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Sample records for denaturing polyacrylamide gel

  1. Detection of enzymes active on various beta-1,3-glucans after denaturing polyacrylamide gel electrophoresis.

    Science.gov (United States)

    Trudel, J; Grenier, J; Asselin, A

    1998-07-01

    Enzymes were assayed for glucanase activity after denaturing sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) in gels containing beta-1,3-glucans embedded as substrate. Lentinan, curdlan, paramylon, baker's yeast alkali-insoluble glucan, baker's yeast alkali-soluble glucan and carboxymethyl (CM)-pachyman were compared to oligomeric laminarin, which is the usual substrate for assaying beta-1,3-glucanase activities. Detecting enzyme activities by aniline blue fluorescent staining was also compared with the staining of released reducing sugars by 2,3,5-triphenyltetrazolium chloride (TTC). For the nonreduced proteins, the Driselase extract exhibited one major band at 32.5 kDa and one less intense band at 23 kDa for most substrates with the two detection procedures. No Lyticase enzyme was detected in either detection procedures for all tested substrates. For barley enzymes, no activity was revealed after aniline blue staining while one undescribed 19 kDa glucanase activity was best shown after TTC staining with curdlan, paramylon and CM-pachyman as substrates. In the case of reduced proteins, the Lyticase extract yielded three bands (33, 36 and 46 kDa) on several substrates with both detection procedures. This was the same for the barley leaf extract (32, 36 and 39 kDa). The Driselase extract showed one 42 kDa band. Many enzymes active on beta-1,3-glucans are thus best revealed when proteins are denatured and reduced and when protein renaturation after SDS-PAGE involves a pH 8.0 treatment and the inclusion of 1 mM cysteine in buffers. However, some enzymes are only detected when proteins are denatured without reduction. Finally, the use of various polymeric beta-1,3-glucan substrates different from oligomeric laminarin is necessary to detect new types of enzymes such as the 19 kDa barley glucanase.

  2. Characterization of Multi-subunit Protein Complexes of Human MxA Using Non-denaturing Polyacrylamide Gel-electrophoresis.

    Science.gov (United States)

    Nigg, Patricia E; Pavlovic, Jovan

    2016-10-28

    The formation of oligomeric complexes is a crucial prerequisite for the proper structure and function of many proteins. The interferon-induced antiviral effector protein MxA exerts a broad antiviral activity against many viruses. MxA is a dynamin-like GTPase and has the capacity to form oligomeric structures of higher order. However, whether oligomerization of MxA is required for its antiviral activity is an issue of debate. We describe here a simple protocol to assess the oligomeric state of endogenously or ectopically expressed MxA in the cytoplasmic fraction of human cell lines by non-denaturing polyacrylamide gel electrophoresis (PAGE) in combination with Western blot analysis. A critical step of the protocol is the choice of detergents to prevent aggregation and/or precipitation of proteins particularly associated with cellular membranes such as MxA, without interfering with its enzymatic activity. Another crucial aspect of the protocol is the irreversible protection of the free thiol groups of cysteine residues by iodoacetamide to prevent artificial interactions of the protein. This protocol is suitable for a simple assessment of the oligomeric state of MxA and furthermore allows a direct correlation of the antiviral activity of MxA interface mutants with their respective oligomeric states.

  3. Silver and Cyanine Staining of Oligonucleotides in Polyacrylamide Gel.

    Directory of Open Access Journals (Sweden)

    Weizhong Tang

    Full Text Available To explore why some oligonucleotides in denaturing polyacrylamide gel could not be silver-stained, 134 different oligonucleotides were analyzed using denaturing polyacrylamide gel electrophoresis stained with silver and asymmetric cyanine. As a result, we found that the sensitivity of oligos (dA, (dC, (dG and (dT to silver staining could be ranged as (dA > (dG > (dC > (dT from high to low. It was unexpected that oligo (dT was hard to be silver-stained. Moreover, the silver staining of an oligonucleotide containing base T could be partially or completely inhibited by base T. The inhibition of silver staining by base T was a competitive inhibition which could be affected by the amounts of the argyrophil nucleobase and base T, the cis-distance between the argyrophil nucleobase and base T, and the gel concentration. The changes of the intensity of an oligonucleotide band caused by the changes of DNA base composition were diverse and interesting. The intensity of some oligonucleotide bands would significantly change when the changes of DNA base composition accumulated to a certain extent (usually ≥ 4 nt. The sensitivity of cyanine staining of ≤ 11-nt long oligonucleotides could be enhanced about 250-fold by fixing the gels with methanol fixing solution.

  4. One-dimensional SDS-polyacrylamide gel electrophoresis (1D SDS-PAGE).

    Science.gov (United States)

    Brunelle, Julie L; Green, Rachel

    2014-01-01

    This protocol describes a denaturing polyacrylamide gel system utilizing sodium dodecyl sulfate (SDS) to separate protein molecules based on size as first described by Laemmli (1970). SDS-PAGE can be used to monitor protein purifications, check the purity of samples, and to estimate molecular weights for unknown proteins. © 2014 Elsevier Inc. All rights reserved.

  5. Electrophoresis of DNA in agarose gels, polyacrylamide gels and in free solution

    Science.gov (United States)

    Stellwagen, Nancy C.

    2009-01-01

    This review describes the electrophoresis of curved and normal DNA molecules in agarose gels, polyacrylamide gels and in free solution. These studies were undertaken to clarify why curved DNA molecules migrate anomalously slowly in polyacrylamide gels but not in agarose gels. Two milestone papers are cited, in which Ferguson plots were used to estimate the effective pore size of agarose and polyacrylamide gels. Subsequent studies on the effect of the electric field on agarose and polyacrylamide gel matrices, DNA interactions with the two gel matrices, and the effect of curvature on the free solution mobility of DNA are also described. The combined results suggest that the anomalously slow mobilities observed for curved DNA molecules in polyacrylamide gels are due primarily to preferential interactions of curved DNAs with the polyacrylamide gel matrix; the restrictive pore size of the matrix is of lesser importance. In free solution, DNA mobilities increase with increasing molecular mass until leveling off at a plateau value of (3.17 ± 0.01) × 10-4 cm2/Vs in 40 mM Tris-acetate-EDTA buffer at 20°C. Curved DNA molecules migrate anomalously slowly in free solution as well as in polyacrylamide gels, explaining why the Ferguson plots of curved and normal DNAs containing the same number of base pairs extrapolate to different mobilities at zero gel concentration. PMID:19517510

  6. Evaluation of wheat by polyacrylamide gel electrophoresis

    African Journals Online (AJOL)

    PRECIOUS

    2010-01-11

    Jan 11, 2010 ... SDS-PAGE gels cluster analysis was performed to check the ... It is concluded that SDS-PAGE analysis of wheat endosperm protein is useful for evaluation of ..... Comparison of phenotypic and molecular marker-based.

  7. Electrically induced gel-sol transition of polyvinyl alcohol/ polyacrylamide semi-interpenetrated hydrogels

    Institute of Scientific and Technical Information of China (English)

    Xia Chao Jin; Yong Min Huang; Hong Lai Liu

    2009-01-01

    Polyvinyl alcohol/polyacrylamide semi-interpenetrated hydrogels were prepared via freeze-thaw process. When a 20 V of DC was applied across the gels, the gels with lower polyacrylamide content underwent a contraction or partly turned into solution, while for the gels with higher polyacrylamide concentration, a complete gel-sol transition was observed in a short time.

  8. Maltodextrin hydrolysis with glucoamylase immobilized in polyacrylamide gel

    Energy Technology Data Exchange (ETDEWEB)

    Yankov, D.; Peeva, L.; Beschkov, V. (Inst. of Chemical Engineering, Bulgarian Academy of Sciences, Sofia (Bulgaria))

    1992-08-01

    The immobilization of glucoamylase by entrapping in a polyacrylamide gel has been studied. The optimum values of pH and temperature for the immobilized enzyme have been determinated. The influence of worked off cycles, substrate concentration and initial degree of hydrolysis on the final conversion of starch have been investigated. (orig.).

  9. Polyacrylamide gel electrophoresis of isoenzymes from Entamoeba species.

    OpenAIRE

    Mathews, H M; Moss, D M; Healy, G R; Visvesvara, G S

    1983-01-01

    In this preliminary report, we describe a polyacrylamide gel electrophoresis technique for the resolution of isoenzyme patterns of four isolates of Entamoeba histolytica and one isolate of Entamoeba coli. Our findings were similar to previous findings for three enzyme systems: maleic enzyme (malate dehydrogenase [EC 1.1.1.40]), hexokinase (EC 2.7.1.1), and phosphoglucomutase (EC 2.7.5.1). We found preliminary evidence that glucosephosphate isomerase (EC 5.3.1.9) may also differentiate invasiv...

  10. Basics and recent advances of two dimensional- polyacrylamide gel electrophoresis

    Science.gov (United States)

    2014-01-01

    Gel- based proteomics is one of the most versatile methods for fractionating protein complexes. Among these methods, two dimensional- polyacrylamide gel electrophoresis (2-DE) represents a mainstay orthogonal approach, which is popularly used to simultaneously fractionate, identify, and quantify proteins when coupled with mass spectrometric identification or other immunological tests. Although 2-DE was first introduced more than three decades ago, several challenges and limitations to its utility still exist. This review discusses the principles of 2-DE as well as both recent methodological advances and new applications. PMID:24735559

  11. Western Blot of Stained Proteins from Dried Polyacrylamide Gels

    Science.gov (United States)

    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.

  12. Scalable lithography from Natural DNA Patterns via polyacrylamide gel

    Science.gov (United States)

    Qu, Jiehao; Hou, Xianliang; Fan, Wanchao; Xi, Guanghui; Diao, Hongyan; Liu, Xiangdon

    2015-12-01

    A facile strategy for fabricating scalable stamps has been developed using cross-linked polyacrylamide gel (PAMG) that controllably and precisely shrinks and swells with water content. Aligned patterns of natural DNA molecules were prepared by evaporative self-assembly on a PMMA substrate, and were transferred to unsaturated polyester resin (UPR) to form a negative replica. The negative was used to pattern the linear structures onto the surface of water-swollen PAMG, and the pattern sizes on the PAMG stamp were customized by adjusting the water content of the PAMG. As a result, consistent reproduction of DNA patterns could be achieved with feature sizes that can be controlled over the range of 40%-200% of the original pattern dimensions. This methodology is novel and may pave a new avenue for manufacturing stamp-based functional nanostructures in a simple and cost-effective manner on a large scale.

  13. Characteristics of polyacrylamide gel with THPC and Turnbull Blue gel dosimeters evaluated using optical tomography

    Science.gov (United States)

    Pilařová (Vávrů), Kateřina; Kozubíková, Petra; Šolc, Jaroslav; Spěváček, Václav

    2014-11-01

    The purpose of this study was to compare characteristics of radiochromic gel - Turnbull Blue gel (TB gel) with polymer gel - polyacrylamide gel and tetrakis hydroxymethyl phosphonium chloride (PAGAT) using optical tomography. Both types of gels were examined in terms of dose sensitivity, dose response linearity and background value of spectrophotometric absorbance. The calibration curve was obtained for 60Co irradiation performed on Gammacell 220 at predefined gamma dose levels between 0 and 140 Gy for TBG and 0-15 Gy for PAGAT. To measure relative dose distributions from stereotactic irradiation, dosimeters were irradiated on Leksell Gamma Knife Perfexion. The cylindrical glass housings filled with gel were attached to the stereotactic frame. They were exposed with single shot and 16 mm collimator by 65 Gy to a 50% prescription isodose for TB gel and 4 Gy to a 50% prescription isodose for PAGAT. Evaluations of dosimeters were performed on an UV-vis Spectrophotometer Helios β and an optical cone beam homemade tomography scanner with a 16-bit astronomy CCD camera with a set of color filters. The advantages and potential disadvantages for both types of gel dosimeters were summarized. Dose distribution in central slice and measured profiles of 16 mm shot shows excellent correspondence with treatment planning system Leksell GammaPlan® for both PAGAT and Turnbull Blue gels. Gel dosimeters are suitable for steep dose gradient verification. An optical tomography evaluation method is successful. Dose response characteristics of TB gel and PAGAT gel are presented.

  14. Improved recovery of DNA from polyacrylamide gels after in situ DNA footprinting

    NARCIS (Netherlands)

    van Keulen, G; Meijer, WG

    Methods used to date for the isolation of DNA from polyacrylamide gels are elution based, time-consuming and with low yield in DNA. This paper describes an improved system employing polyacrylamide gels made of a meltable matrix. The new system was successfully applied to in situ DNA footprinting

  15. Improved recovery of DNA from polyacrylamide gels after in situ DNA footprinting

    NARCIS (Netherlands)

    van Keulen, G; Meijer, WG

    2003-01-01

    Methods used to date for the isolation of DNA from polyacrylamide gels are elution based, time-consuming and with low yield in DNA. This paper describes an improved system employing polyacrylamide gels made of a meltable matrix. The new system was successfully applied to in situ DNA footprinting fol

  16. Non-Gradient Blue Native Polyacrylamide Gel Electrophoresis.

    Science.gov (United States)

    Luo, Xiaoting; Wu, Jinzi; Jin, Zhen; Yan, Liang-Jun

    2017-02-02

    Gradient blue native polyacrylamide gel electrophoresis (BN-PAGE) is a well established and widely used technique for activity analysis of high-molecular-weight proteins, protein complexes, and protein-protein interactions. Since its inception in the early 1990s, a variety of minor modifications have been made to this gradient gel analytical method. Here we provide a major modification of the method, which we call non-gradient BN-PAGE. The procedure, similar to that of non-gradient SDS-PAGE, is simple because there is no expensive gradient maker involved. The non-gradient BN-PAGE protocols presented herein provide guidelines on the analysis of mitochondrial protein complexes, in particular, dihydrolipoamide dehydrogenase (DLDH) and those in the electron transport chain. Protocols for the analysis of blood esterases or mitochondrial esterases are also presented. The non-gradient BN-PAGE method may be tailored for analysis of specific proteins according to their molecular weight regardless of whether the target proteins are hydrophobic or hydrophilic. © 2017 by John Wiley & Sons, Inc. Copyright © 2017 John Wiley & Sons, Inc.

  17. A versatile polyacrylamide gel electrophoresis based sulfotransferase assay

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    Prather Brittany

    2010-02-01

    Full Text Available Abstract Background Sulfotransferases are a large group of enzymes that regulate the biological activity or availability of a wide spectrum of substrates through sulfation with the sulfur donor 3'-phosphoadenosine-5'-phosphosulfate (PAPS. These enzymes are known to be difficult to assay. A convenient assay is needed in order to better understand these enzymes. Results A universal sulfotransferase assay method based on sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE is described. This assay has been successfully applied to substrates as small as α-naphthol and as big as proteoglycans. As examples, we present the assays for recombinant human CHST4, TPST1, CHST3 and HS6ST1. In order to assess whether a small molecule can be applicable to this type of assay, a method to estimate the relative mobility of a molecule to PAPS is also presented. The estimated relative mobilities of various sulfated small molecules generated by SULT1A1, SULT1E1, SULT2A1 and CHST4 are in the range of ± 0.2 of the actual relative mobilities. Conclusion The versatility of the current method comes from the ability that SDS-PAGE can separate proteins and small molecules according to different parameters. While mobilities of proteins during SDS-PAGE are inversely related to their sizes, mobilities of small molecules are positively related to their charge/mass ratios. The predicted relative mobility of a product to PAPS is a good indicator of whether a sulfotransferase can be assayed with SDS-PAGE. Because phosphorylation is most similar to sulfation in chemistry, the method is likely to be applicable to kinases as well.

  18. Denaturing and non-denaturing gel electrophoresis as methods for the detection ofjunctional diversity in rearranged T cell receptor sequences

    NARCIS (Netherlands)

    Offermans, M.T.C.; Sonneveld, R.D.; Bakker, E.; Deutz-Terlouw, P.P.; Geus, B. de; Rozing, J.

    1995-01-01

    Two nucleic acid gel electrophoresis techniques were tested as a possible tool for analyzing junctional diversity in rearranged T cell receptor (TcR) sequences in order to define the extent of T cell heterogeneity. For this purpose denaturing gradient gel electrophoresis (DGGE) as well as

  19. Agarose gel electrophoresis and polyacrylamide gel electrophoresis for visualization of simple sequence repeats.

    Science.gov (United States)

    Anderson, James; Wright, Drew; Meksem, Khalid

    2013-01-01

    In the modern age of genetic research there is a constant search for ways to improve the efficiency of plant selection. The most recent technology that can result in a highly efficient means of selection and still be done at a low cost is through plant selection directed by simple sequence repeats (SSRs or microsatellites). The molecular markers are used to select for certain desirable plant traits without relying on ambiguous phenotypic data. The best way to detect these is the use of gel electrophoresis. Gel electrophoresis is a common technique in laboratory settings which is used to separate deoxyribonucleic acid (DNA) and ribonucleic acid (RNA) by size. Loading DNA and RNA onto gels allows for visualization of the size of fragments through the separation of DNA and RNA fragments. This is achieved through the use of the charge in the particles. As the fragments separate, they form into distinct bands at set sizes. We describe the ability to visualize SSRs on slab gels of agarose and polyacrylamide gel electrophoresis.

  20. A continuous acetic acid system for polyacrylamide gel electrophoresis of gliadins and other prolamines.

    Science.gov (United States)

    Clements, R L

    1988-02-01

    A polyacrylamide gel electrophoresis system buffered by acetic acid alone was developed for electrophoresis of prolamines. When applied to gliadin electrophoresis, the acetic acid system produces more bands than does a conventional aluminum lactate-lactic acid system (using 12% acrylamide gels). The acetic acid system is relatively simple, requiring a single buffer component that is universally available in high purity.

  1. A technique for detecting antifungal activity of proteins separated by polyacrylamide gel electrophoresis.

    Science.gov (United States)

    De Bolle, M F; Goderis, I J; Terras, F R; Cammue, B P; Broekaert, W F

    1991-06-01

    A technique was developed for the detection of antifungal activity of proteins after discontinuous polyacrylamide gel electrophoresis under native conditions. The antifungal activity is detected as growth inhibition zones in a homogeneous fungal lawn, grown in an agar layer spread on top of the polyacrylamide gel. The position of proteins with antifungal activity can be determined on a diffusion blot prepared from the same gel. The technique is illustrated for three antifungal plant proteins, i.e. alpha-purothionin, Urtica dioica agglutinin, and tobacco chitinase.

  2. An improved silver staining procedure for schizodeme analysis in polyacrylamide gradient gels

    Directory of Open Access Journals (Sweden)

    Antonio M. Gonçalves

    1990-03-01

    Full Text Available A simple protocol is described for the silver staining of polyacrylamide gradient gels used for the separation of restriction fragments of kinetoplast DNA [schizodeme analysis of trypanosomatids (Morel et al., 1980]. The method overcomes the problems of non-uniform staining and strong background color which are frequently encountered when conventional protocols for silver staining of linear gels. The method described has proven to be of general applicability for DNA, RNA and protein separations in gradient gels.

  3. Two previously undetected variants of glutamic-pyruvic transaminase found by acidic polyacrylamide gel electrophoresis.

    OpenAIRE

    McLellan, T

    1982-01-01

    Two new electrophoretic variants of glutamic-pyruvic transaminase (GPT) have been found by polyacrylamide gel electrophoresis at acidic pH. They appeared to represent a single allele, GPT 2, by the standard method of starch gel electrophoresis. Studies in families show that they are inherited as codominant alleles at the GPT locus. Population frequencies are about the same as those of other rare GPT variants. Their behavior on gels is consistent with both of them having substitutions of histi...

  4. Congruence between starch gel and polyacrylamide gel electrophoresis in detecting allozyme variation in pulmonate land slugs.

    Science.gov (United States)

    Geenen, Sofie; Jordaens, Kurt; Castilho, Rita; Backeljau, Thierry

    2003-02-01

    The predominantly selfing slug species Arion (Carinarion) fasciatus, A. (C.) silvaticus and A. (C.) circumscriptus are native in Europe and have been introduced into North America, where each species consists of a single, homozygous multilocus genotype (strain), as defined by starch gel electrophoresis (SGE) of allozymes. In Europe, the "one strain per species" hypothesis does not hold since polyacrylamide gel electrophoresis (PAGE) of allozymes uncovered 46 strains divided over the three species. However, electrophoretic techniques may differ in their ability to detect allozyme variation. Therefore, several Carinarion populations from both continents were screened by applying the two techniques simultaneously on the same individual slugs and enzyme loci. SGE and PAGE yielded exactly the same results, so that the different degree of variation in North American and European populations cannot be attributed to differences in resolving power between SGE and PAGE. We found four A. (C.) silvaticus strains in North America indicating that in this region the "one strain per species" hypothesis also cannot be maintained. Hence, the discrepancies between previous electrophoretic studies on Carinarion are most likely due to sampling artefacts and possible founder effects.

  5. Application of denaturing gradient gel electrophoresis (DGGE) to the analysis of endodontic infections.

    Science.gov (United States)

    Siqueira, José F; Rôças, Isabela N; Rosado, Alexandre S

    2005-11-01

    The recent expanding use of cultivation-independent techniques for bacterial identification is reliant on the lack of knowledge of the conditions under which most bacteria are growing in their natural habitat and the difficulty to develop culture media that accurately reproduce these conditions. A molecular method that has been recently used in several areas to examine the bacterial diversity living in diverse environments is the denaturing gradient gel electrophoresis (DGGE). In DGGE, polymerase chain reaction (PCR)-generated DNA fragments of the same length but with different base-pair sequences can be separated. Separation is based on electrophorectic mobility of a partially melted double-strand DNA molecule in polyacrylamide gels, which is decreased when compared with that of the completely helical form of the molecule. Molecules with different sequences may have a different melting behavior and will therefore stop migrating at different positions in the gel. Application of the PCR-DGGE method in endodontic research has revealed that there are significant differences in the predominant bacterial composition between asymptomatic and symptomatic cases. This suggests that the structure of the bacterial community can play a role in the development of symptoms. In addition, new bacterial phylotypes have been disclosed in primary endodontic infections. PCR-DGGE has also confirmed that intra-radicular infections are a common finding in root-filled teeth associated with persistent periradicular lesions. The microbiota in failed cases significantly vary from teeth to teeth, with a mean number of species far higher than previously shown by culturing approaches. Application of the PCR-DGGE technique in endodontic microbiology research has the potential to shed light on several aspects of the different types of endodontic infection as well as on the effects of treatment procedures with regard to infection control.

  6. A Novel Method for Detection of Glycoproteins on Sodium Dodecyl Sulphate Polyacrylamide Gel Using Radio-Iodinated Tyrosine

    DEFF Research Database (Denmark)

    Nalla, Amarnadh; Draz, Hossam M.; Dole, Anita;

    2009-01-01

    The aim of this study is to develop a novel method for detection of glycoproteins on polyacrylamide gel. In this method, radio-iodinated-tyrosine (125I-tyrosine) was conjugated to glycoprotein by schiff's base mechanism on the sodium dodecyl sulfate- polyacrylamide gel. Ovalbumin and Concanavalin...

  7. Denaturing gradient gel electrophoresis profiling of bacterial communities composition in Arabian Sea

    Digital Repository Service at National Institute of Oceanography (India)

    Singh, S.K.; Ramaiah, N.

    Denaturing gradient gel electrophoresis (DGGE) was used to elucidate spatial and temporal variations in bacterial community composition (BCC) from four locations along the central west coast of India. DNA extracts from 36 water samples collected...

  8. Wide frequency rheological modeling of crosslinked polyacrylamide gels

    CERN Document Server

    Abidine, Yara; Michel, Richard; Duperray, Alain; Palade, Liviu Iulian; Verdier, Claude

    2013-01-01

    Gels are known to behave as viscoelastic materials but only a small amount of data is usually provided in the glassy transition. Results concerning the dynamic moduli G′ and G′′ are presented here using AFM in contact oscillatory mode and show good agreement with classical rheological data. Different gels are studied with increasing polymer concentration. G0N, the plateau modulus, is measured at low frequencies, but interestingly another one, G1, is found at high frequencies. A model based on fractional derivatives is proposed, covering the whole frequency range. The relaxation spectrum is recovered, and the physical parameters contain interesting information about the local dynamics of crosslinks.

  9. Separation of Native Allophycocyanin and R-Phycocyanin from Marine Red Macroalga Polysiphonia urceolata by the Polyacrylamide Gel Electrophoresis Performed in Novel Buffer Systems

    OpenAIRE

    Yu Wang; Xueqin Gong; Shumei Wang; Lixue Chen; Li Sun

    2014-01-01

    Three buffer systems of Imidazole-Acetic acid, HEPES-Imidazole/Bis-tris and Bis-tris-HEPES-MES were designed based on the principle of discontinuous polyacrylamide gel electrophoresis (PAGE) for the native PAGE which could be performed in pH 7.0 and 6.5 in order to analyze and prepare the minor components of allophycocyanin (AP) and R-phycocyanin (R-PC) from marine red macroalga Polysiphonia urceolata. These AP and R-PC phycobiliproteins are easily denatured in alkaline environments. The obta...

  10. Thermal denaturation of type I collagen vitrified gels

    Energy Technology Data Exchange (ETDEWEB)

    Xia, Zhiyong, E-mail: zhiyong.xia@jhuapl.edu [The Johns Hopkins University, Applied Physics Laboratory, Laurel, MD 20723 (United States); Calderon-Colon, Xiomara [The Johns Hopkins University, Applied Physics Laboratory, Laurel, MD 20723 (United States); Trexler, Morgana, E-mail: morgana.trexler@jhuapl.edu [The Johns Hopkins University, Applied Physics Laboratory, Laurel, MD 20723 (United States); Elisseeff, Jennifer; Guo, Qiongyu [The Johns Hopkins University, Department of Biomedical Engineering, Baltimore, MD 21231 (United States)

    2012-01-10

    Highlights: Black-Right-Pointing-Pointer We analyzed the denaturation of vitrigels synthesized under different conditions. Black-Right-Pointing-Pointer Overall denaturation kinetics consisted of both reversible and irreversible steps. Black-Right-Pointing-Pointer More stable vitrigels were formed under high level of vitrification. - Abstract: The denaturation kinetics of type I collagen vitrigels synthesized under different vitrification time and temperature were analyzed by the classical Kissinger approach and the advanced model free kinetics (AMFK) using the Vyazovkin algorithm. The AMFK successfully elucidated the overall denaturation into reversible and irreversible processes. Depending on vitrification conditions, the activation energy for the irreversible process ranged from 100 to 200 kJ/mol, and the reversible enthalpy ranged from 250 to 300 kJ/mol. All of these values increased with the vitrification time and temperature, indicating that a more stable and complex structure formed with increased vitrification. The classical Kissinger method predicted the presence of a critical temperate of approximately 60 Degree-Sign C for the transition between reversible and irreversible processes. Scanning electron microscopy revealed the presence of fibril structures in vitrigels both before and after full denaturation; however the fibrils had became thicker and rougher after denaturation.

  11. Tris-acetate polyacrylamide gradient gel electrophoresis for the analysis of protein oligomerization.

    Science.gov (United States)

    Cubillos-Rojas, Monica; Schneider, Taiane; Sánchez-Tena, Susana; Bartrons, Ramon; Ventura, Francesc; Rosa, Jose Luis

    2016-02-01

    Here we report a new approach for studying protein oligomerization in cells using a single electrophoresis gel. We combined the use of a crosslinking reagent for sample preparation, such as glutaraldehyde, with the analysis of oligomers by Tris-acetate polyacrylamide gel electrophoresis. The use of a 3-15% Tris-acetate polyacrylamide gradient gel allows for the simultaneous analysis of proteins of masses ranging from 10 to 500 kDa. We showed the usefulness of this method for analyzing endogenous p53 oligomerization with high resolution and sensitivity in human cells. Oligomerization analysis was dependent on the crosslinker concentration used. We also showed that this method could be used to study the regulation of oligomerization. In all experiments, Tris-acetate polyacrylamide gel electrophoresis proved to be a robust, manageable, and cost- and time-efficient method that provided excellent results using a single gel. This approach can be easily extrapolated to the study of other oligomers. All of these features make this method a highly useful tool for the analysis of protein oligomerization.

  12. Polyacrylamide gels with selective recognition of the tetrameric molecular form of human growth hormone

    Directory of Open Access Journals (Sweden)

    R. Kublickas

    2017-08-01

    Full Text Available Networks of polyacrylamide were studied for the possibility of imprinting of the oligomeric form of human growth hormone. The tetrameric molecular form of human growth hormone was molecularly imprinted for the first time. The results show that approximately 50–70% (w/w of the templates (depending on polymerization conditions could be extracted from the molecularly imprinted acrylamide polymers. The resulting ‘gel antibodies’ against this form of human growth hormone in the form of granules of polyacrylamide were compared with granules of non-imprinted polymer. The selectivity of the artificial gel antibodies was studied. Investigation of the binding to imprinted polymer of the template hormone, other molecular forms of the hormone and other proteins shows the selectivity of the developed artificial gel antibodies.

  13. Analysis of variations in band positions for normalization in across-gel denaturing gradient gel electrophoresis.

    Science.gov (United States)

    Matsushita, Yuko; Yamamura, Kohji; Morimoto, Sho; Bao, Zhihua; Kurose, Daisuke; Sato, Ikuo; Yoshida, Shigenobu; Tsushima, Seiya

    2015-05-01

    Variation in band position between gels is a well-known problem in denaturing gradient gel electrophoresis (DGGE). However, few reports have evaluated the degree of variation in detail. In this study, we investigated the variation in band positions of DNA samples extracted from soil, normalized using reference positions within marker lanes for DGGE in three organismal (bacterial, fungal, and nematode) conditions. For sample lanes, marker DNA (as a control) and sample DNA were used. The test for normality of distribution showed that the position data of a large percentage of bands were normally distributed but not for certain bands. For the normally-distributed data, their variations [standard deviation of marker bands (SDM) and standard deviation of sample bands (SDS), respectively] were assessed. For all organismal conditions, the degree of within-gel variation were similar between SDMs and SDSs, while between-gel variations in SDSs were larger than those in SDMs. Due to the large effect of between-gel variations, the total variations in SDSs were more varied between sample bands, and the mean variations of all sample bands were higher than those in the markers. We found that the total variation in the fungal and nematode SDSs decreased when the intervals between marker bands were narrowed, suggesting that band interval is important for reducing total variation in normalized band positions. For the non-normally distributed data, the distribution was examined in detail. This study provided detailed information on the variation of band positions, which could help to optimize markers for reducing band position variation, and could aid in the accurate identification of bands in across-gel DGGE analyses. Copyright © 2015 Elsevier B.V. All rights reserved.

  14. Using in situ rheology to characterize the microstructure in photopolymerized polyacrylamide gels for DNA electrophoresis.

    Science.gov (United States)

    Wang, Jian; Ugaz, Victor M

    2006-09-01

    Photopolymerized cross-linked polyacrylamide hydrogels are attractive sieving matrix formulations for DNA electrophoresis owing to their rapid polymerization times and the potential to locally tailor the gel pore structure through spatial variation of illumination intensity. This capability is especially important in microfluidic systems, where photopolymerization allows gel matrices to be precisely positioned within complex microchannel networks. Separation performance is also directly related to the nanoscale gel pore structure, which is in turn strongly influenced by polymerization kinetics. Unfortunately, detailed studies of the interplay among polymerization kinetics, mechanical properties, and structural morphology are lacking in photopolymerized hydrogel systems. In this paper, we address this issue by performing a series of in situ dynamic small-amplitude oscillatory shear measurements during photopolymerization of cross-linked polyacrylamide electrophoresis gels to investigate the relationship between rheology and parameters associated with the gelation environment including UV intensity, monomer and cross-linker composition, and reaction temperature. In general, we find that the storage modulus G' increases with increasing initial monomer concentration, cross-linker concentration, and polymerization temperature. The steady-state value of G', however, exhibits a more complex dependence on UV intensity that varies with gel concentration. A simple model based on rubber elasticity theory is used to obtain estimates of the average gel pore size that are in surprisingly good agreement with corresponding data obtained from analysis of DNA electrophoretic mobility in gels cast under identical polymerization conditions.

  15. [THE USE OF THE COMMERCIAL APPARATUS DUAL GEL MODULE FOR THE TWO-DIMENSIONAL POLYACRYLAMIDE GEL ELECTROPHORESIS].

    Science.gov (United States)

    Evteeva, I N; Starkova, T Yu; Artemov, A V; Barlev, N A

    2015-01-01

    Two-dimensional gel electrophoresis, continues to be one of the fundamental methods to study the biological protein diversity. This method described by O'Farrell in 1975 includes two following steps: isoelectric focusing in the first dimension and polyacrylamide gel electrophoretic fractionation of proteins according to their molecular weight in the second dimension. In this manuscript we described several technical parameters of the commercial apparatus Dual Gel Module for the gel electrophoresis by means of which it is possible to accomplish the electrophoretic protein fractionation in both dimensions. The distribution of the highly purified commercial proteins used as molecular standards in the detection system of the apparatus Dual Gel Module was identical to the commercial strips of the device GE Healthcare, USA.

  16. Polyacrylamide gels for invadopodia and traction force assays on cancer cells.

    Science.gov (United States)

    Jerrell, Rachel J; Parekh, Aron

    2015-01-04

    Rigid tumor tissues have been strongly implicated in regulating cancer cell migration and invasion. Invasive migration through cross-linked tissues is facilitated by actin-rich protrusions called invadopodia that proteolytically degrade the extracellular matrix (ECM). Invadopodia activity has been shown to be dependent on ECM rigidity and cancer cell contractile forces suggesting that rigidity signals can regulate these subcellular structures through actomyosin contractility. Invasive and contractile properties of cancer cells can be correlated in vitro using invadopodia and traction force assays based on polyacrylamide gels (PAAs) of different rigidities. Invasive and contractile properties of cancer cells can be correlated in vitro using invadopodia and traction force assays based on polyacrylamide gels (PAAs) of different rigidities. While some variations between the two assays exist, the protocol presented here provides a method for creating PAAs that can be used in both assays and are easily adaptable to the user's specific biological and technical needs.

  17. Preparation of Barley Storage Protein, Hordein, for Analytical Sodium Dodecyl Sulfate-Polyacrylamide Gel Electrophoresis

    DEFF Research Database (Denmark)

    Doll, Hans; Andersen, Bente

    1981-01-01

    The extraction, reduction, and alkylation of barley hordein for routine electrophoresis in sodium dodecyl sulfate-polyacrylamide gels were studied to set up a simple preparation procedure giving well-resolved bands in the electrophoresis gel. Hordein was extracted from single crushed seeds or flour...... by aqueous 50% propan-2-ol containing a Tris-borate buffer, pH 8.6. The presence of the buffer facilitates the consecutive complete reduction of the extracted protein in the alcohol. Reduction and alkylation in the buffer containing propan-2-ol give sharper bands in the electrophoresis than reduction...

  18. Highly sensitive fluorescent stain for detecting lipopolysaccharides in sodium dodecyl sulfate polyacrylamide gel electrophoresis.

