression increases in the suprabasal layer of gingival epithelial cells which are involved in immune inflammatory response in the patients with chronic periodontitis. The findings provide a new target to explore the pathology and the therapy of periodontitis. Copyright © 2014 Elsevier Ltd. All rights reserved.

Spectroscopic analysis of normal and neoplastic (WI-FTC) thyroid tissue.

PubMed

Depciuch, Joanna; Stanek-Widera, Agata; Lange, Dariusz; Biskup-Frużyńska, Magdalena; Stanek-Tarkowska, Jadwiga; Czarny, Wojciech; Cebulski, Jozef

2018-06-07

Thyroid cancer holds the first place of the malignant tumors of the endocrine system. One of the less common thyroid cancers is follicular thyroid carcinoma (FTC), which is very difficult to diagnose because it gives the same image as adenoma, which is benign. Certainty of the diagnosis is gained only when FTC gives metastases. Therefore, it was decided to compare normal and neoplastic (FTC) thyroid tissues with Fourier Transform Infrared (FTIR) spectroscopy. The obtained FTIR spectra and Principal Component Analysis (PCA) allowed us to conclude that there are differences in the FTIR spectrum between normal tissues and those affected by cancer. In addition, the results indicate that there is a decrease in the number of functional groups that build cellular and tissue structures in tumoral tissues. The shifts of wave numbers corresponding to the protein and lipid function group vibrations, as well as the calculated second derivative of the FTIR spectra showed the structural changes in neoplastic tissues. Moreover, the deconvolution of the amide I massif indicates that in cancerous tissues the prevailing secondary structure is β-sheet structure, while in normal tissues it is α-helix. The obtained results allow us to conclude that infrared spectroscopy, in addition to providing information on the composition of tested samples, can be an excellent diagnostic tool contributing to understanding the FTC substrate. Copyright © 2018. Published by Elsevier B.V.

Molecular Diagnosis of Orthopedic-Device-Related Infection Directly from Sonication Fluid by Metagenomic Sequencing

PubMed Central

Sanderson, Nicholas D.; Atkins, Bridget L.; Brent, Andrew J.; Cole, Kevin; Foster, Dona; McNally, Martin A.; Oakley, Sarah; Peto, Leon; Taylor, Adrian; Peto, Tim E. A.; Crook, Derrick W.; Eyre, David W.

2017-01-01

ABSTRACT Culture of multiple periprosthetic tissue samples is the current gold standard for microbiological diagnosis of prosthetic joint infections (PJI). Additional diagnostic information may be obtained through culture of sonication fluid from explants. However, current techniques can have relatively low sensitivity, with prior antimicrobial therapy and infection by fastidious organisms influencing results. We assessed if metagenomic sequencing of total DNA extracts obtained direct from sonication fluid can provide an alternative rapid and sensitive tool for diagnosis of PJI. We compared metagenomic sequencing with standard aerobic and anaerobic culture in 97 sonication fluid samples from prosthetic joint and other orthopedic device infections. Reads from Illumina MiSeq sequencing were taxonomically classified using Kraken. Using 50 derivation samples, we determined optimal thresholds for the number and proportion of bacterial reads required to identify an infection and confirmed our findings in 47 independent validation samples. Compared to results from sonication fluid culture, the species-level sensitivity of metagenomic sequencing was 61/69 (88%; 95% confidence interval [CI], 77 to 94%; for derivation samples 35/38 [92%; 95% CI, 79 to 98%]; for validation samples, 26/31 [84%; 95% CI, 66 to 95%]), and genus-level sensitivity was 64/69 (93%; 95% CI, 84 to 98%). Species-level specificity, adjusting for plausible fastidious causes of infection, species found in concurrently obtained tissue samples, and prior antibiotics, was 85/97 (88%; 95% CI, 79 to 93%; for derivation samples, 43/50 [86%; 95% CI, 73 to 94%]; for validation samples, 42/47 [89%; 95% CI, 77 to 96%]). High levels of human DNA contamination were seen despite the use of laboratory methods to remove it. Rigorous laboratory good practice was required to minimize bacterial DNA contamination. We demonstrate that metagenomic sequencing can provide accurate diagnostic information in PJI. Our findings, combined with the increasing availability of portable, random-access sequencing technology, offer the potential to translate metagenomic sequencing into a rapid diagnostic tool in PJI. PMID:28490492

TECHNICAL BRIEF: Isolation of total DNA from postmortem human eye tissues and quality comparison between iris and retina

PubMed Central

Wang, Jay Ching Chieh; Wang, Aikun; Gao, Jiangyuan; Cao, Sijia; Samad, Idris; Zhang, Dean; Ritland, Carol; Cui, Jing Z.

2012-01-01

Background Recent genomic technologies have propelled our understanding of the mechanisms underlying complex eye diseases such as age-related macular degeneration (AMD). Genotyping postmortem eye tissues for known single nucleotide polymorphisms (SNPs) associated with AMD may prove valuable, especially when combined with information obtained through other methods such as immunohistochemistry, western blot, enzyme-linked immunosorbent assay (ELISA), and proteomics. Initially intending to genotype postmortem eye tissues for AMD-related SNPs, our group became interested in isolating and comparing the quality of DNA from the iris and retina of postmortem donor eyes. Since there is no previously published protocol in the literature on this topic, we present a protocol suitable for isolating high-quality DNA from postmortem eye tissues for genomic studies. Methods DNA from 33 retinal samples and 35 iris samples was extracted using the phenol-chloroform-isoamyl method from postmortem donor eye tissues. The quantity of DNA was measured with a spectrophotometer while the quality was checked using gel electrophoresis. The DNA samples were then amplified with PCR for the complement factor H (CFH) gene. The purified amplified products were then genotyped for the SNPs in the CFH gene. Results Regarding concentration, the retina yielded 936 ng/μl of DNA, while the iris yielded 78 ng/μl of DNA. Retinal DNA was also purer than iris DNA (260/280=1.78 vs. 1.46, respectively), and produced superior PCR results. Retinal tissue yielded significantly more DNA than the iris tissue per mg of sample (21.7 ng/μl/mg vs. 7.42 ng/μl/mg). Retinal DNA can be readily amplified with PCR, while iris DNA can also be amplified by adding bovine serum albumin. Overall, retinal tissues yielded DNA of superior quality, quantity, and suitability for genotyping and genomic studies. Conclusions The protocol presented here provides a clear and reliable method for isolating total DNA from postmortem eye tissues. Retinal tissue provides DNA of excellent quantity and quality for genotyping and downstream genomic studies. However, DNA isolated from iris tissues, and treated with bovine serum albumin, may also be a valuable source of DNA for genotyping and genomic studies. PMID:23288996

Detection of toxoplasmosis in experimentally infected goats by PCR.

PubMed

Sreekumar, C; Rao, J R; Mishra, A K; Ray, D; Joshi, P; Singh, R K

2004-05-15

PCR was used to diagnose toxoplasmosis in two pairs of Barbari goats infected by oral administration of doses of either 10(4) or 10(5) oocysts of Toxoplasma gondii. Blood and lymph node aspirates were collected from the infected goats and control goat at intervals, and tissues were also collected from a fetus that was aborted and a doe that died during the trial. Both processed and unprocessed samples were used for the PCR, using primers directed to the multicopy B1 gene. None of the blood samples was positive, but a specific signal was obtained from the lymph node aspirates after partial DNA extraction. Direct PCR of the lung, muscle and mesenteric lymph node of the doe and lung tissue of the aborted fetus yielded the target fragment. The simplified PCR protocols, including partial DNA extraction and direct assay of lung tissue, were effective for the diagnosis of toxoplasmosis.

Orientational tomography of optical axes directions distributions of multilayer biological tissues birefringent polycrystalline networks

NASA Astrophysics Data System (ADS)

Zabolotna, Natalia I.; Dovhaliuk, Rostyslav Y.

2013-09-01

We present a novel measurement method of optic axes orientation distribution which uses a relatively simple measurement setup. The principal difference of our method from other well-known methods lies in direct approach for measuring the orientation of optical axis of polycrystalline networks biological crystals. Our test polarimetry setup consists of HeNe laser, quarter wave plate, two linear polarizers and a CCD camera. We also propose a methodology for processing of measured optic axes orientation distribution which consists of evaluation of statistical, correlational and spectral moments. Such processing of obtained data can be used to classify particular tissue sample as "healthy" or "pathological". For our experiment we use thin layers of histological section of normal and muscular dystrophy tissue sections. It is shown that the difference between mentioned moments` values of normal and pathological samples can be quite noticeable with relative difference up to 6.26.

Clinical significance of CD44 expression in children with hepatoblastoma.

PubMed

Cai, H-Y; Yu, B; Feng, Z-C; Qi, X; Wei, X-J

2015-10-27

The aim of this study was to investigate the expression of CD44 and its clinical significance in children suffering from hepatoblastoma (HB). CD44 expression was detected with immunohistochemistry staining in 30 samples from hepatoblastoma children and 10 normal liver tissue samples from normal children. The data obtained was statistically analyzed using the chi-square test, using the SPSS (v.11.0) software. The rate of CD44 expression was significantly higher (66.7%) in hepatoblastoma tissues than in normal liver tissues (χ(2) = 4.848, P < 0.05). The rate of CD44 expression was significantly higher in children with stage III or IV hepatoblastoma (83.3%) than that in children with stage I and II hepatoblastoma (χ(2) ＝ 5.625, P < 0.05) (41.7%). Therefore, CD44 expression might play an important role in the pathogenesis, progression, and prognosis of HB in children.

Intraoperative optical coherence tomography of the cerebral cortex using a 7 degree-of freedom robotic arm

NASA Astrophysics Data System (ADS)

Reyes Perez, Robnier; Jivraj, Jamil; Yang, Victor X. D.

2017-02-01

Optical Coherence Tomography (OCT) provides a high-resolution imaging technique with limited depth penetration. The current use of OCT is limited to relatively small areas of tissue for anatomical structure diagnosis or minimally invasive guided surgery. In this study, we propose to image a large area of the surface of the cerebral cortex. This experiment aims to evaluate the potential difficulties encountered when applying OCT imaging to large and irregular surface areas. The current state-of-the-art OCT imaging technology uses scanning systems with at most 3 degrees-of-freedom (DOF) to obtain a 3D image representation of the sample tissue. We propose the use of a 7 DOF industrial robotic arm to increase the scanning capabilities of our OCT. Such system will be capable of acquiring data from large samples of tissue that are too irregular for conventional methods. Advantages and disadvantages of our system are discussed.

Update on the Mechanisms of Pulmonary Inflammation and Oxidative Imbalance Induced by Exercise.

PubMed

Araneda, O F; Carbonell, T; Tuesta, M

2016-01-01

The mechanisms involved in the generation of oxidative damage and lung inflammation induced by physical exercise are described. Changes in lung function induced by exercise involve cooling of the airways, fluid evaporation of the epithelial surface, increased contact with polluting substances, and activation of the local and systemic inflammatory response. The present work includes evidence obtained from the different types of exercise in terms of duration and intensity, the effect of both acute performance and chronic performance, and the influence of special conditions such as cold weather, high altitude, and polluted environments. Levels of prooxidants, antioxidants, oxidative damage to biomolecules, and cellularity, as well as levels of soluble mediators of the inflammatory response and its effects on tissues, are described in samples of lung origin. These samples include tissue homogenates, induced sputum, bronchoalveolar lavage fluid, biopsies, and exhaled breath condensate obtained in experimental protocols conducted on animal and human models. Finally, the need to simultaneously explore the oxidative/inflammatory parameters to establish the interrelation between them is highlighted.

Update on the Mechanisms of Pulmonary Inflammation and Oxidative Imbalance Induced by Exercise

PubMed Central

Araneda, O. F.; Carbonell, T.; Tuesta, M.

2016-01-01

The mechanisms involved in the generation of oxidative damage and lung inflammation induced by physical exercise are described. Changes in lung function induced by exercise involve cooling of the airways, fluid evaporation of the epithelial surface, increased contact with polluting substances, and activation of the local and systemic inflammatory response. The present work includes evidence obtained from the different types of exercise in terms of duration and intensity, the effect of both acute performance and chronic performance, and the influence of special conditions such as cold weather, high altitude, and polluted environments. Levels of prooxidants, antioxidants, oxidative damage to biomolecules, and cellularity, as well as levels of soluble mediators of the inflammatory response and its effects on tissues, are described in samples of lung origin. These samples include tissue homogenates, induced sputum, bronchoalveolar lavage fluid, biopsies, and exhaled breath condensate obtained in experimental protocols conducted on animal and human models. Finally, the need to simultaneously explore the oxidative/inflammatory parameters to establish the interrelation between them is highlighted. PMID:26881028

The impact of luteal phase support on endometrial estrogen and progesterone receptor expression: a randomized control trial

PubMed Central

2012-01-01

Background To assess the impact of luteal phase support on the expression of estrogen receptor (ER) alpha and progesterone receptors B (PR-B) on the endometrium of oocyte donors undergoing controlled ovarian hyperstimulation (COH). Methods A prospective, randomized study was conducted in women undergoing controlled ovarian hyperstimulation for oocyte donation. Participants were randomized to receive no luteal support, vaginal progesterone alone, or vaginal progesterone plus orally administered 17 Beta estradiol. Endometrial biopsies were obtained at 4 time points in the luteal phase and evaluated by tissue microarray for expression of ER alpha and PR-B. Results One-hundred and eight endometrial tissue samples were obtained from 12 patients. No differences were found in expression of ER alpha and PR-B among all the specimens with the exception of one sample value. Conclusions The administration of progesterone during the luteal phase of COH for oocyte donor cycles, either with or without estrogen, does not significantly affect the endometrial expression of ER alpha and PR. PMID:22360924

Laser capture microdissection of embryonic cells and preparation of RNA for microarray assays.

PubMed

Redmond, Latasha C; Pang, Christopher J; Dumur, Catherine; Haar, Jack L; Lloyd, Joyce A

2014-01-01

In order to compare the global gene expression profiles of different embryonic cell types, it is first necessary to isolate the specific cells of interest. The purpose of this chapter is to provide a step-by-step protocol to perform laser capture microdissection (LCM) on embryo samples and obtain sufficient amounts of high-quality RNA for microarray hybridizations. Using the LCM/microarray strategy on mouse embryo samples has some challenges, because the cells of interest are available in limited quantities. The first step in the protocol is to obtain embryonic tissue, and immediately cryoprotect and freeze it in a cryomold containing Optimal Cutting Temperature freezing media (Sakura Finetek), using a dry ice-isopentane bath. The tissue is then cryosectioned, and the microscope slides are processed to fix, stain, and dehydrate the cells. LCM is employed to isolate specific cell types from the slides, identified under the microscope by virtue of their morphology. Detailed protocols are provided for using the currently available ArcturusXT LCM instrument and CapSure(®) LCM Caps, to which the selected cells adhere upon laser capture. To maintain RNA integrity, upon removing a slide from the final processing step, or attaching the first cells on the LCM cap, LCM is completed within 20 min. The cells are then immediately recovered from the LCM cap using a denaturing solution that stabilizes RNA integrity. RNA is prepared using standard methods, modified for working with small samples. To ensure the validity of the microarray data, the quality of the RNA is assessed using the Agilent bioanalyzer. Only RNA that is of sufficient integrity and quantity is used to perform microarray assays. This chapter provides guidance regarding troubleshooting and optimization to obtain high-quality RNA from cells of limited availability, obtained from embryo samples by LCM.

Laser Capture Microdissection of Embryonic Cells and Preparation of RNA for Microarray Assays

PubMed Central

Redmond, Latasha C.; Pang, Christopher J.; Dumur, Catherine; Haar, Jack L.; Lloyd, Joyce A.

2014-01-01

In order to compare the global gene expression profiles of different embryonic cell types, it is first necessary to isolate the specific cells of interest. The purpose of this chapter is to provide a step-by-step protocol to perform laser capture microdissection (LCM) on embryo samples and obtain sufficient amounts of high-quality RNA for microarray hybridizations. Using the LCM/microarray strategy on mouse embryo samples has some challenges, because the cells of interest are available in limited quantities. The first step in the protocol is to obtain embryonic tissue, and immediately cryoprotect and freeze it in a cryomold containing Optimal Cutting Temperature freezing media (Sakura Finetek), using a dry ice–isopentane bath. The tissue is then cryosectioned, and the microscope slides are processed to fix, stain, and dehydrate the cells. LCM is employed to isolate specific cell types from the slides, identified under the microscope by virtue of their morphology. Detailed protocols are provided for using the currently available ArcturusXT LCM instrument and CapSure® LCM Caps, to which the selected cells adhere upon laser capture. To maintain RNA integrity, upon removing a slide from the final processing step, or attaching the first cells on the LCM cap, LCM is completed within 20 min. The cells are then immediately recovered from the LCM cap using a denaturing solution that stabilizes RNA integrity. RNA is prepared using standard methods, modified for working with small samples. To ensure the validity of the microarray data, the quality of the RNA is assessed using the Agilent bioanalyzer. Only RNA that is of sufficient integrity and quantity is used to perform microarray assays. This chapter provides guidance regarding troubleshooting and optimization to obtain high-quality RNA from cells of limited availability, obtained from embryo samples by LCM. PMID:24318813

GC-MS/MS Analyses of Biological Samples in Support of Developmental Toxic Effects on Percutaneous Exposure of Rats to VX

DTIC Science & Technology

2016-07-01

of blood, tissues, and organs (heart, lung, liver, kidney , brain, eye, diaphragm, and skin) that were obtained from rats (postnatal days 42 and 70...of blood, tissues, and organs (heart, lung, liver, kidney , brain, eye, and diaphragm) that were used to quantify the amounts of free and regenerated...Biosamples (brain, diaphragm, eye, heart, lung, liver, and kidney ) were collected at time of death or 48 h post-exposure for survivors. All

Antibodies to myofibril antigens in cosmonauts after spaceflights

NASA Technical Reports Server (NTRS)

Tashpulatov, R. Y.; Danilova, T. A.; Lesnyak, A. T.; Legenkov, V. I.; Znamenskiy, V. S.; Dedyuyeva, Y. Y.

1980-01-01

Serum samples obtained from 15 astronauts before and after spaceflights were studied with the use of the indirect immunofluorescent method. In seven astronauts antibodies to different elements of the human heart muscle appeared after flights. Strong and very strong luminescence of the elements of heart muscle tissue was detected in the astronauts after the third space flight. In a study of the sera on sections of bovine heart muscle tissue the reactions of the sera taken before and after flight were found to show no essential differences.

Molecular Characteristics of the Equine Periodontal Ligament

PubMed Central

Pöschke, Antje; Krähling, Bastian; Failing, Klaus; Staszyk, Carsten

2018-01-01

The equine periodontal ligament (PDL) is a fibrous connective tissue that covers the intra-alveolar parts of the tooth and anchors it to the alveolar bone—it, therefore, provides a similar function to a tendinous structure. While several studies have considered the formation and structure of tendons, there is insufficient information particularly on the molecular composition of the PDL. Especially for the equine PDL, there is limited knowledge concerning the expression of genes commonly regarded as typical for tendon tissue. In this study, the gene expression of, e.g., collagen type 1 alpha 1 (COL1), collagen type 3 alpha 1 (COL3), scleraxis (SCX), and fibrocartilage markers was examined in the functional mature equine PDL compared with immature and mature equine tendon tissue. PDL samples were obtained from incisor, premolar, and molar teeth from seven adult horses. Additionally, tendon samples were collected from four adult horses and five foals at different sampling locations. Analyses of gene expression were performed using real-time quantitative polymerase chain reaction (qRT-PCR). Significantly higher expression levels of COL1 and 3 were found in the mature equine PDL in comparison with mature tendon, indicating higher rates of collagen production and turnover in the mature equine PDL. The expression levels of SCX, a specific marker for tenogenic-differentiated cells, were on a similar level in functional mature PDL and in mature tendon tissue. Evidence of chondrogenic metaplasia, often found in tendon entheses or in pressurized regions of tendons, was not found in the mature equine PDL. The obtained results justify further experiments focused on the possible use of equine PDL cells for cell-based regenerative therapies. PMID:29376061

Development of stereotactic mass spectrometry for brain tumor surgery.

PubMed

Agar, Nathalie Y R; Golby, Alexandra J; Ligon, Keith L; Norton, Isaiah; Mohan, Vandana; Wiseman, Justin M; Tannenbaum, Allen; Jolesz, Ferenc A

2011-02-01

Surgery remains the first and most important treatment modality for the majority of solid tumors. Across a range of brain tumor types and grades, postoperative residual tumor has a great impact on prognosis. The principal challenge and objective of neurosurgical intervention is therefore to maximize tumor resection while minimizing the potential for neurological deficit by preserving critical tissue. To introduce the integration of desorption electrospray ionization mass spectrometry into surgery for in vivo molecular tissue characterization and intraoperative definition of tumor boundaries without systemic injection of contrast agents. Using a frameless stereotactic sampling approach and by integrating a 3-dimensional navigation system with an ultrasonic surgical probe, we obtained image-registered surgical specimens. The samples were analyzed with ambient desorption/ionization mass spectrometry and validated against standard histopathology. This new approach will enable neurosurgeons to detect tumor infiltration of the normal brain intraoperatively with mass spectrometry and to obtain spatially resolved molecular tissue characterization without any exogenous agent and with high sensitivity and specificity. Proof of concept is presented in using mass spectrometry intraoperatively for real-time measurement of molecular structure and using that tissue characterization method to detect tumor boundaries. Multiple sampling sites within the tumor mass were defined for a patient with a recurrent left frontal oligodendroglioma, World Health Organization grade II with chromosome 1p/19q codeletion, and mass spectrometry data indicated a correlation between lipid constitution and tumor cell prevalence. The mass spectrometry measurements reflect a complex molecular structure and are integrated with frameless stereotaxy and imaging, providing 3-dimensional molecular imaging without systemic injection of any agents, which can be implemented for surgical margins delineation of any organ and with a rapidity that allows real-time analysis.

Multimodal optical analysis discriminates freshly extracted human sample of gliomas, metastases and meningiomas from their appropriate controls

NASA Astrophysics Data System (ADS)

Zanello, Marc; Poulon, Fanny; Pallud, Johan; Varlet, Pascale; Hamzeh, H.; Abi Lahoud, Georges; Andreiuolo, Felipe; Ibrahim, Ali; Pages, Mélanie; Chretien, Fabrice; di Rocco, Federico; Dezamis, Edouard; Nataf, François; Turak, Baris; Devaux, Bertrand; Abi Haidar, Darine

2017-02-01

Delineating tumor margins as accurately as possible is of primordial importance in surgical oncology: extent of resection is associated with survival but respect of healthy surrounding tissue is necessary for preserved quality of life. The real-time analysis of the endogeneous fluorescence signal of brain tissues is a promising tool for defining margins of brain tumors. The present study aims to demonstrate the feasibility of multimodal optical analysis to discriminate fresh samples of gliomas, metastases and meningiomas from their appropriate controls. Tumor samples were studied on an optical fibered endoscope using spectral and fluorescence lifetime analysis and then on a multimodal set-up for acquiring spectral, one and two-photon fluorescence images, second harmonic generation signals and two-photon fluorescence lifetime datasets. The obtained data allowed us to differentiate healthy samples from tumor samples. These results confirmed the possible clinical relevance of this real-time multimodal optical analysis. This technique can be easily applied to neurosurgical procedures for a better delineation of surgical margins.

Avian mercury exposure and toxicological risk across western North America: A synthesis

USGS Publications Warehouse

Ackerman, Joshua T.; Eagles-Smith, Collin A.; Herzog, Mark; Hartman, Christopher; Peterson, Sarah; Evers, David C.; Jackson, Allyson K.; Elliott, John E.; Vander Pol, Stacy S.; Bryan, Colleen E.

2016-01-01

Methylmercury contamination of the environment is an important issue globally, and birds are useful bioindicators for mercury monitoring programs. The available data on mercury contamination of birds in western North America were synthesized. Original data from multiple databases were obtained and a literature review was conducted to obtain additional mercury concentrations. In total, 29219 original bird mercury concentrations from 225 species were compiled, and an additional 1712 mean mercury concentrations, representing 19998 individuals and 176 species, from 200 publications were obtained. To make mercury data comparable across bird tissues, published equations of tissue mercury correlations were used to convert all mercury concentrations into blood-equivalent mercury concentrations. Blood-equivalent mercury concentrations differed among species, foraging guilds, habitat types, locations, and ecoregions. Piscivores and carnivores exhibited the greatest mercury concentrations, whereas herbivores and granivores exhibited the lowest mercury concentrations. Bird mercury concentrations were greatest in ocean and salt marsh habitats and lowest in terrestrial habitats. Bird mercury concentrations were above toxicity benchmarks in many areas throughout western North America, and multiple hotspots were identified. Additionally, published toxicity benchmarks established in multiple tissues were summarized and translated into a common blood-equivalent mercury concentration. Overall, 66% of birds sampled in western North American exceeded a blood-equivalent mercury concentration of 0.2 μg/g wet weight (ww; above background levels), which is the lowest-observed effect level, 28% exceeded 1.0 μg/g ww (moderate risk), 8% exceeded 3.0 μg/g ww (high risk), and 4% exceeded 4.0 μg/g ww (severe risk). Mercury monitoring programs should sample bird tissues, such as adult blood and eggs, that are most-easily translated into tissues with well-developed toxicity benchmarks and that are directly relevant to bird reproduction. Results indicate that mercury contamination of birds is prevalent in many areas throughout western North America, and large-scale ecological attributes are important factors influencing bird mercury concentrations.

Avian mercury exposure and toxicological risk across western North America: A synthesis

PubMed Central

Ackerman, Joshua T.; Eagles-Smith, Collin A.; Herzog, Mark P.; Hartman, C. Alex; Peterson, Sarah H.; Evers, David C.; Jackson, Allyson K.; Elliott, John E.; Vander Pol, Stacy S.; Bryan, Colleen E.

2017-01-01

Methylmercury contamination of the environment is an important issue globally and birds are useful bioindicators for mercury monitoring programs. The available data on mercury contamination of birds in western North America were synthesized. Original data from multiple databases were obtained and a literature review was conducted to obtain additional mercury concentrations. In total, 29219 original bird mercury concentrations from 225 species were compiled, and an additional 1712 mean mercury concentrations, representing 19998 individuals and 176 species, from 200 publications were obtained. To make mercury data comparable across bird tissues, published equations of tissue mercury correlations were used to convert all mercury concentrations into blood-equivalent mercury concentrations. Blood-equivalent mercury concentrations differed among species, foraging guilds, habitat types, locations, and ecoregions. Piscivores and carnivores exhibited the greatest mercury concentrations, whereas herbivores and granivores exhibited the lowest mercury concentrations. Bird mercury concentrations were greatest in ocean and salt marsh habitats and lowest in terrestrial habitats. Bird mercury concentrations were above toxicity benchmarks in many areas throughout western North America, and multiple hotspots were identified. Additionally, published toxicity benchmarks established in multiple tissues were summarized and translated into a common blood-equivalent mercury concentration. Overall, 66% of birds sampled in western North American exceeded a blood-equivalent mercury concentration of 0.2 μg/g wet weight (ww; above background levels), which is the lowest-observed effect level, 28% exceeded 1.0 μg/g ww (moderate risk), 8% exceeded 3.0 μg/g ww (high risk), and 4% exceeded 4.0 μg/g ww (severe risk). Mercury monitoring programs should sample bird tissues, such as adult blood and eggs, that are most-easily translated into tissues with well-developed toxicity benchmarks and that are directly relevant to bird reproduction. Results indicate that mercury contamination of birds is prevalent in many areas throughout western North America, and large-scale ecological attributes are important factors influencing bird mercury concentrations. PMID:27093907

Inorganic elements in green sea turtles (Chelonia mydas): relationships among external and internal tissues

USGS Publications Warehouse

Faust, Derek R.; Hooper, Michael J.; Cobb, George P.; Barnes, Melanie; Shaver, Donna; Ertolacci, Shauna; Smith, Philip N.

2014-01-01

Inorganic elements from anthropogenic sources have entered marine environments worldwide and are detectable in marine organisms, including sea turtles. Threatened and endangered classifications of sea turtles have heretofore made assessments of contaminant concentrations difficult because of regulatory restrictions on obtaining samples using nonlethal techniques. In the present study, claw and skin biopsy samples were examined as potential indicators of internal tissue burdens in green sea turtles (Chelonia mydas). Significant relationships were observed between claw and liver, and claw and muscle concentrations of mercury, nickel, arsenic, and selenium (p < 0.05). Similarly, significant relationships were observed between skin biopsy concentrations and those in liver, kidney, and muscle tissues for mercury, arsenic, selenium, and vanadium (p < 0.05). Concentrations of arsenic, barium, chromium, nickel, strontium, vanadium, and zinc in claws and skin biopsies were substantially elevated when compared with all other tissues, indicating that these highly keratinized tissues may represent sequestration or excretion pathways. Correlations between standard carapace length and cobalt, lead, and manganese concentrations were observed (p < 0.05), indicating that tissue concentrations of these elements may be related to age and size. Results suggest that claws may indeed be useful indicators of mercury and nickel concentrations in liver and muscle tissues, whereas skin biopsy inorganic element concentrations may be better suited as indicators of mercury, selenium, and vanadium concentrations in liver, kidney, and muscle tissues of green sea turtles.

