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Sample records for addition northern blot

  1. Multiplexed miRNA northern blots via hybridization chain reaction.

    PubMed

    Schwarzkopf, Maayan; Pierce, Niles A

    2016-09-06

    Northern blots enable detection of a target RNA of interest in a biological sample using standard benchtop equipment. miRNAs are the most challenging targets as they must be detected with a single short nucleic acid probe. With existing approaches, it is cumbersome to perform multiplexed blots in which several RNAs are detected simultaneously, impeding the study of interacting regulatory elements. Here, we address this shortcoming by demonstrating multiplexed northern blotting based on the mechanism of hybridization chain reaction (HCR). With this approach, nucleic acid probes complementary to RNA targets trigger chain reactions in which fluorophore-labeled DNA hairpins self-assemble into tethered fluorescent amplification polymers. The programmability of HCR allows multiple amplifiers to operate simultaneously and independently within a blot, enabling straightforward multiplexing. We demonstrate simultaneous detection of three endogenous miRNAs in total RNA extracted from 293T and HeLa cells. For a given target, HCR signal scales linearly with target abundance, enabling relative and absolute quantitation. Using non-radioactive HCR, sensitive and selective miRNA detection is achieved using 2'OMe-RNA probes. The HCR northern blot protocol takes ∼1.5 days independent of the number of target RNAs.

  2. Detection of Long Non-coding RNA Expression by Non-radioactive Northern Blots

    PubMed Central

    Hu, Xiaowen; Feng, Yi; Hu, Zhongyi; Zhang, Youyou; Yuan, Chao-Xing; Xu, Xiaowei; Zhang, Lin

    2016-01-01

    With the advances in sequencing technology and transcriptome analysis, it is estimated that up to 75% of the human genome is transcribed into RNAs. This finding prompted intensive investigations on the biological functions of non-coding RNAs and led to very exciting discoveries of microRNAs as important players in disease pathogenesis and therapeutic applications. Research on long non-coding RNAs (lncRNAs) is in its infancy, yet a broad spectrum of biological regulations has been attributed to lncRNAs. As a novel class of RNA transcripts, the expression level and splicing variants of lncRNAs are various. Northern blot analysis can help us learn about the identity, size, and abundance of lncRNAs. Here we describe how to use northern blot to determine lncRNA abundance and identify different splicing variants of a given lncRNA. PMID:26721491

  3. An improved method for Southern DNA and Northern RNA blotting using a Mupid-2 Mini-Gel electrophoresis unit.

    PubMed

    Furuya, Hirokazu; Yamada, Takeshi; Ikezoe, Koji; Ohyagi, Yasumasa; Fukumaki, Yasuyuki; Fujii, Naoki

    2006-08-31

    An improved method for Southern DNA and Northern RNA blotting using the Mupid-2 Mini-Gel System is described. We get sharp and clear bands in Southern and Northern blotting after only 30 min short gel electrophoresis instead of the several hours large gel electrophoresis of conventional methods. The high electrical voltage with a pulse-like current of the Mupid-2 Mini-Gel System also allows reduction of the amount of formaldehyde, a harmful reagent, from the gel running buffer in RNA blotting. This minor modification of DNA and RNA blotting technique enables us to perform the complete experimental procedure more quickly economically in less space, than conventional Southern and Northern blotting, as well as using an extremely small amount of formaldehyde in RNA blotting.

  4. Immuno-Northern Blotting: Detection of RNA Modifications by Using Antibodies against Modified Nucleosides

    PubMed Central

    Akiyama, Yasutoshi; Itoh, Kunihiko; Nankumo, Shinnosuke; Shima, Hisato; Kikuchi, Koichi; Takeuchi, Yoichi; Elkordy, Alaa; Suzuki, Takehiro; Niizuma, Kuniyasu; Ito, Sadayoshi; Tomioka, Yoshihisa; Abe, Takaaki

    2015-01-01

    The biological roles of RNA modifications are still largely not understood. Thus, developing a method for detecting RNA modifications is important for further clarification. We developed a method for detecting RNA modifications called immuno-northern blotting (INB) analysis and herein introduce its various capabilities. This method involves the separation of RNAs using either polyacrylamide or agarose gel electrophoresis, followed by transfer onto a nylon membrane and subsequent immunoblotting using antibodies against modified nucleosides for the detection of specific modifications. We confirmed that INB with the antibodies for 1-methyladenosine (m1A), N6-methyladenosine (m6A), pseudouridine, and 5-methylcytidine (m5C) showed different modifications in a variety of RNAs from various species and organelles. INB with the anti-m5C antibody revealed that the antibody cross-reacted with another modification on DNA, suggesting the application of this method for characterization of the antibody for modified nucleosides. Additionally, using INB with the antibody for m1A, which is a highly specific modification in eukaryotic tRNA, we detected tRNA-derived fragments known as tiRNAs under the cellular stress response, suggesting the application for tracking target RNA containing specific modifications. INB with the anti-m6A antibody confirmed the demethylation of m6A by the specific demethylases fat mass and obesity-associated protein (FTO) and ALKBH5, suggesting its application for quantifying target modifications in separated RNAs. Furthermore, INB demonstrated that the knockdown of FTO and ALKBH5 increased the m6A modification in small RNAs as well as in mRNA. The INB method has high specificity, sensitivity, and quantitative capability, and it can be employed with conventional experimental apparatus. Therefore, this method would be useful for research on RNA modifications and metabolism. PMID:26606401

  5. A sensitive non-radioactive northern blot method to detect small RNAs.

    PubMed

    Kim, Sang Woo; Li, Zhihua; Moore, Patrick S; Monaghan, A Paula; Chang, Yuan; Nichols, Mark; John, Bino

    2010-04-01

    The continuing discoveries of potentially active small RNAs at an unprecedented rate using high-throughput sequencing have raised the need for methods that can reliably detect and quantitate the expression levels of small RNAs. Currently, northern blot is the most widely used method for validating small RNAs that are identified by methods such as high-throughput sequencing. We describe a new northern blot-based protocol (LED) for small RNA (approximately 15-40 bases) detection using digoxigenin (DIG)-labeled oligonucleotide probes containing locked nucleic acids (LNA) and 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide for cross-linking the RNA to the membrane. LED generates clearly visible signals for RNA amounts as low as 0.05 fmol. This method requires as little as a few seconds of membrane exposure to outperform the signal intensity using overnight exposure of isotope-based methods, corresponding to approximately 1000-fold improvement in exposure-time. In contrast to commonly used radioisotope-based methods, which require freshly prepared and hazardous probes, LED probes can be stored for at least 6 months, facilitate faster and more cost-effective experiments, and are more environmentally friendly. A detailed protocol of LED is provided in the Supplementary Data.

  6. The mouse prostaglandin E receptor EP2 subtype: cloning, expression, and northern blot analysis.

    PubMed

    Katsuyama, M; Nishigaki, N; Sugimoto, Y; Morimoto, K; Negishi, M; Narumiya, S; Ichikawa, A

    1995-09-25

    A functional cDNA clone for the mouse prostaglandin (PG) E receptor EP2 subtype was isolated from a mouse cDNA library. The mouse EP2 receptor consists of 362 amino acid residues with seven putative transmembrane domains. [3H]PGE2 bound specifically to the membrane of Chinese hamster ovary cells stably expressing the cloned receptor. This binding was displaced by unlabeled prostanoids in the order of PGE2 = PGE1 > iloprost, a stable PGI2 agonist > PGF2 alpha > PGD2. Binding was also inhibited by butaprost (an EP2 agonist) and to a lesser extent by M&B 28767 (an EP3 agonist), but not by sulprostone (an EP1 and EP3 agonist) or SC-19220 (an EP1 antagonist). PGE2 and butaprost increased the cAMP level in the Chinese hamster ovary cells in a concentration-dependent manner. Northern blot analysis revealed that EP2 mRNA is expressed most abundantly in the uterus, followed by the spleen, lung, thymus, ileum, liver, and stomach.

  7. Quantitative Expression Analysis in Brassica napus by Northern Blot Analysis and Reverse Transcription-Quantitative PCR in a Complex Experimental Setting.

    PubMed

    Rumlow, Annekathrin; Keunen, Els; Klein, Jan; Pallmann, Philip; Riemenschneider, Anja; Cuypers, Ann; Papenbrock, Jutta

    Analysis of gene expression is one of the major ways to better understand plant reactions to changes in environmental conditions. The comparison of many different factors influencing plant growth challenges the gene expression analysis for specific gene-targeted experiments, especially with regard to the choice of suitable reference genes. The aim of this study is to compare expression results obtained by Northern blot, semi-quantitative PCR and RT-qPCR, and to identify a reliable set of reference genes for oilseed rape (Brassica napus L.) suitable for comparing gene expression under complex experimental conditions. We investigated the influence of several factors such as sulfur deficiency, different time points during the day, varying light conditions, and their interaction on gene expression in oilseed rape plants. The expression of selected reference genes was indeed influenced under these conditions in different ways. Therefore, a recently developed algorithm, called GrayNorm, was applied to validate a set of reference genes for normalizing results obtained by Northern blot analysis. After careful comparison of the three methods mentioned above, Northern blot analysis seems to be a reliable and cost-effective alternative for gene expression analysis under a complex growth regime. For using this method in a quantitative way a number of references was validated revealing that for our experiment a set of three references provides an appropriate normalization. Semi-quantitative PCR was prone to many handling errors and difficult to control while RT-qPCR was very sensitive to expression fluctuations of the reference genes.

  8. Quantitative Expression Analysis in Brassica napus by Northern Blot Analysis and Reverse Transcription-Quantitative PCR in a Complex Experimental Setting

    PubMed Central

    Rumlow, Annekathrin; Keunen, Els; Klein, Jan; Pallmann, Philip; Riemenschneider, Anja; Cuypers, Ann

    2016-01-01

    Analysis of gene expression is one of the major ways to better understand plant reactions to changes in environmental conditions. The comparison of many different factors influencing plant growth challenges the gene expression analysis for specific gene-targeted experiments, especially with regard to the choice of suitable reference genes. The aim of this study is to compare expression results obtained by Northern blot, semi-quantitative PCR and RT-qPCR, and to identify a reliable set of reference genes for oilseed rape (Brassica napus L.) suitable for comparing gene expression under complex experimental conditions. We investigated the influence of several factors such as sulfur deficiency, different time points during the day, varying light conditions, and their interaction on gene expression in oilseed rape plants. The expression of selected reference genes was indeed influenced under these conditions in different ways. Therefore, a recently developed algorithm, called GrayNorm, was applied to validate a set of reference genes for normalizing results obtained by Northern blot analysis. After careful comparison of the three methods mentioned above, Northern blot analysis seems to be a reliable and cost-effective alternative for gene expression analysis under a complex growth regime. For using this method in a quantitative way a number of references was validated revealing that for our experiment a set of three references provides an appropriate normalization. Semi-quantitative PCR was prone to many handling errors and difficult to control while RT-qPCR was very sensitive to expression fluctuations of the reference genes. PMID:27685087

  9. Analysis of solvent control and 1-nitrosopyrene-induced chinese hamster ovary cell mutants by southern and northern blots and the polymerase chain reaction

    SciTech Connect

    Newton, R.K.; Mittelstaedt, R.A.; Heflich, R.H. )

    1992-01-01

    1-Nitrosopyrene, a metabolite of the tumorigenic environmental pollutant 1-nitropyrene, is a potent mutagen at the hprt locus in Chinese hamster ovary (CHO) cells. A single DNA adduct, N-(deoxyguanosin-8-yl)-l-aminopyrene, is produced in CHO cells treated with 1-nitrosopyrene, and this adduct is found in rats and mice exposed to 1-nitropyrene. In this study, the structure of the hprt gene and the structure and amount of hprt mRNA were analyzed in 43 CHO cell mutants (16 isolated from solvent control cultures and 27 isolated from 1-nitrosopyrene-treated cultures). PstI- and BamHI-digested DNA from the mutants were subjected to Southern blot analysis using a hamster hprt cDNA probe. None of the 1-nitrosopyrene-induced mutants and only one of the control mutants displayed hybridization patterns that were different from the parent CHO cells. Northern blot analysis revealed that two control mutants had truncated hprt mRNAs, while 56% of the control mutants and 78% of the induced mutants had reduced levels of hprt mRNA. Using polymerase chain reaction amplification of cDNA synthesized from RNA, the hprt protein-coding region could be amplified from 23 of the 1-nitrosopyrene-induced mutants and 11 of the control mutants. The amplification products from 3 of the control mutants and 5 of the induced mutants were smaller than that found with RNA from parental CHO cells. These results indicate that the mutagenic DNA damage produced by 1-nitrosopyrene in CHO cells does not cause major structural alterations in the hprt gene and suggest that 1-nitrosopyrene acts as a point mutagen. A large number of both control and 1-nitrosopyrene-induced mutants exhibited a marked reduction in hprt mRNA concentration or possessed truncated mRNA hprt protein coding sequences. These alterations may contribute to the 6-thioguanine-resistant phenotype.

  10. MicroRNA fate upon targeting with anti-miRNA oligonucleotides as revealed by an improved Northern-blot-based method for miRNA detection

    PubMed Central

    Torres, Adrian G.; Fabani, Martin M.; Vigorito, Elena; Gait, Michael J.

    2011-01-01

    MicroRNAs (miRNAs) are small non-coding RNAs involved in fine-tuning of gene regulation. Antisense oligonucleotides (ONs) are promising tools as anti-miRNA (anti-miR) agents toward therapeutic applications and to uncover miRNA function. Such anti-miR ONs include 2′-O-methyl (OMe), cationic peptide nucleic acids like K-PNA-K3, and locked nucleic acid (LNA)-based anti-miRs such as LNA/DNA or LNA/OMe. Northern blotting is a widely used and robust technique to detect miRNAs. However, miRNA quantification in the presence of anti-miR ONs has proved to be challenging, due to detection artifacts, which has led to poor understanding of miRNA fate upon anti-miR binding. Here we show that anti-miR ON bound to miR-122 can prevent the miRNA from being properly precipitated into the purified RNA fraction using the standard RNA extraction protocol (TRI-Reagent), yielding an RNA extract that does not reflect the real cellular levels of the miRNA. An increase in the numbers of equivalents of isopropanol during the precipitation step leads to full recovery of the targeted miRNA back into the purified RNA extract. Following our improved protocol, we demonstrate by Northern blotting, in conjunction with a PNA decoy strategy and use of high denaturing PAGE, that high-affinity anti-miRs (K-PNA-K3, LNA/DNA, and LNA/OMe) sequester miR-122 without causing miRNA degradation, while miR-122 targeting with a lower-affinity anti-miR (OMe) seems to promote degradation of the miRNA. The technical issues explored in this work will have relevance for other hybridization-based techniques for miRNA quantification in the presence of anti-miR ONs. PMID:21441346

  11. Whole genomic sequencing of RT98 mitochondria derived from Oryza rufipogon and northern blot analysis to uncover a cytoplasmic male sterility-associated gene.

    PubMed

    Igarashi, Keisuke; Kazama, Tomohiko; Motomura, Keiji; Toriyama, Kinya

    2013-02-01

    Cytoplasmic male sterility (CMS) is a maternally inherited trait resulting in the failure to produce functional pollen and is often observed when an alien cytoplasm is transferred into a cultivated species. An RT98A CMS line and an RT98C fertility restorer line were obtained by successive backcrossing between Oryza rufipogon W1109 and Oryza sativa cultivar Taichung 65. To uncover the CMS-associated mitochondrial genes, we determined the complete sequence of the RT98-CMS mitochondrial genome using next-generation pyrosequencing, and searched new open reading frames (orfs) absent in a reported mitochondrial genome of O. sativa Nipponbare. Then, six candidates were selected for the CMS-associated genes based on the criteria in which they were chimeric in structure or encoded a peptide with transmembrane domains. One of the candidates, orf113, showed different transcript sizes between RT98A and RT98C on Northern blot analysis. The orf113 gene was shown to be co-transcribed with atp4 and cox3 encoding ATP synthase F0 subunit 4 and Cyt c oxidase subunit 3, respectively, and their transcripts were distinctly processed in the presence of a fertility restorer gene. Our results indicate that orf113 is a CMS-associated gene of RT98-CMS.

  12. The western blot

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Western blotting is a technique that involves the separation of proteins by gel electrophoresis, their blotting or transfer to a membrane, and selective immunodetection of an immobilized antigen. This is an important and routine method for protein analysis that depends on the specificity of antibod...

  13. Additions to the rust fungi (Pucciniales) from northern Oman

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The first compilation of the rust fungi occurring in the Sultanate of Oman is presented based on historical records and numerous recent collections, primarily from agricultural hosts. The study compiles data for 16 species of Pucciniales in northern Oman, along with voucher and sequence data and pre...

  14. The Dot Blot ELISA.

    ERIC Educational Resources Information Center

    Gerbig, Donald G., Jr.; Fenk, Christopher J.; Goodhart, Amy S.

    2000-01-01

    Uses two laboratory techniques, Enzyme Linked Immunosorbent Assay (ELISA) and Western Blot, to demonstrate antibody-antigen binding concepts. Includes a list of required materials and directions for the procedure, and makes suggestions for classroom applications. (Contains 13 references.) (YDS)

  15. The Western Blot.

    PubMed

    Hnasko, Thomas S; Hnasko, Robert M

    2015-01-01

    Western blotting is a technique that involves the separation of proteins by gel electrophoresis, their blotting or transfer to a membrane, and selective immunodetection of an immobilized antigen. This is an important and routine method for protein analysis that depends on the specificity of antibody-antigen interaction and is useful for the qualitative or semiquantitative identification of specific proteins and their molecular weight from a complex mixture. This chapter will outline the requisite steps including gel electrophoresis of a protein sample, transfer of protein from a gel to a membrane support, and immunodetection of a target antigen.

  16. BLOT Ver. 1.65

    SciTech Connect

    MEYERS, RAY; GLICK, III, JOHN; FORSYTHE, CHRISTI; GILKEY, AMY; SJAARDEMA, GREGORY

    2009-03-24

    BLOT is a graphic program for post-processing finite element analyses output in the EXODUS II database format. It is command driven with free-format input and can drive graphics devices supported by the Sandia Virtual Device Interface. BLOT produces mesh plots of the analysis output variables including deformed mesh plots, line contours, filled (painted) contours, vector plots of two/three variables (velocity vectors), and symbol plots of scalar variables (discrete cracks). Features include pathlines of analysis variables drawn on the mesh, element selection by material, element birth and death, multiple views combining several displays on each plot, symmetry mirroring, and node and element numbering. X-Y plots of the analysis variables include time vs. variable plots or variable vs. variable plots, and distance vs. variable plots at selected time steps where distance is the accumulated distance between pairs of nodes or element centers. BLOT is written in as portable a form as possible. Fortran code is written in ANSI Standard FORTRAN-77. Machine-specific routines are limited in number and are grouped together to minimize the time required to adapt them to a new system. SEACAS codes have been ported to several Unix systems

  17. Additional Arctic observations improve weather and sea-ice forecasts for the Northern Sea Route.

    PubMed

    Inoue, Jun; Yamazaki, Akira; Ono, Jun; Dethloff, Klaus; Maturilli, Marion; Neuber, Roland; Edwards, Patti; Yamaguchi, Hajime

    2015-11-20

    During ice-free periods, the Northern Sea Route (NSR) could be an attractive shipping route. The decline in Arctic sea-ice extent, however, could be associated with an increase in the frequency of the causes of severe weather phenomena, and high wind-driven waves and the advection of sea ice could make ship navigation along the NSR difficult. Accurate forecasts of weather and sea ice are desirable for safe navigation, but large uncertainties exist in current forecasts, partly owing to the sparse observational network over the Arctic Ocean. Here, we show that the incorporation of additional Arctic observations improves the initial analysis and enhances the skill of weather and sea-ice forecasts, the application of which has socioeconomic benefits. Comparison of 63-member ensemble atmospheric forecasts, using different initial data sets, revealed that additional Arctic radiosonde observations were useful for predicting a persistent strong wind event. The sea-ice forecast, initialised by the wind fields that included the effects of the observations, skilfully predicted rapid wind-driven sea-ice advection along the NSR.

  18. Additional Arctic observations improve weather and sea-ice forecasts for the Northern Sea Route

    PubMed Central

    Inoue, Jun; Yamazaki, Akira; Ono, Jun; Dethloff, Klaus; Maturilli, Marion; Neuber, Roland; Edwards, Patti; Yamaguchi, Hajime

    2015-01-01

    During ice-free periods, the Northern Sea Route (NSR) could be an attractive shipping route. The decline in Arctic sea-ice extent, however, could be associated with an increase in the frequency of the causes of severe weather phenomena, and high wind-driven waves and the advection of sea ice could make ship navigation along the NSR difficult. Accurate forecasts of weather and sea ice are desirable for safe navigation, but large uncertainties exist in current forecasts, partly owing to the sparse observational network over the Arctic Ocean. Here, we show that the incorporation of additional Arctic observations improves the initial analysis and enhances the skill of weather and sea-ice forecasts, the application of which has socioeconomic benefits. Comparison of 63-member ensemble atmospheric forecasts, using different initial data sets, revealed that additional Arctic radiosonde observations were useful for predicting a persistent strong wind event. The sea-ice forecast, initialised by the wind fields that included the effects of the observations, skilfully predicted rapid wind-driven sea-ice advection along the NSR. PMID:26585690

  19. Effects of Nitrogen Addition on Nitrogen Resorption in Temperate Shrublands in Northern China.

    PubMed

    Zhang, Jianhua; Li, He; Shen, Haihua; Chen, Yahan; Fang, Jingyun; Tang, Zhiyao

    2015-01-01

    Nutrient resorption from senescing leaves is a key mechanism of nutrient conservation for plants. The nutrient resorption efficiency is highly dependent on leaf nutrient status, species identity and soil nutrient availability. Nitrogen is a limiting nutrient in most ecosystems, it is widely reported that nitrogen resorption efficiency (NRE) was highly dependent on the soil nitrogen availability and vary with N deposition. The effects of nitrogen deposition on NRE and nitrogen concentration in green and senescing leaves have been well established for forests and grasslands; in contrast, little is known on how plants in shrublands respond to nitrogen deposition across the world. In this study, we conducted a two-year nitrogen addition manipulation experiment to explore the responses of nitrogen concentration in green and senescing leaves, and NRE of seven dominant species, namely, Vitex negundo, Wikstroemia chamaedaphne, Carex rigescens and Cleistogenes chinensis from the Vitex negundo community, and Spirea trilobata, Armeniaca sibirica, V. negundo, C. rigescens and Spodiopogon sibiricus from the Spirea trilobata community, to nitrogen deposition in two typical shrub communities of Mt. Dongling in northern China. Results showed that NRE varied remarkably among different life forms, which was lowest in shrubs, highest in grasses, and intermediate in forbs, implying that shrubs may be most capable of obtaining nitrogen from soil, grasses may conserve more nitrogen by absorption from senescing leaves, whereas forbs may adopt both mechanisms to compete for limited nitrogen supply from the habitats. As the N addition rate increases, N concentration in senescing leaves ([N]s) increased consistent from all species from both communities, that in green leaves ([N]g) increased for all species from the Vitex negundo community, while no significant responses were found for all species from the Spirea trilobata community; NRE decreased for all species except A. sibirica from the

  20. Effects of Nitrogen Addition on Nitrogen Resorption in Temperate Shrublands in Northern China

    PubMed Central

    Zhang, Jianhua; Li, He; Shen, Haihua; Chen, Yahan; Fang, Jingyun; Tang, Zhiyao

    2015-01-01

    Nutrient resorption from senescing leaves is a key mechanism of nutrient conservation for plants. The nutrient resorption efficiency is highly dependent on leaf nutrient status, species identity and soil nutrient availability. Nitrogen is a limiting nutrient in most ecosystems, it is widely reported that nitrogen resorption efficiency (NRE) was highly dependent on the soil nitrogen availability and vary with N deposition. The effects of nitrogen deposition on NRE and nitrogen concentration in green and senescing leaves have been well established for forests and grasslands; in contrast, little is known on how plants in shrublands respond to nitrogen deposition across the world. In this study, we conducted a two-year nitrogen addition manipulation experiment to explore the responses of nitrogen concentration in green and senescing leaves, and NRE of seven dominant species, namely, Vitex negundo, Wikstroemia chamaedaphne, Carex rigescens and Cleistogenes chinensis from the Vitex negundo community, and Spirea trilobata, Armeniaca sibirica, V. negundo, C. rigescens and Spodiopogon sibiricus from the Spirea trilobata community, to nitrogen deposition in two typical shrub communities of Mt. Dongling in northern China. Results showed that NRE varied remarkably among different life forms, which was lowest in shrubs, highest in grasses, and intermediate in forbs, implying that shrubs may be most capable of obtaining nitrogen from soil, grasses may conserve more nitrogen by absorption from senescing leaves, whereas forbs may adopt both mechanisms to compete for limited nitrogen supply from the habitats. As the N addition rate increases, N concentration in senescing leaves ([N]s) increased consistent from all species from both communities, that in green leaves ([N]g) increased for all species from the Vitex negundo community, while no significant responses were found for all species from the Spirea trilobata community; NRE decreased for all species except A. sibirica from the

  1. Addition of alfalfa (Medicago sativa L.) to lamb diets enhances production and profits in northern China

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The agro-pastoral transitional zone of northern China is an ecologically fragile area with a short growing season and extensive livestock production, where feed shortage is a key constraint to ruminant production, especially in winter. This study was conducted to evaluate the effects of various rat...

  2. Additives

    NASA Technical Reports Server (NTRS)

    Smalheer, C. V.

    1973-01-01

    The chemistry of lubricant additives is discussed to show what the additives are chemically and what functions they perform in the lubrication of various kinds of equipment. Current theories regarding the mode of action of lubricant additives are presented. The additive groups discussed include the following: (1) detergents and dispersants, (2) corrosion inhibitors, (3) antioxidants, (4) viscosity index improvers, (5) pour point depressants, and (6) antifouling agents.

  3. [Effects of nitrogen and water addition on soil bacterial diversity and community structure in temperate grasslands in northern China].

    PubMed

    Yang, Shan; Li, Xiao-bing; Wang, Ru-zhen; Cai, Jiang-ping; Xu, Zhu-wen; Zhang, Yu-ge; Li, Hui; Jiang, Yong

    2015-03-01

    In this study, we measured the responses of soil bacterial diversity and community structure to nitrogen (N) and water addition in the typical temperate grassland in northern China. Results showed that N addition significantly reduced microbial biomass carbon (MBC) and microbial biomass nitrogen (MBN) under regular precipitation treatment. Similar declined trends of MBC and MBN caused by N addition were also found under increased precipitation condition. Nevertheless, water addition alleviated the inhibition by N addition. N addition exerted no significant effects. on bacterial α-diversity indices, including richness, Shannon diversity and evenness index under regular precipitation condition. Precipitation increment tended to increase bacterial α-diversity, and the diversity indices of each N gradient under regular precipitation were much lower than that of the corresponding N addition rate under increased precipitation. Correlation analysis showed that soil moisture, nitrate (NO3(-)-N) and ammonium (NH4+-N) were significantly negatively correlated with bacterial evenness index, and MBC and MBN had a significant positive correlation with bacterial richness and evenness. Non-metric multidimensional scaling (NMDS) ordination illustrated that the bacterial communities were significantly separated by N addition rates, under both water ambient and water addition treatments. Redundancy analysis (RDA) revealed that soil MBC, MBN, pH and NH4+-N were the key environmental factors for shaping bacterial communities.

  4. Problem-Solving Test: Southwestern Blotting

    ERIC Educational Resources Information Center

    Szeberényi, József

    2014-01-01

    Terms to be familiar with before you start to solve the test: Southern blotting, Western blotting, restriction endonucleases, agarose gel electrophoresis, nitrocellulose filter, molecular hybridization, polyacrylamide gel electrophoresis, proto-oncogene, c-abl, Src-homology domains, tyrosine protein kinase, nuclear localization signal, cDNA,…

  5. Detection of Proteins on Blot Membranes.

    PubMed

    Goldman, Aaron; Harper, Sandra; Speicher, David W

    2016-11-01

    Staining of blot membranes enables the visualization of bound proteins. Proteins are usually transferred to blot membranes by electroblotting, by direct spotting of protein solutions, or by contact blots. Staining allows the efficiency of transfer to the membrane to be monitored. This unit describes protocols for staining proteins after electroblotting from polyacrylamide gels to blot membranes such as polyvinylidene difluoride (PVDF), nitrocellulose, or nylon membranes. The same methods can be used if proteins are directly spotted, either manually or using robotics. Protocols are included for seven general protein stains (amido black, Coomassie blue, Ponceau S, colloidal gold, colloidal silver, India ink, and MemCode) and three fluorescent protein stains (fluorescamine, IAEDANS, and SYPRO Ruby). Also included is an in-depth discussion of the different blot membrane types and the compatibility of different protein stains with downstream applications, such as immunoblotting or N-terminal Edman sequencing. © 2016 by John Wiley & Sons, Inc.

  6. Effects of water and nitrogen addition on species turnover in temperate grasslands in northern China.

    PubMed

    Xu, Zhuwen; Wan, Shiqiang; Ren, Haiyan; Han, Xingguo; Li, Mai-He; Cheng, Weixin; Jiang, Yong

    2012-01-01

    Global nitrogen (N) deposition and climate change have been identified as two of the most important causes of current plant diversity loss. However, temporal patterns of species turnover underlying diversity changes in response to changing precipitation regimes and atmospheric N deposition have received inadequate attention. We carried out a manipulation experiment in a steppe and an old-field in North China from 2005 to 2009, to test the hypothesis that water addition enhances plant species richness through increase in the rate of species gain and decrease in the rate of species loss, while N addition has opposite effects on species changes. Our results showed that water addition increased the rate of species gain in both the steppe and the old field but decreased the rates of species loss and turnover in the old field. In contrast, N addition increased the rates of species loss and turnover in the steppe but decreased the rate of species gain in the old field. The rate of species change was greater in the old field than in the steppe. Water interacted with N to affect species richness and species turnover, indicating that the impacts of N on semi-arid grasslands were largely mediated by water availability. The temporal stability of communities was negatively correlated with rates of species loss and turnover, suggesting that water addition might enhance, but N addition would reduce the compositional stability of grasslands. Experimental results support our initial hypothesis and demonstrate that water and N availabilities differed in the effects on rate of species change in the temperate grasslands, and these effects also depend on grassland types and/or land-use history. Species gain and loss together contribute to the dynamic change of species richness in semi-arid grasslands under future climate change.

  7. Mixed lubrication after rewetting of blotted pleural mesothelium.

    PubMed

    Bodega, Francesca; Sironi, Chiara; Porta, Cristina; Pecchiari, Matteo; Zocchi, Luciano; Agostoni, Emilio

    2013-01-15

    Coefficient of kinetic friction (μ) of pleural mesothelium blotted with filter paper, and rewetted with Ringer solution markedly increases; this increase is removed if a sufficient amount of sialomucin or hyaluronan is added to Ringer (Bodega et al., 2012. Respiratory Physiology and Neurobiology 180, 34-39). In this research we found that μ of pleural mesothelium blotted, rewetted, and sliding at physiological velocities and loads, decreased with increase of velocity, mainly at low velocities. Despite this decrease, μ at highest velocity was still double that before blotting. With small concentration of sialomucin or hyaluronan μ was markedly smaller at each velocity, decreased less with increase of velocity, and at highest velocity approached preblotting value. These findings indicate a regime of mixed lubrication in post-blotting Ringer, at variance with boundary lubrication occurring before blotting or postblotting with sufficient macromolecule addition. Greater roughness of mesothelial surface, caused by blotting, likely induces zones of elastohydrodynamic lubrication, which increase with velocity, while contact area decreases.

  8. Non-additive effects of litter diversity on greenhouse gas emissions from alpine steppe soil in Northern Tibet.

    PubMed

    Chen, Youchao; Sun, Jian; Xie, Fangting; Yan, Yan; Wang, Xiaodan; Cheng, Genwei; Lu, Xuyang

    2015-12-04

    While litter decomposition is a fundamental ecological process, previous studies have mainly focused on the decay of single species. In this study, we conducted a litter-mixing experiment to investigate litter diversity effects on greenhouse gas (GHG) emissions from an alpine steppe soil in Northern Tibet. Significant non-additive effects of litter diversity on GHG dynamics can be detected; these non-additive effects were the result of species composition rather than species richness. Synergistic effects were frequent for CO2 and N2O emissions, as they were found to occur in 70.5% and 47.1% of total cases, respectively; antagonistic effects on CH4 uptake predominated in 60.3% of the cases examined. The degree of synergism and antagonism may be significantly impacted by litter chemical traits, such as lignin and N, lignin:N ratio, and total phenols during decomposition (P < 0.05). In addition, the relationship between chemical traits and litter-mixing effects changed over incubation time. Our study provides an opportunity to gain insight into the relationship between litter diversity and soil ecological processes. The results indicate that higher plant diversity may generally enhance CO2 and N2O emissions while inhibiting CH4 uptake; meanwhile, the direction and strength of non-additive effects appear to be related to litter chemical traits.

  9. Streamlined Strategies to Better Visualize Southern Blotting

    ERIC Educational Resources Information Center

    Dean, Derek M.

    2012-01-01

    In this article, I describe an animated slideshow of Southern blotting that I have made freely available to other instructors. My hope is to provide a clear visualization of the logistics behind the technique so that instructors have a solid basis--as well as time freed up--to discuss its applications with students.

  10. Blot hybridisation analysis of genomic DNA.

    PubMed Central

    Vandenplas, S; Wiid, I; Grobler-Rabie, A; Brebner, K; Ricketts, M; Wållis, G; Bester, A; Boyd, C; Måthew, C

    1984-01-01

    Restriction endonuclease analysis of specific gene sequences is proving to be a valuable technique for characterisation and diagnosis of inherited disorders. This paper describes detailed protocols for isolation, restriction, and blot hybridisation of genomic DNA. Problems and alternatives in the procedure are discussed and a troubleshooting guide has been provided to help rectify faults. Images PMID:6086927

  11. TLC-Blot (Far-Eastern Blot) and Its Application to Functional Lipidomics.

    PubMed

    Taki, Takao

    2015-01-01

    A simple method for transfer of lipids-including phospholipids, glycolipids, and neutral lipids-from a high performance thin layer chromatography (HPTLC) plate to a polyvinylidene difluoride (PVDF) membrane, TLC-Blot (Far-Eastern Blot), and its biochemical applications are presented. This chapter presents the conventional procedures for separating lipid from tissue samples, cultured cells, and serum and the subsequent development of TLC. Individual lipids separated on an HPTLC plate can be transferred to the PVDF membrane quantitatively and also isolated from the lipid-blotted membrane by a one-step purification procedure. Immunodetection with monoclonal antibodies and treatment with lipid-metabolizing enzymes on the lipid-blotted membrane are possible. The method for identification of individual lipids transferred on the PVDF membrane using matrix-assisted laser desorption/ionization quadrupole ion trap time-of-flight mass spectrometry (TLC-Blot/MALDI-TOF MS) is shown as a functional lipidomics application.

  12. BLOT II Ver.1.39

    SciTech Connect

    2003-06-03

    BLOT II is a graphic program for post-processing finite element analyses output in the EXODUS II database format. It is command driven with free-format input and can drive graphics devices supported by the Sandia Virtual Device Interface. BLOT produces mesh plots of the analysis output variables including deformed mesh plots, line contours, filled (painted) contours, vector plots of two/three variables (velocity vectors), and symbol plots of scalar variables (discrete cracks). Features include pathlines of analysis variables drawn on the mesh, element selection by material, element birth and death, multiple views combining several displays on each plot, symmetry mirroring, and node and element numbering. X-Y plots of the analysis variables include time vs. variable plots or variable vs. variable plots, and distance vs. variable plots at selected time stips where distance is the accumulated distance between pairs of nodes or element centers.

  13. Mechanism of DNA (Southern) and protein (Western) blotting on cellulose nitrate and other membranes.

    PubMed

    Van Oss, C J; Good, R J; Chaudhury, M K

    1987-03-27

    The transfer of DNA fractions from hydrophilic gels to nitrocellulose membranes (Southern blotting) which was soon followed by the description of an analogous procedure for RNA (Northern blotting), and somewhat later for proteins (Western blotting), has rapidly become an important separation and characterization method in molecular biology, genetic engineering, and immunological detection. Surface tension measurements have shown that the interfacial attraction between DNA and cellulose esters (-delta G132) in aqueous media can be considerable. The weaker binding energy of proteins to cellulose nitrate and to cellulose acetate may be compared to hydrophobic interaction chromatography, as on account of the somewhat lower [-delta G132] values, it often is necessary to "fix" them more tightly onto nitrocellulose by using high salt concentrations. The binding energy of RNA to both cellulose esters also is rather low. In addition to the effect of high ionic strength, the effect of adding methanol, and the effects of denaturation, heating and drying on the energy of attachment of the biopolymers to cellulose esters, have been studied. Cationized nylon membranes have been advocated recently, especially for electrophoretic transfer of nucleic acids (in which process high salt concentrations cannot easily be used). With positively charged nylon membranes, the attachment mainly occurs through the electrostatic attraction between the strongly negatively charged nucleic acids (or proteins) and the positively charged membrane. Also, more apolar membranes (of polyvinyl difluoride) have been proposed, which manifest a strong interfacial (hydrophobic) attraction to all the above biopolymers (regardless of their electrostatic charge). However, with these two novel membrane types it is no longer possible to exploit the large difference in binding energy between DNA and RNA, which makes cellulose nitrate membranes so uniquely suited for RNA-DNA hybridization assays.

  14. Western Blotting Inaccuracies with Unverified Antibodies: Need for a Western Blotting Minimal Reporting Standard (WBMRS).

    PubMed

    Gilda, Jennifer E; Ghosh, Rajeshwary; Cheah, Jenice X; West, Toni M; Bodine, Sue C; Gomes, Aldrin V

    2015-01-01

    Western blotting is a commonly used technique in biological research. A major problem with Western blotting is not the method itself, but the use of poor quality antibodies as well as the use of different experimental conditions that affect the linearity and sensitivity of the Western blot. Investigation of some conditions that are commonly used and often modified in Western blotting, as well as some commercial antibodies, showed that published articles often fail to report critical parameters needed to reproduce the results. These parameters include the amount of protein loaded, the blocking solution and conditions used, the amount of primary and secondary antibodies used, the antibody incubation solutions, the detection method and the quantification method utilized. In the present study, comparison of ubiquitinated proteins in rat heart and liver samples showed different results depending on the antibody utilized. Validation of five commercial ubiquitin antibodies using purified ubiquitinated proteins, ubiquitin chains and free ubiquitin showed that these antibodies differ in their ability to detect free ubiquitin or ubiquitinated proteins. Investigating proteins modified with interferon-stimulated gene 15 (ISG15) in young and old rat hearts using six commercially available antibodies showed that most antibodies gave different semi-quantitative results, suggesting large variability among antibodies. Evidence showing the importance of the Western blot buffer and the concentration of antibody used is presented. Hence there is a critical need for comprehensive reporting of experimental conditions to improve the accuracy and reproducibility of Western blot analysis. A Western blotting minimal reporting standard (WBMRS) is suggested to improve the reproducibility of Western blot analysis.

  15. Aboveground and belowground responses to nutrient additions and herbivore exclusion in Arctic tundra ecosystems in northern Alaska

    NASA Astrophysics Data System (ADS)

    Moore, J. C.; Gough, L.; Simpson, R.; Johnson, D. R.

    2011-12-01

    The Arctic has experienced significant increased regional warming over the past 30 years. Warming generally increases tundra soil nutrient availability by creating a more favorable environment for plant growth, decomposition and nutrient mineralization. Aboveground there has been a "greening" of the Arctic with increased net primary productivity (NPP), and an increase in woody vegetation. Concurrent with the changes aboveground has been an increase in root growth at lower depths and a loss of soil organic C (40 -100 g C m-2 yr-1). Given that arctic soils contain 14% of the global soil C pool, understanding the mechanisms behind shifts of this magnitude that are changing arctic soils from a net sink to a net source of atmospheric C is critical. We took an integrated multi-trophic level approach to examine how altering soil nutrients and mammalian herbivore activity affects vegetation, soil fauna, and microbial communities as well as soil physical characteristics in moist acidic (MAT) and dry heath (DH) tundra. Our work was conducted at the Arctic LTER site in northern Alaska. We sampled the nutrient (controls and annual N+P additions) and herbivore (controls and exclosures) manipulations established in 1996 after 10 years of treatment. Models that incorporated the biomass estimates from the field were used to characterize the trophic structure of the belowground food web and to estimate carbon flux among soil organisms and C-mineralization rates. Both MAT and DH exhibited significant increases in NPP and root growth and changes in vegetation structure with transitions from a mixed community to deciduous shrubs in MAT and from lichens to grasses and shrubs in DH, with nutrient additions and herbivore exclosures. Belowground responses to the treatments were dependent on ecosystem type, but exposed alterations in trophic structure that included changes in microbial biomass, the establishment of microbivorous enchytreaids, increases in root-feeding nematodes, and

  16. Line blot and western blot immunoassays for diagnosis of Mediterranean spotted fever.

    PubMed Central

    Raoult, D; Dasch, G A

    1989-01-01

    The line blot, a new immunoassay in which antigens are placed on nitrocellulose as narrow lines, was evaluated for its sensitivity and specificity relative to the microimmunofluorescence assay for the diagnosis of Mediterranean spotted fever (MSF). The line blot assay was only slightly less sensitive and less specific than the microimmunofluorescence assay for detection of immunoglobulin M (IgM) or IgG in 100 serum specimens from 42 patients with MSF. No line blot reactions were observed among 50 control serum specimens from febrile patients with other illnesses. The line blot assay was largely group reactive for spotted fever rickettsiae, but 26% of the positive serum specimens also cross-reacted by IgM with Rickettsia typhi. Western immunoblotting was used to characterize the antigenic components recognized by 19 MSF serum specimens. For both IgM and IgG, lipopolysaccharide was the cross-reactive group antigen, whereas the high-molecular-weight species-specific protein antigens (SPAs) were the only reactive proteins. Relative to the other nine rickettsiae, Rickettsia bellii was unique both in exhibiting no SPA reactions and in having a lipopolysaccharide with a predominantly high-molecular-weight distribution. Although most of the 19 MSF serum specimens examined by Western blotting exhibited preferential reactivity to SPAs of two strains of R. conorii and weaker reactions to the other rickettsiae, 2 serum specimens exhibited SPA reactions consistent with typhus infections. In comparison with other assays, the line blot and Western blot immunoassays have advantages which may permit an improvement in the general availability and commercialization of assays for the serodiagnosis of rickettsial infections. Images PMID:2506223

  17. Western blot analysis of adhesive interactions under fluid shear conditions: the blot rolling assay.

    PubMed

    Sackstein, Robert; Fuhlbrigge, Robert

    2015-01-01

    Western blotting has proven to be an important technique in the analysis of receptor-ligand interactions (i.e., by ligand blotting) and for identifying molecules mediating cell attachment (i.e., by cell blotting). Conventional ligand blotting and cell blotting methods employ non-dynamic (static) incubation conditions, whereby molecules or cells of interest are placed in suspension and overlaid on membranes. However, many cell-cell and cell-matrix adhesive interactions occur under fluid shear conditions, and shear stress itself mediates and/or facilitates the engagement of these physiologically appropriate receptors and ligands. Notably, shear forces critically influence the adhesion of circulating cells and platelets to vessel walls in physiologic cell migration and hemostasis, as well as in inflammatory and thrombotic disorders, cancer metastasis, and atherosclerosis. Use of non-dynamic blotting conditions to analyze such interactions can introduce bias, overtly missing relevant effectors and/or exaggerating the relative role(s) of non-physiologic adhesion molecules. To address this shortfall, we have developed a new technique for identifying binding interactions under fluid shear conditions, the "blot rolling assay." Using this method, molecules in a complex mixture are resolved by gel electrophoresis, transferred to a membrane that is rendered semitransparent, and the membrane is then incorporated into a parallel-plate flow chamber apparatus. Under controlled flow conditions, cells or particles bearing adhesion proteins of interest are then introduced into the chamber and interactions with individual immobilized molecules (bands) can be visualized in real time. The substrate molecule(s) supporting adhesion under fluid shear can then be identified by staining with specific antibodies or by excising the relevant band(s) and performing mass spectrometry or microsequencing of the isolated material. This method thus allows for the identification, within a complex

  18. Western blot analysis of adhesive interactions under fluid shear conditions: the blot rolling assay.

    PubMed

    Sackstein, Robert; Fuhlbrigge, Robert

    2009-01-01

    Western blotting has proven to be an important technique in analysis of receptor-ligand interactions (i.e., by ligand blotting) and for identifying molecules mediating cell attachment (i.e., by cell blotting). Conventional ligand blotting and cell blotting methods employ nondynamic (static) incubation conditions, whereby molecules or cells of interest are placed in suspension and overlaid on membranes. However, many cell-cell and cell-matrix adhesive interactions occur under fluid shear conditions, and shear stress itself mediates and/or facilitates the engagement of these physiologically appropriate receptors and ligands. Notably, shear forces critically influence the adhesion of circulating cells and platelets to vessel walls in physiologic cell migration and hemostasis, as well as in inflammatory and thrombotic disorders, cancer metastasis, and atherosclerosis. Use of nondynamic blotting conditions to analyze such interactions can introduce bias, overtly missing relevant effectors and/or exaggerating the relative role(s) of nonphysiologic adhesion molecules. To address this shortfall, we have developed a new technique for identifying binding interactions under fluid shear conditions, the "blot rolling assay." Using this method, molecules in a complex mixture are resolved by gel electrophoresis, transferred to a membrane that is rendered semi-transparent, and the membrane is then incorporated into a parallel-plate flow chamber apparatus. Under controlled flow conditions, cells or particles bearing adhesion proteins of interest are then introduced into the chamber and interactions with individual immobilized molecules (bands) can be visualized in real-time. The substrate molecule(s) supporting adhesion under fluid shear can then be identified by staining with specific antibodies or by excising the relevant band(s) and performing mass spectrometry or microsequencing of the isolated material. This method thus allows for the identification, within a complex mixture

  19. Repeated probing of Southwestern blots using alkaline phosphatase stripping.

    PubMed

    Jia, Yinshan; Jiang, Daifeng; Jarrett, Harry W

    2010-11-05

    Southwestern blotting is when a DNA sequence is used to probe DNA-binding proteins on an electrophoretic gel blot. It would be highly desirable to be able to probe a blot repeatedly with different DNA sequences. Alkaline phosphatase can remove 5'-phosphoryl groups from DNA and radiolabeled 5'-(32)P-DNA probes are commonly used in Southwestern blotting. Here is shown that once probed, the radioisotope signal on the blot can be effectively removed by brief digestion with alkaline phosphatase, and the blot can then be repeatedly probed at least six times with different DNA probes. This exceeds the repetitions possible with another commonly used method using SDS. The technique can be used with either one-dimensional or multi-dimensional Southwestern blots and does not have a large effect on the phosphorylation state of the blotted proteins. An alternative method using T4 polynucleotide kinase stripping is also introduced but was less well characterized.

  20. Soil microbial responses to forest floor litter manipulation and nitrogen addition in a mixed-wood forest of northern China.

    PubMed

    Sun, Xiao-Lu; Zhao, Jing; You, Ye-Ming; Jianxin Sun, Osbert

    2016-01-14

    Changes in litterfall dynamics and soil properties due to anthropogenic or natural perturbations have important implications to soil carbon (C) and nutrient cycling via microbial pathway. Here we determine soil microbial responses to contrasting types of litter inputs (leaf vs. fine woody litter) and nitrogen (N) deposition by conducting a multi-year litter manipulation and N addition experiment in a mixed-wood forest. We found significantly higher soil organic C, total N, microbial biomass C (MBC) and N (MBN), microbial activity (MR), and activities of four soil extracellular enzymes, including β-glucosidase (BG), N-acetyl-β-glucosaminidase (NAG), phenol oxidase (PO), and peroxidase (PER), as well as greater total bacteria biomass and relative abundance of gram-negative bacteria (G-) community, in top soils of plots with presence of leaf litter than of those without litter or with presence of only fine woody litter. No apparent additive or interactive effects of N addition were observed in this study. The occurrence of more labile leaf litter stimulated G-, which may facilitate microbial community growth and soil C stabilization as inferred by findings in literature. A continued treatment with contrasting types of litter inputs is likely to result in divergence in soil microbial community structure and function.

  1. Soil microbial responses to forest floor litter manipulation and nitrogen addition in a mixed-wood forest of northern China

    PubMed Central

    Sun, Xiao-Lu; Zhao, Jing; You, Ye-Ming; Jianxin Sun, Osbert

    2016-01-01

    Changes in litterfall dynamics and soil properties due to anthropogenic or natural perturbations have important implications to soil carbon (C) and nutrient cycling via microbial pathway. Here we determine soil microbial responses to contrasting types of litter inputs (leaf vs. fine woody litter) and nitrogen (N) deposition by conducting a multi-year litter manipulation and N addition experiment in a mixed-wood forest. We found significantly higher soil organic C, total N, microbial biomass C (MBC) and N (MBN), microbial activity (MR), and activities of four soil extracellular enzymes, including β-glucosidase (BG), N-acetyl-β-glucosaminidase (NAG), phenol oxidase (PO), and peroxidase (PER), as well as greater total bacteria biomass and relative abundance of gram-negative bacteria (G-) community, in top soils of plots with presence of leaf litter than of those without litter or with presence of only fine woody litter. No apparent additive or interactive effects of N addition were observed in this study. The occurrence of more labile leaf litter stimulated G-, which may facilitate microbial community growth and soil C stabilization as inferred by findings in literature. A continued treatment with contrasting types of litter inputs is likely to result in divergence in soil microbial community structure and function. PMID:26762490

  2. Combined effects of nitrogen addition and litter manipulation on nutrient resorption of Leymus chinensis in a semi-arid grassland of northern China.

    PubMed

    Li, X; Liu, J; Fan, J; Ma, Y; Ding, S; Zhong, Z; Wang, D

    2015-01-01

    Plant growth in semi-arid ecosystems is usually severely limited by soil nutrient availability. Alleviation of these resource stresses by fertiliser application and aboveground litter input may affect plant internal nutrient cycling in such regions. We conducted a 4-year field experiment to investigate the effects of nitrogen (N) addition (10 g N·m(-2) ·year(-1)) and plant litter manipulation on nutrient resorption of Leymus chinensis, the dominant native grass in a semi-arid grassland in northern China. Although N addition had no clear effects on N and phosphorus (P) resorption efficiencies in leaves and culms, N fertilisation generally decreased leaf N resorption proficiency by 54%, culm N resorption proficiency by 65%. Moreover, N fertilisation increased leaf P resorption proficiency by 13%, culm P resorption proficiency by 20%. Under ambient or enriched N conditions, litter addition reduced N and P resorption proficiencies in both leaves and culms. The response of P resorption proficiency to litter manipulation was more sensitive than N resorption proficiency: P resorption proficiency in leaves and culms decreased strongly with increasing litter amount under both ambient and enriched N conditions. In contrast, N resorption proficiency was not significantly affected by litter addition, except for leaf N resorption proficiency under ambient N conditions. Furthermore, although litter addition caused a general decrease of leaf and culm nutrient resorption efficiencies under both ambient and enriched N conditions, litter addition effects on nutrient resorption efficiency were much weaker than the effects of litter addition on nutrient resorption proficiency. Taken together, our results show that leaf and non-leaf organs of L. chinensis respond consistently to altered soil N availability. Our study confirms the strong effects of N addition on plant nutrient resorption processes and the potential role of aboveground litter, the most important natural fertiliser in

  3. BLOTS AND ALL: A HISTORY OF THE RORSCHACH INK BLOT TEST IN BRITAIN.

    PubMed

    Hubbard, Katherine; Hegarty, Peter

    2016-01-01

    Despite the easily recognizable nature of the Rorschach ink blot test very little is known about the history of the test in Britain. We attend to the oft-ignored history of the Rorschach test in Britain and compare it to its history in the US. Prior to the Second World War, Rorschach testing in Britain had attracted advocates and critiques. Afterward, the British Rorschach Forum, a network with a high proportion of women, developed around the Tavistock Institute in London and The Rorschach Newsletter. In 1968, the International Rorschach Congress was held in London but soon after the group became less exclusive, and fell into decline. A comparative account of the Rorschach in Britain demonstrates how different national institutions invested in the 'projective hypothesis' according to the influence of psychoanalysis, the adoption of a nationalized health system, and the social positioning of 'others' throughout the twentieth century. In comparing and contrasting the history of the Rorschach in Britain and the US, we decentralize and particularize the history of North American Psychology.

  4. The line blot assay: problems with titrating first and second antibodies for Western blot and immunohistochemistry assays?

    PubMed

    Rojas-Espinosa, O; Silva-Miranda, M; Wek-Rodriguez, K; Arce-Paredes, P

    2006-01-01

    We describe a technique designed to assess the optimal dilution of primary and secondary antibodies, to be used in Western blot, dot blot, the multi-antigen print immunoassay (MAPIA) and immunohistochemistry assays. The method that we call "line blot" is not an alternative but a practical, complementary tool for the above techniques that assures definitive results are obtained from single assays, so there is no need to repeat the assay. As with most immunoenzymatic assays, the line blot assay is very sensitive, allowing the detection of absolute amounts of antigen as low as 2.5 ng in the 0.5 cm-long segment line (see Results), depending on the strength of the secondary, enzyme-labelled antibody.

  5. Western Blot of Stained Proteins from Dried Polyacrylamide Gels

    NASA Technical Reports Server (NTRS)

    Gruber, Claudia; Stan-Lotter, Helga

    1996-01-01

    Western blotting of proteins is customarily performed following their separation on polyacrylamide gels, either prior to staining (1) or, as recently reported, following staining (2). We describe here Western blotting with stained gels, which had been dried and some of which had been stored for years. This procedure permits immunological analysis of proteins, to which antisera may have become available only later, or where the application of newly developed sensitive detection methods is desired. Once rehydration of the gels is achieved, proteins can be-transferred to blotting membranes by any appropriate protocol. Proteins stained with Coomassie Blue have to be detected with a non-chromogenic method, such as the film-based enhanced chemiluminescence (ECL)2) procedure (3). Silver stained proteins, which transfer in the colorless form, may be visualized by any detection method, although, because of the usually very low amounts of proteins, detection by ECL is preferable. Blotting of stained proteins from rehydrated gels is as rapid and as quantitative as from freshly prepared gels, in contrast to blotting from wet stained gels, which requires extensive washing and results in low transfer efficiency (2). Together with a photographic record of the gel pattern, unambiguous identification of immunoreactive proteins from complex mixtures is possible. Some further applications of this work are discussed.

  6. Relative impact of previous disturbance history on the likelihood of additional disturbance in the Northern United States Forest Service USFS Region

    NASA Astrophysics Data System (ADS)

    Hernandez, A. J.

    2015-12-01

    The Landsat archive is increasingly being used to detect trends in the occurrence of forest disturbance. Beyond information about the amount of area affected, forest managers need to know if and how disturbance regimes change. The National Forest System (NFS) has developed a comprehensive plan for carbon monitoring that requires a detailed temporal mapping of forest disturbances across 75 million hectares. A long-term annual time series that shows the timing, extent, and type of disturbance beginning in 1990 and ending in 2011 has been prepared for several USFS Regions, including the Northern Region. Our mapping starts with an automated detection of annual disturbances using a time series of historical Landsat imagery. Automated detections are meticulously inspected, corrected and labeled using various USFS ancillary datasets. The resulting maps of verified disturbance show the timing and types are fires, harvests, insect activity, disease, and abiotic (wind, drought, avalanche) damage. Also, the magnitude of each change event is modeled in terms of the proportion of canopy cover lost. The sequence of disturbances for every pixel since 1990 has been consistently mapped and is available across the entirety of NFS. Our datasets contain sufficient information to describe the frequency of stand replacement, as well as how often disturbance results in only a partial loss of canopy. This information provides empirical insight into how an initial disturbance may predispose a stand to further disturbance, and it also show a climatic signal in the occurrence of processes such as fire and insect epidemics. Thus, we have the information to model the likelihood of occurrence of certain disturbances after a given event (i.e. if we have a fire in the past what does that do to the likelihood of occurrence of insects in the future). Here, we explore if previous disturbance history is a reliable predictor of additional disturbance in the future and we present results of applying

  7. Effects of Reusing Gel Electrophoresis and Electrotransfer Buffers on Western Blotting

    PubMed Central

    Omotola, Oluwabukola B.; Heda, Rajiv P.; Avery, Jamie

    2016-01-01

    SDS-PAGE and Western blotting are 2 of the most commonly used biochemical methods for protein analysis. Proteins are electrophoretically separated based on their MWs by SDS-PAGE and then electrotransferred to a solid membrane surface for subsequent protein-specific analysis by immunoblotting, a procedure commonly known as Western blotting. Both of these procedures use a salt-based buffer, with the latter procedure consisting of methanol as an additive known for its toxicity. Previous reports present a contradictory view in favor or against reusing electrotransfer buffer, also known as Towbin’s transfer buffer (TTB), with an aim to reduce the toxic waste. In this report, we present a detailed analysis of not only reusing TTB but also gel electrophoresis buffer (EB) on proteins of low to high MW range. Our results suggest that EB can be reused for at least 5 times without compromising the electrophoretic separation of mixture of proteins in an MW standard, BSA, and crude cell lysates. Additionally, reuse of EB did not affect the quality of subsequent Western blots. Successive reuse of TTB, on the other hand, diminished the signal of proteins of different MWs in a protein standard and a high MW membrane protein cystic fibrosis transmembrane-conductance regulator (CFTR) in Western blotting. PMID:27582639

  8. Evaluating strategies to normalise biological replicates of Western blot data.

    PubMed

    Degasperi, Andrea; Birtwistle, Marc R; Volinsky, Natalia; Rauch, Jens; Kolch, Walter; Kholodenko, Boris N

    2014-01-01

    Western blot data are widely used in quantitative applications such as statistical testing and mathematical modelling. To ensure accurate quantitation and comparability between experiments, Western blot replicates must be normalised, but it is unclear how the available methods affect statistical properties of the data. Here we evaluate three commonly used normalisation strategies: (i) by fixed normalisation point or control; (ii) by sum of all data points in a replicate; and (iii) by optimal alignment of the replicates. We consider how these different strategies affect the coefficient of variation (CV) and the results of hypothesis testing with the normalised data. Normalisation by fixed point tends to increase the mean CV of normalised data in a manner that naturally depends on the choice of the normalisation point. Thus, in the context of hypothesis testing, normalisation by fixed point reduces false positives and increases false negatives. Analysis of published experimental data shows that choosing normalisation points with low quantified intensities results in a high normalised data CV and should thus be avoided. Normalisation by sum or by optimal alignment redistributes the raw data uncertainty in a mean-dependent manner, reducing the CV of high intensity points and increasing the CV of low intensity points. This causes the effect of normalisations by sum or optimal alignment on hypothesis testing to depend on the mean of the data tested; for high intensity points, false positives are increased and false negatives are decreased, while for low intensity points, false positives are decreased and false negatives are increased. These results will aid users of Western blotting to choose a suitable normalisation strategy and also understand the implications of this normalisation for subsequent hypothesis testing.

  9. Evaluating Strategies to Normalise Biological Replicates of Western Blot Data

    PubMed Central

    Degasperi, Andrea; Birtwistle, Marc R.; Volinsky, Natalia; Rauch, Jens; Kolch, Walter; Kholodenko, Boris N.

    2014-01-01

    Western blot data are widely used in quantitative applications such as statistical testing and mathematical modelling. To ensure accurate quantitation and comparability between experiments, Western blot replicates must be normalised, but it is unclear how the available methods affect statistical properties of the data. Here we evaluate three commonly used normalisation strategies: (i) by fixed normalisation point or control; (ii) by sum of all data points in a replicate; and (iii) by optimal alignment of the replicates. We consider how these different strategies affect the coefficient of variation (CV) and the results of hypothesis testing with the normalised data. Normalisation by fixed point tends to increase the mean CV of normalised data in a manner that naturally depends on the choice of the normalisation point. Thus, in the context of hypothesis testing, normalisation by fixed point reduces false positives and increases false negatives. Analysis of published experimental data shows that choosing normalisation points with low quantified intensities results in a high normalised data CV and should thus be avoided. Normalisation by sum or by optimal alignment redistributes the raw data uncertainty in a mean-dependent manner, reducing the CV of high intensity points and increasing the CV of low intensity points. This causes the effect of normalisations by sum or optimal alignment on hypothesis testing to depend on the mean of the data tested; for high intensity points, false positives are increased and false negatives are decreased, while for low intensity points, false positives are decreased and false negatives are increased. These results will aid users of Western blotting to choose a suitable normalisation strategy and also understand the implications of this normalisation for subsequent hypothesis testing. PMID:24475266

  10. Additions to Philippine Slender Skinks of the Brachymeles bonitae Complex (Reptilia: Squamata: Scincidae) II: a new species from the northern Philippines.

    PubMed

    Siler, Cameron D; Davis, Drew R; Freitas, Elyse S; Huron, Nicholas A; Geheber, Aaron D; Watters, Jessa L; Penrod, Michelle L; Papeș, Monica; Amrein, Andrew; Anwar, Alyssa; Cooper, Dontae; Hein, Tucker; Manning, Annalisa; Patel, Neeral; Pinaroc, Lauren; Diesmos, Arvin C; Diesmos, Mae L; Oliveros, Carl H; Brown, Rafe M

    2016-06-28

    We describe a new digitless scincid lizard of the genus Brachymeles from northern Luzon and Camiguin Norte islands in the Philippines. This species belongs to the Brachymeles bonitae Complex, and both molecular and morphological data confirm that this species is distinct from all other congeners. Formerly considered to be a single widespread species, this group of species has been the focus of recent systematic reviews. Here we describe a new species in the B. bonitae Complex, recognized currently to constitute five species. Brachymeles ilocandia sp. nov. is the second digitless and the seventeenth non-pentadactyl species in genus. The description of this species brings the total number of species in the genus to 40, and provides new insight into unique distribution patterns of species of the northern Philippines.

  11. Microfluidic Western Blotting of Low-Molecular-Mass Proteins

    PubMed Central

    2015-01-01

    We describe a microfluidic Western blot assay (μWestern) using a Tris tricine discontinuous buffer system suitable for analyses of a wide molecular mass range (6.5–116 kDa). The Tris tricine μWestern is completed in an enclosed, straight glass microfluidic channel housing a photopatterned polyacrylamide gel that incorporates a photoactive benzophenone methacrylamide monomer. Upon brief ultraviolet (UV) light exposure, the hydrogel toggles from molecular sieving for size-based separation to a covalent immobilization scaffold for in situ antibody probing. Electrophoresis controls all assay stages, affording purely electronic operation with no pumps or valves needed for fluid control. Electrophoretic introduction of antibody into and along the molecular sieving gel requires that the probe must traverse through (i) a discontinuous gel interface central to the transient isotachophoresis used to achieve high-performance separations and (ii) the full axial length of the separation gel. In-channel antibody probing of small molecular mass species is especially challenging, since the gel must effectively sieve small proteins while permitting effective probing with large-molecular-mass antibodies. To create a well-controlled gel interface, we introduce a fabrication method that relies on a hydrostatic pressure mismatch between the buffer and polymer precursor solution to eliminate the interfacial pore-size control issues that arise when a polymerizing polymer abuts a nonpolymerizing polymer solution. Combined with a new swept antibody probe plug delivery scheme, the Tris tricine μWestern blot enables 40% higher separation resolution as compared to a Tris glycine system, destacking of proteins down to 6.5 kDa, and a 100-fold better signal-to-noise ratio (SNR) for small pore gels, expanding the range of applicable biological targets. PMID:25268977

  12. A streamlined Western blot exercise: An efficient and greener approach in the laboratory classroom.

    PubMed

    Ness, Traci L; Robinson, Rebekah L; Mojadedi, Wais; Peavy, Lydia; Weiland, Mitch H

    2015-01-01

    SDS-PAGE and western blotting are two commonly taught protein detection techniques in biochemistry and molecular biology laboratory classrooms. A pitfall associated with incorporating these techniques into the laboratory is the significant wait times that do not allow students to obtain timely results. The waiting associated with SDS-PAGE comes from staining and destaining, whereas with western blotting it is the times required for antibody incubations and the numerous wash steps. This laboratory exercise incorporates 2,2,2-trichloroethanol (TCE) into the SDS-PAGE gel allowing for visualization of migrated proteins in a matter of minutes, saving both the time and chemical waste associated with traditional Coomassie staining. Additionally, TCE staining does not affect protein transfer eliminating the requirement for duplicated gels for total protein and western analyses. Protein transfer can be confirmed immediately without the use of Ponceau S staining. Lastly, this western blot procedure has been further shortened by using an HRP-conjugated primary antibody, which eliminates the secondary antibody incubation and washes, and uses a colorimetric detection to allow for visualization by students without the need for specialized equipment.

  13. Differentiation of larva migrans caused by Baylisascaris procyonis and Toxocara species by Western blotting.

    PubMed

    Dangoudoubiyam, Sriveny; Kazacos, Kevin R

    2009-11-01

    Baylisascaris procyonis and Toxocara species are two important causes of larva migrans in humans. Larva migrans caused by Toxocara spp. is well known and is diagnosed serologically by enzyme immunoassay. Over a dozen cases of larva migrans and associated eosinophilic encephalitis caused by B. procyonis have also been reported, and at least a dozen additional cases are known. An enzyme-linked immunosorbent assay (ELISA) using the excretory-secretory (ES) antigen of B. procyonis larvae is currently being used in our laboratory as an aid in the diagnosis of this infection in humans. Clinically affected individuals show very high reactivity (measured as the optical density) on this ELISA; however, a one-way cross-reactivity with Toxocara spp. has been observed. As an approach to differentiate these two infections based on serology, we performed Western blots, wherein the B. procyonis ES antigen was reacted with serum samples from individuals known to be positive for either Toxocara spp. or B. procyonis larva migrans. Western blot results showed that B. procyonis antigens of between 30 and 45 kDa were specifically identified only by the sera from individuals with Baylisascaris larva migrans, thus allowing for differentiation between the two infections. This included human patient serum samples submitted for serologic testing, as well as sera from rabbits experimentally infected with B. procyonis. When used in conjunction with the ELISA, Western blotting could be an efficient tool for diagnosis of this infection in humans.

  14. PCR versus Southern blot detection of somatic mosaicism in fragile X syndrome

    SciTech Connect

    Mueller, O.T.; Amar, M.J.A.; Gallardo, L.A.; Kousseff, B.G.

    1994-09-01

    The incidence of somatic mosaicism in males with fragile X syndrome has been reported to be as high as 17% of all clinically affected males. Mosaic cases usually do not show the cytogenetic fragile site at Xq27.3 and generally are fully affected according to the clinical criteria, although some subjects show somewhat milder symptoms. Detection of mosaicism relies on the identification of multiple distinct sizes of the CGG size anomaly in the 5{prime} untranslated exon of the FMR-1 gene. Some alleles identified are of a premutation size with trinucleotides numbering 50 to 80 X CGG. In addition, there is a greatly expanded allele with well over 200 copies of the CGG repeat detected. We screened 314 subjects for fragile X syndrome over a three year period. Cases were routinely screened by Southern blotting using the StB12.3 probe as well as by polymerase chain reaction. The PCR amplification products were electrophoresed in agarose, blotted and hybridized with a (CGG){sub 5} oligonucleotide followed by chemiluminescent detection. Seventeen males and 16 females were identified with a CGG expansion, including two males with premutations with repeat sizes of between 50 and 100 CGG trinucleotides and three cases exhibiting somatic mosaicism. The mosaic cases had both a premutation-sized allele and one or more expanded alleles of over 250 CGG copies. The mosaic cases were usually undetected with Southern blotting but easily identified with this PCR protocol. The relative proportion of the expanded allele as determined by scanning densitometry were 70%, 35%, and 5% in the three cases. All three cases were cytogenetically negative. The clinical severity of the mosaic cases was variable, with symptoms ranging from severe MR with most of the physical stigmata to mild learning disability. In our experience, Southern blotting allows more accurate sizing of the expanded allele; however, PCR is essential to identify cases that exhibit mosaicism.

  15. Blotting Assisted by Heating and Solvent Extraction for DESI-MS Imaging

    NASA Astrophysics Data System (ADS)

    Cabral, Elaine C.; Mirabelli, Mario F.; Perez, Consuelo J.; Ifa, Demian R.

    2013-06-01

    Imprints of potato sprout ( Solanum tuberosum L.), gingko leaves (Gingko biloba L. ) and strawberries (Fragaria x ananassa Duch. ) were successfully imaged by desorption electrospray ionization mass spectrometry (DESI-MS) on TLC plates through blotting assisted by heating and/or solvent extraction. Ion images showing the distribution of significant compounds such as glycoalkaloid toxins in potato sprout, ginkgolic acids and flavonoids in ginkgo leaves, and sugars and anthocyanidin in strawberry were obtained. Practical implications of this work include analysis of a wide range of irregular or soft materials by different imprinting conditions without requiring the addition of matrices or use of specific kinds of surfaces.

  16. Selection of DNA aptamers against VEGF165 using a protein competitor and the aptamer blotting method.

    PubMed

    Hasegawa, Hijiri; Sode, Koji; Ikebukuro, Kazunori

    2008-05-01

    Two DNA aptamers against a tumor marker protein, human vascular endothelial growth factor (VEGF(165)) were identified. In the screening process, another protein was used as the competitor to isolate those aptamers that have high specificity for the target. In addition, we evaluated the affinities of the enriched library by means of aptamer blotting. The isolated aptamers bound to VEGF(165) with a K(d) value in the range of a few hundred nanomoles, and did not bind to the competitor. This selection method enabled us to efficiently select the specific aptamers against the target protein. These specific aptamers would be useful sensor elements for cancer diagnosis.

  17. Serological Diagnosis of Paracoccidioidomycosis through a Western Blot Technique

    PubMed Central

    Perenha-Viana, M. C. Z.; Gonzales, I. A. A.; Brockelt, S. R.; Machado, L. N. C.

    2012-01-01

    Paracoccidioidomycosis (PCM) is a serious infectious disease that progresses toward death if untreated. Its confirmatory diagnosis is made by the detection of the fungus Paracoccidioides brasiliensis in a direct mycological examination or by histopathology. However, these techniques are of low sensitivity. Serological tests seem to be more promising. The objective of this study was to test Western blot (WB) analysis using sera from patients suspected of PCM to determine whether it represents a safe and sensitive serological technique for a rapid and effective diagnosis for this disease. Sera from 517 patients were analyzed through WB analysis and double-immunodiffusion (DID) techniques using a crude exoantigen of P. brasiliensis 339. DID gave positive reactions for 140 sera (27%) and WB for 250 sera (48.4%). All sera that had a positive reaction by DID also had a positive result with a 43-kDa glycoprotein by WB analysis. Among the 377 samples that were negative by DID, 29.1% were reactive in WB analysis. For the cutoff dilution used (1:400), a positive reaction was not observed with any of the 102 sera from patients with other diseases in regions where such diseases are endemic and 30 healthy individuals tested as negative controls. These results prove WB analysis to be a sensitive technique and suggest its inclusion among routine laboratory assays as a safe method for PCM diagnosis. PMID:22301695

  18. Differential ecophysiological response of deciduous shrubs and a graminoid to long-term experimental snow reductions and additions in moist acidic tundra, Northern Alaska.

    PubMed

    Pattison, Robert R; Welker, Jeffrey M

    2014-02-01

    Changes in winter precipitation that include both decreases and increases in winter snow are underway across the Arctic. In this study, we used a 14-year experiment that has increased and decreased winter snow in the moist acidic tussock tundra of northern Alaska to understand impacts of variation in winter snow depth on summer leaf-level ecophysiology of two deciduous shrubs and a graminoid species, including: instantaneous rates of leaf gas exchange, and δ(13)C, δ(15)N, and nitrogen (N) concentrations of Betula nana, Salix pulchra, and Eriophorum vaginatum. Leaf-level measurements were complemented by measurements of canopy leaf area index (LAI) and depth of thaw. Reductions in snow lowered summer leaf photosynthesis, conductance, and transpiration rates by up to 40% compared to ambient and deep snow conditions for Eriophorum vaginatum, and reduced Salix pulchra conductance and transpiration by up to 49%. In contrast, Betula nana exhibited no changes in leaf gas exchange in response to lower or deeper snow. Canopy LAI increased with added snow, while reduced winter snow resulted in lower growing season soil temperatures and reduced thaw depths. Our findings indicate that the spatial and temporal variability of future snow depth will have individualistic consequences for leaf-level C fixation and water flux by tundra species, and that these responses will be manifested over the longer term by changes in canopy traits, depth of thaw, soil C and N processes, and trace gas (CO2 and H2O) exchanges between the tundra and the atmosphere.

  19. Design, preparation and use of ligated phosphoproteins: a novel approach to study protein phosphatases by dot blot array, ELISA and Western blot assays.

