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Sample records for situ hybridization experiments

  1. Triplex in-situ hybridization

    Science.gov (United States)

    Fresco, Jacques R.; Johnson, Marion D.

    2002-01-01

    Disclosed are methods for detecting in situ the presence of a target sequence in a substantially double-stranded nucleic acid segment, which comprises: a) contacting in situ under conditions suitable for hybridization a substantially double-stranded nucleic acid segment with a detectable third strand, said third strand being capable of hybridizing to at least a portion of the target sequence to form a triple-stranded structure, if said target sequence is present; and b) detecting whether hybridization between the third strand and the target sequence has occured.

  2. Standardization and optimization of fluorescence in situ hybridization (FISH) for HER-2 assessment in breast cancer: A single center experience.

    Science.gov (United States)

    Bogdanovska-Todorovska, Magdalena; Petrushevska, Gordana; Janevska, Vesna; Spasevska, Liljana; Kostadinova-Kunovska, Slavica

    2018-05-20

    Accurate assessment of human epidermal growth factor receptor 2 (HER-2) is crucial in selecting patients for targeted therapy. Commonly used methods for HER-2 testing are immunohistochemistry (IHC) and fluorescence in situ hybridization (FISH). Here we presented the implementation, optimization and standardization of two FISH protocols using breast cancer samples and assessed the impact of pre-analytical and analytical factors on HER-2 testing. Formalin fixed paraffin embedded (FFPE) tissue samples from 70 breast cancer patients were tested for HER-2 using PathVysion™ HER-2 DNA Probe Kit and two different paraffin pretreatment kits, Vysis/Abbott Paraffin Pretreatment Reagent Kit (40 samples) and DAKO Histology FISH Accessory Kit (30 samples). The concordance between FISH and IHC results was determined. Pre-analytical and analytical factors (i.e., fixation, baking, digestion, and post-hybridization washing) affected the efficiency and quality of hybridization. The overall hybridization success in our study was 98.6% (69/70); the failure rate was 1.4%. The DAKO pretreatment kit was more time-efficient and resulted in more uniform signals that were easier to interpret, compared to the Vysis/Abbott kit. The overall concordance between IHC and FISH was 84.06%, kappa coefficient 0.5976 (p characteristics. Differences in the pre-analytical and analytical steps can affect the hybridization quality and efficiency. The use of DAKO pretreatment kit is time-saving and cost-effective.

  3. Standardization and optimization of fluorescence in situ hybridization (FISH for HER-2 assessment in breast cancer: A single center experience

    Directory of Open Access Journals (Sweden)

    Magdalena Bogdanovska-Todorovska

    2018-05-01

    Full Text Available Accurate assessment of human epidermal growth factor receptor 2 (HER-2 is crucial in selecting patients for targeted therapy. Commonly used methods for HER-2 testing are immunohistochemistry (IHC and fluorescence in situ hybridization (FISH. Here we presented the implementation, optimization and standardization of two FISH protocols using breast cancer samples and assessed the impact of pre-analytical and analytical factors on HER-2 testing. Formalin fixed paraffin embedded (FFPE tissue samples from 70 breast cancer patients were tested for HER-2 using PathVysion™ HER-2 DNA Probe Kit and two different paraffin pretreatment kits, Vysis/Abbott Paraffin Pretreatment Reagent Kit (40 samples and DAKO Histology FISH Accessory Kit (30 samples. The concordance between FISH and IHC results was determined. Pre-analytical and analytical factors (i.e., fixation, baking, digestion, and post-hybridization washing affected the efficiency and quality of hybridization. The overall hybridization success in our study was 98.6% (69/70; the failure rate was 1.4%. The DAKO pretreatment kit was more time-efficient and resulted in more uniform signals that were easier to interpret, compared to the Vysis/Abbott kit. The overall concordance between IHC and FISH was 84.06%, kappa coefficient 0.5976 (p < 0.0001. The greatest discordance (82% between IHC and FISH was observed in IHC 2+ group. A standardized FISH protocol for HER-2 assessment, with high hybridization efficiency, is necessary due to variability in tissue processing and individual tissue characteristics. Differences in the pre-analytical and analytical steps can affect the hybridization quality and efficiency. The use of DAKO pretreatment kit is time-saving and cost-effective.

  4. In Situ Hybridization Pada Kanker Payudara

    OpenAIRE

    Diah Witari, Ni Putu

    2014-01-01

    Kesulitan yang dijumpai pada penanganan kanker payudara adalah terjadinya kekambuhan atau relaps. Deteksi status HER2 pada pasien merupakan salah satu upaya untuk mendeteksi terjadinya relaps dan juga untuk menentukan jenis terapi yang ada diberikan. Ekspresi protein HER2 dapat dideteksi dengan immunohistochemistry (IHC), sedangkan mutasi gen HER2 dapat dideteksi dengan teknik in situ hybridization baik berupa fluorescence in situ hybridization (FISH) ataupun chromogenic in situ hy...

  5. The Use of Whole-Mount "in Situ" Hybridization to Illustrate Gene Expression Regulation

    Science.gov (United States)

    Llamusí, Beatriz; Muñoz-Soriano, Verónica; Paricio, Nuria; Artero, Rubén

    2014-01-01

    "In situ" hybridization is a widely used technique for studying gene expression. Here, we describe two experiments addressed to postgraduate genetics students in which the effect of transcription factors on gene expression is analyzed in "Drosophila embryos of different genotypes by whole-mount in situ hybridization. In one of the…

  6. Fluorescence In Situ Hybridization for MDM2 Amplification as a Routine Ancillary Diagnostic Tool for Suspected Well-Differentiated and Dedifferentiated Liposarcomas: Experience at a Tertiary Center

    Directory of Open Access Journals (Sweden)

    Khin Thway

    2015-01-01

    Full Text Available Background. The assessment of MDM2 gene amplification by fluorescence in situ hybridization (FISH has become a routine ancillary tool for diagnosing atypical lipomatous tumor (ALT/well-differentiated liposarcoma and dedifferentiated liposarcoma (WDL/DDL in specialist sarcoma units. We describe our experience of its utility at our tertiary institute. Methods. All routine histology samples in which MDM2 amplification was assessed with FISH over a 2-year period were included, and FISH results were correlated with clinical and histologic findings. Results. 365 samples from 347 patients had FISH for MDM2 gene amplification. 170 were positive (i.e., showed MDM2 gene amplification, 192 were negative, and 3 were technically unsatisfactory. There were 122 histologically benign cases showing a histology:FISH concordance rate of 92.6%, 142 WDL/DDL (concordance 96.5%, and 34 cases histologically equivocal for WDL (concordance 50%. Of 64 spindle cell/pleomorphic neoplasms (in which DDL was a differential diagnosis, 21.9% showed MDM2 amplification. Of the cases with discrepant histology and FISH, all but 3 had diagnoses amended following FISH results. For discrepancies of benign histology but positive FISH, lesions were on average larger, more frequently in “classical” (intra-abdominal or inguinal sites for WDL/DDL and more frequently core biopsies. Discrepancies of malignant histology but negative FISH were smaller, less frequently in “classical” sites but again more frequently core biopsies. Conclusions. FISH has a high correlation rate with histology for cases with firm histologic diagnoses of lipoma or WDL/DDL. It is a useful ancillary diagnostic tool in histologically equivocal cases, particularly in WDL lacking significant histologic atypia or DDL without corresponding WDL component, especially in larger tumors, those from intra-abdominal or inguinal sites or core biopsies. There is a significant group of well-differentiated adipocytic neoplasms

  7. Comparison of Fluorescence In Situ Hybridization and Chromogenic In Situ Hybridization for Low and High Throughput HER2 Genetic Testing

    DEFF Research Database (Denmark)

    Poulsen, Tim S; Espersen, Maiken Lise Marcker; Kofoed, Vibeke

    2013-01-01

    cancer patients with HER2 immunohistochemistry (IHC) results scored as 0/1+, 2+, and 3+. HER2 genetic status was analysed using chromogenic in situ hybridization (CISH) and fluorescence in situ hybridization (FISH). Scoring results were documented through digital image analysis. The cancer region...

  8. Application of Genomic In Situ Hybridization in Horticultural Science

    Directory of Open Access Journals (Sweden)

    Fahad Ramzan

    2017-01-01

    Full Text Available Molecular cytogenetic techniques, such as in situ hybridization methods, are admirable tools to analyze the genomic structure and function, chromosome constituents, recombination patterns, alien gene introgression, genome evolution, aneuploidy, and polyploidy and also genome constitution visualization and chromosome discrimination from different genomes in allopolyploids of various horticultural crops. Using GISH advancement as multicolor detection is a significant approach to analyze the small and numerous chromosomes in fruit species, for example, Diospyros hybrids. This analytical technique has proved to be the most exact and effective way for hybrid status confirmation and helps remarkably to distinguish donor parental genomes in hybrids such as Clivia, Rhododendron, and Lycoris ornamental hybrids. The genome characterization facilitates in hybrid selection having potential desirable characteristics during the early hybridization breeding, as this technique expedites to detect introgressed sequence chromosomes. This review study epitomizes applications and advancements of genomic in situ hybridization (GISH techniques in horticultural plants.

  9. Fathead minnow whole-mount in situ hybridization (WISH)

    Data.gov (United States)

    U.S. Environmental Protection Agency — This study demonstrates the potential of whole-mount in situ hybridization (WISH), in conjunction with quantitative real-time polymerase chain reaction (QPCR)...

  10. Applications of in situ hybridization to plant-improvement

    International Nuclear Information System (INIS)

    Abbasi, F.M.

    2004-01-01

    In situ hybridization is a powerful method for characteristic alien addition and substitution lines. RFLP analysis can identify the presence of a particular individual chromosome, but whether they are as a pair or as a single chromosome cannot be determined. In situ hybridization has become established as an essential method in cell and molecular biology. It is able to link DNA sequences with their organization and physical position. The rate of technology-development in the field of in situ hybridization has been rapid: radioactive probes are now rarely used, while labeling methods, fluorochromes, chromosomes and tissue-preparation methods, microscope and imaging technology have all useful in functional genomics and localization of transgenes on the chromosomes. (author)

  11. In situ hybridization; principles and applications: review article

    Directory of Open Access Journals (Sweden)

    Zahra Nozhat

    2015-06-01

    Full Text Available In situ hybridization (ISH is a method that uses labeled complementary single strand DNA or RNA to localize specific DNA or RNA sequences in an intact cell or in a fixed tissue section. The main steps of ISH consist of: probe selection, tissue or sample preparation, pre-hybridization treatment, hybridization and washing, detection and control procedure. Probe selection is one of the important aspects of successful hybridization. ISH sensitivity and specificity can be influenced by: probe construct, efficiency of labeling, percentage of GC, probe length and signal detection systems. Different methods such as nick translation, random priming, end tailing and T4 DNA polymerase replacement are used for probe generation. Both radioactive and non-radioactive labels can be used in order to probe labeling. Nucleic acid maintenance in samples, prevention of morphological changes of samples and probe penetration into tissue section are the main aims of sample preparation step. Then, a small amount of solution containing probe, is added on slides containing tissue sections for hybridization process, then slides are incubated overnight. Next day, washes are carried out to remove the probes which are not bound to target DNA or RNA. Finally, in order to be sure that the observed labeling is specific to the target sequence, using several control procedures is very important. Various techniques based on ISH consist of: Fluorescence in situ hybridization (FISH, chromogenic in situ hybridization (CISH, genomic in situ hybridization (GISH, comparative genomic hybridization (CGH, spectral karyotyping (SKY and multiplex fluorescence in situ hybridization (MFISH. One of the most common techniques of ISH is fluorescence in situ hybridization. FISH can be used to: 1 detect small deletions and duplications that are not visible using microscope analysis, 2 detect how many chromosomes of a certain type are present in each cell and 3 confirm rearrangements that are

  12. Contribution of fluorescence in situ hybridization to biological dosimetry

    International Nuclear Information System (INIS)

    Sorokine-Durm, I.; Roy, L.; Durand, V.; Voisin, P.

    1995-01-01

    Fluorescence in situ hybridization with composite whole chromosome specific DNA probes for human chromosomes 2, 4 and 12 an α-satellite centromeric DNA probe labelled with biotin were used to measure symmetrical and terminal translocations (dose rate 0.5 Gy/min) and dicentrics (0.1 Gy/min) induced in vitro by 60 Co γ-irradiation (0-5 Gy). The suitability of fluorescence in situ hybridization (F.I.S.H.) technique for dicentrics detection is compared with the conventional technique. Dose-response curves for γ-rays ( 60 Co) for two dose rates are shown (dicentrics and translocations). (authors). 10 refs., 2 figs

  13. Modulation of tyrosine hydroxylase gene expression in the central nervous system visualized by in situ hybridization

    International Nuclear Information System (INIS)

    Berod, A.; Biguet, N.F.; Dumas, S.; Bloch, B.; Mallet, J.

    1987-01-01

    cDNA probe was used for in situ hybridization studies on histological sections through the locus coeruleus, substantia nigra, and the ventral tegmental area of the rat brain. Experimental conditions were established that yielded no background and no signal when pBR322 was used as control probe. Using the tyrosine hydroxylase probe, the authors ascertained the specificity of the labeling over catecholaminergic cells by denervation experiments and comparison of the hybridization pattern with that of immunoreactivity. The use of 35 S-labeled probe enabled the hybridization signal to be resolved at the cellular level. A single injection of reserpine into the rat led to an increase of the intensity of the autoradiographic signal over the locus coeruleus area, confirming an RNA gel blot analysis. The potential of in situ hybridization to analyze patterns of modulation of gene activity as a result of nervous activity is discussed

  14. Supernumerary ring chromosome 20 characterized by fluorescence in situ hybridization

    NARCIS (Netherlands)

    Van Langen, Irene M.; Otter, Mariëlle A.; Aronson, Daniël C.; Overweg-Plandsoen, W.C.G.; Hennekam, Raoul C.M.; Leschot, Nico J.; Hoovers, Jan M.N.

    1996-01-01

    We report on a boy with mild dysmorphic features and developmental delay, in whom karyotyping showed an additional minute ring chromosome in 60% of metaphases. Fluorescence in situ hybridization (FISH) with a centromere specific probe demonstrated that the ring chromosome contained the centromeric

  15. Diagnosis, prognosis and disease management using in situ hybridization

    International Nuclear Information System (INIS)

    Kucheria, K.; Talwar, R.

    2002-01-01

    cytogenetics were identified using ISH technique. The SRY region was detected in few cases of ambiguous genitalia that lacked the Y chromosome. Most leukemia patients who were in complete cytogenetic remission showed evidence of minimal residual disease when analyzed for bcr/abl and PML/RARα fusion genes. Aneuploidies and gene rearrangements could also be detected on interphase (non-dividing) cells obtained from peripheral blood and bone marrow. Further, many targets could be visualized in the same experiment using probes labeled with different coloured fluorescent dyes. To conclude, the present study stresses that for analysis of chromosomal rearrangements at the molecular level, conventional cytogenetics needs to be supplemented with highly sensitive molecular methods like In Situ Hybridization. Further, Interphase cytogenetics is a simple, rapid and efficient technique that can add to the diagnostic utility of conventional cytogenetics by decreasing the time interval between sampling and diagnosis

  16. Fluorescence in situ hybridization with reference to biodosimetry: a review

    International Nuclear Information System (INIS)

    Venkatachalam, P.; Paul, S.F.D.; Jeevanram, R.K.

    1996-01-01

    Many advances have taken place in the field of radiation biodosimetry in the recent past. Measurement of dicentric chromosome aberrations, was first developed and followed by micronuclei scoring. These, however, are unstable type aberrations and the cells carrying such aberrations are eliminated from the body in few years. They are therefore of use primarily in case of accidental exposures. The challenge is to measure the cumulative radiation exposure resulting from normal operations by measuring stable chromosome aberrations. Banding technique can measure stable chromosome aberration but require long time to analyse the banding pattern to study translocations. On the other hand fluorescence in situ hybridization (FISH) technique is sensitive, fast and easy to identify the translocations as the chromosomes involved in translocation are painted with different colours. This review brings out the requirements of various materials, their preparations, method of detection of fluorescence etc. for carrying out FISH. The experience of various laboratories using FISH in the monitoring of radiation absorbed dose is discussed. (author)

  17. Design and optimization of hybrid ex situ/in situ steam generation recovery processes for heavy oil and bitumen

    Energy Technology Data Exchange (ETDEWEB)

    Yang, X.; Gates, I.D. [Calgary Univ., AB (Canada). Dept. of Chemical and Petroleum Engineering; Larter, S.R. [Calgary Univ., AB (Canada). Dept. of Geoscience]|[Alberta Ingenuity Centre for In Situ Energy, Edmonton, AB (Canada)

    2008-10-15

    Hybrid steam-air based oil recovery techniques were investigated using advanced 3-D reactive thermal reservoir simulations. The hybrid techniques combined ex situ steam and in situ steam generation processes in order to raise efficiency, lower natural gas consumption, and reduce gas emissions. The steam-air based processes used 70 per cent of the energy of conventional steam assisted gravity drainage (SAGD) techniques to recover the same amount of oil. The process used an SAGD wellpair arrangement, where steam and air were injected through the top injection well. The kinetic parameters used in the study were developed by history matching a combustion tube experiments with Athabasca bitumen conducted to predict cumulative bitumen and gas production volumes and compositions. A total of 6 SAGD and 6 in situ combustion simulations were conducted with steam oxygen volume ratios set at 50 per cent steam and 50 per cent oxygen. Various case studies were considered over a 5 year period. Carbon dioxide (CO{sub 2}) emissions were also measured as well as cumulative water and methane consumption rates. Results of the study were used to develop an optimized hybrid operation that consisted of a SAGD well pair arrangement operating with cyclic steam-oxygen injection at high pressures. It was concluded that the high pressure operation increased the steam partial pressure within the reservoir and enhanced combustion performance. A 29 per cent improvement in the cumulative energy to oil ratio was obtained. 23 refs., 2 tabs., 9 figs.

  18. Genome reorganization in Nicotiana asymmetric somatic hybrids analysed by in situ hybridization

    International Nuclear Information System (INIS)

    Parokonny, A.S.; Kenton, A.Y.; Gleba, Y.Y.; Bennett, M.D.

    1992-01-01

    In situ hybridization was used to examine genome reorganization in asymmetric somatic hybrids between Nicotiana plumbaginifolia and Nicotiana sylvestris obtained by fusion of gamma-irradiated protoplasts from one of the parents (donor) with non-irradiated protoplasts from the other (recipient). Probing with biotinylated total genomic DNA from either the donor or the recipient species unequivocally identified genetic material from both parents in 31 regenerant plants, each originating from a different nuclear hybrid colony. This method, termed genomic in situ hybridization (GISH), allowed intergenomic translocations containing chromosome segments from both species to be recognized in four regenerants. A probe homologous to the consensus sequence of the Arabidopsis thaliana telomeric repeat (5'-TTTAGGG-3')n, identified telomeres on all chromosomes, including 'mini-chromosomes' originating from the irradiated donor genome. Genomic in situ hybridization to plant chromosomes provides a rapid and reliable means of screening for recombinant genotypes in asymmetric somatic hybrids. Used in combination with other DNA probes, it also contributes to a greater understanding of the events responsible for genomic recovery and restabilization following genetic manipulation in vitro

  19. Detection of dengue group viruses by fluorescence in situ hybridization

    Directory of Open Access Journals (Sweden)

    Raquin Vincent

    2012-10-01

    Full Text Available Abstract Background Dengue fever (DF and dengue hemorrhagic fever (DHF represent a global challenge in public health. It is estimated that 50 to 100 million infections occur each year causing approximately 20,000 deaths that are usually linked to severe cases like DHF and dengue shock syndrome. The causative agent of DF is dengue virus (genus Flavivirus that comprises four distinct serotypes (DENV-1 to DENV-4. Fluorescence in situ hybridization (FISH has been used successfully to detect pathogenic agents, but has not been implemented in detecting DENV. To improve our understanding of DENV infection and dissemination in host tissues, we designed specific probes to detect DENV in FISH assays. Methods Oligonucleotide probes were designed to hybridize with RNA from the broadest range of DENV isolates belonging to the four serotypes, but not to the closest Flavivirus genomes. Three probes that fit the criteria defined for FISH experiments were selected, targeting both coding and non-coding regions of the DENV genome. These probes were tested in FISH assays against the dengue vector Aedes albopictus (Diptera: Culicidae. The FISH experiments were led in vitro using the C6/36 cell line, and in vivo against dissected salivary glands, with epifluorescence and confocal microscopy. Results The three 60-nt oligonucleotides probes DENV-Probe A, B and C cover a broad range of DENV isolates from the four serotypes. When the three probes were used together, specific fluorescent signals were observed in C6/36 infected with each DENV serotypes. No signal was detected in either cells infected with close Flavivirus members West Nile virus or yellow fever virus. The same protocol was used on salivary glands of Ae. albopictus fed with a DENV-2 infectious blood-meal which showed positive signals in the lateral lobes of infected samples, with no significant signal in uninfected mosquitoes. Conclusion Based on the FISH technique, we propose a way to design and use

  20. Double-staining chromogenic in situ hybridization as a useful alternative to split-signal fluorescence in situ hybridization in lymphoma diagnostics

    DEFF Research Database (Denmark)

    van Rijk, A.; Svenstroup-Poulsen, T.; Jones, M.

    2010-01-01

    within the reach of every pathology laboratory. Design and Methods Our study was initiated to determine the consistency between chromogenic in situ hybridization and fluorescence in situ hybridization, both using split-signal probes developed for the detection of chromosomal breaks. Five hundred...... and actual signal were compared to the original fluorescence hybridization results. In addition, hematoxylin background staining intensity and signal intensity of the double-staining chromogenic in situ hybridization procedure were analyzed. Results With respect to the presence or absence of chromosomal...

  1. 21 CFR 866.4700 - Automated fluorescence in situ hybridization (FISH) enumeration systems.

    Science.gov (United States)

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Automated fluorescence in situ hybridization (FISH... Laboratory Equipment and Reagents § 866.4700 Automated fluorescence in situ hybridization (FISH) enumeration... Hybridization (FISH) Enumeration Systems.” See § 866.1(e) for the availability of this guidance document. [70 FR...

  2. In Situ Synthesis of Metal Nanoparticle Embedded Hybrid Soft Nanomaterials.

    Science.gov (United States)

    Divya, Kizhmuri P; Miroshnikov, Mikhail; Dutta, Debjit; Vemula, Praveen Kumar; Ajayan, Pulickel M; John, George

    2016-09-20

    The allure of integrating the tunable properties of soft nanomaterials with the unique optical and electronic properties of metal nanoparticles has led to the development of organic-inorganic hybrid nanomaterials. A promising method for the synthesis of such organic-inorganic hybrid nanomaterials is afforded by the in situ generation of metal nanoparticles within a host organic template. Due to their tunable surface morphology and porosity, soft organic materials such as gels, liquid crystals, and polymers that are derived from various synthetic or natural compounds can act as templates for the synthesis of metal nanoparticles of different shapes and sizes. This method provides stabilization to the metal nanoparticles by the organic soft material and advantageously precludes the use of external reducing or capping agents in many instances. In this Account, we exemplify the green chemistry approach for synthesizing these materials, both in the choice of gelators as soft material frameworks and in the reduction mechanisms that generate the metal nanoparticles. Established herein is the core design principle centered on conceiving multifaceted amphiphilic soft materials that possess the ability to self-assemble and reduce metal ions into nanoparticles. Furthermore, these soft materials stabilize the in situ generated metal nanoparticles and retain their self-assembly ability to generate metal nanoparticle embedded homogeneous organic-inorganic hybrid materials. We discuss a remarkable example of vegetable-based drying oils as host templates for metal ions, resulting in the synthesis of novel hybrid nanomaterials. The synthesis of metal nanoparticles via polymers and self-assembled materials fabricated via cardanol (a bioorganic monomer derived from cashew nut shell liquid) are also explored in this Account. The organic-inorganic hybrid structures were characterized by several techniques such as UV-visible spectroscopy, scanning electron microscopy (SEM), and

  3. In situ hybridization at the electron microscope level: hybrid detection by autoradiography and colloidal gold.

    Science.gov (United States)

    Hutchison, N J; Langer-Safer, P R; Ward, D C; Hamkalo, B A

    1982-11-01

    In situ hybridization has become a standard method for localizing DNA or RNA sequences in cytological preparations. We developed two methods to extend this technique to the transmission electron microscope level using mouse satellite DNA hybridization to whole mount metaphase chromosomes as the test system. The first method devised is a direct extension of standard light microscope level using mouse satellite DNA hybridization to whole mount metaphase chromosomes as the test system. The first method devised is a direct extension of standard light microscope in situ hybridization. Radioactively labeled complementary RNA (cRNA) is hybridized to metaphase chromosomes deposited on electron microscope grids and fixed in 70 percent ethanol vapor; hybridixation site are detected by autoradiography. Specific and intense labeling of chromosomal centromeric regions is observed even after relatively short exposure times. Inerphase nuclei present in some of the metaphase chromosome preparations also show defined paatterms of satellite DNA labeling which suggests that satellite-containing regions are associate with each other during interphase. The sensitivity of this method is estimated to at least as good as that at the light microscope level while the resolution is improved at least threefold. The second method, which circumvents the use of autoradiogrphic detection, uses biotin-labeled polynucleotide probes. After hybridization of these probes, either DNA or RNA, to fixed chromosomes on grids, hybrids are detected via reaction is improved at least threefold. The second method, which circumvents the use of autoradiographic detection, uses biotin-labeled polynucleotide probes. After hybridization of these probes, either DNA or RNA, to fixed chromosomes on grids, hybrids are detected via reaction with an antibody against biotin and secondary antibody adsorbed to the surface of over centromeric heterochromatin and along the associated peripheral fibers. Labeling is on average

  4. Data acquisition. GRAAL experiment. Hybrid reactor experiment. AMS experiment

    International Nuclear Information System (INIS)

    Barancourt, D.; Barbier, G.; Bosson, G.; Bouvier, J.; Gallin-Martel, L.; Meillon, B.; Stassi, P.; Tournier, M.

    1997-01-01

    The main activity of the data acquisition team has consisted in hardware and software developments for the GRAAL experiment with the trigger board, for the 'Reacteurs Hybrides' group with an acquisition board ADCVME8V and for the AMS experiment with the monitoring of the aerogel detector. (authors)

  5. Common Fluorescence In Situ Hybridization Applications in Cytology.

    Science.gov (United States)

    Savic, Spasenija; Bubendorf, Lukas

    2016-12-01

    - Fluorescence in situ hybridization (FISH) is a well-established method for detection of genomic aberrations in diagnostic, prognostic, and predictive marker testing. - To review common applications of FISH in cytology. - The published literature was reviewed. - Cytology is particularly well suited for all kinds of FISH applications, which is highlighted in respiratory tract cytology with an increasing demand for predictive FISH testing in lung cancer. Fluorescence in situ hybridization is the gold standard for detection of predictive anaplastic lymphoma kinase gene (ALK) rearrangements, and the same evaluation criteria as in histology apply to cytology. Several other gene rearrangements, including ROS proto-oncogene 1 receptor tyrosine kinase (ROS1), are becoming clinically important and share the same underlining cytogenetic mechanisms with ALK. MET amplification is one of the most common mechanisms of acquired resistance to epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors and can be targeted by crizotinib. As genomic aberrations are a hallmark of malignant cells, FISH is a valuable objective ancillary diagnostic tool. In urinary tract cytology, atypical urothelial cells equivocal for malignancy are a common diagnostic dilemma and multitarget FISH can help clarify such cells. Diagnosis of malignant mesothelioma remains one of the most challenging fields in effusion cytology, and ancillary FISH is useful in establishing the diagnosis. Fluorescence in situ hybridization is a morphology-based technique, and the prerequisite for reliable FISH results is a targeted evaluation of the cells in question (eg, cancer or atypical cells). Cytopathologists and cytotechnicians should therefore be involved in molecular testing in order to select the best material and to provide their morphologic expertise.

  6. 10p Duplication characterized by fluorescence in situ hybridization

    Energy Technology Data Exchange (ETDEWEB)

    Wiktor, A.; Feldman, G.L.; Van Dyke, D.L.; Kratkoczki, P.; Ditmars, D.M. Jr. [Henry Ford Hospital, Detroit, MI (United States)

    1994-09-01

    We describe a patient with severe failure to thrive, mild-moderate developmental delay, cleft lip and palate, and other anomalies. Routine cytogenetic analysis documented a de novo chromosome rearrangement involving chromosome 4, but the origin of the derived material was unknown. Using chromosome specific painting probes, the karyotype was defined as 46,XY,der(4)t(4;10)(q35;p11.23). Characterization of the dup(10p) by fluorescence in situ hybridization (FISH) analysis provides another example of the usefulness of this technology in identifying small deletions, duplications, or supernumerary marker chromosomes. 19 refs., 4 figs.

  7. A novel fluorescent in situ hybridization technique for detection of Rickettsia spp. in archival samples

    DEFF Research Database (Denmark)

    Svendsen, Claus Bo; Boye, Mette; Struve, Carsten

    2009-01-01

    A novel, sensitive and specific method for detecting Rickettsia spp. in archival samples is described. The method involves the use of fluorescently marked oligonucleotide probes for in situ hybridization. Specific hybridization of Ricekttsia was found without problems of cross-reactions with bact......A novel, sensitive and specific method for detecting Rickettsia spp. in archival samples is described. The method involves the use of fluorescently marked oligonucleotide probes for in situ hybridization. Specific hybridization of Ricekttsia was found without problems of cross...

  8. Regulatory pathway analysis by high-throughput in situ hybridization.

    Directory of Open Access Journals (Sweden)

    Axel Visel

    2007-10-01

    Full Text Available Automated in situ hybridization enables the construction of comprehensive atlases of gene expression patterns in mammals. Such atlases can become Web-searchable digital expression maps of individual genes and thus offer an entryway to elucidate genetic interactions and signaling pathways. Towards this end, an atlas housing approximately 1,000 spatial gene expression patterns of the midgestation mouse embryo was generated. Patterns were textually annotated using a controlled vocabulary comprising >90 anatomical features. Hierarchical clustering of annotations was carried out using distance scores calculated from the similarity between pairs of patterns across all anatomical structures. This process ordered hundreds of complex expression patterns into a matrix that reflects the embryonic architecture and the relatedness of patterns of expression. Clustering yielded 12 distinct groups of expression patterns. Because of the similarity of expression patterns within a group, members of each group may be components of regulatory cascades. We focused on the group containing Pax6, an evolutionary conserved transcriptional master mediator of development. Seventeen of the 82 genes in this group showed a change of expression in the developing neocortex of Pax6-deficient embryos. Electromobility shift assays were used to test for the presence of Pax6-paired domain binding sites. This led to the identification of 12 genes not previously known as potential targets of Pax6 regulation. These findings suggest that cluster analysis of annotated gene expression patterns obtained by automated in situ hybridization is a novel approach for identifying components of signaling cascades.

  9. RNA Imaging with Multiplexed Error Robust Fluorescence in situ Hybridization

    Science.gov (United States)

    Moffitt, Jeffrey R.; Zhuang, Xiaowei

    2016-01-01

    Quantitative measurements of both the copy number and spatial distribution of large fractions of the transcriptome in single-cells could revolutionize our understanding of a variety of cellular and tissue behaviors in both healthy and diseased states. Single-molecule Fluorescence In Situ Hybridization (smFISH)—an approach where individual RNAs are labeled with fluorescent probes and imaged in their native cellular and tissue context—provides both the copy number and spatial context of RNAs but has been limited in the number of RNA species that can be measured simultaneously. Here we describe Multiplexed Error Robust Fluorescence In Situ Hybridization (MERFISH), a massively parallelized form of smFISH that can image and identify hundreds to thousands of different RNA species simultaneously with high accuracy in individual cells in their native spatial context. We provide detailed protocols on all aspects of MERFISH, including probe design, data collection, and data analysis to allow interested laboratories to perform MERFISH measurements themselves. PMID:27241748

  10. In situ synthesis of silver benzene-dithiolate hybrid films

    Energy Technology Data Exchange (ETDEWEB)

    Brenier, Roger, E-mail: roger.brenier@univ-lyon1.fr [Institut Lumière Matière, UMR 5306, Université Lyon 1-CNRS, Université de Lyon, Domaine Scientifique de La Doua, Batiment Kastler, 10 rue Ada Byron, 69622 Villeurbanne, Cedex (France); Piednoir, Agnès, E-mail: agnes.piednoir@univ-lyon1.fr [Institut Lumière Matière, UMR 5306, Université Lyon 1-CNRS, Université de Lyon, Domaine Scientifique de La Doua, Batiment Kastler, 10 rue Ada Byron, 69622 Villeurbanne, Cedex (France); Bertorelle, Franck, E-mail: franck.bertorelle@univ-lyon1.fr [Institut Lumière Matière, UMR 5306, Université Lyon 1-CNRS, Université de Lyon, Domaine Scientifique de La Doua, Batiment Kastler, 10 rue Ada Byron, 69622 Villeurbanne, Cedex (France); Penuelas, José, E-mail: jose.penuelas@ec-lyon.fr [Université de Lyon, Institut des Nanotechnologies de Lyon, Ecole Centrale de Lyon, CNRS, UMR 5270, 36 rue Guy de Collongues, F69134 Ecully (France); Grenet, Geneviève, E-mail: genevieve.grenet@ec-lyon.fr [Université de Lyon, Institut des Nanotechnologies de Lyon, Ecole Centrale de Lyon, CNRS, UMR 5270, 36 rue Guy de Collongues, F69134 Ecully (France)

    2016-02-01

    In this article, a method for in situ synthesis of silver benzene-dithiolate hybrid films is presented. Silver nanoparticles, generated on ZrO{sub 2} films, are transformed into silver benzene 1,4-dithiolate or, partially, into silver benzene 1,2-dithiolate after sample immersion in the corresponding thiol solutions. These transformations occur at room temperature owing to the catalytic action of ZrO{sub 2}. It is also shown that TiO{sub 2} in place of ZrO{sub 2} is very efficient, both for the catalytic generation of silver nanoparticles and for their further transformation in benzene 1,4-dithiolate compound. This latter semiconductor has an optical bandgap of about 3 eV and the film is made of touching nanoparticles in an amorphous state. Our work has potential applications in the electronic and photovoltaic fields. - Highlights: • A method for in situ synthesis of silver benzene-dithiolate hybrid semiconductor films is presented. • Silver nanoparticles are, first, generated on ZrO{sub 2} or on TiO{sub 2} coated silica substrates. • The samples are immersed in benzene dithiol solution for two days at room temperature. • During the immersion, the silver nanoparticles are transformed into silver benzene dithiolate. • The silver benzene dithiolate film is made of amorphous nanoparticles with a banbgap of 3 eV.

  11. In situ hybridization of superparamagnetic iron-biomolecule nanoparticles.

    Science.gov (United States)

    Moghimi, Nafiseh; Donkor, Apraku David; Mohapatra, Mamata; Thomas, Joseph Palathinkal; Su, Zhengding; Tang, Xiaowu Shirley; Leung, Kam Tong

    2014-07-23

    The increase in interest in the integration of organic-inorganic nanostructures in recent years has promoted the use of hybrid nanoparticles (HNPs) in medicine, energy conversion, and other applications. Conventional hybridization methods are, however, often long, complicated, and multistepped, and they involve biomolecules and discrete nanostructures as separate entities, all of which hinder the practical use of the resulting HNPs. Here, we present a novel, in situ approach to synthesizing size-specific HNPs using Fe-biomolecule complexes as the building blocks. We choose an anticancer peptide (p53p, MW 1.8 kDa) and an enzyme (GOx, MW 160 kDa) as model molecules to demonstrate the versatility of the method toward different types of molecules over a large size range. We show that electrostatic interaction for complex formation of metal hydroxide ion with the partially charged side of biomolecule in the solution is the key to hybridization of metal-biomolecule materials. Electrochemical deposition is then used to produce hybrid NPs from these complexes. These HNPs with controllable sizes ranging from 30 nm to 3.5 μm are found to exhibit superparamagnetic behavior, which is a big challenge for particles in this size regime. As an example of greatly improved properties and functionality of the new hybrid material, in vitro toxicity assessment of Fe-GOx HNPs shows no adverse effect, and the Fe-p53p HNPs are found to selectively bind to cancer cells. The superparamagnetic nature of these HNPs (superparamagnetic even above the size regime of 15-20 nm!), their biocompatibility, and the direct integration approach are fundamentally important to biomineralization and general synthesis strategy for bioinspired functional materials.

  12. Comparison of Fluorescence In Situ Hybridization and Chromogenic In Situ Hybridization for Low and High Throughput HER2 Genetic Testing

    Science.gov (United States)

    Poulsen, Tim S.; Espersen, Maiken L. M.; Kofoed, Vibeke; Dabetic, Tanja; Høgdall, Estrid; Balslev, Eva

    2013-01-01

    The purpose was to evaluate and compare 5 different HER2 genetic assays with different characteristics that could affect the performance to analyze the human epidermal growth factor 2 (HER2) gene copy number under low and high throughput conditions. The study included 108 tissue samples from breast cancer patients with HER2 immunohistochemistry (IHC) results scored as 0/1+, 2+, and 3+. HER2 genetic status was analysed using chromogenic in situ hybridization (CISH) and fluorescence in situ hybridization (FISH). Scoring results were documented through digital image analysis. The cancer region of interest was identified from a serial H&E stained slide following tissue cores were transferred to a tissue microarrays (TMA). When using TMA in a routine flow, all patients will be tested for HER2 status with IHC followed by CISH or FISH, thereby providing individual HER2 results. In conclusion, our results show that the differences between the HER2 genetic assays do not have an effect on the analytic performance and the CISH technology is superior to high throughput HER2 genetic testing due to scanning speed, while the IQ-FISH may still be a choice for fast low throughput HER2 genetic testing. PMID:24383005

  13. In situ migration experiment in argillaceous formation

    International Nuclear Information System (INIS)

    Yoshida, Hidekazu

    1990-01-01

    International cooperative R and D has been performed within the five years framework of the bilateral agreement between PNC (Power Reactor and Nuclear Fuel Development Corporation) and SCK/CEN (Studiecentrum voor Kernergie/Centre D'etude de L'energie Nucleaire, Mol, Belgium) which is focused on 'The Migration Experiment in argillaceous formation.' This Tertiary argillaceous formation, called Boom clay, is located at about 230m depth in Mol-Dessel area, Belgium. The argillaceous rock is considered to have a high capability for retardation to radionuclides when they migrate in geosphere because of a high content of clay minerals and dissolved carbon-rich pore water. The main purpose of this collaboration work is to characterize the migration phenomena in sedimentary rock through understanding of the behaviour of radionuclides migration in the argillaceous formation. The present report describes the preliminary results of in situ one-dimensional migration experiment with labelled clay core emplaced in borehole under advective condition. In the experiment, radioactive tracer Sr-85 and Eu-152+154 have been used in order to determine the apparent dispersion coefficient and retardation factor of Boom clay. Finally, the following conclusions were obtained by in situ measurement and calculation based on a appropriate migration model; a) From the Sr-85 experiment, diffusive behavior is interpreted to be a dominant phenomena on radionuclides transportation. b) From the Eu-152+154 experiment, very small non-retarded fraction is observed. (author)

  14. Human cDNA mapping using fluorescence in situ hybridization

    Energy Technology Data Exchange (ETDEWEB)

    Korenberg, J.R.

    1993-03-04

    Genetic mapping is approached using the techniques of high resolution fluorescence in situ hybridization (FISH). This technology and the results of its application are designed to rapidly generate whole genome as tool box of expressed sequence to speed the identification of human disease genes. The results of this study are intended to dovetail with and to link the results of existing technologies for creating backbone YAC and genetic maps. In the first eight months, this approach generated 60--80% of the expressed sequence map, the remainder expected to be derived through more long-term, labor-intensive, regional chromosomal gene searches or sequencing. The laboratory has made significant progress in the set-up phase, in mapping fetal and adult brain and other cDNAs, in testing a model system for directly linking genetic and physical maps using FISH with small fragments, in setting up a database, and in establishing the validity and throughput of the system.

  15. Putting the colours into chromogenic in situ hybridization (CISH).

    Science.gov (United States)

    Shipley, J

    2006-09-01

    Recurrent genomic alterations are the hallmarks of particular cancers. Application of molecular cytogenetic technologies to tumour material in order to detect these alterations has become important for molecular diagnostics and research. A dual-colour chromogenic in situ hybridization (dc-CISH) method described recently in the Journal of Pathology has the advantage of visualizing two probes simultaneously with the ability to discern morphological features. In addition, the bright field microscopy required is readily accessible to many laboratories. The approach has been validated by comparison of results with standard analyses for HER-2 amplification status in formalin-fixed, paraffin-embedded breast cancers and is applicable to the analysis of other clinically relevant genomic aberrations as well as of use in research investigations. Copyright (c) 2006 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

  16. In-situ fabrication of hybrid polyoxometalate nanoparticles composite films

    International Nuclear Information System (INIS)

    Lan Yang; Mao Baodong; Wang Enbo; Song Yonghai; Kang Zhenhui; Wang Chunlei; Tian Chungui; Zhang Chao; Xu Lin; Li Zhuang

    2007-01-01

    Inorganic-organic hybrid nanoparticles multilayer films were fabricated by extending the method of nucleation and growth of particles in polymer assemblies. The polyelectrolyte matrix was constructed by layer-by-layer self-assembly method. Synthesis of polyoxometalate nanoparticles was achieved by alternately dipping the precursor polyelectrolyte matrix into AgNO 3 and H 4 SiW 12 O 40 aqueous solutions. Repeating the above synthesis process, Ag 4 SiW 12 O 40 nanoparticles with controllable diameters of 20 to 77 nm were synthesized in the multilayer films in-situ. UV-vis absorption spectra indicate that the nanoparticles grew gradually in the synthesis process. Transmission electron microscopy was used to observe the size and morphology of the nanoparticles

  17. Heavy ion-induced chromosomal aberrations analyzed by fluorescence in situ hybridization

    International Nuclear Information System (INIS)

    Durante, M.; Gialanella, G.; Grossi, G.; Pugliese, M.; Cella, L.; Greco, O.; George, K.; Yang, T.C.

    1997-01-01

    We have investigated the effectiveness of heavy ions in the induction of chromosomal aberrations in mammalian cells by the recent technique of fluorescence in situ hybridization (FISH) with whole-chromosome probes. FISH-painting was used both in metaphase and interphase (prematurely condensed) chromosomes. The purpose of our experiments was to address the following problems: (a) the ratio of different types of aberrations as a function of radiation quality (search for biomarkers); (b) the ratio between aberrations scored in interphase and metaphase as a function of radiation quality (role of apoptosis); (c) differences between cytogenetic effects produced by different ions at the same LET (role of track structure). (orig./MG)

  18. Heavy ion-induced chromosomal aberrations analyzed by fluorescence in situ hybridization

    Energy Technology Data Exchange (ETDEWEB)

    Durante, M; Gialanella, G; Grossi, G; Pugliese, M [Univ. ` ` Federico II` ` , Naples (Italy). Dept. of Physics; [INFN, Naples (Italy); Cella, L; Greco, O [Univ. ` ` Federico II` ` , Naples (Italy). Dept. of Physics; Furusawa, Y [NIRS, Chiba (Japan); George, K; Yang, T C [NASA Lyndon B. Johnson Space Center, Houston, TX (United States)

    1997-09-01

    We have investigated the effectiveness of heavy ions in the induction of chromosomal aberrations in mammalian cells by the recent technique of fluorescence in situ hybridization (FISH) with whole-chromosome probes. FISH-painting was used both in metaphase and interphase (prematurely condensed) chromosomes. The purpose of our experiments was to address the following problems: (a) the ratio of different types of aberrations as a function of radiation quality (search for biomarkers); (b) the ratio between aberrations scored in interphase and metaphase as a function of radiation quality (role of apoptosis); (c) differences between cytogenetic effects produced by different ions at the same LET (role of track structure). (orig./MG)

  19. Mars Oxygen In-Situ Resource Utilization Experiment

    Data.gov (United States)

    National Aeronautics and Space Administration — The Mars Oxygen In-Situ Resource Utilization Experiment (MOXIE) will be the first in-situ resource utilization (ISRU) technology demonstration on Mars. Competitively...

  20. Exploring the origin of the D genome of oat by fluorescence in situ hybridization.

    Science.gov (United States)

    Luo, Xiaomei; Zhang, Haiqin; Kang, Houyang; Fan, Xing; Wang, Yi; Sha, Lina; Zhou, Yonghong

    2014-09-01

    Further understanding of the origin of cultivated oat would accelerate its genetic improvement. In particular, it would be useful to clarify which diploid progenitor contributed the D genome of this allohexaploid species. In this study, we demonstrate that the landmarks produced by fluorescence in situ hybridization (FISH) of species of Avena using probes derived from Avena sativa can be used to explore the origin of the D genome. Selected sets of probes were hybridized in several sequential experiments performed on exactly the same chromosome spreads, with multiple probes of cytological preparations. Probes pITS and A3-19 showed there might be a similar distribution of pITS between the Ac and D genomes. These results indicated that the Ac genome is closely related to the D genome, and that Avena canariensis (AcAc) could be the D-genome donor of cultivated oat.

  1. Clinical application of fluorescence in situ hybridization for prenatal diagnosis

    Directory of Open Access Journals (Sweden)

    Shu-fang JIANG

    2012-07-01

    Full Text Available Objective To establish and optimize the procedures of fluorescence in situ hybridization(FISH), and evaluate its clinical value in rapid prenatal diagnosis of fetal numerical abnormality of chromosomes 21, 18, 13, X, Y. Methods Amniotic fluid or fetal blood was sampled by routine invasive procedures. After the amniotic fluid cells or fetal blood cells were separated and sequentially processed with hypotonic solution, fixation solution, smear and high temperature, they were hybridized in situ with two panels of specific fluorescence probes to detect numerical abnormality of chromosomes 21, 18, 13, X, Y. All the samples were also cultured and analyzed for their karyotype by conventional methods. Results When it was used as a diagnostic criterion of chromosomal number that the fluorescence signals were observed in ≥90% cells, GLP 13/GLP 21 probe panel showed 2 green/2 red fluorescence signals and CSP18/CSP X/CSP Y probe panel showed 2 blue/2 yellow (female or 2 blue/1 yellow/1 red fluorescence signals (male under normal condition. The test reports of all 196 cases were sent out in 72-96 hours, and 7 cases of Down syndrome, 2 cases of trisomy 18 and 1 case of sex chromosomal numerical abnormality were detected, which were accordant with karyotype analysis results reported one month later. Conclusions FISH has potential for clinical application, and is applicable to rapid prenatal diagnosis of fetal numerical abnormality of chromosomes 21, 18, 13, X, Y. The rapid FISH, together with conventional karyotyping, offer a valuable means for prenatal diagnosis of fetal aneuploidies.

  2. Presence and localization of bacteria in the bovine endometrium postpartum using fluorescence in situ hybridization

    DEFF Research Database (Denmark)

    Karstrup, C. C.; Agerholm, J. S.; Jensen, Tim Kåre

    2017-01-01

    The aim of this study was to investigate bacterial invasiveness of the bovine endometrium during the postpartum period. Fluorescence in situ hybridization was applied to endometrial biopsies using probes for Fusobacterium necrophorum, Porphyromonas levii, Trueperella pyogenes, Escherichia coli...

  3. Study of the frequency of translocations and dicentrics in human spermatozoid using fluorescent in situ hybridization

    International Nuclear Information System (INIS)

    Alvarez, R.; Ponsa, I.; Tusell, L.; Genesca, A.; Miro, R.; Egozcue, J.

    1998-01-01

    Present study has intended to analyze the induction translocations and dicentrics in human sperms irradiated in vitro to the dose 4Gy by means of the use technical in situ hybridization with probes marked fluorescently

  4. Detection of airborne Legionella while showering using liquid impingement and fluorescent in situ hybridization (FISH).

    Science.gov (United States)

    Deloge-Abarkan, Magali; Ha, Thi-Lan; Robine, Enric; Zmirou-Navier, Denis; Mathieu, Laurence

    2007-01-01

    Aerosols of water contaminated with Legionella bacteria constitute the only mode of exposure for humans. However, the prevention strategy against this pathogenic bacteria risk is managed through the survey of water contamination. No relationship linked the Legionella bacteria water concentration and their airborne abundance. Therefore, new approaches in the field of the metrological aspects of Legionella bioaerosols are required. This study was aimed at testing the main principles for bioaerosol collection (solid impaction, liquid impingement and filtration) and the in situ hybridization (FISH) method, both in laboratory and field assays, with the intention of applying such methodologies for airborne Legionella bacteria detection while showering. An aerosolization chamber was developed to generate controlled and reproducible L. pneumophila aerosols. This tool allowed the identification of the liquid impingement method as the most appropriate one for collecting airborne Legionella bacteria. The culturable fraction of airborne L. pneumophila recovered with the liquid impingement principle was 4 and 700 times higher compared to the impaction and filtration techniques, respectively. Moreover, the concentrations of airborne L. pneumophila in the impinger fluid were on average 7.0 x 10(5) FISH-cells m(-3) air with the fluorescent in situ hybridization (FISH) method versus 9.0 x 10(4) CFU m(-3) air with the culture method. These results, recorded under well-controlled conditions, were confirmed during the field experiments performed on aerosols generated by hot water showers in health institutions. This new approach may provide a more accurate characterization of aerobiocontamination by Legionella bacteria.

  5. Comparative genomic and in situ hybridization of germ cell tumors of the infantile testis

    NARCIS (Netherlands)

    Mostert, M; Rosenberg, C; Stoop, H; Schuyer, M; Timmer, A; Oosterhuis, W; Looijenga, L

    Chromosomal information on germ cell tumors of the infantile testis, ie, teratomas and yolk sac tumors, is limited and controversial. We studied two teratomas and four yolk sac tumors using comparative genomic hybridization (CGH) and in situ hybridization. No chromosomal anomalies were found in the

  6. Analysis of experimental mink enteritis virus infection in mink: in situ hybridization, serology, and histopathology

    DEFF Research Database (Denmark)

    Uttenthal, Åse; Larsen, S; Lund, E

    1990-01-01

    Strand-specific hybridization probes were used in in situ hybridization studies to localize cells containing mink enteritis virus (MEV) virion DNA or MEV replicative-form DNA and mRNA. Following the experimental MEV infection of 3-month-old unvaccinated mink, a significant increase in serum antib...

  7. A simple protocol for attenuating the auto-fluorescence of cyanobacteria for optimized fluorescence in situ hybridization (FISH) imaging.

    Science.gov (United States)

    Zeller, Perrine; Ploux, Olivier; Méjean, Annick

    2016-03-01

    Cyanobacteria contain pigments, which generate auto-fluorescence that interferes with fluorescence in situ hybridization (FISH) imaging of cyanobacteria. We describe simple chemical treatments using CuSO4 or H2O2 that significantly reduce the auto-fluorescence of Microcystis strains. These protocols were successfully applied in FISH experiments using 16S rRNA specific probes and filamentous cyanobacteria. Copyright © 2016 Elsevier B.V. All rights reserved.

  8. Identification of bacteria used for microbial enhanced oil recovery process by fluorescence in situ hybridization technique

    Energy Technology Data Exchange (ETDEWEB)

    Fujiwara, K.; Tanaka, S.; Otsuka, M. [Kansai Research Institute, Kyoto (Japan). Lifescience Lab.; Yonebayashi, H. [Japan National Oil Corp., Chiba (Japan). Tech. Research Center; Enomoto, H. [Tohoku University, Sendai (Japan). Dept. of Geoscience and Tech.

    2000-01-01

    A fluorescence in situ hybridization (FISH) technique using 16S rRNA-targeted oligonucleotide probes was developed for rapid detection of microorganisms for use in the microbial enhancement of oil recovery (MEOR) process. Two microorganisms, Enterobacter cloacae TRC-322 and Bacillus licheniformis TRC-18-2-a, were selected from a collection of Enterobacter sp. and Bacillus sp. which were screened in previous studies as candidate microorganisms for injection, and were used for this experiment. Oligonucleotide probes, design based on specific sequences in the 16S rRNA gene were labeled with either fluorescein isothiocyanate (FITC), or 6-car-boxy-X-rhodamine (ROX), and were allowed to hybridize with fixed cells of the two microorganisms noted above. The fluorescence signal emitted from each microorganism cells could clearly be detected by an epifluorescence microscope. Moreover, E. cloacae TRC-322 and B, licheniformis TRC-18-2-a, suspended in actual reservoir brine, including inorganic salts, oil and aboriginal cells of the reservoir brine, could be detected directly by this hybridization method, without the need for cultivation and isolation. (author)

  9. Characterization of a de novo duplication of 11p14----p13, using fluorescent in situ hybridization and southern hybridization

    NARCIS (Netherlands)

    Speleman, F.; Mannens, M.; Redeker, B.; Vercruyssen, M.; van Oostveldt, P.; Leroy, J.; Slater, R.

    1991-01-01

    A de novo 11p+ chromosome was found in a child with mild mental retardation but no other remarkable dysmorphic characteristics. Banding studies suggested a duplication of regions 11p13 and 11p14 or regions 11p14 and 11p15. Using fluorescent in situ hybridization and digital imaging microscopy, we

  10. Fluorescence in situ hybridization on formalin-fixed and paraffin-embedded tissue

    DEFF Research Database (Denmark)

    Laub Petersen, Bodil; Zeuthen, Mette Christa; Pedersen, Sanni

    2004-01-01

    Fluorescence in situ hybridization (FISH) is widely used to study numerical and structural genetic abnormalities in both metaphase and interphase cells. The technique is based on the hybridization of labeled probes to complementary sequences in the DNA or RNA of the cells. Interphase FISH is most...... in time lapse between removal of tissue and fixation, duration of fixation, enzymatic pretreatment, hybridization conditions, and posthybridization washing conditions are important factors in the hybridization. In this study, we have listed the results of a systematic approach to improve FISH on isolated...

  11. Hybrid Wars: Israel’s experience

    Directory of Open Access Journals (Sweden)

    E. N. Grachikov

    2015-01-01

    Full Text Available The article attempts to summarize the experience of Israel in the fight against non-traditional threats, which escalated into a hybrid war. Feature of these wars lies in the fact that they are usually conducted between Western and non-Western countries in the border and poorly controlled space. West opponents favor alliances and associations of non-Western countries and non-state actors. In this case, to justify the war, the norms of international law are used.

  12. Cytogenetic, genomic in situ hybridization (GISH) and agronomic ...

    African Journals Online (AJOL)

    F3 generations of a wheat-Psathyrostachys huashanica intergeneric cross. Their agronomic traits were evaluated in the field and their meiotic behaviors and chromosome composition were analyzed by cytogenetic and GISH (genomic in situ ...

  13. Karyotypes and Distribution of Tandem Repeat Sequences in Brassica nigra Determined by Fluorescence in situ Hybridization

    Czech Academy of Sciences Publication Activity Database

    Wang, G.; He, Q.; Macas, Jiří; Novák, Petr; Neumann, Pavel; Meng, D.; Zhao, H.; Guo, N.; Han, S.; Zong, M.; Jin, W.; Liu, F.

    2017-01-01

    Roč. 152, č. 3 (2017), s. 158-165 ISSN 1424-8581 R&D Projects: GA ČR GBP501/12/G090 Institutional support: RVO:60077344 Keywords : asymmetric somatic hybridization * Fluorescence in situ hybridization * Karyotype * (Peri) centromere Subject RIV: EB - Genetics ; Molecular Biology OBOR OECD: Genetics and heredity (medical genetics to be 3) Impact factor: 1.354, year: 2016

  14. Comparison of Chromogenic In Situ Hybridization and Fluorescence In Situ Hybridization for the Evaluation of MDM2 Amplification in Adipocytic Tumors.

    Science.gov (United States)

    Mardekian, Stacey K; Solomides, Charalambos C; Gong, Jerald Z; Peiper, Stephen C; Wang, Zi-Xuan; Bajaj, Renu

    2015-11-01

    Atypical lipomatous tumor/well-differentiated liposarcoma (ALT-WDLPS) and dedifferentiated liposarcoma (DDLPS) are characterized cytogenetically by a 12q13-15 amplification involving the mouse double minute 2 (MDM2) oncogene. Fluorescence in situ hybridization (FISH) is used frequently to detect this amplification and aid with the diagnosis of these entities, which is difficult by morphology alone. Recently, bright-field in situ hybridization techniques such as chromogenic in situ hybridization (CISH) have been introduced for the determination of MDM2 amplification status. The present study compared the results of FISH and CISH for detecting MDM2 amplification in 41 cases of adipocytic tumors. Amplification was defined in both techniques as a MDM2/CEN12 ratio of 2 or greater. Eleven cases showed amplification with both FISH and CISH, and 26 cases showed no amplification with both methods. Two cases had discordant results between CISH and FISH, and two cases were not interpretable by CISH. CISH is advantageous for allowing pathologists to evaluate the histologic and molecular alterations occurring simultaneously in a specimen. Moreover, CISH is found to be more cost- and time-efficient when used with automation, and the signals do not quench over time. CISH technique is a reliable alternative to FISH in the evaluation of adipocytic tumors for MDM2 amplification. © 2014 Wiley Periodicals, Inc.

  15. Tracking alien chromosome in sativa background by genomic in situ hybridization

    International Nuclear Information System (INIS)

    Abbasi, F.M.; Iqbal, M.; Salim, M.

    2004-01-01

    Genomic in situ hybridization (GISH) was used to look into the genomic constitution of monosomic alien -addition line derived from O. sativa x O. brachyantha. Biotin label genomic DNA from O. brachyantha was used as probe. The probe hybridized to the brachyantha chromosome. No detectable hybridization signal was observed on sativa chromosomes. This differential painting of chromosome enables us to unequivocally discriminate brachyantha chromosome from those of sativa. Results showed the usefulness of GISH in the identification of a single alien chromosome in the sativa background. (author)

  16. Imaging of RNA in situ hybridization by atomic force microscopy

    NARCIS (Netherlands)

    Kalle, W.H.J.; Macville, M.V.E.; van de Corput, M.P.C.; de Grooth, B.G.; Tanke, H.J.; Raap, A.K.

    In this study we investigated the possibility of imaging internal cellular molecules after cytochemical detection with atomic force microscopy (AFM). To this end, rat 9G and HeLa cells were hybridized with haptenized probes for 28S ribosomal RNA, human elongation factor mRNA and cytomegalovirus

  17. Application of Pseudomurein Endoisopeptidase to Fluorescence In Situ Hybridization of Methanogens within the Family Methanobacteriaceae▿

    Science.gov (United States)

    Nakamura, Kohei ; Terada, Takeshi; Sekiguchi, Yuji; Shinzato, Naoya; Meng, Xian-Ying; Enoki, Miho; Kamagata, Yoichi

    2006-01-01

    In situ detection of methanogens within the family Methanobacteriaceae is sometimes known to be unsuccessful due to the difficulty in permeability of oligonucleotide probes. Pseudomurein endoisopeptidase (Pei), a lytic enzyme that specifically acts on their cell walls, was applied prior to 16S rRNA-targeting fluorescence in situ hybridization (FISH). For this purpose, pure cultured methanogens within this family, Methanobacterium bryantii, Methanobrevibacter ruminantium, Methanosphaera stadtmanae, and Methanothermobacter thermautotrophicus together with a Methanothermobacter thermautotrophicus-containing syntrophic acetate-oxidizing coculture, endosymbiotic Methanobrevibacter methanogens within an anaerobic ciliate, and an upflow anaerobic sludge blanket (UASB) granule were examined. Even without the Pei treatment, Methanobacterium bryantii and Methanothermobacter thermautotrophicus cells are relatively well hybridized with oligonucleotide probes. However, almost none of the cells of Methanobrevibacter ruminantium, Methanosphaera stadtmanae, cocultured Methanothermobacter thermautotrophicus, and the endosymbiotic methanogens and the cells within UASB granule were hybridized. Pei treatment was able to increase the probe hybridization ratio in every specimen, particularly in the specimen that had shown little hybridization. Interestingly, the hybridizing signal intensity of Methanothermobacter thermautotrophicus cells in coculture with an acetate-oxidizing H2-producing syntroph was significantly improved by Pei pretreatment, whereas the probe was well hybridized with the cells of pure culture of the same strain. We found that the difference is attributed to the differences in cell wall thicknesses between the two culture conditions. These results indicate that Pei treatment is effective for FISH analysis of methanogens that show impermeability to the probe. PMID:16950902

  18. Fluorescence in situ hybridization on human metaphase chromosomes detected by near-field scanning optical microscopy

    NARCIS (Netherlands)

    Moers, M.H.P.; Moers, M.H.P.; Kalle, W.H.J.; Kalle, W.H.J.; Ruiter, A.G.T.; Wiegant, J.C.A.G.; Raap, A.K.; Greve, Jan; de Grooth, B.G.; van Hulst, N.F.

    1996-01-01

    Fluorescence in situ hybridization o­n human metaphase chromosomes is detected by near-field scanning optical microscopy. This combination of cytochemical and scanning probe techniques enables the localization and identification of several fluorescently labelled genomic DNA fragments o­n a single

  19. Applications of ribosomal in situ hybridization for the study of bacterial cells in the mouse intestine

    DEFF Research Database (Denmark)

    Licht, Tine Rask; Poulsen, Lars Kongsbak; Molin, Søren

    1997-01-01

    Localization of E. coli and S. typhimurium in the large and small intestine of streptomycin-treated mice was visualized by in situ hybridization with specific rRNA target probes and epi-fluorescence microscopy. Growth rates of E. coli BJ4 colonizing the large intestine of streptomycin-treated mic...

  20. Differential in situ hybridization for herpes simplex virus typing in routine skin biopsies

    NARCIS (Netherlands)

    Botma, H. J.; Dekker, H.; van Amstel, P.; Cairo, I.; van den Berg, F. M.

    1995-01-01

    A herpes simplex virus (HSV) type 2 specific recombinant plasmid probe designated pH2S3 was constructed from non-HSV-1 crossreactive regions of the HSV-2 genome. DNA in situ hybridization on in vitro reconstructed tissue samples of sheep collagen matrix impregnated with herpes virus-infected human

  1. Cytogenetic analysis using fluorescence in situ hybridization (FISH) to evaluate occupational exposure to carcinogens

    Czech Academy of Sciences Publication Activity Database

    Šrám, Radim; Beskid, Olena; Binková, Blanka; Rössner, P.; Šmerhovský, Zdeněk

    2004-01-01

    Roč. 149, - (2004), s. 335-344 ISSN 0378-4274 R&D Projects: GA MŽP SI/340/2/00 Institutional research plan: CEZ:AV0Z5039906 Keywords : Chromosomal aberrations * Fluorescence in situ hybridization Subject RIV: DN - Health Impact of the Environment Quality Impact factor: 2.571, year: 2004

  2. Fluorescence in situ hybridization for phytoplasma and endophytic bacteria localization in plant tissues.

    Science.gov (United States)

    Bulgari, Daniela; Casati, Paola; Faoro, Franco

    2011-11-01

    In the present study, we developed a rapid and efficient fluorescence in situ hybridization assay (FISH) in non-embedded tissues of the model plant Catharanthus roseus for co-localizing phytoplasmas and endophytic bacteria, opening new perspectives for studying the interaction between these microorganisms. Copyright © 2011 Elsevier B.V. All rights reserved.

  3. Biomarkers for ALK and ROS1 in Lung Cancer: Immunohistochemistry and Fluorescent In Situ Hybridization.

    Science.gov (United States)

    Luk, Peter P; Selinger, Christina I; Mahar, Annabelle; Cooper, Wendy A

    2018-06-14

    - A small proportion of non-small cell lung cancers harbor rearrangements of ALK or ROS1 genes, and these tumors are sensitive to targeted tyrosine kinase inhibitors. It is crucial for pathologists to accurately identify tumors with these genetic alterations to enable patients to access optimal treatments and avoid unnecessary side effects of less effective agents. Although a number of different techniques can be used to identify ALK- and ROS1-rearranged lung cancers, immunohistochemistry and fluorescence in situ hybridization are the mainstays. - To review the role of immunohistochemistry in assessment of ALK and ROS1 rearrangements in lung cancer, focusing on practical issues in comparison with other modalities such as fluorescence in situ hybridization. - This manuscript reviews the current literature on ALK and ROS1 detection using immunohistochemistry and fluorescence in situ hybridization as well as current recommendations. - Although fluorescence in situ hybridization remains the gold standard for detecting ALK and ROS1 rearrangement in non-small cell lung cancer, immunohistochemistry plays an important role and can be an effective screening method for detection of these genetic alterations, or a diagnostic test in the setting of ALK.

  4. Implementation of the Fluorescent in Situ Hybridization technique in the Faculty of Medicine, UdelaR

    Directory of Open Access Journals (Sweden)

    Andrea Cairus

    2017-11-01

    Full Text Available The Cytogenetic Laboratory of the Faculty of Medicine processes, on average, 300 annual samples of public and private healthcare centers by conventional cytogenetics. It is essential to implement new techniques to improve the quality of the service offered. The purpose of this work was to implement the Fluorescent in situ Hybridization technique (FISH. An observational, cross-sectional, analytical study was performed. Peripheral blood samples from patients with sex chromosomopathies diagnosed by conventional cytogenetics were analyzed. Fluorescent in situ hybridization technique was applied, comparing results with FISH and with conventional cytogenetics. The percentage of mosaicism detected by conventional cytogenetics and Fluorescent in situ Hybridization was studied: 24 samples were analyzed; 19 presented numerical alterations, 3 structural and 2 both. Numerical alterations were Turner syndrome, Klinefelter syndrome, XXX syndrome and XYY syndrome. Concordance in diagnoses was found for both techniques. For Turner syndrome, 8 of 12 samples corresponded to mosaicism, and there were no significant differences between conventional cytogenetics and the technique studied (p0.05. Klinefelter syndrome and XYY were both presented in a non-mosaic karyotype. For XXX syndrome, a normal line (46, XX was observed in three of the samples, in a percentage close to the cut off. From this research, it will be possible to implement Fluorescent in situ Hybridization in this service, to extend it to other pathologies and to enable the training of human resources; consolidating this laboratory as a national academic reference center.

  5. Analysis of hepcidin expression: in situ hybridization and quantitative polymerase chain reaction from paraffin sections.

    Science.gov (United States)

    Sakuraoka, Yuhki; Sawada, Tokihiko; Shiraki, Takayuki; Park, Kyunghwa; Sakurai, Yuhichiro; Tomosugi, Naohisa; Kubota, Keiichi

    2012-07-28

    To establish methods for quantitative polymerase chain reaction (PCR) for hepcidin using RNAs isolated from paraffin-embedded sections and in situ hybridization of hepatocellular carcinoma (HCC). Total RNA from paraffin-embedded sections was isolated from 68 paraffin-embedded samples of HCC. Samples came from 54 male and 14 female patients with a mean age of 66.8 ± 7.8 years. Quantitative PCR was performed. Immunohistochemistry and in situ hybridization for hepcidin were also performed. Quantitative PCR for hepcidin using RNAs isolated from paraffin-embedded sections of HCC was performed successfully. The expression level of hepcidin mRNA in cancer tissues was significantly higher than that in non-cancer tissues. A method of in situ hybridization for hepcidin was established successfully, and this demonstrated that hepcidin mRNA was expressed in non-cancerous tissue but absent in cancerous tissue. We have established novel methods for quantitative PCR for hepcidin using RNAs isolated from paraffin-embedded sections and in situ hybridization of HCC.

  6. In Situ Production of Graphene-Fiber Hybrid Structures

    DEFF Research Database (Denmark)

    Akia, Mandana; Cremar, Lee; Chipara, Mircea

    2017-01-01

    We report a scalable method to obtain a new material where large graphene sheets form webs linking carbon fibers. Film-fiber hybrid nonwoven mats are formed during fiber processing and converted to carbon structures after a simple thermal treatment. This contrasts with multistep methods...... that attempt to mix previously prepared graphene and fibers, or require complicated and costly processes for deposition of graphene over carbon fibers. The developed graphene-fiber hybrid structures have seamless connections between graphene and fibers, and in fact the graphene "veils" extend directly from one...... a capillarity effect that promoted the formation of thin veils, which become graphene sheets upon dehydration by sulfuric acid vapor followed by carbonization (at relatively low temperatures, below 800 °C). These veils extend over several micrometers within the pores of the fiber network, and consist...

  7. Chromosome identification by new molecular markers and genomic in situ hybridization in the Triticum-Secale-Thinopyrum trigeneric hybrids.

    Science.gov (United States)

    Dai, Yi; Duan, Yamei; Chi, Dawn; Liu, Huiping; Huang, Shuai; Cao, Wenguang; Gao, Yong; Fedak, George; Chen, Jianmin

    2017-08-01

    It is very important to use chromosome-specific markers for identifying alien chromosomes in advanced generations of distant hybridization. The chromosome-specific markers of rye and Thinopyrum elongatum, as well as genomic in situ hybridization, were used to identify the alien chromosomes in eight lines that were derived from the crossing between Triticum trititrigia (AABBEE) and triticale (AABBRR). The results showed that four lines contained all rye chromosomes but no Th. elongatum chromosomes. The line RE36-1 contained all of the rye chromosomes except for chromosome 2R. The lines RE33-2 and RE62-1 contained all rye chromosomes and 1E and 5E translocated chromosome, respectively. The line RE24-4 contained 12 rye chromosomes plus a 7E chromosome or 12 rye chromosomes plus one R-E translocated chromosome. Chromosome identification in the above lines was consistent using chromosome-specific markers and genomic in situ hybridization. These chromosome-specific markers provide useful tools for detecting alien chromosomes in trigeneric hybrids, and these lines could be utilized as valuable germplasm in wheat improvement.

  8. Fluorescent in situ hybridization of mitochondrial DNA and RNA

    Czech Academy of Sciences Publication Activity Database

    Alán, Lukáš; Zelenka, Jaroslav; Ježek, Jan; Dlasková, Andrea; Ježek, Petr

    2010-01-01

    Roč. 57, č. 4 (2010), s. 403-408 ISSN 0001-527X R&D Projects: GA ČR GAP302/10/0346; GA ČR GPP304/10/P204; GA AV ČR KJB500110902 Institutional research plan: CEZ:AV0Z50110509 Keywords : mitochondrial DNA and RNA * nucleoids of mitochondrial DNA * molecular beacon fluorescent hybridization probes Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 1.234, year: 2010

  9. [Chromosomal localization of foreign genes in transgenic mice using dual-color fluorescence in situ hybridization].

    Science.gov (United States)

    Lin, Dan; Gong, Xiu-li; Li, Wei; Guo, Xin-bing; Zhu, Yi-wen; Huang, Ying

    2008-02-01

    To establish a highly sensitive and specific dual-color fluorescence in situ hybridization (D-FISH) method used for chromosomal localization of foreign genes in double transgenic mice. Two strains of double transgenic mice were used in this experiment, one was integrated with the herpes simplex virus thymidine kinase (HSV-tk) and the enhanced green fluorescence protein (eGFP), the other was with the short hairpin RNA interference(RNAi) and beta(654). Splenic cells cultured in vitro were arrested in metaphase by colchicine and hybridized with digoxigenin-labeled and biotinylated DNA probes, then detected by rhodamine-conjugated avidin and FITC-conjugated anti-digoxigenin. Dual-color fluorescence signals were detected on the same metaphase in both transgenic mice strains. In HSV-tk/eGFP double transgenic mice, strong green fluorescence for HSV-tk and red for eGFP were observed and localized at 2E5-G3 and 8A2-A4 respectively. In beta(654)/RNAi mice, beta(654) was detected as red fluorescence on chromosome 7D3-E2, and RNAi showed random integration on chromosomes. It was detected as green fluorescence on chromosome 12B1 in one mouse, while on 1E2.3-1F and 3A3 in the other. Highly sensitive and specific D-FISH method was established using the self-prepared DNA probes, and chromosomal localization of the foreign genes was also performed in combination with G-banding in double transgenic mice. This technology will facilitate the researches in transgenic animals and gene therapy models.

  10. Image files of planarians analyzed by in situ hybridication and immunohistochemical staining - Plabrain DB | LSDB Archive [Life Science Database Archive metadata

    Lifescience Database Archive (English)

    Full Text Available List Contact us Plabrain DB Image files of planarians analyzed by in situ hybridication and immunohistochemical... staining Data detail Data name Image files of planarians analyzed by in situ hybridication and immunohistochemical...sion patterns by whole-mount in situ hybridication and also protein distribution by immunohistochemical...Images are displayed in A list of image files of planarians analyzed by in situ hybridication and immunohistochemical...le search URL - Data acquisition method Whole-mount in situ hybridication, immunohistochemical staining Data

  11. Lower hybrid experiments in the PETULA Tokamak

    International Nuclear Information System (INIS)

    Melin, G.; Bardet, R.; Bernard, M.

    1979-01-01

    The present paper deals with the application of high RF power near the lower hybrid (LH) frequency to a Tokamak discharge. A double waveguide grill antenna has been used to launch a 1.25 GHz-500kW-6 ms RF pulse in PETULA. One of the objectives of this study is to investigate the technological problems associated with LH heating and caused by the high level of RF power involved. Two aspects have been carefully studied: grill conditioning and grill coupling. The latter is strongly dependent on the scrape-off layer of the PETULA plasma and the observations are possibly not significant. The former aspect though not really understood is probably to be expected in other LH experiments. Plasma experiments have been carried out with a wide range of electron density 2. 13 cm -3 in both hydrogen and deuterium. For hydrogen this allows theoretically linear mode conversion to take place in the discharge. Although some heating has been observed, the experiments are not, as yet, very conclusive. The actual RF pulse duration is hardly larger than the ion energy confinement time, and the radial plasma displacement caused by the RF is not feedback controlled. The main discharge parameters were the following: plasma radius 14 cm, toroidal magnetic field 27 kG, plasma current 75 kA, loop voltage 1.6 V, Zsub(eff) < 2., 800 < Tsub(e)(o)< 1100eV, 200 < Tsub(i)(o)< 350 eV

  12. Dramatically improved RNA in situ hybridization signals using LNA-modified probes

    DEFF Research Database (Denmark)

    Thomsen, Rune; Nielsen, Peter Stein; Jensen, Torben Heick

    2005-01-01

    . This increases the thermal stability of hybrids formed with RNA. The LNA-based probes detect specific RNAs in fixed yeast cells with an efficiency far better than conventional DNA oligonucleotide probes of the same sequence. Using this probe design, we were also able to detect poly(A)+ RNA accumulation within......In situ detection of RNA by hybridization with complementary probes is a powerful technique. Probe design is a critical parameter in successful target detection. We have evaluated the efficiency of fluorescent DNA oligonucleotides modified to contain locked nucleic acid (LNA) residues...

  13. Chromogenic in situ hybridization (CISH) to detect HER2 gene amplification in breast and gastric cancer: comparison with immunohistochemistry (IHC) and fluorescence in situ hybridization (FISH).

    Science.gov (United States)

    Kiyose, Shinichiro; Igarashi, Hisaki; Nagura, Kiyoko; Kamo, Takaharu; Kawane, Kazunori; Mori, Hiroki; Ozawa, Takachika; Maeda, Matsuyoshi; Konno, Keisuke; Hoshino, Hideaki; Konno, Hiroyuki; Ogura, Hiroyuki; Shinmura, Kazuya; Hattori, Naohiko; Sugimura, Haruhiko

    2012-11-01

    The chromogenic in situ hybridization (CISH) assay, designed to detect the amplification of the HER2 gene in formalin-fixed, paraffin-embedded (FFPE) breast cancer (BC) and gastric cancer (GC) tissue specimens, was evaluated in 125 FFPE BC cases and 198 FFPE GC cases for which the HER2 status had been predetermined using immunohistochemistry (IHC) and fluorescence in situ hybridization (FISH). In the 125 BC cases and the 198 gastric cases, we found a very good concordance (98.4% and 99.0%, respectively) between CISH and FISH. In particular, we evaluated the polysomy cases, as these cases often have ambiguous treatment options in clinical practice. The polysomy of chromosome 17 was defined as the presence of three or more CEP17 signals in at least 10% of the tumor cells. In the 50 BC cases and 54 GC cases displaying chromosome 17 polysomy, the concordance between FISH and CISH was 98.0% and 98.1%, respectively. These results indicate that CISH could provide an accurate and practical alternative to FISH for the clinical diagnosis of HER2 gene amplification in FFPE BC and FFPE GC samples. © 2012 The Authors. Pathology International © 2012 Japanese Society of Pathology and Wiley Publishing Asia Pty Ltd.

  14. Detection of denitrification genes by in situ rolling circle amplification - fluorescence in situ hybridization (in situ RCA-FISH) to link metabolic potential with identity inside bacterial cells

    DEFF Research Database (Denmark)

    Hoshino, Tatsuhiko; Schramm, Andreas

    2010-01-01

    target site. Finally, the RCA product inside the cells was detected by standard fluorescence in situ hybridization (FISH). The optimized protocol showed high specificity and signal-to-noise ratio but low detection frequency (up to 15% for single-copy genes and up to 43% for the multi-copy 16S rRNA gene...... as Candidatus Accumulibacter phosphatis by combining in situ RCA-FISH with 16S rRNA-targeted FISH. While not suitable for quantification because of its low detection frequency, in situ RCA-FISH will allow to link metabolic potential with 16S rRNA (gene)-based identification of single microbial cells.......). Nevertheless, multiple genes (nirS and nosZ; nirS and the 16S rRNA gene) could be detected simultaneously in P. stutzeri. Environmental application of in situ RCA-FISH was demonstrated on activated sludge by the differential detection of two types of nirS-defined denitrifiers; one of them was identified...

  15. In situ fatigue-crack-propagation experiment

    International Nuclear Information System (INIS)

    Ermi, A.M.; Chin, B.A.