Characterization of in vitro healthy and pathological human liver tissue periodicity using backscattered ultrasound signals.

PubMed

Machado, Christiano Bittencourt; Pereira, Wagner Coelho de Albuquerque; Meziri, Mahmoud; Laugier, Pascal

2006-05-01

This work studied the periodicity of in vitro healthy and pathologic liver tissue, using backscattered ultrasound (US) signals. It utilized the mean scatterer spacing (MSS) as a parameter of tissue characterization, estimated by three methods: the spectral autocorrelation (SAC), the singular spectrum analysis (SSA) and the quadratic transformation method (SIMON). The liver samples were classified in terms of tissue status using the METAVIR scoring system. Twenty tissue samples were classified in four groups: F0, F1, F3 and F4 (five samples for each). The Kolmogorov-Smirnov test (applied on group pairs) resulted as nonsignificant (p > 0.05) for two pairs only: F1/F3 (for SSA) and F3/F4 (for SAC). A discriminant analysis was applied using as parameters the MSS mean (MSS) and standard deviation (sigmaMSS), the estimates histogram mode (mMSS), and the speed of US (mc(foie)) in the medium, to evaluate the degree of discrimination among healthy and pathologic tissues. The better accuracy (Ac) with SAC (80%) was with parameter group (MSS, sigmaMSS, mc(foie)), achieving a sensitivity (Ss) of 92.3% and a specificity (Sp) of 57.1%. For SSA, the group with all four parameters showed an Ac of 75%, an Ss of 78.6% and an Sp of 66.70%. SIMON obtained the best Ac of all (85%) with group (MSS, mMSS, mc(foie)), an Ss of 100%, but with an Sp of 50%.

Systemic and mammary gland disposition of enrofloxacin in healthy sheep following intramammary administration.

PubMed

López, Cristina; García, Juan José; Sierra, Matilde; Diez, María José; Pérez, Claudia; Sahagún, Ana Maria; Fernández, Nélida

2015-04-09

Mastitis is one of the most important diseases affecting dairy sheep. Antimicrobial drugs are often administered directly through teat to treat or prevent this disease, but data on drug distribution within glandular tissue are scarce and it cannot be estimated from concentrations in milk. Thus, the aim of this study was to investigate systemic and mammary gland distribution of enrofloxacin after intramammary administration. The drug was administered to 6 healthy lactating Assaf sheep with an injector containing an enrofloxacin preparation (1 g drug/5 g ointment). Blood samples were collected at 0, 30, 60, 90, 120, 150 and 180 min. Animals were then sedated and sacrificed, and glandular tissue samples were obtained from treated udders at 2, 4, 6 and 8 cm height. Enrofloxacin concentrations were measured in plasma and tissue samples by UV high-performed liquid chromatography. Mean enrofloxacin plasma concentrations were below 0.5 μg/mL. Mean tissue concentrations decreased in mammary gland with vertical distance from the teat, ranging from 356.6 μg/g at 2 cm to 95.60 μg/g at the base of the udder. Glandular tissue concentrations best fitted to a decreasing monoexponential model, and showed a good correlation with an ex vivo model previously developed. Enrofloxacin concentrations were effective in the entire glandular tissue against the main pathogens causing mastitis in sheep. These results suggest that this drug may be suitable to treat mastitis in sheep by intramammary administration.

Preservation of Multiple Mammalian Tissues to Maximize Science Return from Ground Based and Spaceflight Experiments.

PubMed

Choi, Sungshin; Ray, Hami E; Lai, San-Huei; Alwood, Joshua S; Globus, Ruth K

2016-01-01

Even with recent scientific advancements, challenges posed by limited resources and capabilities at the time of sample dissection continue to limit the collection of high quality tissues from experiments that can be conducted only infrequently and at high cost, such as in space. The resources and time it takes to harvest tissues post-euthanasia, and the methods and duration of long duration storage, potentially have negative impacts on sample quantity and quality, thereby limiting the scientific outcome that can be achieved. The goals of this study were to optimize methods for both sample recovery and science return from rodent experiments, with possible relevance to both ground based and spaceflight studies. The first objective was to determine the impacts of tissue harvest time post-euthanasia, preservation methods, and storage duration, focusing on RNA quality and enzyme activities in liver and spleen as indices of sample quality. The second objective was to develop methods that will maximize science return by dissecting multiple tissues after long duration storage in situ at -80°C. Tissues of C57Bl/6J mice were dissected and preserved at various time points post-euthanasia and stored at -80°C for up to 11 months. In some experiments, tissues were recovered from frozen carcasses which had been stored at -80°C up to 7 months. RNA quantity and quality was assessed by measuring RNA Integrity Number (RIN) values using an Agilent Bioanalyzer. Additionally, the quality of tissues was assessed by measuring activities of hepatic enzymes (catalase, glutathione reductase and GAPDH). Fresh tissues were collected up to one hour post-euthanasia, and stored up to 11 months at -80°C, with minimal adverse effects on the RNA quality of either livers or RNAlater-preserved spleens. Liver enzyme activities were similar to those of positive controls, with no significant effect observed at any time point. Tissues dissected from frozen carcasses that had been stored for up to 7 months at -80°C had variable results, depending on the specific tissue analyzed. RNA quality of liver, heart, and kidneys were minimally affected after 6-7 months of storage at -80°C, whereas RNA degradation was evident in tissues such as small intestine, bone, and bone marrow when they were collected from the carcasses frozen for 2.5 months. These results demonstrate that 1) the protocols developed for spaceflight experiments with on-orbit dissections support the retrieval of high quality samples for RNA expression and some protein analyses, despite delayed preservation post-euthanasia or prolonged storage, and 2) many additional tissues for gene expression analysis can be obtained by dissection even following prolonged storage of the tissue in situ at -80°C. These findings have relevance both to high value, ground-based experiments when sample collection capability is severely constrained, and to spaceflight experiments that entail on-orbit sample recovery by astronauts.

Endoductal tissue sampling of biliary strictures through endoscopic retrograde cholangiopan creatography (ERCP).

PubMed

Pugliese, V; Antonelli, G; Vincenti, M; Gatteschi, B

1997-01-01

Pathological proof of malignant in biliary strictures is useful in the preoperative setting as it helps define therapeutic planning and prognosis, and reduces the length of the subsequent surgical intervention. However, it is difficult to obtain. The aim of this study was to evaluate the yield of histological and cytological examination of endobiliary samples obtained during endoscopic retrograde cholangiopancreatography (ERCP). Endobiliary forceps biopsy and brush cytology were performed during ERCP examination in 52 consecutive patients, 36 with malignant and 16 with benign strictures. Histology and cytology turned out to have the same sensitivity (53%). The gain in sensitivity achieved by combining the two techniques was limited, reaching a value of 61%. The specificity, however, was 100% for both methods. Most of the few complications observed were due to sphincterotomy and subsided spontaneously or with medical treatment. However, one patient experienced a serous complication and chose to be treated by surgical intervention. The complication was caused by forceps biopsy. This study shows that 1) sampling of biliary strictures during ERCP is the primary approach to tissue diagnosis; 2) brush cytology alone is sufficient in clinical practice; 3) forceps biopsy must always be used to sample intra-ampullary strictures but should be considered as a secondary step to sample strictures located more proximally, in the bile ducta, if previous cytology was negative.

Sapphire implant based neuro-complex for deep-lying brain tumors phototheranostics

NASA Astrophysics Data System (ADS)

Sharova, A. S.; Maklygina, YU S.; Yusubalieva, G. M.; Shikunova, I. A.; Kurlov, V. N.; Loschenov, V. B.

2018-01-01

The neuro-complex as a combination of sapphire implant optical port and osteoplastic biomaterial "Collapan" as an Aluminum phthalocyanine nanoform photosensitizer (PS) depot was developed within the framework of this study. The main goals of such neuro-complex are to provide direct access of laser radiation to the brain tissue depth and to transfer PS directly to the pathological tissue location that will allow multiple optical phototheranostics of the deep-lying tumor region without repeated surgical intervention. The developed complex spectral-optical properties research was carried out by photodiagnostics method using the model sample: a brain tissue phantom. The optical transparency of sapphire implant allows obtaining a fluorescent signal with high accuracy, comparable to direct measurement "in contact" with the tissue.

Advantages of tandem LC-MS for the rapid assessment of tissue-specific metabolic complexity using a pentafluorophenylpropyl stationary phase

PubMed Central

Lv, Haitao; Palacios, Gustavo; Hartil, Kirsten; Kurland, Irwin J.

2014-01-01

In this study a UPLC-tandem (Waters Xevo TQ) MRM based MS method was developed for rapid, broad profiling of hydrophilic metabolites from biological samples, in either positive or negative ion modes without the need for an ion pairing reagent, using a reversed-phase pentafluorophenylpropyl (PFPP) column. The developed method was successfully applied to analyze various biological samples from C57BL/6 mice; including urine, duodenum, liver, plasma, kidney, heart, and skeletal muscle. As result, a total 112 of hydrophilic metabolites were detected within 8 min of running time to obtain a metabolite profile of the biological samples. The analysis of this number of hydrophilic metabolites is significantly faster than previous studies. Classification separation for metabolites from different tissues was globally analyzed by PCA, PLS-DA and HCA biostatistical methods. Overall, most of the hydrophilic metabolites were found to have a “fingerprint” characteristic of tissue dependency. In general, a higher level of most metabolites was found in urine, duodenum and kidney. Altogether, these results suggest that this method has potential application for targeted metabolomic analyzes of hydrophilic metabolites in a wide ranges of biological samples. PMID:21322650

Advantages of tandem LC-MS for the rapid assessment of tissue-specific metabolic complexity using a pentafluorophenylpropyl stationary phase.

PubMed

Lv, Haitao; Palacios, Gustavo; Hartil, Kirsten; Kurland, Irwin J

2011-04-01

In this study, a tandem LC-MS (Waters Xevo TQ) MRM-based MS method was developed for rapid, broad profiling of hydrophilic metabolites from biological samples, in either positive or negative ion modes without the need for an ion pairing reagent, using a reversed-phase pentafluorophenylpropyl (PFPP) column. The developed method was successfully applied to analyze various biological samples from C57BL/6 mice, including urine, duodenum, liver, plasma, kidney, heart, and skeletal muscle. As result, a total 112 of hydrophilic metabolites were detected within 8 min of running time to obtain a metabolite profile of the biological samples. The analysis of this number of hydrophilic metabolites is significantly faster than previous studies. Classification separation for metabolites from different tissues was globally analyzed by PCA, PLS-DA and HCA biostatistical methods. Overall, most of the hydrophilic metabolites were found to have a "fingerprint" characteristic of tissue dependency. In general, a higher level of most metabolites was found in urine, duodenum, and kidney. Altogether, these results suggest that this method has potential application for targeted metabolomic analyzes of hydrophilic metabolites in a wide ranges of biological samples.

Myometrial contractility influences oxytocin receptor (OXTR) expression in term trophoblast cells obtained from the maternal surface of the human placenta.

PubMed

Szukiewicz, Dariusz; Bilska, Anna; Mittal, Tarun Kumar; Stangret, Aleksandra; Wejman, Jaroslaw; Szewczyk, Grzegorz; Pyzlak, Michal; Zamlynski, Jacek

2015-09-16

Oxytocin (OXT) acts through its specific receptor (OXTR) and increased density of OXTR and/or augmented sensitivity to OXT were postulated as prerequisites of normal onset of labor. Expression of OXTR in the placental term trophoblast cells has not yet been analyzed in the context of contractile activity of the uterus. Here we examine comparatively OXT contents in the placental tissue adjacent to the uterine wall and expressions of OXTR in this tissue and corresponding isolated placental trophoblast cells. Twenty eight placentae after normal labors at term (group I, N = 14) and after cesarean sections performed without uterine contractile activity (group II, N = 14) have been collected. Tissue excised from the maternal surface of examined placenta was used for OXT concentration measurement, cytotrophoblast cell cultures preparation and immunohistochemistry of OXTR. Concentration of OXT was estimated in the tissue homogenates by an enzyme immunoassay with colorimetric detection. Cytotrophoblast cells were isolated using Kliman's method based on trypsin, DNase, and a 5-70% Percoll gradient centrifugation. The cultures were incubated for 5 days in normoxia. Both placental specimens and terminated cytotrophoblast cultures were fixed and embedded in paraffin before being immunostained for OXTR. Using light microscopy with computed morphometry for quantitative analysis, OXTR expressions were estimated in calibrated areas of the paraffin sections. There were not significant differences between the groups in respect to the mean OXT concentration. However, in both groups the median value of OXT concentration was significantly (p < 0.05) higher in the tissue obtained from the peripheral regions of the maternal surface of the placenta, compared to the samples from the central region of this surface. In placental tissue the mean expression of OXTR in group I was significantly (p < 0.05) increased by approximately 3.2-fold and 3.45-fold (the samples collected from central and peripheral regions, respectively) compared to the values obtained in group II. In the isolated primary trophoblast cultures the differences were even more evident (p < 0.02) and the mean change in OXTR expression in group I comprised approximately 6.9-fold increase and 6.5-fold increase (the samples collected from central and peripheral regions, respectively) compared to the values obtained in group II. Upregulation of OXTR within placental trophoblast cells localized close or adherent to uterine wall may play a crucial role in labor with efficient contractile activity (vaginal delivery). Further studies may disclose if this local OXT/OXTR signaling is utilized in the third stage of labor to elicit placental detachment or contribute in a more versatile way throughout the labor period.

Determination of the methylation status of MGMT in different regions within glioblastoma multiforme.

PubMed

Hamilton, Mark G; Roldán, Gloria; Magliocco, Anthony; McIntyre, John B; Parney, Ian; Easaw, Jacob C

2011-04-01

Epigenetic silencing of the MGMT gene through promoter methylation correlates with improved survival in Glioblastoma Multiforme (GBM) patients receiving concurrent chemoradiotherapy. Although the clinical benefit is primarily seen in patients with methylated MGMT promoter, some unmethylated patients also respond to Temozolomide. One possible explanation may be intratumoral heterogeneity. This study was designed to assess the methylation status of the MGMT promoter in different areas of GBM and determine if methylation status varied depending on the fixation technique (paraffin-embedding versus fresh frozen) used to store tissue. Using intraoperative navigation, biopsies were obtained from three distinct regions: the enhancing outer area, the non-enhancing inner core, and an area immediately outside the enhancing region. Only patients with GBM were included for evaluation and analysis. Samples taken from each area were divided with half stored by flash freezing and the other half stored using paraffin fixation. Methylation Specific-PCR (MS-PCR) was used for analysis of MGMT promoter methylation. Thirteen patients were included. Ten were male with a median age of 62 years. In each patient, samples were taken from the enhancing rim and the necrotic centre. However, it was not considered safe or feasible to obtain samples from the area immediately adjacent to the enhancing tumor rim in one case. All patients were homogeneous for methylation status throughout their tumor and tissue taken adjacent to it when frozen tissue was used. However, four patients had discrepancies in the MGMT promoter status between the frozen and paraffin-embedded blocks and one patient was not homogeneous within the tumor when paraffin-embedded tissue was used. MGMT promoter methylation status was homogeneous in all GBM tumors. Our observation that methylation status varied depending if the DNA was extracted from paraffin-embedded versus frozen tissue is concerning. Although the reason for this is unclear, we postulate that the timing from resection to fixation or the process of fixation itself may potentially alter methylation status in paraffin-embedded tumors.

Physiological and Pathological Impact of Blood Sampling by Retro-Bulbar Sinus Puncture and Facial Vein Phlebotomy in Laboratory Mice

PubMed Central

Holst, Birgitte; Hau, Jann; Rozell, Björn; Abelson, Klas Stig Peter

2014-01-01

Retro-bulbar sinus puncture and facial vein phlebotomy are two widely used methods for blood sampling in laboratory mice. However, the animal welfare implications associated with these techniques are currently debated, and the possible physiological and pathological implications of blood sampling using these methods have been sparsely investigated. Therefore, this study was conducted to assess and compare the impacts of blood sampling by retro-bulbar sinus puncture and facial vein phlebotomy. Blood was obtained from either the retro-bulbar sinus or the facial vein from male C57BL/6J mice at two time points, and the samples were analyzed for plasma corticosterone. Body weights were measured at the day of blood sampling and the day after blood sampling, and the food consumption was recorded automatically during the 24 hours post-procedure. At the end of study, cheeks and orbital regions were collected for histopathological analysis to assess the degree of tissue trauma. Mice subjected to facial vein phlebotomy had significantly elevated plasma corticosterone levels at both time points in contrast to mice subjected to retro-bulbar sinus puncture, which did not. Both groups of sampled mice lost weight following blood sampling, but the body weight loss was higher in mice subjected to facial vein phlebotomy. The food consumption was not significantly different between the two groups. At gross necropsy, subcutaneous hematomas were found in both groups and the histopathological analyses revealed extensive tissue trauma after both facial vein phlebotomy and retro-bulbar sinus puncture. This study demonstrates that both blood sampling methods have a considerable impact on the animals' physiological condition, which should be considered whenever blood samples are obtained. PMID:25426941

The pituitary gland under infrared light - in search of a representative spectrum for homogeneous regions.

PubMed

Banas, A; Banas, K; Furgal-Borzych, A; Kwiatek, W M; Pawlicki, B; Breese, M B H

2015-04-07

The pituitary gland is a small but vital organ in the human body. It is located at the base of the brain and is often described as the master gland due to its multiple functions. The pituitary gland secretes and stores hormones, such as the thyroid-stimulating hormone (TSH), adrenocorticotropic hormone (ACTH), growth hormone (hGH), prolactin, gonadotropins, and luteinizing hormones, as well as the antidiuretic hormone (ADH). A proper diagnosis of pituitary disorders is of utmost importance as this organ participates in regulating a variety of body functions. Typical histopathological analysis provides much valuable information, but it gives no insight into the biochemical background of the changes that occur within the gland. One approach that could be used to evaluate the biochemistry of tissue sections obtained from pituitary disorders is Fourier Transform Infra-Red (FTIR) spectromicroscopy. In order to collect diagnostically valuable information large areas of tissue must be investigated. This work focuses on obtaining a unique and representative FTIR spectrum characteristic of one type of cell architecture within a sample. The idea presented is based on using hierarchical cluster analysis (HCA) for data evaluation to search for uniform patterns within samples from the perspective of FTIR spectra. The results obtained demonstrate that FTIR spectromicroscopy, combined with proper statistical evaluation, can be treated as a complementary method for histopathological analysis and ipso facto can increase the sensitivity and specificity for detecting various disorders not only for the pituitary gland, but also for other human tissues.

A novel method for preparing histology slides to integrate the teaching of gross and microscopic anatomy.

PubMed

Provo-Klimek, Judy A; Troyer, Deryl L

2002-01-01

The authors have previously reported the development of a novel technique for sampling and preparing tissue slides for routine microscopic examination, without the use of a microtome. Termed "RAMP" (Rapid Adhesive Mediated Procedure), this simple, albeit somewhat crude, technique holds promise as a method that can be used in the field by veterinary practitioners for rapid microscopic evaluations to obtain early preliminary estimates of the nature of a mass or lesion. We incorporated the use of this method into a gross anatomy course in an attempt to gauge its utility for novices in tissue sampling and histology slide preparation. By having each group of students take a tissue sample from their cadaver, the activity simulated an actual necropsy situation in which practitioners in the field might use the technique. Because students were able to follow their specimen from sampling to microscopic examination, the activity provided a valuable integration of their learning of gross and microscopic anatomy. We conducted an evaluation of the process and the resulting slides with two successive classes of students. We conclude that the RAMP method is reasonably successful in the hands of individuals not trained in tissue preparation; was well received by the students as a valuable learning tool; and could potentially yield useful histological information for practicing veterinarians. Limitations of the method are also discussed.

Tumoural specimens for forensic purposes: comparison of genetic alterations in frozen and formalin-fixed paraffin-embedded tissues.

PubMed

Ananian, Viviana; Tozzo, Pamela; Ponzano, Elena; Nitti, Donato; Rodriguez, Daniele; Caenazzo, Luciana

2011-05-01

In certain circumstances, tumour tissue specimens are the only DNA resource available for forensic DNA analysis. However, cancer tissues can show microsatellite instability and loss of heterozygosity which, if concerning the short tandem repeats (STRs) used in the forensic field, can cause misinterpretation of the results. Moreover, though formalin-fixed paraffin-embedded tissues (FFPET) represent a large resource for these analyses, the quality of the DNA obtained from this kind of specimen can be an important limit. In this study, we evaluated the use of tumoural tissue as biological material for the determination of genetic profiles in the forensic field, highlighting which STR polymorphisms are more susceptible to tumour genetic alterations and which of the analysed tumours show a higher genetic variability. The analyses were conducted on samples of the same tissues conserved in different storage conditions, to compare genetic profiles obtained by frozen tissues and formalin-fixed paraffin-embedded tissues. The importance of this study is due to the large number of specimens analysed (122), the large number of polymorphisms analysed for each specimen (39), and the possibility to compare, many years after storage, the same tissue frozen and formalin-fixed paraffin-embedded. In the comparison between the genetic profiles of frozen tumour tissues and FFPET, the same genetic alterations have been reported in both kinds of specimens. However, FFPET showed new alterations. We conclude that the use of FFPET requires greater attention than frozen tissues in the results interpretation and great care in both pre-extraction and extraction processes.

Physical properties of hydrated tissue determined by surface interferometry of laser-induced thermoelastic deformation

NASA Astrophysics Data System (ADS)

Dark, Marta L.; Perelman, Lev T.; Itzkan, Irving; Schaffer, Jonathan L.; Feld, Michael S.

2000-02-01

Knee meniscus is a hydrated tissue; it is a fibrocartilage of the knee joint composed primarily of water. We present results of interferometric surface monitoring by which we measure physical properties of human knee meniscal cartilage. The physical response of biological tissue to a short laser pulse is primarily thermomechanical. When the pulse is shorter than characteristic times (thermal diffusion time and acoustic relaxation time) stresses build and propagate as acoustic waves in the tissue. The tissue responds to the laser-induced stress by thermoelastic expansion. Solving the thermoelastic wave equation numerically predicts the correct laser-induced expansion. By comparing theory with experimental data, we can obtain the longitudinal speed of sound, the effective optical penetration depth and the Grüneisen coefficient. This study yields information about the laser-tissue interaction and determines properties of the meniscus samples that could be used as diagnostic parameters.

Microwave fixation versus formalin fixation of surgical and autopsy tissue.

PubMed

Login, G R

1978-05-01

Microwave irradiation of surgical and autopsy tissue penetrates, fixes, and hardens the tissue almost immediately (the fluid media used in the microwave consisted of saline, ten percent phosphate buffered formalin, and distilled water). Tissue sections from a representative sample of organs were tested. Comparable sections were simultaneously fixed in a phosphate buffered ten percent formalin bath in a vaccum oven as a control. Hematoxylin and eosin were used to stain the sections. Results equal to and superior to the control method were obtained. Saline microwave fixation was superior to formalin microwave fixation. Tissues placed in Zenker's solution and fixed in standard microwave oven (for approximately one minute) yielded results at least equal to conventional Zenker fixation (approximately two hours). No tissue hardening resulted from Zenker microwave fixation. A unique time versus temperature graph (microwave heating curve) reduces individual variation with this technique.

Simultaneous isolation of high-quality DNA, RNA, miRNA and proteins from tissues for genomic applications

PubMed Central

Peña-Llopis, Samuel; Brugarolas, James

2014-01-01

Genomic technologies have revolutionized our understanding of complex Mendelian diseases and cancer. Solid tumors present several challenges for genomic analyses, such as tumor heterogeneity and tumor contamination with surrounding stroma and infiltrating lymphocytes. We developed a protocol to (i) select tissues of high cellular purity on the basis of histological analyses of immediately flanking sections and (ii) simultaneously extract genomic DNA (gDNA), messenger RNA (mRNA), noncoding RNA (ncRNA; enriched in microRNA (miRNA)) and protein from the same tissues. After tissue selection, about 12–16 extractions of DNA/RNA/protein can be obtained per day. Compared with other similar approaches, this fast and reliable methodology allowed us to identify mutations in tumors with remarkable sensitivity and to perform integrative analyses of whole-genome and exome data sets, DNA copy numbers (by single-nucleotide polymorphism (SNP) arrays), gene expression data (by transcriptome profiling and quantitative PCR (qPCR)) and protein levels (by western blotting and immunohistochemical analysis) from the same samples. Although we focused on renal cell carcinoma, this protocol may be adapted with minor changes to any human or animal tissue to obtain high-quality and high-yield nucleic acids and proteins. PMID:24136348

Methods: a comparative analysis of radiography, microcomputed tomography, and histology for bone tissue engineering.

PubMed

Hedberg, Elizabeth L; Kroese-Deutman, Henriette C; Shih, Charles K; Lemoine, Jeremy J; Liebschner, Michael A K; Miller, Michael J; Yasko, Alan W; Crowther, Roger S; Carney, Darrell H; Mikos, Antonios G; Jansen, John A

2005-01-01

This study focused on the assessment of radiography, microcomputed tomography, and histology for the evaluation of bone formation in a 15.0-mm defect in the rabbit radius after the implantation of a tissue-engineered construct. Radiography was found to be useful as a noninvasive method for obtaining images of calcified tissue throughout the time course of the experiment. With this method, however, image quality was low, making it difficult to obtain precise information about the location and quantity of the bone formed. Microcomputed tomography was used to create three-dimensional reconstructions of the bone (25-microm resolution). These reconstructions allowed for greater spatial resolution than the radiography, but did not allow for imaging of the implanted scaffold material or the surrounding, nonmineralized tissue. To visualize all materials within the defect area at the cellular level, histology was used. Histological analysis, however, is a destructive technique that did not allow for any further analysis of the samples. Each technique examined here has its own advantages and limitations, but each yields unique information regarding bone regeneration. It is only through the use of all three techniques that complete characterization of the bone growth and tissue/construct responses after implantation in vivo.

High Quality Genomic Copy Number Data from Archival Formalin-Fixed Paraffin-Embedded Leiomyosarcoma: Optimisation of Universal Linkage System Labelling

PubMed Central

Salawu, Abdulazeez; Ul-Hassan, Aliya; Hammond, David; Fernando, Malee; Reed, Malcolm; Sisley, Karen

2012-01-01

Most soft tissue sarcomas are characterized by genetic instability and frequent genomic copy number aberrations that are not subtype-specific. Oligonucleotide microarray-based Comparative Genomic Hybridisation (array CGH) is an important technique used to map genome-wide copy number aberrations, but the traditional requirement for high-quality DNA typically obtained from fresh tissue has limited its use in sarcomas. Although large archives of Formalin-fixed Paraffin-embedded (FFPE) tumour samples are available for research, the degradative effects of formalin on DNA from these tissues has made labelling and analysis by array CGH technically challenging. The Universal Linkage System (ULS) may be used for a one-step chemical labelling of such degraded DNA. We have optimised the ULS labelling protocol to perform aCGH on archived FFPE leiomyosarcoma tissues using the 180k Agilent platform. Preservation age of samples ranged from a few months to seventeen years and the DNA showed a wide range of degradation (when visualised on agarose gels). Consistently high DNA labelling efficiency and low microarray probe-to-probe variation (as measured by the derivative log ratio spread) was seen. Comparison of paired fresh and FFPE samples from identical tumours showed good correlation of CNAs detected. Furthermore, the ability to macro-dissect FFPE samples permitted the detection of CNAs that were masked in fresh tissue. Aberrations were visually confirmed using Fluorescence in situ Hybridisation. These results suggest that archival FFPE tissue, with its relative abundance and attendant clinical data may be used for effective mapping for genomic copy number aberrations in such rare tumours as leiomyosarcoma and potentially unravel clues to tumour origins, progression and ultimately, targeted treatment. PMID:23209738

Deep learning for tissue microarray image-based outcome prediction in patients with colorectal cancer

NASA Astrophysics Data System (ADS)

Bychkov, Dmitrii; Turkki, Riku; Haglund, Caj; Linder, Nina; Lundin, Johan

2016-03-01

Recent advances in computer vision enable increasingly accurate automated pattern classification. In the current study we evaluate whether a convolutional neural network (CNN) can be trained to predict disease outcome in patients with colorectal cancer based on images of tumor tissue microarray samples. We compare the prognostic accuracy of CNN features extracted from the whole, unsegmented tissue microarray spot image, with that of CNN features extracted from the epithelial and non-epithelial compartments, respectively. The prognostic accuracy of visually assessed histologic grade is used as a reference. The image data set consists of digitized hematoxylin-eosin (H and E) stained tissue microarray samples obtained from 180 patients with colorectal cancer. The patient samples represent a variety of histological grades, have data available on a series of clinicopathological variables including long-term outcome and ground truth annotations performed by experts. The CNN features extracted from images of the epithelial tissue compartment significantly predicted outcome (hazard ratio (HR) 2.08; CI95% 1.04-4.16; area under the curve (AUC) 0.66) in a test set of 60 patients, as compared to the CNN features extracted from unsegmented images (HR 1.67; CI95% 0.84-3.31, AUC 0.57) and visually assessed histologic grade (HR 1.96; CI95% 0.99-3.88, AUC 0.61). As a conclusion, a deep-learning classifier can be trained to predict outcome of colorectal cancer based on images of H and E stained tissue microarray samples and the CNN features extracted from the epithelial compartment only resulted in a prognostic discrimination comparable to that of visually determined histologic grade.