    PubMed

    Sun, Luo; Ghosh, Inca; Barshevsky, Tanya; Kochinyan, Samvel; Xu, Ming-Qun

    2007-07-01

    The study of substrate specificity of protein phosphatases (PPs) is very challenging since it is difficult to prepare a suitable phosphorylated substrate. Phosphoproteins, phosphorylated by a protein kinase, or chemically synthesized phosphopeptides are commonly used substrates for PPs. Both types of these substrates have their advantages and limitations. Phosphoproteins mimic more closely the physiologically relevant PP substrates, but their preparation is technically demanding. Synthetic phosphopeptides present advantages over proteins because they can be easily produced in large quantity and their amino acid sequence can be designed to contain potential determinants of substrate specificity. However, short peptides are less optimal compared to in vivo PP substrates and often display poor and variable binding to different matrices, resulting in low sensitivity in analysis of PP activity on solid support. In this work we utilize the intein-mediated protein ligation (IPL) technique to generate substrates for PPs, combining the advantages of proteins and synthetic peptides in one molecule. The ligation of a synthetic phosphopeptide to an intein-generated carrier protein (CP) with a one-to-one stoichiometry results in the formation of a ligated phosphoprotein (LPP). Three widely used assays, dot blot array, Western blot and ELISA were employed to study the PP activity on LPP substrates. Dephosphorylation was measured by detection of the remaining phosphorylation, or lack of it, with a phospho-specific antibody. The data show the advantage of LPPs over free peptides in assays on solid supports. LPPs exhibited enhanced binding to the matrices used in the study, which significantly improved sensitivity and consistency of the assays. In addition, saturation of the signal was circumvented by serial dilution of the assay samples. This report describes detailed experimental procedures for preparation of LPP substrates and their use in PP assays based on immobilization on

  20. FANCD2 Western blot as a diagnostic tool for Brazilian patients with Fanconi anemia.

    PubMed

    Pilonetto, D V; Pereira, N F; Bitencourt, M A; Magdalena, N I R; Vieira, E R; Veiga, L B A; Cavalli, I J; Ribeiro, R C; Pasquini, R

    2009-03-01

    Fanconi anemia is a rare hereditary disease showing genetic heterogeneity due to a variety of mutations in genes involved in DNA repair pathways, which may lead to different clinical manifestations. Phenotypic variability makes diagnosis difficult based only on clinical manifestations, therefore laboratory tests are necessary. New advances in molecular pathogenesis of this disease led researchers to develop a diagnostic test based on Western blot for FANCD2. The objective of the present study was to determine the efficacy of this method for the diagnosis of 84 Brazilian patients with Fanconi anemia, all of whom tested positive for the diepoxybutane test, and 98 healthy controls. The FANCD2 monoubiquitinated isoform (FANCDS+/FANCD2L-) was not detected in 77 patients (91.7%). In 2 patients (2.4%), there was an absence of both the monoubiquitinated and the non-ubiquitinated proteins (FANCD2S-/FANCD2L-) and 5 patients (5.9%) had both isoforms (FANCD2S+/FANCD2L+). This last phenotype suggests downstream subtypes or mosaicism. All controls were diepoxybutane negative and were also negative on the FANCD2 Western blot. The Western blot for FANCD2 presented a sensitivity of 94% (79/84) and specificity of 100% (98/98). This method was confirmed as an efficient approach to screen Brazilian patients with deleterious mutations on FANCD2 (FANCD2S-/FANCD2L-) or other upstream genes of the FA/BRCA pathway (FANCDS+/FANCD2L-), to confirm the chromosome breakage test and to classify patients according to the level of FA/BRCA pathway defects. However, patients showing both FANCD2 isoforms (FANCD2S+/FANCD2L+) require additional studies to confirm mutations on downstream Fanconi anemia genes or the presence of mosaicism.

  1. Northern Australia

    Atmospheric Science Data Center

    2013-04-16

    article title:  Tropical Northern Australia     View Larger Image ... Imaging SpectroRadiometer (MISR) images of tropical northern Australia were acquired on June 1, 2000 (Terra orbit 2413) during the long dry ...

  2. Gel electrophoresis of polyphenol oxidase with instant identification by in situ blotting.

    PubMed

    Cheng, Tsai-Mu; Huang, Pei-Chen; Pan, Ju-Pin; Lin, Kuan-Yu; Mao, Simon J T

    2007-04-15

    Polyphenol oxidase (PPO) or tyrosinase is an important and ubiquitous enzyme responsible for browning in plants and melanization in animals. The molecular size of the plant PPO is varied among the species and its activity can be enhanced by a variety of anionic detergents. In the present study, we developed a simple method for the first-step identification of PPO in fruit and vegetable extracts. First, 3mm chromatographic paper was immersed in 0.5% (w/v) catechol solution as an immobilized PPO substrate. After running the extract with 10% sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE), one side of the glass plate was removed. The plate was immediately laid on top of the dried catechol-paper. A dark-brown band corresponding to PPO was visualized within 1 min and was further confirmed by a conventional Western blot using an antibody prepared against mushroom PPO. It also reveals that some vegetation (such as tomato, radish, and oriental melon) with low or no detectable activity in a conventional enzyme assay actually possessed marked levels of PPO activity when assessed by PAGE-blot. We propose that an inhibitor is associated with PPO in some plants; the inhibitor, however, is dissociated during the electrophoresis. Therefore, in addition to identify the molecular form of PPO, the present technique may explore the existence of PPO inhibitor(s) in plants. The detail of the method with respect to its relevance for searching a natural PPO inhibitor is described and discussed.

  3. Cell adhesion to proteins separated by lithium dodecyl sulfate-polyacrylamide gel electrophoresis and blotted onto a polyvinylidene difluoride membrane: a new cell-blotting technique.

    PubMed

    Seshi, B

    1994-12-02

    Cell blotting, although conceptually simple, has failed to achieve wide practical application. Described here is a new cell-blotting technique which involves cell adhesion to protein bands after separation by lithium dodecyl sulfate-polyacrylamide gel electrophoresis (LDS-PAGE) and blotting onto polyvinylidene difluoride (PVDF) membrane at 4 degrees C. Cell bands adherent on PVDF are detected using hematoxylin, or propidium iodide (PI) staining followed by viewing under ultraviolet (UV) light. The technique allows quick microscopic visualization of adherent cells composing the bands, without requiring clearing of the membrane. Representative cell adhesion proteins from different sources, i.e., plant lectins (e.g., phytohemagglutinin, PHA; concanavalin A, ConA; and wheat germ agglutinin, WGA); extracellular matrix (ECM) proteins; and integral membrane proteins (e.g., recombinant soluble vascular cell adhesion molecule-1, rs VCAM-1) were tested for cell binding by the new cell-blotting technique using human lymphoid progenitor (NALM-6) and myeloid progenitor (KG1a) cell lines. Cell adhesion proteins retained their adhesion function in all cases tested. Specificity of cell binding on PVDF blot was demonstrated by inhibition of cell adhesion to WGA protein bands using an appropriate sugar, i.e., N-acetyl D-glucosamine. The cell blotting assay was comparable in sensitivity to Coomassie blue staining of protein bands. The ability to conduct protein extraction, separation and blotting at low temperature avoids thermal denaturation, thereby preserving the adhesion properties of the proteins. The electrophoretic/blotting system has unique detergent removal/protein renaturation properties and the ability to preserve functionally active adhesion protein complexes. The cell-blotting technique described is sufficiently robust for routine application in the investigation of novel cell adhesion proteins.

  4. Reversible Ponceau staining as a loading control alternative to actin in Western blots.

    PubMed

    Romero-Calvo, Isabel; Ocón, Borja; Martínez-Moya, Patricia; Suárez, María Dolores; Zarzuelo, Antonio; Martínez-Augustin, Olga; de Medina, Fermín Sánchez

    2010-06-15

    It is becoming standard practice to measure a housekeeping gene, typically actin, in Western blots, as it is the rule in RNA blots. We have applied reversible Ponceau staining to check equal loading of gels and measured actin in parallel under different conditions. Our results show that densitometric analysis is comparable with both techniques. Therefore, routine quantitation of Ponceau staining before antibody probing is validated as an alternative to actin blotting.

  5. Application of a Dot Blot Hybridization Platform to Assess Streptococcus uberis Population Structure in Dairy Herds

    PubMed Central

    Albuquerque, Pedro; Ribeiro, Niza; Almeida, Alexandre; Panschin, Irena; Porfirio, Afonso; Vales, Marta; Diniz, Francisca; Madeira, Helena; Tavares, Fernando

    2017-01-01

    Streptococcus uberis is considered one of the most important pathogens associated with bovine mastitis. While traditionally acknowledged as an environmental pathogen, S. uberis has been shown to adopt a contagious epidemiological pattern in several dairy herds. Since different control strategies are employed depending on the mode of transmission, in-depth studies of S. uberis populations are essential to determine the best practices to control this pathogen. In this work, we optimized and validated a dot blot platform, combined with automatic image analysis, to rapidly assess the population structure of infective S. uberis, and evaluated its efficiency when compared to multilocus sequence analysis (MLSA) genotyping. Two dairy herds with prevalent S. uberis infections were followed in a 6 month period, in order to collect and characterize isolates from cows with persistent infections. These herds, located in Portugal (Barcelos and Maia regions), had similar management practices, with the herd from Barcelos being smaller and having a better milking parlor management, since infected cow segregation was immediate. A total of 54 S. uberis isolates were obtained from 24 different cows from the two herds. To overcome operator-dependent analysis of the dot blots and increase the technique's consistency and reliability, the hybridization signals were converted into probability values, with average probabilities higher than 0.5 being considered positive results. These data allowed to confirm the isolates' identity as S. uberis using taxa-specific markers and to determine the presence of virulence- and antibiotic resistance-related genes. In addition, MLSA allowed to disclose the most prevalent S. uberis clonal lineages in both herds. Seven different clusters were identified, with Barcelos showing a high clonal diversity and Maia a dominant lineage infecting most cows, suggesting distinct epidemiological patterns, with S. uberis displaying an environmental or contagious

  6. A comparison of the Genie and western blot assays in confirmatory testing for HIV-1 antibody.

    PubMed

    Chan, E L; Sidaway, F; Horsman, G B

    1996-03-01

    The Genie HIV-1/2 kit (Sanofi Diagnostics Pasteur, Montreal, Quebec), a synthetic-peptide solid-phase enzyme immunoassay, was evaluated as a confirmatory assay for HIV-1 antibodies in comparison with Western blot (BioRad, Hercules, CA, USA) on 50 stored HIV-1 antibody-positive sera and the 137 sera yielding repeated positive results in the conventional EIA screen out of 13405 fresh patient sera from Saskatchewan in 1993. The stored HIV-1-positive sera were uniformly positive in the Genie test. Of the 137 EIA screen-positive sera, 33 were uniformly positive and 64 were uniformly negative in Genie and Western blot; 36 were Genie-negative and indeterminate by Western blot; and four were Genie indeterminate, of which one was negative and three were indeterminate by Western blot. All HIV-1 Western blot-indeterminate and Genie-interdeterminate sera were negative in radio-immunoprecipitation assay (RIPA) and Western blot for HIV-1 and HIV-2 antibodies performed by a reference laboratory. Genie gave an accurate definitive result for 97% of EIA positive sera compared with 71% for Western blot. There was excellent correlation between Genie, Western blot and RIPA results. However, the Genie assay was faster, less costly and yielded fewer indeterminate results than Western blot in confirmatory testing for HIV-1 antibodies.

  7. Indeterminate HIV-1 Western Blots: Etiology, Natural History and Psychological Reactions

    DTIC Science & Technology

    1991-09-24

    viral-like syndrome in the month between the IWB (p24 antibody only) and the positive Western blot. Case 2 was a woman with a history of autoimmune...Dupont blot two weeks later and by Epitope blot four weeks later. Case 3 was a homosexual man with a v!ral-like syndrome who had p2 4 and weak gpl60...proteins on Western blot. Case 4 was a homosexual man with a mononucleosis-like syndrome with a borderline ETA (R-value 0.9) and a faint p24 and weak

  8. Contribution of a Comparative Western Blot Method to Early Postnatal Diagnosis of Congenital Syphilis

    PubMed Central

    Foschi, Claudio; Capretti, Maria Grazia; Nardini, Paola; Compri, Monica; Corvaglia, Luigi Tommaso; Faldella, Giacomo; Cevenini, Roberto

    2016-01-01

    Serology has a pivotal role in the diagnosis of congenital syphilis (CS), but problems arise because of the passive transfer of IgG antibodies across the placenta. The aim of this study was to assess the diagnostic value of a comparative Western blot (WB) method finalized to match the IgG immunological profiles of mothers and their own babies at birth in order to differentiate between passively transmitted maternal antibodies and antibodies synthesized by the infants against Treponema pallidum. Thirty infants born to mothers with unknown or inadequate treatment for syphilis were entered in a retrospective study, conducted at St. Orsola-Malpighi Hospital, Bologna, Italy. All of the infants underwent clinical, instrumental, and laboratory examinations, including IgM WB testing. For the retrospective study, an IgG WB assay was performed by blotting T. pallidum antigens onto nitrocellulose sheets and incubating the strips with serum specimens from mother-child pairs. CS was diagnosed in 11 out of the 30 enrolled infants; 9/11 cases received the definitive diagnosis within the first week of life, whereas the remaining two were diagnosed later because of increasing serological test titers. The use of the comparative IgG WB testing performed with serum samples from mother-child pairs allowed a correct CS diagnosis in 10/11 cases. The CS diagnosis was improved by a strategy combining comparative IgG WB results with IgM WB results, leading to a sensitivity of 100%. The comparative IgG WB test is thus a welcome addition to the conventional laboratory methods used for CS diagnosis, allowing identification and adequate treatment of infected infants and avoiding unnecessary therapy of uninfected newborns. PMID:26961856

  9. Parafilm-M®, An Available Cost-Effective Alternative for Immuno-blot Pouches.

    PubMed

    Quadri, Syed M S

    2015-01-01

    Commercially available standard immuno-blot pouches do play an efficient role in antibody incubation in performing an immuno-blot, but are not readily available in the laboratory and have to be specifically ordered. We have developed an equally efficient technique to make an immune-blot more cost-effective with more conservation of antibodies by using a common and readily available laboratory product Parafilm-M(®). Parafilm-M(®) which serves as a sealant for various items of laboratory equipment can be used for antibody incubation. Manually made Parafilm-M(®) pouch has a clear advantage over standard immuno-blot pouches in terms of availability, cost-effectiveness, and consumption of antibodies that ultimately reduces the cost of an immuno-blot. We have performed a series of experiments to check the efficacy of both the techniques. Samples with equal amount of protein were analyzed on separate SDS PAGE gels. The proteins were transferred electrophoretically to the nitrocellulose membrane using Trans-Blot(®) Turbo™ Mini Nitrocellulose Transfer Pack. Antibody incubation was done using standard immuno-blot pouch, standard container and Parafilm-M(®) sealed pouch. The expression of protein was determined and the results of immuno-blots were compared. We found that antibodies are binding the membrane in Parafilm-M(®) pouches as efficiently as in container method or in standard immuno-blot pouches. By restricting the membrane, the surface area of the manually made Parafilm-M(®) pouch can be reduced, less diluent is required to cover the membrane as a result less antibodies are consumed. We also calculated that each immuno-blot pouch cost around $0.1906, whereas the cost for Parafilm-M(®) pouch is 0.0695 which is almost one-third the price of an immuno-blot pouch. Thus, Parafilm-M(®) method distinctly provides a cost-effective solution for antibody incubation.

  10. A Laboratory Exercise Illustrating the Sensitivity and Specificity of Western Blot Analysis

    ERIC Educational Resources Information Center

    Chang, Ming-Mei; Lovett, Janice

    2011-01-01

    Western blot analysis, commonly known as "Western blotting," is a standard tool in every laboratory where proteins are analyzed. It involves the separation of polypeptides in polyacrylamide gels followed by the electrophoretic transfer of the separated polypeptides onto a nitrocellulose or polyvinylidene fluoride membrane. A replica of the…

  11. Microfluidic integration of Western blotting is enabled by electrotransfer-assisted sodium dodecyl sulfate dilution.

    PubMed

    Hou, Chenlu; Herr, Amy E

    2013-01-07

    We integrate sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) with subsequent antibody probing in a single, monolithic microdevice to realize microfluidic Western blotting. A hurdle to successful on-chip Western blotting lies in restoring antibody recognition of previously sized (denatured, reduced) proteins. To surmount this hurdle, we locally dilute free SDS from SDS-protein complexes using differential electromigration of the species during electrotransfer between SDS-PAGE and blotting regions of a microchamber. Local dilution of SDS minimizes re-association of SDS with proteins offering means to restore antibody binding affinity to proteins after SDS-PAGE. To achieve automated, programmable operation in a single instrument, we utilize a 1 × 2 mm(2) glass microchamber photopatterned with spatially distinct, contiguous polyacrylamide regions for SDS-PAGE, electrotransfer, and antibody blotting. Optimization of both the SDS-PAGE and electrotransfer conditions yields transfer distances of <1 mm (40 s). The Western blot is completed in 180 s, with fully automated assay operation using programmable voltage control. After SDS-PAGE and electrotransfer, we observe ~80% capture of protein band mass on the blotting region for a model protein, C-reactive protein. This novel microfluidic Western blot approach introduces fine transport control for in-transit protein handling to form the basis for an automated, rapid alternative to conventional slab-gel Western blotting.

  12. Detection of antibodies to Hypoderma lineatum in cattle by Western blotting with recombinant hypodermin C antigen.

    PubMed

    Boldbaatar, D; Xuan, X; Kimbita, E; Huang, X; Igarashi, I; Byambaa, B; Battsetseg, B; Battur, B; Battsetseg, G; Batsukh, Z; Nagasawa, H; Fujisaki, K; Mikami, T

    2001-08-01

    The cDNA encoding the entire mature hypodermin C (HC) of Hypoderma lineatum was cloned and expressed in Escherichia coli as a glutathione S-transferase fusion protein using pGEX vector. The recombinant HC protein (rHC) was tested by Western blotting to detect antibodies to H. lineatum in cattle. Western blotting with rHC as antigen clearly differentiated between H. lineatum-infested cattle sera and normal cattle sera. Forty-six out of forty-eight serum samples from cattle in Central Mongolia were positive, whereas all 30 serum samples from cows in Hokkaido, Japan, were negative by Western blotting. The result of Western blotting was identical to that of a previously developed enzyme-linked immunosorbent assay. These data demonstrated that Western blotting, with rHC expressed in E. coli, might be a useful method for the diagnosis of cattle hypodermosis.

  13. GTP-blot analysis of small GTP-binding proteins. The C-terminus is involved in renaturation of blotted proteins.

    PubMed

    Klinz, F J

    1994-10-01

    Recombinant c-Ha-ras, ralA and rap2, but not rap1A or rap1B proteins retained their ability to bind [alpha-32P]GTP after SDS/PAGE and transfer to nitrocellulose. Recombinant c-Has-ras missing the C-terminal 23 amino acid residues failed to bind [alpha-32P]GTP after the blot, and the ability of recombinant ralA missing the C-terminal 28 amino acid residues to bind [alpha-32P]GTP was decreased many-fold. The presence of nonionic detergents of the polyoxyethylene type such as Tween 20, Triton X-100, Nonidet P40 or Lubrol PX in the incubation buffer was necessary to induce renaturation of blotted recombinant c-Ha-ras protein, whereas other types of detergents were ineffective. We propose that detergents of the polyoxyethylene type induce the refolding of some types of blotted small GTP-binding proteins and that the C-terminus is involved in the refolding process. Membranes from NIH3T3 fibroblasts overexpressing c-Ha-ras protein showed much weaker binding of [alpha-32P]GTP as expected from the level of ras immunoreactivity. Treatment of fibroblasts with lovastatin, an inhibitor of hydroxymethylglutaryl-coenzyme A reductase, caused the accumulation of the unfarnesylated form of c-Ha-ras in the cytosol. Examination of [alpha-32P]GTP-binding and immunoreactivity for cytosolic and membrane-bound c-Ha-ras revealed that binding of [alpha-32P]GTP to unprocessed c-Ha-ras was increased about threefold compared to the same amount of processed c-Ha-ras. Our results demonstrate that detection and quantification of small GTP-binding proteins in eukaryotic cells by GTP-blot analysis is hampered by the fact that these proteins differ strongly in their ability to renature after blotting to nitrocellulose.

  14. Western blotting of formaldehyde-fixed neuropeptides as small as 400 daltons on gelatin-coated nitrocellulose paper.

    PubMed

    Too, C K; Murphy, P R; Croll, R P

    1994-06-01

    A method is described for Western blotting of peptides as small as 400 daltons (Da). Peptides were separated by tricine-sodium dodecyl sulfate electrophoresis and electroblotted to gelatin-coated PH79 nitrocellulose paper (0.1 micron). The electroblotted peptides were fixed to the nitrocellulose paper for 5-10 min in 4% paraformaldehyde solution. Using anti-rabbit FMRF-amide (Phe-Met-Arg-Phe-NH2) as primary antibody, positive immunoreactivity was detected with an amplified alkaline phosphatase assay which was sensitive to at least 0.5 microgram FMRFamide/lane. When immunoreactivity was determined with 125I-protein A, it was possible to amplify and detect weak signals by increasing the autoradiography time. Therefore, using the 125I-protein A detection method, Western blot analysis of brain extracts from Lymnaea stagnalis (pond snail) and Poecilia reticulata (guppy) indicated the presence of four FMRFamide immunoreactive bands after a 7-day exposure to X-ray film. The most abundant peptide coelectrophoresed with the FMRFamide standard (M(r) 598.8 Da). In addition, this Western blotting procedure also detected APGWamide (Ala-Pro-Gly-Try-NH2; 428.5 Da) and [D-Ala2]-Leu-enkephalinamide (568.7 Da) with their respective specific antibodies.

  15. Application of SYPRO Ruby- and Flamingo-stained polyacrylamide gels to Western blot analysis.

    PubMed

    Hagiwara, Makoto; Kobayashi, Ken-Ichi; Tadokoro, Tadahiro; Yamamoto, Yuji

    2010-02-15

    Western blots are widely used for analysis of the expression levels of specific proteins. Blotting is conducted after sodium dodecyl sulfate or native polyacrylamide gel electrophoresis without staining the gel. However, when it is necessary to analyze the gel, duplicate polyacrylamide gels (one of which is stained) usually must be prepared, leading to the consumption of precious sample. Thus, we have developed a convenient and efficient Western blot method using a stained gel. This simple modification should be beneficial for the analysis of samples that are limited in quantity and/or samples for which the stained gel serves as the loading control.

  16. Detection and quantitation of HPV DNA replication by Southern blotting and real-time PCR.

    PubMed

    Morgan, Iain M; Taylor, Ewan R

    2005-01-01

    This provides a brief introduction into the mechanism of DNA replication by the E1 and E2 proteins and describes the traditional Southern blotting technique that is used to monitor E1- and E2-mediated DNA replication. It also includes a novel real-time polymerase chain reaction (PCR) approach for monitoring E1- and E2-mediated DNA replication that has enhanced sensitivity and quantitation compared with Southern blotting, and a discussion of when to use the Southern blotting and real-time PCR techniques.

  17. High-selectivity cytology via lab-on-a-disc western blotting of individual cells.

    PubMed

    Kim, John J; Sinkala, Elly; Herr, Amy E

    2017-02-28

    Cytology of sparingly available cell samples from both clinical and experimental settings would benefit from high-selectivity protein tools. To minimize cell handling losses in sparse samples, we design a multi-stage assay using a lab-on-a-disc that integrates cell handling and subsequent single-cell western blotting (scWestern). As the two-layer microfluidic device rotates, the induced centrifugal force directs dissociated cells to dams, which in turn localize the cells over microwells. Cells then sediment into the microwells, where the cells are lysed and subjected to scWestern. Taking into account cell losses from loading, centrifugation, and lysis-buffer exchange, our lab-on-a-disc device handles cell samples with as few as 200 cells with 75% cell settling efficiencies. Over 70% of microwells contain single cells after the centrifugation. In addition to cell settling efficiency, cell-size filtration from a mixed population of two cell lines is also realized by tuning the cell time-of-flight during centrifugation (58.4% settling efficiency with 6.4% impurity). Following the upstream cell handling, scWestern analysis detects four proteins (GFP, β-TUB, GAPDH, and STAT3) in a glioblastoma cell line. By integrating the lab-on-a-disc cell preparation and scWestern analysis, our platform measures proteins from sparse cell samples at single-cell resolution.

  18. Evaluation of an enzyme-linked immunoelectrotransfer blot test for the confirmatory serodiagnosis of human toxocariasis.

    PubMed

    Roldán, William H; Espinoza, Yrma A

    2009-05-01

    To improve the serodiagnosis of human toxocariasis, a sensitive and specific enzyme-linked immunoelectrotransfer blot (EITB-IgG) test was developed and evaluated using Toxocara canislarvae excretory-secretory antigens for detecting anti-Toxocara IgG antibodies. The EITB-IgG profile of toxocariasis was characterized by comparing 27 sera from patients with toxocariasis, 110 sera from healthy subjects and 186 sera from patients with other helminth diseases (ascariasis, ancylostomiasis, trichuriasis, enterobiasis, strongyloidiasis, hymenolepiasis, diphyllobothriasis, taeniasis, cysticercosis, hydatidosis and fascioliasis). Antigenic bands of 24, 28, 30, 35, 56, 117, 136 and 152 kDa were predominantly recognized in sera from all patients with toxocariasis. However, only bands of 24-35 kDa were highly specific for Toxocara infection (98.3%), whereas other antigenic bands observed displayed cross-reactivity. Additionally, when the results of the EITB-IgG test were compared to those of the ELISA-IgG test, a 100% concordance was observed for positive results in human toxocariasis cases. The concordance for negative results between the two tests for healthy subjects and patients with other helminth diseases were 96.3% and 53.7%, respectively, showing that the EITB-IgG test has a higher specificity than ELISA. In conclusion, the EITB-IgG test is a very useful tool to confirm the serological diagnosis of human toxocariasis.

  19. Multiplexed microfluidic blotting of proteins and nucleic acids by parallel, serpentine microchannels.

    PubMed

    He, Sha; Zhang, Yi; Wang, Pei; Xu, Xingzhi; Zhu, Kui; Pan, Wenying; Liu, Wenwen; Cai, Kaiyong; Sun, Jiashu; Zhang, Wei; Jiang, Xingyu

    2015-01-07

    This work develops a high-throughput, high-efficiency and straightforward microfluidic blotting method for analyzing proteins and nucleic acids. Sample solutions containing antibodies (for protein detection) or hybridization probes (for nucleic acid detection) are introduced into the parallel, serpentine microchannels to specifically recognize the immobilized targets on the substrate, achieving the identification of multiple targets in multiple samples simultaneously. The loading control, molecular weight markers, and antigen/antibody titration are designed and integrated into the microfluidic chip, thus allowing for the quantification of proteins and nucleic acids. Importantly, we could easily distinguish the adjacent blotting bands inside parallel microchannels, which may be difficult to achieve in conventional blotting. The small dimensions of microfluidic channels also help to reduce the amount of probing molecules and to accelerate the biochemical reaction. Our microfluidic blotting could bypass the steps of blocking and washing, further reducing the operation time and complexity.

  20. Automated capillary Western dot blot method for the identity of a 15-valent pneumococcal conjugate vaccine.

    PubMed

    Hamm, Melissa; Ha, Sha; Rustandi, Richard R

    2015-06-01

    Simple Western is a new technology that allows for the separation, blotting, and detection of proteins similar to a traditional Western except in a capillary format. Traditionally, identity assays for biological products are performed using either an enzyme-linked immunosorbent assay (ELISA) or a manual dot blot Western. Both techniques are usually very tedious, labor-intensive, and complicated for multivalent vaccines, and they can be difficult to transfer to other laboratories. An advantage this capillary Western technique has over the traditional manual dot blot Western method is the speed and the automation of electrophoresis separation, blotting, and detection steps performed in 96 capillaries. This article describes details of the development of an automated identity assay for a 15-valent pneumococcal conjugate vaccine, PCV15-CRM197, using capillary Western technology.

  1. Detection of DNA sequence polymorphisms in human genomic DNA by using denaturing gradient gel blots

    SciTech Connect

    Gray, M.R. )

    1992-02-01

    Denaturing gradient gel electrophoresis can detect sequence differences outside restriction-enzyme recognition sites. DNA sequence polymorphisms can be detected as restriction-fragment melting polymorphisms (RFMPs) in genomic DNA by using blots made from denaturing gradient gels. In contrast to the use of Southern blots to find sequence differences, denaturing gradient gel blots can detect differences almost anywhere, not just at 4-6-bp restriction-enzyme recognition sites. Human genomic DNA was digested with one of several randomly selected 4-bp recognition-site restriction enzymes, electrophoresed in denaturing gradient gels, and transferred to nylon membranes. The blots were hydridized with radioactive probes prepared from the factor VIII, type II collagen, insulin receptor, [beta][sub 2]-adrenergic receptor, and 21-hydroxylase genes; in unrelated individuals, several RFM's were found in fragments from every locus tested. No restriction map or sequence information was used to detect RFMP's.

  2. Identification of immunodiagnostic antigens for cerebrospinal filariasis in horses by western blot analysis

    PubMed Central

    TAKESUE, Masataka; OSAKA, Yuki; MURANAKA, Masanori; KATAYAMA, Yoshinari; IKADAI, Hiromi

    2016-01-01

    ABSTRACT In the present study, the serum and cerebrospinal fluid of horses diagnosed with Setaria digitata cerebrospinal filariasis were analyzed by western blot. The results revealed S. digitata protein bands measuring 65, 34, 22, and 18 kDa in molecular weight. In particular, the 18 kDa band is a possible candidate for clinical immunodiagnosis on the basis of western blot findings. PMID:27073332

  3. Accuracy of Reverse Dot-Blot PCR in Detection of Different β-Globin Gene Mutations.

    PubMed

    El-Fadaly, N; Abd-Elhameed, A; Abd-Elbar, E; El-Shanshory, M

    2016-06-01

    Prevention programs for β-thalassemia based on molecular diagnosis of heterozygous carriers and/or patients require the use of reliable mutation screening methods. The aim of this study was to compare between direct DNA sequencing, and reverse dot-blot PCR in detection of different β-globin gene mutations in Egyptian children with β-thalassemia. Forty children with β-thalassemia were subjected to mutation analysis, performed by both direct DNA sequencing and β-globin Strip Assay MED™ (based on reverse dot-blot PCR). The most frequent mutant alleles detected by reverse dot-blot PCR were; IVSI-110 G>A (31.25 %), IVS I-6 T > C (21.25 %), and IVS I-1 G>A (20 %). Relatively less frequent mutant alleles detected by reverse dot-blot PCR were "IVSII-1 G>A (5 %), IVSII-745 C>G (5 %), IVSII-848 C>A (2.5 %), IVSI-5 G>C (2.5 %), -87 C>G(2.5 %), and cd39 C>T (2.5 %)", While the genotypes of three patients (6 alleles 7.5 %) were not detected by reverse dot-blot PCR. Mutant alleles detected by direct DNA sequencing were the same as reverse dot-blot PCR method except it revealed the genotypes of 3 undetected patients (one patient was homozygous IVSI-110 G>A, and two patients were homozygous IVS I-1 G>A. Sensitivity of the reverse dot-blot PCR was 92.5 % when compared to direct DNA sequencing for detecting β-thalassemia mutations. Our results therefore suggest that, direct DNA sequencing may be preferred over reverse dot-blot PCR in critical diagnostic situations like genetic counseling for prenatal diagnosis.

  4. Using a large area CMOS APS for direct chemiluminescence detection in Western blotting electrophoresis

    NASA Astrophysics Data System (ADS)

    Esposito, Michela; Newcombe, Jane; Anaxagoras, Thalis; Allinson, Nigel M.; Wells, Kevin

    2012-03-01

    Western blotting electrophoretic sequencing is an analytical technique widely used in Functional Proteomics to detect, recognize and quantify specific labelled proteins in biological samples. A commonly used label for western blotting is Enhanced ChemiLuminescence (ECL) reagents based on fluorescent light emission of Luminol at 425nm. Film emulsion is the conventional detection medium, but is characterized by non-linear response and limited dynamic range. Several western blotting digital imaging systems have being developed, mainly based on the use of cooled Charge Coupled Devices (CCDs) and single avalanche diodes that address these issues. Even so these systems present key drawbacks, such as a low frame rate and require operation at low temperature. Direct optical detection using Complementary Metal Oxide Semiconductor (CMOS) Active Pixel Sensors (APS)could represent a suitable digital alternative for this application. In this paper the authors demonstrate the viability of direct chemiluminescent light detection in western blotting electrophoresis using a CMOS APS at room temperature. Furthermore, in recent years, improvements in fabrication techniques have made available reliable processes for very large imagers, which can be now scaled up to wafer size, allowing direct contact imaging of full size western blotting samples. We propose using a novel wafer scale APS (12.8 cm×13.2 cm), with an array architecture using two different pixel geometries that can deliver an inherently low noise and high dynamic range image at the same time representing a dramatic improvement with respect to the current western blotting imaging systems.

  5. Immunoconfirmation of central nervous system tuberculosis by blotting: A study of 300 cases.

    PubMed

    Patil, Shripad; Giribhattanavar, Prashant; Patil, Madhusudan; Kumar, Kavitha

    2015-06-01

    Tuberculous meningitis (TBM) is a serious form of disease of the central nervous system. Early and accurate diagnosis of the disease and effective treatment are key important factors to contain the disease. The disease presents as chronic meningitis where other partners such as fungal meningitis, neurosyphilis, cysticercal meningitis, carcinomatous meningitis and partially treated pyogenic meningitis share a similar clinical picture making the diagnosis complicated. Culturing of the pathogen Mycobacterium tuberculosis (MTB) from the cerebrospinal fluid (CSF) sample has shown a poor response. The main immunological method for the immunodiagnosis of TBM is the detection of an antibody response in the CSF. In the present study, total MTB sonicated extract antigen was used for ELISA and Western blot. ELISA shows overall immune response of the test sample, whereas Western blotting reveals the specific reactivity to a particular molecular weight antigen. This would also reveal the immunodominant antigen. A total of 300 CSF samples were analyzed by both ELISA and Western blotting. Of the 240 clinically suspected TBM cases, 111 samples were positive by ELISA and 81 samples by Western blot. A total of 76 CSF samples were positive by both ELISA and Western blot. None of the control samples showed positivity either by ELISA or by Western blot. TBM patients revealed major antibody reactivity to 30-40 kD region, followed by 14 kD region. ELISA is sensitive with mild non-specific binding, but Western blot is specific in detecting the immune response. The findings will be useful in definitive immunodiagnosis of TBM.

  6. Blot-based detection of dehydroalanine-containing glutathione peroxidase with the use of biotin-conjugated cysteamine.

    PubMed

    Rhee, Sue Goo; Cho, Chun-Seok

    2010-01-01

    Dehydroalanine (DHA), alpha,beta-unsaturated amino acid, is found in the position corresponding to the serine, cysteine, and selenocysteine (Sec) residues of various proteins. Proteinaceous Sec is readily oxidized and subsequently undergoes beta-elimination to produce DHA. Glutathione peroxidase (GPx), which contains a Sec at the active site, is irreversibly inactivated by its own substrate as the result of the oxidation of selenium atom followed by the conversion of oxidized Sec to DHA. We developed a convenient method for estimation of the amount of DHA-GPx1 in cell homogenates. This blot-based method depends on specific addition of biotin-conjugated cysteamine to the DHA residue followed by detection of biotinylated protein based on its interaction with streptavidin. The method required an immunoprecipitation of GPx1 before labeling with the cysteamine derivative because many other proteins contain DHA. With the use of this method, we found that conversion of the Sec residue at the active site of GPx1 to DHA occurred during aging of red blood cells (RBCs) in vivo as well as in RBCs exposed to H(2)O(2) generated either externally by glucose oxidase or internally as a result of aniline-induced Hb autoxidation. Accordingly, the content of DHA-GPx1 in each RBC likely reflects total oxidative stress experienced by the cell during its lifetime of 120 days. Previous studies suggested that the activity of GPx1 in RBCs is most influenced by lifestyle and environmental factors such as the use of dietary supplements and smoking habit. Therefore, DHA-GPx1 in RBCs might be a suitable surrogate marker for evaluation of oxidative stress in the body. Our blot-based method for the detection of DHA-GPx1 will be very useful for evaluation of such stress. In addition, similar blot detection method can be devised for other proteins for which immunoprecipitating antibodies are available.

  7. Two-Phase Contiguous Supported Lipid Bilayer Model for Membrane Rafts via Polymer Blotting and Stenciling.

    PubMed

    Richards, Mark J; Daniel, Susan

    2017-02-07

    The supported lipid bilayer has been portrayed as a useful model of the cell membrane compatible with many biophysical tools and techniques that demonstrate its appeal in learning about the basic features of the plasma membrane. However, some of its potential has yet to be realized, particularly in the area of bilayer patterning and phase/composition heterogeneity. In this work, we generate contiguous bilayer patterns as a model system that captures the general features of membrane domains and lipid rafts. Micropatterned polymer templates of two types are investigated for generating patterned bilayer formation: polymer blotting and polymer lift-off stenciling. While these approaches have been used previously to create bilayer arrays by corralling bilayers patches with various types of boundaries impenetrable to bilayer diffusion, unique to the methods presented here, there are no physical barriers to diffusion. In this work, interfaces between contiguous lipid phases define the pattern shapes, with continuity between them allowing transfer of membrane-bound biomolecules between the phases. We examine effectors of membrane domain stability including temperature and cholesterol content to investigate domain dynamics. Contiguous patterning of supported bilayers as a model of lipid rafts expands the application of the SLB to an area with current appeal and brings with it a useful toolset for characterization and analysis. These combined tools should be helpful to researchers investigating lipid raft dynamics and function and biomolecule partitioning studies. Additionally, this patterning technique may be useful for applications such as bioseparations that exploit differences in lipid phase partitioning or creation of membranes that bind species like viruses preferentially at lipid phase boundaries, to name a few.

  8. HTLV-I/II seroindeterminate Western blot reactivity in a cohort of patients with neurological disease.

    PubMed

    Soldan, S S; Graf, M D; Waziri, A; Flerlage, A N; Robinson, S M; Kawanishi, T; Leist, T P; Lehky, T J; Levin, M C; Jacobson, S

    1999-09-01

    The human T-cell lymphotropic virus type I (HTLV-I) is associated with a chronic, progressive neurological disease known as HTLV-I-associated myelopathy/tropical spastic paraparesis. Screening for HTLV-I involves the detection of virus-specific serum antibodies by EIA and confirmation by Western blot. HTLV-I/II seroindeterminate Western blot patterns have been described worldwide. However, the significance of this blot pattern is unclear. We identified 8 patients with neurological disease and an HTLV-I/II seroindeterminate Western blot pattern, none of whom demonstrated increased spontaneous proliferation and HTLV-I-specific cytotoxic T lymphocyte activity. However, HTLV-I tax sequence was amplified from the peripheral blood lymphocytes of 4 of them. These data suggest that patients with chronic progressive neurological disease and HTLV-I/II Western blot seroindeterminate reactivity may harbor either defective HTLV-I, novel retrovirus with partial homology to HTLV-I, or HTLV-I in low copy number.

  9. A guide to modern quantitative fluorescent western blotting with troubleshooting strategies.

    PubMed

    Eaton, Samantha L; Hurtado, Maica Llavero; Oldknow, Karla J; Graham, Laura C; Marchant, Thomas W; Gillingwater, Thomas H; Farquharson, Colin; Wishart, Thomas M

    2014-11-20

    The late 1970s saw the first publicly reported use of the western blot, a technique for assessing the presence and relative abundance of specific proteins within complex biological samples. Since then, western blotting methodology has become a common component of the molecular biologists experimental repertoire. A cursory search of PubMed using the term "western blot" suggests that in excess of two hundred and twenty thousand published manuscripts have made use of this technique by the year 2014. Importantly, the last ten years have seen technical imaging advances coupled with the development of sensitive fluorescent labels which have improved sensitivity and yielded even greater ranges of linear detection. The result is a now truly Quantifiable Fluorescence based Western Blot (QFWB) that allows biologists to carry out comparative expression analysis with greater sensitivity and accuracy than ever before. Many "optimized" western blotting methodologies exist and are utilized in different laboratories. These often prove difficult to implement due to the requirement of subtle but undocumented procedural amendments. This protocol provides a comprehensive description of an established and robust QFWB method, complete with troubleshooting strategies.

  10. Northern Nutrition.

    ERIC Educational Resources Information Center

    Northwest Territories Dept. of Education, Yellowknife.

    This guide contains nutrition information and nutrition education strategies aimed at residents of the Canadian Arctic. Section I: (1) defines nutrition terms; (2) describes the sources and functions of essential nutrients; (3) explains Canada's food guide and special considerations for the traditional northern Native diet and for lactose…

  11. Western blotting using in-gel protein labeling as a normalization control: stain-free technology.

    PubMed

    Gilda, Jennifer E; Gomes, Aldrin V

    2015-01-01

    Western blotting is a commonly used laboratory technique for semi-quantifying protein amounts. It is important when quantifying protein expression to account for differences in the amount of total protein loaded onto the gel using a loading control. Common loading controls include housekeeping proteins, such as β-actin or GAPDH, quantified by Western blot, or total protein, quantified using a stain such as Coomassie Brilliant Blue or Ponceau S. A more recently developed method for total protein quantification utilizes stain-free technology, which has a linear dynamic detection range and allows for protein detection on both gels and membranes. Here, we describe the theory and use of stain-free gels for total protein quantification and normalization of Western blots.

  12. Application of SYPRO Ruby- and Flamingo-stained polyacrylamide gels to Western blot analysis.

    PubMed

    Hagiwara, Makoto; Kobayashi, Ken-Ichi; Tadokoro, Tadahiro; Yamamoto, Yuji

    2009-06-15

    Western blot analysis has been a useful method for analysis of expression levels of specific proteins and is conducted after sodium dodecyl sulfate (SDS) or native polyacrylamide gel electrophoresis without staining the gel. However, when it is necessary to analyze the gel, duplicate polyacrylamide gels usually must be prepared, one of which is stained, leading to the consumption of precious sample. Thus, we developed a convenient and efficient Western blotting method using a stained gel. This simple modification should be beneficial for analyzing samples that are limited in quantity and/or samples for which the stained gel serves as the loading control.

  13. Detection of Sleeping Beauty transposition in the genome of host cells by non-radioactive Southern blot analysis.

    PubMed

    Aravalli, Rajagopal N; Park, Chang W; Steer, Clifford J

    2016-08-26

    The Sleeping Beauty transposon (SB-Tn) system is being used widely as a DNA vector for the delivery of therapeutic transgenes, as well as a tool for the insertional mutagenesis in animal models. In order to accurately assess the insertional potential and properties related to the integration of SB it is essential to determine the copy number of SB-Tn in the host genome. Recently developed SB100X transposase has demonstrated an integration rate that was much higher than the original SB10 and that of other versions of hyperactive SB transposases, such as HSB3 or HSB17. In this study, we have constructed a series of SB vectors carrying either a DsRed or a human β-globin transgene that was encompassed by cHS4 insulator elements, and containing the SB100X transposase gene outside the SB-Tn unit within the same vector in cis configuration. These SB-Tn constructs were introduced into the K-562 erythroid cell line, and their presence in the genomes of host cells was analyzed by Southern blot analysis using non-radioactive probes. Many copies of SB-Tn insertions were detected in host cells regardless of transgene sequences or the presence of cHS4 insulator elements. Interestingly, the size difference of 2.4 kb between insulated SB and non-insulated controls did not reflect the proportional difference in copy numbers of inserted SB-Tns. We then attempted methylation-sensitive Southern blots to assess the potential influence of cHS4 insulator elements on the epigenetic modification of SB-Tn. Our results indicated that SB100X was able to integrate at multiple sites with the number of SB-Tn copies larger than 6 kb in size. In addition, the non-radioactive Southern blot protocols developed here will be useful to detect integrated SB-Tn copies in any mammalian cell type.

  14. Western Blot Analysis of the Exotoxin Components From Bacillus anthracis Separated by Isoelectric Focusing Gel Electrophoresis

    DTIC Science & Technology

    2004-02-14

    Biochemical and Biophysical Research Communications 317 (2004) 294–300 295 Approved for...Ames isolate was originally identified as Iowa, it has subsequently been identified as Texas. 296 S.F. Little / Biochemical and Biophysical Research Communications 317... Biochemical and Biophysical Research Communications 317 (2004) 294–300 297 Approved for public release. Distribution is unlimited.Western blots

  15. Ferulic acid enhances IgE binding to peanut allergens in western blots.

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Phenolic compounds at high concentrations are known to form insoluble complexes with proteins. We hypothesized that this complex formation could interfere with Western blot and ELISA assays for peanut allergens. To verify this, three simple phenolic compounds (ferulic, caffeic, and chlorogenic acids...

  16. Ferulic acid enhances IgE binding to peanut allergens in western blots.

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Because phenolic compounds can precipitate or complex with proteins, we postulated that interactions of phenolics with IgE antibodies help enhance IgE binding to peanut allergens in Western blots. Three different phenolics, such as, ferulic, caffeic and chlorogenic acids were examined. Each was mixe...

  17. Quantitative Immunofluorescent Blotting of the Multidrug Resistance-associated Protein 2 (MRP2)

    PubMed Central

    Gerk, Phillip M.

    2011-01-01

    Introduction Quantitation of the expression levels of proteins involved in drug transport and disposition is needed to overcome limitations of film-based detection of chemiluminescent immunoblots. Purpose The purpose was to describe and validate a quantitative immunofluorescent blotting method for detection of ATP-Binding Cassette Transporter Isoform C2/Multidrug Resistance-associated Protein 2 (ABCC2/MRP2). Methods Western blotting was performed by electrophoresis of membrane vesicle protein isolated from Sf9 cells overexpressing MRP2 subsequently blotting with infrared labeled secondary antibody. The bound complex was detected using the Odyssey Infrared Imaging System (Li-Cor; Lincoln, NE). The images were analyzed using the Odyssey Application Software to obtain the integrated intensities, followed by linear regression of the intensity data. Results The limits of quantitation for the time-insensitive technique described here were from 0.001μg to 0.5μg of total membrane protein, the coefficient of variation of the slope was 8.9%; r2 values were 0.986 ± 0.012. The utility and sensitivity of this technique was demonstrated in quantitating expression of MRP2 in human placental tissue samples, in which MRP2 was present in low abundance. Discussion The immunofluorescent blotting technique described provides sensitive, reproducible, and quantitative determinations of large, integral membrane proteins such as MRP2, all with potential long-term cost savings. PMID:21277982

  18. A Study of Rubisco through Western Blotting and Tissue Printing Techniques

    ERIC Educational Resources Information Center

    Ma, Zhong; Cooper, Cynthia; Kim, Hyun-Joo; Janick-Buckner, Diane

    2009-01-01

    We describe a laboratory exercise developed for a cell biology course for second-year undergraduate biology majors. It was designed to introduce undergraduates to the basic molecular biology techniques of Western blotting and immunodetection coupled with the technique of tissue printing in detecting the presence, relative abundance, and…

  19. A Streamlined Western Blot Exercise: An Efficient and Greener Approach in the Laboratory Classroom

    ERIC Educational Resources Information Center

    Ness, Traci L.; Robinson, Rebekah L.; Mojadedi, Wais; Peavy, Lydia; Weiland, Mitch H.

    2015-01-01

    SDS-PAGE and western blotting are two commonly taught protein detection techniques in biochemistry and molecular biology laboratory classrooms. A pitfall associated with incorporating these techniques into the laboratory is the significant wait times that do not allow students to obtain timely results. The waiting associated with SDS-PAGE comes…

  20. COMPARISONS OF ELISA AND WESTERN BLOT ASSAYS FOR DETECTION OF CRYPTOSPORIDIUM ANTIBODY

    EPA Science Inventory

    A seroprevalence survey was conducted using ELISA and Western blot (WB) assays for antibody to three Cryptosporidium antigens on 380 blood donors in Jackson County, Oregon. The purpose was to determine if either assay could detect serological evidence of an outbreak which occurre...

  1. A Guide to Modern Quantitative Fluorescent Western Blotting with Troubleshooting Strategies

    PubMed Central

    Eaton, Samantha L.; Hurtado, Maica Llavero; Oldknow, Karla J.; Graham, Laura C.; Marchant, Thomas W.; Gillingwater, Thomas H.; Farquharson, Colin; Wishart, Thomas M.

    2014-01-01

    The late 1970s saw the first publicly reported use of the western blot, a technique for assessing the presence and relative abundance of specific proteins within complex biological samples. Since then, western blotting methodology has become a common component of the molecular biologists experimental repertoire. A cursory search of PubMed using the term “western blot” suggests that in excess of two hundred and twenty thousand published manuscripts have made use of this technique by the year 2014. Importantly, the last ten years have seen technical imaging advances coupled with the development of sensitive fluorescent labels which have improved sensitivity and yielded even greater ranges of linear detection. The result is a now truly Quantifiable Fluorescence based Western Blot (QFWB) that allows biologists to carry out comparative expression analysis with greater sensitivity and accuracy than ever before. Many “optimized” western blotting methodologies exist and are utilized in different laboratories. These often prove difficult to implement due to the requirement of subtle but undocumented procedural amendments. This protocol provides a comprehensive description of an established and robust QFWB method, complete with troubleshooting strategies. PMID:25490604

  2. Quantum dot bio-conjugate: as a western blot probe for highly sensitive detection of cellular proteins

    NASA Astrophysics Data System (ADS)

    Kale, Sonia; Kale, Anup; Gholap, Haribhau; Rana, Abhimanyu; Desai, Rama; Banpurkar, Arun; Ogale, Satishchandra; Shastry, Padma

    2012-03-01

    In the present study, we report a quantum dot (QD)-tailored western blot analysis for a sensitive, rapid and flexible detection of the nuclear and cytoplasmic proteins. Highly luminescent CdTe and (CdTe)ZnS QDs are synthesized by aqueous method. High resolution transmission electron microscopy, Raman spectroscopy, fourier transform infrared spectroscopy, fluorescence spectroscopy and X-ray diffraction are used to characterize the properties of the quantum dots. The QDs are functionalized with antibodies of prostate apoptosis response-4 (Par-4), poly(ADP-ribose) polymerases and β actin to specifically bind with the proteins localized in the nucleus and cytoplasm of the cells, respectively. The QD-conjugated antibodies are used to overcome the limitations of conventional western blot technique. The sensitivity and rapidity of protein detection in QD-based approach is very high, with detection limits up to 10 pg of protein. In addition, these labels provide the capability of enhanced identification and localization of marker proteins in intact cells by confocal laser scanning microscopy.

  3. A solid-phase immunoassay of protease-resistant prion protein with filtration blotting involving sodium dodecyl sulfate.

    PubMed

    Kobayashi, Yoshiteru; Kohno, Naoyuki; Wanibe, Shoko; Hirayasu, Kazunari; Uemori, Hitoshi; Tagawa, Yuichi; Yokoyama, Takashi; Shinagawa, Morikazu

    2006-02-15

    The precise diagnosis for bovine spongiform encephalopathy (BSE) is crucial for preventing new transmission to humans. Several testing procedures are reported for determining protease-resistant prion protein in various tissues as a major hallmark of prion diseases such as BSE, scrapie, and Creutzfeldt-Jakob disease. However, contamination of materials from tissues or degradation of the specimens sometimes disturbs the accuracy of the assay. Here, we have developed a novel method for solid-phase immunoassay of the disease-specific conformational isoform, PrP(Sc), using filtration blotting of protein in the presence of sodium dodecyl sulfate (SDS) followed by a filtration-based immunoassay with a single anti-prion protein antibody, together with the improved fractionation procedure involving high concentrations of surfactant/detergent. The SDS/heat treatment renders unfolded PrP(Sc) quantitative retention on a polyvinylidene difluoride filter and allows enhancement of the analyte signal with immunodetection; thus, all of the tested specimens are determined with 100% accuracy. In addition, the immunoassay is completed in approximately 1h, indicating its usefulness not only for the screening of BSE specimens but probably also for the postmortem BSE diagnosis of fallen stock as the antibody recognizes the core part of PrP(Sc). The solid-phase immunoassay method, including the filtration blotting with SDS, would be applicable to determining even more sensitively proteins other than PrP(Sc), especially those having rigid conformations.

  4. Laboratory diagnosis of hymenoptera venom allergy: comparative study between specific IgE, western blot and allergen leukocyte stimulation (CAST).

    PubMed

    Santos, M C Pereira; Carlos, M L Palma; Pedro, E; Carlos, A G Palma

    2002-01-01

    Allergy to hymenoptera venom is a classical IgE mediated disease with a potentially fatal course. Specific venom immunotherapy (SIT) is the most effective mean of treating this serious condition, after the diagnosis has been clearly established by a clinical history, in-vivo and in vitro tests. We have evaluated the usefulness of a cellular test (CAST) which is a recently developed ELISA method based on the evaluation of sulfidoleukotriene secretion by leukocytes stimulated with specific antigen. We also evaluated the correlation of CAST with skin tests, specific IgE (sIgE) and western blot for sIgE for hymenoptera venom sIgE. We have included in this study 14 patients, with a clinical history suggestive of hymenoptera venom allergy. None of them had previously been subjected to immunotherapy. A good correlation was obtained between skin tests, sIgE and western blot. However, there was no correlation between these methods and CAST. We conclude that the positivity of CAST method raises some questions about other mechanisms, which maybe non-IgE dependent. Although the number of patients in this study is quite small, the immunoblot analysis may be a valuable additional method in insect venom allergy.

  5. Biofilm detection by wound blotting can predict slough development in pressure ulcers: A prospective observational study.

    PubMed

    Nakagami, Gojiro; Schultz, Gregory; Gibson, Daniel J; Phillips, Priscilla; Kitamura, Aya; Minematsu, Takeo; Miyagaki, Tomomitsu; Hayashi, Akitatsu; Sasaki, Sanae; Sugama, Junko; Sanada, Hiromi

    2016-12-26

    Bacteria have been found to form multicellular aggregates which have collectively been termed "biofilms." It is hypothesized that biofilm formation is a means to protect bacterial cells including protection form the immune response of humans. This protective mechanism is believed to explain persistent chronic wound infections. At times, the biofilms are abundant enough to see, and remove by simple wiping. However, recent evidence has shown that the removal of these visible portions are not sufficient, and that biofilms can continue to form even with daily wiping. In this work, we tested an approach to detect the biofilms which are present after clinically wiping or sharp wound debridement. Our method is based on a variation of impression cytology in which a nitrocellulose membrane was used to collect surface biofilm components, which were then differentially stained. In this prospective study, members of an interdisciplinary pressure ulcer team at a university hospital tested our method's ability to predict the generation of wound slough in the week that followed each blotting. A total of 70 blots collected from 23 pressure ulcers produced 27 wounds negative for staining and 43 positive. In the negative blots 55.6% were found to have decreased wound slough, while 81.4% with positive staining had either increase or unchanged wound slough generation. These results lead to an odds ratio of positive blotting cases of 9.37 (95% confidence intervals: 2.47-35.5, p = 0.001) for slough formation; suggesting that the changes in wound slough formation can be predicted clinically using a non-invasive wound blotting method.

  6. Detection of human cytomegalovirus by slot-blot hybridization assay employing oligo-primed /sup 32/P-labelled probe

    SciTech Connect

    Agha, S.A.; Coleman, J.C.; Selwyn, S.; Mahmound, L.A.; Abd-Elaal, A.M.; Archard, L.C.

    1988-12-01

    A /sup 32/P-labelled Hind III-0 DNA fragment (nine Kilobases; Kb) from human cytomegalovirus AD-169 (HCMV) was used in slot-blot hybridization assay for the detection of HCMV in clinical samples. The results obtained with DNA hybridization assay (DNA HA) were compared with virus isolation using conventional tube cell culture (CTC) and centrifugation vial culture (CVC), immunofluorescence (IF), and complement fixation test (CFT). Of 15 CTC-positive samples, 13 were positive with DNA HA (sensitivity 86.7%). Also, 14 additional samples were DNA HA-positive but CTC-negative. CVC and/or IF confirmed the diagnosis in nine of 14; the remaining five samples were from three patients who showed fourfold rising antibody titre by CFT. Although DNA HA using /sup 32/P-labelled probes is relatively cumbersome and expensive, it is a valuable test for quantitation of viral shedding in patients with HCMV infections who may benefit from antiviral therapy.

  7. Western blot membrane composed of electrospun polyvinylidene fluoride nanofiber membrane and polyethylene terephthalate sheet.

    PubMed

    Cho, Eugene; Kim, Chan; Hwang, Cheol Ho; Chang, Duck Rye; Kook, Joong-Ki

    2013-06-01

    In a previous study, an electrospun polyvinylidene fluoride (PVDF) nanofiber membrane was developed for Western blotting. The membrane exhibited high sensitivity and high binding capacity for the detection of protein bands that was unlike that observed for conventional, microphase separation-based porous PVDF membranes. Nevertheless, the PVDF nanofiber membrane is quite expensive. The objective of this study was to develop an economical Western blot membrane using a hybrid electrospun PVDF nanofiber and polyethylene terephthalate (PET) sheet. The results showed that the detection sensitivity of the 4 gram per square meter (gsm) membrane was similar to those of the electrospun PVDF nanofiber membrane only, and the 7 gsm PVDF nanofiber membranes on a PET sheet and the electrospun PVDF nanofiber membrane. This means the protein detection sensitivity is not proportional to the thickness of the PVDF nanofiber membrane. The 4 gsm PVDF nanofiber membrane on a PET sheet can be used to detect proteins with high sensitivity and economic efficiency.

  8. Mapping Point Mutations in the Drosophila Rosy Locus Using Denaturing Gradient Gel Blots

    PubMed Central

    Gray, M.; Charpentier, A.; Walsh, K.; Wu, P.; Bender, W.

    1991-01-01

    Mutations within the rosy locus of Drosophila were mapped using blots of genomic DNA fragments separated on denaturing gradient gels. DNA sequence differences between otherwise identical small rosy DNA fragments were detected among the mutants as mobility shifts on the blots. Mutations were mapped to within a few hundred base pairs of rosy sequence in 100 of 130 mutants tested--a 77% detection rate. The sequence changes in 43 rosy mutations are presented; all but six of these were single base changes. Thirty-four of 36 sequenced mutations induced by the alkylating agents N-ethyl-N-nitrosourea and ethyl methanesulfonate were transitions. All of the mutations mapped in the rosy transcription unit. Twenty-three of the 43 sequenced mutations change the predicted rosy gene polypeptide sequence; the remainder would interrupt protein translation (17), or disrupt mRNA processing (3). PMID:1901817

  9. Direct detection of molluscum contagiosum virus in clinical specimens by dot blot hybridization.

    PubMed Central

    Hurst, J W; Forghani, B; Chan, C S; Oshiro, L; Darai, G

    1991-01-01

    A dot blot hybridization protocol was developed for the direct detection of molluscum contagiosum virus (MCV) DNA in clinical specimens submitted for virus isolation. Samples were concentrated by high-speed centrifugation and treated with proteinase K; this was followed by a single phenol-chloroform extraction step. The DNA was denatured, and the entire volume was spotted onto a nitrocellulose membrane. A biotinylated DNA probe specific for the BamHI-C region of MCV type 1 was used for hybridization. Evidence of MCV DNA was visualized by using streptavidin alkaline phosphatase conjugate and 5-bromo-4-chloro-3-indolyl phosphate-nitroblue tetrazolium as the substrate. Results showed that nonspecific hybridization does not occur with herpes simplex virus- or orf virus-infected clinical specimens and that dot blotting is more sensitive and reproducible than electron microscopy. Images PMID:1774321

  10. Mapping of radiolabeled peptides derived from proteolysis of polypeptides bound to nitrocellulose after Western blotting

    SciTech Connect

    Carrey, E.A.; Hardie, D.G.

    1986-11-01

    Sections of nitrocellulose containing bound /sup 32/P-labeled polypeptides were excised from Western blots and exhaustively digested by trypsin in order to analyze the distribution of phosphorylation sites between the products of limited proteolysis of the multifunctional protein CAD. Using the criterion of analytical isoelectric focusing, the /sup 32/P-peptides obtained by this method were found to be similar, although not identical, to peptides obtained by a more conventional digestion of trichloroacetic acid precipitates. Digestion on Western blots is more straightforward than electrophoretic elution of individual gel slices, gives better recoveries than direct digestion of gel slices, and is particularly suitable for peptide mapping of small peptides which bind to nitrocellulose but would diffuse out of polyacrylamide gels during the commonly used fixing and staining procedures.

  11. Western Blot Analysis of the Exotoxins Components from Bacillus anthracis Separated by Isoelectric Focusing Gel Electrophoresis

    DTIC Science & Technology

    2004-07-01

    Biochemical and Biophysical Research Communications 317 (2004) 294–300 BBRC www.elsevier.com/locate/ybbrcWestern blot analysis...pag iden- tified five different point mutations from 26 different S.F. Little / Biochemical and Biophysical Research Communications 317 (2004) 294...identified as Iowa, it has subsequently been identified as Texas. 296 S.F. Little / Biochemical and Biophysical Research Communications 317 (2004)

  12. Serological Differentiation of Murine Typhus and Epidemic Typhus Using Cross-Adsorption and Western Blotting

    PubMed Central

    La Scola, Bernard; Rydkina, Lena; Ndihokubwayo, Jean-Bosco; Vene, Sirkka; Raoult, Didier

    2000-01-01

    Differentiation of murine typhus due to Rickettsia typhi and epidemic typhus due to Rickettsia prowazekii is critical epidemiologically but difficult serologically. Using serological, epidemiological, and clinical criteria, we selected sera from 264 patients with epidemic typhus and from 44 patients with murine typhus among the 29,188 tested sera in our bank. These sera cross-reacted extensively in indirect fluorescent antibody assays (IFAs) against R. typhi and R. prowazekii, as 42% of the sera from patients with epidemic typhus and 34% of the sera from patients with murine typhus exhibited immunoglobulin M (IgM) and/or IgG titers against the homologous antigen (R. prowazekii and R. typhi, respectively) that were more than one dilution higher than those against the heterologous antigen. Serum cross-adsorption studies and Western blotting were performed on sera from 12 selected patients, 5 with murine typhus, 5 with epidemic typhus, and 2 suffering from typhus of undetermined etiology. Differences in IFA titers against R. typhi and R. prowazekii allowed the identification of the etiological agent in 8 of 12 patients. Western blot studies enabled the identification of the etiological agent in six patients. When the results of IFA and Western blot studies were considered in combination, identification of the etiological agent was possible for 10 of 12 patients. Serum cross-adsorption studies enabled the differentiation of the etiological agent in all patients. Our study indicates that when used together, Western blotting and IFA are useful serological tools to differentiate between R. prowazekii and R. typhi exposures. While a cross-adsorption study is the definitive technique to differentiate between infections with these agents, it was necessary in only 2 of 12 cases (16.7%), and the high costs of such a study limit its use. PMID:10882661

  13. Using genomic slot blot hybridization to assess intergeneric Saccharum x Erianthus hybrids (Andropogoneae - Saccharinae).

    PubMed

    Besse, P; McIntyre, C L; Burner, D M; Almeida, C G

    1997-08-01

    The use of genomic slot blot hybridization enabled the differentiation of hybrids from selfs in Saccharum x Erianthus intergeneric crosses in which Saccharum was used as the female parent. Based on the genomic in situ hybridization technique, slot blots of DNA from the parents and the progeny were blocked with the Saccharum parent DNA and hybridized with the labelled male Erianthus genomic DNA. This technique allowed a rapid screening for hybrids and was sensitive enough to detect a 1/20 dilution of Erianthus in Saccharum DNA, which should enable the detection of most partial hybrids. The genomic slot blot hybridization technique was shown to be potentially useful for assessing crosses involving Saccharum species with either Old World Erianthus section Ripidium or North American Erianthus (= Saccharum) species. The effectiveness of the technique was assessed on 144 progeny of a Saccharum officinarum x Erianthus arundinaceus cross, revealing that 43% of the progeny were selfs. The importance of this test as a tool to support intergeneric breeding programs is discussed.

  14. Use of a Western blot technique for the serodiagnosis of glanders

    PubMed Central

    2011-01-01

    Background The in vivo diagnosis of glanders relies on the highly sensitive complement fixation test (CFT). Frequently observed false positive results are troublesome for veterinary authorities and cause financial losses to animal owners. Consequently, there is an urgent need to develop a test with high specificity. Hence, a Western blot assay making use of a partly purified lipopolysaccaride (LPS) containing antigen of three Burkholderia mallei strains was developed. The test was validated investigating a comprehensive set of positive and negative sera obtained from horses and mules from endemic and non endemic areas. Results The developed Western blot assay showed a markedly higher diagnostic specificity when compared to the prescribed CFT and therefore can be used as a confirmatory test. However, the CFT remains the test of choice for routine testing of glanders due to its high sensitivity, its feasibility using standard laboratory equipment and its worldwide distribution in diagnostic laboratories. Conclusions The CFT should be amended by the newly validated Western blot to increase the positive likelihood ratio of glanders serodiagnosis in non endemic areas or areas with low glanders prevalence. Its use for international trade of horses and mules should be implemented by the OIE. PMID:21247488

  15. Specific Western Blot Bands Are Associated with Initial CD4+ Lymphocyte Counts in Human Immunodeficiency Virus Seroconverters.

    DTIC Science & Technology

    1992-09-30

    SPECIFIC WESTERN BLOT BANDS ARE ASSOCIATED WITH INITIAL CD4+ LYMPHOCYTE COUNTS IN HUMAN IMMUNODEFICIENCY VIRUS SEROCONVERTERS DTI C ELEkCTE AUG...NAVAL MEDICAL RESEARCH AND DEVELOPMENT COMMAND BETHESDA, MARYLAND Specific Western Blot Bands Are Associated With Initial CD4+ Lymphocyte Counts in...and Ms. Susan Yu also provided - quality assurance review of Western blot results. Mr. Jerry Talicurian, Mr. Juan Jurado, Mr. David Morgan, and

  16. Fluorescent detection of Southern blots and PCR-based genetic typing tests

    SciTech Connect

    Mansfield, E.S.; Worley, J.M.; Zimmerman, P.A.

    1994-09-01

    The Southern blot is used to study gene organization, to identify disease-causing genomic rearrangements, or for typing RFLP markers in forensic, paternity, or prenatal diagnostic testing. Fluorescence offers a much greater dynamic range and a more linear response than film used in radioactive or chemiluminescent detection of RFLPs. We therefore investigated using the Fluorimager{trademark} 575 (Molecular Dynamics, Inc.) for analyzing Southern blots. Using a single-locus probe to D2S44 (YNH24) (Promega Corp.), we detect as little as 100 ng (0.05 attomole) genomic DNA. The alkaline phosphatase-labeled probe is detected using AttoPhos (JBL Scientific), and the developed membrane is scanned with the Fluorimager. Biotinylated hybridization probes can also be developed using a streptavidin-alkaline phosphatase conjugate and AttoPhos. The instrument scan parameters can be adjusted to prevent overexposure and accompanying loss of resolution in images of blots, gels, or 96-well microplates. We have used these other sample formats in PCR-based genetic typing assays. We use FluorKit DQS (Molecular Dynamics) to accurately quantify PCR template DNA (1-500 ng) in 96-well microplates scanned using the same instrument. Mutation detection assays run include heteroduplex gels (5% polyacrylamide, 2.7 M urea), short tandem repeat (STR) markers, amplified fragment length polymorphisms (AmpFLP), competitive priming PCR, and allele-specific oligotyping. These assays are run using either 1- or 2-color labeling. We detect unlabeled PCR products, such as the AmpFLP marker D1S80 (Perkin-Elmer) by post-staining gels for 10 minutes with SYBR Green 1 (Molecular Probes) and scanning the wet gel. The Fluorimager scans a 20 x 25 cm sample within three minutes, allowing rapid optimization of fluorescent protocols and high sample throughput.

  17. Western Blotting Is an Efficient Tool for Differential Diagnosis of Paracoccidioidomycosis and Pulmonary Tuberculosis

    PubMed Central

    Bertoni, Thâmara Aline; Perenha-Viana, Maysa Cláudia Zolin; Patussi, Eliana Valéria; Cardoso, Rosilene Fressatti

    2012-01-01

    Sputum and sera from 134 patients screened for tuberculosis (TB) were analyzed to investigate TB and paracoccidioidomycosis (PCM). Of these patients, 11 (8.2%) were confirmed to have TB, but six (4.5%) were positive only for PCM. All patients with PCM presented anti-43-kDa-component antibodies in Western blotting (WB) assays, while in the TB-positive patients these antibodies did not appear. This preliminary study suggests WB as a potential tool for differential laboratory diagnosis between TB and PCM. PMID:22971781

  18. Food additives

    PubMed Central

    Spencer, Michael

    1974-01-01

    Food additives are discussed from the food technology point of view. The reasons for their use are summarized: (1) to protect food from chemical and microbiological attack; (2) to even out seasonal supplies; (3) to improve their eating quality; (4) to improve their nutritional value. The various types of food additives are considered, e.g. colours, flavours, emulsifiers, bread and flour additives, preservatives, and nutritional additives. The paper concludes with consideration of those circumstances in which the use of additives is (a) justified and (b) unjustified. PMID:4467857

  19. Northern cardiometeopathies.

    PubMed

    Hasnulin, V I; Sevost'yanova, E V; Hasnulina, A V

    2001-04-01

    Our research in high latitudes has allowed the identification of a special class of deadaptive disorders, northern cardiometeopathies, which integrates cardiovascular functional violations connected to biologically significant modifications of meteorological, geomagnetic, electrical, gravitational, rhythmological and other geo-ecological factors of the North. Cardiac and cerebral disorder complexes, and also some psychoemotional modifications manifest cardiometeopathies. Cardiometeopathies can occur with developing of pathology, and in such a case they can be selected in the independent form of deadaptive disease. At the same time, cardiometeopathies in case where cardiovascular pathology already exists could cause complications and become a particular risk factor for the development of injuries and myocardial heart attack. The most important mechanism of cardiometeopathies' formation is the organism's reaction to modifications of the Earth's electromagnetic field, based on internal electromagnetic fields' (first of all pulsing field of heart) dependence on the magnitude and directness of modification of the external electromagnetic fields. The analysis of the high degree of dependence of the blood circulation effiency during geomagnetic perturbations in the North of the modification of electromagnetic heart activity allows us to speak of the discovery of an earlier unknown electromagnetic blood pump.

  20. Applying phage antibodies to proteomics: selecting single chain Fv antibodies to antigens blotted on nitrocellulose.

    PubMed

    Liu, B; Marks, J D

    2000-11-01

    Two-dimensional gel electrophoresis is a powerful tool for identification of proteins that differ between patients with qualitatively or quantitatively different disease states. Further characterization of these protein differences would be greatly facilitated by the availability of antibodies that could be used to detect and quantitate the temporo-spatial pattern and cellular and tissue location of the different proteins. To generate such antibodies, methods were developed which permit the successful selection of monoclonal phage antibodies from phage display libraries against antigens blotted from SDS-PAGE gels onto nitrocellulose. First, it was determined that nitrocellulose and PVDF membranes gave significantly lower levels of background phage binding than two other membranes studied. Next, it was determined that blocking with fish gelatin and binding in the presence of 0.5 M NaCl could reduce nonspecific binding 10,000-fold and result in enrichment ratios greater than 500-fold with antigen concentrations as low as 1 ng/mm(2). When optimized conditions were applied to phage antibody libraries, panels of monoclonal phage antibodies were generated against the proteins ErbB2 and bovine serum albumin electroblotted from SDS-PAGE gels onto nitrocellulose. Antibodies were obtained with as little as 10 to 1 ng of antigen, depending on whether the libraries displayed single or multiple copies of antibody per phage. The antibodies worked as reagents in both ELISA and Western blotting.

  1. An overview of technical considerations for Western blotting applications to physiological research.

    PubMed

    Bass, J J; Wilkinson, D J; Rankin, D; Phillips, B E; Szewczyk, N J; Smith, K; Atherton, P J

    2017-01-01

    The applications of Western/immunoblotting (WB) techniques have reached multiple layers of the scientific community and are now considered routine procedures in the field of physiology. This is none more so than in relation to skeletal muscle physiology (i.e., resolving the mechanisms underpinning adaptations to exercise). Indeed, the inclusion of WB data is now considered an essential aspect of many such physiological publications to provide mechanistic insight into regulatory processes. Despite this popularity, and due to the ubiquitous and relatively inexpensive availability of WB equipment, the quality of WB in publications and subsequent analysis and interpretation of the data can be variable, perhaps resulting in spurious conclusions. This may be due to poor laboratory technique and/or lack of comprehension of the critical steps involved in WB and what quality control procedures should be in place to ensure robust data generation. The present review aims to provide a detailed description and critique of WB procedures and technicalities, from sample collection through preparation, blotting and detection, to analysis of the data collected. We aim to provide the reader with improved expertise to critically conduct, evaluate, and troubleshoot the WB process, to produce reproducible and reliable blots.