    Science.gov (United States)

    Wang, Xu; Zhou, Ayi; Cai, Wanhui; Yu, Dongdong; Zhu, Zhongxin; Jiang, Chengxi; Jin, Litai

    2015-08-01

    A sensitive and simple technique was developed for the visualization of gel-separated lipopolysaccharides by using a hydrazide derivative, UGF202. As low as 0.5-1 ng total LPS could be detected by UGF202 stain, which is 2- and 16-fold more sensitive than that of the commonly used Pro-Q Emerald 300 and Keenan et al. developed silver stain, respectively. The results indicated that UGF202 stain could be a good choice for LPS determination in polyacrylamide gels. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  19. Detection of connexins in liver cells using sodiumdodecylsulfate polyacrylamide gel electrophoresis and immunoblot analysis

    Science.gov (United States)

    Willebrords, Joost; Maes, Michaël; Yanguas, Sara Crespo; Cogliati, Bruno; Vinken, Mathieu

    2016-01-01

    Summary Since connexin expression is partly regulated at the protein level, immunoblot analysis represents a frequently addressed technique in the connexin research field. The present chapter describes the set-up of an immunoblot procedure, including protein extraction and quantification from biological samples, gel electrophoresis, protein transfer and immunoblotting, which is optimized for analysis of connexins in liver tissue. In essence, proteins are separated on a polyacrylamide gel using sodiumdodecylsulfate followed by transfer of proteins on a nitrocellulose membrane. The latter allows specific detection of connexins with antibodies combined with revelation through enhanced chemiluminescence. PMID:27207285

  20. Resolving Acetylated and Phosphorylated Proteins by Neutral Urea Triton-Polyacrylamide Gel Electrophoresis, NUT-PAGE

    Science.gov (United States)

    Buehl, Christopher J.; Deng, Xiexiong; Liu, Mengyu; Hovde, Stacy; Xu, Xinjing; Kuo, Min-Hao

    2014-01-01

    Protein acetylation and phosphorylation can be key modifications that regulate both normal and pathological protein functions. Current gel systems used to analyze modified proteins require either expensive reagents or time–consuming second dimension electrophoresis. In this manuscript, we present a neutral pH gel system that allows the analysis of acetylated and phosphorylated proteins. This neutral pH urea Triton-polyacrylamide gel electrophoresis system, or NUT-PAGE, separates proteins based on their charge at pH 7 and generates discrete bands from each acetylated and phosphorylated species. In addition, the gel is composed of common and inexpensive laboratory reagents, and requires only a single dimension of electrophoresis. We are able to demonstrate the effectiveness of this system by analyzing phosphorylated species of an acidic protein, α-synuclein, and both acetylated and phosphorylated species of a basic protein, histone H3. NUT-PAGE thus provides a cost-effective alternative to resolving acetylated and phosphorylated proteins, and potentially proteins with other post-translational modifications that alter net charge. Method Summary Here we present a single-dimension neutral pH urea Triton-polyacrylamide gel electrophoresis (NUT-PAGE) system affording high-resolution separation of acetylated and phosphorylated proteins. PMID:25109292

  1. Gel electrophoresis of DNA partially denatured at the ends: what are the dominant conformations?

    Science.gov (United States)

    Sean, David; Slater, Gary W

    2013-03-01

    Gel electrophoresis of a partially denatured dsDNA fragment is studied using Langevin Dynamics computer simulations. For simplicity, the denatured ssDNA sections are placed at the ends of the fragment in a symmetrical fashion. A squid-like conformation is found to sometimes cause the fragment to completely block in the gel. In fact, this conformation is the principal cause of the steep reduction in mobility observed in the simulations. As the field is increased, it is found that the occurrence of this conformation dominates the migration dynamics. Although the squid conformation seems to be more stable at high fields, the field can eventually force the fragments to thread through the gel pores regardless. We qualitatively explore the behavior of this squid-like conformation across a range of fields and degrees of denaturation, and we discuss the relevance of our findings for TGGE. © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  2. Analysis of mucosal mucins separated by SDS-urea agarose polyacrylamide composite gel electrophoresis.

    Science.gov (United States)

    Issa, Samah M A; Schulz, Benjamin L; Packer, Nicolle H; Karlsson, Niclas G

    2011-12-01

    Efficient separation of mucins (200 kDa-2 MDa) was demonstrated using gradient SDS agarose/polyacrylamide composite gel electrophoresis (SDS-AgPAGE). Inclusion of urea (SDS-UAgPAGE) in the gels casting were shown to have no effect on the migration of mucins in the gel and allowed casting of gel at room temperature. This simplified the procedure for multiple casting of agarose polyacrylamide gradients and increased reproducibility of these gels. Hence, the implementation of urea makes the technique applicable for high throughput isolation and screening of mucin oligosaccharides by LC-MS after releasing the oligosaccharides from isolated, blotted mucin subpopulations. It was also shown that the urea addition had no effect on other supporting applications such as western and lectin blotting. In addition, identification of the mucin protein after tryptic digestion and LC-MS was possible and no protein carbamylation due to the presence of urea in the gel was detected. LC-MS software developed for metabolomic analysis was used for O-linked oligosaccharide detection and differential display of various mucin samples. Using this method, heterogeneous glycosylation of mucins and mucin-type molecules isolated by SDS-AgPAGE and SDS-UAgPAGE was shown to consist of more than 80 different components in a single band, and in the extreme cases, up to 300-500 components (MUC5B/AC from saliva and sputum and). Metabolomic software was also used to show that the migration of mucin isoforms within the gel is due to heterogeneous size distribution of the oligosaccharides, with the slower migrating bands enriched in high-molecular-weight oligosaccharides. Copyright © 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  3. Heat denaturation of soy glycinin. Structural characteristics in relation to aggregation and gel formation.

    NARCIS (Netherlands)

    Lakemond, C.M.M.

    2001-01-01

    key words: soy protein; glycinin; thermal stability; pH; ionic strength;genetic variant; solubility; gelationThe main aim of this thesis was to study structural changes of soy glycinin at different conditions (pH and ionic strength) during thermal denaturation and their effect on aggregation and gel

  4. Denaturing gradient gel electrophoresis analysis to study bacterial community structure in pockets of periodontitis patients

    NARCIS (Netherlands)

    Zijnge, V.; Harmsen, H.J.M.; Kleinfelder, J.W.; Rest, M.E. van der; Degener, J.E.; Welling, G.W.

    2003-01-01

    Bacteria are involved in the onset and progression of periodontitis. A promising molecular technique, denaturing gradient gel electrophoresis (DGGE), to study microbial population dynamics in the subgingival pocket is presented. Twenty-three samples were taken from the subgingival pockets of nine pa

  5. Investigating the fermentation of cocoa by correlating denaturing gradient gel electrophoresis profiles and near infrared spectra

    DEFF Research Database (Denmark)

    Nielsen, Dennis Sandris; Snitkjær, Pia; van der Berg, Franciscus Winfried J

    2008-01-01

    of the beans and the chemical processes inside the beans have been carried out previously. Recently it has been shown that Denaturing Gradient Gel Electrophoresis (DGGE) offers an efficient tool for monitoring the microbiological changes taking place during the fermentation of cocoa. Near Infrared (NIR...

  6. A simple remedy against artifactual double bands in denaturing gradient gel electrophoresis

    NARCIS (Netherlands)

    Janse, I.; Bok, J.M.; Zwart, G.

    2004-01-01

    Denaturant gradient gel electrophoresis (DGGE) is a widely used method for mutation analysis and for studies of microbial diversity. Particular combinations of target gene fragments and primers may give rise to erroneous DGGE profiles. We report on a very straightforward means to eliminate the

  7. Polyacrylamide gel electrophoresis-SDS as a tool to study myofibrillar proteins. A review.

    Directory of Open Access Journals (Sweden)

    Perez-Chabela, M. Lourdes

    2015-12-01

    Full Text Available Miofibrillar proteins are part of land and sea animals’ muscle. Nonetheless, even when muscle proteins are the same type of proteins, their structure, rigor mortis time, and biochemical process associated to muscle to meat conversion, are different among animal species. This review has the aim to describe the advantages of SDS-polyacrylamide gel electrophoresis (SDS-PAGE in the study of myofibrillar proteins structure, besides the influence of many parameters on this technique to obtain an electrophoretic profile. Applications of this technique as a diagnostic tool in the food science, ecology and health are described as well.

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

    Science.gov (United States)

    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.

  9. Deciphering metabolic networks by blue native polyacrylamide gel electrophoresis: A functional proteomic exploration

    Directory of Open Access Journals (Sweden)

    Christopher Auger

    2015-06-01

    Full Text Available Metabolism is the consortium of reactions within a cell which directs a variety of processes including energy synthesis, signalling and the behaviour of a biological system. Metabolic networks, and more specifically the activity of enzymes within them, provide an accurate status of how cellular information is being executed. The performance of these networks and their ability to siphon metabolites in a number of directions may be the difference between a healthy and diseased state. Blue native polyacrylamide gel electrophoresis (BN-PAGE, owing to its simplicity and wide-ranging applications, permits the inspection of these nodules. The separation of proteins and enzyme complexes in their native format enables the exploration of enzymatic activity in metabolic networks via in-gel assays. These are quick, specific, and amenable to further studies. This electrophoretic technology not only enables the visualization of enzymatic efficacy but reveals the crosstalk among enzymes and their interactions with other organellar partners.

  10. Polyacrylamide Gel Treatment of Antiretroviral Therapy-induced Facial Lipoatrophy in HIV Patients

    DEFF Research Database (Denmark)

    Mansor, Samreen; Breiting, Vibeke Bro; Dahlstrøm, Karin

    2011-01-01

    been used successfully in HIV patients abroad. This article describes the results of a Danish study. METHODS: Forty HIV patients recruited from two major referral hospitals in the capitol area of Copenhagen, Denmark, each received a series of PAAG gel injections (small deposits in several sessions......) with a 14-day interval. Patient satisfaction, injector's evaluation, evaluation by an external specialist in plastic surgery, and long-term aesthetic effect and complications were registered with follow-up until 2 years. RESULTS: All patients were very satisfied or satisfied with the result. The injector......BACKGROUND: Today, highly active antiretroviral therapy is lifesaving for most HIV-infected patients, but the treatment can result in facial lipoatrophy, which changes the face so radically that patients may develop severe psychological and social problems. Since 2001 polyacrylamide gel (PAAG) has...

  11. Improved detection of amylase activity by sodium dodecyl sulfate-polyacrylamide gel electrophoresis with copolymerized starch.

    Science.gov (United States)

    Martínez, T F; Alarcón, F J; Díaz-López, M; Moyano, F J

    2000-08-01

    An improved method, based on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) for detection of amylase activity is described. This method will allow better characterization of certain amylases than that obtained by the Davis technique. The main features of the technique are: (i) identification of amylase bands and molecular mass determination are possible in the same gel; (ii) the hydrolysis of copolymerized substrate during electrophoretic separation is prevented using very low temperatures instead of inactivating agents such as chelating agents; and (iii) the technique is applicable to reveal amylase activity in a wide range of biological samples. The method is not useful for enzymes sensitive to SDS and for high molecular mass amylases.

  12. MALDI analysis of proteins after extraction from dissolvable ethylene glycol diacrylate cross-linked polyacrylamide gels.

    Science.gov (United States)

    Papasotiriou, Dimitrios G; Markoutsa, Stavroula; Gorka, Jan; Schleiff, Enrico; Karas, Michael; Meyer, Bjoern

    2013-09-01

    Although the extraction of intact proteins from polyacrylamide gels followed by mass spectrometric molecular mass determination has been shown to be efficient, there is room for alternative approaches. Our study evaluates ethylene glycol diacrylate, a cleavable cross-linking agent used for a new type of dissolvable gels. It attains an ester linkage that can be hydrolyzed in alkali conditions. The separation performance of the new gel system was tested by 1D and 2D SDS-PAGE using the outer chloroplast envelope of Pisum sativum as well as a soluble protein fraction of human lymphocytes, respectively. Gel spot staining (CBB), dissolving, and extracting were conducted using a custom-developed workflow. It includes protein extraction with an ammonia-SDS buffer followed by methanol treatment to remove acrylamide filaments. Necessary purification for MALDI-TOF analysis was implemented using methanol-chloroform precipitation and perfusion HPLC. Both cleaning procedures were applied to several standard proteins of different molecular weight as well as 'real' biological samples (8-75 kDa). The protein amounts, which had to be loaded on the gel to detect a peak in MALDI-TOF MS, were in the range of 0.1 to 5 μg, and the required amount increased with increasing mass.

  13. Deposition of Ibuprofen Crystals on Hydroxypropyl Cellulose/Polyacrylamide Gel: Experimental and Mathematic Modeling Releasing

    Directory of Open Access Journals (Sweden)

    Claudia Alicia Castillo-Miranda

    2016-01-01

    Full Text Available The crystallization of nonsteroidal anti-inflammatory drug [2-(4-isobutyl-phenyl propionic acid] ibuprofen (IBP on a hydroxypropyl cellulose (HPC and polyacrylamide (PAAm gel was studied as well as the release kinetics of the drug. The IBP was crystallized on the gel surface of HPC/PAAm. It had a prismatic shape and the growth was made in an aqueous medium; the crystallinity grade of the gels HPC/PAAm and HPC/PAAm-IBU increased to 68% and to 58%, respectively. The release of IBP is performed by two means: by a non-Fickian diffusion process and by relaxation of the chains of the gel; without regard to temperature and the diffusion media, this correlates with the lower critical solution temperature (LCST of the proposed gel. This polymer matrix provides an option for releasing nonsteroidal anti-inflammatory drugs in a temperature range of 35–39°C. Korsmeyer and Peppas mathematical model was simulated for data releases, statistically significant at 95% confidence level.

  14. Characterization of immunoreactive proteins of Setaria cervi isolated by preparative polyacrylamide gel electrophoresis.

    Science.gov (United States)

    Priyadarshi, Priyanka; Dravid, Piyush; Sheikh, Inayat Hussain; Saxena, Sunita; Tandon, Ashish; Kaushal, Deep C; Ali, Shakir; Kaushal, Nuzhat A

    2017-03-01

    Filarial parasites are complex mixtures of antigenic proteins and characterization of these antigenic molecules is essential to identify the diagnostically important filaria-specific antigens. In the present study, we have fractionated the somatic extracts from adults of Setaria cervi (bovine filarial parasite) on preparative SDS-polyacrylamide gel electrophoresis and tested the immunoreactivity of the separated gel fractions with polyclonal antibodies against filarial excretory-secretory antigens as well as filarial patients sera. The SDS-PAGE analysis of gel eluted fractions revealed 1 protein band in F-1 fraction, 2 protein bands in F-2 fraction and 2-3 protein bands in all other fractions (F3- F11). Seven gel eluted fractions (F1, F2, F3, F4, F5, F6 and F11) showed high ELISA reactivity with the polyclonal antibody (against excretory-secretory antigen) and four of these fractions (F-1, F-2, F3 and F6) exhibited high ELISA reactivity with antibodies present in filarial patient sera. The reactivities of the gel fractions (F1 and F2), recognized by filarial patients sera, were also tested with the monoclonal antibody (detecting the filarial circulating antigen). The F1 and F2 gel eluted fractions were found to have the target antigen of monoclonal antibody as evident by high reactivity with the monoclonal antibody in ELISA and immunoblotting. The S. cervi gel eluted F1 fraction (containing single antigen) could detect antibodies in filarial patients sera and not in non-filarial sera thereby suggesting its usefulness for specific serodiagnosis of human filariasis.

  15. Purification of Peptide Components including Melittin from Bee Venom using gel filtration chromatography and propionic acid/urea polyacrylamide gel electrophoresis

    Directory of Open Access Journals (Sweden)

    Young Chon Choi

    2006-06-01

    Full Text Available Objectives : This study was conducted to carry out Purification of Melittin and other peptide components from Bee Venom using gel filtration chromatography and propionic acid/urea polyacrylamide gel electrophoresis Methods : Melittin and other peptide components were separated from bee venom by using gel filtration chromatography on Sephadex G-50 column in 0.05M ammonium acetate buffer. Results : Melittin and other peptide components were separated from bee venom by using gel filtration chromatography on Sephadex G-50 column in 0.05M ammonium acetate buffer. The fractions obtained from gel filtration chromatography was analyzed by using SDS-PAGE and propionic acid/urea polyacrylamide gel electrophoresis. The melittin obtained from the gel filtration contained residual amount of phospholipase A2 and a protein with molecular weight of 6,000. The contaminating proteins were removed by the second gel filtration chromatography. Conclusion : Gel filtration chromatography and propionic acid/urea polyacrylamide gel electrophoresis are useful to separate peptide components including melittin from bee venom.

  16. Detection of Rifampin Resistance in Mycobacterium tuberculosis by Double Gradient-Denaturing Gradient Gel Electrophoresis

    Science.gov (United States)

    Scarpellini, Paolo; Braglia, Sergio; Carrera, Paola; Cedri, Maura; Cichero, Paola; Colombo, Alessia; Crucianelli, Rosella; Gori, Andrea; Ferrari, Maurizio; Lazzarin, Adriano

    1999-01-01

    We applied double gradient-denaturing gradient gel electrophoresis (DG-DGGE) for the rapid detection of rifampin (RMP) resistance from rpoB PCR products of Mycobacterium tuberculosis isolates and clinical samples. The results of this method were fully concordant with those of DNA sequencing and susceptibility testing analyses. DG-DGGE is a valid alternative to the other methods of detecting mutations for predicting RMP resistance. PMID:10508043

  17. Polyacrylamide Gel-Entrapped Maltase: An Excellent Design of Using Maltase in Continuous Industrial Processes.

    Science.gov (United States)

    Nawaz, Muhammad Asif; Aman, Afsheen; Rehman, Haneef Ur; Bibi, Zainab; Ansari, Asma; Islam, Ziaul; Khan, Ishtiaq Ahmad; Qader, Shah Ali Ul

    2016-06-01

    Bacterial maltase catalyzes the hydrolysis of maltose and is known as one of the most significant hydrolases. It has several applications in different industrial processes but widely used in food fermentation technology and alcohol production. In the current study, entrapment technique was comprehensively examined using polyacrylamide gel as a matrix support to improve the stability and catalytic efficiency of maltase for continuous use. Maximum entrapment yield of maltase was achieved at 10 % polyacrylamide concentration with 3.0-mm bead size. Optimized conditions indicated an increase in the reaction temperature from 45 to 55 °C after maltase entrapment while no change was observed in the reaction time and pH. An increase in the K m value of entrapped maltase was attained whereas V max value decreased from 8411.0 to 6813.0 U ml(-1) min(-1) with reference to its free counterpart. Entrapped maltase showed remarkable thermal stability and retained 16 % activity at 70 °C even after 120.0 min. Entrapped maltase also exhibited excellent recycling efficiency up to eight consecutive reaction cycles. With respect to economic feasibility, entrapped maltase indicates its high potential to be used in various biotechnological applications.

  18. Synthesis, Characterization and Electromagnetic Studies on Nanocrystalline Nickel Zinc Ferrite by Polyacrylamide Gel

    Institute of Scientific and Technical Information of China (English)

    Ruiting MA; Yi WANG; Yanwen TIAN; Chunli ZHANG; Xikun LI

    2008-01-01

    Nanocrystalline nickel zinc ferrite powders (NixZn1-xFe2O4, A for x=0, B for x=0.2, C for x=0.5, D for x= 0.8 and E for x= 1) were synthesized by polyacrylamide gel method. X-ray diffraction (XRD), transmission electron microscopy (TEM) and wave-guide were used to characterize the composition. The XRD results show that the dried gel powders are amorphous, and the characteristic peaks of the spinel Ni0.5Zn0.5Fe2O4 appear after the gel is calcined at 400℃ for 1 h. When the calcining temperatures are 600 and 800℃, the average grain sizes are identified by TEM to be 10 and 30 nm, respectively. The NixZn1-xFe2O4 powders have both.dielectric loss and magnetic loss in the frequency range of 8.2-11.0GHz. With the increase of Ni2+ ions content, the dielectric parameters (ε′) and permeability (u′) of the NixZn1-xFe2O4 powders decrease while the dielectric loss (ε″), magnetic loss (u″) and the reflection loss increase.

  19. Functional Characterization of Reductive Dehalogenases by Using Blue Native Polyacrylamide Gel Electrophoresis

    Science.gov (United States)

    Tang, Shuiquan; Chan, Winnie W. M.; Fletcher, Kelly E.; Seifert, Jana; Liang, Xiaoming; Löffler, Frank E.; Adrian, Lorenz

    2013-01-01

    Dehalococcoides mccartyi strains are obligate organohalide-respiring bacteria harboring multiple distinct reductive dehalogenase (RDase) genes within their genomes. A major challenge is to identify substrates for the enzymes encoded by these RDase genes. We demonstrate an approach that involves blue native polyacrylamide gel electrophoresis (BN-PAGE) followed by enzyme activity assays with gel slices and subsequent identification of proteins in gel slices using liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS). RDase expression was investigated in cultures of Dehalococcoides mccartyi strain BAV1 and in the KB-1 consortium growing on chlorinated ethenes and 1,2-dichloroethane. In cultures of strain BAV1, BvcA was the only RDase detected, revealing that this enzyme catalyzes the dechlorination not only of vinyl chloride, but also of all dichloroethene isomers and 1,2-dichloroethane. In cultures of consortium KB-1, five distinct Dehalococcoides RDases and one Geobacter RDase were expressed under the conditions tested. Three of the five RDases included orthologs to the previously identified chlorinated ethene-dechlorinating enzymes VcrA, BvcA, and TceA. This study revealed substrate promiscuity for these three enzymes and provides a path forward to further explore the largely unknown RDase protein family. PMID:23204411

  20. Magnetic resonance imaging on complications of breast augmentation with injected hydrophilic polyacrylamide gel

    Institute of Scientific and Technical Information of China (English)

    XU Li-ying; KONG Xiang-quan; TIAN Zhi-xiong; QIU Da-sheng

    2006-01-01

    @@ The injection augmentation mammaplasty for cosmetic purpose has been popular recently in China. Two kinds of injectable material are used clinically, autologous fat and biomaterial. The fat injection for breast augmentation is in question with the major problems of progressive fat re-absorption,microcalcification, and fat liquefaction.1 Now, the principal alloplastic biomaterial for injection augmentation mammaplasty in China is hydrophilic polyacrylamide gel (HPAAG). Although thousands of breasts have been augmented with HPAAG and it seems to be a good biocompatible material, some complications develop after HPAAG injection augmentation mammaplasty.2-4 The patients had to undergo surgery to remove the injected HPAAG and associated lesions. Ultrasonography or magnetic resonance imaging (MRI) should be taken preoperatively to demonstrate the distribution of injected HPAAG and associated lesions.5 In this report, the diagnostic value and clinical significance of MRI on the complications of HPAAG breast augmentation were discussed.

  1. Combined alcian blue and silver staining of subnanogram quantities of proteoglycans and glycosaminoglycans in sodium dodecyl sulfate-polyacrylamide gels

    DEFF Research Database (Denmark)

    Møller, H J; Heinegård, D; Poulsen, J H

    1993-01-01

    Proteoglycans stain weakly in polyacrylamide gels by traditional protein stains such as coomassie brilliant blue or silver. In the present work preparations of large aggregating proteoglycan from human articular cartilage were used to evaluate a convenient staining method based on successive stai...

  2. Breast Fibromatosis after Hydrophilic Polyacrylamide Gel Injection for Breast Augmentation: a Case Report and Review of the Literature

    Institute of Scientific and Technical Information of China (English)

    Xiao Long; Qun Qiao

    2011-01-01

    @@ BREAST fibromatosis is a rare kind of lesion.The average incidence is about 2-4 per million every year.1 So far there have been about 100 cases reported altogether.2 In this report, we describe a case of breast fibromatosis developed after hydrophilic polyacrylamide gel (HPG) injection for breast augmenta-Received for publication December 10, 2010.

  3. Imaging findings of breast augmentation with injected hydrophilic polyacrylamide gel: Patient reports and literature review

    Energy Technology Data Exchange (ETDEWEB)

    Khedher, Najoua Ben, E-mail: nbenkhedher@gmail.com [Centre d' Investigation et de Recherche Diagnostique des maladies du sein (CRID), Centre hospitalier de l' Universite de Montreal, Hotel-Dieu, 3840 rue Saint-Urbain, Montreal (Quebec), H2W 1T8 (Canada); David, Julie, E-mail: jdavid@vl.videotron.ca [Centre d' Investigation et de Recherche Diagnostique des maladies du sein (CRID), Centre hospitalier de l' Universite de Montreal, Hotel-Dieu, 3840 rue Saint-Urbain, Montreal (Quebec), H2W 1T8 (Canada); Trop, Isabelle, E-mail: itrop@yahoo.com [Centre d' Investigation et de Recherche Diagnostique des maladies du sein (CRID), Centre hospitalier de l' Universite de Montreal, Hotel-Dieu, 3840 rue Saint-Urbain, Montreal (Quebec), H2W 1T8 (Canada); Drouin, Suzanne, E-mail: suedrouin@sympatico.ca [Clinique Radiologique Leger et associees, 1851 rue Sherbrooke Est, Bureau 201, Montreal (Quebec), H2K 4L5 (Canada); Peloquin, Laurence, E-mail: laurence.peloquin@gmail.com [Centre d' Investigation et de Recherche Diagnostique des maladies du sein (CRID), Centre hospitalier de l' Universite de Montreal, Hotel-Dieu, 3840 rue Saint-Urbain, Montreal (Quebec), H2W 1T8 (Canada); Lalonde, Lucie, E-mail: lalucie@sympatico.ca [Centre d' Investigation et de Recherche Diagnostique des maladies du sein (CRID), Centre hospitalier de l' Universite de Montreal, Hotel-Dieu, 3840 rue Saint-Urbain, Montreal (Quebec), H2W 1T8 (Canada)

    2011-04-15

    Hydrophilic polyacrylamide gel (PAAG) is a nonresorbable soft tissue filler that has been used as implant material for breast augmentation in some countries, particularly from the Asian continent. Many complications associated with hydrogel use have been reported in the clinical literature including inflammation, persistent mastodynia, formation of multiple lumps, poor cosmetic results, glandular atrophy, and significant spread of hydrogel into the surrounding tissue. Data on long-term toxicity is currently unavailable. The radiologic features of PAAG injection mammoplasty frequently constitute a diagnostic challenge for radiologists. Indeed, the imaging appearances of uncomplicated PAAG implants may mimic conventional implants on mammography, sonography and MRI, with some distinguishing features. The location and local spread of the injected PAAG, and the eventual detection of local inflammation, are best evaluated by ultrasonography and especially MRI, considered the most sensitive technique for assessment of PAAG mammoplasty. MRI clearly depicts the volume and the distribution of gel within the breast; contrast medium enhancement allows delineation of areas of inflammation and infection. It is important to be familiar with the spectrum of imaging findings in order to make an accurate diagnosis and offer proper management. This paper aims to review the normal and abnormal mammographic, sonographic, and MR imaging characteristics of PAAG augmentation mammoplasty through presented patient reviews of three women having undergone direct PAAG injection.

  4. Growth of BeO Nanograins Synthesized by Polyacrylamide Gel Route

    Institute of Scientific and Technical Information of China (English)

    Xiaofeng Wang; Richu Wang; Chaoqun Peng; Tingting Li; Bing Liu

    2011-01-01

    BeO nanoparticles were synthesized by polyacrylamide gel route. The effects of the processing parameters on the morphology and size of the synthesized BeO nanoparticles were investigated. The calcination temperature of the gel precursor containing beryllium sulfate was determined by thermogravimetry and differential scanning calorimetry (TG-DSC), which is around 690℃ and 160℃ lower than the general temperature. X-ray diffractometry (XRD), transmission electron microscopy (TEM), and specific surface area measurements (BET) showed that the synthesized nanoparticles under 700℃ were pure, globular and about ~5-20 nm with narrow distribution. Interestingly, the nanograins coalesced and grew under higher calcination temperatures and longer calcination time. The influence of calcination temperature on the morphology and growth behavior is greater than that of its duration. The activation energy for grain growth was estimated to be 24.53 kJ/mol, and the dominant growth mechanism was most likely to be related to the vapor transport in pore control mode and grain-rotation-induced grain coalescence (GRIGC) mechanism.

  5. Serum alkaline phosphatase isoenzymes in laboratory beagle dogs detected by polyacrylamide-gel disk electrophoresis.

    Science.gov (United States)

    Hatayama, Kazuhisa; Nishihara, Yoshito; Kimura, Sayaka; Goto, Ken; Nakamura, Daichi; Wakita, Atsushi; Urasoko, Yoshinaka

    2011-10-01

    Serum alkaline phosphatase (ALP) activity is frequently measured in toxicity studies. Itoh et al. (2002) reported that a commercially available polyacrylamide-gel (PAG) disk electrophoresis kit used in humans (AlkPhor System, Jokoh Co., Ltd., Tokyo, Japan) for identifying serum ALP isoenzymes was useful for veterinary clinicopathological diagnosis in mongrel dogs. In the present study, based on the report of Itoh et al. (2002), we tried to expand the application range of this kit to laboratory beagle dogs which are commonly used in toxicity studies. In order to identify the origin of each ALP isoenzyme, tissue ALP extracts from the liver, bone and small intestine and serum samples were treated with neuraminidase, anti-small intestinal ALP antibody, ALP inhibitor levamisole and/or wheat germ agglutinin (WGA). The main serum ALP isoenzymes in 5-month-old intact beagle dogs were bone-derived (bone and atypical ALP: corresponding to human variant bone ALP) and they tended to decrease with age. However, liver-derived ALP isoenzyme greatly increased in the serum of cholestasis model dogs. The cholestasis model dogs also had a large molecular ALP detected in the resolving gel. This ALP could be originated from intestinal ALP or corticosteroid-induced ALP (CALP), because the activity remained even after levamisole inhibition. CALP was observed in intact laboratory beagle dogs with individual differences. These results suggest that the present method is a useful tool for detecting serum ALP isoenzymes in laboratory beagle dogs and concomitant levamisole inhibition with another gel is applicable for the evaluation of organ toxicity.

  6. Agarose and Polyacrylamide Gel Electrophoresis Methods for Molecular Mass Analysis of 5–500 kDa Hyaluronan

    Science.gov (United States)

    Bhilocha, Shardul; Amin, Ripal; Pandya, Monika; Yuan, Han; Tank, Mihir; LoBello, Jaclyn; Shytuhina, Anastasia; Wang, Wenlan; Wisniewski, Hans-Georg; de la Motte, Carol; Cowman, Mary K.

    2011-01-01

    Agarose and polyacrylamide gel electrophoresis systems for the molecular mass-dependent separation of hyaluronan (HA) in the size range of approximately 5–500 kDa have been investigated. For agarose-based systems, the suitability of different agarose types, agarose concentrations, and buffers systems were determined. Using chemoenzymatically synthesized HA standards of low polydispersity, the molecular mass range was determined for each gel composition, over which the relationship between HA mobility and logarithm of the molecular mass was linear. Excellent linear calibration was obtained for HA molecular mass as low as approximately 9 kDa in agarose gels. For higher resolution separation, and for extension to molecular masses as low as approximately 5 kDa, gradient polyacrylamide gels were superior. Densitometric scanning of stained gels allowed analysis of the range of molecular masses present in a sample, and calculation of weight-average and number-average values. The methods were validated for polydisperse HA samples with viscosity-average molecular masses of 112, 59, 37, and 22 kDa, at sample loads of 0.5 µg (for polyacrylamide) to 2.5 µg (for agarose). Use of the methods for electrophoretic mobility shift assays was demonstrated for binding of the HA-binding region of aggrecan (recombinant human aggrecan G1-IGD-G2 domains) to a 150 kDa HA standard. PMID:21684248

  7. Screening for mutations in the uroporphyrinogen decarboxylase gene using denaturing gradient gel electrophoresis

    DEFF Research Database (Denmark)

    Christiansen, L; Ged, C; Hombrados, I

    1999-01-01

    The two porphyrias, familial porphyria cutanea tarda (fPCT) and hepatoerythropoietic porphyria (HEP), are associated with mutations in the gene encoding the enzyme uroporphyrinogen decarboxylase (UROD). Several mutations, most of which are private, have been identified in HEP and fPCT patients......, confirming the heterogeneity of the underlying genetic defects of these diseases. We have established a denaturing gradient gel electrophoresis (DGGE) assay for mutation detection in the UROD gene, enabling the simultaneous screening for known and unknown mutations. The established assay has proved able...

  8. Polyacrylamide Gel-Entrapped Fungal Manganese Peroxidase with Enhanced Catalytic, Stability and Reusability Characteristics.

    Science.gov (United States)

    Bilal, Muhammad; Asgher, Muhammad; Iqbal, Hafiz M N

    2016-07-19

    In the present study, polyacrylamide gel (PAG) was utilized as bolster material for the immobilization of in-house extracted and partially purified manganese peroxidase (MnP) through entrapment technique. The entrapment technique impelled incredibly compelling MnP immobilization (87.3±3.3%) and conferred remarkable stability to the enzyme (37.2±2.4%) following two months of storage at 4°C. The PAG-assisted immobilization expanded the reaction time of MnP after 10 min of response when contrasted with a partially purified free MnP counterpart, which demonstrated the highest activity after 5.0 min. Following PAG-assisted immobilization, an improvement in the optimal temperature and a chemical (an alkaline) shift in the pH optima of MnP were recorded. Moreover, a significant enhancement in the thermo-stability was also observed after immobilization. After 72 h, PAG-entrapped-MnP exhibited 41.2% residual activity at 50°C, whereas the free counterpart lost its activity completely under the same conditions. Furthermore, the PAG-entrapped-MnP also showed an excellent recycling efficiency and retained more than 50% of its initial activity after five consecutive reaction cycles. In conclusion, owing to the economic feasibility, carrier-supported MnP may be a promising candidate for various applications in different sectors of the modern world.

  9. Mechanosensitivity of astrocytes on optimized polyacrylamide gels analyzed by quantitative morphometry

    Science.gov (United States)

    Moshayedi, Pouria; Costa, Luciano da F.; Christ, Andreas; Lacour, Stephanie P.; Fawcett, James; Guck, Jochen; Franze, Kristian

    2010-05-01

    Cells are able to detect and respond to mechanical cues from their environment. Previous studies have investigated this mechanosensitivity on various cell types, including neural cells such as astrocytes. In this study, we have carefully optimized polyacrylamide gels, commonly used as compliant growth substrates, considering their homogeneity in surface topography, mechanical properties, and coating density, and identified several potential pitfalls for the purpose of mechanosensitivity studies. The resulting astrocyte response to growth on substrates with shear storage moduli of G' = 100 Pa and G' = 10 kPa was then evaluated as a function of coating density of poly-D-lysine using quantitative morphometric analysis. Astrocytes cultured on stiff substrates showed significantly increased perimeter, area, diameter, elongation, number of extremities and overall complexity if compared to those cultured on compliant substrates. A statistically significant difference in the overall morphological score was confirmed with an artificial intelligence-based shape analysis. The dependence of the cells' morphology on PDL coating density seemed to be weak compared to the effect of the substrate stiffness and was slightly biphasic, with a maximum at 10-100 µg ml - 1 PDL concentration. Our finding suggests that the compliance of the surrounding tissue in vivo may influence astrocyte morphology and behavior.

  10. Culture-independent analysis of probiotic products by denaturing gradient gel electrophoresis.

    Science.gov (United States)

    Temmerman, R; Scheirlinck, I; Huys, G; Swings, J

    2003-01-01

    In order to obtain functional and safe probiotic products for human consumption, fast and reliable quality control of these products is crucial. Currently, analysis of most probiotics is still based on culture-dependent methods involving the use of specific isolation media and identification of a limited number of isolates, which makes this approach relatively insensitive, laborious, and time-consuming. In this study, a collection of 10 probiotic products, including four dairy products, one fruit drink, and five freeze-dried products, were subjected to microbial analysis by using a culture-independent approach, and the results were compared with the results of a conventional culture-dependent analysis. The culture-independent approach involved extraction of total bacterial DNA directly from the product, PCR amplification of the V3 region of the 16S ribosomal DNA, and separation of the amplicons on a denaturing gradient gel. Digital capturing and processing of denaturing gradient gel electrophoresis (DGGE) band patterns allowed direct identification of the amplicons at the species level. This whole culture-independent approach can be performed in less than 30 h. Compared with culture-dependent analysis, the DGGE approach was found to have a much higher sensitivity for detection of microbial strains in probiotic products in a fast, reliable, and reproducible manner. Unfortunately, as reported in previous studies in which the culture-dependent approach was used, a rather high percentage of probiotic products suffered from incorrect labeling and yielded low bacterial counts, which may decrease their probiotic potential.