    1981-01-01

    An in-reactor fatigue experiment was conducted in the Oak Ridge Research Reactor to determine the effects of dynamic irradiation on fatigue crack propagation. Eight 20% cold-worked 316 stainless steel specimens were precracked to various initial crack lengths, linked together to form a chain, and inserted into a specially designed in-reactor fatigue machine. Test conditions included a maximum temperature of 460 0 C, an environment of sodium, a frequency of 1 cycle/min, and a stress ratio of 0.10. Results indicated that (1) no effects of dynamic irradiation were observed for a fluence of 1.5 x 10 21 n/cm 2 (E > 0.1 MeV); and (2) crack growth rates in elevated temperature sodium were a factor of 3 to 4 lower than in room temperature air

  16. Chromosomal imbalances detected in primary bone tumors by comparative genomic hybridization and interphase fluorescence in situ hybridization

    Directory of Open Access Journals (Sweden)

    Marcelo Razera Baruffi

    2003-01-01

    Full Text Available We applied a combination of comparative genomic hybridization (CGH and fluorescence in situ hybridization (FISH, to characterize the genetic aberrations in three osteosarcomas (OS and one Ewing's sarcoma. CGH identified recurrent chromosomal losses at 10p14-pter and gains at 8q22.3-24.1 in OS. Interphase FISH allowed to confirm 8q gain in two cases. A high amplification level of 11q12-qter was detected in one OS. The Ewing's sarcoma showed gain at 1p32-36.1 as the sole chromosome alteration. These studies demonstrate the value of molecular cytogenetic methods in the characterization of recurrent genomic alterations in bone tumor tissue.

  17. Design of Hybrid Steam-In Situ Combustion Bitumen Recovery Processes

    International Nuclear Information System (INIS)

    Yang Xiaomeng; Gates, Ian D.

    2009-01-01

    Given enormous capital costs, operating expenses, flue gas emissions, water treatment and handling costs of thermal in situ bitumen recovery processes, improving the overall efficiency by lowering energy requirements, environmental impact, and costs of these production techniques is a priority. Steam-assisted gravity drainage (SAGD) is the most widely used in situ recovery technique in Athabasca reservoirs. Steam generation is done on surface and consequently, because of heat losses, the energy efficiency of SAGD can never be ideal with respect to the energy delivered to the sandface. An alternative to surface steam generation is in situ combustion (ISC) where heat is generated within the formation through injection of oxygen at a sufficiently high pressure to initiate combustion of bitumen. In this manner, the heat from the combustion reactions can be used directly to mobilize the bitumen. As an alternative, the heat can be used to generate steam within the formation which then is the agent to move heat in the reservoir. In this research, alternative hybrid techniques with simultaneous and sequential steam-oxygen injection processes are examined to maximize the thermal efficiency of the recovery process. These hybrid processes have the advantage that during ISC, steam is generated within the reservoir from injected and formation water and as a product of oxidation. This implies that ex situ steam generation requirements are reduced and if there is in situ storage of combustion gases, that overall gas emissions are reduced. In this research, detailed reservoir simulations are done to examine the dynamics of hybrid processes to enable design of these processes. The results reveal that hybrid processes can lower emitted carbon dioxide-to-oil ratio by about 46%, decrease the consumed natural gas-to-oil ratio by about 73%, reduce the cumulative energy-to-oil ratio by between 40% and 70% compared to conventional SAGD, and drop water consumption per unit oil produced

  18. Assignment of electron transfer flavoprotein-ubiquinone oxidoreductase (ETF-QO) to human chromosome 4q33 by fluorescence in situ hybridization and somatic cell hybridization.

    Science.gov (United States)

    Spector, E B; Seltzer, W K; Goodman, S I

    1999-08-01

    Electron transfer flavoprotein-ubiquinone oxidoreductase (ETF-QO) is a nuclear-encoded protein located in the inner mitochondrial membrane. Inherited defects of ETF-QO cause glutaric acidemia type II. We here describe the localization of the ETF-QO gene to human chromosome 4q33 by somatic cell hybridization and fluorescence in situ hybridization. Copyright 1999 Academic Press.

  19. Automated Image Analysis for Quantitative Fluorescence In Situ Hybridization with Environmental Samples▿ †

    OpenAIRE

    Zhou, Zhi; Pons, Marie Noëlle; Raskin, Lutgarde; Zilles, Julie L.

    2007-01-01

    When fluorescence in situ hybridization (FISH) analyses are performed with complex environmental samples, difficulties related to the presence of microbial cell aggregates and nonuniform background fluorescence are often encountered. The objective of this study was to develop a robust and automated quantitative FISH method for complex environmental samples, such as manure and soil. The method and duration of sample dispersion were optimized to reduce the interference of cell aggregates. An au...

  20. Submicroscopic deletions at the WAGR locus, revealed by nonradioactive in situ hybridization.

    OpenAIRE

    Fantes, J A; Bickmore, W A; Fletcher, J M; Ballesta, F; Hanson, I M; van Heyningen, V

    1992-01-01

    Fluorescence in situ hybridization (FISH) with biotin-labeled probes mapping to 11p13 has been used for the molecular analysis of deletions of the WAGR (Wilms tumor, aniridia, genitourinary abnormalities, and mental retardation) locus. We have detected a submicroscopic 11p13 deletion in a child with inherited aniridia who subsequently presented with Wilms tumor in a horseshoe kidney, only revealed at surgery. The mother, who has aniridia, was also found to carry a deletion including both the ...

  1. Hybrid Perovskite Thin-Film Photovoltaics: In Situ Diagnostics and Importance of the Precursor Solvate Phases

    KAUST Repository

    Munir, Rahim

    2016-11-07

    Solution-processed hybrid perovskite semiconductors attract a great deal of attention, but little is known about their formation process. The one-step spin-coating process of perovskites is investigated in situ, revealing that thin-film formation is mediated by solid-state precursor solvates and their nature. The stability of these intermediate phases directly impacts the quality and reproducibility of thermally converted perovskite films and their photovoltaic performance.

  2. Using Whole Mount in situ Hybridization to Link Molecular and Organismal Biology

    OpenAIRE

    Jacobs, Nicole L.; Albertson, R. Craig; Wiles, Jason R.

    2011-01-01

    Whole mount in situ hybridization (WISH) is a common technique in molecular biology laboratories used to study gene expression through the localization of specific mRNA transcripts within whole mount specimen. This technique (adapted from Albertson and Yelick, 2005) was used in an upper level undergraduate Comparative Vertebrate Biology laboratory classroom at Syracuse University. The first two thirds of the Comparative Vertebrate Biology lab course gave students the opportunity to study the ...

  3. Detection of Methylobacterium radiotolerans IMBG290 in potato plants by in situ hybridization

    OpenAIRE

    Pirttila A. M.; Kozyrovska N. O.; Ovcharenko L. P.; Podolich O. V.

    2009-01-01

    A new bacterial strain of pink-pigmented facultative methylotroph (M. radiotolerans IMBG290) which was previously isolated from in vitro grown potato plantlets after their inoculation with Pseudomonas fluorescens IMBG163 was detected in tissues by in situ hybridization method (ISH/FISH). The presence of Methylobacterium rRNA was observed in leaves and stems of potato plantlets, whereas no signal was detected in potato roots. The signal was less abundant in the untreated plants than in the pla...

  4. Fluorescence in situ hybridization: an improved method of quantitating chromosome damage and repair

    International Nuclear Information System (INIS)

    Brown, J.M.; Evans, J.W.

    1993-01-01

    The authors combined fluorescence in situ hybridization (FISH) with specific full-length chromosome probes using the premature chromosome condensation (PCC) technique chromosome condensation (PCC) technique to simplify scoring chromosome damage and its repair. They have shown the technique works well and enables breaks and exchanges to be readily detected and scored in individual chromosomes. A chromosome 4 full-length specific library has been used in initial studies. (UK)

  5. Localization of tRNAsup(asp)2 genes from Drosophila melanogaster by 'in situ' hybridization

    International Nuclear Information System (INIS)

    Schmidt, T.; Egg, A.H.; Kubli, E.

    1978-01-01

    Transfer RNAsup(asp) 2 delta was isolated from Drosophila melanogaster by affinity chromatography on concanavalin A-Sepharose. The tRNA was iodinated 'in vitro' with Na[ 125 I] and hybridized 'in situ' to salivary gland chromosomes from Drosophila. Subsequent autoradiography allowed the localization of the genes for tRNAsup(asp) 2 delta to the left arm of the second chromosome in the regions 29 D and E. (orig.) [de

  6. Actinobacillus pleuropneumoniae osteomyelitis in pigs demonstrated by fluorescent in situ hybridization

    DEFF Research Database (Denmark)

    Jensen, Tim Kåre; Boye, Mette; Hagedorn-Olsen, T.

    1999-01-01

    Necrotizing osteomyelitis and fibrinopurulent arthritis with isolation of Actinobacillus pleuropneumoniae serotype 2 is reported in two pigs from a herd with lameness and mild coughing problems among 8 to 12-week-old pigs. Application of fluorescent in situ hybridization targeting 16S ribosomal R......, in joints with arthritis, and in bone necroses including lysis of growth plate and suppurative inflammation in the adjacent trabecular metaphysis, thus demonstrating that well-known infections manifest new, unusual lesions....

  7. Hybrid Perovskite Thin-Film Photovoltaics: In Situ Diagnostics and Importance of the Precursor Solvate Phases

    KAUST Repository

    Munir, Rahim; Sheikh, Arif D.; Abdelsamie, Maged; Hu, Hanlin; Yu, Liyang; Zhao, Kui; Kim, Taesoo; El Tall, Omar; Li, Ruipeng; Smilgies, Detlef M.; Amassian, Aram

    2016-01-01

    Solution-processed hybrid perovskite semiconductors attract a great deal of attention, but little is known about their formation process. The one-step spin-coating process of perovskites is investigated in situ, revealing that thin-film formation is mediated by solid-state precursor solvates and their nature. The stability of these intermediate phases directly impacts the quality and reproducibility of thermally converted perovskite films and their photovoltaic performance.

  8. Chromogenic in situ hybridization for Her-2/neu-oncogene in breast cancer: comparison of a new dual-colour chromogenic in situ hybridization with immunohistochemistry and fluorescence in situ hybridization.

    Science.gov (United States)

    Mayr, Doris; Heim, Sibylle; Weyrauch, Kerstin; Zeindl-Eberhart, Evelyn; Kunz, Anne; Engel, Jutta; Kirchner, Thomas

    2009-12-01

    Her-2/neu testing is used as a marker for Herceptin therapy. The aim was to investigate new dual-colour chromogenic in situ hybridization (CISH), in a large number of breast carcinomas (n = 205) with DNA-specific dual-colour probes (ZytoVision, Bremerhaven, Germany) and to compare the results with immunohistochemistry (n = 205) and fluorescence in situ hybridization (FISH) (n = 129). Paraffin-embedded tissue of 205 patients was used. After immunohistochemistry with a focus on immunohistochemically uncertain cases, Her-2/neu amplification using dual-colour CISH (ZytoVision) was analysed. Validation by FISH was performed. The results were: immunohistochemistry, 27.8% with strong expression, 53.7% with uncertain overexpression and 18.5% with no expression; FISH, 25.6% amplified and 74.4% negative; CISH, 35.6% amplified, 62.9% negative and 1.5% not evaluable. Comparison of immunohistochemistry with CISH: CISH negative in 100% with immunohistochemistry 0/1+, amplified in 82.5% with immunohistochemistry 3+; 5.9% contradictory results: 4.4% immunohistochemistry 3+ and negative by CISH, 1.5% negative in immunohistochemistry but amplified by CISH; FISH (129 cases), 8.5% contradictory results to immunohistochemistry, 6.2% immunohistochemistry 3+ and negative by FISH, 2.3% negative by immunohistochemistry and amplified by FISH; comparison of CISH and FISH, 94.6% same results, 3.9% different ones, 1.6% CISH not analysable. CISH, using dual-colour probes (ZytoVision) is as good as FISH for Her-2/neu analysis. The few discrepant results are likely to be caused by polysomy or tumour heterogeneity.

  9. Ultrastructural localization of human papilloma virus by nonradioactive in situ hybridization on tissue of human cervical intraepithelial neoplasia

    DEFF Research Database (Denmark)

    Multhaupt, H A; Rafferty, P A; Warhol, M J

    1992-01-01

    BACKGROUND: A nonradioactive in situ hybridization was developed to localize human papilloma virus (HPV) at the ultrastructural level. EXPERIMENTAL DESIGN: Cervical biopsies from human uterine cervices clinically suspicious of condyloma were embedded in Lowicryl K4M at low temperature...

  10. TRIO-01 experiment: in-situ tritium-recovery results

    International Nuclear Information System (INIS)

    Clemmer, R.G.; Finn, P.A.; Billone, M.C.

    1983-08-01

    The TRIO-01 experiment is a test of in-situ tritium recovery from γ-LiAlO 2 with test conditions chosen to simulate those anticipated in fusion power reactors. A status report is presented which describes qualitatively the results observed during the irradiation phase of the experiment. Both the rate of tritium release and the chemical forms of tritium were measured using a helium sweep gas which flowed past the breeder material to a gas analysis system

  11. TRIO-01 experiment: in-situ tritium recovery results

    International Nuclear Information System (INIS)

    Clemmer, R.G.; Finn, P.A.; Billone, M.C.

    1983-10-01

    The TRIO-01 experiment is a test of in-situ tritium recovery from γ-LiAlO 2 with test conditions chosen to simulate those anticipated in fusion power reactors. A status report is presented which describes qualitatively the results observed during the irradiation phase of the experiment. Both the rate of tritium release and the chemical forms of tritium were measured using a helium sweep gas which flowed past the breeder material to a gas analysis system

  12. In situ biosynthesis of bacterial nanocellulose-CaCO3 hybrid bionanocomposite: One-step process

    International Nuclear Information System (INIS)

    Mohammadkazemi, Faranak; Faria, Marisa; Cordeiro, Nereida

    2016-01-01

    In this work, a simple and green route to the synthesis of the bacterial nanocellulose-calcium carbonate (BNC/CaCO 3 ) hybrid bionanocomposites using one-step in situ biosynthesis was studied. The CaCO 3 was incorporated in the bacterial nanocellulose structure during the cellulose biosynthesis by Gluconacetobacter xylinus PTCC 1734 bacteria. Hestrin-Schramm (HS) and Zhou (Z) culture media were used to the hybrid bionanocomposites production and the effect of ethanol addition was investigated. Attenuated total reflection Fourier transform infrared spectroscopy, field emission scanning electron microscopy, X-ray diffraction, energy-dispersive X-ray spectroscopy, inverse gas chromatography and thermogravimetric analysis were used to characterize the samples. The experimental results demonstrated that the ethanol and culture medium play an important role in the BNC/CaCO 3 hybrid bionanocomposites production, structure and properties. The BNC/CaCO 3 biosynthesized in Z culture medium revealed higher O/C ratio and amphoteric surface character, which justify the highest CaCO 3 content incorporation. The CaCO 3 was incorporated into the cellulosic matrix decreasing the bacterial nanocellulose crystallinity. This work reveals the high potential of in situ biosynthesis of BNC/CaCO 3 hybrid bionanocomposites and opens a new way to the high value-added applications of bacterial nanocellulose. - Graphical Abstract: Display Omitted - Highlights: • BNC/CaCO 3 hybrid bionanocomposites were produced using in situ biosynthesis process. • Ethanol and culture medium play an important role in the production and properties. • Z-BNC/CaCO 3 bionanocomposites revealed higher O/C ratio and amphoteric surface character. • CaCO 3 incorporated into the BNC decreased crystallinity.

  13. Combined fluorescence in situ hybridization and microautoradiography (FISH-MAR)

    International Nuclear Information System (INIS)

    Ginige, M.P.

    2005-01-01

    Cultivation independent studies have revealed, that due to the complexity of natural ecosystems such as activated sludge, rhizosphere, rumen etc., the pure cultivation of all related micro-organisms in these diverse ecosystems is quite unsuccessful. Accordingly, the small subunit ribosomal RNA (SSU rRNA, i.e. 16S and 18S rRNA) or genes, obtained from these ecosystems without cultivation has become a widely accepted approach to describe the phylogenetic diversity of microbial communities present in these ecosystems. The rapid growth of the rRNA gene (rDNA) sequence data bank, accessible via the Internet (http://www.ncbi.nlm.nih.gov/BLAST/) has enabled us to compare microbial diversities across the globe without cultivation. However, these rRNA gene sequences provide very few direct clues regarding the interactions and the metabolic capabilities of the identified micro-organisms. Accordingly, the knowledge available on the in situ physiology of the inhabitants of most microbial ecosystems is quite remote and the availability of such knowledge will enable our ability to manipulate ecosystems (e.g. activated sludge) to achieve better process performances with the aid of improved mathematical models. In both natural and engineered systems, there are a diverse group of microorganisms cohabiting within matrix-encloses such as bio-films or flocs rather than single planktonic cells. In order to manipulate these systems to obtain higher productivity, it is important to understand population structure, dynamics, functionality and also special distributions in the matrix-encloses. It is an accepted fact that pure cultivation of micro-organisms in these complex ecosystems does not permit to resolve any of the above mentioned issues since (1) rigorous biomass disaggregation which is a common practice in cultivation-based approaches results in a loss of special information; (2) cultivation approaches also introduces significant biases resulting in pronounced population shifts

  14. Chromogenic in situ hybridization (CISH): a novel alternative in screening archival breast cancer tissue samples for HER-2/neu status

    OpenAIRE

    Madrid, Manuelito A; Lo, Raymundo W

    2004-01-01

    Background Chromogenic in situ hybridization (CISH) is emerging as a practical, cost-effective, and valid alternative to fluorescent in situ hybridization in testing for gene alteration, especially in centers primarily working with immunohistochemistry (IHC). Methods We assessed Her-2/neu alteration using CISH on formalin-fixed paraffin-embedded primary invasive ductal carcinoma tumors in which IHC (CB11 antibody) had previously been performed, and we compared the results with IHC. The 160 se...

  15. Evaluation of Myc Gene Amplification in Prostate Cancer Using a Dual Color Chromogenic in-Situ Hybridization (Dual CISH) Assay

    OpenAIRE

    Daniel Lerda; Marta Cabrera; Jorge Flores; Luis Gutierrez; Armando Chierichetti; Martin Revol; Hernan Garcia Onto

    2013-01-01

    Objetive: The overall purpose of the study was to demonstrate applicability of the Dako dual-color chromogenic in situ hybridization (CISH) assay (DAKO Denmark, Glostrup) with respect to fluorescence in situ hybridization (FISH) probes MYC-C. Methods: MYC gene amplification by FISH and Dako dual-color CISH Results: The study showed that the dual-color CISH assay can convert Texas red and fluorescein isothiocyanate (FITC) signals into chromogenic signals. The dual –color CISH assay was p...

  16. In situ biosynthesis of bacterial nanocellulose-CaCO3 hybrid bionanocomposite: One-step process.

    Science.gov (United States)

    Mohammadkazemi, Faranak; Faria, Marisa; Cordeiro, Nereida

    2016-08-01

    In this work, a simple and green route to the synthesis of the bacterial nanocellulose-calcium carbonate (BNC/CaCO3) hybrid bionanocomposites using one-step in situ biosynthesis was studied. The CaCO3 was incorporated in the bacterial nanocellulose structure during the cellulose biosynthesis by Gluconacetobacter xylinus PTCC 1734 bacteria. Hestrin-Schramm (HS) and Zhou (Z) culture media were used to the hybrid bionanocomposites production and the effect of ethanol addition was investigated. Attenuated total reflection Fourier transform infrared spectroscopy, field emission scanning electron microscopy, X-ray diffraction, energy-dispersive X-ray spectroscopy, inverse gas chromatography and thermogravimetric analysis were used to characterize the samples. The experimental results demonstrated that the ethanol and culture medium play an important role in the BNC/CaCO3 hybrid bionanocomposites production, structure and properties. The BNC/CaCO3 biosynthesized in Z culture medium revealed higher O/C ratio and amphoteric surface character, which justify the highest CaCO3 content incorporation. The CaCO3 was incorporated into the cellulosic matrix decreasing the bacterial nanocellulose crystallinity. This work reveals the high potential of in situ biosynthesis of BNC/CaCO3 hybrid bionanocomposites and opens a new way to the high value-added applications of bacterial nanocellulose. Copyright © 2016 Elsevier B.V. All rights reserved.

  17. Fluorescence in situ hybridization of old G-banded and mounted chromosome preparations

    DEFF Research Database (Denmark)

    Gerdes, A M; Pandis, N; Bomme, L

    1997-01-01

    the coverslips detach spontaneously; any mechanical manipulation will jeopardize the results. The success of chromosome painting is improved by excluding the regular RNase treatment step prior to hybridization. Additional changes compared with standard FISH protocols are that the 2 x SSC step is omitted......An improved method for fluorescence in situ hybridization (FISH) investigation of old, previously G-banded, mounted chromosome preparations with chromosome specific painting probes and centromere-specific probes is described. Before hybridization, the slides are incubated in xylene until......, that the amount of added probe is increased approximately 2.5 times, and that the amplification of signals is performed twice. The applicability of the method, which allows double painting with two differently labeled probes using two differently fluorescing colors, was tested on 11 cases involving different...

  18. Hard and transparent hybrid polyurethane coatings using in situ incorporation of calcium carbonate nanoparticles

    International Nuclear Information System (INIS)

    Yao Lu; Yang Jie; Sun Jing; Cai Lifang; He Linghao; Huang Hui; Song Rui; Hao Yongmei

    2011-01-01

    Highlights: → In situ mineralization via gas diffusion was adopted for a good dispersion of calcium carbonate nanoparticles in the polymeric PU matrix. → Hybrid films with high dispersion, transparency, robust and thermal stability can be obtained by controlling the CaCO 3 loading. → The hybrid films display a significant improvement in its water resistance, surface hardness, scratch resistance and flexibility, with the introduction of CaCO 3 , and all coatings exhibited excellent chemical resistance and adhesion. - Abstract: The combination of hardness, scratch resistance, and flexibility is a highly desired feature in many coating applications. The aim of this study is to achieve this goal through the in situ introduction of an unmodified calcium carbonate (CaCO 3 ) into a water-soluble polyurethane (PU) matrix. Smooth and (semi-) transparent films were prepared from both the neat PU and the CaCO 3 -filled composites. As evidenced by the measurements from scanning electron microscopy (SEM), optical microscopy, dynamic mechanical analysis (DMA) and thermogravimetric analysis (TGA), hybrid films with high dispersion, transparency, robustness and thermal stability could be obtained by controlling the CaCO 3 loading. The storage modulus could increase from 441 MPa of neat PU matrix to 1034 MPa of hybrid film containing 2% (w/w) CaCO 3 . In addition, the same hybrid films displayed a significant improvement in its water resistance. In this case, the water-uptake ratio decreased from 41.54% of PU to 2.21% of hybrid film containing 2% (w/w) CaCO 3 . Moreover, with the introduction of CaCO 3 , conventional coating characterization methods demonstrated an increase in the surface hardness, scratch resistance and flexibility, and all coatings exhibited excellent chemical resistance and adhesion.

  19. Geothermal in situ experiments in the Asse salt-mine

    International Nuclear Information System (INIS)

    Kopietz, J.; Jung, R.

    1978-01-01

    The paper presents design and results of in situ experiments carried out by the Bundesanstat fuer Geowissenschaften und Rohstoffe (Federal Institute for Geosciences and Natural Resources, F.R. of Germany) in the Asse salt-mine. With reference to model calculations of the temperature field which is produced in salt formations by radioactive waste, temperature measurements in the area of electrical heating elements and in situ measurements of thermal conductivity have been performed. The measured temperatures are in good accordance with the theoretical prediction. Preliminary results of the thermal conductivity measurements correspond with the data of single NaCl crystals published by Birch and Clark. At present a heating experiment is being conducted in the Asse mine to investigate thermo-mechanical effects of a cylindrical heat source upon the surrounding rock salt. Possible thermal induced fractures monitored by permeability changes and seismoacoustical phenomena are the main objects of this experiment

  20. [HER-2 oncogene amplification assessment in invasive breast cancer by dual-color in situ hybridization (dc-CISH): a comparative study with fluorescent in situ hybridization (FISH)].

    Science.gov (United States)

    Akhdar, Abbas; Bronsard, Marc; Lemieux, Renald; Geha, Sameh

    2011-12-01

    The amplification of the gene encoding for the human epidermal growth factor receptor 2 (HER-2 oncogene), located on chromosome 17 (17q21-q22), or the overexpression of this receptor have prognostic and therapeutic implications in invasive breast cancer. An evaluation of the HER-2 status by immunohistochemistry (IHC) is performed on all invasive breast cancer cases. Fluorescent in situ hybridization (FISH) is considered as the gold standard for the detection of HER-2 gene amplification for IHC equivocal cases (score 2+). A more recent in situ hybridization technique, the dual-color chromogenic in situ hybridization (dc-CISH), has been proposed as an alternative to FISH. The aim of this study was to measure the correlation between dc-CISH and FISH for HER-2 oncogene amplification assessment in invasive breast cancer. We built four tissue micro-array (TMA) blocs with 100 breast invasive cancer cases that had been previously tested by IHC for HER-2 detection: 10 score 0 cases, 10 score 3+cases, 39 score 1+and 41 score 2+cases. Both FISH and dc-CISH techniques were applied on all TMA cases as well as on two additional slides serving as controls. Interpretation of dc-CISH was carried out by a pathologist using an optical microscope. For FISH, the interpretation was done by a professional from the medical genetics department using a fluorescent microscope linked to a computer system for image capturing and analysis. The interpretation of the HER-2/CEN-17 ratio for both tests was in accordance with the values of the updated recommendations from the Canadian National Consensus Meeting on HER-2/neu testing in breast cancer and from the ASCO/CAP. Among the 100 cases initially included in the study, eight were excluded from the analysis due to sampling or technical flaws. From the 92 remaining cases, we obtained a concordance of 97.8% (90/92 cases) between the two techniques (Kappa coefficient 0.97, 95% confidence interval). The correlation coefficient (rho) between ratios

  1. A list of image files of planarians analyzed by in situ hybridication and immunohistochemical staining - Plabrain DB | LSDB Archive [Life Science Database Archive metadata

    Lifescience Database Archive (English)

    Full Text Available List Contact us Plabrain DB A list of image files of planarians analyzed by in situ hybridication and immunohistochemical...tu hybridication and also protein distribution by immunohistochemical staining in intact planarians or plana...planarians analyzed by In situ hybridication and immunohistochemical staining . D..._image#en Data acquisition method Whole-mount in situ hybridication, immunohistochemical...te Policy | Contact Us A list of image files of planarians analyzed by in situ hybridication and immunohistochemical staining - Plabrain DB | LSDB Archive ...

  2. Detection and quantitative analysis of actin mRNA by in situ hybridization with an oligodeoxynucleotide probe

    International Nuclear Information System (INIS)

    Taneja, K.; Singer, R.

    1987-01-01

    In situ hybridization is a useful method for localizing specific nucleic acid sequences intracellularly and for studying regulation of gene expression. Recently synthetic oligonucleotides have been successfully used as probes in this technique. Since they can be made easily to specific nucleic acid regions, they may be the best approach for analysis of a gene family of highly conserved sequences. They have analyzed these probes for the development of an in situ hybridization method. Oligonucleotides were made to different regions of chick beta-actin mRNA and used for detection of these sequences in a culture of chicken fibroblasts and myoblasts. They found that synthetic DNAs have different efficiencies of hybridization, indicating that not all target sequences are equivalent. They have investigated in detail a particular probe to the actin mRNA coding region and have optimized hybridization parameters. When hybridization was quantitated it was found that an oligonucleotide end labelled with 35 S or 32 P was capable of detecting several thousand messages per cell with a signal-to-noise ratio of 10:1. In situ hybridization confirmed the specificity of the hybridization as well as the background level. Increase in the number of oligonucleotides used should increase the signal-to-noise ratio-proportionately. Under particular circumstances the specificity of oligonucleotides make them an important reagent for in situ hybridization

  3. Hierarchically structured transparent hybrid membranes by in situ growth of mesostructured organosilica in host polymer

    Science.gov (United States)

    Vallé, Karine; Belleville, Philippe; Pereira, Franck; Sanchez, Clément

    2006-02-01

    The elaborate performances characterizing natural materials result from functional hierarchical constructions at scales ranging from nanometres to millimetres, each construction allowing the material to fit the physical or chemical demands occurring at these different levels. Hierarchically structured materials start to demonstrate a high input in numerous promising applied domains such as sensors, catalysis, optics, fuel cells, smart biologic and cosmetic vectors. In particular, hierarchical hybrid materials permit the accommodation of a maximum of elementary functions in a small volume, thereby optimizing complementary possibilities and properties between inorganic and organic components. The reported strategies combine sol-gel chemistry, self-assembly routes using templates that tune the material's architecture and texture with the use of larger inorganic, organic or biological templates such as latex, organogelator-derived fibres, nanolithographic techniques or controlled phase separation. We propose an approach to forming transparent hierarchical hybrid functionalized membranes using in situ generation of mesostructured hybrid phases inside a non-porogenic hydrophobic polymeric host matrix. We demonstrate that the control of the multiple affinities existing between organic and inorganic components allows us to design the length-scale partitioning of hybrid nanomaterials with tuned functionalities and desirable size organization from ångström to centimetre. After functionalization of the mesoporous hybrid silica component, the resulting membranes have good ionic conductivity offering interesting perspectives for the design of solid electrolytes, fuel cells and other ion-transport microdevices.

  4. Utility of chromogenic in situ hybridization (CISH) for detection of EGFR amplification in glioblastoma: comparison with fluorescence in situ hybridization (FISH).

    Science.gov (United States)

    Fischer, Ingeborg; de la Cruz, Clarissa; Rivera, Andreana L; Aldape, Kenneth

    2008-12-01

    In this study, we test the reliability of chromogenic in situ hybridization (CISH) for the detection of epidermal growth factor receptor (EGFR) gene amplification in glioblastoma. Earlier reports have described EGFR CISH in glioblastoma multiforme, but a comparison of CISH with a "gold standard" testing method, such as fluorescence in situ hybridization (FISH), has not been described. Therapies targeting the EGFR-signaling pathway might increase the importance of assessment of EGFR-amplification status. CISH is a potential alternative to FISH as a testing method. To test its reliability, EGFR-amplification status by CISH was assessed in 89 cases of glioblastoma and compared with FISH results, and correlated with the protein expression using immunohistochemistry (IHC) for EGFR. FISH was scored as being EGFR-amplified in 47/89 tumors, CISH as being amplified in 43/89 tumors. The CISH and FISH results were in agreement in 83/89 cases (93%). Four glioblastomas were scored as being amplified by FISH, but not by CISH; whereas amplification was detected in 2 tumors by CISH that were not amplified using FISH. Forty-eight of the 89 cases were positive for EGFR expression by IHC. EGFR amplification was highly correlated with protein expression by IHC, as 40/48 (83%) EGFR IHC-positive cases were found to be EGFR-amplified. The high concordance of CISH and FISH for the assessment of EGFR gene-amplification status indicates that CISH is a viable alternative to FISH for the detection of EGFR gene amplification in glioblastoma. Detectable EGFR expression by IHC can occur in the absence of gene amplification, but is uncommon.

  5. The correlation between dual-color chromogenic in situ hybridization and fluorescence in situ hybridization in assessing HER2 gene amplification in breast cancer.

    Science.gov (United States)

    Pedersen, Marianne; Rasmussen, Birgitte Bruun

    2009-06-01

    Fluorescence in situ hybridization (FISH) is regarded as the gold standard method for detecting HER2 gene amplification. Chromogenic in situ hybridization (CISH) is a promising alternative to FISH because CISH has the advantages of being a method evaluated by bright-field microscopy and the generated chromogenic signals are also stable. This study presents a dual color CISH for simultaneous detection of the HER2 gene and chromosome 17. The CISH method performs a chromogenic detection "on top" of the Food and Drug Administration (FDA)-approved HER2 FISH pharmDx method, where the fluorochrome-labeled probes are detected using enzyme-labeled antibodies and visualized by chromogenic enzymatic reactions. The HER2 status (amplified/not amplified and HER2 ratios) was evaluated by the CISH method and compared with results obtained by the FDA-approved FISH method. Of the 72 successfully investigated invasive breast carcinomas, both FISH and CISH detected HER2 amplification in 24 cases and nonamplification was detected in 47 cases. One case showed a discrepancy between FISH and CISH. The concordance between CISH and FISH was found to be almost perfect (98.6%). The correlation between the HER2 ratios obtained by the 2 methods showed excellent correlation (correlation coefficient 0.95). In conclusion, it is possible by dual-color CISH method to demonstrate HER2 genes and chromosome 17 genes, in the same tissue section and reliably assess HER2 status. The CISH method is a very promising alternative to the FISH method.

  6. Dual-colour chromogenic in-situ hybridization is a potential alternative to fluorescence in-situ hybridization in HER2 testing.

    Science.gov (United States)

    Hwang, Cheng-Cheng; Pintye, Mariann; Chang, Liang-Che; Chen, Huang-Yang; Yeh, Kun-Yan; Chein, Hui-Ping; Lee, Nin; Chen, Jim-Ray

    2011-11-01

    Dual-colour chromogenic in-situ hybridization (dc-CISH) is an emerging methodology for characterizing genomic alterations. This study was aimed at evaluating the performance of a dc-CISH kit (ZytoVision) in determining human epidermal growth factor receptor 2 (HER2) status in breast cancer. Two hundred and twenty-eight invasive breast carcinomas arranged in tissue microarrays were analysed in parallel with dc-CISH, fluorescence in-situ hybridization (FISH), and immunohistochemistry. Of 227 tumours with available FISH and dc-CISH results, HER2 amplification and non-amplification were detected in 49 (21.6%) and 178 (78.4%) tumours, respectively, by both assays. The concordance between dc-CISH and FISH results showed 100% agreement (κ-coefficient=1.00). Immunohistochemically, 162 (71%), 25 (11.0%) and 41 (18%) tumours were scored 0/1+, 2+, and 3+, respectively. The corresponding results with both FISH and dc-CISH demonstrated HER2 amplification in two (3.2%), nine (36%) and 38 (93%) tumours, respectively. Complete consensus among these three methods was observed in 197 cases, representing 98% of all 3+ and 0/1+ tumours (κ-coefficient=0.92). Confirmatory testing of 25 2+ tumours showed complete consensus between FISH and dc-CISH. dc-CISH is a promising alternative to FISH in HER2 testing, and the single-institute incidence of HER2 amplification in breast cancer in Taiwan is 21.2%. © 2011 Blackwell Publishing Limited.

  7. Immunoglobulin heavy-chain fluorescence in situ hybridization-chromogenic in situ hybridization DNA probe split signal in the clonality assessment of lymphoproliferative processes on cytological samples.

    Science.gov (United States)

    Zeppa, Pio; Sosa Fernandez, Laura Virginia; Cozzolino, Immacolata; Ronga, Valentina; Genesio, Rita; Salatiello, Maria; Picardi, Marco; Malapelle, Umberto; Troncone, Giancarlo; Vigliar, Elena

    2012-12-25

    The human immunoglobulin heavy-chain (IGH) locus at chromosome 14q32 is frequently involved in different translocations of non-Hodgkin lymphoma (NHL), and the detection of any breakage involving the IGH locus should identify a B-cell NHL. The split-signal IGH fluorescence in situ hybridization-chromogenic in situ hybridization (FISH-CISH) DNA probe is a mixture of 2 fluorochrome-labeled DNAs: a green one that binds the telomeric segment and a red one that binds the centromeric segment, both on the IGH breakpoint. In the current study, the authors tested the capability of the IGH FISH-CISH DNA probe to detect IGH translocations and diagnose B-cell lymphoproliferative processes on cytological samples. Fifty cytological specimens from cases of lymphoproliferative processes were tested using the split-signal IGH FISH-CISH DNA probe and the results were compared with light-chain assessment by flow cytometry (FC), IGH status was tested by polymerase chain reaction (PCR), and clinicohistological data. The signal score produced comparable results on FISH and CISH analysis and detected 29 positive, 15 negative, and 6 inadequate cases; there were 29 true-positive cases (66%), 9 true-negative cases (20%), 6 false-negative cases (14%), and no false-positive cases (0%). Comparing the sensitivity of the IGH FISH-CISH DNA split probe with FC and PCR, the highest sensitivity was obtained by FC, followed by FISH-CISH and PCR. The split-signal IGH FISH-CISH DNA probe is effective in detecting any translocation involving the IGH locus. This probe can be used on different samples from different B-cell lymphoproliferative processes, although it is not useful for classifying specific entities. Cancer (Cancer Cytopathol) 2012;. © 2012 American Cancer Society. Copyright © 2012 American Cancer Society.

  8. Hybrid rocket engine, theoretical model and experiment

    Science.gov (United States)

    Chelaru, Teodor-Viorel; Mingireanu, Florin

    2011-06-01

    The purpose of this paper is to build a theoretical model for the hybrid rocket engine/motor and to validate it using experimental results. The work approaches the main problems of the hybrid motor: the scalability, the stability/controllability of the operating parameters and the increasing of the solid fuel regression rate. At first, we focus on theoretical models for hybrid rocket motor and compare the results with already available experimental data from various research groups. A primary computation model is presented together with results from a numerical algorithm based on a computational model. We present theoretical predictions for several commercial hybrid rocket motors, having different scales and compare them with experimental measurements of those hybrid rocket motors. Next the paper focuses on tribrid rocket motor concept, which by supplementary liquid fuel injection can improve the thrust controllability. A complementary computation model is also presented to estimate regression rate increase of solid fuel doped with oxidizer. Finally, the stability of the hybrid rocket motor is investigated using Liapunov theory. Stability coefficients obtained are dependent on burning parameters while the stability and command matrixes are identified. The paper presents thoroughly the input data of the model, which ensures the reproducibility of the numerical results by independent researchers.

  9. In situ hybridization studies of hepatitis A viral RNA in patients with acute hepatitis A

    Energy Technology Data Exchange (ETDEWEB)

    Taylor, M; Goldin, R D; Ladva, S [Department of Histopathology, St. Mary' s Hospital Medical School, Imperial College of Science, Technology and Medicine, London (United Kingdom); Scheuer, P J [Department of Histopathology, Royal Free Hospital and School of Medicine, London (United Kingdom); Thomas, H C [Department of Medicine, St. Mary' s Hospital Medical School, Imperial College of Science, Technology and Medicine, London (United Kingdom)

    1994-01-01

    In situ hybridization with oligonucleotide probes has been used to localise hepatitis A virus RNA genomic sequences in formalin-fixed and routinely processed human liver biopsies from three patients. Using radiolabelled Sulphur-35 antisense probes, viral genomic sequences were found in all three cases, but signal intensity was greatest in cases 1 and 2 with fulminant hepatitis, and was minimal in the third case of resolving hepatitis biopsied 2 months after acute illness. Localisation showed the viral RNA to be present in hepatocytes, sinusoidal cells and inflammatory cells in and around the portal tracts. Both cases showed signal in similar cell types, but the distribution of staining was predominantly periportal in case 1, whereas lobular staining was more apparent in case 2. Hybridization with sense polarity probes failed to detect any evidence of replicative intermediates of antigenomic viral RNA. The presence of hepatitis A RNA in phagocytic cells was confirmed using immunohistochemistryfor a macrophage marker, CD68, combined with in situ hybridization. In all cases the signal was predominantly cytoplasmic, and this was confirmed with the use of tritiated probes. (au).

  10. In situ hybridization studies of hepatitis A viral RNA in patients with acute hepatitis A

    International Nuclear Information System (INIS)

    Taylor, M.; Goldin, R.D.; Ladva, S.; Scheuer, P.J.; Thomas, H.C.

    1994-01-01

    In situ hybridization with oligonucleotide probes has been used to localise hepatitis A virus RNA genomic sequences in formalin-fixed and routinely processed human liver biopsies from three patients. Using radiolabelled Sulphur-35 antisense probes, viral genomic sequences were found in all three cases, but signal intensity was greatest in cases 1 and 2 with fulminant hepatitis, and was minimal in the third case of resolving hepatitis biopsied 2 months after acute illness. Localisation showed the viral RNA to be present in hepatocytes, sinusoidal cells and inflammatory cells in and around the portal tracts. Both cases showed signal in similar cell types, but the distribution of staining was predominantly periportal in case 1, whereas lobular staining was more apparent in case 2. Hybridization with sense polarity probes failed to detect any evidence of replicative intermediates of antigenomic viral RNA. The presence of hepatitis A RNA in phagocytic cells was confirmed using immunohistochemistryfor a macrophage marker, CD68, combined with in situ hybridization. In all cases the signal was predominantly cytoplasmic, and this was confirmed with the use of tritiated probes. (au)

  11. Fluorescent In Situ Hybridization to Detect Transgene Integration into Plant Genomes

    Science.gov (United States)

    Schwarzacher, Trude

    Fluorescent chromosome analysis technologies have advanced our understanding of genome organization during the last 30 years and have enabled the investigation of DNA organization and structure as well as the evolution of chromosomes. Fluorescent chromosome staining allows even small chromosomes to be visualized, characterized by their composition and morphology, and counted. Aneuploidies and polyploidies can be established for species, breeding lines, and individuals, including changes occurring during hybridization or tissue culture and transformation protocols. Fluorescent in situ hybridization correlates molecular information of a DNA sequence with its physical location on chromosomes and genomes. It thus allows determination of the physical position of sequences and often is the only means to determine the abundance and distribution of DNA sequences that are difficult to map with any other molecular method or would require segregation analysis, in particular multicopy or repetitive DNA. Equally, it is often the best way to establish the incorporation of transgenes, their numbers, and physical organization along chromosomes. This chapter presents protocols for probe and chromosome preparation, fluorescent in situ hybridization, chromosome staining, and the analysis of results.

  12. Fluorescence In situ Hybridization: Cell-Based Genetic Diagnostic and Research Applications.

    Science.gov (United States)

    Cui, Chenghua; Shu, Wei; Li, Peining

    2016-01-01

    Fluorescence in situ hybridization (FISH) is a macromolecule recognition technology based on the complementary nature of DNA or DNA/RNA double strands. Selected DNA strands incorporated with fluorophore-coupled nucleotides can be used as probes to hybridize onto the complementary sequences in tested cells and tissues and then visualized through a fluorescence microscope or an imaging system. This technology was initially developed as a physical mapping tool to delineate genes within chromosomes. Its high analytical resolution to a single gene level and high sensitivity and specificity enabled an immediate application for genetic diagnosis of constitutional common aneuploidies, microdeletion/microduplication syndromes, and subtelomeric rearrangements. FISH tests using panels of gene-specific probes for somatic recurrent losses, gains, and translocations have been routinely applied for hematologic and solid tumors and are one of the fastest-growing areas in cancer diagnosis. FISH has also been used to detect infectious microbias and parasites like malaria in human blood cells. Recent advances in FISH technology involve various methods for improving probe labeling efficiency and the use of super resolution imaging systems for direct visualization of intra-nuclear chromosomal organization and profiling of RNA transcription in single cells. Cas9-mediated FISH (CASFISH) allowed in situ labeling of repetitive sequences and single-copy sequences without the disruption of nuclear genomic organization in fixed or living cells. Using oligopaint-FISH and super-resolution imaging enabled in situ visualization of chromosome haplotypes from differentially specified single-nucleotide polymorphism loci. Single molecule RNA FISH (smRNA-FISH) using combinatorial labeling or sequential barcoding by multiple round of hybridization were applied to measure mRNA expression of multiple genes within single cells. Research applications of these single molecule single cells DNA and RNA FISH

  13. Fluorescence In situ Hybridization: Cell-Based Genetic Diagnostic and Research Applications

    Directory of Open Access Journals (Sweden)

    Chenghua Cui

    2016-09-01

    Full Text Available Fluorescence in situ hybridization (FISH is a macromolecule recognition technology based on the complementary nature of DNA or DNA/RNA double strands. Selected DNA strands incorporated with fluorophore-coupled nucleotides can be used as probes to hybridize onto the complementary sequences in tested cells and tissues and then visualized through a fluorescence microscope or an imaging system. This technology was initially developed as a physical mapping tool to delineate genes within chromosomes. Its high analytical resolution to a single gene level and high sensitivity and specificity enabled an immediate application for genetic diagnosis of constitutional common aneuploidies, microdeletion/microduplication syndromes and subtelomeric rearrangements. FISH tests using panels of gene-specific probes for somatic recurrent losses, gains and translocations have been routinely applied for hematologic and solid tumors and are one of the fastest-growing areas in cancer diagnosis. FISH has also been used to detect infectious microbials and parasites like malaria in human blood cells. Recent advances in FISH technology involve various methods for improving probe labeling efficiency and the use of super resolution imaging systems for direct visualization of intra-nuclear chromosomal organization and profiling of RNA transcription in single cells. Cas9-mediated FISH (CASFISH allowed in situ labeling of repetitive sequences and single-copy sequences without the disruption of nuclear genomic organization in fixed or living cells. Using oligopaint-FISH and super-resolution imaging enabled in situ visualization of chromosome haplotypes from differentially specified single-nucleotide polymorphism loci. Single molecule RNA FISH (smRNA-FISH using combinatorial labeling or sequential barcoding by multiple round of hybridization were applied to measure mRNA expression of multiple genes within single cells. Research applications of these single molecule single cells

  14. Facile In Situ Fabrication of Nanostructured Graphene–CuO Hybrid with Hydrogen Sulfide Removal Capacity

    Institute of Scientific and Technical Information of China (English)

    Sunil P.Lonkar; Vishnu V.Pillai; Samuel Stephen; Ahmed Abdala; Vikas Mittal

    2016-01-01

    A simple and scalable synthetic approach for one-step synthesis of graphene–Cu O(TRGC) nanocomposite by an in situ thermo-annealing method has been developed.Using graphene oxide(GO) and copper hydroxide as a precursors reagent,the reduction of GO and the uniform deposition of in situ formed Cu O nanoparticles on graphene was simultaneously achieved.The method employed no solvents,toxic-reducing agents,or organic modifiers.The resulting nanostructured hybrid exhibited improved H2 S sorption capacity of 1.5 mmol H2S/g-sorbent(3 g S/100 g-sorbent).Due to its highly dispersed sub-20 nm Cu O nanoparticles and large specific surface area,TRGC nanocomposite exhibits tremendous potential for energy and environment applications.

  15. In situ hybridization for the detection of infectious laryngotracheitis virus in sections of trachea from experimentally infected chickens

    DEFF Research Database (Denmark)

    Nielsen, O.L.; Handberg, Kurt; Jørgensen, Poul Henrik

    1998-01-01

    An in situ hybridization procedure for the detection of infectious laryngotracheitis virus (ILTV) in experimentally infected chickens is described. Formalin-fixed, paraffin-embedded sections of trachea, taken from chickens on days 3-10 post-inoculation (p.i.) with ILTV were hybridized with a mixt......An in situ hybridization procedure for the detection of infectious laryngotracheitis virus (ILTV) in experimentally infected chickens is described. Formalin-fixed, paraffin-embedded sections of trachea, taken from chickens on days 3-10 post-inoculation (p.i.) with ILTV were hybridized...... on day 5 p.i. No hybridization was observed in 3 out of 3 chickens examined on day 10 p.i. ILTV nucleic acid was detected in nuclei of degenerated tracheal epithelial cells and in intranuclear inclusion bodies of syncytia....

  16. Localization of insulin receptor mRNA in rat brain by in situ hybridization

    International Nuclear Information System (INIS)

    Marks, J.L.; Porte, D. Jr.; Stahl, W.L.; Baskin, D.G.

    1990-01-01

    Insulin receptor mRNA was demonstrated in rat brain slices by in situ hybridization with three 35 S-oligonucleotide probes and contact film autoradiography. Specificity was confirmed by showing that (a) excess unlabeled probe abolished the signal, (b) an oligonucleotide probe for rat neuropeptide Y mRNA showed a different distribution of hybridization signal, and (c) the distribution of insulin receptor binding was consistent with the distribution of insulin receptor mRNA. Insulin receptor mRNA was most abundant in the granule cell layers of the olfactory bulb, cerebellum and dentate gyrus, in the pyramidal cell body layers of the pyriform cortex and hippocampus, in the choroid plexus and in the arcuate nucleus of the hypothalamus

  17. Nerve growth factor mRNA in brain: localization by in situ hybridization

    International Nuclear Information System (INIS)

    Rennert, P.D.; Heinrich, G.

    1986-01-01

    Nerve Growth Factor is a 118 amino acid polypeptide that plays an important role in the differentiation and survival of neurons. The recent discovery that a mRNA that encodes beta Nerve Growth Factor is present in brain suggests that the Nerve Growth Factor gene may not only regulate gene expression of peripheral but also of central neurons. To identify the site(s) of Nerve Growth Factor mRNA production in the brain and to determine which cells express the Nerve Growth Factor gene, the technique of in situ hybridization was employed. A 32P-labeled RNA probe complementary to Nerve Growth Factor mRNA hybridized to cells in the stratum granulosum of the dentate gyrus and the stratum pyramidale of the hippocampus. These observations identify for the first time cellular sites of Nerve Growth Factor gene expression in the central nervous system, and suggest that Nerve Growth Factor mRNA is produced by neurons

  18. Detection of human papillomavirus in laryngeal lesions by in situ hybridization

    DEFF Research Database (Denmark)

    Multhaupt, H A; Fessler, J N; Warhol, M J

    1994-01-01

    Human papillomavirus (HPV) is associated with human neoplasms of squamous epithelium. Squamous papillomas and verrucous carcinomas are two types of squamous neoplasms of the larynx that present difficult problems in differential diagnosis. Using in situ hybridization with biotinylated DNA probes......, we examined benign squamous papillomas and verrucous squamous carcinomas of the larynx for the presence of HPV. Forty-two biopsy specimens from 18 patients with laryngeal papillomas and 11 biopsy specimens from seven patients with verrucous carcinomas were obtained from the files of Pennsylvania...... Hospital, Philadelphia, PA. Tissue sections were hybridized with an HPV DNA cocktail. The HPV-positive cases then were subtyped further with DNA probes specific for HPV subtypes 6/11, 16/18, and 31/33/35. All benign squamous papillomas (42 of 42) were positive for HPV subtype 6/11. None of the verrucous...

  19. In situ hybridization of somatolactin transcripts in the pituitary glands from acclimatized carp (Cyprinus carpio

    Directory of Open Access Journals (Sweden)

    MAURICIO LÓPEZ

    2001-01-01

    Full Text Available We isolated and cloned a carp somatolactin SL DNA fragment, of which 78% of the nucleotides were identical to the corresponding salmon SL sequence. The results obtained upon Northern blot hybridization of carp pituitary RNA allowed the identification of two transcripts as described for other fish. When the content of SL transcripts in pituitary sections from summer- and winter- acclimatized carp was quantified by in situ hybridization assays, we found no significant differences between the two seasons. In salmonids, plasma SL reaches higher levels in summer than in winter in synchrony with the water temperature cycle; in the eurythermal carp, however, the complex adaptive responses imposed by seasonal environmental changes do not seem to include the regulation of the somatolactin detected with the probe used at the transcriptional level in pituitary glands

  20. Localization of mRNA in vertebrate axonal compartments by in situ hybridization.

    Science.gov (United States)

    Sotelo-Silveira, José Roberto; Calliari, Aldo; Kun, Alejandra; Elizondo, Victoria; Canclini, Lucía; Sotelo, José Roberto

    2011-01-01

    The conclusive demonstration of RNA in vertebrate axons by in situ hybridization (ISH) has been elusive. We review the most important reasons for difficulties, including low concentration of axonal RNAs, localization in specific cortical domains, and the need to isolate axons. We demonstrate the importance of axon micro-dissection to obtain a whole mount perspective of mRNA distribution in the axonal territory. We describe a protocol to perform fluorescent ISH in isolated axons and guidelines for the preservation of structural and molecular integrity of cortical RNA-containing domains (e.g., Periaxoplasmic Ribosomal Plaques, or PARPs) in isolated axoplasm.

  1. Application of locked nucleic acid-based probes in fluorescence in situ hybridization

    DEFF Research Database (Denmark)

    Fontenete, Sílvia; Carvalho, Daniel R; Guimarães, Nuno

    2016-01-01

    of nucleic acid mimics used as mixmers in LNA-based probes strongly influence the efficiency of detection. LNA probes with 10 to 15 mers showed the highest efficiency. Additionally, the combination of 2′-OMe RNA with LNA allowed an increase on the fluorescence intensities of the probes. Overall......Fluorescence in situ hybridization (FISH) employing nucleic acid mimics as probes is becoming an emerging molecular tool in the microbiology area for the detection and visualization of microorganisms. However, the impact that locked nucleic acid (LNA) and 2′-O-methyl (2′-OMe) RNA modifications have...

  2. Detection of Methylobacterium radiotolerans IMBG290 in potato plants by in situ hybridization

    Directory of Open Access Journals (Sweden)

    Pirttila A. M.

    2009-04-01

    Full Text Available A new bacterial strain of pink-pigmented facultative methylotroph (M. radiotolerans IMBG290 which was previously isolated from in vitro grown potato plantlets after their inoculation with Pseudomonas fluorescens IMBG163 was detected in tissues by in situ hybridization method (ISH/FISH. The presence of Methylobacterium rRNA was observed in leaves and stems of potato plantlets, whereas no signal was detected in potato roots. The signal was less abundant in the untreated plants than in the plantlets infected with M. radiotolerans IMBG290.

  3. Transmission electron microscopic method for gene mapping on polytene chromosomes by in situ hybridization

    OpenAIRE

    Wu, Madeline; Davidson, Norman

    1981-01-01

    A transmission electron microscope method for gene mapping by in situ hybridization to Drosophila polytene chromosomes has been developed. As electron-opaque labels, we use colloidal gold spheres having a diameter of 25 nm. The spheres are coated with a layer of protein to which Escherichia coli single-stranded DNA is photochemically crosslinked. Poly(dT) tails are added to the 3' OH ends of these DNA strands, and poly(dA) tails are added to the 3' OH ends of a fragmented cloned Drosophila DN...

  4.   In situ identification of streptococci and other bacteria in initial dental biofilm by confocal laser scanning microscopy and fluorescence in situ hybridization

    DEFF Research Database (Denmark)

    Dige, Irene; Kilian, Mogens; Nilsson, Holger

    2007-01-01

    Confocal laser scanning microscopy (CLSM) has been employed as a method for studying intact natural biofilm. When combined with fluorescence in situ hybridization (FISH) it is possible to analyze spatial relationships and changes of specific members of microbial populations over time. The aim...

  5. Oligonucleotide PIK3CA/Chromosome 3 Dual in Situ Hybridization Automated Assay with Improved Signals, One-Hour Hybridization, and No Use of Blocking DNA.

    Science.gov (United States)

    Zhang, Wenjun; Hubbard, Antony; Baca-Parkinson, Leslie; Stanislaw, Stacey; Vladich, Frank; Robida, Mark D; Grille, James G; Maxwell, Daniel; Tsao, Tsu-Shuen; Carroll, William; Gardner, Tracie; Clements, June; Singh, Shalini; Tang, Lei

    2015-09-01

    The PIK3CA gene at chromosome 3q26.32 was found to be amplified in up to 45% of patients with squamous cell carcinoma of the lung. The strong correlation between PIK3CA amplification and increased phosphatidylinositol 3-kinase (PI3K) pathway activities suggested that PIK3CA gene copy number is a potential predictive biomarker for PI3K inhibitors. Currently, all microscopic assessments of PIK3CA and chromosome 3 (CHR3) copy numbers use fluorescence in situ hybridization. PIK3CA probes are derived from bacterial artificial chromosomes whereas CHR3 probes are derived mainly from the plasmid pHS05. These manual fluorescence in situ hybridization assays mandate 12- to 18-hour hybridization and use of blocking DNA from human sources. Moreover, fluorescence in situ hybridization studies provide limited morphologic assessment and suffer from signal decay. We developed an oligonucleotide-based bright-field in situ hybridization assay that overcomes these shortcomings. This assay requires only a 1-hour hybridization with no need for blocking DNA followed by indirect chromogenic detection. Oligonucleotide probes produced discrete and uniform CHR3 stains superior to those from the pHS05 plasmid. This assay achieved successful staining in 100% of the 195 lung squamous cell carcinoma resections and in 94% of the 33 fine-needle aspirates. This robust automated bright-field dual in situ hybridization assay for the simultaneous detection of PIK3CA and CHR3 centromere provides a potential clinical diagnostic method to assess PIK3CA gene abnormality in lung tumors. Copyright © 2015 American Society for Investigative Pathology and the Association for Molecular Pathology. Published by Elsevier Inc. All rights reserved.

  6. Demonstration of interleukin-1 beta transcripts in acute myeloblastic leukemic cells by in situ hybridization.

    Science.gov (United States)

    Nakamura, M; Kanakura, Y; Furukawa, Y; Ernst, T J; Griffin, J D

    1990-07-01

    The cells from some patients with acute myeloblastic leukemia will secrete autostimulatory cytokines in tissue culture without the addition of stimulators such as phorbol 12-myristate 13-acetate. Production of interleukin-1 beta (IL-1 beta), for example, has been observed in up to 50% of cases. In order to investigate the nature of the cell secreting IL-1 beta in AML, we used an antisense RNA probe to detect specific IL-1 beta transcripts in individual leukemic cells by in situ hybridization. In fresh, uncultured cells, IL-1 beta transcripts were observed in 1-40% of undifferentiated leukemic blast cells in 17 of 19 cases. In situ hybridization was at least as sensitive as Northern blot analysis in detecting IL-1 beta transcripts. No correlation of IL-1 beta transcript expression with FAB classification was observed. Normal blood and bone marrow mononuclear cells did not contain cells expressing IL-1 beta transcripts. These results support the concept that the regulation of cytokine genes in AML cells is aberrant.

  7. Fluorescence In Situ Hybridization for MicroRNA Detection in Archived Oral Cancer Tissues

    Directory of Open Access Journals (Sweden)

    Zonggao Shi

    2012-01-01

    Full Text Available The noncoding RNA designated as microRNA (miRNA is a large group of small single-stranded regulatory RNA and has generated wide-spread interest in human disease studies. To facilitate delineating the role of microRNAs in cancer pathology, we sought to explore the feasibility of detecting microRNA expression in formalin-fixed paraffin-embedded (FFPE tissues. Using FFPE materials, we have compared fluorescent in situ hybridization (FISH procedures to detect miR-146a with (a different synthetic probes: regular custom DNA oligonucleotides versus locked nucleic acid (LNA incorporated DNA oligonucleotides; (b different reporters for the probes: biotin versus digoxigenin (DIG; (c different visualization: traditional versus tyramide signal amplification (TSA system; (d different blocking reagents for endogenous peroxidase. Finally, we performed miR-146a FISH on a commercially available oral cancer tissue microarray, which contains 40 cases of oral squamous cell carcinoma (OSCC and 10 cases of normal epithelia from the human oral cavity. A sample FISH protocol for detecting miR-146a is provided. In summary, we have established reliable in situ hybridization procedures for detecting the expression of microRNA in FFPE oral cancer tissues. This method is an important tool for studies on the involvement of microRNA in oral cancer pathology and may have potential prognostic or diagnostic value.

  8. Resolution-improved in situ DNA hybridization detection based on microwave photonic interrogation.

    Science.gov (United States)

    Cao, Yuan; Guo, Tuan; Wang, Xudong; Sun, Dandan; Ran, Yang; Feng, Xinhuan; Guan, Bai-ou

    2015-10-19

    In situ bio-sensing system based on microwave photonics filter (MPF) interrogation method with improved resolution is proposed and experimentally demonstrated. A microfiber Bragg grating (mFBG) is used as sensing probe for DNA hybridization detection. Different from the traditional wavelength monitoring technique, we use the frequency interrogation scheme for resolution-improved bio-sensing detection. Experimental results show that the frequency shift of MPF notch presents a linear response to the surrounding refractive index (SRI) change over the range of 1.33 to 1.38, with a SRI resolution up to 2.6 × 10(-5) RIU, which has been increased for almost two orders of magnitude compared with the traditional fundamental mode monitoring technique (~3.6 × 10(-3) RIU). Due to the high Q value (about 27), the whole process of DNA hybridization can be in situ monitored. The proposed MPF-based bio-sensing system provides a new interrogation method over the frequency domain with improved sensing resolution and rapid interrogation rate for biochemical and environmental measurement.

  9. Detection of human papillomavirus in oral warts using in situ hybridization

    Directory of Open Access Journals (Sweden)

    Suzana Orsini Machado de Sousa

    2008-01-01

    Full Text Available Objective: The human papillomavirus is a group of DNA epitheliotrophic viruses associated with the etiology of benign and malignant oral warts. More than 100 types have been identified and among them, 24 have been found into the oral cavity. The aim of this study was to analyze human papillomavirus prevalence and its subtypes in 50 oral warts, of which 20 were squamous papillomas, 17 condylomaacuminatum and 13 verruca vulgaris. Method: In situ hybridization was used with biotinylated DNA probes for wide-spectrum HPV and with specific probes for human papillomavirus 6/11, human papillomavirus 16/18 and human papillomavirus 31/33. Results: Human papillomavirus was present in ten (20% of the 50 oral wart cases, 03 (3/20 squamous papillomas, 05 (5/17 condyloma acuminatum and 02 (2/13 verruca vulgaris. Of these, 8 (16% were positive to the HPV probe 6/11 being 5 condyloma acuminatum, 1 squamous papilloma and 2 verruca vulgaris. Three cases (6% demonstrated positivity to the human papillomavirus probe 16/18, with 2 being cases of condyloma and the other a case of squamous papilloma. Of the six positive cases to the human papillomavirus probe 31/33, (12% 4 were condyloma acuminatum and 2 squamous papillomas. Conclusion: The human papillomavirus expression (20% found in this study was low, but within the average found in the literature. Nonetheless, in addition to in situ hybridization, other methods may be necessary for confirming the presence of human papillomavirus.

  10. Fluorescence in situ hybridization and molecular studies in infertile men with dysplasia of the fibrous sheath.

    Science.gov (United States)

    Baccetti, Baccio; Collodel, Giulia; Gambera, Laura; Moretti, Elena; Serafini, Francesca; Piomboni, Paola

    2005-07-01

    To perform fluorescence in situ hybridization (FISH) and molecular analysis in patients with the genetic sperm defect "dysplasia of the fibrous sheath" (DFS). Retrospective study. Regional Referral Center for Male Infertility, Siena, Italy. Twelve infertile patients with DFS sperm defects. Family history, lymphocytic karyotype, physical and hormonal assays, semen analysis. The DFS sperm phenotype was defined by light, fluorescent, and electron microscopy. Sperm chromosomal constitution was examined by FISH. Gene deletions were tested by polymerase chain reaction. The genetic sperm defect DFS was determined by transmission and scanning electron microscopy. Immunofluorescence staining of A-kinase anchoring protein 4 (AKAP4) showed a moderate and diffuse signal, revealing a disorganized and incompletely assembled fibrous sheath. In 11 of 12 DFS patients, polymerase chain reaction for detecting the presence of partial sequence of AKAP4/AKAP3 binding regions gave positive results. Fluorescence in situ hybridization was performed in decondensed sperm nuclei with probes for chromosomes 18, X, and Y. The mean disomy frequency of chromosome 18 was in the normal range, whereas the mean disomy frequencies of sex chromosomes and diploidies were twice those of controls. These results should be considered when DFS sperm are used in assisted reproductive technology, owing to the high risk of transmission of chromosomal unbalance and of DFS sperm defects to male offspring.

  11. Erythrovirus B19 infection in acquired immunodeficiency syndrome: screening by histopathology, immunohistochemistry, and in situ hybridization

    Directory of Open Access Journals (Sweden)

    Sérgio Setúbal

    2006-06-01

    Full Text Available Erythrovirus B19 infects erythrocytic progenitors, transiently interrupting erythropoiesis. In AIDS patients it causes chronic anemia amenable to treatment. We looked for evidences of B19 infection in stored bone marrow material from patients with acquired immunodeficiency syndrome. Histological sections were made from stored paraffin blocks from 33 autopsies (39 blocks and 35 biopsies (45 blocks, 30 patients performed from 1988 to 2002. They were examined after hematoxylin-eosin (HE staining, immunohistochemical (IHC, and in situ hybridization. HE revealed intra-nuclear inclusion bodies ("lantern cells" suggesting B19 infection in 19 sections corresponding to 19 of 63 patients examined with this test. Seven of 78 sections subjected to immunohistochemistry were positive, corresponding to 7 of 58 patients examined with this test. Fourteen sections corresponding to 13 of the 20 HE and/or IHC positive patients were subjected to in situ hybridization, with six positives results. Among the 13 patients subjected to the three techniques, only one gave unequivocal positive results in all and was considered a true positive. The frequency of B19 infection (1/63 patients in the material examined can be deemed low.

  12. In situ green synthesis of antimicrobial carboxymethyl chitosan-nanosilver hybrids with controlled silver release.

    Science.gov (United States)

    Huang, Siqi; Yu, Zhiming; Zhang, Yang; Qi, Chusheng; Zhang, Shifeng

    2017-01-01

    In order to fabricate antimicrobial carboxymethyl chitosan-nanosilver (CMC-Ag) hybrids with controlled silver release, this study demonstrated comparable formation via three synthetic protocols: 1) carboxymethyl chitosan (CMC) and glucose (adding glucose after AgNO 3 ), 2) CMC and glucose (adding glucose before AgNO 3 ), and 3) CMC only. Under principles of green chemistry, the synthesis was conducted in an aqueous medium exposed to microwave irradiation for 10 minutes with nontoxic chemicals. The structure and formation mechanisms of the three CMC-Ag hybrids were explored using X-ray diffraction, ultraviolet-visible spectroscopy, transmission electron microscopy, and Fourier-transform infrared analyses. Additionally, antimicrobial activity and in vitro silver release of the three synthesized hybrids were investigated in detail. The results revealed that a large number of stable, uniform, and small silver nanoparticles (AgNPs) were synthesized in situ on CMC chains via protocol 1. AgNPs were well dispersed with narrow size distribution in the range of 6-20 nm, with mean diameter only 12.22±2.57 nm. The addition of glucose resulted in greater AgNP synthesis. The order of addition of glucose and AgNO 3 significantly affected particle size and size distribution of AgNPs. Compared to CMC alone and commercially available AgNPs, the antimicrobial activities of three hybrids were significantly improved. Of the three hybrids, CMC-Ag1 synthesized via protocol 1 exhibited better antimicrobial activity than CMC-Ag2 and CMC-Ag3, and showed more effective inhibition of Staphylococcus aureus than Escherichia coli . Due to strong coordination and electrostatic interactions between CMC and silver and good steric protection provided by CMC, CMC-Ag1 displayed stable and continuous silver release and better performance in retaining silver for prolonged periods than CMC-Ag2 and CMC-Ag3.

  13. In situ green synthesis of antimicrobial carboxymethyl chitosan–nanosilver hybrids with controlled silver release

    Science.gov (United States)

    Huang, Siqi; Yu, Zhiming; Zhang, Yang; Qi, Chusheng; Zhang, Shifeng

    2017-01-01

    In order to fabricate antimicrobial carboxymethyl chitosan–nanosilver (CMC-Ag) hybrids with controlled silver release, this study demonstrated comparable formation via three synthetic protocols: 1) carboxymethyl chitosan (CMC) and glucose (adding glucose after AgNO3), 2) CMC and glucose (adding glucose before AgNO3), and 3) CMC only. Under principles of green chemistry, the synthesis was conducted in an aqueous medium exposed to microwave irradiation for 10 minutes with nontoxic chemicals. The structure and formation mechanisms of the three CMC-Ag hybrids were explored using X-ray diffraction, ultraviolet-visible spectroscopy, transmission electron microscopy, and Fourier-transform infrared analyses. Additionally, antimicrobial activity and in vitro silver release of the three synthesized hybrids were investigated in detail. The results revealed that a large number of stable, uniform, and small silver nanoparticles (AgNPs) were synthesized in situ on CMC chains via protocol 1. AgNPs were well dispersed with narrow size distribution in the range of 6–20 nm, with mean diameter only 12.22±2.57 nm. The addition of glucose resulted in greater AgNP synthesis. The order of addition of glucose and AgNO3 significantly affected particle size and size distribution of AgNPs. Compared to CMC alone and commercially available AgNPs, the antimicrobial activities of three hybrids were significantly improved. Of the three hybrids, CMC-Ag1 synthesized via protocol 1 exhibited better antimicrobial activity than CMC-Ag2 and CMC-Ag3, and showed more effective inhibition of Staphylococcus aureus than Escherichia coli. Due to strong coordination and electrostatic interactions between CMC and silver and good steric protection provided by CMC, CMC-Ag1 displayed stable and continuous silver release and better performance in retaining silver for prolonged periods than CMC-Ag2 and CMC-Ag3. PMID:28458539

  14. The effect of tissue decalcification on mRNA retention within bone for in-situ hybridization studies.

    Science.gov (United States)

    Walsh, L; Freemont, A J; Hoyland, J A

    1993-06-01

    Tissue decalcification is a routine part of the preparation of bone tissue for histological studies. Although in-situ hybridization has been employed to localize mRNA of collagenous and non-collagenous bone related proteins in skeletal tissue, little is known regarding the effects of decalcifying agents on mRNA retention within tissue. In this study in-situ hybridization using an oligonucleotide probe (i.e. a poly d(T) probe) to detect total messenger RNA has been employed to investigate the effects of the decalcifying agents nitric acid, formic acid and EDTA on mRNA retention compared to undeacalcified tissue. The results show that formalin fixation and EDTA decalcification preserve substantial amounts of mRNA within the tissue. In particular, this study illustrates that it is possible to perform in-situ hybridization on formalin fixed decalcified paraffin embedded tissue.

  15. Fluorescence in situ hybridization evaluation of chromosome deletion patterns in prostate cancer.

    Science.gov (United States)

    Huang, S F; Xiao, S; Renshaw, A A; Loughlin, K R; Hudson, T J; Fletcher, J A

    1996-11-01

    Various nonrandom chromosomal aberrations have been identified in prostate carcinoma. These aberrations include deletions of several chromosome regions, particularly the chromosome 8 short arm. Large-scale numerical aberrations, reflected in aberrant DNA ploidy, are also found in a minority of cases. However, it is unclear whether prostate carcinomas contain aberrations of certain chromosome regions that are deleted frequently in other common types of cancer. In this study, we performed dual-color fluorescence in situ hybridization on intact nuclei from touch preparations of 16 prostate cancers. Chromosome copy number was determined using pericentromeric probes, whereas potential chromosome arm deletions were evaluated using yeast artificial chromosome (YAC) and P1 probes. Two YAC probes targeted chromosome 8 short arm regions known to be deleted frequently in prostate cancer. Other YACs and P1s were for chromosome regions, including 1p22, 3p14, 6q21, 9p21, and 22q12, that are deletion targets in a variety of cancers although not extensively studied in prostate cancer. Hybridization efficiencies and signal intensities were excellent for both repeat sequence (alpha-satellite) and single, copy (YAC and P1) fluorescence in situ hybridization probes. Of 16 prostate cancers, 11 had clonal aberrations of 1 or more of the 13 chromosome regions evaluated, and 10 cases (62.5%) had 8p deletions, including 4 cases with 8p deletion in virtually all cells and aneuploidy in only a subset of those deleted cells. Deletions at 3p14, 6q21, and 22q12 were identified in 2, 1, and 1 case, respectively, and each of those cases had a similarly sized cell population with 8p deletion. These studies confirm 8p deletion in the majority of prostate carcinomas. 8p deletions appear to be early events in prostate tumorigenesis, often antedating aneuploidy. Fluorescence in situ hybridization strategies incorporating pericentromeric and single-copy regional chromosome probes offer a powerful and

  16. An Optimized Small Tissue Handling System for Immunohistochemistry and In Situ Hybridization.

    Directory of Open Access Journals (Sweden)

    Giovanni Anthony

    Full Text Available Recent development in 3D printing technology has opened an exciting possibility for manufacturing 3D devices on one's desktop. We used 3D modeling programs to design 3D models of a tissue-handling system and these models were "printed" in a stereolithography (SLA 3D printer to create precision histology devices that are particularly useful to handle multiple samples with small dimensions in parallel. Our system has been successfully tested for in situ hybridization of zebrafish embryos. Some of the notable features include: (1 A conveniently transferrable chamber with 6 mesh-bottomed wells, each of which can hold dozens of zebrafish embryos. This design allows up to 6 different samples to be treated per chamber. (2 Each chamber sits in a well of a standard 6-well tissue culture plate. Thus, up to 36 different samples can be processed in tandem using a single 6 well plate. (3 Precisely fitting lids prevent solution evaporation and condensation, even at high temperatures for an extended period of time: i.e., overnight riboprobe hybridization. (4 Flat bottom mesh maximizes the consistent treatment of individual tissue samples. (5 A magnet-based lifter was created to handle up to 6 chambers (= 36 samples in unison. (6 The largely transparent resin aids in convenient visual inspection both with eyes and using a stereomicroscope. (7 Surface engraved labeling enables an accurate tracking of different samples. (8 The dimension of wells and chambers minimizes the required amount of precious reagents. (9 Flexible parametric modeling enables an easy redesign of the 3D models to handle larger or more numerous samples. Precise dimensions of 3D models and demonstration of how we use our devices in whole mount in situ hybridization are presented. We also provide detailed information on the modeling software, 3D printing tips, as well as 3D files that can be used with any 3D printer.

  17. Spatial Exploration and Characterization of Endozoicomonas spp. Bacteria in Stylophora pistillata Using Fluorescence In Situ Hybridization

    KAUST Repository

    Alsheikh-­Hussain, Areej

    2011-12-12

    Studies of coral-­associated bacterial communities have repeatedly demonstrated that the microbial assemblages of the coral host are highly specific and complex. In particular, bacterial community surveys of scleractinian and soft corals from geographically diverse reefs continually uncover a high abundance of sequences affiliated with the Gammaproteobacteria genus Endozoicomonas. The role of these bacteria within the complex coral holobiont is currently unknown. In order to localize these cells and gain an understanding of their potential interactions within the coral, we developed a fluorescence in situ hybridization(FISH) approach for reef-­building coral tissues. Using a custom small-­subunit ribosomal RNA gene database, we developed two Endozoicomonas-­specific probes that cover almost all known coral-­associated Endozoicomonas sequences. Probe hybridization conditions were quantitatively evaluated against target and non-­target bacterial cultures using fluorescence microscopy. Using these experimentally tested conditions, probes were then hybridized to the branching coral Stylophora pistillata, obtained from the Red Sea, using whole mount and paraffin embedding techniques. This study allowed preliminary spatial exploration and characterization of Endozoicomonas in coral, which has provided insight into their functional role and interactions within the coral holobiont.

  18. A double labeling technique for performing immunocytochemistry and in situ hybridization in virus infected cell cultures and tissues

    International Nuclear Information System (INIS)

    Gendelman, H.E.; Moench, T.R.; Narayan, O.; Griffin, D.E.; Clements, J.E.

    1985-01-01

    This report describes a combined immunocytochemical and in situ hybridization procedure which allows visualization of cellular or viral antigens and viral RNA in the same cell. Cultures infected with visna or measles virus were fixed in periodate-lysine-paraformaldehyde-glutaraldehyde, stained by the avidin-biotin-peroxidase technique using antibodies to viral or cellular proteins and then incubated with radiolabeled specific DNA probes (in situ hybridization). This technique provides a new approach to the study of viral pathogenesis by: (1) identifying the types of cells which are infected in the host and (2) identifying points of blockade in the virus life cycle during persistent infections. (Auth.)

  19. Expression of proto-oncogenes in non-Hodgkin's lymphomas by in situ hybridization with biotinylated DNA probes

    International Nuclear Information System (INIS)

    Hamatani, Kiyohiro; Yoshida, Kuniko; Abe, Masumi; Shimaoka, Katsutaro; Shiku, Hiroshi; Akiyama, Mitoshi; Kondo, Hisayoshi.

    1989-11-01

    Expression of six proto-oncogenes (fos, myc, myb, Ki-ras, Ha-ras, and N-ras) in 43 cases of non-Hodgkin's lymphoma was analyzed by means of in situ hybridization. Biotinylated DNA probes of the six oncogenes and those of the immunoglobulin H-chain (IgH) gene and the T cell receptor β-chain (TCRβ) gene were used. The results of in situ hybridization performed under blind conditions by IgH and TCRβ gene probes were compatible with those of typing by cell surface markers. The nuclear protein-related proto-oncogenes, fos myc, and myb, were expressed in about 70 % - 80 % of all cases regardless of phenotypes, histology or histologic grade. On the contrary, genes of the ras family were expressed in fewer cases except for the Ki-ras gene which was more frequently expressed by cases of the T cell immunophenotype with a high malignancy grade. The results of dot hybridization with RNA extracted from some cases were compatible with those of in situ hybridization, further demonstrating the specificity of in situ hybridization. (author)

  20. Determining the origin of cells in tissue engineered skin substitutes: a pilot study employing in situ hybridization.

    Science.gov (United States)

    Weber, Andreas Daniel; Pontiggia, Luca; Biedermann, Thomas; Schiestl, Clemens; Meuli, Martin; Reichmann, Ernst

    2011-03-01

    Definitive and high-quality coverage of large and, in particular, massive skin defects remains a significant challenge in burn as well as plastic and reconstructive surgery because of donor site shortage. A novel and promising approach to overcome these problems is tissue engineering of skin. Clearly, before eventual clinical application, engineered skin substitutes of human origin must be grafted and then evaluated in animal models. For the various tests to be conducted it is indispensable to be able to identify human cells as such in culture and also to distinguish between graft and recipient tissue after transplantation. Here we describe a tool to identify human cells in vitro and in vivo. In situ hybridization allows for the detection and localization of specific DNA or RNA sequences in morphologically preserved cells in culture or tissue sections, respectively. We used digoxigenin-labeled DNA probes corresponding to human-specific Alu repeats in order to identify human keratinocytes grown in culture together with rat cells, and also to label split and full thickness skin grafts of human origin after transplantation on immuno-incompetent rats. Digoxigenin-labeled DNA probing resulted in an intensive nuclear staining of human cells, both in culture and after transplantation onto recipient animals, while recipient animal cells (rat cells) did not stain. In situ hybridization using primate-specific Alu probes reliably allows distinguishing between cells of human and non-human origin both in culture as well as in histological sections. This method is an essential tool for those preclinical experiments (performed on non-primate animals) that must be conducted before novel tissue engineered skin substitutes might be introduced into clinical practice.

  1. Subseabed Disposal Program In-Situ Heat Transfer Experiment (ISHTE)

    International Nuclear Information System (INIS)

    Percival, C.M.