Spatial-Resolution Cell Type Proteome Profiling of Cancer Tissue by Fully Integrated Proteomics Technology.

PubMed

Xu, Ruilian; Tang, Jun; Deng, Quantong; He, Wan; Sun, Xiujie; Xia, Ligang; Cheng, Zhiqiang; He, Lisheng; You, Shuyuan; Hu, Jintao; Fu, Yuxiang; Zhu, Jian; Chen, Yixin; Gao, Weina; He, An; Guo, Zhengyu; Lin, Lin; Li, Hua; Hu, Chaofeng; Tian, Ruijun

2018-05-01

Increasing attention has been focused on cell type proteome profiling for understanding the heterogeneous multicellular microenvironment in tissue samples. However, current cell type proteome profiling methods need large amounts of starting materials which preclude their application to clinical tumor specimens with limited access. Here, by seamlessly combining laser capture microdissection and integrated proteomics sample preparation technology SISPROT, specific cell types in tumor samples could be precisely dissected with single cell resolution and processed for high-sensitivity proteome profiling. Sample loss and contamination due to the multiple transfer steps are significantly reduced by the full integration and noncontact design. H&E staining dyes which are necessary for cell type investigation could be selectively removed by the unique two-stage design of the spintip device. This easy-to-use proteome profiling technology achieved high sensitivity with the identification of more than 500 proteins from only 0.1 mm 2 and 10 μm thickness colon cancer tissue section. The first cell type proteome profiling of four cell types from one colon tumor and surrounding normal tissue, including cancer cells, enterocytes, lymphocytes, and smooth muscle cells, was obtained. 5271, 4691, 4876, and 2140 protein groups were identified, respectively, from tissue section of only 5 mm 2 and 10 μm thickness. Furthermore, spatially resolved proteome distribution profiles of enterocytes, lymphocytes, and smooth muscle cells on the same tissue slices and across four consecutive sections with micrometer distance were successfully achieved. This fully integrated proteomics technology, termed LCM-SISPROT, is therefore promising for spatial-resolution cell type proteome profiling of tumor microenvironment with a minute amount of clinical starting materials.

In vitro autoradiography of carcinoembryonic antigen in tissue from patients with colorectal cancer using multifunctional antibody TF2 and 67/68Ga-labeled haptens by pretargeting

PubMed Central

Hall, Håkan; Velikyan, Irina; Blom, Elisabeth; Ulin, Johan; Monazzam, Azita; Påhlman, Lars; Micke, Patrick; Wanders, Alkwin; McBride, William; Goldenberg, David M.; Långström, Bengt

2012-01-01

The carcinoembryonic antigen (CEA) was visualized in vitro in tissue from patients with colorectal cancer with trivalent bispecific antibody TF2 and two hapten molecules, [67/68Ga]Ga-IMP461 and [67/68Ga]Ga-IMP485 by means of pretargeting. Colorectal cancer tissue samples obtained from surgery at Uppsala University Hospital, were frozen fresh and cryosectioned. The two hapten molecules comprising 1,4,7-triazacyclononanetriacetic acid chelate moiety (NOTA) were labeled with 67Ga or 68Ga. The autoradiography was conducted by incubating the tissue samples with the bispecific antibody TF2, followed by washing and incubation with one of the radiolabeled hapten molecules. After washing, drying and exposure to phosphor imager plates, the autoradiograms were analyzed and compared to standard histochemistry (hematoxylin-eosin). Pronounced binding was found in the tissue from colorectal cancer using the bispecific antibody TF2 and either of the haptens [67/68Ga]Ga-IMP461 and [67/68Ga]Ga-IMP485. Distinct binding was also detected in the epithelium of most samples of neighboring tissue, taken at a minimum of 10 cm from the site of the tumor. It is concluded that pretargeting CEA with the bispecific antibody TF2 followed by the addition of 67/68Ga-labeled hapten is extremely sensitive for visualizing this marker for colorectal cancer. This methodology is therefore a very specific complement to other histochemical techniques in the diagnosis of biopsies or in samples taken from surgery. Use of the pretargeting technique in vivo may also be an advance in diagnosing patients with colorectal cancer, either using 67Ga and SPECT or 68Ga and PET. PMID:23133809

Clearance of a persistent Picornavirus infection is associated with enhanced pro-apoptotic and cellular immune responses

USDA-ARS?s Scientific Manuscript database

Immunomodulatory mechanisms associated with clearance versus persistence of foot-and-mouth disease virus (FMDV) in distinct microanatomic compartments of the bovine nasopharynx were investigated using quantitative RT-PCR and whole transcriptome microarray. Analysis of tissue samples obtained during ...

Immunohistochemical evidence for expression of fast-twitch type sarco(endo)plasmic reticulum Ca2+ ATPase (SERCA1) in German shepherd dogs with dilated cardiomyopathy myocardium.

PubMed

Summerfield, Nuala; Peters, Mary E; Hercock, Carol A; Mobasheri, Ali; Young, Iain S

2010-04-01

Dilated cardiomyopathy (DCM) is one of the most common acquired canine heart diseases. It is particularly common in large and giant breed dogs. Although a great deal is known about the clinical progression and manifestations of the disease, the underlying cellular and molecular mechanisms remain poorly understood. One widely held belief is that calcium-handling abnormalities are critically involved in the disease process. This study investigates the changes in expression of the sarco(endo)plasmic reticulum calcium ATPase (SERCA) isoforms in DCM myocardium from German shepherd dogs. Affected tissue samples were obtained from German shepherd dogs with DCM, euthanized for intractable congestive heart failure while normal myocardial tissue samples were obtained from German shepherd dogs, euthanized for non-cardiovascular reasons. Tissue microarrays containing normal and DCM myocardium samples were prepared, immunostained with SERCA1 and SERCA2 antibodies and analyzed. We were able to demonstrate, for the first time, that while there is little change in the expression of the cardiac isoform (SERCA2), there is clear expression of the fast-twitch skeletal muscle isoform SERCA1 in the myocardium of dogs diagnosed with DCM. We propose that SERCA1 expression is evidence of a natural adaptive response to the impaired Ca2+ handling thought to occur in German shepherd dogs with DCM and heart failure. Copyright 2010 Elsevier B.V. All rights reserved.

Precise Detection of IDH1/2 and BRAF Hotspot Mutations in Clinical Glioma Tissues by a Differential Calculus Analysis of High-Resolution Melting Data

PubMed Central

Hatae, Ryusuke; Yoshimoto, Koji; Kuga, Daisuke; Akagi, Yojiro; Murata, Hideki; Suzuki, Satoshi O.; Mizoguchi, Masahiro; Iihara, Koji

2016-01-01

High resolution melting (HRM) is a simple and rapid method for screening mutations. It offers various advantages for clinical diagnostic applications. Conventional HRM analysis often yields equivocal results, especially for surgically obtained tissues. We attempted to improve HRM analyses for more effective applications to clinical diagnostics. HRM analyses were performed for IDH1R132 and IDH2R172 mutations in 192 clinical glioma samples in duplicate and these results were compared with sequencing results. BRAFV600E mutations were analyzed in 52 additional brain tumor samples. The melting profiles were used for differential calculus analyses. Negative second derivative plots revealed additional peaks derived from heteroduplexes in PCR products that contained mutations; this enabled unequivocal visual discrimination of the mutations. We further developed a numerical expression, the HRM-mutation index (MI), to quantify the heteroduplex-derived peak of the mutational curves. Using this expression, all IDH1 mutation statuses matched those ascertained by sequencing, with the exception of three samples. These discordant results were all derived from the misinterpretation of sequencing data. The effectiveness of our approach was further validated by analyses of IDH2R172 and BRAFV600E mutations. The present analytical method enabled an unequivocal and objective HRM analysis and is suitable for reliable mutation scanning in surgically obtained glioma tissues. This approach could facilitate molecular diagnostics in clinical environments. PMID:27529619

EXPERIMENTAL NEEDLE BIOPSY OF THE MYOCARDIUM OF DOGS WITH PARTICULAR REFERENCE TO HISTOLOGIC STUDY BY ELECTRON MICROSCOPY

PubMed Central

Price, Kenneth C.; Weiss, Jules M.; Hata, Daikichi; Smith, John R.

1955-01-01

Our experience with needle biopsy of the heart in dogs indicates that myocardial tissue can be sampled one or more times in each animal with comparative safety. Tamponade, pericarditis, serious arrhythmias, or myocardial infarction due to the interruption of coronary vessels was not observed. Excellent specimens were obtained for critical study by light and electron microscopy. Casten and Marsh (1) have used biochemical techniques to study myocardial tissue obtained in similar fashion. Histochemical methods would also be applicable. Although limited to animal studies at present, the technique may conceivably be adapted to the study of human disease. Myocardial puncture has been carried out (20–22) in patients for the recording of intracardiac pressures and for other diagnostic purposes without apparent harm. Our study of the myocardium of dogs by electron microscopy generally confirms the observations of other workers, except that presence of significant numbers of red blood cells in the extravascular spaces of the heart had not been previously described (and is possibly an artifact). Nevertheless, it is notable that the tissue cells, cellular membranes, and intracellular structures appeared to be intact and undistorted in the tissue specimens which were obtained, fixed, and examined by these methods. PMID:14367689

[Sinusal penetration of amoxicillin-clavulanic acid. Formulation 1 g./125 mg., twice daily versus formulation 500 mg./125 mg., three times daily].

PubMed

Jehl, F; Klossek, J M; Peynegre, R; Serrano, E; Castillo, L; Bobin, S; Desprez, D; Renault, C; Neel, V; Rouffiac, E; Borie, C

2002-10-19

In order to meet the evolution of pneumococcus resistance to beta-lactam antibiotics, a new formulation of amoxicillin (AMX) and clavulanic acid (CA), with twice as much AMX (1 g/125 mg vs. 500 mg/125 mg) was developed for the treatment of acute pneumonia in patients at risk. This formulation can also be used in the treatment of acute maxillary sinusitis using a 1 g/125 mg regimen twice-daily. Compare the sinusal penetration of AMX and CA (1 g/125 mg twice-daily vs. 500 mg/125 mg three times a day) when administered at both regimens to demonstrate equivalent pharmacokinetic and pharmacodynamic behaviour of the former when compared to the latter. Concentrations of AMX and CA were measured in the anterior ethmoid, maxillary, posterior ethmoid sinus and in the middle nasa concha in 62 patients undergoing surgery for nasosinusal polyps. Patients randomised in two groups corresponding to 2 oral regimens, received either 1 g/125 mg twice a day or 500 mg/125 mg three times a day for 4 days. The last dose in both groups was administered 1 h 30, 3, 5 or 8 hrs prior to surgery. Serum samples were taken simultaneously to tissue samples. AMX and CA were measured by high performance liquid chromatography. Exogenous and above all endogenous blood contamination were taken into account with the hematocrit as well as blood and tissue haemoglobin concentrations. Comparisons of tissue concentrations were made for each sampling time, according to values obtained for a specific tissue with both doses on one hand, and on the other to values obtained with a specific dose in different tissues. The calculated pharmacodynamic parameters, which are considered to be predictive for bacteriological and clinical efficacy, result directly from tissue concentrations of AMX. tissue inhibitory quotients (IQtissue = Tissue concentration/MIC). time above MICs for serum and tissue concentrations (T > MIC). As regards AMX, whatever the dose, at 1 h 30 and at 3 hrs, tissue concentrations did not differ significantly whatever the tissue studied (from 1.1 to 2.5 micrograms/g). Conversely, at 5 and 8 hrs, they were greater than after the 1 g/125 mg regimen given twice-daily (0.06-0.7 vs. 0.7-1.8 micrograms/g). If we consider a given dose, the comparison between the various tissues showed identical concentrations in the four tissues studied at each sampling time, except in two cases with the dose of 500 mg/125 mg 3 times a day. T > MIC for serum and tissue showed higher values than those required for AMX/pneumococcus association (40-50%) with, nevertheless, greater tissue values for the 1 g/125 mg dose given twice-daily when MIC was of 1 microgram/ml (40-52% vs. 50-66%). The maximum tissue inhibitory quotients were also greater with the twice-daily 1 g/125 mg dose, when calculated with MIC 50 or 90 of S. Pneumoniae, H. influenzae, M. catarrhalis or S. pyogenes. As for CA, concentrations were equivalent for both doses at each sampling time and greater than those required in vitro during respectively 4 and 5 hours for beta-lactamases H. influenzae and M. catarrhalis. A least an equivalence between both dose regimens was observed, with occasionally a superiority of the twice-daily 1 g/125 mg dose, in terms of pharmacokinetics, tissue penetration and pharmacodynamics for both AMX and CA. This new regimen therefore appears more appropriate for the treatment of acute maxillary sinusitis in adults.

The histopathologic reliability of tissue taken from cadavers within the gross anatomy laboratory.

PubMed

Rae, Guenevere; Newman, William P; McGoey, Robin; Donthamsetty, Supriya; Karpinski, Aryn C; Green, Jeffrey

2018-03-01

The purpose of this study was to examine the histopathologic reliability of embalmed cadaveric tissue taken from the gross anatomy laboratory. Tissue samples from hearts, livers, lungs, and kidneys were collected after the medical students' dissection course was completed. All of the cadavers were embalmed in a formalin-based fixative solution. The tissue was processed, embedded in paraffin, sectioned at six micrometers, and stained with H&E. The microscope slides were evaluated by a board certified pathologist to determine whether the cellular components of the tissues were preserved at a high enough quality to allow for histopathologic diagnosis. There was a statistically significant relationship between ratings and organ groups. Across all organs, there was a smaller proportion of "poor" ratings. The lung group had the highest percentage of "poor" ratings (23.1%). The heart group had the least "poor" ratings (0.0%). The largest percentage of "satisfactory" ratings were in the lung group (52.8%), and the heart group contained the highest percentage of "good" ratings (58.5%) The lung group had the lowest percentage of "good" ratings (24.2%). These results indicate that heart tissue is more reliable than lung, kidney, or liver tissue when utilizing tissue from the gross anatomy laboratory for research and/or educational purposes. This information advises educators and researchers about the quality and histopathologic reliability of tissue samples obtained from the gross anatomy laboratory. Anat Sci Educ 11: 207-214. © 2017 American Association of Anatomists. © 2017 American Association of Anatomists.

Preliminary characterization of IL32 in basal-like/triple negative compared to other types of breast cell lines and tissues

PubMed Central

2014-01-01

Background Triple negative breast cancer (TNBC) and often basal-like cancers are defined as negative for estrogen receptor, progesterone receptor and Her2 gene expression. Over the past few years an incredible amount of data has been generated defining the molecular characteristics of both cancers. The aim of these studies is to better understand the cancers and identify genes and molecular pathways that might be useful as targeted therapies. In an attempt to contribute to the understanding of basal-like/TNBC, we examined the Gene Expression Omnibus (GEO) public datasets in search of genes that might define basal-like/TNBC. The Il32 gene was identified as a candidate. Findings Analysis of several GEO datasets showed differential expression of IL32 in patient samples previously designated as basal and/or TNBC compared to normal and luminal breast samples. As validation of the GEO results, RNA and protein expression levels were examined using MCF7 and MDA MB231 cell lines and tissue microarrays (TMAs). IL32 gene expression levels were higher in MDA MB231 compared to MCF7. Analysis of TMAs showed 42% of TNBC tissues and 25% of the non-TNBC were positive for IL32, while non-malignant patient samples and all but one hyperplastic tissue sample demonstrated lower levels of IL32 protein expression. Conclusion Data obtained from several publically available GEO datasets showed overexpression of IL32 gene in basal-like/TNBC samples compared to normal and luminal samples. In support of these data, analysis of TMA clinical samples demonstrated a particular pattern of IL32 differential expression. Considered together, these data suggest IL32 is a candidate suitable for further study. PMID:25100201

Assessment of molecular markers demonstrates concordance between samples acquired via stereotactic biopsy and open craniotomy in both anaplastic astrocytomas and glioblastomas.

PubMed

Gessler, Florian; Baumgarten, Peter; Bernstock, Joshua D; Harter, Patrick; Lescher, Stephanie; Senft, Christian; Seifert, Volker; Marquardt, Gerhard; Weise, Lutz

2017-06-01

The classification, treatment and prognosis of high-grade gliomas has been shown to correlate with the expression of molecular markers (e.g. MGMT promotor methylation and IDH1 mutations). Acquisition of tumor samples may be obtained via stereotactic biopsy or open craniotomy. Between the years 2009 and 2013, 22 patients initially diagnosed with HGGs via stereotactic biopsy, that ultimately underwent open craniotomy for resection of their tumor were prospectively included in an institutional glioma database. MGMT promotor analysis was performed using methylation-specific (MS)-PCR and IDH1R132H mutation analysis was performed using immunohistochemistry. Three patients (13.7%) exhibited IDH1R132H mutations in samples obtained via stereotactic biopsy. Tissue derived from stereotaxic biopsy was demonstrated to have MGMT promotor methylation in ten patients (45.5%), while a non-methylated MGMT promotor was demonstrated in ten patients (45.5%); inconclusive results were obtained for the remaining two patients (9%) within our cohort. The initial histologic grading, IDH1R132H mutation and MGMT promotor methylation results were confirmed using samples obtained during open craniotomy in all but one patient; here inconclusive MGMT promotor analysis was obtained in contrast to that which was obtained via stereotactic biopsy. Tumor samples acquired via stereotactic biopsy provide accurate information with regard to clinically relevant molecular markers that have been shown to impact patient care decisions. The profile of markers analyzed in our cohort was nearly concordant between those samples obtained via stereotactic biopsy or open craniotomy thereby suggesting that clinical decisions may be based on the molecular profile of the tumor samples obtained via stereotactic biopsy.

In vitro imaging of ophthalmic tissue by digital interference holography

NASA Astrophysics Data System (ADS)

Potcoava, Mariana C.; Kay, Christine N.; Kim, Myung K.; Richards, David W.

2010-01-01

We used digital interference holography (DIH) for in vitro imaging of human optic nerve head and retina. Samples of peripheral retina, macula, and optic nerve head from two formaldehyde-preserved human eyes were dissected and mounted onto slides. Holograms were captured by a monochrome CCD camera (Sony XC-ST50, with 780 × 640 pixels and pixel size of ∼9 µm). Light source was a solid-state pumped dye laser with tunable wavelength range of 560-605 nm. Using about 50 wavelengths in this band, holograms were obtained and numerically reconstructed using custom software based on NI LabView. Tomographic images were produced by superposition of holograms. Holograms of all tissue samples were obtained with a signal-to-noise ratio of approximately 50 dB. Optic nerve head characteristics (shape, diameter, cup depth, and cup width) were quantified with a few micron resolution (4.06-4.8 µm). Multiple layers were distinguishable in cross-sectional images of the macula. To our knowledge, this is the first report of DIH use to image human macular and optic nerve tissue. DIH has the potential to become a useful tool for researchers and clinicians in the diagnosis and treatment of many ocular diseases, including glaucoma and a variety of macular diseases.

Feature space analysis of MRI

NASA Astrophysics Data System (ADS)

Soltanian-Zadeh, Hamid; Windham, Joe P.; Peck, Donald J.

1997-04-01

This paper presents development and performance evaluation of an MRI feature space method. The method is useful for: identification of tissue types; segmentation of tissues; and quantitative measurements on tissues, to obtain information that can be used in decision making (diagnosis, treatment planning, and evaluation of treatment). The steps of the work accomplished are as follows: (1) Four T2-weighted and two T1-weighted images (before and after injection of Gadolinium) were acquired for ten tumor patients. (2) Images were analyed by two image analysts according to the following algorithm. The intracranial brain tissues were segmented from the scalp and background. The additive noise was suppressed using a multi-dimensional non-linear edge- preserving filter which preserves partial volume information on average. Image nonuniformities were corrected using a modified lowpass filtering approach. The resulting images were used to generate and visualize an optimal feature space. Cluster centers were identified on the feature space. Then images were segmented into normal tissues and different zones of the tumor. (3) Biopsy samples were extracted from each patient and were subsequently analyzed by the pathology laboratory. (4) Image analysis results were compared to each other and to the biopsy results. Pre- and post-surgery feature spaces were also compared. The proposed algorithm made it possible to visualize the MRI feature space and to segment the image. In all cases, the operators were able to find clusters for normal and abnormal tissues. Also, clusters for different zones of the tumor were found. Based on the clusters marked for each zone, the method successfully segmented the image into normal tissues (white matter, gray matter, and CSF) and different zones of the lesion (tumor, cyst, edema, radiation necrosis, necrotic core, and infiltrated tumor). The results agreed with those obtained from the biopsy samples. Comparison of pre- to post-surgery and radiation feature spaces confirmed that the tumor was not present in the second study but radiation necrosis was generated as a result of radiation.

Reconstruction of an input function from a dynamic PET water image using multiple tissue curves

NASA Astrophysics Data System (ADS)

Kudomi, Nobuyuki; Maeda, Yukito; Yamamoto, Yuka; Nishiyama, Yoshihiro

2016-08-01

Quantification of cerebral blood flow (CBF) is important for the understanding of normal and pathologic brain physiology. When CBF is assessed using PET with {{\\text{H}}2} 15O or C15O2, its calculation requires an arterial input function, which generally requires invasive arterial blood sampling. The aim of the present study was to develop a new technique to reconstruct an image derived input function (IDIF) from a dynamic {{\\text{H}}2} 15O PET image as a completely non-invasive approach. Our technique consisted of using a formula to express the input using tissue curve with rate constant parameter. For multiple tissue curves extracted from the dynamic image, the rate constants were estimated so as to minimize the sum of the differences of the reproduced inputs expressed by the extracted tissue curves. The estimated rates were used to express the inputs and the mean of the estimated inputs was used as an IDIF. The method was tested in human subjects (n  =  29) and was compared to the blood sampling method. Simulation studies were performed to examine the magnitude of potential biases in CBF and to optimize the number of multiple tissue curves used for the input reconstruction. In the PET study, the estimated IDIFs were well reproduced against the measured ones. The difference between the calculated CBF values obtained using the two methods was small as around  <8% and the calculated CBF values showed a tight correlation (r  =  0.97). The simulation showed that errors associated with the assumed parameters were  <10%, and that the optimal number of tissue curves to be used was around 500. Our results demonstrate that IDIF can be reconstructed directly from tissue curves obtained through {{\\text{H}}2} 15O PET imaging. This suggests the possibility of using a completely non-invasive technique to assess CBF in patho-physiological studies.

Cryopreservation of Cell/Scaffold Tissue-Engineered Constructs

PubMed Central

Costa, Pedro F.; Dias, Ana F.; Reis, Rui L.

2012-01-01

The aim of this work was to study the effect of cryopreservation over the functionality of tissue-engineered constructs, analyzing the survival and viability of cells seeded, cultured, and cryopreserved onto 3D scaffolds. Further, it also evaluated the effect of cryopreservation over the properties of the scaffold material itself since these are critical for the engineering of most tissues and in particular, tissues such as bone. For this purpose, porous scaffolds, namely fiber meshes based on a starch and poly(caprolactone) blend were seeded with goat bone marrow stem cells (GBMSCs) and cryopreserved for 7 days. Discs of the same material seeded with GBMSCs were also used as controls. After this period, these samples were analyzed and compared to samples collected before the cryopreservation process. The obtained results demonstrate that it is possible to maintain cell viability and scaffolds properties upon cryopreservation of tissue-engineered constructs based on starch scaffolds and goat bone marrow mesenchymal cells using standard cryopreservation methods. In addition, the outcomes of this study suggest that the greater porosity and interconnectivity of scaffolds favor the retention of cellular content and cellular viability during cryopreservation processes, when compared with nonporous discs. These findings indicate that it might be possible to prepare off-the-shelf engineered tissue substitutes and preserve them to be immediately available upon request for patients' needs. PMID:22676448

Classification of microscopic images of breast tissue

NASA Astrophysics Data System (ADS)

Ballerini, Lucia; Franzen, Lennart

2004-05-01

Breast cancer is the most common form of cancer among women. The diagnosis is usually performed by the pathologist, that subjectively evaluates tissue samples. The aim of our research is to develop techniques for the automatic classification of cancerous tissue, by analyzing histological samples of intact tissue taken with a biopsy. In our study, we considered 200 images presenting four different conditions: normal tissue, fibroadenosis, ductal cancer and lobular cancer. Methods to extract features have been investigated and described. One method is based on granulometries, which are size-shape descriptors widely used in mathematical morphology. Applications of granulometries lead to distribution functions whose moments are used as features. A second method is based on fractal geometry, that seems very suitable to quantify biological structures. The fractal dimension of binary images has been computed using the euclidean distance mapping. Image classification has then been performed using the extracted features as input of a back-propagation neural network. A new method that combines genetic algorithms and morphological filters has been also investigated. In this case, the classification is based on a correlation measure. Very encouraging results have been obtained with pilot experiments using a small subset of images as training set. Experimental results indicate the effectiveness of the proposed methods. Cancerous tissue was correctly classified in 92.5% of the cases.

Determination of sulfonamides in animal tissues by modified QuEChERS and liquid chromatography tandem mass spectrometry.

PubMed

Wen, Ching-Hsuan; Lin, Shu-Ling; Fuh, Ming-Ren

2017-03-01

In this study, the salting-out solvent extraction and dispersive solid-phase extraction (dSPE) clean-up steps in QuEChERS (quick, easy, cheap, effective, rugged, and safe) method were optimized to reduce matrix effect and efficiently extract target sulfonamides from a variety of edible animal tissues. The extracted sulfonamides were then analyzed using liquid chromatography tandem mass spectrometry (LC-MS/MS). Good extraction recoveries (74.0-100.3% in five different sources of animal tissues; n=3) with acceptable matrix effect (<10%, except for liver samples) were obtained using the proposed method. For the first time, a commercial ND-lipids cartridge was used to remove hydrophobic matrix components from fat-rich animal tissues in the clean-up step of QuEChERS. In addition, good linearity (0.125-12.5ngg -1 ) was observed using matrix-matched calibration (in beef). Limits of detection (LODs) were estimated at 0.01-0.03ngg -1 in beef, pork, and chicken samples. For beef tripe and pig liver samples, the LODs were in the range of 0.02-0.04ngg -1 . Good intra-day/inter-day precision (1.0-10.5%/0.4-8.0%) and accuracy (95.2-107.2%/97.8-102.1%) were also achieved using the modified QuEChERS for sample pretreatment. The applicability of the modified QuEChERS-LC-MS/MS method was demonstrated by determining the occurrence of target sulfonamides in various edible animal tissues for potential food safety analysis. Copyright © 2016 Elsevier B.V. All rights reserved.

Secretory Leukocyte Protease Inhibitor Expression and High-Risk HPV Infection in Anal Lesions of HIV-Positive Patients.

PubMed

Nicol, Alcina F; Brunette, Laurie L; Nuovo, Gerard J; Grinsztejn, Beatriz; Friedman, Ruth K; Veloso, Valdiléa G; Cunha, Cynthia B; Coutinho, José R; Vianna-Andrade, Cecilia; Oliveira, Nathalia S; Woodham, Andrew W; DA Silva, Diane M; Kast, W Martin

2016-09-01

The aim of this study was to evaluate secretory leukocyte protease inhibitor (SLPI) expression in anal biopsies from HIV-positive (HIV+) individuals, and compare that to anal intraepithelial neoplasia (AIN) diagnoses and human papillomavirus (HPV) status. This is a cross-sectional study of a cohort of 54 HIV+ (31 males and 23 females) from an AIDS clinic in Rio de Janeiro, Brazil. The study material consisted of anorectal tissue biopsies obtained from HIV+ subjects, which were used to construct tissue microarray paraffin blocks for immunohistochemical analysis of SLPI expression. Biopsies were evaluated by an expert pathologist and classified as low-grade AIN1, high-grade AIN2/3, or normal squamous epithelium. In addition, DNA from the biopsies was extracted and analyzed for the presence of low- or high-risk HPV DNA. Histologically, normal squamous epithelium from the anorectal region showed strong positive SLPI staining in 17/20 (85%) samples. In comparison, 9/17 (53%) dysplastic squamous epithelial samples from AIN1 patients showed strong SLPI staining, and only 5/17 (29%) samples from AIN2/3 patients exhibited strong SPLI staining, which both were significantly fewer than those from normal tissue (P = 0.005). Furthermore, there was a significantly higher proportion of samples in which oncogenic high-risk HPV genotypes were detected in low SLPI-expressing tissues than that in tissues with high SLPI expression (P = 0.040). Taken together these results suggest that low SLPI expression is associated with high-risk HPV infections in the development of AIN.