  2. Analysis of sperm antigens by sodium dodecyl sulfate gel/protein blot radioimmunobinding method

    SciTech Connect

    Lee, C.Y.G.; Huang, Y.S.; Hu, P.C.; Gomel, V.; Menge, A.C.

    1982-06-01

    A radioimmunobinding method based on the blotting of renatured proteins from sodium dodecyl sulfate gels on to nitrocellulose filter papers was developed to analyze the sperm antigens that elicit serum anti-sperm antibodies. In rabbits, serum anti-sperm antibodies were raised by immunization with homologous epididymal spermatozoa mixed with complete Freund's adjuvant. The raised antisera from either male or female rabbits were shown to react with three major sperm protein bands on sodium dodecyl sulfate gels with the corresponding molecular weights of about 70,000 +/- 5000, 14,000, and 13,000, respectively. In humans, the monoclonal antibodies against human sperm were raised by a hybridoma technique. Out of six independent hybrid cell lines that were generated, three of them were shown to secrete immunoglobulins that react with the same two protein bands on sodium dodecyl sulfate gels, which have the approximate molecular weight of 10,000. The same procedure was also used to analyze human serum samples that were shown to contain anti-sperm antibodies by the known techniques. Unique sperm antigens that elicit anti-sperm antibodies in humans were identified and correlated. The results of this study suggest that sodium dodecyl sulfate gel/protein blot radioimmunobinding method may be a sensitive and useful tool for the study of sperm antigens that elicit autoimmune responses and their association with human infertility.

  3. A simple DNA recombination screening method by RT-PCR as an alternative to Southern blot.

    PubMed

    Albers, Eliene; Sbroggiò, Mauro; Martin-Gonzalez, Javier; Avram, Alexandra; Munk, Stephanie; Lopez-Contreras, Andres J

    2017-01-19

    The generation of genetically engineered mouse models (GEMMs), including knock-out (KO) and knock-in (KI) models, often requires genomic screening of many mouse ES cell (mESC) clones by Southern blot. The use of large targeting constructs facilitates the recombination of exogenous DNA in a specific genomic locus, but limits the detection of its correct genomic integration by standard PCR methods. Genomic Long Range PCR (LR-PCR), using primers adjacent to the homology arms, has been used as an alternative to radioactive-based Southern blot screenings. However, LR-PCRs are often difficult and render many false positive and false negative results. Here, we propose an alternative screening method based on the detection of a genetic modification at the mRNA level, which we successfully optimized in two mouse models. This screening method consists of a reverse-transcription PCR (RT-PCR) using primers that match exons flanking the targeting construct. The detection of the expected modification in this PCR product confirms the integration at the correct genomic location and shows that the mutant mRNA is expressed. This is a simple and sensitive strategy to screen locus-specific recombination of targeting constructs which can also be useful to screen KO and KI mutant mice or cell lines including those generated by CRISPR/Cas9.

  4. Capillary blotting of glycosaminoglycans on nitrocellulose membranes after agarose-gel electrophoresis separation.

    PubMed

    Volpi, Nicola; Maccari, Francesca

    2009-01-01

    A method for the blotting and immobilizing of several nonsulfated and sulfated complex polysaccharides on membranes made hydrophilic and positively charged by cationic detergent after their separation by conventional agarose gel electrophoresis is illustrated. This new approach to the study of glycosaminoglycans (GAGs) utilizes the capacity of agarose gel electrophoresis to separate single species of polysaccharides from mixtures and the membrane technology for further preparative and analytical uses.Nitrocellulose membranes are derivatized with the cationic detergent cetylpyridinium chloride and mixtures of GAGs are capillary blotted after their separation in agarose gel electrophoresis. Single purified species of variously sulfated polysaccharides are transferred on derivatized membranes with an efficiency of 100% and stained with alcian blue (irreversible staining) and toluidine blue (reversible staining). This enables a lower amount limit of detection of 0.1 microg. Nonsulfated polyanions, for example hyaluronic acid, may also be transferred to membranes with a limit of detection of approximately 0.1-0.5 microg after irreversible or reversible staining. The membranes may be stained with reversible staining and the same lanes are used for immunological detection or other applications.

  5. Quantitation of proteoglycans as glycosaminoglycans in biological fluids using an alcian blue dot blot analysis.

    PubMed

    Björnsson, S

    1998-02-15

    A method for quantitation of intact proteoglycans as GAGs in biological fluids (blood plasma, synovial fluid) or 4 M guanidine extracts of tissues has been published previously (S. Björnsson, Anal. Biochem. 210, 282-291, 1993). The method is based on the specific interaction between sulfated polymers and the tetravalent cationic dye Alcian blue at pH 1.5 in 0.4 M guanidine-HCl and in the presence of 0.25% Triton. The absorbance assay has a measuring range of 1-20 microgram of glycosaminoglycan (GAG) which is not sensitive enough to measure the low contents of proteoglycans in blood plasma, urine, or wound fluid. A dot blot assay is now described in which the Alcian blue-GAG complexes are collected on a polyvinylidene fluoride membrane, by filtration in a dot blot apparatus, and the stain is quantitated as reflectance by scanning and densitometry. The assay requires 10 microliter of sample and has a measuring range of 10-800 ng of GAG, corresponding to a concentration of 1-80 mg/liter, suitable for proteoglycans in biological fluids. The procedures for chemistry, scanning, densitometry, and curve fitting were each evaluated separately. The error contributed by chemistry accounted for a minor portion of the imprecision. The imprecision contributed by scanning was the most important source of within-run and between-run imprecision, and was caused by inequalities of the charge-coupled device along the scanning arm. Unexpectedly, curve fitting was also a major source of total imprecision in dot blot quantitation and differed with the type of equation used. The between-run imprecision calculated as CV (SD/mean . 100) was 13.0% at 8 mg/liter. The response of the assay was identical for six different commercial preparations of GAGs (chondroitin-4-sulfate, chondroitin-6-sulfate, dermatan sulfate, keratan sulfate, heparan sulfate, and heparin) despite differences in degree of sulfation known to exist. There was no positive or negative interference by blood plasma, apart

  6. Recombinant Dense Granular Protein (GRA5) for Detection of Human Toxoplasmosis by Western Blot

    PubMed Central

    Ching, Xiao Teng; Lau, Yee Ling; Fong, Mun Yik; Nissapatorn, Veeranoot; Andiappan, Hemah

    2014-01-01

    Toxoplasma gondii infects all warm-blooded animals, including humans, causing serious public health problems and great economic loss for the food industry. Commonly used serological tests require costly and hazardous preparation of whole Toxoplasma lysate antigens from tachyzoites. Here, we have evaluated an alternative method for antigen production, which involved a prokaryotic expression system. Specifically, we expressed T. gondii dense granular protein-5 (GRA5) in Escherichia coli and isolated it by affinity purification. The serodiagnostic potential of the purified recombinant GRA5 (rGRA5) was tested through Western blot analysis against 212 human patient serum samples. We found that rGRA5 protein was 100% specific for analysis of toxoplasmosis-negative human sera. Also, rGRA5 was able to detect acute and chronic T. gondii infections (sensitivities of 46.8% and 61.2%, resp.). PMID:24987700

  7. Western blot analysis of Src kinase assays using peptide substrates ligated to a carrier protein.

    PubMed

    Xu, Jie; Sun, Luo; Ghosh, Inca; Xu, Ming-Qun

    2004-06-01

    We have applied intein-mediated peptide ligation (IPL) to the use of peptide substrates for kinase assays and subsequent Western blot analysis. IPL allows for the efficient ligation of a synthetic peptide with an N-terminal cysteine residue to an intein-generated carrier protein containing a cysteine reactive C-terminal thioester through a native peptide bond. A distinct advantage of this procedure is that each carrier protein molecule ligates only one peptide, ensuring that the ligation product forms a sharp band on sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). We demonstrate the effectiveness of this approach by mutational analysis of peptide substrates derived from human cyclin-dependent kinase, Cdc2, which contains a phosphorylation site of human c-Src protein tyrosine kinase.

  8. Reverse line blot hybridization used to identify hemoprotozoa in Minorcan cattle.

    PubMed

    Almeria, Sonia; Castella, Joaquim; Ferrer, David; Gutierrez, Juan Francisco; Estrada-Pena, Agustin; Sparagano, Olivier

    2002-10-01

    Piroplasmosis, a tick-borne protozoal disease, is an important disease affecting domestic and wild animals. We performed PCR-based reverse line blot hybridization (RLB) assays on blood samples obtained from 133 cattle exposed to ticks in field conditions in Minorca (Balearic Islands, Spain) in three different seasons. The oligonucleotides used were those for Theileria annulata, T. buffeli, T. taurotragi, T. velifera, Babesia bigemina, B. bovis, B. divergens, and B. major. The RLB technique allowed the simultaneous identification of T. annulata, T. buffeli, B. bigemina, and B. bovis as the piroplasms present in cattle in Minorca. Of the 133 animals, only 4 were not infected by any of the studied parasites. The results indicated endemic piroplasm infection in cattle in Minorca; especially important was the presence of T. annulata. The RLB was highly sensitive and allowed the simultaneous detection and identification of the Theileria and Babesia species in carrier cattle, which cannot be achieved by classical identification methods.

  9. Direct identification and characterization of llama (Lama glama L.) whey proteins by microsequencing after western blotting.

    PubMed

    Cantisani, A; Napolitano, L; Giuffrida, M G; Conti, A

    1990-01-01

    Amino acid sequence determination is the most reliable and powerful tool to identify a protein or to classify a new one by comparison of its primary structure with already known sequences. A rapid and simple purification procedure is an essential pre-requisite for routine sequence determination. Structural characterization of llama whey proteins was undertaken for evolutionary as well as economic purposes. N-terminal sequence analyses directly on an immobilon polyvinylidene difluoride (PVDF) membrane, following Western blotting of both native and SDS-denatured llama whey proteins after polyacrylamide gel electrophoresis, revealed three different forms of glycosylated alpha-lactalbumin, and a protein with a high degree of homology with a camel whey protein of unknown function. Furthermore, by immunoblotting techniques, the electrophoretic band corresponding to serum albumin was identified.

  10. Optimization of dot blot method to detect bcr/abl transcripts in chronic myeloid leukemia

    SciTech Connect

    Tharapel, S.A.; Zhao, J.

    1994-09-01

    Detection of abl-bcr fusion transcripts using molecular methodologies is becoming an attractive alternative (or supplement) to traditional cytogenetics in identifying the Philadelphia (Ph) chromosome. Among these methods, RT-PCR technique has provided an extremely powerful tool for improving the detection of bcr/abl translocations through enzymatic amplification of the reverse-transcribed cDNA. The analysis of PCR products can be accomplished by a number of techniques including dot blot following liquid-phase hybridization. In order to render the detection of PCR products more simple, accurate and efficient, and therefore more amenable for the clinical laboratory routine use, we optimized several parameters of the procedure. (1) We discovered that with the starting material of 1 ug of total RNA, the amount of the final PCR amplified products was linear to the PCR cycles between 20 to 30 cycles. Since the dot blot procedure does not separate the amplified products according to their sizes, increased background would increase the false positive rate. (2) If a detection sensitivity of 1 in 10{sup 3} cells is sufficient, then the nested or a second PCR amplification is not necessary. (3) Starting material more than 5 ug of total RNA would decrease the amplification efficiency and therefore compromise the sensitivity. (4) Ten minutes of hybridization gave equal signal intensity as 24 hours. (5) The ionic strength and temperature in the washing step were also tested. Upon optimization of each parameter, the detection procedure was tested on 18 clinical samples. Compared to the procedures that are currently available, our optimized procedure is less time consuming, has higher sensitivity and lower false positive rate. This method has the potential to be automated and therefore can be used as a screening method for Ph chromosome in high volume settings.

  11. Western blot diagnosis of vivax malaria with multiple stage-specific antigens of the parasite

    PubMed Central

    Son, Eui-Sun; Kim, Tong Soo

    2001-01-01

    Western blot analysis was performed to diagnose vivax malaria using stage-specific recombinant antigens. Genomic DNA from the whole blood of a malaria patient was used as templates to amplify the coding regions for the antigenic domains of circumsporozoite protein (CSP-1), merozoite surface protein (MSP-1), apical merozoite antigen (AMA-1), serine repeat antigen (SERA), and exported antigen (EXP-1) of Plasmodium vivax. Each amplified DNA fragment was inserted into a pGEX-4T plasmid to induce the expression of GST fusion protein in Escherichia coli by IPTG. The bacterial cell extracts were separated on 10% SDS-PAGE followed by western blot analysis with patient sera which was confirmed by blood smear examination. When applied with patient sera, 147 (91.9%) out of 160 vivax malaria, 12 (92.3%) out of 13 falciparum malaria, and all 9 vivax/falciparum mixed malaria reacted with at least one antigen, while no reactions occurred with 20 normal uninfected sera. In the case of vivax malaria, CSP-1 reacted with 128 (80.0%) sera, MSP-1 with 102 (63.8%), AMA-1 with 128 (80.0%), SERA with 115 (71.9%), and EXP-1 with 89 (55.6%), respectively. We obtained higher detection rates when using 5 antigens (91.9%) rather than using each antigen solely (55.6-80%), a combination of 2 (76.3-87.5%), 3 (85.6-90.6%), or 4 antigens (89.4-91.3%). This method can be applied to serological diagnosis, mass screening in endemic regions, or safety test in transfusion of prevalent vivax malaria. PMID:11441504

  12. Western blot diagnosis of vivax malaria with multiple stage-specific antigens of the parasite.

    PubMed

    Son, E S; Kim, T S; Nam, H W

    2001-06-01

    Western blot analysis was performed to diagnose vivax malaria using stage-specific recombinant antigens. Genomic DNA from the whole blood of a malaria patient was used as templates to amplify the coding regions for the antigenic domains of circumsporozoite protein (CSP-1), merozoite surface protein (MSP-1), apical merozoite antigen (AMA-1), serine repeat antigen (SERA), and exported antigen (EXP-1) of Plasmodium vivax. Each amplified DNA fragment was inserted into a pGEX-4T plasmid to induce the expression of GST fusion protein in Escherichia coli by IPTG. The bacterial cell extracts were separated on 10% SDS-PAGE followed by western blot analysis with patient sera which was confirmed by blood smear examination. When applied with patient sera, 147 (91.9%) out of 160 vivax malaria, 12 (92.3%) out of 13 falciparum malaria, and all 9 vivax/falciparum mixed malaria reacted with at least one antigen, while no reactions occurred with 20 normal uninfected sera. In the case of vivax malaria, CSP-1 reacted with 128 (80.0%) sera, MSP-1 with 102 (63.8%), AMA-1 with 128 (80.0%), SERA with 115 (71.9%), and EXP-1 with 89 (55.6%), respectively. We obtained higher detection rates when using 5 antigens (91.9%) rather than using each antigen solely (55.6-80%), a combination of 2 (76.3-87.5%), 3 (85.6-90.6%), or 4 antigens (89.4-91.3%). This method can be applied to serological diagnosis, mass screening in endemic regions, or safety test in transfusion of prevalent vivax malaria.

  13. Comparative evaluation of western blotting in hepatic and pulmonary cystic echinococcosis.

    PubMed

    Akisu, C; Delibas, S B; Bicmen, C; Ozkoc, S; Aksoy, U; Turgay, N

    2006-12-01

    Many serological tests are widely used in the diagnosis of cystic echinococcosis (CE), caused by the larval stages of Echinococcus granulosus. The present study was carried for differentiation between hepatic and pulmonary cystic echinococcosis by Western Blotting (WB). A total of 121 sera from patients with hepatic CE (37), pulmonary CE (31) and controls (53; consisting of six healthy, seven Hymenolepis nana infection, 20 hepatic and 20 pulmonary diseases other than CE) were examined. In all of the CE patients, E. gronulosus infection was confirmed by surgical intervention. Sera were previously tested using IHA and ELISA to detect the E. gronulosus specific antibodies. Sera from hepatic cases of CE reacted with 16 polypeptides of 6-116 kDa and sera from pulmonary cases of CE reacted with 14 polypeptides of 4-130 kDa by Western Blotting. The WB test enabled the detection of antibodies in the hepatic CE samples for proteins of 24, 32 34, 44-46 and 52-54 kDa in molecular weight in 78.4%, 75.7%, 78.4% and 89.2% of the patients, respectively. In the pulmonary CE samples sera WB test enabled the detection of antibodies 24, 44-46, 100, 110, 116 and 120 124 kDa in molecular weight in 81.3%, 75.0%, 87.5%, 71.9%, 84.4% and 65.6% of the patients, respectively. We indicated that the antigenic components of high molecular weight can be good candidates for differentiation of hepatic CE from pulmonary CE.

  14. Rapid detection and differentiation of important Campylobacter spp. in poultry samples by dot blot and PCR.

    PubMed

    Fontanot, Marco; Iacumin, Lucilla; Cecchini, Francesca; Comi, Giuseppe; Manzano, Marisa

    2014-10-01

    The detection of Campylobacter, the most commonly reported cause of foodborne gastroenteritis in the European Union, is very important for human health. The most commonly recognised risk factor for infection is the handling and/or consumption of undercooked poultry meat. The methods typically applied to evaluate the presence/absence of Campylobacter in food samples are direct plating and/or enrichment culture based on the Horizontal Method for Detection and Enumeration of Campylobacter spp. (ISO 10272-1B: 2006) and PCR. Molecular methods also allow for the detection of cells that are viable but cannot be cultivated on agar media and that decrease the time required for species identification. The current study proposes the use of two molecular methods for species identification: dot blot and PCR. The dot blot method had a sensitivity of 25 ng for detection of DNA extracted from a pure culture using a digoxigenin-labelled probe for hybridisation; the target DNA was extracted from the enrichment broth at 24 h. PCR was performed using a pair of sensitive and specific primers for the detection of Campylobacter jejuni and Campylobacter coli after 24 h of enrichment in Preston broth. The initial samples were contaminated by 5 × 10 C. jejuni cells/g and 1.5 × 10(2)C. coli cells/g, thus the number of cells present in the enrichment broth at 0 h was 1 or 3 cell/g, respectively.

  15. Food additives

    MedlinePlus

    ... or natural. Natural food additives include: Herbs or spices to add flavor to foods Vinegar for pickling ... Certain colors improve the appearance of foods. Many spices, as well as natural and man-made flavors, ...

  16. Induction of a protective antibody response to FMDV in mice following oral immunization with transgenic Stylosanthes spp. as a feedstuff additive.

    PubMed

    Wang, Dong Mei; Zhu, Jian Bo; Peng, Ming; Zhou, Peng

    2008-12-01

    The expression of antigens in transgenic plants has increasingly been used as an alternative to the classical methodologies for the development of experimental vaccines, and it remains one of the real challenges in this field to use transgenic plant-based vaccines effectively as feedstuff additives. We report herein the development of a new oral immunization system for foot and mouth disease with the structural protein VP1 of the foot and mouth disease virus (FMDV) produced in transgenic Stylosanthes guianensis cv. Reyan II. The transgenic plantlets were identified by polymerase chain reaction (PCR), Southern blotting, and northern blotting; and the production of VP1 protein in transgenic plants was confirmed and quantified by western blotting and enzyme-linked immunosorbent assays (ELISA). Six transformed lines were obtained, and the level of the expressed protein was 0.1-0.5% total soluble protein (TSP). Mice that were orally immunized using studded feedstuff mixed with desiccated powder of the transgenic plants developed a virus-specific immune response to the structural VP1 and intact FMDV particles. To our knowledge, this is the first report of transgenic plants expressing the antigen protein of FMDV as feedstuff additives that has demonstrated the induction of a protective systemic antibody response in animals. These results support the feasibility of producing edible vaccines from transgenic forage plants, and provide proof of the possibility of using plant-based vaccines as feedstuff additives.

  17. Combination of Insecticide Treated Nets and Indoor Residual Spraying in Northern Tanzania Provides Additional Reduction in Vector Population Density and Malaria Transmission Rates Compared to Insecticide Treated Nets Alone: A Randomised Control Trial

    PubMed Central

    Protopopoff, Natacha; Wright, Alexandra; West, Philippa A; Tigererwa, Robinson; Mosha, Franklin W; Kisinza, William; Kleinschmidt, Immo; Rowland, Mark

    2015-01-01

    Indoor residual spraying (IRS) combined with insecticide treated nets (ITN) has been implemented together in several sub-Saharan countries with inconclusive evidence that the combined intervention provides added benefit. The impact on malaria transmission was evaluated in a cluster randomised trial comparing two rounds of IRS with bendiocarb plus universal coverage ITNs, with ITNs alone in northern Tanzania. From April 2011 to December 2012, eight houses in 20 clusters per study arm were sampled monthly for one night with CDC light trap collections. Anopheles gambiae s.l. were identified to species using real time PCR Taq Man and tested for the presence of Plasmodium falciparum circumsporozoite protein. ITN and IRS coverage was estimated from household surveys. IRS coverage was more than 85% in two rounds of spraying in January and April 2012. Household coverage with at least one ITN per house was 94.7% after the universal coverage net campaign in the baseline year and the proportion of household with all sleeping places covered by LLIN was 50.1% decreasing to 39.1% by the end of the intervention year. An.gambiae s.s. comprised 80% and An.arabiensis 18.3% of the anopheline collection in the baseline year. Mean An.gambiae s.l. density in the ITN+IRS arm was reduced by 84% (95%CI: 56%-94%, p = 0.001) relative to the ITN arm. In the stratum of clusters categorised as high anopheline density at baseline EIR was lower in the ITN+IRS arm compared to the ITN arm (0.5 versus 5.4 per house per month, Incidence Rate Ratio: 0.10, 95%CI: 0.01–0.66, p-value for interaction <0.001). This trial provides conclusive evidence that combining carbamate IRS and ITNs produces major reduction in Anopheles density and entomological inoculation rate compared to ITN alone in an area of moderate coverage of LLIN and high pyrethroid resistance in An.gambiae s.s. PMID:26569492

  18. Combination of Insecticide Treated Nets and Indoor Residual Spraying in Northern Tanzania Provides Additional Reduction in Vector Population Density and Malaria Transmission Rates Compared to Insecticide Treated Nets Alone: A Randomised Control Trial.

    PubMed

    Protopopoff, Natacha; Wright, Alexandra; West, Philippa A; Tigererwa, Robinson; Mosha, Franklin W; Kisinza, William; Kleinschmidt, Immo; Rowland, Mark

    2015-01-01

    Indoor residual spraying (IRS) combined with insecticide treated nets (ITN) has been implemented together in several sub-Saharan countries with inconclusive evidence that the combined intervention provides added benefit. The impact on malaria transmission was evaluated in a cluster randomised trial comparing two rounds of IRS with bendiocarb plus universal coverage ITNs, with ITNs alone in northern Tanzania. From April 2011 to December 2012, eight houses in 20 clusters per study arm were sampled monthly for one night with CDC light trap collections. Anopheles gambiae s.l. were identified to species using real time PCR Taq Man and tested for the presence of Plasmodium falciparum circumsporozoite protein. ITN and IRS coverage was estimated from household surveys. IRS coverage was more than 85% in two rounds of spraying in January and April 2012. Household coverage with at least one ITN per house was 94.7% after the universal coverage net campaign in the baseline year and the proportion of household with all sleeping places covered by LLIN was 50.1% decreasing to 39.1% by the end of the intervention year. An.gambiae s.s. comprised 80% and An.arabiensis 18.3% of the anopheline collection in the baseline year. Mean An.gambiae s.l. density in the ITN+IRS arm was reduced by 84% (95%CI: 56%-94%, p = 0.001) relative to the ITN arm. In the stratum of clusters categorised as high anopheline density at baseline EIR was lower in the ITN+IRS arm compared to the ITN arm (0.5 versus 5.4 per house per month, Incidence Rate Ratio: 0.10, 95%CI: 0.01-0.66, p-value for interaction <0.001). This trial provides conclusive evidence that combining carbamate IRS and ITNs produces major reduction in Anopheles density and entomological inoculation rate compared to ITN alone in an area of moderate coverage of LLIN and high pyrethroid resistance in An.gambiae s.s.

  19. Avoiding Pitfalls of Internal Controls: Validation of Reference Genes for Analysis by qRT-PCR and Western Blot throughout Rat Retinal Development

    PubMed Central

    Rocha-Martins, Maurício; Njaine, Brian; Silveira, Mariana S.

    2012-01-01

    Background Housekeeping genes have been commonly used as reference to normalize gene expression and protein content data because of its presumed constitutive expression. In this paper, we challenge the consensual idea that housekeeping genes are reliable controls for expression studies in the retina through the investigation of a panel of reference genes potentially suitable for analysis of different stages of retinal development. Methodology/Principal Findings We applied statistical tools on combinations of retinal developmental stages to assess the most stable internal controls for quantitative RT-PCR (qRT-PCR). The stability of expression of seven putative reference genes (Actb, B2m, Gapdh, Hprt1, Mapk1, Ppia and Rn18s) was analyzed using geNorm, BestKeeper and Normfinder software. In addition, several housekeeping genes were tested as loading controls for Western blot in the same sample panel, using Image J. Overall, for qRT-PCR the combination of Gapdh and Mapk1 showed the highest stability for most experimental sets. Actb was downregulated in more mature stages, while Rn18s and Hprt1 showed the highest variability. We normalized the expression of cyclin D1 using various reference genes and demonstrated that spurious results may result from blind selection of internal controls. For Western blot significant variation could be seen among four putative internal controls (β-actin, cyclophilin b, α-tubulin and lamin A/C), while MAPK1 was stably expressed. Conclusion Putative housekeeping genes exhibit significant variation in both mRNA and protein content during retinal development. Our results showed that distinct combinations of internal controls fit for each experimental set in the case of qRT-PCR and that MAPK1 is a reliable loading control for Western blot. The results indicate that biased study outcomes may follow the use of reference genes without prior validation for qRT-PCR and Western blot. PMID:22916200

  20. Far-western blotting as a solution to the non-specificity of the anti-erythropoietin receptor antibody

    PubMed Central

    Fecková, Barbora; Kimáková, Patrícia; Ilkovičová, Lenka; Szentpéteriová, Erika; Debeljak, Nataša; Solárová, Zuzana; Sačková, Veronika; Šemeláková, Martina; Bhide, Mangesh; Solár, Peter

    2016-01-01

    The erythropoietin receptor (EpoR) is a member of the cytokine receptor family. The interaction between erythropoietin (Epo) and EpoR is important for the production and maturation of erythroid cells, resulting in the stimulation of hematopoiesis. The fact that EpoR was also detected in neoplastic cells has opened the question about the relevance of anemia treatment with recombinant Epo in cancer patients. Numerous studies have reported pro-stimulating and anti-apoptotic effects of Epo in cancer cells, thus demonstrating EpoR functionality in these cells. By contrast, a previous study claims the absence of EpoR in tumor cells. This apparent discrepancy is based, according to certain authors, on the use of non-specific anti-EpoR antibodies. With the aim of bypassing the direct detection of EpoR with an anti-EpoR antibody, the present authors propose a far-western blot methodology, which in addition, confirms the interaction of Epo with EpoR. Applying this technique, the presence of EpoR and its interaction with Epo in human ovarian adenocarcinoma A2780 and normal human umbilical vein endothelial cells was confirmed. Furthermore, modified immunoprecipitation of EpoR followed by matrix-assisted laser desorption/ionization-time-of-flight mass spectrometry analysis confirmed a 57 kDa protein as a human Epo-interacting protein in both cell lines. PMID:27446474

  1. A cost effective non-commercial ECL-solution for Western blot detections yielding strong signals and low background.

    PubMed

    Haan, Claude; Behrmann, Iris

    2007-01-10

    We compared several alternative ECL solutions for Western blot detection of endogenous proteins in whole cell lysates using inexpensive, commercially available reagents. Starting from an existing protocol based on p-coumaric acid (pCA) as enhancer, we found that the ECL solution containing 4-iodophenylboronic acid (4IPBA) generated strong specific signals and low background chemiluminescence. We optimised the luminol, 4IPBA and hydrogenperoxide concentrations of this 4IPBA-ECL solution. The optimised 4IPBA-ECL solution (100 mM Tris/HCl pH 8.8, 1.25 mM luminol, 2 mM 4IPBA, 5.3 mM hydrogenperoxide) shows a greatly increased signal intensity compared to the initial pCA-ECL protocol and to some commercially available ECL solutions. In addition, the optimised 4IPBA-ECL solution also generates much lower background chemiluminescence than other non-commercial ECL solutions using p-coumaric acid or 4-iodophenol as enhancers. The 4IPBA-ECL solution was stable when stored but had the lowest background when prepared freshly from stock solutions. Thus, we present an optimised protocol for a well-performing inexpensive ECL solution which is an alternative to expensive commercial ECL solutions and which achieves a better signal and lower background than the commercial solutions tested.

  2. The diagnosis of proventricular dilatation disease: use of a Western blot assay to detect antibodies against avian Borna virus.

    PubMed

    Villanueva, Itamar; Gray, Patricia; Mirhosseini, Negin; Payne, Susan; Hoppes, Sharman; Honkavuori, Kirsi S; Briese, Thomas; Turner, Debra; Tizard, Ian

    2010-07-14

    Avian Borna virus (ABV) has recently been shown to be the causal agent of proventricular dilatation disease (PDD) a lethal neurologic disease of captive psittacines and other birds. An immunoblot assay was used to detect the presence of antibodies against avian Borna virus in the serum of affected birds. A lysate from ABV-infected duck embryo fibroblasts served as a source of antigen. The assay was used to test for the presence of antibodies to ABV in 117 birds. Thirty of these birds had biopsy or necropsy-confirmed proventricular dilatation disease (PDD), while the remaining 87 birds were apparently healthy or were suffering from diseases other than PDD. Sera from 27 of the 30 PDD cases (90%) contained antibodies to ABV. Seventy-three (84%) of the apparently "healthy" birds were seronegative. Additionally, sera from seven macaws and one parrot trapped in the Peruvian Amazon were seronegative. Positive sera recognized the bornaviral nucleoprotein (N-protein). While the presence of antibodies to ABV largely corresponded with the development of clinical PDD, 14 apparently healthy normal birds possessed detectable antibodies to ABV. The existence of a carrier state was confirmed when 13 of 15 apparently healthy cockatiels were shown by PCR to have detectable ABV RNA in their feces. Western blot assays may be of significant assistance in diagnosing proventricular dilatation disease. Many apparently healthy birds may however be seronegative while, at the same time, shedding ABV in their feces.

  3. Western blot analysis to illustrate relative control levels of the lac and ara promoters in Escherichia coli.

    PubMed

    Nielsen, Brent L; Willis, Van C; Lin, Chin-Yo

    2007-03-01

    The lactose operon and its control is a fundamental transcriptional regulatory concept presented in introductory and many advanced molecular biology courses. Much is known about the positive and negative control mechanisms that govern levels of expression of this operon. One basic principle that is taught about the lac operon is that it is "leaky," meaning that the transcriptional control of the operon is not 100% efficient and that in wild-type cells, transcription from the promoter is never completely "off," but there is always some basal transcription. In contrast, the arabinose operon is often used as an example of a tightly controlled operon, and transcription from the ara promoter is very low in the absence of inducer. The relative levels of control of these two operons can be illustrated using Western blots of proteins expressed in the presence and absence of the appropriate inducers and antibodies against the gene products. Different times of growth and the addition of inducer can also be examined. The results are very dramatic and help to reinforce the principles of promoter control.

  4. Exposure to Sarcocystis spp. in horses from Spain determined by Western blot analysis using Sarcocystis neurona merozoites as heterologous antigen.

    PubMed

    Arias, M; Yeargan, M; Francisco, I; Dangoudoubiyam, S; Becerra, P; Francisco, R; Sánchez-Andrade, R; Paz-Silva, A; Howe, D K

    2012-04-30

    Horses serve as an intermediate host for several species of Sarcocystis, all of which utilize canids as the definitive host. Sarcocystis spp. infection and formation of latent sarcocysts in horses often appears to be subclinical, but morbidity can occur, especially when the parasite burden is large. A serological survey was conducted to determine the presence of antibodies against Sarcocystis spp. in seemingly healthy horses from the Galicia region of Spain. Western blot analyses using Sarcocystis neurona merozoites as heterologous antigen suggested greater than 80% seroprevalance of Sarcocystis spp. in a sample set of 138 horses. The serum samples were further tested with enzyme-linked immunosorbent assays (ELISAs) based on recombinant S. neurona-specific surface antigens (rSnSAGs). As expected for horses from the Eastern Hemisphere, less than 4% of the serum samples were positive when analyzed with either the rSnSAG2 or the rSnSAG4/3 ELISAs. An additional 246 horses were tested using the rSnSAG2 ELISA, which revealed that less than 3% of the 384 samples were seropositive. Collectively, the results of this serologic study suggested that a large proportion of horses from this region of Spain are exposed to Sarcocystis spp. Furthermore, the anti-Sarcocystis seroreactivity in these European horses could be clearly distinguished from anti-S. neurona antibodies using the rSnSAG2 and rSnSAG4/3 ELISAs.

  5. Double-blotting: a solution to the problem of nonspecific binding of secondary antibodies in immunoblotting procedures.

    PubMed

    Lasne, Françoise

    2009-01-01

    Nonspecific interactions between blotted proteins and unrelated secondary antibodies generate false positives in immunoblotting techniques. Some procedures have been developed to reduce this adsorption but they may work in specific applications and be ineffective in other ones. "Double-blotting" has been developed to overcome this problem. It consists of interpolating a second blotting step between the usual probings of the blot membrane with the primary antibody and the secondary antibodies. This step, by isolating the primary antibody from the interfering proteins, guarantees the specificity of the probing with the secondary antibody. This method has been developed for the study of erythropoietin in concentrated urine since a strong nonspecific binding of biotinylated secondary antibodies to some urinary proteins is observed using classical immunoblotting protocols. However, its concept makes it usable in other applications that come up against this kind of problem. This method is expected to be especially useful for investigating proteins that are present in minute amounts in complex biological media.

  6. Double-blotting: a solution to the problem of nonspecific binding of secondary antibodies in immunoblotting procedures.

    PubMed

    Lasne, Françoise

    2015-01-01

    Nonspecific interactions between blotted proteins and unrelated secondary antibodies generate false positives in immunoblotting techniques. Some procedures have been developed to reduce this adsorption, but they may work in specific applications and be ineffective in others. "Double-blotting" has been developed to overcome this problem. It consists of interpolating a second blotting step between the usual probings of the blot membrane with the primary antibody and the secondary antibodies. This step, by isolating the primary antibody from the interfering proteins, guarantees the specificity of the probing with the secondary antibody. This method has been developed for the study of erythropoietin in concentrated urine since a strong nonspecific binding of biotinylated secondary antibodies to some urinary proteins is observed using classical immunoblotting protocols. However, its concept makes it usable in other applications that come up against this kind of problem. This method is expected to be especially useful for investigating proteins that are present in minute amounts in complex biological media.

  7. Potlining Additives

    SciTech Connect

    Rudolf Keller

    2004-08-10

    In this project, a concept to improve the performance of aluminum production cells by introducing potlining additives was examined and tested. Boron oxide was added to cathode blocks, and titanium was dissolved in the metal pool; this resulted in the formation of titanium diboride and caused the molten aluminum to wet the carbonaceous cathode surface. Such wetting reportedly leads to operational improvements and extended cell life. In addition, boron oxide suppresses cyanide formation. This final report presents and discusses the results of this project. Substantial economic benefits for the practical implementation of the technology are projected, especially for modern cells with graphitized blocks. For example, with an energy savings of about 5% and an increase in pot life from 1500 to 2500 days, a cost savings of $ 0.023 per pound of aluminum produced is projected for a 200 kA pot.

  8. Phosphazene additives

    DOEpatents

    Harrup, Mason K; Rollins, Harry W

    2013-11-26

    An additive comprising a phosphazene compound that has at least two reactive functional groups and at least one capping functional group bonded to phosphorus atoms of the phosphazene compound. One of the at least two reactive functional groups is configured to react with cellulose and the other of the at least two reactive functional groups is configured to react with a resin, such as an amine resin of a polycarboxylic acid resin. The at least one capping functional group is selected from the group consisting of a short chain ether group, an alkoxy group, or an aryloxy group. Also disclosed are an additive-resin admixture, a method of treating a wood product, and a wood product.

  9. Identification of Yeast V-ATPase Mutants by Western Blots Analysis of Whole Cell Lysates

    NASA Astrophysics Data System (ADS)

    Parra-Belky, Karlett

    2002-11-01

    A biochemistry laboratory was designed for an undergraduate course to help students better understand the link between molecular engineering and biochemistry. Students identified unknown yeast strains with high specificity using SDS-PAGE and Western blot analysis of whole cell lysates. This problem-solving exercise is a common application of biochemistry in biotechnology research. Three different strains were used: a wild-type and two mutants for the proton pump vacuolar ATPase (V-ATPase). V-ATPases are multisubunit enzymes and the mutants used were deletion mutants; each lacked one structural gene of the complex. After three, three-hour labs, mutant strains were easily identified by the students and distinguished from wild-type cells analyzing the pattern of SDS-PAGE distribution of proteins. Identifying different subunits of one multimeric protein allowed for discussion of the structure and function of this metabolic enzyme, which captured the interest of the students. The experiment can be adapted to other multimeric protein complexes and shows improvement of the described methodology over previous reports, perhaps because the problem and its solution are representative of the type of techniques currently used in research labs.

  10. The prevalence and significance of HTLV-I/II seroindeterminate Western blot patterns.

    PubMed

    Abrams, Anna; Akahata, Yoshimi; Jacobson, Steven

    2011-08-01

    Human T-lymphotropic virus type I (HTLV-I) infects an estimated 15-20 million persons worldwide. A number of diseases have been associated with the virus including adult T-cell leukemia (ATL), HTLV-associated myelopathy/tropical spastic paraparesis (HAM/TSP), HTLV-I uveitis, and HTLV-I-associated infective dermatitis. Once it was shown that there is an increased risk for developing HAM/TSP associated with blood transfusion, screening for HTLV-1 among blood banks was implemented in Japan, United States, France, and the Netherlands. This process includes detection by an enzyme immunoassay (EIA) followed by a confirmatory Western blot (WB) in which recombinant proteins specific for HTLV-I Env glycoproteins are incorporated into WB strips. HTLV-I seropositive results are defined by the presence of antibodies against either gp46 or gp62/68 (both Env protein bands) and either p19, p24, or p53 (one of the gag bands). HTLV-II seropositivity is confirmed by the presence of rgp46-II. However, numerous cases have been documented in which serum samples are reactive by EIA, but an incomplete banding pattern is displayed by subsequent confirmatory WB. Although the significance of these HTLV-I/II seroindeterminates is unclear, it may suggest a much higher incidence of exposure to HTLV-I/II than previously estimated.

  11. Characterization of excretory-secretory antigens of adult Toxocara canis by western blotting.

    PubMed

    Sudhakar, N R; Samanta, S; Sahu, Shivani; Raina, O K; Gupta, S C; Goswami, T K; Lokesh, K M; Kumar, Ashok

    2014-06-01

    Toxocara canis is one of the most common helminth worm of dogs which continues to stimulate both public health concern alongside the higher scientific interest. It may cause visceral and ocular damage in humans especially in children. The identification of specific antigens of T. canis is important so as to develop better diagnostic techniques. Excretory-secretory (ES) antigens were prepared by culturing the adult T. canis worms in RPMI 1640 medium without serum supplementation followed by ammonium sulphate precipitation. These antigens were separated using sodium dodecyl sulphate-electrophoresis (SDS-PAGE). Recovered proteins ranged from 30 to 384 kDa. The specific reactivity of the T. canis excretory-secretory (TC-ES) proteins was checked by western blotting. The immuno-reactivity of the naturally infected dog sera with the TC-ES antigens showed five bands at 43, 57,105, 139 and 175 kDa. The immuno-reactivity of the hyper immune serum raised in rabbits against TC-ES antigens was observed with ten polypeptides of 21, 25, 30, 37, 45, 50, 57, 69, 77 and 105 kDa. Common antigens band were observed at 57 and 105 KDa. These antigens merit further evaluation as candidate for use in diagnosis of toxocariasis in humans and adult dogs.

  12. Diagnosis of cutaneous tuberculosis in biopsy specimens by PCR and southern blotting.

    PubMed Central

    Quirós, E; Maroto, M C; Bettinardi, A; González, I; Piédrola, G

    1996-01-01

    AIMS: To evaluate the use of a gene amplification and hybridisation method for detecting mycobacterial nucleic acid as a possible diagnostic method for cutaneous tuberculosis infection. METHODS: Biopsy specimens from 20 patients with various skin conditions of possible tuberculous aetiology were studied. Six patients had ulcerative nodules, seven lupiform lesions, two non-necrotic granulomas, one scrofulous lichen, one impetigo, one erythematosus lesions, one warty lesions, and one suspected tuberculous lipoma. Biopsy specimens were stained using Ziehl-Neelsen stain and cultured in Lowenstein-Jensen medium. DNA was extracted and then amplified by PCR using primers specific for the Mycobacterium tuberculosis complex. Specificity was confirmed by Southern blotting. RESULTS: Of the specimens, 30% were positive for mycobacteria on staining with Ziehl-Neelsen stain, 60% were culture positive and 85% PCR positive. Only 35.2% of specimens were positive with all three techniques. A further 32.5% were both culture and PCR positive. All PCR negative samples were also negative when cultured or stained with Ziehl-Neelsen stain. Of the PCR positive specimens, 29.4% were negative when cultured or stained. CONCLUSIONS: PCR, using suitable primers, is an efficient and sensitive method for the diagnosis of cutaneous tuberculosis. PMID:8944606

  13. A simple dot-blot-Sirius red-based assay for collagen quantification.

    PubMed

    Rodríguez-Rodríguez, Pilar; Arribas, Silvia M; de Pablo, Angel Luis López; González, M Carmen; Abderrahim, Fatima; Condezo-Hoyos, Luis

    2013-08-01

    The assessment of collagen content in tissues is important in biomedical research, since this protein is altered in numerous diseases. Hydroxyproline and Sirius red based assays are the most common methods for collagen quantification. However, these procedures have some pitfalls, such as the requirement of oxygen-free medium or expensive equipment and large sample size or being unsuitable for hydrolyzed collagen, respectively. Our objective was to develop a specific, versatile, and user-friendly quantitative method applicable to small tissue samples and extracts obtained from elastin purification, therefore, suitable for simultaneous quantification of elastin. This method is based on the binding of Sirius red to collagen present in a sample immobilized on a PVDF membrane, as in the dot-blot technique, and quantified by a scanner and image analysis software. Sample loading, Sirius red concentration, temperature and incubation time, type of standard substance, albumin interference, and quantification time are optimized. The method enabled the quantification of (1) intact collagen in several rat tissue homogenates, including small resistance-sized arteries, (2) partially hydrolyzed collagen obtained from NaOH extracts, compatible with elastin purification, and (3) for the detection of differences in collagen content between hypertensive and normotensive rats. We conclude that the developed technique can be widely used since it is versatile (quantifies intact and hydrolyzed collagen), requires small sample volumes, is user-friendly (low-cost, easy to use, minimum toxic materials, and reduced time of test), and is specific (minimal interference with serum albumin).

  14. HIV-1: absence of infection in subjects with indeterminate western blot.

    PubMed

    Migali, E; Mariotti, D; Lovari, A; Tenani, T; Imperiali, P; Ozzola, G

    1993-01-01

    The search for specific antibodies against the Human Immunodeficiency Virus type-1 (Ab anti HIV 1), using an immunoenzymatic test and a subsequent confirmation test (Western Blot-WB) in patients who were previously positive or borderline at the first test, singled out from about 12500 tested subjects, fifteen patients with indeterminate WB (WBi) for the presence of an abnormal band. The presence of p24 was predominant in those WBi (about 50%); generally p24 was the only band found. In all serum samples with WBi, the viral antigen p24 (Ag p24) was absent. In 3 subjects with WBi, after at least six months from the first test, not only did the same pattern persist but also the search for VIH-1 DNA sequence using the polymerase chain reaction (PCR) gave a negative result. According to our experience and the literature on the subject, we suggest that patients with low risk of infection and whose WBi does not modify with time have a remote possibility of being infected by HIV.

  15. Genotype-specific RNA probes for direct identification of wild polioviruses by blot hybridization.

    PubMed Central

    De, L; Yang, C F; Da Silva, E; Boshell, J; Cáceres, P; Gómez, J R; Pallansch, M; Kew, O

    1997-01-01

    We have developed RNA probes for the direct identification of wild poliovirus isolates by blot hybridization. The probes are complementary to sequences of the first 30 to 32 codons of VP1, which evolve more extensively (approximately 1.5-fold) than the rest of VP1. To illustrate our general approach, we describe the design of probes specific to each of four major genotypes recently endemic (1981 to 1991) to the Americas: Andean type 1, Brazil type 1, Brazil type 3, and Central America-Mexico type 3. A wild isolate of each genotype was selected according to molecular and epidemiologic criteria to be representative of the principal lineages in circulation. Variable VP1 sequences of the representative isolates were amplified by the reverse transcriptase PCR and were inserted into a plasmid vector containing a T7 promoter. The in vitro transcripts, labeled with digoxigenin, served as probes. These formed stable hybrids only with RNAs of isolates of the corresponding genotypes. Hybrids were detected by a sensitive chemiluminescence assay, capable under normal diagnostic conditions of detecting specific wild poliovirus sequences in samples containing up to a 100-fold excess of Sabin vaccine strain-related sequences of the same serotype. PMID:9350743

  16. Digital chemiluminescence imaging of DNA sequencing blots using a charge-coupled device camera.

    PubMed Central

    Karger, A E; Weiss, R; Gesteland, R F

    1992-01-01

    Digital chemiluminescence imaging with a cryogenically cooled charge-coupled device (CCD) camera is used to visualize DNA sequencing fragments covalently bound to a blotting membrane. The detection is based on DNA hybridization with an alkaline phosphatase(AP) labeled oligodeoxyribonucleotide probe and AP triggered chemiluminescence of the substrate 3-(2'-spiro-adamantane)-4-methoxy-4-(3"-phosphoryloxy)phenyl- 1,2-dioxetane (AMPPD). The detection using a direct AP-oligonucleotide conjugate is compared to the secondary detection of biotinylated oligonucleotides with respect to their sensitivity and nonspecific binding to the nylon membrane by quantitative imaging. Using the direct oligonucleotide-AP conjugate as a hybridization probe, sub-attomol (0.5 pg of 2.7 kb pUC plasmid DNA) quantities of membrane bound DNA are detectable with 30 min CCD exposures. Detection using the biotinylated probe in combination with streptavidin-AP was found to be background limited by nonspecific binding of streptavidin-AP and the oligo(biotin-11-dUTP) label in equal proportions. In contrast, the nonspecific background of AP-labeled oligonucleotide is indistinguishable from that seen with 5'-32P-label, in that respect making AP an ideal enzymatic label. The effect of hybridization time, probe concentration, and presence of luminescence enhancers on the detection of plasmid DNA were investigated. Images PMID:1480487

  17. Performance of PCR-reverse blot hybridization assay for detection of rifampicin-resistant Mycobacterium leprae.

    PubMed

    Wang, Hye-young; Kim, Hyunjung; Kim, Yeun; Bang, Hyeeun; Kim, Jong-Pill; Hwang, Joo Hwan; Cho, Sang-Nae; Kim, Tae Ue; Lee, Hyeyoung

    2015-10-01

    Drug resistance in Mycobacterium leprae is a significant problem in countries where leprosy is endemic. A sensitive, specific, and high-throughput reverse blot hybridization assay (REBA) for the detection of genotypic resistance to rifampicin (RIF) was designed and evaluated. It has been shown that resistance to RIF in M. leprae involves mutations in the rpoB gene encoding the -subunit of the RNA polymerase. The PCR-REBA simultaneously detects both 6 wild-type regions and 5 different mutations (507 AGC, 513 GTG, 516 TAT, 531 ATG, and 531 TTC) including the most prevalent mutations at positions 507 and 531. Thirty-one clinical isolates provided by Korea Institute of Hansen-s Disease were analyzed by PCR-REBA with RIF resistance of rpoB gene. As a result, missense mutations at codons 507 AGC and 531 ATG with 2-nucleotide substitutions were found in one sample, and a missense mutation at codon 516 TAT and ΔWT6 (deletion of 530-534) was found in another sample. These cases were confirmed by DNA sequence analysis. This rapid, simple, and highly sensitive assay provides a practical alternative to sequencing for genotypic evaluation of RIF resistance in M. leprae.

  18. Western blot analysis of antibody response to pneumococcal protein antigens in a murine model of pneumonia.

    PubMed Central

    Mouneimne, H; Juvin, M; Beretti, J L; Azoulay-Dupuis, E; Vallee, E; Geslin, P; Petitpretz, P; Berche, P; Gaillard, J L

    1997-01-01

    To detect new antigen candidates for serological tests, we studied the antibody response to pneumococcal protein antigens in mice infected intratracheally with various Streptococcus pneumoniae strains. Sera were tested by Western blotting against whole-cell protein extracts. Mice developed a detectable immunoglobulin G-type response against a small number of polypeptides. The antibody response was strain dependent: sera from individuals infected with the same strain gave similar banding patterns on immunoblots. The banding patterns varied with the strain used for infection. However, a band at 36 to 38 kDa was recognized by all reactive sera. This band appeared to correspond to a polypeptide that was antigenically well conserved among the different S. pneumoniae serotypes. An antibody response to this antigen developed in mice irrespective of the capsular type, the virulence, and the susceptibility to penicillin G of the infecting strain. Thus, this 36- to 38-kDa protein antigen may be of value for the development of a serological test for humans. PMID:9384307

  19. Renaturation of blotted allergens increases the sensitivity of specific IgE detection.

    PubMed

    Muro, M D; Fernández, C; Moneo, I

    1996-01-01

    Several authors have demonstrated that renaturation is an essential step for the appropriate recognition of blotted proteins. The use of nonionic detergents has been described as a useful alternative to enhance the antigenicity in immunoblotting, although elution from proteins by detergents has been observed. To measure the influence of different factors on the sensitivity of specific IgE by immunoblotting, we used twenty human sera from atopic patients who were allergic or nonallergic to a common, reliable allergen (grass pollen mixture). The use of Nonidet-P40 was found to be a useful alternative for the renaturation of the allergens. No elution from the membrane was found when employing this detergent, even at high concentrations (3%), and its use gave better sensitivity than methanol. On the other hand, we detected that methanol possessed renaturing properties. A transfer method using diffusion instead of electric transfer gave the best results and two membranes could be obtained from each gel. Using this method, we found that after NP-40 incubation of the membrane, the use of bovine albumin could be omitted as blocking agent and that its use had even deleterious effects.

  20. Trypanosoma cruzi: variability of stocks from Colombia determined by molecular karyotype and minicircle Southern blot analysis.

    PubMed

    Triana, Omar; Ortiz, Sylvia; Dujardin, Jean-Claude; Solari, Aldo

    2006-05-01

    Nineteen Trypanosoma cruzi stocks, most of them of wild origin, and four Trypanosoma rangeli stocks from Colombia were analysed by molecular karyotype analysis with cloned DNA cruzipain as the probe. Another 27 cloned stocks of T. cruzi from different geographic areas of South America were used as reference for T. cruzi lineages. Phenetic analysis of chromosome size polymorphism demonstrated a great variability of Colombian T. cruzi stocks, suggesting that most belong to lineage I, although two of them belong to lineage II. The 2 lineage II T. cruzi, 17 T. cruzi lineage I, and 3 T. rangeli stocks from Colombia were studied further by Southern blot analysis with a panel of kinetoplast DNA minicircle probes. Hybridisation results indicate that the two T. cruzi II stocks are genetically distant from each other and from T. cruzi lineages IIb, IId, and IIe from Chile. Finally, T. cruzi minicircle probes do not cross-hybridise in any stringency condition tested with T. rangeli minicircles, a clear indication that these parasites can be easily distinguished by this method.

  1. Development of a dot blot assay using gene probes for the detection of enteroviruses in water

    SciTech Connect

    Margolin, A.B.

    1986-01-01

    Enteric viruses are viruses which replicate in the intestinal tract of man and animals. One mode of transmission for enteric viruses is the fecal-oral route. Drinking water which has been contaminated with sewage or sewage effluent has been implicated as a means for the spread of enteric viruses. Current methods for the detection of enteric viruses in water requires the use of animal cell culture. This technique has several drawbacks. More rapid techniques, such as fluorescent antibody or radioimmunoassay do not have the needed sensitivity to detect the low levels of virus found in contaminated water. An alternative technique for the detection of viruses in water was sought. Recent advances in recombinant DNA technology now makes it possible to detect viruses without the use of cell culture or antibodies. Gene probes that hybridize to the RNA of poliovirus and hepatitis A virus were tested for their ability to detect different enteric viruses. The probes were labeled with /sup 32/P dCTP and /sup 32/P dATP to a specific activity greater then 1.0 x 10/sup 9/ cpm/ug DNA. One infectious unit of poliovirus and hepatitis A virus was detected using labeled cDNA probes. Upon comparison, the dot blot assay was as sensitive as tissue culture for the detection of poliovirus in beef extract, secondary effluent, and tap water. Environmental samples, such as secondary effluent, reclaimed wastewater and unchlorinated drinking water were also assayed for poliovirus and hepatitis A virus with the use of gene probes. The results presented here offer an alternative method for screening water samples for the presence of enteric viruses.

  2. Evaluation of the Aspergillus Western Blot IgG Kit for Diagnosis of Chronic Aspergillosis

    PubMed Central

    Oliva, A.; Flori, P.; Hennequin, C.; Dubus, J.-C.; Reynaud-Gaubert, M.; Charpin, D.; Vergnon, J. M.; Gay, P.; Colly, A.; Piarroux, R.; Pelloux, H.

    2014-01-01

    Immunoprecipitin detection (IPD) is the current reference confirmatory technique for anti-Aspergillus antibody detection; however, the lack of standardization is a critical drawback of this assay. In this study, we evaluated the performance of the Aspergillus Western blot (Asp-WB) IgG kit (LDBio Diagnostics, Lyon, France), a recently commercialized immunoblot assay for the diagnosis of various clinical presentations of chronic aspergillosis. Three hundred eight serum samples from 158 patients with aspergillosis sensu lato (s.l.) were analyzed. More specifically, 267 serum samples were derived from patients with Aspergillus disease, including 89 cases of chronic pulmonary aspergillosis, 10 of aspergilloma, and 32 of allergic bronchopulmonary aspergillosis, while 41 samples were from patients with Aspergillus colonization, including 15 cystic fibrosis (CF) and 12 non-CF patients. For blood donor controls, the Asp-WB specificity was 94%, while the kit displayed a sensitivity for the aspergillosis s.l. diagnosis of 88.6%, with a diagnostic odds ratio (DOR) of 119 (95% confidence interval [CI], 57 to 251). The DOR values were 185.22 (95% CI,78.79 to 435.45) and 43.74 (95% CI, 15.65 to 122.20) for the diagnosis of Aspergillus disease and Aspergillus colonization, respectively. Among the patients, the sensitivities of the Asp-WB in the diagnosis of Aspergillus colonization were 100% and 41.7% in CF and non-CF patients, respectively. The Asp-WB yielded fewer false-negative results than did IPD. In conclusion, the Asp-WB kit performed well for the diagnosis of various clinical presentations of aspergillosis in nonimmunocompromised patients, with an enhanced standardization and a higher sensitivity than with IPD, which is the current reference method. PMID:25392351

  3. Evaluation of a Western Blot Test in an Outbreak of Acute Pulmonary Histoplasmosis

    PubMed Central

    Pizzini, Claudia V.; Zancopé-Oliveira, Rosely M.; Reiss, Errol; Hajjeh, Rana; Kaufman, Leo; Peralta, José Mauro

    1999-01-01

    A western blot (WB) test was evaluated for detection of antibodies against native glycosylated and chemically deglycosylated M and H antigens of Histoplasma capsulatum in serum obtained from patients during the acute phase of pulmonary histoplasmosis that occurred during an outbreak. Of 275 serum samples tested by immunodiffusion and complement fixation (CF) samples from 40 patients affected during this outbreak and from 37 negative controls were tested by WB test. A group of patients whose sera were negative for CF antibodies and precipitins early in the acute stage of histoplasmosis but who all seroconverted during convalescence 6 weeks later were tested with the WB test. Antibodies against untreated H and M antigens were detected at a 1:100 dilution by WB test in 45% of the 20 acute-phase serum samples and in all 20 of the convalescent-phase specimens. The WB test’s sensitivity for acute-phase specimens increased to 90% (18 of 20 specimens) when H and M antigens were treated by periodate oxidation to inactivate susceptible carbohydrate epitopes. When native glycosylated antigens were used in the WB test, positive reactions were observed in negative control serum specimens (3 of 37 specimens; 8%) and in serum specimens obtained from asymptomatic persons screened as part of the outbreak investigation (13 of 20 specimens; 65%). These positive reactions were also attributed to glycosidic epitopes since the specificity of the WB test increased from 78 to 100% when periodate-treated H and M antigens were used. WB test with deglycosylated H and M antigens of histoplasmin provides a rapid, sensitive, and specific test to diagnose acute pulmonary histoplasmosis before precipitins can be detected. PMID:9874658

  4. Evaluation of the Aspergillus Western blot IgG kit for diagnosis of chronic aspergillosis.

    PubMed

    Oliva, A; Flori, P; Hennequin, C; Dubus, J-C; Reynaud-Gaubert, M; Charpin, D; Vergnon, J M; Gay, P; Colly, A; Piarroux, R; Pelloux, H; Ranque, S

    2015-01-01

    Immunoprecipitin detection (IPD) is the current reference confirmatory technique for anti-Aspergillus antibody detection; however, the lack of standardization is a critical drawback of this assay. In this study, we evaluated the performance of the Aspergillus Western blot (Asp-WB) IgG kit (LDBio Diagnostics, Lyon, France), a recently commercialized immunoblot assay for the diagnosis of various clinical presentations of chronic aspergillosis. Three hundred eight serum samples from 158 patients with aspergillosis sensu lato (s.l.) were analyzed. More specifically, 267 serum samples were derived from patients with Aspergillus disease, including 89 cases of chronic pulmonary aspergillosis, 10 of aspergilloma, and 32 of allergic bronchopulmonary aspergillosis, while 41 samples were from patients with Aspergillus colonization, including 15 cystic fibrosis (CF) and 12 non-CF patients. For blood donor controls, the Asp-WB specificity was 94%, while the kit displayed a sensitivity for the aspergillosis s.l. diagnosis of 88.6%, with a diagnostic odds ratio (DOR) of 119 (95% confidence interval [CI], 57 to 251). The DOR values were 185.22 (95% CI,78.79 to 435.45) and 43.74 (95% CI, 15.65 to 122.20) for the diagnosis of Aspergillus disease and Aspergillus colonization, respectively. Among the patients, the sensitivities of the Asp-WB in the diagnosis of Aspergillus colonization were 100% and 41.7% in CF and non-CF patients, respectively. The Asp-WB yielded fewer false-negative results than did IPD. In conclusion, the Asp-WB kit performed well for the diagnosis of various clinical presentations of aspergillosis in nonimmunocompromised patients, with an enhanced standardization and a higher sensitivity than with IPD, which is the current reference method.

  5. Electrostatic protein immobilization using charged polyacrylamide gels and cationic detergent microfluidic Western blotting.

    PubMed

    Kim, Dohyun; Karns, Kelly; Tia, Samuel Q; He, Mei; Herr, Amy E

    2012-03-06

    We report a novel protein immobilization matrix for fully integrated microfluidic Western blotting (WB). The electrostatic immobilization gel (EIG) enables immobilization of all proteins sized using cetyl trimethylammonium bromide polyacrylamide gel electrophoresis (CTAB-PAGE), for subsequent electrophoretic probing with detection affinity reagents (e.g., labeled antibodies). The "pan-analyte" capture strategy introduced here uses polyacrylamide gel grafted with concentrated point charges (zwitterionic macromolecules), in contrast to existing microfluidic WB strategies that rely on a sandwich immunoassay format for analyte immobilization and detection. Sandwich approaches limit analyte immobilization to capture of only a priori known targets. A charge interaction mechanism study supports the hypothesis that electrostatic interaction plays a major role in analyte immobilization on the EIG. We note that protein capture efficiency depends on both the concentration of copolymerized charges and ionic strength of the gel buffer. We demonstrate pan-analyte immobilization of sized CTAB-laden model proteins (protein G, ovalbumin, bovine serum albumin, β-galactosidase, lactoferrin) on the EIG with initial capture efficiencies ranging from 21 to 100%. Target proteins fixed on the EIG (protein G, lactoferrin) are detected using antibody probes with signal-to-noise ratios of 34 to 275. The approach advances protein immunoblotting performance through 200× reduction on sample consumption, 12× reduction in assay duration, and automated assay operation, compared to slab-gel WB. Using the microfluidic WB assay, assessment of lactoferrin in human tear fluid is demonstrated with a goal of advancing toward nonbiopsy-based diagnosis of Sjögren's Syndrome, an autoimmune disease.

  6. Antibody responses to Borrelia burgdorferi detected by western blot vary geographically in Canada

    PubMed Central

    Ogden, Nicholas H.; Arsenault, Julie; Hatchette, Todd F.; Mechai, Samir; Lindsay, L. Robbin

    2017-01-01

    Lyme disease is emerging in eastern and central Canada, and most cases are diagnosed using the two-tier serological test (Enzyme Immuno Assay [EIA] followed by Western blot [WB]). Simplification of this algorithm would be advantageous unless it impacts test performance. In this study, accuracy of individual proteins of the IgG WB algorithm in predicting the overall test result in samples from Canadians was assessed. Because Borrelia burgdorferi strains vary geographically in Canada, geographic variations in serological responses were also explored. Metrics of relative sensitivity, specificity and the kappa statistic measure of concordance were used to assess the capacity of responses to individual proteins to predict the overall IgG WB result of 2524 EIA (C6)-positive samples from across Canada. Geographic and interannual variations in proportions of samples testing positive were explored by logistic regression. No one protein was highly concordant with the IgG WB result. Significant variations were found amongst years and geographic regions in the prevalence of samples testing positive using the overall IgG WB algorithm, and for individual proteins of the algorithm. In most cases the prevalence of samples testing positive were highest in Nova Scotia, and lower in samples from Manitoba westwards. These findings suggest that the current two tier test may not be simplified and continued use of the current two-tier test method and interpretation is recommended. Geographic and interannual variations in the prevalence of samples testing positive may be consistent with B. burgdorferi strain variation in Canada, and further studies are needed to explore this. PMID:28182723

  7. Evaluation of immunodominant proteins of Mycobacterium avium paratuberculosis cell wall by Western blot analysis.

    PubMed

    Hashemi, Maryam; Madani, Rasool; Razmi, Nematollah

    2014-04-01

    Mycobacterium avium subspecies paratuberculosis (M. paratuberculosis) is a slow growing mycobactin, whose dependence on mycobacterial species is known to be the causative agent of Johne's disease (paratuberculosis) in all species of domestic ruminants worldwide. The organism is transmitted via close contact, ingestion, or transplacentally from mother to fetus and occurs commonly in grazing domestic animals. Johne's disease (JD) is characterized by gradual weight loss, decreased milk production, and diarrhea due to the chronic, progressive, granulomatous enteritis and lymphadenitis. The disease can cause serious economic damage to the dairy industry due to the loss of milk production and early culling of infected animals. In recent years, researchers have focused on the identification of a specific antigen of M. paratuberculosis to use in diagnosis test and preparation of effective vaccine. The goal of this study is evaluation of the immunodominant proteins of M. paratuberculosis cell wall. The amount of protein was determined with a Lowry assay (22.68 μg/100 μL). For production of polyclonal antibody against proteins of M. paratuberculosis cell wall, a New Zealand white rabbit was immunized with antigen and Freund's adjuvant. After immunization, the rabbit was bled to produce enriched serum. Antibodies were purified from serum with ion exchange chromatography. In the Ouchterlony test, the reactions between antigen and antibodies were seen in dilutions of one quarter for serum, one quarter for Ig, and one half for IgG by clear precipitation lines due to the well immunization of the rabbit. Electrophoresis and Western blot analysis were used and subsequently a sharp band appeared in nitrocellulose paper; these bands were about 25, 37, 50, 75, and 150 kDa molecular weight, which indicated immunodominant proteins.

  8. A rapid chemiluminescent slot blot immunoassay for the detection and quantification of Clostridium botulinum neurotoxin type E, in cultures.

    PubMed

    Cadieux, Brigitte; Blanchfield, Burke; Smith, James P; Austin, John W

    2005-05-01

    A simple, rapid, cost-effective in vitro slot blot immunoassay was developed for the detection and quantification of botulinum neurotoxin type E (BoNT/E) in cultures. Culture supernatants of 36 strains of clostridia, including 12 strains of Clostridium botulinum type E, 12 strains of other C. botulinum neurotoxin serotypes, and 12 strains of other clostridial species were tested. Samples containing BoNT/E were detected using affinity-purified polyclonal rabbit antisera prepared against BoNT/E with subsequent detection of secondary antibodies using chemiluminescence. All strains of C. botulinum type E tested positive, while all non C. botulinum type E strains tested negative. The sensitivity of the slot blot immunoassay for detection of BoNT/E was approximately four mouse lethal doses (MLD). The intensity of chemiluminescence was directly correlated with the concentration of BoNT/E up to 128 MLD, allowing quantification of BoNT/E between 4 and 128 MLD. The slot blot immunoassay was compared to the mouse bioassay for detection of BoNT/E using cultures derived from fish samples inoculated with C. botulinum type E, and cultures derived from naturally contaminated environmental samples. A total of 120 primary enrichment cultures derived from fish samples, of which 103 were inoculated with C. botulinum type E, and 17 were uninoculated controls, were assayed. Of the 103 primary enrichment cultures derived from inoculated fish samples, all were positive by mouse bioassay, while 94 were also positive by slot blot immunoassay, resulting in a 7.5% false-negative rate. All 17 primary enrichment cultures derived from the uninoculated fish samples were negative by both mouse bioassay and slot blot immunoassay. A total of twenty-six primary enrichment cultures derived from environmental samples were tested by mouse bioassay and slot blot immunoassay. Of 13 primary enrichment cultures positive by mouse bioassay, 12 were also positive by slot blot immunoassay, resulting in a 3

  9. Application of Western blot analysis for the diagnosis of Encephalitozoon cuniculi infection in rabbits: example of a quantitative approach.

    PubMed

    Desoubeaux, Guillaume; Pantin, Ana; Peschke, Roman; Joachim, Anja; Cray, Carolyn

    2017-02-01

    Diagnosis of Encephalitozoon cuniculi infection in rabbits remains a major veterinary issue. ELISA or immunofluorescence assays are the current reference standards of serological tests. However, these conventional techniques suffer from a lack of accuracy for distinguishing active from past infections, as a positive serostatus is common in clinically normal rabbits. In this study, we assessed the diagnostic performance of Western blot (WB) to detect both anti-E. cuniculi immunoglobulin G (IgG) and immunoglobulin M (IgM) in comparison with ELISA and to address the intensity of the immune response through a quantitative approach. Positive WB results were highly correlated with the E. cuniculi-related diseased status (P < 0.0001). Although it was more labor intensive and less standardized, quantitative WB provided detailed comparable analysis regarding the humoral response and diagnostic performance similar to ELISA testing with statistically higher sensitivity (88.4 vs. 76.1% for IgG detection and 84.3 vs. 70.4% for IgM, P < 0.01). Several specific WB bands were shown to be significantly associated with concomitant clinical signs, like the one located at 50 kDa (OR = 8.2, [2.4-27.7], P = 0.0008) for IgG and (OR = 27.9, [4.2-187.9], P = 0.0006) for IgM. Therefore, the quantitative WB may have application in veterinary diagnostic laboratories to increase the accuracy of the clinical diagnosis of E. cuniculi infection. In addition, this tool may help to further understand the development and function of the humoral immune response to this infectious agent.

  10. Deep proteomic profiling of vasopressin-sensitive collecting duct cells. I. Virtual Western blots and molecular weight distributions.

    PubMed

    Yang, Chin-Rang; Tongyoo, Pumipat; Emamian, Milad; Sandoval, Pablo C; Raghuram, Viswanathan; Knepper, Mark A

    2015-12-15

    The mouse mpkCCD cell line is a continuous cultured epithelial cell line with characteristics of renal collecting duct principal cells. This line is widely used to study epithelial transport and its regulation. To provide a data resource useful for experimental design and interpretation in studies using mpkCCD cells, we have carried out "deep" proteomic profiling of these cells using three levels of fractionation (differential centrifugation, SDS-PAGE, and HPLC) followed by tandem mass spectrometry to identify and quantify proteins. The analysis of all resulting samples generated 34.6 gigabytes of spectral data. As a result, we identified 6,766 proteins in mpkCCD cells at a high level of stringency. These proteins are expressed over eight orders of magnitude of protein abundance. The data are provided to users as a public data base (https://helixweb.nih.gov/ESBL/Database/mpkFractions/). The mass spectrometry data were mapped back to their gel slices to generate "virtual Western blots" for each protein. For most of the 6,766 proteins, the apparent molecular weight from SDS-PAGE agreed closely with the calculated molecular weight. However, a substantial fraction (>15%) of proteins was found to run aberrantly, with much higher or much lower mobilities than predicted. These proteins were analyzed to identify mechanisms responsible for altered mobility on SDS-PAGE, including high or low isoelectric point, high or low hydrophobicity, physiological cleavage, residence in the lysosome, posttranslational modifications, and expression of alternative isoforms due to alternative exon usage. Additionally, this analysis identified a previously unrecognized isoform of aquaporin-2 with apparent molecular mass <20 kDa.

  11. Qualitative and quantitative evaluation of Simon™, a new CE-based automated Western blot system as applied to vaccine development.

    PubMed

    Rustandi, Richard R; Loughney, John W; Hamm, Melissa; Hamm, Christopher; Lancaster, Catherine; Mach, Anna; Ha, Sha

    2012-09-01

    Many CE-based technologies such as imaged capillary IEF, CE-SDS, CZE, and MEKC are well established for analyzing proteins, viruses, or other biomolecules such as polysaccharides. For example, imaged capillary isoelectric focusing (charge-based protein separation) and CE-SDS (size-based protein separation) are standard replacement methods in biopharmaceutical industries for tedious and labor intensive IEF and SDS-PAGE methods, respectively. Another important analytical tool for protein characterization is a Western blot, where after size-based separation in SDS-PAGE the proteins are transferred to a membrane and blotted with specific monoclonal or polyclonal antibodies. Western blotting analysis is applied in many areas such as biomarker research, therapeutic target identification, and vaccine development. Currently, the procedure is very manual, laborious, and time consuming. Here, we evaluate a new technology called Simple Western™ (or Simon™) for performing automated Western analysis. This new technology is based on CE-SDS where the separated proteins are attached to the wall of capillary by a proprietary photo activated chemical crosslink. Subsequent blotting is done automatically by incubating and washing the capillary with primary and secondary antibodies conjugated with horseradish peroxidase and detected with chemiluminescence. Typically, Western blots are not quantitative, hence we also evaluated the quantitative aspect of this new technology. We demonstrate that Simon™ can quantitate specific components in one of our vaccine candidates and it provides good reproducibility and intermediate precision with CV <10%.

  12. Double-blotting: a solution to the problem of non-specific binding of secondary antibodies in immunoblotting procedures.

    PubMed

    Lasne, F

    2001-07-01

    "Double-blotting" (DB) was developed to overcome the problem of non-specific binding of secondary antibodies in immunoblotting (IB). After it had been probed by the primary antibody, the membrane with the blotted proteins was assembled with a second blank membrane and submitted to a second blotting under acidic conditions. The primary antibody molecules were thus desorbed from their corresponding antigen and transferred onto the second membrane, whereas the antigen and the interfering proteins remained bound to the first one. The second membrane could then be probed by the secondary antibodies without the risk of non-specific binding. This method was developed for the study of erythropoietin (EPO) in concentrated urine since a strong non-specific binding of biotinylated secondary antibodies to some urinary proteins had been observed using classical IB protocols.

  13. Double-blotting: a solution to the problem of nonspecific binding of secondary antibodies in immunoblotting procedures.

    PubMed

    Lasne, Françoise

    2003-05-01

    "Double-blotting" (DB) has been developed to overcome the problem of nonspecific binding of secondary antibodies in immunoblotting (IB). After it has been probed by the primary antibody, the membrane with the blotted proteins is assembled with a second blank membrane and submitted to a second blotting under acidic conditions. The primary antibody molecules are thus desorbed from their corresponding antigen and transferred onto the second membrane, whereas the antigen and the interfering proteins remain bound to the first one. The second membrane can then be probed by the secondary antibodies without the risk of nonspecific binding. This method has been developed for the study of erythropoietin (EPO) in concentrated urine since a strong nonspecific binding of biotinylated secondary antibodies to some urinary proteins is observed using classical IB protocols. However, its concept makes it usable in other applications that come up against this kind of problem.