  11. Influence of pH and Ionic Strength on Heat-Induced Formation and Rheological Properties of Soy Protein Gels in Relation to Denaturation and Their Protein Compositions

    NARCIS (Netherlands)

    Renkema, J.M.S.; Gruppen, H.; Vliet, van T.

    2002-01-01

    The influence of pH and ionic strength on gel formation and gel properties of soy protein isolate (SPI) in relation to denaturation and protein aggregation/precipitation was studied. Denaturation proved to be a prerequisite for gel formation under all conditions of pH and ionic strength studied.

  12. Fiber based optical tweezers for simultaneous in situ force exertion and measurements in a 3D polyacrylamide gel compartment.

    Science.gov (United States)

    Ti, Chaoyang; Thomas, Gawain M; Ren, Yundong; Zhang, Rui; Wen, Qi; Liu, Yuxiang

    2015-07-01

    Optical tweezers play an important role in biological applications. However, it is difficult for traditional optical tweezers based on objective lenses to work in a three-dimensional (3D) solid far away from the substrate. In this work, we develop a fiber based optical trapping system, namely inclined dual fiber optical tweezers, that can simultaneously apply and measure forces both in water and in a 3D polyacrylamide gel matrix. In addition, we demonstrate in situ, non-invasive characterization of local mechanical properties of polyacrylamide gel by measurements on an embedded bead. The fiber optical tweezers measurements agree well with those of atomic force microscopy (AFM). The inclined dual fiber optical tweezers provide a promising and versatile tool for cell mechanics study in 3D environments.

  13. Method for the typing of Clostridium difficile based on polyacrylamide gel electrophoresis of (/sup 35/S)methionine-labeled proteins

    Energy Technology Data Exchange (ETDEWEB)

    Tabaqchali, S.; O' Farrell, S.; Holland, D.; Silman, R.

    1986-01-01

    A typing method for Clostridium difficile based on the incorporation of (/sup 35/S)methionine into cellular proteins, their separation by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and their visualization by autoradiography is described. On analysis of the radiolabeled-protein profiles, nine distinct groups were observed (A to E and W to Z). The method, which is simple, reproducible, and readily expandable, has been applied in epidemiological studies to demonstrate cross-infection and hospital acquisition of C. difficile.

  14. Simple and Specific Dual-Wavelength Excitable Dye Staining for Glycoprotein Detection in Polyacrylamide Gels and Its Application in Glycoproteomics

    Directory of Open Access Journals (Sweden)

    Yu-Hsuan Chiang

    2011-01-01

    Full Text Available In this study, a commercially available fluorescent dye, Lissamine rhodamine B sulfonyl hydrazine (LRSH, was designed to specifically stain the glycoproteins in polyacrylamide gels. Through the periodate/Schiff base mechanism, the fluorescent dye readily attaches to glycoproteins and the fluorescence can be simultaneously observed under either 305 nm or 532 nm excitation therefore, the dye-stained glycoproteins can be detected under a regular UV transilluminator or a more elegant laser-based gel scanner. The specificity and detection limit were examined using a standard protein mixture in polyacrylamide gels in this study. The application of this glycoprotein stain dye was further demonstrated using pregnancy urine samples. The fluorescent spots were further digested in gel and their identities confirmed through LC-MS/MS analysis and database searching. In addition, the N-glycosylation sites of LRSH-labeled uromodulin were readily mapped via in-gel PNGaseF deglycosylation and LC-MS/MS analysis, which indicated that this fluorescent dye labeling does not interfere with enzymatic deglycosylation. Hence, the application of this simple and specific dual-wavelength excitable dye staining in current glycoproteome research is promising.

  15. Linear dichroism studies of tryptophanase and its quasisubstrate complexes oriented in polyacrylamide gel.

    Science.gov (United States)

    Zakomirdina, L N; Sakharova, I S; Torchinsky, Y M

    1990-10-01

    Tryptophanase from Escherichia coli was oriented in a compressed slab of polyacrylamide gel and its linear dichroism (LD) and absorption spectra have been measured. The free enzyme displays four LD bands at 305, 340, 425 and 490 nm. Two bands at 340 and 425 nm belong to the internal coenzyme-lysine aldimine. The 305-nm band apparently belongs to an aromatic amino acid residue. The 490-nm band disappears after treatment with NaBH4 or after incubation with L-alanine and subsequent dialysis. It is suggested that the 490-nm band belongs to a quinonoid enzyme subform. The reaction of tryptophanase with threo-3-phenyl-DL-serine, L-threonine and D-alanine leads to formation of an external aldimine with an intense absorption band at 420-425 nm. The values of reduced LD (delta A/A) in this band strongly differ from that in the 420-nm band of the free enzyme. The LD value of the complex with D-alanine is intermediate between those of the free enzyme and the complex with 3-phenylserine. In the presence of indole the complex with D-alanine displays the same LD as that observed with 3-phenylserine. The reaction of tryptophanase with L-alanine or oxindolyl-L-alanine leads to formation of a quinonoid intermediate with an absorption band near 500 nm. The LD value in this band is close to that of an external aldimine with L-threonine. It is concluded that reorientations of the coenzyme occur in the course of the tryptophanase reaction.

  16. Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE of urinary protein in acute kidney injury

    Directory of Open Access Journals (Sweden)

    Sufi M Suhail

    2011-01-01

    Full Text Available Recent experimental and clinical studies have shown the importance of urinary proteomics in acute kidney injury (AKI. We analyzed the protein in urine of patients with clinical AKI using sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE for its diagnostic value, and followed them up for 40 months to evaluate prognosis. Urine from 31 consecutive cases of AKI was analyzed with SDS-PAGE to determine the low, middle and high molecular weight proteins. Fractional excretion of sodium (FENa was estimated from serum and urine creatinine and sodium (Na. The cases were followed-up for 40 months from the end of the recruitment of study cases. Glomerular protein was higher in the hematuria group when compared with the non-hematuria group (P <0.04 and in the AKI group than in the acute on chronic renal failure (AKI-on-CRF group (P <0.002. Tubular protein was higher in the AKI-on-CRF group (P <0.003 than in the AKI group. Tubular protein correlated with FENa in groups with diabetes mellitus (DM, AKI-on-CRF, and without hematuria (P <0.03, P <0.02 and P <0.004, respectively. Pattern of protein did not differ between groups with and without DM and clinical acute tubular necrosis (ATN. At the end of 40 months follow-up, category with predominantly glomerular protein progressed to chronic renal failure (CRF or end-stage renal failure in higher proportion (P <0.05. In clinical AKI, we observed that glomerular protein dominated in cases with glomerular insult, as indicated by hematuria. Tubular protein was common in the study cases with CRF, DM and cases without hematuria. This indicates tubulo-interstitial injury for AKI in these cases. Patients with predominantly glomerular protein had an adverse outcome.

  17. [Effect of physico-chemical properties of polyacrylamide gel media on the growth of Escherichia coli colonies].

    Science.gov (United States)

    Rodin, V B; Bulatova, R F; Domotenko, L V; Inkovskaia, T E; Artiukhin, V I

    1992-01-01

    We studied the growth of Escherichia coli LE-392 colonies on polyacrylamide gels (PAAG) depending on the physico-chemical properties of the latter, i.e. polymer concentration in the gel, swelling degree, bound water content (fm), spin-lattice relaxation and spin-spin relaxation times of water molecule protons, and modulus of elasticity (G0). S- or R-type colonies formed depending on gel properties; the diametral growth rate of S colonies was 3 times less compared with that on the control agar medium (Tryptose broth). The procedure is proposed for preparation of PAAG which rules out syneresis. Functional relations between the polymer concentrations in uniformly swelling gels and concentrations of copolymers in the reaction mixture, fm and G0 were revealed. The fm and G0 parameters can be used for controlling the quality of PAAG.

  18. New Primers for Denaturing Gradient Gel Electrophoresis Analysis of Nitrate-Reducing Bacterial Community in Soil

    Institute of Scientific and Technical Information of China (English)

    R.PASTORELLI; R.PICCOLO; S.SIMONCINI; S.LANDI

    2013-01-01

    The narG gene is frequently used as a molecular marker for bacterial nitrate-reducing community analysis.In this study,a new set of primers targeting the narG gene was designed and applied to semi-nested polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) assay.The potential of the new primers was verified on DNA directly extracted from soils from five different experimental sites distributed in Central and Southern Italy.Specificity of the primers was determined by excision,amplification,and sequencing of bands resolved by DGGE.A phylogenetic analysis showed the correlation between the sequences retrieved from the soils studied and the narG sequences from β and γ-Proteobacteria.These primers expanded the existing molecular tools for ecological study on the size and diversity of nitrate-reducing bacterial community in soil.

  19. Agarose gel purification of PCR products for denaturing gradient gel electrophoresis results in GC-clamp deletion.

    Science.gov (United States)

    Sun, Guowei; Xiao, Jinzhou; Lu, Man; Wang, Hongming; Chen, Xiaobing; Yu, Yongxin; Pan, Yingjie; Wang, Yongjie

    2015-01-01

    The 16S ribosomal RNA (rRNA) gene of marine archaeal samples was amplified using a nested PCR approach, and the V3 region of 16S rRNA gene of crab gut microbiota (CGM) was amplified using the V3 universal primer pair with a guanine and cytosine (GC)-clamp. Unpurified PCR products (UPPs), products purified from reaction solution (PPFSs), and products purified from gel (PPFGs) of above two DNA samples were used for denaturing gradient gel electrophoresis (DGGE) analysis, respectively. In contrast to almost identical band patterns shared by both the UPP and PPFS, the PPFGs were barely observed on the DGGE gel for both the marine archaea and CGM samples. Both PPFS and PPFG of CGM V3 regions were subjected to cloning. A small amount of positive clones was obtained for PPFS, but no positive clones were observed for PPFG. The melt curve and direct sequencing analysis of PPFS and PPFG of E. coli V3 region indicated that the Tm value of PPFG (82.35 ± 0.19 °C) was less than that of PPFS (83.81 ± 0.11 °C), and the number of shorter GC-clamps was significant higher in PPFG than in PPFS. The ultraviolet exposure experiment indicated that the ultraviolet was not responsible for the deletion of the GC-clamps. We conclude that the gel purification method is not suitable for DGGE PCR products or even other GC-rich DNA samples.

  20. Microbial Community Structure of Korean Cabbage Kimchi and Ingredients with Denaturing Gradient Gel Electrophoresis.

    Science.gov (United States)

    Hong, Sung Wook; Choi, Yun-Jeong; Lee, Hae-Won; Yang, Ji-Hee; Lee, Mi-Ai

    2016-06-28

    Kimchi is a traditional Korean fermented vegetable food, the production of which involves brining of Korean cabbage, blending with various other ingredients (red pepper powder, garlic, ginger, salt-pickled seafood, etc.), and fermentation. Recently, kimchi has also become popular in the Western world because of its unique taste and beneficial properties such as antioxidant and antimutagenic activities, which are derived from the various raw materials and secondary metabolites of the fermentative microorganisms used during production. Despite these useful activities, analysis of the microbial community present in kimchi has received relatively little attention. The objective of this study was to evaluate the bacterial community structure from the raw materials, additives, and final kimchi product using the culture-independent method. Specifically, polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) was used to analyze the 16S rRNA partial sequences of the microflora. One primer set for bacteria, 341F(GC)-518R, reliably produced amplicons from kimchi and its raw materials, and these bands were clearly separated on a 35-65% denaturing gradient gel. Overall, 117 16S rRNA fragments were identified by PCR-DGGE analysis. Pediococcus pentosaceus, Leuconostoc citreum, Leuconostoc gelidum, and Leuconostoc mesenteroides were the dominant bacteria in kimchi. The other strains identified were Tetragenococcus, Pseudomonas, Weissella, and uncultured bacterium. Comprehensive analysis of these microorganisms could provide a more detailed understanding of the biologically active components of kimchi and help improve its quality. PCR-DGGE analysis can be successfully applied to a fermented food to detect unculturable or other species.

  1. Simple and cost effective apparatus for silver staining of polyacrylamide gels with sequential reagents addition and real time monitoring.

    Science.gov (United States)

    Maurye, Praveen; Basu, Arpita; Gupta, Angshuman

    2014-06-01

    Highly reproducible results in molecular biology depend a lot on effective staining and destaining methods. Silver staining of polyacrylamide DNA and protein gel has been adopted widely in the molecular biology laboratories for detecting a very low nanogram range of sample. An efficient staining of a polyacrylamide gel requires a number of well controlled and highly sensitive steps that often becomes tiresome when done manually or when there are a number of gels to be stained simultaneously. Since, silver staining is a multistep procedure that requires proper fixation and exchange of substance, a reliable protocol is necessary and a simple apparatus may be an added advantage to carry out the steps with ease and safety. Here, we describe a simple and cost effective device made from off-the-shelf components for some established silver staining protocols. Staining is done on a tray while six graduated bottles with a liquid delivery stopcock each, is connected to the tray through silicon tubing. The used up solution is drained off completely from the staining tray through a liquid outlet stopcock using vacuum pressure. The system is fixed with a camera connected to a computer for effective control of the staining process in each step. The apparatus provides the researchers with efficient staining and real time monitoring of gels without the need for handling toxic chemicals.

  2. Comprehensive TP53-denaturing gradient gel electrophoresis mutation detection assay also applicable to archival paraffin-embedded tissue

    NARCIS (Netherlands)

    Hayes, V M; Bleeker, W; Verlind, E; Timmer, T; Karrenbeld, A; Plukker, J T; Marx, M P; Hofstra, R M; Buys, C H

    1999-01-01

    A comprehensive mutation detection assay is described for the entire coding region and all splice site junctions of TP53. The assay is based on denaturing gradient gel electrophoresis, which follows either multiplex polymerase chain reaction (PCR) applied to DNA extracted from fresh or frozen tissue

  3. Separation of Native Allophycocyanin and R-Phycocyanin from Marine Red Macroalga Polysiphonia urceolata by the Polyacrylamide Gel Electrophoresis Performed in Novel Buffer Systems

    Science.gov (United States)

    Wang, Yu; Gong, Xueqin; Wang, Shumei; Chen, Lixue; Sun, Li

    2014-01-01

    Three buffer systems of Imidazole−Acetic acid, HEPES−Imidazole/Bis-tris and Bis-tris−HEPES−MES were designed based on the principle of discontinuous polyacrylamide gel electrophoresis (PAGE) for the native PAGE which could be performed in pH 7.0 and 6.5 in order to analyze and prepare the minor components of allophycocyanin (AP) and R-phycocyanin (R-PC) from marine red macroalga Polysiphonia urceolata. These AP and R-PC phycobiliproteins are easily denatured in alkaline environments. The obtained results demonstrated that the PAGE modes performed in the buffer systems of HEPES−Imidazole/Bis-tris and Bis-tris−HEPES−MES gave the satisfactory resolution and separation of AP and R-PC proteins. The absorption and fluorescence spectra of the AP and R-PC proteins which were prepared by the established PAGE modes proved that they maintained natural spectroscopic characteristics. The established PAGE modes may also provide useful references and selections for some other proteins that are sensitive to alkaline environments or are not effectively separated by the classical PAGE modes performed normally in alkaline buffer systems. PMID:25166028

  4. Separation of native allophycocyanin and R-phycocyanin from marine red macroalga Polysiphonia urceolata by the polyacrylamide gel electrophoresis performed in novel buffer systems.

    Directory of Open Access Journals (Sweden)

    Yu Wang

    Full Text Available Three buffer systems of Imidazole-Acetic acid, HEPES-Imidazole/Bis-tris and Bis-tris-HEPES-MES were designed based on the principle of discontinuous polyacrylamide gel electrophoresis (PAGE for the native PAGE which could be performed in pH 7.0 and 6.5 in order to analyze and prepare the minor components of allophycocyanin (AP and R-phycocyanin (R-PC from marine red macroalga Polysiphonia urceolata. These AP and R-PC phycobiliproteins are easily denatured in alkaline environments. The obtained results demonstrated that the PAGE modes performed in the buffer systems of HEPES-Imidazole/Bis-tris and Bis-tris-HEPES-MES gave the satisfactory resolution and separation of AP and R-PC proteins. The absorption and fluorescence spectra of the AP and R-PC proteins which were prepared by the established PAGE modes proved that they maintained natural spectroscopic characteristics. The established PAGE modes may also provide useful references and selections for some other proteins that are sensitive to alkaline environments or are not effectively separated by the classical PAGE modes performed normally in alkaline buffer systems.

  5. Separation of native allophycocyanin and R-phycocyanin from marine red macroalga Polysiphonia urceolata by the polyacrylamide gel electrophoresis performed in novel buffer systems.

    Science.gov (United States)

    Wang, Yu; Gong, Xueqin; Wang, Shumei; Chen, Lixue; Sun, Li

    2014-01-01

    Three buffer systems of Imidazole-Acetic acid, HEPES-Imidazole/Bis-tris and Bis-tris-HEPES-MES were designed based on the principle of discontinuous polyacrylamide gel electrophoresis (PAGE) for the native PAGE which could be performed in pH 7.0 and 6.5 in order to analyze and prepare the minor components of allophycocyanin (AP) and R-phycocyanin (R-PC) from marine red macroalga Polysiphonia urceolata. These AP and R-PC phycobiliproteins are easily denatured in alkaline environments. The obtained results demonstrated that the PAGE modes performed in the buffer systems of HEPES-Imidazole/Bis-tris and Bis-tris-HEPES-MES gave the satisfactory resolution and separation of AP and R-PC proteins. The absorption and fluorescence spectra of the AP and R-PC proteins which were prepared by the established PAGE modes proved that they maintained natural spectroscopic characteristics. The established PAGE modes may also provide useful references and selections for some other proteins that are sensitive to alkaline environments or are not effectively separated by the classical PAGE modes performed normally in alkaline buffer systems.

  6. Separation and recovery of nucleic acids with improved biological activity by acid-degradable polyacrylamide gel electrophoresis.

    Science.gov (United States)

    Kim, Yoon Kyung; Kwon, Young Jik

    2010-05-01

    One of the fundamental challenges in studying biomacromolecules (e.g. nucleic acids and proteins) and their complexes in a biological system is isolating them in their structurally and functionally intact forms. Electrophoresis offers convenient and efficient separation and analysis of biomacromolecules but recovery of separated biomacromolecules is a significant challenge. In this study, DNAs of various sizes were separated by electrophoresis in an acid-degradable polyacrylamide gel. Almost 100% of the nucleic acids were recovered after the identified gel bands were hydrolyzed under a mildly acidic condition and purified using anion exchange resin. Further concentration by centrifugal filtration and a second purification using ion exchange column chromatography yielded 44-84% of DNA. The second conventional (non-degradable) gel electrophoresis confirmed that the nucleic acids recovered from acid-degradable gel bands preserved their electrophoretic properties through acidic gel hydrolysis, purification, and concentration processes. The plasmid DNA recovered from acid-degradable gel transfected cells significantly more efficiently than the starting plasmid DNA (i.e. improved biological activity via acid-degradable PAGE). Separation of other types of nucleic acids such as small interfering RNA using this convenient and efficient technique was also demonstrated.

  7. Screening for TP53 mutations in osteosarcomas using constant denaturant gel electrophoresis (CDGE).

    Science.gov (United States)

    Smith-Sørensen, B; Gebhardt, M C; Kloen, P; McIntyre, J; Aguilar, F; Cerutti, P; Børresen, A L

    1993-01-01

    We have previously developed conditions to screen for TP53 point mutations inside the conserved domains II-V of the gene by using constant denaturant gel electrophoresis (CDGE). The present study reports conditions for screening more of the codons in the frequently mutated region exon 5-8 and for detecting mutations in sequences encoding functional domains in the N- and C-terminal part of the protein. The ability of the CDGE technique to detect mutations was studied using controls with known sequence deviations. The resolution power of the technique to separate different types of mutations was tested by using seven different single base pair mutants all residing in a stretch of four base pairs. All mutants were separated from the wild type. The established CDGE screening strategy was then used to look for mutations in DNA from 28 osteosarcomas. Six (21.5%) of the samples were shown to have a TP53 mutation, and the exact characterization was performed by direct sequencing. All of these were within the frequently reported mutated region exon 5-8.

  8. Monitoring the lactic acid bacterial diversity during shochu fermentation by PCR-denaturing gradient gel electrophoresis.

    Science.gov (United States)

    Endo, Akihito; Okada, Sanae

    2005-03-01

    The presence of lactic acid bacteria (LAB) during shochu fermentation was monitored by PCR-denaturing gradient gel electrophoresis (DGGE) and by bacteriological culturing. No LAB were detected from fermented mashes by PCR-DGGE using a universal bacterial PCR primer set. However, PCR-DGGE using a new primer specific for the 16S rDNA of Lactococcus, Streptococcus, Tetragenococcus, Enterococcus, and Vagococcus and two primers specific for the 16S rDNA of Lactobacillus, Pediococcus, Leuconostoc, and Weissella revealed that Enterococcus faecium, Lactobacillus casei, Lactobacillus fermentum, Lactobacillus nagelii, Lactobacillus plantarum, Lactococcus lactis, Leuconostoc citreum, Leuconostoc mesenteroides, and Weissella cibaria inhabited in shochu mashes. It was also found that the LAB community composition during shochu fermentation changed after the main ingredient and water were added during the fermentation process. Therefore, we confirmed that PCR-DGGE using all three primers specific for groups of LAB together was well suited to the study of the LAB diversity in shochu mashes. The results of DGGE profiles were similar to the results of bacteriological culturing. In conclusion, LAB are present during shochu fermentation but not dominant.

  9. Assessment of microbial populations in methyl ethyl ketone degrading biofilters by denaturing gradient gel electrophoresis.

    Science.gov (United States)

    Li, C; Moe, W M

    2004-05-01

    Denaturing gradient gel electrophoresis (DGGE) analysis of polymerase chain reaction-amplified genes coding for 16S rRNA was used to assess differences in bacterial community structure as a function of spatial location along the height of two biofilters used to treat a model waste gas stream containing methyl ethyl ketone (MEK). One of the laboratory-scale biofilters was operated as a conventional continuous-flow biofilter (CFB) and the other was operated as a sequencing batch biofilter (SBB). Both biofilters, inoculated with an identical starting culture and operated over a period lasting more than 300 days, received the same influent MEK concentration and same mass of MEK on a daily basis. The systems differed, however, in terms of the fraction of time during which contaminated air was supplied and the overall operating strategy employed. DGGE analysis indicated that microbial community structures differed as a function of height in each of the biofilters. The DGGE banding patterns also differed between the two biofilters, suggesting that operating strategies imposed on the biofilters imparted a sufficiently large selective pressure to influence microbial community structures. This may explain, in part, the superior performance of the SBB over the CFB during model transient loading conditions, and it may open new possibilities for purposely manipulating the microbial populations in biofilters treating gas-phase contaminants in a manner that leads to more favorable treatment performance.

  10. Preparation of polyacrylamide gel-filled capillaries with step gradients and low UV-detection background

    Institute of Scientific and Technical Information of China (English)

    陈义

    1997-01-01

    Polyacrylamide- filled capillaries with step gradients were designed and prepared with a newly established method,which is also suitable for producing other sorts of capillaries.The resulting capillaries allow the use of any UV light to approach the most sensitive detection and have the features of fast running speed and high separation efficiency In addition,the capillaries can he used continuously for more than two weeks.

  11. Detection of low-molecular weight allergens resolved on two-dimensional electrophoresis with acid-urea polyacrylamide gel.

    Science.gov (United States)

    Kitta, Kazumi; Ohnishi-Kameyama, Mayumi; Moriyama, Tatsuya; Ogawa, Tadashi; Kawamoto, Shinichi

    2006-04-15

    Two-dimensional electrophoresis with immobilized pH gradient (IPG) followed by acetic acid/urea-polyacrylamide gel electrophoresis (AU-PAGE) was developed for the detection of low-molecular weight food allergens. Wheat proteins were used to test the applicability of AU-PAGE for the analysis of food allergens. Isoelectric focusing (IEF) for first dimension was performed with IPG pH 3-10. AU-PAGE was performed as a second-dimensional electrophoresis and high resolution was obtained, especially for proteins below 15 kDa. For immunodetection, the proteins resolved on AU gel were transferred to a polyvinylidene difluoride membrane. The assembly of semidry electroblotting for AU gel was set reversed as for sodium dodecyl sulfate (SDS)-PAGE gel. The electroblotted membrane was immunolabeled with serum from a radio-allergosorbent test-positive individual for wheat to identify allergenic proteins. Protein spots strongly recognized by the patient's serum were chosen for further analysis. Mass spectrometry analysis revealed that these proteins were alpha-amylase/trypsin inhibitors and lipid transfer protein. The system developed in this study was shown to be useful as a standard protocol for the separation of low-molecular weight proteins. Moreover, the IPG strips on which IEF was performed could be used either for SDS-PAGE or AU-PAGE by only changing equilibrating conditions, allowing for a wide range of allergen analysis.

  12. Production of lignocellulolytic enzymes by Trametes gallica and detection of polysaccharide hydrolase and laccase activities in polyacrylamide gels.

    Science.gov (United States)

    Sun, Xun; Zhang, Renhuai; Zhang, Yizheng

    2004-01-01

    White rot fungus Trametes gallica was studied for the production of lignocellulolytic enzymes: cellulase, xylanase, laccase, manganese-dependent peroxidase (MnP), and lignin peroxidase (LiP). The results demonstrated that low-nitrogen (2.2 mM N) and surface stationary cultivation favored production of extracellular MnP. MnP activity reached 118.1 UL(-1) while T. gallica was grown in a low-nitrogen culture containing phenylalanine. However, laccase levels observed in high-nitrogen (22 mM N) agitated cultures were much greater than those seen in low-nitrogen. The N source experiments seemed to reveal that NH4+ plays an important role in inducing MnP and laccase of the fungus. Results showed that T. gallica produces a series of the lignocellulolytic enzymes, and needs high N to produce all the enzymes during solid-state fermentation of wheat straw. This paper also presents a modified zymogram procedure to detect xylanase and laccase of T. gallica in polyacrylamide gel. Xylanase in crude enzyme of T. gallica was displayed by contacting protein gel strips with xylan substrate gels and by staining with iodine. By immersing the protein gel strips in o-tolidine solution, the blue-green zones representing laccase activity were visualized against a colorless background. Copyright 2004 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim

  13. Association of Streptomyces community composition determined by PCR-denaturing gradient gel electrophoresis with indoor mold status

    OpenAIRE

    Johansson, Elisabet; Reponen, Tiina; Meller, Jarek; Vesper, Stephen; Yadav, Jagjit

    2014-01-01

    Both Streptomyces species and mold species have previously been isolated from moisture-damaged building materials; however, an association between these two groups of microorganisms in indoor environments is not clear. In this study we used a culture-independent method, PCR denaturing gradient gel electrophoresis (PCR-DGGE) to investigate the composition of the Streptomyces community in house dust. Twenty-three dust samples each from two sets of homes categorized as high-mold and low-mold bas...

  14. Isoelectric focusing of human hair keratins: correlation with sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) patterns and effect of cosmetic treatments.

    Science.gov (United States)

    Rodriguez-Calvo, M S; Carracedo, A; Muñoz, I; Concheiro, L

    1992-03-01

    A new isoelectric focusing (IEF) technique in polyacrylamide gels with 6M urea and 1.5% Nonidet P40 has been developed to characterize human hair samples. The phenotypes demonstrated with this procedure has been correlated with the sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) patterns described by other authors. The method described can be applied in the forensic science analysis of a single human hair. Using the same IEF technique we have studied the changes in electrophoretic patterns of cosmetically treated hair. The characteristics of the modifications observed and its utility in forensic science work are also discussed in this paper.

  15. Denaturing gradient gel electrophoresis of neonatal intestinal microbiota in relation to the development of asthma

    Directory of Open Access Journals (Sweden)

    Desager Kristine N

    2011-04-01

    Full Text Available Abstract Background The extended 'hygiene hypothesis' suggests that the initial composition of the infant gut microbiota is a key determinant in the development of atopic disease. Several studies have demonstrated that the microbiota of allergic and non-allergic infants are different even before the development of symptoms, with a critical time window during the first 6 months of life. The aim of the study was to investigate the association between early intestinal colonisation and the development of asthma in the first 3 years of life using DGGE (denaturing gradient gel electrophoresis. Methods In a prospective birth cohort, 110 children were classified according to the API (Asthma Predictive Index. A positive index included wheezing during the first three years of life combined with eczema in the child in the first years of life or with a parental history of asthma. A fecal sample was taken at the age of 3 weeks and analysed with DGGE using universal and genus specific primers. Results The Asthma Predictive Index was positive in 24/110 (22% of the children. Using universal V3 primers a band corresponding to a Clostridum coccoides XIVa species was significantly associated with a positive API. A Bacteroides fragilis subgroup band was also significantly associated with a positive API. A final DGGE model, including both bands, allowed correct classification of 73% (80/110 of the cases. Conclusion Fecal colonisation at age 3 weeks with either a Bacteroides fragilis subgroup or a Clostridium coccoides subcluster XIVa species is an early indicator of possible asthma later in life. These findings need to be confirmed in a new longitudinal follow-up study.

  16. A PCR-denaturing gradient gel electrophoresis approach to assess Fusarium diversity in asparagus.

    Science.gov (United States)

    Yergeau, E; Filion, M; Vujanovic, V; St-Arnaud, M

    2005-02-01

    In North America, asparagus (Asparagus officinalis) production suffers from a crown and root rot disease mainly caused by Fusarium oxysporum f. sp. asparagi and F. proliferatum. Many other Fusarium species are also found in asparagus fields, whereas accurate detection and identification of these organisms, especially when processing numerous samples, is usually difficult and time consuming. In this study, a PCR-denaturing gradient gel electrophoresis (DGGE) method was developed to assess Fusarium species diversity in asparagus plant samples. Fusarium-specific PCR primers targeting a partial region of the translation elongation factor-1 alpha (EF-1 alpha) gene were designed, and their specificity was tested against genomic DNA extracted from a large collection of closely and distantly related organisms isolated from multiple environments. Amplicons of 450 bp were obtained from all Fusarium isolates, while no PCR product was obtained from non-Fusarium organisms. The ability of DGGE to discriminate between Fusarium taxa was tested over 19 different Fusarium species represented by 39 isolates, including most species previously reported from asparagus fields worldwide. The technique was effective to visually discriminate between the majority of Fusarium species and/or isolates tested in pure culture, while a further sequencing step permitted to distinguish between the few species showing similar migration patterns. Total genomic DNA was extracted from field-grown asparagus plants naturally infested with different Fusarium species, submitted to PCR amplification, DGGE analysis and sequencing. The two to four bands observed for each plant sample were all affiliated with F. oxysporum, F. proliferatum or F. solani, clearly supporting the reliability, sensitivity and specificity of this approach for the study of Fusarium diversity from asparagus plants samples.

  17. ISOLATION OF EGG DROP SYNDROME VIRUS AND ITS MOLECULAR CHARACTERIZATION USING SODIUM DODECYL SULPHATE POLYACRYLAMIDE GEL ELECTROPHORESIS

    Directory of Open Access Journals (Sweden)

    M. H. Rasool, S. U. Rahman and M. K. Mansoor

    2005-10-01

    Full Text Available Six isolates of egg drop syndrome (EDS virus were recovered from five different outbreaks of EDS in commercial laying hens in and around Faisalabad. The aberrant eggs were fed to the susceptible laying hens for experimental induction of infection. The samples from infected birds (egg washing, cloacal swabs, oviducts and spleens were collected, processed and inoculated into 11-day old duck embryos. The presence of virus in harvested allanto-amniotic fluid was monitored by spot and microhaemagglutination tests and confirmed by haemagglutination inhibition and agar gel precipitation tests. The EDS virus grew well in duck embryos and agglutinated only avian but not mammalian red blood cells. These isolates were purified through velocity density gradient centrifugation. Protein concentration was determined through Lowry method and sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE was conducted by loading 300 µg protein concentration on 12.5% gel using discontinuous buffer system. All the six isolates showed 13 polypeptides, which were identical to those described in the referral EDS-76 virus (strain-127. The molecular weights of the polypeptides ranged from 6.5 KDa to 126 KDa.

  18. Optimization of a reliable, fast, cheap and sensitive silver staining method to detect SSR markers in polyacrylamide gels

    Directory of Open Access Journals (Sweden)

    Mergeai G.

    2006-01-01

    Full Text Available A reliable, fast, cheap and sensitive silver staining method to detect nucleic acids in polyacrylamide gels was developed from two standard stain procedures. The main differences between the three methods concerned (i the preexposure with formaldehyde during silver nitrate impregnation, (ii the addition of sodium thiosulfate and sodium carbonate instead of sodium hydroxide during development; (iii the removal of the stop reaction or the inclusion of absolute ethanol with acetic acid in the stop solution and (iv the duration of the different reaction steps. All methods allowed the detection of similar polymorphisms for single sequence repeats with different cotton genotypes but important differences regarding the contrast, background and conservation duration of the gels were observed. Two methods gave superior sensitivity. The improved method was sensitive, fast (20 min, gave lower backgrounds, produced gels with long-term conservation ability, and allowed a reutilization of all the solutions used in the staining procedure from fi ve to seven times, making it quite cheap.

  19. Swelling/deswelling of polyacrylamide gels in aqueous NaCl solution: Light scattering and macroscopic swelling study

    Indian Academy of Sciences (India)

    M Sivanantham; B V R Tata

    2012-09-01

    Swelling kinetics of water-swollen polyacrylamide (PAAm) hydrogels (WSG) was investigated in various concentrations of aqueous NaCl by macroscopic swelling measurements. For lower concentration of NaCl, WSG showed exponential swelling whereas at higher concentration of NaCl it underwent deswelling at short times and exponential swelling at long times. From these studies, collective diffusion coefficient, , of the polymer network and polymer–solvent interaction parameter, , were calculated and found to decrease with increase in [NaCl]. Collective diffusion coefficients measured from dynamic light scattering (DLS) and that obtained from macroscopic swelling measurements are found to agree well. Measured ensemble-averaged dynamic structure factor (, ) for WSG and salt-swollen gels (SSG) showed an initial decay followed by a plateau at long times and it can be described by harmonically bound Brownian particle (HBBP) model. Enhanced scattering intensity at low scattering angles using static light scattering (SLS) measurements revealed the presence of inhomogeneities in PAAm gels. The reasons for increased scattering intensity of SSG over WSG gel and the linear decrease of with increase in NaCl concentration are explained.

  20. Assessing CMT cell line stability by two dimensional polyacrylamide gel electrophoresis and mass spectrometry based proteome analysis

    DEFF Research Database (Denmark)

    Zhang, Kelan; Wrzesinski, Krzysztof; Fey, Stephen J;

    2008-01-01

    Two-dimensional polyacrylamide gel electrophoresis (2-D PAGE) followed by mass spectrometric identification of the proteins in the protein spots has become a central tool in proteomics. CMT167(H), CMT64(M) and CMT170(L) cell lines, selected from a spontaneous mouse lung adenocarcinoma, with high......-, middle- or low-metastatic potential have been characterized in vivo. In this study, the comprehensive protein expression profiles of the CMT cell lines were analyzed at passages 5, 15 and 35 in order to assess the cell line stability. During the passages 5 to 15, the expression profiles of CMT cells...... to be a useful tool for assessing differences in cell line stability. This approach provided a tool to select the best cell line and optimal subculture period for studies of cancer related phenomena and for testing the effect of potential anticancer drugs....

  1. Quantitation of yeast total proteins in sodium dodecyl sulfate-polyacrylamide gel electrophoresis sample buffer for uniform loading.