Image analysis of representative food structures: application of the bootstrap method.

PubMed

Ramírez, Cristian; Germain, Juan C; Aguilera, José M

2009-08-01

Images (for example, photomicrographs) are routinely used as qualitative evidence of the microstructure of foods. In quantitative image analysis it is important to estimate the area (or volume) to be sampled, the field of view, and the resolution. The bootstrap method is proposed to estimate the size of the sampling area as a function of the coefficient of variation (CV(Bn)) and standard error (SE(Bn)) of the bootstrap taking sub-areas of different sizes. The bootstrap method was applied to simulated and real structures (apple tissue). For simulated structures, 10 computer-generated images were constructed containing 225 black circles (elements) and different coefficient of variation (CV(image)). For apple tissue, 8 images of apple tissue containing cellular cavities with different CV(image) were analyzed. Results confirmed that for simulated and real structures, increasing the size of the sampling area decreased the CV(Bn) and SE(Bn). Furthermore, there was a linear relationship between the CV(image) and CV(Bn) (.) For example, to obtain a CV(Bn) = 0.10 in an image with CV(image) = 0.60, a sampling area of 400 x 400 pixels (11% of whole image) was required, whereas if CV(image) = 1.46, a sampling area of 1000 x 100 pixels (69% of whole image) became necessary. This suggests that a large-size dispersion of element sizes in an image requires increasingly larger sampling areas or a larger number of images.

WAXS fat subtraction model to estimate differential linear scattering coefficients of fatless breast tissue: Phantom materials evaluation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Tang, Robert Y., E-mail: rx-tang@laurentian.ca; Laamanen, Curtis, E-mail: cx-laamanen@laurentian.ca; McDonald, Nancy, E-mail: mcdnancye@gmail.com

Purpose: Develop a method to subtract fat tissue contributions to wide-angle x-ray scatter (WAXS) signals of breast biopsies in order to estimate the differential linear scattering coefficients μ{sub s} of fatless tissue. Cancerous and fibroglandular tissue can then be compared independent of fat content. In this work phantom materials with known compositions were used to test the efficacy of the WAXS subtraction model. Methods: Each sample 5 mm in diameter and 5 mm thick was interrogated by a 50 kV 2.7 mm diameter beam for 3 min. A 25 mm{sup 2} by 1 mm thick CdTe detector allowed measurements ofmore » a portion of the θ = 6° scattered field. A scatter technique provided means to estimate the incident spectrum N{sub 0}(E) needed in the calculations of μ{sub s}[x(E, θ)] where x is the momentum transfer argument. Values of μ{sup ¯}{sub s} for composite phantoms consisting of three plastic layers were estimated and compared to the values obtained via the sum μ{sup ¯}{sub s}{sup ∑}(x)=ν{sub 1}μ{sub s1}(x)+ν{sub 2}μ{sub s2}(x)+ν{sub 3}μ{sub s3}(x), where ν{sub i} is the fractional volume of the ith plastic component. Water, polystyrene, and a volume mixture of 0.6 water + 0.4 polystyrene labelled as fibphan were chosen to mimic cancer, fat, and fibroglandular tissue, respectively. A WAXS subtraction model was used to remove the polystyrene signal from tissue composite phantoms so that the μ{sub s} of water and fibphan could be estimated. Although the composite samples were layered, simulations were performed to test the models under nonlayered conditions. Results: The well known μ{sub s} signal of water was reproduced effectively between 0.5 < x < 1.6 nm{sup −1}. The μ{sup ¯}{sub s} obtained for the heterogeneous samples agreed with μ{sup ¯}{sub s}{sup ∑}. Polystyrene signals were subtracted successfully from composite phantoms. The simulations validated the usefulness of the WAXS models for nonlayered biopsies. Conclusions: The methodology to measure μ{sub s} of homogeneous samples was quantitatively accurate. Simple WAXS models predicted the probabilities for specific x-ray scattering to occur from heterogeneous biopsies. The fat subtraction model can allow μ{sub s} signals of breast cancer and fibroglandular tissue to be compared without the effects of fat provided there is an independent measurement of the fat volume fraction ν{sub f}. Future work will consist of devising a quantitative x-ray digital imaging method to estimate ν{sub f} in ex vivo breast samples.« less

Simultaneous extraction and clean-up of polychlorinated biphenyls and their metabolites from small tissue samples using pressurized liquid extraction

PubMed Central

Kania-Korwel, Izabela; Zhao, Hongxia; Norstrom, Karin; Li, Xueshu; Hornbuckle, Keri C.; Lehmler, Hans-Joachim

2008-01-01

A pressurized liquid extraction-based method for the simultaneous extraction and in situ clean-up of polychlorinated biphenyls (PCBs), hydroxylated (OH)-PCBs and methylsulfonyl (MeSO2)-PCBs from small (< 0.5 gram) tissue samples was developed and validated. Extraction of a laboratory reference material with hexane:dichloromethane:methanol (48:43:9, v/v) and Florisil as fat retainer allowed an efficient recovery of PCBs (78–112%; RSD: 13–37%), OH-PCBs (46±2%; RSD: 4%) and MeSO2-PCBs (89±21%; RSD: 24%). Comparable results were obtained with an established analysis method for PCBs, OH-PCBs and MeSO2-PCBs. PMID:19019378

GoIFISH: a system for the quantification of single cell heterogeneity from IFISH images.

PubMed

Trinh, Anne; Rye, Inga H; Almendro, Vanessa; Helland, Aslaug; Russnes, Hege G; Markowetz, Florian

2014-08-26

Molecular analysis has revealed extensive intra-tumor heterogeneity in human cancer samples, but cannot identify cell-to-cell variations within the tissue microenvironment. In contrast, in situ analysis can identify genetic aberrations in phenotypically defined cell subpopulations while preserving tissue-context specificity. GoIFISHGoIFISH is a widely applicable, user-friendly system tailored for the objective and semi-automated visualization, detection and quantification of genomic alterations and protein expression obtained from fluorescence in situ analysis. In a sample set of HER2-positive breast cancers GoIFISHGoIFISH is highly robust in visual analysis and its accuracy compares favorably to other leading image analysis methods. GoIFISHGoIFISH is freely available at www.sourceforge.net/projects/goifish/.

Primary cultures of astrocytes from fetal bovine brain.

PubMed

Ballarin, Cristina; Peruffo, Antonella

2012-01-01

We describe here a method to obtain primary cell cultures from the cerebral cortex and the hypothalamus of bovine fetuses. We report how tissue origin, developmental stages, and culture medium conditions influence cell differentiation and the prevalence of glial cells vs. neurons. We compare explants from early, middle, and late stages of development and two different fetal calf serum concentrations (1 and 10%) to identify the best conditions to obtain and grow viable astrocytes in culture. In addition, we describe how to cryopreserve and obtain viable cortical astrocytes from frozen fetal bovine brain samples.

Fetal tissue banking for transplantation: characteristics of the donor population and considerations for donor and tissue screening.

PubMed

Newman-Gage, H; Bravo, D; Holmberg, L; Mason, J; Eisenhower, M; Nekhani, N; Fantel, A

2000-01-01

We initiated this study to evaluate the suitability for therapeutic use in transplantation of tissues obtained from human abortuses. We have developed protocols for the collection, handling and preservation of hepatic stem cells from electively aborted embryos and have developed methods for assessment of the cells so derived and processed. In this paper we present our findings regarding screening of potential donors, acquisition of fetal tissues, and assessment of the tissues for potentially infectious contaminants. We assess the suitability of the tissue donors according to current standards used for donors of commonly transplanted tissues (e.g., bone grafts, skin grafts and heart valves) and present data regarding the real availability of tissues from elective abortion procedures that would meet those standard tissue banking criteria.We specifically evaluated the donor's willingness to provide a blood sample for testing, conducted a detailed interview similar to those used for typical organ and tissue donors, and assessed the type and incidence of contamination in collected tissues. We find that although many women are willing to consent to use of the tissues for transplantation, attrition from the study for various reasons results in few fetal organs ultimately realistically available for transplantation. Typical reasons for attrition include: unwillingness to have a blood sample drawn or tested, positive serology results, social/medical high risk factors for acquisition of transmissible disease, no identifiable organs available, and unacceptable microbial contamination. Thus, although it might seem that due to the numbers of abortions performed annually, that there would be substantial numbers of suitable tissues available, only a small proportion are truly suitable for transplantation.

Lack of concordance in microarray gene expression responses to Phenobarbital in companion aged FFPE and Frozen liver samples

EPA Science Inventory

Despite the immense potential value of public and private biorepositories, direct utilization of archival tissues for molecular profiling has been limited. A major reason for this limited use is the difficulty in obtaining reliable transcriptomic profiles from formalin-fixed par...

75 FR 64986 - Marine Mammals; File No. 14525

Federal Register 2010, 2011, 2012, 2013, 2014

2010-10-21

... captivity, sampled while in captivity, and euthanized at the termination of study to obtain their brains. Whole brains and brain tissues will be imported to the U.S. for anatomical and immunohistochemical studies. The first aim of the project is to correlate the release of major neurotransmitters in the brain...

Human-tissue-related inventions: ownership and intellectual property rights in international collaborative research in developing countries.

PubMed

Andanda, P A

2008-03-01

There are complex unresolved ethical, legal and social issues related to the use of human tissues obtained in the course of research or diagnostic procedures and retained for further use in research. The question of intellectual property rights over commercially viable products or procedures that are derived from these samples and the suitability or otherwise of participants relinquishing their rights to the samples needs urgent attention. The complexity of these matters lies in the fact that the relationship between intellectual property rights and ownership or rights pertaining to the samples on which the intellectual property right is based may either be overlooked or taken for granted. What equally makes the matter complex is that samples may be obtained from participants in developing countries and exported to developed countries for analysis and research. It is important for research ethics committees to tread carefully when reviewing research protocols that raise such issues for purposes of ensuring that appropriate benefit sharing agreements, particularly with developing countries, are in place. This paper attempts to analyse the key questions related to ownership and intellectual property rights in commercially viable products derived from human tissue samples. Patent law is used as a point of reference as opposed to other forms of intellectual property rights such as industrial designs because it is the right that most inventors apply for in respect of human tissue-related inventions. The key questions are formulated following a systematic analysis of peer reviewed journal articles that have reported original investigations into relevant issues in this field. Most of the cases and reported studies that are referred to in this paper do not directly deal with HIV/AIDS research but the underlying principles are helpful in HIV/AIDS research as well. Pertinent questions, which members of ethics review committees should focus on in this regard are discussed and suggestions on appropriate approaches to the issues are proposed in the form of specific questions that an ethics review committee should consider. Specific recommendations regarding areas for further research and action are equally proposed.

Does peripheral quantitative computed tomography ignore tissue density of cancellous bone?

PubMed

Banse, X; Devogelaer, J P

2002-01-01

The purpose of this work was to determine the capacity of peripheral quantitative computed tomography (pQCT) to accurately measure the true physical properties of vertebral cancellous bone samples and to predict their stiffness. pQCT bone mineral density (BMD) was first measured in ideal conditions. Ten cubic specimens of vertebral cancellous bone (10 x 10 x 10 mm) were washed with a water jet, defatted, and scanned in saline after elimination of air bubbles; thirteen slices were obtained. Seventy-one unprepared cylindrical samples were scanned in more realistic conditions, which allow further biomechanical testing. After extraction from the vertebral body, the samples were pushed into a plastic tube (no effort was made to remove the marrow or air bubbles), and only four slices were obtained to reduce the duration of scan. For the 81 samples, the true bone volume fraction (BV/TV, %), true apparent density (rho(app), g/cm(3)), and tissue density (rho(tiss), g/cm(3)) (an indicator of the degree of mineralization of the matrix) were then measured using Archimedes principle. rho(app) was closely correlated to BV/TV (r(2) = 0.97). rho(tiss) (1.58 +/- 0.08 g/cm(2)) was almost constant but had some influence on rho(app) (r(2) = 0.03, p < 0.001). The pQCT BMD predicted accurately rho(app) (r(2) = 0.96) and BV/TV (r(2) = 0.93) for the cylinders. For the cubes, in ideal conditions, the same correlations were even better (r(2) > 0.99, both). Analysis of covariance indicated no difference (p > 0.05) in the regressions due to preparation of the samples. The stiffness was better predicted by the true rho(app) (r(2) = 0.87) than by BV/TV (r(2) = 0.83), indicating that stiffness was influenced by small differences in the tissue density. Consequently, the correlation between pQCT BMD and stiffness was excellent (r(2) = 0.84). The fact that pQCT did not ignore this tissue density information compensated for the inaccuracies linked to realistic scanning conditions of the cylinder.

Composite elastomeric polyurethane scaffolds incorporating small intestinal submucosa for soft tissue engineering.

PubMed

Da, Lincui; Gong, Mei; Chen, Anjing; Zhang, Yi; Huang, Yizhou; Guo, Zhijun; Li, Shengfu; Li-Ling, Jesse; Zhang, Li; Xie, Huiqi

2017-09-01

Although soft tissue replacement has been clinically successful in many cases, the corresponding procedure has many limitations including the lack of resilience and mechanical integrity, significant donor-site morbidity, volume loss with time, and fibrous capsular contracture. These disadvantages can be alleviated by utilizing bio-absorbable scaffolds with high resilience and large strain, which are capable of stimulating natural tissue regeneration. Hence, the chemically crosslinked tridimensional scaffolds obtained by incorporating water-based polyurethane (PU) (which was synthesized from polytetramethylene ether glycol, isophorone diisocyanate, and 2,2-bis(hydroxymethyl) butyric acid) into a bioactive extracellular matrix consisting of small intestinal submucosa (SIS) have been tested in this study to develop a new approach for soft tissue engineering. After characterizing the structure and properties of the produced PU/SIS composites, the strength, Young's modulus, and resilience of wet PU/SIS samples were compared with those of crosslinked PU. In addition, the fabricated specimens were investigated using human umbilical vein endothelial cells to evaluate their ability to enhance cell attachment and proliferation. As a result, the synthesized PU/SIS samples exhibited high resilience and were capable of enhancing cell viability with no evidence of cytotoxicity. Subcutaneous implantation in animals and the subsequent testing conducted after 2, 4, and 8weeks indicated that sound implant integration and vascularization occurred inside the PU/SIS composites, while the presence of SIS promoted cell infiltration, angiogenesis, and ultimately tissue regeneration. The obtained results revealed that the produced PU/SIS composites were characterized by high bioactivity and resilience, and, therefore, could be used for soft tissue engineering applications. Hybrid composites containing synthetic polymers with high mechanical strength and naturally derived components, which create a bio-mimetic environment, are one of the most promising biomaterials. Although synthetic polymer/ECM composites have been previously used for soft tissue repair, their resilience properties were not investigated in sufficient detail, while the development of elastic composites composed of synthetic polymers and ECMs in nontoxic aqueous solutions remains a rather challenging task. In this study, porous PU/SIS composites were fabricated in a non-toxic manner; the obtained materials exhibited sufficient mechanical support, which promote cell growth, angiogenesis, and tissue regeneration. The described method can be adapted for the development of scaffolds with various acellular matrices and subsequently used during the restoration of particular types of tissue. Copyright © 2017 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Determination of oxidation state of iron in normal and pathologically altered human aortic valves

NASA Astrophysics Data System (ADS)

Czapla-Masztafiak, J.; Lis, G. J.; Gajda, M.; Jasek, E.; Czubek, U.; Bolechała, F.; Borca, C.; Kwiatek, W. M.

2015-12-01

In order to investigate changes in chemical state of iron in normal and pathologically altered human aortic valves X-ray absorption spectroscopy was applied. Since Fe is suspected to play detrimental role in aortic valve stenosis pathogenesis the oxidation state of this element has been determined. The experimental material consisted of 10 μm sections of valves excised during routine surgery and from autopsies. The experiment was performed at the MicroXAS beamline of the SLS synchrotron facility in Villigen (Switzerland). The Fe K-edge XANES spectra obtained from tissue samples were carefully analyzed and compared with the spectra of reference compounds containing iron in various chemical structures. The analysis of absorption edge position and shape of the spectra revealed that both chemical forms of iron are presented in valve tissue but Fe3+ is the predominant form. Small shift of the absorption edge toward higher energy in the spectra from stenotic valve samples indicates higher content of the Fe3+ form in pathological tissue. Such a phenomenon suggests the role of Fenton reaction and reactive oxygen species in the etiology of aortic valve stenosis. The comparison of pre-edge regions of XANES spectra for control and stenotic valve tissue confirmed no differences in local symmetry or spin state of iron in analyzed samples.

Extraction and analysis methods for the determination of pyrethroid insecticides in surface water, sediments and biological tissues at environmentally relevant concentrations.

PubMed

Mekebri, A; Crane, D B; Blondina, G J; Oros, D R; Rocca, J L

2008-05-01

The aim of this study was to develop and validate chemical methods for measuring pyrethroid insecticides at environmentally relevant concentrations in different matrices. The analytes included six synthetic pyrethroids with the highest agricultural and commercial structural uses in California: bifenthrin, cyfluthrin, cypermethrin, esfenvalerate/fenvalerate, lambda-cyhalothrin, permethrin, and their corresponding stereoisomers, which includes enantiomers, diastereomers and racemic mixtures. Fortified water samples were extracted for analysis of synthetic pyrethroids using liquid-liquid extraction, while fortified sediment and fish tissue samples were extracted using pressurized fluid extraction followed by gel permeation chromatography (GPC) to remove matrix interferences. A florisil column was used for additional cleanup and fractionation of sediment and tissue extracts. Extracts were analyzed using dual column high resolution gas chromatography with electron capture detection (GC/ECD) and confirmation was obtained with gas chromatography mass spectrometry using a quadrupole ion trap detector in MS-MS mode. Method detection limits (MDLs) have been established for water (1-3 ng/L), sediment (0.5-4 ng/g dry weight) and tissue (1-3 ng/g fresh weight). Mean percent recoveries of fortified blanks and samples ranged from 75 to 115% with relative standard deviation (RSD) values less than 20% for all target compounds.

A comparison study of different excitation wavelengths to determine the relative content of key biomolecules in breast cancer and breast normal tissue

NASA Astrophysics Data System (ADS)

Sordillo, Laura A.; Sordillo, Peter P.; Budansky, Yury; Pu, Yang; Alfano, R. R.

2015-03-01

Fluorescence profiles from breast cancer and breast normal tissue samples with excitation wavelengths at 280 nm and 340 nm were obtained using the conventional LS-50 Perkin-Elmer spectrometer. Fluorescence ratios from these tissue samples, demonstrated by emission peaks at 340 nm, 440 nm and 460 nm and likely representing tryptophan and NADH, show increased relative content of tryptophan in malignant samples. Double ratio (DR) techniques were used to measure the severity of disease. The single excitation double ratio (Single-DR) method utilizes the emission intensity peaks from the spectrum acquired using a single excitation of 280 nm; while the dual excitation double ratio (dual-DR) method utilizes the emission intensity peaks from the spectra acquired using an excitation of 280 nm and 340 nm. Single-DR and dual-DR from 13 patients with breast carcinoma were compared in terms of their efficiency to distinguish high from low/intermediate tumors. Similar results were found with both methods. Results suggest that dual excitation wavelengths may be as effective as single excitation wavelength in calculating the relative content of biomolecules in breast cancer tissue, as well as for the assessment of the malignant potential of these tumors.

Application of simple biomechanical and biochemical tests to heart valve leaflets: implications for heart valve characterization and tissue engineering.

PubMed

Huang, Hsiao-Ying S; Balhouse, Brittany N; Huang, Siyao

2012-11-01

A simple biomechanical test with real-time displacement and strain mapping is reported, which provides displacement vectors and principal strain directions during the mechanical characterization of heart valve tissues. The maps reported in the current study allow us to quickly identify the approximate strain imposed on a location in the samples. The biomechanical results show that the aortic valves exhibit stronger anisotropic mechanical behavior than that of the pulmonary valves before 18% strain equibiaxial stretching. In contrast, the pulmonary valves exhibit stronger anisotropic mechanical behavior than aortic valves beyond 28% strain equibiaxial stretching. Simple biochemical tests are also conducted. Collagens are extracted at different time points (24, 48, 72, and 120 h) at different locations in the samples. The results show that extraction time plays an important role in determining collagen concentration, in which a minimum of 72 h of extraction is required to obtain saturated collagen concentration. This work provides an easy approach for quantifying biomechanical and biochemical properties of semilunar heart valve tissues, and potentially facilitates the development of tissue engineered heart valves.

Evaluation of residual radioactivity in human tissues associated with weapons testing at the nevada test site. Technical report, 1 June 1981-31 March 1989

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wrenn, M.E.; Singh, N.P.; Paschoa, A.S.

1992-11-01

Residual radioactivity consisting of 239 Pu were measured by radiochemistry and alpha spectrometry in samples of bone and soft tissues from 100 autopsies or surgeries from northern and southwestern Utah and from control areas in Colorado and Pennsylvania. Based upon the isotopic ratio 240 Pu/239 Pu contamination was attributable to atmospheric weapons tests at the Nevada Test Site (NTS). In addition, 110 thyroid tissue samples obtained from tissue blocks made at autopsy of veterans dying at the VA hospital in Salt Lake City in the 1940's and 1950's were measured for 129 I (half life 16 million year) and 127more » I (stable) by neutron activation. The results were analyzed by year of death, by periods before and during atmospheric nuclear weapons testing at the NTS and by origin of usual residence. A model was developed to relate thyroid dose from 131 I to the measured 124/127 I ratios, and thyroid dose estimates were made based upon the measured ratios.« less

Optimising the quantification of cytokines present at low concentrations in small human mucosal tissue samples using Luminex assays☆

PubMed Central

Staples, Emily; Ingram, Richard James Michael; Atherton, John Christopher; Robinson, Karen

2013-01-01

Sensitive measurement of multiple cytokine profiles from small mucosal tissue biopsies, for example human gastric biopsies obtained through an endoscope, is technically challenging. Multiplex methods such as Luminex assays offer an attractive solution but standard protocols are not available for tissue samples. We assessed the utility of three commercial Luminex kits (VersaMAP, Bio-Plex and MILLIPLEX) to measure interleukin-17A (IL-17) and interferon-gamma (IFNγ) concentrations in human gastric biopsies and we optimised preparation of mucosal samples for this application. First, we assessed the technical performance, limits of sensitivity and linear dynamic ranges for each kit. Next we spiked human gastric biopsies with recombinant IL-17 and IFNγ at a range of concentrations (1.5 to 1000 pg/mL) and assessed kit accuracy for spiked cytokine recovery and intra-assay precision. We also evaluated the impact of different tissue processing methods and extraction buffers on our results. Finally we assessed recovery of endogenous cytokines in unspiked samples. In terms of sensitivity, all of the kits performed well within the manufacturers' recommended standard curve ranges but the MILLIPLEX kit provided most consistent sensitivity for low cytokine concentrations. In the spiking experiments, the MILLIPLEX kit performed most consistently over the widest range of concentrations. For tissue processing, manual disruption provided significantly improved cytokine recovery over automated methods. Our selected kit and optimised protocol were further validated by measurement of relative cytokine levels in inflamed and uninflamed gastric mucosa using Luminex and real-time polymerase chain reaction. In summary, with proper optimisation Luminex kits (and for IL-17 and IFNγ the MILLIPLEX kit in particular) can be used for the sensitive detection of cytokines in mucosal biopsies. Our results should help other researchers seeking to quantify multiple low concentration cytokines in small tissue samples. PMID:23644159

A liquid chromatography-tandem mass spectrometry-based targeted proteomics assay for monitoring P-glycoprotein levels in human breast tissue.

PubMed

Yang, Ting; Chen, Fei; Xu, Feifei; Wang, Fengliang; Xu, Qingqing; Chen, Yun

2014-09-25

P-glycoprotein (P-gp) can efflux drugs from cancer cells, and its overexpression is commonly associated with multi-drug resistance (MDR). Thus, the accurate quantification of P-gp would help predict the response to chemotherapy and for prognosis of breast cancer patients. An advanced liquid chromatography-tandem mass spectrometry (LC/MS/MS)-based targeted proteomics assay was developed and validated for monitoring P-gp levels in breast tissue. Tryptic peptide 368IIDNKPSIDSYSK380 was selected as a surrogate analyte for quantification, and immuno-depleted tissue extract was used as a surrogate matrix. Matched pairs of breast tissue samples from 60 patients who were suspected to have drug resistance were subject to analysis. The levels of P-gp were quantified. Using data from normal tissue, we suggested a P-gp reference interval. The experimental values of tumor tissue samples were compared with those obtained from Western blotting and immunohistochemistry (IHC). The result indicated that the targeted proteomics approach was comparable to IHC but provided a lower limit of quantification (LOQ) and could afford more reliable results at low concentrations than the other two methods. LC/MS/MS-based targeted proteomics may allow the quantification of P-gp in breast tissue in a more accurate manner. Copyright © 2014 Elsevier B.V. All rights reserved.

Factors for C-Kit Expression in Cardiac Outgrowth Cells and Human Heart Tissue.

PubMed

Matsushita, Satoshi; Minematsu, Kazuo; Yamamoto, Taira; Inaba, Hirotaka; Kuwaki, Kenji; Shimada, Akie; Yokoyama, Yasutaka; Amano, Atsushi

2017-12-12

We determined the factors associated with the expression of c-kit in the heart and the proliferation of c-kit-positive (c-kit pos ) cardiac stem cells among the outgrowth cells cultured from human cardiac explants.Samples of the right atrium (RA), left atrium (LA), and left ventricle obtained from patients during open-heart surgery were processed for cell culture of outgrowth cells and tissue analysis. The total number of growing cells and the population of c-kit pos cells were measured and compared with c-kit expression in native tissues and characteristics of the patients according to the region of the heart.We analyzed 452 samples from 334 patients. Atrial fibrillation (AF) in the patients reduced the number of outgrowth cells from the RA and LA, and aging was a co-factor for the LA. The c-kit pos population from the RA was associated with serum brain natriuretic peptide (BNP). C-kit expression in native tissue was also associated with BNP expression. However, we observed no relationship in expression between outgrowth cells and native tissue. In addition, the RA tissue provided the highest number of c-kit pos cells, and the left ventricle provided the lowest.C-kit was weakly expressed in response to damage. In addition, no correlation between outgrowth cells and native tissue was found for c-kit expression.

Novel biomarker identification using metabolomic profiling to differentiate radiation necrosis and recurrent tumor following Gamma Knife radiosurgery.

PubMed

Lu, Alex Y; Turban, Jack L; Damisah, Eyiyemisi C; Li, Jie; Alomari, Ahmed K; Eid, Tore; Vortmeyer, Alexander O; Chiang, Veronica L

2017-08-01

OBJECTIVE Following an initial response of brain metastases to Gamma Knife radiosurgery, regrowth of the enhancing lesion as detected on MRI may represent either radiation necrosis (a treatment-related inflammatory change) or recurrent tumor. Differentiation of radiation necrosis from tumor is vital for management decision making but remains difficult by imaging alone. In this study, gas chromatography with time-of-flight mass spectrometry (GC-TOF) was used to identify differential metabolite profiles of the 2 tissue types obtained by surgical biopsy to find potential targets for noninvasive imaging. METHODS Specimens of pure radiation necrosis and pure tumor obtained from patient brain biopsies were flash-frozen and validated histologically. These formalin-free tissue samples were then analyzed using GC-TOF. The metabolite profiles of radiation necrosis and tumor samples were compared using multivariate and univariate statistical analysis. Statistical significance was defined as p ≤ 0.05. RESULTS For the metabolic profiling, GC-TOF was performed on 7 samples of radiation necrosis and 7 samples of tumor. Of the 141 metabolites identified, 17 (12.1%) were found to be statistically significantly different between comparison groups. Of these metabolites, 6 were increased in tumor, and 11 were increased in radiation necrosis. An unsupervised hierarchical clustering analysis found that tumor had elevated levels of metabolites associated with energy metabolism, whereas radiation necrosis had elevated levels of metabolites that were fatty acids and antioxidants/cofactors. CONCLUSIONS To the authors' knowledge, this is the first tissue-based metabolomics study of radiation necrosis and tumor. Radiation necrosis and recurrent tumor following Gamma Knife radiosurgery for brain metastases have unique metabolite profiles that may be targeted in the future to develop noninvasive metabolic imaging techniques.