  14. Product-selective blot: a technique for measuring enzyme activities in large numbers of samples and in native electrophoresis gels

    SciTech Connect

    Thompson, G.A.; Davies, H.M.; McDonald, N.

    1985-08-01

    A method termed product-selective blotting has been developed for screening large numbers of samples for enzyme activity. The technique is particularly well suited to detection of enzymes in native electrophoresis gels. The principle of the method was demonstrated by blotting samples from glutaminase or glutamate synthase reactions into an agarose gel embedded with ion-exchange resin under conditions favoring binding of product (glutamate) over substrates and other substances in the reaction mixture. After washes to remove these unbound substances, the product was measured using either fluorometric staining or radiometric techniques. Glutaminase activity in native electrophoresis gels was visualized by a related procedure in which substrates and products from reactions run in the electrophoresis gel were blotted directly into a resin-containing image gel. Considering the selective-binding materials available for use in the image gel, along with the possible detection systems, this method has potentially broad application.

  15. Detection of enterotoxigenic Clostridium perfringens in spices used in Mexico by dot blotting using a DNA probe.

    PubMed

    Rodríguez-Romo, L A; Heredia, N L; Labbé, R G; García-Alvarado, J S

    1998-02-01

    Several reports on the microbiology of spices and herbs indicate the presence of Clostridium perfringens, a spore-forming foodborne pathogen responsible for gastrointestinal disease. In the present study, a total of 380 samples of spices and herbs (cumin seed, black pepper, oregano, garlic powder, and bay leaves) widely used in Mexico were analyzed for the presence of C. perfringens, and the enterotoxigenicity of the isolates was determined by a dot-blot technique using an enterotoxin degoxigenin-labeled DNA probe. C. perfringens counts varied from <100 to 433 CFU/g in garlic powder, from <100 to 200 CFU/g in black pepper, from <100 to 433 CFU/g in cumin seed, from <100 to 340 CFU/g in oregano, and from < 100 to 450 CFU/g in bay leaves. The dot-blot technique detected the enterotoxin gene in 8 (4.25%) of 188 confirmed isolates of C. perfringens. dot-blot.

  16. Stain-free detection as loading control alternative to Ponceau and housekeeping protein immunodetection in Western blotting.

    PubMed

    Rivero-Gutiérrez, B; Anzola, A; Martínez-Augustin, O; de Medina, F Sánchez

    2014-12-15

    It is currently a routine practice to require a measurement of a housekeeping reference, including actin, glyceraldehyde-3-phosphate dehydrogenase, β-tubulin, among others, in Western blots, as it is the rule in RNA blots. Reversible Ponceau staining has been applied successfully to check equal loading of gels. Here we test a new technique, with the Stain-Free gels from Bio-Rad, against both Ponceau staining and housekeeping protein immunodetection under different conditions. Our results show that Stain-Free gels outperform Ponceau staining and that both are more consistent than housekeeping proteins as a loading control.

  17. Stain-Free total protein staining is a superior loading control to β-actin for Western blots.

    PubMed

    Gilda, Jennifer E; Gomes, Aldrin V

    2013-09-15

    Semi-quantification of proteins using Western blots typically involves normalization against housekeeping genes such as β-actin. More recently, Ponceau S and Coomassie blue staining have both been shown to be suitable alternatives to housekeeping genes as loading controls. Stain-Free total protein staining offers the advantage of no staining or destaining steps. Evaluation of the use of Stain-Free staining as an alternative to β-actin or the protein stain Ponceau S showed that Stain-Free staining was superior to β-actin and as good as or better than Ponceau S staining as a loading control for Western blots.

  18. Enzyme-linked immunoelectrotransfer blot analysis of a cryptosporidiosis outbreak on a United States Coast Guard cutter.

    PubMed

    Moss, D M; Bennett, S N; Arrowood, M J; Wahlquist, S P; Lammie, P J

    1998-01-01

    Symptoms consistent with an outbreak of cryptosporidiosis (diarrhea, vomiting, nausea, and abdominal cramps) occurred on a U.S. Coast Guard cutter within 0-18 days after the cutter filled its tanks with Milwaukee, Wisconsin city water in March 1993. At three-weeks postdocking (PD), the suspected water was removed, and serum samples and stool specimens were collected from 47 of the 58 crew members, as well as questionnaire data on their water consumption and symptoms aboard the cutter. At 10-weeks PD and/or at 28-weeks PD, additional serum specimens were collected. Intensitometric data from enzyme-linked immunoelectrotransfer blot (EITB) were obtained on IgA responses to a 17-kD antigen group, IgM responses to a 27-kD antigen group, and IgG responses to 27-, 17-, and 15-kD antigen groups extracted from oocysts. In addition, IgG responses to crude oocyst antigens were obtained by ELISA. Based on reported symptoms, EITB results, and stool examination, the crew members were classified as confirmed (10), probable (10), suspected (22), and noncases (16). Of the 10 confirmed cases (all symptomatic) and the 10 probable cases (eight symptomatic) whose stools were positive and negative, respectively, for Cryptosporidium oocysts by microscopy, all showed changes in EITB intensities to the antigen groups and were considered EITB positive. The remaining 38 crew members, 22 suspected cases (all symptomatic), and 16 noncases (all asymptomatic), if tested, had negative stool examinations and were considered EITB negative. Of the 10 confirmed cases, only four showed a significant change in IgG responses (P < 0.05) between three-weeks PD and follow-up serum specimens by ELISA. Crew members considered confirmed cases consumed significantly more water (P < or = 0.005) aboard the cutter than noncases. Crew members considered EITB positive consumed more water (P < or = 0.04) than crew members considered EITB negative while there was no significant difference in water consumption (P > or

  19. Detection of X chromosome aneuploidy using Southern blot analysis during routine population-based screening for fragile X syndrome.

    PubMed

    Adir, Vardit; Shahak, Elena; Dar, Hanna; Borochowitz, Zvi U

    2003-01-01

    We report herein two cases where detection of X chromosome aneuploidy (cytogenetically proved 45,X/46XX and 47,XXX) was made possible by molecular diagnosis during population-based carrier screening for Fragile X syndrome, using Southern blot analysis. This study emphasizes the value of molecular analysis for gene dosage to suggest chromosomal aneuploidy.

  20. Enrichment of PrPSc in Formalin Fixed Paraffin Embedded Tissues Prior to Analysis by Western Blot

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Diagnosis of prion disease is primarily through immunodetection of the infectious agent. Typically, 2 distinct procedures are recommended for a definitive diagnosis with immunohistochemistry and Western blot providing the most information as to the specific isolate in question. In the past these app...

  1. Human T-cell lymphotropic virus type I (HTLV-I) confirmed by PCR-Southern blot and sequencing analysis in a woman with tropical spastic paraparesis.

    PubMed

    Perandin, F; Cariani, A; Bonfanti, C; Trainini, L; Magoni, M; Manca, N

    2006-09-01

    Human T-cell lymphotropic virus type I (HTLV-I) is a human retrovirus and the aetiological agent of a progressive neurological disease called tropical spastic paraparesis/HTLV-I-associated myelopathy (TSP/HAM), as confirmed by evidence accumulated in HTLV-I seroprevalence studies. TSP/HAM is rarely diagnosed in Italy, given the low prevalence of HTLV-I in the population. TSP/HAM begins insidiously in the fourth decade, mainly with spastic paraparesis of the lower extremities and positive Babinski reflex, as well as interfering with bowel and bladder functions. In this study we report the clinical, virological and haemato chemical data of a 54-year-old woman, born in the Ivory Cost, with symptoms suggestive of TSP. The presence of HTLV-I infection was demonstrated by the detection of antibodies in serum and in cerebrospinal fluid by immunoenzymatic assay and Western blot analysis. In addition, viral isolation was carried out in peripheral blood cells, and the presence of HTLV-I proviral DNA was confirmed by polymerase chain reaction/Southern blot and sequencing analysis. According to our results, HTLV-I testing might be useful when TSP/HAM is suspected.

  2. Positively charged polymer brush-functionalized filter paper for DNA sequence determination following Dot blot hybridization employing a pyrrolidinyl peptide nucleic acid probe.

    PubMed

    Laopa, Praethong S; Vilaivan, Tirayut; Hoven, Voravee P

    2013-01-07

    As inspired by the Dot blot analysis, a well known technique in molecular biology and genetics for detecting biomolecules, a new paper-based platform for colorimetric detection of specific DNA sequences employing peptide nucleic acid (PNA) as a probe has been developed. In this particular study, a pyrrolidinyl PNA bearing a conformationally rigid d-prolyl-2-aminocyclopentanecarboxylic acid backbone (acpcPNA) was used as a probe. The filter paper was modified to be positively charged with grafted polymer brushes of quaternized poly(dimethylamino)ethyl methacrylate (QPDMAEMA) prepared by surface-initiated polymerization of 2-(dimethylamino)ethyl methacrylate from the filter paper via ARGET ATRP followed by quaternization with methyl iodide. Following the Dot blot format, a DNA target was first immobilized via electrostatic interactions between the positive charges of the QPDMAEMA brushes and negative charges of the phosphate backbone of DNA. Upon hybridization with the biotinylated pyrrolidinyl peptide nucleic acid (b-PNA) probe, the immobilized DNA can be detected by naked eye observation of the yellow product generated by the enzymatic reaction employing HRP-labeled streptavidin. It has been demonstrated that this newly developed assay was capable of discriminating between complementary and single base mismatch targets at a detection limit of at least 10 fmol. In addition, the QPDMAEMA-grafted filter paper exhibited a superior performance to the commercial membranes, namely Nylon 66 and nitrocellulose.

  3. A mass screening survey of cystic echinococcosis by ultrasonography, Western blotting, and ELISA among university students in Manisa, Turkey.

    PubMed

    Kilimcioğlu, Ali Ahmet; Girginkardeşler, Nogay; Korkmaz, Metin; Özkol, Mine; Düzgün, Fatih; Östan, Ipek; Pabuşcu, Yüksel; Dinç, Gönül; Ok, Ulgen Zeki

    2013-12-01

    Cystic echinococcosis (CE) is one of the most important zoonotic diseases in a wide geographic area, including Turkey. In the present project, a total of 4275 students from Celal Bayar University, Manisa, Turkey, were screened by ultrasonography (US) and specific antibodies for CE were examined by Western blotting (WB) and ELISA in finger prick blood samples of 2034 of 4275 volunteered students. We aimed to report the apparent prevalence of CE based on different diagnostic procedures and to compare WB and ELISA with US in diagnosis of CE in a mass screening setting. Six new cases were diagnosed as CE by US during the survey. In addition to these cases, three students were also detected to have been previously operated and pathologically confirmed for hepatic CE. US revealed parenchymal changes in these cases in concordance with their operation history; so, the prevalence of CE by US was calculated as 0.21% (9/4275) (95%CI, 0.11-0.39%) among university students in Manisa. Bands were detected at 8, 28, 32, 38, 42, 47, 70 and 90kDa by WB and the cases were considered to be positive for CE when at least three of the bands were seen together. Apparent prevalence of CE by ELISA and WB were found to be 2.11% (43/2034) (95%CI, 1.57-2.83%) and 0.25% (5/2034) (95%CI, 0.10-0.57%), respectively. Of the six US positive cases, WB was positive in only one case with two cysts in the liver. All of four cases with liver involvement were positive by ELISA. The high prevalence of CE among university students in Manisa indicated that CE is a major health problem in this area of Turkey. Our results supported that WB is rather difficult and not feasible as a mass screening test and may not be effective for confirmation especially in asymptomatic cases. As a result, we recommend US to be used initially in mass screening surveys for CE followed by confirmation by ELISA for suspected cases. Further examination primarily by chest X-ray followed by computed tomography and/or magnetic

  4. Rapid Screening of the Epidermal Growth Factor Receptor Phosphosignaling Pathway via Microplate-Based Dot Blot Assays

    PubMed Central

    Cappione, Amedeo; Smith, Janet; Mabuchi, Masaharu; Nadler, Timothy

    2012-01-01

    Expression profiling on a large scale, as is the case in drug discovery, is often accomplished through use of sophisticated solid-phase protein microarrays or multiplex bead technologies. While offering both high-throughput and high-content analysis, these platforms are often too cost prohibitive or technically challenging for many research settings. Capitalizing on the favorable attributes of the standard ELISA and slot blotting techniques, we developed a modified dot blot assay that provides a simple cost-effective alternative for semiquantitative expression analysis of multiple proteins across multiple samples. Similar in protocol to an ELISA, but based in a membrane bound 96-well microplate, the assay takes advantage of vacuum filtration to expedite the tedious process of washing in between binding steps. We report on the optimization of the assay and demonstrate its use in profiling temporal changes in phosphorylation events in the well-characterized EGF-induced signaling cascade of A431 cells. PMID:22934183

  5. Detection of La/SS-B by western blot using nanogold-tagged antibodies and silver enhancement.

    PubMed

    Maier, Shannon; Hubbell, Sherry; Scofield, R Hal

    2009-01-01

    Immunogold staining with silver enhancement is a versatile, sensitive and specific method for immunodetection of diverse protein antigens separated by sodium dodecyl sulfate polyacrylamide gel electrophoresis and transferred to nitrocellulose or polyvinylidene difluoride membranes. "Next-generation" antibodies tagged with nanogold particles have a wide scope of use including but not limited to immunohistochemistry, western blotting, electron microscopy, fluorescent activated cell sorting procedures, and cell isolation and migration studies. Herein, we describe the use of a nanogold-tagged anti-mouse IgG secondary antibody and silver enhancement methodologies coupled with antigen-specific unlabeled primary antibodies for the detection of the La/SS-B autoantigen by western blotting as a useful alternative to chemiluminescent and enzymatic detection methods.

  6. Stabilization of hypoxia-inducible factor-1α in buffer containing cobalt chloride for Western blot analysis.

    PubMed

    Srinivasan, Sathyanarayanan; Dunn, Jeff F

    2011-09-01

    Hypoxia-inducible factor-1α (HIF-1α) is a widely studied protein with significant biomedical impact. Care is needed to stabilize HIF-1α protein during sample preparation for Western blot analysis due to its rapid degradation in the presence of oxygen. Enzyme inhibitor cocktails can be complex and expensive. We present a protease inhibitor-free buffer, containing cobalt chloride, which is effective at stabilizing HIF-1α, while being inexpensive, straightforward, and convenient, and has potential for widespread application.

  7. Differential detection of cytoplasmic Wilms tumor 1 expression by immunohistochemistry, western blotting and mRNA quantification.

    PubMed

    Maki, Takehiro; Ikeda, Hiroaki; Kuroda, Aki; Kyogoku, Noriaki; Yamamura, Yoshiyuki; Tabata, Yukiko; Abiko, Takehiro; Tsuchikawa, Takahiro; Hida, Yasuhiro; Shichinohe, Toshiaki; Tanaka, Eiichi; Kaga, Kichizo; Hatanaka, Kanako; Matsuno, Yoshihiro; Imai, Naoko; Hirano, Satoshi

    2017-01-01

    Wilms tumor 1 (WT1) is considered to be a promising target of cancer treatment because it has been reported to be frequently expressed at high levels in various malignancies. Although WT1-targeted cancer treatment has been initiated, conclusive detection methods for WT1 are not established. The present study aimed to consolidate immunohistochemistry for WT1 with statistical basis. Transfected cells with forced WT1 expression yielded specific western blot bands and nuclear immunostaining; cytoplasmic immunostaining was not specifically recognized. Immunohistochemistry, western blotting, and quantitative reverse transcriptase-polymerase chain reaction were performed in 35 human cell lines using multiple WT1 antibodies and their results were quantified. Relationships among the quantified results were statistically analyzed; the nuclear immunostaining positively correlated with western blot bands and mRNA expression levels, whereas cytoplasmic immunostaining did not. These results indicate that nuclear immunostaining reflects WT1 expression but cytoplasmic immunostaining does not. The nuclear immunostaining was barely (3/541) observed in primary cancer of esophagus, bile duct, pancreas and lung. Although the present study has some limitations, the results indicate that the cytoplasmic immunostaining does not correlate with actual WT1 expression and prompts researchers to carefully evaluate target molecule expression in treatment of cancer.

  8. Development of PCR/dot blot assay for specific detection and differentiation of taeniid cestode eggs in canids.

    PubMed

    Armua-Fernandez, Maria Teresa; Nonaka, Nariaki; Sakurai, Tatsuya; Nakamura, Seita; Gottstein, Bruno; Deplazes, Peter; Phiri, Isaac G K; Katakura, Ken; Oku, Yuzaburo

    2011-01-01

    We report the development of a colourimetric PCR/dot blot assay targeting the mitochondrial gene NADH dehydrogenase subunit 1 (nad1) for differential diagnosis of taeniid eggs. Partial sequences of the cestode nad1 gene were aligned and new primers were designed based on conserved regions. Species-specific oligonucleotide probes (S-SONP) for canine taeniid cestodes were then designed manually based on the variable region between the conserved primers. Specifically, S-SONP were designed for the Taenia crassiceps, T. hydatigena, T. multiceps, T. ovis, T. taeniaeformis, Echinococcus granulosus (genotype 1), E. multilocularis and E. vogeli. Each probe showed high specificity as no cross-hybridisation with any amplified nad1 fragment was observed. We evaluated the assay using 49 taeniid egg-positive samples collected from dogs in Zambia. DNA from 5 to 10 eggs was extracted in each sample. Using the PCR/dot blot assay, the probes successfully detected PCR products from T. hydatigena in 42 samples, T. multiceps in 3 samples, and both species (mixed infection) in the remaining 4 samples. The results indicate that the PCR/dot blot assay is a reliable alternative for differential diagnosis of taeniid eggs in faecal samples.

  9. Investigation of Anti-Toxocara Antibodies in Epileptic Patients and Comparison of Two Methods: ELISA and Western Blotting

    PubMed Central

    Zibaei, Mohammad; Firoozeh, Farzaneh; Bahrami, Parviz; Sadjjadi, Seyed Mahmoud

    2013-01-01

    The relationship between Toxocara infection and epilepsy was previously demonstrated by several case-control studies and case reports. These previous studies were often based on the enzyme-linked immunosorbent assay (ELISA) using Toxocara excretory-secretory antigens, which are not specific due to cross-reactivity with other parasitic infections such as ascariasis, trichuriasis, and anisakiasis. An immunoblot analysis is highly specific and can detect low levels of Toxocara antibodies. Therefore, this assay may be useful in the identification of toxocariasis in epileptic patients. We examined patients who had epilepsy and healthy subjects for seropositivity for Toxocara infection by ELISA and Western blotting. Out of 85 epileptic patients, 10 (11.8%) and 3 (3.5%) persons exhibited Toxocara immunoglobulin G (IgG) antibodies responses by ELISA and by both techniques, respectively. Moreover, in the healthy group (n = 85), 3 (3.5%) persons were positive by ELISA, but none was detected by Western blotting. This study indicates that Toxocara infection is a risk factor for epilepsy in Iran. These findings strongly suggest the need to perform Western blotting immunodiagnosis, as well as the ELISA using Toxocara excretory-secretory antigens, to improve diagnosis of human toxocariasis in patients with epilepsy. PMID:23710354

  10. Rattlesnake Bites in Northern California

    PubMed Central

    Butner, Alfred N.

    1983-01-01

    In a series of 59 cases of rattlesnake bites at two major northern California hospitals, no deaths occurred, no amputations or fasciotomies were required and only one patient had tissue necrosis requiring a graft. Because patients are being seen in major medical facilities earlier, envenomation is encountered in earlier stages. Less specific national standards of treatment, therefore, should receive less emphasis than treatment based on the virulence of the snakes in the particular geographic region. Initial doses of antivenin given intravenously should be based on the degree of envenomation, with additional titration done for worsening symptoms. PMID:6636730

  11. Northern Trust Mines

    EPA Pesticide Factsheets

    The United States and the Navajo Nation entered into settlement agreements that provide funds to conduct investigations and any needed cleanup at 16 of the 46 priority mines, including six mines in the Northern Abandoned Uranium Mine Region.

  12. Tornadoes Strike Northern Wisconsin

    NASA Technical Reports Server (NTRS)

    2007-01-01

    A series of tornadoes ripped through the Upper Midwest region of the United States in the evening of June 7, 2007. At least five different tornadoes touched down in Wisconsin, according to the Associated Press, one of which tore through the Bear Paw Resort in northern Wisconsin. Despite dropping as much as fifteen centimeters (six inches) of rain in some places and baseball-size hail in others, authorities were reporting no deaths attributable to the storm system, and only a smattering of injuries, but considerable property damage in some areas. When the MODIS instrument on NASA's Terra satellite observed the area on June 9, 2007, the track torn through the woods by one of the tornadoes stands out quite clearly. This photo-like image uses data collected by MODIS in the normal human vision range to give a familiar natural-looking appearance. The landscape is largely a checkerboard of farms, towns, roads, and cities. The pale land is predominantly farmland where crops have not fully grown in yet. Dark blue shows the winding path of rivers and lakes dotting the landscape. The large blue lake on the east (right) side of the image is Lake Michigan. Towns and cities, including the city of Green Bay, are gray. To the north side, farmland gives way to dark green as land use shifts from agriculture to the Menominee Indian Reservation and Nicolet National Forest. The diagonal slash through the dark green forested land shows the tornado track. Bare land was revealed where the tornado tore down trees or stripped vegetation off the branches. The high-resolution image provided above is at MODIS' full spatial resolution (level of detail) of 250 meters per pixel. The MODIS Rapid Response System provides this image at additional resolutions.

  13. How to Distinguish Between the Activity of HDAC1-3 and HDAC6 with Western Blot.

    PubMed

    Beyer, Mandy; Kiweler, Nicole; Mahboobi, Siavosh; Krämer, Oliver H

    2017-01-01

    Histone deacetylases (HDACs) catalyze the deacetylation of lysine residues in their target proteins. This biochemical modification can have profound effects on the functions of these proteins and a dysregulation of HDAC activity contributes to severe diseases, including neoplastic transformation. In the following chapter, we present a strategy that allows to distinguish between the inhibition of the class I HDACs HDAC1, 2, and 3 and of the class IIb HDAC HDAC6. This method is based on Western blot and relies on the detection of hyperacetylated substrates of class I or class IIb HDACs in lysates from cells that were treated with histone deacetylase inhibitors (HDACi).

  14. Western blot analysis of virus-specific antibody responses for capripox and contagious pustular dermatitis viral infections in sheep.

    PubMed

    Chand, P; Kitching, R P; Black, D N

    1994-10-01

    This paper reports the development and evaluation of serological tests for the differentiation of antibodies in animals infected with capripox and parapox viruses. Agar-gel immunodiffusion tests using sera from sheep with naturally-acquired infections and from sheep experimentally inoculated with orf or capripox viruses showed cross reactions. Virus-specific antibody responses to structural proteins of the viruses were analysed by Western-blot analysis. This analysis readily differentiated the infections as either capripox or contagious pustular dermatitis. The antibody responses to the 32 kDa and 26 kDa proteins of capripoxvirus provided a firm basis for differentiation.

  15. Simultaneous detection of fourteen Italian cystic fibrosis mutations in seven exons by reverse dot-blot analysis.

    PubMed

    Rady, M; D'Alcamo, E; Seia, M; Iapichino, L; Ferrari, M; Russo, S; Romeo, G; Maggio, A

    1995-10-01

    The increasing number of cystic fibrosis (CF) mutations is a great obstacle to the use of DNA procedures in the detection of gene defects. We describe a fast, cheap and non-radioactive procedure, Reverse dot-blot analysis (RDB), for the simultaneous detection of CF mutations in the Italian population. We used this approach to study seven exons of the CF gene for 14 CF gene defects and were able to characterize 222 of 272 CF chromosomes (80%). The cost of the procedure was $25 per sample analysed.

  16. Detection of Rickettsia in Rhipicephalus sanguineus Ticks and Ctenocephalides felis Fleas from Southeastern Tunisia by Reverse Line Blot Assay

    PubMed Central

    Khrouf, Fatma; M'Ghirbi, Youmna; Znazen, Abir; Ben Jemaa, Mounir; Hammami, Adnene

    2014-01-01

    Ticks (n = 663) and fleas (n = 470) collected from domestic animals from southeastern Tunisia were screened for Rickettsia infection using reverse line blot assay. Evidence of spotted fever group Rickettsia was obtained. We detected Rickettsia felis in fleas, Rickettsia massiliae Bar 29 and the Rickettsia conorii Israeli spotted fever strain in ticks, and Rickettsia conorii subsp. conorii and Rickettsia spp. in both arthropods. The sensitivity of the adopted technique allowed the identification of a new association between fleas and R. conorii subsp. conorii species. The presence of these vector-borne Rickettsia infections should be considered when diagnosing this disease in humans in Tunisia. PMID:24226919

  17. Detection of Mycobacterium avium subsp. paratuberculosis by buoyant density centrifugation, sequence capture PCR and dot blot hybridisation.

    PubMed

    Halldórsdóttir, Stefania; Englund, Stina; Nilsen, Sigrun Fredsvold; Olsaker, Ingrid

    2002-07-22

    Detection of Mycobacterium avium subsp. paratuberculosis (M. paratuberculosis) by polymerase chain reaction (PCR) is often hampered by the lack of efficient methods for sample treatment. We report a protocol for analysis of faecal samples based on buoyant density centrifugation in Percoll and IS900 sequence capture PCR combined with a dot blot assay for detection of low-grade infection of M. paratuberculosis. Serial dilutions of M. paratuberculosis genomic DNA and M. paratuberculosis bacteria were used to assess the sensitivity of the method. The final evaluation was performed with spiked faecal samples, which also were analysed by culture. The presence of PCR inhibitory substances in processed faecal samples was evaluated by including a PCR internal control. By using buoyant density centrifugation, sequence capture PCR, and dot blot hybridisation, we achieved a sensitivity of 10(3)CFU (colony forming units)/g of faeces. The detection limit by culture was assessed to 10(2)CFU/g of faeces. We conclude that the described protocol is a fast and sensitive alternative to bacterial culture of faecal samples.

  18. Nitrotyrosine Density of Rabbit Urinary Bladder Muscle and Mucosa Measured via Western Blotting and 96-Well Plate Analysis.

    PubMed

    Fitzpatrick, Brittany; Schuler, Catherine; Leggett, Robert E; Levin, Robert M

    2012-01-01

    Purpose. Nitrotyrosine was quantitated in rabbit bladder muscle and mucosa using two analytical systems: Western blotting analyses and a 96-well plate quantitative analysis kit. Materials and Methods. Rabbit bladder muscle and mucosa were obtained from control rabbits. For the Western analysis, the samples were loaded into a SDS page gel and then transferred to a PVDF membrane. The optical density was measured using a Kodak Scanner. Using the 96-well plate, the samples and standards were loaded, incubated with primary and secondary antibody, washed and vacuumed with 10x wash buffer three times between each incubation period. Stop buffer was added to the plate and the results were quantified via the plate reader. Results. For both muscle and mucosa tissue, the optical density readings were linear with tissue concentration; the concentration of nitrotyrosine in the mucosa was significantly higher than in the muscle. However, whereas the Western blot analysis is based on relative optical densities, the 96-well plate kit provides a truly quantitative analysis. Discussion. Mucosa tissue displayed a higher density of nitrotyrosine than did detrusor muscle tissue. This may well be due to the significantly higher metabolic activity of the mucosa compared to the muscle.

  19. Phylogenetic distribution of apolipoproteins A-I and E in vertebrates as determined by Western blot analysis.

    PubMed

    Duggan, A E; Callard, I P

    2001-08-01

    A putative apolipoprotein E (apoE) has been identified in the HDL and VHDL fractions of the turtle. This observation is of particular interest considering apoE has been reported absent in the domestic hen (Hermier et al., '95; Biochim Biophys Acta: 105-118, 1995) and thus presumed absent in nonmammalian vertebrates altogether. As a result, partial amino acid sequencing of this protein was performed and revealed that one fragment shared 41% sequence identity to human apoE. Western blot analysis using antisera to apoE demonstrated cross-reactivity to a 34-kDa protein (putative apoE) in turtle plasma. Further investigation using anti-apoE antibody in Western blot analysis detected immunoreactive apoE in the plasma of lamprey, spiny dogfish, skate, and alligator, but not in flounder, newt or python; its absence in several species of birds was confirmed. Using anti-apoA-I antibody, apoA-I was detected in all vertebrate groups except a representative teleost (flounder). Apo-A-I antibody cross-reacted weakly with some putative apoE proteins (chicken, spiny dogfish and skate) and the reverse was true for anti-apoE, which cross-reacted with putative apoA-I in birds, reptiles, and elasmobranchs, confirming the molecular similarity and phylogenetic relatedness of these two proteins.

  20. Identification of Glioblastoma Phosphotyrosine-Containing Proteins with Two-Dimensional Western Blotting and Tandem Mass Spectrometry

    PubMed Central

    Guo, Tianyao; Wang, Xiaowei; Li, Maoyu; Yang, Haiyan; Li, Ling; Peng, Fang

    2015-01-01

    To investigate the presence of, and the potential biological roles of, protein tyrosine phosphorylation in the glioblastoma pathogenesis, two-dimensional gel electrophoresis- (2DGE-) based Western blotting coupled with liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS) analysis was used to detect and identify the phosphotyrosine immunoreaction-positive proteins in a glioblastoma tissue. MS/MS and Mascot analyses were used to determine the phosphotyrosine sites of each phosphopeptide. Protein domain and motif analysis and systems pathway analysis were used to determine the protein domains/motifs that contained phosphotyrosine residue and signal pathway networks to clarify the potential biological functions of protein tyrosine phosphorylation. A total of 24 phosphotyrosine-containing proteins were identified. Each phosphotyrosine-containing protein contained at least one tyrosine kinase phosphorylation motif and a certain structural and functional domains. Those phosphotyrosine-containing proteins were involved in the multiple signal pathway systems such as oxidative stress, stress response, and cell migration. Those data show 2DGE-based Western blotting, MS/MS, and bioinformatics are a set of effective approaches to detect and identify glioblastoma tyrosine-phosphorylated proteome and to effectively rationalize the biological roles of tyrosine phosphorylation in the glioblastoma biological systems. It provides novel insights regarding tyrosine phosphorylation and its potential role in the molecular mechanism of a glioblastoma. PMID:26090378

  1. Mutation analysis of fragile X syndrome by Southern blot, radioactive PCR, silver-stained polyacrylamide gel and DIG DNA

    SciTech Connect

    Lee, Sook-Hwan; Kim, Un-Kyung; Chung-Woong, M.S.

    1994-09-01

    Fragile X syndrome is the most common inherited form of mental retardation. In fragile X syndrome, the underlying mutation is caused by an expansion of the CTG triplet in the 5{prime} untranslated region of the FMR-1 gene located at Xq27.3 and diagnosed by methylation of the associated CpG island. This disorder becomes clinically manifested when the mutation is caused by an expansion of (CGG)n reaching a threshold of about 600bp (200 repeats). The number of inserted repeats increases through the generation. We have analyzed fragile X syndrome by 4 different methods: Southern blot, radioactive PCR, polyacrylamide gel and DIG DNA labeling/detection techniques. Southern blot and DIG DNA labeling/detection by double DNA digestion with EcoRI and EagI reveals both the presence of the mutation and the methylation status. Radioactive PCR and silver-stained polyacrylamide gel is a rapid and sensitive technique to define the unaffected carriers and NTMs, but it is difficult to amplify such a highly GC-rich sequence. Further testing in other fragile X patients is currently in progress.

  2. Applications of an Automated and Quantitative CE-Based Size and Charge Western Blot for Therapeutic Proteins and Vaccines.

    PubMed

    Rustandi, Richard R; Hamm, Melissa; Lancaster, Catherine; Loughney, John W

    2016-01-01

    Capillary Electrophoresis (CE) is a versatile and indispensable analytical tool that can be applied to characterize proteins. In recent years, labor-intensive SDS-PAGE and IEF slab gels have been replaced with CE-SDS (CGE) and CE-IEF methods, respectively, in the biopharmaceutical industry. These two CE-based methods are now an industry standard and are an expectation of the regulatory agencies for biologics characterization. Another important and traditional slab gel technique is the western blot, which detects proteins using immuno-specific reagents after SDS-PAGE separation. This technique is widely used across industrial and academic laboratories, but it is very laborious, manual, time-consuming, and only semi-quantitative. Here, we describe the applications of a relatively new CE-based western blot technology which is automated, fast, and quantitative. We have used this technology for both charge- and size-based CE westerns to analyze biotherapeutic and vaccine products. The size-based capillary western can be used for fast antibody screening, clone selection, product titer, identity, and degradation while the charge-based capillary western can be used to study product charge heterogeneity. Examples using this technology for monoclonal antibody (mAb), Enbrel, CRM197, and Clostridium difficile (C. difficile) vaccine proteins are presented here to demonstrate the utility of the capillary western techniques. Details of sample preparation and experimental conditions for each capillary western mode are described in this chapter.

  3. Diapause-specific gene expression in the northern house mosquito, Culex pipiens L., identified by suppressive subtractive hybridization

    Technology Transfer Automated Retrieval System (TEKTRAN)

    In this study we probe the molecular events underpinning diapause observed in overwintering females of Culex pipiens. Using suppressive subtractive hybridization (SSH) we have identified 40 genes that are either upregulated or downregulated during this seasonal period of dormancy. Northern blot hybr...

  4. Northern Plains 'Crater'

    NASA Technical Reports Server (NTRS)

    2004-01-01

    10 December 2004 The lower left (southwest) corner of this Mars Global Surveyor (MGS) Mars Orbiter Camera (MOC) image shows the location of a somewhat filled and buried meteor impact crater on the northern plains of Mars. The dark dots are boulders. A portion of a similar feature is seen in the upper right (northeast) corner of the image. This picture, showing landforms (including the odd mound north/northeast of the crater) that are typical of the martian northern lowland plains, was obtained as part of the MGS MOC effort to support the search for a landing site for the Phoenix Mars Scout lander. Phoenix will launch in 2007 and land on the northern plains in 2008. This image is located near 68.0oN, 227.4oW, and covers an area approximately 3 km (1.9 mi) wide. The scene is illuminated by sunlight from the lower left.

  5. Northern Plains of Mars

    NASA Technical Reports Server (NTRS)

    2004-01-01

    22 November 2004 This Mars Global Surveyor (MGS) Mars Orbiter Camera (MOC) image shows a typical view of the martian northern plains. Thousands of square kilometers of the northern middle and polar latitudes of Mars look similar to the scene in this image. In late spring and in summer, dust devils crisscross the northern plains, leaving a variety of dark streaks. The streaks do not survive from year to year, indicating their ephemeral nature. The circular features in this image, including the prominent bright circular feature near the bottom, are the locations of buried meteor impact craters. This image is located near 58.1oN, 207.6oW, and covers an area approximately 3 km (1.9 mi) wide. Sunlight illuminates the scene from the lower left.

  6. Detection of haemoparasites in cattle by reverse line blot hybridisation with a note on the distribution of ticks in Sicily.

    PubMed

    Georges, K; Loria, G R; Riili, S; Greco, A; Caracappa, S; Jongejan, F; Sparagano, O

    2001-08-31

    A reverse line blot hybridisation (RLB) of 21 oligonucleotides with polymerase chain reaction (PCR) amplified regions of 16S rRNA (Ehrlichia/Anaplasma group) or 18S rRNA (Babesia/Theileria group) genes of haemoparasites detected Theileria annulata, T. buffeli/orientalis, Babesia bovis, B. bigemina, B. divergens, Ehrlichia bovis, Anaplasma marginale, A. centrale and unknown species within the Rickettsia tribe.A very high prevalence of mixed infections was detected, which indicated that animals infected with Babesia spp. were also infected with Theileria spp. and/or Anaplasma spp. The tick distribution appeared to be seasonal with Hyalomma marginatum as the most frequently observed tick and Boophilus annulatus and Ixodes ricinus as the least frequently observed ticks. Other species identified in the 818 ticks collected during the five sampling periods between April 1998 and November 1999 included H. lusitanicum, Rhipicephalus sanguineus group, R. bursa, Dermacentor marginatus, Haemaphysalis punctata, B. annulatus and I. ricinus.

  7. Eighteen Years of Molecular Genotyping the Hemophilia Inversion Hotspot: From Southern Blot to Inverse Shifting-PCR

    PubMed Central

    Rossetti, Liliana C.; Radic, Claudia P.; Abelleyro, Miguel M.; Larripa, Irene B.; De Brasi, Carlos D.

    2011-01-01

    The factor VIII gene (F8) intron 22 inversion (Inv22) is a paradigmatic duplicon-mediated rearrangement, found in about one half of patients with severe hemophilia A worldwide. The identification of this prevalent cause of hemophilia was delayed for nine years after the F8 characterization in 1984. The aim of this review is to present the wide diversity of practical approaches that have been developed for genotyping the Inv22 (and related int22h rearrangements) since discovery in 1993. The sequence— Southern blot, long distance-PCR and inverse shifting-PCR—for Inv22 genotyping is an interesting example of scientific ingenuity and evolution in order to resolve challenging molecular diagnostic problems. PMID:22072947

  8. Western blot analysis of virus-specific antibody responses for capripox and contagious pustular dermatitis viral infections in sheep.

    PubMed Central

    Chand, P.; Kitching, R. P.; Black, D. N.

    1994-01-01

    This paper reports the development and evaluation of serological tests for the differentiation of antibodies in animals infected with capripox and parapox viruses. Agar-gel immunodiffusion tests using sera from sheep with naturally-acquired infections and from sheep experimentally inoculated with orf or capripox viruses showed cross reactions. Virus-specific antibody responses to structural proteins of the viruses were analysed by Western-blot analysis. This analysis readily differentiated the infections as either capripox or contagious pustular dermatitis. The antibody responses to the 32 kDa and 26 kDa proteins of capripoxvirus provided a firm basis for differentiation. Images Fig. 1 Fig. 2 Fig. 3 Fig. 4 PMID:7925674

  9. Lectin staining and Western blot data showing differential sialylation of nutrient-deprived cancer cells to sialic acid supplementation.

    PubMed

    Badr, Haitham A; AlSadek, Dina M M; Mathew, Mohit P; Li, Chen-Zhong; Djansugurova, Leyla B; Yarema, Kevin J; Ahmed, Hafiz

    2015-12-01

    This report provides data that are specifically related to the differential sialylation of nutrient deprived breast cancer cells to sialic acid supplementation in support of the research article entitled, "Nutrient-deprived cancer cells preferentially use sialic acid to maintain cell surface glycosylation" [1]. Particularly, breast cancer cells, when supplemented with sialic acid under nutrient deprivation, display sialylated glycans at the cell surface, but non-malignant mammary cells show sialylated glycans intracellularly. The impact of sialic acid supplementation under nutrient deprivation was demonstrated by measuring levels of expression and sialylation of two markers, EGFR1 and MUC1. This Data in Brief article complements the main manuscript by providing detailed instructions and representative results for cell-level imaging and Western blot analyses of changes in sialylation during nutrient deprivation and sialic acid supplementation. These methods can be readily generalized for the study of many types of glycosylation and various glycoprotein markers through the appropriate selection of fluorescently-labeled lectins.

  10. Standardisation of Western blotting to detect HTLV-1 antibodies synthesised in the central nervous system of HAM/TSP patients.

    PubMed

    Ribeiro, Luiz Claudio Pereira; Gonçalves, Cassia Cristina Alves; Slater, Carla Maria Sena Andrade; Carvalho, Silvia Maia Farias de; Puccioni-Sohler, Marzia

    2013-09-01

    Intrathecal synthesis of human T-lymphotropic virus type 1 (HTLV-1) antibodies (Abs) represents conclusive evidence of a specific immune response in the central nervous system of HTLV-1 associated myelopathy/tropical spastic paraparesis (HAM/TSP) patients. Western blotting (WB) for HTLV Abs in serum is a confirmatory test for HTLV-1 infection. The aim of this study was to standardise the Western blot to demonstrate the intrathecal pattern of Abs against HTLV-1 proteins in HAM/TSP patients. Paired cerebrospinal fluid (CSF) and serum samples were selected from 20 patients with definite HAM/TSP, 19 HTLV-1 seronegative patients and two HTLV-1 patients without definite HAM/TSP. The presence of reactive bands of greater intensity in the CSF compared to serum (or bands in only the CSF) indicated the intrathecal synthesis of anti-HTLV-1 Abs. All definite HAM/TSP patients presented with an intrathecal synthesis of anti-HTLV-1 Abs; these Abs were not detected in the control patients. The most frequent intrathecal targets of anti-HTLV-1 Abs were GD21, rgp46-I and p24 and, to a lesser extent, p19, p26, p28, p32, p36, p53 gp21 and gp46. The intrathecal immune response against env (GD21 and rgp46-I) and gag (p24) proteins represents the most important humoral pattern in HAM/TSP. This response may be used as a diagnostic marker, considering the frequent association of intrathecal anti-HTLV-1 Ab synthesis with HAM/TSP and the pathogenesis of this neurological disease.

  11. Murine interleukin 1 receptor. Direct identification by ligand blotting and purification to homogeneity of an interleukin 1-binding glycoprotein

    SciTech Connect

    Bird, T.A.; Gearing, A.J.; Saklatvala, J.

    1988-08-25

    Functional receptors (IL1-R) for the proinflammatory cytokine interleukin 1 (IL1) were solubilized from plasma membranes of the NOB-1 subclone of murine EL4 6.1 thymoma cells using the zwitterionic detergent 3((3-cholamidopropyl)dimethylammonio)-1-propanesulfonate (CHAPS). Membrane extracts were subjected to sodium dodecyl sulfate-polyacrylamide gel electrophoresis, transferred to nitrocellulose membranes, and ligand blotted with /sup 125/I-labeled recombinant human IL1 alpha in order to reveal proteins capable of specifically binding IL1. A single polydisperse polypeptide of Mr approximately equal to 80,000 was identified in this way, which bound IL1 alpha and IL1 beta with the same affinity as the IL1-R on intact NOB-1 cells (approximately equal to 10(-10) M). The IL1-binding polypeptide was only seen in membranes from IL1-R-bearing cells and did not react with interleukin 2, tumor necrosis factor alpha, or interferon. IL1-R was purified to apparent homogeneity from solubilized NOB-1 membranes by affinity chromatography on wheat germ agglutinin-Sepharose and IL1 alpha-Sepharose. Gel electrophoresis and silver staining of purified preparations revealed a single protein of Mr approximately equal to 80,000 which reacted positively in the ligand-blotting procedure and which we identify as the ligand-binding moiety of the murine IL1-R. Purified IL1-R exhibited the same affinity and specificity as the receptor on intact cells. The relationship of this protein to proteins identified by covalent cross-linking studies is discussed.

  12. MDR-TB Antibody Response (Western Blot) to Fractions of Isoniazid and Rifampicin Resistant Antigens of Mycobacterium tuberculosis.

    PubMed

    Hadizadeh Tasbiti, Alireza; Yari, Shamsi; Ghanei, Mostafa; Shokrgozar, Mohammad Ali; Bahrmand, Ahmadreza

    2015-12-01

    Drug-resistant TB poses a major threat to control of TB worldwide. Despite progress in the detection of Multidrug-resistant TB (MDR-TB) cases, a major diagnostic gap remains: 55% of reported TB patients estimated to have MDR-TB were not detected in 2013. MDR-TB antigens were conjugated to CNBr-activated Sepharose 4B. Specific polyclonal antibodies against MDR-TB Ags were prepared in rabbits using two boosted injections of the MDR-TB antigen. The antibodies were purified and treated with susceptible TB to remove any non-specific and cross-reactive antibodies. In the present study, comparative analysis of electrophoretic pattern of different antigens of INH/RIF-resistant TB were studied for identifying protein profiles. A RIF-resistant TB antigen was shown here to have different protein profiles from INH-resistant TB isolate. The results of Western blotting analysis showed that in the RIF- and INH-resistant antigenic fractions some bands of 14.4 and 45 kDa as immunogenic were common. Moreover, four bands of RIF-resistant TB antigen fractions (16, 19, 21, and 45 KDa) and one band of INH-resistant TB (about 26 KDa) were detected as diagnostic antigens. This study suggests that the Western blot is an accurate test to survey INH- and RIF-resistant TB antigens of M. tuberculosis infection. These findings indicate that MDR-TB diagnosis (based on Ag detection) could be useful in the identification of disease stages that precede symptomatic and microbiologically positive TB, such as subclinical and incipient TB.

  13. Imprinting mutations in Angelman syndrome detected by Southern blotting using a probe containing exon {alpha} of SNRPN

    SciTech Connect

    Beuten, J.; Sutcliffe, J.S.; Nakao, M.

    1994-09-01

    Prader-Willi syndrome (PWS) and Angelman syndrome (AS) are associated with paternal and maternal deficiencies respectively, of gene expression within human chromosome 15q11-q13, and are caused by deletion, uniparental disomy (UPD), or other mutations. The SNRPN gene maps in this region, is paternally expressed, and is a candidate gene for PWS. Southern blotting using methylation-sensitive enzymes and a genomic DNA probe from the CpG island containing exon {alpha} of the SNRPN gene reveals methylation specific for the maternal allele. In cases of the usual deletions or UPD, the probe detects absence of an unmethylated allele in PWS and absence of a methylated allele in AS. We have analyzed 21 nondeletion/nonUPD AS patients with this probe and found evidence for an imprinting mutation (absence of a methylated allele) in 3 patients. Southern blotting with methylation-sensitive enzymes using the exon {alpha} probe, like use of the PW71 probe, should detect abnormalities in all known PWS cases and in 3 of the 4 forms of AS: deletion, UPD and imprinting mutations. This analysis provides a valuable diagnostic approach for PWS and AS. In efforts to localize the imprinting mutations in AS, one patient was found with failure to inherit a dinucleotide repeat polymorphism near probe 189-1 (D15S13). Analysis of this locus in AS families and CEPH families demonstrates a polymorphism that impairs amplification and a different polymorphism involving absence of hybridization to the 189-1 probe. The functional significance, if any, of deletion of the 189-1 region is unclear.

  14. Northern Arizona University

    ERIC Educational Resources Information Center

    Butcher, Michael F.; Saltonstall, Margot; Bickel, Sarah; Brandel, Rick

    2009-01-01

    Northern Arizona University (NAU) is a public university nestled below the San Francisco Peaks in Flagstaff, Arizona. It enrolls more than 21,000 undergraduate and graduate students at its main campus in Flagstaff, through its 35 statewide sites, and via online program offerings. Within the university organizational system, Student Affairs has a…

  15. Northern Pintail (Anas acuta)

    USGS Publications Warehouse

    Austin, J.E.; Miller, M.R.; Poole, A.; Gill, F.

    1995-01-01

    The Northern Pintail is a medium-sized dabbling duck of slender, elegant lines and conservative plumage coloration. It is circumpolar in distribution and abundant in North America, with core nesting habitat in Alaska and the Prairie Pothole Region of southern Canada and the northern Great Plains. Breeders favor shallow wetlands interspersed throughout prairie grasslands or arctic tundra. An early fall migrant, the species arrives on wintering areas beginning in August, after wing molt, often forming large roosting and feeding flocks on open, shallow wetlands and flooded agricultural fields. The birds consume grains, marsh plant seeds, and aquatic invertebrates throughout the fall and winter. Northern Pintails are among the earliest nesting ducks in North America, beginning shortly after ice-out in many northern areas. Individuals form new pair bonds each winter but are highly promiscuous during the nesting season, with mated and unmated males often involved in vigorous, acrobatic Pursuit Flights. Annual nest success and productivity vary with water conditions, predation, and weather. Females build nests on the ground, often long distances from water. Only the female incubates; her mate leaves shortly after incubation begins. Ducklings hatch together in one day, follow the female to water after a day in the nest, and fledge by July or August. Adults and ducklings consume mainly aquatic invertebrates during the breeding season. Predators and farming operations destroy many thousands of Northern Pintail nests annually; farming has also greatly reduced the amount of quality nesting cover available. Winter habitats are threatened by water shortages, agricultural development, contamination, and urbanization. Periods of extended drought in prairie nesting regions have caused dramatic population declines, usually followed by periods of recovery. Over the long term, however, the continental population of Northern Pintails has declined significantly from 6 million birds in

  16. Identification of Xanthomonas fragariae, Xanthomonas axonopodis pv. phaseoli, and Xanthomonas fuscans subsp. fuscans with Novel Markers and Using a Dot Blot Platform Coupled with Automatic Data Analysis ▿ †

    PubMed Central

    Albuquerque, Pedro; Caridade, Cristina M. R.; Marcal, Andre R. S.; Cruz, Joana; Cruz, Leonor; Santos, Catarina L.; Mendes, Marta V.; Tavares, Fernando

    2011-01-01

    Phytosanitary regulations and the provision of plant health certificates still rely mainly on long and laborious culture-based methods of diagnosis, which are frequently inconclusive. DNA-based methods of detection can circumvent many of the limitations of currently used screening methods, allowing a fast and accurate monitoring of samples. The genus Xanthomonas includes 13 phytopathogenic quarantine organisms for which improved methods of diagnosis are needed. In this work, we propose 21 new Xanthomonas-specific molecular markers, within loci coding for Xanthomonas-specific protein domains, useful for DNA-based methods of identification of xanthomonads. The specificity of these markers was assessed by a dot blot hybridization array using 23 non-Xanthomonas species, mostly soil dwelling and/or phytopathogens for the same host plants. In addition, the validation of these markers on 15 Xanthomonas spp. suggested species-specific hybridization patterns, which allowed discrimination among the different Xanthomonas species. Having in mind that DNA-based methods of diagnosis are particularly hampered for unsequenced species, namely, Xanthomonas fragariae, Xanthomonas axonopodis pv. phaseoli, and Xanthomonas fuscans subsp. fuscans, for which comparative genomics tools to search for DNA signatures are not yet applicable, emphasis was given to the selection of informative markers able to identify X. fragariae, X. axonopodis pv. phaseoli, and X. fuscans subsp. fuscans strains. In order to avoid inconsistencies due to operator-dependent interpretation of dot blot data, an image-processing algorithm was developed to analyze automatically the dot blot patterns. Ultimately, the proposed markers and the dot blot platform, coupled with automatic data analyses, have the potential to foster a thorough monitoring of phytopathogenic xanthomonads. PMID:21705524

  17. Human T lymphotropic virus types I and II western blot seroindeterminate status and its association with exposure to prototype HTLV-I.

    PubMed

    Yao, Karen; Hisada, Michie; Maloney, Elizabeth; Yamano, Yoshihisa; Hanchard, Barrie; Wilks, Rainford; Rios, Maria; Jacobson, Steven

    2006-02-01

    Human T lymphotropic virus types I and II (HTLV-I/II) Western blot (WB) seroindeterminate status, which is defined as an incomplete banding pattern of HTLV protein Gag (p19 or p24) or Env (GD21 or rgp46), is commonly observed. To investigate the significance of this finding, we examined HTLV-I/II serostatus and HTLV-I proviral load in 2 groups of individuals with WB seroindeterminate status. Low proviral loads were detected in 42% of patients with neurologic symptoms and 44% of voluntary blood donors. These data suggest that a subset of WB seroindeterminate individuals may be infected with prototype HTLV-I. To confirm this hypothesis, we evaluated HTLV-I/II serostatus and proviral load in prospectively collected specimens from 66 WB seronegative patients who had received HTLV-I-infected blood products by transfusion. Eight individuals developed WB seroindeterminate profiles after the transfusion. In addition, using a human leukocyte antigen type A*201-restricted HTLV-I Tax11-19 tetramer, we detected virus-specific CD8(+) T cells in peripheral blood mononuclear cells from WB seroindeterminate patients. These CD8(+) T cells were effective at targeting HTLV-I-infected cells. Collectively, the results suggest that HTLV-I/II WB seroindeterminate status may reflect a history of HTLV-I exposure. Our findings warrant further investigation of the possible clinical outcomes associated with WB seroindeterminate status.

  18. Spectrophotometric quantitation of DNA on blots after ethanol-solubilization of the MTT-formazan from anti-digoxigenin-based detection of nucleic acids.

    PubMed

    Colgan, D J

    1993-01-01

    The tetrazolium salt, 3-(4,5-dimethyl-thiazolyl-2)-2,5-diphenyl tetrazolium bromide (MTT) has recently been established as a substitute for Nitro-blue tetrazolium (NBT) in stain mixtures using antibody-conjugated alkaline phosphatase for the location of proteins on Western blots (Heegaard, 1990). Experiments reported here show that MTT is as sensitive as NBT in digoxigenin-labeled probe localization (on nucleic acid blots) utilizing alkaline-phosphatase-labelled, anti-digoxigenin antibodies. Moreover, as the formazan from MTT is soluble in ethanol, it is shown that spectrophotometric quantitation can be used to estimate the amount of target DNA on dot and Southern blots. For dot blotting, pBR328 was used as the probe and pBR322 as target. For Southern blots, human rDNA was used as the probe and total genomic calf DNA as the target. Staining response was linear over at least six twofold DNA dilutions in both types of blot.

  19. High prevalence of human papillomavirus infection in American Indian women of the Northern Plains

    PubMed Central

    Bell, Maria C.; Schmidt-Grimminger, Delf; Patrick, Sarah; Ryschon, Tim; Linz, Laurie; Chauhan, Subhash C.

    2008-01-01

    Objectives Cervical cancer is the leading gynecological malignancy worldwide, and the incidence of this disease is very high in American Indian women. Infection with the Human Papillomavirus (HPV) is responsible for more than 95% of cervical squamous carcinomas. Therefore, the main objective of this study was to analyze oncogenic HPV infections in American Indian women residing in the Northern Plains. Methods Cervical samples were collected from 287 women attending a Northern Plains American Indian reservation outpatient clinic. DNA was extracted from the cervical samples and HPV specific DNA were amplified by polymerase chain reaction (PCR) using the L1 consensus primer sets. The PCR products were hybridized with the Roche HPV Line Blot assay for HPV genotyping to detect 27 different low and high-risk HPV genotypes. The chi-square test was performed for statistical analysis of the HPV infection and cytology diagnosis data. Results Of the total 287 patients, 61 women (21.25%) tested positive for HPV infection. Among all HPV-positive women, 41 (67.2%) were infected with high-risk HPV types. Of the HPV infected women, 41% presented with multiple HPV genotypes. Additionally, of the women infected with oncogenic HPV types, 20 (48.7%) were infected with HPV 16 and 18 and the remaining 21 (51.3%) were infected with other oncogenic types (i.e., HPV59, 39, 73). Women infected with oncogenic HPV types had significantly higher (p=0.001) abnormal Papanicolaou smear tests (Pap test) compared to women who were either HPV negative or positive for non-oncogenic HPV types. The incidence of HPV infection was inversely correlated (p<0.05) with the age of the patients, but there was no correlation (p=0.33) with seasonal variation. Conclusions In this study, we observed a high prevalence of HPV infection in American Indian women residing on Northern Plains Reservations. In addition, a significant proportion of the oncogenic HPV infections were other than HPV16 and 18. PMID:17659767

  20. Northern Plains Patterns

    NASA Technical Reports Server (NTRS)

    2003-01-01

    MGS MOC Release No. MOC2-513, 14 October 2003

    Patterns are common on the northern plains of Mars. Like their terrestrial counterparts in places like Siberia, Alaska, and northern Canada, patterned ground on Mars might be an indicator of the presence of ground ice. Whether it is true that the patterns on Mars are related to ground ice and whether the ice is still present beneath the martian surface are unknown. This Mars Global Surveyor (MGS) Mars Orbiter Camera (MOC) picture shows an example of patterned ground on the martian northern plains near 72.4oN, 252.6oW. The dark dots and lines are low mounds and chains of mounds. The circular feature near the center of the image is the location of a buried meteor impact crater; its presence today is marked only by the dark boulders on its rim and ejecta blanket that have managed to remain uncovered at the martian surface. The area shown is 3 km (1.9 mi) wide and illuminated by sunlight from the lower left.

  1. Dermatophytosis in northern Africa.

    PubMed

    Nweze, E I; Eke, I

    2016-03-01

    Infections caused by dermatophytes are a global problem and a major public health burden in the world today. In Africa, especially in the northern geographical zone, dermatophytic infections are being reported at an alarming rate. This is mostly because of some local but unique cultural practices, socioeconomic and environmental conditions, lack of reliable diagnostic personnel and facilities and ineffective treatment. Interestingly, the pathogen spectrum and the clinical manifestation are most times different from what is seen in other continents. Several epidemiological studies have been performed on the incidence and aetiology of dermatophytoses in northern Africa. However, there is currently no review article with up-to-date information on the relevant findings reported so far in this region. This information is necessary for clinicians who treat dermatophytic infections all over the world since agents of dermatophytes are no longer restricted because of the rapid mobility of humans from one part of the world to another. Moreover, the epidemiology of dermatophytoses is known to change over time, thus requiring the update of information from time to time. A review of relevant studies published on dermatophytoses in northern Africa is presented. This covers all of old Sudan, Algeria, Egypt, Libya, Tunisia and Morocco.

  2. [Optimization of the immunoelectrophoresis technic (western blot) for the confirmation of human immunodeficiency virus infection (HIV) in Panama].

    PubMed

    Pascale, J M; de Austin, E; de Moreno, N O; Ledezma, C; de Márquez, E; Blanco, R; Quiroz, E; Calzada, J; Vincensini, A R; de Martin, M C

    1995-01-01

    The purpose of this study is to report the results of the authors' investigation to apply the western blot technique (WB UP-LCS) in the diagnosis of human immunodeficiency virus type 1 (HIV-1) infection. To do this, the authors separated the proteins of the HIV-1 virus by electrophoresis, based on their molecular weight, in poliacilamide gel with SDS (SDS-PAGE) during 3 hours at 200 volts. Then they electrotransferred these proteins to nitrocellulose paper during four hours at 200 milliamperes, with the aid of external cooling. The nitrocellulose strips were evaluated considering the incubation time (1 and 16 hours), two conjugates (human anti IgG with Peroxidase and human anti IgG Biotin plus Streptatividine with Peroxidase) and two dilutions of the patients' sera (1/50 and 1/100). Based on their results the Authors conclude that, in the first place, the optimal conditions for the test include a dilution of 1/100 of the patients serum, incubation of the serum for 16 hours and the use of the conjugate of anti human IgG with Biotin and Streptavidine with Peroxidase; secondary, that the immunologic reactivity against proteins p24 and gp 160/120 is the most important diagnostic criterion for the confirmation of infection with HIV-1 and that they obtained a diagnostic correlation of 100% at a cost which was 5 to 7 times less than that of the commercial system.

  3. Analysis of cell wall extracts of Candida albicans by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and Western blot techniques.

    PubMed Central

    Ponton, J; Jones, J M

    1986-01-01

    Cell walls of intact yeast- and mycelial-phase Candida albicans B311 were extracted with different compounds: dithiothreitol, dithiothreitol with protease, dithiothreitol with lyticase, and dithiothreitol with protease followed by beta-glucuronidase with chitinase. Extracts were analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and Western blot techniques. Dithiothreitol extracts contained the most satisfactory array of components for study. Analysis of these extracts demonstrated that the outer cell wall layers of Candida blastoconidia and germ tubes contained a complex array of polysaccharides, glycoproteins, and proteins. The proteins contributed to a latticework stabilized by covalent bonds that was important in determining the porosity of the outer cell wall layers. When equivalent weights were analyzed, mycelial-phase extract contained a more varied array of proteins than did yeast-phase extract. Only a portion of proteins in mycelial-phase extract elicited antibody responses in hyperimmunized rabbits or infected humans. A polysaccharide-rich, high-molecular-weight component (migrating at a position that would correspond to proteins having molecular weights of 235,000 to 250,000) and a protein component (molecular weight, 19,000) were readily demonstrable in the mycelial-phase extract but could not be identified in the yeast-phase extract. Images PMID:3527986

  4. Comparison of germ line minisatellite mutation detection at the CEB1 locus by Southern blotting and PCR amplification.

    PubMed

    Taylor, Malcolm; Cieslak, Marcin; Rees, Gwen S; Oojageer, Anthony; Leith, Cheryl; Bristow, Claire; Tawn, E Janet; Winther, Jeanette F; Boice, John D

    2010-07-01

    Identification of de novo minisatellite mutations in the offspring of parents exposed to mutagenic agents offers a potentially sensitive measure of germ line genetic events induced by ionizing radiation and genotoxic chemicals. Germ line minisatellite mutations (GMM) are usually detected by hybridizing Southern blots of unamplified size-fractionated genomic DNA with minisatellite probes. However, this consumes a relatively large amount of DNA, requires several steps and may lack sensitivity. We have developed a polymerase chain reaction (PCR)-based GMM assay, which we applied to the hypermutable minisatellite, CEB1. Here, we compare the sensitivity and specificity of this assay with the conventional Southern hybridization method using DNA from 10 spouse pairs, one parent of each pair being a survivor of cancer in childhood, and their 20 offspring. We report that both methods have similar specificity but that the PCR method uses 250 times less DNA, has fewer steps and is better at detecting GMM with single repeats provided that specific guidelines for allele sizing are followed. The PCR GMM method is easier to apply to families where the amount of offspring DNA sample is limited.

  5. Molecular mass dependence of hyaluronan detection by sandwich ELISA-like assay and membrane blotting using biotinylated hyaluronan binding protein

    PubMed Central

    Yuan, Han; Tank, Mihir; Alsofyani, Abeer; Shah, Naman; Talati, Nishant; LoBello, Jaclyn C; Kim, Jin Ryoun; Oonuki, Yoji; de la Motte, Carol A; Cowman, Mary K

    2013-01-01

    Hyaluronan (HA) is widely detected in biological samples and its concentration is most commonly determined by the use of a labeled specific HA binding protein (aggrecan G1-IGD-G2, HABP), employing membrane blotting and sandwich enzyme-linked immunosorbent assay (ELISA)-like methods. However, the detected signal intensity or the quantified value obtained by using these surface-based methods is related to the molecular mass (M) of HA, especially for HA in the low M range below ∼150 kDa. At the same mass or mass concentration, higher M HA gives a higher signal than lower M HA. We have experimentally determined the quantitative relationship between the M of HA (in the range 20–150 kDa) and the relative signal intensity in comparison with a standard HA, in a sandwich ELISA-like assay. An M-dependent signal correction factor (SCF) was calculated and used to correct the signal intensity, so that the corrected concentration value would more accurately reflect the true HA concentration in solution. The SCF for polydisperse low M HA was also calculated and compared with experimental results. When the molecular mass distribution of an HA sample is determined by a method such as gel electrophoresis, then its appropriately averaged SCF can be calculated and used to correct the signal in sandwich ELISA to obtain a more accurate concentration estimation. The correction method works for HA with M between ∼150 and 20 kDa, but lower M HA is too poorly detected for useful analysis. The physical basis of the M-dependent detection is proposed to be the increase in detector-accessible fraction of each surface-bound molecule as M increases. PMID:23964097

  6. Molecular Detection and Genotyping of Male-Specific Coliphages by Reverse Transcription-PCR and Reverse Line Blot Hybridization

    PubMed Central

    Vinjé, Jan; Oudejans, Sjon J. G.; Stewart, Jill R.; Sobsey, Mark D.; Long, Sharon C.

    2004-01-01

    In recent years, there has been increased interest in the use of male-specific or F+ coliphages as indicators of microbial inputs to source waters. Sero- or genotyping of these coliphages can also be used for microbial source tracking (MST). Among the male-specific coliphages, the F+ RNA (FRNA) viruses are well studied, while little is known about the F+ DNA (FDNA) viruses. We have developed a reverse line blot hybridization (RLB) assay which allows for the simultaneous detection and genotyping of both FRNA as well as FDNA coliphages. These assays included a novel generic duplex reverse transcription-PCR (RT-PCR) assay for FRNA viruses as well as a generic PCR for FDNA viruses. The RT-PCR assays were validated by using 190 field and prototype strains. Subsequent DNA sequencing and phylogenetic analyses of RT-PCR products revealed the classification of six different FRNA clusters, including the well-established subgroups I through IV, and three different FDNA clusters, including one (CH) not previously described. Within the leviviruses, a potentially new subgroup (called JS) including strains having more than 40% nucleotide sequence diversity with the known levivirus subgroups (MS2 and GA) was identified. We designed subgroup-specific oligonucleotides that were able to genotype all nine (six FRNA, three FDNA) different clusters. Application of the method to a panel of 351 enriched phage samples from animal feces and wastewater, including known prototype strains (MS2, GA, Qβ, M11, FI, and SP for FRNA and M13, f1, and fd for FDNA), resulted in successful genotyping of 348 (99%) of the samples. In summary, we developed a novel method for standardized genotyping of F+ coliphages as a useful tool for large-scale MST studies. PMID:15466543

  7. TESA-blot for the diagnosis of Chagas disease in dogs from co-endemic regions for Trypanosoma cruzi, Trypanosoma evansi and Leishmania chagasi.

    PubMed

    Umezawa, E S; Souza, A I; Pinedo-Cancino, V; Marcondes, M; Marcili, A; Camargo, L M A; Camacho, A A; Stolf, A M S; Teixeira, M M G

    2009-07-01

    We standardized serodiagnosis of dogs infected with Trypanosoma cruzi using TESA (trypomastigote excreted-secreted antigen)-blot developed for human Chagas disease. TESA-blot showed 100% sensitivity and specificity. In contrast, ELISA using TESA (TESA-ELISA) or epimastigotes (epi-ELISA) as antigen yielded 100% sensitivity but specificity of 94.1% and 49.4%, respectively. When used in field studies in an endemic region for Chagas disease, visceral leishmaniasis and Trypanosoma evansi (Mato Grosso do Sul state, Central Brazil), positivities were 9.3% for TESA-blot, 10.7% for TESA-ELISA and 32% for epi-ELISA. Dogs from a non-endemic region for these infections (Rondonia state, western Amazonia) where T. cruzi is enzootic showed positivity of 4.5% for TESA-blot and epi-ELISA and 6.8% for TESA-ELISA. Sera from urban dogs from Santos, São Paulo, where these diseases are absent, yielded negative results. TESA-blot was the only method that distinguished dogs infected with T. cruzi from those infected with Leishmania chagasi and/or Trypanosoma evansi.

  8. Human T lymphotropic virus types I and II proviral sequences in Argentinian blood donors with indeterminate Western blot patterns.

    PubMed

    Mangano, A M; Remesar, M; del Pozo, A; Sen, Luisa

    2004-10-01

    Human T-cell lymphotropic virus (HTLV) seroindeterminate blood donors have been reported worldwide including Argentina. To investigate the significance of HTLV-I/II seroindeterminate Western blot (WB) patterns, we conducted an 8-year cross-sectional study. Of 86,238 Argentinian blood donors, 146 sera were reactive by screening tests. The WB results indicated that 20% were HTLV-I reactive, 8% HTLV-II reactive, 61% indeterminate, and 11% negative. The overall seroprevalence was 0.034% for HTLV-I, 0.014% for HTLV-II, and 0.103% for indeterminate. In 57 reactive specimens, HTLV-I/II provirus could be examined by type specific PCR for tax, pol, and env regions. When at least two gene fragments were amplified HTLV-I/II infection was considered confirmed. PCR results confirmed all WB seropositive samples for HTLV-I (n = 15), and HTLV-II (n = 7), and the only WB negative case was also PCR negative, showing a complete concordance between PCR and WB. However, of 34 WB seroindeterminate sera studied by PCR, in 5 was proviral DNA amplified. According to our criteria PCR confirmed one to be HTLV-I, and one HTLV-II, 3 remained indeterminate since only tax sequences were amplified. Among WB indeterminate samples tested by PCR, most of their serological profile showed reactivity to gag codified proteins but lacked env reactivities (70%). One sample with a WB gag pattern showed proviral tax sequences, but of the four samples with reactivity to env proteins GD21 (n = 3) or rgp46II (n = 1) PCR results indicated that one was HTLV-I, one was HTLV-II, and two were indeterminate (only tax sequences). In conclusion, the majority of HTLV-seroindeterminate WB donors exhibited a gag indeterminate profile lacking HTLV provirus, and were thus considered uninfected. However, seroreactivity to env proteins, in particular to GD21, may indicate infection and a follow-up study of each seroreactive blood donor should be considered.

  9. Identification of recombinant human EPO variants in greyhound plasma and urine by ELISA, LC-MS/MS and western blotting: a comparative study.

    PubMed

    Timms, Mark; Steel, Rohan; Vine, John

    2016-02-01

    The recombinant human erythropoietins epoetin alfa (Eprex®), darbepoetin (Aranesp®) and methoxy polyethylene glycol-epoetin beta (Mircera®) were administered to greyhounds for 7, 10 and 14 days respectively. Blood and urine samples were collected and analysed for erythropoietin by ELISA, LC-MS/MS and western blotting. Limits of confirmation in plasma for western blotting and LC-MS/MS methods ranged from a low of 2.5mIU/mL, and closely matched the sensitivity of ELISA screening.

  10. Comparative reproductive and physiological responses of northern bobwhite and scaled quail to water deprivation

    USGS Publications Warehouse

    Giuliano, W.M.; Patino, R.; Lutz, R.S.

    1998-01-01

    We compared reproductive and physiological responses of captive female northern bobwhite (Colinus virginianus) and scaled quail (Callipepla squamata) under control and water deprivation conditions. Scaled quail required less food and water to reproduce successfully under control conditions than northern bobwhite. Additionally, in scaled quail, serum osmolality levels and kidney mass were unaffected by water deprivation, whereas in northern bobwhite, serum osmolality levels increased and kidney mass declined. This finding indicates that scaled quail may have osmoregulatory abilities superior to those of northern bobwhite. Under control conditions, northern bobwhite gained more body mass and produced more but smaller eggs than scaled quail. Under water deprivation conditions, northern bobwhite lost more body mass but had more laying bens with a higher rate of egg production than scaled quail. Our data suggest that northern bobwhite allocated more resources to reproduction than to body maintenance, while scaled quail apparently forego reproduction in favor of body maintenance during water deprivation conditions.

  11. Mitigating Radicalism in Northern Nigeria

    DTIC Science & Technology

    2013-08-01

    radicalization in northern Nigeria. u Active engagement of youth and communities in peacebuilding programs that facilitate interactions among individuals...leaders, sustained development investments in marginalized communities , promotion of values of inclusivity to mitigate the spread of extremist ideology...claiming to have repelled Boko Haram, the militants return, regroup, and seek revenge. As a result, social and economic activities in the northern

  12. Report on Ontario's Northern Schools.

    ERIC Educational Resources Information Center

    2003

    Ontario's funding formula fails to recognize the unique needs of northern school boards, which cover immense geographic areas, have many small schools, and enroll a high proportion of Aboriginal students. This report examines school size, enrollment, and staffing in northern Ontario schools, drawing on 2002-03 tracking reports of provincial…

  13. Northern Sinus Meridiani Stereo

    NASA Technical Reports Server (NTRS)

    2003-01-01

    MGS MOC Release No. MOC2-341, 25 April 2003

    This is a stereo (3-d anaglyph) composite of Mars Global Surveyor (MGS) Mars Orbiter Camera (MOC) wide angle images of northern Sinus Meridiani near 2oN, 0oW. The light-toned materials at the south (bottom) end of the picture are considered to be thick (100-200 meters; 300-600 ft) exposures of sedimentary rock. Several ancient meteor impact craters are being exhumed from within these layered materials. To view in stereo, use '3-d' glasses with red over the left eye, and blue over the right. The picture covers an area approximately 113 km (70 mi) wide; north is up.

  14. Glyphosate in northern ecosystems.

    PubMed

    Helander, Marjo; Saloniemi, Irma; Saikkonen, Kari

    2012-10-01

    Glyphosate is the main nonselective, systemic herbicide used against a wide range of weeds. Its worldwide use has expanded because of extensive use of certain agricultural practices such as no-till cropping, and widespread application of glyphosate-resistant genetically modified crops. Glyphosate has a reputation of being nontoxic to animals and rapidly inactivated in soils. However, recent evidence has cast doubts on its safety. Glyphosate may be retained and transported in soils, and there may be cascading effects on nontarget organisms. These processes may be especially detrimental in northern ecosystems because they are characterized by long biologically inactive winters and short growing seasons. In this opinion article, we discuss the potential ecological, environmental and agricultural risks of intensive glyphosate use in boreal regions.

  15. Profile of the MP Diagnostics HTLV Blot 2.4 test: a supplemental assay for the confirmation and differentiation of antibodies to HTLV-1 and HTLV-2.

    PubMed

    Miller, Liane

    2016-01-01

    As the first US FDA-approved assay for supplemental HTLV testing, the MP Diagnostics HTLV Blot 2.4 is an effective and efficient method for confirming and differentiating HTLV type infection in repeatedly reactive samples. Novel and patented antigens added increased sensitivity in identifying specimens from infected individuals while differentiating those from uninfected individuals with false reactivity.