    1983-05-01

    A heat transfer experiment is being developed in support of the Subseabed Disposal Program. The primary objectives of this experiment are: to provide information on the in situ response of seabed sediment to localized heating; to provide an opportunity to evaluate theoretical models of the response and to observe any unanticipated phenomena which may occur; and to develop and demonstrate the technology necessary to perform waste isolation oriented experiments on the seafloor at depths up to 6000 m. As presently envisaged, the heat transfer experiment will be conducted at a location in the central North Pacific though it could be performed anywhere that the ocean bottom is of the type deemed suitable for the disposal of nuclear waste material. The experiment will be conducted of the seafloor from a recoverable space-frame platform at a depth of approximately 6000 m. A 400-W isotopic heat source will be implanted in the illite sediment and the subsequent response of the sediment to the induced thermal field evaluated. After remote initiation of the experiment, a permanent record of the data obtained will be recorded on board the platform, with selected information transmitted to a surface vessel by acoustic telemetry. The experiment will be operational for one year, after which the entire platform will be recovered. Current plans call for the deployment of the experiment in 1986. Specific activities which will be pursued during the course of the experiment include: measurement of the thermal field; determination of the effective thermal conductivity of the sediment; measurement of pore pressure; evaluation of radionuclide migration processes; pore water sampling; sediment chemistry studies; sediment shear strength measurements; and coring operations in the immediate vicinity of the experiment for postexperiment analysis

  2. Fluorescence in situ hybridization karyotyping reveals the presence of two distinct genomes in the taxon Aegilops tauschii

    OpenAIRE

    Zhao, Laibin; Ning, Shunzong; Yi, Yingjin; Zhang, Lianquan; Yuan, Zhongwei; Wang, Jirui; Zheng, Youliang; Hao, Ming; Liu, Dengcai

    2018-01-01

    Background Aegilops tauschii is the donor of the bread wheat D genome. Based on spike morphology, the taxon has conventionally been subdivided into ssp. tauschii and ssp. strangulata. The present study was intended to address the poor match between this whole plant morphology-based subdivision and genetic relationships inferred from genotyping by fluorescence in situ hybridization karyotyping a set of 31 Ae. tauschii accessions. Results The distribution of sites hybridizing to the two probes ...

  3. In-situ preparation of NaA zeolite/chitosan porous hybrid beads for removal of ammonium from aqueous solution.

    Science.gov (United States)

    Yang, Kai; Zhang, Xiang; Chao, Cong; Zhang, Bing; Liu, Jindun

    2014-07-17

    Inorganic/organic hybrid materials play important roles in removal of contaminants from wastewater. Herein, we used the natural materials of halloysite and chitosan to prepare a new adsorbent of NaA zeolite/chitosan porous hybrid beads by in-situ hydrothermal synthesis method. SEM indicated that the porous hybrid beads were composed of 6-8 μm sized cubic NaA zeolite particles congregated together with chitosan. The adsorption behavior of NH4(+) from aqueous solution onto hybrid beads was investigated at different conditions. The Langmuir and Freundlich adsorption models were applied to describe the equilibrium isotherms. A maximum adsorption capacity of 47.62 mg/g at 298 K was achieved according to Langmuir model. The regenerated or reused experiments indicated that the adsorption capacity of the hybrid beads could maintain in 90% above after 10 successive adsorption-desorption cycles. The high adsorption and reusable ability implied potential application of the hybrid beads for removing NH4(+) pollutants from wastewater. Copyright © 2014 Elsevier Ltd. All rights reserved.

  4. Determination of HER2 gene amplification by chromogenic in situ hybridization (CISH) in archival breast carcinoma.

    Science.gov (United States)

    Zhao, Jianxin; Wu, Rina; Au, Alfred; Marquez, Abbey; Yu, Yibing; Shi, Zuorong

    2002-06-01

    To compare the efficacy of chromogenic in situ hybridization (CISH(TM)) with fluorescence in situ (FISH) hybridization and immunohistochemistry (IHC) in determination of the HER2 status in human breast cancer. HER2 gene amplification was determined on formalin-fixed paraffin-embedded (FFPE) sections of 62 invasive breast cancers by FISH and followed by CISH using a digoxigenin (DIG)-labeled HER2 DNA probe generated by Subtraction Probe Technology (SPT(TM)), and a biotin-labeled chromosome 17 centromeric (chr.17cen) probe. The sections were heat treated and enzyme digested. After in situ hybridization, the HER2 probe was detected with fluorescein (FITC)-anti-DIG for FISH, followed by peroxidase-anti-FITC and diaminobenzidine (DAB) for CISH. The chr.17cen probe was detected with peroxidase-streptavidin and DAB. For CISH application, HER2 gene copies or chromosome 17 centromeres and morphology of cells were easily visualized simultaneously with a 40x objective under bright-field microscope in hematoxylin-counterstained sections. IHC study of HER2 overexpression was performed on adjacent sections using a panel of three HER2 antibodies (TAB 250, CB11, A0485), and staining was scored according to the criteria specified in the HercepTest. HER2 gene amplification detected by CISH was visualized typically as large DAB-stained clusters or by many dots in the nucleus. FISH and CISH identified HER2 gene amplification in 19% of the tumors. Chromosome 17 polysomy was detected in 31% of the tumors. HER2 overexpression was demonstrated in 19% (TAB 250), 23% (CB11), and 36% (A0485) of the tumors. Complete concordance between the results of CISH with FISH, TAB 250, CB11, and A0485 was seen in 100%, 97%, 94%, and 84% of the cases, respectively. By permitting observation of morphology using a bright-field microscope, CISH is an accurate, practical, and economical approach to screen HER2 status in breast cancers. It is a useful methodology for confirming ambiguous IHC results.

  5. Lower hybrid experiments in the Petula tokamak

    International Nuclear Information System (INIS)

    Singh, C.M.; Briand, P.; Dupas, I.

    1978-01-01

    The study of lower hybrid waves on Petula Tokamak addresses itself to the following questions: wave excitation by phased waveguide arrays, technological problems encountered with high power microwave circuitry, and nonlinear processes induced by large amplitude pump waves. An r.f. generator was used to deliver a maximum power of 1.0 MW, 100 μs at 1.25 GHz to a two waveguide grill (Emax=4 kV/cm). The data reported here were taken in a deuterium plasma, n(e)=3.5x10 13 cm -3 , T(e) 500 eV, T(i) 200 eV and B=15 kG. The choice of the plasma parameters and the frequency of the pump was made so as to avoid the linear mode conversion layer in the plasma

  6. Combination of microautoradiography and fluorescence in situ hybridization for identification of microorganisms degrading xenobiotic contaminants.

    Science.gov (United States)

    Yang, Yanru; Zarda, Annatina; Zeyer, Josef

    2003-12-01

    One of the central topics in environmental bioremediation research is to identify microorganisms that are capable of degrading the contaminants of interest. Here we report application of combined microautoradiography (MAR) and fluorescence in situ hybridization (FISH). The method has previously been used in a number of systems; however, here we demonstrate its feasibility in studying the degradation of xenobiotic compounds. With a model system (coculture of Pseudomonas putida B2 and Sphingomonas stygia incubated with [14C] o-nitrophenol), combination of MAR and FISH was shown to be able to successfully identify the microorganisms degrading o-nitrophenol. Compared with the conventional techniques, MAR-FISH allows fast and accurate identification of the microorganisms involved in environmental contaminant degradation.

  7. Analysis of Single-cell Gene Transcription by RNA Fluorescent In Situ Hybridization (FISH)

    DEFF Research Database (Denmark)

    Ronander, Elena; Bengtsson, Dominique C; Joergensen, Louise

    2012-01-01

    Adhesion of Plasmodium falciparum infected erythrocytes (IE) to human endothelial receptors during malaria infections is mediated by expression of PfEMP1 protein variants encoded by the var genes. The haploid P. falciparum genome harbors approximately 60 different var genes of which only one has...... been believed to be transcribed per cell at a time during the blood stage of the infection. How such mutually exclusive regulation of var gene transcription is achieved is unclear, as is the identification of individual var genes or sub-groups of var genes associated with different receptors...... fluorescent in situ hybridization (FISH) analysis of var gene transcription by the parasite in individual nuclei of P. falciparum IE(1). Here, we present a detailed protocol for carrying out the RNA-FISH methodology for analysis of var gene transcription in single-nuclei of P. falciparum infected human...

  8. Identification of supernumerary ring chromosome 1 mosaicism using fluorescence in situ hybridization.

    Science.gov (United States)

    Chen, H; Tuck-Muller, C M; Batista, D A; Wertelecki, W

    1995-03-27

    We report on a 15-year-old black boy with severe mental retardation, multiple congenital anomalies, and a supernumerary ring chromosome mosaicism. Fluorescence in situ hybridization with a chromosome 1 painting probe (pBS1) identified the ring as derived from chromosome 1. The karyotype was 46,XY/47,XY,+r(1)(p13q23). A review showed 8 reports of ring chromosome 1. In 5 cases, the patients had a non-supernumerary ring chromosome 1 resulting in partial monosomies of the short and/or long arm of chromosome 1. In 3 cases, the presence of a supernumerary ring resulted in partial trisomy of different segments of chromosome 1. In one of these cases the supernumerary ring was composed primarily of the centromere and the heterochromatic region of chromosome 1, resulting in normal phenotype. Our patient represents the third report of a supernumerary ring chromosome 1 resulting in abnormal phenotype.

  9. MicroRNA Detection by Whole-Mount In Situ Hybridization in C. elegans.

    Science.gov (United States)

    Andachi, Yoshiki; Kohara, Yuji

    2018-01-01

    MicroRNAs (miRNAs) loaded on argonaute proteins guide RNA-induced silencing complexes to target mRNAs. An excellent method to decipher the spatiotemporal expression patterns of miRNAs is whole-mount in situ hybridization (WISH), which has been successfully used in vertebrate embryos but still remains unavailable for many animal species. Here, we describe a WISH method for miRNA detection in Caenorhabditis elegans at both embryonic and post-embryonic stages. Strategies devised for detection include fixation of animals with carbodiimide at a high temperature and subsequent partial digestion of the fixed animals with an extremely high concentration of proteinase. WISH signals are visualized by staining with a chromogenic substrate or a fluorescent dye.

  10. Analysis of the Ceratitis capitata y chromosome using in situ hybridization to mitotic chromosomes

    International Nuclear Information System (INIS)

    Willhoeft, U.; Franz, G.

    1998-01-01

    In Ceratitis capitata the Y chromosome is responsible for sex-determination. We used fluorescence in situ hybridization (FISH) for cytogenetic analysis of mitotic chromosomes. FISH with the wild-type strain EgyptII and two repetitive DNA probes enabled us to differentiate between the short and the long arm of the Y chromosome and gives a much better resolution than C-banding of mitotic chromosomes. We identified the Y-chromosomal breakpoints in Y-autosome translocations using FISH. Even more complex rearrangements i.e. deletions and insertions in some translocation strains were detected by this method. A strategy for mapping the primary sex determination factor in Ceratitis capitata by FISH is presented. (author)

  11. Chromosome translocations measured by fluorescence in-situ hybridization: A promising biomarker

    International Nuclear Information System (INIS)

    Lucas, J.N.; Straume, T.

    1995-10-01

    A biomarker for exposure and risk assessment would be most useful if it employs an endpoint that is highly quantitative, is stable with time, and is relevant to human risk. Recent advances in chromosome staining using fluorescence in situ hybridization (FISH) facilitate fast and reliable measurement of reciprocal translocations, a kind of DNA damage linked to both prior exposure and risk. In contrast to other biomarkers available, the frequency of reciprocal translocations in individuals exposed to whole-body radiation is stable with time post exposure, has a rather small inter-individual variability, and can be measured accurately at the low levels. Here, the authors discuss results from their studies demonstrating that chromosome painting can be used to reconstruct radiation dose for workers exposed within the dose limits, for individuals exposed a long time ago, and even for those who have been diagnosed with leukemia but not yet undergone therapy

  12. Determination of the ruminant origin of bone particles using fluorescence in situ hybridization (FISH).

    Science.gov (United States)

    Lecrenier, M C; Ledoux, Q; Berben, G; Fumière, O; Saegerman, C; Baeten, V; Veys, P

    2014-07-17

    Molecular biology techniques such as PCR constitute powerful tools for the determination of the taxonomic origin of bones. DNA degradation and contamination by exogenous DNA, however, jeopardise bone identification. Despite the vast array of techniques used to decontaminate bone fragments, the isolation and determination of bone DNA content are still problematic. Within the framework of the eradication of transmissible spongiform encephalopathies (including BSE, commonly known as "mad cow disease"), a fluorescence in situ hybridization (FISH) protocol was developed. Results from the described study showed that this method can be applied directly to bones without a demineralisation step and that it allows the identification of bovine and ruminant bones even after severe processing. The results also showed that the method is independent of exogenous contamination and that it is therefore entirely appropriate for this application.

  13. Diagnostic and Prognostic Utility of Fluorescence In situ Hybridization (FISH) Analysis in Acute Myeloid Leukemia.

    Science.gov (United States)

    Gonzales, Patrick R; Mikhail, Fady M

    2017-12-01

    Acute myeloid leukemia (AML) is a hematologic neoplasia consisting of incompletely differentiated hematopoietic cells of the myeloid lineage that proliferate in the bone marrow, blood, and/or other tissues. Clinical implementation of fluorescence in situ hybridization (FISH) in cytogenetic laboratories allows for high-resolution analysis of recurrent structural chromosomal rearrangements specific to AML, especially in AML with normal karyotypes, which comprises approximately 33-50% of AML-positive specimens. Here, we review the use of several FISH probe strategies in the diagnosis of AML. We also review the standards and guidelines currently in place for use by clinical cytogenetic laboratories in the evaluation of AML. Updated standards and guidelines from the WHO, ACMG, and NCCN have further defined clinically significant, recurring cytogenetic anomalies in AML that are detectable by FISH. FISH continues to be a powerful technique in the diagnosis of AML, with higher resolution than conventional cytogenetic analysis, rapid turnaround time, and a considerable diagnostic and prognostic utility.

  14. Detection of TTV-DNA in PBMC using digoxigenin labelled probe by in situ hybridization

    International Nuclear Information System (INIS)

    Liu Yang; Qi Qige

    2002-01-01

    To determine TTV-DNA in PBMC in patients with viral hepatitis, a study of in situ hybridization using digoxigenin labelled probe by PCR method to the TTV ORF1 region was performed on PBMC. Results showed that the detection rate of TTV-DNA using double-stranded probe in TTV-DNA positive group in sera was 58.06 (18/31), and the detection rate of TTV-DNA using double-stranded probe in TTV-DNA negative group in sera was 27.59 (8/29). For TTV-DNA positive group detected by double- stranded probe, then we use negative- stranded probe to detect their replication. The detection rate was 22.2%(4/18). Conclusions: TTV can infect PBMC and replicate in PBMC

  15. Detection of Bacteria by Fluorescence in Situ Hybridization in Culture-Negative Soft Tissue Filler Lesions

    DEFF Research Database (Denmark)

    Bjarnsholt, Thomas; Tolker-Nielsen, Tim; Givskov, Michael

    2009-01-01

    BACKGROUND Adverse reactions to polyacrylamide gel occur as swellings or nodules, and controversy exists whether these are due to bacterial infection or an autoimmune reaction to the filler. OBJECTIVES Biopsies from culture-negative long-lasting nodules after injection with different types...... of polyacrylamide gel were examined with a combination of Gram stain and fluorescence in situ hybridization. RESULTS Bacteria were detected in biopsies from seven of eight patients. They inhabited gel and intervening tissue and tended to lie in aggregates. CONCLUSION This study supports the assumption...... that infection with bacteria in aggregates causes culture-negative late adverse reactions to polyacrylamide gel, suggesting a biofilm environment. The authors have indicated no significant interest with commercial supporters....

  16. SUPERSENSITIVE IN SITU HYBRIDIZATION BY TYRAMIDE SIGNAL AMPLIFICATION AND NANOGOLD SILVER STAINING: THE CONTRIBUTION OF AUTOMETALLOGRAPHY AND CATALYZED REPORTER DEPOSITION TO THE REJUVENATION OF IN SITU HYBRIDIZATION.

    Energy Technology Data Exchange (ETDEWEB)

    TUBBS,R.R.PETTAY,J.GROGAN,T.CHEUNG,A.L.M.POWELL,R.D.HAINFELD,J.HAUSER-KRONBERGER,C.HACKER,G.W.

    2002-04-17

    It is peculiar that in situ hybridization (ISH), a technique with many similarities to immunohistochemistry (IHC), has not enjoyed the phenomenal growth in both basic research and clinical applications as has its sister technique IHC. Since the late 1970s, when immunoperoxidase techniques began to be applied to routine diagnostic material and to numerous research applications, there has been a natural evolution of the IHC procedure. Namely, only a few primary antibodies were available commercially at the onset, and only one indirect and the peroxidase-antiperoxidase (PAP) technique detection systems were in place. With the advent of avidin-biotin detection systems and monoclonal antibodies, and a viable commercial market, extraordinary growth of the procedure's applications in clinical research and diagnostic pathology occurred during the subsequent two decades. Today, IHC is automated and widely used for research purposes and, to a large extent, has become a routine diagnostic ''special stain'' in most clinical laboratories. During the same period, ISH enjoyed very little growth in both research and diagnostic applications. What has accounted for this lack of maturation of the technique? The success of IHC is part of the reason measuring a gene's encoded protein routinely and inexpensively, particularly as automation evolved, rendered IHC a more viable choice in many instances. Inherent comparative sensitivity of the procedures has also clearly been a factor. Unfortunately, the chromogenic procedures in place are often insufficiently sensitive to detect the relatively low amounts of DNA and RNA levels at which the clinical utility is to be found.

  17. Preimplantation genetic diagnosis by fluorescence in situ hybridization of reciprocal and Robertsonian translocations.

    Science.gov (United States)

    Chen, Chun-Kai; Wu, Dennis; Yu, Hsing-Tse; Lin, Chieh-Yu; Wang, Mei-Li; Yeh, Hsin-Yi; Huang, Hong-Yuan; Wang, Hsin-Shin; Soong, Yung-Kuei; Lee, Chyi-Long

    2014-03-01

    The presence of reciprocal and Robertsonian chromosomal rearrangement is often related to recurrent miscarriage. Using preimplantation genetic diagnosis, the abortion rate can be decreased. Cases treated at our center were reviewed. A retrospective analysis for either Robertsonian or reciprocal translocations was performed on all completed cycles of preimplantation genetic diagnosis at our center since the first reported case in 2004 until the end of 2010. Day 3 embryo biopsies were carried out, and the biopsied cell was checked by fluorescent in situ hybridization using relevant informative probes. Embryos with a normal or balanced translocation karyotype were transferred on Day 4. Thirty-eight preimplantation genetic diagnosis cycles involving 17 couples were completed. A total of 450 (82.6%) of the total oocytes were MII oocytes, and 158 (60.0%) of the two-pronuclei embryos were biopsied. In 41.4% of the fluorescent in situ hybridization analyses, the results were either normal or balanced. Embryos were transferred back after 21 cycles. Three babies were born from Robertsonian translocation carriers and another two from reciprocal translocation carriers. The miscarriage rate was 0%. Among the reciprocal translocation group, the live delivery rate was 8.3% per ovum pick-up cycle and 18.2% per embryo transfer cycle. Among the Robertsonian translocation group, the live delivery rate was 14.3% per ovum pick-up cycle and 20.0% per embryo transfer cycle. There is a trend whereby the outcome for Robertsonian translocation group carriers is better than that for reciprocal translocation group carriers. Aneuploidy screening may possibly be added in order to improve the outcome, especially for individuals with an advanced maternal age. The emergence of an array-based technology should help improve this type of analysis. Copyright © 2014. Published by Elsevier B.V.

  18. Partial trisomy 13 in an infant with a mild phenotype: application of fluorescence in situ hybridization in cytogenetic syndromes.

    Science.gov (United States)

    Begovic, D; Hitrec, V; Lasan, R; Letica, L; Baric, I; Sarnavka, V; Galic, S

    1998-06-01

    We report on a month-old infant with dysmorphic face and several anomalies known to be associated with trisomy 13. Fluorescence in situ hybridization (FISH) studies performed on metaphase cells allowed us to identify an extra material on the short arm of the chromosome 13 as a duplication of 13q22-qter.

  19. HER-2 protein concentrations in breast cancer cells increase before immunohistochemical and fluorescence in situ hybridization analysis turn positive

    DEFF Research Database (Denmark)

    Olsen, Dorte A; Østergaard, Birthe; Bokmand, Susanne

    2007-01-01

    BACKGROUND: The level of HER-2/neu in breast cancer cells is normally measured by immunohistochemistry (IHC) and/or fluorescence in situ hybridization (FISH). It determines whether patients should be treated with trastuzumab (Herceptin). In this study, HER-2 protein in breast cancer tissue...

  20. Detection of genetic changes in Barrett's adenocarcinoma and Barrett's esophagus by DNA in situ hybridization and immunohistochemistry

    NARCIS (Netherlands)

    Krishnadath, K. K.; Tilanus, H. W.; Alers, J. C.; Mulder, A. H.; van Dekken, H.

    1994-01-01

    We have investigated the occurrence of chromosomal DNA and cell cycle-related protein changes in Barrett's epithelium and adenocarcinoma. The presence of numerical chromosomal aberrations was studied by applying nonisotopic in situ hybridization (ISH) with (peri-)centromeric DNA probes, specific for

  1. Expression and localization of ionotropic glutamate receptor subunits in the goldfish retina--an in situ hybridization and immunocytochemical study

    NARCIS (Netherlands)

    Vandenbranden, C. A.; Kamphuis, W.; Nunes Cardozo, B.; Kamermans, M.

    2000-01-01

    The expression and distribution of AMPA, kainate and NMDA glutamate receptor subunits was studied in the goldfish retina. For the immunocytochemical localization of the AMPA receptor antisera against GluR2, GluR2/3 and GluR4 were used, and for in situ hybridization rat specific probes for GluR1 and

  2. Preparation of epoxy/zirconia hybrid materials via in situ polymerization using zirconium alkoxide coordinated with acid anhydride

    International Nuclear Information System (INIS)

    Ochi, Mitsukazu; Nii, Daisuke; Harada, Miyuki

    2011-01-01

    Highlights: → Novel epoxy/zirconia hybrid materials were synthesized via in situ polymerization using zirconium alkoxide coordinated with acid anhydride. → The half-ester compound of acid anhydride desorbed from zirconium played as curing agent of epoxy resin. → The zirconia was uniformly dispersed in the epoxy matrix on the nanometer or sub-nanometer scale by synchronizing the epoxy curing and sol-gel reactions. → The refractive indices of the hybrid materials significantly improved with an increase in the zirconia content. - Abstract: Novel epoxy/zirconia hybrid materials were synthesized using a bisphenol A epoxy resin (diglycidyl ether of bisphenol A; DGEBA), zirconium(IV)-n-propoxide (ZTNP), and hexahydrophthalic anhydride (HHPA) via in situ polymerization. HHPA played two roles in this system: it acted as a modifier to control the hydrolysis and condensation reactions of zirconium alkoxide and also as a curing agent - the half-ester compound of HHPA desorbed from zirconium reacted with the epoxy resin to form the epoxy network. As a result, both the sol-gel reaction and epoxy curing occurred simultaneously in a homogeneous solution, and organic-inorganic hybrid materials were readily obtained. Further, the zirconia produced by the in situ polymerization was uniformly dispersed in the epoxy matrix on the nanometer or sub-nanometer scale; thus, hybrid materials that exhibited excellent optical transparency were obtained. Furthermore, the heat resistance of the hybrid materials could be improved by hybridization with zirconia. And, the refractive indices of the hybrid materials significantly improved with an increase in the zirconia content.

  3. Multicolor fluorescent in situ hybridization to define abutting and overlapping gene expression in the embryonic zebrafish brain

    Directory of Open Access Journals (Sweden)

    Hauptmann Giselbert

    2011-04-01

    Full Text Available Abstract Background In recent years, mapping of overlapping and abutting regulatory gene expression domains by chromogenic two-color in situ hybridization has helped define molecular subdivisions of the developing vertebrate brain and shed light on its basic organization. Despite the benefits of this technique, visualization of overlapping transcript distributions by differently colored precipitates remains difficult because of masking of lighter signals by darker color precipitates and lack of three-dimensional visualization properties. Fluorescent detection of transcript distributions may be able to solve these issues. However, despite the use of signal amplification systems for increasing sensitivity, fluorescent detection in whole-mounts suffers from rapid quenching of peroxidase (POD activity compared to alkaline phosphatase chromogenic reactions. Thus, less strongly expressed genes cannot be efficiently detected. Results We developed an optimized procedure for fluorescent detection of transcript distribution in whole-mount zebrafish embryos using tyramide signal amplification (TSA. Conditions for hybridization and POD-TSA reaction were optimized by the application of the viscosity-increasing polymer dextran sulfate and the use of the substituted phenol compounds 4-iodophenol and vanillin as enhancers of POD activity. In combination with highly effective bench-made tyramide substrates, these improvements resulted in dramatically increased signal-to-noise ratios. The strongly enhanced signal intensities permitted fluorescent visualization of less abundant transcripts of tissue-specific regulatory genes. When performing multicolor fluorescent in situ hybridization (FISH experiments, the highly sensitive POD reaction conditions required effective POD inactivation after each detection cycle by glycine-hydrochloric acid treatment. This optimized FISH procedure permitted the simultaneous fluorescent visualization of up to three unique transcripts

  4. Homology of polytene elements between Drosophila and Zaprionus determined by in situ hybridization in Zaprionus indianus.

    Science.gov (United States)

    Campos, S R C; Rieger, T T; Santos, J F

    2007-05-09

    The drosophilid Zaprionus indianus due to its economical importance as an insect pest in Brazil deserves more investigation into its genetics. Its mitotic karyotype and a line-drawing map of its polytene chromosomes are already available. This paper presents a photomap of Z. indianus polytene chromosomes, which was used as the reference map for identification of sections marked by in situ hybridization with gene probes. Hybridization signals for Hsp70 and Hsr-omega were detected, respectively, in sections 34B and 32C of chromosome V of Z. indianus, which indicates its homology to the chromosomal arm 3R of Drosophila melanogaster and, therefore, to Muller's element E. The main signal for Hsp83 gene probe hybridization was in section 17C of Z. indianus chromosome III, suggesting its homology to arm 3L of D. melanogaster and to element D of Muller. The Ubi probe hybridized in sections 10C of chromosome II and 17A of chromosome III. Probably the 17A is the polyubiquitin locus, with homology to arm 3L of D. melanogaster and to the mullerian D element, as suggested also by Hsp83 gene location. The Br-C gene was mapped in section 1D, near the tip of the X chromosome, indicating its homology to the X chromosome of D. melanogaster and to mullerian element A. The Dpp gene probe hybridized mainly in the section 32A of chromosome V and, at lower frequencies to other sections, although no signal was observed as expected in the correspondent mullerian B element. This result led to the suggestion of a rearrangement including the Dpp locus in Z. indianus, the secondary signals possibly pointing to related genes of the TGF-beta family. In conclusion, the results indicate that chromosomes X, III, V of Z. indianus are respectively correspondents to elements A, D, and E of Muller. At least chromosome V of Z. indianus seems to share synteny with the 3R arm of D. melanogaster, as indicated by the relative positions of Hsp70 and Hsr-omega, although the Dpp gene indicates a disruption of

  5. Fluorescence in situ hybridization detection of cytogenetic abnormalities and prognosis in multiple myeloma

    Directory of Open Access Journals (Sweden)

    Zhou Xu

    2015-01-01

    Full Text Available We evaluated the prognosis of patients with newly diagnosed multiple myeloma (MM and attempted to find a suitable treatment strategy for them. Interphase fluorescence in situ hybridization (FISH detection was performed on 57 patients with MM. The following probes: IgH, p53, 1q21, RB1, and D13S319 specific for the rearrangements of 14q32, 17p13, 1q21 and 13q14 were used. Fluorescent hybridization signals were observed using an Olympus BX60 epifluorescence microscope equipped with filters for detecting fluoroisothiocyanate (FITC, Texas red, and 4'-6-Diamidino-2-phenylindole (DAPI. Triple color clone-specific images were captured using a Quips XL genetic workstation. The mortalities in patients with moderate prognosis (66.7% and poor prognosis (50% were significantly higher compared with that in patients with good prognosis (15%, P<0.05. All the patients in good and moderate prognosis groups achieved complete remission (CR/very good partial remission (VGPR/partial remission (PR, whereas only half of the cases in the poor prognosis group reached this level. Patients 2 supported by autologous hematopoietic stem-cell transplantation presented CR/PR and long survival. For those with poor prognosis, a proper therapeutic regimen and timely transplantation, especially tandem transplantation, was necessary due to the rapid progression and complications.

  6. Thermoelectric properties of graphene nanosheets-modified polyaniline hybrid nanocomposites by an in situ chemical polymerization

    International Nuclear Information System (INIS)

    Lu, Yan; Song, Ying; Wang, Fuping

    2013-01-01

    A hybrid material of polyaniline protonated with hydrochloric acid and conductive graphene nanosheets (PANi/GNs) has been prepared by an in situ chemical polymerization method. The interactions between PANi and GNs in the hybrid composites are investigated by utilizing XRD, FT-IR, UV–vis and Raman. It is found that the PANi are adsorbed on the surface of the GNs, and the morphology of PANi transforms from twist structure to extended structure after the GNs are introduced. The thermoelectric (TE) properties of PANi/GNs composites have been investigated in the range from 323 K to 453 K. The electrical conductivity and the Seebeck coefficient of PANi/GNs composites are obviously higher than those of the PANi, while the thermal conductivity of the composites still keeps relatively low values even with high GNs content, resulting in the increase in dimensionless figure of merit (ZT). A highest ZT value of 1.95 × 10 −3 has been obtained for the composite containing 30 wt % GNs at 453 K, which is about 70 times higher than that obtained from the PANi. - Highlights: ► PANi adsorbed on the surface of the GNs possesses more extended structure. ► Electrical conductivity and Seebeck coefficient of PANi/GNs composites are superior to those of PANi. ► Thermal conductivity of the composites still keeps relatively low values

  7. In Situ Hybridization of Pulp Fibers Using Mg-Al Layered Double Hydroxides

    Directory of Open Access Journals (Sweden)

    Carl-Erik Lange

    2015-04-01

    Full Text Available Inorganic Mg2+ and Al3+ containing layered double hydroxide (LDH particles were synthesised in situ from aqueous solution onto chemical pulp fibers of pine (Pinus sylvestris. High super saturated (hss solution with sodium carbonate produced LDH particles with an average diameter of 100–200 nm. Nano-size (70 nm LDH particles were found from fibers external surface and, to a lesser degree, from the S2 cell wall after synthesis via low super saturated (lss route. The synthesis via slow urea hydrolysis (Uhyd yielded micron and clay sized LDH (2–5 μm and enabled efficient fiber densification via mineralization of S2 fiber wall layer as indicated by TEM and compliance analysis. The Uhyd method decreased fiber compliance up to 50%. Reduction in the polymerisation degree of cellulose was observed with capillary viscometry. Thermogravimetric analysis showed that the hybridization with LDH reduced the exothermic heat, indicating, that this material can be incorporated in flame retardant applications. Fiber charge was assessed by Fibers 2015, 3 104 adsorption expermients with methylene blue (MB and metanil yellow (MY. Synthesis via lss route retained most of the fibres original charge and provided the highest capacity (10 μmol/g for anionic MY, indicating cationic character of hybrid fibers. Our results suggested that mineralized fibers can be potentially used in advanced applications such as biocomposites and adsorbent materials.

  8. Localization of the Norrie disease gene mRNA by in situ hybridization.

    Science.gov (United States)

    Hartzer, M K; Cheng, M; Liu, X; Shastry, B S

    1999-07-15

    Norrie disease is a rare X-linked recessive neurodevelopmental disorder. The affected males manifest congenital blindness, which is often associated with hearing loss, mental retardation and psychiatric problems. Genetic linkage studies have localized the gene to the short arm of the X-chromosome and the gene has been isolated recently. The encoded protein is a member of the superfamily of growth factors containing a cystine knot motif and may be involved in cell adhesion and neurodevelopment. Molecular genetic analysis revealed a large number of missense, nonsense, deletion, and splice-site mutations among Norrie patients. In order to further determine the role of the Norrie disease gene, we studied the distribution pattern of its mRNA in the retina and in brain by in situ hybridization. The results show abundant hybridization signals in outer nuclear, inner nuclear, and ganglion cell layers of the retina in all three species (mice, rabbit, and human) examined. There was no significant expression in the vitreous body, lens, and rod outer segment. High expression levels were also observed in the cerebellar granular layer, hippocampus, olfactory bulb, cortex, and epithelium of the rabbit brain. These data suggest that the Norrie disease gene could play a critical role in the differentiation or maintenance of the differentiated state of the retina.

  9. In situ DNA-RNA hybridization using in vitro 125I-labeled ribosomal RNA of higher plant

    International Nuclear Information System (INIS)

    Sato, Seiichi; Kikuchi, Tadatoshi; Ishida, M.R.; Tanaka, Ryuso.

    1975-01-01

    In situ hybridization using 125 I-labeled ribosomal RNA was applied to plant cells. Cytoplasmic 25 s rRNA, which was eluted from acrylamide gels after electrophoretic separation, was labeled in vitro with carrier-free 125 I and hybridized with the interphase nuclei in root tips of Vicia faba. In most of the preparations, the nucleoli were more heavily labeled than the other regions within nuclei, and several types of grain distribution were observed on the nucleoli. From these results, it was confirmed that in situ hybridization using 125 I-labeled rRNA can be used very effectively to detect the annealing sites of different molecular species of rRNA within the nuclei of plant cells, for which it is not as easy to obtain high specific radioactive rRNA in vivo as it is in the case of cultured animal cells. (auth.)

  10. Intracellular distribution of histone mRNAs in human fibroblasts studied by in situ hybridization

    International Nuclear Information System (INIS)

    Lawrence, J.B.; Singer, R.H.; Villnave, C.A.; Stein, J.L.; Stein, G.S.

    1988-01-01

    We have used in situ hybridization to study the intracellular distribution of mRNAs for cell cycle-dependent core and H1 histone proteins in human WI-38 fibroblasts. Because histones are abundant nuclear proteins and histone mRNA expression is tightly coupled to DNA synthesis, it was of interest to determine whether histone mRNAs are localized near the nucleus. Cells were hybridized with tritiated DNA probes specific for either histone H1, histone H4, actin, or poly(A)+ mRNA and were processed for autoradiography. In exponentially growing cultures, the fraction of histone mRNA-positive cells correlated well with the fraction of cells in S phase and was eliminated by hydroxyurea inhibition of DNA synthesis. Within individual cells the label for histone mRNA was widely distributed throughout the cytoplasm and did not appear to be more heavily concentrated near the nucleus. However, histone mRNA appeared to exhibit patchy, nonhomogeneous localization, and a quantitative evaluation confirmed that grain distributions were not as uniform as they were after hybridizations to poly(A)+ mRNA. Actin mRNA in WI-38 cells was also widely distributed throughout the cytoplasm but differed from histone mRNA in that label for actin mRNA was frequently most dense at the outermost region of narrow cell extensions. The localization of actin mRNA was less pronounced but qualitatively very similar to that previously described for chicken embryonic myoblasts and fibroblasts. We conclude that localization of histones in WI-38 cells is not facilitated by localization of histone protein synthesis near the nucleus and that there are subtle but discrete and potentially functional differences in the distributions of histone, actin, and poly(A)+ mRNAs

  11. The City's new hybrid experience spaces

    DEFF Research Database (Denmark)

    Andersson, Lasse; Kiib, Hans

    2007-01-01

    In a series of workshops in Danish cities during the last couple of years, we have been establishing new ways of working with a clear local perspective in the new global discourse on culture, creativity and urbanity - urban innovation and urban branding in what could be called the experience city...... serve as frameworks for traditional functions, while simultaneously taking on new roles, new meanings and new narratives. This article serves as the first tentative reflection on results from a workshop at the ‘Skanok 05'; a conference on the experience economy held in Aalborg, October 2005.In...

  12. A safer, urea-based in situ hybridization method improves detection of gene expression in diverse animal species.

    Science.gov (United States)

    Sinigaglia, Chiara; Thiel, Daniel; Hejnol, Andreas; Houliston, Evelyn; Leclère, Lucas

    2018-02-01

    In situ hybridization is a widely employed technique allowing spatial visualization of gene expression in fixed specimens. It has greatly advanced our understanding of biological processes, including developmental regulation. In situ protocols are today routinely followed in numerous laboratories, and although details might change, they all include a hybridization step, where specific antisense RNA or DNA probes anneal to the target nucleic acid sequence. This step is generally carried out at high temperatures and in a denaturing solution, called hybridization buffer, commonly containing 50% (v/v) formamide - a hazardous chemical. When applied to the soft-bodied hydrozoan medusa Clytia hemisphaerica, we found that this traditional hybridization approach was not fully satisfactory, causing extensive deterioration of morphology and tissue texture which compromised our observation and interpretation of results. We thus tested alternative solutions for in situ detection of gene expression and, inspired by optimized protocols for Northern and Southern blot analysis, we substituted the 50% formamide with an equal volume of 8M urea solution in the hybridization buffer. Our new protocol not only yielded better morphologies and tissue consistency, but also notably improved the resolution of the signal, allowing more precise localization of gene expression and reducing aspecific staining associated with problematic areas. Given the improved results and reduced manipulation risks, we tested the urea protocol on other metazoans, two brachiopod species (Novocrania anomala and Terebratalia transversa) and the priapulid worm Priapulus caudatus, obtaining a similar reduction of aspecific probe binding. Overall, substitution of formamide by urea during in situ hybridization offers a safer alternative, potentially of widespread use in research, medical and teaching contexts. We encourage other workers to test this approach on their study organisms, and hope that they will also

  13. Hybrid aerogels and bioactive aerogels under uniaxial compression: an in situ SAXS study

    Directory of Open Access Journals (Sweden)

    Esquivias, L.

    2010-12-01

    Full Text Available The complex structure of hybrid organic/inorganic aerogels is composed by an inorganic phase covalently bonded to an organic chain forming a copolymer. Conventional hybrid aerogels were studied as well as bioactive hybrid aerogels, that is, aerogels with a calcium active phase added. In this work, the relationship between mechanical response and nanostructure was studied, using a specifically designed sample-holder for in situ uniaxial compression obtaining at the same time the small-angle X-ray pattern from synchrotron radiation (SAXS. Structural elements can be described as a particulated silica core surrounded by the organic chains. These chains are compressed on the direction parallel to the load, and a relationship between macroscopic uniaxial compression and particle and pore deformations can be established.

    La compleja estructura de los aerogeles híbridos orgánico/inorgánicos está compuesta por una fase inorgánica de sílice, unida mediante enlaces covalentes a una red de cadenas orgánicas. Se han estudiado composites híbridos convencionales y bioactivos, esto es, con una fase activa de calcio añadida. En este trabajo se ha investigado la relación entre la respuesta mecánica y la nanoestructura, con ayuda de un portamuestras específicamente diseñado para el estudio in situ de muestras bajo compresión uniaxial, a la vez que se obtiene el espectro de rayos-X a bajo-ángulo de radiación sincrotrón (SAXS. Los elementos estructurales se pueden describir como núcleos particulados de sílice rodeados de las cadenas orgánicas. Estas, se comprimen en la dirección paralela a la carga pudiéndose establecer una relación entre la compresión uniaxial macroscópica y la deformación de las partículas y poros que forman la estructura.

  14. Hybrid Experience Space for Cultural Heritage Communication

    DEFF Research Database (Denmark)

    Veirum, Niels Einar; Christensen, Mogens Fiil; Mayerhofer, Mikkel

    2006-01-01

    by daily use of experience products like computer-games, IMAX cinemas and theme parks featuring virtual reality installations. “It’s a question of stone-axe displays versus Disney-power installations” as one of the involved museum professionals point it, “but we don’t want any of these possibilities...

  15. Lower hybrid heating experiments in tokamaks: an overview

    International Nuclear Information System (INIS)

    Porkolab, M.

    1985-10-01

    Lower hybrid wave propagation theory relevant to heating fusion grade plasmas (tokamaks) is reviewed. A brief discussion of accessibility, absorption, and toroidal ray propagation is given. The main part of the paper reviews recent results in heating experiments on tokamaks. Both electron and ion heating regimes will be discussed. The prospects of heating to high temperatures in reactor grade plasmas will be evaluated

  16. Students' Positive and Negative Experiences in Hybrid and Online Classes

    Science.gov (United States)

    El Mansour, Bassou; Mupinga, Davison M.

    2007-01-01

    As higher education institutions struggle to meet the growing demand for education from non-traditional students, many are turning to hybrid and online courses. These courses, free up classroom space, allow faculty to reach a wider audience using technology; and are therefore cost effective. But, what learning experiences do these courses provide…

  17. Distribution of 45S rDNA in Modern Rose Cultivars (Rosa hybrida), Rosa rugosa, and Their Interspecific Hybrids Revealed by Fluorescence in situ Hybridization.

    Science.gov (United States)

    Ding, Xiao-Liu; Xu, Ting-Liang; Wang, Jing; Luo, Le; Yu, Chao; Dong, Gui-Min; Pan, Hui-Tang; Zhang, Qi-Xiang

    2016-01-01

    To elucidate the evolutionary dynamics of the location and number of rDNA loci in the process of polyploidization in the genus Rosa, we examined 45S rDNA sites in the chromosomes of 6 modern rose cultivars (R. hybrida), 5 R. rugosa cultivars, and 20 hybrid progenies by fluorescence in situ hybridization. Variation in the number of rDNA sites in parents and their interspecific hybrids was detected. As expected, 4 rDNA sites were observed in the genomes of 4 modern rose cultivars, while 3 hybridization sites were observed in the 2 others. Two expected rDNA sites were found in 2 R. rugosa cultivars, while in the other 3 R. rugosa cultivars 4 sites were present. Among the 20 R. hybrida × R. rugosa offspring, 13 carried the expected number of rDNA sites, and 1 had 6 hybridization sites, which exceeded the expected number by far. The other 6 offspring had either 2 or 3 hybridization sites, which was less than expected. Differences in the number of rDNA loci were observed in interspecific offspring, indicating that rDNA loci exhibit instability after distant hybridization events. Abnormal chromosome pairing may be the main factor explaining the variation in rDNA sites during polyploidization. © 2016 S. Karger AG, Basel.

  18. Efficient in situ growth of enzyme-inorganic hybrids on paper strips for the visual detection of glucose.

    Science.gov (United States)

    Li, WanYun; Lu, ShiYu; Bao, ShuJuan; Shi, ZhuanZhuan; Lu, Zhisong; Li, ChangMing; Yu, Ling

    2018-01-15

    A visual colorimetric microfluidic paper-based analytical device (μPAD) was constructed following the direct synthesis of enzyme-inorganic hybrid nanomaterials on the paper matrix. An inorganic solution of MnSO 4 and KH 2 PO 4 containing a diluted enzyme (glucose oxidase, GOx) was subsequently pipetted onto cellulose paper for the in situ growth of GOx@Mn 3 (PO 4 ) 2 hybrid functional materials. The characterization of the morphology and chemical composition validated the presence of hybrid materials roots in the paper fiber, while the Mn 3 (PO 4 ) 2 of the hybrid provided both a surface for enzyme anchoring and a higher peroxidase-like catalytic activity as compared to the Mn 3 (PO 4 ) 2 crystal that was synthesized without enzyme modulation. This new approach for the in situ growth of an enzyme-inorganic hybrid on a paper matrix eliminates centrifugation and the dry process by casting the solution on paper. The sensing material loading was highly reproducible because of the accuracy and stability of pipetting, which eventually contributed to the reliability of the μPAD. The self-assembled natural and artificial enzyme hybrid on the μPADs specifically detected glucose from a group of interferences, which shows great specificity using this method. Moreover, the colorimetric signal exhibited detection limitation for glucose is 0.01mM, which lies in the physiological range of glucose in biological samples. Copyright © 2017 Elsevier B.V. All rights reserved.

  19. Single molecule localization imaging of telomeres and centromeres using fluorescence in situ hybridization and semiconductor quantum dots.

    Science.gov (United States)

    Wang, Le; Zong, Shenfei; Wang, Zhuyuan; Lu, Ju; Chen, Chen; Zhang, Ruohu; Cui, Yiping

    2018-07-13

    Single molecule localization microscopy (SMLM) is a powerful tool for imaging biological targets at the nanoscale. In this report, we present SMLM imaging of telomeres and centromeres using fluorescence in situ hybridization (FISH). The FISH probes were fabricated by decorating CdSSe/ZnS quantum dots (QDs) with telomere or centromere complementary DNA strands. SMLM imaging experiments using commercially available peptide nucleic acid (PNA) probes labeled with organic fluorophores were also conducted to demonstrate the advantages of using QDs FISH probes. Compared with the PNA probes, the QDs probes have the following merits. First, the fluorescence blinking of QDs can be realized in aqueous solution or PBS buffer without thiol, which is a key buffer component for organic fluorophores' blinking. Second, fluorescence blinking of the QDs probe needs only one excitation light (i.e. 405 nm). While fluorescence blinking of the organic fluorophores usually requires two illumination lights, that is, the activation light (i.e. 405 nm) and the imaging light. Third, the high quantum yield, multiple switching times and a good optical stability make the QDs more suitable for long-term imaging. The localization precision achieved in telomeres and centromeres imaging experiments is about 30 nm, which is far beyond the diffraction limit. SMLM has enabled new insights into telomeres or centromeres on the molecular level, and it is even possible to determine the length of telomere and become a potential technique for telomere-related investigation.

  20. Hybrid Fictionality and Vicarious Narrative Experience

    DEFF Research Database (Denmark)

    Hatavara, Mari Annukka; Mildorf, Jarmila

    2017-01-01

    This article discusses the recent trends in Fictionality Studies and argues for a point of view focusing more on the narrative dimension of fictionality than on the fictive story content. With the analysis of two case studies, where a non-fictional third person narrator represents the experience...... with other minds travel between fictional and nonfictional narratives, and between stories artistically designed and those occurring in conversational or documentary environments....

  1. Identification of Cannabis sativa L. using the 1-kbTHCA synthase-fluorescence in situ hybridization probe.

    Science.gov (United States)

    Jeangkhwoa, Pattraporn; Bandhaya, Achirapa; Umpunjun, Puangpaka; Chuenboonngarm, Ngarmnij; Panvisavas, Nathinee

    2017-03-01

    This study reports a successful application of fluorescence in situ hybridization (FISH) technique in the identification of Cannabis sativa L. cells recovered from fresh and dried powdered plant materials. Two biotin-16-dUTP-labeled FISH probes were designed from the Cannabis-specific tetrahydrocannabinolic acid synthase (THCAS) gene and the ITS region of the 45S rRNA gene. Specificity of probe-target hybridization was tested against the target and 4 non-target plant species, i.e., Humulus lupulus, Mitragyna speciosa, Papaver sp., and Nicotiana tabacum. The 1-kb THCA synthase hybridization probe gave Cannabis-specific hybridization signals, unlike the 700-bp Cannabis-ITS hybridization probe. Probe-target hybridization was also confirmed against 20 individual Cannabis plant samples. The 1-kb THCA synthase and 700-bp Cannabis-ITS hybridization probes clearly showed 2 hybridization signals per cell with reproducibility. The 1-kb THCA synthase probe did not give any FISH signal when tested against H. lupulus, its closely related member of the Canabaceae family. It was also showed that 1-kb THCA synthase FISH probe can be applied to identify small amount of dried powdered Cannabis material with an addition of rehydration step prior to the experimental process. This study provided an alternative identification method for Cannabis trace. Copyright © 2016. Published by Elsevier B.V.

  2. Experiences from the Roadrunner petascale hybrid systems

    Energy Technology Data Exchange (ETDEWEB)

    Kerbyson, Darren J [Los Alamos National Laboratory; Pakin, Scott [Los Alamos National Laboratory; Lang, Mike [Los Alamos National Laboratory; Sancho Pitarch, Jose C [Los Alamos National Laboratory; Davis, Kei [Los Alamos National Laboratory; Barker, Kevin J [Los Alamos National Laboratory; Peraza, Josh [Los Alamos National Laboratory

    2010-01-01

    The combination of flexible microprocessors (AMD Opterons) with high-performing accelerators (IBM PowerXCell 8i) resulted in the extremely powerful Roadrunner system. Many challenges in both hardware and software were overcome to achieve its goals. In this talk we detail some of the experiences in achieving performance on the Roadrunner system. In particular we examine several implementations of the kernel application, Sweep3D, using a work-queue approach, a more portable Thread-building-blocks approach, and an MPI on the accelerator approach.

  3. Detection of human papillomavirus type 6/11 DNA in conjunctival papillomas by in situ hybridization with radioactive probes

    International Nuclear Information System (INIS)

    McDonnell, P.J.; McDonnell, J.M.; Kessis, T.; Green, W.R.; Shah, K.V.

    1987-01-01

    Twenty-three conjunctival papillomas and 28 conjunctival dysplasias were examined for human papillomavirus (HPV)-DNA sequences by in situ hybridization with nick-translated 35 S-labeled HPV probes. Adjacent paraffin sections were hybridized with HPV type 2, 6, 16, and 18 probes at Tm - 17 degrees C. Fifteen tissues, all papillomas, displayed positive hybridization with the HPV-6 probe. Infection with HPV-6 (or the closely related HPV-11) appeared to be responsible for most of the conjunctival papillomas of children and young adults. The presence of genital tract HPV-6 in these lesions suggests that some of the infections were acquired during passage through an infected birth canal. The lack of hybridization in adult conjunctival dysplasias indicates either that HPVs are not associated with this condition or that the probes and the technique utilized were not adequate for demonstration of this association

  4. PV-Diesel Hybrid SCADA Experiment Network Design

    Science.gov (United States)

    Kalu, Alex; Durand, S.; Emrich, Carol; Ventre, G.; Wilson, W.; Acosta, R.

    1999-01-01

    The essential features of an experimental network for renewable power system satellite based supervisory, control and data acquisition (SCADA) are communication links, controllers, diagnostic equipment and a hybrid power system. Required components for implementing the network consist of two satellite ground stations, to satellite modems, two 486 PCs, two telephone receivers, two telephone modems, two analog telephone lines, one digital telephone line, a hybrid-power system equipped with controller and a satellite spacecraft. In the technology verification experiment (TVE) conducted by Savannah State University and Florida Solar Energy Center, the renewable energy hybrid system is the Apex-1000 Mini-Hybrid which is equipped with NGC3188 for user interface and remote control and the NGC2010 for monitoring and basic control tasks. This power system is connected to a satellite modem via a smart interface, RS232. Commands are sent to the power system control unit through a control PC designed as PC1. PC1 is thus connected to a satellite model through RS232. A second PC, designated PC2, the diagnostic PC is connected to both satellite modems via separate analog telephone lines for checking modems'health. PC2 is also connected to PC1 via a telephone line. Due to the unavailability of a second ground station for the ACTS, one ground station is used to serve both the sending and receiving functions in this experiment. Signal is sent from the control PC to the Hybrid system at a frequency f(sub 1), different from f(sub 2), the signal from the hybrid system to the control PC. f(sub l) and f(sub 2) are sufficiently separated to avoid interference.

  5. FISHIS: fluorescence in situ hybridization in suspension and chromosome flow sorting made easy.

    Directory of Open Access Journals (Sweden)

    Debora Giorgi

    Full Text Available The large size and complex polyploid nature of many genomes has often hampered genomics development, as is the case for several plants of high agronomic value. Isolating single chromosomes or chromosome arms via flow sorting offers a clue to resolve such complexity by focusing sequencing to a discrete and self-consistent part of the whole genome. The occurrence of sufficient differences in the size and or base-pair composition of the individual chromosomes, which is uncommon in plants, is critical for the success of flow sorting. We overcome this limitation by developing a robust method for labeling isolated chromosomes, named Fluorescent In situ Hybridization In suspension (FISHIS. FISHIS employs fluorescently labeled synthetic repetitive DNA probes, which are hybridized, in a wash-less procedure, to chromosomes in suspension following DNA alkaline denaturation. All typical A, B and D genomes of wheat, as well as individual chromosomes from pasta (T. durum L. and bread (T. aestivum L. wheat, were flow-sorted, after FISHIS, at high purity. For the first time in eukaryotes, each individual chromosome of a diploid organism, Dasypyrum villosum (L. Candargy, was flow-sorted regardless of its size or base-pair related content. FISHIS-based chromosome sorting is a powerful and innovative flow cytogenetic tool which can develop new genomic resources from each plant species, where microsatellite DNA probes are available and high quality chromosome suspensions could be produced. The joining of FISHIS labeling and flow sorting with the Next Generation Sequencing methodology will enforce genomics for more species, and by this mightier chromosome approach it will be possible to increase our knowledge about structure, evolution and function of plant genome to be used for crop improvement. It is also anticipated that this technique could contribute to analyze and sort animal chromosomes with peculiar cytogenetic abnormalities, such as copy number variations

  6. FISHIS: fluorescence in situ hybridization in suspension and chromosome flow sorting made easy.

    Science.gov (United States)

    Giorgi, Debora; Farina, Anna; Grosso, Valentina; Gennaro, Andrea; Ceoloni, Carla; Lucretti, Sergio

    2013-01-01

    The large size and complex polyploid nature of many genomes has often hampered genomics development, as is the case for several plants of high agronomic value. Isolating single chromosomes or chromosome arms via flow sorting offers a clue to resolve such complexity by focusing sequencing to a discrete and self-consistent part of the whole genome. The occurrence of sufficient differences in the size and or base-pair composition of the individual chromosomes, which is uncommon in plants, is critical for the success of flow sorting. We overcome this limitation by developing a robust method for labeling isolated chromosomes, named Fluorescent In situ Hybridization In suspension (FISHIS). FISHIS employs fluorescently labeled synthetic repetitive DNA probes, which are hybridized, in a wash-less procedure, to chromosomes in suspension following DNA alkaline denaturation. All typical A, B and D genomes of wheat, as well as individual chromosomes from pasta (T. durum L.) and bread (T. aestivum L.) wheat, were flow-sorted, after FISHIS, at high purity. For the first time in eukaryotes, each individual chromosome of a diploid organism, Dasypyrum villosum (L.) Candargy, was flow-sorted regardless of its size or base-pair related content. FISHIS-based chromosome sorting is a powerful and innovative flow cytogenetic tool which can develop new genomic resources from each plant species, where microsatellite DNA probes are available and high quality chromosome suspensions could be produced. The joining of FISHIS labeling and flow sorting with the Next Generation Sequencing methodology will enforce genomics for more species, and by this mightier chromosome approach it will be possible to increase our knowledge about structure, evolution and function of plant genome to be used for crop improvement. It is also anticipated that this technique could contribute to analyze and sort animal chromosomes with peculiar cytogenetic abnormalities, such as copy number variations or cytogenetic

  7. Comparative cytogenetic characterization of primary canine melanocytic lesions using array CGH and fluorescence in situ hybridization.

    Science.gov (United States)

    Poorman, Kelsey; Borst, Luke; Moroff, Scott; Roy, Siddharth; Labelle, Philippe; Motsinger-Reif, Alison; Breen, Matthew

    2015-06-01

    Melanocytic lesions originating from the oral mucosa or cutaneous epithelium are common in the general dog population, with up to 100,000 diagnoses each year in the USA. Oral melanoma is the most frequent canine neoplasm of the oral cavity, exhibiting a highly aggressive course. Cutaneous melanocytomas occur frequently, but rarely develop into a malignant form. Despite the differential prognosis, it has been assumed that subtypes of melanocytic lesions represent the same disease. To address the relative paucity of information about their genomic status, molecular cytogenetic analysis was performed on the three recognized subtypes of canine melanocytic lesions. Using array comparative genomic hybridization (aCGH) analysis, highly aberrant distinct copy number status across the tumor genome for both of the malignant melanoma subtypes was revealed. The most frequent aberrations included gain of dog chromosome (CFA) 13 and 17 and loss of CFA 22. Melanocytomas possessed fewer genome wide aberrations, yet showed a recurrent gain of CFA 20q15.3-17. A distinctive copy number profile, evident only in oral melanomas, displayed a sigmoidal pattern of copy number loss followed immediately by a gain, around CFA 30q14. Moreover, when assessed by fluorescence in situ hybridization (FISH), copy number aberrations of targeted genes, such as gain of c-MYC (80 % of cases) and loss of CDKN2A (68 % of cases), were observed. This study suggests that in concordance with what is known for human melanomas, canine melanomas of the oral mucosa and cutaneous epithelium are discrete and initiated by different molecular pathways.

  8. Detection of Alien Oryza punctata Kotschy Chromosomes in Rice, Oryza sativa L., by Genomic in situ Hybridization

    OpenAIRE

    Yasui, Hideshi; Nonomura, Ken-ichi; Iwata, Nobuo; 安井, 秀; 野々村, 賢一; 岩田, 伸夫

    1997-01-01

    Genomic in situ hybridization (GIS H) using total Oryza punctata Kotschy genomic DNA as a probe was applied to detect alien chromosomes transferred from O. punctata (W1514: 2n=2x=24: BB) to O. sativa Japonica cultivar, Nipponbare (2n=2x=24: AA). Only 12 chromosomes in the interspecific hybrids (2n=3x=36: AAB) between autotetraploid of O. sativa cultivar Nipponbare and a diploid strain of O. punctata (W1514) showed intense staining by FITC in mitotic metaphase spreads. Only one homologous pair...

  9. Determination of HER2 amplification in primary breast cancer using dual-colour chromogenic in situ hybridization is comparable to fluorescence in situ hybridization: a European multicentre study involving 168 specimens

    Science.gov (United States)

    García-Caballero, Tomás; Grabau, Dorthe; Green, Andrew R; Gregory, John; Schad, Arno; Kohlwes, Elke; Ellis, Ian O; Watts, Sarah; Mollerup, Jens

    2010-01-01

    García-Caballero T, Grabau D, Green A R, Gregory J, Schad A, Kohlwes E, Ellis I O, Watts S & Mollerup J (2010) Histopathology56, 472–480 Determination of HER2 amplification in primary breast cancer using dual-colour chromogenic in situ hybridization is comparable to fluorescence in situ hybridization: a European multicentre study involving 168 specimens Aims: Fluorescence in situ hybridization (FISH) can be used to reveal several genomic imbalances relevant to proper cancer diagnosis and to the correct treatment regime. However, FISH requires expensive and advanced fluorescence microscopes in addition to expertise in fluorescence microscopy. To determine whether a newly developed dual-colour chromogenic in situ hybridization (CISH) method is a suitable alternative to FISH, we analysed the human epidermal growth factor receptor 2 gene (HER2) amplification level of 168 breast cancer specimens using dual-colour CISH and FISH and compared the results. Methods and results: We found 100% agreement between HER2 status determined by FISH and dual-colour CISH. Furthermore, we observed that the time used to score slides was significantly reduced by 28% in dual-colour CISH compared with the FISH protocol. Concordance between HER2 protein status and dual-colour CISH or FISH was equally good with an overall agreement of 96.8%. Correlation between the HER2/centromere 17 gene ratios obtained with dual-colour CISH and FISH was highly significant with an overall correlation coefficient (ρ) of 0.96. Conclusions: We conclude that dual-colour CISH and bright field microscopy are excellent alternatives to FISH when analysing the HER2 status of primary breast cancer. PMID:20459554

  10. High sensitive diagnostic technique for virus gene using radioisotope. Development of PCR in situ hybridization and its application

    Energy Technology Data Exchange (ETDEWEB)

    Iwasaki, Takuya; Sada, Tetsutaro; Terai, Masanori; Sato, Yuko; Kurata, Takeshi [National Inst. of Infectious Diseases, Tokyo (Japan); Yamaguchi, Kazuyoshi; Yanagisawa, Akio; Sakai, Yuzo

    1998-02-01

    An animal model, experimentally induced squamous cell carcinoma was produced in hamster mucosa to establish a carcinogenic system inducible by interaction of a virus and chemical(s). Human hydatid mole and cholioepithelioma were chosen as the target subjects. Several tumor cells and host cells were isolated under microscopy and DNA was extracted from these cells to indentify the respective origins (maternal, paternal or host origin). The base sequence of HLADRB region was analyzed by PCR using synthetic primer and the tissue localization was examined by PCR in situ hybridization. A PCR product of 82 bp was found in 15 of 17 samples and there were 2 samples in which the product was not detectable with the constructed primer and 6 samples were negative. While significant differences were not observed by in situ hybridization compared with the negative control. (M.N.)

  11. High sensitive diagnostic technique for virus gene using radioisotope. Development of PCR in situ hybridization and its application

    International Nuclear Information System (INIS)

    Iwasaki, Takuya; Sada, Tetsutaro; Terai, Masanori; Sato, Yuko; Kurata, Takeshi; Yamaguchi, Kazuyoshi; Yanagisawa, Akio; Sakai, Yuzo

    1998-01-01

    An animal model, experimentally induced squamous cell carcinoma was produced in hamster mucosa to establish a carcinogenic system inducible by interaction of a virus and chemical(s). Human hydatid mole and cholioepithelioma were chosen as the target subjects. Several tumor cells and host cells were isolated under microscopy and DNA was extracted from these cells to indentify the respective origins (maternal, paternal or host origin). The base sequence of HLADRB region was analyzed by PCR using synthetic primer and the tissue localization was examined by PCR in situ hybridization. A PCR product of 82 bp was found in 15 of 17 samples and there were 2 samples in which the product was not detectable with the constructed primer and 6 samples were negative. While significant differences were not observed by in situ hybridization compared with the negative control. (M.N.)

  12. MMP activity in the hybrid layer detected with in situ zymography.

    Science.gov (United States)

    Mazzoni, A; Nascimento, F D; Carrilho, M; Tersariol, I; Papa, V; Tjäderhane, L; Di Lenarda, R; Tay, F R; Pashley, D H; Breschi, L

    2012-05-01

    Dentinal proteases are believed to play an important role in the degradation of hybrid layers (HL). This study investigated the HL gelatinolytic activity by in situ zymography and functional enzyme activity assay. The hypotheses were that HLs created by an etch-and-rinse adhesive exhibit active gelatinolytic activity, and MMP-2 and -9 activities in dentin increase during adhesive procedures. Etched-dentin specimens were bonded with Adper Scotchbond 1XT and restored with composite. Adhesive/dentin interface slices were placed on microscope slides, covered with fluorescein-conjugated gelatin, and observed with a multi-photon confocal microscope after 24 hrs. Human dentin powder aliquots were prepared and assigned to the following treatments: A, untreated; B, etched with 10% phosphoric acid; or C, etched with 10% phosphoric acid and mixed with Scotchbond 1XT. The MMP-2 and -9 activities of extracts of dentin powder were measured with functional enzyme assays. Intense and continuous enzyme activity was detected at the bottom of the HL, while that activity was more irregular in the upper HL. Both acid-etching and subsequent adhesive application significantly increased MMP-2 and -9 activities (p < 0.05). The results demonstrate, for the first time, intrinsic MMP activity in the HL, and intense activation of matrix-bound MMP activity with both etching and adhesive application.

  13. Monosomy 3 by chromogenic in situ hybridization (CISH) in Iranian patients with uveal melanoma.

    Science.gov (United States)

    Naseripour, Masood; Mehrazma, Mitra; Pourmatin, Rama; Kashkouli, Mohsen Bahmani; Sedaghat, Ahad; Gheytanchi, Elmira

    2015-01-01

    The aim of this study was to investigate the rate of monosomy 3 by CISH technique in Iranian patients with uveal melanoma (UM) and its correlation with clinical and histopathological features. Archival formalin fixed, paraffin-embedded material from 50 patients who had undergone enucleation for large uveal melanomas was obtained. Monosomy of chromosome 3 alteration by chromogenic in situ hybridization (CISH) was investigated. Clinical and histopathological features of tumors were collected. The patients had a mean age of 56.6±7.6 years. Mean basal diameter and thickness of tumors were 14.1 mm and 10.2 mm, respectively. Four patients (8%) were identified to harbor monosomy of chromosome 3. In the mean follow-up of 5.3 years (range, 3.2-9.5 y), only one case with monosomy 3 died of UM metastasis. The most common type of cellularity was mixed cell (86%). There was not any statistically significant correlation between monosomy of chromosome 3 and type of cellularity, ciliary body involvement, and largest basal diameter. The low rate of monosomy chromosome 3 and the consequent low rate of mortality may be indicative of good prognosis in Iranian patients with uveal melanoma.

  14. [Molecular classification of breast cancer patients obtained through the technique of chromogenic in situ hybridization (CISH)].

    Science.gov (United States)

    Fernández, Angel; Reigosa, Aldo

    2013-12-01

    Breast cancer is a heterogeneous disease composed of a growing number of biological subtypes, with substantial variability of the disease progression within each category. The aim of this research was to classify the samples object of study according to the molecular classes of breast cancer: luminal A, luminal B, HER2 and triple negative, as a result of the state of HER2 amplification obtained by the technique of chromogenic in situ hybridization (CISH). The sample consisted of 200 biopsies fixed in 10% formalin, processed by standard techniques up to paraffin embedding, corresponding to patients diagnosed with invasive ductal carcinoma of the breast. These biopsies were obtained from patients from private practice and the Institute of Oncology "Dr. Miguel Pérez Carreño", for immunohistochemistry (IHC) of hormone receptors and HER2 made in the Hospital Metropolitano del Norte, Valencia, Venezuela. The molecular classification of the patient's tumors considering the expression of estrogen and progesterone receptors by IHC and HER2 amplification by CISH, allowed those cases originally classified as unknown, since they had an indeterminate (2+) outcome for HER2 expression by IHC, to be grouped into the different molecular classes. Also, this classification permitted that some cases, initially considered as belonging to a molecular class, were assigned to another class, after the revaluation of the HER2 status by CISH.

  15. Chromosome translocations in chinese medical X-ray workers analyzed by fluorescence in situ hybridization

    International Nuclear Information System (INIS)

    Sun Yuanming; Li Jin; Wang Qin; Tang Weisheng; Wang Zhiquan

    2002-01-01

    Objective: To study long-term radiation effect in occupational workers exposed to low dose X-rays using the method of fluorescence in situ hybridization (FISH). Method: Chromosome translocations of 25 medical X-ray workers were analyzed by FISH with chromosome No. 4 and No. 7 probes according to PAINT (The Protocol for Aberration Identification and Nomenclature Terminology) system. Results: The frequency of genome translocation in X-ray workers was (13.14 ± 1.23)/1000 cells. The rate of complete and incomplete translocation was 1:1.7. According to the calendar year of entry before/after the year of 1965 as the border, the data showed that the incomplete translocation of the after 1965 group was obviously higher than those of the controls (P < 0.01 and P < 0.05, respectively). Conclusion: The chromosome translocation in early Chinese medical X-ray workers is mainly the incomplete one, the frequency of translocation does not dependent on chromosomal DNA content, and incomplete and complete ones increase along with prolongation of working years in their position

  16. Fluorescent in situ hybridization (FISH as a diagnostic tool for Williams-Beuren syndrome

    Directory of Open Access Journals (Sweden)

    Deise Helena de Souza

    2007-01-01

    Full Text Available Fluorescent in situ hybridization (FISH with commercial probes covering the elastin gene (ELN was used to determine the frequency of the 7q11.23 deletion in 18 children clinically diagnosed with Williams-Beuren syndrome (WBS. A de novo deletion was detected in 15 of the children (83%. Diagnostic investigation for WBS started late in childhood (median = 5.8 years. All the children showed facial features typical of the syndrome, mental retardation and developmental delay. Over-friendliness was observed in the majority of cases. Clinodactyly of the 5th finger (n = 13, cardiovascular disease (n = 9, loquacity (n = 9, low birthweight (n = 8, and failure to thrive (n = 9 were observed only in those children with the deletion. Respiratory problems (n = 9, though not previously reported in the literature, was a common finding in the group studied. Our results confirmed that FISH is useful in identifying 7q11.23 deletions in cases of WBS. Clinical manifestations were more evident in the deletion-positive children.

  17. Accuracy Assessment of Interphase Fluorescence In-Situ Hybridization on Uncultured Amniotic Fluid Cells

    Directory of Open Access Journals (Sweden)

    Hamideh Karimi

    2007-01-01

    Full Text Available Background: Parental anxiety while waiting for the results of amniocentesis has been investigatedby many authors. It seems that the implementation of faster techniques such as fluorescence in-situhybridization (FISH will have some benefits in reducing this anxiety. Besides the patients' attitudesto choosing this method, gynecologists who are the persons responsible for treatment, must feelcomfortable about prescribing FISH techniques.Materials and Methods: This study, using a simple methodology, was undertaken to evaluate theresults of FISH tests on the amniotic fluid from 40 pregnant women undergoing cesarean surgery.Two sets of probes including X/Y cocktail and 13, 21 and 18 were applied on different slides.Results: The results of FISH tests were compared with the reports of the pediatrician about thehealth condition of the newborn. Complete conformity between the two sets of findings, haveconvinced our gynecologists of the benefit of prescribing this method to reduce the anxiety ofpatients at risk of having abnormal offspring due to chromosomal anuploidies.Conclusion: As has been documented by many authors, conventional chromosome analysis hasgreat advantages over fluorescence in situ hybridization of interphase amniocytes, but reducing theanxiety of parents is a good reason for employing the FISH technique.

  18. Localization of glucocorticoid receptor mRNA in the male rat brain by in situ hybridization

    International Nuclear Information System (INIS)

    Aronsson, M.; Fuxe, K.; Dong, Y.; Agnati, L.F.; Okret, S.; Gustafsson, J.A.

    1988-01-01

    The localization and distribution of mRNA encoding the glucocorticoid receptor (GR) was investigated in tissue sections of the adult male rat brain by in situ hybridization and RNA blot analysis. GR mRNA levels were measured by quantitative autoradiography with 35S- and 32P-labeled RNA probes, respectively. Strong labeling was observed within the pyramidal nerve cells of the CA1 and CA2 areas of the hippocampal formation, in the granular cells of the dentate gyrus, in the parvocellular nerve cells of the paraventricular hypothalamic nucleus, and in the cells of the arcuate nucleus, especially the parvocellular part. Moderate labeling of a large number of nerve cells was observed within layers II, III, and VI of the neocortex and in many thalamic nuclei, especially the anterior and ventral nuclear groups as well as several midline nuclei. Within the cerebellar cortex, strong labeling was observed all over the granular layer. In the lower brainstem, strong labeling was found within the entire locus coeruleus and within the mesencephalic raphe nuclei rich in noradrenaline and 5-hydroxytryptamine cell bodies, respectively. A close correlation was found between the distribution of GR mRNA and the distribution of previously described GR immunoreactivity. These studies open the possibility of obtaining additional information on in vivo regulation of GR synthesis and how the brain may alter its sensitivity to circulating glucocorticoids

  19. Metaphase FISH on a Chip: Miniaturized Microfluidic Device for Fluorescence in situ Hybridization

    Directory of Open Access Journals (Sweden)

    Niels Tommerup

    2010-11-01

    Full Text Available Fluorescence in situ Hybridization (FISH is a major cytogenetic technique for clinical genetic diagnosis of both inherited and acquired chromosomal abnormalities. Although FISH techniques have evolved and are often used together with other cytogenetic methods like CGH, PRINS and PNA-FISH, the process continues to be a manual, labour intensive, expensive and time consuming technique, often taking over 3–5 days, even in dedicated labs. We have developed a novel microFISH device to perform metaphase FISH on a chip which overcomes many shortcomings of the current laboratory protocols. This work also introduces a novel splashing device for preparing metaphase spreads on a microscope glass slide, followed by a rapid adhesive tape-based bonding protocol leading to rapid fabrication of the microFISH device. The microFISH device allows for an optimized metaphase FISH protocol on a chip with over a 20-fold reduction in the reagent volume. This is the first demonstration of metaphase FISH on a microfluidic device and offers a possibility of automation and significant cost reduction of many routine diagnostic tests of genetic anomalies.

  20. A whole-mount in situ hybridization method for microRNA detection in Caenorhabditis elegans.

    Science.gov (United States)

    Andachi, Yoshiki; Kohara, Yuji

    2016-07-01

    Whole-mount in situ hybridization (WISH) is an outstanding method to decipher the spatiotemporal expression patterns of microRNAs (miRNAs) and provides important clues for elucidating their functions. The first WISH method for miRNA detection was developed in zebrafish. Although this method was quickly adapted for other vertebrates and fruit flies, WISH analysis has not been successfully used to detect miRNAs in Caenorhabditis elegans Here, we show a novel WISH method for miRNA detection in C. elegans Using this method, mir-1 miRNA was detected in the body-wall muscle where the expression and roles of mir-1 miRNA have been previously elucidated. Application of the method to let-7 family miRNAs, let-7, mir-48, mir-84, and mir-241, revealed their distinct but partially overlapping expression patterns, indicating that miRNAs sharing a short common sequence were distinguishably detected. In pash-1 mutants that were depleted of mature miRNAs, signals of mir-48 miRNA were greatly reduced, suggesting that mature miRNAs were detected by the method. These results demonstrate the validity of WISH to detect mature miRNAs in C. elegans. © 2016 Andachi and Kohara; Published by Cold Spring Harbor Laboratory Press for the RNA Society.

  1. JC virus chromogenic in situ hybridization in brain biopsies from patients with and without PML.

    Science.gov (United States)

    Procop, Gary W; Beck, Rose C; Pettay, James D; Kohn, Debra J; Tuohy, Marion J; Yen-Lieberman, Belinda; Prayson, Richard A; Tubbs, Raymond R

    2006-06-01

    Progressive multifocal leukoencephalopathy (PML) is a demyelinating disease of the central nervous system caused by the JC polyoma virus. Electron microscopy and immunohistochemistry are the traditional methods of confirming the presence of the virus in brain biopsies from these patients. We studied the brain biopsies from 7 patients with PML and 6 patients without PML with chromogenic in situ hybridization (CISH) for the JC polyoma virus using a commercially available probe. The biopsies from the patients with the PML cases were proven to contain the JC polyoma virus by traditional and molecular methods. The CISH findings were compared with the known state of infection. All (7/7) of the biopsies from patients with PML were positive for the presence of polyoma virus by CISH, whereas the biopsies from patients without PML were uniformly negative. CISH seems to be a useful tool for the detection of the JC virus in brain biopsies from patients with PML, and is more accessible because a commercial probe is available.

  2. Evaluation of fluorescence in situ hybridization for the detection of bacteria in feline inflammatory liver disease.

    Science.gov (United States)

    Twedt, David C; Cullen, John; McCord, Kelly; Janeczko, Stephanie; Dudak, Julie; Simpson, Kenny

    2014-02-01

    The etiopathogenesis of feline inflammatory liver disease (ILD) is unclear. Therefore, we sought to determine the presence and distribution of bacteria within the livers of cats with ILD using eubacterial fluorescence in situ hybridization (FISH). Histopathology from 39 cats with ILD and 19 with histologically normal livers (C) were classified using World Small Animal Veterinary Association guidelines. Hepatic sections were examined by 16 and 23S ribosomal RNA FISH. Antibodies against cytokeratins and factor VIIIa were used to distinguish bile ducts and vascular structures. Histopathologic findings included non-specific reactive hepatitis (12), neutrophilic cholangitis (NC; 12), lymphocytic cholangitis (seven), cholestasis/obstruction (three), probable lymphoma (three) and acute hepatitis (two). Bacteria were observed in 21/39 ILD and 3/19 C (P = 0.0054). In 8/39 ILD and 2/19 C bacteria were restricted to the outer liver capsule (P = 0.29) and may represent contaminants. The prevalence of intrahepatic bacteria was higher (P = 0.008) in ILD (13/31) than C (1/17). Bacteria in ILD were more frequently (P bacteria. Bacterial culture was positive (predominantly E coli and Enterococcus species) in 11/23 (48%) samples, and concurred with FISH in 15/23 cases. The presence of intrahepatic bacteria in 13/31 (41%) cats with ILD suggests a role in etiopathogenesis. The distribution of bacteria within the liver supports the possibility of colonization via either enteric translocation or hematogenous seeding.

  3. In situ hybridization of bat chromosomes with human (TTAGGGn probe, after previous digestion with Alu I

    Directory of Open Access Journals (Sweden)

    Karina de Cassia Faria

    2002-01-01

    Full Text Available The purpose of this work was to verify the ability of the enzyme Alu I to cleave and/or remove satellite DNA sequences from heterochromatic regions in chromosomes of bats, by identifying the occurrence of modifications in the pattern of fluorescence in situ hybridization with telomeric DNA. The localization and fluorescence intensity of the telomeric DNA sites of the Alu-digested and undigested chromosomes of species Eumops glaucinus, Carollia perspicillata, and Platyrrhinus lineatus were analyzed. Telomeric sequences were detected at the termini of chromosomes of all three species, although, in C. perspicillata, the signals were very faint or absent in most chromosomes. This finding was interpreted as being due to a reduced number of copies of the telomeric repeat, resulting from extensive telomeric association and/or rearrangements undergone by the chromosomes of Carollia. Fluorescent signals were also observed in centromeric and pericentromeric regions in several two-arm chromosomes of E. glaucinus and C. perspicillata. In E. glaucinus and P. lineatus, some interstitial and terminal telomeric sites were observed to be in association with regions of constitutive heterochromatin and ribosomal DNA (NORs. After digestion, these telomeric sites showed a significant decrease in signal intensity, indicating that enzyme Alu I cleaves and/or removes part of the satellite DNA present in these regions. These results suggest that the telomeric sequence is a component of the heterochromatin, and that the C-band- positive regions of bat chromosomes have a different DNA composition.

  4. Mechanisms of induction of chromosomal aberrations and their detection by fluorescence in situ hybridization

    International Nuclear Information System (INIS)

    Natarajan, A.T.

    2002-01-01

    Recently introduced fluorescence in situ hybridization (FISH) technique employing chromosome specific DNA libraries as well as region specific DNA probes (e.g., centromere, telomere) have helped to analyse chromosomal aberrations in great detail and thus have given some new insights into the mechanisms of induction of chromosomal aberrations. The relative proportion of induction of translocations and dicentrics by ionising radiation was studied in human, mice and Chinese hamster cells. Many of the studies point to the differences between the mechanisms of induction of dicentrics and translocations. Preliminary results obtained in our laboratory using arm specific probes for human chromosomes 1 and 3 indicate that the aberrations between the arms appear to be more than expected on a random basis. By employing telomeric probes the frequencies of interstitial deletions were found to be high and similar to the frequencies of dicentrics both in human and mouse lymphocytes. A recent study with human chromosome specific probes clearly shows variation of sensitivity of chromosomes for the induction of exchange aberrations. Radiation response studies with Chinese hamster cells using telomeric probes, suggest that telomeric sequences, especially interstitial ones appear to be an important factor in the origin of both spontaneous and induced chromosomal aberrations

  5. Ulex Europaeus Agglutinin-1 Is a Reliable Taste Bud Marker for In Situ Hybridization Analyses.

    Science.gov (United States)

    Yoshimoto, Joto; Okada, Shinji; Kishi, Mikiya; Misaka, Takumi

    2016-03-01

    Taste signals are received by taste buds. To better understand the taste reception system, expression patterns of taste-related molecules are determined by in situ hybridization (ISH) analyses at the histological level. Nevertheless, even though ISH is essential for determining mRNA expression, few taste bud markers can be applied together with ISH. Ulex europaeus agglutinin-1 (UEA-1) appears to be a reliable murine taste bud marker based on immunohistochemistry (IHC) analyses. However, there is no evidence as to whether UEA-1 can be used for ISH. Thus, the present study evaluated UEA-1 using various histochemical methods, especially ISH. When lectin staining was performed after ISH procedures, UEA-1 clearly labeled taste cellular membranes and distinctly indicated boundaries between taste buds and the surrounding epithelial cells. Additionally, UEA-1 was determined as a taste bud marker not only when used in single-colored ISH but also when employed with double-labeled ISH or during simultaneous detection using IHC and ISH methods. These results suggest that UEA-1 is a useful marker when conducting analyses based on ISH methods. To clarify UEA-1 staining details, multi-fluorescent IHC (together with UEA-1 staining) was examined, resulting in more than 99% of cells being labeled by UEA-1 and overlapping with KCNQ1-expressing cells. © 2016 The Histochemical Society.