Study of the intra-arterial distribution of Fe₃O₄ nanoparticles in a model of colorectal neoplasm induced in rat liver by MRI and spectrometry.

PubMed

Echevarria-Uraga, José J; García-Alonso, Ignacio; Plazaola, Fernando; Insausti, Maite; Etxebarria, Néstor; Saiz-López, Alberto; Fernández-Ruanova, Begoña

2012-01-01

To evaluate, in an experimental model, the reliability of MRI for determining whether a higher iron concentration was obtained in tumor tissue than in normal liver parenchyma after intra-arterial administration of Fe₃O₄ lipophilic nanoparticles. WAG/RijCrl rats were inoculated in the left hepatic lobe with 25,000 syngeneic CC-531 colon adenocarcinoma cells, after which they were randomized into two groups: control (CG) and infused (IG). After confirming tumor induction, the IG rats received intra-arterial suspensions of Fe₃O₄ nanoparticles (2.6 mg) in Lipiodol® (0.15 mL). To calculate the iron concentration, [Fe], in the tumor and liver tissues of both groups of rats, measurements of signal intensity from the tumors, healthy liver tissue, and paravertebral muscles were made on a 1.5T MRI system in gradient-echo DP* and T2*-weighted sequences. In addition, samples were collected to quantify the [Fe] by inductively coupled plasma-mass spectrometry (ICP-MS), as well as for histological analysis. Statistical analysis was performed with non-parametric tests, and Bland-Altman plots were produced; P values <0.05 were considered significant. In the CG rats (n = 23), the mean [Fe] values estimated by MRI and ICP-MS were 13.2 μmol·g⁻¹ and 5.9 μmol·g⁻¹, respectively, in the tumors, and 19.0 μmol ·g⁻¹ and 11.7 μmol·g⁻¹, respectively, in the hepatic tissue. In the IG rats (n = 19), the values obtained by MRI and ICP-MS were 148.9 μmol·g⁻¹ and 9.4 μmol · g⁻¹, respectively, in the tumors, and 115.3 μmol·g⁻¹ and 11.6 μmol·g⁻¹, respectively, in the healthy liver tissue. The IG results revealed a clear disagreement between MRI and ICP-MS. In the comparative analysis between the groups regarding the [Fe] values obtained by ICP-MS, significant differences were found for the tumor samples (P < 0.001), but not for the hepatic tissue (P = 0.92). Under microscopy, scattered intravascular deposits of nanoparticles were observed, especially in the tumors. ICP-MS demonstrated significant uptake of exogenous iron in tumor tissue. MRI was useful for quantifying the [Fe] in the different tissues in the CG animals, but not in the IG animals. Although the irregular distribution of nanoparticles caused an important bias in the measurements obtained by MRI, the relative increase in iron content inside the tumor was suggested.

Study of the intra-arterial distribution of Fe3O4 nanoparticles in a model of colorectal neoplasm induced in rat liver by MRI and spectrometry

PubMed Central

Echevarria-Uraga, José J; García-Alonso, Ignacio; Plazaola, Fernando; Insausti, Maite; Etxebarria, Néstor; Saiz-López, Alberto; Fernández-Ruanova, Begoña

2012-01-01

Purpose To evaluate, in an experimental model, the reliability of MRI for determining whether a higher iron concentration was obtained in tumor tissue than in normal liver parenchyma after intra-arterial administration of Fe3O4 lipophilic nanoparticles. Materials and methods WAG/RijCrl rats were inoculated in the left hepatic lobe with 25,000 syngeneic CC-531 colon adenocarcinoma cells, after which they were randomized into two groups: control (CG) and infused (IG). After confirming tumor induction, the IG rats received intra-arterial suspensions of Fe3O4 nanoparticles (2.6 mg) in Lipiodol® (0.15 mL). To calculate the iron concentration, [Fe], in the tumor and liver tissues of both groups of rats, measurements of signal intensity from the tumors, healthy liver tissue, and paravertebral muscles were made on a 1.5T MRI system in gradient-echo DP* and T2*-weighted sequences. In addition, samples were collected to quantify the [Fe] by inductively coupled plasma-mass spectrometry (ICP-MS), as well as for histological analysis. Statistical analysis was performed with non-parametric tests, and Bland–Altman plots were produced; P values <0.05 were considered significant. Results In the CG rats (n = 23), the mean [Fe] values estimated by MRI and ICP-MS were 13.2 μmol·g−1 and 5.9 μmol·g−1, respectively, in the tumors, and 19.0μmol ·g−1 and 11.7 μmol·g−1, respectively, in the hepatic tissue. In the IG rats (n = 19), the values obtained by MRI and ICP-MS were 148.9 μmol·g−1 and 9.4 μmol · g−1, respectively, in the tumors, and 115.3 μmol·g−1 and 11.6 μmol·g−1, respectively, in the healthy liver tissue. The IG results revealed a clear disagreement between MRI and ICP-MS. In the comparative analysis between the groups regarding the [Fe] values obtained by ICP-MS, significant differences were found for the tumor samples (P < 0.001), but not for the hepatic tissue (P = 0.92). Under microscopy, scattered intravascular deposits of nanoparticles were observed, especially in the tumors. Conclusion ICP-MS demonstrated significant uptake of exogenous iron in tumor tissue. MRI was useful for quantifying the [Fe] in the different tissues in the CG animals, but not in the IG animals. Although the irregular distribution of nanoparticles caused an important bias in the measurements obtained by MRI, the relative increase in iron content inside the tumor was suggested. PMID:22661893

A preliminary result of three-dimensional microarray technology to gene analysis with endoscopic ultrasound-guided fine-needle aspiration specimens and pancreatic juices

PubMed Central

2010-01-01

Background Analysis of gene expression and gene mutation may add information to be different from ordinary pathological tissue diagnosis. Since samples obtained endoscopically are very small, it is desired that more sensitive technology is developed for gene analysis. We investigated whether gene expression and gene mutation analysis by newly developed ultra-sensitive three-dimensional (3D) microarray is possible using small amount samples from endoscopic ultrasound-guided fine-needle aspiration (EUS-FNA) specimens and pancreatic juices. Methods Small amount samples from 17 EUS-FNA specimens and 16 pancreatic juices were obtained. After nucleic acid extraction, the samples were amplified with labeling and analyzed by the 3D microarray. Results The analyzable rate with the microarray was 46% (6/13) in EUS-FNA specimens of RNAlater® storage, and RNA degradations were observed in all the samples of frozen storage. In pancreatic juices, the analyzable rate was 67% (4/6) in frozen storage samples and 20% (2/10) in RNAlater® storage. EUS-FNA specimens were classified into cancer and non-cancer by gene expression analysis and K-ras codon 12 mutations were also detected using the 3D microarray. Conclusions Gene analysis from small amount samples obtained endoscopically was possible by newly developed 3D microarray technology. High quality RNA from EUS-FNA samples were obtained and remained in good condition only using RNA stabilizer. In contrast, high quality RNA from pancreatic juice samples were obtained only in frozen storage without RNA stabilizer. PMID:20416107

LASER METHODS IN BIOLOGY: Optical anisotropy of fibrous biological tissues: analysis of the influence of structural properties

NASA Astrophysics Data System (ADS)

Zimnyakov, D. A.; Sinichkin, Yu P.; Ushakova, O. V.

2007-08-01

The results of theoretical analysis of the optical anisotropy of multiply scattering fibrillar biological tissues based on the model of an effective anisotropic medium are compared with the experimental in vivo birefringence data for the rat derma obtained earlier in spectral polarisation measurements of rat skin samples in the visible region. The disordered system of parallel dielectric cylinders embedded into an isotropic dielectric medium was considered as a model medium. Simulations were performed taking into account the influence of a partial mutual disordering of the bundles of collagen and elastin fibres in derma on birefringence in samples. The theoretical optical anisotropy averaged over the spectral interval 550-650 nm for the model medium with parameters corresponding to the structural parameters of derma is in good agreement with the results of spectral polarisation measurements of skin samples in the corresponding wavelength range.

Experiment kits for processing biological samples inflight on SLS-2

NASA Technical Reports Server (NTRS)

Savage, P. D.; Hinds, W. E.; Jaquez, R.; Evans, J.; Dubrovin, L.

1995-01-01

This paper describes development of an innovative, modular approach to packaging the instruments used to obtain and preserve the inflight rodent tissue and blood samples associated with hematology experiments on the Spacelab Life Sciences-2 (SLS-2) mission. The design approach organized the multitude of instruments into twelve 5- x 6- x l-in. kits which were each used for a particular experiment. Each kit contained the syringes, vials, microscope slides, etc., necessary for processing and storing blood and tissue samples for one rat on a particular day. A total of 1245 components, packaged into 128 kits and stowed in 17 Zero(registered trademark) boxes, were required. Crewmembers found the design easy to use and laid out in a logical, simple configuration which minimized chances for error during the complex procedures in flight. This paper also summarizes inflight performance of the kits on SLS-2.

High Resolution Tissue Imaging Using the Single-probe Mass Spectrometry under Ambient Conditions

NASA Astrophysics Data System (ADS)

Rao, Wei; Pan, Ning; Yang, Zhibo

2015-06-01

Ambient mass spectrometry imaging (MSI) is an emerging field with great potential for the detailed spatial analysis of biological samples with minimal pretreatment. We have developed a miniaturized sampling and ionization device, the Single-probe, which uses in-situ surface micro-extraction to achieve high detection sensitivity and spatial resolution during MSI experiments. The Single-probe was coupled to a Thermo LTQ Orbitrap XL mass spectrometer and was able to create high spatial and high mass resolution MS images at 8 ± 2 and 8.5 μm on flat polycarbonate microscope slides and mouse kidney sections, respectively, which are among the highest resolutions available for ambient MSI techniques. Our proof-of-principle experiments indicate that the Single-probe MSI technique has the potential to obtain ambient MS images with very high spatial resolutions with minimal sample preparation, which opens the possibility for subcellular ambient tissue MSI to be performed in the future.

A method to quantify at late imaging a release rate of 18F-FDG in tissues.

PubMed

Laffon, Eric; Allard, Michèle; Marthan, Roger; Ducassou, Dominique

2005-08-01

This theoretical work shows that the rate constant for the (18)F-FDG release in tissues can be assessed without needing any arterial blood sampling. The method requires that the clearance of (18)F-FDG from plasma has occurred, whereas (18)F-FDG is still present in the tissue. This condition can be met dating from 3 h after (18)F-FDG injection, when hydration and/or phlorizin injection are applied after the routine static acquisition. The release rate constant can be obtained from a graphical analysis performed at the later decreasing phase of the tissue tracer activity. A two-compartment and a three-compartment model are developed, both in accordance with one another. To cite this article: E. Laffon et al., C. R. Biologies 328 (2005).

Detection and typing of human papillomaviruses (HPV) in malignant, dysplastic, nondysplastic and normal oral epithelium by nested polymerase chain reaction, immunohistochemistry and transitional electron microscopy in patients of northern Greece.

PubMed

Blioumi, E; Chatzidimitriou, D; Pazartzi, Ch; Katopodi, Th; Tzimagiorgis, G; Emmanouil-Nikoloussi, E-N; Markopoulos, A; Kalekou, C; Lazaridis, N; Diza, E; Antoniades, D

2014-09-01

To evaluate the role of HPV in oral carcinogenesis, we examined the prevalence of HPV in malignant, potentially malignant and normal oral epithelium and studied the relation of HPV prevalence with other factors obtained from the patient's records. Our material consisted of 291 tissue specimens from 258 individuals. From every individual formalin fixed and paraffin embedded tissues were examined by nested Polymerase Chain Reaction (NPCR) for the detection of HPV DNA and by immunohistochemistry (IHC) for the in situ detection of HPV L1 protein. Positive PCR products were sequenced in order to type HPVs. Also 33 fresh tissues were obtained, fixed and used to detect HPV particles by transitional electron microscopy (TEM). HPV was detected in 32.9% of the tissue specimens by NPCR, in 4.7% by immunohistochemistry and in 28.1% by TEM. In detail, by nested PCR HPV L1 DNA was detected in 40% of normal tissues, 40% of fibromas, 35.8% of non-dysplastic leukoplakias, 31.6% of dysplastic leukoplakias and 22.2% of oral squamous cell carcinomas. The HPV viral load of 96.5% of the samples was very low (1 viral copy per 10(2)-10(4) cells). HPV16 prevails in all histological groups in 89-100%. We conclude that HPV does not seem, from the specific sample examined, to play a substantial role in oral carcinogenesis. However, it cannot be excluded that HPV could be involved in oral carcinogenesis only in cases with high viral load or at early stages of carcinogenesis possibly through the hit-and-run mechanism. Copyright © 2014 Elsevier Ltd. All rights reserved.

Investigation of procalcitonin, IL-6, oxidative stress index (OSI) plasma and tissue levels in experimental mild and severe pancreatitis in rats.

PubMed

Soyalp, M; Yalcin, M; Oter, V; Ozgonul, A

2017-01-01

This study was planned to evaluate blood and tissue levels of procalcitonin, IL-6 and Oxidative Stress Index (OSI), which have a role in the pathophysiology of inflammation, in rats with experimental mild and severe pancreatitis. Thirty male Wistar Albino rats were included in the study and the rats were equally divided into the three groups. After 1, 5 and 24 hours, blood was obtained from the tail of all rats and samples were taken from pancreas tissue after 24 hours. Amylase, lipase, AST, ALT, WBC, LDH, glucose, total bilirubin, direct bilirubin, GGT, ALP and TNF-α, Procalcitonin and IL-6 levels were evaluated from blood and tissue specimens. The mild pancreatitis group (group 1) and the severe pancreatitis group (group 3) were compared according to the OSI, Amylase, Lipase, Pct, IL-6, AST, ALT, Glucose, WBC, LDH and Tnf-α levels. In the severe pancreatitis group, a statistically significant increase was detected compared to the mild pancreatitis group (p < 0.05). There was no statistically significant difference in TOS, T.Bil, D.Bil, GGT and ALP values (p > 0.05). Statistically significant increase was observed in severe pancreatitis group according to OSI, Amylase, Lipase, Pct, IL-6, LDH, WBC and TNF-α levels samples obtained from pancreatic tissues, compared to mild pancreatitis group (p < 0.05). In conclusion, our study showed that OSI, TNF-α, IL-6 and procalcitonin levels increases proportionally with the severity of pancreatitis. This also suggests that OSI, TNF-α, IL-6 and procalcitonin are the guiding substances in acute pancreatitis, and that this damage increases as the duration and severity of the pathological process increase. However, this result must be confirmed with more detailed and broadly planned future studies (Fig. 3, Ref. 25).

Diamond, titanium dioxide, titanium silicon oxide, and barium strontium titanium oxide nanoparticles as matrixes for direct matrix-assisted laser desorption/ionization mass spectrometry analysis of carbohydrates in plant tissues.

PubMed

Gholipour, Yousef; Giudicessi, Silvana L; Nonami, Hiroshi; Erra-Balsells, Rosa

2010-07-01

Nanoparticles (NPs) of diamond, titanium dioxide, titanium silicon oxide, barium strontium titanium oxide, and silver (Ag) were examined for their potential as MALDI matrixes for direct laser desorption/ionization of carbohydrates, especially fructans, from plant tissue. Two sample preparation methods including solvent-assisted and solvent-free (dry) NPs deposition were performed and compared. All examined NPs except for Ag could desorb/ionize standard sucrose and fructans in positive and in negative ion mode. Ag NPs yielded good signals only for nonsalt-doped samples that were measured in the negative ion mode. In the case of in vivo studies, except for Ag, all NPs studied could desorb/ionize carbohydrates from tissue in both the positive and negative ion modes. Furthermore, compared to the results obtained with soluble sugars extracted from plant tissues, fructans with higher molecular weight intact molecular ions could be detected when the plant tissues were directly profiled. The limit of detection (LOD) of fructans and the ratios between signal intensities and fructan concentrations were analyzed. NPs had similar LODs for standard fructan triose (1-kestose) in the positive ion mode and better LODs in the negative ion mode when compared with the common crystalline organic MALDI matrixes used for carbohydrates (2,5-dihydroxybenzoic acid and nor-harmane) or carbon nanotubes. Solvent-free NP deposition on tissues partially improves the signal acquisition. Although lower signal-to-noise ratio sugar signals were acquired from the tissues when compared to the solvent-assisted method, the reproducibility averaged over all sample was more uniform.

A naive Bayes algorithm for tissue origin diagnosis (TOD-Bayes) of synchronous multifocal tumors in the hepatobiliary and pancreatic system.

PubMed

Jiang, Weiqin; Shen, Yifei; Ding, Yongfeng; Ye, Chuyu; Zheng, Yi; Zhao, Peng; Liu, Lulu; Tong, Zhou; Zhou, Linfu; Sun, Shuo; Zhang, Xingchen; Teng, Lisong; Timko, Michael P; Fan, Longjiang; Fang, Weijia

2018-01-15

Synchronous multifocal tumors are common in the hepatobiliary and pancreatic system but because of similarities in their histological features, oncologists have difficulty in identifying their precise tissue clonal origin through routine histopathological methods. To address this problem and assist in more precise diagnosis, we developed a computational approach for tissue origin diagnosis based on naive Bayes algorithm (TOD-Bayes) using ubiquitous RNA-Seq data. Massive tissue-specific RNA-Seq data sets were first obtained from The Cancer Genome Atlas (TCGA) and ∼1,000 feature genes were used to train and validate the TOD-Bayes algorithm. The accuracy of the model was >95% based on tenfold cross validation by the data from TCGA. A total of 18 clinical cancer samples (including six negative controls) with definitive tissue origin were subsequently used for external validation and 17 of the 18 samples were classified correctly in our study (94.4%). Furthermore, we included as cases studies seven tumor samples, taken from two individuals who suffered from synchronous multifocal tumors across tissues, where the efforts to make a definitive primary cancer diagnosis by traditional diagnostic methods had failed. Using our TOD-Bayes analysis, the two clinical test cases were successfully diagnosed as pancreatic cancer (PC) and cholangiocarcinoma (CC), respectively, in agreement with their clinical outcomes. Based on our findings, we believe that the TOD-Bayes algorithm is a powerful novel methodology to accurately identify the tissue origin of synchronous multifocal tumors of unknown primary cancers using RNA-Seq data and an important step toward more precision-based medicine in cancer diagnosis and treatment. © 2017 UICC.

A comparison of liver sampling techniques in dogs.

PubMed

Kemp, S D; Zimmerman, K L; Panciera, D L; Monroe, W E; Leib, M S; Lanz, O I

2015-01-01

The liver sampling technique in dogs that consistently provides samples adequate for accurate histopathologic interpretation is not known. To compare histopathologic results of liver samples obtained by punch, cup, and 14 gauge needle to large wedge samples collected at necropsy. Seventy dogs undergoing necropsy. Prospective study. Liver specimens were obtained from the left lateral liver lobe with an 8 mm punch, a 5 mm cup, and a 14 gauge needle. After sample acquisition, two larger tissue samples were collected near the center of the left lateral lobe to be used as a histologic standard for comparison. Histopathologic features and numbers of portal triads in each sample were recorded. The mean number of portal triads obtained by each sampling method were 2.9 in needle samples, 3.4 in cup samples, 12 in punch samples, and 30.7 in the necropsy samples. The diagnoses in 66% of needle samples, 60% of cup samples, and 69% of punch samples were in agreement with the necropsy samples, and these proportions were not significantly different from each other. The corresponding kappa coefficients were 0.59 for needle biopsies, 0.52 for cup biopsies, and 0.62 for punch biopsies. The histopathologic interpretation of a liver sample in the dog is unlikely to vary if the liver biopsy specimen contains at least 3-12 portal triads. However, in comparison large necropsy samples, the accuracy of all tested methods was relatively low. Copyright © 2014 by the American College of Veterinary Internal Medicine.

Identification and validation of differentially expressed transcripts by RNA-sequencing of formalin-fixed, paraffin-embedded (FFPE) lung tissue from patients with Idiopathic Pulmonary Fibrosis.

PubMed

Vukmirovic, Milica; Herazo-Maya, Jose D; Blackmon, John; Skodric-Trifunovic, Vesna; Jovanovic, Dragana; Pavlovic, Sonja; Stojsic, Jelena; Zeljkovic, Vesna; Yan, Xiting; Homer, Robert; Stefanovic, Branko; Kaminski, Naftali

2017-01-12

Idiopathic Pulmonary Fibrosis (IPF) is a lethal lung disease of unknown etiology. A major limitation in transcriptomic profiling of lung tissue in IPF has been a dependence on snap-frozen fresh tissues (FF). In this project we sought to determine whether genome scale transcript profiling using RNA Sequencing (RNA-Seq) could be applied to archived Formalin-Fixed Paraffin-Embedded (FFPE) IPF tissues. We isolated total RNA from 7 IPF and 5 control FFPE lung tissues and performed 50 base pair paired-end sequencing on Illumina 2000 HiSeq. TopHat2 was used to map sequencing reads to the human genome. On average ~62 million reads (53.4% of ~116 million reads) were mapped per sample. 4,131 genes were differentially expressed between IPF and controls (1,920 increased and 2,211 decreased (FDR < 0.05). We compared our results to differentially expressed genes calculated from a previously published dataset generated from FF tissues analyzed on Agilent microarrays (GSE47460). The overlap of differentially expressed genes was very high (760 increased and 1,413 decreased, FDR < 0.05). Only 92 differentially expressed genes changed in opposite directions. Pathway enrichment analysis performed using MetaCore confirmed numerous IPF relevant genes and pathways including extracellular remodeling, TGF-beta, and WNT. Gene network analysis of MMP7, a highly differentially expressed gene in both datasets, revealed the same canonical pathways and gene network candidates in RNA-Seq and microarray data. For validation by NanoString nCounter® we selected 35 genes that had a fold change of 2 in at least one dataset (10 discordant, 10 significantly differentially expressed in one dataset only and 15 concordant genes). High concordance of fold change and FDR was observed for each type of the samples (FF vs FFPE) with both microarrays (r = 0.92) and RNA-Seq (r = 0.90) and the number of discordant genes was reduced to four. Our results demonstrate that RNA sequencing of RNA obtained from archived FFPE lung tissues is feasible. The results obtained from FFPE tissue are highly comparable to FF tissues. The ability to perform RNA-Seq on archived FFPE IPF tissues should greatly enhance the availability of tissue biopsies for research in IPF.

Whole slide imaging of unstained tissue using lensfree microscopy

NASA Astrophysics Data System (ADS)

Morel, Sophie Nhu An; Hervé, Lionel; Bordy, Thomas; Cioni, Olivier; Delon, Antoine; Fromentin, Catherine; Dinten, Jean-Marc; Allier, Cédric

2016-04-01

Pathologist examination of tissue slides provides insightful information about a patient's disease. Traditional analysis of tissue slides is performed under a binocular microscope, which requires staining of the sample and delays the examination. We present a simple cost-effective lensfree imaging method to record 2-4μm resolution wide-field (10 mm2 to 6 cm2) images of unstained tissue slides. The sample processing time is reduced as there is no need for staining. A wide field of view (10 mm2) lensfree hologram is recorded in a single shot and the image is reconstructed in 2s providing a very fast acquisition chain. The acquisition is multispectral, i.e. multiple holograms are recorded simultaneously at three different wavelengths, and a dedicated holographic reconstruction algorithm is used to retrieve both amplitude and phase. Whole tissue slides imaging is obtained by recording 130 holograms with X-Y translation stages and by computing the mosaic of a 25 x 25 mm2 reconstructed image. The reconstructed phase provides a phase-contrast-like image of the unstained specimen, revealing structures of healthy and diseased tissue. Slides from various organs can be reconstructed, e.g. lung, colon, ganglion, etc. To our knowledge, our method is the first technique that enables fast wide-field lensfree imaging of such unlabeled dense samples. This technique is much cheaper and compact than a conventional phase contrast microscope and could be made portable. In sum, we present a new methodology that could quickly provide useful information when a rapid diagnosis is needed, such as tumor margin identification on frozen section biopsies during surgery.

Real time analysis of brain tissue by direct combination of ultrasonic surgical aspiration and sonic spray mass spectrometry.

PubMed

Schäfer, Karl-Christian; Balog, Júlia; Szaniszló, Tamás; Szalay, Dániel; Mezey, Géza; Dénes, Júlia; Bognár, László; Oertel, Matthias; Takáts, Zoltán

2011-10-15

Direct combination of cavitron ultrasonic surgical aspirator (CUSA) and sonic spray ionization mass spectrometry is presented. A commercially available ultrasonic surgical device was coupled to a Venturi easy ambient sonic-spray ionization (V-EASI) source by directly introducing liquified tissue debris into the Venturi air jet pump. The Venturi air jet pump was found to efficiently nebulize the suspended tissue material for gas phase ion production. The ionization mechanism involving solely pneumatic spraying was associated with that of sonic spray ionization. Positive and negative ionization spectra were obtained from brain and liver samples reflecting the primary application areas of the surgical device. Mass spectra were found to feature predominantly complex lipid-type constituents of tissues in both ion polarity modes. Multiply charged peptide anions were also detected. The influence of instrumental settings was characterized in detail. Venturi pump geometry and flow parameters were found to be critically important in ionization efficiency. Standard solutions of phospholipids and peptides were analyzed in order to test the dynamic range, sensitivity, and suppression effects. The spectra of the intact tissue specimens were found to be highly specific to the histological tissue type. The principal component analysis (PCA) and linear discriminant analysis (LDA) based data analysis method was developed for real-time tissue identification in a surgical environment. The method has been successfully tested on post-mortem and ex vivo human samples including astrocytomas, meningeomas, metastatic brain tumors, and healthy brain tissue. © 2011 American Chemical Society

Inorganic elements in green sea turtles (Chelonia mydas): relationships among external and internal tissues.

PubMed

Faust, Derek R; Hooper, Michael J; Cobb, George P; Barnes, Melanie; Shaver, Donna; Ertolacci, Shauna; Smith, Philip N

2014-09-01

Inorganic elements from anthropogenic sources have entered marine environments worldwide and are detectable in marine organisms, including sea turtles. Threatened and endangered classifications of sea turtles have heretofore made assessments of contaminant concentrations difficult because of regulatory restrictions on obtaining samples using nonlethal techniques. In the present study, claw and skin biopsy samples were examined as potential indicators of internal tissue burdens in green sea turtles (Chelonia mydas). Significant relationships were observed between claw and liver, and claw and muscle concentrations of mercury, nickel, arsenic, and selenium (p < 0.05). Similarly, significant relationships were observed between skin biopsy concentrations and those in liver, kidney, and muscle tissues for mercury, arsenic, selenium, and vanadium (p < 0.05). Concentrations of arsenic, barium, chromium, nickel, strontium, vanadium, and zinc in claws and skin biopsies were substantially elevated when compared with all other tissues, indicating that these highly keratinized tissues may represent sequestration or excretion pathways. Correlations between standard carapace length and cobalt, lead, and manganese concentrations were observed (p < 0.05), indicating that tissue concentrations of these elements may be related to age and size. Results suggest that claws may indeed be useful indicators of mercury and nickel concentrations in liver and muscle tissues, whereas skin biopsy inorganic element concentrations may be better suited as indicators of mercury, selenium, and vanadium concentrations in liver, kidney, and muscle tissues of green sea turtles. © 2014 SETAC.

Tissue temperature distribution measurement by MRI and laser immunology for cancer treatment

NASA Astrophysics Data System (ADS)

Chen, Yichao; Gnyawali, Surya C.; Wu, Feng; Liu, Hong; Tesiram, Yasvir A.; Abbott, Andrew; Towner, Rheal A.; Chen, Wei R.

2007-02-01

In cancer treatment and immune response enhancement research, Magnetic Resonance Imaging (MRI) is an ideal method for non-invasive, three-dimensional temperature measurement. We used a 7.1-Tesla magnetic resonance imager for ex vivo tissues and small animal to determine temperature distribution of target tissue during laser irradiation. The feasibility of imaging is approved with high spatial resolution and high signal-noise- ratio. Tissue-simulating gel phantom gel, biological tissues, and tumor-bearing animals were used in the experiments for laser treatment and MR imaging. Thermal couple measurement of temperature in target samples was used for system calibration. An 805-nm laser was used to irradiate the samples with a laser power in the range of 1 to 2.5 watts. Using the MRI system and a specially developed processing algorithm, a clear temperature distribution matrix in the target tissue and surrounding tissue was obtained. The temperature profiles show that the selective laser photothermal effect could result in tissue temperature elevation in a range of 10 to 45 °C. The temperature resolution of the measurement was about 0.37°C including the total system error. The spatial resolution was 0.4 mm (128x128 pixels with field of view of 5.5x5.5 cm). The temperature distribution provided in vivo thermal information and future reference for optimizing dye concentration and irradiation parameters to achieve optimal thermal effects in cancer treatment.