  16. Resolution and identification of major peanut allergens using a combination of fluorescence two-dimensional differential gel electrophoresis, western blotting and Q-TOF mass spectrometry.

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Peanut allergy is triggered by several proteins known as allergens. The matching resolution and identification of major peanut allergens in 2D protein maps, was accomplished by the use of fluorescence two-dimensional differential gel electrophoresis (2D DIGE), Western blotting and quadrupole time-of...

  17. Additive Similarity Trees

    ERIC Educational Resources Information Center

    Sattath, Shmuel; Tversky, Amos

    1977-01-01

    Tree representations of similarity data are investigated. Hierarchical clustering is critically examined, and a more general procedure, called the additive tree, is presented. The additive tree representation is then compared to multidimensional scaling. (Author/JKS)

  18. Qualitative and quantitative analyses of Epstein-Barr virus early antigen diffuse component by western blotting enzyme-linked immunosorbent assay with a monoclonal antibody.

    PubMed Central

    Lin, J C; Choi, E I; Pagano, J S

    1985-01-01

    We report the use of monoclonal antibody against the early antigen diffuse component (anti-EA-D) of Epstein-Barr virus (EBV) to analyze, both qualitatively and quantitatively, the expression of EA-D in various human lymphoblastoid cell lines activated by chemical inducers. The kinetics of synthesis of EA-D in P3HR-1, B95-8, and Ramos/AW cells were similar in that they all reached the peak of synthesis on day 5 after induction. Surprisingly, no expression of EA-D was found in induced BJAB/GC, an EBV-genome-containing cell line. EBV-negative cell lines, BJAB and Ramos, were negative for EA-D. Raji cells had no detectable EA-D but responded rapidly to induction, reaching a peak on day 3. Superinfection of Raji cells also resulted in marked induction of EA-D, which reached a plateau between 8 to 12 h postinfection. Western blotting coupled with the enzyme-linked immunosorbent assay was employed to identify polypeptides representing EA-D. A family of four polypeptides with molecular weights of 46,000 (46K protein), 49,000, 52,000, and 55,000 were identified to be reactive with monoclonal anti-EA-D antiserum. The pattern of EA-D polypeptides expressed in each cell line was different. Of particular interest was the expression of a large quantity of 46K protein both in induced Raji and P3HR-1 cells, but not in superinfected Raji cells. A 49K doublet was expressed in activated p3HR-1, B95-8, and Ramos/AW cells and in superinfected Raji cells. In addition, two distinct 52K and 55K polypeptides were expressed in induced Ramos/AW and superinfected Raji cells. However, none of these EA-D polypeptides was detectable in BJAB/GC, BJAB, Ramos, and mock-infected Raji cells. To approximate relative concentrations of EA-D in cell extracts, we employed the enzyme-linked immunosorbent assay and immunoblot dot methods by using one of the purified EA-D components to construct a standard curve. Depending upon the cell lines, it was estimated that ca. 1 to 3% (determined by the enzyme

  19. Polyimide processing additives

    NASA Technical Reports Server (NTRS)

    Pratt, J. R.; St. Clair, T. L.; Burks, H. D.; Stoakley, D. M.

    1987-01-01

    A method has been found for enhancing the melt flow of thermoplastic polyimides during processing. A high molecular weight 422 copoly(amic acid) or copolyimide was fused with approximately 0.05 to 5 pct by weight of a low molecular weight amic acid or imide additive, and this melt was studied by capillary rheometry. Excellent flow and improved composite properties on graphite resulted from the addition of a PMDA-aniline additive to LARC-TPI. Solution viscosity studies imply that amic acid additives temporarily lower molecular weight and, hence, enlarge the processing window. Thus, compositions containing the additive have a lower melt viscosity for a longer time than those unmodified.

  20. [Food additives and healthiness].

    PubMed

    Heinonen, Marina

    2014-01-01

    Additives are used for improving food structure or preventing its spoilage, for example. Many substances used as additives are also naturally present in food. The safety of additives is evaluated according to commonly agreed principles. If high concentrations of an additive cause adverse health effects for humans, a limit of acceptable daily intake (ADI) is set for it. An additive is a risk only when ADI is exceeded. The healthiness of food is measured on the basis of nutrient density and scientifically proven effects.

  1. MISR Views Northern Australia

    NASA Technical Reports Server (NTRS)

    2000-01-01

    MISR images of tropical northern Australia acquired on June 1, 2000 (Terra orbit 2413) during the long dry season. Left: color composite of vertical (nadir) camera blue, green, and red band data. Right: multi-angle composite of red band data only from the cameras viewing 60 degrees aft, 60 degrees forward, and nadir. Color and contrast have been enhanced to accentuate subtle details. In the left image, color variations indicate how different parts of the scene reflect light differently at blue, green, and red wavelengths; in the right image color variations show how these same scene elements reflect light differently at different angles of view. Water appears in blue shades in the right image, for example, because glitter makes the water look brighter at the aft camera's view angle. The prominent inland water body is Lake Argyle, the largest human-made lake in Australia, which supplies water for the Ord River Irrigation Area and the town of Kununurra (pop. 6500) just to the north. At the top is the southern edge of Joseph Bonaparte Gulf; the major inlet at the left is Cambridge Gulf, the location of the town of Wyndham (pop. 850), the port for this region. This area is sparsely populated, and is known for its remote, spectacular mountains and gorges. Visible along much of the coastline are intertidal mudflats of mangroves and low shrubs; to the south the terrain is covered by open woodland merging into open grassland in the lower half of the pictures.

    MISR was built and is managed by NASA's Jet Propulsion Laboratory, Pasadena, CA, for NASA's Office of Earth Science, Washington, DC. The Terra satellite is managed by NASA's Goddard Space Flight Center, Greenbelt, MD. JPL is a division of the California Institute of Technology.

  2. Isolated Northern Dunes

    NASA Technical Reports Server (NTRS)

    2005-01-01

    [figure removed for brevity, see original site]

    Our topic for the weeks of April 4 and April 11 is dunes on Mars. We will look at the north polar sand sea and at isolated dune fields at lower latitudes. Sand seas on Earth are often called 'ergs,' an Arabic name for dune field. A sand sea differs from a dune field in two ways: 1) a sand sea has a large regional extent, and 2) the individual dunes are large in size and complex in form.

    This VIS image was taken at 81 degrees North latitude during Northern spring. In this region, the dunes are isolated from each other. The dunes are just starting to emerge from the winter frost covering appearing dark with bright crests. These dunes are located on top of ice.

    Image information: VIS instrument. Latitude 82.1, Longitude 191.3 East (168.7 West). 19 meter/pixel resolution.

    Note: this THEMIS visual image has not been radiometrically nor geometrically calibrated for this preliminary release. An empirical correction has been performed to remove instrumental effects. A linear shift has been applied in the cross-track and down-track direction to approximate spacecraft and planetary motion. Fully calibrated and geometrically projected images will be released through the Planetary Data System in accordance with Project policies at a later time.

    NASA's Jet Propulsion Laboratory manages the 2001 Mars Odyssey mission for NASA's Office of Space Science, Washington, D.C. The Thermal Emission Imaging System (THEMIS) was developed by Arizona State University, Tempe, in collaboration with Raytheon Santa Barbara Remote Sensing. The THEMIS investigation is led by Dr. Philip Christensen at Arizona State University. Lockheed Martin Astronautics, Denver, is the prime contractor for the Odyssey project, and developed and built the orbiter. Mission operations are conducted jointly from Lockheed Martin and from JPL, a division of the California Institute of Technology in Pasadena.

  3. Northern Sand Sea

    NASA Technical Reports Server (NTRS)

    2005-01-01

    [figure removed for brevity, see original site]

    Our topic for the weeks of April 4 and April 11 is dunes on Mars. We will look at the north polar sand sea and at isolated dune fields at lower latitudes. Sand seas on Earth are often called 'ergs,' an Arabic name for dune field. A sand sea differs from a dune field in two ways: 1) a sand sea has a large regional extent, and 2) the individual dunes are large in size and complex in form.

    This VIS image was taken at 82 degrees North latitude during Northern spring. The image is completely dominated by dunes. In sand seas, it is very common for a single type of dune to occur, and for a single predominate wind to control the alignment of the dunes.

    Image information: VIS instrument. Latitude 82.2, Longitude 152.5 East (207.5 West). 19 meter/pixel resolution.

    Note: this THEMIS visual image has not been radiometrically nor geometrically calibrated for this preliminary release. An empirical correction has been performed to remove instrumental effects. A linear shift has been applied in the cross-track and down-track direction to approximate spacecraft and planetary motion. Fully calibrated and geometrically projected images will be released through the Planetary Data System in accordance with Project policies at a later time.

    NASA's Jet Propulsion Laboratory manages the 2001 Mars Odyssey mission for NASA's Office of Space Science, Washington, D.C. The Thermal Emission Imaging System (THEMIS) was developed by Arizona State University, Tempe, in collaboration with Raytheon Santa Barbara Remote Sensing. The THEMIS investigation is led by Dr. Philip Christensen at Arizona State University. Lockheed Martin Astronautics, Denver, is the prime contractor for the Odyssey project, and developed and built the orbiter. Mission operations are conducted jointly from Lockheed Martin and from JPL, a division of the California Institute of Technology in Pasadena.

  4. Dual infection of chickens with pox and infectious laryngotracheitis (ILT) confirmed with specific pox and ILT DNA dot-blot hybridization assays.

    PubMed

    Fatunmbi, O O; Reed, W M; Schwartz, D L; Tripathy, D N

    1995-01-01

    Dual infection with fowl pox (FP) and infectious laryngotracheitis (ILT) was diagnosed as the cause of acute mortality in a flock of three age groups of Hy-Line leghorn layers. The affected chickens had not been previously vaccinated against either FP or ILT. The diagnosis was confirmed by virus isolation, histopathology, and the use of specific pox and ILT genomic DNA probes in a dot-blot hybridization assay. FP and ILT vaccinations were recommended to control mortality. The use of FP- and ILT-specific DNA dot-blot hybridization may be used as a routine diagnostic tool to differentiate between the two diseases, especially in atypical cases of either infection or to confirm the existence of the two diseases as a mixed infection in a flock of chickens.

  5. Polylactides in additive biomanufacturing.

    PubMed

    Poh, Patrina S P; Chhaya, Mohit P; Wunner, Felix M; De-Juan-Pardo, Elena M; Schilling, Arndt F; Schantz, Jan-Thorsten; van Griensven, Martijn; Hutmacher, Dietmar W

    2016-12-15

    New advanced manufacturing technologies under the alias of additive biomanufacturing allow the design and fabrication of a range of products from pre-operative models, cutting guides and medical devices to scaffolds. The process of printing in 3 dimensions of cells, extracellular matrix (ECM) and biomaterials (bioinks, powders, etc.) to generate in vitro and/or in vivo tissue analogue structures has been termed bioprinting. To further advance in additive biomanufacturing, there are many aspects that we can learn from the wider additive manufacturing (AM) industry, which have progressed tremendously since its introduction into the manufacturing sector. First, this review gives an overview of additive manufacturing and both industry and academia efforts in addressing specific challenges in the AM technologies to drive toward AM-enabled industrial revolution. After which, considerations of poly(lactides) as a biomaterial in additive biomanufacturing are discussed. Challenges in wider additive biomanufacturing field are discussed in terms of (a) biomaterials; (b) computer-aided design, engineering and manufacturing; (c) AM and additive biomanufacturing printers hardware; and (d) system integration. Finally, the outlook for additive biomanufacturing was discussed.

  6. Additive Manufactured Product Integrity

    NASA Technical Reports Server (NTRS)

    Waller, Jess; Wells, Doug; James, Steve; Nichols, Charles

    2017-01-01

    NASA is providing key leadership in an international effort linking NASA and non-NASA resources to speed adoption of additive manufacturing (AM) to meet NASA's mission goals. Participants include industry, NASA's space partners, other government agencies, standards organizations and academia. Nondestructive Evaluation (NDE) is identified as a universal need for all aspects of additive manufacturing.

  7. Characterization of Corynebacterium diphtheriae isolates from infected skin lesions in the Northern Territory of Australia.

    PubMed

    Gordon, Claire L; Fagan, Peter; Hennessy, Jann; Baird, Robert

    2011-11-01

    Corynebacterium diphtheriae is commonly isolated from cutaneous skin lesions in the Northern Territory of Australia. We prospectively assessed 32 recent isolates from infected skin lesions, in addition to reviewing 192 isolates collected over 5 years for toxin status. No isolates carried the toxin gene. Toxigenic C. diphtheriae is now a rare occurrence in the Northern Territory.

  8. Anti-RAINBOW dye-specific antibodies as universal tools for the visualization of prestained protein molecular weight markers in Western blot analysis

    PubMed Central

    Schüchner, Stefan; Andorfer, Peter; Mudrak, Ingrid; Ogris, Egon

    2016-01-01

    Western blotting is one of the most widely used techniques in molecular biology and biochemistry. Prestained proteins are used as molecular weight standards in protein electrophoresis. In the chemiluminescent Western blot analysis, however, these colored protein markers are invisible leaving researchers with the unsatisfying situation that the signal for the protein of interest and the signal for the markers are not captured simultaneously and have to be merged in an error-prone step. To allow the simultaneous detection of marker proteins we generated monoclonal antibodies specific for the protein dyes. To elicit a dye rather than protein specific immune response we immunized mice sequentially with dye-carrier protein complexes, in which a new carrier protein was used for each subsequent immunization. Moreover, by sequentially immunizing with dye-carrier protein complexes, in which different but structurally related dyes were used, we could also generate an antibody, termed anti-RAINBOW, that cross-reacted even with structurally related dyes not used in the immunizations. Our novel antibodies represent convenient tools for the simultaneous Western blot detection of commercially available prestained marker proteins in combination with the detection of any specific protein of interest. These antibodies will render obsolete the anachronistic tradition of manually charting marker bands on film. PMID:27531616

  9. Polyimide processing additives

    NASA Technical Reports Server (NTRS)

    Fletcher, James C. (Inventor); Pratt, J. Richard (Inventor); St.clair, Terry L. (Inventor); Stoakley, Diane M. (Inventor); Burks, Harold D. (Inventor)

    1992-01-01

    A process for preparing polyimides having enhanced melt flow properties is described. The process consists of heating a mixture of a high molecular weight poly-(amic acid) or polyimide with a low molecular weight amic acid or imide additive in the range of 0.05 to 15 percent by weight of additive. The polyimide powders so obtained show improved processability, as evidenced by lower melt viscosity by capillary rheometry. Likewise, films prepared from mixtures of polymers with additives show improved processability with earlier onset of stretching by TMA.

  10. Polyimide processing additives

    NASA Technical Reports Server (NTRS)

    Pratt, J. Richard (Inventor); St.clair, Terry L. (Inventor); Stoakley, Diane M. (Inventor); Burks, Harold D. (Inventor)

    1993-01-01

    A process for preparing polyimides having enhanced melt flow properties is described. The process consists of heating a mixture of a high molecular weight poly-(amic acid) or polyimide with a low molecular weight amic acid or imide additive in the range of 0.05 to 15 percent by weight of the additive. The polyimide powders so obtained show improved processability, as evidenced by lower melt viscosity by capillary rheometry. Likewise, films prepared from mixtures of polymers with additives show improved processability with earlier onset of stretching by TMA.

  11. An Environmental Scan of Northern Alameda County. Revised.

    ERIC Educational Resources Information Center

    Peralta Community Coll. District, Oakland, CA. Office of Research, Planning and Development.

    An overview is provided of the demographic and economic characteristics of the geographical area defined by the Peralta Community College District's boundaries in Northern Alameda County, California. In addition, projections are presented concerning the population and economy of the county until the year 2005. Highlighted findings of the…

  12. Food Additives and Hyperkinesis

    ERIC Educational Resources Information Center

    Wender, Ester H.

    1977-01-01

    The hypothesis that food additives are causally associated with hyperkinesis and learning disabilities in children is reviewed, and available data are summarized. Available from: American Medical Association 535 North Dearborn Street Chicago, Illinois 60610. (JG)

  13. Smog control fuel additives

    SciTech Connect

    Lundby, W.

    1993-06-29

    A method is described of controlling, reducing or eliminating, ozone and related smog resulting from photochemical reactions between ozone and automotive or industrial gases comprising the addition of iodine or compounds of iodine to hydrocarbon-base fuels prior to or during combustion in an amount of about 1 part iodine per 240 to 10,000,000 parts fuel, by weight, to be accomplished by: (a) the addition of these inhibitors during or after the refining or manufacturing process of liquid fuels; (b) the production of these inhibitors for addition into fuel tanks, such as automotive or industrial tanks; or (c) the addition of these inhibitors into combustion chambers of equipment utilizing solid fuels for the purpose of reducing ozone.

  14. Family Medicine/Northern Medical Services Involvement in Northern Saskatchewan

    PubMed Central

    Irvine, James

    1988-01-01

    To address the problems of recruitment and retention of family physicians in various remote locations in northern Saskatchewan, the University of Saskatchewan became involved through Northern Medical Services, a division of the Department of Family Medicine. The University's involvement consists of the provision of resident family-physician services, visiting consultant services, family-practise resident training, research, and a consulting role of the Medical Health Officer. This paper reviews the context in which this program was created, its role in health care in the area, and its involvement with the communities in health promotion and research. PMID:21253032

  15. Group Sparse Additive Models

    PubMed Central

    Yin, Junming; Chen, Xi; Xing, Eric P.

    2016-01-01

    We consider the problem of sparse variable selection in nonparametric additive models, with the prior knowledge of the structure among the covariates to encourage those variables within a group to be selected jointly. Previous works either study the group sparsity in the parametric setting (e.g., group lasso), or address the problem in the nonparametric setting without exploiting the structural information (e.g., sparse additive models). In this paper, we present a new method, called group sparse additive models (GroupSpAM), which can handle group sparsity in additive models. We generalize the ℓ1/ℓ2 norm to Hilbert spaces as the sparsity-inducing penalty in GroupSpAM. Moreover, we derive a novel thresholding condition for identifying the functional sparsity at the group level, and propose an efficient block coordinate descent algorithm for constructing the estimate. We demonstrate by simulation that GroupSpAM substantially outperforms the competing methods in terms of support recovery and prediction accuracy in additive models, and also conduct a comparative experiment on a real breast cancer dataset.

  16. Additive Manufacturing Infrared Inspection

    NASA Technical Reports Server (NTRS)

    Gaddy, Darrell

    2014-01-01

    Additive manufacturing is a rapid prototyping technology that allows parts to be built in a series of thin layers from plastic, ceramics, and metallics. Metallic additive manufacturing is an emerging form of rapid prototyping that allows complex structures to be built using various metallic powders. Significant time and cost savings have also been observed using the metallic additive manufacturing compared with traditional techniques. Development of the metallic additive manufacturing technology has advanced significantly over the last decade, although many of the techniques to inspect parts made from these processes have not advanced significantly or have limitations. Several external geometry inspection techniques exist such as Coordinate Measurement Machines (CMM), Laser Scanners, Structured Light Scanning Systems, or even traditional calipers and gages. All of the aforementioned techniques are limited to external geometry and contours or must use a contact probe to inspect limited internal dimensions. This presentation will document the development of a process for real-time dimensional inspection technique and digital quality record of the additive manufacturing process using Infrared camera imaging and processing techniques.

  17. Current Status and Future Earth System Studies in Northern Eurasia

    NASA Astrophysics Data System (ADS)

    Groisman, Pavel; Gordov, Evgeny; Maksyutov, Shamil

    2013-12-01

    Northern Eurasia is a sensitive and rapidly changing area with the signal of climate change effects already observed in many components of the Earth's system. The ongoing warming in the Northern Eurasia Earth Science Partnership Initiative (NEESPI) domain was substantially larger than for the globe and during the past 50 years, with a rate of the annual temperature increase of 0.33°C per decade. In addition, this region experienced impacts of abrupt institutional and economic changes in the former Soviet Union countries, east Europe, Mongolia, and China.

  18. Phenylethynyl Containing Reactive Additives

    NASA Technical Reports Server (NTRS)

    Connell, John W. (Inventor); Smith, Joseph G., Jr. (Inventor); Hergenrother, Paul M. (Inventor)

    2002-01-01

    Phenylethynyl containing reactive additives were prepared from aromatic diamine, containing phenylethvnvl groups and various ratios of phthalic anhydride and 4-phenylethynviphthalic anhydride in glacial acetic acid to form the imide in one step or in N-methyl-2-pvrrolidinone to form the amide acid intermediate. The reactive additives were mixed in various amounts (10% to 90%) with oligomers containing either terminal or pendent phenylethynyl groups (or both) to reduce the melt viscosity and thereby enhance processability. Upon thermal cure, the additives react and become chemically incorporated into the matrix and effect an increase in crosslink density relative to that of the host resin. This resultant increase in crosslink density has advantageous consequences on the cured resin properties such as higher glass transition temperature and higher modulus as compared to that of the host resin.

  19. Fused Lasso Additive Model

    PubMed Central

    Petersen, Ashley; Witten, Daniela; Simon, Noah

    2016-01-01

    We consider the problem of predicting an outcome variable using p covariates that are measured on n independent observations, in a setting in which additive, flexible, and interpretable fits are desired. We propose the fused lasso additive model (FLAM), in which each additive function is estimated to be piecewise constant with a small number of adaptively-chosen knots. FLAM is the solution to a convex optimization problem, for which a simple algorithm with guaranteed convergence to a global optimum is provided. FLAM is shown to be consistent in high dimensions, and an unbiased estimator of its degrees of freedom is proposed. We evaluate the performance of FLAM in a simulation study and on two data sets. Supplemental materials are available online, and the R package flam is available on CRAN. PMID:28239246

  20. Phenylethynyl Containing Reactive Additives

    NASA Technical Reports Server (NTRS)

    Connell, John W. (Inventor); Smith, Joseph G., Jr. (Inventor); Hergenrother, Paul M. (Inventor)

    2002-01-01

    Phenylethynyl containing reactive additives were prepared from aromatic diamines containing phenylethynyl groups and various ratios of phthalic anhydride and 4-phenylethynylphthalic anhydride in glacial acetic acid to form the imide in one step or in N-methyl-2-pyrrolidi none to form the amide acid intermediate. The reactive additives were mixed in various amounts (10% to 90%) with oligomers containing either terminal or pendent phenylethynyl groups (or both) to reduce the melt viscosity and thereby enhance processability. Upon thermal cure, the additives react and become chemically incorporated into the matrix and effect an increase in crosslink density relative to that of the host resin. This resultant increase in crosslink density has advantageous consequences on the cured resin properties such as higher glass transition temperature and higher modulus as compared to that of the host resin.

  1. Additives in plastics.

    PubMed Central

    Deanin, R D

    1975-01-01

    The polymers used in plastics are generally harmless. However, they are rarely used in pure form. In almost all commercial plastics, they are "compounded" with monomeric ingredients to improve their processing and end-use performance. In order of total volume used, these monomeric additives may be classified as follows: reinforcing fibers, fillers, and coupling agents; plasticizers; colorants; stabilizers (halogen stabilizers, antioxidants, ultraviolet absorbers, and biological preservatives); processing aids (lubricants, others, and flow controls); flame retardants, peroxides; and antistats. Some information is already available, and much more is needed, on potential toxicity and safe handling of these additives during processing and manufacture of plastics products. PMID:1175566

  2. Additives in plastics.

    PubMed

    Deanin, R D

    1975-06-01

    The polymers used in plastics are generally harmless. However, they are rarely used in pure form. In almost all commercial plastics, they are "compounded" with monomeric ingredients to improve their processing and end-use performance. In order of total volume used, these monomeric additives may be classified as follows: reinforcing fibers, fillers, and coupling agents; plasticizers; colorants; stabilizers (halogen stabilizers, antioxidants, ultraviolet absorbers, and biological preservatives); processing aids (lubricants, others, and flow controls); flame retardants, peroxides; and antistats. Some information is already available, and much more is needed, on potential toxicity and safe handling of these additives during processing and manufacture of plastics products.

  3. Spatial models of Northern Bobwhite populations for conservation planning

    USGS Publications Warehouse

    Twedt, Daniel J.; Wilson, R. Randy; Keister, Amy S.

    2007-01-01

    Since 1980, northern bobwhite (Colinus virginianus) range-wide populations declined 3.9% annually. Within the West Gulf Coastal Plain Bird Conservation Region in the south-central United States, populations of this quail species have declined 6.8% annually. These declines sparked calls for land use change and prompted implementation of various conservation practices. However, to effectively reverse these declines and restore northern bobwhite to their former population levels, habitat conservation and management efforts must target establishment and maintenance of sustainable populations. To provide guidance for conservation and restoration of habitat capable of supporting sustainable northern bobwhite populations in the West Gulf Coastal Plain, we modeled their spatial distribution using landscape characteristics derived from 1992 National Land Cover Data and bird detections, from 1990 to 1994, along 10-stop Breeding Bird Survey route segments. Four landscape metrics influenced detections of northern bobwhite: detections were greater in areas with more grassland and increased aggregation of agricultural lands, but detections were reduced in areas with increased density of land cover edge and grassland edge. Using these landscape metrics, we projected the abundance and spatial distribution of northern bobwhite populations across the entire West Gulf Coastal Plain. Predicted populations closely approximated abundance estimates from a different cadre of concurrently collected data but model predictions did not accurately reflect bobwhite detections along species-specific call-count routes in Arkansas and Louisiana. Using similar methods, we also projected northern bobwhite population distribution circa 1980 based on Land Use Land Cover data and bird survey data from 1976 to 1984. We compared our 1980 spatial projections with our spatial estimate of 1992 populations to identify areas of population change. Additionally, we used our projection of the spatial

  4. Biobased lubricant additives

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Fully biobased lubricants are those formulated using all biobased ingredients, i.e. biobased base oils and biobased additives. Such formulations provide the maximum environmental, safety, and economic benefits expected from a biobased product. Currently, there are a number of biobased base oils that...

  5. More Than Additional Space...

    ERIC Educational Resources Information Center

    CEFP Journal, 1973

    1973-01-01

    A much needed addition to the Jamestown Elementary School turned out to be more than an expansion of walls for more space. A new educational program, a limited budget, and a short time line were tackled on a team approach basis and were successfully resolved. (Author)

  6. Trends in Northern Hemisphere surface cyclone frequency and intensity

    USGS Publications Warehouse

    McCabe, G.J.; Clark, M.P.; Serreze, M.C.

    2001-01-01

    One of the hypothesized effects of global warming from increasing concentrations of greenhouse gases is a change in the frequency and/or intensity of extratropical cyclones. In this study, winter frequencies and intensities of extratropical cyclones in the Northern Hemisphere for the period 1959-97 are examined to determine if identifiable trends are occurring. Results indicate a statistically significant decrease in midlatitude cyclone frequency and a significant increase in high-latitude cyclone frequency. In addition, storm intensity has increased in both the high and midlatitudes. The changes in storm frequency correlate with changes in winter Northern Hemisphere temperature and support hypotheses that global warming may result in a northward shift of storm tracks in the Northern Hemisphere.

  7. Northern Eurasian Heat Waves and Droughts

    NASA Technical Reports Server (NTRS)

    Schubert, Siegfried; Wang, Hailan; Koster, Randal; Suarez, Max; Groisman, Pavel

    2013-01-01

    This article reviews our understanding of the characteristics and causes of northern Eurasian summertime heat waves and droughts. Additional insights into the nature of temperature and precipitation variability in Eurasia on monthly to decadal time scales and into the causes and predictability of the most extreme events are gained from the latest generation of reanalyses and from supplemental simulations with the NASA GEOS-5 AGCM. Key new results are: 1) the identification of the important role of summertime stationary Rossby waves in the development of the leading patterns of monthly Eurasian surface temperature and precipitation variability (including the development of extreme events such as the 2010 Russian heat wave), 2) an assessment of the mean temperature and precipitation changes that have occurred over northern Eurasia in the last three decades and their connections to decadal variability and global trends in SST, and 3) the quantification (via a case study) of the predictability of the most extreme simulated heat wave/drought events, with some focus on the role of soil moisture in the development and maintenance of such events. A literature survey indicates a general consensus that the future holds an enhanced probability of heat waves across northern Eurasia, while there is less agreement regarding future drought, reflecting a greater uncertainty in soil moisture and precipitation projections. Substantial uncertainties remain in our understanding of heat waves and drought, including the nature of the interactions between the short-term atmospheric variability associated with such extremes and the longer-term variability and trends associated with soil moisture feedbacks, SST anomalies, and an overall warming world.

  8. Discovery of novel hematopoietic cell adhesion molecules from human bone marrow stromal cell membrane protein extracts by a new cell-blotting technique.

    PubMed

    Seshi, B

    1994-05-01

    In an attempt to define the role of cell adhesion molecules (CAMs) within the bone marrow (BM) microenvironment in normal hematopoiesis and in leukemia development, a novel cell-blotting technique that involved cell adhesion to protein bands after separation by lithium dodecyl sulfate-polyacrylamide gel electrophoresis (LDS-PAGE) and blotting onto polyvinylidene difluoride (PVDF) membrane has been developed. Human BM stromal cell membrane fractions have been prepared from Dexter-type cultures after cell lysis by sonification and differential centrifugations of the sonification contents. The 20,000 g pellets representing membrane fractions have been solubilized by 2% Triton X-100, 0.575% LDS, and 8 mol/L urea in sequential order. The protein extracts are fractionated by LDS-PAGE and screened for CAMs by the new cell-blotting technique. This led to identification of nine protein bands in lanes containing LDS extracts showing adhesion of KG1a (CD34+ progenitor myeloid) cells. Evidence that the BM proteins exhibiting KG1a cell adhesion are novel CAMs is based on the observations that these proteins, in comparison with known CAMs, specifically VCAM-1, CD54, and CD44, show (1) contrasting detergent-solubility properties, (2) different temperature requirement for mediating cell adhesion function, and (3) markedly distinct electrophoretic mobilities. The various cell types tested, notably KG1a, NALM-6, WIL-2, Ramos, HS-Sultan, K562, JY B lymphoblastoid cells, and T lymphoblasts, showed distinctive patterns of binding to different subsets of BM CAMs. These results demonstrate a new approach to studies of molecular mechanisms that may determine specificity of hematopoietic cellular localization within BM microenvironment and may play an important role in controlling hematopoiesis.

  9. Reverse Cross Blot Hybridization Assay for Rapid Detection of PCR-Amplified DNA from Candida Species, Cryptococcus neoformans, and Saccharomyces cerevisiae in Clinical Samples

    PubMed Central

    Posteraro, Brunella; Sanguinetti, Maurizio; Masucci, Luca; Romano, Lucio; Morace, Giulia; Fadda, Giovanni

    2000-01-01

    A PCR-based assay was developed to detect and identify medically important yeasts in clinical samples. Using a previously described set of primers (G. Morace et al., J. Clin. Microbiol. 35:667–672, 1997), we amplified a fragment of the ERG11 gene for cytochrome P-450 lanosterol 14α-demethylase, a crucial enzyme in the biosynthesis of ergosterol. The PCR product was analyzed in a reverse cross blot hybridization assay with species-specific probes directed to a target region of the ERG11 gene of Candida albicans (pCal), C. guilliermondii (pGui), C. (Torulopsis) glabrata (pGla), C. kefyr (pKef), C. krusei (pKru), C. parapsilosis (pPar), C. tropicalis (pTro), the newly described species C. dubliniensis (pDub), Saccharomyces cerevisiae (pSce), and Cryptococcus neoformans (pCry). The PCR-reverse cross blot hybridization assay correctly identified multiple isolates of each species tested. No cross-hybridization was detected with any other fungal, bacteria, or human DNAs tested. The method was tested against conventional identification on 140 different clinical samples, including blood and cerebrospinal fluid, from patients with suspected fungal infections. The results agreed with those of culture and phenotyping for all but six specimens (two of which grew yeasts not included in the PCR panel of probes and four in which PCR positivity-culture negativity was justified by clinical findings). Species identification time was reduced from a mean of 4 days with conventional identification to 7 h with the molecular method. The PCR-reverse cross blot hybridization assay is a rapid method for the direct detection and identification of yeasts in clinical samples. PMID:10747151

  10. The epidemiology of tick-borne haemoparasites as determined by the reverse line blot hybridization assay in an intensively studied cohort of calves in western Kenya

    PubMed Central

    Njiiri, Nyawira E.; Bronsvoort, B. Mark deC.; Collins, Nicola E.; Steyn, Helena C.; Troskie, Milana; Vorster, Ilse; Thumbi, S.M.; Sibeko, Kgomotso P.; Jennings, Amy; van Wyk, Ilana Conradie; Mbole-Kariuki, Mary; Kiara, Henry; Poole, E. Jane; Hanotte, Olivier; Coetzer, Koos; Oosthuizen, Marinda C.; Woolhouse, Mark; Toye, Philip

    2015-01-01

    The development of sensitive surveillance technologies using PCR-based detection of microbial DNA, such as the reverse line blot assay, can facilitate the gathering of epidemiological information on tick-borne diseases, which continue to hamper the productivity of livestock in many parts of Africa and elsewhere. We have employed a reverse line blot assay to detect the prevalence of tick-borne parasites in an intensively studied cohort of indigenous calves in western Kenya. The calves were recruited close to birth and monitored for the presence of infectious disease for up to 51 weeks. The final visit samples from 453 calves which survived for the study period were analyzed by RLB. The results indicated high prevalences of Theileria mutans (71.6%), T. velifera (62.8%), Anaplasma sp. Omatjenne (42.7%), A. bovis (39.9%), Theileria sp. (sable) (32.7%), T. parva (12.9%) and T. taurotragi (8.5%), with minor occurrences of eight other haemoparasites. The unexpectedly low prevalence of the pathogenic species Ehrlichia ruminantium was confirmed by a species-specific PCR targeting the pCS20 gene region. Coinfection analyses of the seven most prevalent haemoparasites indicated that they were present as coinfections in over 90% of the cases. The analyses revealed significant associations between several of the Theileria parasites, in particular T. velifera with Theileria sp. sable and T. mutans, and T. parva with T. taurotragi. There was very little coinfection of the two most common Anaplasma species, although they were commonly detected as coinfections with the Theileria parasites. The comparison of reverse line blot and serological results for four haemoparasites (T. parva, T. mutans, A. marginale and B. bigemina) indicated that, except for the mostly benign T. mutans, indigenous cattle seem capable of clearing infections of the three other, pathogenic parasites to below detectable levels. Although the study site was located across four agroecological zones, there was

  11. Titan's Stratospheric Condensibles at High Northern Latitudes During Northern Winter

    NASA Technical Reports Server (NTRS)

    Anderson, Carrie; Samuelson, R.; Achterberg, R.

    2012-01-01

    The Infrared Interferometer Spectrometer (IRIS) instrument on board Voyager 1 caught the first glimpse of an unidentified particulate feature in Titan's stratosphere that spectrally peaks at 221 per centimeter. Until recently, this feature that we have termed 'the haystack,' has been seen persistently at high northern latitudes with the Composite Infrared Spectrometer (CIRS) instrument onboard Cassini, The strength of the haystack emission feature diminishes rapidly with season, becoming drastically reduced at high northern latitudes, as Titan transitions from northern winter into spring, In contrast to IRIS whose shortest wavenumber was 200 per centimeter, CIRS extends down to 10 per centimeter, thus revealing an entirely unexplored spectral region in which nitrile ices have numerous broad lattice vibration features, Unlike the haystack, which is only found at high northern latitudes during northern winter/early northern spring, this geometrically thin nitrile cloud pervades Titan's lower stratosphere, spectrally peaking at 160 per centimeter, and is almost global in extent spanning latitudes 85 N to 600 S, The inference of nitrile ices are consistent with the highly restricted altitude ranges over which these features are observed, and appear to be dominated by a mixture of HCN and HC3N, The narrow range in altitude over which the nitrile ices extend is unlike the haystack, whose vertical distribution is significantly broader, spanning roughly 70 kilometers in altitude in Titan's lower stratosphere, The nitrile clouds that CIRS observes are located in a dynamically stable region of Titan's atmosphere, whereas CH4 clouds, which ordinarily form in the troposphere, form in a more dynamically unstable region, where convective cloud systems tend to occur. In the unusual situation where Titan's tropopause cools significantly from the HASI 70.5K temperature minimum, CH4 should condense in Titan's lower stratosphere, just like the aforementioned nitrile clouds, although

  12. Titan's stratospheric condensibles at high northern latitudes during northern winter

    NASA Astrophysics Data System (ADS)

    Anderson, C.; Samuelson, R.; Achterberg, R.

    2012-04-01

    The Infrared Interferometer Spectrometer (IRIS) instrument on board Voyager 1 caught the first glimpse of an unidentified particulate feature in Titan’s stratosphere that spectrally peaks at 221 cm-1. Until recently, this feature that we have termed ‘the haystack,’ has been seen persistently at high northern latitudes with the Composite Infrared Spectrometer (CIRS) instrument onboard Cassini. The strength of the haystack emission feature diminishes rapidly with season, becoming drastically reduced at high northern latitudes, as Titan transitions from northern winter into spring. In contrast to IRIS whose shortest wavenumber was 200 cm-1, CIRS extends down to 10 cm-1, thus revealing an entirely unexplored spectral region in which nitrile ices have numerous broad lattice vibration features. Unlike the haystack, which is only found at high northern latitudes during northern winter/early northern spring, this geometrically thin nitrile cloud pervades Titan’s lower stratosphere, spectrally peaking at 160 cm-1, and is almost global in extent spanning latitudes 85°N to 60°S. The inference of nitrile ices are consistent with the highly restricted altitude ranges over which these features are observed, and appear to be dominated by a mixture of HCN and HC3N. The narrow range in altitude over which the nitrile ices extend is unlike the haystack, whose vertical distribution is significantly broader, spanning roughly 70 km in altitude in Titan’s lower stratosphere. The nitrile clouds that CIRS observes are located in a dynamically stable region of Titan’s atmosphere, whereas CH4 clouds, which ordinarily form in the troposphere, form in a more dynamically unstable region, where convective cloud systems tend to occur. In the unusual situation where Titan’s tropopause cools significantly from the HASI 70.5K temperature minimum, CH4 should condense in Titan’s lower stratosphere, just like the aforementioned nitrile clouds, although in significantly larger

  13. Vinyl capped addition polyimides

    NASA Technical Reports Server (NTRS)

    Vannucci, Raymond D. (Inventor); Malarik, Diane C. (Inventor); Delvigs, Peter (Inventor)

    1991-01-01

    Polyimide resins (PMR) are generally useful where high strength and temperature capabilities are required (at temperatures up to about 700 F). Polyimide resins are particularly useful in applications such as jet engine compressor components, for example, blades, vanes, air seals, air splitters, and engine casing parts. Aromatic vinyl capped addition polyimides are obtained by reacting a diamine, an ester of tetracarboxylic acid, and an aromatic vinyl compound. Low void materials with improved oxidative stability when exposed to 700 F air may be fabricated as fiber reinforced high molecular weight capped polyimide composites. The aromatic vinyl capped polyimides are provided with a more aromatic nature and are more thermally stable than highly aliphatic, norbornenyl-type end-capped polyimides employed in PMR resins. The substitution of aromatic vinyl end-caps for norbornenyl end-caps in addition polyimides results in polymers with improved oxidative stability.

  14. Electrophilic addition of astatine

    SciTech Connect

    Norseev, Yu.V.; Vasaros, L.; Nhan, D.D.; Huan, N.K.

    1988-03-01

    It has been shown for the first time that astatine is capable of undergoing addition reactions to unsaturated hydrocarbons. A new compound of astatine, viz., ethylene astatohydrin, has been obtained, and its retention numbers of squalane, Apiezon, and tricresyl phosphate have been found. The influence of various factors on the formation of ethylene astatohydrin has been studied. It has been concluded on the basis of the results obtained that the univalent cations of astatine in an acidic medium is protonated hypoastatous acid.

  15. Functional Generalized Additive Models.

    PubMed

    McLean, Mathew W; Hooker, Giles; Staicu, Ana-Maria; Scheipl, Fabian; Ruppert, David

    2014-01-01

    We introduce the functional generalized additive model (FGAM), a novel regression model for association studies between a scalar response and a functional predictor. We model the link-transformed mean response as the integral with respect to t of F{X(t), t} where F(·,·) is an unknown regression function and X(t) is a functional covariate. Rather than having an additive model in a finite number of principal components as in Müller and Yao (2008), our model incorporates the functional predictor directly and thus our model can be viewed as the natural functional extension of generalized additive models. We estimate F(·,·) using tensor-product B-splines with roughness penalties. A pointwise quantile transformation of the functional predictor is also considered to ensure each tensor-product B-spline has observed data on its support. The methods are evaluated using simulated data and their predictive performance is compared with other competing scalar-on-function regression alternatives. We illustrate the usefulness of our approach through an application to brain tractography, where X(t) is a signal from diffusion tensor imaging at position, t, along a tract in the brain. In one example, the response is disease-status (case or control) and in a second example, it is the score on a cognitive test. R code for performing the simulations and fitting the FGAM can be found in supplemental materials available online.

  16. Primary Science in Northern Ireland

    ERIC Educational Resources Information Center

    McAllister, Peter

    2005-01-01

    Since 1990 the science curriculum in Northern Ireland has gone through three major changes. In the beginning, fifteen attainment targets were introduced to an unsuspecting and largely unprepared teaching population: these were eventually reduced to five in 1993 and then to the present two in 1996. Unlike in England, technology has never stood as…

  17. "Subtractive" Bilingualism in Northern Belize.

    ERIC Educational Resources Information Center

    Rubinstein, Robert A.

    "Subtractive" bilingualism in Northern Belize is analyzed based on an extension of a model by Wallace Lambert. The impact of English language instruction on Spanish speaking children in Corozal Town, the northernmost urban center in the British colony of Belize, Central America, is described. This description extends an earlier account…

  18. Motivating Learners in Northern Communities.

    ERIC Educational Resources Information Center

    Swanson, Sharon

    2003-01-01

    A teacher at a northern Ontario adult Native literacy program describes how she cultivates student motivation. The teacher-student relationship is the most influential factor for motivating students. By focusing on cultural awareness, cultural teaching practices, and a sense of community, teachers can help students be successful. Program…

  19. Opsin gene repertoires in northern archaic hominids.

    PubMed

    Taylor, John S; Reimchen, Thomas E

    2016-08-01

    The Neanderthals' northern distribution, hunting techniques, and orbit breadths suggest that they were more active in dim light than modern humans. We surveyed visual opsin genes from four Neanderthals and two other archaic hominids to see if they provided additional support for this hypothesis. This analysis was motivated by the observation that alleles responsible for anomalous trichromacy in humans are more common in northern latitudes, by data suggesting that these variants might enhance vision in mesopic conditions, and by the observation that dim light active species often have fewer opsin genes than diurnal relatives. We also looked for evidence of convergent amino acid substitutions in Neanderthal opsins and orthologs from crepuscular or nocturnal species. The Altai Neanderthal, the Denisovan, and the Ust'-Ishim early modern human had opsin genes that encoded proteins identical to orthologs in the human reference genome. Opsins from the Vindija Cave Neanderthals (three females) had many nonsynonymous substitutions, including several predicted to influence colour vision (e.g., stop codons). However, the functional implications of these observations were difficult to assess, given that "control" loci, where no substitutions were expected, differed from humans to the same extent. This left unresolved the test for colour vision deficiencies in Vindija Cave Neanderthals.

  20. Northern New Mexico regional airport market feasibility

    SciTech Connect

    Drake, R.H.; Williams, D.S.

    1998-06-01

    This report is about the market for airline travel in northern New Mexico. Interest in developing a northern New Mexico regional airport has periodically surfaced for a number of years. The New Mexico State Legislature passed a memorial during the 1998 Second Session calling for the conduct of a study to determine the feasibility of building a new regional airport in NNM. This report is a study of the passenger market feasibility of such an airport. In addition to commercial passenger market feasibility, there are other feasibility issues dealing with siting, environmental impact, noise, economic impact, intermodal transportation integration, region-wide transportation services, airport engineering requirements, and others. These other feasibility issues are not analyzed in any depth in this report although none were discovered to be show-stoppers as a by-product of the authors doing research on the passenger market itself. Preceding the need for a detailed study of these other issues is the determination of the basic market need for an airport with regular commercial airline service in the first place. This report is restricted to an in-depth look at the market for commercial passenger air service in NNM. 20 figs., 8 tabs.

  1. The Northern Walker Lane Seismic Refraction Experiment

    NASA Astrophysics Data System (ADS)

    Louie, J. N.; Smith, S. B.; Thelen, W.; Scott, J. B.; Clark, M.

    2002-12-01

    We are developing a three-dimensional reference seismic velocity model for the western Great Basin region of Nevada and eastern California. The northern Walker Lane had not been characterized well by previous work. In May 2002 we collected a new crustal refraction profile from Battle Mountain, Nev. across western Nevada, the Reno area, Lake Tahoe, and the northern Sierra to Auburn, Calif. Mine blasts and earthquakes were recorded by 199 Texan instruments (loaned by the PASSCAL Instrument Center) extending across this more than 450-km-long transect. The seismic sources at the eastern end were mining blasts at Barrick's GoldStrike pit. We recorded additional blasts at the Florida Canyon and other mines between Lovelock and Battle Mountain, Nevada. The GoldStrike mine produced several ripple-fired blasts using 10,000-40,000 kg of ANFO each. First arrivals from the larger blasts are obvious to distances exceeding 250 km in the raw records. A M2.4 earthquake near Bridgeport, Calif. also produced pickable P-wave arrivals across at least half the transect, providing fan-shot data. We recorded only during working hours, and so missed an M4 earthquake that occurred at night. Events of M2 occurred during our recording to the west on the San Andreas fault near Pinnacles, Calif.; M3 events occurred near Portola and Mammoth Lakes, Calif. Arrivals from M5 events in the Mariana and Kuril Islands also appear in the records. Time-picks from these earthquakes may be possible after more work on synthetic-time modeling, data filtering, and display. We plan to record blasts at quarries in the western Sierra in future experiments, for a direct refraction reversal. We will compare our time picks against times generated from regional velocity models, to identify potential crustal and upper-mantle velocity anomalies. Such anomalies may be associated with the Battle Mountain heat-flow high, the northern Walker Lane belt, or the northern Sierran block.

  2. Northern Border Pipeline Company NPDES Permit

    EPA Pesticide Factsheets

    Under NPDES permit MT-0030791, the Northern Border Pipeline Company is authorized to discharge from locations along the Northern Border Gas Transmission Pipeline located within the exterior boundaries of the Fort Peck Indian Reservation, Montana.

  3. Multiplexed quantification of plant thylakoid proteins on Western blots using lanthanide-labeled antibodies and laser ablation inductively coupled plasma mass spectrometry (LA-ICP-MS).

    PubMed

    de Bang, Thomas Christian; Pedas, Pai; Schjoerring, Jan Kofod; Jensen, Poul Erik; Husted, Søren

    2013-05-21

    We have developed a novel calibration method that allows concurrent quantification of multiple proteins by laser ablation inductively coupled plasma mass spectrometry (LA-ICP-MS) after Western blotting. Calibrants were made of nitrocellulose membranes doped with lanthanide standards. Excellent linearity was obtained in the interval from 0 to 24 ng lanthanide cm(-2). Cerium-labeled lysozyme was introduced as an internal reference protein, enabling correction for up to 50% difference in transfer efficiency during the blotting of membranes. The sensitivity of the LA-ICP-MS method was comparable to state-of-the-art chemiluminescence detection and was further improved by a factor of 20, using a polymer tag. Our method allowed reproducible and multiplexed quantification of five thylakoid proteins extracted from chloroplasts of the plant species Arabidopsis thaliana (relative standard deviation (RSD) of ≤ 5% in three independent analytical series). The method was capable of measuring the L subunit in photosystem I of an Arabidopsis mutant containing <5% of this particular protein, relative to the wild type. We conclude that the developed calibration method is highly suited for multiplexed and comparative protein studies, allowing for intermembrane comparisons with high sensitivity and reproducibility.

  4. Assessment and modification of a Western blot assay for detection of central nervous system tissue in meat products in the United States.

    PubMed

    Salman, M D; Jemmi, T; Triantis, J; Dewell, R D

    2005-08-01

    Health hazards associated with meat contaminated by the bovine spongiform encephalopathy agent have led to the development of tests for the presence of this agent. The objective of this study was to optimize a neuron-specific enolase Western blot assay for use in the United States. We compared the original test with a modified protocol to evaluate the detection limit for the presence of central nervous system (CNS) tissue in experimentally inoculated samples and compared and evaluated the utility of these tests for detecting CNS tissue in retail sausages. Sensitivity and specificity of the original and modified protocols were evaluated using the kappa statistic to assess agreement between the results of the two protocols. The original protocol resulted in 100% specificity and 92% sensitivity for raw samples and 92% specificity and 72% sensitivity for cooked samples. The modified protocol resulted in 92% specificity and 89% sensitivity for raw samples and 83% specificity and 75% sensitivity for cooked samples. The kappa statistic for protocol comparison was 0.94 for raw samples and 0.74 for cooked samples. Both protocols correctly identified CNS tissue in positive controls for each replicate. Although the Western blot technique should be considered for screening for the presence of bovine CNS tissue in meat samples, the techniques should be further optimized to address problems of low sensitivity. A test with higher sensitivity is needed to protect consumers from food safety threats associated with bovine CNS tissue.

  5. Polymerase chain reaction and Southern blot-based analysis of the C9orf72 hexanucleotide repeat in different motor neuron diseases.

    PubMed

    Hübers, Annemarie; Marroquin, Nicolai; Schmoll, Birgit; Vielhaber, Stefan; Just, Marlies; Mayer, Benjamin; Högel, Josef; Dorst, Johannes; Mertens, Thomas; Just, Walter; Aulitzky, Anna; Wais, Verena; Ludolph, Albert C; Kubisch, Christian; Weishaupt, Jochen H; Volk, Alexander E

    2014-05-01

    The GGGGCC-hexanucleotide repeat expansion in C9orf72 is the most common genetic cause of familial amyotrophic lateral sclerosis and frontotemporal dementia. This study determined the frequency of C9orf72 repeat expansions in different motor neuron diseases (amyotrophic lateral sclerosis (ALS), motor neuron diseases affecting primarily the first or the second motor neuron and hereditary spastic paraplegia). Whereas most studies on C9orf72 repeat expansions published so far rely on a polymerase chain reaction-based screening, we applied both polymerase chain reaction-based techniques and Southern blotting. Furthermore, we determined the sensitivity and specificity of Southern blotting of the C9orf72 hexanucleotide repeat in DNA derived from lymphoblastoid cell lines. C9orf72 repeat expansions were found in 27.1% out of 166 familial ALS patients, only once in 68 sporadic ALS patients, and not in 61 hereditary spastic paraplegia patients or 52 patients with motor neuron diseases affecting clinically primarily either the first or the second motor neuron. We found hints for a correlation between C9orf72 repeat length and the age of onset. Somatic instability of the C9orf72 repeat was observed in lymphoblastoid cell lines compared with DNA derived from whole blood from the same patient and therefore caution is warranted for repeat length determination in immortalized cell lines.

  6. A Comparison of Antibacterial Activity of Selected Thyme (Thymus) Species by Means of the Dot Blot Test with Direct Bioautographic Detection.

    PubMed

    Orłowska, Marta; Kowalska, Teresa; Sajewicz, Mieczysław; Jesionek, Wioleta; Choma, Irena M; Majer-Dziedzic, Barbara; Szymczak, Grażyna; Waksmundzka-Hajnos, Monika

    2015-01-01

    Bioautography carried out with the aid of thin-layer chromatographic adsorbents can be used to assess antibacterial activity in samples of different origin. It can either be used as a simple and cost-effective detection method applied to a developed chromatogram, or to the dot blot test performed on a chromatographic plate, where total antibacterial activity of a sample is scrutinized. It was an aim of this study to compare antibacterial activity of 18 thyme (Thymus) specimens and species (originating from the same gardening plot and harvested in the same period of time) by means of a dot blot test with direct bioautography. A two-step extraction of herbal material was applied, and at step two the polar fraction of secondary metabolites was obtained under the earlier optimized extraction conditions [methanol-water (27+73, v/v), 130°C]. This fraction was then tested for its antibacterial activity against Bacillus subtilis bacteria. It was established that all investigated extracts exhibited antibacterial activity, yet distinct differences were perceived in the size of the bacterial growth inhibition zones among the compared thyme species. Based on the results obtained, T. citriodorus "golden dwarf" (sample No. 5) and T. marschallianus (sample No. 6) were selected as promising targets for further investigations and possible inclusion in a herbal pharmacopeia, which is an essential scientific novelty of this study.

  7. Analysis of the insulin receptor gene in noninsulin-dependent diabetes mellitus by denaturing gradient gel blots: A clinical research center study

    SciTech Connect

    Magre, J.; Goldfine, A.B.; Warram, J.H.

    1995-06-01

    We have used a new technique of denaturing gradient gel blotting to determine the prevalence of alterations in the intracellular domain of the insulin receptor in normal individuals and subjects with non-insulin-dependent diabetes mellitus (NIDDM). This method detects DNA sequence differences as restriction fragment melting polymorphisms (RFMP) and is sensitive to changes in sequence at both restriction sites and within the fragments themselves. Using restriction digests with AluI, HaeIII, HinfI, RsaI, Sau3A, and Sau96, 12 RFMPs were found to localize to the region of the {beta}-subunit of the insulin receptor gene. Using exon-specific probes, these RFMPs could be localized to specific regions surrounding individual exons, including exons, 14, 15, 16, 18, 20, and 22. In general, linkage disequilibrium between polymorphisms was inversely related to their distance in the gene structure, although there was a {open_quotes}hot spot{close_quotes} for recombination between exons 19 and 20. No difference in melting temperatures or allele frequency was observed between NIDDM patients and controls. These data indicate that the region of the insulin receptor gene coding for the intracellular portion of the {beta}-subunit is highly polymorphic and that polymorphisms surrounding specific exons can be identified by denaturing gradient gel blotting, but there is no evidence that variation at this locus contributes to NIDDM susceptibility in most individuals. 36 refs., 3 figs., 3 tabs.

  8. Characterization of antigenic property of Toxocara canis and Toxascaris leonina adults and larvae through immunodiagnostic electrophoresis (SDS-PAGE) and western blot technique.

    PubMed

    el-Massry, A A

    1999-08-01

    Differential molecular studies were performed by sodium dodecyle sulphate polyacrylamide gel electrophoresis--SDS-PAGE--between somatic antigen of Toxocara canis and Toxascaris leonina, adults and larvae, recovered from dogs. SDS-PAGE of both adults somatic antigen showed two closely similar bands (90.00, 91.95 KDa and 69.25-70.56 KDa). Each parasite had characteristic bands clustered in different molecular weights. While for their larval antigen, T. canis showed a very different antigenic profile when analysed in comparison to T. leonina antigen except at one band (66.85-66.89 KDa). The Western blot analysis showed four prominent bands represented immunoreaction between the separated somatic antigen of T. canis adults and experimentally immunized rabbit with the corresponding parasite (125.37, 117.73, 90.00 and 69.25 KDa). While separated antigen of T. leonina adults immune reacted with the corresponding hyperimmune rabbit sera at 119.04, 91.95 and 70.56 KDa. The Western blot showed cross reactive immune bands between T. canis and T. leonina adults somatic antigen at two bands (90.00, 91.95 KDa and 69.25-70.56 KDa). The polypeptide bands reacted at 125.37 KDa and 117.73 KDa can be used as specific finger print for T. canis adults while that at 119.04 KDa was specific for T. leonina adult worm.

  9. Siloxane containing addition polyimides

    NASA Technical Reports Server (NTRS)

    Maudgal, S.; St. Clair, T. L.

    1984-01-01

    Addition polyimide oligomers have been synthesized from bis(gamma-aminopropyl) tetramethyldisiloxane and 3, 3', 4, 4'-benzophenonetetracarboxylic dianhydride using a variety of latent crosslinking groups as endcappers. The prepolymers were isolated and characterized for solubility (in amide, chlorinated and ether solvents), melt flow and cure properties. The most promising systems, maleimide and acetylene terminated prepolymers, were selected for detailed study. Graphite cloth reinforced composites were prepared and properties compared with those of graphite/Kerimid 601, a commercially available bismaleimide. Mixtures of the maleimide terminated system with Kerimid 601, in varying proportions, were also studied.

  10. Platelet additive solution - electrolytes.

    PubMed

    Azuma, Hiroshi; Hirayama, Junichi; Akino, Mitsuaki; Ikeda, Hisami

    2011-06-01

    Recent attention to solutions that replace most or all plasma in platelet concentrates, while maintaining satisfactory platelet function, is motivated by the potential of plasma reduction or depletion to mitigate various transfusion-related adverse events. This report considers the electrolytic composition of previously described platelet additive solutions, in order to draw general conclusions about what is required for platelet function and longevity. The optimal concentrations of Na(+) and Cl(-) are 69-115 mM. The presence of both K(+) and Mg(2+) in platelet suspension at nearly physiological concentrations (3-5mM and 1.5-3mM, respectively) is indispensable for good preservation capacity because both electrolytes are required to prevent platelet activation. In contrast to K(+) and Mg(2+), Ca(2+) may not be important because no free Ca(2+) is available in M-sol, which showed excellent platelet preservation capacity at less than 5% plasma concentration. The importance of bicarbonate (approximately 40 mM) can be recognized when the platelets are suspended in additive solution under less than 5% residual plasma concentration.

  11. Additive composition, for gasoline

    SciTech Connect

    Vataru, M.

    1989-01-10

    An admixture is described that comprises Diesel fuel and an additive composition added thereto which is between about 0.05 to about 2.0 percent by weight of the fuel, the composition comprising: (a) between about 0.05 and 25% relative weight parts of an organic peroxide, and (b) between about 0.1 and 25% relative weight parts of detergent selected from the component group that consists of: (i) fatty amines; (ii) ethoxylated and propoxylated derivatives of fatty amines; (iii) fatty diamines; (iv) fatty imidazlines; (v) polymeric amines and derivatives thereof; (vi) combination of one or more of the (i) through (v) components with carboxylic acid or acids having from three to forth carbon atoms, (c) from about 99.0 to about 50% by weight of a hydrocarbon solvent.

  12. Teardrop bladder: additional considerations

    SciTech Connect

    Wechsler, R.J.; Brennan, R.E.

    1982-07-01

    Nine cases of teardrop bladder (TDB) seen at excretory urography are presented. In some of these patients, the iliopsoas muscles were at the upper limit of normal in size, and additional evaluation of the perivesical structures with computed tomography (CT) was necessary. CT demonstrated only hypertrophied muscles with or without perivesical fat. The psoas muscles and pelvic width were measured in 8 patients and compared with the measurements of a control group of males without TDB. Patients with TDB had large iliopsoas muscles and narrow pelves compared with the control group. The psoas muscle width/pelvic width ratio was significantly greater (p < 0.0005) in patients with TDB than in the control group, with values of 1.04 + 0.05 and 0.82 + 0.09, respectively. It is concluded that TDB is not an uncommon normal variant in black males. Both iliopsoas muscle hypertrophy and a narrow pelvis are factors that predispose a patient to TDB.

  13. New addition curing polyimides

    NASA Technical Reports Server (NTRS)

    Frimer, Aryeh A.; Cavano, Paul

    1991-01-01

    In an attempt to improve the thermal-oxidative stability (TOS) of PMR-type polymers, the use of 1,4-phenylenebis (phenylmaleic anhydride) PPMA, was evaluated. Two series of nadic end-capped addition curing polyimides were prepared by imidizing PPMA with either 4,4'-methylene dianiline or p-phenylenediamine. The first resulted in improved solubility and increased resin flow while the latter yielded a compression molded neat resin sample with a T(sub g) of 408 C, close to 70 C higher than PME-15. The performance of these materials in long term weight loss studies was below that of PMR-15, independent of post-cure conditions. These results can be rationalized in terms of the thermal lability of the pendant phenyl groups and the incomplete imidization of the sterically congested PPMA. The preparation of model compounds as well as future research directions are discussed.

  14. Perspectives on Additive Manufacturing

    NASA Astrophysics Data System (ADS)

    Bourell, David L.

    2016-07-01

    Additive manufacturing (AM) has skyrocketed in visibility commercially and in the public sector. This article describes the development of this field from early layered manufacturing approaches of photosculpture, topography, and material deposition. Certain precursors to modern AM processes are also briefly described. The growth of the field over the last 30 years is presented. Included is the standard delineation of AM technologies into seven broad categories. The economics of AM part generation is considered, and the impacts of the economics on application sectors are described. On the basis of current trends, the future outlook will include a convergence of AM fabricators, mass-produced AM fabricators, enabling of topology optimization designs, and specialization in the AM legal arena. Long-term developments with huge impact are organ printing and volume-based printing.

  15. Sewage sludge additive

    NASA Technical Reports Server (NTRS)

    Kalvinskas, J. J.; Mueller, W. A.; Ingham, J. D. (Inventor)

    1980-01-01

    The additive is for a raw sewage treatment process of the type where settling tanks are used for the purpose of permitting the suspended matter in the raw sewage to be settled as well as to permit adsorption of the dissolved contaminants in the water of the sewage. The sludge, which settles down to the bottom of the settling tank is extracted, pyrolyzed and activated to form activated carbon and ash which is mixed with the sewage prior to its introduction into the settling tank. The sludge does not provide all of the activated carbon and ash required for adequate treatment of the raw sewage. It is necessary to add carbon to the process and instead of expensive commercial carbon, coal is used to provide the carbon supplement.

  16. A theoretical timeline for myocardial infarction: immunohistochemical evaluation and western blot quantification for Interleukin-15 and Monocyte chemotactic protein-1 as very early markers

    PubMed Central

    2014-01-01

    Background Experimental and human studies have demonstrated that innate immune mechanisms and consequent inflammatory reaction play a critical role in cardiac response to ischemic injury. Thus, the detection of immuno-inflammatory and cellular phenomena accompanying cardiac alterations during the early inflammatory phase of myocardial infarction (MI) may be an excellent diagnostic tool. Current knowledge of the chronology of the responses of myocardial tissue following the occurrence of ischemic insult, as well as the existence of numerous studies aiming to identify reliable markers in dating MI, induced us to investigate the myocardial specimens of MI fatal cases in order to better define the age of MI. Methods We performed an immunohistochemical study and a Western blot analysis to evaluate detectable morphological changes in myocardial specimens of fatal MI cases and to quantify the effects of cardiac expression of inflammatory mediators (CD15, IL-1β, IL-6, TNF-α, IL-15, IL-8, MCP-1, ICAM-1, CD18, tryptase) and structural and functional cardiac proteins. Results We observed a biphasic course of MCP-1: it was strongly expressed in the very early phase (0-4 hrs), to diminish in the early period (after 6-8 hrs). Again, our choice of IL-15 is explained by the synergism with neutrophilic granulocytes (CD15) and our study shows the potential for striking cytokine synergy in promoting fast, local neutrophil response in damaged tissues. A progressively stronger immunoreaction for the CD15 antibody was visible in the areas where the margination of circulating inflammatory cells was detectable, up to very strong expression in the oldest ones (>12 hours). Further, the induction of CD15, IL-15, MCP-1 expression levels was quantified by Western blot analysis. The results were as follows: IL-15/β-actin 0.80, CD15/β-actin 0.30, and MCP-1/β-actin 0.60, matching perfectly with the results of immunohistochemistry. Control hearts from traumatic death cases did not show any

  17. Detection of immunoglobulin M antibodies to glycoprotein G-2 by western blot (immunoblot) for diagnosis of initial herpes simplex virus type 2 genital infections.

    PubMed Central

    Ho, D W; Field, P R; Irving, W L; Packham, D R; Cunningham, A L

    1993-01-01

    Western blots (immunoblots) for the detection of immunoglobulin M (IgM) antibodies specific for herpes simplex virus type 1 (HSV-1) and HSV-2 in patients' sera were developed. The locations of the type-specific glycoprotein G (gpG-2) of HSV-2 (92- and 140-kDa forms) and glycoprotein C of HSV-1 (gpC-1), which carries mostly type-specific antigenic epitopes, were checked with specific monoclonal antibodies. Western blot assays for IgM antibody to gpC-1 or gpG-2 were performed after depletion of IgG by precipitation with anti-human IgG. In patients with primary HSV-2 genital infections, seroconversion of IgM and IgG antibodies to both the 92- and 140-kDa forms of gpG-2 was observed, although both antibodies appeared in convalescent-phase serum after the first week. IgM and IgG antibodies to low-molecular-size polypeptides (40 to 65 kDa) were the first antibodies observed in patients with primary infection, but these antibodies were cross-reactive with HSV-1 and HSV-2. However, in patients with recurrent HSV-2 infections, IgG antibodies to both forms of gpG-2 and the low-molecular-size polypeptides were found no matter how early after onset the patient was bled, and IgM to gpG-2 did not appear. In patients with nonprimary initial genital HSV-2 infections, IgG antibody to HSV-1 was demonstrated in the first serum specimen, and HSV-2-specific IgM was found in 39% of the serum specimens. Hence, the Western blot assay can be used to test for IgM antibody to gpG-2, allowing for the retrospective diagnosis of inital HSV-2 infections and its use as a supplementary test to the gpG-2 IgG enzyme-linked immunosorbent assays developed elsewhere. In contrast, IgM antibody to gpG-2 is not usually detected in patients with recurrent HSV-2 infections. Images PMID:7508453

  18. A simple, inexpensive, robust and sensitive dot-blot assay for equal detection of the nonstructural-1 glycoprotein of all dengue virus serotypes

    PubMed Central

    2013-01-01

    Background Detection of dengue virus (DENV) soluble/excreted (s/e) form of the nonstructural-1 (NS1) glycoprotein in patient acute-phase sera is ideal for diagnosis. The commercially-available detection assays are, however, too expensive for routine use and have low specificity, particularly for the s/e NS1 glycoprotein of DENV-2 and DENV-4, which are important causes of lethal human disease worldwide. Methods Mouse monoclonal antibodies (MAbs) were generated and screened against s/e NS1 glycoprotein purified from each DENV serotype to obtain those that reacted equally with each serotype, but not with yellow fever virus (YFV) s/e NS1 glycoprotein or human serum proteins. One MAb, MAb 2C4.6, was further tested against these DENV glycoproteins in human sera using simple, peroxidase-labelled secondary antibody/substrate-developed dot-blot assays. Results Optimal quenching of endogenous human serum peroxidases was attained using 3% H2O2 in H20 for 5 min. MAb 2C4.6 showed an acceptable detection sensitivity of < 32 ng/ml for the s/e NS1 glycoprotein of each DENV serotype but did not cross-react with the YFV s/e NS1 glycoprotein or human serum proteins. By contrast, the LX1 epitope-specific MAb, 3D1.4, showed similar detection sensitivity against only the DENV-1 NS1 glycoprotein, consistent with results from commercial DENV s/e NS1 glycoprotein detection assays. DENV s/e NS1 glycoproteins were stable in human sera after drying on the nitrocellulose membranes and storage for one month at ambient temperature (28°C) before being processed. The total assay time was reduced to 3 h without any loss of detection sensitivity. This dot-blot format was ideal for the circulating immune complex disruption step, which is required for increased DENV s/e NS1 glycoprotein detection. Conclusions This is the first study to determine the detection sensitivity of MAbs against known concentrations of s/e NS1 glycoprotein from each DENV serotype. The preparation of patient serum samples for

  19. Additive lattice kirigami.

    PubMed

    Castle, Toen; Sussman, Daniel M; Tanis, Michael; Kamien, Randall D

    2016-09-01

    Kirigami uses bending, folding, cutting, and pasting to create complex three-dimensional (3D) structures from a flat sheet. In the case of lattice kirigami, this cutting and rejoining introduces defects into an underlying 2D lattice in the form of points of nonzero Gaussian curvature. A set of simple rules was previously used to generate a wide variety of stepped structures; we now pare back these rules to their minimum. This allows us to describe a set of techniques that unify a wide variety of cut-and-paste actions under the rubric of lattice kirigami, including adding new material and rejoining material across arbitrary cuts in the sheet. We also explore the use of more complex lattices and the different structures that consequently arise. Regardless of the choice of lattice, creating complex structures may require multiple overlapping kirigami cuts, where subsequent cuts are not performed on a locally flat lattice. Our additive kirigami method describes such cuts, providing a simple methodology and a set of techniques to build a huge variety of complex 3D shapes.

  20. Ceramics with Different Additives

    NASA Astrophysics Data System (ADS)

    Wang, Juanjuan; Feng, Lajun; Lei, Ali; Zhao, Kang; Yan, Aijun

    2014-09-01

    Li2CO3, MgCO3, BaCO3, and Bi2O3 dopants were introduced into CaCu3Ti4O12 (CCTO) ceramics in order to improve the dielectric properties. The CCTO ceramics were prepared by conventional solid-state reaction method. The phase structure, microstructure, and dielectric behavior were carefully investigated. The pure structure without any impurity phases can be confirmed by the x-ray diffraction patterns. Scanning Electron Microscopy (SEM) analysis illuminated that the grains of Ca0.90Li0.20Cu3Ti4O12 ceramics were greater than that of pure CCTO. It was important for the properties of the CCTO ceramics to study the additives in complex impedance spectroscopy. It was found that the Ca0.90Li0.20Cu3Ti4O12 ceramics had the higher permittivity (>45000), the lower dielectric loss (<0.025) than those of CCTO at 1 kHz at room temperature and good temperature stability from -30 to 75 °C.

  1. Additive lattice kirigami

    PubMed Central

    Castle, Toen; Sussman, Daniel M.; Tanis, Michael; Kamien, Randall D.

    2016-01-01

    Kirigami uses bending, folding, cutting, and pasting to create complex three-dimensional (3D) structures from a flat sheet. In the case of lattice kirigami, this cutting and rejoining introduces defects into an underlying 2D lattice in the form of points of nonzero Gaussian curvature. A set of simple rules was previously used to generate a wide variety of stepped structures; we now pare back these rules to their minimum. This allows us to describe a set of techniques that unify a wide variety of cut-and-paste actions under the rubric of lattice kirigami, including adding new material and rejoining material across arbitrary cuts in the sheet. We also explore the use of more complex lattices and the different structures that consequently arise. Regardless of the choice of lattice, creating complex structures may require multiple overlapping kirigami cuts, where subsequent cuts are not performed on a locally flat lattice. Our additive kirigami method describes such cuts, providing a simple methodology and a set of techniques to build a huge variety of complex 3D shapes. PMID:27679822

  2. Investigation of cross-reactions against Trichinella spiralis antigens by enzyme-linked immunosorbent assay and enzyme-linked immunoelectrotransfer blot assay in patients with various diseases.

    PubMed Central

    De-la-Rosa, J L; Alcantara, P; Correa, D

    1995-01-01

    Data regarding cross-reactions against Trichinella spiralis in humans are scarce and controversial. For this reason, we tested serum samples from patients with typhoid fever, brucellosis, toxoplasmosis, amoebiasis, cysticercosis, trichocephaliasis, ascariasis, and onchocerciasis against an antigenic extract of T. spiralis infective larvae in an enzyme-linked immunosorbent assay (ELISA) and an enzyme-linked immunoelectrotransfer blot (EITB) assay. All except one serum sample from the group of patients with onchocerciasis were negative in the ELISA; in the EITB assay, only faint bands were observed with the samples from patients with onchocerciasis and ascariasis and negative results were obtained with the samples from patients with other diseases. In conclusion, cross-reactions were found only in the groups of patients with other nematode infections and were of very low magnitude, most of them virtually negative. PMID:7719905

  3. Detection of Cryptosporidium parvum oocysts by dot-blotting using monoclonal antibodies to Cryptosporidium parvum virus 40-kDa capsid protein.

    PubMed

    Jenkins, Mark C; O'Brien, Celia N; Trout, James M

    2008-02-01

    Monoclonal antibodies (MAb) were prepared against the 40-kDa capsid protein of Cryptosporidium parvum virus (CPV) by immunizing mice with purified recombinant CPV40 protein. In immunoblotting analysis, MAbCPV40-1 bound to a 40-kDa protein in extracts of C. parvum oocysts. This 40-kDa protein was localized in the sporozoite cytoplasm by immunofluorescence (IFA) staining with MAbCPV40-1. In a dot-blot assay, MAbCPV40-1 was capable of detecting 10(2) non-bleach-treated and 10(2)-10(3) bleach-treated C. parvum oocysts. MAbCPV40-1 was capable of detecting CPV40 antigen in both soluble and total C. parvum oocyst protein extracts, indicating a potential use for detecting this parasite in environmental samples.

  4. Identification of Helicobacter pylori by immunological dot blot method based on reaction of a species-specific monoclonal antibody with a surface-exposed protein.