  6. In situ 3D nanoprinting of free-form coupling elements for hybrid photonic integration

    Science.gov (United States)

    Dietrich, P.-I.; Blaicher, M.; Reuter, I.; Billah, M.; Hoose, T.; Hofmann, A.; Caer, C.; Dangel, R.; Offrein, B.; Troppenz, U.; Moehrle, M.; Freude, W.; Koos, C.

    2018-04-01

    Hybrid photonic integration combines complementary advantages of different material platforms, offering superior performance and flexibility compared with monolithic approaches. This applies in particular to multi-chip concepts, where components can be individually optimized and tested. The assembly of such systems, however, requires expensive high-precision alignment and adaptation of optical mode profiles. We show that these challenges can be overcome by in situ printing of facet-attached beam-shaping elements. Our approach allows precise adaptation of vastly dissimilar mode profiles and permits alignment tolerances compatible with cost-efficient passive assembly techniques. We demonstrate a selection of beam-shaping elements at chip and fibre facets, achieving coupling efficiencies of up to 88% between edge-emitting lasers and single-mode fibres. We also realize printed free-form mirrors that simultaneously adapt beam shape and propagation direction, and we explore multi-lens systems for beam expansion. The concept paves the way to automated assembly of photonic multi-chip systems with unprecedented performance and versatility.

  7. In situ hybridization for the detection of rust fungi in paraffin embedded plant tissue sections.

    Science.gov (United States)

    Ellison, Mitchell A; McMahon, Michael B; Bonde, Morris R; Palmer, Cristi L; Luster, Douglas G

    2016-01-01

    Rust fungi are obligate pathogens with multiple life stages often including different spore types and multiple plant hosts. While individual rust pathogens are often associated with specific plants, a wide range of plant species are infected with rust fungi. To study the interactions between these important pathogenic fungi and their host plants, one must be able to differentiate fungal tissue from plant tissue. This can be accomplished using the In situ hybridization (ISH) protocol described here. To validate reproducibility using the ISH protocol, samples of Chrysanthemum × morifolium infected with Puccinia horiana, Gladiolus × hortulanus infected with Uromyces transversalis and Glycine max infected with Phakopsora pachyrhizi were tested alongside uninfected leaf tissue samples. The results of these tests show that this technique clearly distinguishes between rust pathogens and their respective host plant tissues. This ISH protocol is applicable to rust fungi and potentially other plant pathogenic fungi as well. It has been shown here that this protocol can be applied to pathogens from different genera of rust fungi with no background staining of plant tissue. We encourage the use of this protocol for the study of plant pathogenic fungi in paraffin embedded sections of host plant tissue.

  8. Fluorescence in situ hybridization in combination with the comet assay and micronucleus test in genetic toxicology

    Directory of Open Access Journals (Sweden)

    Hovhannisyan Galina G

    2010-09-01

    Full Text Available Abstract Comet assay and micronucleus (MN test are widely applied in genotoxicity testing and biomonitoring. While comet assay permits to measure direct DNA-strand breaking capacity of a tested agent MN test allows estimating the induced amount of chromosome and/or genome mutations. The potential of these two methods can be enhanced by the combination with fluorescence in situ hybridization (FISH techniques. FISH plus comet assay allows the recognition of targets of DNA damage and repairing directly. FISH combined with MN test is able to characterize the occurrence of different chromosomes in MN and to identify potential chromosomal targets of mutagenic substances. Thus, combination of FISH with the comet assay or MN test proved to be promising techniques for evaluation of the distribution of DNA and chromosome damage in the entire genome of individual cells. FISH technique also permits to study comet and MN formation, necessary for correct application of these methods. This paper reviews the relevant literature on advantages and limitations of Comet-FISH and MN-FISH assays application in genetic toxicology.

  9. A Case of Renal Primitive Neuroectodermal Tumor Confirmed by Fluorescence in situ Hybridization

    Directory of Open Access Journals (Sweden)

    Toshiki Etani

    2015-04-01

    Full Text Available Primitive neuroectodermal tumor (PNET is a member of the Ewing's sarcoma family of tumors (ESFT. We report a case of PNET in a 66-year-old male who presented with a large solid tumor within the parenchyma of the middle pole of the left kidney with metastases to the left adrenal gland and right ischium. A fine-needle biopsy was performed and showed a small round cell tumor. Results of immunohistochemical staining suggested this tumor belonged to ESFT. Preoperative VDC-IE (combined vincristine, doxorubicin and cyclophosphamide followed by another combination of ifosfamide and etoposide chemotherapy and left radical nephrectomy and adrenalectomy were performed. The histopathological findings of the resected tumor were similar to those in the biopsy specimen, but the results of AE1/AE3 were different. For the diagnosis, fluorescence in situ hybridization was performed. Split signals of the EWSR1 gene were detected, and transmission electron microscopy showed neuroendocrine granules and microtubules. The final diagnosis of this tumor was PNET of the kidney.

  10. Combined Confocal and Wide-Field High-Resolution Cytometry of Fluorescent In Situ Hybridization-Stained Cells

    Czech Academy of Sciences Publication Activity Database

    Kozubek, Michal; Kozubek, Stanislav; Lukášová, Emilie; Bártová, Eva; Skalníková, M.; Matula, Pa.; Matula, Pe.; Jirsová, Pavla; Cafourková, Alena; Koutná, Irena

    2001-01-01

    Roč. 45, č. 1 (2001), s. 1-12 ISSN 0196-4763 R&D Projects: GA MŠk VS97031; GA ČR GA202/99/P008; GA AV ČR IBS5004010 Institutional research plan: CEZ:AV0Z5004920 Keywords : high-resolution cytometry * fluorescence in situ hybridization * interphase nuclei Subject RIV: BO - Biophysics Impact factor: 2.220, year: 2001

  11. Application of Fluorescence In Situ Hybridization (FISH) Technique for the Detection of Genetic Aberration in Medical Science

    OpenAIRE

    Ratan, Zubair Ahmed; Zaman, Sojib Bin; Mehta, Varshil; Haidere, Mohammad Faisal; Runa, Nusrat Jahan; Akter, Nasrin

    2017-01-01

    Fluorescence in situ hybridization (FISH) is a macromolecule recognition technique, which is considered as a new advent in the field of cytology.?Initially, it was developed as a physical mapping tool to delineate genes within chromosomes. The accuracy and versatility of FISH were subsequently capitalized upon in biological and medical research. This visually appealing technique provides an intermediate degree of resolution between DNA analysis and chromosomal investigations. FISH consists of...

  12. Correlation between HER2 gene amplification and protein overexpression through fluorescence in situ hybridization and immunohistochemistry in breast carcinoma patients

    OpenAIRE

    R N Makroo; Mohit Chowdhry; Manoj Kumar; Priyanka Srivastava; Richa Tyagi; Preeti Bhadauria; Sumaid Kaul; Ramesh Sarin; P K Das; Harsh Dua

    2012-01-01

    Background : In India, the incidence of breast cancer has increased in the urban population, with 1 in every 22 women diagnosed with breast cancer. It is important to know the HER2/neu gene status for a better prognostication of these patients. Aim : The aim of this study was to compare the efficacy of fluorescence in situ hybridization (FISH) and immunohistochemistry (IHC) for determining HER2/neu alteration in breast carcinoma. Materials and Methods : A total of 188 histologically proven br...

  13. Evidence for Integrity of Parental Genomes in the Diploid Hybridogenetic Water Frog Pelophylax esculentus by Genomic in situ Hybridization

    Czech Academy of Sciences Publication Activity Database

    Zalésna, A.; Choleva, Lukáš; Ogielska, M.; Rábová, Marie; Marec, František; Ráb, Petr

    2011-01-01

    Roč. 134, č. 3 (2011), s. 206-212 ISSN 1424-8581 R&D Projects: GA MŠk LC06073; GA ČR GA523/09/2106 Institutional research plan: CEZ:AV0Z50450515; CEZ:AV0Z50070508 Keywords : Amphibia * Chromosomes * Genomic in situ hybridization (GISH) Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 1.533, year: 2011

  14. In situ biosynthesis of bacterial nanocellulose-CaCO{sub 3} hybrid bionanocomposite: One-step process

    Energy Technology Data Exchange (ETDEWEB)

    Mohammadkazemi, Faranak, E-mail: f_mkazemi@sbu.ac.ir [Department of Cellulose and Paper Technology, Faculty of New Technologies Engineering, Shahid Beheshti University, Science and Research Campus, Zirab, Savadkooh, Mazandaran (Iran, Islamic Republic of); Faria, Marisa; Cordeiro, Nereida [Faculty of Exact Science and Engineering, University of Madeira, Funchal (Portugal)

    2016-08-01

    In this work, a simple and green route to the synthesis of the bacterial nanocellulose-calcium carbonate (BNC/CaCO{sub 3}) hybrid bionanocomposites using one-step in situ biosynthesis was studied. The CaCO{sub 3} was incorporated in the bacterial nanocellulose structure during the cellulose biosynthesis by Gluconacetobacter xylinus PTCC 1734 bacteria. Hestrin-Schramm (HS) and Zhou (Z) culture media were used to the hybrid bionanocomposites production and the effect of ethanol addition was investigated. Attenuated total reflection Fourier transform infrared spectroscopy, field emission scanning electron microscopy, X-ray diffraction, energy-dispersive X-ray spectroscopy, inverse gas chromatography and thermogravimetric analysis were used to characterize the samples. The experimental results demonstrated that the ethanol and culture medium play an important role in the BNC/CaCO{sub 3} hybrid bionanocomposites production, structure and properties. The BNC/CaCO{sub 3} biosynthesized in Z culture medium revealed higher O/C ratio and amphoteric surface character, which justify the highest CaCO{sub 3} content incorporation. The CaCO{sub 3} was incorporated into the cellulosic matrix decreasing the bacterial nanocellulose crystallinity. This work reveals the high potential of in situ biosynthesis of BNC/CaCO{sub 3} hybrid bionanocomposites and opens a new way to the high value-added applications of bacterial nanocellulose. - Graphical Abstract: Display Omitted - Highlights: • BNC/CaCO{sub 3} hybrid bionanocomposites were produced using in situ biosynthesis process. • Ethanol and culture medium play an important role in the production and properties. • Z-BNC/CaCO{sub 3} bionanocomposites revealed higher O/C ratio and amphoteric surface character. • CaCO{sub 3} incorporated into the BNC decreased crystallinity.

  15. Chromogenic in situ hybridization compared with other approaches to evaluate HER2/neu status in breast carcinomas

    Energy Technology Data Exchange (ETDEWEB)

    Rosa, F.E.; Santos, R.M. [Departamento de Patologia, Hospital das Clínicas, Faculdade de Medicina, Universidade Estadual Paulista, Botucatu, SP (Brazil); Rogatto, S.R. [Departamento de Urologia, Faculdade de Medicina, Universidade Estadual Paulista, Botucatu, SP (Brazil); Hospital A.C. Camargo, CIPE, São Paulo, SP (Brazil); Domingues, M.A.C. [Departamento de Patologia, Faculdade de Medicina, Universidade Estadual Paulista, Botucatu, SP (Brazil)

    2013-03-19

    Human epidermal growth factor receptor 2 (HER2) has been evaluated in breast cancer patients to identify those most likely to benefit from herceptin-targeted therapy. HER2 amplification, detected in 20-30% of invasive breast tumors, is associated with reduced survival and metastasis. The most frequently used technique for evaluating HER2 protein status as a routine procedure is immunohistochemistry (IHC). HER2 copy number alterations have also been evaluated by fluorescence in situ hybridization (FISH) in moderate immunoexpression (IHC 2+) cases. An alternative procedure to evaluate gene amplification is chromogenic in situ hybridization (CISH), which has some advantages over FISH, including the correlation between HER2 status and morphological features. Other methodologies have also been used, such as silver-enhanced in situ hybridization (SISH) and quantitative real-time RT-PCR, to determine the number of HER2 gene copies and expression, respectively. Here we will present a short and comprehensive review of the current advances concerning HER2 evaluation in human breast cancer.

  16. Chromogenic in situ hybridization compared with other approaches to evaluate HER2/neu status in breast carcinomas

    Science.gov (United States)

    Rosa, F.E.; Santos, R.M.; Rogatto, S.R.; Domingues, M.A.C.

    2013-01-01

    Human epidermal growth factor receptor 2 (HER2) has been evaluated in breast cancer patients to identify those most likely to benefit from herceptin-targeted therapy. HER2 amplification, detected in 20-30% of invasive breast tumors, is associated with reduced survival and metastasis. The most frequently used technique for evaluating HER2 protein status as a routine procedure is immunohistochemistry (IHC). HER2 copy number alterations have also been evaluated by fluorescence in situ hybridization (FISH) in moderate immunoexpression (IHC 2+) cases. An alternative procedure to evaluate gene amplification is chromogenic in situ hybridization (CISH), which has some advantages over FISH, including the correlation between HER2 status and morphological features. Other methodologies have also been used, such as silver-enhanced in situ hybridization (SISH) and quantitative real-time RT-PCR, to determine the number of HER2 gene copies and expression, respectively. Here we will present a short and comprehensive review of the current advances concerning HER2 evaluation in human breast cancer. PMID:23558859

  17. Chromogenic in situ hybridization compared with other approaches to evaluate HER2/neu status in breast carcinomas

    Directory of Open Access Journals (Sweden)

    F.E. Rosa

    Full Text Available Human epidermal growth factor receptor 2 (HER2 has been evaluated in breast cancer patients to identify those most likely to benefit from herceptin-targeted therapy. HER2 amplification, detected in 20-30% of invasive breast tumors, is associated with reduced survival and metastasis. The most frequently used technique for evaluating HER2 protein status as a routine procedure is immunohistochemistry (IHC. HER2 copy number alterations have also been evaluated by fluorescence in situ hybridization (FISH in moderate immunoexpression (IHC 2+ cases. An alternative procedure to evaluate gene amplification is chromogenic in situ hybridization (CISH, which has some advantages over FISH, including the correlation between HER2 status and morphological features. Other methodologies have also been used, such as silver-enhanced in situ hybridization (SISH and quantitative real-time RT-PCR, to determine the number of HER2 gene copies and expression, respectively. Here we will present a short and comprehensive review of the current advances concerning HER2 evaluation in human breast cancer.

  18. Chromogenic in situ hybridization compared with other approaches to evaluate HER2/neu status in breast carcinomas

    Directory of Open Access Journals (Sweden)

    F.E. Rosa

    2013-03-01

    Full Text Available Human epidermal growth factor receptor 2 (HER2 has been evaluated in breast cancer patients to identify those most likely to benefit from herceptin-targeted therapy. HER2 amplification, detected in 20-30% of invasive breast tumors, is associated with reduced survival and metastasis. The most frequently used technique for evaluating HER2 protein status as a routine procedure is immunohistochemistry (IHC. HER2 copy number alterations have also been evaluated by fluorescence in situ hybridization (FISH in moderate immunoexpression (IHC 2+ cases. An alternative procedure to evaluate gene amplification is chromogenic in situ hybridization (CISH, which has some advantages over FISH, including the correlation between HER2 status and morphological features. Other methodologies have also been used, such as silver-enhanced in situ hybridization (SISH and quantitative real-time RT-PCR, to determine the number of HER2 gene copies and expression, respectively. Here we will present a short and comprehensive review of the current advances concerning HER2 evaluation in human breast cancer.

  19. Chromogenic in situ hybridization compared with other approaches to evaluate HER2/neu status in breast carcinomas

    International Nuclear Information System (INIS)

    Rosa, F.E.; Santos, R.M.; Rogatto, S.R.; Domingues, M.A.C.

    2013-01-01

    Human epidermal growth factor receptor 2 (HER2) has been evaluated in breast cancer patients to identify those most likely to benefit from herceptin-targeted therapy. HER2 amplification, detected in 20-30% of invasive breast tumors, is associated with reduced survival and metastasis. The most frequently used technique for evaluating HER2 protein status as a routine procedure is immunohistochemistry (IHC). HER2 copy number alterations have also been evaluated by fluorescence in situ hybridization (FISH) in moderate immunoexpression (IHC 2+) cases. An alternative procedure to evaluate gene amplification is chromogenic in situ hybridization (CISH), which has some advantages over FISH, including the correlation between HER2 status and morphological features. Other methodologies have also been used, such as silver-enhanced in situ hybridization (SISH) and quantitative real-time RT-PCR, to determine the number of HER2 gene copies and expression, respectively. Here we will present a short and comprehensive review of the current advances concerning HER2 evaluation in human breast cancer

  20. Identification of MYCN gene amplification in neuroblastoma using chromogenic in situ hybridization (CISH): an alternative and practical method.

    Science.gov (United States)

    Bhargava, Rohit; Oppenheimer, Orit; Gerald, William; Jhanwar, Suresh C; Chen, Beiyun

    2005-06-01

    Chromogenic in situ hybridization (CISH) is a recently developed technique, which utilizes the general principles of in situ hybridization and a detection system similar to immunohistochemistry. To assess the utility of CISH for analysis of MYCN gene amplification, we compared this assay with established diagnostic assays such as Southern blot analysis (SB) and fluorescent in situ hybridization (FISH). CISH was performed on 67 cases of neuroblastoma using tissue microarray (65 cases) and whole tissue sections (2 cases). Unequivocal, high-level amplification (> or =10 gene copies per tumor nucleus) was identified in 19 of 67 (28.4%) tumors. Two (3%) tumors showed low-level amplification (6-9 gene copies per tumor nucleus). No amplification was seen in 46 of 67 (68.6%) tumors. SB data were available in 44 tumors. Forty-one of the 44 tumors (93%) showed concordant results between CISH and SB. Three tumors showed MYCN amplification by CISH but no amplification by SB, most likely due to dilution effect of nonneoplastic tissue in the test samples. Two of these three tumors also showed MYCN amplification by FISH, and the third tumor was not analyzed by FISH. FISH data were available in total of 30 tumors. All 30 tumors showed concordant results between CISH and FISH for classifying a tumor as MYCN amplified or not amplified. We conclude that CISH is an accurate method for determining MYCN gene amplification, with added advantages that make it a more practically useful method.

  1. Human Papilloma Virus 16 and 18 Association in Cervical Intraepithelial Lesions and Cervical Cancers by In Situ Hybridization

    Directory of Open Access Journals (Sweden)

    Mohanty Manisa

    2017-03-01

    Full Text Available Objective: To correlate the association of high risk Human Papilloma Virus (HPV 16, 18 in cervical intraepithelial lesions and cervical cancers by in-situ hybridization (ISH technique. Study Group: Cervical biopsy and hysterectomy specimen of 78 young and adult women, attending Hi-Tech Medical College and Hospital, Bhubaneswar, who were clinically or cytologically suspected of cervical intraepithelial lesion or cervical cancer were taken as source of target viral DNA. Material: Formalin 10% as fixative H & E stain as routine staining agent In-situ hybridization kit for HPV 16 and 18 DNA. Method: After following standard protocol for surgical grossing, HPV 16, 18 In-situ hybridization kit was used on paraffin embedded tissue sections. Results: The percentage of positive cases was highest in cervical cancer patients followed by cervical intraepithelial lesions, high grade, and low grade. Conclusion: This study has been carried out for the first in our state and our results show high degree of positivity of HPV 16/18 in females with cervical intraepithelial lesions and cervical cancers attending our tertiary care hospital.

  2. Detection of a variable number of ribosomal DNA loci by fluorescent in situ hybridization in Populus species.

    Science.gov (United States)

    Prado, E A; Faivre-Rampant, P; Schneider, C; Darmency, M A

    1996-10-01

    Fluorescent in situ hybridization (FISH) was applied to related Populus species (2n = 19) in order to detect rDNA loci. An interspecific variability in the number of hybridization sites was revealed using as probe an homologous 25S clone from Populus deltoides. The application of image analysis methods to measure fluorescence intensity of the hybridization signals has enabled us to characterize major and minor loci in the 18S-5.8S-25S rDNA. We identified one pair of such rDNA clusters in Populus alba; two pairs, one major and one minor, in both Populus nigra and P. deltoides; and three pairs in Populus balsamifera, (two major and one minor) and Populus euroamericana (one major and two minor). FISH results are in agreement with those based on RFLP analysis. The pBG13 probe containing 5S sequence from flax detected two separate clusters corresponding to the two size classes of units that coexist within 5S rDNA of most Populus species. Key words : Populus spp., fluorescent in situ hybridization, FISH, rDNA variability, image analysis.

  3. Structure and Optical Properties of Titania-PDMS Hybrid Nanocomposites Prepared by In Situ Non-Aqueous Synthesis

    Directory of Open Access Journals (Sweden)

    Antoine R. M. Dalod

    2017-12-01

    Full Text Available Organic-inorganic hybrid materials are attractive due to the combination of properties from the two distinct types of materials. In this work, transparent titania-polydimethylsiloxane hybrid materials with up to 15.5 vol. % TiO2 content were prepared by an in situ non-aqueous method using titanium (IV isopropoxide and hydroxy-terminated polydimethylsiloxane as precursors. Spectroscopy (Fourier transform infrared, Raman, Ultraviolet-visible, ellipsometry and small-angle X-ray scattering analysis allowed to describe in detail the structure and the optical properties of the nanocomposites. Titanium alkoxide was successfully used as a cross-linker and titania-like nanodomains with an average size of approximately 4 nm were shown to form during the process. The resulting hybrid nanocomposites exhibit high transparency and tunable refractive index from 1.42 up to 1.56, depending on the titania content.

  4. Novel RNA hybridization method for the in situ detection of ETV1, ETV4, and ETV5 gene fusions in prostate cancer.

    Science.gov (United States)

    Kunju, Lakshmi P; Carskadon, Shannon; Siddiqui, Javed; Tomlins, Scott A; Chinnaiyan, Arul M; Palanisamy, Nallasivam

    2014-09-01

    The genetic basis of 50% to 60% of prostate cancer (PCa) is attributable to rearrangements in E26 transformation-specific (ETS) (ERG, ETV1, ETV4, and ETV5), BRAF, and RAF1 genes and overexpression of SPINK1. The development and validation of reliable detection methods are warranted to classify various molecular subtypes of PCa for diagnostic and prognostic purposes. ETS gene rearrangements are typically detected by fluorescence in situ hybridization and reverse-transcription polymerase chain reaction methods. Recently, monoclonal antibodies against ERG have been developed that detect the truncated ERG protein in immunohistochemical assays where staining levels are strongly correlated with ERG rearrangement status by fluorescence in situ hybridization. However, specific antibodies for ETV1, ETV4, and ETV5 are unavailable, challenging their clinical use. We developed a novel RNA in situ hybridization-based assay for the in situ detection of ETV1, ETV4, and ETV5 in formalin-fixed paraffin-embedded tissues from prostate needle biopsies, prostatectomy, and metastatic PCa specimens using RNA probes. Further, with combined RNA in situ hybridization and immunohistochemistry we identified a rare subset of PCa with dual ETS gene rearrangements in collisions of independent tumor foci. The high specificity and sensitivity of RNA in situ hybridization provides an alternate method enabling bright-field in situ detection of ETS gene aberrations in routine clinically available PCa specimens.

  5. Automated image analysis for quantitative fluorescence in situ hybridization with environmental samples.

    Science.gov (United States)

    Zhou, Zhi; Pons, Marie Noëlle; Raskin, Lutgarde; Zilles, Julie L

    2007-05-01

    When fluorescence in situ hybridization (FISH) analyses are performed with complex environmental samples, difficulties related to the presence of microbial cell aggregates and nonuniform background fluorescence are often encountered. The objective of this study was to develop a robust and automated quantitative FISH method for complex environmental samples, such as manure and soil. The method and duration of sample dispersion were optimized to reduce the interference of cell aggregates. An automated image analysis program that detects cells from 4',6'-diamidino-2-phenylindole (DAPI) micrographs and extracts the maximum and mean fluorescence intensities for each cell from corresponding FISH images was developed with the software Visilog. Intensity thresholds were not consistent even for duplicate analyses, so alternative ways of classifying signals were investigated. In the resulting method, the intensity data were divided into clusters using fuzzy c-means clustering, and the resulting clusters were classified as target (positive) or nontarget (negative). A manual quality control confirmed this classification. With this method, 50.4, 72.1, and 64.9% of the cells in two swine manure samples and one soil sample, respectively, were positive as determined with a 16S rRNA-targeted bacterial probe (S-D-Bact-0338-a-A-18). Manual counting resulted in corresponding values of 52.3, 70.6, and 61.5%, respectively. In two swine manure samples and one soil sample 21.6, 12.3, and 2.5% of the cells were positive with an archaeal probe (S-D-Arch-0915-a-A-20), respectively. Manual counting resulted in corresponding values of 22.4, 14.0, and 2.9%, respectively. This automated method should facilitate quantitative analysis of FISH images for a variety of complex environmental samples.

  6. Longitudinal fluorescence in situ hybridization reveals cytogenetic evolution in myeloma relapsing after autologous transplantation

    Science.gov (United States)

    Merz, Maximilian; Jauch, Anna; Hielscher, Thomas; Mai, Elias K.; Seckinger, Anja; Hose, Dirk; Bertsch, Uta; Neben, Kai; Raab, Marc S.; Salwender, Hans; Blau, Igor W.; Lindemann, Hans-Walter; Schmidt-Wolf, Ingo; Scheid, Christof; Haenel, Mathias; Weisel, Katja; Goldschmidt, Hartmut; Hillengass, Jens

    2017-01-01

    To investigate cytogenetic evolution after upfront autologous stem cell transplantation for newly diagnosed myeloma we retrospectively analyzed fluorescence in situ hybridization results of 128 patients with paired bone marrow samples from the time of primary diagnosis and at relapse. High-risk cytogenetic abnormalities (deletion 17p and/or gain 1q21) occurred more frequently after relapse (odds ratio: 6.33; 95% confidence interval: 1.86–33.42; P<0.001). No significant changes were observed for defined IGH translocations [t(4;14); t(11;14); t(14;16)] or hyperdiploid karyotypes between primary diagnosis and relapse. IGH translocations with unknown partners occurred more frequently at relapse. New deletion 17p and/or gain 1q21 were associated with cytogenetic heterogeneity, since some de novo lesions with different copy numbers were present only in subclones. No distinct baseline characteristics were associated with the occurrence of new high-risk cytogenetic abnormalities after progression. Patients who relapsed after novel agent-based induction therapy had an increased risk of developing high-risk aberrations (odds ratio 10.82; 95% confidence interval: 1.65–127.66; P=0.03) compared to those who were treated with conventional chemotherapy. Survival analysis revealed dismal outcomes regardless of whether high-risk aberrations were present at baseline (hazard ratio, 3.53; 95% confidence interval: 1.53–8.14; P=0.003) or developed at relapse only (hazard ratio, 3.06; 95% confidence interval: 1.09–8.59; P=0.03). Our results demonstrate cytogenetic evolution towards high-risk disease after autologous transplantation and underline the importance of repeated genetic testing in relapsed myeloma (EudraCT number of the HD4 trial: 2004-000944-26). PMID:28495913

  7. Diagnosis of BK viral nephropathy in the renal allograft biopsy: role of fluorescence in situ hybridization.

    Science.gov (United States)

    Wang, Zhen; Portier, Bryce P; Hu, Bo; Chiesa-Vottero, Andres; Myles, Jonathan; Procop, Gary W; Tubbs, Raymond R

    2012-09-01

    Early recognition of BK viral nephropathy is essential for successful management. Our aim in this study was to evaluate a novel fluorescence in situ hybridization (FISH) assay for detection of BK virus in renal transplant biopsies in the context of standard detection methods. Renal allograft biopsies (n = 108) were analyzed via H&E, immunohistochemistry (IHC) for simian virus 40, and FISH for BK virus. BK virus was detected in 16 (14.8%) cases by H&E, 13 (12%) cases by IHC, 18 (16.6%) cases by FISH, and 19 (17.6%) cases by real-time PCR; 24 of 108 showed a discrepancy in ≥1 testing modalities. Comparison of H&E, IHC, and FISH showed no statistical difference in detection of BK virus. However, performing comparisons between the different tissue-based assays in the context of plasma or urine real-time PCR results showed significant improvement in detection of BK by FISH over H&E (P = 0.02) but not IHC (P = 0.07). This novel FISH-based approach for BK virus identification in renal allograft biopsy tissue mirrored real-time PCR results and showed superior performance to detection of inclusions by H&E. Therefore, use of FISH for BK virus detection in the setting of renal allograft biopsy is a useful and sensitive detection method and could be adopted in any laboratory that currently performs FISH analysis. Copyright © 2012 American Society for Investigative Pathology and the Association for Molecular Pathology. Published by Elsevier Inc. All rights reserved.

  8. Endothelin: Visualization of mRNAs by in situ hybridization provides evidence for local action

    International Nuclear Information System (INIS)

    MacCumber, M.W.; Ross, C.A.; Glaser, B.M.; Snyder, S.H.

    1989-01-01

    Endothelin (ET) is a recently identified vasoactive peptide with three isoforms for which three genes have been cloned. The cellular sites of synthesis of this peptide have not yet been identified in vivo. Using Northern blot analysis, we have detected two forms of ET mRNA in rat tissues: a 3.7-kilobase form in the kidney, eye, and brain, a 2.5-kilobase form in the intestine, and both forms in the lung. We have localized these forms of ET mRNA in several rat tissues using in situ hybridization. In the 19-day rat fetus, ET mRNA is highest in the lung, intestine, and meninges. At high resolution, ET mRNA is localized in the lung to respiratory epithelial cells of bronchioles and apparently in blood vessels. In adult tissues, ET mRNA is present throughout the lung, in the renal medulla vasa recta, and in the iris of the eye. ET mRNA is synthesized in close proximity to ET binding sites in many organs (e.g., lung, kidney, intestine, and eye), suggesting a local action of this peptide. However, in other areas (e.g., heart and renal cortex), ET binding sites are present in the absence of ET mRNA, suggesting an action of ET from the bloodstream or from neurons. Northern blot analysis of ET mRNA in microvascular endothelial cells in culture indicates that ET is synthesized in small blood vessels and regulated similarly to its regulation in large vessels. Our results provide evidence that ET, like other regulatory peptides, may serve in several tissues as a neuromodulator or local hormone

  9. Rapid-prenatal diagnosis through fluorescence in situ hybridization for preventing aneuploidy related birth defects.

    Science.gov (United States)

    Fauzdar, Ashish; Chowdhry, Mohit; Makroo, R N; Mishra, Manoj; Srivastava, Priyanka; Tyagi, Richa; Bhadauria, Preeti; Kaul, Anita

    2013-01-01

    Women with high-risk pregnancies are offered prenatal diagnosis through amniocentesis for cytogenetic analysis of fetal cells. The aim of this study was to evaluate the effectiveness of the rapid fluorescence in situ hybridization (FISH) technique for detecting numerical aberrations of chromosomes 13, 21, 18, X and Y in high-risk pregnancies in an Indian scenario. A total of 163 samples were received for a FISH and/or a full karyotype for prenatal diagnosis from high-risk pregnancies. In 116 samples both conventional culture techniques for getting karyotype through G-banding techniques were applied in conjunction to FISH test using the AneuVysion kit (Abbott Molecular, Inc.), following standard recommended protocol to compare the both the techniques in our setup. Out of 116 patients, we got 96 normal for the five major chromosome abnormality and seven patients were found to be abnormal (04 trisomy 21, 02 monosomy X, and 01 trisomy 13) and all the FISH results correlated with conventional cytogenetics. To summarize the results of total 163 patients for the major chromosomal abnormalities analyzed by both/or cytogenetics and FISH there were 140 (86%) normal, 9 (6%) cases were abnormal and another 4 (2.5%) cases were suspicious mosaic and 10 (6%) cases of culture failure. The diagnostic detection rate with FISH in 116 patients was 97.5%. There were no false-positive and false-negative autosomal or sex chromosomal results, within our established criteria for reporting FISH signals. Rapid FISH is a reliable and prompt method for detecting numerical chromosomal aberrations and has now been implemented as a routine diagnostic procedure for detection of fetal aneuploidy in India.

  10. Fluorescence In Situ Hybridization (FISH Signal Analysis Using Automated Generated Projection Images

    Directory of Open Access Journals (Sweden)

    Xingwei Wang

    2012-01-01

    Full Text Available Fluorescence in situ hybridization (FISH tests provide promising molecular imaging biomarkers to more accurately and reliably detect and diagnose cancers and genetic disorders. Since current manual FISH signal analysis is low-efficient and inconsistent, which limits its clinical utility, developing automated FISH image scanning systems and computer-aided detection (CAD schemes has been attracting research interests. To acquire high-resolution FISH images in a multi-spectral scanning mode, a huge amount of image data with the stack of the multiple three-dimensional (3-D image slices is generated from a single specimen. Automated preprocessing these scanned images to eliminate the non-useful and redundant data is important to make the automated FISH tests acceptable in clinical applications. In this study, a dual-detector fluorescence image scanning system was applied to scan four specimen slides with FISH-probed chromosome X. A CAD scheme was developed to detect analyzable interphase cells and map the multiple imaging slices recorded FISH-probed signals into the 2-D projection images. CAD scheme was then applied to each projection image to detect analyzable interphase cells using an adaptive multiple-threshold algorithm, identify FISH-probed signals using a top-hat transform, and compute the ratios between the normal and abnormal cells. To assess CAD performance, the FISH-probed signals were also independently visually detected by an observer. The Kappa coefficients for agreement between CAD and observer ranged from 0.69 to 1.0 in detecting/counting FISH signal spots in four testing samples. The study demonstrated the feasibility of automated FISH signal analysis that applying a CAD scheme to the automated generated 2-D projection images.

  11. Fluorescence in situ hybridization (CARD-FISH) of microorganisms in hydrocarbon contaminated aquifer sediment samples.

    Science.gov (United States)

    Tischer, Karolin; Zeder, Michael; Klug, Rebecca; Pernthaler, Jakob; Schattenhofer, Martha; Harms, Hauke; Wendeberg, Annelie

    2012-12-01

    Groundwater ecosystems are the most important sources of drinking water worldwide but they are threatened by contamination and overexploitation. Petroleum spills account for the most common source of contamination and the high carbon load results in anoxia and steep geochemical gradients. Microbes play a major role in the transformation of petroleum hydrocarbons into less toxic substances. To investigate microbial populations at the single cell level, fluorescence in situ hybridization (FISH) is now a well-established technique. Recently, however, catalyzed reporter deposition (CARD)-FISH has been introduced for the detection of microbes from oligotrophic environments. Nevertheless, petroleum contaminated aquifers present a worst case scenario for FISH techniques due to the combination of high background fluorescence of hydrocarbons and the presence of small microbial cells caused by the low turnover rates characteristic of groundwater ecosystems. It is therefore not surprising that studies of microorganisms from such sites are mostly based on cultivation techniques, fingerprinting, and amplicon sequencing. However, to reveal the population dynamics and interspecies relationships of the key participants of contaminant degradation, FISH is an indispensable tool. In this study, a protocol for FISH was developed in combination with cell quantification using an automated counting microscope. The protocol includes the separation and purification of microbial cells from sediment particles, cell permeabilization and, finally, CARD-FISH in a microwave oven. As a proof of principle, the distribution of Archaea and Bacteria was shown in 60 sediment samples taken across the contaminant plume of an aquifer (Leuna, Germany), which has been heavily contaminated with several ten-thousand tonnes of petroleum hydrocarbons since World War II. Copyright © 2012 Elsevier GmbH. All rights reserved.

  12. Use of Hybridization Chain Reaction-Fluorescent In Situ Hybridization To Track Gene Expression by Both Partners during Initiation of Symbiosis.

    Science.gov (United States)

    Nikolakakis, K; Lehnert, E; McFall-Ngai, M J; Ruby, E G

    2015-07-01

    The establishment of a productive symbiosis between Euprymna scolopes, the Hawaiian bobtail squid, and its luminous bacterial symbiont, Vibrio fischeri, is mediated by transcriptional changes in both partners. A key challenge to unraveling the steps required to successfully initiate this and many other symbiotic associations is characterization of the timing and location of these changes. We report on the adaptation of hybridization chain reaction-fluorescent in situ hybridization (HCR-FISH) to simultaneously probe the spatiotemporal regulation of targeted genes in both E. scolopes and V. fischeri. This method revealed localized, transcriptionally coregulated epithelial cells within the light organ that responded directly to the presence of bacterial cells while, at the same time, provided a sensitive means to directly show regulated gene expression within the symbiont population. Thus, HCR-FISH provides a new approach for characterizing habitat transition in bacteria and for discovering host tissue responses to colonization. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  13. Distribution and evolution of repeated sequences in genomes of Triatominae (Hemiptera-Reduviidae inferred from genomic in situ hybridization.

    Directory of Open Access Journals (Sweden)

    Sebastian Pita

    Full Text Available The subfamily Triatominae, vectors of Chagas disease, comprises 140 species characterized by a highly homogeneous chromosome number. We analyzed the chromosomal distribution and evolution of repeated sequences in Triatominae genomes by Genomic in situ Hybridization using Triatoma delpontei and Triatoma infestans genomic DNAs as probes. Hybridizations were performed on their own chromosomes and on nine species included in six genera from the two main tribes: Triatomini and Rhodniini. Genomic probes clearly generate two different hybridization patterns, dispersed or accumulated in specific regions or chromosomes. The three used probes generate the same hybridization pattern in each species. However, these patterns are species-specific. In closely related species, the probes strongly hybridized in the autosomal heterochromatic regions, resembling C-banding and DAPI patterns. However, in more distant species these co-localizations are not observed. The heterochromatic Y chromosome is constituted by highly repeated sequences, which is conserved among 10 species of Triatomini tribe suggesting be an ancestral character for this group. However, the Y chromosome in Rhodniini tribe is markedly different, supporting the early evolutionary dichotomy between both tribes. In some species, sex chromosomes and autosomes shared repeated sequences, suggesting meiotic chromatin exchanges among these heterologous chromosomes. Our GISH analyses enabled us to acquire not only reliable information about autosomal repeated sequences distribution but also an insight into sex chromosome evolution in Triatominae. Furthermore, the differentiation obtained by GISH might be a valuable marker to establish phylogenetic relationships and to test the controversial origin of the Triatominae subfamily.

  14. Transit experience with hydrogen fueled hybrid electric buses

    International Nuclear Information System (INIS)

    Scott, P.B.; Mazaika, D.M.; Levin, J.; Edwards, T.

    2006-01-01

    Both AC Transit and SunLine Transit operate hybrid electric hydrogen fueled buses in their transit service. ACT presently operates three fuel cell buses in daily revenue service, and SunLine operates a fuel cell bus and a HHICE (Hybrid Hydrogen Internal Combustion Engine) bus. All these buses use similar electric drive train and electric accessories, although the detailed design differs notably between the fuel cell and the hybrid ICE buses. The fuel cell buses use a 120kW UTC fuel cell and a Van Hool Chassis, whereas the HHICE bus uses a turbocharged Ford engine which is capable of 140kW generator output in a New Flyer Chassis. The HHICE bus was the first in service, and has been subjected to both winter testing in Manitoba, Canada and summer testing in the Palm Springs, CA region. The winter testing included passenger sampling using questionnaires to ascertain passenger response. The fuel cell buses were introduced to service at the start of 2006. All five buses are in daily revenue service use. The paper will describe the buses and the experience of the transit properties in operating the buses. (author)

  15. Recent progress in lower hybrid current drive theory and experiments

    International Nuclear Information System (INIS)

    Barbato, E.

    1998-01-01

    In this paper lower hybrid current drive (LHCD) experimental milestones paving the way for future experiments are briefly summarized. The current drive efficiency scaling with the electron temperature is discussed. The role of wave propagation in determining the power deposition profile is stressed and methods are discussed to control the current density profile. Modelling of negative central shear configurations, experimentally obtained by LHCD, are reported. A good agreement is found between the modelling results and the experimental findings, thus showing that a good degree of understanding has been achieved in LHCD theory. (author)

  16. Microstructure and wear of in-situ Ti/(TiN + TiB) hybrid composite layers produced using liquid phase process

    Energy Technology Data Exchange (ETDEWEB)

    Yazdi, R., E-mail: ryazdi@ut.ac.ir; Kashani-Bozorg, S.F.

    2015-02-15

    Tungsten inert gas (TIG) technique was conducted on commercially pure (CP)-Ti substrate, which was coated with h-BN-based powder mixture prior to the treatment. The treated surfaces were evaluated and characterized by means of scanning electron microscope (SEM), X-ray diffraction analysis, and electron dispersive spectrometry (EDS). The microhardness and wear experiment were also performed by using a microhardness machine and pin-on-disk tribometer. As h-BN reacted with titanium, an in-situ hybrid composite layer was formed showing near stoichiometric dendrites of TiN, platelets of TiB and interdendritic regions of α′-Ti martensite crystal structures. The population level of TiN and TiB regions were found to increase using a pre-placed powder mixture with greater h-BN content. However, the fabricated layers exhibited cracking and porosity; these were minimized by adjusting arc energy density and h-BN content of powder mixture. The microhardness value of the fabricated hybrid composite layers was found to be in the range of ∼650 HV{sub 0.2}–1000 HV{sub 0.2}; this is three to five times higher than that of the untreated CP-Ti substrate. In addition, the in-situ hybrid composite layers exhibited superior wear behavior over CP-Ti substrate; this is attributed to the formation of newly formed ceramic phases in the solidified surface layers and good coherent interface between the composite layer and CP-substrate. Meanwhile, severe adhesive wear mechanism of CP-titanium surface changed to mild abrasive one as a result of surface treatment. - Highlights: • In-situ Ti/(TiN + TiB) hybrid composite layers were synthesized by TIG processing on commercially pure titanium. • The microstructure features were characterized by several methods. • Microhardness enhanced three to five times higher than that of the CP-Ti substrate after surface modification. • The fabricated composite layers improved wear resistance of CP-titanium. • Severe adhesive wear mechanism of

  17. Enhanced lower hybrid current drive experiments on HT-7 tokamak

    International Nuclear Information System (INIS)

    Shen Weici; Kuang Guangli; Liu Yuexiu; Ding Bojiang; Shi Yaojiang

    2003-01-01

    Effective Lower Hybrid Current Driving (LHCD) and improved confinement experiments in higher plasma parameters (I p >200 kA, n e >2 x 10 13 cm -3 , T e ≥1 keV) have been curried out in optimized LH wave spectrum and plasma parameters in HT-7 superconducting tokamak. The dependence of current driving efficiency on LH power spectrum, plasma density (anti n e ) and toroidal magnetic field B T has been obtained under optimal conditions. A good CD efficiency was obtained at higher plasma current and higher electron density. The improvement of the energy confinement time is accompanied with the increase in line averaged electron density, and in ion and electron temperatures. The highest current driving efficiency reached η CD =I p (anti n e )R/P RF ≅1.05 x 10 19 Am -2 /W. Wave-plasma coupling was sustained in a good state and the reflective coefficient was less than 5%. The experiments have also demonstrated the ability of LH wave in the start-up and ramp-up of the plasma current. The measurement of the temporal distribution of plasma parameter shows that lower hybrid leads to a broader profile in plasma parameter. The LH power deposition profile and the plasma current density profile were modeled with a 2D Fokker-Planck code corresponding to the evolution process of the hard x-ray detector array

  18. Lower hybrid heating experiment in JFT-2 tokamak

    International Nuclear Information System (INIS)

    Uchara, K.; Nagashima, T.

    1982-01-01

    Lower hybrid heating experiments in JFT-2 are reviewed. Good maintenance and controlling of the coupling structure are very important in the injection of RF power before heating experiments. Accessibility of waves and the existence of the mode conversion region are necessary for ion heating in the main plasma. Parametric instabilities which may bring undesirable power deposition are suppressed by enough electron heating in the boundary region. Optimizing the Nsub(z) spectrum and the improvement of the plasma confinement may lead the electron heating in the high density region. Current generation by use of quasi-linear Landau damping is confirmed and is suggested to bring the improvement of plasma confinement. High power and long pulse klystrons may be expected to open a frontier toward a stational reactor plasma in tokamaks. (author)

  19. Fluorescence in situ hybridization is superior for monitoring Epstein Barr viral load in infectious mononucleosis patients.

    Science.gov (United States)

    Cao, Pengfei; Zhang, Meili; Wang, Wei; Dai, Yafei; Sai, Buqing; Sun, Jun; Wang, Lujuan; Wang, Fan; Li, Guiyuan; Xiang, Juanjuan

    2017-05-03

    Epstein Barr virus (EBV) plays a causal role in some diseases, including infectious mononucleosis, lymphoproliferative diseases and nasopharyngeal carcinoma. Detection of EBV infection has been shown to be a useful tool for diagnosing EBV-related diseases. In the present study, we compared the performance of molecular tests, including fluorescence in situ hybridization (FISH) and EBV real-time PCR, to those of serological assays for the detection of EBV infection. Thirty-eight patients with infectious mononucleosis (IM) were enrolled, of whom 31 were diagnosed with a mild type, and seven were diagnosed with IM with haemophagocytic lymphohistiocytosis and chronic active EBV infection. Twenty healthy controls were involved in the study. The atypical lymphocytes in peripheral blood were detected under a microscope and the percentage of positive cells was calculated. EBV DNA load in peripheral blood was detected using real-time PCR. The FISH assay was developed to detect the EBV genome from peripheral blood mononuclear cells (PBMC). Other diagnosis methods including the heterophil agglutination (HA) test and EBV-VCA-IgM test, to detect EBV were also compared. SPSS17.0 was used for statistical analysis. In all, 5-41% atypical lymphocytes were found among the PBMC in mild IM patients, whereas 8-51% atypical lymphocytes were found in IM patients with haemophagocytic lymphohistiocytosis and chronic active EBV infection patients. There was no significant difference in the ratios of atypical lymphoma between patients of the different types. We observed that 71.2% of mild IM patients and 85.7% of IM patients with haemophagocytic lymphohistiocytosis and chronic active EBV infection patients were positive for EBV-VCA-IgM. EBV-VCA-IgM was negative in all healthy control subjects. In addition, 67.1% of mild IM patients tested heterophile antibody positive, whereas 71.4% of IM patients with haemophagocytic lymphohistiocytosis and chronic active EBV infection tested positive. EBV

  20. Potential actionable targets in appendiceal cancer detected by immunohistochemistry, fluorescent in situ hybridization, and mutational analysis

    Science.gov (United States)

    Millis, Sherri Z.; Kimbrough, Jeffery; Doll, Nancy; Von Hoff, Daniel; Ramanathan, Ramesh K.

    2017-01-01

    Background Appendiceal cancers are rare and consist of carcinoid, mucocele, pseudomyxoma peritonei (PMP), goblet cell carcinoma, lymphoma, and adenocarcinoma histologies. Current treatment involves surgical resection or debulking, but no standard exists for adjuvant chemotherapy or treatment for metastatic disease. Methods Samples were identified from approximately 60,000 global tumors analyzed at a referral molecular profiling CLIA-certified laboratory. A total of 588 samples with appendix primary tumor sites were identified (male/female ratio of 2:3; mean age =55). Sixty-two percent of samples were adenocarcinomas (used for analysis); the rest consisted of 9% goblet cell, 15% mucinous; 6% pseudomyxoma, and less than 5% carcinoids and 2% neuroendocrine. Tests included sequencing [Sanger, next generation sequencing (NGS)], protein expression/immunohistochemistry (IHC), and gene amplification [fluorescent in situ hybridization (FISH) or CISH]. Results Profiling across all appendiceal cancer histological subtypes for IHC revealed: 97% BRCP, 81% MRP1, 81% COX-2, 71% MGMT, 56% TOPO1, 5% PTEN, 52% EGFR, 40% ERCC1, 38% SPARC, 35% PDGFR, 35% TOPO2A, 25% RRM1, 21% TS, 16% cKIT, and 12% for TLE3. NGS revealed mutations in the following genes: 50.4% KRAS, 21.9% P53, 17.6% GNAS, 16.5% SMAD4, 10% APC, 7.5% ATM, 5.5% PIK3CA, 5.0% FBXW7, and 1.8% BRAF. Conclusions Appendiceal cancers show considerable heterogeneity with high levels of drug resistance proteins (BCRP and MRP1), which highlight the difficulty in treating these tumors and suggest an individualized approach to treatment. The incidence of low TS (79%) could be used as a backbone of therapy (using inhibitors such as 5FU/capecitabine or newer agents). Therapeutic options includeTOPO1 inhibitors (irinotecan/topotecan), EGFR inhibitors (erlotinib, cetuximab), PDGFR antagonists (regorafenib, axitinib), MGMT (temozolomide). Clinical trials targeting pathways involving KRAS, p53, GNAS, SMAD4, APC, ATM, PIK3CA, FBXW7, and

  1. Establishment of 60Co dose calibration curve using fluorescent in situ hybridization assay technique: Result of preliminary study

    International Nuclear Information System (INIS)

    Rahimah Abdul Rahim; Noriah Jamal; Noraisyah Mohd Yusof; Juliana Mahamad Napiah; Nelly Bo Nai Lee

    2010-01-01

    This study aims at establishing an in-vitro 60 Co dose calibration curve using Fluorescent In-Situ Hybridization assay technique for the Malaysian National Bio dosimetry Laboratory. Blood samples collected from a female healthy donor were irradiated with several doses of 60 Co radiation. Following culturing of lymphocytes, microscopic slides are prepared, denatured and hybridized. The frequencies of translocation are estimated in the metaphases. A calibration curve was then generated using a regression technique. It shows a good fit to a linear-quadratic model. The results of this study might be useful in estimating absorbed dose for the individual exposed to ionizing radiation retrospectively. This information may be useful as a guide for medical treatment for the assessment of possible health consequences. (author)

  2. In situ hybridization of nucleus basalis neurons shows increased β-amyloid mRNA in Alzheimer disease

    International Nuclear Information System (INIS)

    Cohen, M.L.; Golde, T.E.; Usiak, M.F.; Younkin, L.H.; Younkin, S.G.

    1988-01-01

    To determine which cells within the brain produce β-amyloid mRNA and to assess expression of the β-amyloid gene in Alzheimer disease, the authors analyzed brain tissue from Alzheimer and control patients by in situ hybridization. The results demonstrate that β-amyloid mRNA is produced by neurons in the nucleus basalis of Meynert and cerebral cortex and that nuclues basalis perikarya from Alzheimer patients consistently hybridize more β-amyloid probe than those from controls. These observations support the hypothesis that increased expression of the β-amyloid gene plays an important role in the deposition of amyloid in the brains of patients with Alzheimer disease

  3. Small supernumerary marker chromosome derived from proximal p-arm of chromosome 2: identification by fluorescent in situ hybridization.

    Science.gov (United States)

    Lasan Trcić, Ruzica; Hitrec, Vlasta; Letica, Ljiljana; Cuk, Mario; Begović, Davor

    2003-08-01

    Conventional cytogenetics detected an interstitial deletion of proximal region of p-arm of chromosome 2 in a 6-month-old boy with a phenotype slightly resembling Down's syndrome. The deletion was inherited from the father, whose karyotype revealed a small ring-shaped marker chromosome, in addition to interstitial deletion. Fluorescence in situ hybridization identified the marker, which consisted of the proximal region of the p-arm of chromosome 2, including a part of its centromere. This case shows that molecular cytogenetic analysis can reveal the mechanism of the formation of the marker chromosome.

  4. Assignment of Alzheimer's presenilin-2 (PS-2) gene to 1q42.1 by fluorescence in situ hybridization.

    Science.gov (United States)

    Takano, T; Sahara, N; Yamanouchi, Y; Mori, H

    1997-01-17

    Presenilin-2 (PS-2) was suggested to be localized on 1q31-42 based on linkage analysis and cDNA cloning. The final identification of PS-2 as the causal gene for early-onset familial Alzheimer's disease in Voga-German pedigrees was concluded based on the point mutation found in the candidate cDNA isolated from this familial AD. We present evidence of its physical genome mapping of PS-2 on chromosome 1q42.1 by fluorescence in situ hybridization method.

  5. Electron microscopic in situ hybridization and autoradiography: Localization and transcription of rDNA in human lymphocyte nucleoli

    International Nuclear Information System (INIS)

    Wachtler, F.; Mosgoeller, W.S.; Schwarzacher, H.G.

    1990-01-01

    The distribution of ribosomal DNA (rDNA) in the nucleoli of human lymphocytes was revealed by in situ hybridization with a nonautoradiographic procedure at the electron microscopic level. rDNA is located in the dense fibrillar component of the nucleolus but not in the fibrillar centers. In the same cells the incorporation of tritiated uridine takes place in the dense fibrillar component of the nucleolus as seen by autoradiography followed by gold latensification. From these findings it can be concluded that the transcription of ribosomal DNA takes place in the dense fibrillar component of the nucleolus

  6. In situ hybridization of iodinated 5S and 18/25S RNA to Vicia faba metaphase chromosomes

    International Nuclear Information System (INIS)

    Schubert, I.; Baeumlein, H.; Wobus, U.

    1978-01-01

    In vitro labelled 125 I ribosomal RNA fractions (18/25S and 5S) were in situ hybridized to metaphase chromosomes of a reconstructed karyotype of Vicia faba (characterized by two translocations and one pericentric inversion, each being present homozygously). The sites of 18S and 25S RNA were found to be confined to the nucleolus organizing secondary constriction. Two loci of 5S RNA were recognized on the satellite of nucleolus bearing chromosome. Possible correlations between the location of ribosomal genes, heterochromatic G-bands and clusters of mutagen induced chromatid aberrations are discussed. (author)

  7. Association of Serpulina hyodysenteriae with the colonic mucosa in experimental swine dysentery studied by fluorescent in situ hybridization

    DEFF Research Database (Denmark)

    Jensen, Tim Kåre; Boye, Mette; Møller, Kristian

    1998-01-01

    The localization of Serpulina hyodysenteriae in experimental swine dysentery was studied by fluorescent in situ hybridization (FISH) using an oligonucleotide probe targeting the 23S rRNA of S. hyodysenteriae. Nine 8-week-old pigs were challenged. Seven of the pigs were intragastrically dosed with 1......x10(9) cfu S. hyodysenteriae for 3 consecutive days, whereas two pigs were infected by contact. Six non-challenged pigs served as negative controls. The challenged pigs developed clinical swine dysentery from 8 to 14 days postinfection with typical gross lesions. By FISH S. hyodysenteriae cells...

  8. One-step synthesis of graphene/polyaniline hybrids by in situ intercalation polymerization and their electromagnetic properties

    Science.gov (United States)

    Chen, Xiangnan; Meng, Fanchen; Zhou, Zuowan; Tian, Xin; Shan, Liming; Zhu, Shibu; Xu, Xiaoling; Jiang, Man; Wang, Li; Hui, David; Wang, Yong; Lu, Jun; Gou, Jihua

    2014-06-01

    A new method is introduced for the preparation of graphene/polyaniline hybrids using a one-step intercalation polymerization of aniline inside the expanded graphite. The structural and morphological characterizations were performed by X-ray diffraction analysis, transmission electron microscopy and field emission scanning electron microscopy. Both the experimental and first-principles simulated results show that the aniline cation formed by aniline and H+ tends to be drawn towards the electron-enriched zone and to intercalate into the interlayer of graphite. Subsequently, an in situ polymerization leads to the separation of graphite into graphene sheet, resulting from the exothermic effect and more vigorous movements of the chain molecules of polyaniline. The interactions between polyaniline and graphene were confirmed by Fourier transform infrared spectroscopy and Raman spectra. In addition, the graphene/polyaniline hybrid exhibited a breakthrough in the improvement of microwave absorption.A new method is introduced for the preparation of graphene/polyaniline hybrids using a one-step intercalation polymerization of aniline inside the expanded graphite. The structural and morphological characterizations were performed by X-ray diffraction analysis, transmission electron microscopy and field emission scanning electron microscopy. Both the experimental and first-principles simulated results show that the aniline cation formed by aniline and H+ tends to be drawn towards the electron-enriched zone and to intercalate into the interlayer of graphite. Subsequently, an in situ polymerization leads to the separation of graphite into graphene sheet, resulting from the exothermic effect and more vigorous movements of the chain molecules of polyaniline. The interactions between polyaniline and graphene were confirmed by Fourier transform infrared spectroscopy and Raman spectra. In addition, the graphene/polyaniline hybrid exhibited a breakthrough in the improvement of

  9. Role of chromogenic in situ hybridization (CISH) in the evaluation of HER2 status in breast carcinoma: comparison with immunohistochemistry and FISH.

    Science.gov (United States)

    Li-Ning-T, Elsa; Ronchetti, Ruben; Torres-Cabala, Carlos; Merino, Maria J

    2005-10-01

    We report our experience with Chromogenic in Situ Hybridization (CISH) for the evaluation of HER2 amplification on 55 cases of formalin-fixed, paraffin-embedded invasive breast carcinomas of different histology. All the results were corrected for chromosome 17 aneusomy and compared with immunohistochemistry (IHC); a subset of cases was compared to FISH. Thirty-one of 32 cases in which FISH and CISH were performed yielded the same results. CISH and IHC showed a good concordance in the 0/1+ and 3+ category, while a poor agreement with weakly protein overexpression was confirmed. Chromosome 17 analysis was necessary in cases with a low number of HER2 gene copies. CISH is a useful tool to evaluate breast cancer HER2 status that can be easily implemented in a laboratory of surgical pathology.

  10. Whole-slide imaging is a robust alternative to traditional fluorescent microscopy for fluorescence in situ hybridization imaging using break-apart DNA probes.

    Science.gov (United States)

    Laurent, Camille; Guérin, Maxime; Frenois, François-Xavier; Thuries, Valérie; Jalabert, Laurence; Brousset, Pierre; Valmary-Degano, Séverine

    2013-08-01

    Fluorescence in situ hybridization is an indispensable technique used in routine pathology and for theranostic purposes. Because fluorescence in situ hybridization techniques require sophisticated microscopic workstations and long procedures of image acquisition with sometimes subjective and poorly reproducible results, we decided to test a whole-slide imaging system as an alternative approach. In this study, we used the latest generation of Pannoramic 250 Flash digital microscopes (P250 Flash digital microscopes; 3DHISTECH, Budapest, Hungary) to digitize fluorescence in situ hybridization slides of diffuse large B cells lymphoma cases for detecting MYC rearrangement. The P250 Flash digital microscope was found to be precise with better definition of split signals in cells containing MYC rearrangement with fewer truncated signals as compared to traditional fluorescence microscopy. This digital technique is easier thanks to the preview function, which allows almost immediate identification of the tumor area, and the panning and zooming functionalities as well as a shorter acquisition time. Moreover, fluorescence in situ hybridization analyses using the digital technique appeared to be more reproducible between pathologists. Finally, the digital technique also allowed prolonged conservation of photos. In conclusion, whole-slide imaging technologies represent rapid, robust, and highly sensitive methods for interpreting fluorescence in situ hybridization slides with break-apart probes. In addition, these techniques offer an easier way to interpret the signals and allow definitive storage of the images for pathology expert networks or e-learning databases. Copyright © 2013 Elsevier Inc. All rights reserved.

  11. Reliability of chromogenic in situ hybridization for epidermal growth factor receptor gene copy number detection in non-small-cell lung carcinomas: a comparison with fluorescence in situ hybridization study.

    Science.gov (United States)

    Yoo, Seol Bong; Lee, Hyun Ju; Park, Jung Ok; Choe, Gheeyoung; Chung, Doo Hyun; Seo, Jeong-Wook; Chung, Jin-Haeng

    2010-03-01

    Fluorescence in situ hybridization (FISH) has been known to be the most representative and standardized test for assessing gene amplification. However, FISH requires a fluorescence microscope, the signals are labile and rapidly fade over time. Recently, chromogenic in situ hybridization (CISH) has emerged as a potential alternative to FISH. The aim of this study is to test the reliability of CISH technique for the detection of epidermal growth factor receptor (EGFR) gene amplification in non-small-cell lung carcinomas (NSCLC), to compare CISH results with FISH. A total of 277 formalin-fixed and paraffin embedded NSCLC tissue samples were retrieved from the surgical pathology archives at Seoul National University Bundang Hospital. CISH and FISH examinations were performed to test EGFR gene amplification status. There was high concordance in the assessment of EGFR gene copy number between CISH and FISH tests (Kappa coefficient=0.83). Excellent concordance was shown between two observers on the interpretation of the CISH results (Kappa coefficient=0.90). In conclusion, CISH result is highly reproducible, accurate and practical method to determine EGFR gene amplification in NSCLC. In addition, CISH allows a concurrent analysis of histological features of the tumors and gene copy numbers.

  12. Fluoroscence in situ hybridization of chicken intestinal samples with bacterial rRNA targeted oligonucleotide probes

    DEFF Research Database (Denmark)

    Olsen, Katja Nyholm; Francesch, M.; Christensen, Henrik

    2006-01-01

    were hybridized for 24-72h, centrifuged, washed with pre-heated hybridization buffer, centrifuged and resuspended in Millipore quality water before filtration onto a 0.22 µm black polycarbonate filter. The probes used in this study were, LGC354A, LGC354B, LGC354C, Strc493, Bacto1080, Sal3, Chis150, EUB...

  13. Overview, Progress, and Plans for the Compact Toroidal Hybrid Experiment

    Science.gov (United States)

    Hartwell, G. J.; Allen, N. R.; Ennis, D. A.; Hanson, J. D.; Howell, E. C.; Johnson, C. A.; Knowlton, S. F.; Kring, J. D.; Ma, X.; Maurer, D. A.; Ross, K. G.; Schmitt, J. C.; Traverso, P. J.; Williamson, E. N.

    2017-10-01

    The Compact Toroidal Hybrid (CTH) is an l = 2 , m = 5 torsatron/tokamak hybrid (R0 = 0.75 m, ap 0.2 m, and | B | disruption studies. The main goals of the CTH experiment are to study disruptive behavior as a function of applied 3D magnetic shaping, and to test and advance the V3FIT reconstruction code and NIMROD modeling of CTH. The disruptive density limit is observed to exceed the Greenwald limit as the vacuum transform is increased with no observed threshold for avoidance. Low-q operations (1.1 routine, with disruptions ceasing if the vacuum transform is raised above 0.07. Sawteeth are observed in CTH and have a similar phenomenology to tokamak sawteeth despite employing a 3D confining field. Application of vacuum transform has been demonstrated to reduce and eliminate the vertical drift of elongated discharges. Internal SXR diagnostics, in conjunction with external magnetics, extend the range of reconstruction accuracy into the plasma core. This work is supported by U.S. Department of Energy Grant No. DE-FG02-00ER54610.

  14. Time resolved electron microscopy for in situ experiments

    International Nuclear Information System (INIS)

    Campbell, Geoffrey H.; McKeown, Joseph T.; Santala, Melissa K.

    2014-01-01

    Transmission electron microscopy has functioned for decades as a platform for in situ observation of materials and processes with high spatial resolution. Yet, the dynamics often remain elusive, as they unfold too fast to discern at these small spatial scales under traditional imaging conditions. Simply shortening the exposure time in hopes of capturing the action has limitations, as the number of electrons will eventually be reduced to the point where noise overtakes the signal in the image. Pulsed electron sources with high instantaneous current have successfully shortened exposure times (thus increasing the temporal resolution) by about six orders of magnitude over conventional sources while providing the necessary signal-to-noise ratio for dynamic imaging. We describe here the development of this new class of microscope and the principles of its operation, with examples of its application to problems in materials science

  15. Time resolved electron microscopy for in situ experiments

    Energy Technology Data Exchange (ETDEWEB)

    Campbell, Geoffrey H., E-mail: ghcampbell@llnl.gov; McKeown, Joseph T.; Santala, Melissa K. [Lawrence Livermore National Laboratory, Livermore, California 94550 (United States)

    2014-12-15

    Transmission electron microscopy has functioned for decades as a platform for in situ observation of materials and processes with high spatial resolution. Yet, the dynamics often remain elusive, as they unfold too fast to discern at these small spatial scales under traditional imaging conditions. Simply shortening the exposure time in hopes of capturing the action has limitations, as the number of electrons will eventually be reduced to the point where noise overtakes the signal in the image. Pulsed electron sources with high instantaneous current have successfully shortened exposure times (thus increasing the temporal resolution) by about six orders of magnitude over conventional sources while providing the necessary signal-to-noise ratio for dynamic imaging. We describe here the development of this new class of microscope and the principles of its operation, with examples of its application to problems in materials science.

  16. Experiments at Scale with In-Situ Visualization Using ParaView/Catalyst in RAGE

    Energy Technology Data Exchange (ETDEWEB)

    Kares, Robert John [Los Alamos National Lab. (LANL), Los Alamos, NM (United States)

    2014-10-31

    In this paper I describe some numerical experiments performed using the ParaView/Catalyst in-situ visualization infrastructure deployed in the Los Alamos RAGE radiation-hydrodynamics code to produce images from a running large scale 3D ICF simulation on the Cielo supercomputer at Los Alamos. The detailed procedures for the creation of the visualizations using ParaView/Catalyst are discussed and several images sequences from the ICF simulation problem produced with the in-situ method are presented. My impressions and conclusions concerning the use of the in-situ visualization method in RAGE are discussed.

  17. Fluorescent whole-mount RNA in situ hybridization (F-WISH) in plant germ cells and the fertilized ovule.

    Science.gov (United States)

    Bleckmann, Andrea; Dresselhaus, Thomas

    2016-04-01

    First evidence on gene function and regulation is provided by the cellular expression pattern in complex tissues. However, to understand the activity of a specific gene, it is essential to analyze the regulatory network, which controls the spatio-temporal translation pattern during the entire life span of the transcribed mRNA. To explore mechanisms which control mRNA abundance and localization in space and time, it is necessary to visualize mRNAs quantitatively with a subcellular resolution, without sectioning the tissues. We have adapted and optimized a protocol for colorimetric whole-mount RNA in situ hybridization (WISH) using egg cell-specific digoxigenin (DIG) labeled probes (Hejátko et al., 2006) [1] on ovules and early seeds of Arabidopsis. Furthermore, we established a fluorescent whole-mount RNA in situ hybridization (F-WISH) protocol, which allows mRNA visualization on a subcellular level. The polar localized mRNA of SBT4.13, encoding a subtilase, was identified using this protocol. Both methods are described and discussed in detail. Additionally a (F)-WISH flow-chart is provided along with a troubleshooting table. Copyright © 2015 Elsevier Inc. All rights reserved.

  18. In situ hybridization analysis of the temporospatial expression of the midkine/pleiotrophin family in rat embryonic pituitary gland.

    Science.gov (United States)

    Fujiwara, Ken; Maliza, Rita; Tofrizal, Alimuddin; Batchuluun, Khongorzul; Ramadhani, Dini; Tsukada, Takehiro; Azuma, Morio; Horiguchi, Kotaro; Kikuchi, Motoshi; Yashiro, Takashi

    2014-07-01

    Pituitary gland development is controlled by numerous signaling molecules, which are produced in the oral ectoderm and diencephalon. A newly described family of heparin-binding growth factors, namely midkine (MK)/pleiotrophin (PTN), is involved in regulating the growth and differentiation of many tissues and organs. Using in situ hybridization with digoxigenin-labeled cRNA probes, we detected cells expressing MK and PTN in the developing rat pituitary gland. At embryonic day 12.5 (E12.5), MK expression was localized in Rathke's pouch (derived from the oral ectoderm) and in the neurohypophyseal bud (derived from the diencephalon). From E12.5 to E19.5, MK mRNA was expressed in the developing neurohypophysis, and expression gradually decreased in the developing adenohypophysis. To characterize MK-expressing cells, we performed double-staining of MK mRNA and anterior pituitary hormones. At E19.5, no MK-expressing cells were stained with any hormone. In contrast, PTN was expressed only in the neurohypophysis primordium during all embryonic stages. In situ hybridization clearly showed that MK was expressed in primitive (immature/undifferentiated) adenohypophyseal cells and neurohypophyseal cells, whereas PTN was expressed only in neurohypophyseal cells. Thus, MK and PTN might play roles as signaling molecules during pituitary development.

  19. Evaluation of Myc Gene Amplification in Prostate Cancer Using a Dual Color Chromogenic in-Situ Hybridization (Dual CISH Assay

    Directory of Open Access Journals (Sweden)

    Daniel Lerda

    2013-04-01

    Full Text Available Objetive: The overall purpose of the study was to demonstrate applicability of the Dako dual-color chromogenic in situ hybridization (CISH assay (DAKO Denmark, Glostrup with respect to fluorescence in situ hybridization (FISH probes MYC-C. Methods: MYC gene amplification by FISH and Dako dual-color CISH Results: The study showed that the dual-color CISH assay can convert Texas red and fluorescein isothiocyanate (FITC signals into chromogenic signals. The dual –color CISH assay was performed on 40 cases of prostate cancer. Amplification was identified in 12 of 40 (30% tumors. No amplification was seen in 28 of 40 (70% tumors. FISH data were available in total of 40 tumors. All tumors showed concordant results between dual-color CISH and FISH for classifying a tumor as MYC amplified or not amplified. Conclusions: We conclude that dual-color Dako CISH assay is an accurate method for determining MYC gene amplification with added advantages that make it a more practically useful method. [J Interdiscipl Histopathol 2013; 1(2.000: 81-84

  20. Development of an in situ hybridization assay for the detection of ostreid herpesvirus type 1 mRNAs in the Pacific oyster, Crassostrea gigas.

    Science.gov (United States)

    Corbeil, Serge; Faury, Nicole; Segarra, Amélie; Renault, Tristan

    2015-01-01

    An in situ hybridization protocol for detecting mRNAs of ostreid herpesvirus type 1 (OsHV-1) which infects Pacific oysters, Crassostrea gigas, was developed. Three RNA probes were synthesized by cloning three partial OsHV-1 genes into plasmids using three specific primer pairs, and performing a transcription in the presence of digoxigenin dUTP. The RNA probes were able to detect the virus mRNAs in paraffin sections of experimentally infected oysters 26 h post-injection. The in situ hybridization showed that the OsHV-1 mRNAs were mainly present in connective tissues in gills, mantle, adductor muscle, digestive gland and gonads. DNA detection by in situ hybridization using a DNA probe and viral DNA quantitation by real-time PCR were also performed and results were compared with those obtained using RNA probes. Copyright © 2014 Elsevier B.V. All rights reserved.

  1. Using whole mount in situ hybridization to link molecular and organismal biology.

    Science.gov (United States)

    Jacobs, Nicole L; Albertson, R Craig; Wiles, Jason R

    2011-03-31

    Whole mount in situ hybridization (WISH) is a common technique in molecular biology laboratories used to study gene expression through the localization of specific mRNA transcripts within whole mount specimen. This technique (adapted from Albertson and Yelick, 2005) was used in an upper level undergraduate Comparative Vertebrate Biology laboratory classroom at Syracuse University. The first two thirds of the Comparative Vertebrate Biology lab course gave students the opportunity to study the embryology and gross anatomy of several organisms representing various chordate taxa primarily via traditional dissections and the use of models. The final portion of the course involved an innovative approach to teaching anatomy through observation of vertebrate development employing molecular techniques in which WISH was performed on zebrafish embryos. A heterozygous fibroblast growth factor 8 a (fgf8a) mutant line, ace, was used. Due to Mendelian inheritance, ace intercrosses produced wild type, heterozygous, and homozygous ace/fgf8a mutants in a 1:2:1 ratio. RNA probes with known expression patterns in the midline and in developing anatomical structures such as the heart, somites, tailbud, myotome, and brain were used. WISH was performed using zebrafish at the 13 somite and prim-6 stages, with students performing the staining reaction in class. The study of zebrafish embryos at different stages of development gave students the ability to observe how these anatomical structures changed over ontogeny. In addition, some ace/fgf8a mutants displayed improper heart looping, and defects in somite and brain development. The students in this lab observed the normal development of various organ systems using both external anatomy as well as gene expression patterns. They also identified and described embryos displaying improper anatomical development and gene expression (i.e., putative mutants). For instructors at institutions that do not already own the necessary equipment or where

  2. QUANTITATIVE IMAGING AND STATISTICAL ANALYSIS OF FLUORESCENCE IN SITU HYBRIDIZATION (FISH) OF AUREOBASIDIUM PULLULANS. (R823845)

    Science.gov (United States)

    AbstractImage and multifactorial statistical analyses were used to evaluate the intensity of fluorescence signal from cells of three strains of A. pullulans and one strain of Rhodosporidium toruloides, as an outgroup, hybridized with either a universal o...

  3. Space Storable Hybrid Rockets for Orbit Insertion or In Situ Resource Utilization Applications

    Data.gov (United States)

    National Aeronautics and Space Administration — This research effort will pave the way towards a Mars Sample Return (MSR) campaign and potentially, future human exploration of Mars. Hybrid rockets utilize a solid...

  4. Fluorescence in situ hybridization techniques (FISH) to detect changes in CYP19a gene expression of Japanese medaka (Oryzias latipes)

    International Nuclear Information System (INIS)

    Park, June-Woo; Tompsett, Amber; Zhang, Xiaowei; Newsted, John L.; Jones, Paul D.; Au, Doris; Kong, Richard; Wu, Rudolf S.S.; Giesy, John P.; Hecker, Markus

    2008-01-01

    The aim of this study was to develop a sensitive in situ hybridization methodology using fluorescence-labeled riboprobes (FISH) that allows for the evaluation of gene expression profiles simultaneously in multiple target tissues of whole fish sections of Japanese medaka (Oryzias latipes). To date FISH methods have been limited in their application due to autofluorescence of tissues, fixatives or other components of the hybridization procedure. An optimized FISH method, based on confocal fluorescence microscopy was developed to reduce the autofluorescence signal. Because of its tissue- and gender-specific expression and relevance in studies of endocrine disruption, gonadal aromatase (CYP19a) was used as a model gene. The in situ hybridization (ISH) system was validated in a test exposure with the aromatase inhibitor fadrozole. The optimized FISH method revealed tissue-specific expression of the CYP19a gene. Furthermore, the assay could differentiate the abundance of CYP19a mRNA among cell types. Expression of CYP19a was primarily associated with early stage oocytes, and expression gradually decreased with increasing maturation. No expression of CYP19a mRNA was observed in other tissues such as brain, liver, or testes. Fadrozole (100 μg/L) caused up-regulation of CYP19a expression, a trend that was confirmed by RT-PCR analysis on excised tissues. In a combination approach with gonad histology, it could be shown that the increase in CYP19a expression as measured by RT-PCR on a whole tissue basis was due to a combination of both increases in numbers of CYP19a-containing cells and an increase in the amount of CYP19a mRNA present in the cells

  5. In situ corrosion monitoring of PC structures with distributed hybrid carbon fiber reinforced polymer sensors

    Science.gov (United States)

    Yang, C. Q.; Wu, Z. S.

    2007-08-01

    Firstly, the fabrication and sensing properties of hybrid carbon fiber reinforced polymer (HCFRP) composite sensors are addressed. In order to provide a distributed sensing manner, the HCFRP sensors were divided into multi-zones with electrodes, and each zone was regarded as a separate sensor. Secondly, their application is studied to monitor the steel corrosion of prestressed concrete (PC) beams. The HCFRP sensors with different gauge lengths were mounted on a PC tendon, steel bar and embedded in tensile and compressive sides of the PC beam. The experiment was carried out under an electric accelerated corrosion and a constant load of about 54 kN. The results reveal that the corrosion of the PC tendon can be monitored through measuring the electrical resistance (ER) change of the HCFRP sensors. For the sensors embedded in tensile side of the PC beam, their ER increases as the corrosion progresses, whereas for the sensors embedded in compressive side, their ER decreases with corrosion time. Moreover, the strains due to the corrosion can be obtained based on the ER change and calibration curves of HCFRP sensors. The strains measured with traditional strain gauges agree with the strains calculated from the ER changes of HCFRP sensors. The electrical behavior of the zones where the corrosion was performed is much different from those of the other zones. In these zones, either there exist jumps in ER, or the ER increases with a much larger rate than those of the other zones. Distributed corrosion monitoring for PC structures is thus demonstrated with the application of HCFRP sensors through a proper installation of multi-electrodes.

  6. In-situ fabrication of halloysite nanotubes/silica nano hybrid and its application in unsaturated polyester resin

    Science.gov (United States)

    Lin, Jing; Zhong, Bangchao; Jia, Zhixin; Hu, Dechao; Ding, Yong; Luo, Yuanfang; Jia, Demin

    2017-06-01

    Silica nanoparticles was in-situ grown on the surface of halloysite nanotubes (HNTs) by a facile one-step approach to prepare a unique nano-structured hybrid (HNTs-g-Silica). The structure, morphology and composition of HNTs-g-Silica were investigated. It was confirmed that silica nanoparticles with the diameter of 10-20 nm were chemically grafted through Sisbnd O bonds and uniformly dispersed onto the surface of HNTs, leading to the formation of nano-protrusions on the nanotube surface. Due to the significantly improved interface strength between HNTs-g-Silica and polymer matrix, HNTs-g-Silica effectively toughened unsaturated polyester resin (UPE) and endowed UPE with superior thermal stability compared to HNTs. Based on the unique hybrid architecture and the improved properties of UPE nanocomposites, it is envisioned that HNTs-g-Silica may be a promising filler for more high performance and functional polymers composites and the fabrication method may have implications in the synthesis of nano hybrid materials.

  7. Subcellular localization of low-abundance human immunodeficiency virus nucleic acid sequences visualized by fluorescence in situ hybridization

    International Nuclear Information System (INIS)

    Lawrence, J.B.; Marselle, L.M.; Byron, K.S.; Johnson, C.V.; Sullivan, J.L.; Singer, R.H.