    Science.gov (United States)

    Sheen, Hyukho

    2016-04-01

    Proteins in sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) sample buffer are difficult to quantitate due to SDS and reducing agents being in the buffer. Although acetone precipitation has long been used to clean up proteins from detergents and salts, previous studies showed that protein recovery from acetone precipitation varies from 50 to 100% depending on the samples tested. Here, this article shows that acetone precipitates proteins highly efficiently from SDS-PAGE sample buffer and that quantitative recovery is achieved in 5 min at room temperature. Moreover, precipitated proteins are resolubilized with urea/guanidine, rather than with SDS. Thus, the resolubilized samples are readily quantifiable with Bradford reagent without using SDS-compatible assays.

  2. A case of alcoholic liver injury with an unusual polyacrylamide-gel disc-electrophoretic pattern of serum lipoproteins.

    Directory of Open Access Journals (Sweden)

    Watanabe,Makoto

    1979-06-01

    Full Text Available An unusual lipoprotein pattern on polyacrylamide-gel disc-electrophoresis was observed in 37 year-old male diagnosed as alcoholic liver injury. The electrophoretic lipoprotein pattern consisted of a major band of pre-beta mobility and minor intermediate, fast-beta and slow-alpha bands. The normal beta band was virtually absent and the alpha band was diminished. The abnormal lipoprotein pattern was observed one week after discontinuing alcohol consumption when marked hypertriglyceridemia demonstrated earlier had already normalized leaving a moderate hypercholesterolemia with reduced esterified cholesterol and abnormal liver function tests. The lipoprotein abnormalities were completely normal one month later. The appearance of a major pre-beta band with normal triglyceride and high cholesterol levels is discussed in relation to the formation of larger triglyceride-rich LDL particles in recovery from alcoholic hepatitis.

  3. Rheological and mechanical behavior of polyacrylamide hydrogels chemically crosslinked with allyl agarose for two-dimensional gel electrophoresis.

    Science.gov (United States)

    Suriano, R; Griffini, G; Chiari, M; Levi, M; Turri, S

    2014-02-01

    Two-dimensional (2-D) gel electrophoresis currently represents one of the most standard techniques for protein separation. In addition to the most commonly employed polyacrylamide crosslinked hydrogels, acrylamide-agarose copolymers have been proposed as promising systems for separation matrices in 2-D electrophoresis, because of the good resolution of both high and low molecular mass proteins made possible by careful control and optimization of the hydrogel pore structure. As a matter of fact, a thorough understanding of the nature of the hydrogel pore structure as well as of the parameters by which it is influenced is crucial for the design of hydrogel systems with optimal sieving properties. In this work, a series of acrylamide-based hydrogels covalently crosslinked with different concentrations of allyl agarose (0.2-1%) is prepared and characterized by creep-recovery measurements, dynamic rheology and tensile tests, in the attempt to gain a clearer understanding of structure-property relationships in crosslinked polyacrylamide-based hydrogels. The rheological and mechanical properties of crosslinked acrylamide-agarose hydrogels are found to be greatly affected by crosslinker concentration. Dynamic rheological tests show that hydrogels with a percentage of allyl agarose between 0.2% and 0.6% have a low density of elastically effective crosslinks, explaining the good separation of high molecular mass proteins in 2-D gel electrophoresis. Over the same range of crosslinker concentration, creep-recovery measurements reveal the presence of non-permanent crosslinks in the hydrogel network that justifies the good resolution of low molecular mass proteins as well. In tensile tests, the hydrogel crosslinked with 0.4% of allyl agarose exhibits the best results in terms of mechanical strength and toughness. Our results show how the control of the viscoelastic and the mechanical properties of these materials allow the design of mechanically stable hydrogels with improved

  4. Sodium dodecyl sulfate-polyacrylamide gel electrophoretic method for assessing the quaternary state and comparative thermostability of avidin and streptavidin.

    Science.gov (United States)

    Bayer, E A; Ehrlich-Rogozinski, S; Wilchek, M

    1996-08-01

    Avidin, a positively charged egg-white protein, aggregates extensively when mixed at ambient temperatures with anionic detergents, such as sodium dodecyl sulfate (SDS). The resultant aggregates fail to penetrate the stacking gel during polyacrylamide gel electrophoresis (PAGE). To prevent the formation of such aggregates, avidin was acetylated and the pI was thus reduced. Acetylated avidin was found to behave in a manner similar to that of streptavidin; under nondenaturing conditions (i.e., incubation of samples at room temperature), both proteins normally migrated mainly as tetramers with a tendency to form oligomers of the tetramer. When samples were boiled, both proteins migrated mainly as the monomer. The comparative stability properties of avidin and streptavidin were also examined using SDS-PAGE by heating samples and determining the extent of dissociation of tetramers to monomers as a function of temperature. A distinctive transition temperature could be defined for individual samples. Using this assay, it was determined that, in the absence of biotin, the quaternary structure of streptavidin is more stable than that of avidin. Biotin appears to stabilize structures of both avidin and streptavidin to a similar degree. Acetylation of avidin thus provides a simple means to analyze the quaternary structure of the molecule using SDS-PAGE.

  5. Antigenic profile of heat-killed versus thimerosal-treated Leishmania major using sodium dodecyl sulfate-polyacrylamide gel electrophoresis

    Directory of Open Access Journals (Sweden)

    Reza Arjmand

    2015-01-01

    Full Text Available Background: Leishmania is a parasitic protozoan of trypanosomatidae family which causes a wide spectrum of diseases ranging from self-healing cutaneous lesions to deadly visceral forms. In endemic areas, field trials of different preparations of Leishmania total antigen were tested as leishmaniasis vaccine. Two preparations of killed Leishmania major were produced In Iran, which were heat-killed vaccine called autoclaved L. major (ALM and thimerosal-treated freeze-thawed vaccine called killed L. major (KLM. In this study, the protein content of both ALM and KLM were compared with that of freshly harvested intact L. major promastigotes using sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE. Materials and Methods: L. major (MRHO/IR/75/ER from pre-infected Balb/c mice was isolated with modified Novy-MacNeal-Nicolle (NNN medium and then subcultured in liquid RPMI 1640 medium supplemented with fetal calf serum (FCS 20% for mass production. Two preparations of KLM and ALM were produced by Razi Vaccine and Serum Research Institute, Iran, under WHO/TDR supervision. Electrophoresis was performed by SDS-PAGE method and the gel was stained by Coomassie brilliant blue dye. The resultant unit bands were compared using standard molecular proteins. Results: Electrophoresis of the two preparations produced many bands from 10 kDa to 100 kDa. KLM bands were much like those of freshly harvested intact L. major. Conclusion: It is concluded that although there are similar bands in the three forms of Leishmania antigens, there are some variations which might be considered for identification and purification of protective immunogens in a total crude antigen, and detection of their stability is essential for the production and marketing of a putative vaccine.

  6. Electrochemical characterizations on MnO2 supercapacitors with potassium polyacrylate and potassium polyacrylate-co-polyacrylamide gel polymer electrolytes

    KAUST Repository

    Lee, Kuang-Tsin

    2009-11-01

    MnO2·nH2O supercapacitors with potassium polyacrylate (PAAK) and potassium polyacrylate-co-polyacrylamide (PAAK-co-PAAM) gel polymer electrolytes (GPEs) having the weight compositions of polymer:KCl:H2O = 9%:6.7%:84.3% have been characterized for their electrochemical performance. Compared with the liquid electrolyte (LE) counterpart, the GPE cells exhibit remarkable (∼50-130%) enhancement in specific capacitance of the oxide electrode, and the extent of the enhancement increases with increasing amount of the carboxylate groups in the polymers as well as with increasing oxide/electrolyte interfacial area. In situ X-ray absorption near-edge structure (XANES) analysis indicates that the oxide electrodes of the GPE cells possess higher Mn-ion valences and are subjected to greater extent of valence variation than that of the LE cell upon charging/discharging over the same potential range. Copolymerization of PAAK with PAAM greatly improves the cycling stability of the MnO2·nH2O electrode, and the improvement is attributable to the alkaline nature of the amino groups. Both GPEs exhibit ionic conductivities greater than 1.0 × 10-1 S cm-1 and are promising for high-rate applications. © 2009 Elsevier Ltd. All rights reserved.

  7. Polyacrylamide Gel-Entrapped Fungal Manganese Peroxidase from Ganoderma lucidum IBL-05 with Enhanced Catalytic, Stability, and Reusability Characteristics.

    Science.gov (United States)

    Bilal, Muhammad; Asgher, Muhammad; Iqbal, Hafiz M N

    2016-01-01

    In the present study, polyacrylamide gel (PAG) was utilized as bolster material for the immobilization of in-house extracted and partially purified manganese peroxidase (MnP) through an entrapment technique yielding significant MnP immobilization (87.3±3.3 %) and remarkable stability of the enzyme (37.2±2.4 %) after a storage period of two months at 4°C. The immobilization also increased the optimal temperature by 10 °C and provided an alkaline shift of the pH optimum. Moreover, a significant enhancement in the thermo-stability was observed. After an incubation period of 72 h at 50°C, the PAG-entrapped-MnP still exhibited 41.2 % of the initial activity, whereas the free enzyme was completely inactive. Furthermore, PAG-entrapped-MnP showed an excellent recycling efficiency and retained more than 50% of its initial activity after five consecutive reaction cycles. In conclusion, owing to the economic feasibility, carrier-supported MnP may be a promising candidate for various applications in different industrial sectors.

  8. Sample Preparation and Staining Methods for Two-Dimensional Polyacrylamide Gel Electrophoresis of Proteins from Animal Tissues

    Directory of Open Access Journals (Sweden)

    Levente Czegledi

    2010-05-01

    Full Text Available Proteomics in animal science as well as in other biological sciences is a significant tool in the post-genomic era. In proteomic studies the presence and relative abundance of expressed proteins of a cell, tissue or biological fluid is studied. Recently, the whole genome of more and more domestic animal species is known, but genes and the transcribed mRNA have no direct effect on biological systems as they are regulated by proteins, which explain the importance of proteomics. The most common tool in proteomic approach is the two-dimensional polyacrylamide gel electrophoresis (2D PAGE, when proteins are separated by their isoelectric point followed by their mass separation as a second dimension. In this study authors used different sample preparation and protein staining methods on meat,  liver and blood plasma and carried out 2D PAGE experiments. The most appropriate sample preparation methods are described in this paper. We concluded that depletion of major proteins in plasma is required but not necessary for meat and liver samples.

  9. Identification of coagulase-negative staphylococci by using sodium dodecyl sulfate-polyacrylamide gel electrophoresis and rRNA restriction patterns.

    OpenAIRE

    Pennington, T. H.; Harker, C.; Thomson-Carter, F

    1991-01-01

    A total of 1,417 staphylococcal and micrococcal strains were collected from the beards and scalps of 10 subjects over a period of 8 months. Sixteen strains identified as Staphylococcus epidermidis with an API system had distinctive yellow colonies on nutrient agar plates and sodium dodecyl sulfate-polyacrylamide gel electrophoresis whole-cell polypeptide profiles similar to those of Staphylococcus capitis; this identification was confirmed by analysis of rRNA gene restriction patterns.

  10. Synthesis of YAG:Ce3+ Phosphor by Polyacrylamide Gel Method and Promoting Action of α-Al2O3 Seed Crystal on Phase Formation

    Institute of Scientific and Technical Information of China (English)

    Li Yongxiu; Li Yinyi; Min Yulin; Wu Yanli; Cheng Changming; Zhou Xuezhen; Gu Ziying

    2005-01-01

    YAG:Ce3+ phosphor particles were prepared using polyacrylamide gel method. The structure evolution of powders during annealing process was followed by X-ray diffraction determination. It is found that some intermediate phases, including θ-Al2O3, YAM and YAP, are formed when calcining polyacrylamide gel, however, the pure YAG phase can be formed directly when calcining polyacrylamide gel with α-Al2O3 as seed crystal. These facts show that the existence of α-Al2O3 seed crystal can block the formation of θ-Al2O3, YAM and YAP, and accelerate its reaction with Y2O3 to form YAG phase directly at lower temperature. The emission peak of prepared YAG:Ce3+ phosphor is wide with maximum at 550 nm and the exitation band has two peaks, the major one is around at 460 nm, which matches the blue emission of GaN LED and is suitable for the assemble of white LED. Some fluxes can enhance the photoluminescence intensity of phosphor particles, that can be attributed both to the improvement of crystallization processes of YAG and to the stabilization of trivalence cerium ion in YAG:Ce3+.

  11. Serum sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis of patients with membranous nephropathy and focal and segmental glomerulosclerosis

    Directory of Open Access Journals (Sweden)

    Pragya Pant

    2016-01-01

    Full Text Available Diagnosis of membranous nephropathy (MN and focal and segmental glomerulo- sclerosis (FSGS needs a renal biopsy, which is an invasive procedure with potentially serious complications. Proteomics may be applied for the development of a biomarker for these diseases which will obviate the need of biopsy. Serum sodium dodecyl sulfate-polyacrylamide gel electro-phoresis (SDS-PAGE analysis gives an idea of the various proteins with different molecular weights (MWs in a given sample. This study was conducted to analyze proteins with different MWs in patients with MN and FSGS and to compare the two groups with regard to their protein profile. This was a comparative, experimental study performed from June 2013 to July 2014 in the Department of Nephrology, Sir Sunderlal Hospital, Banaras Hindu University, Varanasi. Twenty-three histologically diagnosed cases of primary MN and 25 cases of FSGS were included in the study. Patients were categorized as having mild, moderate, and severe proteinuria with 24 h urinary protein levels of <4, 4- 8 and ≥8 g/24 h, respectively. SDS-PAGE analysis was performed by the method of Laemmli and revealed a significantly higher number of patients with FSGS (80% having a protein corresponding to 29 kDa MW, than those with MN (39.1% (P = 0.004. Protein of 5 kDa MW was present in a significantly higher number of patients with moderate (80% and severe (100% proteinuria than those with mild proteinuria (25% (P <0.001. Thus, protein of MW 29 kDa may be a marker for FSGS and needs further characterization. Similarly, 5 kDa protein, present in patients with moderate and severe proteinuria, might be either contributing to or be a marker of severe illness.

  12. Serum sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis of patients with membranous nephropathy and focal and segmental glomerulosclerosis.

    Science.gov (United States)

    Pant, Pragya; Singh, R G; Singh, Santosh K; Singh, Vijay P; Doley, Prodip K; Sivasankar, M

    2016-05-01

    Diagnosis of membranous nephropathy (MN) and focal and segmental glomerulo- sclerosis (FSGS) needs a renal biopsy, which is an invasive procedure with potentially serious complications. Proteomics may be applied for the development of a biomarker for these diseases which will obviate the need of biopsy. Serum sodium dodecyl sulfate-polyacrylamide gel electro-phoresis (SDS-PAGE) analysis gives an idea of the various proteins with different molecular weights (MWs) in a given sample. This study was conducted to analyze proteins with different MWs in patients with MN and FSGS and to compare the two groups with regard to their protein profile. This was a comparative, experimental study performed from June 2013 to July 2014 in the Department of Nephrology, Sir Sunderlal Hospital, Banaras Hindu University, Varanasi. Twenty-three histologically diagnosed cases of primary MN and 25 cases of FSGS were included in the study. Patients were categorized as having mild, moderate, and severe proteinuria with 24 h urinary protein levels of <4, 4- 8 and ≥8 g/24 h, respectively. SDS-PAGE analysis was performed by the method of Laemmli and revealed a significantly higher number of patients with FSGS (80%) having a protein corresponding to 29 kDa MW, than those with MN (39.1%) (P = 0.004). Protein of 5 kDa MW was present in a significantly higher number of patients with moderate (80%) and severe (100%) proteinuria than those with mild proteinuria (25%) (P <0.001). Thus, protein of MW 29 kDa may be a marker for FSGS and needs further characterization. Similarly, 5 kDa protein, present in patients with moderate and severe proteinuria, might be either contributing to or be a marker of severe illness.

  13. Comparison of Different Denaturing Gradient Gel Electrophoresis Primer Sets for the Study of Marine Bacterioplankton Communities▿ †

    Science.gov (United States)

    Sánchez, Olga; Gasol, Josep M.; Massana, Ramon; Mas, Jordi; Pedrós-Alió, Carlos

    2007-01-01

    An annual seasonal cycle of composition of a bacterioplankton community in an oligotrophic coastal system was studied by denaturing gradient gel electrophoresis (DGGE) using five different primer sets. Analysis of DGGE fingerprints showed that primer set 357fGC-907rM grouped samples according to seasons. Additionally, we used the set of 16S rRNA genes archived in the RDPII database to check the percentage of perfect matches of each primer for the most abundant bacterial groups inhabiting coastal plankton communities. Overall, primer set 357fGC-907rM was the most suitable for the routine use of PCR-DGGE analyses in this environment. PMID:17660308

  14. Utilization of denaturing gradient gel electrophoresis for diagnosis of {beta}-thalassemia and ascertainment of new mutations

    Energy Technology Data Exchange (ETDEWEB)

    Ngo, K.Y.; Liu, D.; Lee, J. [Univ. of California, San Diego (United States)] [and others

    1994-09-01

    During the past two years we have tested 2,300 Southeast Asians for alpha- and beta-thaleassemia mutations. We found the incidence of hemoglobin E ({beta}{sup 26}) to be 47% among Laotians and 38% among Cambodians. The incidence of beta thalassemia trait is 9% for Laotians and 6% for Cambodians. Thus, the risk for hemoglobin E/{beta}{sup 26} thalassemia, a transfusion-dependent disorder, is increased in these two population groups. Denaturing gradient gel electrophoresis (DGGE) has proven to be useful in testing for beta-thalassemia carriers and identifying new mutations in the beta globin gene. DNA was extracted from venous blood obtained from patients with elevated Hgb A2 (>4%). Five DNA fragments, encompassing the beta globin gene cluster, were amplified by PCR and analyzed, along with known beta gene mutations as controls, by DGGE using different denaturing gradient concentrations. Different mutations at the same nucleotide position can be distinguished by migration pattern on the DGGE (e.g., in IVS-I-1, G{r_arrow}A and T). Compound heterozygotes for {beta}-thalassemia can be detected on the same gel (e.g., HbE/mutation codon 17). New mutations are identified by their migration pattern compared with controls and determined by subsequent sequencing. We have identified three new mutations: codon 82 CAA{r_arrow}AAA in one Cambodian patient; IVS-II-667, T{r_arrow}C and IVS-II-672, A{r_arrow}C in two Laotian patients. When the parent`s genotypes are known, prenatal diagnosis can be obtained within 24 hours. Thus, PCR/DGGE combination is a rapid and reliable diagnostic approach to clinically significant {beta}-thalassemia. The most important steps are carefully designed primers and predetermined gradient concentrations for DGGE.

  15. SU-E-T-105: Development of 3D Dose Verification System for Volumetric Modulated Arc Therapy Using Improved Polyacrylamide-Based Gel Dosimeter

    Energy Technology Data Exchange (ETDEWEB)

    Ono, K; Fujimoto, S; Akagi, Y; Hirokawa, Y [Hiroshima Heiwa Clinic, Hiroshima (Japan); Hayashi, S [Hiroshima International University, Hiroshima (Japan); Miyazawa, M [R-TECH.INC, Toukyo (Japan)

    2014-06-01

    Purpose: The aim of this dosimetric study was to develop 3D dose verification system for volumetric modulated arc therapy (VMAT) using polyacrylamide-based gel (PAGAT) dosimeter improved the sensitivity by magnesium chloride (MgCl{sub 2}). Methods: PAGAT gel containing MgCl{sub 2} as a sensitizer was prepared in this study. Methacrylic-acid-based gel (MAGAT) was also prepared to compare the dosimetric characteristics with PAGAT gel. The cylindrical glass vials (4 cm diameter, 12 cm length) filled with each polymer gel were irradiated with 6 MV photon beam using Novalis Tx linear accelerator (Varian/BrainLAB). The irradiated polymer gel dosimeters were scanned with Signa 1.5 T MRI system (GE), and dose calibration curves were obtained using T{sub 2} relaxation rate (R{sub 2} = 1/T{sub 2}). Dose rate (100-600 MU min{sup −1}) and fractionation (1-8 fractions) were varied. In addition, a cubic acrylic phantom (10 × 10 × 10 cm{sup 3}) filled with improved PAGAT gel inserted into the IMRT phantom (IBA) was irradiated with VMAT (RapidArc). C-shape structure was used for the VMAT planning by the Varian Eclipse treatment planning system (TPS). The dose comparison of TPS and measurements with the polymer gel dosimeter was accomplished by the gamma index analysis, overlaying the dose profiles for a set of data on selected planes using in-house developed software. Results: Dose rate and fractionation dependence of improved PAGAT gel were smaller than MAGAT gel. A high similarity was found by overlaying the dose profiles measured with improved PAGAT gel dosimeter and the TPS dose, and the mean pass rate of the gamma index analysis using 3%/3 mm criteria was achieved 90% on orthogonal planes for VMAT using improved PAGAT gel dosimeter. Conclusion: In-house developed 3D dose verification system using improved polyacrylamide-based gel dosimeter had a potential as an effective tool for VMAT QA.

  16. Comparison of Fecal Methanogenic Archaeal Community Between Erhualian and Landrace Pigs Using Denaturing Gradient Gel Electrophoresis and Real-Time PCR Analysis

    NARCIS (Netherlands)

    Su, Y.; Smidt, H.; Zhu, W.Y.

    2014-01-01

    Erhualian and Landrace breeds are typical genetically obese and lean pigs, respectively. To compare the fecal methanogenic Archaeal community between these two pig breeds, fecal samples from different growth phase pigs were collected and used for PCR-denaturing gradient gel electrophoresis (DGGE) wi

  17. Analysis of bacterial communities in soil by use of denaturing gradient gel electrophoresis and clone libraries, as influenced by different reverse primers

    NARCIS (Netherlands)

    Brons, Jolanda; van Elsas, J.D.

    2008-01-01

    To assess soil bacterial diversity, PCR systems consisting of several slightly different reverse primers together with forward primer F968-GC were used along with subsequent denaturing gradient gel electrophoresis (DGGE) or clone library analyses. In this study, a set of 13 previously used and novel

  18. A New Standard-Based Polynomial Interpolation (SBPIn) method to address gel-to-gel variability for the comparison of multiple denaturing gradient gel electrophoresis profile matrices.

    Science.gov (United States)

    Valentín-Vargas, Alexis; Chorover, Jon; Maier, Raina M

    2013-02-15

    The Standard-Based Polynomial Interpolation (SBPIn) method is a new simple three-step protocol proposed to address common gel-to-gel variations for the comparison of sample profiles across multiple DGGE gels. The advantages of this method include no requirement for additional software or modification of the standard DGGE protocol.

  19. A New Standard-Based Polynomial Interpolation (SBPIn) Method to Address Gel-to-Gel Variability for the Comparison of Multiple Denaturing Gradient Gel Electrophoresis Profile Matrices

    Science.gov (United States)

    Valentín-Vargas, Alexis; Chorover, Jon; Maier, Raina M.

    2013-01-01

    The Standard-Based Polynomial Interpolation (SBPIn) method is a new simple three-step protocol proposed to address common gel-to-gel variations for the comparison of sample profiles across multiple DGGE gels. The advantages of this method include no requirement for additional software or modification of the standard DGGE protocol. PMID:23234884

  20. Evaluation of mutation screening by heteroduplex analysis in acute intermittent porphyria: comparison with denaturing gradient gel electrophoresis.

    Science.gov (United States)

    Tchernitchko, D; Lamoril, J; Puy, H; Robreau, A M; Bogard, C; Rosipal, R; Gouya, L; Deybach, J C; Nordmann, Y

    1999-01-01

    Acute intermittent porphyria is the major autosomal dominant form of acute hepatic porphyrias. The disease is due to mutations in the gene encoding for porphobilinogen deaminase (PBGD). Many different strategies have been developed to screen for mutations. However the high prevalence (0.6 per thousand) of PBGD gene defect, the large allelic heterogeneity of mutations (n = 130), and the limitations of the PBGD enzymatic assay for asymptomatic patients' detection, require for diagnosis an efficient and easy to handle strategy for locating mutations within the PBGD gene. In a recent study the sensitivity of the denaturing gradient gel electrophoresis (DGGE) technique was 100%. However DGGE requires the preparation of gradient gels and the use of primers with long GC-clamps; thus alternative methods should be preferable in the clinical laboratory. We have compared the detection rate of DGGE with heteroduplex analysis (HA) using 16 characterized PBGD gene mutations. Six different HA conditions were used to determine the efficiency of the method, including: (1) MDE (mutation detection enhancement) gel concentration; (2) addition of urea and sodium dodecyl sulfate (SDS); (3) radioactive labelling. The sensitivity of each HA condition varied from 31 to 81% vs. 100% in DGGE analysis. HA using 1 x MDE with 15% urea with or without 0.55% SDS was the most sensitive condition. This first comparative study of DGGE and HA mutation screening methods suggests that DGGE is a more sensitive screening assay than optimized HA. However, because of its simplicity HA should be considered as an efficient alternative mutation screening method.

  1. Identification and Population Dynamics of Yeasts in Sourdough Fermentation Processes by PCR-Denaturing Gradient Gel Electrophoresis

    Science.gov (United States)

    Meroth, Christiane B.; Hammes, Walter P.; Hertel, Christian

    2003-01-01

    Four sourdoughs (A to D) were produced under practical conditions, using a starter obtained from a mixture of three commercially available sourdough starters and baker's yeast. The doughs were continuously propagated until the composition of the microbiota remained stable. A fungi-specific PCR-denaturing gradient gel electrophoresis (DGGE) system was established to monitor the development of the yeast biota. The analysis of the starter mixture revealed the presence of Candida humilis, Debaryomyces hansenii, Saccharomyces cerevisiae, and Saccharomyces uvarum. In sourdough A (traditional process with rye flour), C. humilis dominated under the prevailing fermentation conditions. In rye flour sourdoughs B and C, fermented at 30 and 40°C, respectively, S. cerevisiae became predominant in sourdough B, whereas in sourdough C the yeast counts decreased within a few propagation steps below the detection limit. In sourdough D, which corresponded to sourdough C in temperature but was produced with rye bran, Candida krusei became dominant. Isolates identified as C. humilis and S. cerevisiae were shown by randomly amplified polymorphic DNA-PCR analysis to originate from the commercial starters and the baker's yeast, respectively. The yeast species isolated from the sourdoughs were also detected by PCR-DGGE. However, in the gel, additional bands were visible. Because sequencing of these PCR fragments from the gel failed, cloning experiments with 28S rRNA amplicons obtained from rye flour were performed, which revealed Cladosporium sp., Saccharomyces servazii, S. uvarum, an unculturable ascomycete, Dekkera bruxellensis, Epicoccum nigrum, and S. cerevisiae. The last four species were also detected in sourdoughs A, B, and C. PMID:14660398

  2. Analysis of Soluble Proteins in Natural Cordyceps sinensis from Different Producing Areas by Sodium Dodecyl Sulfate-Polyacrylamide Gel Electrophoresis and Two-dimensional Electrophoresis.

    Science.gov (United States)

    Li, Chun-Hong; Zuo, Hua-Li; Zhang, Qian; Wang, Feng-Qin; Hu, Yuan-Jia; Qian, Zheng-Ming; Li, Wen-Jia; Xia, Zhi-Ning; Yang, Feng-Qing

    2017-01-01

    As one of the bioactive components in Cordyceps sinensis (CS), proteins were rarely used as index components to study the correlation between the protein components and producing areas of natural CS. Protein components of 26 natural CS samples produced in Qinghai, Tibet, and Sichuan provinces were analyzed and compared to investigate the relationship among 26 different producing areas. Proteins from 26 different producing areas were extracted by Tris-HCl buffer with Triton X-100, and separated using sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and two-dimensional electrophoresis (2-DE). The SDS-PAGE results indicated that the number of protein bands and optical density curves of proteins in 26 CS samples was a bit different. However, the 2-DE results showed that the numbers and abundance of protein spots in protein profiles of 26 samples were obviously different and showed certain association with producing areas. Based on the expression values of matched protein spots, 26 batches of CS samples can be divided into two main categories (Tibet and Qinghai) by hierarchical cluster analysis. The number of protein bands and optical density curves of proteins in 26 Cordyceps sinensis samples were a bit different on the sodium dodecyl sulfate-polyacrylamide gel electrophoresis protein profilesNumbers and abundance of protein spots in protein profiles of 26 samples were obvious different on two-dimensional electrophoresis mapsTwenty-six different producing areas of natural Cordyceps sinensis samples were divided into two main categories (Tibet and Qinghai) by Hierarchical cluster analysis based on the values of matched protein spots. Abbreviations Used: SDS-PAGE: Sodium dodecyl sulfate polyacrylamide gel electrophoresis, 2-DE: Two-dimensional electrophoresis, Cordyceps sinensis: CS, TCMs: Traditional Chinese medicines.

  3. Determination of amino acid compositions and NH2-terminal sequences of peptides electroblotted onto PVDF membranes from tricine-sodium dodecyl sulfate-polyacrylamide gel electrophoresis

    DEFF Research Database (Denmark)

    Ploug, M; Jensen, A L; Barkholt, V.

    1989-01-01

    The combination of high-resolution Tricine-Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (H. Schägger and G. von Jagow (1987) Anal. Biochem. 166, 368-379) and electroblotting onto polyvinylidene difluoride (PVDF) membranes represents a powerful technique for the isolation of small...... amounts of peptides and protein fragments (Mr 1000-20,000) in a suitable form for amino acid sequencing, directly on the blotting membrane. Conditions for electrophoresis and electroblotting were optimized with respect to high transfer yield and suitability for both amino acid analysis and sequence...

  4. Nondenaturing polyacrylamide gel electrophoresis to study the dissociation of the p53·MDM2/X complex by potentially anticancer compounds.

    Science.gov (United States)

    Sgammato, Roberta; Desiderio, Doriana; Lamberti, Anna; Raimo, Gennaro; Novellino, Ettore; Carotenuto, Alfonso; Masullo, Mariorosario

    2015-12-01

    A new analytical method to study the dissociation of the complexes between the oncosuppressor p53 and its negative modulators murine double-minute protein 2 (MDM2) or MDMX, is proposed. This technique is reliable to determine the dissociative power exerted by small molecules on the complex taking advantage of the appearance of migrating MDM2 or MDMX in a native polyacrylamide gel, when inhibitors are added to the complex mixture. Therefore, we propose this new approach to easily screen library of compounds, with potential pharmacological anticancer activity. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  5. Microbial Diversity during Fermentation of Sweet Paste, a Chinese Traditional Seasoning, Using PCR-Denaturing Gradient Gel Electrophoresis.

    Science.gov (United States)

    Mao, Ping; Hu, Yuanliang; Liao, Tingting; Wang, Zhaoting; Zhao, Shumiao; Liang, Yunxiang; Hu, Yongmei

    2017-04-28

    The aim of this study was to elucidate the changes in the microbial community and biochemical properties of a traditional sweet paste during fermentation. PCR-denaturing gradient gel electrophoresis (DGGE) analysis showed that Aspergillus oryzae was the predominant species in the koji (the fungal mixture), and the majority of the fungi isolated belonged to two Zygosaccharomyces species in the mash. The bacterial DGGE profiles revealed the presence of Bacillus subtilis during fermentation, and Lactobacillus acidipiscis, Lactobacillus pubuzihii, Lactobacillus sp., Staphylococcus kloosi, and several uncultured bacteria were also detected in the mash after 14 days of main fermentation. Additionally, during main fermentation, amino-type nitrogen and total acid increased gradually to a maximum of 6.77 ± 0.25 g/kg and 19.10 ± 0.58 g/kg (30 days) respectively, and the concentration of reducing sugar increased to 337.41 ± 3.99 g/kg (7 days). The 180-day fermented sweet paste contained 261.46 ± 19.49 g/kg reducing sugar and its pH value remained at around 4.65. This study has used the PCR-DGGE technique to demonstrate the microbial community (including bacteria and fungi) in sweet paste and provides useful information (biochemical properties) about the assessment of the quality of sweet paste throughout fermentation.

  6. Detection and differentiation of Entamoeba histolytica and Entamoeba dispar in clinical samples through PCR-denaturing gradient gel electrophoresis

    Directory of Open Access Journals (Sweden)

    P. López-López

    Full Text Available Amebiasis is one of the twenty major causes of disease in Mexico; however, the diagnosis is difficult due to limitations of conventional microscopy-based techniques. In this study, we analyzed stool samples using polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE to differentiate between Entamoeba histolytica (pathogenic and E. dispar (non-pathogenic. The target for the PCR amplification was a small region (228 bp of the adh112 gene selected to increase the sensitivity of the test. The study involved 62 stool samples that were collected from individuals with complaints of gastrointestinal discomfort. Of the 62 samples, 10 (16.1% were positive for E. histolytica while 52 (83.9% were negative. No sample was positive for E. dispar. These results were validated by nested PCR-RFLP (restriction fragment length polymorphism and suggest that PCR-DGGE is a promising tool to differentiate among Entamoeba infections, contributing to determine the specific treatment for patients infected with E. histolytica, and therefore, avoiding unnecessary treatment of patients infected with the non-pathogenic E. dispar.

  7. Analysis of microbial diversity on deli slicers using polymerase chain reaction and denaturing gradient gel electrophoresis technologies.

    Science.gov (United States)

    Koo, O K; Mertz, A W; Akins, E L; Sirsat, S A; Neal, J A; Morawicki, R; Crandall, P G; Ricke, S C

    2013-02-01

    Cross-contamination of pathogenic and spoilage bacteria from food-contact surfaces to food products is a serious public health issue. Bacteria may survive and attach to food-contact surfaces by residual food components and/or background bacteria which may subsequently transfer to other food products. Deli slicers, generally used for slicing ready-to-eat products, can serve as potential sources for considerable bacterial transfer. The objective of this study was to assess the extent and distribution of microbial diversity of deli slicers by identification of pathogenic and background bacteria. Slicer-swab samples were collected from restaurants in Arkansas and Texas in the United States. Ten surface areas for each slicer were swabbed using sterile sponges. Denaturing gradient gel electrophoresis (DGGE) was applied to investigate the fingerprint of samples, and each band was further identified by sequence analysis. Pseudomonads were identified as the dominant bacteria followed by Enterobacteriaceae family, and lactic acid bacteria such as Lactococcus lactis and Streptococcus thermophilus were also found. Bacterial distribution was similar for all surface areas, while the blade guard exhibited the greatest diversity. This study provides a profile of the microbial ecology of slicers using DGGE to develop more specific sanitation practices and to reduce cross-contamination during slicing.

  8. Monitoring the bacterial population dynamics in sourdough fermentation processes by using PCR-denaturing gradient gel electrophoresis.

    Science.gov (United States)

    Meroth, Christiane B; Walter, Jens; Hertel, Christian; Brandt, Markus J; Hammes, Walter P

    2003-01-01

    Four sourdoughs (A to D) were produced under practical conditions by using a starter mixture of three commercially available sourdough starters and a baker's yeast constitutively containing various species of lactic acid bacteria (LAB). The sourdoughs were continuously propagated until the composition of the LAB flora remained stable. Two LAB-specific PCR-denaturing gradient gel electrophoresis (DGGE) systems were established and used to monitor the development of the microflora. Depending on the prevailing ecological conditions in the different sourdough fermentations, only a few Lactobacillus species were found to be competitive and became dominant. In sourdough A (traditional process with rye flour), Lactobacillus sanfranciscensis and a new species, L. mindensis, were detected. In rye flour sourdoughs B and C, which differed in the process temperature, exclusively L. crispatus and L. pontis became the predominant species in sourdough B and L. crispatus, L. panis, and L. frumenti became the predominant species in sourdough C. On the other hand, in sourdough D (corresponding to sourdough C but produced with rye bran), L. johnsonii and L. reuteri were found. The results of PCR-DGGE were consistent with those obtained by culturing, except for sourdough B, in which L. fermentum was also detected. Isolates of the species L. sanfranciscensis and L. fermentum were shown by randomly amplified polymorphic DNA-PCR analysis to originate from the commercial starters and the baker's yeast, respectively.