Optical coherence elastography (OCE) as a method for identifying benign and malignant prostate biopsies

NASA Astrophysics Data System (ADS)

Li, Chunhui; Guan, Guangying; Ling, Yuting; Lang, Stephen; Wang, Ruikang K.; Huang, Zhihong; Nabi, Ghulam

2015-03-01

Objectives. Prostate cancer is the most frequently diagnosed malignancy in men. Digital rectal examination (DRE) - a known clinical tool based on alteration in the mechanical properties of tissues due to cancer has traditionally been used for screening prostate cancer. Essentially, DRE estimates relative stiffness of cancerous and normal prostate tissue. Optical coherence elastography (OCE) are new optical imaging techniques capable of providing cross-sectional imaging of tissue microstructure as well as elastogram in vivo and in real time. In this preliminary study, OCE was used in the setting of the human prostate biopsies ex vivo, and the images acquired were compared with those obtained using standard histopathologic methods. Methods. 120 prostate biopsies were obtained by TRUS guided needle biopsy procedures from 9 patients with clinically suspected cancer of the prostate. The biopsies were approximately 0.8mm in diameter and 12mm in length, and prepared in Formalin solution. Quantitative assessment of biopsy samples using OCE was obtained in kilopascals (kPa) before histopathologic evaluation. The results obtained from OCE and standard histopathologic evaluation were compared provided the cross-validation. Sensitivity, specificity, and positive and negative predictive values were calculated for OCE (histopathology was a reference standard). Results. OCE could provide quantitative elasticity properties of prostate biopsies within benign prostate tissue, prostatic intraepithelial neoplasia, atypical hyperplasia and malignant prostate cancer. Data analysed showed that the sensitivity and specificity of OCE for PCa detection were 1 and 0.91, respectively. PCa had significantly higher stiffness values compared to benign tissues, with a trend of increasing in stiffness with increasing of malignancy. Conclusions. Using OCE, microscopic resolution elastogram is promising in diagnosis of human prostatic diseases. Further studies using this technique to improve the detection and staging of malignant cancer of the prostate are ongoing.

THE PASSAGE OF MARKED IONS INTO TISSUES AND FLUIDS OF THE REPRODUCTIVE TRACT OF PREGNANT RABBITS

DOE Office of Scientific and Technical Information (OSTI.GOV)

Bennett, J.P.; Boursnell, J.C.; Lutwak-Mann, C.

1959-10-31

Rapid changes were demonstrated in the uptake of labeled ions both in the developing embryo and in the endometrium, mesodermal placental folds, and other closely associated tissues and fluids following the intravenous injection of labeled ions in pregnant rabbits. Phosphorus-32, sulfur-85, sodium-24, iodine- 131, and potassium-42 were used as tracers. A number of new techniques were developed to obtain, weigh, and handle the extremely small samples. The influence of exogenous materials on the early development of fetuses is discussed briefly. (C.R.)

Low-grade infection after a total knee arthroplasty caused by Actinomyces naeslundii.

PubMed

Hedke, J; Skripitz, R; Ellenrieder, M; Frickmann, H; Köller, T; Podbielski, A; Mittelmeier, W

2012-08-01

Here, we present a case of an 85-year-old woman with a low-grade-infection caused by Actinomyces naeslundii after total-knee arthroplasty (TKA) followed by septic loosening. Actinomyces naeslundii was cultured from a tissue sample from the knee joint capsule/synovial tissue obtained after the initial TKA. A review of the literature revealed two cases of periprosthetic infection and another three cases of arthritis due to Actinomyces naeslundii. So far, no standard treatment for periprosthetic infections caused by Actinomyces species has been established.

Id-1 and Id-2 genes and products as markers of epithelial cancer

DOEpatents

Desprez, Pierre-Yves [El Cerrito, CA; Campisi, Judith [Berkeley, CA

2008-09-30

A method for detection and prognosis of breast cancer and other types of cancer. The method comprises detecting expression, if any, for both an Id-1 and an Id-2 genes, or the ratio thereof, of gene products in samples of breast tissue obtained from a patient. When expressed, Id-1 gene is a prognostic indicator that breast cancer cells are invasive and metastatic, whereas Id-2 gene is a prognostic indicator that breast cancer cells are localized and noninvasive in the breast tissue.

Id-1 and Id-2 genes and products as markers of epithelial cancer

DOEpatents

Desprez, Pierre-Yves [El Cerrito, CA; Campisi, Judith [Berkeley, CA

2011-10-04

A method for detection and prognosis of breast cancer and other types of cancer. The method comprises detecting expression, if any, for both an Id-1 and an Id-2 genes, or the ratio thereof, of gene products in samples of breast tissue obtained from a patient. When expressed, Id-1 gene is a prognostic indicator that breast cancer cells are invasive and metastatic, whereas Id-2 gene is a prognostic indicator that breast cancer cells are localized and noninvasive in the breast tissue.

Hard tissue as a composite material. I - Bounds on the elastic behavior.

NASA Technical Reports Server (NTRS)

Katz, J. L.

1971-01-01

Recent determination of the elastic moduli of hydroxyapatite by ultrasonic methods permits a re-examination of the Voigt or parallel model of the elastic behavior of bone, as a two phase composite material. It is shown that such a model alone cannot be used to describe the behavior of bone. Correlative data on the elastic moduli of dentin, enamel and various bone samples indicate the existence of a nonlinear dependence of elastic moduli on composition of hard tissue. Several composite models are used to calculate the bounds on the elastic behavior of these tissues. The limitations of these models are described, and experiments to obtain additional critical data are discussed.

Imaging the hard/soft tissue interface.

PubMed

Bannerman, Alistair; Paxton, Jennifer Z; Grover, Liam M

2014-03-01

Interfaces between different tissues play an essential role in the biomechanics of native tissues and their recapitulation is now recognized as critical to function. As a consequence, imaging the hard/soft tissue interface has become increasingly important in the area of tissue engineering. Particularly as several biotechnology based products have made it onto the market or are close to human trials and an understanding of their function and development is essential. A range of imaging modalities have been developed that allow a wealth of information on the morphological and physical properties of samples to be obtained non-destructively in vivo or via destructive means. This review summarizes the use of a selection of imaging modalities on interfaces to date considering the strengths and weaknesses of each. We will also consider techniques which have not yet been utilized to their full potential or are likely to play a role in future work in the area.

Aescin formation in calli and embryoids from cotyledon and stem explants of Aesculus hippocastanum L.

PubMed

Profumo, P; Caviglia, A M; Gastaldo, P

1994-11-01

Aescin in calli and embryoids obtained from both cotyledon and stem explants of Aesculus hippocastanum were investigated by HPLC. Determinations were carried out on tissues cultured in agarized medium supplemented with growth substances (2,4-dichlorophenoxyacetic acid; kinetin; 1-naphthaleneacetic acid). The results indicate that aescin was produced in all the analysed samples. The amount of active principle present in some samples was higher than that found in horse-chestnut seeds.

Monitoring corneal crosslinking using phase-decorrelation OCT (Conference Presentation)

NASA Astrophysics Data System (ADS)

Blackburn, Brecken J.; Gu, Shi; Jenkins, Michael W.; Rollins, Andrew M.

2017-02-01

Viscosity is often a critical characteristic of biological fluids such as blood and mucus. However, traditional rheology is often inadequate when only small quantities of sample are available. A robust method to measure viscosity of microquantities of biological samples could lead to a better understanding and diagnosis of diseases. Here, we present a method to measure viscosity by observing particle Brownian motion within a sample. M-mode optical coherence tomography (OCT) imaging, obtained with a phase-sensitive 47 kHz spectral domain system, yields a viscosity measurement from multiple 200-1000 microsecond frames. This very short period of continuous acquisition, as compared to laser speckle decorrelation, decreases sensitivity to bulk motion, thereby potentially enabling in vivo and in situ applications. The theory linking g(1) first-order image autocorrelation to viscosity is derived from first principles of Brownian motion and the Stokes-Einstein relation. To improve precision, multiple windows acquired over 500 milliseconds are analyzed and the resulting linear fit parameters are averaged. Verification experiments were performed with 200 µL samples of glycerol and water with polystyrene microbeads. Lateral bulk motion up to 2 mm/s was tolerated and accurate viscosity measurements were obtained to a depth of 400 µm or more. Additionally, the method measured a significant decrease of the apparent diffusion constant of soft tissue after formalin fixation, suggesting potential for mapping tissue stiffness over a volume.

A New Method for Endoscopic Sampling of Submucosal Tissue in the Gastrointestinal Tract: A Comparison of the Biopsy Forceps and a New Drill Instrument.

PubMed

Walther, Charles; Jeremiasen, Martin; Rissler, Pehr; Johansson, Jan L M; Larsson, Marie S; Walther, Bruno S C S

2016-12-01

Background Sampling of submucosal lesions in the gastrointestinal tract through a flexible endoscope is a well-recognized clinical problem. One technique often used is endoscopic ultrasound-guided fine-needle aspiration, but it does not provide solid tissue biopsies with preserved architecture for histopathological evaluation. To obtain solid tissue biopsies from submucosal lesions, we have constructed a new endoscopic biopsy tool and compared it in a crossover study with the standard double cupped forceps. Methods Ten patients with endoscopically verified submucosal lesions were sampled. The endoscopist selected the position for the biopsies and used the instrument selected by randomization. After a biopsy was harvested, the endoscopist chose the next site for a biopsy and again used the instrument picked by randomization. A total of 6 biopsies, 3 with the forceps and 3 with the drill instrument, were collected in every patient. Results The drill instrument resulted in larger total size biopsies (mm 2 ; Mann-Whitney U test, P = .048) and larger submucosal part (%) of the biopsies (Mann-Whitney U test, P = .003) than the forceps. Two patients were observed because of chest pain and suspicion of bleeding in 24 hours. No therapeutic measures were necessary to be taken. Conclusion The new drill instrument for flexible endoscopy can safely deliver submucosal tissue samples from submucosal lesions in the upper gastrointestinal tract. © The Author(s) 2016.

Extraction of high-quality DNA from ethanol-preserved tropical plant tissues.

PubMed

Bressan, Eduardo A; Rossi, Mônica L; Gerald, Lee T S; Figueira, Antonio

2014-04-24

Proper conservation of plant samples, especially during remote field collection, is essential to assure quality of extracted DNA. Tropical plant species contain considerable amounts of secondary compounds, such as polysaccharides, phenols, and latex, which affect DNA quality during extraction. The suitability of ethanol (96% v/v) as a preservative solution prior to DNA extraction was evaluated using leaves of Jatropha curcas and other tropical species. Total DNA extracted from leaf samples stored in liquid nitrogen or ethanol from J. curcas and other tropical species (Theobroma cacao, Coffea arabica, Ricinus communis, Saccharum spp., and Solanum lycopersicon) was similar in quality, with high-molecular-weight DNA visualized by gel electrophoresis. DNA quality was confirmed by digestion with EcoRI or HindIII and by amplification of the ribosomal gene internal transcribed spacer region. Leaf tissue of J. curcas was analyzed by light and transmission electron microscopy before and after exposure to ethanol. Our results indicate that leaf samples can be successfully preserved in ethanol for long periods (30 days) as a viable method for fixation and conservation of DNA from leaves. The success of this technique is likely due to reduction or inactivation of secondary metabolites that could contaminate or degrade genomic DNA. Tissue conservation in 96% ethanol represents an attractive low-cost alternative to commonly used methods for preservation of samples for DNA extraction. This technique yields DNA of equivalent quality to that obtained from fresh or frozen tissue.

Extraction of high-quality DNA from ethanol-preserved tropical plant tissues

PubMed Central

2014-01-01

Background Proper conservation of plant samples, especially during remote field collection, is essential to assure quality of extracted DNA. Tropical plant species contain considerable amounts of secondary compounds, such as polysaccharides, phenols, and latex, which affect DNA quality during extraction. The suitability of ethanol (96% v/v) as a preservative solution prior to DNA extraction was evaluated using leaves of Jatropha curcas and other tropical species. Results Total DNA extracted from leaf samples stored in liquid nitrogen or ethanol from J. curcas and other tropical species (Theobroma cacao, Coffea arabica, Ricinus communis, Saccharum spp., and Solanum lycopersicon) was similar in quality, with high-molecular-weight DNA visualized by gel electrophoresis. DNA quality was confirmed by digestion with EcoRI or HindIII and by amplification of the ribosomal gene internal transcribed spacer region. Leaf tissue of J. curcas was analyzed by light and transmission electron microscopy before and after exposure to ethanol. Our results indicate that leaf samples can be successfully preserved in ethanol for long periods (30 days) as a viable method for fixation and conservation of DNA from leaves. The success of this technique is likely due to reduction or inactivation of secondary metabolites that could contaminate or degrade genomic DNA. Conclusions Tissue conservation in 96% ethanol represents an attractive low-cost alternative to commonly used methods for preservation of samples for DNA extraction. This technique yields DNA of equivalent quality to that obtained from fresh or frozen tissue. PMID:24761774

Ion Beam Sputtered Coatings of Bioglass

NASA Technical Reports Server (NTRS)

Hench, Larry L.; Wilson, J.; Ruzakowski, Patricia Henrietta Anne

1982-01-01

The ion beam sputtering technique available at the NASA-Lewis was used to apply coatings of bioglass to ceramic, metallic, and polymeric substrates. Experiments in vivo and in vitro described investigate these coatings. Some degree of substrate masking was obtained in all samples although stability and reactivity equivalent to bulk bioglass was not observed in all coated samples. Some degree of stability was seen in all coated samples that were reacted in vitro. Both metallic and ceramic substrates coated in this manner failed to show significantly improved coatings over those obtained with existing techniques. Implantation of the coated ceramic substrate samples in bone gave no definite bonding as seen with bulk glass; however, partial and patchy bonding was seen. Polymeric substrates in these studies showed promise of success. The coatings applied were sufficient to mask the underlying reactive test surface and tissue adhesion of collagen to bioglass was seen. Hydrophilic, hydrophobic, charged, and uncharged polymeric surfaces were successfully coated.

Quantitative analysis of drug distribution by ambient mass spectrometry imaging method with signal extinction normalization strategy and inkjet-printing technology.

PubMed

Luo, Zhigang; He, Jingjing; He, Jiuming; Huang, Lan; Song, Xiaowei; Li, Xin; Abliz, Zeper

2018-03-01

Quantitative mass spectrometry imaging (MSI) is a robust approach that provides both quantitative and spatial information for drug candidates' research. However, because of complicated signal suppression and interference, acquiring accurate quantitative information from MSI data remains a challenge, especially for whole-body tissue sample. Ambient MSI techniques using spray-based ionization appear to be ideal for pharmaceutical quantitative MSI analysis. However, it is more challenging, as it involves almost no sample preparation and is more susceptible to ion suppression/enhancement. Herein, based on our developed air flow-assisted desorption electrospray ionization (AFADESI)-MSI technology, an ambient quantitative MSI method was introduced by integrating inkjet-printing technology with normalization of the signal extinction coefficient (SEC) using the target compound itself. The method utilized a single calibration curve to quantify multiple tissue types. Basic blue 7 and an antitumor drug candidate (S-(+)-deoxytylophorinidine, CAT) were chosen to initially validate the feasibility and reliability of the quantitative MSI method. Rat tissue sections (heart, kidney, and brain) administered with CAT was then analyzed. The quantitative MSI analysis results were cross-validated by LC-MS/MS analysis data of the same tissues. The consistency suggests that the approach is able to fast obtain the quantitative MSI data without introducing interference into the in-situ environment of the tissue sample, and is potential to provide a high-throughput, economical and reliable approach for drug discovery and development. Copyright © 2017 Elsevier B.V. All rights reserved.

FIB-SEM imaging of carbon nanotubes in mouse lung tissue.

PubMed

Købler, Carsten; Saber, Anne Thoustrup; Jacobsen, Nicklas Raun; Wallin, Håkan; Vogel, Ulla; Qvortrup, Klaus; Mølhave, Kristian

2014-06-01

Ultrastructural characterisation is important for understanding carbon nanotube (CNT) toxicity and how the CNTs interact with cells and tissues. The standard method for this involves using transmission electron microscopy (TEM). However, in particular, the sample preparation, using a microtome to cut thin sample sections for TEM, can be challenging for investigation of regions with agglomerations of large and stiff CNTs because the CNTs cut with difficulty. As a consequence, the sectioning diamond knife may be damaged and the uncut CNTs are left protruding from the embedded block surface excluding them from TEM analysis. To provide an alternative to ultramicrotomy and subsequent TEM imaging, we studied focused ion beam scanning electron microscopy (FIB-SEM) of CNTs in the lungs of mice, and we evaluated the applicability of the method compared to TEM. FIB-SEM can provide serial section volume imaging not easily obtained with TEM, but it is time-consuming to locate CNTs in the tissue. We demonstrate that protruding CNTs after ultramicrotomy can be used to locate the region of interest, and we present FIB-SEM images of CNTs in lung tissue. FIB-SEM imaging was applied to lung tissue from mice which had been intratracheally instilled with two different multiwalled CNTs; one being short and thin, and the other longer and thicker. FIB-SEM was found to be most suitable for detection of the large CNTs (Ø ca. 70 nm), and to be well suited for studying CNT agglomerates in biological samples which is challenging using standard TEM techniques.

Psychiatric Brain Banking: Three Perspectives on Current Trends and Future Directions

PubMed Central

Deep-Soboslay, Amy; Benes, Francine M.; Haroutunian, Vahram; Ellis, Justin K.; Kleinman, Joel E.; Hyde, Thomas M.

2011-01-01

Introduction The study of postmortem human brain tissue is central to the advancement of the neurobiological studies of psychiatric illness, particularly for the study of brain-specific isoforms and molecules. Methods The state-of-the-art methods and recommendations for maintaining a successful brain bank for psychiatric disorders are discussed, using the convergence of viewpoints from three brain collections, the National Institute of Mental Health Brain Collection (NIMH), the Harvard Brain Tissue Resource Center (HBTRC), and the Mt. Sinai School of Medicine Brain Bank (MSSM-BB), with diverse research interests and divergent approaches to tissue acquisition. Results While the NIMH obtains donations from medical examiners for its collection, and places particular emphasis on clinical diagnosis, toxicology, and building lifespan control cohorts, the HBTRC is uniquely designed as a repository whose sole purpose is to collect large-volume, high quality brain tissue from community-based donors based on relationships across an expansive nationwide network, and places emphasis on the accessibility of its bank in disseminating tissue and related data to research groups worldwide. The MSSM-BB collection has shown that, with dedication, prospective recruitment is a successful approach to tissue donation, and places particular emphasis on rigorous clinical diagnosis through antemortem contact with donors. The MSSM-BB places great importance on stereological tissue sampling methods for neuroanatomical studies, and frozen tissue sampling approaches that enable multiple assessments (RNA, DNA, protein, enzyme activity, binding, etc.) of the same tissue block. Promising scientific approaches for elucidating the molecular and cellular pathways in brain that may contribute to schizophrenia and/or bipolar disorder, such as cell culture techniques and microarray-based gene expression and genotyping studies are briefly discussed. Conclusions Despite unique perspectives from three established brain collections, there is a consensus that (1) diverse strategies for tissue acquisition, (2) rigor in tissue and diagnostic characterization, (3) the importance of sample accessibility, and (4) continual application of innovative scientific approaches to the study of brain tissue are all integral to the success and future of psychiatric brain banking. The future of neuropsychiatric research depends upon in the availability of high quality brain specimens from large numbers of subjects, including non-psychiatric controls. PMID:20673875

Presence of human papilloma virus, herpes simplex virus and Epstein-Barr virus DNA in oral biopsies from Sudanese patients with regard to toombak use.

PubMed

Jalouli, Jamshid; Ibrahim, Salah O; Sapkota, Dipak; Jalouli, Miranda M; Vasstrand, Endre N; Hirsch, Jan M; Larsson, Per-Anders

2010-09-01

Using PCR/DNA sequencing, we investigated the prevalence of human papillomavirus (HPV), herpes simplex virus (HSV) and Epstein-Barr virus (EBV) DNA in brush biopsies obtained from 150 users of Sudanese snuff (toombak) and 25 non-users of toombak in formalin-fixed paraffin-embedded tissue samples obtained from 31 patients with oral dysplasias (25 toombak users and 6 non-users), and from 217 patients with oral cancers (145 toombak users and 72 non-users). In the brush tissue samples from toombak users, HPV was detected in 60 (40%), HSV in 44 (29%) and EBV in 97 (65%) of the samples. The corresponding figures for the 25 samples from non-users were 17 (68%) positive for HPV, 6 (24%) positive for HSV and 21 (84%) for EBV. The formalin-fixed samples with oral dysplasias were all negative for HPV. In the 145 oral cancer samples from toombak users, HPV was detected in 39 (27%), HSV in 15 (10%) and EBV in 53 (37%) of the samples. The corresponding figures for the samples from non-users were 15 (21%) positive for HPV, 5 (7%) for HSV and 16 (22%) for EBV. These findings illustrate that prevalence of HSV, HPV and EBV infections are common and may influence oral health and cancer development. It is not obvious that cancer risk is increased in infected toombak users. These observations warrant further studies involving toombak-associated oral lesions, to uncover the possible mechanisms of these viral infections in the development of oral cancer, and the influence of toombak on these viruses. © 2010 John Wiley & Sons A/S.

Concordance of human papillomavirus types detected on the surface and in the tissue of genital lesions in men.

PubMed

Anic, Gabriella M; Messina, Jane L; Stoler, Mark H; Rollison, Dana E; Stockwell, Heather; Villa, Luisa L; Lazcano-Ponce, Eduardo; Gage, Christine; Silva, Roberto Jose C; Baggio, Maria L; Salmerón, Jorge; Giuliano, Anna R

2013-09-01

Swabbing the surface of a genital lesion to obtain a sample for HPV DNA testing is less invasive than a biopsy, but may not represent HPV types present in the lesion tissue. The objective of this study was to examine the concordance of HPV types detected in swab and biopsy samples from 165 genital lesions from men ages 18-70. Lesions included 90 condyloma, 10 penile intraepithelial neoplasia (PeIN), 23 non-condyloma with a known histology, and 42 lesions with an undetermined histology. All lesions were sampled by swabbing the surface of the lesion with a pre-wetted Dacron swab and taking a shave biopsy. HPV genotyping was performed using Linear Array for swab samples and INNO-LiPA for biopsy samples. The kappa and McNemar statistics were used to compare the concordance of detecting HPV types in swab and biopsy samples. Both sampling methods had high agreement for detection of HPV DNA in condyloma (87.8% agreement) and PeIN (100% agreement). There was also high concordance for detection of HPV16 (kappa = 1.00) and HPV18 (kappa = 1.00) in PeIN, however, agreement was low to moderate for detecting HPV6 (kappa = 0.31) and HPV11 (kappa = 0.56) in condyloma. Low to moderate agreement was also observed between sampling methods for detecting individual HPV types in the non-condyloma and lesions with an indefinite histology. The results suggest that obtaining a biopsy in addition to swabbing the surface of a lesion may provide additional information about specific HVP types associated with male genital lesions. Copyright © 2013 Wiley Periodicals, Inc.

5-ALA-induced fluorescence as a marker for diagnostic tissue in stereotactic biopsies of intracranial lymphomas: experience in 41 patients.

PubMed

Kiesel, Barbara; Millesi, Matthias; Woehrer, Adelheid; Furtner, Julia; Bavand, Anahita; Roetzer, Thomas; Mischkulnig, Mario; Wolfsberger, Stefan; Preusser, Matthias; Knosp, Engelbert; Widhalm, Georg

2018-06-01

OBJECTIVE Stereotactic needle biopsies are usually performed for histopathological confirmation of intracranial lymphomas to guide adequate treatment. During biopsy, intraoperative histopathology is an effective tool to avoid acquisition of nondiagnostic samples. In the last years, 5-aminolevulinic acid (5-ALA)-induced fluorescence has been increasingly used for visualization of diagnostic brain tumor tissue during stereotactic biopsies. Recently, visible fluorescence was reported in the first cases of intracranial lymphomas as well. The aim of this study is thus to investigate the technical and clinical utility of 5-ALA-induced fluorescence in a large series of stereotactic biopsies for intracranial lymphoma. METHODS This prospective study recruited adult patients who underwent frameless stereotactic needle biopsy for a radiologically suspected intracranial lymphoma after oral 5-ALA administration. During biopsy, samples from the tumor region were collected for histopathological analysis, and presence of fluorescence (strong, vague, or no fluorescence) was assessed with a modified neurosurgical microscope. In tumors with available biopsy samples from at least 2 different regions the intratumoral fluorescence homogeneity was additionally investigated. Furthermore, the influence of potential preoperative corticosteroid treatment or immunosuppression on fluorescence was analyzed. Histopathological tumor diagnosis was established and all collected biopsy samples were screened for diagnostic lymphoma tissue. RESULTS The final study cohort included 41 patients with intracranial lymphoma. Stereotactic biopsies with assistance of 5-ALA were technically feasible in all cases. Strong fluorescence was found as maximum level in 30 patients (75%), vague fluorescence in 2 patients (4%), and no visible fluorescence in 9 patients (21%). In 28 cases, samples were obtained from at least 2 different tumor regions; homogenous intratumoral fluorescence was found in 16 of those cases (57%) and inhomogeneous intratumoral fluorescence in 12 (43%). According to histopathological analysis, all samples with strong or vague fluorescence contained diagnostic lymphoma tissue, resulting in a positive predictive value of 100%. Analysis showed no influence of preoperative corticosteroids or immunosuppression on fluorescence. CONCLUSIONS The data obtained in this study demonstrate the technical and clinical utility of 5-ALA-induced fluorescence in stereotactic biopsies of intracranial lymphomas. Thus, 5-ALA can serve as a useful tool to select patients not requiring intraoperative histopathology, and its application should markedly reduce operation time and related costs in the future.

Optimised laser microdissection of the human ocular surface epithelial regions for microarray studies

PubMed Central

2013-01-01

Background The most important challenge of performing insitu transcriptional profiling of the human ocular surface epithelial regions is obtaining samples in sufficient amounts, without contamination from adjacent tissue, as the region of interest is microscopic and closely apposed to other tissues regions. We have effectively collected ocular surface (OS) epithelial tissue samples from the Limbal Epithelial Crypt (LEC), limbus, cornea and conjunctiva of post-mortem cadaver eyes with laser microdissection (LMD) technique for gene expression studies with spotted oligonucleotide microarrays and Gene 1.0 ST arrays. Methods Human donor eyes (4 pairs for spotted oligonucleotide microarrays, 3 pairs for Gene 1.0 ST arrays) consented for research were included in this study with due ethical approval of the Nottingham Research Ethics Committee. Eye retrieval was performed within 36 hours of post-mortem period. The dissected corneoscleral buttons were immersed in OCT media and frozen in liquid nitrogen and stored at −80°C till further use. Microscopic tissue sections of interest were taken on PALM slides and stained with Toluidine Blue for laser microdissection with PALM microbeam systems. Optimisation of the laser microdissection technique was crucial for efficient and cost effective sample collection. Results The starting concentration of RNA as stipulated by the protocol of microarray platforms was taken as the cut-off concentration of RNA samples in our studies. The area of LMD tissue processed for spotted oligonucleotide microarray study ranged from 86,253 μm2 in LEC to 392,887 μm2 in LEC stroma. The RNA concentration of the LMD samples ranged from 22 to 92 pg/μl. The recommended starting concentration of the RNA samples used for Gene 1.0 ST arrays was 6 ng/5 μl. To achieve the desired RNA concentration the area of ocular surface epithelial tissue sample processed for the Gene 1.0 ST array experiments was approximately 100,0000 μm2 to 130,0000 μm2. RNA concentration of these samples ranged from 10.88 ng/12 μl to 25.8 ng/12 μl, with the RNA integrity numbers (RIN) for these samples from 3.3 to 7.9. RNA samples with RIN values below 2, that had failed to amplify satisfactorily were discarded. Conclusions The optimised protocol for sample collection and laser microdissection improved the RNA yield of the insitu ocular surface epithelial regions for effective microarray studies on spotted oligonucleotide and affymetrix platforms. PMID:24160452

Mammary stem cell and macrophage markers are enriched in normal tissue adjacent to inflammatory breast cancer.