    1990-01-01

    Detection and subcellular localization of human immunodeficiency virus (HIV) were investigated using sensitive high-resolution in situ hybridization methodology. Lymphocytes infected with HIV in vitro or in vivo were detected by fluorescence after hybridization with either biotin or digoxigenin-labeled probes. At 12 hr after infection in vitro, a single intense signal appeared in the nuclei of individual cells. Later in infection, when cytoplasmic fluorescence became intense, multiple nuclear foci frequently appeared. The nuclear focus consisted of newly synthesized HIV RNA as shown by hybridization in the absence of denaturation and by susceptibility to RNase and actinomycin D. Virus was detected in patient lymphocytes and it was shown that a singular nuclear focus also characterizes cells infected in vivo. The cell line 8E5/LAV containing one defective integrated provirus revealed a similar focus of nuclear RNA, and the single integrated HIV genome was unequivocally visualized on a D-group chromosome. This demonstrates an extremely sensitive single-cell assay for the presence of a single site of HIV transcription in vitro and in vivo and suggests that it derives from one (or very few) viral genomes per cell. In contrast, productive Epstein-Barr virus infection exhibited many foci of nuclear RNA per cell

  8. Efficiency of fluorescence in situ hybridization for bacterial cell identification in temporary river sediments with contrasting water content.

    Science.gov (United States)

    Fazi, Stefano; Amalfitano, Stefano; Pizzetti, Ilaria; Pernthaler, Jakob

    2007-09-01

    We studied the efficiency of two hybridization techniques for the analysis of benthic bacterial community composition under varying sediment water content. Microcosms were set up with sediments from four European temporary rivers. Wet sediments were dried, and dry sediments were artificially rewetted. The percentage of bacterial cells detected by fluorescence in situ hybridization with fluorescently monolabeled probes (FISH) significantly increased from dry to wet sediments, showing a positive correlation with the community activity measured via incorporation of (3)H leucine. FISH and signal amplification by catalyzed reporter deposition (CARD-FISH) could significantly better detect cells with low activity in dried sediments. Through the application of an optimized cell permeabilization protocol, the percentage of hybridized cells by CARD-FISH showed comparable values in dry and wet conditions. This approach was unrelated to (3)H leucine incorporation rates. Moreover, the optimized protocol allowed a significantly better visualization of Gram-positive Actinobacteria in the studied samples. CARD-FISH is, therefore, proposed as an effective technique to compare bacterial communities residing in sediments with contrasting water content, irrespective of differences in the activity state of target cells. Considering the increasing frequencies of flood and drought cycles in European temporary rivers, our approach may help to better understand the dynamics of microbial communities in such systems.

  9. In situ experiments for disposal of radioactive wastes in deep geological formations

    International Nuclear Information System (INIS)

    1987-12-01

    This report reviews the current status of in-situ experiments undertaken to assess various concepts for disposal of spent fuel and reprocessed high-level waste in deep geological formations. Specifically it describes in-situ experiments in three geological formations - clay, granite and domed salt. The emphasis in this report is on the in-situ experiments which deal with the various issues related to the near-field effects in a repository and the geological environment immediately surrounding the repository. These near-field effects are due to the disturbance caused by both the construction of the repository and the waste itself. The descriptions are drawn primarily from four underground research facilities: the Underground Experimental Facility, Belgium (clay), the Stripa Project, Sweden and the Underground Research Laboratory, Canada (granite) and the Asse Mine, Federal Republic of Germany (salt). 54 refs, figs and tab

  10. The Newfoundland in-situ Oil Burn Experiment - NOBE

    International Nuclear Information System (INIS)

    Fingas, M.; Li, K.; Ackerman, F.; Bissonnette, M.C.; Lambert, P.; Halley, G.; Nelson, R.; Belanger, J.; Pare, J.R.P.; Campagna, P.R.

    1993-01-01

    A group of over 25 Canadian and US agencies conducted a major offshore oil spill burn near Newfoundland. Over 20 vesels, 7 aircraft, and 230 people were involved in this test, the largest of its kind ever conducted. The burn involved release of two oil spills of ca 50 tons each into a towed fireproof boom. Each burn lasted over an hour. The burn plume was sampled using remote-controlled helicopters and a blimp, and air emissions were monitored downwind from remote controlled boats which also took water samples and temperatures. Over 200 sensors or samplers were used; these will yield data on over 2,000 parameters or substances. Preliminary results are reported. Burning occurred outside the boom due to some initial oil splashover, but this did not result in sheening or significant oil loss. The scaling of burns from test tanks to on-sea burns did not always hold true. Quantitative analytical data showed that emissions from this in-situ fire were less than expected; all measured compounds and parameters were below health concern levels beyond ca 150 m from the fire and very little was detected beyond 500 m. Polycyclic aromatic hydrocarbons (PAH) were found to be lower in the soot than in the starting oil and were consumed by the fire to a large degree. Particulates were found to be of concern only up to 150 m downwind at sea level. Combustion gases did not reach levels of concern and volatile organics were in high concentrations but less than those emitted from a non-burning spill. No compounds of concern could be detected in the water samples. Burn residue samples had lower PAH levels than the starting oil. Generally, burning oil spills at sea was found to be feasible and practical. 2 refs., 9 figs., 3 tabs

  11. mathFISH, a web tool that uses thermodynamics-based mathematical models for in silico evaluation of oligonucleotide probes for fluorescence in situ hybridization.

    Science.gov (United States)

    Yilmaz, L Safak; Parnerkar, Shreyas; Noguera, Daniel R

    2011-02-01

    Mathematical models of RNA-targeted fluorescence in situ hybridization (FISH) for perfectly matched and mismatched probe/target pairs are organized and automated in web-based mathFISH (http://mathfish.cee.wisc.edu). Offering the users up-to-date knowledge of hybridization thermodynamics within a theoretical framework, mathFISH is expected to maximize the probability of success during oligonucleotide probe design.

  12. In situ SAXS experiment during DNA and liposome complexation

    Energy Technology Data Exchange (ETDEWEB)

    Gasperini, A.A.; Cavalcanti, L.P. [Laboratorio Nacional de Luz Sincrotron (LNLS), Campinas, SP (Brazil); Balbino, T.A.; Torre, L.G. de la [Universidade Estadual de Campinas (UNICAMP), SP (Brazil); Oliveira, C.L.P. [Universidade de Sao Paulo (USP), Sao Paulo, SP (Brazil)

    2012-07-01

    Full text: Gene therapy is an exciting research area that allows the treatment of different diseases. Basically, an engineered DNA that codes a protein is the therapeutic drug that has to be delivered to the cell nucleus. After that, the DNA transfection process allows the protein production using the cell machinery. However, the efficient delivery needs DNA protection against nucleases and interstitial fluids. In this context, the use of cationic liposome/DNA complexes is a promising strategy for non-viral gene therapy. Liposomes are lipid systems that self-aggregate in bilayers and the use of cationic lipids allows the electrostatic complexation with DNA. In this work, we used SAXS technique to study the complexation kinetics between cationic liposomes and plasmid DNA and evaluate the liposome structural modifications in the presence of DNA. Liposomes were prepared according to [1] using as plasmid DNA vector model a modified version of pVAX1-GFP with luciferase as reporter gene [2]. The complexation was promoted in a SAXS sample holder containing a microchannel to get access to the compartment between two mica windows where the X-ray beam could cross through [3]. We obtained in situ complexation using such sample holder coupled to a fed-batch reactor through a peristaltic pump. The scattering curves were recorded each 30 seconds during the cycles. The DNA was added until a certain final ratio between surface charges previously determined. We studied the form and structure factor model for the liposome bilayer to fit the scattering curves [4]. Structural information such as the bilayer electronic density profiles, number of bilayers and fluidity were determined as a function of the complexation with DNA. These differences can reflect in singular in vitro and in vivo effects. [1] L. G. de la Torre et al. Colloids and Surfaces B: Biointerfaces, 73, 175 (2009) [2] A. R. Azzoni et al. The Journal of Gene Medicine, 9, 392 (2007) [3] L. P. Cavalcanti et al. Review of

  13. Detection of ALK gene rearrangement in non-small cell lung cancer: a comparison of fluorescence in situ hybridization and chromogenic in situ hybridization with correlation of ALK protein expression.

    Science.gov (United States)

    Kim, Hyojin; Yoo, Seol-Bong; Choe, Ji-Young; Paik, Jin Ho; Xu, Xianhua; Nitta, Hiroaki; Zhang, Wenjun; Grogan, Thomas M; Lee, Choon-Taek; Jheon, Sanghoon; Chung, Jin-Haeng

    2011-08-01

    Accurate determination of ALK rearrangement is important in lung cancer patients, especially in determining their eligibility for crizotinib therapy. Fluorescence in situ hybridization (FISH) has been regarded as the gold standard method for detecting ALK rearrangement. However, FISH requires a fluorescence microscope, and the signals are labile and rapidly fade over time. This study evaluates the concordance between ALK gene rearrangement in non-small cell lung cancer assessed by ALK FISH and a newly developed ALK chromogenic in situ hybridization (CISH) and correlates the results with ALK protein expression assessed by immunohistochemistry. A total of 465 formalin-fixed, paraffin-embedded non-small cell lung cancer samples were analyzed by ALK FISH (PathVysion, Vysis, Abbott) and ALK CISH. For comparison, all specimens were stained by immunohistochemistry (clone 5A4, Novocastra) and interobserver reproducibility was assessed. We found that agreement between the pathologists on the CISH-determined ALK status was achieved in 449 patients (96.6%), and ALK rearrangement was identified in 18 patients (4.0%) in CISH method. Among these cases, 443 cases (95.3%) had results matching the corresponding FISH results: 17 rearranged, 425 wild types, and 1 discordant case. There was high concordance in the assessment of ALK gene rearrangement between FISH and CISH techniques (κ = 0.92) and between observers (κ = 0.97). In addition, there was high concordance in the ALK gene status and ALK protein expression between CISH and IHC tests (κ = 0.82). CISH is a highly reproducible and practical method to detect ALK gene rearrangement and correlated well with ALK protein expression. Here, we present a diagnostic algorithm (Chung's SNUBH ALK protocol) to detect lung cancer with ALK rearrangements using IHC, FISH and CISH. Because CISH allows a concurrent analysis of histological features of the tumors and gene rearrangement, it appears to be a useful method in determining ALK gene

  14. Elucidating the Burden of HIV in Tissues Using Multiplexed Immunofluorescence and In Situ Hybridization: Methods for the Single-Cell Phenotypic Characterization of Cells Harboring HIV In Situ.

    Science.gov (United States)

    Vasquez, Joshua J; Hussien, Rajaa; Aguilar-Rodriguez, Brandon; Junger, Henrik; Dobi, Dejan; Henrich, Timothy J; Thanh, Cassandra; Gibson, Erica; Hogan, Louise E; McCune, Joseph; Hunt, Peter W; Stoddart, Cheryl A; Laszik, Zoltan G

    2018-02-01

    Persistent tissue reservoirs of HIV present a major barrier to cure. Defining subsets of infected cells in tissues is a major focus of HIV cure research. Herein, we describe a novel multiplexed in situ hybridization (ISH) (RNAscope) protocol to detect HIV-DNA (vDNA) and HIV-RNA (vRNA) in formalin-fixed paraffin-embedded (FFPE) human tissues in combination with immunofluorescence (IF) phenotyping of the infected cells. We show that multiplexed IF and ISH (mIFISH) is suitable for quantitative assessment of HIV vRNA and vDNA and that multiparameter IF phenotyping allows precise identification of the cellular source of the ISH signal. We also provide semi-quantitative data on the impact of various tissue fixatives on the detectability of vDNA and vRNA with RNAscope technology. Finally, we describe methods to quantitate the ISH signal on whole-slide digital images and validation of the quantitative ISH data with quantitative real-time PCR for vRNA. It is our hope that this approach will provide insight into the biology of HIV tissue reservoirs and to inform strategies aimed at curing HIV.

  15. Human chromosome-specific changes in a human-hamster hybrid cell line (AL) assessed by fluorescent in situ hybridization (fish)

    International Nuclear Information System (INIS)

    Geard, Charles R.; Jenkins, Gloria

    1995-01-01

    Purpose: To quantitatively assess all gamma-ray induced chromosomal changes confined to one human chromosome using fluorescence microscopy and in situ hybridization with a fluorescently labeled human chromosome specific nucleic acid probe. Methods and Materials: Synchronized human-hamster hybrid cells containing human chromosome 11 were obtained by a modified mitotic shake-off procedure. G1 phase cells (> 95%) were irradiated with 137 Cs gamma rays (0, 0.5, 1.0, 1.5, 2.0, 4.0, 6.0, 8.0, and 10.0 Gy) at a dose rate of 1.1 Gy/min and mitotic cells collected 16-20 h later; chromosomal spreads were prepared, denatured, and hybridized with a fluorescein-tagged nucleic acid probe against total human DNA. Chromosomes were examined by fluorescence microscopy and all categories of change involving the human chromosome 11 as target, recorded. Results: Overall, of the 3104 human-hamster hybrid cells examined, 82.1% were euploid, of which 88.6% contained one copy of human chromosome 11, 6.2% contained two copies, and 5.2% contained 0 copies. This is compatible with mitotic nondisjunction in a small fraction of cells. Of the remaining 17.9% of cells, 85.2% were tetraploid cells with two copies of human chromosome 11. For all aberrations involving human chromosome 11 there was a linear relationship between yield and absorbed dose of 0.1 aberrations per chromosome per Gy. The yield of dicentrics, translocations, and terminal deletions that involve one lesion on the human chromosome was linear, while the yield of interstitial deletions that arise from two interacting lesions on the human chromosome was curvilinear. The frequencies of dicentrics and translocations were about equal, while there was a high (40-60%) incidence of incomplete exchanges between human and hamster chromosomes. Conclusions: Fluorescent in situ hybridization (FISH) procedures allow for the efficient detection of a broad range of induced changes in target chromosomes. Symmetrical exchanges induced in G1

  16. Human cDNA mapping using fluorescence in situ hybridization. Progress report, April 1, 1992--December 31, 1992

    Energy Technology Data Exchange (ETDEWEB)

    Korenberg, J.R.

    1993-03-04

    Genetic mapping is approached using the techniques of high resolution fluorescence in situ hybridization (FISH). This technology and the results of its application are designed to rapidly generate whole genome as tool box of expressed sequence to speed the identification of human disease genes. The results of this study are intended to dovetail with and to link the results of existing technologies for creating backbone YAC and genetic maps. In the first eight months, this approach generated 60--80% of the expressed sequence map, the remainder expected to be derived through more long-term, labor-intensive, regional chromosomal gene searches or sequencing. The laboratory has made significant progress in the set-up phase, in mapping fetal and adult brain and other cDNAs, in testing a model system for directly linking genetic and physical maps using FISH with small fragments, in setting up a database, and in establishing the validity and throughput of the system.

  17. Application of fluorescence in situ hybridization technique in the diagnosis of acute promyelocytic leukemia with abnormal immunophenotype

    International Nuclear Information System (INIS)

    Chen Leilei; Sun Xuemei; Chen Junhao; Zhang Le

    2005-01-01

    To evaluate the utilization of fluorescence in situ hybridization (FISH) technique in the diagnosis of acute promyelocytic leukemia(APL) with abnormal immunophenotype, flow cytometry was used to detect the immunophenotype of mononuclear cells in APL patients and PML/RARα fusion gene was detected by FISH. The mononuclear cells of several APL patients showed abnormal immunophenotype: CD13 + , CD33 + , CD34 - , HLA-DR + and PML/RARα fusion gene was also detected, which was different from the regular result of APL: HLA- DR - , PML/RARα + . Therefore, the detection of immunophenotype in APL patients should not be regarded as the sole accurate target for diagnosing leukemia. FISH ,associated with traditional FAB classification, is a simple, rapid, accurate and direct method. It can be used to help confirm the diagnosis, to guide the formulation of a reasonable chemotherapy scheme and to supervise the efficacy of the treatment in patients with leukemia. (authors)

  18. Linkage of cDNA expression profiles of mesencephalic dopaminergic neurons to a genome-wide in situ hybridization database

    Directory of Open Access Journals (Sweden)

    Simon Horst H

    2009-01-01

    Full Text Available Abstract Midbrain dopaminergic neurons are involved in control of emotion, motivation and motor behavior. The loss of one of the subpopulations, substantia nigra pars compacta, is the pathological hallmark of one of the most prominent neurological disorders, Parkinson's disease. Several groups have looked at the molecular identity of midbrain dopaminergic neurons and have suggested the gene expression profile of these neurons. Here, after determining the efficiency of each screen, we provide a linked database of the genes, expressed in this neuronal population, by combining and comparing the results of six previous studies and verification of expression of each gene in dopaminergic neurons, using the collection of in situ hybridization in the Allen Brain Atlas.

  19. Persistence of translocations detected by means of fluorescence in situ hybridization in peripheral lymphocytes of accidentally exposed radiation workers

    International Nuclear Information System (INIS)

    Pressl, S.; Stephan, G.

    1997-01-01

    The translocation frequency in lymphocytes of radiation workers accidentally exposed a number of years earlier was determined by means of fluorescence in situ hybridization. Chromosomes 2, 4 and 8 were painted, and simultaneously the centromeres. The genomic frequency of translocations is between 1.7 and 17.9 per 1000 cells. This variation is not significantly different from the level in healthy control subjects. Therefore, no radiation exposure could be detected retrospectively. On the other hand, the frequency of dicentrics in these radiation workers measured by means of fluorescence plus Giemsa staining shortly after the exposure was significantly increased, and whole body doses between 0.2 and 0.3 Gy could be calculated. Consequently, it would seem that dicentrics measured shortly after an exposure are a more sensitive indicator than translocations which are determined years later. (author)

  20. Whole-mount in situ hybridization in the rotifer Brachionus plicatilis representing a basal branch of lophotrochozoans.

    Science.gov (United States)

    Boell, Louis A; Bucher, Gregor

    2008-08-01

    In order to broaden the comparative scope of evolutionary developmental biology and to refine our picture of animal macroevolution, it is necessary to establish new model organisms, especially from previously underrepresented groups, like the Lophotrochozoa. We have established the culture and protocols for molecular developmental biology in the rotifer species Brachionus plicatilis Müller (Rotifera, Monogononta). Rotifers are nonsegmented animals with enigmatic basal position within the lophotrochozoans and marked by several evolutionary novelties like the wheel organ (corona), the median eye, and the nonpaired posterior foot. The expression of Bp-Pax-6 is shown using whole-mount in situ hybridization. The inexpensive easy culture and experimental tractability of Brachionus as well as the range of interesting questions to which it holds the key make it a promising addition to the "zoo" of evo-devo model organisms.

  1. Fluorescence photooxidation with eosin: a method for high resolution immunolocalization and in situ hybridization detection for light and electron microscopy

    Science.gov (United States)

    1994-01-01

    A simple method is described for high-resolution light and electron microscopic immunolocalization of proteins in cells and tissues by immunofluorescence and subsequent photooxidation of diaminobenzidine tetrahydrochloride into an insoluble osmiophilic polymer. By using eosin as the fluorescent marker, a substantial improvement in sensitivity is achieved in the photooxidation process over other conventional fluorescent compounds. The technique allows for precise correlative immunolocalization studies on the same sample using fluorescence, transmitted light and electron microscopy. Furthermore, because eosin is smaller in size than other conventional markers, this method results in improved penetration of labeling reagents compared to gold or enzyme based procedures. The improved penetration allows for three-dimensional immunolocalization using high voltage electron microscopy. Fluorescence photooxidation can also be used for high resolution light and electron microscopic localization of specific nucleic acid sequences by in situ hybridization utilizing biotinylated probes followed by an eosin-streptavidin conjugate. PMID:7519623

  2. Filter-Adapted Fluorescent In Situ Hybridization (FA-FISH) for Filtration-Enriched Circulating Tumor Cells.

    Science.gov (United States)

    Oulhen, Marianne; Pailler, Emma; Faugeroux, Vincent; Farace, Françoise

    2017-01-01

    Circulating tumor cells (CTCs) may represent an easily accessible source of tumor material to assess genetic aberrations such as gene-rearrangements or gene-amplifications and screen cancer patients eligible for targeted therapies. As the number of CTCs is a critical parameter to identify such biomarkers, we developed fluorescent in situ hybridization (FISH) for CTCs enriched on filters (filter-adapted-FISH, FA-FISH). Here, we describe the FA-FISH protocol, the combination of immunofluorescent staining (DAPI/CD45) and FA-FISH techniques, as well as the semi-automated microscopy method that we developed to improve the feasibility and reliability of FISH analyses in filtration-enriched CTC.

  3. Localization of introduced genes on the chromosomes of transgenic barley, wheat and triticale by fluorescence in situ hybridization

    DEFF Research Database (Denmark)

    Pedersen, C.; Zimny, J.; Becker, D.

    1997-01-01

    Using fluorescence in situ hybridization (FISH) we localized introduced genes on metaphase chromosomes of barley, wheat, and triticale transformed by microprojectile bombardment of microspores and scutellar tissue with the pDB1 plasmid containing the uidA and bar genes. Thirteen integration sites...... of single-copy integrations. There was a slight tendency towards the localization of transgenes in distal chromosome regions. Using the GAA-satellite sequence for chromosome banding, the chromosomes containing the inserted genes were identified in most cases. Two barley lines derived from the same...... transformant showed a totally different integration pattern. Southern analysis confirmed that the inserted genes were segregating independently, resulting in different integration patterns among the progeny lines. The application of the FISH technique for the analysis of transgenic plants is discussed....

  4. Scanning electron microscopy and fluorescent in situ hybridization of experimental Brachyspira (Serpulina) pilosicoli infection in growing pigs

    DEFF Research Database (Denmark)

    Jensen, Tim Kåre; Møller, Kristian; Boye, Mette

    2000-01-01

    Two groups of six 8-week-old pigs were challenged with 1X10(9) cfu Brachyspira (Serpulina) pilosicoli or Serpulina intermedia daily for 3 consecutive days to study the pathology of porcine colonic spirochetosis by scanning electron microscopy (SEM) and fluorescent in situ hybridization (FISH......; however, only two pigs developed transient watery diarrhea. S. intermedia was reisolated from four of the inoculated pigs, but clinical signs were not observed. Gross examination of the B. pilosicoli-infected pigs revealed dilated large intestines with a hyperemic mucosa, whereas the large intestines...... of the S. intermedia-inoculated pigs and the control pigs appeared normal. SEM examination of B. pilosicoli-infected pigs revealed degenerated epithelial cells and spirochetal colonization of the colonic mucosa in four pigs. By FISH, B. pilosicoli cells were found colonizing and invading the surface...

  5. In-situ SEM microchip setup for electrochemical experiments with water based solutions

    International Nuclear Information System (INIS)

    Jensen, E.; Købler, C.; Jensen, P.S.; Mølhave, K.

    2013-01-01

    Studying electrochemical (EC) processes with electron microscopes offers the possibility of achieving much higher resolution imaging of nanoscale processes in real time than with optical microscopes. We have developed a vacuum sealed liquid sample electrochemical cell with electron transparent windows, microelectrodes and an electrochemical reference electrode. The system, called the EC-SEM Cell, is used to study electrochemical reactions in liquid with a standard scanning electron microscope (SEM). The central component is a microfabricated chip with a thin (50 nm) Si-rich silicon nitride (SiNx) window with lithographically defined platinum microelectrodes. We show here the design principles of the EC-SEM system, its detailed construction and how it has been used to perform a range of EC experiments, two of which are presented here. It is shown that the EC-SEM Cell can survive extended in-situ EC experiments. Before the EC experiments we characterized the beam current being deposited in the liquid as this will affect the experiments. The first EC experiment shows the influence of the electron-beam (e-beam) on a nickel solution by inducing electroless nickel deposition on the window when increasing the current density from the e-beam. The second experiment shows electrolysis in EC-SEM Cell, induced by the built-in electrodes. - Highlights: • New in-situ SEM system for electrochemistry. • In-situ Beam current measurements through liquid. • In-situ SEM E-beam-induced electroless deposition of Ni. • In-situ electrolysis

  6. In-situ SEM microchip setup for electrochemical experiments with water based solutions

    Energy Technology Data Exchange (ETDEWEB)

    Jensen, E., E-mail: eric.jensen@nanotech.dtu.dk [DTU Nanotech, Technical University of Denmark, Ørsteds Plads, Building 345E, 2800 Kongens Lyngby (Denmark); DTU CEN, Technical University of Denmark, Fysikvej, Building 307, 2800 Kongens Lyngby (Denmark); Købler, C., E-mail: carsten.kobler@nanotech.dtu.dk [DTU Nanotech, Technical University of Denmark, Ørsteds Plads, Building 345E, 2800 Kongens Lyngby (Denmark); DTU CEN, Technical University of Denmark, Fysikvej, Building 307, 2800 Kongens Lyngby (Denmark); Jensen, P.S., E-mail: psj@kemi.dtu.dk [DTU Kemi, Technical University of Denmark, Kemitorvet, Building 207, 2800 Kongens Lyngby (Denmark); Mølhave, K., E-mail: kristian.molhave@nanotech.dtu.dk [DTU Nanotech, Technical University of Denmark, Ørsteds Plads, Building 345E, 2800 Kongens Lyngby (Denmark)

    2013-06-15

    Studying electrochemical (EC) processes with electron microscopes offers the possibility of achieving much higher resolution imaging of nanoscale processes in real time than with optical microscopes. We have developed a vacuum sealed liquid sample electrochemical cell with electron transparent windows, microelectrodes and an electrochemical reference electrode. The system, called the EC-SEM Cell, is used to study electrochemical reactions in liquid with a standard scanning electron microscope (SEM). The central component is a microfabricated chip with a thin (50 nm) Si-rich silicon nitride (SiNx) window with lithographically defined platinum microelectrodes. We show here the design principles of the EC-SEM system, its detailed construction and how it has been used to perform a range of EC experiments, two of which are presented here. It is shown that the EC-SEM Cell can survive extended in-situ EC experiments. Before the EC experiments we characterized the beam current being deposited in the liquid as this will affect the experiments. The first EC experiment shows the influence of the electron-beam (e-beam) on a nickel solution by inducing electroless nickel deposition on the window when increasing the current density from the e-beam. The second experiment shows electrolysis in EC-SEM Cell, induced by the built-in electrodes. - Highlights: • New in-situ SEM system for electrochemistry. • In-situ Beam current measurements through liquid. • In-situ SEM E-beam-induced electroless deposition of Ni. • In-situ electrolysis.

  7. Validation of interphase fluorescence in situ hybridization (iFISH for multiple myeloma using CD138 positive cells

    Directory of Open Access Journals (Sweden)

    Renata Kiyomi Kishimoto

    2016-06-01

    Full Text Available ABSTRACT BACKGROUND: Multiple myeloma is a plasma cell neoplasm with acquired genetic abnormalities of clinical and prognostic importance. Multiple myeloma differs from other hematologic malignancies due to a high fraction of low proliferating malignant plasma cells and the paucity of plasma cells in bone marrow aspiration samples, making cytogenetic analysis a challenge. An abnormal karyotype is found in only one-third of patients with multiple myeloma and interphase fluorescence in situ hybridization is the most useful test for studying the chromosomal abnormalities present in almost 90% of cases. However, it is necessary to study the genetic abnormalities in plasma cells after their identification or selection by morphology, immunophenotyping or sorting. Other challenges are the selection of the most informative FISH panel and determining cut-off levels for FISH probes. This study reports the validation of interphase fluorescence in situ hybridization using CD138 positive cells, according to proposed guidelines published by the European Myeloma Network (EMN in 2012. METHOD: Bone marrow samples from patients with multiple myeloma were used to standardize a panel of five probes [1q amplification, 13q14 deletion, 17p deletion, t(4;14, and t(14;16] in CD138+ cells purified by magnetic cell sorting. RESULTS: This test was validated with a low turnaround time and good reproducibility. Five of six samples showed genetic abnormalities. Monosomy/deletion 13 plus t(4;14 were found in two cases. CONCLUSION: This technique together with magnetic cell sorting is effective and can be used in the routine laboratory practice. In addition, magnetic cell sorting provides a pure plasma cell population that allows other molecular and genomic studies.

  8. Chromogenic in situ hybridization is a reliable assay for detection of ALK rearrangements in adenocarcinomas of the lung.

    Science.gov (United States)

    Schildhaus, Hans-Ulrich; Deml, Karl-Friedrich; Schmitz, Katja; Meiboom, Maren; Binot, Elke; Hauke, Sven; Merkelbach-Bruse, Sabine; Büttner, Reinhard

    2013-11-01

    Reliable detection of anaplastic lymphoma kinase (ALK) rearrangements is a prerequisite for personalized treatment of lung cancer patients, as ALK rearrangements represent a predictive biomarker for the therapy with specific tyrosine kinase inhibitors. Currently, fluorescent in situ hybridization (FISH) is considered to be the standard method for assessing formalin-fixed and paraffin-embedded tissue for ALK inversions and translocations. However, FISH requires a specialized equipment, the signals fade rapidly and it is difficult to detect overall morphology and tumor heterogeneity. Chromogenic in situ hybridization (CISH) has been successfully introduced as an alternative test for the detection of several genetic aberrations. This study validates a newly developed ALK CISH assay by comparing FISH and CISH signal patterns in lung cancer samples with and without ALK rearrangements. One hundred adenocarcinomas of the lung were included in this study, among them 17 with known ALK rearrangement. FISH and CISH were carried out and evaluated according to the manufacturers' recommendations. For both assays, tumors were considered positive if ≥15% of tumor cells showed either isolated 3' signals or break-apart patterns or a combination of both. A subset of tumors was exemplarily examined by using a novel EML4 (echinoderm microtubule-associated protein-like 4) CISH probe. Red, green and fusion CISH signals were clearcut and different signal patterns were easily recognized. The percentage of aberrant tumor cells was statistically highly correlated (PCISH. On the basis of 86 samples that were evaluable by ALK CISH, we found a 100% sensitivity and 100% specificity of this assay. Furthermore, EML4 rearrangements could be recognized by CISH. CISH is a highly reliable, sensitive and specific method for the detection of ALK gene rearrangements in pulmonary adenocarcinomas. Our results suggest that CISH might serve as a suitable alternative to FISH, which is the current gold

  9. Chromogenic in situ hybridization (CISH): a novel alternative in screening archival breast cancer tissue samples for HER-2/neu status.

    Science.gov (United States)

    Madrid, Manuelito A; Lo, Raymundo W

    2004-01-01

    Chromogenic in situ hybridization (CISH) is emerging as a practical, cost-effective, and valid alternative to fluorescent in situ hybridization in testing for gene alteration, especially in centers primarily working with immunohistochemistry (IHC). We assessed Her-2/neu alteration using CISH on formalin-fixed paraffin-embedded primary invasive ductal carcinoma tumors in which IHC (CB11 antibody) had previously been performed, and we compared the results with IHC. The 160 selected cases were equally stratified randomly into the four IHC categories (scores of 0, 1+, 2+, and 3+). We also compared age at diagnosis and tumor histologic grade with IHC and CISH Her-2/neu. We were able to perform and evaluate CISH successfully on all cases. The agreement between 3+ IHC and CISH-amplified cases as well as between all IHC and CISH Her-2/neu negative cases was 100%, and the concordance on all positive cases was 72.50%, with an overall agreement of 86.25%. All the discordant cases had 2+ IHC scores. Although we noted Her-2/neu positivity more in premenopausal women, the age at diagnosis was not significantly associated with IHC or CISH results. Similarly, although the small group of well-differentiated tumors was apparently Her-2/neu negative in both tests, no significant association was noted between any tumor histologic grade and either IHC or CISH results. CISH is easily integrated into routine testing in our laboratory. It is a necessary adjunct in determining the subset of non-amplified IHC-positive invasive tumors that will not benefit from trastuzumab therapy. Those cases with 2+ IHC results will be triaged and subjected to CISH. Her-2/neu testing should be done on all breast cancer cases regardless of age at presentation and tumor histologic grade.

  10. Chromogenic in situ hybridization (CISH): a novel alternative in screening archival breast cancer tissue samples for HER-2/neu status

    International Nuclear Information System (INIS)

    Madrid, Manuelito A; Lo, Raymundo W

    2004-01-01

    Chromogenic in situ hybridization (CISH) is emerging as a practical, cost-effective, and valid alternative to fluorescent in situ hybridization in testing for gene alteration, especially in centers primarily working with immunohistochemistry (IHC). We assessed Her-2/neu alteration using CISH on formalin-fixed paraffin-embedded primary invasive ductal carcinoma tumors in which IHC (CB11 antibody) had previously been performed, and we compared the results with IHC. The 160 selected cases were equally stratified randomly into the four IHC categories (scores of 0, 1+, 2+, and 3+). We also compared age at diagnosis and tumor histologic grade with IHC and CISH Her-2/neu. We were able to perform and evaluate CISH successfully on all cases. The agreement between 3+ IHC and CISH-amplified cases as well as between all IHC and CISH Her-2/neu negative cases was 100%, and the concordance on all positive cases was 72.50%, with an overall agreement of 86.25%. All the discordant cases had 2+ IHC scores. Although we noted Her-2/neu positivity more in premenopausal women, the age at diagnosis was not significantly associated with IHC or CISH results. Similarly, although the small group of well-differentiated tumors was apparently Her-2/neu negative in both tests, no significant association was noted between any tumor histologic grade and either IHC or CISH results. CISH is easily integrated into routine testing in our laboratory. It is a necessary adjunct in determining the subset of non-amplified IHC-positive invasive tumors that will not benefit from trastuzumab therapy. Those cases with 2+ IHC results will be triaged and subjected to CISH. Her-2/neu testing should be done on all breast cancer cases regardless of age at presentation and tumor histologic grade

  11. Fluorescence in situ hybridization as adjunct to cytology improves the diagnosis and directs estimation of prognosis of malignant pleural effusions

    Directory of Open Access Journals (Sweden)

    Han Jingquan

    2012-11-01

    Full Text Available Abstract Background The identification of malignant cells in effusions by conventional cytology is hampered by its limited sensitivity and specificity. The aim of this study was to investigate the value of fluorescence in situ hybridization (FISH as adjuncts to conventional cytologic examination in patients with malignant pleural effusions. Methods We conducted a retrospective cohort study of 93 inpatients with pleural effusions (72 malignant pleural effusions metastatic from 11 different organs and 21 benign over 23 months. All the patients came from Chinese northeast areas. Aspirated pleural fluid underwent cytologic examination and fluorescence in situ hybridization (FISH for aneuploidy. We used FISH in single-colour or if appropriate in dual-colour evaluation to detect chromosomal aberrations (chromosomes 7, 11, and 17 in effusion cells as markers of malignancy, to raise the diagnostic yield and identified the efficiency by diagnostic biopsy. Predominant cytogenetic anomalies and patterns of intratumor cytogenetic heterogeneity were brought in relation to overall survival rate. Results Cytology alone confirmed malignant pleural effusions in 45 of 72 patients (sensitivity 63%, whereas FISH alone positively identified 48 of 72 patients (sensitivity 67%. Both tests had high specificity in predicting benign effusions. If cytology and FISH were considered together, they exhibited 88% sensitivity and 94.5% specificity in discriminating benign and malignant effusions. Combined, the two assays were more sensitive than either test alone. Although the positive predictive value of each test was 94.5%, the negative predictive value of cytology and FISH combined was 78%, better than 47% and 44% for FISH and cytology alone, respectively. There was a significantly prolonged survival rate for patients with aneuploidy for chromosome 17. Conclusions FISH in combination with conventional cytology is a highly sensitive and specific diagnostic tool for detecting

  12. Spatial gene expression quantification: a tool for analysis of in situ hybridizations in sea anemone Nematostella vectensis

    Directory of Open Access Journals (Sweden)

    Botman Daniel

    2012-10-01

    Full Text Available Abstract Background Spatial gene expression quantification is required for modeling gene regulation in developing organisms. The fruit fly Drosophila melanogaster is the model system most widely applied for spatial gene expression analysis due to its unique embryonic properties: the shape does not change significantly during its early cleavage cycles and most genes are differentially expressed along a straight axis. This system of development is quite exceptional in the animal kingdom. In the sea anemone Nematostella vectensis the embryo changes its shape during early development; there are cell divisions and cell movement, like in most other metazoans. Nematostella is an attractive case study for spatial gene expression since its transparent body wall makes it accessible to various imaging techniques. Findings Our new quantification method produces standardized gene expression profiles from raw or annotated Nematostella in situ hybridizations by measuring the expression intensity along its cell layer. The procedure is based on digital morphologies derived from high-resolution fluorescence pictures. Additionally, complete descriptions of nonsymmetric expression patterns have been constructed by transforming the gene expression images into a three-dimensional representation. Conclusions We created a standard format for gene expression data, which enables quantitative analysis of in situ hybridizations from embryos with various shapes in different developmental stages. The obtained expression profiles are suitable as input for optimization of gene regulatory network models, and for correlation analysis of genes from dissimilar Nematostella morphologies. This approach is potentially applicable to many other metazoan model organisms and may also be suitable for processing data from three-dimensional imaging techniques.

  13. Detection of c-myc amplification in formalin-fixed paraffin-embedded tumor tissue by chromogenic in situ hybridization (CISH).

    Science.gov (United States)

    Todorović-Raković, Nataša

    2013-01-01

    In situ hybridization (ISH) allows evaluation of genetic abnormalities, such as changes in chromosome number, chromosome translocations or gene amplifications, by hybridization of tagged DNA (or RNA) probes with complementary DNA (or RNA) sequences in interphase nuclei of target tissue. However, chromogenic in situ hybridization (CISH) is also applicable to formalin-fixed, paraffin-embedded (FFPE) tissues, besides metaphase chromosome spreads. CISH is similar to fluorescent in situ hybridization (FISH) regarding pretreatments and hybridization protocols but differs in the way of visualization. Indeed, CISH signal detection is similar to that used in immunohistochemistry, making use of a peroxidase-based chromogenic reaction instead of fluorescent dyes. In particular, tagged DNA probes are indirectly detected using an enzyme-conjugated antibody targeting the tags. The enzymatic reaction of the chromogenic substrate leads to the formation of strong permanent brown signals that can be visualized by bright-field microscopy at 40 × magnification. The advantage of CISH is that it allows the simultaneous observation of gene amplification and tissue morphology and the slides can be stored for a long time.

  14. In situ hybridization of phytoplankton using fluorescently labeled rRNA probes

    OpenAIRE

    Groben, R.; Medlin, Linda

    2005-01-01

    Fluorescently-labelled molecular probes were used to identify and characterise phytoplankton species using in situ hybridisation coupled with fluorescence microscopy and flow cytometry. The application of this technique is sometimes problematic, because of the many different species with which this method is to be used. Problems that may occur are: probe penetration versus maintanance of cell stability, strong autofluorescence and/or cell lost during the sample processing. Here we present a m...

  15. Seven-hour fluorescence in situ hybridization technique for enumeration of Enterobacteriaceae in food and environmental water sample.

    Science.gov (United States)

    Ootsubo, M; Shimizu, T; Tanaka, R; Sawabe, T; Tajima, K; Ezura, Y

    2003-01-01

    A fluorescent in situ hybridization (FISH) technique using an Enterobacteriaceae-specific probe (probe D) to target 16S rRNA was improved in order to enumerate, within a single working day, Enterobacteriaceae present in food and environmental water samples. In order to minimize the time required for the FISH procedure, each step of FISH with probe D was re-evaluated using cultured Escherichia coli. Five minutes of ethanol treatment for cell fixation and hybridization were sufficient to visualize cultured E. coli, and FISH could be performed within 1 h. Because of the difficulties in detecting low levels of bacterial cells by FISH without cultivation, a FISH technique for detecting microcolonies on membrane filters was investigated to improve the bacterial detection limit. FISH with probe D following 6 h of cultivation to grow microcolonies on a 13 mm diameter membrane filter was performed, and whole Enterobacteriaceae microcolonies on the filter were then detected and enumerated by manual epifluorescence microscopic scanning at magnification of x100 in ca 5 min. The total time for FISH with probe D following cultivation (FISHFC) was reduced to within 7 h. FISHFC can be applied to enumerate cultivable Enterobacteriaceae in food (above 100 cells g-1) and environmental water samples (above 1 cell ml-1). Cultivable Enterobacteriaceae in food and water samples were enumerated accurately within 7 h using the FISHFC method. A FISHFC method capable of evaluating Enterobacteriaceae contamination in food and environmental water within a single working day was developed.

  16. Facile fabrication of ultrathin hybrid membrane for highly flexible supercapacitors via in-situ phase separation of polyethersulfone

    Science.gov (United States)

    Zhao, Xiaoning; Ran, Fen; Shen, Kuiwen; Yang, Yunlong; Wu, Jiayu; Niu, Xiaoqin; Kong, Lingbin; Kang, Long; Chen, Shaowei

    2016-10-01

    In this article, a facile method based on in-situ phase-separation was developed for the fabrication of ultrathin hybrid membranes for highly flexible supercapacitors. The structures and morphologies of the prepared electrodes were characterized by scanning electron microscopy (SEM), Fourier-transformed infrared spectroscopy (FTIR), and thermogravimetric analysis (TGA) measurements; and the electrochemical behaviors were examined in 2 M KOH solution. SEM and FTIR characterizations reveal that activated carbon was imbedded into the polymer membrane of polyethersulfone to form a uniform and flexible hybrid membrane. When the thin polymer-carbon membrane (PCM) was used as an electrode material for supercapacitor, a high specific capacitance of 169.4 Fg-1 was obtained at a current density of 0.5 Ag-1 along with good long-term cycle life of 94.6% capacity retention after 2000 charging-discharging cycles. Benefiting from these merits, the as-fabricated PCM//PCM cell shows an excellent electrochemical property. These results suggest a promising route towards the fabrication of highly flexible electrodes for high-performance supercapacitors.

  17. Application of Fluorescence In Situ Hybridization (FISH) Technique for the Detection of Genetic Aberration in Medical Science

    Science.gov (United States)

    Ratan, Zubair Ahmed; Zaman, Sojib Bin; Haidere, Mohammad Faisal; Runa, Nusrat Jahan; Akter, Nasrin

    2017-01-01

    Fluorescence in situ hybridization (FISH) is a macromolecule recognition technique, which is considered as a new advent in the field of cytology. Initially, it was developed as a physical mapping tool to delineate genes within chromosomes. The accuracy and versatility of FISH were subsequently capitalized upon in biological and medical research. This visually appealing technique provides an intermediate degree of resolution between DNA analysis and chromosomal investigations. FISH consists of a hybridizing DNA probe, which can be labeled directly or indirectly. In the case of direct labeling, fluorescent nucleotides are used, while indirect labeling is incorporated with reporter molecules that are subsequently detected by fluorescent antibodies or other affinity molecules. FISH is applied to detect genetic abnormalities that include different characteristic gene fusions or the presence of an abnormal number of chromosomes in a cell or loss of a chromosomal region or a whole chromosome. It is also applied in different research applications, such as gene mapping or the identification of novel oncogenes. This article reviews the concept of FISH, its application, and its advantages in medical science.  PMID:28690958

  18. Pre-implantation genetic screening using fluorescence in situ hybridization in couples of Indian ethnicity: Is there a scope?

    Directory of Open Access Journals (Sweden)

    Shailaja Gada Saxena

    2014-01-01

    Full Text Available Context: There is a high incidence of numerical chromosomal aberration in couples with repeated in vitro fertilization (IVF failure, advanced maternal age, repeated unexplained abortions, severe male factor infertility and unexplained infertility. Pre-implantation genetic screening (PGS, a variant of pre-implantation genetic diagnosis, screens numerical chromosomal aberrations in couples with normal karyotype, experiencing poor reproductive outcome. The present study includes the results of the initial pilot study on 9 couples who underwent 10 PGS cycles. Aim: The aim of the present study was to evaluate the beneficial effects of PGS in couples with poor reproductive outcome. Settings and Design: Data of initial 9 couples who underwent 10 PGS for various indications was evaluated. Subjects and Methods: Blastomere biopsy was performed on cleavage stage embryos and subjected to two round fluorescence in situ hybridization (FISH testing for chromosomes 13, 18, 21, X and Y as a two-step procedure. Results: Six of the 9 couples (10 PGS cycles conceived, including a twin pregnancy in a couple with male factor infertility, singleton pregnancies in a couple with secondary infertility, in three couples with adverse obstetric outcome in earlier pregnancies and in one couple with repeated IVF failure. Conclusion: In the absence of availability of array-comparative genomic hybridization in diagnostic clinical scenario for PGS and promising results with FISH based PGS as evident from the current pilot study, it is imperative to offer the best available services in the present scenario for better pregnancy outcome for patients.

  19. Fluorescence In Situ Hybridization with Peptide Nucleic Acid Probes for Rapid Identification of Candida albicans Directly from Blood Culture Bottles

    Science.gov (United States)

    Rigby, Susan; Procop, Gary W.; Haase, Gerhard; Wilson, Deborah; Hall, Geraldine; Kurtzman, Cletus; Oliveira, Kenneth; Von Oy, Sabina; Hyldig-Nielsen, Jens J.; Coull, James; Stender, Henrik

    2002-01-01

    A new fluorescence in situ hybridization (FISH) method that uses peptide nucleic acid (PNA) probes for identification of Candida albicans directly from positive-blood-culture bottles in which yeast was observed by Gram staining (herein referred to as yeast-positive blood culture bottles) is described. The test (the C. albicans PNA FISH method) is based on a fluorescein-labeled PNA probe that targets C. albicans 26S rRNA. The PNA probe is added to smears made directly from the contents of the blood culture bottle and hybridized for 90 min at 55°C. Unhybridized PNA probe is removed by washing of the mixture (30 min), and the smears are examined by fluorescence microscopy. The specificity of the method was confirmed with 23 reference strains representing phylogenetically related yeast species and 148 clinical isolates covering the clinically most significant yeast species, including C. albicans (n = 72), C. dubliniensis (n = 58), C. glabrata (n = 5), C. krusei (n = 2), C. parapsilosis (n = 4), and C. tropicalis (n = 3). The performance of the C. albicans PNA FISH method as a diagnostic test was evaluated with 33 routine and 25 simulated yeast-positive blood culture bottles and showed 100% sensitivity and 100% specificity. It is concluded that this 2.5-h method for the definitive identification of C. albicans directly from yeast-positive blood culture bottles provides important information for optimal antifungal therapy and patient management. PMID:12037084

  20. Morphological spot counting from stacked images for automated analysis of gene copy numbers by fluorescence in situ hybridization.

    Science.gov (United States)

    Grigoryan, Artyom M; Dougherty, Edward R; Kononen, Juha; Bubendorf, Lukas; Hostetter, Galen; Kallioniemi, Olli

    2002-01-01

    Fluorescence in situ hybridization (FISH) is a molecular diagnostic technique in which a fluorescent labeled probe hybridizes to a target nucleotide sequence of deoxyribose nucleic acid. Upon excitation, each chromosome containing the target sequence produces a fluorescent signal (spot). Because fluorescent spot counting is tedious and often subjective, automated digital algorithms to count spots are desirable. New technology provides a stack of images on multiple focal planes throughout a tissue sample. Multiple-focal-plane imaging helps overcome the biases and imprecision inherent in single-focal-plane methods. This paper proposes an algorithm for global spot counting in stacked three-dimensional slice FISH images without the necessity of nuclei segmentation. It is designed to work in complex backgrounds, when there are agglomerated nuclei, and in the presence of illumination gradients. It is based on the morphological top-hat transform, which locates intensity spikes on irregular backgrounds. After finding signals in the slice images, the algorithm groups these together to form three-dimensional spots. Filters are employed to separate legitimate spots from fluorescent noise. The algorithm is set in a comprehensive toolbox that provides visualization and analytic facilities. It includes simulation software that allows examination of algorithm performance for various image and algorithm parameter settings, including signal size, signal density, and the number of slices.

  1. Direct fluorescence in situ hybridization on human metaphase chromosomes using quantum dot-platinum labeled DNA probes

    Energy Technology Data Exchange (ETDEWEB)

    Hwang, Gyoyeon [Chemical Kinomics Research Center, Future Convergence Research Division, Korea Institute of Science and Technology, Hwarangno 14-gil 5, Seongbuk-gu, Seoul 136-791 (Korea, Republic of); Biological Chemistry, Korea University of Science and Technology, 217, Gajeong-ro, Yuseong-gu, Deajeon (Korea, Republic of); Lee, Hansol [Chemical Kinomics Research Center, Future Convergence Research Division, Korea Institute of Science and Technology, Hwarangno 14-gil 5, Seongbuk-gu, Seoul 136-791 (Korea, Republic of); Lee, Jiyeon, E-mail: jylee@kist.re.kr [Chemical Kinomics Research Center, Future Convergence Research Division, Korea Institute of Science and Technology, Hwarangno 14-gil 5, Seongbuk-gu, Seoul 136-791 (Korea, Republic of); Biological Chemistry, Korea University of Science and Technology, 217, Gajeong-ro, Yuseong-gu, Deajeon (Korea, Republic of)

    2015-11-13

    The telomere shortening in chromosomes implies the senescence, apoptosis, or oncogenic transformation of cells. Since detecting telomeres in aging and diseases like cancer, is important, the direct detection of telomeres has been a very useful biomarker. We propose a telomere detection method using a newly synthesized quantum dot (QD) based probe with oligonucleotide conjugation and direct fluorescence in situ hybridization (FISH). QD-oligonucleotides were prepared with metal coordination bonding based on platinum-guanine binding reported in our previous work. The QD-oligonucleotide conjugation method has an advantage where any sequence containing guanine at the end can be easily bound to the starting QD-Pt conjugate. A synthesized telomeric oligonucleotide was bound to the QD-Pt conjugate successfully and this probe hybridized specifically on the telomere of fabricated MV-4-11 and MOLT-4 chromosomes. Additionally, the QD-telomeric oligonucleotide probe successfully detected the telomeres on the CGH metaphase slide. Due to the excellent photostability and high quantum yield of QDs, the QD-oligonucleotide probe has high fluorescence intensity when compared to the organic dye-oligonucleotide probe. Our QD-oligonucleotide probe, conjugation method of this QD probe, and hybridization protocol with the chromosomes can be a useful tool for chromosome painting and FISH. - Highlights: • We prepared a probe linked between QD and telomeric oligonucleotide with platinum-guanine bonding. • Telomeres were detected by our new telomere probes successfully in three different human metaphase chromosomes. • QDPt-DNA probe has high fluorescence intensity in comparison with organic dye-DNA probe.

  2. Enhanced functional connectivity properties of human brains during in-situ nature experience

    Directory of Open Access Journals (Sweden)

    Zheng Chen

    2016-07-01

    Full Text Available In this study, we investigated the impacts of in-situ nature and urban exposure on human brain activities and their dynamics. We randomly assigned 32 healthy right-handed college students (mean age = 20.6 years, SD = 1.6; 16 males to a 20 min in-situ sitting exposure in either a nature (n = 16 or urban environment (n = 16 and measured their Electroencephalography (EEG signals. Analyses revealed that a brief in-situ restorative nature experience may induce more efficient and stronger brain connectivity with enhanced small-world properties compared with a stressful urban experience. The enhanced small-world properties were found to be correlated with “coherent” experience measured by Perceived Restorativeness Scale (PRS. Exposure to nature also induces stronger long-term correlated activity across different brain regions with a right lateralization. These findings may advance our understanding of the functional activities during in-situ environmental exposures and imply that a nature or nature-like environment may potentially benefit cognitive processes and mental well-being.

  3. Enhanced functional connectivity properties of human brains during in-situ nature experience.

    Science.gov (United States)

    Chen, Zheng; He, Yujia; Yu, Yuguo

    2016-01-01

    In this study, we investigated the impacts of in-situ nature and urban exposure on human brain activities and their dynamics. We randomly assigned 32 healthy right-handed college students (mean age = 20.6 years, SD = 1.6; 16 males) to a 20 min in-situ sitting exposure in either a nature (n = 16) or urban environment (n = 16) and measured their Electroencephalography (EEG) signals. Analyses revealed that a brief in-situ restorative nature experience may induce more efficient and stronger brain connectivity with enhanced small-world properties compared with a stressful urban experience. The enhanced small-world properties were found to be correlated with "coherent" experience measured by Perceived Restorativeness Scale (PRS). Exposure to nature also induces stronger long-term correlated activity across different brain regions with a right lateralization. These findings may advance our understanding of the functional activities during in-situ environmental exposures and imply that a nature or nature-like environment may potentially benefit cognitive processes and mental well-being.

  4. Automated brightfield dual-color in situ hybridization for detection of mouse double minute 2 gene amplification in sarcomas.

    Science.gov (United States)

    Zhang, Wenjun; McElhinny, Abigail; Nielsen, Alma; Wang, Maria; Miller, Melanie; Singh, Shalini; Rueger, Ruediger; Rubin, Brian P; Wang, Zhen; Tubbs, Raymond R; Nagle, Raymond B; Roche, Pat; Wu, Ping; Pestic-Dragovich, Lidija

    2011-01-01

    The human homolog of the mouse double minute 2 (MDM2) oncogene is amplified in about 20% of sarcomas. The measurement of the MDM2 amplification can aid in classification and may provide a predictive value for recently formulated therapies targeting MDM2. We have developed and validated an automated bright field dual-color in situ hybridization application to detect MDM2 gene amplification. A repeat-depleted MDM2 probe was constructed to target the MDM2 gene region at 12q15. A chromosome 12-specific probe (CHR12) was generated from a pα12H8 plasmid. The in situ hybridization assay was developed by using a dinitrophenyl-labeled MDM2 probe and a digoxigenin-labeled CHR12 probe on the Ventana Medical Systems' automated slide-staining platforms. The specificity of the MDM2 and CHR12 probes was shown on metaphase spreads and further validated against controls, including normal human tonsil and known MDM2-amplified samples. The assay performance was evaluated on a cohort of 100 formalin-fixed, paraffin-embedded specimens by using a conventional bright field microscope. Simultaneous hybridization and signal detection for MDM2 and CHR12 showed that both DNA targets were present in the same cells. One hundred soft tissue specimens were stained for MDM2 and CHR12. Although 26 of 29 lipomas were nonamplified and eusomic, MDM2 amplification was noted in 78% of atypical lipomatous tumors or well-differentiated liposarcomas. Five of 6 dedifferentiated liposarcoma cases were amplified for MDM2. MDM2 amplification was observed in 1 of 8 osteosarcomas; 3 showed CHR12 aneusomy. MDM2 amplification was present in 1 of 4 chondrosarcomas. Nine of 10 synovial sarcomas displayed no evidence of MDM2 amplification in most tumor cells. In pleomorphic sarcoma, not otherwise specified (pleomorphic malignant fibrous histiocytoma), MDM2 was amplified in 38% of cases, whereas 92% were aneusomic for CHR12. One alveolar rhabdomyosarcoma and 2 embryonal rhabdomyosarcomas displayed low-level aneusomy

  5. Establishment of a new human pleomorphic malignant fibrous histiocytoma cell line, FU-MFH-2: molecular cytogenetic characterization by multicolor fluorescence in situ hybridization and comparative genomic hybridization

    Directory of Open Access Journals (Sweden)

    Isayama Teruto

    2010-11-01

    Full Text Available Abstract Background Pleomorphic malignant fibrous histiocytoma (MFH is one of the most frequent malignant soft tissue tumors in adults. Despite the considerable amount of research on MFH cell lines, their characterization at a molecular cytogenetic level has not been extensively analyzed. Methods and results We established a new permanent human cell line, FU-MFH-2, from a metastatic pleomorphic MFH of a 72-year-old Japanese man, and applied multicolor fluorescence in situ hybridization (M-FISH, Urovysion™ FISH, and comparative genomic hybridization (CGH for the characterization of chromosomal aberrations. FU-MFH-2 cells were spindle or polygonal in shape with oval nuclei, and were successfully maintained in vitro for over 80 passages. The histological features of heterotransplanted tumors in severe combined immunodeficiency mice were essentially the same as those of the original tumor. Cytogenetic and M-FISH analyses displayed a hypotriploid karyotype with numerous structural aberrations. Urovysion™ FISH revealed a homozygous deletion of the p16INK4A locus on chromosome band 9p21. CGH analysis showed a high-level amplification of 9q31-q34, gains of 1p12-p34.3, 2p21, 2q11.2-q21, 3p, 4p, 6q22-qter, 8p11.2, 8q11.2-q21.1, 9q21-qter, 11q13, 12q24, 15q21-qter, 16p13, 17, 20, and X, and losses of 1q43-qter, 4q32-qter, 5q14-q23, 7q32-qter, 8p21-pter, 8q23, 9p21-pter, 10p11.2-p13, and 10q11.2-q22. Conclusion The FU-MFH-2 cell line will be a particularly useful model for studying molecular pathogenesis of human pleomorphic MFH.

  6. Principle, function, experiences, hybrid chilled ceilings; Prinzip, Funktion, Erfahrungen. Hybrid-Kuehldecken

    Energy Technology Data Exchange (ETDEWEB)

    Schulz, Uwe W. [Hochschule Luzern (Switzerland). Technik und Architektur

    2010-12-15

    Hybrid chilled ceilings combine a radiation chilled ceiling with thermal activations of a concrete ceiling. The activation permits the utilization of alternative and/or small cold generators for the efficient night cooling. The traditional chilled ceiling grants a speedy reaction to load variations as well as a reduction of the reverberation periods. In addition, the combination supplies a draught-free insertion of air. Beside established solutions, MWH Barcol-Air (Staefa, Switzerland) also offers project-specific hybrid chilled ceilings.

  7. Automated processing of fluorescence in-situ hybridization slides for HER2 testing in breast and gastro-esophageal carcinomas.

    Science.gov (United States)

    Tafe, Laura J; Allen, Samantha F; Steinmetz, Heather B; Dokus, Betty A; Cook, Leanne J; Marotti, Jonathan D; Tsongalis, Gregory J

    2014-08-01

    HER2 fluorescence in-situ hybridization (FISH) is used in breast and gastro-esophageal carcinoma for determining HER2 gene amplification and patients' eligibility for HER2 targeted therapeutics. Traditional manual processing of the FISH slides is labor intensive because of multiple steps that require hands on manipulation of the slides and specifically timed intervals between steps. This highly manual processing also introduces inter-run and inter-operator variability that may affect the quality of the FISH result. Therefore, we sought to incorporate an automated processing instrument into our FISH workflow. Twenty-six cases including breast (20) and gastro-esophageal (6) cancer comprising 23 biopsies and three excision specimens were tested for HER2 FISH (Pathvysion, Abbott) using the Thermobrite Elite (TBE) system (Leica). Up to 12 slides can be run simultaneously. All cases were previously tested by the Pathvysion HER2 FISH assay with manual preparation. Twenty cells were counted by two observers for each case; five cases were tested on three separate runs by different operators to evaluate the precision and inter-operator variability. There was 100% concordance in the scoring between the manual and TBE methods as well as among the five cases that were tested on three runs. Only one case failed due to poor probe hybridization. In total, seven cases were positive for HER2 amplification (HER2:CEP17 ratio >2.2) and the remaining 19 were negative (HER2:CEP17 ratio <1.8) utilizing the 2007 ASCO/CAP scoring criteria. Due to the automated denaturation and hybridization, for each run, there was a reduction in labor of 3.5h which could then be dedicated to other lab functions. The TBE is a walk away pre- and post-hybridization system that automates FISH slide processing, improves work flow and consistency and saves approximately 3.5h of technologist time. The instrument has a small footprint thus occupying minimal counter space. TBE processed slides performed

  8. A new adaptive hybrid electromagnetic damper: modelling, optimization, and experiment

    International Nuclear Information System (INIS)

    Asadi, Ehsan; Ribeiro, Roberto; Behrad Khamesee, Mir; Khajepour, Amir

    2015-01-01

    This paper presents the development of a new electromagnetic hybrid damper which provides regenerative adaptive damping force for various applications. Recently, the introduction of electromagnetic technologies to the damping systems has provided researchers with new opportunities for the realization of adaptive semi-active damping systems with the added benefit of energy recovery. In this research, a hybrid electromagnetic damper is proposed. The hybrid damper is configured to operate with viscous and electromagnetic subsystems. The viscous medium provides a bias and fail-safe damping force while the electromagnetic component adds adaptability and the capacity for regeneration to the hybrid design. The electromagnetic component is modeled and analyzed using analytical (lumped equivalent magnetic circuit) and electromagnetic finite element method (FEM) (COMSOL ® software package) approaches. By implementing both modeling approaches, an optimization for the geometric aspects of the electromagnetic subsystem is obtained. Based on the proposed electromagnetic hybrid damping concept and the preliminary optimization solution, a prototype is designed and fabricated. A good agreement is observed between the experimental and FEM results for the magnetic field distribution and electromagnetic damping forces. These results validate the accuracy of the modeling approach and the preliminary optimization solution. An analytical model is also presented for viscous damping force, and is compared with experimental results The results show that the damper is able to produce damping coefficients of 1300 and 0–238 N s m −1 through the viscous and electromagnetic components, respectively. (paper)

  9. In situ observation of a soap-film catenoid-a simple educational physics experiment

    International Nuclear Information System (INIS)

    Ito, Masato; Sato, Taku

    2010-01-01

    The solution to the Euler-Lagrange equation is an extremal functional. To understand that the functional is stationary at local extrema (maxima or minima), we propose a physics experiment that involves using a soap film to form a catenoid. A catenoid is a surface that is formed between two coaxial circular rings and is classified mathematically as a minimal surface. Using the soap film, we create catenoids between two rings and characterize the catenoid in situ while varying the distance between the rings. The shape of the soap film is very interesting and can be explained using dynamic mechanics. By observing the catenoid, physics students can observe local extrema phenomena. We stress that in situ observation of soap-film catenoids is an appropriate physics experiment that combines theory and experimentation.

  10. In situ brine migration experiments at the Avery Island salt mine

    International Nuclear Information System (INIS)

    Krause, W.B.; Van Sambeek, L.L.; Stickney, R.G.

    1980-01-01

    An in situ brine movement study was conducted at the Avery Island Salt Mine of the International Salt Company in southwestern Louisiana. The objective of the in situ experiments was to relate field measurements to previously determined laboratory and analytical results for the purpose of determining the rate and amount of brine movement through dome salt when subjected to heating. The heating in the experiments was provided by electrical heaters emplaced in the salt mine floor. An understanding of thermally induced brine movement is essential from the standpoint of identifying conditions which may influence the physical integrity of the nuclear waste canisters or impede the functional performance of the waste package system in a nuclear waste repository in geologic salt. 28 refs

  11. Annual report for fiscal 1995, Kamaishi in-situ experiments (phase 2)

    Energy Technology Data Exchange (ETDEWEB)

    Aoki, Kazuhiro; Araki, Ryusuke; Koide, Kaoru; Sawada, Atsushi; Shimizu, Isao; Fujita, Asao; Yoshida, Eiichi [Power Reactor and Nuclear Fuel Development Corp., Tokyo (Japan)

    1996-04-01

    The Kamaishi in-situ experiments (Phase 2) have strived to ascertain geological characteristics of the deep underground and the various phenomenon occurring therein and to improve technologies and methodologies required for such studies since fiscal 1993. Fiscal 1995 is the third year of Phase 2. The in-situ experiments are conducted at the northern most end of the Kamaishi mine in order to minimize the effect of the already excavated drifts totaling approximately 140 km long. The studies are conducted in Kurihashi granodiorite of Early Cretaceous widely distributed in this area. Major activities performed in this fiscal year are summarized below: (1) TASK 1 (Characterization of the deep underground geological environment). (2) TASK 2 (Study of excavation disturbance in fractured rock). (3) TASK 3 (Study of groundwater flow and solute transport in crystalline rock). (4) TASK 4 (Study of engineered barrier). (5) TASK 5 (Study of earthquakes). (J.P.N.)

  12. In situ observation of a soap-film catenoid-a simple educational physics experiment

    Energy Technology Data Exchange (ETDEWEB)

    Ito, Masato; Sato, Taku [Department of Physics, Aichi University of Education, Kariya, 448-8542 (Japan)], E-mail: mito@auecc.aichi-edu.ac.jp

    2010-03-15

    The solution to the Euler-Lagrange equation is an extremal functional. To understand that the functional is stationary at local extrema (maxima or minima), we propose a physics experiment that involves using a soap film to form a catenoid. A catenoid is a surface that is formed between two coaxial circular rings and is classified mathematically as a minimal surface. Using the soap film, we create catenoids between two rings and characterize the catenoid in situ while varying the distance between the rings. The shape of the soap film is very interesting and can be explained using dynamic mechanics. By observing the catenoid, physics students can observe local extrema phenomena. We stress that in situ observation of soap-film catenoids is an appropriate physics experiment that combines theory and experimentati0008.

  13. Rapid detection of rRNA group I pseudomonads in contaminated metalworking fluids and biofilm formation by fluorescent in situ hybridization.

    Science.gov (United States)

    Saha, Ratul; Donofrio, Robert S; Goeres, Darla M; Bagley, Susan T

    2012-05-01

    Metalworking fluids (MWFs), used in different machining operations, are highly prone to microbial degradation. Microbial communities present in MWFs lead to biofilm formation in the MWF systems, which act as a continuous source of contamination. Species of rRNA group I Pseudomonas dominate in contaminated MWFs. However, their actual distribution is typically underestimated when using standard culturing techniques as most fail to grow on the commonly used Pseudomonas Isolation Agar. To overcome this, fluorescent in situ hybridization (FISH) was used to study their abundance along with biofilm formation by two species recovered from MWFs, Pseudomonas fluorescens MWF-1 and the newly described Pseudomonas oleovorans subsp. lubricantis. Based on 16S rRNA sequences, a unique fluorescent molecular probe (Pseudo120) was designed targeting a conserved signature sequence common to all rRNA group I Pseudomonas. The specificity of the probe was evaluated using hybridization experiments with whole cells of different Pseudomonas species. The probe's sensitivity was determined to be 10(3) cells/ml. It successfully detected and enumerated the abundance and distribution of Pseudomonas indicating levels between 3.2 (± 1.1) × 10(6) and 5.0 (± 2.3) × 10(6) cells/ml in four different industrial MWF samples collected from three different locations. Biofilm formation was visualized under stagnant conditions using high and low concentrations of cells for both P. fluorescens MWF-1 and P. oleovorans subsp. lubricantis stained with methylene blue and Pseudo120. On the basis of these observations, this molecular probe can be successfully be used in the management of MWF systems to monitor the levels and biofilm formation of rRNA group I pseudomonads.

  14. Dual color chromogenic in situ hybridization for determination of HER2 status in breast cancer: a large comparative study to current state of the art fluorescence in situ hybridization

    Directory of Open Access Journals (Sweden)

    Mollerup Jens

    2012-02-01

    Full Text Available Abstract Background Chromogenic in situ hybridization (CISH is fast becoming a well established technique for easy and sensitive determination of HER2 gene status in breast cancer. However, for the chromogenic method to achieve status as a safe and reliable technique, the method needs to be validated against already known and validated FISH techniques. Methods Here it is reported from a comparative study where HER2 gene status obtained by HER2 CISH pharmDx™ Kit was compared to HER2 gene status obtained by the FDA approved HER2 FISH pharmDx™ Kit and the PathVysion HER-2 DNA probe Kit. The study included 365 formalin fixed and paraffin-embedded invasive breast cancer tissue specimens collected consecutively at a US reference laboratory. Results The data obtained revealed an overall HER2 status concordance of approximately 98% for comparisons of HER2 CISH pharmDx™ Kit to both HER2 FISH pharmDx™ Kit and PathVysion HER-2 DNA Probe Kit. Conclusions The concordance between results obtained using the recently FDA approved HER2 CISH pharmDx™ Kit with previously FDA approved FISH techniques for HER2 gene status determination indicate that the HER2 CISH pharmDx™ Kit is a reliable chromogenic alternative to fluorescence-based methods.

  15. Dual color chromogenic in situ hybridization for determination of HER2 status in breast cancer: a large comparative study to current state of the art fluorescence in situ hybridization

    Science.gov (United States)

    2012-01-01

    Background Chromogenic in situ hybridization (CISH) is fast becoming a well established technique for easy and sensitive determination of HER2 gene status in breast cancer. However, for the chromogenic method to achieve status as a safe and reliable technique, the method needs to be validated against already known and validated FISH techniques. Methods Here it is reported from a comparative study where HER2 gene status obtained by HER2 CISH pharmDx™ Kit was compared to HER2 gene status obtained by the FDA approved HER2 FISH pharmDx™ Kit and the PathVysion HER-2 DNA probe Kit. The study included 365 formalin fixed and paraffin-embedded invasive breast cancer tissue specimens collected consecutively at a US reference laboratory. Results The data obtained revealed an overall HER2 status concordance of approximately 98% for comparisons of HER2 CISH pharmDx™ Kit to both HER2 FISH pharmDx™ Kit and PathVysion HER-2 DNA Probe Kit. Conclusions The concordance between results obtained using the recently FDA approved HER2 CISH pharmDx™ Kit with previously FDA approved FISH techniques for HER2 gene status determination indicate that the HER2 CISH pharmDx™ Kit is a reliable chromogenic alternative to fluorescence-based methods. PMID:22333181

  16. Feasibility of mapping low-multiplicity genes by in situ hybridization. [/sup 125/I and tritium tracers

    Energy Technology Data Exchange (ETDEWEB)

    Atwood, K C; Henderson, A S; Kacian, D; Eicher, E M

    1975-01-01

    The assignment of human hemoglobin loci to 2q and Bq was followed by objections based on the low specific activity of their /sup 3/H-mRNA, estimated as somewhere between 100 and 1000 dpm/..mu..g. With that preparation, the time required for one disintegration per molecule is between 80,000 and 8000 years. The consideration that globin loci may contain as many as 10 copies does not help. In view of these incontestably valid objections, it is instructive to compare the foregoing experiment with another in which the chromosomes were hybridized with cDNA copied from human reticulocyte mRNA by means of the reverse transcriptase from avian myeloblastosis virus. Our preparation had a specific activity of 1.4 x 10/sup 8/ dpm/..mu..g, requiring less than a month per disintegration per molecule. Despite the rather more favorable anticipated exposure time, successful localization was by no means expected. The use of cDNA precluded the enzymatic removal of background; the concentration of DNA applied to the slides, 0.03 ..mu..g/ml, was unfavorable for hybridization; and the copying reaction, primed with oligo-dT, probably attached stretches up to T/sub 20/ to the structural sequence, with the possibility that these might hybridize to uninteresting chromosomal regions. Results indicate that the original assignments of the human globin loci are correct.

  17. In-Situ Visualization Experiments with ParaView Cinema in RAGE

    Energy Technology Data Exchange (ETDEWEB)

    Kares, Robert John [Los Alamos National Lab. (LANL), Los Alamos, NM (United States)

    2015-10-15

    A previous paper described some numerical experiments performed using the ParaView/Catalyst in-situ visualization infrastructure deployed in the Los Alamos RAGE radiation-hydrodynamics code to produce images from a running large scale 3D ICF simulation. One challenge of the in-situ approach apparent in these experiments was the difficulty of choosing parameters likes isosurface values for the visualizations to be produced from the running simulation without the benefit of prior knowledge of the simulation results and the resultant cost of recomputing in-situ generated images when parameters are chosen suboptimally. A proposed method of addressing this difficulty is to simply render multiple images at runtime with a range of possible parameter values to produce a large database of images and to provide the user with a tool for managing the resulting database of imagery. Recently, ParaView/Catalyst has been extended to include such a capability via the so-called Cinema framework. Here I describe some initial experiments with the first delivery of Cinema and make some recommendations for future extensions of Cinema’s capabilities.

  18. Chromogenic in situ Hybridization Compared with Real time Quantitative Polymerase Chain Reaction to Evaluate HER2/neu Status in Breast Cancer.

    Science.gov (United States)

    Ayatollahi, Hossein; Fani, Azar; Ghayoor Karimiani, Ehsan; Homaee, Fateme; Shajiei, Arezoo; Sheikh, Maryam; Shakeri, Sepideh; Shams, Seyyede Fatemeh

    2017-01-01

    The assessment of human epidermal growth factor receptor 2 (HER2) status has become of great importance in the diagnosis of breast cancer. The aim of this study was to investigate the diagnostic value of quantitative Polymerase Chain Reaction (qPCR) and Chromogenic In Situ Hybridization (CISH) to assess HER2 status of biopsy specimens. To elucidate the status of HER2 gene amplification, biopsies of breast carcinoma from 120 patients with 2+ IHC status were analyzed by qPCR and CISH. The results of the two experiments were compared, and it was depicted that the concordance rate between CISH and qPCR assays was 88.1%.The quantification of HER2 gene with CISH and qPCR showed that there was a significant correlation (p value= 0.0001 and r= 0.808). The results of this research support the idea that qPCR is a precise and reproducible technique, which can be employed as a supplementary method to evaluate HER2 status.

  19. Mercury's plasma belt: hybrid simulations results compared to in-situ measurements

    Science.gov (United States)

    Hercik, D.; Travnicek, P. M.; Schriver, D.; Hellinger, P.

    2012-12-01

    The presence of plasma belt and trapped particles region in the Mercury's inner magnetosphere has been questionable due to small dimensions of the magnetosphere of Mercury compared to Earth, where these regions are formed. Numerical simulations of the solar wind interaction with Mercury's magnetic field suggested that such a structure could be found also in the vicinity of Mercury. These results has been recently confirmed also by MESSENGER observations. Here we present more detailed analysis of the plasma belt structure and quasi-trapped particle population characteristics and behaviour under different orientations of the interplanetary magnetic field.The plasma belt region is constantly supplied with solar wind protons via magnetospheric flanks and tail current sheet region. Protons inside the plasma belt region are quasi-trapped in the magnetic field of Mercury and perform westward drift along the planet. This region is well separated by a magnetic shell and has higher average temperatures and lower bulk proton current densities than surrounding area. On the day side the population exhibits loss cone distribution function matching the theoretical loss cone angle. Simulations results are also compared to in-situ measurements acquired by MESSENGER MAG and FIPS instruments.

  20. In Situ Caging of Biomolecules in Graphene Hybrids for Light Modulated Bioactivity.

    Science.gov (United States)

    Cheng, Gong; Han, Xiao-Hui; Hao, Si-Jie; Nisic, Merisa; Zheng, Si-Yang

    2018-01-31

    Remote and noninvasive modulation of protein activity is essential for applications in biotechnology and medicine. Optical control has emerged as the most attractive approach owing to its high spatial and temporal resolutions; however, it is challenging to engineer light responsive proteins. In this work, a near-infrared (NIR) light-responsive graphene-silica-trypsin (GST) nanoreactor is developed for modulating the bioactivity of trypsin molecules. Biomolecules are spatially confined and protected in the rationally designed compartment architecture, which not only reduces the possible interference but also boosts the bioreaction efficiency. Upon NIR irradiation, the photothermal effect of the GST nanoreactor enables the ultrafast in situ heating for remote activation and tuning of the bioactivity. We apply the GST nanoreactor for remote and ultrafast proteolysis of proteins, which remarkably enhances the proteolysis efficiency and reduces the bioreaction time from the overnight of using free trypsin to seconds. We envision that this work not only provides a promising tool of ultrafast and remotely controllable proteolysis for in vivo proteomics in study of tissue microenvironment and other biomedical applications but also paves the way for exploring smart artificial nanoreactors in biomolecular modulation to gain insight in dynamic biological transformation.

  1. The ISHTE [In-Situ Heat Transfer Experiment] lander: Final report

    International Nuclear Information System (INIS)

    Olson, L.O.; Harrison, J.G.

    1986-12-01

    This report describes the design and development of a sea floor lander constructed to support the In-Situ Heat Transfer Experiment (ISHTE). The work entailed fabricating and testing a steel space frame that would support and accurately position delicate instruments which would monitor a heat source driven into the sediments of the deep ocean. This lander is capable of being (1) transported from Seattle to Hawaii and back several times; (2) deployed from a ship at sea; (3) operated on the sea floor to field delicate experimental equipment; and (4) recovered for retrofit to support a one-year experiment on the sea floor

  2. In Situ Hybridization Method Reveals (Pro)renin Receptor Expressing Cells in the Pituitary Gland of Rats: Correlation with Anterior Pituitary Hormones.

    Science.gov (United States)

    Takahashi, Kazuhiro; Yatabe, Megumi; Fujiwara, Ken; Hirose, Takuo; Totsune, Kazuhito; Yashiro, Takashi

    2013-02-28

    Expression of (pro)renin receptor ((P)RR), a specific receptor for renin and prorenin, was studied in rat pituitary gland. In situ hybridization showed that cells expressing (P)RR mRNA were widely distributed in the anterior lobe and intermediate lobe of the pituitary gland. Double-staining using in situ hybridization for (P)RR mRNA and immunohistochemistry for the pituitary hormones showed that (P)RR mRNA was expressed in most of the GH cells and ACTH cells in the anterior lobe. (P)RR mRNA was also expressed in a few prolactin cells and TSH cells, but not in LH cells. The present study has shown for the first time the distribution of (P)RR mRNA expressing cells in the rat pituitary gland. These findings suggest that (P)RR plays physiological roles in the pituitary gland, such as the modulation of the pituitary hormone secretion.

  3. In Situ Hybridization Method Reveals (Pro)renin Receptor Expressing Cells in the Pituitary Gland of Rats: Correlation with Anterior Pituitary Hormones

    International Nuclear Information System (INIS)

    Takahashi, Kazuhiro; Yatabe, Megumi; Fujiwara, Ken; Hirose, Takuo; Totsune, Kazuhito; Yashiro, Takashi

    2013-01-01

    Expression of (pro)renin receptor ((P)RR), a specific receptor for renin and prorenin, was studied in rat pituitary gland. In situ hybridization showed that cells expressing (P)RR mRNA were widely distributed in the anterior lobe and intermediate lobe of the pituitary gland. Double-staining using in situ hybridization for (P)RR mRNA and immunohistochemistry for the pituitary hormones showed that (P)RR mRNA was expressed in most of the GH cells and ACTH cells in the anterior lobe. (P)RR mRNA was also expressed in a few prolactin cells and TSH cells, but not in LH cells. The present study has shown for the first time the distribution of (P)RR mRNA expressing cells in the rat pituitary gland. These findings suggest that (P)RR plays physiological roles in the pituitary gland, such as the modulation of the pituitary hormone secretion

  4. Human cDNA mapping using fluorescence in situ hybridization. Progress report, April 1--December 31, 1992

    Energy Technology Data Exchange (ETDEWEB)

    Korenberg, J.R.