  9. Detection and differentiation of Entamoeba histolytica and Entamoeba dispar in clinical samples through PCR-denaturing gradient gel electrophoresis.

    Science.gov (United States)

    López-López, P; Martínez-López, M C; Boldo-León, X M; Hernández-Díaz, Y; González-Castro, T B; Tovilla-Zárate, C A; Luna-Arias, J P

    2017-04-03

    Amebiasis is one of the twenty major causes of disease in Mexico; however, the diagnosis is difficult due to limitations of conventional microscopy-based techniques. In this study, we analyzed stool samples using polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) to differentiate between Entamoeba histolytica (pathogenic) and E. dispar (non-pathogenic). The target for the PCR amplification was a small region (228 bp) of the adh112 gene selected to increase the sensitivity of the test. The study involved 62 stool samples that were collected from individuals with complaints of gastrointestinal discomfort. Of the 62 samples, 10 (16.1%) were positive for E. histolytica while 52 (83.9%) were negative. No sample was positive for E. dispar. These results were validated by nested PCR-RFLP (restriction fragment length polymorphism) and suggest that PCR-DGGE is a promising tool to differentiate among Entamoeba infections, contributing to determine the specific treatment for patients infected with E. histolytica, and therefore, avoiding unnecessary treatment of patients infected with the non-pathogenic E. dispar.

  10. Denaturing gradient gel electrophoresis (DGGE as a powerful novel alternative for differentiation of epizootic ISA virus variants.

    Directory of Open Access Journals (Sweden)

    Marisela Carmona

    Full Text Available Infectious Salmon Anemia is a devastating disease critically affecting world-wide salmon production. Chile has been particularly stricken by this disease which in all cases has been directly related with its causative agent, a novel orthomyxovirus which presents specific and distinctive infective features. Among these, two molecular markers have been directly associated with pathogenicity in two of the eight RNA sub genomic coding units of the virus: an insertion hot spot region present in viral segment 5 and a Highly Polymorphic Region (HPR located in viral segment 6. Here we report the successful adaptation of a PCR-dependent denaturing gel electrophoresis technique (DGGE, which enables differentiation of selected reported HPR epizootic variants detected in Chile. At the same time, the technique allows us to distinguish one nucleotide differences in sequences associated with the intriguing, and still not well-understood, insertion events which tend to occur on RNA Segment 5. Thus, the versatility of the technique opens new opportunities for improved understanding of the complex biology of all ISA variants as well as possible applications to other highly variable pathogens.

  11. Seven new mutations in hMSH2, an HNPCC Gene, identified by denaturing gradient-gel electrophoresis

    Energy Technology Data Exchange (ETDEWEB)

    Wijnen, J.; Vasen, H.; Khan, P.M.; Klift, H. van der; Leeuwen, C. van; Broek, M. van den; Leeuwen-Cornelisse, I. van; Fodde, R.; Menko, F.H. [Univ. Medical Center, Leiden (Netherlands); Nagengast, F. [Free Univ. Hospital, Amsterdam (Netherlands)

    1995-05-01

    Hereditary nonpolyposis colorectal cancer (HNPCC) is a relatively common autosomal dominant cancer-susceptibility condition. The recent isolation of the DNA mismatch repair genes (hMSH2, hMLH1, hPMS1, and hPMS2) responsible for HNPCC has allowed the search for germ-line mutations in affected individuals. In this study we used denaturing gradient-gel electrophoresis to screen for mutations in the hMSH2 gene. Analysis of all the 16 exons of HMSH2, in 34 unrelated HNPCC kindreds, has revealed seven novel pathogenic germ-line mutations resulting in stop codons either directly or through frameshifts. Additionally, nucleotide substitutions giving rise to one missense, two silent, and one useful polymorphism have been identified. The proportion of families in which hMSH2 mutations were found is 21%. Although the spectrum of mutations spread at the hMSH2 gene among HNPCC patients appears extremely heterogeneous, we were not able to establish any correlation between the site of the individual mutations and the corresponding tumor spectrum. Our results indicate that, given the genomic size and organization of the hMSH2 gene and the heterogeneity of its mutation spectrum, a rapid and efficient mutation detection procedure is necessary for routine molecular diagnosis and presymptomatic detection of the disease in a clinical setup. 34 refs., 2 figs., 3 tabs.

  12. Variations among Japanese of the factor IX gene (F9) detected by PCR-denaturing gradient gel electrophoresis

    Energy Technology Data Exchange (ETDEWEB)

    Satoh, Chiyoko; Takahashi, Norio; Asakawa, Junichi; Hiyama, Keiko; Kodaira, Meiko (Radiation Effects Research Foundation, Hiroshima (Japan))

    1993-01-01

    In the course of feasibility studies to examine the efficiencies and practicalities of various techniques for screening for genetic variations, the human coagulation factor IX (F9) genes of 63 Japanese families were examined by PCR-denaturing gradient gel electrophoresis (PCR-DGGE). Four target sequences with lengths of 983-2,891 bp from the F9 genes of 126 unrelated individuals from Hiroshima and their 100 children were amplified by PCR, digested with restriction enzymes to approximately 500-bp fragments, and examined by DGGE - a total of 6,724 bp being examined per individual. GC-rich sequences (GC-clamps) of 40 bp were attached to both ends of the target sequences, as far as was feasible. Eleven types of new nucleotide substitutions were detected in the population, none of which produced RFLPs or caused hemophilia B. By examining two target sequences in a single lane, approximately 8,000 bp in a diploid individual could be examined. This approach is very effective for the detection of variations in DNA and is applicable to large-scale population studies. 46 refs., 3 figs., 1 tab.

  13. A comparative study of the 'rhodochrous' complex and related taxa by delayed-type skin reactions on guinea pigs and by polyacrylamide gel electrophoresis.

    Science.gov (United States)

    Hyman, I S; Chaparas, S D

    1977-06-01

    Cell extracts prepared by ultrasonic disruption of 17 strains of the 'rhodochrous' complex and related taxa were compared by polyacrylamide gel electrophoresis and for immunologic relatedness, by skin test reactions. Two organisms, Jensenia canicruria and Nocardia calcarea, gave similar gel patterns and skin test reactions, and are considered to be identical. Extracts of nocardia rubra showed a strong antigenic relationship with those of three Nocardia pellegrino organisms (N325, N324 and N420) previously assigned to the 'rhodochrous' complex. Two Gordona organisms appeared to be less antigenically related to the 'rhodochrous' complex. Extracts of three of four organisms designated Lspi (Rhodococcus coprophilus Rowbotham & Cross 1976) elicited skin test reactions similar to those of the 'rhodochrous' strains. One Lspi strain, N650, showed striking similarities to the 'rhodochrous' complex strain N420 (Nocardia pellegrino).

  14. Characterization of the aerosol produced by infrared femtosecond laser ablation of polyacrylamide gels for the sensitive inductively coupled plasma mass spectrometry detection of selenoproteins

    Energy Technology Data Exchange (ETDEWEB)

    Claverie, Fanny [Laboratoire de Chimie Analytique Bio-Inorganique et Environnement, Institut Pluridisciplinaire de Recherche sur l' Environnement et les Materiaux, UMR 5254 CNRS- Universite de Pau et des Pays de l' Adour, Helioparc Pau-Pyrenees, 2 Avenue du President Angot, 64053 Pau Cedex 9 (France); Novalase SA, Z.I de la Briqueterie, 6 Impasse du bois de la Grange, 33610 Canejan (France); Pecheyran, Christophe [Laboratoire de Chimie Analytique Bio-Inorganique et Environnement, Institut Pluridisciplinaire de Recherche sur l' Environnement et les Materiaux, UMR 5254 CNRS- Universite de Pau et des Pays de l' Adour, Helioparc Pau-Pyrenees, 2 Avenue du President Angot, 64053 Pau Cedex 9 (France)], E-mail: Christophe.pecheyran@univ-pau.fr; Mounicou, Sandra [Laboratoire de Chimie Analytique Bio-Inorganique et Environnement, Institut Pluridisciplinaire de Recherche sur l' Environnement et les Materiaux, UMR 5254 CNRS- Universite de Pau et des Pays de l' Adour, Helioparc Pau-Pyrenees, 2 Avenue du President Angot, 64053 Pau Cedex 9 (France); Ballihaut, Guillaume [Laboratoire de Chimie Analytique Bio-Inorganique et Environnement, Institut Pluridisciplinaire de Recherche sur l' Environnement et les Materiaux, UMR 5254 CNRS- Universite de Pau et des Pays de l' Adour, Helioparc Pau-Pyrenees, 2 Avenue du President Angot, 64053 Pau Cedex 9 (France); Laboratoire d' Ecologie Moleculaire (Microbiologie), UMR 5254 CNRS- Universite de Pau et des Pays de l' Adour, avenue de l' Universite, B.P. 1155, F-64013 Pau (France); Fernandez, Beatriz [Laboratoire de Chimie Analytique Bio-Inorganique et Environnement, Institut Pluridisciplinaire de Recherche sur l' Environnement et les Materiaux, UMR 5254 CNRS- Universite de Pau et des Pays de l' Adour, Helioparc Pau-Pyrenees, 2 Avenue du President Angot, 64053 Pau Cedex 9 (France); Alexis, Joel [Laboratoire Genie de Production, Ecole Nationale d' Ingenieurs de Tarbes, 47 avenue d' Azereix BP 1629, 65016 Tarbes (France)] (and others)

    2009-07-15

    A 2D high repetition rate femtosecond laser ablation strategy (2-mm wide lane) previously developed for the detection of selenoproteins in gel electrophoresis by inductively coupled plasma mass spectrometry was found to increase signal sensitivity by a factor of 40 compared to conventional nanosecond ablation (0.12-mm wide lane) [G. Ballihaut, F. Claverie, C. Pecheyran, S. Mounicou, R. Grimaud and R. Lobinski, Sensitive Detection of Selenoproteins in Gel Electrophoresis by High Repetition Rate Femtosecond Laser Ablation-Inductively Coupled Plasma Mass Spectrometry, Anal. Chem. 79 (2007) 6874-6880]. Such improvement couldn't be explained solely by the difference of amount of material ablated, and then, was attributed to the aerosol properties. In order to validate this hypothesis, the characterization of the aerosol produced by nanosecond and high repetition rate femtosecond laser ablation of polyacrylamide gels was investigated. Our 2D high repetition rate femtosecond laser ablation strategy of 2-mm wide lane was found to produce aerosols of similar particle size distribution compared to nanosecond laser ablation of 0.12-mm wide lane, with 38% mass of particles < 1 {mu}m. However, at high repetition rate, when the ablated surface was reduced, the particle size distribution was shifted toward thinner particle diameter (up to 77% for a 0.12-mm wide lane at 285 {mu}m depth). Meanwhile, scanning electron microscopy was employed to visualize the morphology of the aerosol. In the case of larger ablation, the fine particles ejected from the sample were found to form agglomerates due to higher ablation rate and then higher collision probability. Additionally, investigations of the plasma temperature changes during the ablation demonstrated that the introduction of such amount of polyacrylamide gel particles had very limited impact on the ICP source ({delta}T{approx} 25 {+-} 5 K). This suggests that the cohesion forces between the thin particles composing these large

  15. IMMOBILIZATION OF Saccharomyces Cerevisiae USING POLY(ACRYLAMIDE) GEL FOR ASYMMETRIC SYNTHESIS OF R(-)-MANDELIC ACID

    Institute of Scientific and Technical Information of China (English)

    LI Zhongqin; GUO Daiping; HUANG Xinghua; YANG Kai; XU Xiaoping

    2006-01-01

    In this paper, the poly(acrylamide) hydrogel used to immobilize saccharomyces cerevisiae for asymmetric synthesis of R(-)-mandelic acid was prepared with free radical ploymerization in deionized water at room temperature under nitrogen atmosphere. The influence of the composition of hydrogel, loading amount of cells and culture conditions on the asymmetric synthesis was investigated. Results show that PAAm hydrogel is a feasible carrier for immobilization of cells which is a potential alternative method to prepare enantiomerically pure R(-)-mandelic acid.

  16. Comparative Analysis of Denaturing Gradient Gel Electrophoresis and Temporal Temperature Gradient Gel Electrophoresis Profiles as a Tool for the Differentiation of Candida Species.

    Science.gov (United States)

    Mohammadi, Parisa; Hamidkhani, Aida; Asgarani, Ezat

    2015-10-01

    Candida species are usually opportunistic organisms that cause acute to chronic infections when conditions in the host are favorable. Accurate identification of Candida species is an essential pre-requisite for improved therapeutic strategy. Identification of Candida species by conventional methods is time-consuming with low sensitivity, yet molecular approaches have provided an alternative way for early diagnosis of invasive candidiasis. Denaturing gradient gel electrophoresis (DGGE) and temporal temperature gradient gel electrophoresis (TTGE) are polymerase chain reaction (PCR)-based approaches that are used for studying the community structure of microorganisms. By using these methods, simultaneous identification of multiple yeast species will be possible and reliable results will be obtained quickly. In this study, DGGE and TTGE methods were set up and evaluated for the detection of different Candida species, and their results were compared. Five different Candida species were cultured on potato dextrose agar medium for 24 hours. Next, total DNA was extracted by the phenol-chloroform method. Two sets of primers, ITS3-GC/ITS4 and NL1-GC/LS2 were applied to amplify the desired regions. The amplified fragments were then used to analyze DGGE and TTGE profiles. The results showed that NL1-GC/LS2 primer set could yield species-specific amplicons, which were well distinguished and allowed better species discrimination than that generated by the ITS3-GC/ITS4 primer set, in both DGGE and TTGE profiles. All five Candida species were discriminated by DGGE and TTGE using the NL1-GC/LS2 primer set. Comparison of DGGE and TTGE profiles obtained from NL1-GC/LS2 amplicons exhibited the same patterns. Although both DGGE and TTGE techniques are capable of detecting Candida species, TTGE is recommended because of easier performance and lower costs.

  17. A Comparison Between Denaturing Gradient Gel Electrophoresis and Denaturing High Performance Liquid Chromatography in Detecting Mutations in Genes Associated with Hereditary Non-Polyposis Colorectal Cancer (HNPCC and the Identification of 9 New Mutations Previously Unidentified by DGGE

    Directory of Open Access Journals (Sweden)

    Meldrum Cliff J

    2003-12-01

    Full Text Available Abstract Denaturing high performance liquid chromatography is a relatively new method by which heteroduplex structures formed during the PCR amplification of heterozygote samples can be rapidly identified. The use of this technology for mutation detection in hereditary non-polyposis colorectal cancer (HNPCC has the potential to appreciably shorten the time it takes to analyze genes associated with this disorder. Prior to acceptance of this method for screening genes associated with HNPCC, assessment of the reliability of this method should be performed. In this report we have compared mutation and polymorphism detection by denaturing gradient gel electrophoresis (DGGE with denaturing high performance liquid chromatography (DHPLC in a set of 130 families. All mutations/polymorphisms representing base substitutions, deletions, insertions and a 23 base pair inversion were detected by DHPLC whereas DGGE failed to identify four single base substitutions and a single base pair deletion. In addition, we show that DHPLC has been used for the identification of 5 different mutations in exon 7 of hMSH2 that could not be detected by DGGE. From this study we conclude that DHPLC is a more effective and rapid alternative to the detection of mutations in hMSH2 and hMLH1 with the same or better accuracy than DGGE. Furthermore, this technique offers opportunities for automation, which have not been realised for the majority of other methods of gene analysis.

  18. Species identification of cooked fish by urea isoelectric focusing and sodium dodecylsulfate polyacrylamide gel electrophoresis : a collaborative study

    DEFF Research Database (Denmark)

    Rehbein, H.; Kundiger, R.; Yman, I.M.

    1999-01-01

    of gels to be run in the same flat bed electrophoresis chamber. By strictly following optimised standard operation procedures (SOPs), five unknown cooked samples had to be identified with each technique using a set of 10 raw reference samples. With urea IEF, only one out of 35 identifications......The suitability and reliability of urea IEF and SDS-PAGE for the identification of cooked fish flesh was tested by a collaborative study among nine laboratories. Urea IEF was performed with CleanGels as well as with ImmobilineGels, and ExcelGels were used for SDS-PAGE, enabling all three types...

  19. Monitoring of microbial community structure and succession in the biohydrogen production reactor by denaturing gradient gel electrophoresis (DGGE)

    Institute of Scientific and Technical Information of China (English)

    XING; Defeng; REN; Nanqi; GONG; Manli; LI; Jianzheng; LI; Q

    2005-01-01

    To study the structure of microbial communities in the biological hydrogen production reactor and determine the ecological function of hydrogen producing bacteria, anaerobic sludge was obtained from the continuous stirred tank reactor (CSTR) in different periods of time, and the diversity and dynamics of microbial communities were investigated by denaturing gradient gel electrophoresis (DGGE). The results of DGGE demonstrated that an obvious shift of microbial population happened from the beginning of star-up to the 28th day, and the ethanol type fermentation was established. After 28 days the structure of microbial community became stable, and the climax community was formed. Comparative analysis of 16S rDNA sequences from reamplifying and sequencing the prominent bands indicated that the dominant population belonged to low G+C Gram-positive bacteria (Clostridium sp. And Ethanologenbacterium sp.), β- proteobacteria (Acidovorax sp.), γ-proteobacteria (Kluyvera sp.), Bacteroides (uncultured bacterium SJA-168), and Spirochaetes (uncultured eubacterium E1-K13), respectively. The hydrogen production rate increased obviously with the increase of Ethanologenbacterium sp., Clostridium sp. And uncultured Spirochaetes after 21 days, meanwhile the succession of ethanol type fermentation was formed. Throughout the succession the microbial diversity increased however it decreased after 21 days. Some types of Clostridium sp. Acidovorax sp., Kluyvera sp., and Bacteroides were dominant populations during all periods of time. These special populations were essential for the construction of climax community. Hydrogen production efficiency was dependent on both hydrogen producing bacteria and other populations. It implied that the co-metabolism of microbial community played a great role of biohydrogen production in the reactors.

  20. Monitoring of microbial community structure and succession in the biohydrogen production reactor by denaturing gradient gel electrophoresis (DGGE).

    Science.gov (United States)

    Xing, Defeng; Ren, Nanqi; Gong, Manli; Li, Jianzheng; Li, Qiubo

    2005-04-01

    To study the structure of microbial communities in the biological hydrogen production reactor and determine the ecological function of hydrogen producing bacteria, anaerobic sludge was obtained from the continuous stirred tank reactor (CSTR) in different periods of time, and the diversity and dynamics of microbial communities were investigated by denaturing gradient gel electrophoresis (DGGE). The results of DGGE demonstrated that an obvious shift of microbial population happened from the beginning of star-up to the 28th day, and the ethanol type fermentation was established. After 28 days the structure of microbial community became stable, and the climax community was formed. Comparative analysis of 16S rDNA sequences from reamplifying and sequencing the prominent bands indicated that the dominant population belonged to low G+C Gram-positive bacteria (Clostridium sp. and Ethanologenbacterium sp.), beta-proteobacteria (Acidovorax sp.), gamma-proteobacteria (Kluyvera sp.), Bacteroides (uncultured bacterium SJA-168), and Spirochaetes (uncultured eubacterium E1-K13), respectively. The hydrogen production rate increased obviously with the increase of Ethanologenbacterium sp., Clostridium sp. and uncultured Spirochaetes after 21 days, meanwhile the succession of ethanol type fermentation was formed. Throughout the succession the microbial diversity increased however it decreased after 21 days. Some types of Clostridium sp. Acidovorax sp., Kluyvera sp., and Bacteroides were dominant populations during all periods of time. These special populations were essential for the construction of climax community. Hydrogen production efficiency was dependent on both hydrogen producing bacteria and other populations. It implied that the co-metabolism of microbial community played a great role of biohydrogen production in the reactors.

  1. Diversity and dynamics of antibiotic-resistant bacteria in cheese as determined by PCR denaturing gradient gel electrophoresis.

    Science.gov (United States)

    Flórez, Ana Belén; Mayo, Baltasar

    2015-12-01

    This work reports the composition and succession of tetracycline- and erythromycin-resistant bacterial communities in a model cheese, monitored by polymerase chain reaction denaturing gradient gel electrophoresis (PCR-DGGE). Bacterial 16S rRNA genes were examined using this technique to detect structural changes in the cheese microbiota over manufacturing and ripening. Total bacterial genomic DNA, used as a template, was extracted from cultivable bacteria grown without and with tetracycline or erythromycin (both at 25 μg ml(-1)) on a non-selective medium used for enumeration of total and viable cells (Plate Count agar with Milk; PCA-M), and from those grown on selective and/or differential agar media used for counting various bacterial groups; i.e., lactic acid bacteria (de Man, Rogosa and Sharpe agar; MRSA), micrococci and staphylococci (Baird-Parker agar; BPA), and enterobacteria (Violet Red Bile Glucose agar; VRBGA). Large numbers of tetracycline- and erythromycin-resistant bacteria were detected in cheese samples at all stages of ripening. Counts of antibiotic-resistant bacteria varied widely depending on the microbial group and the point of sampling. In general, resistant bacteria were 0.5-1.0 Log10 units fewer in number than the corresponding susceptible bacteria. The PCR-DGGE profiles obtained with DNA isolated from the plates for total bacteria and the different bacterial groups suggested Escherichia coli, Lactococcus lactis, Enterococcus faecalis and Staphylococcus spp. as the microbial types resistant to both antibiotics tested. This study shows the suitability of the PCR-DGGE technique for rapidly identifying and tracking antibiotic resistant populations in cheese and, by extension, in other foods.

  2. Dynamic changes of yak ( gut microbiota during growth revealed by polymerase chain reaction-denaturing gradient gel electrophoresis and metagenomics

    Directory of Open Access Journals (Sweden)

    Yuanyang Nie

    2017-07-01

    Full Text Available Objective To understand the dynamic structure, function, and influence on nutrient metabolism in hosts, it was crucial to assess the genetic potential of gut microbial community in yaks of different ages. Methods The denaturing gradient gel electrophoresis (DGGE profiles and Illumina-based metagenomic sequencing on colon contents of 15 semi-domestic yaks were investigated. Unweighted pairwise grouping method with mathematical averages (UPGMA clustering and principal component analysis (PCA were used to analyze the DGGE fingerprint. The Illumina sequences were assembled, predicted to genes and functionally annotated, and then classified by querying protein sequences of the genes against the Kyoto encyclopedia of genes and genomes (KEGG database. Results Metagenomic sequencing showed that more than 85% of ribosomal RNA (rRNA gene sequences belonged to the phylum Firmicutes and Bacteroidetes, indicating that the family Ruminococcaceae (46.5%, Rikenellaceae (11.3%, Lachnospiraceae (10.0%, and Bacteroidaceae (6.3% were dominant gut microbes. Over 50% of non-rRNA gene sequences represented the metabolic pathways of amino acids (14.4%, proteins (12.3%, sugars (11.9%, nucleotides (6.8%, lipids (1.7%, xenobiotics (1.4%, coenzymes, and vitamins (3.6%. Gene functional classification showed that most of enzyme-coding genes were related to cellulose digestion and amino acids metabolic pathways. Conclusion Yaks’ age had a substantial effect on gut microbial composition. Comparative metagenomics of gut microbiota in 0.5-, 1.5-, and 2.5-year-old yaks revealed that the abundance of the class Clostridia, Bacteroidia, and Lentisphaeria, as well as the phylum Firmicutes, Bacteroidetes, Lentisphaerae, Tenericutes, and Cyanobacteria, varied more greatly during yaks’ growth, especially in young animals (0.5 and 1.5 years old. Gut microbes, including Bacteroides, Clostridium, and Lentisphaeria, make a contribution to the energy metabolism and synthesis of amino

  3. Association of Streptomyces community composition determined by PCR-denaturing gradient gel electrophoresis with indoor mold status.

    Science.gov (United States)

    Johansson, Elisabet; Reponen, Tiina; Meller, Jarek; Vesper, Stephen; Yadav, Jagjit

    2014-12-01

    Both Streptomyces species and mold species have previously been isolated from moisture-damaged building materials; however, an association between these two groups of microorganisms in indoor environments is not clear. In this study, we used a culture-independent method, PCR-denaturing gradient gel electrophoresis (PCR-DGGE), to investigate the composition of the Streptomyces community in house dust. Twenty-three dust samples each from two sets of homes categorized as high-mold and low-mold based on mold-specific quantitative PCR analysis were used in the study. Taxonomic identification of prominent bands was performed by cloning and sequencing. Associations between DGGE amplicon band intensities and home mold status were assessed using univariate analyses as well as multivariate recursive partitioning (decision trees) to test the predictive value of combinations of bands intensities. In the final classification tree, a combination of two bands was significantly associated with mold status of the home (p = 0.001). The sequence corresponding to one of the bands in the final decision tree matched a group of Streptomyces species that included Streptomyces coelicolor and Streptomyces sampsonii, both of which have been isolated from moisture-damaged buildings previously. The closest match for the majority of sequences corresponding to a second band consisted of a group of Streptomyces species that included Streptomyces hygroscopicus, an important producer of antibiotics and immunosuppressors. Taken together, the study showed that DGGE can be a useful tool for identifying bacterial species that may be more prevalent in mold-damaged buildings.

  4. Characterization of Sm14 related components in different helminths by sodium dodecyl sulphate-polyacrylamide gel electrophoresis and Western blotting analysis

    Directory of Open Access Journals (Sweden)

    Nilton Thaumaturgo

    2002-10-01

    Full Text Available Sm14 was the first fatty acid-binding protein homologue identified in helminths. Thereafter, members of the same family were identified in several helminth species, with high aminoacid sequence homology between them. In addition, immune crossprotection was also reported against Fasciola hepatica infection, in animals previously immunized with the Schistosoma mansoni vaccine candidate, r-Sm14. In the present study, data on preliminary sodium dodecyl sulphate-polyacrylamide gel electrophoresis and Western blotting analysis of nine different helminth extracts focusing the identification of Sm14 related proteins, is reported. Out of these, three extracts - Ascaris suum (males and females, Echinostoma paraensei, and Taenia saginata - presented components that comigrated with Sm14 in SDS-PAGE, and that were recognized by anti-rSm14 policlonal serum, in Western blotting tests.

  5. Synthesis, Characterization and Electromagnetic Studies on Nanocrystalline Co0.5Zn0.5Fe2O4 Synthesized by Polyacrylamide Gel

    Institute of Scientific and Technical Information of China (English)

    Ruiting MA; Yanwen TIAN; Haitao ZHAO; Gang ZHANG; Hui ZHAO

    2008-01-01

    Nanocrystalline Co0.5Zn0.5Fe2O4 ferrite was synthesized by polyacrylamide gel method. The electromagnetic and microwave absorption properties of the ferrite were investigated. The results indicated that calcining temperature of the ferrite had a significant influence on the effective properties of the ferrite. When the calcining temperature was 500,600 and 700℃, the average size of particles was 10, 30 and 80 nm, respectively. The dielectric loss (ε") and magnetic loss (u") of the ferrite was around 0.65 and 0.29 at 8.2 GHz, respectively. Microwave absorption properties of the ferrites were simultaneously influenced due to the strong correlation between reflection loss and electromagnetic parameters of the ferrite.

  6. Studies on middle silkgland proteins of cocoon colour sex-limited silkworm (Bombyx mori L.) using two-dimensional polyacrylamide gel electrophoresis

    Indian Academy of Sciences (India)

    Yuan-Xiang Jin; Yu-Yin Chen; Meng-Kui Xu; Yong-Huang Jiang

    2004-03-01

    Qualitative and quantitative differences in proteins expressed in the middle silkglands of male and female silkworm larvae that differ in silk colour were investigated by high resolution two-dimensional polyacrylamide gel electrophoresis (2-D PAGE), followed by computer assisted image analysis. About 1000 protein spots were resolved in both the sexes and most proteins were shown to be distributed in the area from 15 kDa to 70 kDa and pH 4–8. It was found that some proteins displayed higher expression in yellow cocoon, while two proteins were only expressed in female silkworm silkgland tissue through the comparison and analysis by two-D software. These proteins especially existed in female silkworm middle silkgland tissue of yellow cocoon. Furthermore, these proteins might be involved in the expression of cocoon colour phenotype.

  7. Micelle-induced curvature in a water-insoluble HIV-1 Env peptide revealed by NMR dipolar coupling measurement in stretched polyacrylamide gel.

    Science.gov (United States)

    Chou, James J; Kaufman, Joshua D; Stahl, Stephen J; Wingfield, Paul T; Bax, Ad

    2002-03-20

    The structure of a water-insoluble fragment encompassing residues 282-304 of the HIV envelope protein gp41 is studied when solubilized by dihexanoyl phosphatidylcholine (DHPC) and by small bicelles, consisting of a 4:1 molar ratio of DHPC and dimyristoyl phosphatidylcholine (DMPC). Weak alignment with the magnetic field was accomplished in a stretched polyacrylamide gel, permitting measurement of one-bond (1)H-(15)N, (13)Ca-(13)C', and (13)C'-(15)N dipolar couplings, which formed the basis for determining the peptide structure. In both detergent systems, the peptide adopts an alpha-helical conformation from residue 4 through 18. In the presence of the DHPC micelles the helix is strongly curved towards the hydrophobic surface, whereas in the presence of bicelles a much weaker curvature in the opposite direction is observed.

  8. Efficacy and compatibility with mass spectrometry of methods for elution of proteins from sodium dodecyl sulfate-polyacrylamide gels and polyvinyldifluoride membranes

    DEFF Research Database (Denmark)

    Jørgensen, C.S.; Jagd, M.; Sørensen, B.K.;

    2004-01-01

    for recovering intact proteins from polyacrylamide gels and electroblotting membranes to define efficient methods compatible with MS. These methods complement in situ digestion protocols and allow determination of the molecular mass of whole proteins separated by SDS-PAGE. Passive elution of proteins from SDS...... acid (TFA) or combinations of 8 M urea and 1% Triton X-100, 1% Tween 20, or 40% isopropanol. The same result was obtained using nitrocellulose membranes, except that these were incompatible with organic solvent and TFA. Elution by TFA was compatible with matrix-assisted laser desorption/ionization MS...... (MALDI-MS) but was complicated by a high degree of trifluoroacetylation of the proteins. Alternatively, elution by 8 M urea + 1% Triton X-100, 1% Tween 20, or 40% isopropanol was compatible with both SELDI-MS and MALDI-MS. Eluted proteins were identified in MS experiments by intact mass determination...

  9. Analysis of polyacrylamide gels for trace metals using diffusive gradients in thin films and laser ablation inductively coupled plasma mass spectrometry.

    Science.gov (United States)

    Warnken, Kent W; Zhang, Hao; Davison, William

    2004-10-15

    A simple method for the analysis of polyacrylamide diffusive gradients in thin film (DGT) gels by laser ablation inductively coupled plasma mass spectrometry (LA-ICPMS), employing a novel use of (115)In internal standardization, has been developed. This method allows the determination of Co, Ni, Cu, Zn, Cd, and Pb concentrations (at the DGT filter face) or fluxes in sediments at a spatial resolution of 100 microm. Single-layered gels, using an optimized laser defocus of 4000 microm at 400 mJ power, showed high precision (generally approximately 10%) and a linear response during solution deployment. Of the elements Sc, In, Ba, La, Ce, and Tb, Ba most closely tracked variations in laser energy and showed the highest analytical precision but could not be used as an internal standard due to its elevated presence in natural sediments. Therefore, internal standardization, necessary to normalize data collected on different days, was carried out using (115)In contained within a second layer of backing gel and dried along with the analyte layer as a dual-gel disk. This multilayered gel standard required a laser defocus setting of 1000 microm and a laser power of approximately 800 mJ. Analytical precision for a 64-spot ablation grid at 100-microm spacing was approximately 10%. Verification of this method was carried out on DGT sediment probes deployed in Priest Pot (English Lake District). Results obtained by conventional slicing techniques and aqueous elution agreed with laser ablation results when the different sampling areas were considered. The elution results varied by a factor of laser ablation technique showed a variability of approximately 4, indicating localized elevated concentrations of Co. This higher resolution LA-ICPMS method could ultimately lead to an improved understanding of the geochemical processes responsible for metal uptake and release in sediments.

  10. Detection of metals in proteins by means of polyacrylamide gel electrophoresis and laser ablation-inductively coupled plasma-mass spectrometry: application to selenium.

    Science.gov (United States)

    Chéry, Cyrille C; Günther, Detlef; Cornelis, Rita; Vanhaecke, Frank; Moens, Luc

    2003-10-01

    The capabilities of laser ablation-inductively coupled plasma-mass spectrometry for the detection of trace elements in a gel after gel electrophoresis were systematically studied. Figures of merit, such as limit of detection, linearity, and repeatability, were evaluated for various elements (Li, V, Cr, Mn, Ni, Cu, Zn, As, Se, Mo, Pd, Ag, Cd, Pt, Tl, Pb). Two ablation strategies were followed: single hole drilling, relevant for ablation of spots after two-dimensional (2-D) separations, and ablation with translation, i.e., on a line, relevant for one-dimensional (1-D) separations. This technique was applied to the detection of selenoproteins in red blood cells extracts after a 1-D separation (sodium dodecyl sulfate-polyacrylamide gel electrophoresis) and the detection of selenium-containing proteins in yeast after 2-D electrophoresis (2-DE). The detection procedure was further improved by using the dynamic reaction cell technology, which allowed the removal of the Ar_2(+) interference and hence the use of the most abundant Se isotope, (80)Se. Reaction gases were compared (methane, carbon monoxide, ammonia, oxygen and the combination of argon (collision gas) and hydrogen (reaction gas)). In each instance, the reaction cell parameters were optimized in order to obtain the lowest detection limit for Se (as (80)Se(+), (82)Se(+) or (77)Se(+); and as (80)Se(16)O(+), (82)Se(16)O(+) or (77)Se(16)O(+) with O(2) as the reaction gas). Carbon monoxide was found to offer the best performance. The detection limit with the use of DRC and He as transport gas was 0.07 microg Se g(-1) gel with single hole drilling and 0.15 microg Se g(-1) gel for ablation with translation.

  11. Evaluation of polyacrylamide gels with accelerator ammonium salts for water shutoff in ultralow temperature reservoirs: Gelation performance and application recommendations

    Directory of Open Access Journals (Sweden)

    Hu Jia

    2016-03-01

    Full Text Available Water shutoff in ultralow temperature reservoirs has received great attention in recent years. In previous study, we reported a phenol-formaldehyde-based gel formula with ammonium salt which can provide a gelation time between 2 hrs and 2 days at 25 °C. However, systematic evaluation and field recommendations of this gel formula when encountering complex reservoirs environment are not addressed. In this paper, how and why such practical considerations as water composition, temperature, pH, weight ratio of formaldehyde to resorcinol and contaminant Fe3+ to affect the gelation performance are examined. Brookfield DV-III and scanning electron microscopy (SEM are employed respectively for viscosity measurement and microstructure analysis. SEM results further illustrate the mechanism of the effect of salinity on gelation performance. It reveals that crosslinking done by covalent bond has great advantage for gel stability under high salinity environment. The target gel formula can provide desirable gelation time below 60 °C, perfect for 15–45 °C, while it is unfeasible to use high salinity to delay gelation at 60 °C. We summarized the effect of salinity on gelation performance of different gel formulas from the present study and published literature. The summarized data can provide important guideline for gel formula design before conducting any kinds of experiments. The variation of gelation performance at different salinity may be dominated by the interaction between crosslinker-salt-polymer, not only limited to “charge-screening effect” and “ion association” proposed by several authors. We hope the analysis encouraging further investigations. Some recommendations for field application of this gel are given in the end of this paper.

  12. An optical comparator for measuring two-dimensional polyacrylamide gel electrophoresis records using an on-line microcomputer.