    PubMed Central

    Bölin, I; Lönroth, H; Svennerholm, A M

    1995-01-01

    Monoclonal antibodies (MAbs) against membrane preparations of Helicobacter pylori were produced. One MAb was found to be specific for H. pylori, because it did not react with a number of other bacterial species, including Helicobacter felis and Campylobacter jejuni. This MAb reacted with a 30-kDa protein found in outer membrane preparations of H. pylori. The protein was also detected on the cell surface on intact bacteria when analyzed by immunoelectron microscopy. To facilitate the identification of H. pylori isolates after culturing of biopsies, an immunodot blot assay based on the reaction of this MAb was developed. This assay was found to be highly specific for H. pylori. Sixty-six clinical isolates typed as H. pylori by conventional biochemical tests were found to be positive, whereas no other bacterial species tested gave a positive result. By this method, reliable and rapid identification of H. pylori could be accomplished. PMID:7714196

  5. Serum detection of IgG antibodies against Demodex canis by western blot in healthy dogs and dogs with juvenile generalized demodicosis.

    PubMed

    Ravera, Ivan; Ferreira, Diana; Gallego, Laia Solano; Bardagí, Mar; Ferrer, Lluís

    2015-08-01

    The aim of this study was to investigate the presence of canine immunoglobulins (Ig) G against Demodex proteins in the sera of healthy dogs and of dogs with juvenile generalized demodicosis (CanJGD) with or without secondary pyoderma. Demodex mites were collected from dogs with CanJGD. Protein concentration was measured and a western blot technique was performed. Pooled sera from healthy dogs reacted mainly with antigen bands ranging from 55 to 72 kDa. Pooled sera from dogs with CanJGD without secondary pyoderma reacted either with 10 kDa antigen band or 55 to 72 kDa bands. Pooled sera from dogs with CanJGD with secondary pyoderma reacted only with a 10 kDa antigen band. The results of this study suggest that both healthy dogs and dogs with CanJGD develop a humoral response against different proteins of Demodex canis.

  6. Application of glyco-blotting for identification of structures of polysaccharides causing membrane fouling in a pilot-scale membrane bioreactor treating municipal wastewater.

    PubMed

    Kimura, Katsuki; Nishimura, Shin-Ichiro; Miyoshi, Risho; Hoque, Asiful; Miyoshi, Taro; Watanabe, Yoshimasa

    2015-03-01

    A new approach for the analysis of polysaccharides in membrane bioreactor (MBR) is proposed in this study. Enrichment of polysaccharides by glyco-blotting, in which polysaccharides are specifically collected via interactions between the aldehydes in the polysaccharides and aminooxy groups on glycoblotting beads, enabled MALDI-TOF/MS analysis at a high resolution. Structures of polysaccharides extracted from fouled membranes used in a pilot-scale MBR treating municipal wastewater and those in the supernatant of the mixed liquor suspension in the MBR were investigated. It was found that the overlap between polysaccharides found in the supernatants and those extracted from the fouled membrane was rather limited, suggesting that polysaccharides that dominate in supernatants may not be important in membrane fouling in MBRs. Analysis using a bacterial carbohydrate database suggested that capsular polysaccharides (CPS) and/or lipo-polysaccharides (LPS) produced by gram-negative bacteria are key players in the evolution of membrane fouling in MBRs.

  7. Detection and chromosomal assignment of SV40-DNA integration in Chinese hamster cell lines by chromosome sorting and dot blot hybridization.

    PubMed

    Hutter, K J; Klefenz, H; Goerttler, K

    1990-01-01

    A combination of cytometric (chromosome sorting), molecular (dot blot hybridization using radio-active and/or biotinylated DNA probes) and cytogenetic (G-banding) evaluation is described which allows the rapid identification of single copy and repetitive viral integrates and their assignment to chromosome groups or even individual chromosomes. In the case of Chinese hamster cell line CO 631 it could be demonstrated that SV40 DNA was solely integrated into a submetacentric marker chromosome. Such a cytometric/molecular/cytogenetic "identogram" may prove to be a useful tool in many areas of cell and tumor biology. Furthermore, amounts of chromosomes sufficient for analysis as well as subsequent cloning experiments can be accumulated.

  8. Hammersley Range, northern Western Australia

    NASA Technical Reports Server (NTRS)

    1990-01-01

    The oval shaped basin of the sedimentary rocks of the Hammersley Range, northern Western Australia (23.0S, 119.0E) dominates the center of this near nadir view. The Fortescue River is the remarkably straight, fault controlled feature bordering the Hammersley on the north. Sand dunes are the main surface features in the northeast and southwest. Many dry lakebeds can be seen to the east as light grey colored patches along the watercourses.

  9. A new and frequent human T-cell leukemia virus indeterminate Western blot pattern: epidemiological determinants and PCR results in central African inhabitants.

    PubMed

    Filippone, Claudia; Bassot, Sylviane; Betsem, Edouard; Tortevoye, Patricia; Guillotte, Micheline; Mercereau-Puijalon, Odile; Plancoulaine, Sabine; Calattini, Sara; Gessain, Antoine

    2012-05-01

    Human T-cell leukemia virus (HTLV) indeterminate Western blot (WB) serological patterns are frequently observed in plasma/serum from persons living in intertropical areas. In the framework of ongoing projects on HTLV-1/2 and related viruses in Central Africa, we systematically analyzed plasma from villagers living in South Cameroon by WB. The group included 1,968 individuals (mean age, 44 years; age range, 5 to 90 years; 978 women/990 men), both Bantus (1,165) and Pygmies (803). Plasma samples were tested by WB analysis (MPD HTLV Blot 2.4) and interpreted according to the manufacturer's instructions. Only clear bands were considered in the analysis. Among the 1,968 plasma samples, 38 (1.93%) were HTLV-1, 13 (0.66%) were HTLV-2, and 6 (0.3%) were HTLV WB seropositive. Furthermore, 1,292 (65.65%) samples were WB sero-indeterminate, including 104 (5.28%) with an HTLV-1 Gag-indeterminate pattern (HGIP) and 68 (3.45%) with a peculiar yet unreported pattern exhibiting mostly a strong shifted GD21 and a p28. The other 619 (31.45%) samples were either WB negative or exhibited other patterns, mostly with unique p19 or p24 bands. DNA, extracted from peripheral blood buffy coat, was subjected to PCR using several primer pairs known to detect HTLV-1/2/3/4. Most DNAs from HTLV-1- and HTLV-seropositive individuals were PCR positive. In contrast, all the others, from persons with HTLV-2, HGIP, new WB, and other indeterminate patterns, were PCR negative. Epidemiological determinant analysis of the persons with this new peculiar WB pattern revealed that seroprevalence was independent from age, sex, or ethnicity, thus resembling the indeterminate profile HGIP rather than HTLV-1. Moreover, this new pattern persists over time.

  10. GAPDH and β-actin protein decreases with aging, making Stain-Free technology a superior loading control in Western blotting of human skeletal muscle.

    PubMed

    Vigelsø, Andreas; Dybboe, Rie; Hansen, Christina Neigaard; Dela, Flemming; Helge, Jørn W; Guadalupe Grau, Amelia

    2015-02-01

    Reference proteins (RP) or the total protein (TP) loaded is used to correct for uneven loading and/or transfer in Western blotting. However, the signal sensitivity and the influence of physiological conditions may question the normalization methods. Therefore, three widely used reference proteins [β-actin, glyceraldehyde 3-phosphate dehydrogenase (GAPDH), and α-tubulin], as well as TP loaded measured by Stain-Free technology (SF) as normalization tool were tested. This was done using skeletal muscle samples from men subjected to physiological conditions often investigated in applied physiology where the intervention has been suggested to impede normalization (ageing, muscle atrophy, and different muscle fiber type composition). The linearity of signal and the methodological variation coefficient was obtained. Furthermore, the inter- and intraindividual variation in signals obtained from SF and RP was measured in relation to ageing, muscle atrophy, and different muscle fiber type composition, respectively. A stronger linearity of SF and β-actin compared with GAPDH and α-tubulin was observed. The methodological variation was relatively low in all four methods (4-11%). Protein level of β-actin and GAPDH was lower in older men compared with young men. In conclusion, β-actin, GAPDH, and α-tubulin may not be used for normalization in studies that include subjects with a large age difference. In contrast, the RPs may not be affected in studies that include muscle wasting and differences in muscle fiber type. The novel SF technology adds lower variation to the results compared with the existing methods for correcting for loading inaccuracy in Western blotting of human skeletal muscle in applied physiology.

  11. Development and validation of a novel protein extraction methodology for quantitation of protein expression in formalin-fixed paraffin-embedded tissues using western blotting.

    PubMed

    Nirmalan, Niroshini J; Harnden, Patricia; Selby, Peter J; Banks, Rosamonde E

    2009-03-01

    The development of efficient formaldehyde cross-link reversal strategies will make the vast diagnostic tissue archives of pathology departments amenable to prospective and retrospective translational research, particularly in biomarker-driven proteomic investigations. Heat-induced antigen retrieval strategies (HIARs) have achieved varying degrees of cross-link reversal, potentially enabling archival tissue usage for proteomic applications outside its current remit of immunohistochemistry (IHC). While most successes achieved so far have been based on retrieving tryptic peptide fragments using shot-gun proteomic approaches, attempts at extracting full-length, non-degraded, immunoreactive proteins from archival tissue have proved challenging. We have developed a novel heat-induced antigen retrieval strategy using SDS-containing Laemmli buffer for efficient intact protein recovery from formalin-fixed tissues for subsequent analysis by western blotting. Protocol optimization and comparison of extraction efficacies with frozen tissues and current leader methodology is presented. Quantitative validation of methodology was carried out in a cohort of matched tumour/normal, frozen/FFPE renal tissue samples from 10 patients, probed by western blotting for a selected panel of seven proteins known to be differentially expressed in renal cancer. Our data show that the protocol enables efficient extraction of non-degraded, full-length, immunoreactive protein, with tumour versus normal differential expression profiles for a majority of the panel of proteins tested being comparable to matched frozen tissue controls (rank correlation, r = 0.7292, p < 1.825e-09). However, the variability observed in extraction efficacies for some membrane proteins emphasizes the need for cautious interpretation of quantitative data from this subset of proteins. The method provides a viable, cost-effective quantitative option for the validation of potential biomarker panels through a range of clinical

  12. Localization and Distribution of 'Candidatus Liberibacter asiaticus’ in Citrus and Periwinkle by Direct Tissue Blot Immuno Assay with an Anti-OmpA Polyclonal Antibody

    PubMed Central

    Ding, Fang; Duan, Yongping; Paul, Cristina; Brlansky, Ronald H.; Hartung, John S.

    2015-01-01

    ‘Candidatus Liberibacter asiaticus’ (CaLas), a non-cultured member of the α-proteobacteria, is the causal agent of citrus Huanglongbing (HLB). Due to the difficulties of in vitro culture, antibodies against CaLas have not been widely used in studies of this pathogen. We have used an anti-OmpA polyclonal antibody based direct tissue blot immunoassay to localize CaLas in different citrus tissues and in periwinkle leaves. In citrus petioles, CaLas was unevenly distributed in the phloem sieve tubes, and tended to colonize in phloem sieve tubes on the underside of petioles in preference to the upper side of petioles. Both the leaf abscission zone and the junction of the petiole and leaf midrib had fewer CaLas bacteria compared to the main portions of the petiole and the midribs. Colonies of CaLas in phloem sieve tubes were more frequently found in stems with symptomatic leaves than in stems with asymptomatic leaves with an uneven distribution pattern. In serial sections taken from the receptacle to the peduncle, more CaLas were observed in the peduncle sections adjacent to the stem. In seed, CaLas was located in the seed coat. Many fewer CaLas were found in the roots, as compared to the seeds and petioles when samples were collected from trees with obvious foliar symptoms. The direct tissue blot immuno assay was adapted to whole periwinkle leaves infected by CaLas. The pathogen was distributed throughout the lateral veins and the results were correlated with results of qPCR. Our data provide direct spatial and anatomical information for CaLas in planta. This simple and scalable method may facilitate the future research on the interaction of CaLas and host plant. PMID:25946013

  13. Localization and Distribution of 'Candidatus Liberibacter asiaticus' in Citrus and Periwinkle by Direct Tissue Blot Immuno Assay with an Anti-OmpA Polyclonal Antibody.

    PubMed

    Ding, Fang; Duan, Yongping; Paul, Cristina; Brlansky, Ronald H; Hartung, John S

    2015-01-01

    'Candidatus Liberibacter asiaticus' (CaLas), a non-cultured member of the α-proteobacteria, is the causal agent of citrus Huanglongbing (HLB). Due to the difficulties of in vitro culture, antibodies against CaLas have not been widely used in studies of this pathogen. We have used an anti-OmpA polyclonal antibody based direct tissue blot immunoassay to localize CaLas in different citrus tissues and in periwinkle leaves. In citrus petioles, CaLas was unevenly distributed in the phloem sieve tubes, and tended to colonize in phloem sieve tubes on the underside of petioles in preference to the upper side of petioles. Both the leaf abscission zone and the junction of the petiole and leaf midrib had fewer CaLas bacteria compared to the main portions of the petiole and the midribs. Colonies of CaLas in phloem sieve tubes were more frequently found in stems with symptomatic leaves than in stems with asymptomatic leaves with an uneven distribution pattern. In serial sections taken from the receptacle to the peduncle, more CaLas were observed in the peduncle sections adjacent to the stem. In seed, CaLas was located in the seed coat. Many fewer CaLas were found in the roots, as compared to the seeds and petioles when samples were collected from trees with obvious foliar symptoms. The direct tissue blot immuno assay was adapted to whole periwinkle leaves infected by CaLas. The pathogen was distributed throughout the lateral veins and the results were correlated with results of qPCR. Our data provide direct spatial and anatomical information for CaLas in planta. This simple and scalable method may facilitate the future research on the interaction of CaLas and host plant.

  14. The northern Egyptian continental margin

    NASA Astrophysics Data System (ADS)

    Badawy, Ahmed; Mohamed, Gad; Omar, Khaled; Farid, Walid

    2015-01-01

    Africa displays a variety of continental margin structures, tectonics and sedimentary records. The northern Egyptian continental margin represents the NE portion of the North African passive continental margin. Economically, this region is of great importance as a very rich and productive hydrocarbon zone in Egypt. Moreover, it is characterized by remarkable tectonic setting accompanied by active tectonic processes from the old Tethys to recent Mediterranean. In this article, seismicity of the northern Egyptian continental margin has been re-evaluated for more than 100-years and the source parameters of three recent earthquakes (October 2012, January 2013 and July 2013) have been estimated. Moment tensor inversions of 19th October 2012 and 17th January 2013 earthquakes reveal normal faulting mechanism with strike-slip component having seismic moment of 3.5E16 N m and 4.3E15 N m respectively. The operation of the Egyptian National Seismic Network (ENSN) since the end of 1997 has significantly enhanced the old picture of earthquake activity across northern Egyptian continental margin whereas; the record-ability (annual rate) has changed from 2-events/year to 54-event/year before and after ENSN respectively. The spatial distribution of earthquakes foci indicated that the activity tends to cluster at three zones: Mediterranean Ridge (MR), Nile Cone (NC) and Eratosthenes Seamount (ERS). However, two seismic gaps are reported along Levant Basin (LEV) and Herodotus Basin (HER).

  15. Geologic map of the northern plains of Mars

    USGS Publications Warehouse

    Tanaka, Kenneth L.; Skinner, James A.; Hare, Trent M.

    2005-01-01

    The northern plains of Mars cover nearly a third of the planet and constitute the planet's broadest region of lowlands. Apparently formed early in Mars' history, the northern lowlands served as a repository both for sediments shed from the adjacent ancient highlands and for volcanic flows and deposits from sources within and near the lowlands. Geomorphic evidence for extensive tectonic deformation and reworking of surface materials through release of volatiles occurs throughout the northern plains. In the polar region, Planum Boreum contains evidence for the accumulation of ice and dust, and surrounding dune fields suggest widespread aeolian transport and erosion. The most recent regional- and global-scale maps describing the geology of the northern plains are largely based on Viking Orbiter image data (Dial, 1984; Witbeck and Underwood, 1984; Scott and Tanaka, 1986; Greeley and Guest, 1987; Tanaka and Scott, 1987; Tanaka and others, 1992a; Rotto and Tanaka, 1995; Crumpler and others, 2001; McGill, 2002). These maps reveal highland, plains, volcanic, and polar units based on morphologic character, albedo, and relative ages using local stratigraphic relations and crater counts. This geologic map of the northern plains is the first published map that covers a significant part of Mars using topography and image data from both the Mars Global Surveyor and Mars Odyssey missions. The new data provide a fresh perspective on the geology of the region that reveals many previously unrecognizable units, features, and temporal relations. In addition, we adapted and instituted terrestrial mapping methods and stratigraphic conventions that we think result in a clearer and more objective map. We focus on mapping with the intent of reconstructing the history of geologic activity within the northern plains, including deposition, volcanism, erosion, tectonism, impact cratering, and other processes with the aid of comprehensive crater-density determinations. Mapped areas include all

  16. Beta-thalassaemia mutations in northern India (Delhi).

    PubMed

    Madan, N; Sharma, S; Rusia, U; Sen, S; Sood, S K

    1998-03-01

    The present study was undertaken to define beta-thalassaemia mutations prevalent in northern India (Delhi). Forty six children of beta-thalassaemia major and their families were investigated. DNA was extracted from leucocytes and screened for mutations prevalent in the Indian population. These mutations included 619bp deletion, IVS 1-1 (G-T), IVS 1-5 (G-C), frameshift mutations FS 8/9 (+G), FS 41/42 (-CTTT), Codon 16(-C), Codon 15 (G-A), codon 30 (G-C), IVS 1-110 (G-A) and -88 (C-T). 619 bp deletion mutation was detected directly by amplification of DNA by PCR followed by agarose gel electrophoresis. Other mutations were studied by DNA amplification and dot blot hybridization using synthetic normal and mutant oligonucleotide probes labelled at 5' end with gamma-32 P-ATP. Five mutations accounted for all the chromosomes in 46 patients. 619 bp deletion mutation was found to be the commonest mutation (34.8%) followed by IVS 1-5 (G-C) in 22.8 per cent, IVS 1-1 (G-T) in 19.6 per cent, FS 8/9 (+G) in 13 per cent and FS 41/42 (-CTTT) in 9.8 per cent. Nineteen (41.3%) patients were homozygous and 27 (58.7%) double heterozygous for different beta-thalassaemia mutations. This observation of limited number of mutations is significant and will be useful in planning strategies for prenatal diagnosis of beta-thalassaemia in northern India.

  17. Diversity of avian haemosporidians in arid zones of northern Venezuela.

    PubMed

    Belo, Nayara O; Rodríguez-Ferraro, Adriana; Braga, Erika M; Ricklefs, Robert E

    2012-07-01

    Arid zones of northern Venezuela are represented by isolated areas, important from an ornithological and ecological perspective due to the occurrence of restricted-range species of birds. We analysed the prevalence and molecular diversity of haemosporidian parasites of wild birds in this region by screening 527 individuals (11 families and 20 species) for parasite mitochondrial DNA. The overall prevalence of parasites was 41%, representing 17 mitochondrial lineages: 7 of Plasmodium and 10 of Haemoproteus. Two parasite lineages occurred in both the eastern and western regions infecting a single host species, Mimus gilvus. These lineages are also present throughout northern and central Venezuela in a variety of arid and mesic habitats. Some lineages found in this study in northern Venezuela have also been observed in different localities in the Americas, including the West Indies. In spite of the widespread distributions of some of the parasite lineages found in northern Venezuela, several, including some that are relatively common (e.g. Ven05 and Ven06), have not been reported from elsewhere. Additional studies are needed to characterize the host and geographical distribution of avian malaria parasite lineages, which will provide a better understanding of the influence of landscape, vector abundance and diversity, and host identity on haemosporidian parasite diversity and prevalence.

  18. Geothermal systems of northern Nevada

    USGS Publications Warehouse

    Hose, Richard Kenneth; Taylor, Bruce Edward

    1974-01-01

    Hot springs are numerous and nearly uniformly distributed in northern Nevada. Most occur on the flanks of basins, along Basin and Range (late Miocene to Holocene) faults, while some occur in the inner parts of the basins. Surface temperatures of the springs range from slightly above ambient to, boiling; some springs are superheated. Maximum subsurface water temperatures calculated on the basis of quartz solubility range as high as 252?C, although most are below 190?C. Flows range from a trickle to several hundred liters per minute. The Nevada geothermal systems differ markedly from the power-producing system at The Geysers, Calif., and from those areas with a high potential, for power production (e.g., Yellowstone Park, Wyo.; Jemez Mountains, N. Mex.). These other systems are associated with Quaternary felsic volcanic rocks and probably derive their heat from cooling magma rather high in the crust. In northern Nevada, however, felsic volcanic rocks are virtually all older than 10 million years, and. analogous magmatic heat sources are, therefore, probably lacking. Nevada is part of an area of much higher average heat flow than the rest of the United States. In north-central Nevada, geothermal gradients are as great as 64?C per kilometer in bedrock and even higher in basin fill. The high gradients probably result from a combination of thin crust and high temperature upper mantle. We suggest that the geothermal systems of northern Nevada result from circulation of meteoric waters along Basin and Range faults and that their temperature chiefly depends upon (1) depth of circulation and (2) the geothermal gradient near the faults.

  19. Geothermal development plan: northern Arizona

    SciTech Connect

    White, D.H.; Goldstone, L.A.

    1981-01-01

    Much of the northern counties (Apache, Coconino, Gila, Mohave, Navajo and Yavapai) is located in the Colorado Plateau province, a region of low geothermal potential. Two areas that do show some potential are the Flagstaff - San Francisco Peaks area and the Springerville area. Flagstaff is rapidly becoming the manufacturing center of Arizona and will have many opportunities to use geothermal energy to satisfy part of its increasing need for energy. Using a computer simulation model, projections of geothermal energy on line as a function of time are made for both private and city-owned utility development of a resource.

  20. Comparative ability of northern pintails, gadwalls, and northern shovelers to metabolize foods

    USGS Publications Warehouse

    Miller, M.R.

    1984-01-01

    Feeding trials were used to compare the ability of northern pintails (Anas acuta), gadwalls (A. strepera), and northern shovelers (A. clypeata) to metabolize energy from a turkey starter ration, alfalfa pellets, and common barnyardgrass (Echinochloa crusgalli) seeds. No differences (P > 0.05) were detected among the three species for any of the three foods (kg body weight/day basis), for dry matter intake (DMI), body weight gain (BWG), apparent metabolizable energy (AME), or metabolized energy (MEE) on any given diet consumed in quantities large enough to promote body weight gain. The AME content of alfalfa was 57% less than the value for turkey starter and 50% less than for barnyardgrass seeds. All three species metabolized more energy and gained weight faster when fed turkey starter. Energy modeling may be facilitated if additional research verifies that all species of dabbling ducks have equal ability to obtain energy from foods available to them in the wild. Behavioral and morphological factors may be more important in defining feeding niches than digestive physiology, at least for the three duck species tested, at the time of year of the experiments, and within the limits of the quality of foods used.

  1. Western blot seroindeterminate individuals for human T-lymphotropic virus I/II (HTLV-I/II) in Fortaleza (Brazil): a serological and molecular diagnostic and epidemiological approach.

    PubMed

    Santos, Terezinha de Jesus Teixeira; Costa, Carlos Maurício de Castro; Goubau, Patrick; Vandamme, Anne-Mieke; Desmyter, Jan; Van Doren, Sonia; Mota, Rosa M S; de Castro Costa, Francine Bovy; Oliveira, Ana C S; Barreto, Vania; Gomes, A F; Carneiro-Proietti, Anna B; de Bruin, Veralice Meireles Sales; de Sousa, Francisca C F; Oriá, Reinaldo Barreto

    2003-06-01

    How to handle Western blot (WB) seroindeterminate individuals for Human T-lymphotropic Virus 1/2 (HTLV-1/2) constitutes a challenge for blood banks and families. We made a cross-sectional study of 191 enzyme linked immunoassay (EIA) reactive individuals from the hematological center (HEMOCE) of Fortaleza (Brazil), examining their serological (WB) and molecular (PCR) diagnosis, and demographic profiles, as well as a possible association of their condition with other infectious pathologies and risk factors. Ethical institutional approval and personal consent were obtained. Out of 191 EIA reactive individuals, 118 were WB seroindeterminate and 73 were seropositive for HTLV-1/2. In the PCR analysis of 41 WB seroindeterminate individuals, 9 (22%) were positive and 32 (78%) were negative for HTLV-1/2. The demographic analysis indicated a trend towards a predominance of males among the seroindeterminate individuals and females in the seropositive ones. The seroindeterminate individuals were younger than the seropositive ones. We did not find any association of these conditions with syphilis, Chagas disease or HIV or hepatitis, and with risk factors such as breast-feeding, blood transfusion, STD (syphilis) and IDU.

  2. Molecular characterization of MT3 antigens by two-dimensional gel electrophoresis, NH2-terminal amino acid sequence analysis, and southern blot analysis.

    PubMed Central

    Sorrentino, R; Lillie, J; Strominger, J L

    1985-01-01

    The monoclonal antibody 109d6, which recognizes major histocompatibility antigen MT3-like serologic determinants, has been used to characterize the molecules bearing this determinant in HLA-DR4 and -DR7 homozygous cell lines by two-dimensional gel and sequencing analyses. By these two criteria, these molecules are identical to each other. Southern blot analysis of genomic DNA from HLA-DR1 through -DR7 homozygous cell lines with DR beta-chain gene probes reveals a striking similarity in the pattern of hybridizing fragments between DR4 and DR7 haplotypes and among DR3, DR5, and DRw6 haplotypes reminiscent of the MT3/MT2 allodeterminant distribution. The sharing of the MT2 determinant between DR3, DR5, and DRw6 haplotypes and of the MT3 determinant between DR4 and DR7 haplotypes is part of a broader "homology," which may be a consequence of more recent separation of the haplotypes sharing the MT2 determinant on the one hand and the haplotypes sharing the MT3 determinant on the other hand. Images PMID:2582424

  3. Identification of liver protein targets modified by tienilic acid metabolites using a two-dimensional Western blot-mass spectrometry approach

    NASA Astrophysics Data System (ADS)

    Methogo, Ruth Menque; Dansette, Patrick M.; Klarskov, Klaus

    2007-12-01

    A combined approach based on two-dimensional electrophoresis-immuno-blotting and nanoliquid chromatography coupled on-line with electrospray ionization mass spectrometry (nLC-MS/MS) was used to identify proteins modified by a reactive intermediate of tienilic acid (TA). Liver homogenates from rats exposed to TA were fractionated using ultra centrifugation; four fractions were obtained and subjected to 2D electrophoresis. Following transfer to PVDF membranes, modified proteins were visualized after India ink staining, using an anti-serum raised against TA and ECL detection. Immuno-reactive spots were localized on the PVDF membrane by superposition of the ECL image, protein spots of interest were excised, digested on the membrane with trypsin followed by nLC-MS/MS analysis and protein identification. A total of 15 proteins were identified as likely targets modified by a TA reactive metabolite. These include selenium binding protein 2, senescence marker protein SMP-30, adenosine kinase, Acy1 protein, adenosylhomocysteinase, capping protein (actin filament), protein disulfide isomerase, fumarylacetoacetase, arginase chain A, ketohexokinase, proteasome endopeptidase complex, triosephosphate isomerase, superoxide dismutase, dna-type molecular chaperone hsc73 and malate dehydrogenase.

  4. Utility of Schistosoma bovis Adult Worm Antigens for Diagnosis of Human Schistosomiasis by Enzyme-Linked Immunosorbent Assay and Electroimmunotransfer Blot Techniques

    PubMed Central

    Pardo, J.; Carranza, C.; Turrientes, M. C.; Arellano, J. L. Pérez; Vélez, R. López; Ramajo, V.; Muro, A.

    2004-01-01

    Immunodiagnostic methods based on the detection of antibodies continue to be the most effective and practical methods for the diagnosis of imported schistosomiasis. Schistosoma bovis is a species whose final natural hosts are bovines, ovines, caprines, and small wild ruminants. Different studies have demonstrated the analogies existing between S. bovis and other Schistosoma species which affect humans. The objective of this work was to evaluate the utility of S. bovis adult worm antigens (AWA) for the diagnosis of imported human schistosomiasis by enzyme-linked immunosorbent assay (ELISA) and electroimmunotransfer blotting (EITB) techniques. By detecting eggs, the ELISA for S. bovis AWA was able to definitively detect imported cases with a sensitivity of 94%. The specificity of the ELISA for S. bovis AWA was 97%. There were no differences between the results of the S. bovis AWA ELISA for patients infected with Schistosoma mansoni and those infected with Schistosoma haematobium. The EITB technique showed bands of 85, 37, and 20 kDa, which are characteristic of infections with Schistosoma spp. Specific bands to indicate infection by different species of Schistosoma have not been detected. The combined use of the ELISA for S. bovis AWA and EITB increased the global sensitivity of the study to 97%. Our findings suggest that the ELISA for S. bovis AWA is a useful test for the immunodiagnosis of imported schistosomiasis and that EITB for detecting S. bovis AWA permits the confirmation of diagnosis when the ELISA for S. bovis AWA is positive. PMID:15539523

  5. Differential staining of Western blots of human secreted glycoproteins from serum, milk, saliva, and seminal fluid using lectins displaying diverse sugar specificities.

    PubMed

    Gilboa-Garber, Nechama; Lerrer, Batya; Lesman-Movshovich, Efrat; Dgani, Orly

    2005-12-01

    Human milk, serum, saliva, and seminal fluid glycoproteins (gps) nourish and protect newborn and adult tissues. Their saccharides, which resemble cell membrane components, may block pathogen adhesion and infection. In the present study, they were examined by a battery of lectins from plants, animals, and bacteria, using hemagglutination inhibition and Western blot analyses. The lectins included galactophilic ones from Aplysia gonad, Erythrina corallodendron, Maclura pomifera (MPL), peanut, and Pseudomonas aeruginosa (PA-IL); fucose-binding lectins from Pseudomonas aeruginosa (PA-IIL), Ralstonia solanacearum (RSL), and Ulex europaeus (UEA-I), and mannose/glucose-binding Con A. The results demonstrated the chosen lectin efficiency for differential analysis of human secreted gps as compared to CBB staining. They unveiled the diversity of these body fluid gp glycans (those of the milk and seminal fluid being highest): the milk gps interacted most strongly with PA-IIL, followed by RSL; the saliva gps with RSL, followed by PA-IIL and MPL; the serum gps with Con A and MPL, followed by PA-IIL and RSL, and the seminal plasma gps with RSL and MPL, followed by UEA-I and PA-IIL. The potential usage of these lectins as probes for scientific, industrial, and medical purposes, and for quality control of the desired gps is clearly indicated.

  6. Utility of Schistosoma bovis adult worm antigens for diagnosis of human schistosomiasis by enzyme-linked immunosorbent assay and electroimmunotransfer blot techniques.

    PubMed

    Pardo, J; Carranza, C; Turrientes, M C; Pérez Arellano, J L; López Vélez, R; Ramajo, V; Muro, A

    2004-11-01

    Immunodiagnostic methods based on the detection of antibodies continue to be the most effective and practical methods for the diagnosis of imported schistosomiasis. Schistosoma bovis is a species whose final natural hosts are bovines, ovines, caprines, and small wild ruminants. Different studies have demonstrated the analogies existing between S. bovis and other Schistosoma species which affect humans. The objective of this work was to evaluate the utility of S. bovis adult worm antigens (AWA) for the diagnosis of imported human schistosomiasis by enzyme-linked immunosorbent assay (ELISA) and electroimmunotransfer blotting (EITB) techniques. By detecting eggs, the ELISA for S. bovis AWA was able to definitively detect imported cases with a sensitivity of 94%. The specificity of the ELISA for S. bovis AWA was 97%. There were no differences between the results of the S. bovis AWA ELISA for patients infected with Schistosoma mansoni and those infected with Schistosoma haematobium. The EITB technique showed bands of 85, 37, and 20 kDa, which are characteristic of infections with Schistosoma spp. Specific bands to indicate infection by different species of Schistosoma have not been detected. The combined use of the ELISA for S. bovis AWA and EITB increased the global sensitivity of the study to 97%. Our findings suggest that the ELISA for S. bovis AWA is a useful test for the immunodiagnosis of imported schistosomiasis and that EITB for detecting S. bovis AWA permits the confirmation of diagnosis when the ELISA for S. bovis AWA is positive.

  7. Detection of genetically modified crops using multiplex asymmetric polymerase chain reaction and asymmetric hyperbranched rolling circle amplification coupled with reverse dot blot.

    PubMed

    Wang, Xiumin; Teng, Da; Guan, Qingfeng; Tian, Fang; Wang, Jianhua

    2015-04-15

    To meet the ever-increasing demand for detection of genetically modified crops (GMCs), low-cost, high-throughput and high-accuracy detection assays are needed. The new multiplex asymmetric polymerase chain reaction and asymmetric hyper-branched rolling circle amplification coupled with reverse dot blot (RDB) systems were developed to detect GMCs. Thirteen oligonucleotide probes were designed to identify endogenous targets (Lec1, Hmg and Sad1), event-specific targets (RRS-5C, RRS-3C, Bt176-3C and MON810-3C), screening targets (35S promoter and NOS terminator), and control targets (18S and PLX). Optimised conditions were as follows: tailed hybridization probes (1-2 pmol/l) were immobilized on a membrane by baking for 2h, and a 10:1 ratio of forward to reverse primers was used. The detection limits were 0.1 μg/l of 2% RRS and 0.5 ng/l of DNA from genetically modified (GM) soybean. These results indicate that the RDB assay could be used to detect multiplex target genes of GMCs rapidly and inexpensively.

  8. A comparison of DNA extraction protocols from blood spotted on FTA cards for the detection of tick-borne pathogens by Reverse Line Blot hybridization.

    PubMed

    Hailemariam, Zerihun; Ahmed, Jabbar Sabir; Clausen, Peter-Henning; Nijhof, Ard Menzo

    2017-01-01

    An essential step in the molecular detection of tick-borne pathogens (TBPs) in blood is the extraction of DNA. When cooled storage of blood under field conditions prior to DNA extraction in a dedicated laboratory is not possible, the storage of blood on filter paper forms a promising alternative. We evaluated six DNA extraction methods from blood spotted on FTA Classic(®) cards (FTA cards), to determine the optimal protocol for the subsequent molecular detection of TBPs by PCR and the Reverse Line Blot hybridization assay (RLB). Ten-fold serial dilutions of bovine blood infected with Babesia bovis, Theileria mutans, Anaplasma marginale or Ehrlichia ruminantium were made by dilution with uninfected blood and spotted on FTA cards. Subsequently, DNA was extracted from FTA cards using six different DNA extraction protocols. DNA was also isolated from whole blood dilutions using a commercial kit. PCR/RLB results showed that washing of 3mm discs punched from FTA cards with FTA purification reagent followed by DNA extraction using Chelex(®) resin was the most sensitive procedure. The detection limit could be improved when more discs were used as starting material for the DNA extraction, whereby the use of sixteen 3mm discs proved to be most practical. The presented best practice method for the extraction of DNA from blood spotted on FTA cards will facilitate epidemiological studies on TBPs. It may be particularly useful for field studies where a cold chain is absent.

  9. Effects of histoplasmin M antigen chemical and enzymatic deglycosylation on cross-reactivity in the enzyme-linked immunoelectrotransfer blot method.

    PubMed Central

    Zancopé-Oliveira, R M; Bragg, S L; Reiss, E; Wanke, B; Peralta, J M

    1994-01-01

    The enzyme-linked immunoelectrotransfer blot (EITB) method was evaluated as a suitable method for detecting antibodies against M antigen of Histoplasma capsulatum by use of both glycosylated and deglycosylated M protein of histoplasmin (HMIN). Sera from patients with histoplasmosis, paracoccidioidomycosis, blastomycosis, coccidioidomycosis, and aspergillosis were tested by the EITB with glycosylated M protein of HMIN. This assay demonstrated 100% sensitivity with histoplasmosis serum samples, all of which reacted with the 94-kDa glycoprotein (M antigen). Although the EITB is highly sensitive, it is not specific for histoplasmosis when glycosylated M protein is used as an antigen. A total of 81% of paracoccidioidomycosis, 25% of blastomycosis, 33% of coccidioidomycosis, 73% of aspergillosis, and 16% of tuberculosis serum samples cross-reacted with M protein of HMIN and yielded patterns indistinguishable from those obtained with histoplasmosis serum samples. The EITB reactions with both untreated M antigen and M antigen altered by periodate oxidation or by deglycosylation with endoglycosidases were compared. Cross-reactions with heterologous sera in the EITB could be attributed to periodate-sensitive carbohydrate epitopes, as reflected by the increase in the test specificity from 46.1 to 91.2% after periodate treatment of M protein. The EITB for the detection of antibodies to M antigen is a potential diagnostic test for histoplasmosis, provided that periodate-treated M protein is used as an antigen. Images PMID:8556474

  10. Estimation of hepatitis E virus (HEV) pig seroprevalence using ELISA and Western blot and comparison between human and pig HEV sequences in Belgium.

    PubMed

    Thiry, Damien; Mauroy, Axel; Saegerman, Claude; Thomas, Isabelle; Wautier, Magali; Miry, Cora; Czaplicki, Guy; Berkvens, Dirk; Praet, Nicolas; van der Poel, Wim; Cariolet, Roland; Brochier, Bernard; Thiry, Etienne

    2014-08-27

    Zoonotic transmission of hepatitis E virus (HEV) is of special concern, particularly in high income countries were waterborne infections are less frequent than in developing countries. High HEV seroprevalences can be found in European pig populations. The aims of this study were to obtain prevalence data on HEV infection in swine in Belgium and to phylogenetically compare Belgian human HEV sequences with those obtained from swine. An ELISA screening prevalence of 73% (95% CI 68.8-77.5) was determined in Belgian pigs and a part of the results were re-evaluated by Western blot (WB). A receiver operating characteristic curve analysis was performed and scenarios varying the ELISA specificity relative to WB were analysed. The seroprevalences estimated by the different scenarios ranged between 69 and 81% and are in agreement with the high exposure of the European pig population to HEV. Pig HEV sequences were genetically compared to those detected in humans in Belgium and a predominance of genotype 3 subtype f was shown in both swine and humans. The high HEV seroprevalence in swine and the close phylogenetic relationships between pig and human HEV sequences further support the risk for zoonotic transmission of HEV between humans and pigs.

  11. Inverse PCR and Quantitative PCR as Alternative Methods to Southern Blotting Analysis to Assess Transgene Copy Number and Characterize the Integration Site in Transgenic Woody Plants.

    PubMed

    Stefano, Biricolti; Patrizia, Bogani; Matteo, Cerboneschi; Massimo, Gori

    2016-06-01

    One of the major unanswered questions with respect to the commercial use of genetic transformation in woody plants is the stability of the transgene expression over several decades within the same individual. Gene expression is strongly affected by the copy number which has been integrated into the plant genome and by the local DNA features close to the integration sites. Because woody plants cannot be subjected to selfing or backcrossing to modify the transgenic allelic structure without affecting the valuable traits of the cultivar, molecular characterization of the transformation event is therefore crucial. After assessing the transgene copy number of a set of apple transgenic clones with Southern blotting, we describe two alternative methods: the first is based on inverse PCR (i-PCR) and the second on the quantitative PCR (q-PCR). The methods produced comparable results with the exception of the data regarding a high copy number clone, but while the q-PCR-based system is rapid and easily adaptable to high throughput systems, the i-PCR-based method can provide information regarding the transformation event and the characteristics of the sequences flanking the transgenic construct.

  12. Rapid detection of six common Mediterranean and three non-Mediterranean alpha-thalassemia point mutations by reverse dot blot analysis.

    PubMed

    Foglietta, Enrica; Bianco, Ida; Maggio, Aurelio; Giambona, Antonino

    2003-11-01

    We describe the implementation of reverse dot blot (RDB) hybridization as a rapid nonradioactive method for the identification of six frequent globin gene point mutations in the Mediterranean population: alpha(Hph)alpha: alpha2 IVS I donor site GGTGAGG --> GG-----; alpha(NcoI)alpha: alpha2 initiation codon ATG --> ACG; alpha(TSaudi)alpha: alpha2Poly A signal AATAA --> AATAAG; alpha(Icaria)alpha: alpha2 termination codon TAA --> AAA (Ter --> LYS); alpha(CS)alpha: alpha2 termination codon TAA --> CAA (Ter --> gly); alphaalpha(NcoI): alpha1 initiation codon ATG --> GTG; and three alpha2 globin gene point mutations found in immigrants in Italy: alpha(T-Quongsze)alpha: alpha2 codon 12 CTG --> CCG (Leu --> Pro); alpha(Seal Rock)alpha: alpha2 termination codon TAA --> GAA (TER --> GLU); and alpha(Koyadora)alpha: alpha2 termination codon TAA --> TCA (TER --> SER). The method uses the principle of allele-specific oligonucleotide (ASO) hybridization, but it is a nonradioactive method and permits rapid and simultaneous typing of point mutations and small deletions.

  13. An optimized xylene-free protein extraction method adapted to formalin-fixed paraffin embedded tissue sections for western blot analysis.

    PubMed

    Mansour, Anthony G; Khalil, Pamela Abou; Bejjani, Noha; Chatila, Rajaa; Dagher-Hamalian, Carole; Faour, Wissam H

    2017-03-01

    Deparaffinization of formalin-fixed paraffin embedded (FFPE) tissues with xylene currently remains a major challenge to the biomedical community. We developed an efficient xylene-free protocol to isolate proteins from archived FFPE human tissue sections. A total of 79 different types of FFPE tissue sections of 8 µm thickness were obtained from various archived FFPE specimens. Deparaffinization was conducted by gently washing each section with around 1 ml of hot distilled water (≈80°C). The deparaffinized tissues were homogenized in lysis buffer, and the isolated proteins were quantified and efficiently resolved using western blot analysis for the presence of Protein kinase B (PKB/AKT) and β-actin. Moreover, a significant amount of proteins was successfully isolated with an average of 2.31 µg/µl. The migration pattern of AKT and β-actin obtained from the specimens was similar to the positive control obtained from protein lysates prepared from in vitro cultured MDA231 cancer cell lines. AKT was successfully identified in all specimens, and β-actin protein was resolved with an efficiency higher than 80%. The entire extraction procedure requires only 20 minutes. This newly developed technique is an efficient, safe, cost-effective, and rapid method to isolate proteins from FFPE tissue sections adequate for molecular analysis.

  14. A Western blot-based investigation of the yeast secretory pathway designed for an intermediate-level undergraduate cell biology laboratory.

    PubMed

    Hood-Degrenier, Jennifer K

    2008-01-01

    The movement of newly synthesized proteins through the endomembrane system of eukaryotic cells, often referred to generally as the secretory pathway, is a topic covered in most intermediate-level undergraduate cell biology courses. An article previously published in this journal described a laboratory exercise in which yeast mutants defective in two distinct steps of protein secretion were differentiated using a genetic reporter designed specifically to identify defects in the first step of the pathway, the insertion of proteins into the endoplasmic reticulum (Vallen, 2002). We have developed two versions of a Western blotting assay that serves as a second way of distinguishing the two secretory mutants, which we pair with the genetic assay in a 3-wk laboratory module. A quiz administered before and after students participated in the lab activities revealed significant postlab gains in their understanding of the secretory pathway and experimental techniques used to study it. A second survey administered at the end of the lab module assessed student perceptions of the efficacy of the lab activities; the results of this survey indicated that the experiments were successful in meeting a set of educational goals defined by the instructor.

  15. Immunohistochemical and Western blot analysis of two protein tyrosine phosphatase receptors, R and Z1, in colorectal carcinoma, colon adenoma and normal colon tissues.

    PubMed

    Woźniak, Marta; Gamian, Elżbieta; Łaczmańska, Izabela; Sąsiadek, Maria M; Duś-Szachniewicz, Kamila; Ziółkowski, Piotr

    2014-05-01

    Two classes of proteins, namely tyrosine kinases (PTK) and phosphatases (PTP), play an important role in cell proliferation and differentiation, thus leading to an acceleration or inhibition of tumour growth. The role of the above proteins in colorectal carcinoma (CRC) growth is a well-known event. In this study we carried out immunohistochemical and Western blot analysis of colorectal carcinoma, adenoma and normal colon tissue in relation to two protein tyrosine phosphatase receptors, R and Z1. Twenty-five cases of CRC were analyzed and the results were compared with similar data obtained in non-malignant tissues. High expression of both PTP receptors was observed in all examined cases of CRC, adenoma and normal colon tissue in this study. These results are not in line with recently published data, showing that genetic coding for PTPRR and PTPRZ1 were hypermethylated in CRC's. We presume that the protein tyrosine phosphatase overexpression in colorectal carcinoma is not enough to protect from the progression of disease.

  16. Flood of April and May 2008 in Northern Maine

    USGS Publications Warehouse

    Lombard, Pamela J.

    2010-01-01

    Severe flooding occurred in Aroostook and Penobscot Counties in northern Maine between April 28 and May 1, 2008, and was most extreme in the town of Fort Kent. Peak streamflows in northern Aroostook County were the result of a persistent heavy snowpack that caused high streamflows when it quickly melted during the third week of April 2008. Snowmelt was followed by from two to four inches of rainfall over a 2-day period in northern Maine. Peak water-surface elevations resulting from the flood were obtained from 13 continuous-record streamgages and 63 surveyed high-water marks in Aroostook and Penobscot Counties. Peak streamflows were obtained from 20 sites on 15 streams through stage/discharge rating curves or hydraulic flow models. Peak water-surface elevations and streamflows were the highest ever recorded at seven continuous-record streamgages, which had between 25 and 84 years of record in northern Aroostook County. The annual exceedance probability (the percent chance of exceeding the streamflow recorded during the April/May 2008 flood during any given year) at six streamgages in northern Maine was equal to or less than 1 percent. Data from flood-insurance studies published by the Federal Emergency Management Agency were available for five of the locations analyzed for the April/May 2008 flood and were compared to streamflows and observed peak water-surface elevations from the 2008 flood. Water-surface elevations that would be expected given the observed flow as applied to the effective flood insurance studies ranged from between 1 and 4 feet from the water-surface elevations observed during the 2008 flood. Differences were likely the result of up to 30 years of additional data for the calculation of recurrence intervals and the fact that hydraulic models used for the models had not previously been calibrated to a flood of this magnitude.

  17. Water Ice Albedo Variations on the Martian Northern Polar Cap

    NASA Technical Reports Server (NTRS)

    Hale, A. S.; Bass, D. S.; Tamppari, L. K.

    2003-01-01

    The Viking Orbiters determined that the surface of Mars northern residual cap is water ice. Many researchers have related observed atmospheric water vapor abundances to seasonal exchange between reservoirs such as the polar caps, but the extent to which the exchange between the surface and the atmosphere remains uncertain. Early studies of the ice coverage and albedo of the northern residual Martian polar cap using Mariner 9 and Viking images reported that there were substantial internannual differences in ice deposition on the polar cap, a result which suggested a highly variable Martian climate. However, some of the data used in these studies were obtained at differing values of heliocentric solar longitude (L(sub s)). Reevaluation of this dataset indicated that the residual cap undergoes seasonal brightening throughout the summer, and indicated that this process repeats from year to year. In this study we continue to compare Mariner 9 and Viking Orbiter imaging observations and thermal data of the north residual polar cap to data acquired with Mars Global Surveyor s Mars Orbiter Camera (MOC) instrument. In the current study, our goal is to examine all released data from MGS MOC in the northern summer season, along with applicable TES data in order to better understand the albedo variations in the northern summer and their implications on water transport. To date, work has focused primarily on the MOC dataset. In 1999, data acquisition of the northern polar regions began at L(sub s) = 107, although there was little north polar data acquired from L(sub s)= 107 to L(sub s) = 109. We examined a total of 409 images from L(sub s) = 107 to L(sub s)=148. We have also examined data from 2000 from L(sub s)= 93 to L(sub s)= 110; additional progress is ongoing. Here we present a progress report of our observations, and continue to determine their implications for the Martian water cycle.

  18. Groundwater management in northern Iraq

    NASA Astrophysics Data System (ADS)

    Stevanovic, Zoran; Iurkiewicz, Adrian

    2009-03-01

    Groundwater is vital and the sole resource in most of the studied region of northern Iraq. It has a significant role in agriculture, water supply and health, and the elimination of poverty in rural areas. Although Iraq is currently dramatically disturbed by complex political and socio-economic problems, in its northern part, i.e. the Kurdish-inhabited region, fast urbanization and economic expansion are visible everywhere. Monitoring and water management schemes are necessary to prevent aquifer over-exploitation in the region. Artificial recharge with temporary runoff water, construction of subsurface dams and several other aquifer management and regulation measures have been designed, and some implemented, in order to improve the water situation. Recommendations, presented to the local professionals and decision-makers in water management, include creation of Water Master Plans and Water User Associations, synchronization of drilling programmes, rehabilitation of the existing well fields, opening of new well fields, and the incorporation of new spring intakes in some areas with large groundwater reserves, as well as construction of numerous small-scale schemes for initial in situ water treatment where saline groundwater is present.

  19. Structural features offshore northern Taiwan

    NASA Astrophysics Data System (ADS)

    Yicheng Yang, Eason; Liu, Char-Shine; Chang, Jih-Hsin; Chiu, Chien-Hsuan

    2016-04-01

    The area offshore northern Taiwan is the place where East China Sea Shelf extends into the Southern Okinawa Trough, and where pre-Pleistocene arc-continental collision had occurred. Comparison between fault distribution in the area with previously published results suggests that the fault distribution and regional structural framework are still controversial. Using marine multichannel seismic reflection data collected in 3 marine geophysical survey cruises, we remapped the fault distribution in the northern offshore area of Taiwan. By analyzing all the seismic profiles using the KINGDOM suite (a seismic interpretation software), a new fault distribution map is presented, and a subsurface unconformity PRSB (Pliocene reflection sequence boundary) is identified. Six major NE-SW trending high-angle normal faults cut the PRSB can be traced to the fault systems on land northernmost Taiwan. These normal faults are located between the Southern Okinawa Trough and the East China Sea continental shelf basin, and have been suggested to be reactivated from pre-existing reverse faults. The offsets of fault ramps in PRSB increase toward southeast. The isopach map of the study area compiled shows that sediment strata overlying PRSB thin toward northwest.

  20. Illuminating Northern California's Active Faults

    NASA Astrophysics Data System (ADS)

    Prentice, Carol S.; Crosby, Christopher J.; Whitehill, Caroline S.; Arrowsmith, J. Ramón; Furlong, Kevin P.; Phillips, David A.

    2009-02-01

    Newly acquired light detection and ranging (lidar) topographic data provide a powerful community resource for the study of landforms associated with the plate boundary faults of northern California (Figure 1). In the spring of 2007, GeoEarthScope, a component of the EarthScope Facility construction project funded by the U.S. National Science Foundation, acquired approximately 2000 square kilometers of airborne lidar topographic data along major active fault zones of northern California. These data are now freely available in point cloud (x, y, z coordinate data for every laser return), digital elevation model (DEM), and KMZ (zipped Keyhole Markup Language, for use in Google Earth™ and other similar software) formats through the GEON OpenTopography Portal (http://www.OpenTopography.org/data). Importantly, vegetation can be digitally removed from lidar data, producing high-resolution images (0.5- or 1.0-meter DEMs) of the ground surface beneath forested regions that reveal landforms typically obscured by vegetation canopy (Figure 2).

  1. The 3' addition of CCA to mitochondrial tRNASer(AGY) is specifically impaired in patients with mutations in the tRNA nucleotidyl transferase TRNT1.

    PubMed

    Sasarman, Florin; Thiffault, Isabelle; Weraarpachai, Woranontee; Salomon, Steven; Maftei, Catalina; Gauthier, Julie; Ellazam, Benjamin; Webb, Neil; Antonicka, Hana; Janer, Alexandre; Brunel-Guitton, Catherine; Elpeleg, Orly; Mitchell, Grant; Shoubridge, Eric A

    2015-05-15

    Addition of the trinucleotide cytosine/cytosine/adenine (CCA) to the 3' end of transfer RNAs (tRNAs) is essential for translation and is catalyzed by the enzyme TRNT1 (tRNA nucleotidyl transferase), which functions in both the cytoplasm and mitochondria. Exome sequencing revealed TRNT1 mutations in two unrelated subjects with different clinical features. The first presented with acute lactic acidosis at 3 weeks of age and developed severe developmental delay, hypotonia, microcephaly, seizures, progressive cortical atrophy, neurosensorial deafness, sideroblastic anemia and renal Fanconi syndrome, dying at 21 months. The second presented at 3.5 years with gait ataxia, dysarthria, gross motor regression, hypotonia, ptosis and ophthalmoplegia and had abnormal signals in brainstem and dentate nucleus. In subject 1, muscle biopsy showed combined oxidative phosphorylation (OXPHOS) defects, but there was no OXPHOS deficiency in fibroblasts from either subject, despite a 10-fold-reduction in TRNT1 protein levels in fibroblasts of the first subject. Furthermore, in normal controls, TRNT1 protein levels are 10-fold lower in muscle than in fibroblasts. High resolution northern blots of subject fibroblast RNA suggested incomplete CCA addition to the non-canonical mitochondrial tRNA(Ser(AGY)), but no obvious qualitative differences in other mitochondrial or cytoplasmic tRNAs. Complete knockdown of TRNT1 in patient fibroblasts rendered mitochondrial tRNA(Ser(AGY)) undetectable, and markedly reduced mitochondrial translation, except polypeptides lacking Ser(AGY) codons. These data suggest that the clinical phenotypes associated with TRNT1 mutations are largely due to impaired mitochondrial translation, resulting from defective CCA addition to mitochondrial tRNA(Ser(AGY)), and that the severity of this biochemical phenotype determines the severity and tissue distribution of clinical features.

  2. Importance of coastal primary production in the northern Baltic Sea.

    PubMed

    Ask, Jenny; Rowe, Owen; Brugel, Sonia; Strömgren, Mårten; Byström, Pär; Andersson, Agneta

    2016-10-01

    In this study, we measured depth-dependent benthic microalgal primary production in a Bothnian Bay estuary to estimate the benthic contribution to total primary production. In addition, we compiled data on benthic microalgal primary production in the entire Baltic Sea. In the estuary, the benthic habitat contributed 17 % to the total annual primary production, and when upscaling our data to the entire Bothnian Bay, the corresponding value was 31 %. This estimated benthic share (31 %) is three times higher compared to past estimates of 10 %. The main reason for this discrepancy is the lack of data regarding benthic primary production in the northern Baltic Sea, but also that past studies overestimated the importance of pelagic primary production by not correcting for system-specific bathymetric variation. Our study thus highlights the importance of benthic communities for the northern Baltic Sea ecosystem in general and for future management strategies and ecosystem studies in particular.

  3. A Right to Be Heard: Learning from Learners with Additional Needs in Literacy

    ERIC Educational Resources Information Center

    McPhillips, Therese; Shevlin, Michael; Long, Louise

    2012-01-01

    This study aimed to explore the consultation experiences of pupils who have additional needs in literacy. An opportunistic sample of eight schools--four in Northern Ireland and four in the Republic of Ireland--were chosen by the researchers; selected pupils were receiving additional literacy support. Focus group discussions and arts-based creative…

  4. Nitrogen Inputs via Nitrogen Fixation in Northern Plants and Soils

    NASA Astrophysics Data System (ADS)

    Thorp, N. R.; Wieder, R. K.; Vile, M. A.

    2015-12-01

    Dominated by cold and often acidic water logged environments, mineralization of organic matter is slow in the majority of northern ecosystems. Measures of extractable ammonium and nitrate are generally low and can be undetectable in peat pore waters. Despite this apparent nitrogen limitation, many of these environments produce deep deposits of soil organic matter. Biological nitrogen fixation carried out by autotrophic and heterotrophic diazotrophs associated with cryptograms provides the majority of known nitrogen inputs in these northern ecosystems. Nitrogen fixation was assessed in a variety of northern soils within rhizospheres of dominant plant communities. We investigated the availability of this newly fixed nitrogen to the vascular plant community in nitrogen limited northern plant communities. We tracked nitrogen flow from 15N2 gas fixed in Sphagnum mosses into tissues of two native vascular plant species, boreal cranberry (Vaccinium oxycoccus) and black spruce (Picea mariana). 15N-labeled Sphagnum microcosms were grown within variable mesh size exclusion/inclusion fabrics in a nitrogen addition experiment in situ in order to investigate the role of mycorrhizal fungi in the uptake of newly fixed nitrogen. Up to 24% of daily fixed 15N label was transferred to vascular plant tissues during 2 months. Nitrogen addition resulted in decreased N2 fixation rates; however, with higher nitrogen availability there was a higher rate of 15N label uptake into the vascular plants, likely the result of increased production of dissolved organic nitrogen. Reliance on mycorrhizal networks for nitrogen acquisition was indicated by nitrogen isotope fractionation patterns. Moreover, N2 fixation activities in mosses were stimulated when vascular plants were grown in moss microcosms versus "moss only" treatments. Results indicate that bog vascular plants may derive considerable nitrogen from atmospheric N2 biologically fixed within Sphagnum mosses. This work demonstrates that

  5. Northern Desegregation: A Tale of Two Cities

    ERIC Educational Resources Information Center

    Danns, Dionne

    2011-01-01

    Studies on northern desegregation have focused on political strategies, the role of the courts, the responsibility of the federal government (HEW), and barriers to northern desegregation. Some have conducted individual case studies and comparative studies, and others have examined a number of cities. This article examines the way school…

  6. The Alcoholism Situation in a Northern City

    ERIC Educational Resources Information Center

    Martynov, M. Iu.; Martynova, D. Iu.

    2012-01-01

    Alcohol abuse in Russia has been increasing in recent years, especially in northern regions, as has the incidence of alcohol-related disease rates. A survey was conducted in Surgut (the Khanty-Mansi autonomous okrug) that determined the factors lending to the prevalence of alcohol abuse among the population of the northern city and assessed the…

  7. Environmental overview of geothermal development: northern Nevada

    SciTech Connect

    Slemmons, D.B.; Stroh, J.M.; Whitney, R.A.

    1980-08-01

    Regional environmental problems and issues associated with geothermal development in northern Nevada are studied to facilitate environmental assessment of potential geothermal resources. The various issues discussed are: environmental geology, seismicity of northern Nevada, hydrology and water quality, air quality, Nevada ecosystems, noise effects, socio-economic impacts, and cultural resources and archeological values. (MHR)

  8. Northern Parkway PTA Makes Health a Habit

    ERIC Educational Resources Information Center

    Ferdinand, Marilyn

    2011-01-01

    Health and fitness have been on the agenda of Northern Parkway Elementary School for quite some time, thanks to the concerted efforts of its involved and active PTA officers and members. For the past five years, the Northern Parkway PTA has held a popular and well-attended Family Fun and Fitness Night and has complemented the activities and…

  9. Bullying in Schools: A Northern Ireland Study

    ERIC Educational Resources Information Center

    Collins, Katrina; McAleavy, Gerry; Adamson, Gary

    2004-01-01

    Northern Ireland, unlike the Republic of Ireland or England, has no province-wide information on bullying in schools. This study provided baseline information on this complex issue across 120 schools in all five Education and Library Boards in Northern Ireland, comprising 60 primary and 60 post-primary schools, 1079 primary pupils (Year 6) and…

  10. Meeting Northern Arizona's Supported Employment Training Needs.

    ERIC Educational Resources Information Center

    Martin, William E., Jr.; And Others

    In 1989 Northern Arizona University established a Supported Employment Training Center (SETC) to increase the number of trained job coaches in northern Arizona and provide knowledge and skills in supported employment to personnel from cooperating schools and agencies. First-year SETC activities focused on assessment of the training needs of…

  11. Detection of Y chromosome sequences in a 45,X/46,XXq - patient by Southern blot analysis of PCR-amplified DNA and fluorescent in situ hybridization (FISH)

    SciTech Connect

    Kocova, M.; Siegel, S.F.; Wenger, S.L.

    1995-02-13

    In some cases of gonadal dysgenesis, cytogenetic analysis seems to be discordant with the phenotype of the patients. We have applied techniques such as Southern blot analysis and fluorescent in situ hybridization (FISH) to resolve the phenotype/genotype discrepancy in a patient with ambiguous genitalia in whom the peripheral blood karotype was 45,X. Gonadectomy at age 7 months showed the gonadal tissue to be prepubertal testis on the left side and a streak gonad on the right. The karyotype obtained from the left gonad was 45,X/46,XXq- and that from the right gonad was 45,X. Three different techniques, PCR amplification, FISH, and chromosome painting for X and Y chromosomes, confirmed the presence of Y chromosome sequences. Five different tissues were evaluated. The highest percentage of Y chromosome positive cells were detected in the left gonad, followed by the peripheral blood lymphocytes, skin fibroblasts, and buccal mucosa. No Y chromosomal material could be identified in the right gonad. Since the Xq- chromosome is present in the left gonad (testis), it is likely that the Xq- contains Y chromosomal material. Sophisticated analysis in this patient showed that she has at least 2 cell lines, one of which contains Y chromosomal material. These techniques elucidated the molecular basis of the genital ambiguity for this patient. When Y chromosome sequences are present in patients with Ullrich-Turner syndrome or gonadal dysgenesis, the risk for gonadal malignancy is significantly increased. Hence, molecular diagnostic methods to ascertain for the presence of Y chromosome sequences may expedite the evaluation of patients with the ambiguous genitalia. 21 refs., 4 figs., 2 tabs.

  12. Evaluation of the Bio-Rad Geenius HIV 1/2 Confirmation Assay as an Alternative to Western Blot in the Korean Population: A Multi-Center Study.

    PubMed

    Moon, Hee-Won; Huh, Hee Jin; Oh, Gwi Young; Lee, Sang Gon; Lee, Anna; Yun, Yeo-Min; Hur, Mina

    2015-01-01

    Recently updated recommendations for diagnosis of HIV infection suggest a new diagnostic algorithm including HIV-1/HIV-2 antibody differentiation immunoassay instead of western blot (WB) as a confirmatory testing. We evaluated Bio-Rad Geenius HIV1/2 confirmation assay as a simple and reliable alternative to WB in the Korean population with low HIV prevalence. The Geenius HIV1/2 was performed in a total of 192 serum specimens (140 reactive and 52 nonreactive specimens by ARCHITECT HIV Ag/Ab Combo assay) that were prospectively collected from five institutions. HIV-1 nucleic acid amplification test (NAT) was performed in negative or indeterminate specimens by Geenius HIV1/2 or WB. Among 140 reactive specimens by HIV Ag/Ab assay, 82 (58.6%) were positive for HIV-1 Ab by Geenius HIV1/2. Among 58 negative or indeterminate specimens by Geenius HIV1/2, four specimens (6.9%) were positive by HIV-1 NAT. The sensitivity and specificity of Geenius HIV1/2 were 95.3% and 100.0%, respectively. When we considered only WB, the sensitivity and specificity of Geenius HIV1/2 were 100.0% and 99.1%, respectively. Agreement between Geenius HIV1/2 and WB was excellent (weighted Kappa = 0.89). The Geenius HIV1/2 is simple and time-saving compared with WB. It has an excellent performance and can be a reliable alternative to WB. HIV-1 NAT should be performed in negative or indeterminate specimens by Geenius HIV1/2 to detect acute HIV infection as recommended in new HIV testing algorithms.

  13. Human T-Cell Lymphotropic Virus Type 1 Gag Indeterminate Western Blot Patterns in Central Africa: Relationship to Plasmodium falciparum Infection

    PubMed Central

    Mahieux, Renaud; Horal, Peter; Mauclère, Philippe; Mercereau-Puijalon, Odile; Guillotte, Micheline; Meertens, Laurent; Murphy, Edward; Gessain, Antoine

    2000-01-01

    To gain insight on the significance of human T-cell lymphotropic virus type 1 (HTLV-1) indeterminate serological reactivities, we studied villagers of South Cameroon, focusing on a frequent and specific HTLV-1 Gag indeterminate profile (HGIP) pattern (gag p19, p26, p28, and p30 without p24 or Env gp21 and gp46). Among the 102 sera studied, 29 from all age groups had a stable HGIP pattern over a period of 4 years. There was no epidemiological evidence for sexual or vertical transmission of HGIP. Seventy-five percent of HGIP sera reacted positively on MT2 HTLV-1-infected cells by immunofluorescence assay. However, we could not isolate any HTLV-1 virus or detect the presence of p19 Gag protein in cultures of peripheral blood mononuclear cells obtained from individuals with strong HGIP reactivity. PCR experiments conducted with primers for HTLV-1 and HTLV-2 (HTLV-1/2 primers) encompassing different regions of the virus did not yield HTLV-1/2 proviral sequences from individuals with HGIP. Using 11 peptides corresponding to HTLV-1 or HTLV-2 immunodominant B epitopes in an enzyme-linked immunosorbent assay, one epitope corresponding to the Gag p19 carboxyl terminus was identified in 75% of HGIP sera, while it was recognized by only 41% of confirmed HTLV-1-positive sera. A positive correlation between HTLV-1 optical density values and titers of antibody to Plasmodium falciparum was also demonstrated. Finally, passage of sera through a P. falciparum-infected erythrocyte-coupled column was shown to specifically abrogate HGIP reactivity but not the HTLV-1 pattern, suggesting the existence of cross-reactivity between HTLV-1 Gag proteins and malaria-derived antigens. These data suggest that in Central Africa, this frequent and specific Western blot is not caused by HTLV-1 infection but could instead be associated with P. falciparum infection. PMID:11060067

  14. Low Proviral Load is Associated with Indeterminate Western Blot Patterns in Human T-Cell Lymphotropic Virus Type 1 Infected Individuals: Could Punctual Mutations be Related?

    PubMed Central

    Cánepa, Camila; Salido, Jimena; Ruggieri, Matías; Fraile, Sindy; Pataccini, Gabriela; Berini, Carolina; Biglione, Mirna

    2015-01-01

    Background: indeterminate Western blot (WB) patterns are a major concern for diagnosis of human T-cell lymphotropic virus type 1 (HTLV-1) infection, even in non-endemic areas. Objectives: (a) to define the prevalence of indeterminate WB among different populations from Argentina; (b) to evaluate if low proviral load (PVL) is associated with indeterminate WB profiles; and (c) to describe mutations in LTR and tax sequence of these cases. Results: Among 2031 samples, 294 were reactive by screening. Of them, 48 (16.3%) were WB indeterminate and of those 15 (31.3%) were PCR+. Quantitative real-time PCR (qPCR) was performed to 52 HTLV-1+ samples, classified as Group 1 (G1): 25 WB+ samples from individuals with pathologies; Group 2 (G2): 18 WB+ samples from asymptomatic carriers (AC); and Group 3 (G3): 9 seroindeterminate samples from AC. Median PVL was 4.78, 2.38, and 0.15 HTLV-1 copies/100 PBMCs, respectively; a significant difference (p=0.003) was observed. Age and sex were associated with PVL in G1 and G2, respectively. Mutations in the distal and central regions of Tax Responsive Elements (TRE) 1 and 2 of G3 were observed, though not associated with PVL.The 8403A>G mutation of the distal region, previously related to high PVL, was absent in G3 but present in 50% of WB+ samples (p = 0.03). Conclusions: indeterminate WB results confirmed later as HTLV-1 positive may be associated with low PVL levels. Mutations in LTR and tax are described;  their functional relevance remains to be determined. PMID:26516904

  15. Simultaneous detection of Anaplasma and Ehrlichia species in ruminants and detection of Ehrlichia ruminantium in Amblyomma variegatum ticks by reverse line blot hybridization.

    PubMed

    Bekker, Cornelis P J; de Vos, Sander; Taoufik, Amar; Sparagano, Olivier A E; Jongejan, Frans

    2002-10-22

    The detection of Anaplasma and Ehrlichia species is usually based on species-specific PCR assays, since no assay is yet available which can detect and identify these species simultaneously. To this end, we developed a reverse line blot (RLB) assay for simultaneous detection and identification of Anaplasma and Ehrlichia species in domestic ruminants and ticks. In a PCR the hypervariable V1 region of the 16S ribosomal RNA (rRNA) gene was amplified with a set of primers unique for members of the genera Anaplasma and Ehrlichia [Int. J. Syst. Evol. Microbiol. 51 (2001) 2145]. Amplified PCR products from blood of domestic ruminants or Amblyomma variegatum tick samples were hybridized onto a membrane to which eight species-specific oligonucleotide probes and one Ehrlichia and Anaplasma catch-all oligonucleotide probe were covalently linked. No DNA was amplified from uninfected blood, nor from other hemoparasites such as Theileria annulata, or Babesia bigemina. The species-specific probes did not cross-react with DNA amplified from other species. E. ruminantium, A. ovis and another Ehrlichia were identified by RLB in blood samples collected from small ruminants in Mozambique. Finally, A. variegatum ticks were tested after feeding on E. ruminantium infected sheep. E. ruminantium could be detected in adult ticks even if feeding of nymphs was carried out 3.5 years post-infection. In conclusion, the developed species-specific oligonucleotide probes used in an RLB assay can simultaneously detect and identify several Ehrlichia and Anaplasma species. However, as no quantitative data for the detection limit are available yet, only positive results are interpretable at this stage.

  16. Development of a Rapid Reverse Blot Hybridization Assay for Detection of Clinically Relevant Antibiotic Resistance Genes in Blood Cultures Testing Positive for Gram-Negative Bacteria.

    PubMed

    Wang, Hye-Young; Yoo, Gilsung; Kim, Juwon; Uh, Young; Song, Wonkeun; Kim, Jong Bae; Lee, Hyeyoung

    2017-01-01

    Rapid and accurate identification of the causative pathogens of bloodstream infections is crucial for the prompt initiation of appropriate antimicrobial therapy to decrease the related morbidity and mortality rates. The aim of this study was to evaluate the performance of a newly developed PCR-reverse blot hybridization assay (REBA) for the rapid detection of Gram-negative bacteria (GNB) and their extended-spectrum β-lactamase (ESBL), AmpC β-lactamase, and carbapenemase resistance genes directly from the blood culture bottles. The REBA-EAC (ESBL, AmpC β-lactamase, carbapenemase) assay was performed on 327 isolates that were confirmed to have an ESBL producer phenotype, 200 positive blood culture (PBCs) specimens, and 200 negative blood culture specimens. The concordance rate between the results of REBA-EAC assay and ESBL phenotypic test was 94.2%. The sensitivity, specificity, positive predictive value, and negative predictive value of the REBA-EAC assay for GNB identification in blood culture specimens were 100% (95% CI 0.938-1.000, P < 0.001), 100% (95% CI 0.986-1.000, P < 0.001), 100% (95% CI 0.938-1.000, P < 0.001), and 100% (95% CI 0.986-1.000, P < 0.001), respectively. All 17 EAC-producing GNB isolates from the 73 PBCs were detected by the REBA-EAC assay. The REBA-EAC assay allowed easy differentiation between EAC and non-EAC genes in all isolates. Moreover, the REBA-EAC assay was a rapid and reliable method for identifying GNB and their β-lactamase resistance genes in PBCs. Thus, this assay may provide essential information for accelerating therapeutic decisions to achieve earlier appropriate antibiotic treatment during the acute phase of bloodstream infection.

  17. Identification of Babesia species infecting dogs using reverse line blot hybridization for six canine piroplasms, and evaluation of co-infection by other vector-borne pathogens.

    PubMed

    Yisaschar-Mekuzas, Yael; Jaffe, Charles L; Pastor, Josep; Cardoso, Luís; Baneth, Gad

    2013-01-31

    Canine infection by vector-borne hemoparasites is frequent in tropical and sub-tropical areas where exposure to hematophageous ectoparasites is intensive. A reverse line blot (RLB) assay was designed to improve the simultaneous detection of all named canine piroplasm species combined with other vector-borne pathogens of dogs including Ehrlichia canis, Hepatozoon canis and Leishmania infantum common in the Mediterranean basin. Blood samples of 110 dogs from Spain (n=21), Portugal (n=14) and Israel (n=75) were analyzed. The study evaluated 2 groups of dogs, 49 dogs with piroplasm infection detected by blood smear microscopy from Portugal, Spain and Israel, and 61 dogs surveyed from rural areas in Israel, for which infection status with vector-borne pathogens was unknown. Among the dogs previously diagnosed with piroplasmosis, infection with Babesia canis, Babesia vogeli, Babesia gibsoni and Theileria annae was detected in the Iberian dogs while only B. vogeli was found in Israeli dogs. These differences are attributed to the absence of tick vectors for some piroplasm species such as Dermacentor reticulatus in Israel. Eleven (79%) of the Babesia-positive dogs from Portugal were co-infected with other pathogens including L. infantum, H. canis and E. canis. Eight of 61 (13%) rural Israeli dogs were co-infected with two or more pathogens including B. vogeli, L. infantum, E. canis, and H. canis. Triple infections were demonstrated in 2 dogs. The RLB detection limit for Babesia was 50-fold lower than that of PCR. This study presents a RLB to simultaneously detect and separate the major vector-borne dog pathogens in southern Europe and the Middle East.