    1993-12-31

    The ultimate goal of this proposal is to create a cDNA map of the human genome. Mapping is approached using the techniques of high resolution fluorescence in situ hybridization (FISH). This technology and the results of its application are designed to rapidly generate whole genome as tool box of expressed sequence to speed the identification of human disease genes. The results of this study are intended to dovetail with and to link the results of existing technologies for creating backbone YAC and genetic maps. In the first eight months, this approach will generate 60--80% of the expressed sequence map, the remainder expected to be derived through more long-term, labor-intensive, regional chromosomal gene searches or sequencing. The laboratory has made significant progress in the set-up phase, in mapping fetal and adult brain and other cDNAs, in testing a model system for directly linking genetic and physical maps using FISH with small fragments, in setting up a database, and in establishing the validity and throughput of the system.

  5. Development of a biological dosimeter for translocation scoring based on two-color fluorescence in situ hybridization of chromosome subsets

    Energy Technology Data Exchange (ETDEWEB)

    Popp, S; Cremer, T [Heidelberg Univ. (Germany). Inst. of Human Genetics and Anthropology

    1992-03-01

    Recently fluorescence in situ hybridization protocols have been developed which allow the paining of individual chromosomes using DNA-libraries from sorted human chromosomes. This approach has the particular advantage that radiation induced chromosome translocations can be easily detected, if chromosomes of distinctly different colors take part in the translocation event. To enhance the sensitivity of this approach two metaphase chromosome subsets A and B (A: chromosome 1, 2, 4, 8, 16; B: 3, 5, 9, 10, 13) were simultaneously painted in green and red color. Counterstaining of the chromosomes with DAPI resulted in a third subset which exhibited blue fluorescence only. Green-red, green-blue and red-blue translocation chromosomes could be easily detected after irradiation of lymphocyte cultures with {sup 137}Cs-{gamma}-rays. Analyses of painted chromosomes can be combined with conventional GTG-banding analyses. This new biological dosimeter should become useful to monitor both long term effects of single irradiation events and the cumulative effects of multiple or chronic irradiation exposure. In contrast to translocation scoring based on the analysis of banded chromosomes, this new approach has the particular advantage that a rapid, automated scoring of translocations can now be envisaged. (author).

  6. Hybrid Electrodes by In-Situ Integration of Graphene and Carbon-Nanotubes in Polypyrrole for Supercapacitors

    Science.gov (United States)

    Aphale, Ashish; Maisuria, Krushangi; Mahapatra, Manoj K.; Santiago, Angela; Singh, Prabhakar; Patra, Prabir

    2015-09-01

    Supercapacitors also known as electrochemical capacitors, that store energy via either Faradaic or non-Faradaic processes, have recently grown popularity mainly because they complement, and can even replace, conventional energy storage systems in variety of applications. Supercapacitor performance can be improved significantly by developing new nanocomposite electrodes which utilizes both the energy storage processes simultaneously. Here we report, fabrication of the freestanding hybrid electrodes, by incorporating graphene and carbon nanotubes (CNT) in pyrrole monomer via its in-situ polymerization. At the scan rate of 5 mV s-1, the specific capacitance of the polypyrrole-CNT-graphene (PCG) electrode film was 453 F g-1 with ultrahigh energy and power density of 62.96 W h kg-1 and 566.66 W kg-1 respectively, as shown in the Ragone plot. A nanofibrous membrane was electrospun and effectively used as a separator in the supercapacitor. Four supercapacitors were assembled in series to demonstrate the device performance by lighting a 2.2 V LED.

  7. Immunohistochemical her-2/ neu expression with gene amplification by fluorescence in situ hybridization for assessment in breast carcinomas

    International Nuclear Information System (INIS)

    Moatter, T.; Zahida, Z.U.D.; Kayani, N.; Pervez, S.

    2007-01-01

    To compare gene amplification of HER-2/neu gene by fluorescence in situ hybridization (FISH) in moderate to strong immunohistochemically (IHS) positive HER-2/neu cases of invasive breast carcinomas. Forty one (41) diagnosed cases of invasive breast carcinomas were included in this study in which already determined immunohistochemical HER-2/neu expression was scored as either 2+ or 3+, based on the intensity of membranous staining. These cases were further evaluated for gene amplification by FISH. For gene amplification, a ratio of HER-2/CEP z 2 was accepted as positive gene amplification. Out of a total 41 cases, which were scored as 2+ and 3+ by IHC, 14 cases (34.1%, 95% confidence interval: 19% - 49.3% ) showed gene amplification by FISH. Proportion of FISH positivity in IHC 2+ cases alone was found to be 25% (95% confidence interval: 10.5% - 41%). In contrast, a majority of IHC 3+ cases (5 of 6) were positive by FISH studies. IHC is appropriate for initial HER-2/neu assessment and patients with tumors scored as 3+ may be treated alone based on this information provided strict quality control and 95% concordance with FISH assays; however, patients with tumors interpreted as 2+, would benefit from gene amplification by FISH studies for more accurate assessment to avoid inaccurate prognostication and treatment. (author)

  8. induced chromosome aberrations analyzed by fluorescence in situ hybridization. Eight years follow up of the Goiania radiation accident victims

    International Nuclear Information System (INIS)

    Natarajan, A.T.; Santos, S.J.; Darroudi, F.; Hadjidikova, V.; Vermeulen, S.; Chatterjee, S.; Van de Berg, M.; Grigorova, M.; Sakamoto-Hojo, E.T.; Granath, F.; Ramalho, A.T.; Curado, M.P.

    1998-01-01

    The radiation accident in focus here occurred in a section of Goiania (Brazil) where more than a hundred individuals were contaminated with on September 1987. In order to estimate the absorbed radiation doses, initial frequencies of dicentrics and rings were determined in 129 victims [A.T. Ramalho, PhD Thesis, Subsidios a tecnica de dosimetria citogenetica gerados a partir da analise de resultados obtidos com o acidente radiologico de Goiania, Universidade Federal do Rio de Janeiro, Rio de Janeiro, Brazil, 1992]. We have followed some of these victims cytogenetically over the years seeking for parameters that could be used as basis for retrospective radiation dosimetry. Our data on translocation frequencies obtained by fluorescence in situ hybridization (FISH) could be directly compared to the baseline frequencies of dicentrics available for those same victims. Our results provided valuable information on how precise these estimates are. The frequencies of translocations observed years after the radiation exposure were two to three times lower than the initial dicentrics frequencies, the differences being larger at higher doses (>1 Gy). The accuracy of such dose estimates might be increased by scoring sufficient amount of cells. However, factors such as the persistence of translocation carrying lymphocytes, translocation levels not proportional to chromosome size, and inter-individual variation reduce the precision of these estimates

  9. Induced chromosome aberrations analyzed by fluorescence in situ hybridization. Eight years follow up of the Goiania radiation accident victims

    Energy Technology Data Exchange (ETDEWEB)

    Natarajan, A.T.; Santos, S.J.; Darroudi, F.; Hadjidikova, V.; Vermeulen, S.; Chatterjee, S.; Van de Berg, M.; Grigorova, M. [Leiden University Medical Centrum LUMC, Department of Radiation Genetics and Chemical Mutagenesis, Wassenaarseweg 72, 2333 AL Leiden (Netherlands); Sakamoto-Hojo, E.T. [Department of Biology, Faculty of Philosophy, Sciences and Letters of Ribeirao Preto, University of Sao Paulo, Ribeirao Preto (Brazil); Granath, F. [Department of Mathematical Statistics, Stockholm University, Stockholm (Sweden); Ramalho, A.T. [Institute of Radioprotection and Dosimetry, National Commission of Nuclear Energy, Rio de Janeiro (Brazil); Curado, M.P. [Foundation Leide das Neves Ferreira, Goiania (Brazil)

    1998-05-25

    The radiation accident in focus here occurred in a section of Goiania (Brazil) where more than a hundred individuals were contaminated with on September 1987. In order to estimate the absorbed radiation doses, initial frequencies of dicentrics and rings were determined in 129 victims [A.T. Ramalho, PhD Thesis, Subsidios a tecnica de dosimetria citogenetica gerados a partir da analise de resultados obtidos com o acidente radiologico de Goiania, Universidade Federal do Rio de Janeiro, Rio de Janeiro, Brazil, 1992]. We have followed some of these victims cytogenetically over the years seeking for parameters that could be used as basis for retrospective radiation dosimetry. Our data on translocation frequencies obtained by fluorescence in situ hybridization (FISH) could be directly compared to the baseline frequencies of dicentrics available for those same victims. Our results provided valuable information on how precise these estimates are. The frequencies of translocations observed years after the radiation exposure were two to three times lower than the initial dicentrics frequencies, the differences being larger at higher doses (>1 Gy). The accuracy of such dose estimates might be increased by scoring sufficient amount of cells. However, factors such as the persistence of translocation carrying lymphocytes, translocation levels not proportional to chromosome size, and inter-individual variation reduce the precision of these estimates

  10. Feasibility of using fluorescence in situ hybridization (FISH) to detect early gene changes in sputum cells from uranium miners

    Energy Technology Data Exchange (ETDEWEB)

    Neft, R.E.; Rogers, J.L.; Belinsky, S.A. [and others

    1995-12-01

    Epidemiological studies have shown that combined exposure to radon progeny and tobacco smoke produce a greater than additive or synergistic increase in lung cancer risk. Lung cancer results from multiple genetic changes over a long period of time. An early change that occurs in lung cancer is trisomy 7 which is found in 50% of non-small cell lung cancer and in the far margins of resected lung tumors. The 80% mortality associated with lung cancer is in part related to the high proportion of patients who present with an advanced, unresectable tumor. Therefore, early detection of patients at risk for tumor development is critical to improve treatment of this disease. Currently, it is difficult to detect lung cancer early while it is still amendable by surgery. Saccomanno, G. has shown that premalignant cytologic changes in sputum cells collected from uranium miners can be detected by a skilled, highly trained cytopathologist. A more objective alternative for identifying premalignant cells in sputum may be to determine whether an early genetic change such as trisomy 7 is present in these cells. Fluorescence in situ hybridization (FISH) can be used to identify cells with trisomy 7. The results of this investigation indicate that FISH may prove to be an accurate, efficient method to test at-risk individuals for genetic alterations in bronchial epithelial cells from sputum.

  11. Detection of MYCN Gene Amplification in Neuroblastoma by Fluorescence In Situ Hybridization: A Pediatric Oncology Group Study

    Directory of Open Access Journals (Sweden)

    Prasad Mathew

    2001-01-01

    Full Text Available To assess the utility of fluorescence in situ hybridization (FISH for analysis of MYCN gene amplification in neuroblastoma, we compared this assay with Southern blot analysis using tumor specimens collected from 232 patients with presenting characteristics typical of this disease. The FISH technique identified MYCN amplification in 47 cases, compared with 39 by Southern blotting, thus increasing the total number of positive cases by 21%. The major cause of discordancy was a low fraction of tumor cells (≤30% replacement in clinical specimens, which prevented an accurate estimate of MYCN copy number by Southern blotting. With FISH, by contrast, it was possible to analyze multiple interphase nuclei of tumor cells, regardless of the proportion of normal peripheral blood, bone marrow, or stromal cells in clinical samples. Thus, FISH could be performed accurately with very small numbers of tumor cells from touch preparations of needle biopsies. Moreover, this procedure allowed us to discern the heterogeneous pattern of MYCN amplification that is characteristic of neuroblastoma. We conclude that FISH improves the detection of MYCN gene amplification in childhood neuroblastomas in a clinical setting, thus facilitating therapeutic decisions based on the presence or absence of this prognostically important biologic marker.

  12. Detection of chromosomal aberrations by fluorescence in situ hybridization in the first three postirradiation divisions of human lymphocytes

    International Nuclear Information System (INIS)

    Boei, J.J.W.A.; Vermeulen, S.; Natarajan, A.T.

    1996-01-01

    Chromosomal aberrations in human lymphocytes were analyzed by fluorescence in situ hybridization (FISH) in the first 3 postirradiation (0 and 2 Gy) divisions. Cells were grown in the presence of BrdU, collected at different sampling times (47, 70 and 91 h) and analyzed using an alphoid centromeric probe and PCR amplified DNA libraries for chromosomes 2 and 8. Following differential staining of sister chromatids, the analyzed cells were identified to be either in the first, second or third mitosis after irradiation. The frequencies of both dicentrics and fragments showed a reduction of about 50% after each cell generation, whereas translocations were more persistent. Cells within the same postirradiation division showed higher aberration frequencies when derived from later sampling times, indicating a delay in progression of aberrant cells. As a result, the frequencies for dicentrics and fragments remained rather constant at different sampling times if the cell cycle parameter was not taken into account. Thus, the average generation time of the lymphocytes had a clear effect on the obtained aberration frequencies. The described method allows the study of the persistence of chromosome damage using the FISH technique during 3 subsequent cell divisions in vitro

  13. Detection of Epstein Barr Virus by Chromogenic In Situ Hybridization in cases of extra-hepatic biliary atresia

    Directory of Open Access Journals (Sweden)

    Farahmand Fatemeh

    2008-04-01

    Full Text Available Abstract Introduction Extra-hepatic biliary atresia (EHBA is an important cause of neonatal cholestasis. Several infectious agents have been proposed as etiologic factors such as Rotavirus and Reovirus. There is limited data on the role of Epstein Barr virus (EBV infection in EHBA, so we decided to study the presence of EBV virus in a series of 16 proven EHBA cases by Chromogenic in situ hybridization (CISH technique. Methods In the current study a total of 16 liver wedge biopsies of proven cases of EHBA were selected in a period of 4 years. CISH staining for EBV-encoded RNA (EBER transcript was performed. Results The review of H&E-stained slides of liver biopsies revealed fibrosis and marked ductular proliferation. In CISH-stained slides, EBV trace was observed in hepatocytes in two cases and in biliary epithelium in one case of EHBA. Discussion Considering the association of hepatitis with the Epstein-Barr virus in later life, it is likely that EBV hepatitis and its complications occur in the neonatal/perinatal period. Since EHBA is a relatively rare disease, a similar study on wedge biopsies of this number of proven cases of EHBA has not been performed to date. Current observation proposes the need for a study of larger series and employing other methods for confirming the etiologic role of EBV in EHBA cases.

  14. Gain of chromosome 7 by chromogenic in situ hybridization (CISH) in chordomas is correlated to c-MET expression.

    Science.gov (United States)

    Walter, Beatriz A; Begnami, Maria; Valera, Vladimir A; Santi, Mariarita; Rushing, Elisabeth J; Quezado, Martha

    2011-01-01

    Chordomas are low to intermediate grade malignancies that arise from remnants of embryonic notochord. They often recur after surgery and are highly resistant to conventional adjuvant therapies. Recently, the development of effective targeted molecular therapy has been investigated in chordomas that show receptors for tyrosine kinase (RTKs) activation. Expression of specific RTKs such as Epidermal Growth Factor Receptor (EGFR) and Mesenchymal-epithelial transition factor (c-MET) in chordomas may offer valuable therapeutic options. We investigated changes in copy number of chromosome 7 and correlated it with EGFR gene status and EGFR and c-MET protein expression in 22 chordoma samples. Chromosome 7 copy number was evaluated by chromogenic in situ hybridization (CISH) and protein expression of EGFR and c-MET by immunohistochemistry. Tumors mostly showed conventional histopathologic features and were found mainly in sacral (41%) and cranial sites (54.5%). Aneusomy of chromosome 7 was seen in 73% of the samples, 62% of primary tumors and in all recurrent chordomas. EGFR and c-MET were both expressed, but only c-MET protein expression was significantly correlated with chromosome 7 aneusomy (P ≤ 0.001). c-MET overexpression may represent an early chromosome 7 alteration that could play an important role during chordoma pathogenesis. c-MET overexpression shows promise as a molecular marker of response to targeted molecular therapy in the treatment of chordomas.

  15. Prognostic value of HER2 gene amplification detected by chromogenic in situ hybridization (CISH) in metastatic breast cancer.

    Science.gov (United States)

    Todorović-Raković, Natasa; Jovanović, Danica; Nesković-Konstantinović, Zora; Nikolić-Vukosavljević, Dragica

    2007-06-01

    After so many years of research, clinical value of HER2 (Human epidermal growth factor receptor 2) is unclear. Perhaps the main reason is variability of testing methods that produce controversial results. There is a lack of studies regarding prognostic value of CISH especially in metastatic breast cancer (MBC) when risk evaluation is based on different parameters than for primary breast cancer. Aim of this study was to compare prognostic relevance of HER2 status in MBC tested by two different methods i.e. immunohistochemistry (IHC) and chromogenic in situ hybridization (CISH). HER2 status of the same group of 107 MBC patients was determined by IHC (protein overexpression) and by CISH (gene amplification). HER2 results obtained by IHC and CISH showed significant correlation, beside the existence of discrepancies. Beside the significant correlation in two methods, there was a difference in prognostic values of compared methods during the course of metastatic disease. There was a significant difference in progression-free interval (PFI) between HER2 non-amplified and HER2 amplified cases determined by CISH, in postmenopausal subgroup and node-positive subgroup, but no significant difference for IHC stratified MBC patients. CISH seems to be accurate and more informative method than IHC regarding prognostic value of HER2 in metastatic breast cancer.

  16. Radiation induced wheat-rye chromosomal translocations in triticale. Optimizing the dose using fluorescence in situ hybridization

    International Nuclear Information System (INIS)

    Ahmad, F.; Comeau, A.; Chen, Q.; Collin, J.; St-Pierre, C.A.

    2000-01-01

    Fluorescent in situ hybridization (FISH) was utilized to monitor the level of ionizing radiation ( 60 Co source) in their ability to cause intra- and intergeneric chromosomal aberrations in triticale seeds. Seeds were irradiated with 0, 20, 50, 100, 200, 300, 400, 500 and 1000 Gy doses. The root growth of irradiated seeds was greatly inhibited at 200 Gy and above. Various types of aberrations including wheat-rye, wheat-wheat, rye-rye, wheat-rye-wheat, rye-wheat-rye translocations and acentric fragments with or without translocations were observed. There was a consistent increase in proportion of aberrations per cell with an increase in radiation dose. It was concluded that for an optimal level of chromosomal translocation and least number of acentric fragments, a 20 Gy dose was quite sufficient for inducing a desirable level of wheat-rye chromosomal translocations. The excellent efficiency and importance of utilizing FISH in such studies of alien-introgression via chromosomal translocations are discussed. (author)

  17. Fluorescence in situ hybridization of TP53 for the detection of chromosome 17 abnormalities in myelodysplastic syndromes.

    Science.gov (United States)

    Sánchez-Castro, Judit; Marco-Betés, Víctor; Gómez-Arbonés, Xavier; García-Cerecedo, Tomás; López, Ricard; Talavera, Elisabeth; Fernández-Ruiz, Sara; Ademà, Vera; Marugan, Isabel; Luño, Elisa; Sanzo, Carmen; Vallespí, Teresa; Arenillas, Leonor; Marco Buades, Josefa; Batlle, Ana; Buño, Ismael; Martín Ramos, María Luisa; Blázquez Rios, Beatriz; Collado Nieto, Rosa; Vargas, Ma Teresa; González Martínez, Teresa; Sanz, Guillermo; Solé, Francesc

    2015-01-01

    Conventional G-banding cytogenetics (CC) detects chromosome 17 (chr17) abnormalities in 2% of patients with de novo myelodysplastic syndromes (MDS). We used CC and fluorescence in situ hybridization (FISH) (LSI p53/17p13.1) to assess deletion of 17p in 531 patients with de novo MDS from the Spanish Group of Hematological Cytogenetics. FISH detected - 17 or 17p abnormalities in 13 cases (2.6%) in whom no 17p abnormalities were revealed by CC: 0.9% of patients with a normal karyotype, 0% in non-informative cytogenetics, 50% of patients with a chr17 abnormality without loss of 17p and 4.7% of cases with an abnormal karyotype not involving chr17. Our results suggest that applying FISH of 17p13 to identify the number of copies of the TP53 gene could be beneficial in patients with a complex karyotype. We recommend using FISH of 17p13 in young patients with a normal karyotype or non-informative cytogenetics, and always in isolated del(17p).

  18. Feasibility of using fluorescence in situ hybridization (FISH) to detect early gene changes in sputum cells from uranium miners

    International Nuclear Information System (INIS)

    Neft, R.E.; Rogers, J.L.; Belinsky, S.A.

    1995-01-01

    Epidemiological studies have shown that combined exposure to radon progeny and tobacco smoke produce a greater than additive or synergistic increase in lung cancer risk. Lung cancer results from multiple genetic changes over a long period of time. An early change that occurs in lung cancer is trisomy 7 which is found in 50% of non-small cell lung cancer and in the far margins of resected lung tumors. The 80% mortality associated with lung cancer is in part related to the high proportion of patients who present with an advanced, unresectable tumor. Therefore, early detection of patients at risk for tumor development is critical to improve treatment of this disease. Currently, it is difficult to detect lung cancer early while it is still amendable by surgery. Saccomanno, G. has shown that premalignant cytologic changes in sputum cells collected from uranium miners can be detected by a skilled, highly trained cytopathologist. A more objective alternative for identifying premalignant cells in sputum may be to determine whether an early genetic change such as trisomy 7 is present in these cells. Fluorescence in situ hybridization (FISH) can be used to identify cells with trisomy 7. The results of this investigation indicate that FISH may prove to be an accurate, efficient method to test at-risk individuals for genetic alterations in bronchial epithelial cells from sputum

  19. Hybrid Electrodes by In-Situ Integration of Graphene and Carbon-Nanotubes in Polypyrrole for Supercapacitors.

    Science.gov (United States)

    Aphale, Ashish; Maisuria, Krushangi; Mahapatra, Manoj K; Santiago, Angela; Singh, Prabhakar; Patra, Prabir

    2015-09-23

    Supercapacitors also known as electrochemical capacitors, that store energy via either Faradaic or non-Faradaic processes, have recently grown popularity mainly because they complement, and can even replace, conventional energy storage systems in variety of applications. Supercapacitor performance can be improved significantly by developing new nanocomposite electrodes which utilizes both the energy storage processes simultaneously. Here we report, fabrication of the freestanding hybrid electrodes, by incorporating graphene and carbon nanotubes (CNT) in pyrrole monomer via its in-situ polymerization. At the scan rate of 5 mV s(-1), the specific capacitance of the polypyrrole-CNT-graphene (PCG) electrode film was 453 F g(-1) with ultrahigh energy and power density of 62.96 W h kg(-1) and 566.66 W kg(-1) respectively, as shown in the Ragone plot. A nanofibrous membrane was electrospun and effectively used as a separator in the supercapacitor. Four supercapacitors were assembled in series to demonstrate the device performance by lighting a 2.2 V LED.

  20. Correlation between Photoluminescence and Carrier Transport and a Simple In Situ Passivation Method for High-Bandgap Hybrid Perovskites.

    Science.gov (United States)

    Stoddard, Ryan J; Eickemeyer, Felix T; Katahara, John K; Hillhouse, Hugh W

    2017-07-20

    High-bandgap mixed-halide hybrid perovskites have higher open-circuit voltage deficits and lower carrier diffusion lengths than their lower-bandgap counterparts. We have developed a ligand-assisted crystallization (LAC) technique that introduces additives in situ during the solvent wash and developed a new method to dynamically measure the absolute intensity steady-state photoluminescence and the mean carrier diffusion length simultaneously. The measurements reveal four distinct regimes of material changes and show that photoluminescence brightening often coincides with losses in carrier transport, such as in degradation or phase segregation. Further, the measurements enabled optimization of LAC on the 1.75 eV bandgap FA 0.83 Cs 0.17 Pb(I 0.66 Br 0.34 ) 3 , resulting in an enhancement of the photoluminescence quantum yield (PLQY) of over an order of magnitude, an increase of 80 meV in the quasi-Fermi level splitting (to 1.29 eV), an increase in diffusion length by a factor of 3.5 (to over 1 μm), and enhanced open-circuit voltage and short-circuit current from photovoltaics fabricated from the LAC-treated films.

  1. Comparative cytogenetics of six Indo-Pacific moray eels (Anguilliformes: Muraenidae) by chromosomal banding and fluorescence in situ hybridization.

    Science.gov (United States)

    Coluccia, E; Deidda, F; Cannas, R; Lobina, C; Cuccu, D; Deiana, A M; Salvadori, S

    2015-09-01

    A comparative cytogenetic analysis, using both conventional staining techniques and fluorescence in situ hybridization, of six Indo-Pacific moray eels from three different genera (Gymnothorax fimbriatus, Gymnothorax flavimarginatus, Gymnothorax javanicus, Gymnothorax undulatus, Echidna nebulosa and Gymnomuraena zebra), was carried out to investigate the chromosomal differentiation in the family Muraenidae. Four species displayed a diploid chromosome number 2n = 42, which is common among the Muraenidae. Two other species, G. javanicus and G. flavimarginatus, were characterized by different chromosome numbers (2n = 40 and 2n = 36). For most species, a large amount of constitutive heterochromatin was detected in the chromosomes, with species-specific C-banding patterns that enabled pairing of the homologous chromosomes. In all species, the major ribosomal genes were localized in the guanine-cytosine-rich region of one chromosome pair, but in different chromosomal locations. The (TTAGGG)n telomeric sequences were mapped onto chromosomal ends in all muraenid species studied. The comparison of the results derived from this study with those available in the literature confirms a substantial conservation of the diploid chromosome number in the Muraenidae and supports the hypothesis that rearrangements have occurred that have diversified their karyotypes. Furthermore, the finding of two species with different diploid chromosome numbers suggests that additional chromosomal rearrangements, such as Robertsonian fusions, have occurred in the karyotype evolution of the Muraenidae. © 2015 The Fisheries Society of the British Isles.

  2. Fluorescent in situ hybridization of pre-incubated blood culture material for the rapid diagnosis of histoplasmosis.

    Science.gov (United States)

    da Silva, Roberto Moreira; da Silva Neto, João Ricardo; Santos, Carla Silvana; Cruz, Kátia Santana; Frickmann, Hagen; Poppert, Sven; Koshikene, Daniela; de Souza, João Vicente Braga

    2015-02-01

    Fluorescence in situ hybridization (FISH) has been shown to be useful for the detection of Candida and Cryptococcus species in blood culture materials. FISH procedures for the detection of Histoplasma capsulatum var. capsulatum have not been reported so far. This study describes the development and evaluation of fluorescently labeled rRNA-targeting FISH probes to detect and identify H. capsulatum in blood cultures. All three analyzed H. capsulatum reference strains and clinical isolates showed positive signals with the newly designed specific oligonucleotide probes for H. capsulatum, whereas negative reactions were observed for all three nontarget yeast species and the two nontarget bacteria. The assay was also successfully applied for detections of H. capsulatum cells in pre-incubated blood culture samples of patients with clinical suspicion of histoplasmosis (n = 33). The described FISH-based assay was shown to be easy to apply, sensitive, and specific (compared to polymerase chain reaction) for the detection and identification of H. capsulatum in this proof-of-principle analysis. Larger multicentric assessments are recommended for a thorough diagnostic evaluation of the procedure. © The Author 2014. Published by Oxford University Press on behalf of The International Society for Human and Animal Mycology. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  3. Frequency of chromosome 17 aneuploidy in primary and recurrent pterygium by interphase-fluorescence in situ hybridization.

    Science.gov (United States)

    Kamis, Umit; Kerimoglu, Hurkan; Ozkagnici, Ahmet; Acar, Hasan

    2006-01-01

    To investigate chromosome 17 numerical aberrations by using fluorescence in situ hybridization (FISH) in pterygia and to find out whether there is any association between chromosome 17 aneuploidy and recurrent pterygia. Pterygium tissue samples were taken from 21 patients by surgical excision. Eighteen of them had primary and 3 had recurrent pterygium. Peripheral whole blood interphase cells obtained from 11 healthy subjects were assigned as control group. The cells from pterygium tissue and peripheral blood were incubated with a hypotonic solution and fixed in order to obtain interphase nuclei. FISH analysis with chromosome-17-specific alpha-satellite DNA probe was performed on both the interphase nuclei of pterygium tissue (of patients) and peripheral whole blood cells of controls. The mean percentage of chromosome 17 aneuploidy was 4.71% for the pterygia group and 4.41% for the controls. No significant difference of chromosome 17 aneuploidy was observed between the patients and the controls. When the group of patients with recurrences was compared with the group without recurrences, there was a significant difference in the frequency of chromosome 17 aneuploidy (U = 17, p = 0.029). Chromosome 17 aneuploidy is probably not an important factor in the formation of pterygium, but it may be related to recurrence.

  4. Rapid Identification of Staphylococcus aureus Directly from Blood Cultures by Fluorescence In Situ Hybridization with Peptide Nucleic Acid Probes

    Science.gov (United States)

    Oliveira, Kenneth; Procop, Gary W.; Wilson, Deborah; Coull, James; Stender, Henrik

    2002-01-01

    A new fluorescence in situ hybridization (FISH) method with peptide nucleic acid (PNA) probes for identification of Staphylococcus aureus directly from positive blood culture bottles that contain gram-positive cocci in clusters (GPCC) is described. The test (the S. aureus PNA FISH assay) is based on a fluorescein-labeled PNA probe that targets a species-specific sequence of the 16S rRNA of S. aureus. Evaluations with 17 reference strains and 48 clinical isolates, including methicillin-resistant and methicillin-susceptible S. aureus species, coagulase-negative Staphylococcus species, and other clinically relevant and phylogenetically related bacteria and yeast species, showed that the assay had 100% sensitivity and 96% specificity. Clinical trials with 87 blood cultures positive for GPCC correctly identified 36 of 37 (97%) of the S. aureus-positive cultures identified by standard microbiological methods. The positive and negative predictive values were 100 and 98%, respectively. It is concluded that this rapid method (2.5 h) for identification of S. aureus directly from blood culture bottles that contain GPCC offers important information for optimal antibiotic therapy. PMID:11773123

  5. Correlation between HER2 gene amplification and protein overexpression through fluorescence in situ hybridization and immunohistochemistry in breast carcinoma patients.

    Science.gov (United States)

    Makroo, R N; Chowdhry, Mohit; Kumar, Manoj; Srivastava, Priyanka; Tyagi, Richa; Bhadauria, Preeti; Kaul, Sumaid; Sarin, Ramesh; Das, P K; Dua, Harsh

    2012-01-01

    In India, the incidence of breast cancer has increased in the urban population, with 1 in every 22 women diagnosed with breast cancer. It is important to know the HER2/neu gene status for a better prognostication of these patients. The aim of this study was to compare the efficacy of fluorescence in situ hybridization (FISH) and immunohistochemistry (IHC) for determining HER2/neu alteration in breast carcinoma. A total of 188 histologically proven breast carcinoma cases between the years 2007 and 2011 were retrospectively analyzed on the paraffin tissue sections by both IHC and FISH techniques. FISH for HER2/neu gene amplification was performed on cases where the IHC status was already known and the results were compared. A total of 64 (30%) patients were found to be amplified and the remaining 124 (65.9%) cases were found to be unamplified through FISH. Patients observed with 3+ reading on IHC were later confirmed as unamplified in 29.5% cases through FISH. It has been confirmed with the present study that IHC is a prudent first-step technique to screen tissue samples for HER2/neu gene status, but should be supplemented with the FISH technique especially in equivocal cases.

  6. Correlation between HER2 gene amplification and protein overexpression through fluorescence in situ hybridization and immunohistochemistry in breast carcinoma patients

    Directory of Open Access Journals (Sweden)

    R N Makroo

    2012-01-01

    Full Text Available Background : In India, the incidence of breast cancer has increased in the urban population, with 1 in every 22 women diagnosed with breast cancer. It is important to know the HER2/neu gene status for a better prognostication of these patients. Aim : The aim of this study was to compare the efficacy of fluorescence in situ hybridization (FISH and immunohistochemistry (IHC for determining HER2/neu alteration in breast carcinoma. Materials and Methods : A total of 188 histologically proven breast carcinoma cases between the years 2007 and 2011 were retrospectively analyzed on the paraffin tissue sections by both IHC and FISH techniques. FISH for HER2/neu gene amplification was performed on cases where the IHC status was already known and the results were compared. Results : A total of 64 (30% patients were found to be amplified and the remaining 124 (65.9% cases were found to be unamplified through FISH. Patients observed with 3+ reading on IHC were later confirmed as unamplified in 29.5% cases through FISH. Conclusion : It has been confirmed with the present study that IHC is a prudent first-step technique to screen tissue samples for HER2/neu gene status, but should be supplemented with the FISH technique especially in equivocal cases.

  7. Next-generation in situ hybridization approaches to define and quantify HIV and SIV reservoirs in tissue microenvironments.

    Science.gov (United States)

    Deleage, Claire; Chan, Chi N; Busman-Sahay, Kathleen; Estes, Jacob D

    2018-01-09

    The development of increasingly safe and effective antiretroviral treatments for human immunodeficiency virus (HIV) over the past several decades has led to vastly improved patient survival when treatment is available and affordable, an outcome that relies on uninterrupted adherence to combination antiretroviral therapy for life. Looking to the future, the discovery of an elusive 'cure' for HIV will necessitate highly sensitive methods for detecting, understanding, and eliminating viral reservoirs. Next-generation, in situ hybridization (ISH) approaches offer unique and complementary insights into viral reservoirs within their native tissue environments with a high degree of specificity and sensitivity. In this review, we will discuss how modern ISH techniques can be used, either alone or in conjunction with phenotypic characterization, to probe viral reservoir establishment and maintenance. In addition to focusing on how these techniques have already furthered our understanding of HIV reservoirs, we discuss potential avenues for how high-throughput, next-generation ISH may be applied. Finally, we will review how ISH could allow deeper phenotypic and contextual insights into HIV reservoir biology that should prove instrumental in moving the field closer to viral reservoir elimination needed for an 'HIV cure' to be realized.

  8. Radiation induced wheat-rye chromosomal translocations in triticale. Optimizing the dose using fluorescence in situ hybridization

    Energy Technology Data Exchange (ETDEWEB)

    Ahmad, F. [Brandon Univ., Manitoba (Canada); Comeau, A.; Chen, Q.; Collin, J.; St-Pierre, C.A.

    2000-03-01

    Fluorescent in situ hybridization (FISH) was utilized to monitor the level of ionizing radiation ({sup 60}Co source) in their ability to cause intra- and intergeneric chromosomal aberrations in triticale seeds. Seeds were irradiated with 0, 20, 50, 100, 200, 300, 400, 500 and 1000 Gy doses. The root growth of irradiated seeds was greatly inhibited at 200 Gy and above. Various types of aberrations including wheat-rye, wheat-wheat, rye-rye, wheat-rye-wheat, rye-wheat-rye translocations and acentric fragments with or without translocations were observed. There was a consistent increase in proportion of aberrations per cell with an increase in radiation dose. It was concluded that for an optimal level of chromosomal translocation and least number of acentric fragments, a 20 Gy dose was quite sufficient for inducing a desirable level of wheat-rye chromosomal translocations. The excellent efficiency and importance of utilizing FISH in such studies of alien-introgression via chromosomal translocations are discussed. (author)

  9. Expression of a serine protease (motopsin PRSS12) mRNA in the mouse brain: in situ hybridization histochemical study.

    Science.gov (United States)

    Iijima, N; Tanaka, M; Mitsui, S; Yamamura, Y; Yamaguchi, N; Ibata, Y

    1999-03-20

    Serine proteases are considered to play several important roles in the brain. In an attempt to find novel brain-specific serine proteases (BSSPs), motopsin (PRSS-12) was cloned from a mouse brain cDNA library by polymerase chain reaction (PCR). Northern blot analysis demonstrated that the postnatal 10-day mouse brain contained the most amount of motopsin mRNA. At this developmental stage, in situ hybridization histochemistry showed that motopsin mRNA was specifically expressed in the following regions: cerebral cortical layers II/III, V and VIb, endopiriform cortex and the limbic system, particularly in the CA1 region of the hippocampal formation. In addition, in the brainstem, the oculomotor nucleus, trochlear nucleus, mecencephalic and motor nuclei of trigeminal nerve (N), abducens nucleus, facial nucleus, nucleus of the raphe pontis, dorsoral motor nucleus of vagal N, hypoglossal nucleus and ambiguus nucleus showed motopsin mRNA expression. Expression was also found in the anterior horn of the spinal cord. The above findings strongly suggest that neurons in almost all motor nuclei, particularly in the brainstem and spinal cord, express motopsin mRNA, and that motopsin seems to have a close relation to the functional role of efferent neurons. Copyright 1999 Elsevier Science B.V.

  10. Automated Image Analysis of HER2 Fluorescence In Situ Hybridization to Refine Definitions of Genetic Heterogeneity in Breast Cancer Tissue.

    Science.gov (United States)

    Radziuviene, Gedmante; Rasmusson, Allan; Augulis, Renaldas; Lesciute-Krilaviciene, Daiva; Laurinaviciene, Aida; Clim, Eduard; Laurinavicius, Arvydas

    2017-01-01

    Human epidermal growth factor receptor 2 gene- (HER2-) targeted therapy for breast cancer relies primarily on HER2 overexpression established by immunohistochemistry (IHC) with borderline cases being further tested for amplification by fluorescence in situ hybridization (FISH). Manual interpretation of HER2 FISH is based on a limited number of cells and rather complex definitions of equivocal, polysomic, and genetically heterogeneous (GH) cases. Image analysis (IA) can extract high-capacity data and potentially improve HER2 testing in borderline cases. We investigated statistically derived indicators of HER2 heterogeneity in HER2 FISH data obtained by automated IA of 50 IHC borderline (2+) cases of invasive ductal breast carcinoma. Overall, IA significantly underestimated the conventional HER2, CEP17 counts, and HER2/CEP17 ratio; however, it collected more amplified cells in some cases below the lower limit of GH definition by manual procedure. Indicators for amplification, polysomy, and bimodality were extracted by factor analysis and allowed clustering of the tumors into amplified, nonamplified, and equivocal/polysomy categories. The bimodality indicator provided independent cell diversity characteristics for all clusters. Tumors classified as bimodal only partially coincided with the conventional GH heterogeneity category. We conclude that automated high-capacity nonselective tumor cell assay can generate evidence-based HER2 intratumor heterogeneity indicators to refine GH definitions.

  11. Evaluation of intratumoral HER-2 heterogeneity by fluorescence in situ hybridization in invasive breast cancer: a single institution study.

    Science.gov (United States)

    Lee, Sarah; Jung, Woohee; Hong, Soon-Won; Koo, Ja Seung

    2011-08-01

    This study aimed to determine the incidence and characteristics of HER-2 gene heterogeneity in invasive breast cancer in a single institution. Included were 971 cases of primary invasive breast cancer diagnosed between 2008 and 2010. Fluorescence in situ hybridization (FISH) image files were retrospectively reviewed and HER-2 gene heterogeneity was defined as more than 5% but less than 50% of analyzed invasive tumor cells with a HER-2/Chr17 ratio higher than 2.2, according to the College of American Pathologists guidelines. HER-2 gene heterogeneity was identified in 24 (2.5%) cases. The mean proportion of invasive tumor cells with a HER-2/chromosome 17 ratio higher than 2.2 was 11.6% (range: 5%-25%). Of 24 cases, HER-2 gene status was not amplified in 8, showed borderline amplification in 2, and amplification in 14. All HER-2 amplification cases were low-grade. In conclusion, HER-2 gene heterogeneity of invasive breast cancer is identified in routine FISH examination. This may affect the results of HER-2 gene amplification status in FISH studies.

  12. Quantitative fluorescence in situ hybridization measurement of telomere length in skin with/without sun exposure or actinic keratosis.

    Science.gov (United States)

    Ikeda, Hiroyuki; Aida, Junko; Hatamochi, Atsushi; Hamasaki, Yoichiro; Izumiyama-Shimomura, Naotaka; Nakamura, Ken-Ichi; Ishikawa, Naoshi; Poon, Steven S; Fujiwara, Mutsunori; Tomita, Ken-Ichiro; Hiraishi, Naoki; Kuroiwa, Mie; Matsuura, Masaaki; Sanada, Yukihiro; Kawano, Youichi; Arai, Tomio; Takubo, Kaiyo

    2014-03-01

    Chromosomal and genomic instability due to telomere dysfunction is known to play an important role in carcinogenesis. To study telomere shortening in the epidermis surrounding actinic keratosis, we measured telomere lengths of basal, parabasal, and suprabasal cells in epidermis with actinic keratosis (actinic keratosis group, n = 18) and without actinic keratosis (sun-protected, n = 15, and sun-exposed, n = 13 groups) and in actinic keratosis itself as well as in dermal fibroblasts in the 3 groups, using quantitative fluorescence in situ hybridization. Among the 3 cell types, telomeres of basal cells were not always the longest, suggesting that tissue stem cells are not necessarily located among basal cells. Telomeres of basal cells in the sun-exposed group were shorter than those in the sun-protected group. Telomeres in the background of actinic keratosis and in actinic keratosis itself and those of fibroblasts in actinic keratosis were significantly shorter than those in the controls. Our findings demonstrate that sun exposure induces telomere shortening and that actinic keratosis arises from epidermis with shorter telomeres despite the absence of any histologic atypia. © 2014.

  13. EnVision+, a new dextran polymer-based signal enhancement technique for in situ hybridization (ISH).

    Science.gov (United States)

    Wiedorn, K H; Goldmann, T; Henne, C; Kühl, H; Vollmer, E

    2001-09-01

    Seventy paraffin-embedded cervical biopsy specimens and condylomata were tested for the presence of human papillomavirus (HPV) by conventional in situ hybridization (ISH) and ISH with subsequent signal amplification. Signal amplification was performed either by a commercial biotinyl-tyramide-based detection system [GenPoint (GP)] or by the novel two-layer dextran polymer visualization system EnVision+ (EV), in which both EV-horseradish peroxidase (EV-HRP) and EV-alkaline phosphatase (EV-AP) were applied. We could demonstrate for the first time, that EV in combination with preceding ISH results in a considerable increase in signal intensity and sensitivity without loss of specificity compared to conventional ISH. Compared to GP, EV revealed a somewhat lower sensitivity, as measured by determination of the integrated optical density (IOD) of the positively stained cells. However, EV is easier to perform, requires a shorter assay time, and does not raise the background problems that may be encountered with biotinyl-tyramide-based amplification systems. (J Histochem Cytochem 49:1067-1071, 2001)

  14. Rapid molecular cytogenetic analysis of X-chromosomal microdeletions: Fluorescence in situ hybridization (FISH) for complex glycerol kinase deficiency

    Energy Technology Data Exchange (ETDEWEB)

    Worley, K.C.; Lindsay, E.A.; McCabe, E.R.B. [Baylor College of Medicine, Houston, TX (United States)] [and others

    1995-07-17

    Diagnosis of X-chromosomal microdeletions has relied upon the traditional methods of Southern blotting and DNA amplification, with carrier identification requiring time-consuming and unreliable dosage calculations. In this report, we describe rapid molecular cytogenetic identification of deleted DNA in affected males with the Xp21 contiguous gene syndrome (complex glycerol kinase deficiency, CGKD) and female carriers for this disorder. CGKD deletions involve the genes for glycerol kinase, Duchenne muscular dystrophy, and/or adrenal hypoplasia congenita. We report an improved method for diagnosis of deletions in individuals with CGKD and for identification of female carriers within their families using fluorescence in situ hybridization (FISH) with a cosmid marker (cosmid 35) within the glycerol kinase gene. When used in combination with an Xq control probe, affected males demonstrate a single signal from the control probe, while female carriers demonstrate a normal chromosome with two signals, as well as a deleted chromosome with a single signal from the control probe. FISH analysis for CGKD provides the advantages of speed and accuracy for evaluation of submicroscopic X-chromosome deletions, particularly in identification of female carriers. In addition to improving carrier evaluation, FISH will make prenatal diagnosis of CGKD more readily available. 17 refs., 2 figs.

  15. Analysis of methanogenic activity in a thermophilic-dry anaerobic reactor: Use of fluorescent in situ hybridization

    International Nuclear Information System (INIS)

    Montero, B.; Garcia-Morales, J.L.; Sales, D.; Solera, R.

    2009-01-01

    Methanogenic activity in a thermophilic-dry anaerobic reactor was determined by comparing the amount of methane generated for each of the organic loading rates with the size of the total and specific methanogenic population, as determined by fluorescent in situ hybridization. A high correlation was evident between the total methanogenic activity and retention time [-0.6988Ln(x) + 2.667] (R 2 0.8866). The total methanogenic activity increased from 0.04 x 10 -8 mLCH 4 cell -1 day -1 to 0.38 x 10 -8 mLCH 4 cell -1 day -1 while the retention time decreased, augmenting the organic loading rates. The specific methanogenic activities of H 2 -utilizing methanogens and acetate-utilizing methanogens increased until they stabilised at 0.64 x 10 -8 mLCH 4 cell -1 day -1 and 0.33 x 10 -8 mLCH 4 cell -1 day -1 , respectively. The methanogenic activity of H 2 -utilizing methanogens was higher than acetate-utilizing methanogens, indicating that maintaining a low partial pressure of hydrogen does not inhibit the acetoclastic methanogenesis or the anaerobic process

  16. Fluorescent in situ hybridization (FISH) on corneal impression cytology specimens (CICS): study of epithelial cell survival after keratoplasty.

    Science.gov (United States)

    Catanese, Muriel; Popovici, Cornel; Proust, Hélène; Hoffart, Louis; Matonti, Frédéric; Cochereau, Isabelle; Conrath, John; Gabison, Eric E

    2011-02-22

    To assess corneal epithelial cell survival after keratoplasty. Corneal impression cytology (CIC) was performed on sex-mismatched corneal transplants. Fluorescent in situ hybridization (FISH) with sex chromosome-specific probes was performed to identify epithelial cell mosaicism and therefore allocate the donor or recipient origin of the cells. Twenty-four samples of corneal epithelial cells derived from 21 transplanted patients were analyzed. All patients received post-operative treatment using dexamethasone eye drops, with progressive tapering over 18 months, and nine patients also received 2% cyclosporine eye drops. Out of the 24 samples reaching quality criteria, sex mosaicism was found in 13, demonstrating the presence of donor-derived cells at the center of the graft for at least 211 days post keratoplasty. Kaplan-Meier analysis established a median survival of donor corneal epithelial cells of 385 days. Although not statistically significant, the disappearance of donor cells seemed to be delayed and the average number of persistent cells appeared to be greater when 2% cyclosporine was used topically as an additional immunosuppressive therapy. The combination of corneal impressions and FISH analysis is a valuable tool with negligible side effects to investigate the presence of epithelial cell mosaicism in sex-mismatched donor transplants. Epithelial cells survived at the center of the graft with a median survival of more than one year, suggesting slower epithelial turnover than previously described.

  17. Exploring polycythaemia vera with fluorescence in situ hybridization: additional cryptic 9p is the most frequent abnormality detected.

    Science.gov (United States)

    Najfeld, Vesna; Montella, Lya; Scalise, Angela; Fruchtman, Steven

    2002-11-01

    Between 1986 and 2001, 220 patients with polycythaemia vera (PV) were studied using conventional cytogenetics. Of 204 evaluable patients, 52 (25.4%) had clonal abnormalities. The recurrent chromosomal rearrangements were those of chromosome 9 (21.1%), del(20q) (19.2%), trisomy 8 (19.2%), rearrangements of 13q (13.4%), abnormalities of 1q (11.5%), and of chromosomes 5 and 7 (9.6%). Subsequent analysis of 32 patients, performed at follow-up of up to 14.8 years, revealed new clonal abnormalities in five patients and the disappearance of an abnormal clone in four. Eleven patients remained normal up to 11.5 years and seven patients maintained an abnormality for over 10 years. Fifty-three patients were studied retrospectively using interphase fluorescence in situ hybridization (I-FISH), utilizing probes for centromere enumeration of chromosomes 8 and 9, and for 13q14 and 20q12 loci. Conventional cytogenetics demonstrated clonal chromosome abnormalities in 23% of these 53 patients. The addition of I-FISH increased the detection of abnormalities to 29% and permitted clarification of chromosome 9 rearrangements in an additional 5.6% of patients. FISH uncovered rearrangements of chromosome 9 in 53% of patients with an abnormal FISH pattern, which represented the most frequent genomic alteration in this series.

  18. Analysis of the nitrifying bacterial community in BioCube sponge media using fluorescent in situ hybridization (FISH) and microelectrodes.

    Science.gov (United States)

    Chae, Kyu-Jung; Rameshwar, T; Jang, Am; Kim, Sung H; Kim, In S

    2008-09-01

    There is growing interest in the development of more cost-effective and retrofit technologies for the upgrade and expansion of existing wastewater treatment plants with extreme space constraints. A free-floating sponge media (BioCube) process, using a 24 L lab scale reactor, was operated to study the nitrification profiles and microbial community. The COD removal efficiencies were maintained, at an average of 95%, with the mixed liquor suspended solids (MLSS) inside the BioCube sponge media maintained at 12,688 mg/L. The nitrification removal efficiencies were between 92% and 100%, with an average value of 99%. From the results of microelectrode measurements, the ammonium ion concentration was found to rapidly decrease from the surface of the BioCube sponge media to a depth of 2mm due to chemical reactions carried out by ammonia oxidizing bacteria (AOB) species. Multi-fluorescence in situ hybridization (FISH) has been used to investigate the spatial distributions of various microbial activities within reactors. Microbial communities were targeted using different oligonucleotide probes specific to AOB and nitrite oxidizing bacteria (NOB). There were a large number of AOB populations, but these were not uniformly distributed in the biofilm compared to the NOB populations.

  19. Comprehensive Analysis of ETS Family Members in Melanoma by Fluorescence In Situ Hybridization Reveals Recurrent ETV1 Amplification

    Science.gov (United States)

    Mehra, Rohit; Dhanasekaran, Saravana M; Palanisamy, Nallasivam; Vats, Pankaj; Cao, Xuhong; Kim, Jung H; Kim, David SL; Johnson, Timothy; Fullen, Douglas R; Chinnaiyan, Arul M

    2013-01-01

    E26 transformation-specific (ETS) transcription factors are known to be involved in gene aberrations in various malignancies including prostate cancer; however, their role in melanoma oncogenesis has yet to be fully explored. We have completed a comprehensive fluorescence in situ hybridization (FISH)-based screen for all 27 members of the ETS transcription factor family on two melanoma tissue microarrays, representing 223 melanomas, 10 nevi, and 5 normal skin tissues. None of the melanoma cases demonstrated ETS fusions; however, 6 of 114 (5.3%) melanomas were amplified for ETV1 using a break-apart FISH probe. For the six positive cases, locus-controlled FISH probes revealed that two of six cases were amplified for the ETV1 region, whereas four cases showed copy gains of the entire chromosome 7. The remaining 26 ETS family members showed no chromosomal aberrations by FISH. Quantitative polymerase chain reaction showed an average 3.4-fold (P value = .00218) increased expression of ETV1 in melanomas, including the FISH ETV1-amplified cases, when compared to other malignancies (prostate, breast, and bladder carcinomas). These data suggest that a subset of melanomas overexpresses ETV1 and amplification of ETV1 may be one mechanism for achieving high gene expression. PMID:23908683

  20. Metastatic hidradenocarcinoma with demonstration of Her-2/neu gene amplification by fluorescence in situ hybridization: potential treatment implications.

    Science.gov (United States)

    Nash, Jason W; Barrett, Terry L; Kies, Merrill; Ross, Merrick I; Sneige, Nour; Diwan, A Hafeez; Lazar, Alexander J F

    2007-01-01

    A 44-year-old man was referred for a right chest nodule of 3 months duration. A 'benign' nodule had been excised from this location 8 years prior. On examination, palpable nodes were noted in the right axilla. Radiographic studies were significant only for right axillary lymphadenopathy. Histologically, a nodular dermal proliferation composed of poorly differentiated epithelioid cells in nests and focally forming ducts with pseudopapillary architecture comprised the primary tumor. Features of a clear cell hidradenoma were noted focally. Immunohistochemical (IHC) analysis revealed reactivity for HMW cytokeratins, CK5 and CK7, p53, p63, CEA (focal), androgen receptor, EGFR, estrogen receptor (ER), MUC5AC, and strong/diffuse membranous staining for Her-2/neu. Negative stains included villin, TTF-1, CDX2, S-100 protein, vimentin, gross cystic disease fluid protein 15 (GCDFP-15), mammoglobulin, and MUC2. A wide local excision and axillary node dissection was performed. Metastatic tumor involved nine of 28 nodes. Interphase fluorescence in situ hybridization (FISH) demonstrated chromosomal amplification of the Her-2/neu locus within the tumor and a nodal metastasis. The patient has completed adjuvant and radiotherapy, including trastuzumab, and is asymptomatic. We believe this to be the first demonstration of Her-2/neu amplification in a malignant skin adnexal tumor. In analogy to breast carcinoma, these findings suggest the applicability of trastuzumab for patients with metastatic adnexal carcinomas demonstrating Her-2/neu amplification.

  1. Detection and quantitation of HER-2/neu gene amplification in human breast cancer archival material using fluorescence in situ hybridization.

    Science.gov (United States)

    Pauletti, G; Godolphin, W; Press, M F; Slamon, D J

    1996-07-04

    Amplification and overexpression of the HER-2/neu gene occurs in 25-30% of human breast cancers. This genetic alteration is associated with a poor clinical prognosis in women with either node negative or node positive breast cancers. The initial studies testing this association were somewhat controversial and this controversy was due in large part to significant heterogeneity in both the methods and/or reagents used in testing archival material for the presence of the alteration. These methods included a number of solid matrix blotting techniques for DNA, RNA and protein as well as immunohistochemistry. Fluorescence in situ hybridization (FISH) represents the newest methodologic approach for testing for this genetic alteration. In this study, FISH is compared to Southern, Northern and Western blot analyses as well as immunohistochemistry in a large cohort of archival human breast cancer specimens. FISH was found to be superior to all other methodologies tested in assessing formalin fixed, paraffin embedded material for HER-2/neu amplification. The results from this study also confirm that overexpression of HER-2/neu rarely occurs in the absence of gene amplification in breast cancer (approximately 3% of cases). This method of analysis is rapid, reproducible and extremely reliable in detecting presence of HER-2/neu gene amplification and should have clinical utility.

  2. Chromosome analysis of nuclear power plant workers using fluorescence in situ hybridization and Giemsa assay

    International Nuclear Information System (INIS)

    Hristova, Rositsa; Hadjidekova, Valeria; Grigorova, Mira; Nikolova, Teodora; Bulanova, Minka; Popova, Ljubomira; Staynova, Albena; Benova, Donka

    2013-01-01

    The aim of this study was to evaluate the genotoxic effects of ionizing radiation in vivo in exposed Bulgarian nuclear power plant workers by using classical cytogenetic and molecular cytogenetic analyses of peripheral lymphocytes. Chromosome analysis using fluorescence in situ hybrydization (FISH) and Giemsa techniques was undertaken on 63 workers and 45 administrative staff controls from the Bulgarian Nuclear Power Plant. Using the Giemsa method, the frequencies of cells studied with chromosome aberrations, dicentrics plus rings and chromosome fragments in the radiation workers were significantly higher compared with the control group (P=0.044, P=0.014, and P=0.033, respectively). A significant association between frequencies of dicentrics plus rings and accumulated doses was registered (P<0.01). In the present study, a FISH cocktail of whole chromosome paints for chromosomes 1, 4 and 11 was used. A significant association between frequency of translocations and accumulated doses was also observed (P<0.001). Within the control group, a correlation was found between age and the spontaneous frequency of translocations. No correlation was found between smoking status and frequency of translocations. When compared with the control group, workers with accumulated doses up to 100 mSv showed no increase in genome translocation frequency, whereas workers with accumulated doses from 101 to 200 mSv showed a statistically significant doubling of genome translocation frequency (P=0.009). Thus, in cases of chronic exposure and for purposes of retrospective dosimetry, the genome frequency of translocations is a more useful marker for evaluation of genotoxic effects than dicentric frequency. (author)

  3. Multicenter Evaluation of a New Shortened Peptide Nucleic Acid Fluorescence In Situ Hybridization Procedure for Species Identification of Select Gram-Negative Bacilli from Blood Cultures▿

    Science.gov (United States)

    Morgan, Margie; Marlowe, Elizabeth; Della-Latta, Phyllis; Salimnia, Hossein; Novak-Weekley, Susan; Wu, Fann; Crystal, Benjamin S.

    2010-01-01

    A shortened protocol for two peptide nucleic acid fluorescence in situ hybridization (PNA FISH) assays for the detection of Gram-negative bacilli from positive blood cultures was evaluated in a multicenter trial. There was 100% concordance between the two protocols for each assay (368 of 368 and 370 of 370 results) and 99.7% (367 of 368 and 369 of 370 results) agreement with routine laboratory techniques. PMID:20357212

  4. In Silico and Fluorescence In Situ Hybridization Mapping Reveals Collinearity between the Pennisetum squamulatum Apomixis Carrier-Chromosome and Chromosome 2 of Sorghum and Foxtail Millet

    OpenAIRE

    Sapkota, Sirjan; Conner, Joann A.; Hanna, Wayne W.; Simon, Bindu; Fengler, Kevin; Deschamps, St?phane; Cigan, Mark; Ozias-Akins, Peggy

    2016-01-01

    Apomixis, or clonal propagation through seed, is a trait identified within multiple species of the grass family (Poaceae). The genetic locus controlling apomixis in Pennisetum squamulatum (syn Cenchrus squamulatus) and Cenchrus ciliaris (syn Pennisetum ciliare, buffelgrass) is the apospory-specific genomic region (ASGR). Previously, the ASGR was shown to be highly conserved but inverted in marker order between P. squamulatum and C. ciliaris based on fluorescence in situ hybridization (FISH) a...

  5. The Potassium-Argon Laser Experiment (KArLE): In Situ Geochronology for Planetary Robotic Missions

    Science.gov (United States)

    Cohen, Barbara

    2016-01-01

    The Potassium (K) - Argon (Ar) Laser Experiment (KArLE) will make in situ noble-gas geochronology measurements aboard planetary robotic landers and roverss. Laser-Induced Breakdown Spectroscopy (LIBS) is used to measure the K abun-dance in a sample and to release its noble gases; the evolved Ar is measured by mass spectrometry (MS); and rela-tive K content is related to absolute Ar abundance by sample mass, determined by optical measurement of the ablated volume. KArLE measures a whole-rock K-Ar age to 10% or better for rocks 2 Ga or older, sufficient to resolve the absolute age of many planetary samples. The LIBS-MS approach is attractive because the analytical components have been flight proven, do not require further technical development, and provide complementary measurements as well as in situ geochronology.

  6. Transit experience with hydrogen fueled hybrid electric buses

    Energy Technology Data Exchange (ETDEWEB)

    Scott, P.B.; Mazaika, D.M. [ISE Corp., Poway, CA (United States)

    2006-07-01

    Mass transit buses are ideal candidates for hydrogen implementation due to their capability of carrying 30 to 60 kg of hydrogen. ISE Corporation is a supplier of hydrogen fueled buses, including the first hybrid electric fuel cell bus which was commercialized in 2002, the hybrid electric fuel cell bus, and the hybrid hydrogen internal combustion engine (HHICE) bus which was commercialized in 2004. The configuration of a HHICE bus was illustrated with reference to its engine, control system, energy storage, generator, drive motor, inverter and accessories. Although these vehicles are expensive, the cost is amortized over a large base of hours used and passengers carried. The buses are operated primarily in urban areas where quiet and clean operation is needed the most. ISE has established a joint venture with Thor industries to develop a series of fuel cell buses equipped with a 60 kW PEM fuel cell. A schematic illustrating the energy flow in HHICE bus was also presented. It was shown that regenerative braking recovers the energy of motion. When using regenerative braking, most of the braking energy is saved in the battery. ISE drive systems convert 30 per cent or more of the bus energy to electrical energy to be used in later acceleration. Reduced fuel consumption also reduces the vehicle emissions. Testing of HHICE buses in both summer and winter operating conditions have shown that the range needs to be improved along with engine component reliability and durability. Fuel supply is also a major issue. A comparison with a fuel cell hybrid system was also presented. In the United States, more than 100,000 miles have been logged for the use of hydrogen hybrid buses, fuel cell buses and HHICE buses. The HHICE bus offers low capital cost, familiar technologies, but some NOx. CAT absorber technology offers the possibility of near zero emission capability. The fuel cell bus was found to be more fuel efficient, and can travel nearly twice as far per unit energy as

  7. Dealing with the problem of non-specific in situ mRNA hybridization signals associated with plant tissues undergoing programmed cell death

    Directory of Open Access Journals (Sweden)

    Jokela Anne

    2010-02-01

    Full Text Available Abstract Background In situ hybridization is a general molecular method typically used for the localization of mRNA transcripts in plants. The method provides a valuable tool to unravel the connection between gene expression and anatomy, especially in species such as pines which show large genome size and shortage of sequence information. Results In the present study, expression of the catalase gene (CAT related to the scavenging of reactive oxygen species (ROS and the polyamine metabolism related genes, diamine oxidase (DAO and arginine decarboxylase (ADC, were localized in developing Scots pine (Pinus sylvestris L. seeds. In addition to specific signals from target mRNAs, the probes continually hybridized non-specifically in the embryo surrounding region (ESR of the megagametophyte tissue, in the remnants of the degenerated suspensors as well as in the cells of the nucellar layers, i.e. tissues exposed to cell death processes and extensive nucleic acid fragmentation during Scots pine seed development. Conclusions In plants, cell death is an integral part of both development and defence, and hence it is a common phenomenon in all stages of the life cycle. Our results suggest that extensive nucleic acid fragmentation during cell death processes can be a considerable source of non-specific signals in traditional in situ mRNA hybridization. Thus, the visualization of potential nucleic acid fragmentation simultaneously with the in situ mRNA hybridization assay may be necessary to ensure the correct interpretation of the signals in the case of non-specific hybridization of probes in plant tissues.

  8. Spacecraft Hybrid (Mixed-Actuator) Attitude Control Experiences on NASA Science Missions

    Science.gov (United States)

    Dennehy, Cornelius J.

    2014-01-01

    There is a heightened interest within NASA for the design, development, and flight implementation of mixed-actuator hybrid attitude control systems for science spacecraft that have less than three functional reaction wheel actuators. This interest is driven by a number of recent reaction wheel failures on aging, but what could be still scientifically productive, NASA spacecraft if a successful hybrid attitude control mode can be implemented. Over the years, hybrid (mixed-actuator) control has been employed for contingency attitude control purposes on several NASA science mission spacecraft. This paper provides a historical perspective of NASA's previous engineering work on spacecraft mixed-actuator hybrid control approaches. An update of the current situation will also be provided emphasizing why NASA is now so interested in hybrid control. The results of the NASA Spacecraft Hybrid Attitude Control Workshop, held in April of 2013, will be highlighted. In particular, the lessons learned captured from that workshop will be shared in this paper. An update on the most recent experiences with hybrid control on the Kepler spacecraft will also be provided. This paper will close with some future considerations for hybrid spacecraft control.

  9. Experimental Investigation on Mechanical and Turning Behavior of Al 7075/x% wt. TiB2-1% Gr In Situ Hybrid Composite

    Directory of Open Access Journals (Sweden)

    K. R. Ramkumar

    2015-01-01

    Full Text Available The present research work involves the study of AA 7075-TiB2-Gr in situ composite through stir casting route. This in situ method involves formation of reinforcements within the matrix by the chemical reaction of two or more compounds which also produces some changes in the matrix material within the vicinity. Titanium Diboride (TiB2 and graphite were the reinforcement in a matrix of AA 7075 alloy. The composite was prepared with the formation of the reinforcement inside the molten matrix by adding salts of Potassium Tetrafluoroborate (KBF4 and Potassium Hexafluorotitanate (K2TiF6. The samples were taken under casted condition and the properties of the composite were tested by conducting characterization using X-ray diffraction (XRD, hardness test, flexural strength by using three-point bend test, scanning electron microscope (SEM, optical microstructure, grain size analysis, and surface roughness. It was found that good/excellent mechanical properties were obtained in AA 7075-TiB2-Gr reinforced in situ hybrid composite compared to alloy due to particulate strengthening of ceramic particles of TiB2 in the matrix. Further, Al 7075-3% TiB2-1% Gr hybrid in situ composite exhibited improved machinability over the alloy and composites due to self-lubricating property given by the Gr particles in the materials.

  10. Synthesis and luminescence properties of hybrid organic-inorganic transparent titania thin film activated by in-situ formed lanthanide complexes

    International Nuclear Information System (INIS)

    Wang Yige; Wang Li; Li Huanrong; Liu Peng; Qin Dashan; Liu Binyuan; Zhang Wenjun; Deng Ruiping; Zhang Hongjie

    2008-01-01

    Stable transparent titania thin films were fabricated at room temperature by combining thenoyltrifluoroacetone (TTFA)-modified titanium precursors with amphiphilic triblock poly(ethylene oxide)-poly(propylene oxide)-poly(ethylene oxide) (PEO-PPO-PEO, P123) copolymers. The obtained transparent titania thin films were systematically investigated by IR spectroscopy, PL emission and excitation spectroscopy and transmission electron microscopy. IR spectroscopy indicates that TTFA coordinates the titanium center during the process of hydrolysis and condensation. Luminescence spectroscopy confirms the in-situ formation of lanthanide complexes in the transparent titania thin film. TEM image shows that the in-situ formed lanthanide complexes were homogeneously distributed throughout the whole thin film. The quantum yield and the number of water coordinated to lanthanide metal center have been theoretically determined based on the luminescence data. - Graphical abstract: Novel stable luminescent organic-inorganic hybrid titania thin film with high transparency activated by in-situ formed lanthanide complexes have been obtained at room temperature via a simple one-pot synthesis approach by using TTFA-modified titanium precursor with amphiphilic triblock copolymer P123. The obtained hybrid thin film displays bright red (or green), near-monochromatic luminescence due to the in-situ formed lanthanide complex

  11. Specific identification of human papillomavirus type in cervical smears and paraffin sections by in situ hybridization with radioactive probes: a preliminary communication

    International Nuclear Information System (INIS)

    Gupta, J.; Gendelman, H.E.; Naghashfar, Z.; Gupta, P.; Rosenshein, N.; Sawada, E.; Woodruff, J.D.; Shah, K.

    1985-01-01

    Cervical Papanicolaou smears and paraffin sections of biopsy specimens obtained from women attending dysplasia clinics were examined for viral DNA sequences by in situ hybridization technique using 35 S-labeled cloned recombinant DNA probes of human papillomavirus (HPV) types 6, 11, and 16. These and one unrelated DNA probe complementary to measles virus RNA were labeled by nick translation using either one or two 35 S-labeled nucleotides. Paraffin sections and cervical smears were collected on pretreated slides, hybridized with the probes under stringent or nonstringent conditions for 50 h, and autoradiographed. Additional cervical specimens from the same women were examined for the presence of genus-specific papillomavirus capsid antigen by the immunoperoxidase technique. Preliminary results may be summarized as follows. The infecting virus could be identified in smears as well as in sections. Viral DNA sequences were detected only when there were condylomatous cells in the specimen and in only a proportion of the condylomatous cells. Even under stringent conditions, some specimens reacted with both HPV-6 and HPV-11. In some instances, the cells did not hybridize with any of the three probes even when duplicate specimens contained frankly condylomatous, capsid antigen-positive cells. In situ hybridization of Papanicolaou smears or of tissue sections is a practical method for diagnosis and follow-up of specific papillomavirus infection using routinely collected material

  12. In situ experiments on the performance of near-field for nuclear waste repository at KURT

    Energy Technology Data Exchange (ETDEWEB)

    Cho, Won-Jin, E-mail: wjcho@kaeri.re.kr [Korea Atomic Energy Research Institute, P.O. Box 105, Yuseong, Daejeon 305-600 (Korea, Republic of); Kim, Jin-Sub; Lee, Changsoo [Korea Atomic Energy Research Institute, P.O. Box 105, Yuseong, Daejeon 305-600 (Korea, Republic of); Kwon, Sangki [Inha University, Department of Energy Resources Engineering, 253 Yonghyun-dong, Nam-gu, Incheon 402-751 (Korea, Republic of); Choi, Jong-Won [Korea Atomic Energy Research Institute, P.O. Box 105, Yuseong, Daejeon 305-600 (Korea, Republic of)

    2012-11-15

    Highlights: Black-Right-Pointing-Pointer Results of in situ experiments on the near-field of a repository are summarized. Black-Right-Pointing-Pointer In BHT, the rock temperatures at 0.3 m from heater showed 40-50 Degree-Sign C increase. Black-Right-Pointing-Pointer EDZ size measured from the in situ test was in the range of 0.6-1.8 m. Black-Right-Pointing-Pointer Maximum errors in estimating the location of AE source were 0.6-1.0 m. Black-Right-Pointing-Pointer Permeability in the EDZ increased up to 2 orders compared with the intact rock. - Abstract: To obtain the information on the design and performance assessment of a geological repository for nuclear waste, several in situ experiments on the performance of the near-field have been conducted for 5 years in the small-scale underground research laboratory, KURT. This paper summarizes the results from the in situ experiments. In the borehole heater test, the rock temperature at 0.3 m distance from the heater hole with 90 Degree-Sign C showed a 40 Degree-Sign C increase over initial temperature. After heating with 120 Degree-Sign C, the rock temperature at 0.3 m distance from the heater hole increased up to 50 Degree-Sign C over initial temperature. The EDZ size measured from the in situ test was in the range of 0.6-1.8 m, and was higher than that from the laboratory tests, which was estimated to be around 1.1-1.5 m. The maximum errors in estimating the location of acoustic emission source were 1.0 m in EDZ and 0.6 m in an intact rock zone, respectively. The damping ratios of the EDZ and intact rock were 0.091 and 0.005, respectively. The permeability of rock increased with decreasing distance from the tunnel wall because of the EDZ. The permeability in the EDZ seems to be increased up to 2 orders compared with that in the intact rock.

  13. Long Term Sorption Diffusion Experiment (LTDE-SD). Performance of main in situ experiment and results from water phase measurements

    International Nuclear Information System (INIS)

    Widestrand, Henrik; Byegaard, Johan; Nilsson, Kersti; Hoeglund, Susanne; Gustafsson, Erik; Kronberg, Magnus

    2010-12-01

    The LTDE-SD experiment, (Long Term Sorption Diffusion Experiment) aimed at increasing the scientific knowledge of sorption and diffusion under in situ conditions and to provide data for performance and safety assessment calculations. Performance and results of the in situ experiment phase are presented in the report. In total, 21 radionuclide trace elements and one stable trace element were injected, circulated and sampled for ∼6.5 months in a closed borehole section. The trace elements represented non-sorbing tracers and sorbing tracers for which the sorption was dominated by a cation exchange mechanism, a surface complexation mechanism, or dependent on an electrochemical reduction in order to reach the tetravalent state (oxidation state IV) considered very strongly sorbing. The borehole section in contact with the tracer labelled groundwater consisted in part of a natural fracture surface and a borehole section in the unaltered matrix rock, devoid of natural fractures. Water samples were regularly extracted and analysed for trace element concentration and a few ion exchange speciation and filtered samplings were also conducted. Independent colloid filtering and chemical speciation calculations were performed in support the evaluation. Sorption was demonstrated for a series of elements present in the experiment. The amounts lost of the different respective tracers from the aqueous phase follow very well the general understanding of the relative sorption strength of the tracers, as inferred from e.g. batch sorption experiments and dynamic in situ tracer experiments. The chemical speciation calculations of the different tracers were in line with the results of the ion exchange speciation performed during the experiment. With the exception of UO 2 2+ carbonate complexes formed, no strong indications were obtained that aqueous complexation prevents adsorption under the chemical conditions of the experiment. The 20 nm filtered sampling indicated that radionuclide

  14. Long Term Sorption Diffusion Experiment (LTDE-SD). Performance of main in situ experiment and results from water phase measurements

    Energy Technology Data Exchange (ETDEWEB)

    Widestrand, Henrik; Byegaard, Johan; Nilsson, Kersti; Hoeglund, Susanne; Gustafsson, Erik (Geosigma AB, Uppsala (Sweden)); Kronberg, Magnus (Swedish Nuclear Fuel and Waste Management Co. (Sweden))

    2010-12-15

    The LTDE-SD experiment, (Long Term Sorption Diffusion Experiment) aimed at increasing the scientific knowledge of sorption and diffusion under in situ conditions and to provide data for performance and safety assessment calculations. Performance and results of the in situ experiment phase are presented in the report. In total, 21 radionuclide trace elements and one stable trace element were injected, circulated and sampled for approx6.5 months in a closed borehole section. The trace elements represented non-sorbing tracers and sorbing tracers for which the sorption was dominated by a cation exchange mechanism, a surface complexation mechanism, or dependent on an electrochemical reduction in order to reach the tetravalent state (oxidation state IV) considered very strongly sorbing. The borehole section in contact with the tracer labelled groundwater consisted in part of a natural fracture surface and a borehole section in the unaltered matrix rock, devoid of natural fractures. Water samples were regularly extracted and analysed for trace element concentration and a few ion exchange speciation and filtered samplings were also conducted. Independent colloid filtering and chemical speciation calculations were performed in support the evaluation. Sorption was demonstrated for a series of elements present in the experiment. The amounts lost of the different respective tracers from the aqueous phase follow very well the general understanding of the relative sorption strength of the tracers, as inferred from e.g. batch sorption experiments and dynamic in situ tracer experiments. The chemical speciation calculations of the different tracers were in line with the results of the ion exchange speciation performed during the experiment. With the exception of UO{sub 2} 2+ carbonate complexes formed, no strong indications were obtained that aqueous complexation prevents adsorption under the chemical conditions of the experiment. The 20 nm filtered sampling indicated that

  15. Development of the Potassium-Argon Laser Experiment (KArLE) Instrument for In Situ Geochronology

    Science.gov (United States)

    Cohen, Barbara A.; Li, Z.-H.; Miller, J. S.; Brinckerhoff, W. B.; Clegg, S. M.; Mahaffy, P. R.; Swindle, T. D.; Wiens, R. C.

    2012-01-01

    Absolute dating of planetary samples is an essential tool to establish the chronology of geological events, including crystallization history, magmatic evolution, and alteration. Traditionally, geochronology has only been accomplishable on samples from dedicated sample return missions or meteorites. The capability for in situ geochronology is highly desired, because it will allow one-way planetary missions to perform dating of large numbers of samples. The success of an in situ geochronology package will not only yield data on absolute ages, but can also complement sample return missions by identifying the most interesting rocks to cache and/or return to Earth. In situ dating instruments have been proposed, but none have yet reached TRL 6 because the required high-resolution isotopic measurements are very challenging. Our team is now addressing this challenge by developing the Potassium (K) - Argon Laser Experiment (KArLE) under the NASA Planetary Instrument Definition and Development Program (PIDDP), building on previous work to develop a K-Ar in situ instrument [1]. KArLE uses a combination of several flight-proven components that enable accurate K-Ar isochron dating of planetary rocks. KArLE will ablate a rock sample, determine the K in the plasma state using laser-induced breakdown spectroscopy (LIBS), measure the liberated Ar using quadrupole mass spectrometry (QMS), and relate the two by the volume of the ablated pit using an optical method such as a vertical scanning interferometer (VSI). Our preliminary work indicates that the KArLE instrument will be capable of determining the age of several kinds of planetary samples to +/-100 Myr, sufficient to address a wide range of geochronology problems in planetary science.

  16. Spot counting on fluorescence in situ hybridization in suspension images using Gaussian mixture model

    Science.gov (United States)

    Liu, Sijia; Sa, Ruhan; Maguire, Orla; Minderman, Hans; Chaudhary, Vipin

    2015-03-01

    Cytogenetic abnormalities are important diagnostic and prognostic criteria for acute myeloid leukemia (AML). A flow cytometry-based imaging approach for FISH in suspension (FISH-IS) was established that enables the automated analysis of several log-magnitude higher number of cells compared to the microscopy-based approaches. The rotational positioning can occur leading to discordance between spot count. As a solution of counting error from overlapping spots, in this study, a Gaussian Mixture Model based classification method is proposed. The Akaike information criterion (AIC) and Bayesian information criterion (BIC) of GMM are used as global image features of this classification method. Via Random Forest classifier, the result shows that the proposed method is able to detect closely overlapping spots which cannot be separated by existing image segmentation based spot detection methods. The experiment results show that by the proposed method we can obtain a significant improvement in spot counting accuracy.

  17. BACCHUS 2: an in situ backfill hydration experiment for model validation

    International Nuclear Information System (INIS)

    Volckaert, G.; Bernier, F.; Alonso, E.; Gens, A.

    1995-01-01

    The BACCHUS 2 experiment is an in situ backfill hydration test performed in the HADES underground research facility situated in the plastic Boom clay layer at 220 m depth. The experiment aims at the optimization and demonstration of an installation procedure for a clay based backfill material. The instrumentation has been optimized in such a way that the results of the experiments can be used for the validation of hydro-mechanical codes such a NOSAT developed at the University of Catalunya Spain (UPC). The experimental set-up consists in a bottom flange and a central filter around which the backfill material was applied. The backfill material consist of a mixture of high density clay pellets and clay powder. The experimental set-up and its instrumentation are described in detail. The results of the hydro-mechanical characterization of the backfill material is summarized. (authors). 8 refs., 16 figs., 1 tab

  18. CHEMLAB. A probe for in-situ radionuclide experiments. Diffusion studies

    Energy Technology Data Exchange (ETDEWEB)

    Jansson, Mats; Eriksen, Trygve E. [Royal Inst. of Technology, Stockholm (Sweden). Dept. of Chemistry

    2001-06-01

    CHEMLAB is a borehole laboratory built into a probe, in which in situ experiments can be carried out under ambient conditions with respect to pressure and temperature with the use of natural groundwater from the surrounding rock. The first CHEMLAB experiments, diffusion of cations and anions in compacted bentonite clay, have been carried out in borehole KA2512A at a depth of 320 m in Aespoe Hard Rock Laboratory. Diffusant solutions of Co{sup 2+}, Sr{sup 2+}, Cs{sup +}, I{sup -} and TcO{sup 4-} with {sup 57}Co, {sup 85}Sr, {sup 134}Cs, {sup 131}I and {sup 99}Tc as tracers were used. measured profiles for the radionuclides in the bentonite are in good agreement with profiles predicted from modelling based on apparent diffusivities and sorption coefficients obtained in laboratory experiments with clay compacted to corresponding dry density and synthetic ground water with the same composition as in borehole KA2512A.

  19. CHEMLAB. A probe for in-situ radionuclide experiments. Diffusion studies

    International Nuclear Information System (INIS)

    Jansson, Mats; Eriksen, Trygve E.