PubMed

Reddy, Jay P; Atkinson, Rachel L; Larson, Richard; Burks, Jared K; Smith, Daniel; Debeb, Bisrat G; Ruffell, Brian; Creighton, Chad J; Bambhroliya, Arvind; Reuben, James M; Van Laere, Steven J; Krishnamurthy, Savitri; Symmans, William F; Brewster, Abenaa M; Woodward, Wendy A

2018-06-01

We hypothesized that breast tissue not involved by tumor in inflammatory breast cancer (IBC) patients contains intrinsic differences, including increased mammary stem cells and macrophage infiltration, which may promote the IBC phenotype. Normal breast parenchyma ≥ 5 cm away from primary tumors was obtained from mastectomy specimens. This included an initial cohort of 8 IBC patients and 60 non-IBC patients followed by a validation cohort of 19 IBC patients and 25 non-IBC patients. Samples were immunostained for either CD44 + CD49f + CD133/2 + mammary stem cell markers or the CD68 macrophage marker and correlated with IBC status. Quantitation of positive cells was determined using inForm software from PerkinElmer. We also examined the association between IBC status and previously published tumorigenic stem cell and IBC tumor signatures in the validation cohort samples. 8 of 8 IBC samples expressed isolated CD44 + CD49f + CD133/2 + stem cell marked cells in the initial cohort as opposed to 0/60 non-IBC samples (p = 0.001). Similarly, the median number of CD44 + CD49f + CD133/2 + cells was significantly higher in the IBC validation cohort as opposed to the non-IBC validation cohort (25.7 vs. 14.2, p = 0.007). 7 of 8 IBC samples expressed CD68 + histologically confirmed macrophages in initial cohort as opposed to 12/48 non-IBC samples (p = 0.001). In the validation cohort, the median number of CD68 + cells in IBC was 3.7 versus 1.0 in the non-IBC cohort (p = 0.06). IBC normal tissue was positively associated with a tumorigenic stem cell signature (p = 0.02) and with a 79-gene IBC signature (p < 0.001). Normal tissue from IBC patients is enriched for both mammary stem cells and macrophages and has higher association with both a tumorigenic stem cell signature and IBC-specific tumor signature. Collectively, these data suggest that IBC normal tissue differs from non-IBC tissue. Whether these changes occur before the tumor develops or is induced by tumor warrants further investigation.

Clinical application of Lin's biopsy grasper for intrauterine targeted biopsy and polypectomy during office hysteroscopy.

PubMed

Cheng, Hsin-Yi; Lin, Bao-Liang; Tseng, Jen-Yu; Ueno, Kazunori; Nakada, Sakura

2018-06-01

Hysteroscopy has widely been used for diagnosis of the uterine cavity; however, target biopsy has often been difficult in part to the inherent limitations of ancillary instruments. Lin's biopsy grasper was specifically designed to work in conjunction with a flexible hysteroscope to obtain intrauterine biopsy under transabdominal sonography. Herein, we share our clinical experience in the management of endometrial abnormalities with the use of Lin's biopsy grasper during office-based hysteroscopy. From February 2006 to November 2016, the use of Lin's biopsy grasper for tissue biopsy was attempted on 126 cases. We retrospectively recorded and analyzed the patients' preoperative characteristics and biopsy outcomes to demonstrate the feasibility and efficacy of Lin's biopsy grasper. Out of the one hundred and twenty-six enrolled patients, satisfactory targeted biopsies were achieved; including high diagnostic rate (92.1%, with 116 cases confirmed histologically) and adequate tissue retrieval (77.8%, with 98 cases obtaining optimal specimen volume). All patients tolerated the procedure without analgesics or anesthesia. Diagnostic flexible hysteroscopy combined with the use of Lin's biopsy grasper has proven to be an effective tool for intrauterine evaluation and obtaining tissue sample. Copyright © 2018. Published by Elsevier B.V.

Metabolomics studies in brain tissue: A review.

PubMed

Gonzalez-Riano, Carolina; Garcia, Antonia; Barbas, Coral

2016-10-25

Brain is still an organ with a composition to be discovered but beyond that, mental disorders and especially all diseases that curse with dementia are devastating for the patient, the family and the society. Metabolomics can offer an alternative tool for unveiling new insights in the discovery of new treatments and biomarkers of mental disorders. Until now, most of metabolomic studies have been based on biofluids: serum/plasma or urine, because brain tissue accessibility is limited to animal models or post mortem studies, but even so it is crucial for understanding the pathological processes. Metabolomics studies of brain tissue imply several challenges due to sample extraction, along with brain heterogeneity, sample storage, and sample treatment for a wide coverage of metabolites with a wide range of concentrations of many lipophilic and some polar compounds. In this review, the current analytical practices for target and non-targeted metabolomics are described and discussed with emphasis on critical aspects: sample treatment (quenching, homogenization, filtration, centrifugation and extraction), analytical methods, as well as findings considering the used strategies. Besides that, the altered analytes in the different brain regions have been associated with their corresponding pathways to obtain a global overview of their dysregulation, trying to establish the link between altered biological pathways and pathophysiological conditions. Copyright © 2016 Elsevier B.V. All rights reserved.

Molecular genotyping of Echinococcus granulosus using formalin-fixed paraffin-embedded preparations from human isolates in unusual tissue sites.

PubMed

Hizem, A; M'rad, S; Oudni-M'rad, M; Mestiri, S; Hammedi, F; Mezhoud, H; Zakhama, A; Mokni, M; Babba, H

2016-07-01

Cystic echinococcosis (CE) caused by Echinococcus granulosus remains a serious problem worldwide for issues relating to public health and the economy. The most predominantly affected sites are the liver and the lungs, but other organs such as the heart, the spleen and the peritoneum can also be infected. Access to cysts from uncommon sites has limited genomic and molecular investigations. In the present study, genotypes of E. granulosus sensu lato were identified from formalin-fixed paraffin-embedded tissues (FF-PETs) implicated in human CE. Tissue samples were obtained from 57 patients with histologically confirmed CE. DNA samples were analysed using Egss 1 polymerase chain reaction (PCR) specific to the mitochondrial 12S rRNA gene of E. granulosus sensu stricto. All cysts were typed as E. granulosus sensu stricto with up to 35% of the liver and 16.6% of lungs being the most frequently infected, and up to 48.4% of samples being from rare sites. No correlation was found between cyst site and either the gender or the age of patients. This study demonstrates the possibility of exploiting atypical cysts using FF-PET samples and highlights the predominance of E. granulosus sensu stricto species in the Tunisian population, even in unusual infection sites.

Characterization of human oral tissues based on quantitative analysis of optical coherence tomography images

NASA Astrophysics Data System (ADS)

Salehi, Hassan S.; Kosa, Ali; Mahdian, Mina; Moslehpour, Saeid; Alnajjar, Hisham; Tadinada, Aditya

2017-02-01

In this paper, five types of tissues, human enamel, human cortical bone, human trabecular bone, muscular tissue, and fatty tissue were imaged ex vivo using optical coherence tomography (OCT). The specimens were prepared in blocks of 5 x 5 x 3 mm (width x length x height). The OCT imaging system was a swept source OCT system operating at wavelengths ranging between 1250 nm and 1360 nm with an average power of 18 mW and a scan rate of 50 to 100 kHz. The imaging probe was placed on top of a 2 x 2 cm stabilizing device to maintain a standard distance from the samples. Ten image samples from each type of tissue were obtained. To acquire images with minimum inhomogeneity, imaging was performed multiple times at different points. Based on the observed texture differences between OCT images of soft and hard tissues, spatial and spectral features were quantitatively extracted from the OCT images. The Radon transform from angles of 0 deg to 90 deg was computed, averaged over all the angles, normalized to peak at unity, and then fitted with Gaussian function. The mean absolute values of the spatial frequency components of the OCT image were considered as a feature, where 2-D fast Fourier transform (FFT) was done to OCT images. These OCT features can reliably differentiate between a range of hard and soft tissues, and could be extremely valuable in assisting dentists for in vivo evaluation of oral tissues and early detection of pathologic changes in tissues.

Water Potential in Excised Leaf Tissue

PubMed Central

Nelsen, Charles E.; Safir, Gene R.; Hanson, Andrew D.

1978-01-01

Leaf water potential (Ψleaf) determinations were made on excised leaf samples using a commercial dew point hygrometer (Wescor Inc., Logan, Utah) and a thermocouple psychrometer operated in the isopiestic mode. With soybean leaves (Glycine max L.), there was good agreement between instruments; equilibration times were 2 to 3 hours. With cereals (Triticum aestivum L. and Hordeum vulgare L.), agreement between instruments was poor for moderately wilted leaves when 7-mm-diameter punches were used in the hygrometer and 20-mm slices were used in the psychrometer, because the Ψleaf values from the dew point hygrometer were too high. Agreement was improved by replacing the 7-mm punch samples in the hygrometer by 13-mm slices, which had a lower cut edge to volume ratio. Equilibration times for cereals were normally 6 to 8 hours. Spuriously high Ψleaf values obtained with 7-mm leaf punches may be associated with the ion release and reabsorption that occur upon tissue excision; such errors evidently depend both on the species and on tissue water status. PMID:16660227

Boron concentration measurements by alpha spectrometry and quantitative neutron autoradiography in cells and tissues treated with different boronated formulations and administration protocols.

PubMed

Bortolussi, Silva; Ciani, Laura; Postuma, Ian; Protti, Nicoletta; Luca Reversi; Bruschi, Piero; Ferrari, Cinzia; Cansolino, Laura; Panza, Luigi; Ristori, Sandra; Altieri, Saverio

2014-06-01

The possibility to measure boron concentration with high precision in tissues that will be irradiated represents a fundamental step for a safe and effective BNCT treatment. In Pavia, two techniques have been used for this purpose, a quantitative method based on charged particles spectrometry and a boron biodistribution imaging based on neutron autoradiography. A quantitative method to determine boron concentration by neutron autoradiography has been recently set-up and calibrated for the measurement of biological samples, both solid and liquid, in the frame of the feasibility study of BNCT. This technique was calibrated and the obtained results were cross checked with those of α spectrometry, in order to validate them. The comparisons were performed using tissues taken form animals treated with different boron administration protocols. Subsequently the quantitative neutron autoradiography was employed to measure osteosarcoma cell samples treated with BPA and with new boronated formulations. © 2013 Published by Elsevier Ltd.

Dye-Enhanced Multimodal Confocal Imaging of Brain Cancers

NASA Astrophysics Data System (ADS)

Wirth, Dennis; Snuderl, Matija; Sheth, Sameer; Curry, William; Yaroslavsky, Anna

2011-04-01

Background and Significance: Accurate high resolution intraoperative detection of brain tumors may result in improved patient survival and better quality of life. The goal of this study was to evaluate dye enhanced multimodal confocal imaging for discriminating normal and cancerous brain tissue. Materials and Methods: Fresh thick brain specimens were obtained from the surgeries. Normal and cancer tissues were investigated. Samples were stained in methylene blue and imaged. Reflectance and fluorescence signals were excited at 658nm. Fluorescence emission and polarization were registered from 670 nm to 710 nm. The system provided lateral resolution of 0.6 μm and axial resolution of 7 μm. Normal and cancer specimens exhibited distinctively different characteristics. H&E histopathology was processed from each imaged sample. Results and Conclusions: The analysis of normal and cancerous tissues indicated clear differences in appearance in both the reflectance and fluorescence responses. These results confirm the feasibility of multimodal confocal imaging for intraoperative detection of small cancer nests and cells.

The potential influence of CO2, as an agent for euthanasia, on the pharmacokinetics of basic compounds in rodents.

PubMed

Angus, Derek W; Baker, James A; Mason, Rona; Martin, Iain J

2008-02-01

Rodent tissue distribution and pharmacokinetic studies were performed on basic compounds Org A and Org B in support of central nervous system drug discovery programs. A consistent observation from these studies was that drug concentrations in plasma obtained by cardiac puncture after CO(2) euthanasia were markedly higher compared with those from other sampling methods (serial sampling, isoflurane anesthesia, or cervical dislocation). Further investigations demonstrated that CO(2) euthanasia led to a reduction in blood pH in both rats and mice, which was not observed with the other sampling methods. The use of CO(2) euthanasia resulted in a decrease in the brain/plasma ratio of Org B, largely as a result of increased plasma concentrations. The pharmacokinetics of a basic drug, raloxifene, in rat were also influenced by sampling technique. CO(2) euthanasia before sampling, resulted in a 2- to 3-fold increase in the area under the drug concentration-time curve, a decrease in plasma clearance, and a decrease in the steady-state volume of distribution compared with isoflurane anesthesia. It is proposed that a decrease in the pH of blood relative to that of other tissues, as a consequence of CO(2) exposure, results in a redistribution of basic compounds out of the tissues, leading to higher concentrations in plasma.

Acoustic property reconstruction of a pygmy sperm whale (Kogia breviceps) forehead based on computed tomography imaging.

PubMed

Song, Zhongchang; Xu, Xiao; Dong, Jianchen; Xing, Luru; Zhang, Meng; Liu, Xuecheng; Zhang, Yu; Li, Songhai; Berggren, Per

2015-11-01

Computed tomography (CT) imaging and sound experimental measurements were used to reconstruct the acoustic properties (density, velocity, and impedance) of the forehead tissues of a deceased pygmy sperm whale (Kogia breviceps). The forehead was segmented along the body axis and sectioned into cross section slices, which were further cut into sample pieces for measurements. Hounsfield units (HUs) of the corresponding measured pieces were obtained from CT scans, and regression analyses were conducted to investigate the linear relationships between the tissues' HUs and velocity, and HUs and density. The distributions of the acoustic properties of the head at axial, coronal, and sagittal cross sections were reconstructed, revealing that the nasal passage system was asymmetric and the cornucopia-shaped spermaceti organ was in the right nasal passage, surrounded by tissues and airsacs. A distinct dense theca was discovered in the posterior-dorsal area of the melon, which was characterized by low velocity in the inner core and high velocity in the outer region. Statistical analyses revealed significant differences in density, velocity, and acoustic impedance between all four structures, melon, spermaceti organ, muscle, and connective tissue (p < 0.001). The obtained acoustic properties of the forehead tissues provide important information for understanding the species' bioacoustic characteristics.

A Prospective Evaluation of Circulating Tumor Cells and Cell-Free DNA in EGFR-Mutant Non-Small Cell Lung Cancer Patients Treated with Erlotinib on a Phase II Trial.

PubMed

Yanagita, Masahiko; Redig, Amanda J; Paweletz, Cloud P; Dahlberg, Suzanne E; O'Connell, Allison; Feeney, Nora; Taibi, Myriam; Boucher, David; Oxnard, Geoffrey R; Johnson, Bruce E; Costa, Daniel B; Jackman, David M; Jänne, Pasi A

2016-12-15

Genotype-directed therapy is the standard of care for advanced non-small cell lung cancer (NSCLC), but obtaining tumor tissue for genotyping remains a challenge. Circulating tumor cell (CTC) or cell-free DNA (cfDNA) analysis may allow for noninvasive evaluation. This prospective trial evaluated CTCs and cfDNA in EGFR-mutant NSCLC patients treated with erlotinib until progression. EGFR-mutant NSCLC patients were enrolled in a phase II trial of erlotinib. Blood was collected at baseline, every 2 months on study, and at disease progression. Plasma genotyping was performed by droplet digital PCR for EGFR19del, L858R, and T790M. CTCs were isolated by CellSave, enumerated, and analyzed by immunofluorescence for CD45 and pan-cytokeratin and EGFR and MET FISH were also performed. Rebiopsy was performed at disease progression. Sixty patients were enrolled; 44 patients discontinued therapy for disease progression. Rebiopsy occurred in 35 of 44 patients (80%), with paired CTC/cfDNA analysis in 41 of 44 samples at baseline and 36 of 44 samples at progression. T790M was identified in 23 of 35 (66%) tissue biopsies and 9 of 39 (23%) cfDNA samples. CTC analysis at progression identified MET amplification in 3 samples in which tissue analysis could not be performed. cfDNA analysis identified T790M in 2 samples in which rebiopsy was not possible. At diagnosis, high levels of cfDNA but not high levels of CTCs correlated with progression-free survival. cfDNA and CTCs are complementary, noninvasive assays for evaluation of acquired resistance to first-line EGFR TKIs and may expand the number of patients in whom actionable genetic information can be obtained at acquired resistance. Serial cfDNA monitoring may offer greater clinical utility than serial monitoring of CTCs. Clin Cancer Res; 22(24); 6010-20. ©2016 AACR. ©2016 American Association for Cancer Research.

EBUS-Guided Cautery-Assisted Transbronchial Forceps Biopsies: Safety and Sensitivity Relative to Transbronchial Needle Aspiration

PubMed Central

Bramley, Kyle; Pisani, Margaret A.; Murphy, Terrence E.; Araujo, Katy; Homer, Robert; Puchalski, Jonathan

2016-01-01

Background EBUS-guided transbronchial needle aspiration (TBNA) is important in the evaluation of thoracic lymphadenopathy. Reliably providing excellent diagnostic yield for malignancy, its diagnosis of sarcoidosis is inconsistent. Furthermore, when larger “core” biopsy samples of malignant tissue are required, TBNA may not suffice. The primary objective of this study was to determine if the sequential use of TBNA and a novel technique called cautery-assisted transbronchial forceps biopsies (ca-TBFB) was safe. Secondary outcomes included sensitivity and successful acquisition of tissue. Methods Fifty unselected patients undergoing convex probe EBUS were prospectively enrolled. Under EBUS guidance, all lymph nodes ≥ 1 cm were sequentially biopsied using TBNA and ca-TBFB. Safety and sensitivity were assessed at the nodal level for 111 nodes. Results of each technique were also reported on a per-patient basis. Results There were no significant adverse events. In nodes determined to be malignant, TBNA provided higher sensitivity (100%) than ca-TBFB (78%). However, among nodes with granulomatous inflammation, ca-TBFB exhibited higher sensitivity (90%) than TBNA (33%). For analysis based on patients rather than nodes, 6 of the 31 patients with malignancy would have been missed or understaged if the diagnosis was based on samples obtained by ca-TBFB. On the other hand, 3 of 8 patients with sarcoidosis would have been missed if analysis was based only on TBNA samples. In some cases only ca-TBFB acquired sufficient tissue for the core samples needed in clinical trials of malignancy. Conclusions The sequential use of TBNA and ca-TBFB appears to be safe. The larger samples obtained from ca-TBFB increased its sensitivity to detect granulomatous disease and provided specimens for clinical trials of malignancy when needle biopsies were insufficient. For thoracic surgeons and advanced bronchoscopists, we advocate ca-TBFB as an alternative to TBNA in select clinical scenarios. PMID:26912301

Endobronchial Ultrasound-Guided Cautery-Assisted Transbronchial Forceps Biopsies: Safety and Sensitivity Relative to Transbronchial Needle Aspiration.

PubMed

Bramley, Kyle; Pisani, Margaret A; Murphy, Terrence E; Araujo, Katy L; Homer, Robert J; Puchalski, Jonathan T

2016-05-01

Endobronchial ultrasound (EBUS)-guided transbronchial needle aspiration (TBNA) is important in the evaluation of thoracic lymphadenopathy. Reliably providing excellent diagnostic yield for malignancy, its diagnosis of sarcoidosis is inconsistent. Furthermore, TBNA may not suffice when larger "core biopsy" samples of malignant tissue are required. The primary objective of this study was to determine if the sequential use of TBNA and a novel technique called cautery-assisted transbronchial forceps biopsy (ca-TBFB) was safe. Secondary outcomes included sensitivity and successful acquisition of tissue. The study prospectively enrolled 50 unselected patients undergoing convex-probe EBUS. All lymph nodes exceeding 1 cm were sequentially biopsied under EBUS guidance using TBNA and ca-TBFB. Safety and sensitivity were assessed at the nodal level for 111 nodes. Results of each technique were also reported for each patient. There were no significant adverse events. In nodes determined to be malignant, TBNA provided higher sensitivity (100%) than ca-TBFB (78%). However, among nodes with granulomatous inflammation, ca-TBFB exhibited higher sensitivity (90%) than TBNA (33%). On the one hand, for analysis based on patients rather than nodes, 6 of the 31 patients with malignancy would have been missed or understaged if the diagnosis were based on samples obtained by ca-TBFB. On the other hand, 3 of 8 patients with sarcoidosis would have been missed if analysis were based only on TBNA samples. In some patients, only ca-TBFB acquired sufficient tissue for the core samples needed in clinical trials of malignancy. The sequential use of TBNA and ca-TBFB appears to be safe. The larger samples obtained from ca-TBFB increased its sensitivity to detect granulomatous disease and provided adequate specimens for clinical trials of malignancy when specimens from needle biopsies were insufficient. For thoracic surgeons and advanced bronchoscopists, we advocate ca-TBFB as an alternative to TBNA in select clinical scenarios. Copyright © 2016 The Society of Thoracic Surgeons. Published by Elsevier Inc. All rights reserved.

Environmental DNA Marker Development with Sparse Biological Information: A Case Study on Opossum Shrimp (Mysis diluviana).

PubMed

Carim, Kellie J; Christianson, Kyle R; McKelvey, Kevin M; Pate, William M; Silver, Douglas B; Johnson, Brett M; Galloway, Benjamin T; Young, Michael K; Schwartz, Michael K

2016-01-01

The spread of Mysis diluviana, a small glacial relict crustacean, outside its native range has led to unintended shifts in the composition of native fish communities throughout western North America. As a result, biologists seek accurate methods of determining the presence of M. diluviana, especially at low densities or during the initial stages of an invasion. Environmental DNA (eDNA) provides one solution for detecting M. diluviana, but building eDNA markers that are both sensitive and species-specific is challenging when the distribution and taxonomy of closely related non-target taxa are poorly understood, published genetic data are sparse, and tissue samples are difficult to obtain. To address these issues, we developed a pair of independent eDNA markers to increase the likelihood of a positive detection of M. diluviana when present and reduce the probability of false positive detections from closely related non-target species. Because tissue samples of closely-related and possibly sympatric, non-target taxa could not be obtained, we used synthetic DNA sequences of closely related non-target species to test the specificity of eDNA markers. Both eDNA markers yielded positive detections from five waterbodies where M. diluviana was known to be present, and no detections in five others where this species was thought to be absent. Daytime samples from varying depths in one waterbody occupied by M. diluviana demonstrated that samples near the lake bottom produced 5 to more than 300 times as many eDNA copies as samples taken at other depths, but all samples tested positive regardless of depth.

Metastasis Infiltration: An Investigation of the Postoperative Brain-Tumor Interface

DOE Office of Scientific and Technical Information (OSTI.GOV)

Raore, Bethwel; Schniederjan, Matthew; Prabhu, Roshan

Purpose: This study aims to evaluate brain infiltration of metastatic tumor cells past the main tumor resection margin to assess the biological basis for the use of stereotactic radiosurgery treatment of the tumor resection cavity and visualized resection edge or clinical target volume. Methods and Materials: Resection margin tissue was obtained after gross total resection of a small group of metastatic lesions from a variety of primary sources. The tissue at the border of the tumor and brain tissue was carefully oriented and processed to evaluate the presence of tumor cells within brain tissue and their distance from the resectionmore » margin. Results: Microscopic assessment of the radially oriented tissue samples showed no tumor cells infiltrating the surrounding brain tissue. Among the positive findings were reactive astrocytosis observed on the brain tissue immediately adjacent to the tumor resection bed margin. Conclusions: The lack of evidence of metastatic tumor cell infiltration into surrounding brain suggests the need to target only a narrow depth of the resection cavity margin to minimize normal tissue injury and prevent treatment size-dependent stereotactic radiosurgery complications.« less

Rescue karyotyping: a case series of array-based comparative genomic hybridization evaluation of archival conceptual tissue

PubMed Central

2014-01-01

Background Determination of fetal aneuploidy is central to evaluation of recurrent pregnancy loss (RPL). However, obtaining this information at the time of a miscarriage is not always possible or may not have been ordered. Here we report on “rescue karyotyping”, wherein DNA extracted from archived paraffin-embedded pregnancy loss tissue from a prior dilation and curettage (D&C) is evaluated by array-based comparative genomic hybridization (aCGH). Methods A retrospective case series was conducted at an academic medical center. Patients included had unexplained RPL and a prior pregnancy loss for which karyotype information would be clinically informative but was unavailable. After extracting DNA from slides of archived tissue, aCGH with a reduced stringency approach was performed, allowing for analysis of partially degraded DNA. Statistics were computed using STATA v12.1 (College Station, TX). Results Rescue karyotyping was attempted on 20 specimens from 17 women. DNA was successfully extracted in 16 samples (80.0%), enabling analysis at either high or low resolution. The longest interval from tissue collection to DNA extraction was 4.2 years. There was no significant difference in specimen sufficiency for analysis in the collection-to-extraction interval (p = 0.14) or gestational age at pregnancy loss (p = 0.32). Eight specimens showed copy number variants: 3 trisomies, 2 partial chromosomal deletions, 1 mosaic abnormality and 2 unclassified variants. Conclusions Rescue karyotyping using aCGH on DNA extracted from paraffin-embedded tissue provides the opportunity to obtain critical fetal cytogenetic information from a prior loss, even if it occurred years earlier. Given the ubiquitous archiving of paraffin embedded tissue obtained during a D&C and the ease of obtaining results despite long loss-to-testing intervals or early gestational age at time of fetal demise, this may provide a useful technique in the evaluation of couples with recurrent pregnancy loss. PMID:24589081

Secretory leukocyte protease inhibitor expression and high-risk HPV infection in anal lesions of HIV positive patients

PubMed Central

NUOVO, Gerard J.; GRINSZTEJN, Beatriz; FRIEDMAN, Ruth K.; VELOSO, Valdiléa G.; CUNHA, Cynthia B.; COUTINHO, José R.; VIANNA-ANDRADE, Cecilia; OLIVEIRA, Nathalia S.; WOODHAM, Andrew W.; DA SILVA, Diane M.; KAST, W. Martin

2016-01-01

Objective The aim of the current study was to evaluate secretory leukocyte protease inhibitor (SLPI) expression in anal biopsies from HIV-positive (HIV+) individuals, and compare that to anal intraepithelial neoplasia (AIN) diagnoses and human papillomavirus (HPV) status. Design This is a cross-sectional study of a cohort of 54 HIV+ (31 males and 23 females) from an AIDS clinic in Rio de Janeiro, Brazil. Methods The study material consisted of anorectal tissue biopsies obtained from HIV+ subjects, which were used to construct tissue microarray paraffin blocks for immunohistochemical analysis of SLPI expression. Biopsies were evaluated by an expert pathologist and classified as low-grade anal intraepithelial neoplasia (AIN1), high-grade anal intraepithelial neoplasia (AIN2/3), or normal squamous epithelium. Additionally, DNA from the biopsies was extracted and analyzed for the presence of low- or high-risk HPV DNA. Results Histologically normal squamous epithelium from the anorectal region showed strong positive SLPI staining in 17/20 (85%) samples. In comparison, 9/17 (53%) dysplastic squamous epithelial samples from AIN1 patients showed strong SLPI staining, and only 5/17 (29%) samples from AIN2-3 patients exhibited strong SPLI staining, which both were significantly fewer than those from normal tissue (p=0.005). Furthermore, there was a significantly higher proportion of samples in which oncogenic high-risk HPV genotypes were detected in low SLPI expressing tissues than that in tissues with high SLPI expression (p=0.040). Conclusion Taken together these results suggest that low SLPI expression is associated with high-risk HPV infections in the development of AIN. PMID:27149102

Utility of bronchial lavage fluids for epithelial growth factor receptor mutation assay in lung cancer patients: Comparison between cell pellets, cell blocks and matching tissue specimens

PubMed Central

Asaka, Shiho; Yoshizawa, Akihiko; Nakata, Rie; Negishi, Tatsuya; Yamamoto, Hiroshi; Shiina, Takayuki; Shigeto, Shohei; Matsuda, Kazuyuki; Kobayashi, Yukihiro; Honda, Takayuki

2018-01-01

The detection of epidermal growth factor receptor (EGFR) mutations is necessary for the selection of suitable patients with non-small cell lung cancer (NSCLC) for treatment with EGFR tyrosine kinase inhibitors. Cytology specimens are known to be suitable for EGFR mutation detection, although tissue specimens should be prioritized; however, there are limited studies that examine the utility of bronchial lavage fluid (BLF) in mutation detection. The purpose of the present study was to investigate the utility of BLF specimens for the detection of EGFR mutations using a conventional quantitative EGFR polymerase chain reaction (PCR) assay. Initially, quantification cycle (Cq) values of cell pellets, cell-free supernatants and cell blocks obtained from three series of 1% EGFR mutation-positive lung cancer cell line samples were compared for mutation detection. In addition, PCR analysis of BLF specimens obtained from 77 consecutive NSCLC patients, detecting EGFR mutations was validated, and these results were compared with those for the corresponding formalin-fixed paraffin-embedded (FFPE) tissue specimens obtained by surgical resection or biopsy of 49 of these patients. The Cq values for mutation detection were significantly lower in the cell pellet group (average, 29.58) compared with the other groups, followed by those in cell-free supernatants (average, 34.15) and in cell blocks (average, 37.12) for all three series (P<0.05). Mutational status was successfully analyzed in 77 BLF specimens, and the results obtained were concordant with those of the 49 matching FFPE tissue specimens. Notably, EGFR mutations were even detected in 10 cytological specimens that contained insufficient tumor cells. EGFR mutation testing with BLF specimens is therefore a useful and reliable method, particularly when sufficient cancer cells are not obtained. PMID:29399190

Iron biomineralization of brain tissue and neurodegenerative disorders

NASA Astrophysics Data System (ADS)

Mikhaylova (Mikhailova), Albina

The brain is an organ with a high concentration of iron in specific areas, particularly in the globus pallidus, the substantia nigra, and the red nucleus. In certain pathological states, such as iron overload disease and neurodegenerative disorders, a disturbed iron metabolism can lead to increased accumulation of iron not only in these areas, but also in the brain regions that are typically low in iron content. Recent studies of the physical and magnetic properties of metalloproteins, and in particular the discovery of biogenic magnetite in human brain tissue, have raised new questions about the role of biogenic iron formations in living organisms. Further investigations revealed the presence of magnetite-like crystalline structures in human ferritin, and indicated that released ferritin iron might act as promoter of oxidative damage to tissue, therefore contributing to pathogenesis of neurodegenerative disorders such as Alzheimer's, Parkinson's and Huntington's diseases. The purpose of this work was to examine the elemental composition and structure of iron deposits in normal brain tissue as well as tissue affected by neurodegenerative disorders. Employing the methods of X-ray microfocus fluorescence mapping, X-ray Absorption Near Edge Structure (XANES), X-ray Absorption Fine Structure spectroscopy (XAFS), and light and electron microscopic examinations allows one to obtain qualitative as well as quantitative data with respect to the cellular distribution and chemical state of iron at levels not detected previously. The described tissue preparation technique allows not only satisfactory XAS iron elemental imaging in situ but also multimodal examination with light and electron microscopes of the same samples. The developed protocol has assured consistent and reproducible results on relatively large sections of flat-embedded tissue. The resulting tissue samples were adequate for XAS examination as well as sufficiently well-preserved for future microscopy studies. The continued development of this technique should lead to major advances in mapping iron anomalies and the related chemical and structural information directly to cells and tissue structures in human brain tissue. At present this is done primarily by iron staining methods and any information on the relationship between iron distribution and cellular structures obtained this way is limited. Iron staining also offers no information on the specific compounds of iron that are present. This can be vitally important as the form of iron [including its oxidation state] in the human body can determine whether it plays a detrimental or beneficial role in neurophysiological processes.