  18. Collagen chains detected by western blotting using a /sup 125/I-labeled 45K fragment of fibronectin (45K FN)

    SciTech Connect

    Ristagno, R.; Heimer, R.; Fishman, A.P.; Sampson, P.M.

    1987-05-01

    The objective was to improve the sensitivity and specificity of detection of unlabeled collagen chains in biologic fluids. Chains of Types I,II,III,IV and XI (1..cap alpha..2..cap alpha..3..cap alpha..) collagen were separated by SDS PAGE. Their complete transfer to nitrocellulose was obtained by electrophoresis for 16 h at 150 mA with 10 mM Tris, 117 mM glycine, 100 mM cysteine, 0.1% SDS and 10% methanol. The 45K FN was prepared by chymotryptic digestion of fibronectin adsorbed to gelatin-Sepharose, followed by elution with 1.2 M urea, 1 M Tris-NaCl, pH 8.3 and iodination. When exposed to the nitrocellulose transblot at pH 9.5 and 4/sup 0/C, 45K FN did not react with IgG, fibrinogen, myosin, albumin or carbonic anhydrase. These proteins interfere in the assay under conditions of lower pH and higher temperature. The autoradiographs of the transblots were evaluated by densitometry and reflected results also obtained by dot blotting, that chains of collagen Types I,II,III were detectable at 4 ng and those of collagen Type IV at 12 ng. Generally, ..cap alpha..,BETA, and ..gamma.. chains were detectable. The 45K FN reacted equally with ..cap alpha..1(I) and ..cap alpha..2(I), but for Type XI the 1..cap alpha.. chain had considerably more reactivity than 2..cap alpha.. or 3..cap alpha... As the 45K FN was specific for collagens added to plasma, the authors method appears useful for qualitative and quantitative assays of unlabeled collagens in biologic fluids.

  19. Development of a Rapid Reverse Blot Hybridization Assay for Detection of Clinically Relevant Antibiotic Resistance Genes in Blood Cultures Testing Positive for Gram-Negative Bacteria

    PubMed Central

    Wang, Hye-young; Yoo, Gilsung; Kim, Juwon; Uh, Young; Song, Wonkeun; Kim, Jong Bae; Lee, Hyeyoung

    2017-01-01

    Rapid and accurate identification of the causative pathogens of bloodstream infections is crucial for the prompt initiation of appropriate antimicrobial therapy to decrease the related morbidity and mortality rates. The aim of this study was to evaluate the performance of a newly developed PCR-reverse blot hybridization assay (REBA) for the rapid detection of Gram-negative bacteria (GNB) and their extended-spectrum β-lactamase (ESBL), AmpC β-lactamase, and carbapenemase resistance genes directly from the blood culture bottles. The REBA-EAC (ESBL, AmpC β-lactamase, carbapenemase) assay was performed on 327 isolates that were confirmed to have an ESBL producer phenotype, 200 positive blood culture (PBCs) specimens, and 200 negative blood culture specimens. The concordance rate between the results of REBA-EAC assay and ESBL phenotypic test was 94.2%. The sensitivity, specificity, positive predictive value, and negative predictive value of the REBA-EAC assay for GNB identification in blood culture specimens were 100% (95% CI 0.938–1.000, P < 0.001), 100% (95% CI 0.986–1.000, P < 0.001), 100% (95% CI 0.938–1.000, P < 0.001), and 100% (95% CI 0.986–1.000, P < 0.001), respectively. All 17 EAC-producing GNB isolates from the 73 PBCs were detected by the REBA-EAC assay. The REBA-EAC assay allowed easy differentiation between EAC and non-EAC genes in all isolates. Moreover, the REBA-EAC assay was a rapid and reliable method for identifying GNB and their β-lactamase resistance genes in PBCs. Thus, this assay may provide essential information for accelerating therapeutic decisions to achieve earlier appropriate antibiotic treatment during the acute phase of bloodstream infection. PMID:28232823

  20. The prevalence of urogenital micro-organisms detected by a multiplex PCR-reverse line blot assay in women attending three sexual health clinics in Sydney, Australia.

    PubMed

    McKechnie, Michelle L; Hillman, Richard J; Jones, Rachel; Lowe, Penelope C; Couldwell, Deborah L; Davies, Stephen C; King, Fiona; Kong, Fanrong; Gilbert, Gwendolyn L

    2011-07-01

    This study used a previously described multiplex PCR-based reverse line blot (mPCR/RLB) assay to assess the prevalence and distribution of 14 urogenital pathogens or putative pathogens, namely Neisseria gonorrhoeae, Chlamydia trachomatis, Mycoplasma genitalium, Mycoplasma hominis, Trichomonas vaginalis, Gardnerella vaginalis, Ureaplasma parvum, Ureaplasma urealyticum, Neisseria meningitidis, Streptococcus pneumoniae, Haemophilus influenzae, herpes simplex virus types 1 and 2, and human adenovirus. First-voided urine specimens and endocervical and self-collected vaginal swabs from each of 216 women attending three sexual health clinics in Sydney, Australia, were tested and the results were compared with those of reference methods for each organism. One hundred and sixty-eight women (77.7 %) had at least one and 105 (48.6 %) had more than one target organism, most commonly G. vaginalis and Ureaplasma spp. The prevalence of each of the four known sexually transmissible pathogens was <5 %. Of the 216 women, 111 (51.4 %) reported at least one symptom consistent with genital or urethral infection, including discharge, pain or discomfort. Only G. vaginalis was detected more frequently in women with symptoms (P = 0.05). The specificity of the mPCR/RLB assay compared with that of the reference methods for each organism and for all specimen types was 100 %. The mean sensitivities of the mPCR/RLB assay compared with those of the reference methods for self-collected vaginal swabs, cervical swabs and first-voided urine specimens for all organisms were 99.3, 98.1 and 84.6 %, respectively; however, these differences were not significant. There were no differences in sensitivities between specimen types for C. trachomatis, N. gonorrhoeae, T. vaginalis and H. influenzae, although all were found infrequently. Overall, the mPCR/RLB platform was found to be an accurate testing platform in a sexual health clinic setting.

  1. Localization of α-synuclein in teleost central nervous system: immunohistochemical and Western blot evidence by 3D5 monoclonal antibody in the common carp, Cyprinus carpio.

    PubMed

    Vaccaro, Rosa; Toni, Mattia; Casini, Arianna; Vivacqua, Giorgio; Yu, Shun; D'este, Loredana; Cioni, Carla

    2015-05-01

    Alpha synuclein (α-syn) is a 140 amino acid vertebrate-specific protein, highly expressed in the human nervous system and abnormally accumulated in Parkinson's disease and other neurodegenerative disorders, known as synucleinopathies. The common occurrence of α-syn aggregates suggested a role for α-syn in these disorders, although its biological activity remains poorly understood. Given the high degree of sequence similarity between vertebrate α-syns, we investigated this proteins in the central nervous system (CNS) of the common carp, Cyprinus carpio, with the aim of comparing its anatomical and cellular distribution with that of mammalian α-syn. The distribution of α-syn was analyzed by semiquantitative western blot, immunohistochemistry, and immunofluorescence by a novel monoclonal antibody (3D5) against a fully conserved epitope between carp and human α-syn. The distribution of 3D5 immunoreactivity was also compared with that of choline acetyltransferase (ChAT), tyrosine hydroxylase (TH), and serotonin (5HT) by double immunolabelings. The results showed that a α-syn-like protein of about 17 kDa is expressed to different levels in several brain regions and in the spinal cord. Immunoreactive materials were localized in neuronal perikarya and varicose fibers but not in the nucleus. The present findings indicate that α-syn-like proteins may be expressed in a few subpopulations of catecholaminergic and serotoninergic neurons in the carp brain. However, evidence of cellular colocalization 3D5/TH or 3D5/5HT was rare. Differently, the same proteins appear to be coexpressed with ChAT by cholinergic neurons in several motor and reticular nuclei. These results sustain the functional conservation of the α-syn expression in cholinergic systems and suggest that α-syn modulates similar molecular pathways in phylogenetically distant vertebrates.

  2. Evaluation of the Punch-it™ NA-Sample kit for detecting microbial DNA in blood culture bottles using PCR-reverse blot hybridization assay.

    PubMed

    Kim, Jungho; Wang, Hye-Young; Kim, Seoyong; Park, Soon Deok; Yu, Kwangmin; Kim, Hyo Youl; Uh, Young; Lee, Hyeyoung

    2016-09-01

    DNA extraction efficiency affects the success of PCR-based method applications. The Punch-it™ NA-Sample kit for extracting DNA by using paper chromatography is technically easy to use and requires just two reagents and only 10min to complete. The Punch-it™ NA-Sample kit could be offered as a rapid, accurate, and convenient method for extracting bacterial and fungal DNA from blood culture bottles. We compared the efficiencies of the commercial kit (Punch-it™ NA-Sample kit) and an in-house conventional boiling method with Chelex-100 resin for DNA extraction from blood culture bottles. The efficiency of the two DNA extraction methods was assessed by PCR-reverse blot hybridization assay (PCR-REBA, REBA Sepsis-ID) for detecting Gram positive (GP) bacteria, Gram negative (GN) bacteria, and Candida species with 196 positive and 200 negative blood culture bottles. The detection limits of the two DNA extraction methods were 10(3)CFU/mL for GP bacteria, 10(3)CFU/mL for GN bacteria, and 10(4)CFU/mL for Candida. The sensitivity and specificity of the Punch-it™ NA-Sample kit by REBA Sepsis-ID were 95.4% (187/196) and 100% (200/200), respectively. The overall agreement of the two DNA extraction methods was 98.9% (392/396). Three of four samples showing discrepant results between the two extraction methods were more accurately matched up with the Punch-it™ NA-Sample kit based on conventional culture methods. The results indicated that the Punch-it™ NA-Sample kit extracted bacterial and fungal DNA in blood culture bottles and allowed extracted DNA to be used in molecular assay.

  3. Farmer's lung in Northern Ireland.

    PubMed Central

    Stanford, C F; Hall, G; Chivers, A; Martin, B; Nicholls, D P; Evans, J

    1990-01-01

    A total of 381 farmers in Northern Ireland were studied using a questionnaire, pulmonary function tests, and antibody levels to Micropolyspora faena to assess the incidence of farmer's lung. Twenty (4.9%) had a history of a previous diagnosis of farmer's lung by their doctor. Forty four (10.4%) had delayed onset symptoms compatible with farmer's lung, 32 (7.9%) had precipitant antibody, and 61 (15%) had raised antibody by the enzyme linked immunosorbent (ELISA) method. Restricted lungs were present physiologically in 40 (9.8%). A confirmation of delayed symptoms and precipitant antibody was present in seven (1.7%) whereas delayed symptoms and ELISA antibody was present in nine (2.2%). Using either antibody method only two (0.5%) had a combination of antibody to M faenae, delayed onset symptoms, and restricted pulmonary physiology. PMID:2357452

  4. Farmer's lung in Northern Ireland.

    PubMed

    Stanford, C F; Hall, G; Chivers, A; Martin, B; Nicholls, D P; Evans, J

    1990-05-01

    A total of 381 farmers in Northern Ireland were studied using a questionnaire, pulmonary function tests, and antibody levels to Micropolyspora faena to assess the incidence of farmer's lung. Twenty (4.9%) had a history of a previous diagnosis of farmer's lung by their doctor. Forty four (10.4%) had delayed onset symptoms compatible with farmer's lung, 32 (7.9%) had precipitant antibody, and 61 (15%) had raised antibody by the enzyme linked immunosorbent (ELISA) method. Restricted lungs were present physiologically in 40 (9.8%). A confirmation of delayed symptoms and precipitant antibody was present in seven (1.7%) whereas delayed symptoms and ELISA antibody was present in nine (2.2%). Using either antibody method only two (0.5%) had a combination of antibody to M faenae, delayed onset symptoms, and restricted pulmonary physiology.

  5. Electrical Fatalities in Northern Ireland

    PubMed Central

    Lucas, James

    2009-01-01

    A review of autopsy reports in cases of electrocution in Northern Ireland revealed that there were 50 accidental electrocutions and 9 suicidal electrocutions over a 22 year period (1982 – 2003). No cases of homicidal electrocution were detected in this jurisdiction. Analysis of the cohort of accidental electrocutions showed that there was a clear skew towards young and middle-aged male adults with deaths occurring more frequently in the summer months. Almost 60% of individuals were engaged in occupational tasks when they were accidentally electrocuted. High and low voltage-related deaths occurred with similar frequency and electrical appliances were found to be responsible for approximately one third of accidental electrocutions. The potential hazards of electricity must continue to be stressed in public safety campaigns if these relatively uncommon but tragic deaths are to be prevented. PMID:19252729

  6. Ultraviolet resources over Northern Eurasia.

    PubMed

    Chubarova, Natalia; Zhdanova, Yekaterina

    2013-10-05

    We propose a new climatology of UV resources over Northern Eurasia, which includes the assessments of both detrimental (erythema) and positive (vitamin D synthesis) effects of ultraviolet radiation on human health. The UV resources are defined by using several classes and subclasses - UV deficiency, UV optimum, and UV excess - for 6 different skin types. To better quantifying the vitamin D irradiance threshold we accounted for an open body fraction S as a function of effective air temperature. The spatial and temporal distribution of UV resources was estimated by radiative transfer (RT) modeling (8 stream DISORT RT code) with 1×1° grid and monthly resolution. For this purpose special datasets of main input geophysical parameters (total ozone content, aerosol characteristics, surface UV albedo, UV cloud modification factor) have been created over the territory of Northern Eurasia. The new approaches were used to retrieve aerosol parameters and cloud modification factor in the UV spectral region. As a result, the UV resources were obtained for clear-sky and mean cloudy conditions for different skin types. We show that the distribution of UV deficiency, UV optimum and UV excess is regulated by various geophysical parameters (mainly, total ozone, cloudiness and open body fraction) and can significantly deviate from latitudinal dependence. We also show that the UV optimum conditions can be simultaneously observed for people with different skin types (for example, for 4-5 skin types at the same time in spring over Western Europe). These UV optimum conditions for different skin types occupy a much larger territory over Europe than that over Asia.

  7. North Atlantic sea surface temperature, solar activity and the climate of Northern Fennoscandia

    NASA Astrophysics Data System (ADS)

    Ogurtsov, M.; Lindholm, M.; Jalkanen, R.; Veretenenko, S. V.

    2017-02-01

    Seven proxies of summer temperature in Northern Fennoscandia, sea surface temperature in the North Atlantic and solar activity were analyzed over AD 1567-1986. A stable and significant positive correlation between summer temperatures in Northern Fennoscandia and sea surface temperature in the North Atlantic is shown to exist during the entire time interval. In addition, a significant correlation between solar activity and (a) summer temperature in Northern Fennoscandia as well as (b) surface temperature in the North Atlantic was found during AD 1715-1986. Throughout 1567-1715 correlation is less significant and has an opposite sign. Thus we show that the variation of sea surface temperature in the North Atlantic could be a physical agent, which transferred solar influence on Northern Fennoscandian temperature at least during AD 1715-1986.

  8. Use of the Falcon assay screening test--enzyme-linked immunosorbent assay (FAST-ELISA) and the enzyme-linked immunoelectrotransfer blot (EITB) to determine the prevalence of human fascioliasis in the Bolivian Altiplano.

    PubMed

    Hillyer, G V; Soler de Galanes, M; Rodriguez-Perez, J; Bjorland, J; Silva de Lagrava, M; Ramirez Guzman, S; Bryan, R T

    1992-05-01

    A collaborative study between the University of Puerto Rico School of Medicine, the Centers for Disease Control, the Bolivian Ministry of Health, and private voluntary organizations (Foster Parents Plan International and Danchurchaid) working in Bolivia has identified a region in the northwestern Altiplano of Bolivia near Lake Titicaca as harboring the highest prevalence of human fascioliasis in the world reported to date. Two serologic techniques (the Falcon assay screening test-enzyme-linked immunosorbent assay [FAST-ELISA] and the enzyme-linked immunoelectrotransfer blot [EITB]) were used in the determination of its prevalence. One hundred serum samples and 73 stool samples were obtained from Aymara Indians from Corapata, Bolivia. Antibody absorbance levels to Fasciola hepatica excretion-secretion antigens were compared with EITB banding patterns using the same antigen preparation. A positive FAST-ELISA result was defined as an absorbance value greater than the mean plus three standard deviations of two sets of normal negative controls (Puerto Rican and Bolivian). Using this criterion, 53 of 100 sera tested were found positive by this technique. Within this group, 19 (95%) of 20 individuals who were parasite positive were also positive by FAST-ELISA. An additional 24 individuals who were negative for F. hepatica eggs and 10 individuals for whom no specimens were received were also positive by FAST-ELISA. Among the 53 individuals negative for F. hepatica eggs, 29 were also negative by FAST-ELISA. The EITB analysis of the sera from confirmed infected individuals revealed at least three F. hepatica (Fh) bands with molecular weights of 12, 17, and 63 kD, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)

  9. How Safe Are Color Additives?

    MedlinePlus

    ... Home For Consumers Consumer Updates How Safe are Color Additives? Share Tweet Linkedin Pin it More sharing ... Consumer Updates RSS Feed Download PDF (380 K) Color additives give the red tint to your fruit ...

  10. Middle Cretaceous dinosaur assemblages from northern Brazil and northern Africa and their implications for northern Gondwanan composition

    NASA Astrophysics Data System (ADS)

    Candeiro, Carlos Roberto A.

    2015-08-01

    Dinosaurs are one of the most dominant groups in Cretaceous reptilian faunas. A summary of their record in northern Brazil and northern Africa during the middle of the Cretaceous Period (Aptian-Cenomanian) is presented here. Dinosaurs are represented by 32 species (three ornithischians, six sauropods and 23 theropods) from Brazil, Egypt, Lybia, Morocco, Niger, Sudan and Tunisia. These dinosaur assemblages provide fundamental data about distribution and composition of sauropods and theropods in northern Gondwana during the middle of the Cretaceous Period and confirm these assemblages to be among the most important dinosaur faunas in the north Gondwana areas.

  11. The northern light. From mythology to space research.

    NASA Astrophysics Data System (ADS)

    Brekke, A.; Egeland, A.

    Contents: The northern light in folklore and mythology. The northern light in Norse literature. The northern light - a source of inspiration. Accounts of northern lights in Scandinavia - from the Viking era to the Renaissance. The northern light in Scandinavia during the eighteenth century. Scientific auroral experiments beginning in the nineteenth century. Norwegian auroral pioneers in the dawn of our century. The northern lights as weather signs - and the auroral sound. Northern lights and geomagnetic disturbances - their influence on daily life. Auroral research as a tool to study the upper atmosphere and near space. The first systematic observations of the northern light in Norway. Summary and concluding remarks.

  12. Detergent Additive for Lubricating Oils,

    DTIC Science & Technology

    The Russian patent pertains to a method of producing additives for lubricating oils . A method is known for producing an antiwear additive for... lubricating oils by processing phenols with phosphorus oxychloride, phosphoric acid esters are obtained. In order to give the additive detergent properties

  13. Improving Seismic Monitoring in Northern Switzerland

    NASA Astrophysics Data System (ADS)

    Plenkers, Katrin; Husen, Stephan; Schnellmann, Michael

    2013-04-01

    We currently densify the existing Swiss Digital Seismic Network in northern Switzerland by additional 10 stations. The new network aims at observing seismicity in northern Switzerland with a completeness of Mc = 1.0 and a location error < 0.5 km in epicenter and < 2 km in focal depth. Monitoring of weak seismic events in this region is challenging, because the area of interest is densely populated and geology is dominated by the Swiss molasse basin. A optimal network-design and a thoughtful choice for station-sites is, therefore, mandatory. To help with decision making we developed a multi-step assessment-scheme that takes into account local noise level, geology, infrastructure, and costs necessary to realize the station. The assessment scheme is weighting the different parameters and the most promising sites are identified. In a first step, all potential sites are classified based on information from topographic maps and site inspection. In a second step, local noise conditions are measured at selected sites. Finally, the most promising sites are classified taking into account results from the test measurements and updated information on local geology, availability of electricity and data transmission, and installation costs. The first station went into operation in September 2012. In this study, we present lessons learnt during the network realization. We review the proposed assessment-scheme and demonstrate potentials and limitations for seismic monitoring in such an environment. We find that the step-wise search for the optimum installation site was successful. Test measurements conducted and analyzed for the assessment-scheme are representative for the performance of the later installed stations. In one case a significantly improved installation was possible, because the test measurement revealed noise sources that could be diminished. The assessment scheme allowed to compare station from different regions, which was especially helpful in judging, whether the

  14. Oak ecosystem succession of the Northern Caucasus

    NASA Astrophysics Data System (ADS)

    Chalaya, Elena; Slepykh, Victor; Efimenko, Natalia; Povolotskaya, Nina

    2014-05-01

    English oak (Quercus robur L.) along with its well-known good properties has a high sanitary-hygienic and curative potential. Its volatile metabolites (VM) influence bacteriostatically Staphylococcus aureus 209r, oppressing it in vitro by 85% compared with the control, and Escherichia coli by 45%. There is the least amount of epiphytic microorganisms on the leaves of Q. robur L. compared with some tree species [1]. In addition, VM of Q. robur L. have direct milieu (hypotensive) effects on the organism under its canopy, lowering blood pressure by 20-25 mm Hg [2]. A.P. Kazankin (1993) [4] calculated the prehistoric formula of forest species composition of Caucasian Mineral Waters region (Northern Caucasus): 6Qr3Crp1Fr which was based on the theory of calcium-magnesium absorption complex. According to the theory, underground mineral water, soil, forest litter and the leaves of ground vegetation of the area have the same ratio of the cations Ca and Mg - calcium-magnesium index [3]. Hence oldgrowth in the region consisted of oak (Qr) by 60%, hornbeam (Carpinus-Crp) by 30% and ash (Fraxinus-Fr) only by 10%. Currently, the formula of the forests of the region has been changed by man: 5Fr3Crp2Qr. The proportion of oak forests has decreased to 20%, the proportion of ash has increased by 50%, but the proportion of hornbeam hasn't changed. So it is relevant to restore oak forests of the region in the former ratio to other forest-forming species - ash and hornbeam. Taking into consideration the change of economic formation of society in Russia, it is extremely important to restore natural seed oak forests. Therein the luminance of surface areal is a limiting factor. We have calculated that the natural recovery of oak forests is possible providing observation of optimal moisture standards and soil fertility in combination with solar light within 10-24% from the light of open space. Measures for promotion of oak natural regeneration in mountain resorts of the Northern Caucasus

  15. Identification in gelada baboons (Theropithecus gelada) of a distinct simian T-cell lymphotropic virus type 3 with a broad range of Western blot reactivity.

    PubMed

    Van Dooren, Sonia; Shanmugam, Vedapuri; Bhullar, Vinod; Parekh, Bharat; Vandamme, Anne-Mieke; Heneine, Walid; Switzer, William M

    2004-02-01

    Antibodies to simian T-cell lymphotropic virus (STLV) were found in serum or plasma from 12 of 23 (52.2 %) gelada baboons (Theropithecus gelada) captive in US zoos. A variety of Western blot (WB) profiles was seen in the 12 seroreactive samples, including human T-cell lymphotropic virus (HTLV)-1-like (n=5, 41.7 %), HTLV-2-like (n=1, 8.3 %), HTLV-untypable (n=4, 33.3 %) and indeterminate (n=2, 16.6 %) profiles. Phylogenetic analysis of tax or env sequences that had been PCR amplified from peripheral blood lymphocyte DNA available from nine seropositive geladas showed that four were infected with identical STLV-1s; these sequences clustered with STLV-1 from Celebes macaques and probably represent recent cross-species infections. The tax sequences from the five remaining geladas were also identical and clustered with STLV-3. Analysis of the complete STLV-3 genome (8917 bp) from one gelada, TGE-2117, revealed that it is unique, sharing only 62 % similarity with HTLV-1/ATK and HTLV-2/Mo. STLV-3/TGE-2117 was closest genetically to STLV-3 from an Eritrean baboon (STLV-3/PH969, 95.6 %) but more distant from STLV-3s from red-capped mangabeys from Cameroon and Nigeria (STLV-3/CTO-604, 87.7 %, and STLV-3/CTO-NG409, 87.2 %, respectively) and Senegalese baboons (STLV-3/PPA-F3, 88.4 %). The genetic relatedness of STLV-3/TGE-2117 to STLV-3 was confirmed by phylogenetic analysis of a concatenated gag-pol-env-tax sequence (6795 bp). An ancient origin of 73 628-109 809 years ago for STLV-3 was estimated by molecular clock analysis of third-codon positions of gag-pol-env-tax sequences. LTR sequences from five STLV-3-positive geladas were >99 % identical and clustered with that from a Papio anubisxP. hamadryas hybrid Ethiopian baboon, suggesting a common source of STLV-3 in these sympatric animals. LTR sequences obtained 20 years apart from a mother-infant pair were identical, providing evidence of both mother-to-offspring transmission and a high genetic stability of STLV-3. Since

  16. Demographic response of northern spotted owls to barred owl removal

    USGS Publications Warehouse

    Diller, V. Lowell; Hamm, Keith A; Early, Desiree A; Lamphear, David W; Katie Dugger,; Yackulic, Charles B.; Schwarz, Carl J.; Carlson, Peter C.; McDonald, Trent L.

    2016-01-01

    the treated areas and 0.822 for the untreated areas. Derived estimates of population change on both study areas showed the same general decline before removal with an estimated slope of –0.0036 per year. Following removal, the rate of population change on the treated areas increased to an average of 1.029 but decreased to an average of 0.870 on the untreated areas. The results from this first experiment demonstrated that lethal removal of barred owls allowed the recovery of northern spotted owl populations in the treated portions of our study area. If additional federally funded barred owl removal experiments provide similar results, this could be the foundation for development of a long-term conservation strategy for northern spotted owls.

  17. Localization of eight additional genes in the human major histocompatibility complex, including the gene encoding the casein kinase II {beta} subunit (CSNK2B)

    SciTech Connect

    Albertella, M.R.; Jones, H.; Thomson, W.

    1996-09-01

    A wide range of autoimmune and other diseases are known to be associated with the major histocompatibility complex. Many of these diseases are linked to the genes encoding the polymorphic histocompatibility complex. Many of these diseases are linked to the genes encoding the polymorphic histocompatibility antigens in the class I and class II regions, but some appear to be more strongly associated with genes in the central 1100-kb class III region, making it important to characterize this region fully for the presence of novel genes. An {approximately}220-kb segment of DNA in the class III region separating the Hsp70 (HSPA1L) and BAT1 (D6S8IE) genes, which was previously known to contain 14 genes. Genomic DNA fragments spanning the gaps between the known genes were used as probes to isolate cDNAs corresponding to five new genes within this region. Evidence from Northern blot analysis and exon trapping experiments that suggested the presence of at least two more new genes was also obtained. Partial cDNA and complete exonic genomic sequencing of one of the new genes has identified it as the casein kinase II{beta} subunit (CSNK2B). Two of the other novel genes lie within a region syntenic to that implicated in susceptibility to experimental allergic orchitis in the mouse, an autoimmune disease of the testis, and represent additional candidates for the Orch-1 locus associated with this disease. In addition, characterization of the 13-kb intergenic gap separating the RD (D6545) and G11 (D6S60E) genes has revealed the presence of a gene encoding a 1246-amino-acid polypeptide that shows significant sequence similarity to the yeast anti-viral Ski2p gene product. 49 refs., 8 figs.

  18. Incorporation of additives into polymers

    DOEpatents

    McCleskey, T. Mark; Yates, Matthew Z.

    2003-07-29

    There has been invented a method for incorporating additives into polymers comprising: (a) forming an aqueous or alcohol-based colloidal system of the polymer; (b) emulsifying the colloidal system with a compressed fluid; and (c) contacting the colloidal polymer with the additive in the presence of the compressed fluid. The colloidal polymer can be contacted with the additive by having the additive in the compressed fluid used for emulsification or by adding the additive to the colloidal system before or after emulsification with the compressed fluid. The invention process can be carried out either as a batch process or as a continuous on-line process.

  19. Coal bed methane potential of northern Appalachian basin

    SciTech Connect

    Kelafant, J.; Wicks, D.; Kuuskraa, V.

    1987-09-01

    Coal-bed methane in the northern part of the Appalachian basin represents a significant potential source of natural gas for the northeastern US. The numerous, relatively thick and areally extensive coal beds of attractive rank and depth offer the promise of large, low-cost reserves of natural gas. The proximity of the supply source to northeastern markets and the location of the basin within an established pipeline infrastructure further enhance its economic outlook. Lewin and Associations, Inc., under the sponsorship of the Gas Research Institute and with participation of numerous basin geologists, completed a resource economics study of coal bed methane in the northern part of the Appalachian basin. As part of this study, six major coal seams and groups have been targeted as holding the most potential for containing large volumes of gas: the Brookville-Clarion, Kittanning, Freeport, Pittsburgh, Sewickley, and Waynesburg coals. In addition to these major seams and groups, several less areally extensive coals are economical on a local level, particularly the Brush Creek and Mahoning coals in Pennsylvania. The preliminary estimate of gas in place for the northern part of the Appalachian basin is between 50-100 tcf, with a considerable portion of this resource potentially economic at today's gas prices. The coal seams considered by this resource assessment study are at 12 in. thick and with greater than 400 ft of overburden.

  20. Tourism, biodiversity and protected areas--Review from northern Fennoscandia.

    PubMed

    Tolvanen, Anne; Kangas, Katja

    2016-03-15

    Tourist numbers in northern Fennoscandia outweigh those in other northern boreal - arctic regions, which creates a specific need to evaluate the impacts of tourism. This review 1) identifies patterns and trends in the vegetation and wildlife of northern Fennoscandian terrestrial ecosystems as a consequence of tourism and recreation, 2) discusses the implications of findings in terms of the intensity, area and magnitude of impacts, changing climate and management needs under increasing tourist pressure, and 3) identifies research gaps. The reviewed studies show negative environmental and biodiversity impacts that are most pronounced near tourist resorts. The most sensitive plants, birds and mammals decline or disappear from the disturbed sites, and the species composition shifts from 'wild' species to cultural and human associated species. There is little research on the spread of alien species, but the few examples show that alien species can be promoted by tourism activities. Impacts of the use of motorized vehicles have not been widely studied either, despite the extensive track network which can cause disturbance to wildlife. The integrated impacts of tourism and climate change on the vegetation and wildlife was not addressed directly in any of the reviewed studies. In addition, little research has been done on carrying out restoration at tourist areas. Scientific research on these topics is needed to prevent, minimize or restore the most negative ecological impacts of tourism and recreation.

  1. The puzzling origin of the Martian Northern Lowlands.

    NASA Astrophysics Data System (ADS)

    Altieri, F.; Carrozzo, G.; Carli, C.; Geminale, A.; Bellucci, G.

    Surface studies of the northern lowlands of Mars have shown that this region has undergone a complex history including volcanism, sedimentary deposition and secondary modification by climate change. Despite these analyses, the origin and the evolution of this region are still debated. No clear and definitive evidences have been found so far to conclude whether these plains were formed by a giant impact, were once covered by an ocean or were filled by a large quantity of lavas. In the visible and infrared spectral range, the northern lowlands differ from southern terrains in the NIR negative slope while they exhibit VNIR spectra similar to the southern pyroxene-rich areas (Carrozzo et al., 2012). These observations, combined with both recent detection of mafic minerals at higher spatial resolution by CRISM (Salvatore et al., 2010) and recent results of Horgan and Bell (2012), supports that their mineralogy is linked to weathered basalts with a glassy component. In addition to this, the spectral similarity of Acidalia area with the northern circumpolar sand dunes, apart from the hydration features, suggests that the weathering processes that took place there could be related to past glacial activity, in agreement with superficial morphology showing glacial structures. Aim of this work is to combine the OMEGA mineralogical maps with morphological features (Tanaka et al., 2011) and investigate possible terrestrial analogues in order to give some constrains on the composition and origin of these puzzling Martian terrains.

  2. Northern Terra Meridiani's 'Monument Valley'

    NASA Technical Reports Server (NTRS)

    2002-01-01

    Northern Terra Meridiani, near the intersection of the martian equator and prime meridian, is a region of vast exposures of layered rock. A thermal image from the Phobos 2 orbiter in 1989 showed these materials to be anomalously cool during the daytime, an observation very suggestive of dense, hardened materials like rock. Mars Global Surveyor (MGS) Mars Orbiter Camera (MOC) images of this region show layered material exposed in cliffs, buttes, and mesas that in some ways resemble the rock outcrops of northern Arizona and southeastern Utah in North America (e.g., Monument Valley, Canyonlands, Zion National Park, Four Corners). MGS MOC Extended Mission operations have included several hundred opportunities for the spacecraft to be rolled off-nadir (i.e., at an angle other than 'straight down') to take pictures that repeat earlier MOC coverage. These repeat images, because they are taken from a different angle, can be combined with the original picture to produce a stereoscopic ('3-D') view. The image shown here is a composite of two pictures, the first taken October 23, 2000, the second acquired by pointing the spacecraft off-nadir on May 15, 2001. This view shows four buttes and a pinnacle (near left-center) composed of eroded, layered rock. The four buttes are each capped by the remains of a single layer of rock that is harder than the materials beneath it. It is the presence of this cap rock that has permitted these buttes to remain standing after surrounding materials were eroded away. Like the buttes of Monument Valley in the Navajo Nation on the Arizona/Utah border, these are believed to consist of sedimentary rocks, perhaps deposited in water or by wind, though some scientists have speculated that they could be made of thick accumulations of volcanic ash. The image covers an area approximately 3 km (1.9 mi) across and is illuminated by sunlight from the left. To see the image in 3-D, red (left-eye) and blue (right-eye) '3-D' glasses are required.

  3. Additive manufacturing of optical components

    NASA Astrophysics Data System (ADS)

    Heinrich, Andreas; Rank, Manuel; Maillard, Philippe; Suckow, Anne; Bauckhage, Yannick; Rößler, Patrick; Lang, Johannes; Shariff, Fatin; Pekrul, Sven

    2016-08-01

    The development of additive manufacturing methods has enlarged rapidly in recent years. Thereby, the work mainly focuses on the realization of mechanical components, but the additive manufacturing technology offers a high potential in the field of optics as well. Owing to new design possibilities, completely new solutions are possible. This article briefly reviews and compares the most important additive manufacturing methods for polymer optics. Additionally, it points out the characteristics of additive manufactured polymer optics. Thereby, surface quality is of crucial importance. In order to improve it, appropriate post-processing steps are necessary (e.g. robot polishing or coating), which will be discussed. An essential part of this paper deals with various additive manufactured optical components and their use, especially in optical systems for shape metrology (e.g. borehole sensor, tilt sensor, freeform surface sensor, fisheye lens). The examples should demonstrate the potentials and limitations of optical components produced by additive manufacturing.

  4. The Northern Bering Sea: An Arctic Ecosystem in Change

    NASA Astrophysics Data System (ADS)

    Grebmeier, J. M.; Cooper, L. W.

    2004-12-01

    Arctic systems can be rich and diverse habitats for marine life in spite of the extreme cold environment. Benthic faunal populations and associated biogeochemical cycling processes are influenced by sea-ice extent, seawater hydrography (nutrients, salinity, temperature, currents), and water column production. Benthic organisms on the Arctic shelves and margins are long-term integrators of overlying water column processes. Because these organisms have adapted to living at cold extremes, it is reasonable to expect that these communities will be among the most susceptible to climate warming. Recent observations show that Arctic sea ice in the North American Arctic is melting and retreating northward earlier in the season and the timing of these events can have dramatic impacts on the biological system. Changes in overlying primary production, pelagic-benthic coupling, and benthic production and community structure can have cascading effects to higher trophic levels, particularly benthic feeders such as walruses, gray whales, and diving seaducks. Recent indicators of contemporary Arctic change in the northern Bering Sea include seawater warming and reduction in ice extent that coincide with our time-series studies of benthic clam population declines in the shallow northern Bering shelf in the 1990's. In addition, declines in benthic amphipod populations have also likely influenced the movement of feeding gray whales to areas north of Bering Strait during this same time period. Finally a potential consequence of seawater warming and reduced ice extent in the northern Bering Sea could be the northward movement of bottom feeding fish currently in the southern Bering Sea that prey on benthic fauna. This would increase the feeding pressure on the benthic prey base and enhance competition for this food source for benthic-feeding marine mammals and seabirds. This presentation will outline recent biological changes observed in the northern Bering Sea ecosystem as documented in

  5. Genetically distinct coelacanth population off the northern Tanzanian coast.

    PubMed

    Nikaido, Masato; Sasaki, Takeshi; Emerson, J J; Aibara, Mitsuto; Mzighani, Semvua I; Budeba, Yohana L; Ngatunga, Benjamin P; Iwata, Masamitsu; Abe, Yoshitaka; Li, Wen-Hsiung; Okada, Norihiro

    2011-11-01

    Since the sensational discovery of a living coelacanth off the east coast of South Africa, the geographic distribution of viable coelacanth populations has been a subject of debate. In the past, the coelacanths off the African mainland were thought to be strays from the Comoros because most coelacanths captured were caught in the waters surrounding the Comoros archipelagos. However, in recent years, a large number of coelacanths were captured off the coast of Tanzania, including nine living specimens observed in a remotely operated vehicles survey. Thus, it is possible that there is a reproducing population inhabiting waters off the Tanzania coast. We have sequenced the complete mitochondrial genomes of 21 Tanzanian and 2 Comoran coelacanths and analyzed these sequences together with two additional full mitochondrial genomes and 47 d-loop sequences from the literature. We found that the coelacanth population off the northern Tanzanian coast is genetically differentiated from those of the southern Tanzania coast and the Comoros, whereas no significant genetic differentiation occurs between the latter two localities. The differentiation between the northern and southern Tanzanian coast populations is consistent with the hypothesis that the existence of northward-flowing ocean current along the Tanzanian coast may reduce or prevent gene flow from the northern to the southern population. Finally, we estimated that the population localized to the southern Tanzanian coast and the Comoros diverged from other coelacanths at least 200,000 y ago. These results indicate that the coelacanths off the northern Tanzania coast are not strays but a genetically distinct group. Our study provides important information for the conservation of this threatened "living fossil."

  6. Decomposition in northern Minnesota peatlands

    SciTech Connect

    Farrish, K.W.

    1985-01-01

    Decomposition in peatlands was investigated in northern Minnesota. Four sites, an ombrotrophic raised bog, an ombrotrophic perched bog and two groundwater minerotrophic fens, were studied. Decomposition rates of peat and paper were estimated using mass-loss techniques. Environmental and substrate factors that were most likely to be responsible for limiting decomposition were monitored. Laboratory incubation experiments complemented the field work. Mass-loss over one year in one of the bogs, ranged from 11 percent in the upper 10 cm of hummocks to 1 percent at 60 to 100 cm depth in hollows. Regression analysis of the data for that bog predicted no mass-loss below 87 cm. Decomposition estimates on an area basis were 2720 and 6460 km/ha yr for the two bogs; 17,000 and 5900 kg/ha yr for the two fens. Environmental factors found to limit decomposition in these peatlands were reducing/anaerobic conditions below the water table and cool peat temperatures. Substrate factors found to limit decomposition were low pH, high content of resistant organics such as lignin, and shortages of available N and K. Greater groundwater influence was found to favor decomposition through raising the pH and perhaps by introducing limited amounts of dissolved oxygen.

  7. Northern Prairie Wildlife Research Center

    USGS Publications Warehouse

    ,

    2009-01-01

    The Northern Prairie Wildlife Research Center (NPWRC) conducts integrated research to fulfill the Department of the Interior's responsibilities to the Nation's natural resources. Located on 600 acres along the James River Valley near Jamestown, North Dakota, the NPWRC develops and disseminates scientific information needed to understand, conserve, and wisely manage the Nation's biological resources. Research emphasis is primarily on midcontinental plant and animal species and ecosystems of the United States. During the center's 40-year history, its scientists have earned an international reputation for leadership and expertise on the biology of waterfowl and grassland birds, wetland ecology and classification, mammalian behavior and ecology, grassland ecosystems, and application of statistics and geographic information systems. To address current science challenges, NPWRC scientists collaborate with researchers from other U.S. Geological Survey centers and disciplines (Biology, Geography, Geology, and Water) and with biologists and managers in the Department of the Interior (DOI), other Federal agencies, State agencies, universities, and nongovernmental organizations. Expanding upon its scientific expertise and leadership, the NPWRC is moving in new directions, including invasive plant species, restoration of native habitats, carbon sequestration and marketing, and ungulate management on DOI lands.

  8. Enantioselective Michael addition of water.

    PubMed

    Chen, Bi-Shuang; Resch, Verena; Otten, Linda G; Hanefeld, Ulf

    2015-02-09

    The enantioselective Michael addition using water as both nucleophile and solvent has to date proved beyond the ability of synthetic chemists. Herein, the direct, enantioselective Michael addition of water in water to prepare important β-hydroxy carbonyl compounds using whole cells of Rhodococcus strains is described. Good yields and excellent enantioselectivities were achieved with this method. Deuterium labeling studies demonstrate that a Michael hydratase catalyzes the water addition exclusively with anti-stereochemistry.

  9. Enantioselective Michael Addition of Water

    PubMed Central

    Chen, Bi-Shuang; Resch, Verena; Otten, Linda G; Hanefeld, Ulf

    2015-01-01

    The enantioselective Michael addition using water as both nucleophile and solvent has to date proved beyond the ability of synthetic chemists. Herein, the direct, enantioselective Michael addition of water in water to prepare important β-hydroxy carbonyl compounds using whole cells of Rhodococcus strains is described. Good yields and excellent enantioselectivities were achieved with this method. Deuterium labeling studies demonstrate that a Michael hydratase catalyzes the water addition exclusively with anti-stereochemistry. PMID:25529526

  10. LEECHES (ANNELIDA: EUHIRUDINEA) OF NORTHERN ARKANSAS

    EPA Science Inventory

    Twenty-one lotic and lentic environments throughout central and northern Arkansas were surveyed for the presence of leeches during June 2004, and April, July - October, 2005. Fourteen species of leeches (Desserobdella cryptobranchii, Desserobdella phalera, Desserobdella picta, H...

  11. Huge Filament Rises From Sun's Northern Hemisphere

    NASA Video Gallery

    On August 1, 2010 following a C3-class solar flare from sunspot 1092, an enormous magnetic filament stretching across the sun's northern hemisphere erupted. This 304 angstrom video shows that filam...

  12. Comparison of the efficiency of two commercial kits - ELFA and Western blot in estimating the phase of Toxoplasma gondii infection in pregnant women.

    PubMed

    Sroka, Jacek; Wójcik-Fatla, Angelina; Zając, Violetta; Sawczyn, Anna; Cisak, Ewa; Karamon, Jacek; Dutkiewicz, Jacek; Bojar, Iwona

    2016-12-23

    Sera of 89 pregnant women were selected according to the results of ELFA IgM, IgG and avidity IgG, and tested with commercial tests IgM, IgG and avidity IgG Western Blot (WB) to compare the efficacy of both techniques in determining the phase of T. gondii infection. In total, 81 of 89 tested sera (91.0%) were classified as positive, both in the ELFA and WB tests for the presence of anti-Toxoplasma antibodies of class IgG, indicating a past infection, while the prevalence of anti-Toxoplasma positive reactions associated with the antibodies of class IgM indicating a recent infection was much lower - 31.5% and 20.2%, respectively. Sera of 81 women were also tested in the ELFA and WB tests for avidity, e.g. ability of forming high-molecular IgG antibody complexes. Low or medium results in these tests (in this study all classified as low), indicating a recent infection, were detected by ELFA and WB in 22.2% and 45.7% of the total examined samples, respectively. The Spearman's rank test for correlation, performed for recognition of quantitative data of the ELFA and WB tests (index, units or points), revealed a highly significant correlation between the ELFA and WB tests for homologous classes of antibodies, both for IgM and IgG (p<0.00001). In contrast, the ELFA and WB tests for detection of anti-Toxoplasma IgM antibodies were not correlated with the ELFA and WB tests for detection of anti-Toxoplasma IgG antibodies (p>0.05), except for the WB test for IgM antibodies, which showed a significant correlation with the ELFA test for IgG antibodies (p<0.01). A highly significant negative correlation between the ELFA and WB test for IgM antibodies and ELFA and WB tests for IgG avidity was demonstrated (p<0.01), except for a relationship between the WB test for IgM and WB for avidity, which was not significant. Such negative correlations are theoretically expected, as strong complexes with the participation of IgG antibodies are absent in the early phase of toxoplasmosis when

  13. Long-term variability in Northern Hemisphere snow cover and associations with warmer winters

    USGS Publications Warehouse

    McCabe, Gregory J.; Wolock, David M.

    2010-01-01

    A monthly snow accumulation and melt model is used with gridded monthly temperature and precipitation data for the Northern Hemisphere to generate time series of March snow-covered area (SCA) for the period 1905 through 2002. The time series of estimated SCA for March is verified by comparison with previously published time series of SCA for the Northern Hemisphere. The time series of estimated Northern Hemisphere March SCA shows a substantial decrease since about 1970, and this decrease corresponds to an increase in mean winter Northern Hemisphere temperature. The increase in winter temperature has caused a decrease in the fraction of precipitation that occurs as snow and an increase in snowmelt for some parts of the Northern Hemisphere, particularly the mid-latitudes, thus reducing snow packs and March SCA. In addition, the increase in winter temperature and the decreases in SCA appear to be associated with a contraction of the circumpolar vortex and a poleward movement of storm tracks, resulting in decreased precipitation (and snow) in the low- to mid-latitudes and an increase in precipitation (and snow) in high latitudes. If Northern Hemisphere winter temperatures continue to warm as they have since the 1970s, then March SCA will likely continue to decrease.

  14. Long-term variability in Northern Hemisphere snow cover and associations with warmer winters

    USGS Publications Warehouse

    McCabe, G.J.; Wolock, D.M.

    2010-01-01

    A monthly snow accumulation and melt model is used with gridded monthly temperature and precipitation data for the Northern Hemisphere to generate time series of March snow-covered area (SCA) for the period 1905 through 2002. The time series of estimated SCA for March is verified by comparison with previously published time series of SCA for the Northern Hemisphere. The time series of estimated Northern Hemisphere March SCA shows a substantial decrease since about 1970, and this decrease corresponds to an increase in mean winter Northern Hemisphere temperature. The increase in winter temperature has caused a decrease in the fraction of precipitation that occurs as snow and an increase in snowmelt for some parts of the Northern Hemisphere, particularly the mid-latitudes, thus reducing snow packs and March SCA. In addition, the increase in winter temperature and the decreases in SCA appear to be associated with a contraction of the circumpolar vortex and a poleward movement of storm tracks, resulting in decreased precipitation (and snow) in the low- to mid-latitudes and an increase in precipitation (and snow) in high latitudes. If Northern Hemisphere winter temperatures continue to warm as they have since the 1970s, then March SCA will likely continue to decrease. ?? 2009 Springer Science+Business Media B.V.

  15. Relationship between vegetation coverage and spring dust storms over northern China

    NASA Astrophysics Data System (ADS)

    Zou, Xukai K.; Zhai, Panmao M.

    2004-02-01

    On the basis of normalized difference vegetation index (NDVI) data from 1982 to 2001 and dust storm observations in China the relationship between vegetation and spring dust storms over northern China is discussed. The results show that poor vegetation coverage in northern China is one important factor for the frequent occurrence of spring dust storms. In addition, vegetation cover plays an important role in interannual variations of dust storms. In general, a negative correlation is noted between vegetation coverage and occurrence of dust storms in northern China for spring during the period 1982-2001. The correlation coefficient between vegetation coverage and areas affected by dust storms is -0.59, which is statistically meaningful at 99% confidence level. The sharp decrease of spring vegetation coverage in recent years is one of the major contributors to frequent spring dust storms over northern China specifically during 2000 and 2001. A negative correlation is especially significant in the eastern part of northern China, mainly in central and eastern Inner Mongolia. When vegetation decreases (increases), the occurrence of dust storms increases (decreases). Furthermore, statistics show that abundant vegetation in previous seasons could help reduce dust storms in the coming spring. The effect of prior summer vegetation on the variation of spring dust storms is particularly evident in the central and eastern part of northern China. Because of the presence of little to no vegetation in the desert areas of northwest China the variation in occurrence of spring dust storms seems unrelated to the vegetation.

  16. Predation by northern squawfish on live and dead juvenile Chinook salmon

    USGS Publications Warehouse

    Gadomski, Dena M.; Hall-Griswold, Judy A.

    1992-01-01

    Northern squafish Ptychocheilus oregonensis is a major predator of juvenile salmonids Oncorhynchus spp. migrating downstream through the Columbia River. High predation rates occur just below dams. If northern squafish selectively consume salmonids killed or injured during dam passage, previous estimates of predation mortality may be too high. We conducted laboratory experiments that indicate northern squafish prefer dead juvenile chinook salmon O. tshawytscha over live individuals. When equal numbers of dead and live chinook salmon were offered to northern squafish maintained on a natural photoperiod (15 h light: 9 h darkness), significantly more (P < 0.05) dead than live fish were consumed, both in 1,400-L circular tanks and in an 11,300-L raceway (62% and 79% of prey consumed were dead, respectively). When dead and live juvenile chinook salmon were provided in proportions more similar to those below dams (20% dead, 80% live), northern squafish still selected for dead prey (36% of fish consumed were dead). In additional experiments, northern squafish were offered a proportion of 20% dead juvenile chinook salmon during 4-h periods of either light or darkness. The predators were much more selective for dead chinook salmon during bright light (88% of fish consumed were dead) than during darkness (31% were dead).

  17. Predation by northern squawfish on live and dead juvenile chinook salmon

    SciTech Connect

    Gadomski, D.M.; Hall-Griswold, J.A. )

    1992-09-01

    Northern squawfish Ptychocheilus oregonensis is a major predator of juvenile salmonids Oncorhynchus spp. migrating downstream through the Columbia River. High predation rates occur just below dams. If northern squawfish selectively consume salmonids killed or injured during dam passage, previous estimates of predation mortality may be too high. We conducted laboratory experiments that indicate northern squawfish prefer dead juvenile chinook salmon O. tshawytscha over live individuals. When equal numbers of dead and live chinook salmon were offered to northern squawfish maintained on a natural photoperiod (15 h light: 9 h darkness), significantly more (P < 0.05) dead than live fish were consumed, both in 1,400-L circular tanks and in an 11,300-L raceway (62% and 79% of prey consumed were dead, respectively). When dead and live juvenile chinook salmon were provided in proportions more similar to those below dams (20% dead, 80% live), northern squawfish still selected for dead prey (36% of fish consumed were dead). In additional experiments, northern squawfish were offered a proportion of 20% dead juvenile chinook salmon during 4-h periods of either light or darkness. The predators were much more selective for dead chinook salmon during bright light (88% of fish consumed were dead) than during darkness (31% were dead).

  18. A geographical cluster of progressive supranuclear palsy in northern France

    PubMed Central

    Golbe, Lawrence I.; Deramecourt, Vincent; Maurage, Claude-Alain; Huin, Vincent; Buée-Scherrer, Valerie; Obriot, Helene; Sablonnière, Bernard; Caparros, Francois; Buée, Luc; Lees, Andrew J.

    2015-01-01

    Objective: To describe a cluster of progressive supranuclear palsy (PSP) in northern France. PSP has not been reported in geographical, temporal, or occupational clusters. A unit of Neurology and Neurogeriatrics opened in 2005 at the Centre Hospitalier de Wattrelos, serving the population of Wattrelos and Leers (combined population 51,551) and parts of neighboring towns. For most of the 20th century, this area was a center for chromate and phosphate ore processing, textile dyeing, and tanning. Significant industrial waste persists close to residential areas. Methods: From 2005 to 2014, 92 patients with PSP at Centre Hospitalier de Wattrelos were identified and studied. Detailed residential data were available in the medical records. Eighty cases have had magnetic resonance head scanning and 60 have died, of whom 13 have been examined neuropathologically. Results: The ratio of observed to expected PSP incidence over the period 2005 to 2012 was 12.3 (95% confidence interval: 7.4–35.9). Mean onset age was 74.3 years. The Richardson syndrome/PSP-parkinsonism ratio was 43%/42%. Four other phenotypes each occurred in 2% to 5%. Onset was gait/balance difficulty in 52%. None of the 92 affected patients were relatives and 7 were of North African ancestry. MRI was compatible with a clinical diagnostic of PSP in all cases. Histopathologic examination confirmed neurofibrillary degeneration and tufted astrocytes in all autopsied cases. Western blots revealed a typical tau 4R doublet. The tau H1 haplotype occurred in 95.8% of cases' chromosomes. Conclusions: We have identified a cluster of PSP in a geographical area with severe environmental contamination by industrial metals. PMID:26354981

  19. Color Addition and Subtraction Apps

    ERIC Educational Resources Information Center

    Ruiz, Frances; Ruiz, Michael J.

    2015-01-01

    Color addition and subtraction apps in HTML5 have been developed for students as an online hands-on experience so that they can more easily master principles introduced through traditional classroom demonstrations. The evolution of the additive RGB color model is traced through the early IBM color adapters so that students can proceed step by step…

  20. Additive Effects on Asymmetric Catalysis.

    PubMed

    Hong, Liang; Sun, Wangsheng; Yang, Dongxu; Li, Guofeng; Wang, Rui

    2016-03-23

    This review highlights a number of additives that can be used to make asymmetric reactions perfect. Without changing other reaction conditions, simply adding additives can lead to improved asymmetric catalysis, such as reduced reaction time, improved yield, or/and increased selectivity.

  1. 40 CFR 81.354 - Northern Mariana Islands.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 40 Protection of Environment 18 2014-07-01 2014-07-01 false Northern Mariana Islands. 81.354... § 81.354 Northern Mariana Islands. Northern Mariana Islands—TSP Designated area Does not meet primary... 1 X 1 EPA designation only. Northern Mariana Islands—1971 Sulfur Dioxide NAAQS (Primary...

  2. 40 CFR 81.354 - Northern Mariana Islands.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 40 Protection of Environment 18 2012-07-01 2012-07-01 false Northern Mariana Islands. 81.354... § 81.354 Northern Mariana Islands. Northern Mariana Islands—TSP Designated area Does not meet primary... 1 X 1 EPA designation only. Northern Mariana Islands—SO2 Designated area Does not meet...

  3. 40 CFR 81.354 - Northern Mariana Islands.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 40 Protection of Environment 18 2013-07-01 2013-07-01 false Northern Mariana Islands. 81.354... § 81.354 Northern Mariana Islands. Northern Mariana Islands—TSP Designated area Does not meet primary... 1 X 1 EPA designation only. Northern Mariana Islands—SO2 Designated area Does not meet...

  4. 40 CFR 81.354 - Northern Mariana Islands.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 40 Protection of Environment 17 2010-07-01 2010-07-01 false Northern Mariana Islands. 81.354... § 81.354 Northern Mariana Islands. Northern Mariana Islands—TSP Designated area Does not meet primary... 1 X 1 EPA designation only. Northern Mariana Islands—SO2 Designated area Does not meet...

  5. 40 CFR 81.354 - Northern Mariana Islands.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 40 Protection of Environment 17 2011-07-01 2011-07-01 false Northern Mariana Islands. 81.354... § 81.354 Northern Mariana Islands. Northern Mariana Islands—TSP Designated area Does not meet primary... 1 X 1 EPA designation only. Northern Mariana Islands—SO2 Designated area Does not meet...

  6. Albian ammonites from northern Pakistan

    NASA Astrophysics Data System (ADS)

    Kennedy, William James; Fatmi, Ali N.

    2014-03-01

    The occurrence of rich Albian ammonite faunas in what is now northern Pakistan has been known for more than 80 years, but there has been no comprehensive account of the assemblages present. A total of 36 taxa are described below. The middle part of the Lumshiwal Formation yields Upper Aptian ammonites south of the Samana Range. Elsewhere, it yields Douvilleiceras leightonense Casey, 1962, of the lower Lower Albian Leymeriella regularis Zone and the Sonneratia perinflata and S. kitchini Subzones of the Sonneratia chalensis Zone of the northwest European sequence. The top one to two metres of the Lumshiwal yields an abundant fauna of rolled and phosphatised ammonites that includes elements from much of the Albian. Of these, Prolyelliceras gevreyi (Jacob, 1907) first appears in the lower Lower Albian Leymeriella tardefurcata Zone. The commonest ammonite is Douvilleiceras mammillatum (Schlotheim, 1813) sensu lato, which ranges from the perinflata Subzone of the chalensis Zone to the Otohoplites bulliensis Subzone of the O. auritiformis Zone of the Lower Albian. The presence of Lyelliceras pseudolyelli (Parona and Bonarelli, 1897) indicates the uppermost, pseudolyelli Subzone of the auritiformis Zone. The presence of Lyelliceras lyelli (d'Orbigny, 1841) indicates the basal Middle Albian lyelli Subzone of the Hoplites dentatus Zone. There is no evidence for the higher parts of the Middle Albian. Dipoloceras (Rhytidoceras ) sp. indicates the presence of lower Upper Albian, possibly the pricei Zone. There is evidence, in the form of specifically indeterminate Mortoniceras (Mortoniceras) sp., of a level within the inflatum to fallax Zone inteval from a single locality, but no evidence of the succeeding parts of the upper Upper Albian. The base of the Kawagarth Formation that succeeds the Lumshiwal yields lower Upper Albian Mortoniceras (M.) geometricum Spath, 1932 of the Mortoniceras pricei Zone, northwest of Darmasand in the Samana range.

  7. Multiple Landslide-Hazard Scenarios Modeled for the Oakland-Berkeley Area, Northern California

    USGS Publications Warehouse

    Pike, Richard J.; Graymer, Russell W.

    2008-01-01

    With the exception of Los Angeles, perhaps no urban area in the United States is more at risk from landsliding, triggered by either precipitation or earthquake, than the San Francisco Bay region of northern California. By January each year, seasonal winter storms usually bring moisture levels of San Francisco Bay region hillsides to the point of saturation, after which additional heavy rainfall may induce landslides of various types and levels of severity. In addition, movement at any time along one of several active faults in the area may generate an earthquake large enough to trigger landslides. The danger to life and property rises each year as local populations continue to expand and more hillsides are graded for development of residential housing and its supporting infrastructure. The chapters in the text consist of: *Introduction by Russell W. Graymer *Chapter 1 Rainfall Thresholds for Landslide Activity, San Francisco Bay Region, Northern California by Raymond C. Wilson *Chapter 2 Susceptibility to Deep-Seated Landsliding Modeled for the Oakland-Berkeley Area, Northern California by Richard J. Pike and Steven Sobieszczyk *Chapter 3 Susceptibility to Shallow Landsliding Modeled for the Oakland-Berkeley Area, Northern California by Kevin M. Schmidt and Steven Sobieszczyk *Chapter 4 Landslide Hazard Modeled for the Cities of Oakland, Piedmont, and Berkeley, Northern California, from a M=7.1 Scenario Earthquake on the Hayward Fault Zone by Scott B. Miles and David K. Keefer *Chapter 5 Synthesis of Landslide-Hazard Scenarios Modeled for the Oakland-Berkeley Area, Northern California by Richard J. Pike The plates consist of: *Plate 1 Susceptibility to Deep-Seated Landsliding Modeled for the Oakland-Berkeley Area, Northern California by Richard J. Pike, Russell W. Graymer, Sebastian Roberts, Naomi B. Kalman, and Steven Sobieszczyk *Plate 2 Susceptibility to Shallow Landsliding Modeled for the Oakland-Berkeley Area, Northern California by Kevin M. Schmidt and Steven

  8. Color Addition and Subtraction Apps

    NASA Astrophysics Data System (ADS)

    Ruiz, Frances; Ruiz, Michael J.

    2015-10-01

    Color addition and subtraction apps in HTML5 have been developed for students as an online hands-on experience so that they can more easily master principles introduced through traditional classroom demonstrations. The evolution of the additive RGB color model is traced through the early IBM color adapters so that students can proceed step by step in understanding mathematical representations of RGB color. Finally, color addition and subtraction are presented for the X11 colors from web design to illustrate yet another real-life application of color mixing.

  9. Adverse reactions to drug additives.

    PubMed

    Simon, R A

    1984-10-01

    There is a long list of additives used by the pharmaceutical industry. Most of the agents used have not been implicated in hypersensitivity reactions. Among those that have, only reactions to parabens and sulfites have been well established. Parabens have been shown to be responsible for rare immunoglobulin E-mediated reactions that occur after the use of local anesthetics. Sulfites, which are present in many drugs, including agents commonly used to treat asthma, have been shown to provoke severe asthmatic attacks in sensitive individuals. Recent studies indicate that additives do not play a significant role in "hyperactivity." The role of additives in urticaria is not well established and therefore the incidence of adverse reactions in this patient population is simply not known. In double-blind, placebo-controlled studies, reactions to tartrazine or additives other than sulfites, if they occur at all, are indeed quite rare for the asthmatic population, even for the aspirin-sensitive subpopulation.

  10. Radiation Therapy: Additional Treatment Options

    MedlinePlus

    ... SNIPEND SNIPSTART Find A Radiation Oncologist SNIPEND Additional Treatment Options SNIPSTART A A SNIPEND Chemotherapy Medicines prescribed ... such as antibodies, to fight cancer. Novel Targeted Therapies Cancer doctors now know much more about how ...

  11. Calculators and Computers: Graphical Addition.

    ERIC Educational Resources Information Center

    Spero, Samuel W.

    1978-01-01

    A computer program is presented that generates problem sets involving sketching graphs of trigonometric functions using graphical addition. The students use calculators to sketch the graphs and a computer solution is used to check it. (MP)

  12. Selliguea kachinensis (Polypodiaceae), a new fern species of uncertain affinity from Northern Myanmar

    PubMed Central

    Khine, Phyo Kay; Lindsay, Stuart; Fraser-Jenkins, Christopher; Kluge, Jürgen; Kyaw, Myint; Hovenkamp, Peter

    2016-01-01

    Abstract We describe Selliguea kachinensis as a new species from Northern Myanmar and discuss its generic placement in either Selliguea or Arthromeris. The conservation status is assessed as Data Deficient. In addition, we make the new combination Selliguea erythrocarpa (Mett. ex Kuhn) Hovenkamp, S. Linds., Fraser-Jenk. PMID:27212883

  13. Northern Polar Spring in IR

    NASA Technical Reports Server (NTRS)

    2004-01-01

    [figure removed for brevity, see original site]

    Released 12 March 2004

    The Odyssey spacecraft has completed a full Mars year of observations of the red planet. For the next several weeks the Image of the Day will look back over this first mars year. It will focus on four themes: 1) the poles - with the seasonal changes seen in the retreat and expansion of the caps; 2) craters - with a variety of morphologies relating to impact materials and later alteration, both infilling and exhumation; 3) channels - the clues to liquid surface flow; and 4) volcanic flow features. While some images have helped answer questions about the history of Mars, many have raised new questions that are still being investigated as Odyssey continues collecting data as it orbits Mars.

    Infrared images taken during the daytime exhibit both the morphological and thermophysical properties of the surface of Mars. Morphologic details are visible due to the effect of sun-facing slopes receiving more energy than antisun-facing slopes. This creates a warm (bright) slope and cool (dark) slope appearance that mimics the light and shadows of a visible wavelength image. Thermophysical properties are seen in that dust heats up more quickly than rocks. Thus dusty areas are bright and rocky areas are dark.

    This image was collected October 19, 2002 during the northern spring season. The top half of this daytime IR image shows the North Polar sand sea.

    Image information: IR instrument. Latitude 76.2, Longitude 226.8 East (133.2 West). 19 meter/pixel resolution.

    Note: this THEMIS visual image has not been radiometrically nor geometrically calibrated for this preliminary release. An empirical correction has been performed to remove instrumental effects. A linear shift has been applied in the cross-track and down-track direction to approximate spacecraft and planetary motion. Fully calibrated and geometrically projected images will be released through the Planetary Data System in

  14. Food additives and preschool children.

    PubMed

    Martyn, Danika M; McNulty, Breige A; Nugent, Anne P; Gibney, Michael J

    2013-02-01

    Food additives have been used throughout history to perform specific functions in foods. A comprehensive framework of legislation is in place within Europe to control the use of additives in the food supply and ensure they pose no risk to human health. Further to this, exposure assessments are regularly carried out to monitor population intakes and verify that intakes are not above acceptable levels (acceptable daily intakes). Young children may have a higher dietary exposure to chemicals than adults due to a combination of rapid growth rates and distinct food intake patterns. For this reason, exposure assessments are particularly important in this age group. The paper will review the use of additives and exposure assessment methods and examine factors that affect dietary exposure by young children. One of the most widely investigated unfavourable health effects associated with food additive intake in preschool-aged children are suggested adverse behavioural effects. Research that has examined this relationship has reported a variety of responses, with many noting an increase in hyperactivity as reported by parents but not when assessed using objective examiners. This review has examined the experimental approaches used in such studies and suggests that efforts are needed to standardise objective methods of measuring behaviour in preschool children. Further to this, a more holistic approach to examining food additive intakes by preschool children is advisable, where overall exposure is considered rather than focusing solely on behavioural effects and possibly examining intakes of food additives other than food colours.

  15. Phototaxis of larval and juvenile northern pike

    USGS Publications Warehouse

    Zigler, S.J.; Dewey, M.R.

    1995-01-01

    Age- Phi northern pike Esox lucius prefer vegetated habitats that are difficult to sample with standard towed gears. Light traps can be effective for sampling larval fishes in dense vegetation, given positive phototaxis of fish. We evaluated the phototactic response of young northern pike by comparing the catches of larvae and juveniles obtained with plexiglass traps deployed with a chemical light stick versus traps deployed without a light source (controls) in a laboratory raceway and in a vegetated pond. In the laboratory tests, catches of protolarvae and mesolarvae in lighted traps were 11-35 times greater than catches in control traps. The catches of juvenile northern pike in field and laboratory experiments were 3-15 times greater in lighted traps than in control traps, even though the maximum body width of the larger juveniles was similar to the width of the entrance slots of the traps (5 mm). Larval and juvenile northern pike were photopositive; thus, light traps should effectively sample age-0 northern pike for at least 6 weeks after hatching.

  16. Evaluation of certain food additives.

    PubMed

    2015-01-01

    This report represents the conclusions of a Joint FAO/WHO Expert Committee convened to evaluate the safety of various food additives, including flavouring agents, and to prepare specifications for identity and purity. The first part of the report contains a general discussion of the principles governing the toxicological evaluation of and assessment of dietary exposure to food additives, including flavouring agents. A summary follows of the Committee's evaluations of technical, toxicological and dietary exposure data for eight food additives (Benzoe tonkinensis; carrageenan; citric and fatty acid esters of glycerol; gardenia yellow; lutein esters from Tagetes erecta; octenyl succinic acid-modified gum arabic; octenyl succinic acid-modified starch; paprika extract; and pectin) and eight groups of flavouring agents (aliphatic and alicyclic hydrocarbons; aliphatic and aromatic ethers; ionones and structurally related substances; miscellaneous nitrogen-containing substances; monocyclic and bicyclic secondary alcohols, ketones and related esters; phenol and phenol derivatives; phenyl-substituted aliphatic alcohols and related aldehydes and esters; and sulfur-containing heterocyclic compounds). Specifications for the following food additives were revised: citric acid; gellan gum; polyoxyethylene (20) sorbitan monostearate; potassium aluminium silicate; and Quillaia extract (Type 2). Annexed to the report are tables summarizing the Committee's recommendations for dietary exposures to and toxicological evaluations of all of the food additives and flavouring agents considered at this meeting.

  17. [INVITED] Lasers in additive manufacturing

    NASA Astrophysics Data System (ADS)

    Pinkerton, Andrew J.

    2016-04-01

    Additive manufacturing is a topic of considerable ongoing interest, with forecasts predicting it to have major impact on industry in the future. This paper focusses on the current status and potential future development of the technology, with particular reference to the role of lasers within it. It begins by making clear the types and roles of lasers in the different categories of additive manufacturing. This is followed by concise reviews of the economic benefits and disadvantages of the technology, current state of the market and use of additive manufacturing in different industries. Details of these fields are referenced rather than expanded in detail. The paper continues, focusing on current indicators to the future of additive manufacturing. Barriers to its development, trends and opportunities in major industrial sectors, and wider opportunities for its development are covered. Evidence indicates that additive manufacturing may not become the dominant manufacturing technology in all industries, but represents an excellent opportunity for lasers to increase their influence in manufacturing as a whole.

  18. Additive manufacturing of hybrid circuits

    SciTech Connect

    Bell, Nelson S.; Sarobol, Pylin; Cook, Adam; Clem, Paul G.; Keicher, David M.; Hirschfeld, Deidre; Hall, Aaron Christopher

    2016-03-26

    There is a rising interest in developing functional electronics using additively manufactured components. Considerations in materials selection and pathways to forming hybrid circuits and devices must demonstrate useful electronic function; must enable integration; and must complement the complex shape, low cost, high volume, and high functionality of structural but generally electronically passive additively manufactured components. This article reviews several emerging technologies being used in industry and research/development to provide integration advantages of fabricating multilayer hybrid circuits or devices. First, we review a maskless, noncontact, direct write (DW) technology that excels in the deposition of metallic colloid inks for electrical interconnects. Second, we review a complementary technology, aerosol deposition (AD), which excels in the deposition of metallic and ceramic powder as consolidated, thick conformal coatings and is additionally patternable through masking. As a result, we show examples of hybrid circuits/devices integrated beyond 2-D planes, using combinations of DW or AD processes and conventional, established processes.

  19. Tougher Addition Polyimides Containing Siloxane

    NASA Technical Reports Server (NTRS)

    St. Clair, T. L.; Maudgal, S.

    1986-01-01

    Laminates show increased impact resistances and other desirable mechanical properties. Bismaleamic acid extended by reaction of diaminosiloxane with maleic anhydride in 1:1 molar ratio, followed by reaction with half this molar ratio of aromatic dianhydride. Bismaleamic acid also extended by reaction of diaminosiloxane with maleic anhydride in 1:2 molar ratio, followed by reaction with half this molar ratio of aromatic diamine (Michael-addition reaction). Impact resistances improved over those of unmodified bismaleimide, showing significant increase in toughness. Aromatic addition polyimides developed as both matrix and adhesive resins for applications on future aircraft and spacecraft.

  20. Stratigraphy of the Martian northern plains

    NASA Technical Reports Server (NTRS)

    Tanaka, K. L.

    1993-01-01

    The northern plains of Mars are roughly defined as the large continuous region of lowlands that lies below Martian datum, plus higher areas within the region that were built up by volcanism, sedimentation, tectonism, and impacts. These northern lowlands span about 50 x 10(exp 6) km(sup 2) or 35 percent of the planet's surface. The age and origin of the lowlands continue to be debated by proponents of impact and tectonic explanations. Geologic mapping and topical studies indicate that volcanic, fluvial, and eolian deposition have played major roles in the infilling of this vast depression. Periglacial, glacial, fluvial, eolian, tectonic, and impact processes have locally modified the surface. Because of the northern plains' complex history of sedimentation and modification, much of their stratigraphy was obscured. Thus the stratigraphy developed is necessarily vague and provisional: it is based on various clues from within the lowlands as well as from highland areas within and bordering the plains. The results are summarized.

  1. Identification of immunodominant VP1 linear epitope of enterovirus 71 (EV71) using synthetic peptides for detecting human anti-EV71 IgG antibodies in Western blots.

    PubMed

    Foo, D G W; Ang, R X; Alonso, S; Chow, V T K; Quak, S H; Poh, C L

    2008-03-01

    A major IgG-specific immunodominant VP1 linear epitope of enterovirus 71 (EV71) strain 41 (5865/SIN/00009), defined by the core sequence LEGTTNPNG, was identified by Pepscan analysis. Oligonucleotides corresponding to the amino-acid sequence of synthetic peptide SP32 were cloned and over-expressed in Escherichia coli as a recombinant glutathione-S-transferase (GST)-SP32 fusion protein. In ELISAs, this protein did not react with human anti-EV71 IgG antibodies, but there was significant immunoreactivity according to western blot analysis. The amino-acid sequence of SP32 was highly specific for detecting EV71 strains in western blot analysis, and showed no immunoreactivity with monoclonal antibodies raised against other enteroviruses, e.g., CA9 and Echo 6.

  2. The Additive Property of Energy.

    ERIC Educational Resources Information Center

    Tsaoussis, Dimitris S.

    1995-01-01

    Presents exercises that analyze the additive property of energy. Concludes that if a body has more than one component of energy depending on the same physical quantity, the body's total energy will be the algebraic sum of the components if a linear relationship exists between the energy components and that physical quantity. (JRH)

  3. Tetrasulfide extreme pressure lubricant additives

    SciTech Connect

    Gast, L.E.; Kenney, H.E.; Schwab, A.W.

    1980-08-19

    A novel class of compounds has been prepared comprising the tetrasulfides of /sup 18/C hydrocarbons, /sup 18/C fatty acids, and /sup 18/C fatty and alkyl and triglyceride esters. These tetrasulfides are useful as extreme pressure lubricant additives and show potential as replacements for sulfurized sperm whale oil.