    Science.gov (United States)

    Spragg, S P; Jones, M I; Hill, B J

    1983-03-01

    A comparator which makes it possible to compare two wet gels or photographic negatives or autoradiograms through a flickering light system has been built. The system consists of two special-purpose projectors which combine the images on a digitizing platform. When the lights are switched On and off out of phase, the positions of the common components remain unchanged, whereas those that are spatially displaced appear to jump from side to side and those present in one image but not the other switch on and off. This produces a flickering image in which differences are readily seen. Commercial camera lenses were used to construct the projectors and the overall specifications for the system are given. The coordinates of both the displaced components, as well as the selected standards from the two images, are digitized and entered automatically into an on-line microcomputer. By using an iterative procedure for collecting records from several superimposable records of the gel, it is possible to compensate for the lack of total reproducibility over the whole gels. These coordinates are then normalized and superimposed on a master map through a television display using a curser to adjust the coordinates. The whole procedure can be repeated for many gels using a common reference gel in the comparator, and the result is a set of normalized coordinates which can be plotted on a single map to provide a final record of the experiments.

  13. Development of a sodium dodecyl sulfate-polyacrylamide gel electrophoresis reference method for the analysis and identification of fish species in raw and heat-processed samples : A collaborative study

    DEFF Research Database (Denmark)

    Pineiro, C.; Barros-Velazquez, J.; Perez-Martin, R.I.;

    1999-01-01

    A collaborative study was carried out in seven European labs with the aim of achieving a sodium dodecyl sulfate- polyacrylamide gel electrophoresis (SDS-PAGE) standard operation procedure to identify fish species in raw and cooked samples. Urea and SDS-containing solutions were evaluated as extra...

  14. Development of a sodium dodecyl sulfate-polyacrylamide gel electrophoresis reference method for the analysis and identification of fish species in raw and heat-processed samples : A collaborative study

    DEFF Research Database (Denmark)

    Pineiro, C.; Barros-Velazquez, J.; Perez-Martin, R.I.

    1999-01-01

    A collaborative study was carried out in seven European labs with the aim of achieving a sodium dodecyl sulfate- polyacrylamide gel electrophoresis (SDS-PAGE) standard operation procedure to identify fish species in raw and cooked samples. Urea and SDS-containing solutions were evaluated as extra...

  15. Comparison of diazo-coupling, formazan, and silver staining techniques for visualizing alkaline phosphatase isoenzymes after electrophoresis in homogeneous-pore and gradient-pore polyacrylamide gels.

    Science.gov (United States)

    Hodson, A W; Skillen, A W

    1988-03-01

    Three techniques for visualization of alkaline phosphatase after polyacrylamide-gel electrophoresis are compared. These are diazo-dye simultaneous coupling with the substrate sodium naphthyl phosphate and 5-chloro-2-toluene diazonium chloride; formazan precipitation with the substrate 5-bromo-4-chloro-3-indolyl phosphate and 3-[4,5-dimethylthiazolyl-2)-2,5-diphenyltetrazolium bromide; and silver staining with the substrate sodium glycerophosphate. Each staining technique was tested with gradient-pore and homogeneous-pore acrylamide-gel electrophoresis. The main factors assessed are sensitivity; separation of the human serum alkaline phosphatase isoenzymes of the liver, bone, and intestinal types; and differences in substrate affinity, as well as the complexity of each technique. Using the three techniques only minor differences in substrate affinity are evident. There is some nonspecific staining with the diazo-coupling technique but not with the formazan and silver staining techniques. The differences, in the mobility of the liver, bone, and intestinal isoenzymes achieved by homogeneous-pore gel electrophoresis are sufficient to allow them to be clearly distinguished. However, only very small differences in mobility are found with gradient-pore gel electrophoresis, but the sharper bands in this medium allow much smaller amounts of activity to be detected. As little as 160 microU of enzyme can be visualized by the diazo technique. Silver staining gives an approximately fourfold increase in sensitivity over the formazan technique, which in turn gives a fourfold increase over the diazo technique. An important aspect of the silver staining technique is the potential of increasing sensitivity much further by improvements in the photographic physical development stage.(ABSTRACT TRUNCATED AT 250 WORDS)

  16. Species identification of cooked fish by urea isoelectric focusing and sodium dodecylsulfate polyacrylamide gel electrophoresis : a collaborative study

    DEFF Research Database (Denmark)

    Rehbein, H.; Kundiger, R.; Yman, I.M.;

    1999-01-01

    of gels to be run in the same flat bed electrophoresis chamber. By strictly following optimised standard operation procedures (SOPs), five unknown cooked samples had to be identified with each technique using a set of 10 raw reference samples. With urea IEF, only one out of 35 identifications...

  17. Concentration of acrylamide in a polyacrylamide gel affects VP4 gene coding assignment of group A equine rotavirus strains with P[12] specificity

    Science.gov (United States)

    2010-01-01

    Background It is universally acknowledged that genome segment 4 of group A rotavirus, the major etiologic agent of severe diarrhea in infants and neonatal farm animals, encodes outer capsid neutralization and protective antigen VP4. Results To determine which genome segment of three group A equine rotavirus strains (H-2, FI-14 and FI-23) with P[12] specificity encodes the VP4, we analyzed dsRNAs of strains H-2, FI-14 and FI-23 as well as their reassortants by polyacrylamide gel electrophoresis (PAGE) at varying concentrations of acrylamide. The relative position of the VP4 gene of the three equine P[12] strains varied (either genome segment 3 or 4) depending upon the concentration of acrylamide. The VP4 gene bearing P[3], P[4], P[6], P[7], P[8] or P[18] specificity did not exhibit this phenomenon when the PAGE running conditions were varied. Conclusions The concentration of acrylamide in a PAGE gel affected VP4 gene coding assignment of equine rotavirus strains bearing P[12] specificity. PMID:20573245

  18. Detection of the End Point Temperature of Thermal Denatured Protein in Fish and Chicken Meat Through SDS-PAGE Electrophoresis

    Institute of Scientific and Technical Information of China (English)

    GAO Hongwei; MAO Mao; LIANG Chengzhu; LIN Chao; XIANG Jianhai

    2009-01-01

    Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) was applied in the detection of the end point temperature (EPT) of thermal denatured protein in fish and meat in this study. It was also used in studying the thermal denatured temperature range of proteins in salmon and chicken meat. The results show that the temperature ranges of denatured proteins were from 65℃ to 75℃, and these temperature ranges were influenced by the processing methods. Through SDS-PAGE, the features of repeated heating thermal denatured proteins under the same temperature and processing time were studied. The electrophoresis pat-terns of thermal denatured proteins determined through repeated heating at the same temperature did not exhibit any change. For the detection of cooked fish and meat samples, they were subjected to applying the SDS-PAGE method, which revealed an EPT ranging from 60℃ to 80℃.

  19. Semi-crosslinked polyacrylamides as high-resolution and dynamic self-coating sieving matrices for protein capillary electrophoresis

    Institute of Scientific and Technical Information of China (English)

    ZHOU Jin; XU JianDong; XIE Yao; QU Feng; DENG YuLin; GENG LiNa

    2008-01-01

    This paper describes non-gel capillary sieving electrophoresis employing semi-crosslinked poly-acrylamide as a high performance and low viscous replaceable separation matrix for separation of non-denatured protein separation. Arising from the fine sieving and dynamic coating ability of this polymer, a mixture of basic proteins lysozyme, cytochrome C, ribonuclease A, and trypsin was resolved with excellent reproducibility. Mixing different semi-crosslinked polyacrylamides together further im-proves the separation. The separtion mechanism was analyzed. With network structure developed to an intermediate state between crosslinked gel and linear polymer solutions, these semi-crosslinked polyacrylamide polymers demonstrate a promise as a new class of size sieving separation medium, not only in capillary electrophoresis, but also in microfluidic chip separation schemes.

  20. A rapid and simple 8-quinolinol-based fluorescent stain of phosphoproteins in polyacrylamide gel after electrophoresis.

    Science.gov (United States)

    Wang, Xu; Hwang, Sun-Young; Cong, Wei-Tao; Jin, Li-Tai; Choi, Jung-Kap

    2015-10-01

    In order to obtain an easy and rapid protocol to visualize phosphoproteins in SDS-PAGE, a fluorescent detection method named 8-Quinolinol (8-Q) stain is described. 8-Q can form ternary complexes in the gel matrix contributed by the affinity of aluminum ion (Al(3+) ) to the phosphate groups on the proteins and the metal chelating property of 8-Quinolinol, exhibiting strong fluorescence in ultraviolet light. It can visualize as little as 4∼8 ng of α-casein and β-casein, 16∼32 ng of ovalbumin and κ-casein which is more sensitive than Stains-All but less sensitive than Pro-Q Diamond. The protocol of 8-Q requires only 70 min in 0.75 mm mini-size or 1.0 mm large-size gels with five changes of solutions without destaining step; Pro-Q takes at least 250 min with 11 changes of solutions. In addition, the new method was confirmed by the study of dephosphorylation and LC-MS/MS, respectively. The approach to visualize phosphoprotein utilizing 8-Q could be an alternative to simplify the analytical operations for phosphoproteomics research. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  1. Assessment of the yeast species composition of cocoa bean fermentations in different cocoa-producing regions using denaturing gradient gel electrophoresis.

    Science.gov (United States)

    Papalexandratou, Zoi; De Vuyst, Luc

    2011-11-01

    The yeast species composition of 12 cocoa bean fermentations carried out in Brazil, Ecuador, Ivory Coast and Malaysia was investigated culture-independently. Denaturing gradient gel electrophoresis of 26S rRNA gene fragments, obtained through polymerase chain reaction with universal eukaryotic primers, was carried out with two different commercial apparatus (the DCode and CBS systems). In general, this molecular method allowed a rapid monitoring of the yeast species prevailing during fermentation. Under similar and optimal denaturing gradient gel electrophoresis conditions, the CBS system allowed a better separated band pattern than the DCode system and an unambiguous detection of the prevailing species present in the fermentation samples. The most frequent yeast species were Hanseniaspora sp., followed by Pichia kudriavzevii and Saccharomyces cerevisiae, independent of the origin of the cocoa. This indicates a restricted yeast species composition of the cocoa bean fermentation process. Exceptionally, the Ivorian cocoa bean box fermentation samples showed a wider yeast species composition, with Hyphopichia burtonii and Meyerozyma caribbica among the main representatives. Yeasts were not detected in the samples when the temperature inside the fermenting cocoa pulp-bean mass reached values higher than 45 °C or under early acetic acid production conditions.

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

    Energy Technology Data Exchange (ETDEWEB)

    Magre, J.; Goldfine, A.B.; Warram, J.H. [Harvard Medical School, Boston, MA (United States)] [and others

    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.

  3. Development of a low-cost, high-throughput native polyacrylamide gel electrophoresis (N-PAGE) protocol for lipoprotein sub-fractionation using Quality by Design approach.

    Science.gov (United States)

    Pathak, Mili; Chaudhary, Neha; Rathore, Anurag S

    2014-04-01

    Ratio of low density to high density lipoprotein concentration is critical for normal functioning of human body. Deviation in this ratio has been linked to various diseases, many of which are fatal if not diagnosed at early stages. For example, cardiovascular diseases (CVD) have been linked to the level of low density lipoprotein (LDL). Henceforth, detection of the lipoprotein subtractions is crucial for health of an individual. To date, methods like ultracentrifugation, nuclear magnetic resonance (NMR), high performance liquid chromatography (HPLC) and gradient gel electrophoresis (GGE) have been used for separation and identification of lipoprotein types and subtypes. However, these methods are expensive, time consuming and require specialized equipments and expertise. This paper aims to propose a low-cost, high-throughput native polyacrylamide gel electrophoresis (N-PAGE) based protocol for analysis of lipoproteins. Quality by Design (QbD) based approach has been utilized. The initial screening of parameters was followed by a fractional factorial design to optimize the protocol. The lipoprotein subtractions obtained by the optimized protocol were compared with the commercially available and commonly used Lipoprint(®) Lipoprotein Subfractions Testing System from Quantimetrix. The proposed method gave comparable results to those obtained with the commercial system. The proposed method is capable of analysis of up to forty different samples in two hours at a cost of approximately 2$/sample. This is an order of magnitude better than the present cost of 265$/sample when using the commercial system. We think that the proposed method would be of particular interest to the developing and under-developed economies of the world, where this cost differential would be deemed quite significant and would make testing affordable to the majority of the population. Copyright © 2014 Elsevier B.V. All rights reserved.

  4. Synthesis and electrical field-assisted sintering behaviour of yttria-stabilized tetragonal ZrO$_2$ nanopowders by polyacrylamide gel method

    Indian Academy of Sciences (India)

    XINGHUA SU; BENPAN WANG; JIE ZHOU; HAOYU SUN

    2016-06-01

    The tetragonal ZrO$_2$ nanopowders stabilized with 3 mol% Y$_2$O$_3$ (3YSZ) were synthesized using a polyacrylamide gel method. The mean particle size of the 3YSZ nanopowders was found to decrease with increasing molecular ratio of monomer to the precursor salt. The 3YSZ nanopowders with mean particle size of 12 nm can be densified in 1 h at 800$^{\\circ}$C, by the application of a d.c. electrical field. Under a constant d.c. electrical field, the current density through the specimen of 3YSZ rose rapidly when the temperature increased to a certain value. In the sintering process, the current density was restricted when the sharp increase occurred. By limiting current density to different values for one hour, it was found that current density was the most important factor in electrical field assisted sintering process. The grain size of 3YSZ bulk increased with the enhanced current density. The stable stageof electrical field-assisted sintering process can be explained by Joule heating. Corresponding real temperature of specimens is estimated by applying black body radiation theory.

  5. Characterization of bacterial populations in Danish raw milk cheeses made with different starter cultures by denaturating gradient gel electrophoresis and pyrosequencing

    DEFF Research Database (Denmark)

    Masoud, Wafa Mahmoud Hasan; Takamiya, Monica K Wik; Vogensen, Finn Kvist;

    2011-01-01

    The bacterial populations in Danish raw milk cheeses were identified using denaturating gradient gel electrophoresis (DGGE) of PCR amplicons of the V3 region of the 16S rRNA gene and pyrosequencing of tagged amplicons of the V3 and V4 regions of the 16S rRNA gene. Both DNA and RNA extracted from...... cheeses were studied in order to determine the metabolically active bacteria. The main bacteria, which included Lactococcus, Lactobacillus and Streptococcus, were detected by pyrosequencing and DGGE in both 16S rDNA and cDNA obtained from cheeses indicating their viability and contribution to cheese...... ripening. Other bacteria like Corynebacterium, Halomonas, Pediococcus, Micrococcus and Staphylococcus, which were encountered in some cheese samples at low percentages compared with the total bacterial populations, were only detected by pyrosequencing. 16S rRNA gene pyrosequencing is an efficient method...

  6. Direct detection and identification of lactic acid bacteria in a food processing plant and in meat products using denaturing gradient gel electrophoresis.

    Science.gov (United States)

    Takahashi, Hajime; Kimura, Bon; Yoshikawa, Miwako; Gotou, Seitaro; Watanabe, Itaru; Fujii, Tateo

    2004-11-01

    We established a novel system using denaturing gradient gel electrophoresis (DGGE) to quickly identify bacteria known to be responsible for spoilage in meat processing plants and meat products. We extracted bacterial DNA from swabbed samples at various locations in the plant and from meat products and performed PCR amplification, targeting 16S rDNA from the dominant organisms. The amplification products were subjected to DGGE, and the contaminating bacteria in the meat products and the plant were analyzed. This analysis indicated that lactic acid bacteria and spoilage-causing bacteria are widely distributed within the meat processing plant. We developed molecular size markers to identify the dominant organisms obtained from the plant and meat products. The establishment of the present method allows quick and simple identification of bacteria causing the possible deterioration of products and contamination and thus permits constant monitoring of any harmful bacteria within meat processing plants.

  7. Analysis of phosphate-accumulating organisms cultivated under different carbon sources with polymerase chain reaction-denaturing gradient gel electrophoresis assay

    Institute of Scientific and Technical Information of China (English)

    YU Shui-li; LIU Ya-nan; JING Guo-lin; ZHAO Bing-jie; GUO Si-yuan

    2005-01-01

    To investigate the microbial communities of microorganisms cultivated under different carbon sources, three sequencing batch reactors were operated. They were supplied with sewage, glucose and sodium acetate as carbon sources respectively and showed high phosphorus removal performance. The results of denaturing gradient gel electrophoresis(DGGE) of polymerase chain reaction-amplified (PCR) 16S rDNA fragments demonstrated that β-protebacteria, Actinomyces sp. and γ-protebacteria only exited in 1 # reactor. The microbiological diversity of 1 # reactor exceeded the other two reactors. Flavobacterium, Bacillales, Actinomyces, Actinobacteridae and uncultured bacteria(AF527584, AF502204, AY592749, AB076862, AJ619051, AF495454 and AY133070) could be detected in the biological phosphorus removal reactors.

  8. Comparasion of polyacrylamide gel electrophoresis (PAGE, immuno-electron microscopy (IEM and enzyme immunoassay (EIA for the rapid diagnosis of rotavirus infection in children

    Directory of Open Access Journals (Sweden)

    H. G. Pereira

    1983-12-01

    Full Text Available Detection of rotavirus RNA by polyacrylamide gel electrophoresis (PAGE proved to be a highly sensitive and rapid diagnostic test. A comparison of this assay with immuno-electron microscopy (IEM and enzyme immunoassay (EIA in 245 faeces from children with gastroenteritis revealed complete agreement between the three assays in 238 (97.14% samples. Among 75 samples positive in at least one of the three assays, negative results were observed in 5 (6.48% by PAGE, in 6 (6.76% by EIA and in none by IEM. Silver staining greatly increased the sensitivity of the PAGE assay. We conclude that although IEM remains the most sensitive and rapid rotavirus diagnostic assay, the PAGE technique has many advantages in its favour, including the non-requirement of expensive equipment, the use of only chemically defined reagents and the capacity to distinguish virus subgroup and variants and to detect non-crossreactive rotaviruses which are missed in serological assays.A evidenciação da presença de ácido ribonucleico (ARN viral por eletroforese em gel de policrilamida (EGPA foi comprovada como um método altamente sensível e rápido para o diagnóstico de infecções por rotavirus. Uma comparação desta prova com a imunomicroscopia eletrônica (IEM e com o ensaio imunoenzimático (EIE no exame de 245 fezes de crianças com gastroenterite revelou completa concordancia entre os três ensaios em 238 (97.14% amostras. Entre 75 amostras positivas pelo menos em um dos três ensaios, resultados negativos foram observados em 5 (6.48% por EGPA, em 6 (6.76% por EIE e em nenhum por IEM. Coloração pela prata aumentou consideravelmente a sensibilidade do ensaio por EGPA. Concluímos que embora a IEM ainda seja a prova mais sensível e rápida para o diagnóstico de infecções por rotavirus, o ensaio por EGPA tem muitas vantagens em seu favor, sendo as principais as de não necessitar equipamentos caros, de empregar exclusivamente reagentes quimicamente definidos, de

  9. EXAFS analysis of a human Cu,Zn SOD isoform focused using non-denaturing gel electrophoresis

    Energy Technology Data Exchange (ETDEWEB)

    Chevreux, Sylviane; Roudeau, Stephane; Deves, Guillaume; Ortega, Richard [Laboratoire de Chimie Nucleaire Analytique et Bioenvironnementale, CNRS UMR5084, Universite Bordeaux 1, Chemin du Solarium, F-33175 Gradignan cedex (France); Solari, Pier Lorenzo [Synchrotron SOLEIL, L' Orme des Merisiers, BP 48, F-91192 Gif-sur-Yvette cedex, Saint-Aubin (France); Alliot, Isabelle; Testemale, Denis; Hazemann, Jean Louis, E-mail: ortega@cenbg.in2p3.f [FAME, ESRF, 6 rue Jules Horowitz, BP220, F-38043 Grenoble cedex (France)

    2009-11-15

    Isoelectric point isoforms of a metalloprotein, copper-zinc superoxide dismutase (CuZnSOD), separated on electrophoresis gels were analyzed using X-ray Absorption Spectroscopy. Mutations of this protein are involved in familial cases of amyotrophic lateral sclerosis. The toxicity of mutants could be relied to defects in the metallation state. Our purpose is to establish analytical protocols to study metallation state of protein isoforms such as those from CuZnSOD. We previously highlighted differences in the copper oxidation state between CuZnSOD isoforms using XANES. Here, we present the first results for EXAFS analyses performed at Cu and Zn K-edge on the majoritary expressed isoform of human CuZnSOD separated on electrophoresis gels.

  10. EXAFS analysis of a human Cu,Zn SOD isoform focused using non-denaturing gel electrophoresis

    Science.gov (United States)

    Chevreux, Sylviane; Solari, Pier Lorenzo; Roudeau, Stéphane; Deves, Guillaume; Alliot, Isabelle; Testemale, Denis; Hazemann, Jean Louis; Ortega, Richard

    2009-11-01

    Isoelectric point isoforms of a metalloprotein, copper-zinc superoxide dismutase (CuZnSOD), separated on electrophoresis gels were analyzed using X-ray Absorption Spectroscopy. Mutations of this protein are involved in familial cases of amyotrophic lateral sclerosis. The toxicity of mutants could be relied to defects in the metallation state. Our purpose is to establish analytical protocols to study metallation state of protein isoforms such as those from CuZnSOD. We previously highlighted differences in the copper oxidation state between CuZnSOD isoforms using XANES. Here, we present the first results for EXAFS analyses performed at Cu and Zn K-edge on the majoritary expressed isoform of human CuZnSOD separated on electrophoresis gels.

  11. Application of nested PCR-DGGE (denaturing gradient gel electrophoresis) for the analysis of ciliate communities in soils.

    Science.gov (United States)

    Shimano, Satoshi; Sambe, Mitsuo; Kasahara, Yasuhiro

    2012-01-01

    Ciliates play important roles as prey and predators in ecosystems. Changes in the ciliate community can affect the composition and population of microfauna and microflora in ecosystems. To investigate the structure of ciliate communities, we developed a nested PCR-DGGE method, which combines a universal eukaryotic-specific primer set in the first PCR step with a ciliate-specific primer set in the second PCR step, to amplify 18S rRNA genes from ciliates. The 300 bp DGGE fragments generated more bands on the gel than the 600 bp DGGE fragments. Prior to bead beating, DNA extraction of ciliates from soil samples was optimized with a combination of freeze-thaw cycles and ultrasonication. We applied this nested PCR-DGGE method to agricultural soils amended with 0, 120, 300, and 600 t ha⁻¹ year⁻¹ of livestock slurry. The results from the DGGE profiles and principal component analysis (PCA) revealed that the supplement of slurry to soils influenced the ciliate communities. From phylogenetic analysis, 108 DGGE bands were assigned to six classes, which included Spirotrichea and Colpodea, of the subphylum Intramacronucleata, and one class of the subphylum Postciliodesmatophora. These results indicated that a wide variety of taxonomic groups were detected by DGGE profiling. Thus, the nested PCR-DGGE method described here could clearly differentiate between ciliate communities within soil samples and allowed for the phylogenetic identification of these ciliates at the class level.

  12. Parallel characterization of anaerobic toluene- and ethylbenzene-degrading microbial consortia by PCR-denaturing gradient gel electrophoresis, RNA-DNA membrane hybridization, and DNA microarray technology

    Science.gov (United States)

    Koizumi, Yoshikazu; Kelly, John J.; Nakagawa, Tatsunori; Urakawa, Hidetoshi; El-Fantroussi, Said; Al-Muzaini, Saleh; Fukui, Manabu; Urushigawa, Yoshikuni; Stahl, David A.

    2002-01-01

    A mesophilic toluene-degrading consortium (TDC) and an ethylbenzene-degrading consortium (EDC) were established under sulfate-reducing conditions. These consortia were first characterized by denaturing gradient gel electrophoresis (DGGE) fingerprinting of PCR-amplified 16S rRNA gene fragments, followed by sequencing. The sequences of the major bands (T-1 and E-2) belonging to TDC and EDC, respectively, were affiliated with the family Desulfobacteriaceae. Another major band from EDC (E-1) was related to an uncultured non-sulfate-reducing soil bacterium. Oligonucleotide probes specific for the 16S rRNAs of target organisms corresponding to T-1, E-1, and E-2 were designed, and hybridization conditions were optimized for two analytical formats, membrane and DNA microarray hybridization. Both formats were used to characterize the TDC and EDC, and the results of both were consistent with DGGE analysis. In order to assess the utility of the microarray format for analysis of environmental samples, oil-contaminated sediments from the coast of Kuwait were analyzed. The DNA microarray successfully detected bacterial nucleic acids from these samples, but probes targeting specific groups of sulfate-reducing bacteria did not give positive signals. The results of this study demonstrate the limitations and the potential utility of DNA microarrays for microbial community analysis.

  13. PCR-denaturing gradient gel electrophoresis analysis of microbial community in soy-daddawa, a Nigerian fermented soybean (Glycine max (L.) Merr.) condiment.

    Science.gov (United States)

    Ezeokoli, Obinna T; Gupta, Arvind K; Mienie, Charlotte; Popoola, Temitope O S; Bezuidenhout, Cornelius C

    2016-03-02

    Soy-daddawa, a fermented soybean (Glycine max (L.) Merr.) condiment, plays a significant role in the culinary practice of West Africa. It is essential to understand the microbial community of soy-daddawa for a successful starter culture application. This study investigated the microbial community structure of soy-daddawa samples collected from Nigerian markets, by PCR-denaturing gradient gel electrophoresis (DGGE) targeting the V3-V5 region of the 16S rRNA gene of bacteria and internal transcribed spacer 2 (ITS2) region of fungi. Six bacterial and 16 fungal (nine yeasts and seven molds) operational taxonomic units (OTUs)/species were obtained at 97% sequence similarity. Taxonomic assignments revealed that bacterial OTUs belonged to the phyla Firmicutes and Actinobacteria, and included species from the genera Atopostipes, Bacillus, Brevibacterium and Nosocomiicoccus. Densitometric analysis of DGGE image/bands revealed that Bacillus spp. were the dominant OTU/species in terms of population numbers. Fungal OTUs belonged to the phyla Ascomycota and Zygomycota, and included species from the genera, Alternaria, Aspergillus, Candida, Cladosporium, Dokmaia, Issatchenkia, Kodamaea, Lecythophora, Phoma, Pichia, Rhizopus, Saccharomyces and Starmerella. The majority of fungal species have not been previously reported in soy-daddawa. Potential opportunistic human pathogens such as Atopostipes suicloacalis, Candida rugosa, Candida tropicalis, and Kodamaea ohmeri were detected. Variation in soy-daddawa microbial communities amongst samples and presence of potential opportunistic pathogens emphasises the need for starter culture employment and good handling practices in soy-daddawa processing.

  14. Denaturing gradient gel electrophoresis fingerprinting of soil bacteria in the vicinity of the Chinese Great Wall Station, King George Island, Antarctica.

    Science.gov (United States)

    Pan, Qi; Wang, Feng; Zhang, Yang; Cai, Minghong; He, Jianfeng; Yang, Haizhen

    2013-08-01

    Bacterial diversity was investigated in soil samples collected from 13 sites around the Great Wall Station, Fildes Peninsula, King George Island, Antarctica, using denaturing gradient gel electrophoresis (DGGE) of 16S rRNA genes. The classes alpha-, beta-, and gamma-Proteobacteria, as well as the phylum Actinobacteria, were found to be the dominant bacteria in the soils around the Great Wall Station. Although the selected samples were not contaminated by oil, a relationship between soil parameters, microbial biodiversity, and human impact was still seen. Sample sites in human impacted areas showed lower bacterial biodiversity (average H' = 2.65) when compared to non-impacted sites (average H' = 3.05). There was no statistically significant correlation between soil bacterial diversity and total organic carbon (TOC), total nitrogen, or total phosphorus contents of the soil. Canonical correlation analysis showed that TOC content was the most important factor determining bacterial community profiles among the measured soil parameters. In conclusion, microbial biodiversity and community characteristics within relatively small scales (1.5 km) were determined as a function of local environment parameters and anthropogenic impact.

  15. Denaturing gradient gel electrophoresis fingerprinting of soil bacteria in the vicinity of the Chinese Great Wall Station, King George Island, Antarctica

    Institute of Scientific and Technical Information of China (English)

    Qi Pan; Feng Wang; Yang Zhang; Minghong Cai; Jianfeng He; Haizhen Yang

    2013-01-01

    Bacterial diversity was investigated in soil samples collected from 13 sites around the Great Wall Station,Fildes Peninsula,King George Island,Antarctica,using denaturing gradient gel electrophoresis (DGGE) of 16S rRNA genes.The classes α-,β-,and γ-Proteobacteria,as well as the phylum Actinobacteria,were found to be the dominant bacteria in the soils around the Great Wall Station.Although the selected samples were not contaminated by oil,a relationship between soil parameters,microbial biodiversity,and human impact was still seen.Sample sites in human impacted areas showed lower bacterial biodiversity (average H' =2.65) when compared to non-impacted sites (average H' =3.05).There was no statistically significant correlation between soil bacterial diversity and total organic carbon (TOC),total nitrogen,or total phosphorus contents of the soil.Canonical correlation analysis showed that TOC content was the most important factor determining bacterial community profiles among the measured soil parameters.In conclusion,microbial biodiversity and community characteristics within relatively small scales (1.5 km) were determined as a function of local environment parameters and anthropogenic impact.

  16. Comparison of Fecal Methanogenic Archaeal Community Between Erhualian and Landrace Pigs Using Denaturing Gradient Gel Electrophoresis and Real-Time PCR Analysis

    Institute of Scientific and Technical Information of China (English)

    SU Yong; Hauke Smidt; ZHU Wei-Yun

    2014-01-01

    Erhualian and Landrace breeds are typical genetically obese and lean pigs, respectively. To compare the fecal methanogenic Archaeal community between these two pig breeds, fecal samples from different growth phase pigs were collected and used for PCR-denaturing gradient gel electrophoresis (DGGE) with two primer pairs (344fGC/519r and 519f/915rGC) and real-time PCR analysis. Results showed that a better separation and higher quality of bands pattern were obtained in DGGE proifles using primers 344fGC/519r as compared with primers 519f/915rGC. Sequencing of DGGE bands showed that the predominant methanogens in the feces of Erhualian and Landrace pigs belonged to Methanobrevibacter spp. and Methanosphaera spp. Real-time PCR analysis revealed that there was no signiifcant difference in the numbers of fecal total methanogens between Erhualian and Landrace pigs;however, pig growth phase affected the numbers of 16S rRNA genes of total methanogens and Methanobrevibacter smithii. Dissociation curves of methyl coenzyme-M reductase subunit A (mcrA) gene fragments ampliifed with real-time PCR showed all samples possessed a single peak at 82°C, which might be associated with M. smithii. Samples from the same growth phase of each breed showed good replicative dissociation curves. The results suggest that the growth phase (including diet factor) other than genotype of pig may affect the fecal methanogenic Archaeal community of pigs.

  17. Soil clone library analyses to evaluate specificity and selectivity of PCR primers targeting fungal 18S rDNA for denaturing-gradient gel electrophoresis (DGGE).

    Science.gov (United States)

    Takada Hoshino, Yuko; Morimoto, Sho

    2010-01-01

    We evaluated the fungal specificity and detection bias of four fungal 18S rRNA gene (18S rDNA) primer sets for denaturing-gradient gel electrophoresis (DGGE). We constructed and compared clone libraries amplified from upland and paddy field soils with each primer set (1, NS1/GCFung; 2, FF390/FR1-GC; 3, NS1/FR1-GC; and 4, NS1/EF3 for the first PCR and NS1/FR1-GC for the second PCR). Primer set 4 (for nested PCR) showed the highest specificity for fungi but biased specific sequences. Sets 1, 2, and 3 (for single PCR) amplified non-fungal eukaryotic sequences (from 7 to 16% for upland soil and from 20 to 31% for paddy field soil) and produced libraries with similar distributions of fungal 18S rDNA sequences at both the phylum and the class level. Set 2 tended to amplify more diverse fungal sequences, maintaining higher specificity for fungi. In addition, clone analyses revealed differences among primer sets in the frequency of chimeras. In upland field soil, the libraries amplified with primer sets 3 and 4, which targeted long fragments, contained many chimeric 18S rDNA sequences (18% and 48%, respectively), while the libraries obtained with sets 1 and 2, which targeted short fragments, contained fewer chimeras (5% and 10%, respectively).

  18. Approach to Analyze the Diversity of Myxobacteria in Soil by Semi-Nested PCR-Denaturing Gradient Gel Electrophoresis (DGGE) Based on Taxon-Specific Gene

    Science.gov (United States)

    Li, Baiyuan; Yao, Qing; Zhu, Honghui

    2014-01-01

    The genotypic diversity of insoluble macromolecules degraded myxobacteria, provided an opportunity to discover new bacterial resources and find new ecological functions. In this study, we developed a semi-nested-PCR-denaturing gradient gel electrophoresis (DGGE) strategy to determine the presence and genotypic diversity of myxobacteria in soil. After two rounds of PCR with myxobacteria-specific primers, an 194 bp fragment of mglA, a key gene involved in gliding motility, suitable for DGGE was obtained. A large number of bands were observed in DGGE patterns, indicating diverse myxobacteria inhabiting in soils. Furthermore, sequencing and BLAST revealed that most of the bands belonged to the myxobacteria-group, and only three of the twenty-eight bands belonged to other group, i.e., Deinococcus maricopensis. The results verified that myxobacterial strains with discrepant sequence compositions of gene mglA could be discriminated by DGGE with myxobacteria-specific primers. Collectively, the developed semi-nested-PCR-DGGE strategy is a useful tool for studying the diversity of myxobacteria. PMID:25280065

  19. Comparative microbiota assessment of wilted Italian ryegrass, whole crop corn, and wilted alfalfa silage using denaturing gradient gel electrophoresis and next-generation sequencing.

    Science.gov (United States)

    Ni, Kuikui; Minh, Tang Thuy; Tu, Tran Thi Minh; Tsuruta, Takeshi; Pang, Huili; Nishino, Naoki

    2017-02-01

    The microbiota of pre-ensiled crop and silage were examined using denaturing gradient gel electrophoresis (DGGE) and next-generation sequencing (NGS). Wilted Italian ryegrass (IR), whole crop corn (WC), and wilted alfalfa (AL) silages stored for 2 months were examined. All silages contained lactic acid as a predominant fermentation product. Across the three crop species, DGGE detected 36 and 28 bands, and NGS identified 253 and 259 genera in the pre-ensiled crops and silages, respectively. The NGS demonstrated that, although lactic acid bacteria (LAB) became prevalent in all silages after 2 months of storage, the major groups were different between crops: Leuconostoc spp. and Pediococcus spp. for IR silage, Lactobacillus spp. for WC silage, and Enterococcus spp. for AL silage. The predominant silage LAB genera were also detected by DGGE, but the presence of diverse non-LAB species in pre-ensiled crops was far better detected by NGS. Likewise, good survival of Agrobacterium spp., Methylobacterium spp., and Sphingomonas spp. in IR and AL silages was demonstrated by NGS. The diversity of the microbiota described by principal coordinate analysis was similar between DGGE and NGS. Our finding that analysis of pre-ensiled crop microbiota did not help predict silage microbiota was true for both DGGE and NGS.