    2001-06-01

    CHEMLAB is a borehole laboratory built into a probe, in which in situ experiments can be carried out under ambient conditions with respect to pressure and temperature with the use of natural groundwater from the surrounding rock. The first CHEMLAB experiments, diffusion of cations and anions in compacted bentonite clay, have been carried out in borehole KA2512A at a depth of 320 m in Aespoe Hard Rock Laboratory. Diffusant solutions of Co 2+ , Sr 2+ , Cs + , I - and TcO 4- with 57 Co, 85 Sr, 134 Cs, 131 I and 99 Tc as tracers were used. measured profiles for the radionuclides in the bentonite are in good agreement with profiles predicted from modelling based on apparent diffusivities and sorption coefficients obtained in laboratory experiments with clay compacted to corresponding dry density and synthetic ground water with the same composition as in borehole KA2512A

  20. GTS-LCS, in-situ experiment 2. Modeling of tracer test 09-03

    International Nuclear Information System (INIS)

    Manette, M.; Saaltink, M.W.; Soler, J.M.

    2015-02-01

    Within the framework of the GTS-LCS project (Grimsel Test Site - Long-Term Cement Studies), an in-situ experiment lasting about 5 years was started in 2009 to study water-cement-rock interactions in a fractured granite. Prior to the experiment, a tracer test was performed to characterize the initial flow and transport properties of the rock around the experimental boreholes. This study reports on the model interpretation of tracer test 09-03. The calculations were performed by means of a two-dimensional model (homogeneous fracture plane including 3 boreholes) using the Retraso-CodeBright software package. In the tracer test, Grimsel groundwater containing the tracer (uranine) was circulated in the emplacement borehole during 43 days (zero injection flow rate). Circulation continued without tracer afterwards. Water was extracted at the observation and extraction boreholes. Results from a model sensitivity analysis comparing model results with measured tracer concentrations showed 3 cases where the evolution of tracer concentrations in the 3 different boreholes was satisfactory. In these cases a low-permeability skin affected the emplacement and observation boreholes. No skin appeared to affect the extraction borehole. The background hydraulic gradient seems to have no effect on the results of the tracer test. These results will be applied in the calculation of the initial flow field for the reactive transport phase of in-situ experiment 2 (interaction between pre-hardened cement and fractured granite at Grimsel). (orig.)

  1. Identification of Fetal Inflammatory Cells in Eosinophilic/T-cell Chorionic Vasculitis Using Fluorescent In Situ Hybridization.

    Science.gov (United States)

    Katzman, Philip J; Li, LiQiong; Wang, Nancy

    2015-01-01

    Eosinophilic/T-cell chorionic vasculitis (ETCV) is an inflammatory lesion of placental fetal vessels. In contrast to acute chorionic vasculitis, inflammation in ETCV is seen in chorionic vessel walls opposite the amnionic surface. It is not known whether inflammation in ETCV consists of maternal cells from the intervillous space or fetal cells migrating from the vessel. We used fluorescent in situ hybridization (FISH) to differentiate fetal versus maternal cells in ETCV. Placentas with ETCV, previously identified for a published study, were used. Infant sex in each case was identified using the electronic medical record. For male infants, 3-μm sections were cut from archived tissue blocks from placentas involving ETCV and stained with fluorescent X- and Y-chromosome centromeric probes. A consecutive hematoxylin/eosin-stained section was used for correlation. FISH analysis was performed on 400 interphase nuclei at the site of ETCV to determine the proportion of XX, XY, X, and Y cells. Of 31 ETCV cases, 20 were female and 10 were male (1 sex not recorded). Six of 10 cases with male infants had recuts with visible ETCV. In these 6 cases the average percentages (ranges) of XY cells, X-only cells, and Y-only cells in the region of inflammation were 81 (70-90), 11 (6-17), and 8 (2-14), respectively. There was a 2:1 female:male infant ratio in ETCV. Similar to acute chorionic vasculitis, the inflammation in ETCV is of fetal origin. It is still unknown, however, whether the stimulus for ETCV is of fetal or maternal origin.

  2. 6q deletion detected by fluorescence in situ hybridization using bacterial artificial chromosome in chronic lymphocytic leukemia.

    Science.gov (United States)

    Dalsass, Alessia; Mestichelli, Francesca; Ruggieri, Miriana; Gaspari, Paola; Pezzoni, Valerio; Vagnoni, Davide; Angelini, Mario; Angelini, Stefano; Bigazzi, Catia; Falcioni, Sadia; Troiani, Emanuela; Alesiani, Francesco; Catarini, Massimo; Attolico, Immacolata; Scortechini, Ilaria; Discepoli, Giancarlo; Galieni, Piero

    2013-07-01

    Deletions of the long arm of chromosome 6 are known to occur at relatively low frequency (3-6%) in chronic lymphocytic leukemia (CLL), and they are more frequently observed in 6q21. Few data have been reported regarding other bands on 6q involved by cytogenetic alterations in CLL. The cytogenetic study was performed in nuclei and metaphases obtained after stimulation with a combination of CpG-oligonucleotide DSP30 and interleukin-2. Four bacterial artificial chromosome (BAC) clones mapping regions in bands 6q16, 6q23, 6q25, 6q27 were used as probes for fluorescence in situ hybridization in 107 CLL cases in order to analyze the occurrence and localization of 6q aberrations. We identified 11 cases (10.2%) with 6q deletion of 107 patients studied with CLL. The trends of survival curves and the treatment-free intervals (TFI) of patients with deletion suggest a better outcome than the other cytogenetic risk groups. We observed two subgroups with 6q deletion as the sole anomaly: two cases with 6q16 deletion, and three cases with 6q25.2-27 deletion. There were differences of age, stage, and TFI between both subgroups. By using BAC probes, we observed that 6q deletion has a higher frequency in CLL and is linked with a good prognosis. In addition, it was observed that the deletion in 6q16 appears to be the most frequent and, if present as the only abnormality, it could be associated with a most widespread disease. © 2013 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  3. Flat mount preparation for observation and analysis of zebrafish embryo specimens stained by whole mount in situ hybridization.

    Science.gov (United States)

    Cheng, Christina N; Li, Yue; Marra, Amanda N; Verdun, Valerie; Wingert, Rebecca A

    2014-07-17

    The zebrafish embryo is now commonly used for basic and biomedical research to investigate the genetic control of developmental processes and to model congenital abnormalities. During the first day of life, the zebrafish embryo progresses through many developmental stages including fertilization, cleavage, gastrulation, segmentation, and the organogenesis of structures such as the kidney, heart, and central nervous system. The anatomy of a young zebrafish embryo presents several challenges for the visualization and analysis of the tissues involved in many of these events because the embryo develops in association with a round yolk mass. Thus, for accurate analysis and imaging of experimental phenotypes in fixed embryonic specimens between the tailbud and 20 somite stage (10 and 19 hours post fertilization (hpf), respectively), such as those stained using whole mount in situ hybridization (WISH), it is often desirable to remove the embryo from the yolk ball and to position it flat on a glass slide. However, performing a flat mount procedure can be tedious. Therefore, successful and efficient flat mount preparation is greatly facilitated through the visual demonstration of the dissection technique, and also helped by using reagents that assist in optimal tissue handling. Here, we provide our WISH protocol for one or two-color detection of gene expression in the zebrafish embryo, and demonstrate how the flat mounting procedure can be performed on this example of a stained fixed specimen. This flat mounting protocol is broadly applicable to the study of many embryonic structures that emerge during early zebrafish development, and can be implemented in conjunction with other staining methods performed on fixed embryo samples.

  4. Detection of Tritrichomonas foetus and Pentatrichomonas hominis in intestinal tissue specimens of cats by chromogenic in situ hybridization.

    Science.gov (United States)

    Mostegl, Meike M; Wetscher, Andreas; Richter, Barbara; Nedorost, Nora; Dinhopl, Nora; Weissenböck, Herbert

    2012-02-10

    In this retrospective study 102 cats were analyzed for the presence of trichomonads in intestinal tissue sections using chromogenic in situ hybridization (CISH). Two intestinal trichomonad species are described in cats: Pentatrichomonas hominis and Tritrichomonas foetus. While P. hominis is considered a mere commensal, T. foetus has been found to be the causative agent of feline large-bowel diarrhea. For the detection of both agents within intestinal tissue CISH assays using three different probes were performed. In the first CISH run a probe specific for all relevant members of the order Trichomonadida (OT probe) was used. In a second CISH run all positive samples were further examined on three consecutive tissue sections using the OT probe, a probe specific for the family of Tritrichomonadidae (Tritri probe) and a newly designed probe specifically detecting P. hominis (Penta hom probe). In total, four of the 102 cats were found to be positive with the OT probe. Thereof, one cat gave a positive reaction with the P. hominis probe and three cats were positive with the T. foetus probe. All Trichomonas-positive cats were pure-bred and between 8 and 32 weeks of age. In one cat positive for T. foetus large amounts of parasites were found in the gut lumen and invading the intestinal mucosa. The species of the detected trichomonads were confirmed by polymerase chain reaction and nucleotide sequencing of a part of the 18S ribosomal RNA gene. In this study, the usefulness of CISH to detect intestinal trichomonads within feline tissue samples was shown. Additionally, the specific detection of P. hominis using CISH was established. Generally, it was shown that CISH is well suited for detection and differentiation of trichomonosis in retrospective studies using tissue samples. Copyright © 2011 Elsevier B.V. All rights reserved.

  5. Tubular and endothelial chimerism in renal allografts using fluorescence and chromogenic in situ hybridization (FISH, CISH) technology.

    Science.gov (United States)

    Varga, Zsuzsanna; Gaspert, Ariana; Behnke, Silvia; von Teichman, Adriana; Fritzsche, Florian; Fehr, Thomas

    2012-04-01

    The role of endothelial and tubular chimerism in renal allograft adaptation and rejection varies in different studies. We addressed the correlation between different clinico-pathological settings and sex-chromosomal endothelial and/or tubular chimerism in renal allografts. We examined the presence or absence of the X and Y chromosomes by fluorescence and chromogenic in situ hybridization (FISH, CISH) methodology on paraffin embedded kidney biopsies in 16 gender mismatched renal transplants (1 to 12 years post-transplantation). Twelve patients were male, four female. Four groups were selected: (i) Vascular calcineurin inhibitor toxicity without rejection; (ii) T-cell mediated vascular rejection; (iii) antibody mediated rejection; and (iv) C4d-positivity in AB0-incompatible transplants with or without rejection. Twelve non-transplant kidney biopsies (8 female, 4 male) were used as controls. Tubular chimerism was detected more frequently (69%) than endothelial chimerism (12%) in renal transplants. One of 12 control patients had tubular and endothelial chimeric cells (8%). The Y chromosome occurred in 8/12 male recipients (67%) in tubular epithelial cells and in 5/12 male recipients (42%) in endothelial cells. Double X chromosomes were detected in 3/4 female recipients in tubular epithelium. Tubular chimerism occurred more often with endothelial chimerism and capillaritis without correlation with other parameters, such as rejection. Combined Y chromosomal tubular and lymphatic endothelial chimerism correlated with T-cell mediated vascular rejection in two out of three patients (66%). Combined Y chromosomal tubular and peritubular capillary chimerism correlated with antibody mediated C4d+ rejection in one out of two patients (50%). Tubular and/or endothelial chimerism occur frequently in gender mismatched renal allografts and, when combined, this is associated with T-cell mediated rejection. © 2012 The Authors. Pathology International © 2012 Japanese Society of

  6. Determining the correlation of Epstein-Barr virus with diffuse large B-cell lymphoma by chromogenic in situ hybridization

    Directory of Open Access Journals (Sweden)

    Kosari F

    2012-09-01

    Full Text Available Background: Diffuse large B-cell lymphoma (DLBCL is the most common type of lymphoma. There are various types of DLBCL including immunoblastic and centroblastic. Epstein-Barr virus (EBV is a member of Herpes virus family found in all human populations inducing different lymphoproliferative disorders. The role of EBV in the development of DLBCL is known. Multiple laboratory methods are available for detecting EBV. This study was conducted to determine the correlation of EBV with DLBCL in samples referred to pathology ward in Shariati and Sina Hospitals by chromogenic in situ hybridization (CISH method.Methods: In this case/control study, pathological specimens of 50 patients with DLBCL as well as 50 reactive lymph nodes and tonsils (control group were collected from archives of Shariati and Sina Hospitals and were evaluated for EBV encoded RNA (EBER expression based on CISH method. A peptide nucleic acid (PNA EBV probe (Dakocytomatin was used while all the processes were done in RNAase-free conditions using RNAase-free water, sterile gloves and samplers. Results: Out of fifty specimens in the case group, eight were positive for EBER in comparison with two in the control group (P=0.046. No statistically significant difference was observed between intranodal or extranodal samples (P=0.736 or between males and females (P=0.0746.Conclusion: Our study showed that EBV positivity for EBER in patient with DLBCL could be determined more effectively by CISH method than immunohistochemistry (IHC. Comparative analysis between CISH, PCR and IHC methods is recommended.

  7. Rapid differentiation of Francisella species and subspecies by fluorescent in situ hybridization targeting the 23S rRNA

    Directory of Open Access Journals (Sweden)

    Trebesius Karlheinz

    2010-03-01

    Full Text Available Abstract Background Francisella (F. tularensis is the causative agent of tularemia. Due to its low infectious dose, ease of dissemination and high case fatality rate, F. tularensis was the subject in diverse biological weapons programs and is among the top six agents with high potential if misused in bioterrorism. Microbiological diagnosis is cumbersome and time-consuming. Methods for the direct detection of the pathogen (immunofluorescence, PCR have been developed but are restricted to reference laboratories. Results The complete 23S rRNA genes of representative strains of F. philomiragia and all subspecies of F. tularensis were sequenced. Single nucleotide polymorphisms on species and subspecies level were confirmed by partial amplification and sequencing of 24 additional strains. Fluorescent In Situ Hybridization (FISH assays were established using species- and subspecies-specific probes. Different FISH protocols allowed the positive identification of all 4 F. philomiragia strains, and more than 40 F. tularensis strains tested. By combination of different probes, it was possible to differentiate the F. tularensis subspecies holarctica, tularensis, mediasiatica and novicida. No cross reactivity with strains of 71 clinically relevant bacterial species was observed. FISH was also successfully applied to detect different F. tularensis strains in infected cells or tissue samples. In blood culture systems spiked with F. tularensis, bacterial cells of different subspecies could be separated within single samples. Conclusion We could show that FISH targeting the 23S rRNA gene is a rapid and versatile method for the identification and differentiation of F. tularensis isolates from both laboratory cultures and clinical samples.

  8. Fluorescence In Situ Hybridization and Immunohistochemistry as Diagnostic Methods for ALK Positive Non-Small Cell Lung Cancer Patients

    Science.gov (United States)

    Martinez, Pablo; Hernández-Losa, Javier; Cedrés, Susana; Castellví, Josep; Martinez-Marti, Alex; Tallada, Natalia; Murtra-Garrell, Nuria; Navarro-Mendivill, Alejandro; Rodriguez-Freixinos, Victor; Canela, Mercedes; Ramon y Cajal, Santiago; Felip, Enriqueta

    2013-01-01

    Background Anaplastic Lymphoma Kinase (ALK) positivity represents a novel molecular target in a subset of Non-Small Cell Lung Cancers (NSCLC). We explore Fluorescence in situ Hybridization (FISH) and Immunohistochemistry (IHC) as diagnostic methods for ALK positive patients and to describe its prevalence and outcomes in a population of NSCLC patients. Methods NSCLC patients previously screened for Epidermal Growth Factor Receptor (EGFR) at our institution were selected. ALK positive patients were identified by FISH and the value of IHC (D5F3) was explored. Results ninety-nine patients were identified. Median age was 61.5 years (range 35–83), all were caucasians, eighty percent were adenocarcinomas, fifty-one percent were male and thirty-eight percent were current smokers. Seven (7.1%) patients were ALK positive by FISH, thirteen (13.1%) were EGFR mutant, and 65 (65.6%) were negative/Wild Type (WT) for both ALK and EGFR. ALK positivity and EGFR mutations were mutually exclusive. ALK positive patients tend to be younger than EGFR mutated or wt patients. ALK positive patients were predominantly never smokers (71.4%) and adenocarcinoma (71.4%). ALK positive and EGFR mutant patients have a better outcome than negative/WT. All patients with ALK FISH negative tumours were negative for ALK IHC. Out of 6 patients positive for ALK FISH with more tissue available, 5 were positive for ALK IHC and 1 negative. Conclusions ALK positive patients represent 7.1% of a population of selected NSCLC. ALK positive patients have different clinical features and a better outcome than EGFR WT and ALK negative patients. IHC is a promising method for detecting ALK positive NSCLC patients. PMID:23359795

  9. Evaluation of fluorescent in-situ hybridization technique for diagnosis of malaria in Ahero Sub-County hospital, Kenya.

    Science.gov (United States)

    Kandie, Regina; Ochola, Rachel; Njaanake, Kariuki

    2018-01-08

    Malaria is a major cause of morbidity and mortality. Treatment of malaria in a timely manner could avert deaths. Treatment ultimately relies on the rapid and accurate diagnosis. Fluorescence in situ hybridization (FISH), a cytogenetic technique based on detection of specific nucleic acid, has the potential to address the limitations of the current diagnostic approaches. This study investigates further the performance of FISH for the diagnosis of malaria in a rural setting in Western Kenya. Blood samples from 302 patients presenting with fever (temperature ≥ 37.5 °C) were examined for malaria using the Giemsa microscopy (GM), rapid diagnostic test (RDT), polymerase chain reaction (PCR) and FISH. The sensitivity and specificity of FISH was 85.6% and 96.2% respectively, while the corresponding values for GM were 82.2% and 100% respectively. RDT and PCR had sensitivities of 91.1% and 98.9%, respectively with their specificities being 89.6 and 100%, respectively. The positive predictive values for RDT, GM, FISH and PCR were 78.8%, 100%, 90.6% and 100%, respectively. The negative predictive values for RDT, GM, FISH and PCR were 96.0%, 93.0%, 94.0% and 99.5%, respectively. Their respective diagnostic accuracies were 90.1%, 94.7% 93.0% and 99.7%. The present study demonstrates that the specificity and reproducibility of FISH assays are high, thus adding to the growing evidence on the potential of the technique as an effective tool for the detection of malaria parasites in remote settings.

  10. Role of Tetrasomy for the Diagnosis of Urothelial Carcinoma Using UroVysion Fluorescent In Situ Hybridization.

    Science.gov (United States)

    Zhou, Amy G; Liu, Yuxin; Cyr, Maryann St; Garver, Joanne; Woda, Bruce A; Cosar, Ediz F; Hutchinson, Lloyd M

    2016-06-01

    -UroVysion fluorescent in situ hybridization (FISH) is routinely used to detect urothelial carcinoma (UC). A positive threshold is defined as chromosome polysomy in 4 or more cells, which also includes tetrasomy, a natural product of cell division. -To evaluate tetrasomy for UC detection and explore the relation to the surgical diagnosis or patient history. -The FISH was performed on 1532 urine samples from patients with cytology results and 4 or more years of follow-up. We created separate polysomy and tetrasomy categories and constructed receiver operating curves to determine appropriate thresholds using biopsy (n = 194) as the gold standard. Standard FISH and a novel assay integrating cytomorphology and FISH (Target-FISH) were compared. Matching tissue biopsies of urine samples with 10 or more tetrasomy cells were analyzed. -No significant threshold was found for tetrasomy cells. Exclusion of tetrasomy from the polysomy category changed the threshold from 8.5 to 4.5 cells, increased specificity (59.2% to 78.9%), but reduced sensitivity (78.9% to 65.9%). In Target-FISH, the same approach yielded a specificity of 93.7% and sensitivity of 65.2%. Similarly, specificity improved significantly for low- and high-grade UC, but sensitivity decreased for low-grade UC. No evidence of UC was observed in 95% (52 of 55) of the patients referred for screening who had 10 or more tetrasomy cells by FISH. Matching biopsies for urines containing 10 or more tetrasomy cells showed few or no tetrasomy cells. -Tetrasomy is a nonspecific finding frequently encountered in urine FISH and should be excluded from the polysomy classification. Target-FISH is an optimal approach, offering the ability to detect rare tetrasomy tumors.

  11. INTERRUPTED IN-SITU COMPRESSIVE DEFORMATION EXPERIMENTS ON MMC FOAMS IN AN XCT: EXPERIMENTS AND ESTIMATION OF DISPLACEMENT FIELDS

    Directory of Open Access Journals (Sweden)

    Katharina Losch

    2014-05-01

    Full Text Available The mechanical properties of a metal-matrix composite foam are investigated by interrupted in-situ compressive deformation experiments within an X-ray computed tomography device (XCT. Each in-situ experiment generates a sequence of reconstructed 3D images of the foam microstructure. From these data, the deformation field is estimated by registring the images corresponding to three consecutive steps. To this end, the generic registration framework of the itk software suite is exploited and combined with several image preprocessing steps. Both segmented (binary images having just two grey values for foreground (strut structure and background (pore space and the result of the Euclidean distance transform (EDT on pore space and solid phase are used. The estimation quality is evaluated based on a sequence of synthetic data sets, where the foam’s microstructure is modelled by a random Laguerre tessellation. For large deformations, a combination of non-rigid registration for the EDT images and partwise-rigid registration on strongly deformed regions of the binary images, yields surprisingly small estimation errors.

  12. Lower Hybrid Current Drive Experiments in Alcator C-Mod

    International Nuclear Information System (INIS)

    Wilson, J. R.; Bonoli, P.; Hubbard, A.; Parker, R.; Schmidt, A.; Wallace, G.; Wright, J.; Bernabei, S.

    2007-01-01

    A Lower Hybrid Current Drive (LHCD) system has been installed on the Alcator C-MOD tokamak at MIT. Twelve klystrons at 4.6 GHz feed a 4x22 waveguide array. This system was designed for maximum flexibility in the launched parallel wave-number spectrum. This flexibility allows tailoring of the lower hybrid deposition under a variety of plasma conditions. Power levels up to 900 kW have been injected into the tokomak. The parallel wave number has been varied over a wide range, n parallel ∼1.6-4. Driven currents have been inferred from magnetic measurements by extrapolating to zero loop voltage and by direct comparison to Fisch-Karney theory, yielding an efficiency of n 20 IR/P∼0.3. Modeling using the CQL3D code supports these efficiencies. Sawtooth oscillations vanish, accompanied with peaking of the electron temperature (T e0 rises from 2.8 to 3.8 keV). Central q is inferred to rise above unity from the collapse of the sawtooth inversion radius, indicating off-axis cd as expected. Measurements of non-thermal x-ray and electron cyclotron emission confirm the presence of a significant fast electron population that varies with phase and plasma density. The x-ray emission is observed to be radialy broader than that predicted by simple ray tracing codes. Possible explanations for this broader emission include fast electron diffusion or broader deposition than simple ray tracing predictions (perhaps due to diffractive effects)

  13. One-pot in situ redox synthesis of hexacyanoferrate/conductive polymer hybrids as lithium-ion battery cathodes.

    Science.gov (United States)

    Wong, Min Hao; Zhang, Zixuan; Yang, Xianfeng; Chen, Xiaojun; Ying, Jackie Y

    2015-09-14

    An efficient and adaptable method is demonstrated for the synthesis of lithium hexacyanoferrate/conductive polymer hybrids for Li-ion battery cathodes. The hybrids were synthesized via a one-pot method, involving a redox-coupled reaction between pyrrole monomers and the Li3Fe(CN)6 precursor. The hybrids showed much better cyclability relative to reported Prussian Blue (PB) analogs.

  14. USING THE US EXPERIENCE OF ONLINE AND HYBRID EDUCATION IN UKRAINIAN UNIVERSITIES

    Directory of Open Access Journals (Sweden)

    Ірина Задорожна

    2015-12-01

    Full Text Available The article investigates the best practices of the US universities on providing hybrid and online education that can be implemented at Ukrainian universities which experience lack of finances and a decrease in student enrollment. Basic factors of online and hybrid courses popularity are analysed (flexibility; accessibility, especially for students with special needs; saving time; convenience; motivation as students often feel less stressed in a virtual classroom than in a face-to-face environment. The main challenges of online and hybrid learning (faculty training and professional development for online education (in terms of pedagogy, communication and technology, supporting student learning in the online environment, and creating an efficient and interactive online learning community are defined in the article. Hybrid courses are regarded as such that promote some equivalence between digital and live communication.

  15. TNB Experience in Developing Solar Hybrid Station at RPS Kemar, Gerik, Perak Darul Ridzuan

    International Nuclear Information System (INIS)

    Aziz, K A; Shamsudin, K N

    2013-01-01

    This paper will discuss on TNB experience in developing Solar Hybrid Station at RPS Kemar, Gerik, Perak. TNB has been approached by KKLW to submit proposal to provide electricity in the rural area namely RPS Kemar. Looking at area and source available, Solar Hybrid System was the best method in order to provide electricity at this area. This area is far from national grid sources. Solar Hybrid System is the best method to produce electrical power using the renewable energy from Solar PV, Battery and Diesel Generator Set. Nowadays, price of petroleum is slightly high due to higher demand from industry. Solar energy is good alternative in this country to practice in order to reduce cost for produce of electrical energy. Generally, Solar will produce energy during daytime and when become cloudy and dark, automatically battery and diesel generator set will recover the system through the hybrid controller system.

  16. Utility and impact of early t(15;17) identification by Fluorescence In Situ Hybridization (FISH) in clinical decision making for patients in Acute Promyelocytic Leukemia (APL).

    Science.gov (United States)

    Kolhe, R; Mangaonkar, A; Mansour, J; Clemmons, A; Shaw, J; Dupont, B; Walczak, L; Mondal, A; Rojiani, A; Jillella, A; Kota, V

    2015-08-01

    Acute Promyelocytic Leukemia (APL) is a curable malignancy with studies showing above 90% survival. However, population-based studies looking at survival suggest that approximately 30% of patients with APL die during induction. Early demonstration of t(15;17) will lead to accurate decision making regarding treatment. The aim of this project was to validate earlier time frames for the Abbott Molecular Vysis LSI promyelocytic leukemia (PML)/ retinoic acid receptor alpha (RARA) fluorescence in situ hybridization (FISH) probe (ASR 6-16 h). Twenty patients (15 APL cases and five non-APL cases) were selected for validating various hybridization times for the FISH probe. Expected normal signal pattern was two red and two green signals (2R2G), and the most common expected abnormal signal pattern was two fusion (yellow) signals, one red and one green (2F1R1G) and/or one fusion, one red and one green (1F1R1G). The specificity of the probe ranged from 84% at 2 h, 86% at 4 h, 84% at 6 h, and 87% for overnight hybridization. The sensitivity increased from 79% at 2 h, 80% at 4 h, 81% at 6 h to 87% for overnight hybridization. Based on the validation studies, we recommend reading of FISH results at the 4-h incubation mark for a preliminary diagnosis and confirmation with overnight hybridization. © 2015 John Wiley & Sons Ltd.

  17. Lower hybrid current drive experiments with graphite limiters in the HT-7 superconducting tokamak

    International Nuclear Information System (INIS)

    Liu, J.; Gao, X.; Hu, L.Q.; Asif, M.; Chen, Z.Y.; Ding, B.J.; Zhou, Q.; Liu, H.Q.; Jie, Y.X.; Kong, W.; Lin, S.Y.; Ding, Y.H.; Gao, L.; Xu, Q.

    2006-01-01

    Recent progress of lower hybrid (LH) experiments with new graphite limiters configuration in the HT-7 tokamak is presented. The lower hybrid current drive (LHCD) efficiency can be determined by fitting based on experimental data. Improved particle confinement was observed via LHCD (P LHW >300 kW) characterized by the particle confinement time τ p increased about 1.56 times. It is found that runaways are suppressed during loop voltage is decreasing at the flat-top phase of LH discharges. The main limitations of pulse length are presented in long-pulse experiments with new limiter configuration

  18. In situ TEM experiments of electrochemical lithiation and delithiation of individual nanostructures

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Xiao Hua; Liu, Yang; Huang, Jian Yu [Center for Integrated Nanotechnologies (CINT), Sandia National Laboratories, Albuquerque, New Mexico (United States); Kushima, Akihiro; Li, Ju [Department of Nuclear Science and Engineering and Department of Materials Science and Engineering, Massachusetts Institute of Technology, Cambridge, Massachusetts (United States); Zhang, Sulin [Department of Engineering Science and Mechanics, Pennsylvania State University, University Park, Pennsylvania (United States); Zhu, Ting [Woodruff School of Mechanical Engineering, Georgia Institute of Technology, Atlanta, Georgia (United States)

    2012-07-15

    O, but not of SnO{sub 2}, after a small amount of lithium insertion. In metallic nanowires such as Al, delithiation causes pulverization, and the product nanoparticles are held in place by the surface Li-Al-O glass tube, suggesting possible strategies for improving electrode cyclability by coatings. In addition, a new in situ chemical lithiation method is introduced for fast screening of battery materials by conventional TEM. Evidently, in situ nanobattery experiments inside TEM are a powerful approach for advancing the fundamental understanding of electrochemical reactions and materials degradation and therefore pave the way toward rational design of high-performance LIBs. (Copyright copyright 2012 WILEY-VCH Verlag GmbH and Co. KGaA, Weinheim)

  19. In-situ experiments to investigate rock matrix retention properties in ONKALO, Olkiluoto, Finland

    Energy Technology Data Exchange (ETDEWEB)

    Voutilainen, Mikko; Helariutta, Kerttuli [Helsinki Univ. (Finland). Dept. of Chemistry; Poteri, Antti [Technical Research Centre of Finland VTT (Finland); and others

    2015-07-01

    Spent nuclear fuel from nuclear power plants, owned by TVO (Teollisuuden Voima Oy) and Fortum, is planned to be disposed to a repository at a depth of more than 400 meters in the bedrock of Olkiluoto (Eurajoki, Finland). The repository system of multiple release barriers consists of both manmade and natural barriers. The surrounding rock acts as the last barrier if other barriers fail during passage of the millennia. Therefore, safe disposal of spent nuclear fuel requires information on the radionuclide transport and retention properties within the porous and water-containing rock matrix along the water conducting flow paths. To this end, various types of experiments are being performed and planned within ONKALO, the underground rock characterization facility in Olkiluoto, as part of the project @''rock matrix REtention PROperties'' (REPRO). The research site is located at a depth of 420 meters close to the repository site. The aim is to study the diffusion and sorption properties of nuclear compounds in the rock matrix under real in-situ conditions. The first in-situ experiment was performed during 2012 using HTO, Na-22, Cl-36 and I-125 as tracer nuclides. Breakthrough curves show retention and asymptotic behavior that are in-line with those caused by matrix diffusion and sorption were observed in their breakthrough curves. Weak sorption was also observed in the breakthrough curves of Na-22 and I-125.

  20. Development of a 16S rRNA-targeted fluorescence in situ hybridization probe for quantification of the ammonia-oxidizer Nitrosotalea devanaterra and its relatives.

    Science.gov (United States)

    Restrepo-Ortiz, C X; Merbt, S N; Barrero-Canossa, J; Fuchs, B M; Casamayor, E O

    2018-04-28

    The Thaumarchaeota SAGMCG-1 group and, in particular, members of the genus Nitrosotalea have high occurrence in acidic soils, the rhizosphere, groundwater and oligotrophic lakes, and play a potential role in nitrogen cycling. In this study, the specific oligonucleotide fluorescence in situ hybridization probe SAG357 was designed for this Thaumarchaeota group based on the available 16S rRNA gene sequences in databases, and included the ammonia-oxidizing species Nitrosotalea devanaterra. Cell permeabilization for catalyzed reporter deposition fluorescence in situ detection and the hybridization conditions were optimized on enrichment cultures of the target species N. devanaterra, as well as the non-target ammonia-oxidizing archaeon Nitrosopumilus maritimus. Probe specificity was improved with a competitor oligonucleotide, and fluorescence intensity and cell visualization were enhanced by the design and application of two adjacent helpers. Probe performance was tested in soil samples along a pH gradient, and counting results matched the expected in situ distributions. Probe SAG357 and the CARD-FISH protocol developed in the present study will help to improve the current understanding of the ecology and physiology of N. devanaterra and its relatives in natural environments. Copyright © 2018 Elsevier GmbH. All rights reserved.

  1. Advanced approach to the analysis of a series of in-situ nuclear forward scattering experiments

    Energy Technology Data Exchange (ETDEWEB)

    Vrba, Vlastimil, E-mail: vlastimil.vrba01@upol.cz [Department of Experimental Physics, Faculty of Science, Palacký University Olomouc, 17. listopadu 12, 771 46 Olomouc (Czech Republic); Procházka, Vít, E-mail: v.prochazka@upol.cz [Department of Experimental Physics, Faculty of Science, Palacký University Olomouc, 17. listopadu 12, 771 46 Olomouc (Czech Republic); Smrčka, David, E-mail: david.smrcka@upol.cz [Department of Experimental Physics, Faculty of Science, Palacký University Olomouc, 17. listopadu 12, 771 46 Olomouc (Czech Republic); Miglierini, Marcel, E-mail: marcel.miglierini@stuba.sk [Slovak University of Technology in Bratislava, Faculty of Electrical Engineering and Information Technology, Institute of Nuclear and Physical Engineering, Ilkovicova 3, 812 19 Bratislava (Slovakia); Department of Nuclear Reactors, Faculty of Nuclear Science and Physical Engineering, Czech Technical University in Prague, V Holešovičkách 2, 180 00 Prague (Czech Republic)

    2017-03-01

    This study introduces a sequential fitting procedure as a specific approach to nuclear forward scattering (NFS) data evaluation. Principles and usage of this advanced evaluation method are described in details and its utilization is demonstrated on NFS in-situ investigations of fast processes. Such experiments frequently consist of hundreds of time spectra which need to be evaluated. The introduced procedure allows the analysis of these experiments and significantly decreases the time needed for the data evaluation. The key contributions of the study are the sequential use of the output fitting parameters of a previous data set as the input parameters for the next data set and the model suitability crosscheck option of applying the procedure in ascending and descending directions of the data sets. Described fitting methodology is beneficial for checking of model validity and reliability of obtained results.

  2. Advanced approach to the analysis of a series of in-situ nuclear forward scattering experiments

    International Nuclear Information System (INIS)

    Vrba, Vlastimil; Procházka, Vít; Smrčka, David; Miglierini, Marcel

    2017-01-01

    This study introduces a sequential fitting procedure as a specific approach to nuclear forward scattering (NFS) data evaluation. Principles and usage of this advanced evaluation method are described in details and its utilization is demonstrated on NFS in-situ investigations of fast processes. Such experiments frequently consist of hundreds of time spectra which need to be evaluated. The introduced procedure allows the analysis of these experiments and significantly decreases the time needed for the data evaluation. The key contributions of the study are the sequential use of the output fitting parameters of a previous data set as the input parameters for the next data set and the model suitability crosscheck option of applying the procedure in ascending and descending directions of the data sets. Described fitting methodology is beneficial for checking of model validity and reliability of obtained results.

  3. In situ water and gas injection experiments performed in the Hades Underground Research Facility

    Energy Technology Data Exchange (ETDEWEB)

    Volckaert, G.; Ortiz, L.; Put, M. [SCK-CEN, Mol (Belgium). Geological Waste Disposal Unit

    1995-12-31

    The movement of water and gas through plastic clay is an important subject in the research at SCK-CEN on the possible disposal of high level radioactive waste in the Boom clay layer at Mol. Since the construction of the Hades underground research facility in 1983, SCK-CEN has developed and installed numerous piezometers for the geohydrologic characterization and for in situ radionuclide migration experiments. In situ gas and water injection experiments have been performed at two different locations in the underground laboratory. The first location is a multi filter piezometer installed vertically at the bottom of the shaft in 1986. The second location is a three dimensional configuration of four horizontal multi piezometers installed from the gallery. This piezometer configuration was designed for the MEGAS (Modelling and Experiments on GAS migration through argillaceous rocks) project and installed in 1992. It contains 29 filters at distances between 10 m and 15 m from the gallery in the clay. Gas injection experiments show that gas breakthrough occurs at a gas overpressure of about 0.6 MPa. The breakthrough occurs by the creation of gas pathways along the direction of lowest resistance i.e. the zone of low effective stress resulting from the drilling of the borehole. The water injections performed in a filter -- not used for gas injection -- show that the flow of water is also influenced by the mechanical stress conditions. Low effective stress leads to higher hydraulic conductivity. However, water overpressures up to 1.3 MPa did not cause hydrofracturing. Water injections performed in a filter previously used for gas injections, show that the occluded gas hinders the water flow and reduces the hydraulic conductivity by a factor two.

  4. Initial experience in hybrid PET-MRI for evaluation of refractory focal onset epilepsy.

    Science.gov (United States)

    Shin, Hae W; Jewells, Valerie; Sheikh, Arif; Zhang, Jingwen; Zhu, Hongtu; An, Hongyu; Gao, Wei; Shen, Dinggang; Hadar, Eldad; Lin, Weili

    2015-09-01

    We aim to evaluate the utility/improved accuracy of hybrid PET/MR compared to current practice separate 3T MRI and PET-CT imaging for localization of seizure foci. In a pilot study, twenty-nine patients undergoing epilepsy surgery evaluation were imaged using PET/MR. This subject group had 29 previous clinical 3T MRI as well as 12 PET-CT studies. Prior clinical PET and MR images were read sequentially while the hybrid PET/MR was concurrently read. The median interval between hybrid PET/MR and prior imaging studies was 5 months (range 1-77 months). In 24 patients, there was no change in the read between the clinical exams and hybrid PET/MR while new anatomical or functional lesions were identified by hybrid PET/MR in 5 patients without significant clinical change. Four new anatomical MR lesions were seen with concordant PET findings. The remaining patient revealed a new abnormal PET lesion without an MR abnormality. All new PET/MR lesions were clinically significant with concordant EEG and/or SPECT results as potential epileptic foci. Our initial hybrid PET-MRI experience increased diagnostic yields for detection of potential epileptic lesions. This may be due to the unique advantage of improved co-registration and simultaneous review of both structural and functional data. Copyright © 2015 British Epilepsy Association. Published by Elsevier Ltd. All rights reserved.

  5. Hybridization experiments indicate incomplete reproductive isolating mechanism between Fasciola hepatica and Fasciola gigantica.

    Science.gov (United States)

    Itagaki, T; Ichinomiya, M; Fukuda, K; Fusyuku, S; Carmona, C

    2011-09-01

    Experiments on hybridization between Fasciola hepatica and Fasciola gigantica were carried out to clarify whether a reproductive isolating mechanism appears between the two Fasciola species. Molecular evidence for hybridization was based on the DNA sequence of the internal transcribed spacer 1 (ITS1) region in nuclear ribosomal DNA, which differs between the species. The results suggested that there were not pre-mating but post-mating isolating mechanisms between the two species. However, viable adults of the hybrids F1 and F2 were produced from both parental F. hepatica and F. gigantica. The hybrids inherited phenotypic characteristics such as ratio of body length and width and infectivity to rats from parental Fasciola hepatica and F. gigantica. These findings suggest that reproductive isolation is incomplete between Fasciola hepatica and F. gigantica. Adults of the hybrids F1 and F2 were completely different in mode of reproduction from aspermic Fasciola forms that occur in Asia and seem to be offspring originated from hybridization between F. hepatica and F. gigantica and to reproduce parthenogenetically.

  6. Interfacing with in-Situ Data Networks during the Arctic Boreal Vulnerability Experiment (ABoVE)

    Science.gov (United States)

    McInerney, M.; Griffith, P. C.; Duffy, D.; Hoy, E.; Schnase, J. L.; Sinno, S.; Thompson, J. H.

    2014-12-01

    The Arctic Boreal Vulnerability Experiment (ABoVE) is designed to improve understanding of the causes and impacts of ecological changes in Arctic/boreal regions, and will integrate field-based studies, modeling, and data from airborne and satellite remote sensing. ABoVE will result in a fuller understanding of ecosystem vulnerability and resilience to environmental change in the Arctic and boreal regions of western North America, and provide scientific information required to develop options for societal responses to the impacts of these changes. The studies sponsored by NASA during ABoVE will be coordinated with research and in-situ monitoring activities being sponsored by a number of national and international partners. The NASA Center for Climate Simulation at the Goddard Space Flight Center has partnered with the NASA Carbon Cycle & Ecosystems Office to create a science cloud designed for this field campaign - the ABoVE Science Cloud (ASC). The ASC combines high performance computing with emerging technologies to create an environment specifically designed for large-scale modeling, analysis of remote sensing data, copious disk storage with integrated data management, and integration of core variables from in-situ networks identified by the ABoVE Science Definition Team. In this talk, we will present the scientific requirements driving the development of the ABoVE Science Cloud, discuss the necessary interfaces, both computational and human, with in-situ monitoring networks, and show examples of how the ASC is being used to meet the needs of the ABoVE campaign.

  7. A hybrid experiment to search for beauty particles

    International Nuclear Information System (INIS)

    Aoki, S.; Chiba, K.; Hoshino, K.; Kaway, T.; Kobayashi, M.; Kodama, K.; Miyanishi, M.; Nakamura, M.; Nakamura, Y.; Niu, K.; Niwa, K.; Ohashi, M.; Sasaki, H.; Tajima, H.; Tomita, Y.; Yamakawa, O.; Yanagisawa, Y.; Baroni, G.; Cecchetti, A.M.; Dell'Uomo, S.; De Vincenzi, M.; Di Liberto, S.; Frenkel, A.; Manfredini, A.; Marini, G.; Martellotti, G.; Mazzoni, M.A.; Meddi, F.; Nigro, A.; Penso, G.; Pistilli, P.; Sciubla, A.; Sgarbi, C.; Barth, M.; Bertrand, D.; Bertrand-Coremans, G.; Roosen, R.; Bartley, J.H.; Davis, D.H.; Duff, B.G.; Esten, M.J.; Heymann, F.F.; Imrie, D.C.; Lush, G.J.; Tovee, D.N.; Breslin, A.C.; Donnelly, W.; Montwill, A.; Coupland, M.; Trent, P.; Hazama, M.; Isokane, Y.; Tsuneoka, Y.; Kazuno, M.; Minakawa, F.; Shibuya, H.; Watanabe, S.

    1989-01-01

    We give here a detailed description of experiment WA75, which was performed at CERN to search for beauty particles. Events containing at least one muon with a high momentum transverse to the beam direction were selected; then the primary interactions and decay vertices, located in stacks of nuclear research emulsions, were examined and analysed. The various parts of the apparatus are described and the off-line analysis and search in emulsion are discussed. An estimate is made of the sensitivity of the experiment to beauty- and charmed-particle production. (orig.)

  8. In-situ tuff water migration/heater experiment: experimental plan

    International Nuclear Information System (INIS)

    Johnstone, J.K.

    1980-08-01

    Tuffs on the Nevada Test Site (NTS) are currently under investigation as a potential isolation medium for heat-producing nuclear wastes. The National Academy of Sciences has concurred in our identification of the potentially large water content (less than or equal to 40 vol %) of tuffs as one of the important issues affecting their suitability for a repository. This Experimental Plan describes an in-situ experiment intended as an initial assessment of water generation/migration in response to a thermal input. The experiment will be conducted in the Grouse Canyon Welded Tuff in Tunnel U12g (G-Tunnel) located in the north-central region of the NTS. While the Grouse Canyon Welded Tuff is not a potential repository medium, it has physical, thermal, and mechanical properties very similar to those tuffs currently under consideration and is accessible at depth (400 m below the surface) in an existing facility. Other goals of the experiment are to support computer-code and instrumentation development, and to measure in-situ thermal properties. The experimental array consists of a central electrical heater, 1.2 m long x 10.2 cm diameter, surrounded by three holes for measuring water-migration behavior, two holes for measuring temperature profiles, one hole for measuring thermally induced stress in the rock, and one hole perpendicular to the heater to measure displacement with a laser. This Experimental Plan describes the experimental objectives, the technical issues, the site, the experimental array, thermal and thermomechanical modeling results, the instrumentation, the data-acquisition system, posttest characterization, and the organizational details

  9. In situ and laboratory migration experiments through boom clay at Mol

    International Nuclear Information System (INIS)

    Preter, P. de; Put, M.; Canniere, P. de

    1993-01-01

    Physico-chemical characterization and migration studies in the Boom clay, envisaged as a potential host sediment for high level waste disposal in Belgium, were started some 15 years ago. A synthesis study of this experimental work has recently been conducted to compile all available data. From a comparison of the available migration data and the data requirements as derived from the performance assessment studies PAGIS (1988) and PACOMA (1991) the new migration programme (1991-1995) was defined. The critical radionuclides, both with relation to dose rates to man and to missing or unreliable migration data, turned out to be 14 C, 99 Tc. 135 Cs and 237 Np. A second group of radionuclides was found to be possibly critical: 79 Se, 93 Zr, 107 Pd, U - , Am - , Cm - , and Pu-isotopes. This report concentrates on the experimental results as obtained from the migration experiments started in the previous migration programme. Some of the reported radionuclides e.g. 90 Sr) have lost their critical character and will not be further studied within the new programme. New experimental data from laboratory tests have become available for Np, Cs, Sr and C (as HC0 3 - ) and the first results on the migration of organic molecules dissolved in the interstitial Boom clay water are reported. The hydraulic parameters (the hydraulic conductivity K and the storage coefficient S o ) were calculated from both laboratory percolation experiments and in situ piezometric measurements. Conclusions concerning Boom clay anisotropy are drawn. Finally, a short description of the ongoing in situ HTO injection experiment is given and the experimental data are analyzed and discussed. 10 refs., 4 figs., 1 tab

  10. Establishing a protocol for immunocytochemical staining and chromogenic in situ hybridization of Giemsa and Diff-Quick prestained cytological smears

    Directory of Open Access Journals (Sweden)

    Elsa Beraki

    2012-01-01

    Full Text Available Background: Protocols for immunocytochemical staining (ICC and in situ hybridization (ISH of air-dried Diff-Quick or May-Grünwald Giemsa (MGG-stained smears have been difficult to establish. An increasing need to be able to use prestained slides for ICC and ISH in specific cases led to this study, aiming at finding a robust protocol for both methods. Materials and Methods: The material consisted of MGG- and Diff-Quick-stained smears. After diagnosis, one to two diagnostic smears were stored in the department. Any additional smear(s containing diagnostic material were used for this study. The majority were fine needle aspirates (FNAC from the breast, comprising materials from fibroadenomas, fibrocystic disease, and carcinomas. A few were metastatic lesions (carcinomas and malignant melanomas. There were 64 prestained smears. Ten smears were Diff-Quick stained, and 54 were MGG stained. The antibodies used for testing ICC were Ki-67, ER, and PgR, CK MNF116 (pancytokeratin and E-cadherin. HER-2 Dual SISH was used to test ISH. Citrate, TRS, and TE buffers at pH6 and pH9 were tested, as well as, different heating times, microwave powers and antibody concentrations. The ICC was done on the Dako Autostainer (Dako®, Glostrup, Denmark, and HER-2 Dual SISH was done on the Ventana XT-machine (Ventana / Roche® , Strasbourg, France. Results: Optimal results were obtained with the TE buffer at pH 9, for both ICC and ISH. Antibody concentrations generally had to be higher than in the immunohistochemistry (IHC. The optimal microwave heat treatment included an initial high power boiling followed by low power boiling. No post fixation was necessary for ICC, whereas, 20 minutes post fixation in formalin (4% was necessary for ISH. Conclusions: Microwave heat treatment, with initial boiling at high power followed by boiling at low power and TE buffer at pH 9 were the key steps in the procedure. Antibody concentrations has to be adapted for each ICC marker. Post

  11. Constitutional von Hippel-Lindau (VHL) gene deletions detected in VHL families by fluorescence in situ hybridization.

    Science.gov (United States)

    Pack, S D; Zbar, B; Pak, E; Ault, D O; Humphrey, J S; Pham, T; Hurley, K; Weil, R J; Park, W S; Kuzmin, I; Stolle, C; Glenn, G; Liotta, L A; Lerman, M I; Klausner, R D; Linehan, W M; Zhuang, Z

    1999-11-01

    von Hippel-Lindau (VHL) disease is an autosomal dominantly inherited cancer syndrome predisposing to a variety of tumor types that include retinal hemangioblastomas, hemangioblastomas of the central nervous system, renal cell carcinomas, pancreatic cysts and tumors, pheochromocytomas, endolymphatic sac tumors, and epididymal cystadenomas [W. M. Linehan et al., J. Am. Med. Assoc., 273: 564-570, 1995; E. A. Maher and W. G. Kaelin, Jr., Medicine (Baltimore), 76: 381-391, 1997; W. M. Linehan and R. D. Klausner, In: B. Vogelstein and K. Kinzler (eds.), The Genetic Basis of Human Cancer, pp. 455-473, McGraw-Hill, 1998]. The VHL gene was localized to chromosome 3p25-26 and cloned [F. Latif et al., Science (Washington DC), 260: 1317-1320, 1993]. Germline mutations in the VHL gene have been detected in the majority of VHL kindreds. The reported frequency of detection of VHL germline mutations has varied from 39 to 80% (J. M. Whaley et al., Am. J. Hum. Genet., 55: 1092-1102, 1994; Clinical Research Group for Japan, Hum. Mol. Genet., 4: 2233-2237, 1995; F. Chen et al., Hum. Mutat., 5: 66-75, 1995; E. R. Maher et al., J. Med. Genet., 33: 328-332, 1996; B. Zbar, Cancer Surv., 25: 219-232, 1995). Recently a quantitative Southern blotting procedure was found to improve this frequency (C. Stolle et al., Hum. Mutat., 12: 417-423, 1998). In the present study, we report the use of fluorescence in situ hybridization (FISH) as a method to detect and characterize VHL germline deletions. We reexamined a group of VHL patients shown previously by single-strand conformation and sequencing analysis not to harbor point mutations in the VHL locus. We found constitutional deletions in 29 of 30 VHL patients in this group using cosmid and P1 probes that cover the VHL locus. We then tested six phenotypically normal offspring from four of these VHL families: two were found to carry the deletion and the other four were deletion-free. In addition, germline mosaicism of the VHL gene was identified in

  12. Evaluation of HER-2/neu status in breast cancer specimens using immunohistochemistry (IHC) & fluorescence in-situ hybridization (FISH) assay.

    Science.gov (United States)

    Goud, Kalal Iravathy; Dayakar, Seetha; Vijayalaxmi, Kolanupaka; Babu, Saidam Jangu; Reddy, P Vijay Anand

    2012-03-01

    Fluorescence in situ hybridization (FISH) is increasingly being recognized as the most accurate and predictive test for HER 2/neu gene amplification and response to therapy in breast cancer. In the present study we investigated HER-2/neu gene amplification by FISH in breast carcinoma tissue specimens and compared the results with that of immunohistochemical (IHC) analysis. A total of 90 breast carcinoma tissue samples were used for immunohistochemical (IHC) and FISH analysis. IHC was performed by using mouse monoclonal antibody to the intracellular domain of HER-2/neu protein. Each slide was scored in a blinded fashion by two pathologists according to the manufacturer's recommended criteria. FISH analysis was performed on paraffin embedded breast tumour tissue sections. The polysomy for centromere 17 (Spec green signal) was read as green signals less than 4 as moderate polysomy, and more than 4 as highly polysomy. Thirty of the 90 patients had negative results by IHC and FISH. Of the 28 patients with the score of 2+ by IHC, 20 were FISH positive for HER-2/neu gene amplification, three were FISH negative and five patients showed equivocal (1.8-2.2) results by FISH. These five cases were retested for IHC and FISH on different paraffin embedded tissue blocks, and all five were found positive for HER-2/neu gene amplification. Twenty five patients with the score of 3+ by IHC were FISH positive for HER-2/neu gene amplification (>2.2). Seven cases with the score of 3+ by IHC were FISH negative for HER-2/neu gene amplification (>2.2), and showed polysomy of chromosome number 17 high polysomy > 4. Our results indicated that HER-2/neu status by FISH should be performed in all cases of breast tumour with a 2+ score by IHC. Cases demonstrating a 3+ score by IHC may be subjected to FISH to rule out polysomy of chromosome 17 which could be falsely interpreted as HER-2/neu overexpression by IHC analysis. There is also a need for establishing a clinically validated cut-off value

  13. A Comprehensive Evaluation Of HER-2/NEU In Human Breast Cancer By Immunohistochemistry And Fluorescence In Situ Hybridization

    International Nuclear Information System (INIS)

    Abdel Fattah, H.I.; Seif, A.A.; El Hadidi, E.S.; Mohamed, A.A.; El Shinawi, M.E.; Shabaan, M.A.M.

    2012-01-01

    Background: Accurate diagnostic assessment of human epidermal growth factor receptor-2 (HER2/neu) is essential and a prerequisite for appropriate application of the humanized anti-HER-2 monoclonal antibody trastuzumab (Herceptin) to the treatment of patients with breast cancer. The food drug administration (FDA) approved immunohistochemistry (IHC) and Fluorescence in situ hybridization (FISH) for HER2/neu evaluation. IHC is the most widely applicable diagnostic modality in studying HER-2 status. FISH is used for HER2/neu assessment in cases with an equivocal IHC status (score 2+). Objectives: The aim of this study was to detect the amplification and/or expression of HER2/neu in breast cancer using FISH and IHC techniques and to evaluate these applied techniques for their potential and clinical application, with special consideration of equivocal cases by IHC (2+). Subjects and Methods: Assessment of HER2/neu gene expression was made by FISH analysis using the HER2/CEP dual color probe (Vysis) in paraffin-embedded tissue sections of 32 breast cancer patients who were grouped into stages 1+, 2+ or 3+ based on IHC (CBII monoclonal antibody), 4 were classified as IHC 0, 4 were classified as IHC 1 +, 22 were classified as the borderline group; IHC 2+, and 2 were classified as IHC 3+. ER, PR, CEA and CA15-3 were performed to all cases. Survival data was obtained from 25 patients only. Results: The cut-off suggested for HER2/neu amplification by FISH ratio was > 1.3. No statistical significance was found between HER2/neu -by either FISH or IHC- and the different prognostic parameters. The overall survival for the studied patients -in average 3 years- was 16/25 (64%). A significant statistical association was revealed between breast cancer patients’ survival outcome and HER2/neu amplification (p < 0.05) using chi square test. Conclusion: All breast cancer patients should be assessed for HER2/neu status. IHC is a well established method for assessing HER2/neu status in

  14. Demonstration of constant upregulation of the telomerase RNA component in human gastric carcinomas using in situ hybridization.

    Science.gov (United States)

    Heine, B; Hummel, M; Demel, G; Stein, H

    1998-06-01

    Upregulation of the ribonucleoprotein telomerase seems to be a prerequisite for immortality, a feature of malignant cells. Using a polymerase chain reaction (PCR)-based assay, it is possible to demonstrate telomerase activity (TA) in specimens of most human malignancies, whereas it is absent from most normal tissues. It remains unclear, however, why between 5 and 50 per cent of various malignant tumour samples give negative results when TA is measured by the telomeric repeat amplification protocol (TRAP). The expectation that reverse transcription (RT)-PCR for detection of the telomerase RNA component (hTR) would be able to complement or to replace the TRAP assay failed, since malignant as well as non-malignant tissue samples gave positive results in most instances. In the present study, in situ hybridization (ISH) was developed to demonstrate the RNA component of human telomerase at the single cell level. With this method, 13 specimens of fresh frozen gastric carcinoma and four of normal, dysplastic, or inflamed gastric mucosa were investigated and the results were compared with those obtained by RT-PCR and the TRAP assay. In addition, ISH was performed on formalin-fixed sections of the same cases. The TRAP assay revealed positive results in 8 out of 13 gastric carcinomas and was negative in all non-malignant tissues. RT-PCR led to amplification of the telomerase RNA component in all specimens tested, irrespective of the presence or absence of malignant cells. By ISH, all gastric carcinomas showed strong telomerase RNA component-specific signals over malignant cells, whereas only a few grains were detectable over some types of normal somatic cells, including activated lymphocytes. In conclusion, high expression of the telomerase RNA component was restricted to the malignant cells of all the gastric carcinomas investigated, as shown by ISH. This indicates that the absence of TA in a proportion of carcinomas is due to methodological problems of the TRAP assay and is

  15. Kinetics of Hydrolysis of Acetic Anhydride by In-Situ FTIR Spectroscopy: An Experiment for the Undergraduate Laboratory

    Science.gov (United States)

    Haji, Shaker; Erkey, Can

    2005-01-01

    A reaction kinetics experiment for the chemical engineering undergraduate laboratory course was developed in which in-situ Fourier Transfer Infrared spectroscopy was used to measure reactant and product concentrations. The kinetics of the hydrolysis of acetic anhydride was determined by experiments carried out in a batch reactor. The results…

  16. The application of fluorescence in situ hybridization (FISH technique for studying the microbial communities in intestinal tissues of white shrimp (Penaeus vannamei

    Directory of Open Access Journals (Sweden)

    Supamattaya, K.

    2005-02-01

    Full Text Available Fluorescence in situ hybridization technique is very useful for the evaluation of microbial communities in various environments. It is possible to apply this technique to study the intestinal microflora in white shrimp (Penaeus vannamei. Different fixatives and storage temperature were tested in this technique. It was found that fixation with 10% buffered formalin for 12 hours and changed to 70% ethanol shown positive results when compared to the fixation with Davidson's fixative or RF fixative. The best signaling was obtainedfrom the samples which were stored in -20ºC. By using the DNA probe targeted to the Eubacteria domain (EUB338 probe, 5′-GCT GCC TCC CGT AGG AGT-3′ labeled with fluorescein as a hybridizing probe, it was found that most intestinal microflora were aggregated with the intestinal contents, or dispersed in the lumen. There was not evidence of the attachment of the microflora with the intestinal epithelium in this study.

  17. Genetic analysis of tumorigenesis: XXXII. Localization of constitutionally amplified KRAS sequences to Chinese hamster chromosomes X and Y by in situ hybridization.

    Science.gov (United States)

    Stenman, G; Anisowicz, A; Sager, R

    1988-11-01

    The KRAS gene is constitutionally amplified in the Chinese hamster. We have mapped the amplified sequences by in situ hybridization to two major sites on the X and Y chromosomes, Xq4 and Yp2. No autosomal site was detected despite a search under relaxed hybridization conditions. KRAS DNA is amplified about 50-fold compared to a human cell line known to have a diploid number of KRAS sequences, whereas mRNA expression is 5- to 10-fold lower than in normal human cells. While mRNA expression levels do not necessarily parallel gene copy number, the low expression level strongly suggests that the amplified sequences are transcriptionally silent. It is suggested that the amplified sequences arose from the original KRAS gene on chromosome 8 and that the KRAS sequences on the Y chromosome arose by X-Y recombination.

  18. Rapid detection of chromosome rearrangement in medical diagnostic X-ray workers by using fluorescence in situ hybridization and study on dose estimation

    International Nuclear Information System (INIS)

    Wang Zhiquan; Sun Yuanming; Li Jin

    1998-01-01

    Objective: Biological doses were estimated for medical diagnostic X-ray workers. Methods: Chromosome rearrangements in X-ray workers were analysed by fluorescence in situ hybridization (FISH) with composite whole chromosome paintings number 4 and number 7. Results: The frequency of translocation in medical diagnostic X-ray workers was much higher than that in control group (P<0.01). The biological doses to individual X-ray workers were calculated by their translocation frequency. The translocation frequencies of both FISH and G-banding were in good agreement. Conclusion: The biological doses to X-ray workers are estimated by FISH first when their dosimetry records are not documented

  19. Detection of Group B Streptococci in Lim Broth by Use of Group B Streptococcus Peptide Nucleic Acid Fluorescent In Situ Hybridization and Selective and Nonselective Agars▿

    Science.gov (United States)

    Montague, Naomi S.; Cleary, Timothy J.; Martinez, Octavio V.; Procop, Gary W.

    2008-01-01

    The sensitivity, specificity, and positive and negative predictive values for the detection of group B streptococci from Lim enrichment broth with sheep blood agar (SBA), with selective Streptococcus agar (SSA), and by a peptide nucleic acid fluorescent in situ hybridization (PNA FISH) assay were as follows: for culture on SBA, 68.4%, 100%, 100%, and 87.9%, respectively; for culture on SSA, 85.5%, 100%, 100%, and 94.1%, respectively; and for the PNA FISH assay, 97.4%, 98.3%, 96.1%, and 98.9%, respectively. PMID:18667597

  20. Detection of group B streptococci in Lim broth by use of group B streptococcus peptide nucleic acid fluorescent in situ hybridization and selective and nonselective agars.

    Science.gov (United States)

    Montague, Naomi S; Cleary, Timothy J; Martinez, Octavio V; Procop, Gary W

    2008-10-01

    The sensitivity, specificity, and positive and negative predictive values for the detection of group B streptococci from Lim enrichment broth with sheep blood agar (SBA), with selective Streptococcus agar (SSA), and by a peptide nucleic acid fluorescent in situ hybridization (PNA FISH) assay were as follows: for culture on SBA, 68.4%, 100%, 100%, and 87.9%, respectively; for culture on SSA, 85.5%, 100%, 100%, and 94.1%, respectively; and for the PNA FISH assay, 97.4%, 98.3%, 96.1%, and 98.9%, respectively.

  1. The Diagnosis of Gastric Mucosa-associated Lymphoid Tissue Lymphoma by Flow Cytometry and Fluorescence in situ Hybridization of Biopsy Specimens.

    Science.gov (United States)

    Matsueda, Katsunori; Omote, Sizuma; Sakata, Masahiro; Fujita, Isao; Horii, Jouichiro; Toyokawa, Tatsuya

    2018-04-15

    Mucosa-associated lymphoid tissue (MALT) lymphoma and reactive inflammatory lymphoid changes are frequently difficult to distinguish based on a routine histological differential diagnosis. We were unable to diagnose gastric MALT lymphoma histologically using specimens obtained by endoscopy, although a flow cytometry (FCM) analysis demonstrated clonality of neoplastic cells by separating cells by CD45 gating. Furthermore, a fluorescence in situ hybridization (FISH) analysis showed trisomy 18. We therefore diagnosed gastric MALT lymphoma with trisomy 18. We recommend that FCM and FISH analyses of biopsy specimens be considered for diagnosing gastric MALT lymphoma if this diagnosis is suspected based on endoscopic findings.

  2. Locked Nucleic Acid-Based In Situ Hybridization Reveals miR-7a as a Hypothalamus-Enriched MicroRNA with a Distinct Expression Pattern

    DEFF Research Database (Denmark)

    Herzer, S; Silahtaroglu, A; Meister, B

    2012-01-01

    , a part of the brain that controls vital bodily functions, we employed locked nucleic acid (LNA) - fluorescent in situ hybridization (FISH). The expression pattern of the mature miRNAs miR-7a, miR-7b, miR-137 and miR-153 in mouse brain tissue sections was investigated. Whereas all studied miRNAs were......R-7a expression was particularly prominent in the subfornical organ, suprachiasmatic, paraventricular, periventricular, supraoptic, dorsomedial and arcuate nuclei. Identical expression patterns for miR-7a was seen in mouse and rat hypothalamus. By combining LNA-FISH with immunohistochemistry...

  3. Frequency of reciprocal translocations and dicentrics induced in human blood lymphocytes by X-irradiation as determined by fluorescence in situ hybridization

    International Nuclear Information System (INIS)

    Nakano, M.; Nakashima, E.; Pawel, D.; Kodama, Y.; Awa, A.

    1993-01-01

    This study was designed to test the scoring efficiency of reciprocal translocations and dicentrics induced by X-irradiation in vitro using the fluorescence in situ hybridization (FISH) technique. An excess was found in the frequencies of reciprocal translocations relative to those of dicentrics by measurement with FISH at the first cell division after irradiation (translocation:dicentric ≅ 60:40). However, when the same metaphases were also evaluated sequentially by a conventional staining method, the ratio of about 50:50 was restored. This was due in part to misclassification of certain dicentrics as reciprocal translocations by the FISH technique. (author)

  4. Fluorescence in situ hybridization (FISH screening for the 22q11.2 deletion in patients with clinical features of velocardiofacial syndrome but without cardiac anomalies

    Directory of Open Access Journals (Sweden)

    Paula Sandrin-Garcia

    2007-01-01

    Full Text Available The velocardiofacial syndrome (VCFS, a condition associated with 22q11.2 deletions, is characterized by a typical facies, palatal anomalies, learning disabilities, behavioral disturbances and cardiac defects. We investigated the frequency of these chromosomal deletions in 16 individuals with VCFS features who presented no cardiac anomalies, one of the main characteristics of VCFS. Fluorescent in situ hybridization (FISH with the N25 (D22S75; 22q11.2 probe revealed deletions in ten individuals (62%. Therefore, even in the absence of cardiac anomalies testing for the 22q11.2 microdeletions in individuals showing other clinical features of this syndrome is recommended.

  5. Responses of nutrients and mobile carbohydrates in Quercus variabilis seedlings to environmental variations using in situ and ex situ experiments.

    Directory of Open Access Journals (Sweden)

    Jing-Pin Lei

    Full Text Available Forest tree species distributed across a wide range of geographical areas are subjected to differential climatic and edaphic conditions and long-term selection, leading to genotypes with morphological and physiological adaptation to the local environment. To test the ability of species to cope with changing environmental conditions, we studied the ecophysiological features of Quercus variabilis using seedlings grown in geographically widely isolated populations (Exp. I, in situ and in a common garden (Exp. II, ex situ using seedlings originating from those populations. We found that Q. variabilis plants grown in different locations along a south-north gradient had different levels of nutrients (N, P, K and carbon-physiological performance (photosynthesis, non-structural carbohydrates, such as soluble sugars and starch, and that these physiological differences were not correlated with local soil properties. These geographic variations of plant physiology disappeared when plants from different locations were grown in the same environment. Our results indicate that the physiological performance of Q. variabilis plants is mainly determined by the climatic variations across latitude rather than by their soils or by genetic differentiation. The adaptive ability of Q. variabilis found in the present study suggests that this species has the potential to cope, at least to some extent, with changing environmental conditions.

  6. The effect of aging of formalin-fixed paraffin-embedded tissues on the in situ hybridization and immunohistochemistry signals in cervical lesions.

    Science.gov (United States)

    Nuovo, Allison J; Garofalo, Michela; Mikhail, Alexandria; Nicol, Alcina F; Vianna-Andrade, Cecilia; Nuovo, Gerard J

    2013-09-01

    Formalin-fixed, paraffin-embedded tissues are widely used in biomedical research but little is known about the effect of the age of the block or unstained slides on the in situ hybridization or immunohistochemistry signal. We compared the in situ-based and immunohistochemistry-based signals for cervical intraepithelial neoplasia samples that ranged from 0 to 15 years of age. There was a progressive and statistically significant decrease in the strength of the p16 signal when comparing tissues prepared from recent unstained slides (0 to 1 y old, mean score of 92%) to those of intermediate age (5 to 7 y old, mean score of 49%) to old unstained slides (cut 13 to 15 y ago, mean score of 10%). Equivalent, progressive, and significant decreases in the intensity of the signals for microRNAs, CD45, and human papillomavirus DNA were seen in tissues stored on slides from 5 to 7 years and 13 to 15 years, respectively. However, the diminution of signal was much less, although still statistically significant, if the sections from the 13- to 15-year-old paraffin blocks were prepared in 2012. The data likely does not represent degradation of the targets as extraction of several microRNA from the old blocks showed no detectable degradation, despite the markedly weakened in situ hybridization signal. It is concluded that in situ-based signal for DNA, microRNAs, and proteins in paraffin-embedded tissues are significantly reduced over time, especially when stored long term on glass slides which, in turn, can lead to a significant underestimation of the amount and presence of the nucleic acid or protein target.

  7. Multi-objective optimization of in-situ bioremediation of groundwater using a hybrid metaheuristic technique based on differential evolution, genetic algorithms and simulated annealing

    Directory of Open Access Journals (Sweden)

    Kumar Deepak

    2015-12-01

    Full Text Available Groundwater contamination due to leakage of gasoline is one of the several causes which affect the groundwater environment by polluting it. In the past few years, In-situ bioremediation has attracted researchers because of its ability to remediate the contaminant at its site with low cost of remediation. This paper proposed the use of a new hybrid algorithm to optimize a multi-objective function which includes the cost of remediation as the first objective and residual contaminant at the end of the remediation period as the second objective. The hybrid algorithm was formed by combining the methods of Differential Evolution, Genetic Algorithms and Simulated Annealing. Support Vector Machines (SVM was used as a virtual simulator for biodegradation of contaminants in the groundwater flow. The results obtained from the hybrid algorithm were compared with Differential Evolution (DE, Non Dominated Sorting Genetic Algorithm (NSGA II and Simulated Annealing (SA. It was found that the proposed hybrid algorithm was capable of providing the best solution. Fuzzy logic was used to find the best compromising solution and finally a pumping rate strategy for groundwater remediation was presented for the best compromising solution. The results show that the cost incurred for the best compromising solution is intermediate between the highest and lowest cost incurred for other non-dominated solutions.

  8. Comparative mapping of DNA probes derived from the V{sub k} immunoglobulin gene regions on human and great ape chromosomes by fluorescence in situ hybridization

    Energy Technology Data Exchange (ETDEWEB)

    Arnold, N.; Wienberg, J.; Ermert, K. [Universitaet Muenchen (Germany)] [and others

    1995-03-01

    Fluorescence in situ hybridization (FISH) of cosmid clones of human V{sub K} gene regions to human and primate chromosomes contributed to the dating of chromosome reorganizations in evolution. A clone from the K locus at 2p11-p12 (cos 106) hybridized to the assumed homologous chromosome bands in the chimpanzees Pan troglodytes (PTR) and P. paniscus (PPA), the Gorilla gorilla (GGO), and the orangutan Pongo Pygmaeus (PPY). Human and both chimpanzees differed from gorilla and orangutan by the mapping of cos 170, a clone derived from chromosome 2cen-q11.2; the transposition of this orphon to the other side of the centromere can, therefore, be dated after the human/chimpanzee and gorilla divergence. Hybridization to homologous bands was also found with a cosmid clone containing a V{sub K}I orphon located on chromosome 1 (cos 115, main signal at 1q31-q32), although the probe is not fully unique. Also, a clone derived from the orphon V{sub K} region on chromosome 22q11 (cos 121) hybridized to the homologous bands in the great apes. This indicates that the orphons on human chromosomes 1 and 22 had been translocated early in primate evolution. 18 refs., 2 figs.

  9. Experience with restoration of ore-bearing aquifers after in situ leach uranium mining

    International Nuclear Information System (INIS)

    Yazikov, V.G.; Zabaznov, V.U.

    2002-01-01

    In many cases the most important environmental issue for in situ leach uranium mining technology is the impact on groundwater. Usually the greatest issue is the chemical condition of the ore bearing aquifer following the completion of leaching. Based on experience gained during post leach monitoring, it has been found that in properly selected sites the impact following leaching is greatly reduced because of the process of self restoration, otherwise known as natural attenuation. This paper provides ground water monitoring data from 1985 to 1997 following completion of leaching at the Irkol uranium deposit, Kazakhstan. It shows the evolution of the pH, and other chemical parameters over this period. The monitoring results demonstrate that at this site the process of natural attenuation appears to have effectively reduced the impact on groundwater at the site, as well as to keep contaminated leaching fluids from moving more than a few hundreds of metres from the wellfield. (author)

  10. Pretest thermal analysis of the Tuff Water Migration/In-Situ Heater Experiment

    International Nuclear Information System (INIS)

    Bulmer, B.M.

    1980-02-01

    This report describes the pretest thermal analysis for the Tuff Water Migration/In-Situ Heater Experiment to be conducted in welded tuff in G-tunnel, Nevada Test Site. The parametric thermal modeling considers variable boiling temperature, tuff thermal conductivity, tuff emissivity, and heater operating power. For nominal tuff properties, some near field boiling is predicted for realistic operating power. However, the extent of boiling will be strongly determined by the ambient (100% water saturated) rock thermal conductivity. In addition, the thermal response of the heater and of the tuff within the dry-out zone (i.e., bounded by boiling isotherm) is dependent on the temperature variation of rock conductivity as well as the extent of induced boiling

  11. Lower hybrid current drive experiments in Tore Supra

    International Nuclear Information System (INIS)

    Moreau, D.

    1991-12-01

    Two large multi junction launchers have allowed to couple up to 6 MW to the plasma. The current drive efficiency was about 2 x 10 19 Am -2 /W with LH power alone at a volume-averaged electron temperature of 1.4 keV. Experiments with combined LHCD and ion cyclotron resonant heating allowed to inject up to 7.5 MW into the plasma. The electron energy content followed fairly well the Rebut-Lallia scaling law. At n e = 1.5 x 10 19 m -3 , sawteeth were suppressed and m = 1 MHD oscillations appeared. The central electron temperature then reached 8 keV for 3.6 MW injected. LH power modulation experiments performed at n e = 4 x 10 19 m -3 showed a delayed central electron heating despite the off-axis creation of suprathermal electrons, thus ruling out the possibility of direct heating through central wave absorption. Successful pellet fueling of a partially LH driven plasma was obtained, in which 28 successive pellets could penetrate up to half radius as in ohmic discharges, with 50 to 80% of the pellet content deposited in the plasma. First attempts to combine LHCD with ergodic divertor discharges showed that, when the plasma edge was subject to a radial magnetic perturbation smaller than the ergodicity threshold, a strong stationary radiation (MARFE) was triggered, locked near the inner wall. The radiated power then amounted to 90% of the total input power with no indication of a radiative collapse

  12. In situ micro-focused X-ray beam characterization with a lensless camera using a hybrid pixel detector

    International Nuclear Information System (INIS)

    Kachatkou, Anton; Marchal, Julien; Silfhout, Roelof van

    2014-01-01

    Position and size measurements of a micro-focused X-ray beam, using an X-ray beam imaging device based on a lensless camera that collects radiation scattered from a thin foil placed in the path of the beam at an oblique angle, are reported. Results of studies on micro-focused X-ray beam diagnostics using an X-ray beam imaging (XBI) instrument based on the idea of recording radiation scattered from a thin foil of a low-Z material with a lensless camera are reported. The XBI instrument captures magnified images of the scattering region within the foil as illuminated by the incident beam. These images contain information about beam size, beam position and beam intensity that is extracted during dedicated signal processing steps. In this work the use of the device with beams for which the beam size is significantly smaller than that of a single detector pixel is explored. The performance of the XBI device equipped with a state-of-the-art hybrid pixel X-ray imaging sensor is analysed. Compared with traditional methods such as slit edge or wire scanners, the XBI micro-focused beam characterization is significantly faster and does not interfere with on-going experiments. The challenges associated with measuring micrometre-sized beams are described and ways of optimizing the resolution of beam position and size measurements of the XBI instrument are discussed

  13. Electromagnetic radiation by parametric decay of upper hybrid waves in ionospheric modification experiments

    International Nuclear Information System (INIS)

    Leyser, T.B.

    1994-01-01

    A nonlinear dispersion relation for the parametric decay of an electrostatic upper hybrid wave into an ordinary mode electromagnetic wave, propagating parallel to the ambient magnetic field, and an electrostatic low frequency wave, being either a lower hybrid wave or a high harmonic ion Bernstein wave, is derived. The coherent and resonant wave interaction is considered to take place in a weakly magnetized and collisionless Vlasov plasma. The instability growth rate is computed for parameter values typical of ionospheric modification experiments, in which a powerful high frequency electromagnetic pump wave is injected into the ionospheric F-region from ground-based transmitters. The electromagnetic radiation which is excited by the decaying upper hybrid wave is found to be consistent with the prominent and commonly observed downshifted maximum (DM) emission in the spectrum of stimulated electromagnetic emission

  14. Pretest parametric calculations for the heated pillar experiment in the WIPP In-Situ Experimental Area

    International Nuclear Information System (INIS)

    Branstetter, L.J.

    1983-03-01

    Results are presented for a pretest parametric study of several configurations and heat loads for the heated pillar experiment (Room H) in the Waste Isolation Pilot Plant (WIPP) In Situ Experimental Area. The purpose of this study is to serve as a basis for selection of a final experiment geometry and heat load. The experiment consists of a pillar of undisturbed rock salt surrounded by an excavated annular room. The pillar surface is covered by a blanket heat source which is externally insulated. A total of five thermal and ten structural calculations are described in a four to five year experimental time frame. Results are presented which include relevant temperature-time histories, deformations, rock salt stress component and effective stress profiles, and maximum stresses in anhydrite layers which are in close proximity to the room. Also included are predicted contours of a conservative post-processed measure of potential salt failure. Observed displacement histories are seen to be highly dependent on pillar and room height, but insensitive to other geometrical variations. The use of a tensile cutoff across slidelines is seen to produce more accurate predictions of anhydrite maximum stress, but to have little effect on rock salt stresses. The potential for salt failure is seen to be small in each case for the time frame of interest, and is only seen at longer times in the center of the room floor

  15. Household preferences of hybrid home heating systems – A choice experiment application

    International Nuclear Information System (INIS)

    Ruokamo, Enni

    2016-01-01

    The residential heating sector presents considerable energy savings potential, as numerous heating solutions for reducing electricity consumption and utilizing renewable energy sources are available in the market. The aim of this paper is to examine determinants of household heating system choices and to use this information for policy planning purposes. This paper investigates residential homeowner attitudes regarding innovative hybrid home heating systems (HHHS) with choice experiment. Heating system scenarios are designed to represent the most relevant primary and supplementary heating alternatives currently available in Finland. The choice sets include six main heating alternatives (district heat, solid wood, wood pellet, electric storage heating, ground heat pump and exhaust air heat pump) that are described by five attributes (supplementary heating systems, investment costs, operating costs, comfort of use and environmental friendliness). The results imply that HHHSs generally appear to be accepted among households; however, several factors affect perceptions of these technologies. The results reveal differing household attitudes toward the main heating alternatives and show that such views are affected by socio-demographic characteristics (age, living environment, education, etc.). The results suggest that households view supplementary heating systems (especially solar-based) favorably. The other attributes studied also play a significant role in decision making. - Highlights: •Study of hybrid heating where supplementary and main heating systems are combined. •Choice experiment is applied to study the determinants of hybrid heating adoption. •Hybrid heating appears to be generally accepted among households. •Households exhibit differing attitudes toward hybrid heating. •Policy makers should not underestimate the potential of hybrid heating.

  16. Overview and First Results of an In-situ Stimulation Experiment in Switzerland

    Science.gov (United States)

    Amann, F.; Gischig, V.; Doetsch, J.; Jalali, M.; Valley, B.; Evans, K. F.; Krietsch, H.; Dutler, N.; Villiger, L.