Gene expression of stem cells at different stages of ontological human development.

PubMed

Allegra, Adolfo; Altomare, Roberta; Curcio, Patrizia; Santoro, Alessandra; Lo Monte, Attilio I; Mazzola, Sergio; Marino, Angelo

2013-10-01

To compare multipotent mesenchymal stem cells (MSCs) obtained from chorionic villi (CV), amniotic fluid (AF) and placenta, with regard to their phenotype and gene expression, in order to understand if MSCs derived from different extra-embryonic tissues, at different stages of human ontological development, present distinct stemness characteristics. MSCs obtained from 30 samples of CV, 30 of AF and 10 placentas (obtained from elective caesarean sections) were compared. MSCs at second confluence cultures were characterized by immunophenotypic analysis with flow cytometry using FACS CANTO II. The expression of the genes Oct-4 (Octamer-binding transcription factor 4, also known as POU5F1), Sox-2 (SRY box-containing factor 2), Nanog, Rex-1 (Zfp-42) and Pax-6 (Paired Box Protein-6), was analyzed. Real-time quantitative PCR was performed by ABI Prism 7700, after RNA isolation and retro-transcription in cDNA. Statistical analysis was performed using non-parametric test Kruskal-Wallis (XLSTAT 2011) and confirmed by REST software, to estimate fold changes between samples. Each gene was defined differentially expressed if p-value was <0.05. Cells from all samples were negative for haematopoietic antigens CD45, CD34, CD117 and CD33 and positive for the typical MSCs antigens CD13, CD73 and CD90. Nevertheless, MSCs from AF and placentas showed different fluorescence intensity, reflecting the heterogeneity of these tissues. The gene expression of OCT-4, SOX-2, NANOG was not significantly different among the three groups. In AF, REX-1 and PAX-6 showed a higher expression in comparison to CV. MSCs of different extra-embryonic tissues showed no differences in immunophenotype when collected from second confluence cultures. The expression of OCT-4, NANOG and SOX-2 was not significantly different, demonstrating that all fetal sources are suitable for obtaining MSCs. These results open new possibilities for the clinical use of MSCs derived from easily accessible sources, in order to develop new protocols for clinical and experimental research. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

Gelatinase activity in synovial fluid and synovium obtained from healthy and osteoarthritic joints of dogs.

PubMed

Volk, Susan W; Kapatkin, Amy S; Haskins, Mark E; Walton, Raquel M; D'Angelo, Marina

2003-10-01

To determine matrix metalloproteinase (MMP) activity in synovial fluid (SF) obtained from the joints of dogs with degenerative joint disease (DJD) secondary to various underlying conditions. 35 samples of SF obtained from 18 clinically normal (control) dogs and 34 samples of SF obtained from 17 dogs with DJD; dogs with DJD were from 2 populations (client-owned dogs and research dogs that had DJD secondary to the lysosomal storage disease mucopolysaccharidosis VII). MMP activity in samples of SF was semiquantitatively examined by use of gelatin or casein zymography. Western blot analysis was performed by use of antibodies for MMP-2 and MMP-9. In addition, in situ MMP activity was observed in sections of synovial membrane obtained from healthy and osteoarthritic joints. Samples of SF from osteoarthritic joints had higher MMP-2 activity and dramatically increased MMP-9 activity, compared with values for healthy joints. Substrate-overlay analyses indicated minimal gelatin-degrading activity in synoviocytes obtained from control dogs, whereas greater activity was seen in osteoarthritic synoviocytes, with additional activity in the underlying tissue. Higher MMP-2 activity and dramatic increases in MMP-9 activity were associated with the osteoarthritic state, even though MMP-2 activity was detected in healthy joints. This study expands information on MMP production in SF of osteoarthritic joints in other species and documents the similarity of MMP activity patterns regardless of the cause of DJD.

Recent progress in tissue optical clearing for spectroscopic application

NASA Astrophysics Data System (ADS)

Sdobnov, A. Yu.; Darvin, M. E.; Genina, E. A.; Bashkatov, A. N.; Lademann, J.; Tuchin, V. V.

2018-05-01

This paper aims to review recent progress in optical clearing of the skin and over naturally turbid biological tissues and blood using this technique in vivo and in vitro with multiphoton microscopy, confocal Raman microscopy, confocal microscopy, NIR spectroscopy, optical coherence tomography, and laser speckle contrast imaging. Basic principles of the technique, its safety, advantages and limitations are discussed. The application of optical clearing agent on a tissue allows for controlling the optical properties of tissue. Optical clearing-induced reduction of tissue scattering significantly facilitates the observation of deep-located tissue regions, at the same time improving the resolution and image contrast for a variety of optical imaging methods suitable for clinical applications, such as diagnostics and laser treatment of skin diseases, mucosal tumor imaging, laser disruption of pathological abnormalities, etc. Structural images of different skin layers obtained ex vivo for porcine ear skin samples at application of Omnipaque™ and glycerol solutions during 60 min. Red color corresponds to TPEAF signal channel. Green color corresponds to SHG signal channel.

Imaging of the interaction of low frequency electric fields with biological tissues by optical coherence tomography

NASA Astrophysics Data System (ADS)

Peña, Adrian F.; Devine, Jack; Doronin, Alexander; Meglinski, Igor

2014-03-01

We report the use of conventional Optical Coherence Tomography (OCT) for visualization of propagation of low frequency electric field in soft biological tissues ex vivo. To increase the overall quality of the experimental images an adaptive Wiener filtering technique has been employed. Fourier domain correlation has been subsequently applied to enhance spatial resolution of images of biological tissues influenced by low frequency electric field. Image processing has been performed on Graphics Processing Units (GPUs) utilizing Compute Unified Device Architecture (CUDA) framework in the frequencydomain. The results show that variation in voltage and frequency of the applied electric field relates exponentially to the magnitude of its influence on biological tissue. The magnitude of influence is about twice more for fresh tissue samples in comparison to non-fresh ones. The obtained results suggest that OCT can be used for observation and quantitative evaluation of the electro-kinetic changes in biological tissues under different physiological conditions, functional electrical stimulation, and potentially can be used non-invasively for food quality control.

Multiphoton gradient index endoscopy for evaluation of diseased human prostatic tissue ex vivo

NASA Astrophysics Data System (ADS)

Huland, David M.; Jain, Manu; Ouzounov, Dimitre G.; Robinson, Brian D.; Harya, Diana S.; Shevchuk, Maria M.; Singhal, Paras; Xu, Chris; Tewari, Ashutosh K.

2014-11-01

Multiphoton microscopy can instantly visualize cellular details in unstained tissues. Multiphoton probes with clinical potential have been developed. This study evaluates the suitability of multiphoton gradient index (GRIN) endoscopy as a diagnostic tool for prostatic tissue. A portable and compact multiphoton endoscope based on a 1-mm diameter, 8-cm length GRIN lens system probe was used. Fresh ex vivo samples were obtained from 14 radical prostatectomy patients and benign and malignant areas were imaged and correlated with subsequent H&E sections. Multiphoton GRIN endoscopy images of unfixed and unprocessed prostate tissue at a subcellular resolution are presented. We note several differences and identifying features of benign versus low-grade versus high-grade tumors and are able to identify periprostatic tissues such as adipocytes, periprostatic nerves, and blood vessels. Multiphoton GRIN endoscopy can be used to identify both benign and malignant lesions in ex vivo human prostate tissue and may be a valuable diagnostic tool for real-time visualization of suspicious areas of the prostate.

Structural and molecular interrogation of intact biological systems

PubMed Central

Chung, Kwanghun; Wallace, Jenelle; Kim, Sung-Yon; Kalyanasundaram, Sandhiya; Andalman, Aaron S.; Davidson, Thomas J.; Mirzabekov, Julie J.; Zalocusky, Kelly A.; Mattis, Joanna; Denisin, Aleksandra K.; Pak, Sally; Bernstein, Hannah; Ramakrishnan, Charu; Grosenick, Logan; Gradinaru, Viviana; Deisseroth, Karl

2014-01-01

Obtaining high-resolution information from a complex system, while maintaining the global perspective needed to understand system function, represents a key challenge in biology. Here we address this challenge with a method (termed CLARITY) for the transformation of intact tissue into a nanoporous hydrogel-hybridized form (crosslinked to a three-dimensional network of hydrophilic polymers) that is fully assembled but optically transparent and macromolecule-permeable. Using mouse brains, we show intact-tissue imaging of long-range projections, local circuit wiring, cellular relationships, subcellular structures, protein complexes, nucleic acids and neurotransmitters. CLARITY also enables intact-tissue in situ hybridization, immunohistochemistry with multiple rounds of staining and de-staining in non-sectioned tissue, and antibody labelling throughout the intact adult mouse brain. Finally, we show that CLARITY enables fine structural analysis of clinical samples, including non-sectioned human tissue from a neuropsychiatric-disease setting, establishing a path for the transmutation of human tissue into a stable, intact and accessible form suitable for probing structural and molecular underpinnings of physiological function and disease. PMID:23575631

MicroRNA expression in melanocytic nevi: the usefulness of formalin-fixed, paraffin-embedded material for miRNA microarray profiling.

PubMed

Glud, Martin; Klausen, Mikkel; Gniadecki, Robert; Rossing, Maria; Hastrup, Nina; Nielsen, Finn C; Drzewiecki, Krzysztof T

2009-05-01

MicroRNAs (miRNAs) are small, noncoding RNA molecules that regulate cellular differentiation, proliferation, and apoptosis. MiRNAs are expressed in a developmentally regulated and tissue-specific manner. Aberrant expression may contribute to pathological processes such as cancer, and miRNA may therefore serve as biomarkers that may be useful in a clinical environment for diagnosis of various diseases. Most miRNA profiling studies have used fresh tissue samples. However, in some types of cancer, including malignant melanoma, fresh material is difficult to obtain from primary tumors, and most surgical specimens are formalin fixed and paraffin embedded (FFPE). To explore whether FFPE material would be suitable for miRNA profiling in melanocytic lesions, we compared miRNA expression patterns in FFPE versus fresh frozen samples, obtained from 15 human melanocytic nevi. Out of microarray data, we identified 84 miRNAs that were expressed in both types of samples and represented an miRNA profile of melanocytic nevi. Our results showed a high correlation in miRNA expression (Spearman r-value of 0.80) between paired FFPE and fresh frozen material. The data were further validated by quantitative RT-PCR. In conclusion, FFPE specimens of melanocytic lesions are suitable as a source for miRNA microarray profiling.

Near-infrared optical properties of ex-vivo human skin and subcutaneous tissues using reflectance and transmittance measurements

NASA Astrophysics Data System (ADS)

Simpson, Rebecca; Laufer, Jan G.; Kohl-Bareis, Matthias; Essenpreis, Matthias; Cope, Mark

1997-08-01

The vast majority of 'non-invasive' measurements of human tissues using near infrared spectroscopy rely on passing light through the dermis and subdermis of the skin. Accurate knowledge of the optical properties of these tissues is essential to put into models of light transport and predict the effects of skin perfusion on measurements of deep tissue. Additionally, the skin could be a useful accessible organ for non-invasively determining the constituents of blood flowing through it. Samples of abdominal human skin (including subdermal tissue) were obtained from either post mortem examinations or plastic surgery. The samples were separated into a dermal layer (epidermis and dermis, 1.5 to 2 mm tick), and a sub-cutaneous layer comprised largely of fat. They were enclosed between two glass coverslips and placed in an integrating sphere to measure their reflectance and transmittance over a range of wavelengths from 600 to 1000 nm. The reflectance and transmittance values were converted into average absorption and reduced scattering coefficients by comparison with a Monte Carlo model of light transport. Improvements to the Monte Carlo model and measurement technique removed some previous uncertainties. The results show excellent separation of reduced scattering and absorption coefficient, with clear absorption peaks of hemoglobin, water and lipid. The effect of tissue storage upon measured optical properties was investigated.

Quantitative assessment of soft tissue deformation using digital speckle pattern interferometry: studies on phantom breast models.

PubMed

Karuppanan, Udayakumar; Unni, Sujatha Narayanan; Angarai, Ganesan R

2017-01-01

Assessment of mechanical properties of soft matter is a challenging task in a purely noninvasive and noncontact environment. As tissue mechanical properties play a vital role in determining tissue health status, such noninvasive methods offer great potential in framing large-scale medical screening strategies. The digital speckle pattern interferometry (DSPI)-based image capture and analysis system described here is capable of extracting the deformation information from a single acquired fringe pattern. Such a method of analysis would be required in the case of the highly dynamic nature of speckle patterns derived from soft tissues while applying mechanical compression. Soft phantoms mimicking breast tissue optical and mechanical properties were fabricated and tested in the DSPI out of plane configuration set up. Hilbert transform (HT)-based image analysis algorithm was developed to extract the phase and corresponding deformation of the sample from a single acquired fringe pattern. The experimental fringe contours were found to correlate with numerically simulated deformation patterns of the sample using Abaqus finite element analysis software. The extracted deformation from the experimental fringe pattern using the HT-based algorithm is compared with the deformation value obtained using numerical simulation under similar conditions of loading and the results are found to correlate with an average %error of 10. The proposed method is applied on breast phantoms fabricated with included subsurface anomaly mimicking cancerous tissue and the results are analyzed.

Rapid and accurate peripheral nerve detection using multipoint Raman imaging (Conference Presentation)

NASA Astrophysics Data System (ADS)

Kumamoto, Yasuaki; Minamikawa, Takeo; Kawamura, Akinori; Matsumura, Junichi; Tsuda, Yuichiro; Ukon, Juichiro; Harada, Yoshinori; Tanaka, Hideo; Takamatsu, Tetsuro

2017-02-01

Nerve-sparing surgery is essential to avoid functional deficits of the limbs and organs. Raman scattering, a label-free, minimally invasive, and accurate modality, is one of the best candidate technologies to detect nerves for nerve-sparing surgery. However, Raman scattering imaging is too time-consuming to be employed in surgery. Here we present a rapid and accurate nerve visualization method using a multipoint Raman imaging technique that has enabled simultaneous spectra measurement from different locations (n=32) of a sample. Five sec is sufficient for measuring n=32 spectra with good S/N from a given tissue. Principal component regression discriminant analysis discriminated spectra obtained from peripheral nerves (n=863 from n=161 myelinated nerves) and connective tissue (n=828 from n=121 tendons) with sensitivity and specificity of 88.3% and 94.8%, respectively. To compensate the spatial information of a multipoint-Raman-derived tissue discrimination image that is too sparse to visualize nerve arrangement, we used morphological information obtained from a bright-field image. When merged with the sparse tissue discrimination image, a morphological image of a sample shows what portion of Raman measurement points in arbitrary structure is determined as nerve. Setting a nerve detection criterion on the portion of "nerve" points in the structure as 40% or more, myelinated nerves (n=161) and tendons (n=121) were discriminated with sensitivity and specificity of 97.5%. The presented technique utilizing a sparse multipoint Raman image and a bright-field image has enabled rapid, safe, and accurate detection of peripheral nerves.

Nanobacteria may be linked to calcification in placenta.

PubMed

Lu, He; Guo, Ya-nan; Liu, Sheng-nan; Zhang, De-chun

2012-05-01

Placental calcification is a common pathologic condition in obstetrics. To detect the bacteria infection mechanisms for calcification, an experiment was performed to isolate, culture, and identify the nanobacteria in placental calcification. Sixteen cases of placental calcification of pregnant women were collected for the purpose of the isolation of nanobacteria, cultivation, and identification of 16S rDNA sequence. Under transmission electron microscope, novel oval-shape nanobacteria-like particles (NLP) in extracellular matrix of calcified placenta tissues were found with 50-500 nm in diameter, and among hydroxyapatite crystals aggregation existed. After about 4 weeks of culturing and isolating NLP from these calcified tissues, all calcified placental tissue samples and one adjacent tissue of calcified placental tissue samples showed white granular depositions, which were firmly attached to the bottom of the culture tubes and visible to the naked eyes. In the control group they could not be seen. After PCR was amplified a 1407-bp fragment was obtained and submitted to GenBank after sequencing with accession number JN029830. The 16S rDNA sequence homology between the isolation strain and strain nanobacteria (X98418) was 92% in GenBank. For the first time isolated, cultured, and identified nanobacteria in placental calcification indicated that nanobacteria infection is related to placental calcification.

Tc17 cells in patients with uterine cervical cancer.

PubMed

Zhang, Yan; Hou, Fei; Liu, Xin; Ma, Daoxin; Zhang, Youzhong; Kong, Beihua; Cui, Baoxia

2014-01-01

The existence of Tc17 cells was recently shown in several types of infectious and autoimmune diseases, but their distribution and functions in uterine cervical cancer (UCC) have not been fully elucidated. The frequency of Tc17 cells in peripheral blood samples obtained from UCC patients, cervical intraepithelial neoplasia (CIN) patients and healthy controls was determined by flow cytometry. Besides, the prevalence of Tc17 cells and their relationships to Th17 cells and Foxp3-expressing T cells as well as microvessels in tissue samples of the patients were assessed by immunohistochemistry staining. Compared to controls, patients with UCC or CIN had a higher proportion of Tc17 cells in both peripheral blood and cervical tissues, but the level of Tc17 cells in UCC tissues was significantly higher than that in CIN tissues. Besides, the increased level of Tc17 in UCC patients was associated with the status of pelvic lymph node metastases and increased microvessel density. Finally, significant correlations of infiltration between Tc17 cells and Th17 cells or Foxp3-expressing T cells were observed in UCC and CIN tissues. This study indicates that Tc17 cell infiltration in cervical cancers is associated with cancer progression accompanied by increased infiltrations of Th17 cells and regulatory T cells as well as promoted tumor vasculogenesis.

A comparative study of three tissue-cultured Dendrobium species and their wild correspondences by headspace gas chromatography-mass spectrometry combined with chemometric methods.

PubMed

Chen, Nai-Dong; You, Tao; Li, Jun; Bai, Li-Tao; Hao, Jing-Wen; Xu, Xiao-Yuan

2016-10-01

Plant tissue culture technique is widely used in the conservation and utilization of rare and endangered medicinal plants and it is crucial for tissue culture stocks to obtain the ability to produce similar bioactive components as their wild correspondences. In this paper, a headspace gas chromatography-mass spectrometry method combined with chemometric methods was applied to analyze and evaluate the volatile compounds in tissue-cultured and wild Dendrobium huoshanense Cheng and Tang, Dendrobium officinale Kimura et Migo and Dendrobium moniliforme (Linn.) Sw. In total, 63 volatile compounds were separated, with 53 being identified from the three Dendrobium spp. Different provenances of Dendrobiums had characteristic chemicals and showed remarkable quantity discrepancy of common compositions. The similarity evaluation disclosed that the accumulation of volatile compounds in Dendrobium samples might be affected by their provenance. Principal component analysis showed that the first three components explained 85.9% of data variance, demonstrating a good discrimination between samples. Gas chromatography-mass spectrometry techniques, combined with chemometrics, might be an effective strategy for identifying the species and their provenance, especially in the assessment of tissue-cultured Dendrobium quality for use in raw herbal medicines. Copyright © 2016. Published by Elsevier B.V.

The lipid-reactive oxygen species phenotype of breast cancer. Raman spectroscopy and mapping, PCA and PLSDA for invasive ductal carcinoma and invasive lobular carcinoma. Molecular tumorigenic mechanisms beyond Warburg effect.

PubMed

Surmacki, Jakub; Brozek-Pluska, Beata; Kordek, Radzislaw; Abramczyk, Halina

2015-04-07

Vibrational signatures of human breast tissue (invasive ductal carcinoma and invasive lobular carcinoma) were used to identify, characterize and discriminate structures in normal (noncancerous) and cancerous tissues by confocal Raman imaging, Raman spectroscopy and IR spectroscopy. The most important differences between normal and cancerous tissues were found in regions characteristic for vibrations of carotenoids, fatty acids, proteins, and interfacial water. Particular attention was paid to the role played by unsaturated fatty acids and their derivatives. K-means clustering and basis analysis followed by PCA and PLSDA is employed to analyze Raman spectroscopic maps of human breast tissue and for a statistical analysis of the samples (82 patients, 164 samples). Raman maps successfully identify regions of carotenoids, fatty acids, and proteins. The intensities, frequencies and profiles of the average Raman spectra differentiate the biochemical composition of normal and cancerous tissues. The paper demonstrates that Raman imaging has reached a clinically relevant level in regard to breast cancer diagnosis applications. The sensitivity and specificity obtained directly from PLSLD and cross validation are equal to 90.5% and 84.8% for calibration and 84.7% and 71.9% for cross-validation respectively.

1300 nm and 890 nm OCT images of oral cancer tissue engineered models and biopsy samples offer complimentary performance (Conference Presentation)

NASA Astrophysics Data System (ADS)

Boadi, Joseph; Byers, Robert A.; Fernandes, Jon; Mittar, Shweta; Hearnden, Vanessa; Lu, Zenghai; MacNeil, Sheila; Thornhill, Martin; Murdoch, Craig; Hunter, Keith D.; McKechnie, Alasdair; Matcher, Stephen J.

2016-02-01

OCT has demonstrated great potential to non-invasively detect oral epithelial cancers, potentially guiding biopsy and surgical resection. On non-ophthalmic tissues the preferred illumination wavelength is 1300 nm. Previous studies on skin have shown that useful image data can also be obtained at shorter wavelengths, with systems at 1060 nm and 820 nm offering reduced depth penetration but higher contrast. Here we apply a similar comparison to tissue engineered models of oral cancer and also to human biopsy samples, generally finding a similar trend. 1300 nm multi-beam OCT (Michelson Diagnostics EX1301) visualises stromal structures and surface keratin more clearly, providing useful image contrast down to around 1 mm. This system was compared with an ultra-high resolution home-built system operating at 890 nm (2.5 micron resolution vs 7.5 micron axial resolution for the EX1301). The UHR system reveals epithelial features more clearly, especially in the DOK pre-invasive cell line model and the biopsy samples. The relative effects of center wavelength vs axial resolution in generating the differential, wavelength-dependent contrast are assessed and the OCT biopsy images are compared with contemporary histology.

Simultaneous determination of quinolones in fish by liquid chromatography coupled with fluorescence detection: comparison of sub-2 microm particles and conventional C18 columns.

PubMed

Zhang, Hong; Chen, Si; Lu, Yanbin; Dai, Zhiyuan

2010-07-01

A simple and effective multi-residue analysis method is presented for the extraction and determination of eleven quinolones (pipemidic acid, enoxacin, norfloxacin, ciprofloxacin, lomefloxacin, enrofloxacin, gatifloxacin, difloxacin, oxolinic acid, nalidixic acid and flumequine) in fish tissues. In this study, multi-residue separations on four columns packed with 5 microm or sub-2 microm particles were simultaneously developed for the purpose of comparison. Various gradients were optimized and best resolutions were achieved on each column. A short and sub-2 microm particle-sized HPLC column was chosen for its advantages in analysis time and column performance. Additionally, considering the matrix effect of the complex crude fish tissue, an effective extraction protocol was also established for sample pre-treatment procedure. Good recoveries (71-98%) were obtained from samples fortified with a mix of eleven quinolones at three levels, with satisfactory relative standard deviations and limits of detection. As a result, the sub-2 microm HPLC column and proposed analytical procedures have been evaluated and applied to the analysis of different fish tissues. Detectable residues were observed in 8 of 30 samples, at concentrations ranging from 4.74 to 23.27 microg/kg.

Transjugular Liver Biopsy

PubMed Central

Behrens, George; Ferral, Hector

2012-01-01

Liver biopsy is considered the gold standard for the evaluation of acute and chronic liver disorders. Transjugular liver biopsy (TJLB) was described by Dotter in 1964 and clinically performed for the first time by Hanafee in 1967. TJLB consists of obtaining liver tissue through a rigid cannula introduced into one of the hepatic veins, typically using jugular venous access. The quality of the TJLB specimens has improved so much that the samples obtained by this method are comparable with those obtained with the percutaneous technique. TJLB is indicated for patients with coagulopathy, ascites, peliosis hepatis, morbid obesity, liver transplant, or in patients undergoing a transjugular intrahepatic portosystemic shunt procedure. The technical success rate for a TJLB procedure ranges from 87 to 97%. Sample fragmentation has been reported in 14 to 25% of the TJLB samples. The complication rates are low and range between 1.3% and 6.5%. The purpose of this article is to provide a review of the fundamental aspects of the TJLB procedure, including technique, indications, contraindications, results, and complications. PMID:23729981

Expression of proteins in serum, synovial fluid, synovial membrane, and articular cartilage samples obtained from dogs with stifle joint osteoarthritis secondary to cranial cruciate ligament disease and dogs without stifle joint arthritis.

PubMed

Garner, Bridget C; Kuroki, Keiichi; Stoker, Aaron M; Cook, Cristi R; Cook, James L

2013-03-01

To identify proteins with differential expression between healthy dogs and dogs with stifle joint osteoarthritis secondary to cranial cruciate ligament (CCL) disease. Serum and synovial fluid samples obtained from dogs with stifle joint osteoarthritis before (n = 10) and after (8) surgery and control dogs without osteoarthritis (9) and archived synovial membrane and articular cartilage samples obtained from dogs with stifle joint osteoarthritis (5) and dogs without arthritis (5). Serum and synovial fluid samples were analyzed via liquid chromatography-tandem mass spectrometry; results were compared against a nonredundant protein database. Expression of complement component 3 in archived tissue samples was determined via immunohistochemical methods. No proteins had significantly different expression between serum samples of control dogs versus those of dogs with stifle joint osteoarthritis. Eleven proteins (complement component 3 precursor, complement factor I precursor, apolipoprotein B-100 precursor, serum paraoxonase and arylesterase 1, zinc-alpha-2-glycoprotein precursor, serum amyloid A, transthyretin precursor, retinol-binding protein 4 precursor, alpha-2-macroglobulin precursor, angiotensinogen precursor, and fibronectin 1 isoform 1 preproprotein) had significantly different expression (> 2.0-fold) between synovial fluid samples obtained before surgery from dogs with stifle joint osteoarthritis versus those obtained from control dogs. Complement component 3 was strongly expressed in all (5/5) synovial membrane samples of dogs with stifle joint osteoarthritis and weakly expressed in 3 of 5 synovial membrane samples of dogs without stifle joint arthritis. Findings suggested that the complement system and proteins involved in lipid and cholesterol metabolism may have a role in stifle joint osteoarthritis, CCL disease, or both.