  20. Study of microbial community structures in UASB sludge treating municipal wastewater by denaturing gradient gel electrophoresis of 16S rDNA

    Institute of Scientific and Technical Information of China (English)

    ZHANG Ying; Sunny Aiyuk; XU Hui; CHEN Guanxiong; Willy Verstraete

    2005-01-01

    The structures of microbial communities in lab-scale upflow anaerobic sludge blanket (UASB) reactors for treating municipal wastewater with different ratios of COD soluble/COD total were studied using denaturing gradient gel electrophoresis (DGGE) of 16S rRNA genes.The microbial structure of the inoculum sludge obtained from a full-scale UASB reactor of treating potato processing wastewater was compared with the structures of sludges collected from three lab-scale UASB reactors after eight months feeding with raw municipal wastewater, with CEPS (chemically enhanced primary sedimentation) pretreated municipal wastewater, and with a synthetic municipal sewage, respectively. Computer-aided numerical analysis of the DGGE fingerprints showed that the bacterial community underwent major changes. The sludges for treating raw and CEPS pretreated wastewater had very similar bacterial and archaeal communities (82%and 96% similarity) but were different from that for treating the synthetic sewage. Hence, despite similar % COD in the particulate form in the synthetic and the real wastewater, the two wastewaters were selected for different microbial communities. Prominent DGGE bands of Bacteria and Archaea were purified and sequenced. The 16S rRNA gene sequences of the dominant archaeal bands found in the inoculum, and UASB sludge fed with raw sewage, CEPS pretreated wastewater, and synthetic sewage were closely associated with Methanosaeta concilii. In the UASB sludge fed with synthetic sewage, another dominant band associated with an uncultured archaeon 39-2 was found together with M. concilii.

  1. Association of oral streptococci community dynamics with severe early childhood caries as assessed by PCR-denaturing gradient gel electrophoresis targeting the rnpB gene.

    Science.gov (United States)

    Tao, Ye; Zhou, Yan; Ouyang, Yong; Lin, Huan Cai

    2015-08-01

    This study sought to investigate the possible association between the dynamics of oral streptococci community profiles and severe early childhood caries (S-ECC) development, compared with caries-free (CF) controls. Supragingival plaque samples were evaluated from 8-32-month-old children who had previously been assessed for overall profiles of their oral microbial community. Twelve children were in each group. Bacterial genomic DNA was extracted and amplified using rnpB-specific primers for streptococci; the products were then subjected to denaturing gradient gel electrophoresis (DGGE) and sequence analysis. We observed that the mean values for species richness (N) and diversity of oral streptococci (H') were significantly lower in the S-ECC group than in the CF group (N = 1.25 ± 4.14 vs 14.92 ± 2.84; H' = 1.41 ± 0.29 vs 1.64 ± 0.18) at 32  months of age (P diversity of oral streptococci, that the detection rates of S. sanguinis and S. gordonii have negative correlations with S-ECC; and that there are high levels of intra-individual similarity for the oral streptococci community over time.

  2. Characterization of depth-related microbial communities in lake sediment by denaturing gradient gel electrophoresis of amplified 16S rRNA fragments

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    The characterization of microbial communities of different depth sediment samples was examined by a culture-independent method and compared with physicochemical parameters, those are organic matter (OM), total nitrogen (TN), total phosphorus (TP), pH and redox potential (Eh). Total genomic DNA was extracted from samples derived from different depths. After they were amplified with the GC-341f/907r primer sets of partial bacterial 16S rRNA genes, the products were separated by denaturing gradient gel electrophoresis (DGGE). The profile of DGGE fingerprints of different depth sediment samples revealed that the community structure remained relatively stable along the entire 45 cm sediment core, however, principal-component analysis of DGGE patterns revealed that at greater sediment depths, successional shifts in community structure were evident. The principle coordinates analysis suggested that the bacterial communities along the sediment core could be separated into two groups, which were located 0-20 cm and 21-45 cm, respectively. The sequencing dominant bands demonstrated that the major phylogenetic groups identified by DGGE belonged to Bacillus, Bacterium, Brevibacillus, Exiguobacterium, γ-Proteobacterium, Acinetobacter sp. And some uncultured or unidentified bacteria. The results indicated the existence of highly diverse bacterial community in the lake sediment core.

  3. Molecular Fingerprinting by PCR-Denaturing Gradient Gel Electrophoresis Reveals Differences in the Levels of Microbial Diversity for Musty-Earthy Tainted Corks ▿

    Science.gov (United States)

    Prat, Chantal; Ruiz-Rueda, Olaya; Trias, Rosalia; Anticó, Enriqueta; Capone, Dimitra; Sefton, Mark; Bañeras, Lluís

    2009-01-01

    The microbial community structure of cork with marked musty-earthy aromas was analyzed using denaturing gradient gel electrophoresis of amplified ribosomal DNA. Cork stoppers and discs were used for DNA extraction and were analyzed by using selective primers for bacteria and fungi. Stoppers clearly differed from discs harboring a different fungal community. Moreover, musty-earthy samples of both types were shown to have a specific microbiota. The fungi Penicillium glabrum and Neurospora spp. were present in all samples and were assumed to make only a small contribution to off-odor development. In contrast, Penicillium islandicum and Penicillium variabile were found almost exclusively in 2,4,6-trichloroanisole (TCA) tainted discs. Conversely, Rhodotorula minuta and Rhodotorula sloofiae were most common in cork stoppers, where only small amounts of TCA were detected. Alpha- and gammaproteobacteria were the most commonly found bacteria in either control or tainted cork stoppers. Specific Pseudomonas and Actinobacteria were detected in stoppers with low amounts of TCA and 2-methoxy-3,5-dimethylpyrazine. These results are discussed in terms of biological degradation of taint compounds by specific microorganisms. Reliable and straightforward microbial identification methods based on a molecular approach provided useful data to determine and evaluate the risk of taint formation in cork. PMID:19201983

  4. Molecular fingerprinting by PCR-denaturing gradient gel electrophoresis reveals differences in the levels of microbial diversity for musty-earthy tainted corks.

    Science.gov (United States)

    Prat, Chantal; Ruiz-Rueda, Olaya; Trias, Rosalia; Anticó, Enriqueta; Capone, Dimitra; Sefton, Mark; Bañeras, Lluís

    2009-04-01

    The microbial community structure of cork with marked musty-earthy aromas was analyzed using denaturing gradient gel electrophoresis of amplified ribosomal DNA. Cork stoppers and discs were used for DNA extraction and were analyzed by using selective primers for bacteria and fungi. Stoppers clearly differed from discs harboring a different fungal community. Moreover, musty-earthy samples of both types were shown to have a specific microbiota. The fungi Penicillium glabrum and Neurospora spp. were present in all samples and were assumed to make only a small contribution to off-odor development. In contrast, Penicillium islandicum and Penicillium variabile were found almost exclusively in 2,4,6-trichloroanisole (TCA) tainted discs. Conversely, Rhodotorula minuta and Rhodotorula sloofiae were most common in cork stoppers, where only small amounts of TCA were detected. Alpha- and gammaproteobacteria were the most commonly found bacteria in either control or tainted cork stoppers. Specific Pseudomonas and Actinobacteria were detected in stoppers with low amounts of TCA and 2-methoxy-3,5-dimethylpyrazine. These results are discussed in terms of biological degradation of taint compounds by specific microorganisms. Reliable and straightforward microbial identification methods based on a molecular approach provided useful data to determine and evaluate the risk of taint formation in cork.

  5. Diagnosis of cutaneous T-cell lymphoma detecting T-cell receptor gamma chain gene monoclonality by denaturing gradient gel electrophoresis.

    Science.gov (United States)

    Lapière, K; Dhaene, K; Matthieu, L; Hübner, R; Lambert, J; Van Marck, E

    1999-04-01

    Cutaneous T-cell lymphomas represent a group of malignant lymphoproliferative disorders characterised by the occurrence of a monoclonal population of T-lymphocytes. Diagnosis of early stages of this disease is a difficult challenge for both the dermatologist and the dermatopathologist. With the aid of the polymerase chain reaction it is possible to amplify specific regions of the T-cell receptor gamma gene. The amplification products can then be separated by denaturing gradient gel electrophoresis in order to detect a monoclonal population of T-lymphocytes in the infiltrate. We studied 4 patients with the clinicopathologic diagnosis of mycosis fungoides and 2 patients diagnosed as large plaque parapsoriasis. A monoclonal population was detected in 3 of the 4 mycosis fungoides cases and in 1 of the patients with large plaque parapsoriasis. This indicates that our analysis can help us establishing a diagnosis, and it can also help us to identify patients with a possible early stage of the disease, which clinically or histologically is not yet recognised as such.

  6. Co-monitoring bacterial and dinoflagellates communities by denaturing gradient gel electrophoresis (DGGE) and SSU rRNA sequencing during a dinoflagellates bloom

    Institute of Scientific and Technical Information of China (English)

    KANJinjun; CHENFeng

    2004-01-01

    Dinoflagellates are unicellular eukaryotic protists that dominate in all coastal waters, and are also present in oceanic waters. Despite the central importance of dinoflagellates in global primary production, the relationship between dinoflagellates and bacteria are still poorly understood. In order to understand the ecological interaction between bacterial and dinoflageUates communities, denaturing gradient gel electrophoresis (DGGE) and SSU rRNA sequencing were applied to monitoring the population dynamics of bacteria and dinoflagellates from the onset to disappearance of a dinoflagellates bloom occurred in Baltimore Inner Harbor, from April 15 to 24, 2002. Although Prorocentrum minimum was the major bloom forming species under the light microscopy, DGGE method with dinoflagellate specific primers demonstrated that Prorocentrum micans, Gymnodinium galatheanum and Gyrodinium uncatenum were also present during the bloom. Population shifts among the minor dinoflagellate groups were observed. DGGE of PCR-amplified 16S rRNA gene fragments indicated that cyanobacteria, α, β, γ-proteobacteria, FlavobacteriumBacteroides-Cytophaga (FBC), and Planctomcetes were the major components of bacterial assemblages during the bloom. DGGE analysis showed that Cytophagales and α-proteobacteria played important roles at different stages of dinoflagellates bloom. DGGE can be used as a rapid tool to simultaneously monitor population dynamics of both bacterial and dinoflagellates communities in aquatic environments, which is demonstrated here.

  7. International Conference on Harmonisation; Guidance on Q4B Evaluation and Recommendation of Pharmacopoeial Texts for Use in the International Conference on Harmonisation Regions; Annex 10 on Polyacrylamide Gel Electrophoresis General Chapter; availability. Notice.

    Science.gov (United States)

    2010-04-12

    The Food and Drug Administration (FDA) is announcing the availability of a guidance entitled "Q4B Evaluation and Recommendation of Pharmacopoeial Texts for Use in the ICH Regions; Annex 10: Polyacrylamide Gel Electrophoresis General Chapter. The guidance was prepared under the auspices of the International Conference on Harmonisation of Technical Requirements for Registration of Pharmaceuticals for Human Use (ICH). The guidance provides the results of the ICH Q4B evaluation of the Polyacrylamide Gel Electrophoresis General Chapter harmonized text from each of the three pharmacopoeias (United States, European, and Japanese) represented by the Pharmacopoeial Discussion Group (PDG). The guidance conveys recognition of the three pharmacopoeial methods by the three ICH regulatory regions and provides specific information regarding the recognition. The guidance is intended to recognize the interchangeability between the local regional pharmacopoeias, thus avoiding redundant testing in favor of a common testing strategy in each regulatory region. In the Federal Register of February 21, 2008 (73 FR 9575), FDA made available a guidance on the Q4B process entitled "Q4B Evaluation and Recommendation of Pharmacopoeial Texts for Use in the ICH Regions."

  8. [Denaturalized psychoanalysts].

    Science.gov (United States)

    Peglau, Andreas

    2011-01-01

    This paper presents hitherto unknown material from the German Foreign Office referring to the denaturalization of Therese Benedek, Bruno Bettelheim, Adolf Storfer and Wilhelm Reich by Nazi Germany. It corroborates the finding that nobody was persecuted by the Nazis solely on the basis of psychoanalytic activities or membership in a psychoanalytic organization.

  9. Denaturation and Oxidative Stability of Hemp Seed (Cannabis sativa L.) Protein Isolate as Affected by Heat Treatment.

    Science.gov (United States)

    Raikos, Vassilios; Duthie, Garry; Ranawana, Viren

    2015-09-01

    The present study investigated the impact of heat treatments on the denaturation and oxidative stability of hemp seed protein during simulated gastrointestinal digestion (GID). Heat-denatured hemp protein isolate (HPI) solutions were prepared by heating HPI (2 mg/ml, pH 6.8) to 40, 60, 80 and 100 °C for 10 min. Heat-induced denaturation of the protein isolates was monitored by polyacrylamide gel electrophoresis. Heating HPI at temperatures above 80 °C significantly reduced solubility and led to the formation of large protein aggregates. The isolates were then subjected to in vitro GID and the oxidative stability of the generated peptides was investigated. Heating did not significantly affect the formation of oxidation products during GID. The results suggest that heat treatments should ideally remain below 80 °C if heat stability and solubility of HPI are to be preserved.

  10. Application of PCR-denaturing-gradient gel electrophoresis (DGGE) method to examine microbial community structure in asparagus fields with growth inhibition due to continuous cropping.

    Science.gov (United States)

    Urashima, Yasufumi; Sonoda, Takahiro; Fujita, Yuko; Uragami, Atsuko

    2012-01-01

    Growth inhibition due to continuous cropping of asparagus is a major problem; the yield of asparagus in replanted fields is low compared to that in new fields, and missing plants occur among young seedlings. Although soil-borne disease and allelochemicals are considered to be involved in this effect, this is still controversial. We aimed to develop a technique for the biological field diagnosis of growth inhibition due to continuous cropping. Therefore, in this study, fungal community structure and Fusarium community structure in continuously cropped fields of asparagus were analyzed by polymerase chain reaction/denaturing-gradient gel electrophoresis (PCR-DGGE). Soil samples were collected from the Aizu region of Fukushima Prefecture, Japan. Soil samples were taken from both continuously cropped fields of asparagus with growth inhibition and healthy neighboring fields of asparagus. The soil samples were collected from the fields of 5 sets in 2008 and 4 sets in 2009. We were able to distinguish between pathogenic and non-pathogenic Fusarium by using Alfie1 and Alfie2GC as the second PCR primers and PCR-DGGE. Fungal community structure was not greatly involved in the growth inhibition of asparagus due to continuous cropping. By contrast, the band ratios of Fusarium oxysporum f. sp. asparagi in growth-inhibited fields were higher than those in neighboring healthy fields. In addition, there was a positive correlation between the band ratios of Fusarium oxysporum f. sp. asparagi and the ratios of missing asparagus plants. We showed the potential of biological field diagnosis of growth inhibition due to continuous cropping of asparagus using PCR-DGGE.

  11. Diversity of lactic acid bacteria from modified atmosphere packaged sliced cooked meat products at sell-by date assessed by PCR-denaturing gradient gel electrophoresis.

    Science.gov (United States)

    Audenaert, Kris; D'Haene, Klaas; Messens, Kathy; Ruyssen, Tony; Vandamme, Peter; Huys, Geert

    2010-02-01

    The predominant lactic acid bacteria (LAB) microbiota associated with three types of modified atmosphere packaged (MAP) sliced cooked meat products (i.e. ham, turkey and chicken) was analyzed at sell-by date using a combination of culturing and molecular population fingerprinting. Likewise routine analyses during industrial MAP production, meat samples were plated on the general heterotrophic Plate Count Agar (PCA) and on the LAB-specific de Man, Rogosa, Sharpe (MRS) agar under different temperature and atmosphere conditions. Subsequently, community DNA extracts were prepared from culturable bacterial fractions harvested from both media and used for PCR targeting the V3 hyper-variable region of the 16S rRNA gene followed by denaturing gradient gel electrophoresis (DGGE) of PCR amplicons (PCR-DGGE). Irrespective of aerobic or anaerobic incubation conditions, V3-16S rDNA DGGE fingerprints of culturable fractions from PCA and MRS medium displayed a high level of similarity indicating that LAB constituted the most dominant group in the culturable bacterial community. Comparison of DGGE profiles of fractions grown at 20, 28 or 37 degrees C indicated that part of the culturable community consisted of psychrotrophs. Four DGGE bands were common among cooked ham, turkey and chicken products, suggesting that these represent the microbiota circulating in the plant where all three MAP product types were sliced and packaged. Based on band sequencing and band position analysis using LAB reference strains, these four bands could be assigned to Lactobacillus sakei and/or the closely related Lactobacillus fuchuensis, Lactobacillus curvatus, Carnobacterium divergens and Leuconostoc carnosum. In conclusion, the PCR-DGGE approach described in this study allows to discriminate, identify and monitor core and occasional LAB microbiota of MAP sliced cooked meat products and provides valuable complementary information to the current plating procedures routinely used in industrial plants.

  12. Detection of Helicobacter species in liver and stomach tissues of patients with chronic liver diseases using polymerase chain reaction-denaturing gradient gel electrophoresis and immunohistochemistry.

    Science.gov (United States)

    Stalke, Piotr; Al-Soud, Waleed Abu; Bielawski, Krzysztof P; Bakowska, Alicja; Trocha, Hanna; Stepinski, Jan; Wadström, Torkel

    2005-09-01

    Helicobacter DNA has been detected in the hepatobiliary tree of patients with chronic liver diseases (CLD). The presence of H. pylori in the stomach compared with in the liver of the same patients with CLD has not been studied, therefore to the aim of this study was to investigate the presence of Helicobacter DNA and antigens in the liver and stomach of Polish patients with chronic liver diseases using molecular and immunological methods. Gastric mucosa and liver tissue samples and sera were collected from 97 Polish patients with CLD. Anti-H. pylori antibodies were detected by enzyme immunoassay (EIA), and H. pylori-like antigens detected by immunohistochemistry. Helicobacter DNA was detected in stomach and liver samples using a semi-nested Helicobacter genus-specific polymerase chain reaction (PCR) assay, and Helicobacter species identified by denaturing gradient gel electrophoresis (DGGE) and sequencing analysis of amplified PCR products. H. pylori was identified by DGGE and sequence analysis in 60/62 (97%) and 25/25 (100%) of the gastric and liver Helicobacter genus-positive samples, respectively, whereas DNA of H. heilmannii was detected in 2/62 (3%) of the Helicobacter genus-positive gastric samples. H. pylori cagA gene was detected in 23/62 (36%) and 3/25 (12%) gastric and liver tissue samples, respectively. H. pylori-like antigens were detected in 61/97 (63%) gastric mucosa and in 40/97 (41%) liver tissue samples. H. pylori-like organisms appeared to dominate the gastric mucosa and liver tissue of Polish patients with CLD. The prevalence of the cagA gene was higher in stomach compared with liver samples, which suggests a possible role of cagA negative H. pylori-like organisms in CLD. On the other hand, no significant correlation was found between the presence of H. pylori-like DNA and antigens in the liver and liver function tests.

  13. Stability of capillary gels for automated sequencing of DNA.

    Science.gov (United States)

    Swerdlow, H; Dew-Jager, K E; Brady, K; Grey, R; Dovichi, N J; Gesteland, R

    1992-08-01

    Recent interest in capillary gel electrophoresis has been fueled by the Human Genome Project and other large-scale sequencing projects. Advances in gel polymerization techniques and detector design have enabled sequencing of DNA directly in capillaries. Efforts to exploit this technology have been hampered by problems with the reproducibility and stability of gels. Gel instability manifests itself during electrophoresis as a decrease in the current passing through the capillary under a constant voltage. Upon subsequent microscopic examination, bubbles are often visible at or near the injection (cathodic) end of the capillary gel. Gels have been prepared with the polyacrylamide matrix covalently attached to the silica walls of the capillary. These gels, although more stable, still suffer from problems with bubbles. The use of actual DNA sequencing samples also adversely affects gel stability. We examined the mechanisms underlying these disruptive processes by employing polyacrylamide gel-filled capillaries in which the gel was not attached to the capillary wall. Three sources of gel instability were identified. Bubbles occurring in the absence of sample introduction were attributed to electroosmotic force; replacing the denaturant urea with formamide was shown to reduce the frequency of these bubbles. The slow, steady decline in current through capillary sequencing gels interferes with the ability to detect other gel problems. This phenomenon was shown to be a result of ionic depletion at the gel-liquid interface. The decline was ameliorated by adding denaturant and acrylamide monomers to the buffer reservoirs. Sample-induced problems were shown to be due to the presence of template DNA; elimination of the template allowed sample loading to occur without complications.(ABSTRACT TRUNCATED AT 250 WORDS)

  14. Protein and glycoprotein abnormalities in platelets from human Chediak-Higashi syndrome: polyacrylamide gel electrophoretic study of platelets from five patients.

    Science.gov (United States)

    Ledezma, E; Apitz-Castro, R

    1985-10-01

    Polyacrylamide electrophoretic analysis of proteins and Tritium-labelled glycoproteins of the platelets from five patients with Chediak-Higashi Syndrome shows the existence of marked quantitative differences when compared to normal platelets. While the glycoprotein abnormalities are solely related to the plasma membrane, some of the abnormalities detected in the Coomasie blue pattern are probably representative of defects related to the dense bodies and the alpha-granules. Some of the abnormalities found may, in part, explain the variability of aggregatory responses described in these patients, as well as the marked tendency towards desaggregation exhibited by platelets from humans with the Chediak-Higashi Syndrome.

  15. Separation of metalloproteins using a novel metal ion contaminant sweeping technique and detection of protein-bound copper by a metal ion probe in polyacrylamide gel electrophoresis: distribution of copper in human serum.

    Science.gov (United States)

    Saito, Shingo; Kawashima, Mitsuyoshi; Ohshima, Hiroki; Enomoto, Kazuki; Sato, Makoto; Yoshimura, Hajime; Yoshimoto, Keitaro; Maeda, Mizuo; Shibukawa, Masami

    2013-10-21

    A polyacrylamide gel electrophoresis (PAGE)-based method has been developed, consisting of two types of gel electrophoresis, to obtain an accurate distribution of protein-bound metal ions in biological samples. First, proteins are separated by PAGE without the uptake of contaminant metal ions in the separation field and dissociation of metal ions from the proteins. This is followed by another PAGE for the separation and detection of protein-bound metal ions in small volume samples with high sensitivity in the ppt range using a fluorescent metal probe. The former is a new technique using blue-native (BN) PAGE to electrophoretically sweep all metal contaminants by employing two kinds of chelating agents. These agents form complexes with contaminants in the gel and the separation buffer solution, which migrate towards opposite pole directions, thus lowering the contaminants to below the ppt level during separation. This is termed "Metal Ion Contaminant Sweeping BN-PAGE (MICS-BN-PAGE)". After the separation of proteins under these first metal-free conditions, the metal ions in the gel fractions are eluted, followed by derivatization of copper ions into the metal probe complexes to be separated and determined by fluorescence detection in the second PAGE. In this PAGE-based method, the copper ions bound to ceruloplasmin and superoxide dismutase were quantitatively determined, in addition to the exchangeable albumin-bound copper ions. This system successfully provided distribution maps of protein-copper in human serum. The precise distribution of copper in human serum was investigated, and found to be different from that which is widely accepted.

  16. Study on preparation and performance of removing fluorine of grafted and modified polyacrylamide-gel%接枝改性聚丙烯酰胺凝胶的制备与除氟性能研究

    Institute of Scientific and Technical Information of China (English)

    蔡莉; 刘建; 胡希华

    2012-01-01

    A new material, with ligand exchange properties, of removing fluoride from the fluorine-containing water was obtained. This material was prepared, according to Mannich reaction, from polyacrylamide gel ( PAM-Gel) , formaldehyde and iminodiacetic acid (IDA). With that, it was modified by chelating Fe3+ ions in (NH4) Fe(SO4)2 solution. Preparation conditions and performance of using this modified gel to remove fluoride from high fluoride water was studied. The results showed that the synthetic process of the gel was easy to operate,and its capability of removing fluorine was obvious and rapid. In the fluorine-containing water of 100 mg/L F- concentration, the saturated adsorption capacity was up to 31. 70 mgF-/g-gel. As 5 mg/L F- concentration, through in twice adsorption, total removal rate of fluoride was up to 83. 43% and the rest concentration of F- can meet with China national standards for drinking water. The test results showed as well, common-ions in water did not make affect on adsorbing fluor ions by the gel,and high selection in process of removing flours has been demonstrated.%以聚丙烯酰胺凝胶(PAM-Gel)为骨架,以氨基二乙酸(IDA)为接枝螯合剂,在甲醛参与下,依据Mannich反应,制得螯合基团接枝凝胶,并通过在( NH4) Fe( SO4)2溶液中螯合Fe3+离子,使凝胶获得改性,得到一种新的具有配体交换性能的从含氟水中除氟材料.研究了该种改性凝胶除氟材料制备条件及在高氟水中除氟的效率,结果表明,该除氟剂具有良好的除氟性能,在含氟100 mg/L溶液中,饱和吸附量可达31.70 mgF -/g-gel;对含氟5 mg/L的含氟水,经二次除氟,总除氟率可达83.43%,水中剩余氟离子浓度可达国家饮用水标准.实验表明,水中常见共存离子对氟的吸附率无影响,表现出该种材料对氟吸附的高度选择性.

  17. Effect of detergents and chemicals on purified vaccinia virus: analysis by SDS polyacrylamide gel electrophoresis and electron microscopy1,2.

    Science.gov (United States)

    Boisvert, J; Yamamoto, T

    1977-03-01

    Vaccinia virus particles were dissociated into their constituent polypeptides and analysed by sodium dodecyl sulfate (SDS) gel electrophoresis. Thirty-three distinct polypeptide bands were identified and their molecular weights ranged between 11 000 and 150 000 daltons. Specific staining of gels containing polypeptides of dissociated virions revealed the presence of eight glycopeptides. No lipopeptides were detected. Analysis of chemical extracts (urea, guanidine hydrochloride, and alkali treatment) of the virus by SDS gel electrophoresis indicated that a total of 10 to 14 different polypeptides ranging in molecular weights from 11 000 to 70 000 daltons were solubilized. Analysis of detergent extracts and of the remains of extracted viral particles has shown that the detergent Nonidet P-40 (NP-40) solubilized a total of 11 polypeptides of which 6 were glycopeptides. The other detergents sodium deoxycholate (SDC) and cetyl trimethyl ammonium bromide (CTAB) were not as selective. both solubilizing more than 25 of the polypeptides composing the virus. Gel electrophoresis results also indicated that most of the small molecular weight (11 000-70 000 daltons) polypeptides were readily solubilized by NP-40, SDC, and CTAB, while those with molecular weights of 70 000 daltons and higher were not well solubilized. The effects of detergents were also analysed by electron microscopy. Evidence was obtained for subpopulations of viral particles having different susceptibility to detergent extraction.

  18. Research on Nonionic Polyacrylamide Zirconium Gel Fracturing Fluids in Coalbed Methane Gas Wells%煤层气井用非离子聚丙烯酰胺锆冻胶压裂液优选

    Institute of Scientific and Technical Information of China (English)

    赵辉; 戴彩丽; 梁利; 王欣; 赵福麟

    2012-01-01

    Active water and guar gum are widely used for coalbed methane fracturing in China. Nevertheless, the application of these two types of fracturing fluid was limited due to poor rheological properties of active water and severe formation damage of guar gum fracturing fluid. Considering the temperature and low permeability of coalbed, the formulation of nonionic polyacrylamide zirconium gel fracturing fluids suitable for coalbed mechane fracturing was selected based on the analysis on the effects of different factors on the zirconium gel. The formulation is 0. 4%PAM+0. 035%ZrOCl2. The performance of the optimized formulation of nonionic polyacrylamide zirconium gel fracturing fluids was evaluated through laboratory experiments, and the results showed that,the formulation had the characteristics of shearing resistance, low filter loss, easy breaking, good proppant carrying capacity, non-residue, low formation damage and easy flowback. It is suitable for the coalbed gas reservoirs fracturing.%活性水、瓜胶压裂液是国内煤层气井压裂最常用的压裂液,但活性水压裂液流变性能差,瓜胶压裂液破胶残渣含量高对煤层的伤害大,限制了这两类压裂液在煤层压裂中的应用.为此,针对煤层温度和渗透率低的特点,在分析影响锆冻胶压裂液性能因素的基础上,优选出了适用于煤层气井压裂的非离子型聚丙烯酰胺锆冻胶压裂液配方(0.400%PAM+0.035%ZrOCl2).通过室内试验对优选出的非离子型聚丙烯酰胺锆冻胶压裂液的性能进行了评价,结果表明,该压裂液具有耐剪切、滤失量低、易破胶、携砂性能好、无残渣、对煤层伤害低、易返排的特点,适用于低温煤层压裂.

  19. 双连续相微乳液制备连续孔聚丙烯酰胺凝胶%Preparation and properties of continuous porous polyacrylamide gels from bicontinuous microemulsions

    Institute of Scientific and Technical Information of China (English)

    周应学; 范晓东; 孙乐; 陈卫星; 于德海

    2011-01-01

    Organogel, hydrogel and an organo/hydro hybrid gel were prepared by bicontinuous microemulsion (BME) template of sodium dodecyl sulfate (SDS)/toluene/ 2-butanol/saline. 12-Hydroxystearic acid (12-HA)was organogeltor. These gels were self-supported, and aqueous and oil microphases bicontinuously coexisted and were immobilized in the gels. Particles dimenters and polydispersion of BME solution were detected by static light scattering (SLS) and dimenter was 8-800nm. Morphology constructure of gels were observated by scanning electrinic microscopy (SEM) and was continuous porous networks. Several peaks were measured of gels tested by differential scanning calorimetry (DSC) and thermogravity (TG) for thermal properties due to bicontinuous microemulsion composites. Compared to pure cross-linked polyacrylamide, the swelling ratio and swelling saturated times of BME hydrogel were instrictly decreased.%以十二烷基硫酸钠/甲苯/2-丁醇/盐水双连续微乳液相为模板,制备了3种组分双连续有机凝胶,水凝胶和有机/水杂化凝胶.12-羟基硬脂酸(12-HA)为有机凝胶剂.它们固定了水和油并使水和油微相共存于凝胶中.以静态光散射(SLS)测定了各双连续相微乳液的粒径,结果显示值在8~80nm.扫描电子显微镜观察了各凝胶的形貌,表明凝胶为连续孔网络结构.差示扫描量热(DSC)和热重(TG)研究了凝胶的热性能,由于双连续结构3种凝胶显示多个吸热和失重峰.与纯的闭孔聚丙烯酰胺相比,双连续水凝胶的溶胀率明显降低,饱和溶胀所需时间大大减小.

  20. Viscoelasticity of mixed polyacrylamide solution

    Institute of Scientific and Technical Information of China (English)

    徐丽娜

    2008-01-01

    The viscoelastic behavior of polyacrylamide solution is crucial for its application in various industries.The mixed polyacrylamide solution was prepared by mixing polyacrylamide with different relative molecular masses according to the defined mass fraction.The viscosity and elasticity of mixed polyacrylamide solution were separately tested with RS150 rheometer and capillary breakup extensional rheometer and compared with those of the single polyacrylamide solution which is directly provided by manufacturer without any mixing.The results indicate that the mixed and single polyacrylamide solutions have the same shear viscosity and intrinsic viscosity.However,some mixed polyacrylamide solutions have higher elasticity than single polyacrylamide solution.The flow resistance of mixed polyacrylamide with higher elasticity is also greater than that of single polyacrylamide solution in porous medium.This paper presents an effective method of mixing polyacrylamides with different relative molecular masses,which can enhance the elasticity of polyacrylamide solution and flowing resistance through porous medium.

  1. Formation reason of gel-like impurities of polyacrylamide%聚丙烯酰胺胶状杂质的形成因为

    Institute of Scientific and Technical Information of China (English)

    关淑霞; 邰永娜; 于海南; 宋春红

    2011-01-01

    With a view to the emerge of a great deal of black and yellow strip substances with pungent putrid odor in the filter-bag of percolator at the site where polyacrylamide (PAM) was prepared as well as severe blockage of the percolator and corrosion of equipments caused by the impurities, analyses were made in relation to bacteria and inorganic ions so as to search the reasons leading to the impurities. It was found that, due to the increase of amount of bacteria, the content of Fe3+ increased, and the side amide chain of PAM was degraded to form carboxyl. This, in association with tangling and cross-linking of PAM solution containing increased content of metal ions, resulted in the strip substances. At the same time, PAM was biodegraded to generate NH3, and sulfate reducing bacteria (SRB) reduced sulfate to form H2S by making use of H2 produced inside cell membrane, resulting in putrid odor.%在聚丙烯酰胺(PAM)配制现场发现过滤器滤袋内存在大量黑色和黄色且有刺鼻腐臭气味的胶状条形物;这类杂质造成过滤装置严重堵塞和设备腐蚀.鉴于此,从细菌和无机离子含量两方面着手分析了聚丙烯酰胺胶状条形杂质的形成因为.结果表明:由于细菌和金属离子Fe3+含量的增加,PAM的侧酰胺基降解成羧基;而含大量金属离子的PAM溶液经缠结交联形成条形物.与此同时,PAM经生物降解产生NH3,硫酸盐还原菌(SRB)利用细胞膜内产生的氢将硫酸盐还原为H2S,从而产生腐臭气味.

  2. Characterisation of ribosomal proteins from HeLa and Krebs II mouse ascites tumor cells by different two-dimensional polyacrylamide gel electrophoresis techniques

    DEFF Research Database (Denmark)

    Issinger, O G; Beier, H

    1978-01-01

    electrophoresis; 2. two-dimensional gel electrophoresis at pH 4.K/pH 8.6 in SDS. The molecular weights for 40S proteins ranged from 10,000 to 39,000 dalton (number average molecular weight: 21,000). The molecular weights for the 60S proteins ranged from 14,000 to 44,000 dalton (number average molecular weight: 23......Electrophoresis of ribosomal proteins according to Kaltschmidt and Wittmann, 1970a, b (pH 8.6/pH 4.5 urea system) yielded 29 proteins for the small subunits and 35 and 37 proteins for the large subunits of Krebs II ascites and HeLa ribosomes, respectively. Analysis of the proteins according...... to a modified technique by Mets and Bogorad (1974) (pH 4.5/pH 8.6 SDS system) revealed 28 and 29 proteins in the small subunits and 37 and 38 proteins in the large subunits of Krebs II ascites and HeLa ribosomes. The molecular weights of the individual proteins were determined by: 1. "three-dimensional" gel...

  3. Analysis of the MRI Features of the Complications after Augmentation Mammoplasty with Polyacrylamide Hydrophilic Gel Injection%PAHG注射隆乳术后并发症的MRI表现分析

    Institute of Scientific and Technical Information of China (English)

    梁雯雯; 张雪林; 张海捷

    2011-01-01

    目的 探讨聚丙烯酰胺水凝胶(PAHG)注射隆乳术后并发症的MRI特点及其诊断价值.方法 回顾性分析32例PAHG注射隆乳术后患者的临床和MRI资料.结果 32例中,17例(30只)乳腺假体包膜破裂,表现为多发的团块状长T1长T2信号,内可见低信号影分隔;6例(12只)硬结形成,表现为皮下、腺体内或肌间隙内散在的T2WI明亮高信号结节影;2例(4只)乳腺MRI可见假体严重变形、不对称;2例(4只)乳腺伴发无菌性炎症,表现为乳腺腺体结构紊乱,腺体内可见线条样稍长T1长T2信号影,增强扫描未见异常强化灶.MRI表现与临床表现相符.结论 聚丙烯酰胺水凝胶隆乳术后各种并发症的MRI表现有一定特点,乳腺MRI扫描可指导临床诊断和治疗.%Objective To investigate the clinical value of MRI in the diagnosis of the complications after breast augmentation with polyacrylamide hydrophilic gel(PAHG) injection. Methods The MR imaging and clinical data of 32 patients who had breast augmentation with polyacrylamide hydrophilic gel injection were retrospcctivcly revicwed. Results In the 32 patients,there were 17 cases (30 breasts) with envelope rupture, which appeared as mass- like long T1 and long T2 signal with hypointcnse septa; 6 cases (12 breasts) with callosity,the images showed sporadic nodules with high signal on T2 WI; prosthesis asymmetry occurred in 2 cases( 4 breasts ) ; aseptic inflammation in 2 cases ( 4 breasts) with pieces of slightly long T1 and long T2 signals , and none abnormal reinforcement was seen after enhancement. MRI findings were corresponded to the clinic manifcstations. Conclusion Since each complication after breast augmentation with PAHG has its own MR features,breast imaging plays important role in guiding clinical diagnosis and treatment for the complications after mammoplasty.