  4. Out of bounds additive manufacturing

    SciTech Connect

    Holshouser, Chris; Newell, Clint; Palas, Sid; Love, Lonnie J.; Kunc, Vlastimil; Lind, Randall F.; Lloyd, Peter D.; Rowe, John C.; Blue, Craig A.; Duty, Chad E.; Peter, William H.; Dehoff, Ryan R.

    2013-03-01

    Lockheed Martin and Oak Ridge National Laboratory are working on an additive manufacturing system capable of manufacturing components measured not in terms of inches or feet, but multiple yards in all dimensions with the potential to manufacture parts that are completely unbounded in size.

  5. Climate impacts on northern Canada: regional background.

    PubMed

    Prowse, Terry D; Furgal, Chris; Bonsal, Barrie R; Peters, Daniel L

    2009-07-01

    Understanding the implications of climate change on northern Canada requires a background about the size and diversity of its human and biogeophysical systems. Occupying an area of almost 40% of Canada, with one-third of this contained in Arctic islands, Canada's northern territories consist of a diversity of physical environments unrivaled around the circumpolar north. Major ecozones composed of a range of landforms, climate, vegetation, and wildlife include: Arctic, boreal and taiga cordillera; boreal and taiga plains; taiga shield; and northern and southern Arctic. Although generally characterized by a cold climate, there is an enormous range in air temperature with mean annual values being as high as -5 degrees C in the south to as low as -20 degrees C in the high Arctic islands. A similar contrast characterizes precipitation, which can be > 700 mm y(-1) in some southern alpine regions to as low as 50 mm y(-1) over islands of the high Arctic. Major freshwater resources are found within most northern ecozones, varying from large glaciers or ice caps and lakes to extensive wetlands and peat lands. Most of the North's renewable water, however, is found within its major river networks and originates in more southerly headwaters. Ice covers characterize the freshwater systems for multiple months of the year while permafrost prevails in various forms, dominating the terrestrial landscape. The marine environment, which envelops the Canadian Arctic Archipelago, is dominated by seasonal to multiyear sea ice often several meters thick that plays a key role in the regional climate. Almost two-thirds of northern Canadian communities are located along coastlines with the entire population being just over 100 000. Most recent population growth has been dominated by an expansion of nonaboriginals, primarily the result of resource development and the growth of public administration. The economies of northern communities, however, remain quite mixed with traditional land

  6. Geochemistry and 40Ar/39Ar geochronology of the ophiolite in Northern Xinjiang

    NASA Astrophysics Data System (ADS)

    Qinqin, Xu; Jianqing, Ji

    2010-05-01

    As the remnant of ancient oceanic crust, ophiolite is extremely important to reconstruct the framework of ancient ocean and continent. There are several ophiolite belts in Northern Xinjiang, assuming planar distribution, and they are generally related to different faults since Paleozoic in the outcrops. This paper presents a systematic study of geochemistry and geochronology of the gabbro, diabase and basalt of ophiolite collected from different regions in Northern Xinjiang. The intermediate and basic rocks in ophiolite show similar patterns in primitive mantle-normalized REE and trace elements diagrams with MORB and different from IAT, indicating that the ophiolite may be formed in mid-ocean ridge. Their Sr, Nd and Pb isotopic compositions also show that they have mid-ocean ridge environment affinity. Additionally, their Sr, Nd and Pb isotopic ratios are similar respectively, suggesting the same source area, and they also have positive epsilon Nd values(4.40 ~ 8.04), which indicates they likely originate from the depleted mantle. The previous and the author's researches show that the forming time of ophiolite in Northern Xinjiang is early Paleozoic, but 40Ar/39Ar ages of gabbro, diabase and basalt are in the range of 396 ~ 226Ma (mainly from 350 to 250Ma), and no ages of early Paleozoic are gained. Moreover, this time coincides with the timing of late Paleozoic post-collisional plutonism, indicating the ophiolite in Northern Xinjiang was reworked by the late thermal events. The ophiolite widely exposed in Northern Xinjiang have similar characteristics of occurrence, lithologic association and isotopes in spite of diverse special features, which indicates that a relatively uniform and integrated source region has existed in Northern Xinjiang since Paleozoic. It is likely to infer that this source region is related with the long lasting remnant oceanic basin and the related lithosphere since Paleozoic in North Xinjiang. Key Words: Ophiolite, 40Ar/39Ar age

  7. ABCGheritage project - promoting geotourism in northern Finland, northern Norway and the Kola Peninsula

    NASA Astrophysics Data System (ADS)

    Pihlaja, Jouni; Johansson, Peter; Lauri, Laura S.

    2014-05-01

    Nature tourism has been a growing business sector in the Barents area during the recent decades. With the purpose to develop nature tourism in a sustainable way, a cooperation project ABCGheritage - Arctic Biological, Cultural and Geological Heritage has been carried out. Project has received partial funding from the EU Kolarctic ENPI program. In the geoheritage part of the project the main activities were aimed to develop pro-environmental ways of geotourism in the area. The three main participants in the geoheritage part of the project are the Geological Survey of Finland, Northern Finland Office, the Geological Institute of the Kola Science Centre of the Russian Academy of Sciences and Bioforsk Soil and Environment from northeastern Norway. The duration of the project is 2012-2014 and most of the work has already been completed even if most of the results are not published yet. Totally ten different tasks have been implemented in the geological part of the project. The largest task has been the preparation of a geological outdoor map and guide book of the Khibiny Tundra locating in the central part of the Kola Peninsula. In Finland already 11 such maps have been published, and the experiences gained during their production have been used in this project, too. Geological heritage trails to the Khibiny Tundra have also been created and they will be drawn on the map. The second concrete result is the Barents Tour for Geotourist -guide, which will be published as a guide book, web pages and an exhibition. The route comprises ca 35 best geological demonstration sites along the circle route from northern Finland to northeastern Norway, from there to Kola Peninsula and then back to Finland. Information of the route will be available for all interested travelers. In addition to the geological outdoor map of the Khibiny Tundra and "Barents Tour for Geotourists"-guide, the primary outputs of the project are the geological nature trails on the field, geological

  8. Evaluation of certain food additives.

    PubMed

    2012-01-01

    This report represents the conclusions of a Joint FAO/WHO Expert Committee convened to evaluate the safety of various food additives, including flavouring agents, with a view to concluding as to safety concerns and to preparing specifications for identity and purity. The first part of the report contains a general discussion of the principles governing the toxicological evaluation of and assessment of dietary exposure to food additives, including flavouring agents. A summary follows of the Committee's evaluations of technical, toxicological and dietary exposure data for five food additives (magnesium dihydrogen diphosphate; mineral oil (medium and low viscosity) classes II and III; 3-phytase from Aspergillus niger expressed in Aspergillus niger; serine protease (chymotrypsin) from Nocardiopsis prasina expressed in Bacillus licheniformis; and serine protease (trypsin) from Fusarium oxysporum expressed in Fusarium venenatum) and 16 groups of flavouring agents (aliphatic and aromatic amines and amides; aliphatic and aromatic ethers; aliphatic hydrocarbons, alcohols, aldehydes, ketones, carboxylic acids and related esters, sulfides, disulfides and ethers containing furan substitution; aliphatic linear alpha,beta-unsaturated aldehydes, acids and related alcohols, acetals and esters; amino acids and related substances; epoxides; furfuryl alcohol and related substances; linear and branched-chain aliphatic, unsaturated, unconjugated alcohols, aldehydes, acids and related esters; miscellaneous nitrogen-containing substances; phenol and phenol derivatives; pyrazine derivatives; pyridine, pyrrole and quinoline derivatives; saturated aliphatic acyclic branched-chain primary alcohols, aldehydes and acids; simple aliphatic and aromatic sulfides and thiols; sulfur-containing heterocyclic compounds; and sulfur-substituted furan derivatives). Specifications for the following food additives were revised: ethyl cellulose, mineral oil (medium viscosity), modified starches and titanium

  9. Rural telemedicine project in northern New Mexico

    SciTech Connect

    Zink, S.; Hahn, H.; Rudnick, J.; Snell, J.; Forslund, D.; Martinez, P.

    1998-12-31

    A virtual electronic medical record system is being deployed over the Internet with security in northern New Mexico using TeleMed, a multimedia medical records management system that uses CORBA-based client-server technology and distributed database architecture. The goal of the NNM Rural Telemedicine Project is to implement TeleMed into fifteen rural clinics and two hospitals within a 25,000 square mile area of northern New Mexico. Evaluation of the project consists of three components: job task analysis, audit of immunized children, and time motion studies. Preliminary results of the evaluation components are presented.

  10. Petroleum resource potential GIS of northern Afghanistan

    USGS Publications Warehouse

    Steinshouer, Douglas W.; Klett, Timothy R.; Ulmishek, Gregory F.; Wandrey, Craig J.; Wahl, Ronald R.; Hill, Ronald J.; Pribil, Michael J.; Pawlewicz, Mark J.; King, J. David; Agena, Warren F.; Taylor, David J.; Amirzada, Abdulla; Selab, Amir Mohammad; Mutteh, Abdul-Salam; Haidari, Ghulam Naqshband; Wardak, Moeengul Gullabudeen

    2006-01-01

    The CD-ROM contains an ESRI ArcReader format GIS project presenting the results of a petroleum resource assessment of Northern Afghanistan, and other data used in the petroleum assessment. Geologic, structural, field, well, political, and other GIS layers covering Afghanistan, Northern Afghanistan and adjacent areas, along with associated geochemical and other data tables pertinent to a petroleum assessment are included. The purpose of this GIS is to provide the basic data layers and tables required to support the petroleum assessment, data for further exploration and development, and an index of known data.

  11. Coastal geomorphology of the Martian northern plains

    NASA Technical Reports Server (NTRS)

    Parker, Timothy J.; Gorsline, Donn S.; Saunders, Stephen R.; Pieri, David C.; Schneeberger, Dale M.

    1993-01-01

    The paper considers the question of the formation of the outflow channels and valley networks discovered on the Martian northern plains during the Mariner 9 mission. Parker and Saunders (1987) and Parker et al. (1987, 1989) data are used to describe key features common both in the lower reaches of the outflow channels and within and along the margins of the entire northern plains. It is suggested, that of the geological processes capable of producing similar morphologies on earth, lacustrine or marine deposition and subsequent periglacial modification offer the simplest and most consistent explanation for the suit of features found on Mars.

  12. Geoenvironmental Investigations of the Humboldt River Basin, Northern Nevada

    USGS Publications Warehouse

    Stillings, Lisa L.

    2003-01-01

    Northern Nevada is one of the world's foremost regions of gold production. The Humboldt River Basin (HRB) covers 43,500 km2 in northern Nevada (Crompton, 1995), and it is home to approximately 18 active gold and silver mines (Driesner and Coyner, 2001) among at least 55 significant metallic mineral deposits (Long and others, 1998). Many of the gold mines are along the Carlin trend in the east-central portion of the HRB, and together they have produced 50 million ounces of gold from 1962 (when the Carlin mine first opened) through April 2002 (Nevada Mining Association, 2002). Mining is not new to the region, however. Beginning in 1849, mining has taken place in numerous districts that cover 39 percent of the land area in the HRB (Tingley, 1998). In addition to gold and silver, As, Ba, Cu, Fe, Hg, Li, Mn, Mo, Pb, S, Sb, V, W, Zn, and industrial commodities such as barite, limestone, fluorite, sand and gravel, gypsum, gemstones, pumice, zeolites, and building stone, have been extracted from the HRB (McFaul and others, 2000). All papers within this series of investigations can be found as lettered chapters of USGS Bulletin 2210, Geoenvironmental Investigations of the Humboldt River Basin, Northern Nevada. Each chapter is available separately online. The data and software utilized in this product (Chapter F) permit the user to view and analyze the geographic relationships among chemistry of stream sediments and surface waters, geology, and various cartographic base information such as but not limited to cities, county boundaries, and land ownership. Data for this product were compiled and or produced as part of a mineral and environmental assessment of the Humboldt River basin conducted by the U.S. Geological Survey between 1995 - 2000.

  13. Analysis of the serologic relationship among San Miguel sea lion virus and vesicular exanthema of swine virus isolates. Application of the western blot assay for detection of antibodies in swine sera to these virus types.

    PubMed

    Seal, B S; House, J A; Whetstone, C A; Neill, J D

    1995-04-01

    Caliciviruses are positive-sense single-stranded RNA viruses with a single capsid protein. The serotypes of the marine mammal calicivirus, San Miguel sea lion virus (SMSV), are antigenically related to vesicular exanthema of swine virus (VESV) and are potentially hazardous to swine. Western blot assays using purified SMSV serotypes 1 and 4 were used to further examine the serologic relationship among SMSV and VESV isolates. With the exception of SMSV 8 and SMSV 12, rabbit polyclonal antisera generated against all the available SMSV and VESV isolates reacted positively, as assessed by western blot, with purified capsid protein from SMSV 1 and SMSV 4. Consequently, the SMSV 8 and SMSV 12 virus isolates may not be members of the SMSV/VESV calicivirus group. Using antisera from pigs experimentally inoculated with SMSV and VESV as positive controls, a western blot assay for these virus types was utilized to check for the presence of antibodies to calciviruses in swine sera. Sera from colostrum-deprived gnotobiotic pigs were used as a negative control in all experiments. Examination of sera from domestic and feral swine collected in Iowa, California, and Florida was completed using this technique. The presence of antibodies to these virus types was not detected in any of the porcine sera tested.

  14. The Influence of European Pollution on Ozone in the Near East and Northern Africa

    NASA Technical Reports Server (NTRS)

    Duncan, B. N.; West, J. J.; Yoshida, Y.; Fiore, A. M.; Ziemke, J. R.

    2008-01-01

    We present a modeling study of the long-range transport of pollution from Europe, showing that European emissions regularly elevate surface ozone by as much as 20 ppbv in summer in northern Africa and the Near East. European emissions cause 50-150 additional violations per year (i.e. above those that would occur without European pollution) of the European health standard for ozone (8-h average greater than 120 micrograms per cubic meters or approximately 60 ppbv) in northern Africa and the Near East. We estimate that European ozone pollution is responsible for 50 000 premature mortalities globally each year, of which the majority occurs outside of Europe itself, including 37% (19 000) in northern Africa and the Near East. Much of the pollution from Europe is exported southward at low altitudes in summer to the Mediterranean Sea, northern Africa and the Near East, regions with favorable photochemical environments for ozone production. Our results suggest that assessments of the human health benefits of reducing ozone precursor emissions in Europe should include effects outside of Europe, and that comprehensive planning to improve air quality in northern Africa and the Near East likely needs to address European emissions.

  15. Genetics Show Current Decline and Pleistocene Expansion in Northern Spotted Owls

    USGS Publications Warehouse

    Funk, W. Chris; Forsman, Eric D.; Mullins, Thomas D.; Haig, Susan M.

    2008-01-01

    The northern spotted owl (Strix occidentalis caurina) is one of the most controversial threatened subspecies ever listed under the U.S. Endangered Species Act. Because of concern for persistence of the subspecies, logging on Federal lands in the U.S. Pacific Northwest was dramatically reduced under the Northwest Forest Plan in 1994. Despite protection of its remaining forest habitat, recent field studies show continued demographic declines of northern spotted owls. One potential threat to northern spotted owls that has not yet been shown is loss of genetic variation from population bottlenecks that can increase inbreeding depression and decrease adaptive potential. Here, we show recent genetic bottlenecks in northern spotted owls using a large genetic dataset (352 individuals from across the subspecies' range and 11 microsatellite loci). The signature of bottlenecks was strongest in Washington State, in agreement with field data. Interestingly, we also found a genetic signature of Pleistocene expansion in the same study areas where recent bottlenecks were shown. Our results provide independent evidence that northern spotted owls have recently declined, and suggest that loss of genetic variation is an emerging threat to the subspecies' persistence. Reduced effective population size (Ne), shown here in addition to field evidence for demographic decline, highlights the increasing vulnerability of this bird to extinction.

  16. Cardiovascular risk factors in hypertensive patients born in Northern Africa and living in France.

    PubMed

    Lepoutre-Lussey, Charlotte; Plouin, Pierre-François; Steichen, Olivier

    2010-04-01

    Northern Africans constitute one of the largest immigrant groups in France; however, limited data are available on their cardiovascular risk factors. We carried out a retrospective cross-sectional study in a French hypertension unit, comparing 719 patients born in Northern Africa to 3558 controls born in Europe, individually matched for age and sex. Using a Bonferroni adjusted alpha-level=0.001, we found no significant difference between the groups for blood pressure levels, anti-hypertensive treatment, prevalence of target organ damage or the proportion of patients with secondary hypertension. However, patients of both sexes born in Northern Africa were less likely to take regular physical exercise than their controls. In addition, women born in Northern Africa were less often current or former smokers than their European counterparts (19.9% vs 30.5%, p < 0.001), but had a higher body mass index (28.5 vs 26.8 kg/m(2), p < 0.001) and a higher prevalence of diabetes (19.1% vs 8.9%, p < 0.001 after adjusting for BMI). These results suggest that targeted lifestyle interventions, including regular physical exercise, could be proposed to prevent weight gain and decrease the incidence of diabetes in hypertensive women born in Northern Africa and living in western countries.

  17. The Frontiers of Additive Manufacturing

    SciTech Connect

    Grote, Christopher John

    2016-03-03

    Additive manufacturing, more commonly known as 3-D printing, has become a ubiquitous tool in science for its precise control over mechanical design. For additive manufacturing to work, a 3-D structure is split into thin 2D slices, and then different physical properties, such as photo-polymerization or melting, are used to grow the sequential layers. The level of control allows not only for devices to be made with a variety of materials: e.g. plastics, metals, and quantum dots, but to also have finely controlled structures leading to other novel properties. While 3-D printing is widely used by hobbyists for making models, it also has industrial applications in structural engineering, biological tissue scaffolding, customized electric circuitry, fuel cells, security, and more.

  18. Decontamination formulation with sorbent additive

    DOEpatents

    Tucker; Mark D. , Comstock; Robert H.

    2007-10-16

    A decontamination formulation and method of making that neutralizes the adverse health effects of both chemical and biological compounds, especially chemical warfare (CW) and biological warfare (BW) agents, and toxic industrial chemicals. The formulation provides solubilizing compounds that serve to effectively render the chemical and biological compounds, particularly CW and BW compounds, susceptible to attack, and at least one reactive compound that serves to attack (and detoxify or kill) the compound. The formulation includes at least one solubilizing agent, a reactive compound, a bleaching activator, a sorbent additive, and water. The highly adsorbent, water-soluble sorbent additive (e.g., sorbitol or mannitol) is used to "dry out" one or more liquid ingredients, such as the liquid bleaching activator (e.g., propylene glycol diacetate or glycerol diacetate) and convert the activator into a dry, free-flowing powder that has an extended shelf life, and is more convenient to handle and mix in the field.

  19. Additive Manufacturing Integrated Energy Demonstration

    SciTech Connect

    Jackson, Roderick; Lee, Brian; Love, Lonnie; Mabe, Gavin; Keller, Martin; Curran, Scott; Chinthavali, Madhu; Green, Johney; Sawyer, Karma; Enquist, Phil

    2016-02-05

    Meet AMIE - the Additive Manufacturing Integrated Energy demonstration project. Led by Oak Ridge National Laboratory and many industry partners, the AMIE project changes the way we think about generating, storing, and using electrical power. AMIE uses an integrated energy system that shares energy between a building and a vehicle. And, utilizing advanced manufacturing and rapid innovation, it only took one year from concept to launch.

  20. Robust stability under additive perturbations

    NASA Technical Reports Server (NTRS)

    Bhaya, A.; Desoer, C. A.

    1985-01-01

    A MIMO linear time-invariant feedback system 1S(P,C) is considered which is assumed to be U-stable. The plant P is subjected to an additive perturbation Delta P which is proper but not necessarily stable. It is proved that the perturbed system is U-stable if and only if Delta P(I + Q x Delta P) exp -1 is U-stable.

  1. Additive Manufacturing Integrated Energy Demonstration

    ScienceCinema

    Jackson, Roderick; Lee, Brian; Love, Lonnie; Mabe, Gavin; Keller, Martin; Curran, Scott; Chinthavali, Madhu; Green, Johney; Sawyer, Karma; Enquist, Phil

    2016-07-12

    Meet AMIE - the Additive Manufacturing Integrated Energy demonstration project. Led by Oak Ridge National Laboratory and many industry partners, the AMIE project changes the way we think about generating, storing, and using electrical power. AMIE uses an integrated energy system that shares energy between a building and a vehicle. And, utilizing advanced manufacturing and rapid innovation, it only took one year from concept to launch.

  2. Nanoengineered Additives for Active Coatings

    DTIC Science & Technology

    2007-04-01

    commercial ad bial activ component from the coating, leading to eventual depletion of the film. Small TPU samples were evaluated using a Kirby - Bauer ...7 Table 5. Summary of 24-hr ASTM E 2180 tests with 1 weight-percent additive in PUr (solvent dispersible) based on 6-log loading of...Noveon X-1150). The ASTM E 2180 test is run in triplicate (Note that alternative ro 1° amines) was suspended in dry tetrahydrofuran (THF) (150 mL) in

  3. Reversible Oxidative Addition at Carbon.

    PubMed

    Eichhorn, Antonius F; Fuchs, Sonja; Flock, Marco; Marder, Todd B; Radius, Udo

    2017-04-07

    The reactivity of N-heterocyclic carbenes (NHCs) and cyclic alkyl amino carbenes (cAACs) with arylboronate esters is reported. The reaction with NHCs leads to the reversible formation of thermally stable Lewis acid/base adducts Ar-B(OR)2 ⋅NHC (Add1-Add6). Addition of cAAC(Me) to the catecholboronate esters 4-R-C6 H4 -Bcat (R=Me, OMe) also afforded the adducts 4-R-C6 H4 Bcat⋅cAAC(Me) (Add7, R=Me and Add8, R=OMe), which react further at room temperature to give the cAAC(Me) ring-expanded products RER1 and RER2. The boronate esters Ar-B(OR)2 of pinacol, neopentylglycol, and ethyleneglycol react with cAAC at RT via reversible B-C oxidative addition to the carbene carbon atom to afford cAAC(Me) (B{OR}2 )(Ar) (BCA1-BCA6). NMR studies of cAAC(Me) (Bneop)(4-Me-C6 H4 ) (BCA4) demonstrate the reversible nature of this oxidative addition process.

  4. Additive manufacturing of hybrid circuits

    DOE PAGES

    Bell, Nelson S.; Sarobol, Pylin; Cook, Adam; ...

    2016-03-26

    There is a rising interest in developing functional electronics using additively manufactured components. Considerations in materials selection and pathways to forming hybrid circuits and devices must demonstrate useful electronic function; must enable integration; and must complement the complex shape, low cost, high volume, and high functionality of structural but generally electronically passive additively manufactured components. This article reviews several emerging technologies being used in industry and research/development to provide integration advantages of fabricating multilayer hybrid circuits or devices. First, we review a maskless, noncontact, direct write (DW) technology that excels in the deposition of metallic colloid inks for electrical interconnects.more » Second, we review a complementary technology, aerosol deposition (AD), which excels in the deposition of metallic and ceramic powder as consolidated, thick conformal coatings and is additionally patternable through masking. As a result, we show examples of hybrid circuits/devices integrated beyond 2-D planes, using combinations of DW or AD processes and conventional, established processes.« less

  5. Lithostratigraphical analysis in northern Sinus Meridiani, Mars.

    NASA Astrophysics Data System (ADS)

    Baioni, D.; Sgavetti, M.; Tampella, G.; Wezel, F. C.

    2012-04-01

    In the Terra Meridiani- Arabia Terra region of Mars, layered rock units with a range of stratal patterns have been widely reported in the literature, and the record of the spectral signatures indicative of hydrate minerals suggests the existence of aqueous activity in at least some of the stratigraphic units. . In this work we investigated the area in the northern region of Sinus Meridiani located between 2°N to 3°N latitude and 2°W to 1°E longitude, using multiple remote sensing data sets to identify, characterize in detail, and understand the origin and evolution of the selected units in this region of Mars. MOLA and HiRISE data were used in combination to classify and correlate surface units recorded in distinct stratigraphic logs, based on planimetric configuration, topography, morphological and litho-facies characteristics. In particular, detailed analyses were focused on the layered units discovered in the walls of two impact craters located at a distance of about 100 km to each other within the study area. The stratigraphic interval analysed here corresponds to the upper part of the well known "light-toned layered deposits" identified by several authors. Distinct competent layers are observed in the internal slopes of both craters. Our observations indicate that the lithostratigraphic sections can be divided in three units. The lower unit consists of very thick bodies of light-toned bouldery breccia deposits. In the HiRISE images these white-coloured beds are composed of irregularly-shaped, white nodules which are either isolated or coalescent, distributed between bedding planes. The middle unit mainly consists of alternating thin strong layers and thicker sequences of relatively weak layers, horizontally bedded. The strong, competent layers maintain steeper erosional profiles and play a major role in controlling the overall shape and geomorphology of the wall slopes. The upper unit can be traced laterally in the surrounding ground level, displaying mesa

  6. Subduction zone guided waves in Northern Chile

    NASA Astrophysics Data System (ADS)

    Garth, Thomas; Rietbrock, Andreas

    2016-04-01

    Guided wave dispersion is observed in subduction zones as high frequency energy is retained and delayed by low velocity structure in the subducting slab, while lower frequency energy is able to travel at the faster velocities associated with the surrounding mantle material. As subduction zone guided waves spend longer interacting with the low velocity structure of the slab than any other seismic phase, they have a unique capability to resolve these low velocity structures. In Northern Chile, guided wave arrivals are clearly observed on two stations in the Chilean fore-arc on permanent stations of the IPOC network. High frequency (> 5 Hz) P-wave arrivals are delayed by approximately 2 seconds compared to the low frequency (< 2 Hz) P-wave arrivals. Full waveform finite difference modelling is used to test the low velocity slab structure that cause this P-wave dispersion. The synthetic waveforms produced by these models are compared to the recorded waveforms. Spectrograms are used to compare the relative arrival times of different frequencies, while the velocity spectra is used to constrain the relative amplitude of the arrivals. Constraining the waveform in these two ways means that the full waveform is also matched, and the low pass filtered observed and synthetic waveforms can be compared. A combined misfit between synthetic and observed waveforms is then calculated following Garth & Rietbrock (2014). Based on this misfit criterion we constrain the velocity model by using a grid search approach. Modelling the guided wave arrivals suggest that the observed dispersion cannot be solely accounted for by a single low velocity layer as suggested by previous guided wave studies. Including dipping low velocity normal fault structures in the synthetic model not only accounts for the observed strong P-wave coda, but also produces a clear first motion dispersion. We therefore propose that the lithospheric mantle of the subducting Nazca plate is highly hydrated at intermediate

  7. Hydrocarbon Induced Seismicity in Northern Netherlands

    NASA Astrophysics Data System (ADS)

    Dost, B.; Spetzler, J.; Kraaijpoel, D.; Caccavale, M.

    2015-12-01

    The northern Netherlands has been regarded aseismic until the first earthquakes started in 1986, after more than 25 years of gas production from the one of the largest on-shore gas-fields in the World, the Groningen field, and accompanying smaller gas fields. Due to the shallow sources, at approximately 3 km depth, even small magnitude events cause considerable damage to buildings in the region. Since the largest recorded event in the Groningen field in 2012 with ML= 3,6, more than 30.000 damage claims were received by the mining company. Since 1995 a seismic monitoring network is operational in the region, consisting of 8 200m deep boreholes with 4 levels of 3C 4,5 Hz geophones. The network was designed for a location threshold of ML=1,5 over a 40x 80 km region. Average station separation was 20 km. At the end of 2014, 245 events have been recorded with ML ≥ 1,5, out of a total of 1100. Since 2003 a new mining law is in place in the Netherlands, which requires for each gas field in production a seismic risk analysis. Initially, due to the small number of events for specific fields, a general hazard (PSHA) was calculated for all gas-fields and a maximum magnitude was estimated at ML = 3,9. Since 2003 an increase in the activity rate is observed for the Groningen field, leading to the development of new models and a re-assessment of parameters like the maximum magnitude. More recently these models are extended to seismic risk, where also the fragility of the regional buildings is taken into account. Understanding the earthquake process is essential in taking mitigation measures. Continued research is focused on reducing the uncertainties in the hazard and risk models and is accompanied by an upgrade of the monitoring network. In 2014 a new dense network was designed to monitor the Groningen gas field in this region (30*40 km) with an average separation of 4 km. This allows an improved location threshold (M>0,5) and location accuracy (50-100m). A detailed P- and S

  8. Fire-Retardant Polymeric Additives

    NASA Technical Reports Server (NTRS)

    Williams, Martha K.; Smith, Trent M.

    2011-01-01

    Polyhydroxyamide (PHA) and polymethoxyamide (PMeOA) are fire-retardant (FR) thermoplastic polymers and have been found to be useful as an additive for imparting fire retardant properties to other compatible, thermoplastic polymers (including some elastomers). Examples of compatible flammable polymers include nylons, polyesters, and acrylics. Unlike most prior additives, PHA and PMeOA do not appreciably degrade the mechanical properties of the matrix polymer; indeed, in some cases, mechanical properties are enhanced. Also, unlike some prior additives, PHA and PMeOA do not decompose into large amounts of corrosive or toxic compounds during combustion and can be processed at elevated temperatures. PMeOA derivative formulations were synthesized and used as an FR additive in the fabrication of polyamide (PA) and polystyrene (PS) composites with notable reduction (>30 percent for PS) in peak heat release rates compared to the neat polymer as measured by a Cone Calorimeter (ASTM E1354). Synergistic effects were noted with nanosilica composites. These nanosilica composites had more than 50-percent reduction in peak heat release rates. In a typical application, a flammable thermoplastic, thermoplastic blend, or elastomer that one seeks to render flame-retardant is first dry-mixed with PHA or PMeOA or derivative thereof. The proportion of PHA or PMeOA or derivative in the mixture is typically chosen to lie between 1 and 20 weight percent. The dry blend can then be melt-extruded. The extruded polymer blend can further be extruded and/or molded into fibers, pipes, or any other of a variety of objects that may be required to be fire-retardant. The physical and chemical mechanisms which impart flame retardancy of the additive include inhibiting free-radical oxidation in the vapor phase, preventing vaporization of fuel (the polymer), and cooling through the formation of chemical bonds in either the vapor or the condensed phase. Under thermal stress, the cyclic hydroxyl/ methoxy

  9. Evaluation of certain food additives.

    PubMed

    2009-01-01

    This report represents the conclusions of a Joint FAO/WHO Expert Committee convened to evaluate the safety of various food additives, including flavouring agents, with a view to recommending acceptable daily intakes (ADIs) and to preparing specifications for identity and purity. The first part of the report contains a general discussion of the principles governing the toxicological evaluation and assessment of intake of food additives (in particular, flavouring agents). A summary follows of the Committee's evaluations of technical, toxicological and intake data for certain food additives (asparaginase from Aspergillus niger expressed in A. niger, calcium lignosulfonate (40-65), ethyl lauroyl arginate, paprika extract, phospholipase C expressed in Pichia pastoris, phytosterols, phytostanols and their esters, polydimethylsiloxane, steviol glycosides and sulfites [assessment of dietary exposure]) and 10 groups of related flavouring agents (aliphatic branched-chain saturated and unsaturated alcohols, aldehydes, acids and related esters; aliphatic linear alpha,beta-unsaturated aldehydes, acids and related alcohols, acetals and esters; aliphatic secondary alcohols, ketones and related esters; alkoxy-substituted allylbenzenes present in foods and essential oils and used as flavouring agents; esters of aliphatic acyclic primary alcohols with aliphatic linear saturated carboxylic acids; furan-substituted aliphatic hydrocarbons, alcohols, aldehydes, ketones, carboxylic acids and related esters, sulfides, disulfides and ethers; miscellaneous nitrogen-containing substances; monocyclic and bicyclic secondary alcohols, ketones and related esters; hydroxy- and alkoxy-substituted benzyl derivatives; and substances structurally related to menthol). Specifications for the following food additives were revised: canthaxanthin; carob bean gum and carob bean gum (clarified); chlorophyllin copper complexes, sodium and potassium salts; Fast Green FCF; guar gum and guar gum (clarified

  10. Male-associated polypeptide (MAP) expression in different compartments of the reproductive system of the mussel Mytilus galloprovincialis: immunocytochemical and western blot study.

    PubMed

    Torrado, M; Mikhailov, A T

    1998-12-01

    Mytilus mussels are characterized by annually repeated reproduction which is associated with subsequent growth, morphogenesis, breakdown and redevelopment of the gonad and reproductive tract into mantle mesenchyme. We present a description of the expression of the male-associated polypeptide (MAP; see Mikhailov et al. 1995) in different compartments of the male reproductive system as well as in mantle gonad-supporting tissue. MAP is expressed in both gonad and mantle structures in dynamic patterns that show a substantial overlap in terms of dependence on the stage of gonad development/involution. In general, the total MAP concentration directly correlates with the volume of gonad tubule/duct structures but inversely correlates with mantle connective tissue cell fraction. A maximum of MAP expression is reached in the fully ripe male gonad. MAP is localized around gonad tubules/ducts, in the gonoduct epithelium, membranes of follicle-like structures as well as in the extracellular fiber-like structures of the mantle. However, we also demonstrate unique sites of MAP accumulation in the lumen of gonad follicle-like tubules and in ductal fluid. The latter is characterized by a very high MAP concentration. MAP is also detected in sperm-containing cell suspension obtained by gonad biopsy which we interpret as a result of the adsorption of MAP on mature spermatozoa. The results obtained should be taken into consideration in the interpretation of possible MAP functions since they seem to point to MAP as a major component of ductal (seminal) fluid of the male reproductive tract. It is likely that MAP is able to complement the processes of sperm terminal differentiation and maturation. In addition, we demonstrate that the male-predominant character of MAP expression is restricted by gonad-containing tissues (i.e., mantle and visceral mass) only, although the polypeptide is also detected in other somatic organs in both males and females.

  11. The Divorced Chicana of Northern New Mexico.

    ERIC Educational Resources Information Center

    Fuller, Mary Lou; Casaus, Luis

    County demographic records and the voluntary responses of 80 Chicanas to a questionnaire and a self-concept examination were analyzed to identify the patterns, problems, and dynamics in the lives of divorced Northern New Mexico Chicanas. The women, half of whom were divorced and most of whom were Roman Catholics, were from urban Bernalillo County…

  12. United Kingdom (Northern Ireland): Health system review.

    PubMed

    O'Neill, Ciaran; McGregor, Pat; Merkur, Sherry

    2012-01-01

    The political context within which Northern Irelands integrated health and social care system operates has changed since the establishment of a devolved administration (the Northern Ireland Assembly, set up in 1998 but suspended between 2002 and 2007). A locally elected Health Minister now leads the publicly financed system and has considerable power to set policy and, in principle, to determine the operation of other health and social care bodies. The system underwent major reform following the passing of the Health and Social Care (Reform) Act (Northern Ireland) in 2009. The reform maintained the quasi purchaser provider split already in place but reduced the number and increased the size of many of the bodies involved in purchasing (known locally as commissioning) and delivering services. Government policy has generally placed greater emphasis on consultation and cooperation among health and social care bodies (including the department, commissioners and care providers) than on competition. The small size of the population (1.8 million) and Northern Irelands geographical isolation from the rest of the United Kingdom provide a rationale for eschewing a more competitive model. Without competition, effective control over the system requires information and transparency to ensure provider challenge, and a body outside the system to hold it to account. The restoration of the locally elected Assembly in 2007 has created such a body, but it remains to be seen how effectively it will exercise accountability.

  13. Equality in Higher Education in Northern Ireland

    ERIC Educational Resources Information Center

    Osborne, R.D.

    2005-01-01

    The higher education sector in Northern Ireland has been fully involved in the public policies designed to enhance equality. Starting with measures designed to secure greater employment between Catholics and Protestants, known as fair employment, the policies are now designed to promote equality of opportunity across nine designated groups…

  14. A Grammar of Northern and Southern Gumuz

    ERIC Educational Resources Information Center

    Ahland, Colleen Anne

    2012-01-01

    Gumuz is a Nilo-Saharan dialect cluster spoken in the river valleys of northwestern Ethiopia and the southeastern part of the Republic of the Sudan. There are approximately 200,000 speakers, the majority of which reside in Ethiopia. This study is a phonological and grammatical analysis of two main dialects/languages: Northern Gumuz and Southern…

  15. Nutritional condition of Northern Yellowstone Elk

    USGS Publications Warehouse

    Cook, R.C.; Cook, J.G.; Mech, L.D.

    2004-01-01

    Ultrasonography and body condition scoring was used to estimate nutritional condition of northern Yellowstone elk in late winter. Probability of pregnancy was related to body fat, and lactating cows had 50% less fat than non-lactating cows. For mild to normal winters, most of the elk were in good condition.

  16. Religion, Education and Conflict in Northern Ireland

    ERIC Educational Resources Information Center

    Barnes, L. Philip

    2005-01-01

    The aim of this article is to re-evaluate and reaffirm the contribution of the churches and of Christianity to the realization in Northern Ireland schools of legitimate and progressive educational values such as the cultivation of tolerance, moral integrity and civic virtue. Implicit in this is a critique of educational initiatives that seek to…

  17. The Northern Ireland Resource File and Aspire

    ERIC Educational Resources Information Center

    Ryan, David; Montgomery, Brenda

    2016-01-01

    The paper begins by identifying issues as to how initial teacher training and in-service training for teachers inadequately prepares them for teaching the pupils with special educational needs (SEN). The paper then provides a brief legislative background to SEN in the Northern Ireland context, before describing two elements of educational reform…

  18. From Poetry to Music: "Northern Lullaby"

    ERIC Educational Resources Information Center

    Cardany, Audrey Berger

    2011-01-01

    Nancy White Carlstrom's children's book, "Northern Lullaby," conjures through poetry the beauty of the Alaskan landscape in the evening. The book provides an opportunity for music teachers to help their students transform text and visual images to music. The author describes connections for reading comprehension in the general music…

  19. Variation and Change in Northern Bavarian Quantity

    ERIC Educational Resources Information Center

    Drake, Derek

    2013-01-01

    This dissertation presents new research on the "Bavarian Quantity Law" (the BQL) in the northern Bavarian dialect of Hahnbach. Building upon earlier investigation of the BQL (cf. Bannert 1976a,b for Central Bavarian) this study examines the historical, phonological, and phonetic motivations for this feature as well the variability in its…

  20. VHF radar measurements over Andoya (Northern Norway)

    NASA Technical Reports Server (NTRS)

    Czechowsky, P.; Reid, I. M.; Ruester, R.; Schmidt, G.

    1989-01-01

    The Mobile SOUSY Radar was operated during the MAP/WINE, the MAC/SINE, and MAC/Epsilon campaigns at Andoya in Northern Norway. A comparison between summer and winter results is presented, in particular the generation and development of the scattering regions, the different power spectral densities and the aspect sensitivities which were derived from six different beam directions.

  1. Winter in Northern Europe (WINE) Project

    NASA Technical Reports Server (NTRS)

    Vonzahn, U.

    1982-01-01

    The scientific aims, work plan, and organization of the Middle Atmosphere Program winter in northern Europe (MAP/WINE) are described. Proposed contributions to the MAP/WINE program from various countries are enumerated. Specific atmospheric parameters to be examined are listed along with the corresponding measurement technique.

  2. Seasonal prevailing surface winds in Northern Serbia

    NASA Astrophysics Data System (ADS)

    Tošić, Ivana; Gavrilov, Milivoj B.; Marković, Slobodan B.; Ruman, Albert; Putniković, Suzana

    2017-02-01

    Seasonal prevailing surface winds are analyzed in the territory of Northern Serbia, using observational data from 12 meteorological stations over several decades. In accordance with the general definition of prevailing wind, two special definitions of this term are used. The seasonal wind roses in 16 directions at each station are analyzed. This study shows that the prevailing winds in Northern Serbia have northwestern and southeastern directions. Circulation weather types over Serbia are presented in order to determine the connections between the synoptic circulations and prevailing surface winds. Three controlling pressure centers, i.e., the Mediterranean cyclone, Siberian high, and the Azores anticyclone, appear as the most important large-scale factors that influence the creation of the prevailing winds over Northern Serbia. Beside the synoptic cause of the prevailing winds, it is noted that the orography of the eastern Balkans has a major influence on the winds from the second quadrant. It was found that the frequencies of circulation weather types are in agreement with those of the prevailing winds over Northern Serbia.

  3. Gas hydrate resources of northern Alaska

    USGS Publications Warehouse

    Collett, T.S.

    1997-01-01

    Large amounts of natural gas, composed mainly of methane, can occur in arctic sedimentary basins in the form of gas hydrates under appropriate temperature and pressure conditions. Gas hydrates are solids, composed of rigid cages of water molecules that trap molecules of gas. These substances are regarded as a potential unconventional source of natural gas because of their enormous gas-storage capacity. Most published gas hydrate resource estimates are highly simplified and based on limited geological data. The gas hydrate resource assessment for northern Alaska presented in this paper is based on a "play analysis" scheme, in which geological factors controlling the accumulation and preservation of gas hydrates are individually evaluated and risked for each hydrate play. This resource assessment identified two gas hydrate plays; the in-place gas resources within the gas hydrates of northern Alaska are estimated to range from 6.7 to 66.8 trillion cubic metres of gas (236 to 2,357 trillion cubic feet of gas), at the 0.50 and 0.05 probability levels respectively. The mean in-place hydrate resource estimate for northern Alaska is calculated to be 16.7 trillion cubic metres of gas (590 trillion cubic feet of gas). If this assessment is valid, the amount of natural gas stored as gas hydrates in northern Alaska could be almost seven times larger then the estimated total remaining recoverable conventional natural gas resources in the entire United States.

  4. Racism and Citizenship Education in Northern Ireland

    ERIC Educational Resources Information Center

    Gallagher, Eamonn

    2007-01-01

    Racist attitudes towards, and attacks on, the minority ethnic populations in Northern Ireland (NI) have increased dramatically over the last number of years. Despite the increased media attention regarding racist attacks, the fallacy that racism is not a major problem in NI is an enduring one. However, there is a growing recognition that minority…

  5. Northern Russian "People" Games for Children

    ERIC Educational Resources Information Center

    Krasil'nikov, Valeriy P.

    2006-01-01

    In this article, the author states that people games and contests have always occupied a special place in the traditional physical training of the native people in northern Russia. This article is prepared on the basis of materials obtained as a result of expeditions conducted from 1990 through 1993 into Siberia's "Khanty-Mansyyski" and…

  6. Governance and Aboriginal Claims in Northern Canada.

    ERIC Educational Resources Information Center

    Cozzetto, Don

    1990-01-01

    Focuses on problems of organization and governance that may follow settlement of Canadian aboriginal land claims. Compares financial problems, cultural issues such as subsistence lifestyles, and intergovernmental relations following the Alaska Native Claims Settlement Act, James Bay and Northern Quebec Agreement, and Western Arctic (Inuvialuit)…

  7. Carbon dioxide emissions from estuaries of northern and northeastern Brazil.

    PubMed

    Noriega, Carlos; Araujo, Moacyr

    2014-08-22

    The carbon dioxide flux through the air-water interface of coastal estuarine systems must be quantified to understand the regional balance of carbon and its transport through adjacent coastal regions. We estimated and calculated the emissions of carbon dioxide (FCO₂) and the partial pressure of CO₂ (pCO₂) values in 28 estuarine environments at a variety of spatial scales in the northern and northeastern regions of Brazil. The results showed a mean FCO₂ (water to air) of 55 ± 45 mmol·m(-2)·d(-1). Additionally, a negative correlation between dissolved oxygen saturation and pCO₂ was observed, indicating a control by biological processes and especially by organic matter degradation. This leads to increased dissolved CO₂ concentration in estuarine waters which results in a pCO₂ that reached 8,638 μatm. Our study suggests that northern and northeastern Brazilian estuaries act as sources of atmospheric CO₂. The range of pCO₂ observed were similar to those found in inner estuaries in other places around the world, with the exception of a few semi-arid estuaries (Köppen climate classification - BSh) in which record low levels of pCO₂ have been detected.

  8. Incidence of ovine abortion by Coxiella burnetii in northern Spain.

    PubMed

    Oporto, B; Barandika, J F; Hurtado, A; Aduriz, G; Moreno, B; Garcia-Perez, A L

    2006-10-01

    The infectious causes of ovine abortion occurring in 148 farms in northern Spain between 1999 and 2003 were investigated. Laboratory analysis included microbiological, serological, pathological and molecular techniques. Border disease was diagnosed in 16% of the flocks, toxoplasmosis in 15%, chlamydiosis in 12%, salmonellosis in 10%, Q fever in 3%, miscellaneous infections in 7% (Yersinia spp., Listeria spp., Brucella spp.), and inflammatory lesions compatible with an infectious cause were seen in 7% of the flock. In an additional 1% of the flocks non-infectious causes were identified, and a diagnosis was not reached in 38% of the flocks. When a PCR retrospective study was carried out to investigate the possible implication of Coxiella burnetii in the cases without diagnosis, including those with inflammatory lesions, the prevalence of this pathogen increased from 3% up to 9% of the flocks, revealing the importance of this zoonotic pathogen as a small-ruminant abortifacient agent. Placenta was the most commonly positive sample, but other fetal tissues were also of value for C. burnetii DNA detection. The present results update information about the situation of abortion in sheep farms in northern Spain, and highlight the relevance of molecular diagnostic tools in routine laboratory analysis of abortions by C. burnetii.

  9. Tracking rainfall in the northern Mediterranean borderlands during sapropel deposition

    NASA Astrophysics Data System (ADS)

    Toucanne, Samuel; Angue Minto'o, Charlie Morelle; Fontanier, Christophe; Bassetti, Maria-Angela; Jorry, Stephan J.; Jouet, Gwenaël

    2015-12-01

    The role of mid-latitude precipitation in the hydrological forcing leading to the deposition of sapropels in the Mediterranean Sea remains unclear. The new GDEC-4-2 borehole, East Corsica margin (northern Tyrrhenian Sea), provides the first precisely dated evidence for enhanced rainfall in the Western Mediterranean during warm intervals of interglacial periods over the last 547 kyr. Comparison of GDEC-4-2 proxy records with pollen sequences and speleothems from the central and eastern Mediterranean reveals that these pluvial events were regional in character and occurred probably in response to the intensification of the Mediterranean storm track along the northern Mediterranean borderlands in autumn/winter. Our dataset suggests that the timing of maxima of the Mediterranean autumn/winter storm track precipitation coincide with that of the North African summer monsoon and sapropel deposition. Besides highlighting a close coupling between mid- and low-latitude hydrological changes, our findings suggest that during warm intervals of interglacial periods the reduced sea-surface water salinities, together with the high flux of nutrient and organic matter, produced by the monsoonal Nile (and wadi-systems) floods, were maintained throughout the winter by the Mediterranean rainfall. This provides an important additional constraint on the hydrological perturbation causing sapropel formation.

  10. Landscape Features Shape Genetic Structure in Threatened Northern Spotted Owls

    USGS Publications Warehouse

    Funk, W. Chris; Forsman, Eric D.; Mullins, Thomas D.; Haig, Susan M.

    2008-01-01

    Several recent studies have shown that landscape features can strongly affect spatial patterns of gene flow and genetic variation. Understanding landscape effects on genetic variation is important in conservation for defining management units and understanding movement patterns. The landscape may have little effect on gene flow, however, in highly mobile species such as birds. We tested for genetic breaks associated with landscape features in the northern spotted owl (Strix occidentalis caurina), a threatened subspecies associated with old forests in the U.S. Pacific Northwest and extreme southwestern Canada. We found little evidence for distinct genetic breaks in northern spotted owls using a large microsatellite dataset (352 individuals from across the subspecies' range genotyped at 10 loci). Nonetheless, dry low-elevation valleys and the Cascade and Olympic Mountains restrict gene flow, while the Oregon Coast Range facilitates it. The wide Columbia River is not a barrier to gene flow. In addition, inter-individual genetic distance and latitude were negatively related, likely reflecting northward colonization following Pleistocene glacial recession. Our study shows that landscape features may play an important role in shaping patterns of genetic variation in highly vagile taxa such as birds.

  11. Carbon, nitrogen, and phosphorus mineralization in northern wetlands

    SciTech Connect

    Bridgham, S.D.; Updegraff, K.; Pastor, J.

    1998-07-01

    The authors examined rates of C, N, and P mineralization in soils from 16 northern Minnesota wetlands that occur across an ombrotrophic-minerotrophic gradient. Soils were incubated at 30 C under aerobic and anaerobic conditions for 59 wk, and the results were fit with a two-pool kinetic model. Additionally, 39 different soil quality variables were used in a principal components analysis (PCA) to predict mineralization rates. Mineralization of C, N, and P differed significantly among wetland types, aeration status (aerobic vs. anaerobic), and their interaction term. Despite low total soil N and P, there was a rapid turnover of the nutrient pools in ombrotrophic sites, particularly under aerobic conditions. On a volumetric basis, C and N mineralization increased in a predictable manner across the ombrotrophic-minerotrophic gradient, largely due to increasing soil bulk density. However, P mineralization per cubic centimeter remained relatively high in the bogs. The higher total P content of more minerotrophic soils appears to be offset by greater P immobilization due to geochemical sorption, yielding overall lower availability. Large differences in mineralization rates in northern wetland communities demonstrate that climate change models should not consider these areas as homogeneous entities.

  12. Jurassic extension and Alpine inversion of the northern Morocco

    SciTech Connect

    Zizi, M. )

    1993-09-01

    The lower Mesozoic half grabens of northern Morocco form part of an extensional system that is related to the opening of the western Tethys. They appear to be somewhat younger than the Triassic-Jurassic systems associated with the opening the Atlantic Ocean. During the Tertiary and as consequence of the Alpine collision of Africa with Europe, these half graben systems were inverted as shown by the High and the Middle Atlas mountains. Seismic illustrations of similar but smaller inversion structures are available from the Guercif area and the [open quotes]Rides Prerifaines[close quotes] of northern Morocco. These seismic profiles serve as small models for the much larger Atlas Mountains. In the Guercif area, the inversions are limited in scope, but in the [open quotes]Ride Prerifaines[close quotes] are extensive decollement systems that sole out in the Triassic evaporites. These systems evolve into complex thrust faults and associated lateral ramps that are strongly influenced by the configuration of the Jurassic transtensional systems. Significant hydrocarbon accumulation have been known for some time from the [open quotes]Rides Prerifaines.[close quotes] A review of the geometry of the inverted half-graben systems, combined with detailed stratigraphic studies, is likely to lead to the discovery of additional reserves in the area.

  13. Potential cretaceous play in the Rharb basin of northern Morocco

    SciTech Connect

    Jobidon, G.P. )

    1993-09-01

    The autochthonous Cretaceous in the Rharb basin of northern Morocco is located underneath a cover of neogene sediments and of the Prerif nappe olistostrome, which was emplaced during the Tortonian 7 m.y. The presence of infranappe Cretaceous sediments is documented in a few onshore wells in the Rharb basin and in the adjacent Prerif Rides area, as well as in the Rif Mountains. Their presence in the deeper portion of the Rharb basin is difficult to detail because of poor seismic resolution data beneath dispersive prerif nappe. A recent study of offshore seismic data acquired by PCIAC in 1987 indicates that the infranappe interval can be more than 1500 m thick in some of the offshore Kenitra area. These sediments have seismic signatures that would correspond to Middle Cretaceous transgressions, culminating with a Turonian highstand. Their deposition systems were located on the northern and western flanks of the Meseta and were followed by a hiatus lasting until the Miocene. Regional studies of gravity and magnetic data provide and additional understanding of the Rif province, its evolution, and the possible presence of autochthonous Cretaceous sediments below the prerif nappe cover. The infranappe of Rharb basin has a good potential to develop into a major hydrocarbon play with the presence of middle Cretaceous reservoir rocks, Turonian-Cenomanian black shale source rocks, as well as the timely combination of trap formation, source rock maturation, and hydrocarbon migration.

  14. Carbon dioxide emissions from estuaries of northern and northeastern Brazil

    PubMed Central

    Noriega, Carlos; Araujo, Moacyr

    2014-01-01

    The carbon dioxide flux through the air–water interface of coastal estuarine systems must be quantified to understand the regional balance of carbon and its transport through adjacent coastal regions. We estimated and calculated the emissions of carbon dioxide (FCO2) and the partial pressure of CO2 (pCO2) values in 28 estuarine environments at a variety of spatial scales in the northern and northeastern regions of Brazil. The results showed a mean FCO2 (water to air) of 55 ± 45 mmol·m−2·d−1. Additionally, a negative correlation between dissolved oxygen saturation and pCO2 was observed, indicating a control by biological processes and especially by organic matter degradation. This leads to increased dissolved CO2 concentration in estuarine waters which results in a pCO2 that reached 8,638 μatm. Our study suggests that northern and northeastern Brazilian estuaries act as sources of atmospheric CO2. The range of pCO2 observed were similar to those found in inner estuaries in other places around the world, with the exception of a few semi-arid estuaries (Köppen climate classification – BSh) in which record low levels of pCO2 have been detected. PMID:25145418

  15. 68. Photocopy of drawing (from print, Burlington Northern Engineering Office) ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    68. Photocopy of drawing (from print, Burlington Northern Engineering Office) STRESS DIAGRAM - Burlington Northern Railroad Bridge, Spanning Willamette River at River Mile 6.9, Portland, Multnomah County, OR

  16. 64. Photocopy of drawing (from print, Burlington Northern Engineering Office, ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    64. Photocopy of drawing (from print, Burlington Northern Engineering Office, Seattle) STRESS SHEET - Burlington Northern Railroad Bridge, Spanning Willamette River at River Mile 6.9, Portland, Multnomah County, OR

  17. 49. Photocopy of photograph (from polaroid snapshot in Burlington Northern ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    49. Photocopy of photograph (from polaroid snapshot in Burlington Northern Railroad correspondence files, 1957) PIER III DAMAGE FROM M/V KORSHOLMA COLLISION - Burlington Northern Railroad Bridge, Spanning Willamette River at River Mile 6.9, Portland, Multnomah County, OR

  18. 47. Photocopy of photograph (from polaroid snapshot in Burlington Northern ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    47. Photocopy of photograph (from polaroid snapshot in Burlington Northern Railroad correspondence files, October, 1957) SHEAR FENCE DAMAGE FROM M/V KORSHOLMA - Burlington Northern Railroad Bridge, Spanning Willamette River at River Mile 6.9, Portland, Multnomah County, OR

  19. Aerial view of reroofing of northern LTA ship hangar, circa ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    Aerial view of re-roofing of northern LTA ship hangar, circa 1957. - Marine Corps Air Station Tustin, Northern Lighter Than Air Ship Hangar, Meffett Avenue & Maxfield Street, Tustin, Orange County, CA

  20. The Mozart Effect: Additional Data.

    PubMed

    Hughes, John R.

    2002-04-01

    After the review of the Mozart effect was published in this journal (Hughes JR. Epilepsy Behav 2001;2:369-417), additional data from the music of Haydn and Liszt have been analyzed that may account for the decrease in seizure activity originally reported during Mozart music. Even with these added data Mozart music continued to score significantly higher than the selections from the other six composers in one of the important characteristics of this music, namely, the repetition of the melody. However Haydn's values were second highest among Mozart, J. S. Bach, Wagner, Beethoven, Chopin, and Liszt.

  1. Water based drilling mud additive

    SciTech Connect

    McCrary, J.L.

    1983-12-13

    A water based fluid additive useful in drilling mud used during drilling of an oil or gas well is disclosed, produced by reacting water at temperatures between 210/sup 0/-280/sup 0/ F. with a mixture comprising in percent by weight: gilsonite 25-30%, tannin 7-15%, lignite 25-35%, sulfonating compound 15-25%, water soluble base compound 5-15%, methylene-yielding compound 1-5%, and then removing substantially all of the remaining water to produce a dried product.

  2. Metal Additive Manufacturing: A Review

    NASA Astrophysics Data System (ADS)

    Frazier, William E.

    2014-06-01

    This paper reviews the state-of-the-art of an important, rapidly emerging, manufacturing technology that is alternatively called additive manufacturing (AM), direct digital manufacturing, free form fabrication, or 3D printing, etc. A broad contextual overview of metallic AM is provided. AM has the potential to revolutionize the global parts manufacturing and logistics landscape. It enables distributed manufacturing and the productions of parts-on-demand while offering the potential to reduce cost, energy consumption, and carbon footprint. This paper explores the material science, processes, and business consideration associated with achieving these performance gains. It is concluded that a paradigm shift is required in order to fully exploit AM potential.

  3. Theatre fleet's vital additional capacity.

    PubMed

    2012-11-01

    Vanguard Healthcare's fleet of mobile surgical facilities has been deployed to healthcare sites throughout Europe and beyond for over a decade, providing vital additional clinical capacity when existing buildings are refurbished or upgraded, in the event of flood or fire, or simply to help hospitals cater for rising demand. It is a combination of careful planning, teamwork, and the specialist expertise of Vanguard's personnel--many with a clinical background--that ensures not only each unit's successful installation, but equally its subsequent running, servicing, and maintenance, the company explains.

  4. Shale JP-4 Additive Evaluation

    DTIC Science & Technology

    1986-10-01

    8217. •% . , ’ ,,,r ,% . -- - ,.-. ’ ’ 4,w% %’. " - ,’ . . . * ’, .* . TABLE OF CONTENTS .4q ,4 . * SECTION PAGE I. INTRODUCTION 1 II. TEST PARAMETERS 2 1...42 PRECEDING PAGE BLANK TABLE OF CONTENTS (CON’T) SECT ION PAGE V. CONCLUSIONS 44 REFERENCES 46 APPENDIX A Drum to Test Sample Relationship 47 APPENDIX...B.O.C.L.E. Results 40 vii LIST OF TABLES TABLE PAGE 1 Antioxidants 3 2 Raw Shale/Petroleum Fuel Properties 10 3 Drum Sample Additive Content 13 4

  5. Caribbean basin framework, 2: Northern Central America

    SciTech Connect

    Tyburski, S.A.; Gordon, M.B.; Mann, P. )

    1991-03-01

    There are four Jurassic to Recent basin-forming periods in northern Central America (honduras, Honduran Borderlands, Belize, Guatemala, northern Nicaragua): (1) Middle Jurassic-Early Cretaceous rifting and subsidence along normal faults in Honduras and Guatemala; rifts are suggested but are not well defined in Honduras by the distribution of clastic sediments and associated volcanic rocks. Rifting is attributed to the separation of Central America from the southern margin of the North American plate; (2) Cretaceous subsidence recorded by the development of a Cretaceous carbonate platform in Honduras, Guatemala, and Belize; subsidence is attributed to thermal subsidence of the rifted margins of the various blocks; (3) Late Cretaceous-Recent development of a volcanic arc along the western margin of Middle America and the northern margin of Honduras; (4) Late Cretaceous large-scale folding in Honduras, ophiolite obduction, and formation of a foredeep basin in Guatemala (Sepur trough); deformation is attributed to the collision between a north-facing arc in northern Honduras and the Nicaraguan Rise and the passive margin of Guatemala and Belize; and (5) Eocene to Recent strike-slip faulting along the present-day North American-Caribbean plate boundary in Guatemala, northern Honduras, and Belize. Strike-slip faults and basins form a California-type borderlands characterized by elongate basins that appear as half-grabens in profile. Counterclockwise rotation of the central honduras plateau, a thicker and topographically higher-than-average block within the plate boundary zone, is accommodated by rifting or strike-slip faults at its edges.

  6. AmeriFlux CA-Man Manitoba - Northern Old Black Spruce (former BOREAS Northern Study Area)

    SciTech Connect

    Amiro, Brian

    2016-01-01

    This is the AmeriFlux version of the carbon flux data for the site CA-Man Manitoba - Northern Old Black Spruce (former BOREAS Northern Study Area). Site Description - 55.880° N, 98.481° W, elevation of 259 m, Boreal coniferous: Black spruce; occasional larch present in poorly-drained areas. Groundcover is moss (feathermosses and Sphagnum), Labrador Tea, Vaccinium, and willows are a main component of the understory. It was established in 1993 as a BOREAS site.

  7. High Flow Addition Curing Polyimides

    NASA Technical Reports Server (NTRS)

    Chuang, Kathy C.; Vannucci, Raymond D.; Ansari, Irfan; Cerny, Lawrence L.; Scheiman, Daniel A.

    1994-01-01

    A new series of high flow PMR-type addition curing polyimides was developed, which employed the substitution of 2,2'-bis (trifluoromethyl) -4,4'-diaminobiphenyl (BTDB) for p-phenylenediamine (p -PDA) in a PMR-IL formulation. These thermoset polyimides, designated as 12F resins, were prepared from BTDB and the dimethyl ester of 4,4'- (hexafluo- roisopropylidene) -diphthalic acid (HFDE) with either nadic ester (NE) or p-aminostyrene (PAS) as the endcaps for addition curing. The 12F prepolymers displayed lower melting temperatures in DSC analysis, and higher melt flow in rheological studies than the cor- responding PMR-11 polyimides. Long-term isothermal aging studies showed that BTDB- based 12F resins exhibited comparable thermo-oxidative stability to P-PDA based PMR-11 polyimides. The noncoplanar 2- and 2'-disubstituted biphenyldiamine (BTDB) not only lowered the melt viscosities of 12F prepolymers, but also retained reasonable thermal sta- bility of the cured resins. The 12F polyimide resin with p-aminostyrene endcaps showed the best promise for long-term, high-temperature application at 343 C (650 F).

  8. Fuel Additives: Canada bans MMT

    SciTech Connect

    Sissell, K.

    1997-04-16

    The Canadian Senate voted late last week to ban use of the manganese-based fuel additive MMT, produced only in the US by Ethyl. MMT, which has been sold in Canada for the past 20 years and accounts for about half of Ethyl`s Canadian sales, has been criticized by environmentalists, who have raised public health concerns, and automakers, who say it harms emission control systems. {open_quotes}Canada`s vote is a great victory for public health and the environment,{close_quotes} says Environmental Defense Fund executive director Fred Krupp. {open_quotes}The US should move swiftly to follow suit and suspend sales of MMT until adequate toxicity testing on the additive is completed.{close_quotes} EPA had refused to approve MMT for sale because of health concerns but was compelled to do so by a December 1995 court ruling. Ethyl asserts the ban violates Canada`s obligations under Nafta and says it will file a damage claim with the Nafta arbitration panel.

  9. Additive interaction between heterogeneous environmental ...

    EPA Pesticide Factsheets

    BACKGROUND Environmental exposures often occur in tandem; however, epidemiological research often focuses on singular exposures. Statistical interactions among broad, well-characterized environmental domains have not yet been evaluated in association with health. We address this gap by conducting a county-level cross-sectional analysis of interactions between Environmental Quality Index (EQI) domain indices on preterm birth in the Unites States from 2000-2005.METHODS: The EQI, a county-level index constructed for the 2000-2005 time period, was constructed from five domain-specific indices (air, water, land, built and sociodemographic) using principal component analyses. County-level preterm birth rates (n=3141) were estimated using live births from the National Center for Health Statistics. Linear regression was used to estimate prevalence differences (PD) and 95% confidence intervals (CI) comparing worse environmental quality to the better quality for each model for a) each individual domain main effect b) the interaction contrast and c) the two main effects plus interaction effect (i.e. the “net effect”) to show departure from additive interaction for the all U.S counties. Analyses were also performed for subgroupings by four urban/rural strata. RESULTS: We found the suggestion of antagonistic interactions but no synergism, along with several purely additive (i.e., no interaction) associations. In the non-stratified model, we observed antagonistic interac

  10. Additive manufacturing of RF absorbers

    NASA Astrophysics Data System (ADS)

    Mills, Matthew S.

    The ability of additive manufacturing techniques to fabricate integrated electromagnetic absorbers tuned for specific radio frequency bands within structural composites allows for unique combinations of mechanical and electromagnetic properties. These composites and films can be used for RF shielding of sensitive electromagnetic components through in-plane and out-of-plane RF absorption. Structural composites are a common building block of many commercial platforms. These platforms may be placed in situations in which there is a need for embedded RF absorbing properties along with structural properties. Instead of adding radar absorbing treatments to the external surface of existing structures, which adds increased size, weight and cost; it could prove to be advantageous to integrate the microwave absorbing properties directly into the composite during the fabrication process. In this thesis, a method based on additive manufacturing techniques of composites structures with prescribed electromagnetic loss, within the frequency range 1 to 26GHz, is presented. This method utilizes screen printing and nScrypt micro dispensing to pattern a carbon based ink onto low loss substrates. The materials chosen for this study will be presented, and the fabrication technique that these materials went through to create RF absorbing structures will be described. The calibration methods used, the modeling of the RF structures, and the applications in which this technology can be utilized will also be presented.

  11. 76 FR 44625 - Northern Lights Variable Trust, et al.;

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-07-26

    ... COMMISSION Northern Lights Variable Trust, et al.; Notice of Application July 19, 2011. AGENCY: Securities.... APPLICANTS: Northern Lights Variable Trust (the ``Fund'') and Gemini Fund Services, LLC (``Gemini.... Northern Lights Variable Trust, c/o Emile Molineaux, Esquire, Gemini Fund Services, LLC, 450...

  12. 78 FR 8501 - Northern Natural Gas Company; Notice of Application

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-02-06

    ... Energy Regulatory Commission Northern Natural Gas Company; Notice of Application Take notice that on January 18, 2013, Northern Natural Gas Company (Northern), 1111 South 103rd Street, Omaha, Nebraska 68124, filed in Docket No. CP13-53-000, an application pursuant to section 7(c) of the Natural Gas Act...

  13. 75 FR 35779 - Northern Natural Gas Company; Notice of Application

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-06-23

    ... Energy Regulatory Commission Northern Natural Gas Company; Notice of Application June 16, 2010. Take notice that on June 2, 2010, Northern Natural Gas Company (Northern), 1111 South 103rd Street, Omaha... Natural Gas Act, for a certificate of public convenience and necessity authorizing the increase...

  14. 78 FR 51716 - Northern Natural Gas Company; Notice of Application

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-08-21

    ... Energy Regulatory Commission Northern Natural Gas Company; Notice of Application Take notice that on August 1, 2013, Northern Natural Gas Company (Northern), 1111 South 103rd Street, Omaha, Nebraska 68124, filed an application pursuant to section 7(c) of the Natural Gas Act and part 157 of the...

  15. 76 FR 12721 - Northern Natural Gas Company; Notice of Application

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-03-08

    ... Energy Regulatory Commission Northern Natural Gas Company; Notice of Application Take notice that on February 18, 2011, Northern Natural Gas Company (Northern), 1111 South 103 Street, Omaha, Nebraska 68124-1000, filed in Docket No. CP11-98-000, an application pursuant to section 7(b) of the Natural Gas...

  16. 77 FR 35958 - Northern Natural Gas Company; Notice of Application

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-06-15

    ... Energy Regulatory Commission Northern Natural Gas Company; Notice of Application Take notice that on May 30, 2012, Northern Natural Gas Company (Northern), 1111 South 103rd Street, Omaha, Nebraska 68124... regulations and section 7(b) of the Natural Gas Act, to abandon by sale to DKM Enterprises, LLC (DKM)...

  17. Canada: Native Land Rights and Northern Development. IWGIA Document 26.

    ERIC Educational Resources Information Center

    Cumming, Peter A.

    Presenting the critical elements for a new and meaningful relationship between the Inuit and the emerging industrial society of Northern Canada, this publication includes: (1) Canada as a Nation State and Northern Development; (2) Northern Development and Institutions in Decision-Making; (3) The Northwest Territories and Yukon Territory as…

  18. Neutron Characterization for Additive Manufacturing

    NASA Technical Reports Server (NTRS)

    Watkins, Thomas; Bilheux, Hassina; An, Ke; Payzant, Andrew; DeHoff, Ryan; Duty, Chad; Peter, William; Blue, Craig; Brice, Craig A.

    2013-01-01

    Oak Ridge National Laboratory (ORNL) is leveraging decades of experience in neutron characterization of advanced materials together with resources such as the Spallation Neutron Source (SNS) and the High Flux Isotope Reactor (HFIR) shown in Fig. 1 to solve challenging problems in additive manufacturing (AM). Additive manufacturing, or three-dimensional (3-D) printing, is a rapidly maturing technology wherein components are built by selectively adding feedstock material at locations specified by a computer model. The majority of these technologies use thermally driven phase change mechanisms to convert the feedstock into functioning material. As the molten material cools and solidifies, the component is subjected to significant thermal gradients, generating significant internal stresses throughout the part (Fig. 2). As layers are added, inherent residual stresses cause warping and distortions that lead to geometrical differences between the final part and the original computer generated design. This effect also limits geometries that can be fabricated using AM, such as thin-walled, high-aspect- ratio, and overhanging structures. Distortion may be minimized by intelligent toolpath planning or strategic placement of support structures, but these approaches are not well understood and often "Edisonian" in nature. Residual stresses can also impact component performance during operation. For example, in a thermally cycled environment such as a high-pressure turbine engine, residual stresses can cause components to distort unpredictably. Different thermal treatments on as-fabricated AM components have been used to minimize residual stress, but components still retain a nonhomogeneous stress state and/or demonstrate a relaxation-derived geometric distortion. Industry, federal laboratory, and university collaboration is needed to address these challenges and enable the U.S. to compete in the global market. Work is currently being conducted on AM technologies at the ORNL

  19. Additives in fibers and fabrics.

    PubMed Central

    Barker, R H

    1975-01-01

    The additives and contaminants which occur in textile fibers vary widely, depending on the type of fiber and the pretreatment which it has received. Synthetic fibers such as nylon and polyester contain trace amounts of contaminants such as catalysts and catalyst deactivators which remain after the synthesis of the basic polymers. In addition, there are frequently a number of materials which are added to perform specific functions in almost all man-made fibers. Examples of these would include traces of metals or metal salts used as tracers for identification of specific lots of fiber, TiO2 or similar materials added as delustrants, and a host of organic species added for such special purposes as antistatic agents or flame retardants. There may also be considerable quantities of residual monomer or small oligomers dissolved in the polymer matrix. The situation becomes even more complex after the fibers are converted into fabric form. Numerous materials are applied at various stages of fabric preparation to act as lubricants, sizing agents, antistats, bleaches, and wetting agents to facilitate the processing, but these are normally removed before the fabric reaches the cutters of the ultimate consumers and therefore usually do not constitute potential hazards. However, there are many other chemical agents which are frequently added during the later stages of fabric preparation and which are not designed to be removed. Aside from dyes and printing pigments, the most common additive for apparel fabrics is a durable press treatment. This generally involves the use of materials capable of crosslinking cellulosics by reacting through such functions as N-methylolated amides or related compounds such as ureas and carbamates. These materials pose some potential hazards due to both the nitrogenous bases and the formaldehyde which they usually release. There is usually also some residual catalyst in fabrics which have received such treatments. Other types of chemical treatments

  20. Ground ice conditions in Salluit, Northern Quebec

    NASA Astrophysics Data System (ADS)

    Allard, M.; Fortier, R.; Calmels, F.; Gagnon, O.; L'Hérault, E.

    2011-12-01

    Salluit in Northern Québec (ca. 1300 inhabitants) faces difficult ground ice conditions for its development. The village is located in a U-shaped valley, along a fjord that was deglaciated around 8000 cal BP. The post-glacial marine limit is at the current elevation of 150 m ASL. Among the mapped surficial geology units, three contain particularly ice-rich permafrost: marine clays, till and silty colluviums. A diamond drill was used to extract 10 permafrost cores down to 23 m deep. In addition, 18 shallow cores (to 5 m deep) were extracted with a portable drill. All the frozen cores were shipped to Québec city where ground ice contents were measured and cryostructures were imaged by CT-Scanning. Water contents, grain-size and pore water salinity were measured. Refraction seismic profiles were run to measure the depth to bedrock. GPR and electrical resistivity surveys helped to map ice-rich areas. Three cone penetration tests (CPT) were run in the frozen clays to depths ranging from 8 to 21 m. Maximum clay thickness is ca. 50 m deep near the shoreline. The cone penetration tests and all the cores in clays revealed large amounts of both segregated and aggradational ice (volumetric contents up to 93% over thicknesses of one meter) to depths varying between 2.5 and 4 m, below which the ice content decreases and the salinity increases (values measured up to 42 gr/L between 4.5 and 6 m deep). Chunks of organic matter buried below the actual active layer base indicate past cryoturbations under a somewhat warmer climate, most probably associated with intense frost boil action, as widely observed today. The stony till has developed large quantities of segregation ice which can be seen in larger concentrations and as thicker lenses under boulders and in matrix rich (≥ 50% sand and silt) parts of the glacial sediment. As digging for a sewage pond was undertaken in winter 2008 by blasting, the clast-influenced cryostructure of the till could be observed in cuts and in