    2017-12-01

    A decameter-scale in-situ stimulation and circulation (ISC) experiment is currently being conducted at the Grimsel Test Site in Switzerland with the objective of improving our understanding of key seismo-hydro-mechanical coupled processes associated with high pressure fluid injections in a moderately fractured crystalline rock mass. The ISC experiment activities aim to support the development of EGS technology by 1) advancing the understanding of fundamental processes that occur within the rock mass in response to relatively large-volume fluid injections at high pressures, 2) improving the ability to estimate and model induced seismic hazard and risks, 3) assessing the potential of different injection protocols to keep seismic event magnitudes below an acceptable threshold, 4) developing novel monitoring and imaging techniques for pressure, temperature, stress, strain and displacement as well as geophysical methods such as ground penetration radar, passive and active seismic and 5) generating a high-quality benchmark datasets that facilitates the development and validation of numerical modelling tools. The ISC experiment includes six fault slip and five hydraulic fracturing experiments at an intermediate scale (i.e. 20*20*20m) at 480m depth, which allows high resolution monitoring of the evolution of pore pressure in the stimulated fault zone and the surrounding rock matrix, fault dislocations including shear and dilation, and micro-seismicity in an exceptionally well characterized structural setting. In February 2017 we performed the fault-slip experiments on interconnected faults. Subsequently an intense phase of post-stimulation hydraulic characterization was performed. In Mai 2017 we performed hydraulic fracturing tests within test intervals that were free of natural fractures. In this contribution we give an overview and show first results of the above mentioned stimulation tests.

  17. Development and use of fluorescent 16S rRNA-targeted probes for the specific detection of Methylophaga species by in situ hybridization in marine sediments.

    Science.gov (United States)

    Janvier, Monique; Regnault, Béatrice; Grimont, Patrick

    2003-09-01

    Methylotrophic bacteria are widespread in nature. They may play an important role in the cycling of carbon and in the metabolism of dimethylsulfide in a marine environment. Bacteria belonging to the genus Methylophaga are a unique group of aerobic, halophilic, non-methane-utilizing methylotrophs. Two 16S rRNA-targeted oligonucleotide probes were developed for the specific detection of Methylophaga species, marine methylobacteria, by fluorescence in situ hybridization. Probe MPH-730 was highly specific for all members of the genus Methylophaga while probe MPHm-994 targeted exclusively M. marina. The application of these probes were demonstrated by the detection of Methylophaga species in enrichment cultures from various marine sediments. All isolates recovered were visualized by using the genus specific probe MPH-730. The results were confirmed by 16S rDNA sequencing which demonstrated that all selected isolates belong to Methylophaga. Five isolates could be detected by the M. marina-specific probe MPHm-994 and were confirmed by rRNA gene restriction pattern (ribotyping). With the development of these specific probes, fluorescence in situ hybridization shows that the genus Methylophaga is widespread in marine samples.

  18. Comparison of in situ hybridization methods for the assessment of HER-2/neu gene amplification status in breast cancer using a tissue microarray.

    Science.gov (United States)

    Malicka-Durczak, Anna; Korski, Konstanty; Ibbs, Matthew

    2012-01-01

    This project compared HER-2/neu gene status in breast cancers, as demonstrated by FISH (fluorescent in situ hybridization) and CISH (chromogenic in situ hybridization) and using a tissue microarray (TMA). The study also aimed to show whether the TMA technique could be used in clinical diagnostics, rather than remain a scientific tool. A TMA was constructed using 121 breast cancer specimens, 6 cores from each specimen. Demonstration and assessment of HER-2/neu gene status was by FISH (Vysis Path) and CISH (DAKO Duo CISH). The 121 breast cancer specimens were divided into 3 groups by HER-2 status, as determined by immunohistochemistry. In the HER-2 negative group no amplification was observed in 36 out of 40 cases. 3 cases showed amplification by both methods and one by CISH alone. The equivocal HER-2 group showed no amplification in 30 out of 41 cases and amplification in 9 cases. One case was FISH negative CISH positive and one was discarded. In the HER-2 positive group, amplification was confirmed in 37 of the 40 cases by both methods. 3 cases were unsuitable for assessment. This study indicated that CISH is a sensitive alternative to FISH in detecting HER2 gene amplification and may replace FISH in HER2 testing. Good agreement was observed between methods (98.5% - 119 out of 121 cases). Furthermore, as only 4 out of 121 cases were unsuitable for assessment (no signal or missing TMA cores) - it may be feasible to use TMA in diagnostics.

  19. Evaluation of HER2 gene amplification in invasive breast cancer using a dual-color chromogenic in situ hybridization (dual CISH).

    Science.gov (United States)

    Kato, Nobuaki; Itoh, Hitoshi; Serizawa, Akihiko; Hatanaka, Yutaka; Umemura, Shinobu; Osamura, R Yoshiyuki

    2010-07-01

    Fluorescence in situ hybridization (FISH) assay is considered the 'gold standard' for evaluation of HER2/neu (HER2) gene status, however, it is difficult to recognize morphologic features of tumors using fluorescence microscopy. Thus, chromogenic in situ hybridization (CISH) has been proposed as an alternative method to evaluate HER2 gene amplification. Here, we examined the dual color CISH (dual CISH) method which provides information regarding the copy number of the HER2 gene and chromosome 17 centromere from a single slide. We examined 40 cases of invasive ductal carcinomas of the breast that were resected surgically. HER2 gene status was assessed with FISH (Abbott) and dual CISH (Dako). HER2 gene amplification status was classified according to the guidelines of the American Society of Clinical Oncology and College of American Pathologists (ASCO/CAP). Comparison of the cut-off values for HER2/chromosome 17 centromere copy number ratio obtained by dual CISH and FISH showed that there was almost perfect agreement between two methods (Kappa coefficient 0.96). The results of the two commercial products were almost consistent for evaluation of HER2 gene counts on the sections. The current study proved that dual CISH is comparable with FISH for evaluating HER2 gene status.

  20. Chromosomal instability detected by interphase fluorescence in situ hybridization and its relation to p3 alteration in prostate carcinoma in Saudi patients

    International Nuclear Information System (INIS)

    Al-Maghrabi, Jaudah A.

    2005-01-01

    Chromosomal instability (CIN) is a feature of human neoplasm. The p53 mutation has been shown to be associated with CIN in many human dysplastic and neoplastic lesions. The objective of this study was to examine CIN and p53 mutations in prostate carcinoma (Pca) resected from Saudi patients. Testing of p53 alterations using immunohistochemistry was performed on 28 archived prostatic carcinoma specimens containing Pca foci from Saudi patients seen at King Abdul-Aziz University Hospital, Jeddah, Kingdom of Saudi Arabia. Chrosomal instability was evaluated in the same tissues by interphase in situ hybridization (IFISH) using centromere probes for chromosomes 7 and 8. Immunochemistry and IFISH were performed at Princess Margaret Hospital, University Health Network, Toronto, Ontario, Canada in 2001. The p53 immunoreactivity was found in 29% in Pca and 0% in benign epithelium. Interphase in situ hybridization revealed numerical chromosomal alterations in keeping with CIN in 63% of p53 positive and 20% p53 negative Pca. No evidence of CIN was seen in non-neoplastic epithelium. We concluded that CIN as determined by IFISH is present in Pca from Saudi patients similarly to those reported in western countries. The p53 mutation occurs relatively infrequently in Pca and associated with the presence of CIN at least in a subset of Pca. (author)

  1. An Optimized Set of Fluorescence In Situ Hybridization Probes for Detection of Pancreatobiliary Tract Cancer in Cytology Brush Samples.

    Science.gov (United States)

    Barr Fritcher, Emily G; Voss, Jesse S; Brankley, Shannon M; Campion, Michael B; Jenkins, Sarah M; Keeney, Matthew E; Henry, Michael R; Kerr, Sarah M; Chaiteerakij, Roongruedee; Pestova, Ekaterina V; Clayton, Amy C; Zhang, Jun; Roberts, Lewis R; Gores, Gregory J; Halling, Kevin C; Kipp, Benjamin R

    2015-12-01

    Pancreatobiliary cancer is detected by fluorescence in situ hybridization (FISH) of pancreatobiliary brush samples with UroVysion probes, originally designed to detect bladder cancer. We designed a set of new probes to detect pancreatobiliary cancer and compared its performance with that of UroVysion and routine cytology analysis. We tested a set of FISH probes on tumor tissues (cholangiocarcinoma or pancreatic carcinoma) and non-tumor tissues from 29 patients. We identified 4 probes that had high specificity for tumor vs non-tumor tissues; we called this set of probes pancreatobiliary FISH. We performed a retrospective analysis of brush samples from 272 patients who underwent endoscopic retrograde cholangiopancreatography for evaluation of malignancy at the Mayo Clinic; results were available from routine cytology and FISH with UroVysion probes. Archived residual specimens were retrieved and used to evaluate the pancreatobiliary FISH probes. Cutoff values for FISH with the pancreatobiliary probes were determined using 89 samples and validated in the remaining 183 samples. Clinical and pathologic evidence of malignancy in the pancreatobiliary tract within 2 years of brush sample collection was used as the standard; samples from patients without malignancies were used as negative controls. The validation cohort included 85 patients with malignancies (46.4%) and 114 patients with primary sclerosing cholangitis (62.3%). Samples containing cells above the cutoff for polysomy (copy number gain of ≥2 probes) were classified as positive in FISH with the UroVysion and pancreatobiliary probes. Multivariable logistic regression was used to estimate associations between clinical and pathology findings and results from FISH. The combination of FISH probes 1q21, 7p12, 8q24, and 9p21 identified cancer cells with 93% sensitivity and 100% specificity in pancreatobiliary tissue samples and were therefore included in the pancreatobiliary probe set. In the validation cohort of

  2. Biogeochemical processes in a clay formation in situ experiment: Part F - Reactive transport modelling

    Energy Technology Data Exchange (ETDEWEB)

    Tournassat, Christophe, E-mail: c.tournassat@brgm.fr [BRGM, French Geological Survey, Orleans (France); Alt-Epping, Peter [Rock-Water Interaction Group, Institute of Geological Sciences, University of Bern (Switzerland); Gaucher, Eric C. [BRGM, French Geological Survey, Orleans (France); Gimmi, Thomas [Rock-Water Interaction Group, Institute of Geological Sciences, University of Bern (Switzerland)] [Laboratory for Waste Management, Paul Scherrer Institut, Villigen (Switzerland); Leupin, Olivier X. [NAGRA, CH-5430 Wettingen (Switzerland); Wersin, Paul [Gruner Ltd., CH-4020 Basel (Switzerland)

    2011-06-15

    Highlights: > Reactive transport modelling was used to simulate simultaneously solute transport, thermodynamic reactions, ion exchange and biodegradation during an in-situ experiment in a clay-rock formation. > Opalinus clay formation has a high buffering capacity in terms of chemical perturbations caused by bacterial activity. > Buffering capacity is mainly attributed to the carbonate system and to the reactivity of clay surfaces (cation exchange, pH buffering). - Abstract: Reactive transport modelling was used to simulate solute transport, thermodynamic reactions, ion exchange and biodegradation in the Porewater Chemistry (PC) experiment at the Mont Terri Rock Laboratory. Simulations show that the most important chemical processes controlling the fluid composition within the borehole and the surrounding formation during the experiment are ion exchange, biodegradation and dissolution/precipitation reactions involving pyrite and carbonate minerals. In contrast, thermodynamic mineral dissolution/precipitation reactions involving alumo-silicate minerals have little impact on the fluid composition on the time-scale of the experiment. With the accurate description of the initial chemical condition in the formation in combination with kinetic formulations describing the different stages of bacterial activities, it has been possible to reproduce the evolution of important system parameters, such as the pH, redox potential, total organic C, dissolved inorganic C and SO{sub 4} concentration. Leaching of glycerol from the pH-electrode may be the primary source of organic material that initiated bacterial growth, which caused the chemical perturbation in the borehole. Results from these simulations are consistent with data from the over-coring and demonstrate that the Opalinus Clay has a high buffering capacity in terms of chemical perturbations caused by bacterial activity. This buffering capacity can be attributed to the carbonate system as well as to the reactivity of

  3. Development of an in-situ heat transfer experiment (ISHTE) for illite clays of MPG-1

    International Nuclear Information System (INIS)

    Olson, L.O.; Ewart, T.E.

    1981-01-01

    During 1979 the Applied Physics Laboratory of the University of Washington has been working on the design and development of components for an In-Situ Heat Transfer Experiment (ISHTE). The goal of the experiment is to test the validity of using laboratory experiments and analytical studies to predict the thermal response of red clay sediments in the deep ocean. All areas of the conceptual ISHTE developed during 1978 have been investigated this year at some level, with the greatest emphasis on the key components. Because the platform tracking and acoustic communication system is the most crucial component of the experiment, the major effort has been spent on developing this equipment. Hardware and software for the system are nearly operational. Some preliminary testing to evaluate transducers has been performed in Puget Sound and at shallow ocean depths. The transducers purchased initially were not adequate, but units from another company have been purchased and tested with good results. Deep ocean transponder tests will be run in the spring of 1980, and tests of the full tracking system will be conducted in July of 1980 at MPG-1. Several other hardware systems developed this year were an electric heater to simulate the isotope heatsource for system testing, the heater implantment arm, the heater's thermal sensor system, the thermal sensors for the sediment, and a 1/5 scale model of the planned ISHTE platform for visulization of component interaction. The hydrostatic corers are being designed but no construction will be started on prototypes until late 1980. No severe problems have been found thus far with the conceptualized system for the ISHTE platform, and work is progressing on building and testing all of these component systems before the seafloor platform is designed in detail. Present plans are to start designing the platform in FY 81

  4. Assessment of In Situ Time Resolved Shock Experiments at Synchrotron Light Sources*

    Science.gov (United States)

    Belak, J.; Ilavsky, J.; Hessler, J. P.

    2005-07-01

    Prior to fielding in situ time resolved experiments of shock wave loading at the Advanced Photon Source, we have performed feasibility experiments assessing a single photon bunch. Using single and poly-crystal Al, Ti, V and Cu shock to incipient spallation on the gas gun, samples were prepared from slices normal to the spall plane of thickness 100-500 microns. In addition, single crystal Al of thickness 500 microns was shocked to incipient spallation and soft recovered using the LLNL e-gun mini-flyer system. The e-gun mini-flyer impacts the sample target producing a 10's ns flat-top shock transient. Here, we present results for imaging, small-angle scattering (SAS), and diffraction. In particular, there is little SAS away from the spall plane and significant SAS at the spall plane, demonstrating the presence of sub-micron voids. * Use of the Advanced Photon Source was supported by the U. S. Department of Energy, Office of Science, Office of Basic Energy Sciences, under Contract No. W-31-109-Eng-38 and work performed under the auspices of the U.S. Department of Energy by University of California, Lawrence Livermore National Laboratory under Contract W-7405-Eng-48.

  5. Luminescent hybrid lanthanide sulfates and lanthanide sulfonate-carboxylates with 1,10-phenanthroline involving in-situ oxidation of 2-mercaptonbenzoic acid

    International Nuclear Information System (INIS)

    Zhong, Jie-Cen; Wan, Fang; Sun, Yan-Qiong; Chen, Yi-Ping

    2015-01-01

    A series of lanthanide sulfates and lanthanide sulfonate-carboxylates, [Ln 2 (phen) 2 (SO 4 ) 3 (H 2 O) 2 ] n (I:Ln=Nd(1a), Sm(1b), Eu(1c), phen=1,10-phenanthroline) and [Ln(phen)(2-SBA)(BZA)] n (II: Ln=Sm(2a), Eu(2b), Dy(2c), 2-SBA=2-sulfobenzoate, BZA=benzoate) have been hydrothermally synthesized from lanthanide oxide, 2-mercaptonbenzoic acid with phen as auxiliary ligand and characterized by single-crystal X-ray diffraction, elemental analyses, IR spectra, TG analyses and luminescence spectroscopy. Interestingly, SO 4 2− anions in I came from the in situ deep oxidation of thiol groups of 2-mercaptonbenzoic acid while 2-sulfobenzoate and benzoate ligands in II from the middle oxidation and desulfuration reactions of 2-mercaptonbenzoic acid. Compounds I are organic–inorganic hybrid lanthanide sulfates, which have rare one-dimensional column-like structures. Complexes II are binuclear lanthanide sulfonate-carboxylates with 2-sulfobenzoate and benzoate as bridges and 1,10-phenanthroline as terminal. Photoluminescence studies reveal that complexes I and II exhibit strong lanthanide characteristic emission bands in the solid state at room temperature. - Graphical abstract: Lanthanide sulfates and lanthanide sulfonate-carboxylates have been hydrothermally synthesized. Interestingly, sulfate anions, 2-sulfobenzoate and benzoate ligands came from the in situ oxidation and desulfuration reactions of 2-mercaptonbenzoic acid. - Highlights: • In situ oxidation and desulfuration reactions of 2-mercaptonbenzoic acid. • The organic–inorganic hybrid lanthanide sulfates with one-dimensional column-like structure. • The dinuclear lanthanide sulfonate-carboxylates. • The emission spectra exhibit the characteristic transition of 5 D 0 → 7 F J (J=0–4) of the Eu(III)

  6. Detection and quantification of Epstein-Barr virus EBER1 in EBV-infected cells by fluorescent in situ hybridization and flow cytometry

    Science.gov (United States)

    Stowe, R. P.; Cubbage, M. L.; Sams, C. F.; Pierson, D. L.; Barrett, A. D.

    1998-01-01

    A rapid and highly sensitive fluorescent in situ hybridization (FISH) assay was developed to detect Epstein Barr virus (EBV)-infected cells in peripheral blood. Multiple fluorescein-labeled antisense oligonucleotide probes were designed to hybridize to the EBER1 transcript, which is highly expressed in latently infected cells. After a rapid (30 min) hybridization, the cells were analyzed by flow cytometry. EBER1 was detected in several positive control cell lines that have variable numbers of EBV genome copies. No EBER1 was detected in two known EBV-negative cell lines. Northern blot analyses confirmed the presence and quantity of EBER1 transcripts in each cell line. This method was used to quantify the number of EBV-infected cells in peripheral blood from a patient with chronic mononucleosis. These results indicate that EBV-infected cells can be detected at the single cell level, and that this assay can be used to quantify the number of EBV-infected cells in clinical samples.

  7. In-situ preparation of poly(ethylene oxide)/Li3PS4 hybrid polymer electrolyte with good nanofiller distribution for rechargeable solid-state lithium batteries

    Science.gov (United States)

    Chen, Shaojie; Wang, Junye; Zhang, Zhihua; Wu, Linbin; Yao, Lili; Wei, Zhenyao; Deng, Yonghong; Xie, Dongjiu; Yao, Xiayin; Xu, Xiaoxiong

    2018-05-01

    Nano-sized fillers in a polymer matrix with good distribution can play a positive role in improving polymer electrolytes in the aspects of ionic conductivity, mechanical property and electrochemical performance of Li-ion cells. Herein, polyethylene oxide (PEO)/Li3PS4 hybrid polymer electrolyte is prepared via a new in-situ approach. The ionic conductivities of the novel hybrid electrolytes with variable proportions are measured, and the optimal electrolyte of PEO-2%vol Li3PS4 presents a considerable ionic conductivity of 8.01 × 10-4 S cm-1 at 60 °C and an electrochemical window up to 5.1 V. The tests of DSC and EDXS reveal that the Li3PS4 nanoparticles with better distribution, as active fillers scattering in the PEO, exhibit a positive effect on the transference of lithium ion and electrochemical interfacial stabilities. Finally, the assembled solid-state LiFePO4/Li battery presents a decent cycling performance (80.9% retention rate after 325 cycles at 60 °C) and excellent rate capacities with 153, 143, 139 and 127 mAh g-1 at the discharging rate of 0.1 C, 0.2 C, 0.5 C and 1 C at 60 °C. It is fully proved that it is an advanced strategy to preparing the new organic/inorganic hybrid electrolytes for lithium-ion batteries applications.

  8. Considerations in the use of fluorescence in situ hybridization (FISH) and confocal laser scanning microscopy to characterize rumen methanogens and define their spatial distributions.

    Science.gov (United States)

    Valle, Edith R; Henderson, Gemma; Janssen, Peter H; Cox, Faith; Alexander, Trevor W; McAllister, Tim A

    2015-06-01

    In this study, methanogen-specific coenzyme F420 autofluorescence and confocal laser scanning microscopy were used to identify rumen methanogens and define their spatial distribution in free-living, biofilm-, and protozoa-associated microenvironments. Fluorescence in situ hybridization (FISH) with temperature-controlled hybridization was used in an attempt to describe methanogen diversity. A heat pretreatment (65 °C, 1 h) was found to be a noninvasive method to increase probe access to methanogen RNA targets. Despite efforts to optimize FISH, 16S rRNA methanogen-specific probes, including Arch915, bound to some cells that lacked F420, possibly identifying uncharacterized Methanomassiliicoccales or reflecting nonspecific binding to other members of the rumen bacterial community. A probe targeting RNA from the methanogenesis-specific methyl coenzyme M reductase (mcr) gene was shown to detect cultured Methanosarcina cells with signal intensities comparable to those of 16S rRNA probes. However, the probe failed to hybridize with the majority of F420-emitting rumen methanogens, possibly because of differences in cell wall permeability among methanogen species. Methanogens were shown to integrate into microbial biofilms and to exist as ecto- and endosymbionts with rumen protozoa. Characterizing rumen methanogens and defining their spatial distribution may provide insight into mitigation strategies for ruminal methanogenesis.

  9. In situ chemical osmosis experiment in the Boom Clay at the Mol underground research laboratory

    Science.gov (United States)

    Garavito, A. M.; De Cannière, P.; Kooi, H.

    Studies on the compatibility of Boom Clay with large amounts of nitrate- bearing bituminized radioactive waste have recently raised a particular interest for osmosis-induced effects in this reference formation in Belgium. Indeed, water flow and solute transport may be associated with several types of driving forces, or gradients (chemical, electrical, thermal), in addition to the hydraulic forces, resulting in the so-called coupled flows. Fluid flow caused by driving forces different than hydraulic gradients is referred to as osmosis. Chemical osmosis, the water flow induced by a chemical gradient across a semi-permeable membrane, can generate pressure increase. The question thus arises if there is a risk to create high pore pressures that could damage the near-field of medium-level waste (MLW) galleries, if osmotically driven water flows towards the galleries are produced by the release of large amounts of NaNO 3 (750 t) in the formation. To what extent a low-permeability clay formation such as the Boom Clay acts as an osmotic membrane is thus a key issue to assess the relevance of osmosis phenomena for the disposal of medium-level waste. An in situ osmosis experiment has been conducted at the H ADES underground research laboratory to determine the osmotic efficiency of Boom Clay at the field scale. A recently developed chemical osmosis flow continuum model has been used to design the osmosis experiment, and to interpret the water pressure measurements. Experimental data could be reproduced quite accurately by the model, and the inferred parameter values are consistent with independent determinations for Boom Clay. A rapid water pressure increase (but limited to about a 2 m water column) was observed after 12 h in the filter containing the more saline water. Then, the osmotically induced water pressure slowly decays on several months. So, the experimental results obtained in situ confirm the occurrence of non-hydraulic flow phenomena (chemical osmosis) in a low

  10. Pre-WIPP in-situ experiments in salt. Part I. Executive summary. Part II. Program description

    Energy Technology Data Exchange (ETDEWEB)

    Sattler, A.R.; Hunter, T.O.

    1979-08-01

    This document presents plans for in-situ experiments in a specific location in southeastern New Mexico. Schedule and facility design were based on features of a representative local potash mine and on contract negotiations with mine owners. Subsequent WIPP program uncertainties have required a delay in the implementation of the activities discussed here; however, the relative schedule for various activities are appropriate for future planning. The document represents a matrix of in-situ activities to address relevant technical issues prior to the availability of a bedded salt repository.

  11. Pre-WIPP in-situ experiments in salt. Part I. Executive summary. Part II. Program description

    International Nuclear Information System (INIS)

    Sattler, A.R.; Hunter, T.O.

    1979-08-01

    This document presents plans for in-situ experiments in a specific location in southeastern New Mexico. Schedule and facility design were based on features of a representative local potash mine and on contract negotiations with mine owners. Subsequent WIPP program uncertainties have required a delay in the implementation of the activities discussed here; however, the relative schedule for various activities are appropriate for future planning. The document represents a matrix of in-situ activities to address relevant technical issues prior to the availability of a bedded salt repository

  12. Hybrid Reynolds-Averaged/Large-Eddy Simulations of a Coaxial Supersonic Free-Jet Experiment

    Science.gov (United States)

    Baurle, Robert A.; Edwards, Jack R.

    2010-01-01

    Reynolds-averaged and hybrid Reynolds-averaged/large-eddy simulations have been applied to a supersonic coaxial jet flow experiment. The experiment was designed to study compressible mixing flow phenomenon under conditions that are representative of those encountered in scramjet combustors. The experiment utilized either helium or argon as the inner jet nozzle fluid, and the outer jet nozzle fluid consisted of laboratory air. The inner and outer nozzles were designed and operated to produce nearly pressure-matched Mach 1.8 flow conditions at the jet exit. The purpose of the computational effort was to assess the state-of-the-art for each modeling approach, and to use the hybrid Reynolds-averaged/large-eddy simulations to gather insight into the deficiencies of the Reynolds-averaged closure models. The Reynolds-averaged simulations displayed a strong sensitivity to choice of turbulent Schmidt number. The initial value chosen for this parameter resulted in an over-prediction of the mixing layer spreading rate for the helium case, but the opposite trend was observed when argon was used as the injectant. A larger turbulent Schmidt number greatly improved the comparison of the results with measurements for the helium simulations, but variations in the Schmidt number did not improve the argon comparisons. The hybrid Reynolds-averaged/large-eddy simulations also over-predicted the mixing layer spreading rate for the helium case, while under-predicting the rate of mixing when argon was used as the injectant. The primary reason conjectured for the discrepancy between the hybrid simulation results and the measurements centered around issues related to the transition from a Reynolds-averaged state to one with resolved turbulent content. Improvements to the inflow conditions were suggested as a remedy to this dilemma. Second-order turbulence statistics were also compared to their modeled Reynolds-averaged counterparts to evaluate the effectiveness of common turbulence closure

  13. MEDUSA: The ExoMars experiment for in-situ monitoring of dust and water vapour

    Science.gov (United States)

    Colangeli, L.; Lopez-Moreno, J. J.; Nørnberg, P.; Della Corte, V.; Esposito, F.; Mazzotta Epifani, E.; Merrison, J.; Molfese, C.; Palumbo, P.; Rodriguez-Gomez, J. F.; Rotundi, A.; Visconti, G.; Zarnecki, J. C.; The International Medusa Team

    2009-07-01

    Dust and water vapour are fundamental components of the Martian atmosphere. In view of tracing the past environmental conditions on Mars, that possibly favoured the appearing of life forms, it is important to study the present climate and its evolution. Here dust and water vapour have (and have had) strong influence. Of major scientific interest is the quantity and physical, chemical and electrical properties of dust and the abundance of water vapour dispersed in the atmosphere and their exchange with the surface. Moreover, in view of the exploration of the planet with automated systems and in the future by manned missions, it is of primary importance to analyse the hazards linked to these environmental factors. The Martian Environmental Dust Systematic Analyser (MEDUSA) experiment, included in the scientific payload of the ESA ExoMars mission, accommodates a complement of sensors, based on optical detection and cumulative mass deposition, that aims to study dust and water vapour in the lower Martian atmosphere. The goals are to study, for the first time, in-situ and quantitatively, physical properties of the airborne dust, including the cumulative dust mass flux, the dust deposition rate, the physical and electrification properties, the size distribution of sampled particles and the atmospheric water vapour abundance versus time.

  14. In situ gaseous tracer diffusion experiments and predictive modeling at the Greater Confinement Disposal Test

    International Nuclear Information System (INIS)

    Olson, M.C.

    1985-07-01

    The Greater Confinement Disposal Test (GCDT) at the Nevada Test Site is a research project investigating the feasibility of augered shaft disposal of low-level radioactive waste considered unsuitable for shallow land burial. The GCDT contains environmentally mobile and high-specific-activity sources. Research is focused on providing a set of analytically derived hydrogeologic parameters and an empirical database for application in a multiphase, two-dimensional, transient, predictive performance model. Potential contaminant transport processes at the GCDT are identified and their level of significance is detailed. Nonisothermal gaseous diffusion through alluvial sediments is considered the primary waste migration process. Volatile organic tracers are released in the subsurface and their migration is monitored in situ to determine media effective diffusion coefficients, tortuosity, and sorption-corrected porosity terms. The theoretical basis for volatile tracer experiments is presented. Treatment of thermal and liquid flow components is discussed, as is the basis for eliminating several negligible transport processes. Interpretive techniques include correlation, power spectra, and least squares analysis, a graphical analytical solution, and inverse numerical modeling. Model design and application to the GCDT are discussed. GCDT structural, analytical, and computer facilities are detailed. The status of the current research program is reviewed, and temperature and soil moisture profiles are presented along with results of operational tests on the analytical system. 72 refs., 39 figs., 2 tabs

  15. In situ intercalation strategies for device-quality hybrid inorganic-organic self-assembled quantum wells

    Science.gov (United States)

    Pradeesh, K.; Baumberg, J. J.; Prakash, G. Vijaya

    2009-07-01

    Thin films of self-organized quantum wells of inorganic-organic hybrid perovskites of (C6H9C2H4NH3)2PbI4 are formed from a simple intercalation strategy to yield well-ordered uniform films over centimeter-size scales. These films compare favorably with traditional solution-chemistry-synthesized thin films. The hybrid films show strong room-temperature exciton-related absorption and photoluminescence, which shift with fabrication protocol. We demonstrate the potential of this method for electronic and photonic device applications.

  16. NORDA contribution to the in-situ heat transfer experiment (ISHTE): FY84 annual report

    International Nuclear Information System (INIS)

    Valent, P.J.; Bennett, R.H.; Li, H.; Burns, J.T.

    1986-01-01

    The Subseabed Disposal Program (SDP) of the DOE, managed by Sandia National Laboratories, Albuquerque (SNLA), is studying the feasibility of disposing of high-level radioactive wastes by burial in fine-grained deep-sea sediments. The thermo-mechanical response of these sediments to the thermal gradient and temperatures generated by the decaying radionucleides in a buried waste container is being determined by the SDP-supported In Situ Heat Transfer Experiment (ISHTE). The Naval Ocean Research and Development Activity (NORDA) is responsible for the development and fielding of piezometer probes for measuring the pore water pressure gradients induced by the thermal gradient in the sediment. Pore pressure gradients measured in ISHTE will permit validation of theoretical models predicting the rate of radionucleide leakage from a buried waste container to the overlying seawater column. This report of describes the results of a laboratory simulation of ISHTE, conducted at SNLA, in which sediment cracking due to probe insertion was determined to not be a problem to the experiment. Specialized equipment developed for ISHTE, in particular a pressure transducer calibrator for ambient pressures to 69 MPa (10,000 psi), is described. Preliminary results gleaned from excess pore pressure data obtained during the ISHTE component test cruise, Sept 84, confirm that sediment cracking due to probe insertion will be slight and will have an insignificant influence on the measured excess pore pressure dissipation rates. Excess preenerated by insertion of the piezometer probes vary by a factor of two. No significant faults in the NORDA piezometer system were detected either in the laboratory stimulation or in the test in 5800 m water depth north of Hawaii

  17. Identification of parental chromosomes in hybridogenetic water frog Pelophylax esculentus (Rana esculenta) by genomic in situ hybridization (GISH)

    Czech Academy of Sciences Publication Activity Database

    Zalésna, A.; Choleva, Lukáš; Ogielska, M.; Rábová, Marie; Marec, František; Ráb, Petr

    2010-01-01

    Roč. 18, č. 16 (2010), s. 754-755 ISSN 0967-3849. [19th International Colloquium on animal cytogenetics and gene mapping. 06.06.-09.06.2010, Krakow] Institutional research plan: CEZ:AV0Z50450515; CEZ:AV0Z50070508 Keywords : parental chromosomes * Pelophylax esculentus * hybridization Subject RIV: EB - Genetics ; Molecular Biology

  18. Stationary magnetic shear reversal during Lower Hybrid experiments in Tore Supra

    International Nuclear Information System (INIS)

    Litaudon, X.; Arslanbekov, R.; Hoang, G.T.; Joffrin, E.; Kazarian-Vibert, F.; Moreau, D.; Peysson, Y.; Bibet, P.

    1996-01-01

    Stable and stationary states with hollow current density profiles have been achieved with Lower Hybrid Current Drive (LHCD) during Lower Hybrid (LH) wave accessibility experiments. By analysing the bounded propagation domain in phase space which naturally limits the central penetration and absorption of the waves, off-axis LH power deposition has been realized in a reproducible manner. The resulting current density profile modifications have led to a global confinement enhancement attributed to the formation of an internal 'transport barrier' in the central reversed shear region where the electron thermal diffusivity is reduced to its neoclassical collisional level. The multiple-pass LH wave propagation in the weak Landau damping and reversed magnetic shear regime is also investigated in the framework of a statistical theory and the experimental validation of this theory is discussed. (author)

  19. Frequencies of X-ray and fast neutron induced chromosome translocations in human peripheral blood lymphocytes as detected by in situ hybridization using chromosome specific DNA libraries

    International Nuclear Information System (INIS)

    Natarajan, A.T.; Darroudi, F.; Vermeulen, S.; Wiegant, J.

    1992-01-01

    DNA libraries of six human chromosomes were used to detect translocations in human lymphocytes induced by different doses of X-rays and fast neutrons. Results show that with X-rays, one can detect about 1.5 to 2.0 fold more translocations in comparison to dicentrics, whereas following fast neutron irradiation, the difference between these two classes of aberrations are significantly different at high doses. In addition, triple fluorescent in situ hybridization technique was used to study the frequencies of radiation-induced translocations involving a specific chromosome. Chromosome number 1 was found to be involved in translocations more frequently than chromosomes number 2, 3, 4, 8 and X. (author). 10 refs., 1 fig., 2 tabs

  20. In situ ZnO nanowire growth to promote the PVDF piezo phase and the ZnO-PVDF hybrid self-rectified nanogenerator as a touch sensor.

    Science.gov (United States)

    Li, Zetang; Zhang, Xu; Li, Guanghe

    2014-03-28

    A PVDF-ZnO nanowires (NWs) hybrid generator (PZHG) was designed. A simple, cost effective method to produce the PVDF β phase by nano force is introduced. With the ZnO NWs growing, the in situ nano extension force promotes the phase change. A theoretical analysis of the ZnO NWs acting as a self-rectifier of the nano generator is established. The ZnO NWs acted as a self-adjustment diode to control the current output of the PZHG by piezo-electric and semi-conductive effects. Based on the self-controllability of the piezoelectric output, three kinds of finger touching are distinguished by the output performances of the PZHG, which is applicable to an LCD touch pad.

  1. A populational survey of 45S rDNA polymorphism in the Jefferson salamander Ambystoma jeffersonianum revealed by fluorescence in situ hybridization (FISH

    Directory of Open Access Journals (Sweden)

    Jinzhong FU

    2009-04-01

    Full Text Available The chromosomal localization of 45S ribosomal RNA genes in Ambystoma jeffersonianum was determined by fluorescence in situ hybridization with 18S rDNA fragment as a probe (FISH-rDNA. Our results revealed the presence of rDNA polymorphism among A.jeffersonianum populations in terms of number, location and FISH signal intensity on the chromosomes. Nine rDNA cytotypes were found in ten geographically isolated populations and most of them contained derivative rDNA sites. Our preliminary study provides strong indication of karyotypic diversification of A.jeffersonianum that is demonstrated by intraspecific variation of 45S rDNA cytotypes. rDNA cytotype polymorphism has been described in many other caudate amphibians. We predict that habitat isolation, low dispersal ability and decline of effective population size could facilitate the fixation and accumulation of variable rDNA cytotypes during their chromosome evolution.

  2. Interphase fluorescent in situ hybridization deletion analysis of the 9p21 region and prognosis in childhood acute lymphoblastic leukaemia (ALL)

    DEFF Research Database (Denmark)

    Kuchinskaya, Ekaterina; Heyman, Mats; Nordgren, Ann

    2011-01-01

    Interphase fluorescent in situ hybridization (FISH) was applied on diagnostic BM smears from 519 children with acute lymphoblastic leukaemia (ALL) in order to establish the frequency and prognostic importance of 9p21 deletion in children enrolled in the Nordic Society of Paediatric Haematology...... and Oncology (NOPHO) - 2000 treatment protocol. Among the patients, 452 were diagnosed with B-cell precursor (BCP)-ALL and 66 with T-ALL. A higher incidence of 9p21 deletions was found in T-ALL (38%) compared to BCP-ALL (15·7%). Homozygous deletions were found in 19·7% of T-ALL and 4·0% of BCP-ALL; hemizygous...

  3. Application of a Novel and Automated Branched DNA in Situ Hybridization Method for the Rapid and Sensitive Localization of mRNA Molecules in Plant Tissues

    Directory of Open Access Journals (Sweden)

    Andrew J. Bowling

    2014-04-01

    Full Text Available Premise of the study: A novel branched DNA detection technology, RNAscope in situ hybridization (ISH, originally developed for use on human clinical and animal tissues, was adapted for use in plant tissue in an attempt to overcome some of the limitations associated with traditional ISH assays. Methods and Results: Zea mays leaf tissue was formaldehyde fixed and paraffin embedded (FFPE and then probed with the RNAscope ISH assay for two endogenous genes, phosphoenolpyruvate carboxylase (PEPC and phosphoenolpyruvate carboxykinase (PEPCK. Results from both manual and automated methods showed tissue- and cell-specific mRNA localization patterns expected from these well-studied genes. Conclusions: RNAscope ISH is a sensitive method that generates high-quality, easily interpretable results from FFPE plant tissues. Automation of the RNAscope method on the Ventana Discovery Ultra platform allows significant advantages for repeatability, reduction in variability, and flexibility of workflow processes.

  4. Detection of distribution of avian influenza H5N1 virus by immunohistochemistry, chromogenic in situ hybridization and real-time PCR techniques in experimentally infected chickens.

    Science.gov (United States)

    Chamnanpood, Chanpen; Sanguansermsri, Donruedee; Pongcharoen, Sutatip; Sanguansermsri, Phanchana

    2011-03-01

    Ten specific pathogen free (SPF) chickens were inoculated intranasally with avian influenza virus subtype H5N1. Evaluation revealed distribution of the virus in twelve organs: liver, intestine, bursa, lung, trachea, thymus, heart, pancreas, brain, spleen, kidney, and esophagus. Immunohistochemistry (IHC), chromogenic in situ hybridization (CISH), and real-time polymerase chain reaction (PCR) were developed and compared for detection of the virus from the organs. The distribution of avian influenza H5N1 in chickens varied by animal and detecting technique. The heart, kidneys, intestines, lungs, and pancreas were positive with all three techniques, while the others varied by techique. The three techniques can be used to detect avian influenza effectively, but the pros and cons of each technique need to be determined. The decision of which technique to use depends on the objective of the examination, budget, type and quality of samples, laboratory facilities and technician skills.

  5. Conventional and fluorescence in situ hybridization analysis of three-way complex BCR-ABL rearrangement in a chronic myeloid leukemia patient

    Directory of Open Access Journals (Sweden)

    Ganguly Bani

    2007-01-01

    Full Text Available Chromosomal analysis was carried out in bone marrow sample of an 11-year-old girl suspected of myeloproliferative disorder. Conventional G-banding study detected a complex three-way translocation involving 7, 9 and 22, which has resulted in the formation of a variant Philadelphia chromosome causing rearrangement of abl and bcr genes in 87% cells. Fluorescence in situ hybridization (FISH confirmed the fusion of bcr-abl oncogene. Thus the bone marrow karyotype was observed as 46,XX (13% / 46,XX,t(7;9;22(q11;q34;q11 (87%. Hyperdiploidy was present in two cells. In this study, both conventional cytogenetic and FISH diagnosis proved to be significant to identify the variant nature of the Philadelphia chromosome and hyperdiploid condition for introduction of a suitable treatment regimen and estimation of life expectancy of the young girl.

  6. Localization of pig Na[sup +], K[sup +]-ATPase [alpha] and [beta] subunit genes to chromosome 4 by radioactive in situ hybridization

    Energy Technology Data Exchange (ETDEWEB)

    Lahbib-Mansais, Y.; Yerle, M.; Dalens, M.; Chevalet, C.; Gellin, J. (Centre de Recherches de Toulouse (France))

    1993-01-01

    Two genes coding for Na[sup +],K[sup +] -ATPase [alpha] and [beta] subunits are localized on pig chromosome 4, to the q1.6[yields]q2.3 and 1.3[yields]q2.1 regions, respectively, by radioactive in situ hybridization. According to nucleotide and amino acid sequence comparisons with different human isoforms of Na[sup +] ,K[sup +]-ATPase, these pig [alpha] and [beta] ATPase genes show strong homologies with human [alpha]1 and [beta] subunit ATPase genes, respectively. These results are discussed with respect to comparative mapping data of conserved genes in mammalian species. We showed that the pig cDNA probes encoding ATPase [alpha] and, [beta] genes reveal DNA polymorphism in Meishan an Large White pigs. 35 refs., 4 figs., 2 tabs.

  7. Double hypernuclei experiment with hybrid emulsion method at J-PARC

    International Nuclear Information System (INIS)

    Ekawa, Hiroyuki

    2015-01-01

    Double hypernuclei are important probes to study the system with strangeness S = -2. Several emulsion experiments had been performed to search for them. We are planning a new experiment to search for double hypernuclei at the K1.8 beam line in the Hadron Experimental Facility (J-PARC E07 experiment). Ξ"- tracks in the emulsion plates and SSD will be automatically connected by a hybrid method. The estimated Ξ"- stopped statistics is 10 times as high as that of the KEK E373 experiment. Discoveries of 10 new double hypernuclear species are expected, which enable us to discuss binding energy in terms of mass number dependence. On the other hand, we will also observe X rays from Ξ"- atoms with a germanium detector array installed close to the emulsion plates by tagging Ξ"- stopped events. This will be the first measurement to give information on the Ξ"- potential at the nuclear surface region. (author)

  8. Quantitative real-time RT-PCR and chromogenic in situ hybridization: precise methods to detect HER-2 status in breast carcinoma

    Directory of Open Access Journals (Sweden)

    Soares Fernando A

    2009-03-01

    Full Text Available Abstract Background HER-2 gene testing has become an integral part of breast cancer patient diagnosis. The most commonly used assay in the clinical setting for evaluating HER-2 status is immunohistochemistry (IHC and fluorescence in situ hybridization (FISH. These procedures permit correlation between HER-2 expression and morphological features. However, FISH signals are labile and fade over time, making post-revision of the tumor difficult. CISH (chromogenic in situ hybridization is an alternative procedure, with certain advantages, although still limited as a diagnostic tool in breast carcinomas. Methods To elucidate the molecular profile of HER-2 status, mRNA and protein expression in 75 invasive breast carcinomas were analyzed by real time quantitative RT-PCR (qRT-PCR and IHC, respectively. Amplifications were evaluated in 43 of these cases by CISH and in 11 by FISH. Results The concordance rate between IHC and qRT-PCR results was 78.9%, and 94.6% for qRT-PCR and CISH. Intratumoral heterogeneity of HER-2 status was identified in three cases by CISH. The results of the three procedures were compared and showed a concordance rate of 83.8%; higher discordances were observed in 0 or 1+ immunostaining cases, which showed high-level amplification (15.4% and HER-2 transcript overexpression (20%. Moreover, 2+ immunostaining cases presented nonamplified status (50% by CISH and HER-2 downexpression (38.5% by qRT-PCR. In general, concordance occurred between qRT-PCR and CISH results. A high concordance was observed between CISH/qRT-PCR and FISH. Comparisons with clinicopathological data revealed a significant association between HER-2 downexpression and the involvement of less than four lymph nodes (P = 0.0350. Conclusion Based on these findings, qRT-PCR was more precise and reproducible than IHC. Furthermore, CISH was revealed as an alternative and useful procedure for investigating amplifications involving the HER-2 gene.

  9. Quantitative real-time RT-PCR and chromogenic in situ hybridization: precise methods to detect HER-2 status in breast carcinoma

    International Nuclear Information System (INIS)

    Rosa, Fabíola E; Silveira, Sara M; Silveira, Cássia GT; Bérgamo, Nádia A; Neto, Francisco A Moraes; Domingues, Maria AC; Soares, Fernando A; Caldeira, José RF; Rogatto, Silvia R

    2009-01-01

    HER-2 gene testing has become an integral part of breast cancer patient diagnosis. The most commonly used assay in the clinical setting for evaluating HER-2 status is immunohistochemistry (IHC) and fluorescence in situ hybridization (FISH). These procedures permit correlation between HER-2 expression and morphological features. However, FISH signals are labile and fade over time, making post-revision of the tumor difficult. CISH (chromogenic in situ hybridization) is an alternative procedure, with certain advantages, although still limited as a diagnostic tool in breast carcinomas. To elucidate the molecular profile of HER-2 status, mRNA and protein expression in 75 invasive breast carcinomas were analyzed by real time quantitative RT-PCR (qRT-PCR) and IHC, respectively. Amplifications were evaluated in 43 of these cases by CISH and in 11 by FISH. The concordance rate between IHC and qRT-PCR results was 78.9%, and 94.6% for qRT-PCR and CISH. Intratumoral heterogeneity of HER-2 status was identified in three cases by CISH. The results of the three procedures were compared and showed a concordance rate of 83.8%; higher discordances were observed in 0 or 1+ immunostaining cases, which showed high-level amplification (15.4%) and HER-2 transcript overexpression (20%). Moreover, 2+ immunostaining cases presented nonamplified status (50%) by CISH and HER-2 downexpression (38.5%) by qRT-PCR. In general, concordance occurred between qRT-PCR and CISH results. A high concordance was observed between CISH/qRT-PCR and FISH. Comparisons with clinicopathological data revealed a significant association between HER-2 downexpression and the involvement of less than four lymph nodes (P = 0.0350). Based on these findings, qRT-PCR was more precise and reproducible than IHC. Furthermore, CISH was revealed as an alternative and useful procedure for investigating amplifications involving the HER-2 gene

  10. Fluorescent nanodiamond and lanthanide labelled in situ hybridization for the identification of RNA transcripts in fixed and CLARITY-cleared central nervous system tissues (Conference Presentation)

    Science.gov (United States)

    Parker, Lindsay M.; Staikopoulos, Vicky; Cordina, Nicole M.; Sayyadi, Nima; Hutchinson, Mark R.; Packer, Nicolle H.

    2016-03-01

    Despite significant advancement in the methodology used to conjugate, incorporate and visualize fluorescent molecules at the cellular and tissue levels, biomedical imaging predominantly relies on the limitations of established fluorescent molecules such as fluorescein, cyanine and AlexaFluor dyes or genetic incorporation of fluorescent proteins by viral or other means. These fluorescent dyes and conjugates are highly susceptible to photobleaching and compete with cellular autofluorescence, making biomedical imaging unreliable, difficult and time consuming in many cases. In addition, some proteins have low copy numbers and/or poor antibody recognition, further making detection and imaging difficult. We are developing better methods for imaging central nervous system neuroinflammatory markers using targeted mRNA transcripts labelled with fluorescent nanodiamonds or lanthanide chelates. These tags have increased signal and photostability and can also discriminate against tissue/cell autofluorescence. Brains and spinal cords from BALB/c mice with a chronic constriction model of neuropathic pain (neuroinflammation group) or that have undergone sham surgeries (control group) were collected. A subset of brains and spinal cords were perfused and fixed with paraformaldehyde (n=3 sham and n=3 pain groups) prior to sectioning and in situ hybridization using nanodiamond or lanthanide chelate conjugated complementary RNA probes. Another subset of brains and spinal cords from the same cohort of animals were perfused and processed for CLARITY hydrogel based clearing prior to in situ hybridization with the same probes. We will present our findings on the photostability, sensitivity and discrimination from background tissue autofluorescence of our novel RNA probes, compared to traditional fluorophore tags.

  11. Gene protein detection platform--a comparison of a new human epidermal growth factor receptor 2 assay with conventional immunohistochemistry and fluorescence in situ hybridization platforms.

    Science.gov (United States)

    Stålhammar, Gustav; Farrajota, Pedro; Olsson, Ann; Silva, Cristina; Hartman, Johan; Elmberger, Göran

    2015-08-01

    Human epidermal growth factor receptor 2 (HER2) immunohistochemistry (IHC) and fluorescence in situ hybridization (FISH) are widely used semiquantitative assays for selecting breast cancer patients for HER2 antibody therapy. However, both techniques have been shown to have disadvantages. Our aim was to test a recent automated technique of combined IHC and brightfield dual in situ hybridization-gene protein detection platform (GPDP)-in breast cancer HER2 protein, gene, and chromosome 17 centromere status evaluations, comparing the results in accordance to the American Society of Clinical Oncology/College of American Pathologists recommendations for HER2 testing in breast cancer from both 2007 and 2013. The GPDP technique performance was evaluated on 52 consecutive whole slide invasive breast cancer cases with HER2 IHC 2/3+ scoring results. Applying in turns the American Society of Clinical Oncology/College of American Pathologists recommendations for HER2 testing in breast cancer from 2007 and 2013 to both FISH and GPDP DISH assays, the HER2 gene amplification results showed 100% concordance among amplified/nonamplified cases, but there was a shift in 4 cases toward positive from equivocal results and toward equivocal from negative results. This might be related to the emphasis on the average HER2 copy number in the 2013 criteria. HER2 expression by IVD market IHC kit (Pathway®) has a strong correlation with GPDP HER2 protein, including a full concordance for all cases scored as 3+ and a reduction from 2+ to 1+ in 7 cases corresponding to nonamplified cases. Gene protein detection platform HER2 protein "solo" could have spared the need for 7 FISH studies. In addition, the platform offered advantages on interpretation reassurance including selecting areas for counting gene signals paralleled with protein IHC expression, on heterogeneity detection, interpretation time, technical time, and tissue expense. Copyright © 2015 Elsevier Inc. All rights reserved.

  12. Expression and distribution of octopus gonadotropin-releasing hormone in the central nervous system and peripheral organs of the octopus (Octopus vulgaris) by in situ hybridization and immunohistochemistry.

    Science.gov (United States)

    Iwakoshi-Ukena, Eiko; Ukena, Kazuyoshi; Takuwa-Kuroda, Kyoko; Kanda, Atshuhiro; Tsutsui, Kazuyoshi; Minakata, Hiroyuki

    2004-09-20

    We recently purified a peptide with structural features similar to vertebrate gonadotropin-releasing hormone (GnRH) from the brain of Octopus vulgaris, cloned a cDNA encoding the precursor protein, and named it oct-GnRH. In the current study, we investigated the expression and distribution of oct-GnRH throughout the central nervous system (CNS) and peripheral organs of Octopus by in situ hybridization on the basis of the cDNA sequence and by immunohistochemistry using a specific antiserum against oct-GnRH. Oct-GnRH mRNA-expressing cell bodies were located in 10 of 19 lobes in the supraesophageal and subesophageal parts of the CNS. Several oct-GnRH-like immunoreactive fibers were seen in all the neuropils of the CNS lobes. The sites of oct-GnRH mRNA expression and the mature peptide distribution were consistent with each other as judged by in situ hybridization and immunohistochemistry. In addition, many immunoreactive fibers were distributed in peripheral organs such as the heart, the oviduct, and the oviducal gland. Modulatory effects of oct-GnRH on the contractions of the heart and the oviduct were demonstrated. The results suggested that, in the context of reproduction, oct-GnRH is a key peptide in the subpedunculate lobe and/or posterior olfactory lobe-optic gland-gonadal axis, an octopus analogue of the hypothalamo-hypophysial-gonadal axis. It may also act as a modulatory factor in controlling higher brain functions such as feeding, memory, movement, maturation, and autonomic functions

  13. Y-chromosome status identification suggests a recipient origin of posttransplant non-small cell lung carcinomas: chromogenic in situ hybridization analysis.

    Science.gov (United States)

    Chen, Wei; Brodsky, Sergey V; Zhao, Weiqiang; Otterson, Gregory A; Villalona-Calero, Miguel; Satoskar, Anjali A; Hasan, Ayesha; Pelletier, Ronald; Ivanov, Iouri; Ross, Patrick; Nadasdy, Tibor; Shilo, Konstantin

    2014-05-01

    Owing to the need of lifelong immunosuppression, solid-organ transplant recipients are known to have an increased risk of posttransplant malignancies including lung cancer. Posttransplant neoplastic transformation of donor-derived cells giving rise to hematopoietic malignancies, Kaposi sarcoma, and basal cell carcinoma in nongraft tissues has been reported. The goal of this study was to assess the cell origin (donor versus recipient derived) of posttransplant non-small cell lung carcinomas (NSCLCs) in kidney and heart transplant recipients. An institutional database search identified 2557 kidney and heart transplant recipients in 8 consecutive years. Among this cohort, 20 (0.8%) renal and 18 (0.7%) heart transplant recipients developed NSCLC. The study cohort comprised 6 of 38 NSCLCs arising in donor-recipient sex-mismatched transplant patients. The tumor cell origin was evaluated by chromogenic in situ hybridization with Y-chromosome probe on formalin-fixed, paraffin-embedded tissues. Y-chromosome was identified in 97% ± 1% (range from 92% to 99%) of all types of nucleated cells in male control tissues. In all 5 NSCLCs from male recipients of female donor organ, Y-chromosome was identified in 97% ± 2% (range from 92% to 100%) of tumor cells, statistically equivalent to normal control (P recipient of male kidney. These findings suggest a recipient derivation of NSCLC arising in kidney and heart transplant recipients. A combination of histologic evaluation and chromogenic in situ hybridization with Y-chromosome analysis allows reliable determination of tissue origin in sex-mismatched solid-organ transplant recipients and may aid in management of posttransplant malignancy in such cases. Copyright © 2014 Elsevier Inc. All rights reserved.

  14. HM modelling of in-situ gas injection tests in bentonite and argillite: the PGZ experiment

    International Nuclear Information System (INIS)

    Gerard, P.; Charlier, R.; Radu, J.P.; La Vaissiere, R. de; Talandier, J.; Collin, F.

    2010-01-01

    Document available in extended abstract form only. During long-term repository of high and intermediate level nuclear waste in deep argillaceous geological formation, steel containers will corroded and organic material will be irradiated. The two processes lead mostly to hydrogen production. This study deals with the numerical modelling of the gas migration in both the host formation and a bentonite plug, with an emphasis on coupling between the gas transfer and the mechanical strains and stresses. More particularly the study aims to support the design of the PGZ in situ experiment that will be performed by Andra in its underground laboratory at Bure. The objective of the experiment is the analysis of the dynamics of the bentonite plug re-saturation, studying the competition between the liquid water coming from the argillite and a gas injection. The modelled experiment consists of a borehole drilled in rock clay, inside which a plug of MX-80 bentonite is set. The bentonite is naturally re-saturated by water coming from the host formation. At the same time a gas pressure, higher than the initial water pressure in the host rock, is imposed at both ends of the plug. The developed model takes into account the coupling between the mechanical behaviour and the water and gas transfers in undisturbed geo-materials. It manages explicitly liquid and vapour water, gaseous and dissolved hydrogen. Elastoplastic and non-linear elastic model are used to model the behaviour of, respectively, the argillaceous rock and the bentonite. The numerical results show the small desaturation obtained in bentonite and argillite. The influence of the coupling of the mechanic on the water and gas transfers is thus limited (due to the Bishop's effective stress). The swelling of the bentonite plug is not hindered by the gas migration and the confining effect of the engineered barrier is maintained. An analysis is made of the influence of the main transfer rock properties on the gas pressure

  15. Savannah River Site Experiences in In Situ Field Measurements of Radioactive Materials

    International Nuclear Information System (INIS)

    Moore, F.S.

    1999-01-01

    This paper discusses some of the field gamma-ray measurements made at the Savannah River Site, the equipment used for the measurements, and lessons learned during in situ identification and characterization of radioactive materials

  16. Construction of g-C_3N_4/Al_2O_3 hybrids via in-situ acidification and exfoliation with enhanced photocatalytic activity

    International Nuclear Information System (INIS)

    Wang, Xiao-jing; Liu, Chao; Li, Xu-li; Li, Fa-tang; Li, Yu-pei; Zhao, Jun; Liu, Rui-hong

    2017-01-01

    Highlights: • Ultrathin g-C_3N_4/Al_2O_3 hybrids are prepared via in-situ reaction. • The structure modification role of in-situ formed HNO_3 for g-C_3N_4 is found. • The ultrathin g-C_3N_4 nanosheets are formed by the acidified melamine and Al(OH)_3. • In-situ calcination of melamine and Al(OH)_3 benefits the contact of C_3N_4 and Al_2O_3. • The activity of g-C_3N_4/Al_2O_3 is 16.6 times that of pristine g-C_3N_4 in degrading RhB. - Abstract: Homogeneous ultrathin g-C_3N_4 nanosheets/Al_2O_3 heterojunctions are synthesized using melamine and Al(NO_3)_3 via in-situ reaction and the following thermal polymerization approach. The in-situ reaction between melamine and Al(NO_3)_3 results in the existence of HNO_3-acidified melamine and Al(OH)_3 aggregates via the hydrolysis of Al(NO_3)_3. After thermal polymerization, the aggregates are converted to g-C_3N_4/Al_2O_3 composites. The thermal polymerization of acidified melamine and the support effect of aluminum hydroxide for g-C_3N_4 during the calcination process lead to highly dispersed amrophous Al_2O_3 on ultrathin g-C_3N_4 nanosheets, which is beneficial for the separation of photogenerated electron-hole pairs in the heterojunction. The degradation rate for Rhodamine B (RhB) over the most activie sample is 16.6 times than that of pristine g-C_3N_4 under visible light irradiation, which can be attributed to the high specific surface area, highly dispersion of amorphous Al_2O_3 on ultrathin g-C_3N_4 nanosheet, and the effective electrons transfer from g-C_3N_4 to the amorphous Al_2O_3.

  17. Combining in situ transmission electron microscopy irradiation experiments with cluster dynamics modeling to study nanoscale defect agglomeration in structural metals

    International Nuclear Information System (INIS)

    Xu Donghua; Wirth, Brian D.; Li Meimei; Kirk, Marquis A.

    2012-01-01

    We present a combinatorial approach that integrates state-of-the-art transmission electron microscopy (TEM) in situ irradiation experiments and high-performance computing techniques to study irradiation defect dynamics in metals. Here, we have studied the evolution of visible defect clusters in nanometer-thick molybdenum foils under 1 MeV krypton ion irradiation at 80 °C through both cluster dynamics modeling and in situ TEM experiments. The experimental details are reported elsewhere; we focus here on the details of model construction and comparing the model with the experiments. The model incorporates continuous production of point defects and/or small clusters, and the accompanying interactions, which include clustering, recombination and loss to the surfaces that result from the diffusion of the mobile defects. To account for the strong surface effect in thin TEM foils, the model includes one-dimensional spatial dependence along the foil depth, and explicitly treats the surfaces as black sinks. The rich amount of data (cluster number density and size distribution at a variety of foil thickness, irradiation dose and dose rate) offered by the advanced in situ experiments has allowed close comparisons with computer modeling and permitted significant validation and optimization of the model in terms of both physical model construct (damage production mode, identities of mobile defects) and parameterization (diffusivities of mobile defects). The optimized model exhibits good qualitative and quantitative agreement with the in situ TEM experiments. The combinatorial approach is expected to bring a unique opportunity for the study of radiation damage in structural materials.

  18. Synthesis and luminescence properties of hybrid organic-inorganic transparent titania thin film activated by in- situ formed lanthanide complexes

    Science.gov (United States)

    Wang, Yige; Wang, Li; Li, Huanrong; Liu, Peng; Qin, Dashan; Liu, Binyuan; Zhang, Wenjun; Deng, Ruiping; Zhang, Hongjie

    2008-03-01

    Stable transparent titania thin films were fabricated at room temperature by combining thenoyltrifluoroacetone (TTFA)-modified titanium precursors with amphiphilic triblock poly(ethylene oxide)-poly(propylene oxide)-poly(ethylene oxide) (PEO-PPO-PEO, P123) copolymers. The obtained transparent titania thin films were systematically investigated by IR spectroscopy, PL emission and excitation spectroscopy and transmission electron microscopy. IR spectroscopy indicates that TTFA coordinates the titanium center during the process of hydrolysis and condensation. Luminescence spectroscopy confirms the in-situ formation of lanthanide complexes in the transparent titania thin film. TEM image shows that the in-situ formed lanthanide complexes were homogeneously distributed throughout the whole thin film. The quantum yield and the number of water coordinated to lanthanide metal center have been theoretically determined based on the luminescence data.

  19. In situ one-year burial experiments with simulated nuclear waste glasses

    International Nuclear Information System (INIS)

    Hench, L.L.; Spilman, D.; Buonaquisti, T.; Werme, L.

    1985-01-01

    Two simulated nuclear waste glasses were corroded in an in-situ experiment in the Stripa mine up to one year at 90 degree C and ambient temperature. Changes in compositional in-depth profiles were measured using Fourier transform infrared reflection spectroscopy, SIMS and Rutherford back-scattering. For glass/glass interfaces, both glasses showed depletion of Na, Cs and B, but for the more corrosion resistant glass, the lower depletion is ascribed to the formation of a thin (0.2 nm) coherent and dense outer layer enriched in Mg, Ca, Sr, Ba, Zn-Al and Si, which impedes both ion exchange and network attack of the bulk underneath. For the bentonite interfaces, cation exchange of Ca, Mg, Al and Fe from the bentonite for primarily Na and B is found to produce a glass surface that has three silicate-rich layers. The larger concentrations of M/super2+/ and M/super3+/ cation and the high silica content of the reaction layers result in a considerably retarded rate of ion exchange after the formation of these layers during the first three months of burial. The granite interfaces showed the lowest rate of attack. This appears to be due to a large increase of Fe and Al within the glass surfaces exposed to granite. The results obtained using Rutheford back-scattering confirm the results obtained using the other techniques for surface analysis. Analysis of burial samples cast in steel mini-canisters show no significant effects associated with the steel canister-glass interface. (author)

  20. One-step route to a hybrid TiO2/TixW1−xN nanocomposite by in situ selective carbothermal nitridation

    Directory of Open Access Journals (Sweden)

    Zoë Schnepp, Martin J Hollamby, Masahiko Tanaka, Yoshitaka Matsushita, Yoshio Katsuya and Yoshio Sakka

    2012-01-01

    Full Text Available Metal oxide/nitride nanocomposites have many existing and potential applications, e.g. in energy conversion or ammonia synthesis. Here, a hybrid oxide/nitride nanocomposite (anatase/TixW1−xN was synthesized by an ammonia-free sol–gel route. Synchrotron x-ray diffraction, complemented with electron microscopy and thermogravimetric analysis, was used to study the structure, composition and mechanism of formation of the nanocomposite. The nanocomposite contained nanoparticles (<5 nm diameter of two highly intermixed phases. This was found to arise from controlled nucleation and growth of a single oxide intermediate from the gel precursor, followed by phase separation and in situ selective carbothermal nitridation. Depending on the preparation conditions, the composition varied from anatase/TixW1−xN at low W content to an isostructural mixture of Ti-rich and W-rich TixW1−xN at high W content. In situ selective carbothermal nitridation offers a facile route to the synthesis of nitride-oxide nanocomposites. This conceptually new approach is a significant advance from previous methods, which generally require ammonolysis of a pre-synthesized oxide.

  1. Determination of HER-2 status on FNAC material from breast carcinomas using in situ hybridization with dual chromogen visualization with silver enhancement (dual SISH

    Directory of Open Access Journals (Sweden)

    Beraki Elsa

    2010-01-01

    Full Text Available During the last years, HER-2 status kits and protocols for chromogen visualization of hybridization signals have come on the market. The first generation using chromogen visualization used single color probes. The second generation, now emerging on the market, uses dual chromogen visualization. The aim of this study has been to test a new dual color chromogen kit (Ventana INFORM HER2 Dual Colour ISH Roche ® and compare the results with our in-house method(s. The material consisted primarily of cytological material from invasive breast carcinomas in 49 women. Dual SISH was done on all 49 cytological and histological specimens. The histological specimens were treated according to the manufacturer′s recommendations. The procedure was modified in several steps in order to adapt it to the cytological material. Hybridization failed in two cytological specimens. Dual SISH showed concordant results on cytological and histological material as to amplified/not amplified. The included cases had the same HER-2 expression in the invasive and the in situ components on histology. Four IDC showed HER-2 amplification (8.5%. Polysomy was found in two cases. All dual SISH results except for one concurred with the results of the in-house method(s (1/47=2.1%. The dual SISH is suitable for cytological examination of HER-2 status. The protocol must be optimized for cytological material.

  2. A combination of direct viable count and fluorescence in situ hybridization for specific enumeration of viable Lactobacillus delbrueckii subsp.bulgaricus and Streptococcus thermophilus.

    Science.gov (United States)

    García-Hernández, J; Moreno, Y; Amorocho, C M; Hernández, M

    2012-03-01

    We have developed a direct viable count (DVC)-FISH procedure for quickly and easily discriminating between viable and nonviable cells of Lactobacillus delbrueckii subsp. bulgaricus and Streptococcus thermophilus strains, the traditional yogurt bacteria. direct viable count method has been modified and adapted for Lact. delbrueckii subsp. bulgaricus and Strep. thermophilus analysis by testing different times of incubation and concentrations of DNA-gyrase inhibitors. DVC procedure has been combined with fluorescent in situ hybridization (FISH) for the specific detection of viable cells of both bacteria with specific rRNA oligonucleotide probes (DVC-FISH). Of the four antibiotics tested (novobiocin, nalidixic acid, pipemidic acid and ciprofloxacin), novobiocin was the most effective for DVC method and the optimum incubation time was 7 h for both bacteria. The number of viable cells was obtained by the enumeration of specific hybridized cells that were elongated at least twice their original length for Lactobacillus and twice their original size for Streptococcus. This technique was successfully applied to detect viable cells in inoculated faeces. Results showed that this DVC-FISH procedure is a quick and culture-independent useful method to specifically detect viable Lact. delbrueckii subsp. bulgaricus and Strep. thermophilus in different samples, being applied for the first time to lactic acid bacteria. © 2011 The Authors. Letters in Applied Microbiology © 2011 The Society for Applied Microbiology.

  3. Fluctuation localization imaging-based fluorescence in situ hybridization (fliFISH) for accurate detection and counting of RNA copies in single cells

    Energy Technology Data Exchange (ETDEWEB)

    Cui, Yi; Hu, Dehong; Markillie, Lye Meng; Chrisler, William B.; Gaffrey, Matthew J.; Ansong, Charles; Sussel, Lori; Orr, Galya

    2017-10-04

    Quantitative gene expression analysis in intact single cells can be achieved using single molecule- based fluorescence in situ hybridization (smFISH). This approach relies on fluorescence intensity to distinguish between true signals, emitted from an RNA copy hybridized with multiple FISH sub-probes, and background noise. Thus, the precision in smFISH is often compromised by partial or nonspecific binding of sub-probes and tissue autofluorescence, limiting its accuracy. Here we provide an accurate approach for setting quantitative thresholds between true and false signals, which relies on blinking frequencies of photoswitchable dyes. This fluctuation localization imaging-based FISH (fliFISH) uses blinking frequency patterns, emitted from a transcript bound to multiple sub-probes, which are distinct from blinking patterns emitted from partial or nonspecifically bound sub-probes and autofluorescence. Using multicolor fliFISH, we identified radial gene expression patterns in mouse pancreatic islets for insulin, the transcription factor, NKX2-2, and their ratio (Nkx2-2/Ins2). These radial patterns, showing higher values in β cells at the islet core and lower values in peripheral cells, were lost in diabetic mouse islets. In summary, fliFISH provides an accurate, quantitative approach for detecting and counting true RNA copies and rejecting false signals by their distinct blinking frequency patterns, laying the foundation for reliable single-cell transcriptomics.

  4. Facile in situ synthesis of hierarchical porous Ni/Ni(OH)₂ hybrid sponges with excellent electrochemical energy-storage performances for supercapacitors.

    Science.gov (United States)

    Wang, Wanren; Wang, Wenhua; Wang, Mengjiao; Guo, Xiaohui

    2014-09-01

    Herein, we report the in situ growth of single-crystalline Ni(OH)2 nanoflakes on a Ni support by using facile hydrothermal processes. The as-prepared Ni/Ni(OH)2 sponges were well-characterized by using X-ray diffraction (XRD), SEM, TEM, and X-ray photoelectron spectroscopy (XPS) techniques. The results revealed that the nickel-skeleton-supported Ni(OH)2 rope-like aggregates were composed of numerous intercrossed single-crystal Ni(OH)2 flake-like units. The Ni/Ni(OH)2 hybrid sponges served as electrodes and displayed ultrahigh specific capacitance (SC=3247 F g(-1)) and excellent rate-capability performance, likely owing to fast electron and ion transport, sufficient Faradic redox reaction, and robust structural integrity of the Ni/Ni(OH)2 hybrid electrode. These results support the promising application of Ni(OH)2 nanoflakes as advanced pseudocapacitor materials. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  5. 5-bp Classical Satellite DNA Loci from Chromosome-1 Instability in Cervical Neoplasia Detected by DNA Breakage Detection/Fluorescence in Situ Hybridization (DBD-FISH).

    Science.gov (United States)

    Cortés-Gutiérrez, Elva I; Ortíz-Hernández, Brenda L; Dávila-Rodríguez, Martha I; Cerda-Flores, Ricardo M; Fernández, José Luis; López-Fernández, Carmen; Gosálvez, Jaime

    2013-02-19

    We aimed to evaluate the association between the progressive stages of cervical neoplasia and DNA damage in 5-bp classical satellite DNA sequences from chromosome-1 in cervical epithelium and in peripheral blood lymphocytes using DNA breakage detection/fluorescence in situ hybridization (DBD-FISH). A hospital-based unmatched case-control study was conducted in 2011 with a sample of 30 women grouped according to disease stage and selected according to histological diagnosis; 10 with low-grade squamous intraepithelial lesions (LG-SIL), 10 with high-grade SIL (HG-SIL), and 10 with no cervical lesions, from the Unidad Medica de Alta Especialidad of The Mexican Social Security Institute, IMSS, Mexico. Specific chromosome damage levels in 5-bp classical satellite DNA sequences from chromosome-1 were evaluated in cervical epithelium and peripheral blood lymphocytes using the DBD-FISH technique. Whole-genome DNA hybridization was used as a reference for the level of damage. Results of Kruskal-Wallis test showed a significant increase according to neoplastic development in both tissues. The instability of 5-bp classical satellite DNA sequences from chromosome-1 was evidenced using chromosome-orientation FISH. In conclusion, we suggest that the progression to malignant transformation involves an increase in the instability of 5-bp classical satellite DNA sequences from chromosome-1.

  6. 5-bp Classical Satellite DNA Loci from Chromosome-1 Instability in Cervical Neoplasia Detected by DNA Breakage Detection/Fluorescence in Situ Hybridization (DBD-FISH

    Directory of Open Access Journals (Sweden)

    Jaime Gosálvez

    2013-02-01

    Full Text Available We aimed to evaluate the association between the progressive stages of cervical neoplasia and DNA damage in 5-bp classical satellite DNA sequences from chromosome-1 in cervical epithelium and in peripheral blood lymphocytes using DNA breakage detection/fluorescence in situ hybridization (DBD-FISH. A hospital-based unmatched case-control study was conducted in 2011 with a sample of 30 women grouped according to disease stage and selected according to histological diagnosis; 10 with low-grade squamous intraepithelial lesions (LG-SIL, 10 with high-grade SIL (HG-SIL, and 10 with no cervical lesions, from the Unidad Medica de Alta Especialidad of The Mexican Social Security Institute, IMSS, Mexico. Specific chromosome damage levels in 5-bp classical satellite DNA sequences from chromosome-1 were evaluated in cervical epithelium and peripheral blood lymphocytes using the DBD-FISH technique. Whole-genome DNA hybridization was used as a reference for the level of damage. Results of Kruskal-Wallis test showed a significant increase according to neoplastic development in both tissues. The instability of 5-bp classical satellite DNA sequences from chromosome-1 was evidenced using chromosome-orientation FISH. In conclusion, we suggest that the progression to malignant transformation involves an increase in the instability of 5-bp classical satellite DNA sequences from chromosome-1.

  7. Numerical and structural genomic aberrations are reliably detectable in tissue microarrays of formalin-fixed paraffin-embedded tumor samples by fluorescence in-situ hybridization.

    Directory of Open Access Journals (Sweden)

    Heike Horn

    Full Text Available Few data are available regarding the reliability of fluorescence in-situ hybridization (FISH, especially for chromosomal deletions, in high-throughput settings using tissue microarrays (TMAs. We performed a comprehensive FISH study for the detection of chromosomal translocations and deletions in formalin-fixed and paraffin-embedded (FFPE tumor specimens arranged in TMA format. We analyzed 46 B-cell lymphoma (B-NHL specimens with known karyotypes for translocations of IGH-, BCL2-, BCL6- and MYC-genes. Locus-specific DNA probes were used for the detection of deletions in chromosome bands 6q21 and 9p21 in 62 follicular lymphomas (FL and six malignant mesothelioma (MM samples, respectively. To test for aberrant signals generated by truncation of nuclei following sectioning of FFPE tissue samples, cell line dilutions with 9p21-deletions were embedded into paraffin blocks. The overall TMA hybridization efficiency was 94%. FISH results regarding translocations matched karyotyping data in 93%. As for chromosomal deletions, sectioning artefacts occurred in 17% to 25% of cells, suggesting that the proportion of cells showing deletions should exceed 25% to be reliably detectable. In conclusion, FISH represents a robust tool for the detection of structural as well as numerical aberrations in FFPE tissue samples in a TMA-based high-throughput setting, when rigorous cut-off values and appropriate controls are maintained, and, of note, was superior to quantitative PCR approaches.

  8. In situ preparation of Fe3O4 in a carbon hybrid of graphene nanoscrolls and carbon nanotubes as high performance anode material for lithium-ion batteries

    Science.gov (United States)

    Liu, Yuewen; Hassan Siddique, Ahmad; Huang, Heran; Fang, Qile; Deng, Wei; Zhou, Xufeng; Lu, Huanming; Liu, Zhaoping

    2017-11-01

    A new conductive carbon hybrid combining both reduced graphene nanoscrolls and carbon nanotubes (rGNSs-CNTs) is prepared, and used to host Fe3O4 nanoparticles through an in situ synthesis method. As an anode material for LIBs, the obtained Fe3O4@rGNSs-CNTs shows good electrochemical performance. At a current density of 0.1 A g-1, the anode material shows a high reversible capacity of 1232.9 mAh g-1 after 100 cycles. Even at a current density of 1 A g-1, it still achieves a high reversible capacity of 812.3 mAh g-1 after 200 cycles. Comparing with bare Fe3O4 and Fe3O4/rGO composite anode materials without nanoscroll structure, Fe3O4@rGNSs-CNTs shows much better rate capability with a reversible capacity of 605.0 and 500.0 mAh g-1 at 3 and 5 A g-1, respectively. The excellent electrochemical performance of the Fe3O4@rGNSs-CNTs anode material can be ascribed to the hybrid structure of rGNSs-CNTs, and their strong interaction with Fe3O4 nanoparticles, which on one hand provides more pathways for lithium ions and electrons, on the other hand effectively relieves the volume change of Fe3O4 during the charge-discharge process.

  9. Development of species-specific rDNA probes for Giardia by multiple fluorescent in situ hybridization combined with immunocytochemical identification of cyst wall antigens.

    Science.gov (United States)

    Erlandsen, Stanley L; Jarroll, Edward; Wallis, Peter; van Keulen, Harry

    2005-08-01

    In this study, we describe the development of fluorescent oligonucleotide probes to variable regions in the small subunit of 16S rRNA in three distinct Giardia species. Sense and antisense probes (17-22 mer) to variable regions 1, 3, and 8 were labeled with digoxygenin or selected fluorochomes (FluorX, Cy3, or Cy5). Optimal results were obtained with fluorochome-labeled oligonucleotides for detection of rRNA in Giardia cysts. Specificity of fluorescent in situ hybridization (FISH) was shown using RNase digestion and high stringency to diminish the hybridization signal, and oligonucleotide probes for rRNA in Giardia lamblia, Giardia muris, and Giardia ardeae were shown to specifically stain rRNA only within cysts or trophozoites of those species. The fluorescent oligonucleotide specific for rRNA in human isolates of Giardia was positive for ten different strains. A method for simultaneous FISH detection of cysts using fluorescent antibody (genotype marker) and two oligonucleotide probes (species marker) permitted visualization of G. lamblia and G. muris cysts in the same preparation. Testing of an environmental water sample revealed the presence of FISH-positive G. lamblia cysts with a specific rDNA probe for rRNA, while negative cysts were presumed to be of animal or bird origin.

  10. Distribution of herpes simplex virus types 1 and 2 genomes in human spinal ganglia studied by PCR and in situ hybridization.

    Science.gov (United States)

    Obara, Y; Furuta, Y; Takasu, T; Suzuki, S; Suzuki, H; Matsukawa, S; Fujioka, Y; Takahashi, H; Kurata, T; Nagashima, K

    1997-06-01

    Clinical data indicate that the recurring herpes simplex virus (HSV) from oro-labial lesions is HSV subtype 1 and that the virus from genital lesions is HSV-2. This suggests that HSV-1 and HSV-2 reside in latent forms in the trigeminal ganglia and sacral ganglia, respectively. However, the distribution of latent HSV-1 and HSV-2 infections in human spinal ganglia has not been fully examined. This report concerns the application of polymerase chain reaction (PCR) and in situ hybridization (ISH) to such a study. By using PCR and employing the respective primers, HSV-1 and HSV-2 DNAs were detected in 207 of 524 samples from 262 spinal ganglia (from the cervical to the sacral ganglia) examined on both sides. The percentages of HSV-1 and HSV-2 detected in a given set of ganglia were similar, indicating an absence of site preference. By ISH, few but positive hybridization signals were detected evenly in sacral ganglia sections. The data suggest that regional specificity of recurrent HSV infections is not due to regional distribution of latent virus, but that local host factors may be important for recurrences.

  11. High-Temperature Tensile and Tribological Behavior of Hybrid (ZrB2+Al