  4. 双向电泳分析鸢尾绿白嵌合叶片的蛋白质%Two-Dimensional Polyacrylamide Gel Electrophoresis Analysis of Proteins from Albino-Green Chimeric Leaves of Iris japonica

    Institute of Scientific and Technical Information of China (English)

    胡金勇; 曾英; 桑玉英

    2002-01-01

    Proteins extracted from albino-green chimeric leave s of Iris japonica were separated by two-dimensional polyacrylamide gel ele ctrophoresis and relative molecular weights and isoelectric points were determin ed.Approximately 400 protein spots were identified in each 2-D PAGE gel.Altered expression was apparent for at least 13 proteins.The results showed that protein expression patterns changed obviously in the green and white leaf strips.In com parison with the 2-D PAGE map from Arabidopsis thaliana in SWISS-2DPAGE data base, it revealed that the expression pattern of large subunit of RuBisCO and the proteins labeled W and T was closely related to the mechanism involved in chimeric variations of Iris japonic.%利用双向聚丙烯酰胺凝胶电泳对鸢尾(Iris japonica )绿白嵌合叶片的蛋白质进行分离,并初步鉴定了蛋白质的相对分子量和等电点.每个电泳图谱共检测到400余个蛋白点,其中至少13个蛋白的表达变化明显;结果表明,嵌合叶片的绿色与白色叶组织具有明显不同的蛋白质双向电泳图谱.与数据库中拟南芥双向电泳图谱相比较,发现Rubisco大亚基、标记为W和T蛋白的表达变化与产生绿白嵌合叶片的表型密切相关.

  5. Histone H4 N-terminal acetylation in Kasumi-1 cells treated with depsipeptide determined by acetic acid-urea polyacrylamide gel electrophoresis, amino acid coded mass tagging, and mass spectrometry.

    Science.gov (United States)

    Zhang, Liwen; Su, Xiaodan; Liu, Shujun; Knapp, Amy R; Parthun, Mark R; Marcucci, Guido; Freitas, Michael A

    2007-01-01

    Disrupted patterns of acetylation and deacetylation of core histones play an important role in silencing transcription of hematopoietic important genes in acute myeloid leukemia (AML). A thorough investigation of these mechanisms and the response to pharmacologic modifiers will provide a better understanding of the role of histone acetylation in leukemogenesis. We describe here an analytical approach that combines acid urea polyacrylamide gel electrophoresis (AU-PAGE), amino acid coded mass tagging (AACM), and mass spectrometry (MS) for the investigation of histone acetylation patterns. The combined approach was used to follow the dynamics of H4 acetylation in Kasumi-1 cells harboring the fusion gene AML1/ETO shown to aberrantly recruit histone deacetylases (HDACs). The histones in Kasumi-1 cells were labeled by growing the cells in media in which lysine was replaced with stable isotope-labeled lysine (Lys-D4). Labeled and unlabeled cells were treated with depsipeptide and analyzed at different time points (0, 4, 8, 12, 24, and 48 h). The cells were mixed, the histone was extracted, and acetylated H4 isoforms were separated using AU-PAGE before in-gel trypsin digestion. The digests were analyzed by MALDI-TOF MS. Peptides were identified by mass and isotope pattern. LC-MS/MS of Arg-C digests were also performed to verify the acetylation pattern for H4. The major pattern of acetylation was determined as follows: initial acetylation at K16, followed by acetylation at K12, and finally acetylation of either K8 and/or K5.

  6. Speciation analysis of inorganic arsenic in river water by Amberlite IRA 910 resin immobilized in a polyacrylamide gel as a selective binding agent for As(V) in diffusive gradient thin film technique.

    Science.gov (United States)

    Rolisola, Ana M C M; Suárez, Carlos A; Menegário, Amauri A; Gastmans, Didier; Kiang, Chang H; Colaço, Camila D; Garcez, Daniel L; Santelli, Ricardo E

    2014-09-07

    In this study, a method is proposed for the selective retention of As(V) using diffusive gradient in thin film (DGT) samplers containing a strongly basic anion exchange resin (Amberlite IRA 910) supported on a polyacrylamide gel. In addition, the total arsenic content is determined by ferrihydrite gel discs. Subsequently, the concentration of As(III) was obtained by determining the difference between the total As and As(V). DGT experiments showed linear accumulation of As(V) (up to 280 ng) until a deployment time of 8 h deployment (R(2) > 0.99). The retention of As(V) was appropriate (97.9-112.3%) between pH 5 and 9. For a solution with an ionic strength ranging from 0.001 to 0.05 mol L(-1), the As(V) uptake ranged from 90-120%. The proposed method was applied for the speciation of arsenic in river water. For the analysis of spiked samples collected at the Furnas stream, the recoveries of total arsenic content ranged between 103.9% and 118.8%. However, the recoveries of As(III) and As(V) were 43.3-75.2% and 147.3-153.4%, respectively. These differences were probably because of the oxidation of As(III) to As(V) during deployments. For spiked samples collected at the Ribeirão Claro, the recoveries of dissolved As(III), As(V) and As(T) were 103.1%, 108.0% and 106.3%, respectively. Thus, the DGT technique with Amberlite IRA 910 resin as the binding phase can be employed for the in situ redox speciation of inorganic arsenic.

  7. Association of triglyceride-rich lipoproteins-related markers and low-density lipoprotein heterogeneity with cardiovascular risk: effectiveness of polyacrylamide-gel electrophoresis as a method of determining low-density lipoprotein particle size.

    Science.gov (United States)

    Tani, Shigemasa; Matsumoto, Michiaki; Nagao, Ken; Hirayama, Atsushi

    2014-01-01

    Despite well-controlled low-density lipoprotein cholesterol (LDL-C), hypertriglyceridemia is an independent predictor of coronary events. We investigated the risk of atherosclerotic cardiovascular disease through examining the relation between triglyceride (TG) metabolism and LDL-heterogeneity as assessed by polyacrylamide-gel electrophoresis (PAGE). Estimated LDL-particle size [relative LDL migration (LDL-Rm value)] measured by PAGE with the LipoPhor system (Joko, Tokyo, Japan) was evaluated in 645 consecutive patients with one additional risk factor for atherosclerotic cardiovascular disease.Multivariate regression analysis after adjustments for traditional risk factors revealed an elevated triglyceride-rich lipoproteins (TRLs)-related markers [TG, remnant-like particle cholesterol (RLP-C), very LDL (VLDL) fraction, apolipoprotein (apo) C-II, and apo C-III] level to be an independent predictor of smaller-size LDL-particle size, both in the overall population, and in a subset of patients with serum LDL-C cardiovascular disease, it may be of particular importance to pay attention not only to the quantitative change in the serum LDL-C, but also TG-metabolism associated with LDL-heterogeneity. Combined evaluation of TRLs-related markers and LDL-Rm value may be useful for assessing the risk of atherosclerotic cardiovascular disease. Copyright © 2013 Japanese College of Cardiology. Published by Elsevier Ltd. All rights reserved.

  8. Metal imaging in non-denaturating 2D electrophoresis gels by laser ablation inductively coupled plasma mass spectrometry (LA-ICP-MS) for the detection of metalloproteins.

    Science.gov (United States)

    Becker, J Susanne; Lobinski, Ryszard; Becker, J Sabine

    2009-01-01

    Laser ablation inductively coupled plasma mass spectrometry (LA-ICP-MS) was developed as a powerful analytical technique for metal imaging of 2D gels for the detection of metalloproteins in rat kidney after electrophoretic separation. Protein complexes, extracted with water, were separated in their native state in the first and second dimension by blue native gel electrophoresis (BN-PAGE). Essential and toxic metals, such as zinc, copper, iron, manganese and lead, were monitored by LA-ICP-MS after gel ablation by a focused laser beam in a way that the total surface of a selected fragment of the gel was totally ablated. The metal distribution of this part of the gel was then constructed by plotting the metal (isotope) signal intensity as a function of the x,y (isoelectric point, molecular mass) coordinates of the gel. The proteins at locations rich in metals were cut out, digested with trypsin and analyzed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS).

  9. Combining blue native polyacrylamide gel electrophoresis with liquid chromatography tandem mass spectrometry as an effective strategy for analyzing potential membrane protein complexes of Mycobacterium bovis bacillus Calmette-Guérin

    Directory of Open Access Journals (Sweden)

    Li Weijun

    2011-01-01

    Full Text Available Abstract Background Tuberculosis is an infectious bacterial disease in humans caused primarily by Mycobacterium tuberculosis, and infects one-third of the world's total population. Mycobacterium bovis bacillus Calmette-Guérin (BCG vaccine has been widely used to prevent tuberculosis worldwide since 1921. Membrane proteins play important roles in various cellular processes, and the protein-protein interactions involved in these processes may provide further information about molecular organization and cellular pathways. However, membrane proteins are notoriously under-represented by traditional two-dimensional polyacrylamide gel electrophoresis (2-D PAGE and little is known about mycobacterial membrane and membrane-associated protein complexes. Here we investigated M. bovis BCG by an alternative proteomic strategy coupling blue native PAGE to liquid chromatography tandem mass spectrometry (LC-MS/MS to characterize potential protein-protein interactions in membrane fractions. Results Using this approach, we analyzed native molecular composition of protein complexes in BCG membrane fractions. As a result, 40 proteins (including 12 integral membrane proteins, which were organized in 9 different gel bands, were unambiguous identified. The proteins identified have been experimentally confirmed using 2-D SDS PAGE. We identified MmpL8 and four neighboring proteins that were involved in lipid transport complexes, and all subunits of ATP synthase complex in their monomeric states. Two phenolpthiocerol synthases and three arabinosyltransferases belonging to individual operons were obtained in different gel bands. Furthermore, two giant multifunctional enzymes, Pks7 and Pks8, and four mycobacterial Hsp family members were determined. Additionally, seven ribosomal proteins involved in polyribosome complex and two subunits of the succinate dehydrogenase complex were also found. Notablely, some proteins with high hydrophobicity or multiple transmembrane

  10. Mapping and identification of HeLa cell proteins separated by immobilized pH-gradient two-dimensional gel electrophoresis and construction of a two-dimensional polyacrylamide gel electrophoresis database

    DEFF Research Database (Denmark)

    Shaw, AC; Rossel Larsen, M; Roepstorff, P

    1999-01-01

    improvements of two-dimensional gel electrophoresis with immobilized pH gradients (IPG) compared to isoelectric focusing with carrier ampholytes, a highly reproducible method for examining global changes in HeLa cell protein expression due to different stimuli is now available. Therefore, we have initiated...

  11. Mapping and identification of HeLa cell proteins separated by immobilized pH-gradient two-dimensional gel electrophoresis and construction of a two-dimensional polyacrylamide gel electrophoresis database

    DEFF Research Database (Denmark)

    Shaw, AC; Rossel Larsen, M; Roepstorff, P

    1999-01-01

    improvements of two-dimensional gel electrophoresis with immobilized pH gradients (IPG) compared to isoelectric focusing with carrier ampholytes, a highly reproducible method for examining global changes in HeLa cell protein expression due to different stimuli is now available. Therefore, we have initiated...

  12. 家蝇抗菌物质诱导表达后的电泳分析%Induction of antibacterial peptide from Musca domestica and preliminary isolation by polyacrylamide gel electrophoresis

    Institute of Scientific and Technical Information of China (English)

    许兵红; 曾莉萍; 董卫华

    2011-01-01

    目的 探讨家蝇抗菌肽的诱导及电泳分离方法.方法 家蝇三龄幼虫针刺体壁诱导后,第48h提取其血淋巴,置于100℃、80℃水浴10s、30s、lmin、3min、5min等不同时间后,观察不同温度处理不同时间后的去杂蛋白效果和对溶壁微球菌的抑菌效果,并对各样品进行电泳分离.结果 血淋巴100℃水浴10s、30s、1min仍有抑菌活性,处理3min和5min无抑菌活性;80℃水浴30s-5min均有抑菌活性,电泳显示100℃处理3min和5min的样品与其他相比缺少2条蛋白区带.结论 针刺诱导的家蝇抗菌肽分离时血淋巴预处理采用80℃水浴lmin去杂蛋白效果较好,诱导的血淋巴100℃处理3min和5min即丧失抗菌活性,电泳显示缺少2条区带,此2条差异区带可能为抗菌肽组份.%The objective of this study was to induce and preliminarily isolate antibacterial peptide from Musca domestica by mean of polyacrylamide gel electrophoresis. After being induced by pricking for 48 hours, the hemolymph was collected and tested against Micrococcus lysodeikticus, and then the bacteriostatic effects were observed and the diameters of clear inhibitory ring were recorded respectively. After being bathed at 100℃ and 80℃ for 10sec, 30sec, lmin, 3min and 5min, each hemolymph was centrifuged at 10000r/min for 10min to get rid of unwanted proteins, and the supernatant was tested against M. lysodeikticus and displayed by polyacrylamide gel electrophoresis. The hemolymph didn't possess antibacterial activity against M. lysodeikticus after being bathed at 100℃ for 3min and 5min, but possessed antibacterial activity in others. It was also found that the two samples which were bathed at 100℃ for 3min and 5min were lack of two special bands from the others. It implies that bathing at 80℃ for 1min and then centrifuging could get rid of unwanted proteins in a large extent, which indicated that the two special bands are the components of the antibacterial peptides induced

  13. Identification and In Silico Analysis of Major Redox Modulated Proteins from Brassica juncea Seedlings Using 2D Redox SDS PAGE (2-Dimensional Diagonal Redox Sodium Dodecyl Sulfate Polyacrylamide Gel Electrophoresis).

    Science.gov (United States)

    Chaurasia, Satya Prakash; Deswal, Renu

    2017-02-01

    The thiol-disulphide exchange regulates the activity of proteins by redox modulation. Many studies to analyze reactive oxygen species (ROS), particularly, hydrogen peroxide (H2O2) induced changes in the gene expression have been reported, but efforts to detect H2O2 modified proteins are comparatively few. Two-dimensional diagonal redox sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS PAGE) was used to detect polypeptides which undergo thiol-disulphide exchange in Brassica juncea seedlings following H2O2 (10 mM) treatment for 30 min. Eleven redox responsive polypeptides were identified which included cruciferin, NLI [Nuclear LIM (Lin11, Isl-1 & Mec-3 domains)] interacting protein phosphatase, RuBisCO (ribulose-1,5-bisphosphate carboxylase/oxygenase) large subunit, and myrosinase. Redox modulation of RuBisCO large subunit was further confirmed by western blotting. However, the small subunit of RuBisCO was not affected by these redox changes. All redox modulated targets except NLI interacting protein (although it contains two cysteines) showed oxidation sensitive cysteines by in silico analysis. Interestingly, interactome of cruciferin and myrosinase indicated that they may have additional function(s) beside their well-known roles in the seedling development and abiotic stress respectively. Cruciferin showed interactions with stress associated proteins like defensing-like protein 192 and 2-cys peroxiredoxin. Similarly, myrosinase showed interactions with nitrilase and cytochrome p450 which are involved in nitrogen metabolism and/or hormone biosynthesis. This simple procedure can be used to detect major stress mediated redox changes in other plants.

  14. 高分子网络凝胶法制备Nd∶YAG纳米粉体及其表征%Preparation and Characterization of Nd ∶ YAG Nano-powders by Polyacrylamide Gel Method

    Institute of Scientific and Technical Information of China (English)

    徐晶晶; 冯欢欢; 孙丛立; 陈寰; 罗建勇; 晋勇; 焦志峰; 龚敏; 孙小松

    2013-01-01

    以Al(NO3)3·9H2O、Y(NO3)3·6H2O和Nd(NO3)3·6H2O为原材料,丙烯酰胺和N'N-亚甲基双丙烯酰胺为单体和网络剂,采用高分子网络凝胶法制备了Nd∶YAG纳米粉体.分别用热重-差热分析、X射线衍射、扫描电子显微镜、透射电子显微镜和荧光分光光度计对所得样品进行表征.结果表明,钇铝石榴石晶相的形成温度为880℃,粉体呈珊瑚虫状,粒径随着温度的升高而增大.Nd∶YAG纳米粉在401 nm处有强烈的发光峰,可以作为紫光光源的考虑对象;另外在研究中还发现了Nd3+在YAG中的浓度猝灭现象.%Using Al(NO3)3 · 9H2O,Y(NO3)3 · 6H2O and Nd(NO3)3 · 6H2O,acrylamide and N' N-methylene-bis-acrylamide as the starting materials,monomer and lattice reagent,respectively,the Nd ∶ YAG powders were prepared by polyacrylamide gel method.The prepared samples were characterized by thermo-gravimetric-differential scanning calorimetry,XRD,scanning electron microscopy,transmission electron microscopy,and photoluminescence spectroscopy.The results indicate that the formation temperature of pure YAG phase is 880 ℃.The YAG powders exhibit coral shape,and the particle size increases with the rise of temperature.The luminescence spectrum of YAG ∶ Nd3+ exhibits the strongest emission peak at 401 nm.Concentration quenching effect of Nd3+ in YAG was observed in the study.

  15. Analysis of bacterial diversity during the fermentation of inyu, a high-temperature fermented soy sauce, using nested PCR-denaturing gradient gel electrophoresis and the plate count method.

    Science.gov (United States)

    Wei, Chia-Li; Chao, Shiou-Huei; Tsai, Wen-Bin; Lee, Pei-Shan; Tsau, Nai-Hung; Chen, Jhih-Shan; Lai, Wen-Lin; Tu, James Ching-Yueh; Tsai, Ying-Chieh

    2013-04-01

    The diversity of bacteria associated with the fermentation of inyu, also known as black soy sauce, was studied through the nested PCR-denaturing gradient gel electrophoresis (DGGE) of samples collected from the fermentation stages of the inyu production process. The DGGE profiles targeted the bacterial 16S rDNA and revealed the presence of Citrobacter farmeri, Enterobacter cloacae, Enterobacter hormaechei, Enterococcus faecium, Klebsiella pneumoniae, Pantoea agglomerans, Salmonella enterica, Serratia marcescens, Staphylococcus sciuri and Weissella confusa. The bacterial compositions of 4 fermented samples were further elucidated using the plate count method. The bacteria isolated from the koji-making stage exhibited the highest diversity; Brachybacterium rhamnosum, E. hormaechei, K. pneumoniae, Kurthia gibsonii, Pantoea dispersa, Staphylococcus gallinarum, Staphylococcus kloosii and S. sciuri were identified. Koji collected during the preincubation stage presented the largest cell counts, and E. hormaechei, K. pneumoniae, E. cloacae and Enterobacter pulveris were identified. In brine samples aged for 7 and 31 days, the majority of the bacteria isolated belonged to 4 Bacillus species, but 4 Staphylococcus species and Delftia tsuruhatensis were also detected. This study demonstrates the benefits of using a combined approach to obtain a more complete picture of microbial populations and provides useful information for the control or development of bacterial flora during inyu fermentation.

  16. An assessment of the use of native and denatured forms of okra seed proteins as coagulants in drinking water treatment.

    Science.gov (United States)

    Jones, Alfred Ndahi; Bridgeman, John

    2016-10-01

    The effects of temperature, storage time and water pH on the coagulation performance of okra seed protein in water treatment were assessed. In a jar test experiment, okra salt extract achieved a notable improvement in treatment efficiency with storage time and showed good performance in quality after thermal treatment at 60, 97 and 140 °C temperatures for 6, 4 and 2 hours, respectively. The performance improvement of more than 8% is considered to be due to the denaturation and subsequent removal of coagulation-hindering proteins in okra seed. Furthermore, the results of a sodium dodecyl sulphate polyacrylamide gel electrophoresis analysis show two distinctive bands of protein responsible for the coagulation process after denaturation. It was further shown that at optimal coagulant dose, the pH of the treated water remained unaffected as a result of the protein's buffering capability during coagulation. Therefore, denatured okra seed exhibited improved performance compared to the native crude extract and offers clear benefits as a water treatment coagulant.

  17. 聚丙烯酰胺凝胶法制备NiFe2O4纳米颗粒及性能%Preparation and Properties of NiFe2O4 Nanoparticles by a Polyacrylamide Gel Route

    Institute of Scientific and Technical Information of China (English)

    赵东方; 杨华; 县涛; 王伟鹏; 魏智强; 李瑞山; 冯旺军; 姜金龙

    2013-01-01

    采用聚丙烯酰胺凝胶法制备了NiFe2O4纳米颗粒,利用XRD、SEM、紫外-可见漫反射光谱、FTIR、XPS、VSM等对样品进行了表征分析.结果表明:分别以EDTA和乙酸作络合剂时,在600℃烧结温度下可制得单相NiFe2O4纳米颗粒;两个样品的颗粒形貌较为规整,主要以类球形为主,粒度分布较为均匀,平均粒径分别为55 nm和75nm.根据紫外-可见漫反射光谱求得样品的带隙为1.85 eV.Fe2p3/2和Ni2p3/2的XPS谱分析表明:样品为反尖晶石型结构,其化学通式可表示为(Fe3+)[Ni2+ Fe3+]O4.磁滞回线测量结果表明:样品具有良好的软磁特性,颗粒尺寸较小的样品其饱和磁化强度、剩余磁化强度和矫顽力相对较小.%A polyacrylamide gel route was used to prepare NiFe2O4 nanoparticles. The prepared samples were characterized by X-ray diffraction (XRD) , scanning electron microscope (SEM) , ultraviolet-visible diffuse reflectance spectroscopy, fourier transform infrared spectroscopy (FTIR) , X-ray photoelectron spectroscopy (XPS) , and vibrating sample magnetometer (VSM). It is demonstrated that single-phase NiFe2O4 nanoparticles can be prepared separately using ethylenediamine-tetraacetic acid (EDTA) and acetic acid as the chelating agent at a calcining temperature of 600 ℃. The two as-prepared samples have regular spherical particle shapes and uniform particle sizes with an average diameter of 55 run and 75 nm, respectively. The energy bandgap of the samples is estimated to be 1.85 eV by ultraviolet-visible diffuse reflectance spectroscopy. Fe 2p3/2 and Ni 2p3/2 XPS spectra reveal that the samples have an inverse spinel-type structure with the chemical formula (Fe3+ ) [Ni2+Fe3+ ]O4. Magnetic hysteresis loop measurements indicate that the samples exhibit good soft magnetic properties, and their saturation magnetization, remanent magnetization and coercivity have a decrease with decrease in particle size.

  18. 影响聚丙烯酰胺水凝胶取出的因素及术后乳房整形策略探讨%Mammaplasty after polyacrylamide hydrophilic gel removal from breast

    Institute of Scientific and Technical Information of China (English)

    陈保国; 乔群; 黄渭清; 张海林; 朱琳; 曾昂

    2010-01-01

    目的 探讨聚丙烯酰胺水凝胶取出与乳房整形策略的关系. 方法 2003年2月至2009年8月,共收治聚丙烯酰胺水凝胶注射隆乳术后患者130例,于术前常规行乳腺B超或MRI检查,明确注射物的分布层次和周围组织浸润等情况,并在注射物取出术后根据是否植入假体或何时植入假体,将患者分为一期植入假体、二期植入假体和不植入假体3种情况,分别给予相应治疗. 结果 本组患者随访最长时间3个月,满意、基本满意和不满意率分别为83例(63.84%)、41例(31.53%)、6例(4.63%).基本满意组中5例6只乳房出现轻度包膜挛缩(Baker Ⅰ级);3例患者注射物残留过多,但仍执意要求植入假体,2例术后假体出现不平,但患者可以接受.不满意组皆为患者自行选择乳房假体,术后感到乳房外形与年龄不符,再次行假体取出术.除上述并发症外,无假体疝出、感染、切口裂开及双侧乳房不对称等畸形. 结论 根据聚丙烯酰胺水凝胶取出术后乳房畸形特点及乳房修复条件,选择恰当的再隆乳策略,既可改善胸部外观,又可缓解心理压力.%Objective To explore the relationship between mammaplasty and results after polyacrylamide hydrophilic gel(PAHG)removal from breast. Methods From Feb.2003 to Aug.2009,130 patients with bilateral breast augmentation by PAHG injection were treated.Preoperative ultrasound examination and MRI were performed to know the distribution of PAHG and infiltration at the surrounding tissue.According to the conditions after removal,the patients were received implant augmentationimmediately,or at the second stage,or no implant. Results The patients were followed up for 3 months at the most with a very satisfactory rate of 63.84%(83/120),a satisfactory rate of 31.53%(41/120)and a dissatisfactory rate of 4.63%(6/120).Slight capsular contracture(Baker Ⅰ)occurred in 5 cases with 6 breasts in satisfactory group.All the patients in dissatisfactory

  19. Bacterial communities in Agaricus bisporus compost analysed by denatu-ring gradient gel electrophoresis%双孢蘑菇培养料发酵过程中细菌群落结构分析

    Institute of Scientific and Technical Information of China (English)

    王琳; 李敏; 魏启舜; 张洪海; 周影; 赵荷娟

    2015-01-01

    PCR-denaturing gradient gel electrophoresis ( DGGE) was emloyed to analyze the V3 regions of bacterial 16S ribosomal DNA collected from the compost for Agaricus bisporus cultivation in spring and winter. The dominant DNA were cloned, sequenced and analyzed. Bacillus sp, Flavobacterium sp. , Lysinibacillus sp. , Thermus thermophilus,Pseudomonas fragi, Solibacillus silvestris,Thermobifida fusca, and two kinds of uncultured bacteria were identified in the compost. The bac-teria in spring and winter showed similar changing trend during phase I composting and similar community at the end of com-posting. The bacterial community structure differed at initial composting, phase I composting and phase II composting. The community diversity was environmental temperature-dependant. The bacteria existing in the compost all through the process of composting shared high sequence similarity with that of the bacteria degrading lignocellulose, which was favorable for the com-post to be applied as culture matrix.%为探明双孢蘑菇培养料发酵过程中细菌群落多样性和组成的演变,获得相关优势菌落的信息,本研究利用变性梯度凝胶电泳( Denaturing gradient gel electrophoresis,DGGE)对不同季节不同发酵阶段样品的的细菌进行特异性扩增,并选取主要DNA条带进行克隆、测序和生物信息学分析。结果显示,不同发酵时期带谱差异明显。发酵过程中,培养料中主要有芽孢杆菌属、黄杆菌属、杆菌属、假单胞菌属、Solibacillus、嗜热裂孢菌属、高温双歧菌属和未知分类地位的不可培养细菌。不同季节(春季、冬季)的培养料一次发酵过程中细菌多样性变化趋势相似,且发酵结束时细菌菌落结构相似。双孢蘑菇培养料在发酵过程中细菌群落结构至少经历了3个阶段的演替,即建堆初期、一次发酵阶段和二次发酵阶段。双孢蘑菇培养料发酵过程中细菌种群丰富,并随着发酵的不同阶段发生演

  20. Evaluation of a polyacrylamide hydrogel in the treatment of induced osteoarthritis in a goat model

    DEFF Research Database (Denmark)

    Tnibar, Aziz; Persson, Ann; Jensen, Henrik Elvang;

    2014-01-01

    Polyacrylamide hydrogel (PAAG) is an inert, non-degradable, non-immunogenic polymer gel with high viscoelasticity consisting of 97.5% sterile water and 2.5% cross-linked polyacrylamide. Its biocompatibility in soft tissues has been demonstrated. PAAG has recently been tested for the treatment...

  1. Combination of native and denaturing PAGE for the detection of protein binding regions in long fragments of genomic DNA

    Directory of Open Access Journals (Sweden)

    Metsis Madis

    2008-06-01

    Full Text Available Abstract Background In a traditional electrophoresis mobility shift assay (EMSA a 32P-labeled double-stranded DNA oligonucleotide or a restriction fragment bound to a protein is separated from the unbound DNA by polyacrylamide gel electrophoresis (PAGE in nondenaturing conditions. An extension of this method uses the large population of fragments derived from long genomic regions (approximately 600 kb for the identification of fragments containing protein binding regions. With this method, genomic DNA is fragmented by restriction enzymes, fragments are amplified by PCR, radiolabeled, incubated with nuclear proteins and the resulting DNA-protein complexes are separated by two-dimensional PAGE. Shifted DNA fragments containing protein binding sites are identified by using additional procedures, i. e. gel elution, PCR amplification, cloning and sequencing. Although the method allows simultaneous analysis of a large population of fragments, it is relatively laborious and can be used to detect only high affinity protein binding sites. Here we propose an alternative and straightforward strategy which is based on a combination of native and denaturing PAGE. This strategy allows the identification of DNA fragments containing low as well as high affinity protein binding regions, derived from genomic DNA ( Results We have combined an EMSA-based selection step with subsequent denaturing PAGE for the localization of protein binding regions in long (up to10 kb fragments of genomic DNA. Our strategy consists of the following steps: digestion of genomic DNA with a 4-cutter restriction enzyme (AluI, BsuRI, TruI, etc, separation of low and high molecular weight fractions of resultant DNA fragments, 32P-labeling with Klenow polymerase, traditional EMSA, gel elution and identification of the shifted bands (or smear by denaturing PAGE. The identification of DNA fragments containing protein binding sites is carried out by running the gel-eluted fragments alongside

  2. Improved high-throughput DNA fragment analyzer employing horizontal ultrathin gel electrophoresis

    Science.gov (United States)

    Brumley, Robert L., Jr.; Luckey, John A.

    1996-04-01

    We are currently developing a significantly improved gel electrophoresis and detection system that will allow more than an order of magnitude enhancement in the speed of DNA fragment analysis. This system is based upon the technique of horizontal ultrathin gel electrophoresis (HUGE) which employs denaturing polyacrylamide gels that are 75 microns thick. Because of the thinness of the gel, very high electric field strengths may be applied without deleterious thermal effects on resolution. Our proprietary fluorescence detector that scans the gel during electrophoresis allows for the simultaneous detection of up to four fluorophores. Because of the efficiency of the system of light collection, the gel can be scanned at speeds fast enough to generate high resolution gel images despite the high speed of separations. In addition, we are able to increase sample density by collecting 500 datapoints across the width of the gel. The resulting instrument has the capability to separate and resolve single-stranded DNA molecules that are between 25 and 300 bases in length from each of 60 lanes in less than 45 minutes. With the advent of 96 lane gels and attendant automation, this instrument will have the ability to analyze 18,432 genotypes per day.

  3. 牙周牙髓联合病变菌群的PCR-DGGE分析%Bacterial analysis of combined periodontal-endodontic lesions using polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE)

    Institute of Scientific and Technical Information of China (English)

    夏明慧; 亓庆国

    2012-01-01

    目的 利用变性梯度凝胶电泳(DGGE)技术观察牙周牙髓联合病变患牙牙周组织和根管中原始菌群分布状况及其异同,并通过克隆测序技术来试图探讨两部位可能存在的优势菌.方法 从13例牙周牙髓联合病变患牙分别采集患牙根尖1/3处牙周细菌和根管牙髓细菌,提取细菌总DNA,扩增16S rRNA基因可变区,再进行变性梯度凝胶电泳.应用SPSS17.0软件和Quantity One软件对DGGE图谱菌种条带进行统计学分析和聚类分析.对DGGE凝胶中有代表性的条带进行回收和克隆测序.结果 两取菌部位的菌种条带数间有明显的统计学差异(P<0.01),但二者之间无正相关性.二者间的相似系数为13.1% ~62.5%.牙周牙髓联合病变患牙根尖区1/3处牙周菌属可能有弯曲菌属(Campylobacter)、梭杆菌属(Fusobacterium)、奈瑟菌属(Neisseria)等,该处对应根管中菌属可能有优杆菌属(Mogibacterium)、棒状杆菌属(Corynebacterium)、放线菌属(Actinomyces)等.结论 牙周牙髓联合病变(牙周来源)中牙周组织和邻近根管牙髓组织中菌种在数目和结构上有明显不同.该病变牙周组织和根管中可能存在目前尚未被认知的优势菌种.%Objective To compare the bacterial community profiles present in periodontium and root canals of the same tooth diagnosed as combined periodontal-endodontic lesions by using denaturing gradient gel electrophoresis (DGGE).Methods Samples were collected from 13 extracted teeth with advanced periodontitis,endodontic samples from root tip 1/3 root canal,and periodontal samples from the corresponding neighboring periodontium.Genomic DNA was collected for the following universal bacterial primersPCR.The PCR products were then loaded on the DGGE gels to gain separate bands.The typical DGGE bands were excised,PCR-cloned and sequenced.Results The number of bands,which was indicative of the number of bacterial species,was compared intra-group (periodontal and

  4. [A method for determining DNA sequence by labeling the end of the molecule and cleaving at the base. Isolation of DNA fragments, end-labeling, cleavage, electrophoresis in polyacrylamide gel and analysis of results].

    Science.gov (United States)

    Maxam, A M; Gilbert, W

    1986-01-01

    We elaborate basic chemical principles and current laboratory procedures for sequencing end-labeled DNA by partial cleavage and gel electrophoresis (A. M. Maxam and W. Gilbert, Proc. Natl. Acad. Sci. USA, 1977, v. 74, p. 560-564). We provide step-by-step protocols for 32P-labeling DNA ends, segregating the labeled ends by cutting with a second restriction enzyme or separating strands, partially cleaving the DNA at specific bases with reagents, electrophoresing the labeled products of cleavage on sequencing gels, and interpreting sequencing band patterns. Many of these procedures have been condensed, to make them faster and easier, and some are new. We also discuss sequencing strategies, and suggest a technique which will reduce plasmid or viral DNA to a collection of singly-end-labeled fragments in one day, for efficient sequencing of these chromosomes in 250-nucleotide blocks.

  5. Heat-induced whey protein gels: protein-protein interactions and functional properties.

    Science.gov (United States)

    Havea, Palatasa; Watkinson, Philip; Kuhn-Sherlock, Barbara

    2009-02-25

    Heat-induced gelation (80 degrees C for 30 min or 85 degrees C for 60 min) of whey protein concentrate (WPC) solutions was studied using small deformation dynamic rheology, small and large deformation compression, and polyacrylamide gel electrophoresis (PAGE). The WPC solutions (15% w/w, pH 6.9) were prepared by dispersing WPC powder in water (control), 1% (w/w) sodium dodecyl sulfate (SDS) solution, and N-ethylmaleimide (NEM) solution at a protein/NEM molar ratio of 1:1 or in 10 mM dithiothreitol (DTT) solution. PAGE analyses showed that the heat treatment of control solutions contained both disulfide and non-covalent linkages between denatured protein molecules. Only disulfide linkages were formed in heated SDS-WPC solutions, whereas only non-covalent linkages were formed in DTT-WPC and NEM-WPC solutions during heating. In heated NEM-WPC solutions, the pre-existing disulfide linkages remained unaltered. Small deformation rheology measurements showed that the storage modulus (G') values, compared with those of the control WPC gels (approximately 14000 Pa), were 3 times less for the SDS-WPC gels (approximately 4000 Pa), double for the NEM-WPC gels (approximately 24000 Pa), and even higher for the DTT-WPC gels (approximately 30000 Pa). Compression tests suggested that the rubberiness (fracture strain) of the WPC gels increased as the degree of disulfide linkages within the gels increased, whereas the stiffness (modulus) of the gels increased as the degree of non-covalent associations among the denatured protein molecules increased.

  6. Borehole testing methods using a new temporary polyacrylamide packers technology

    Science.gov (United States)

    Klepikova, Maria V.; Roques, Clement; Selker, John

    2017-04-01

    Range of options for investigation of hydraulic behavior of aquifers from boreholes has been limited to rigid, cumbersome packers, and inflatable sleeves. Here we propose a new temporary polyacrylamide packers (TAMP) technology that uses soft grains of polyacrylamide gel as a borehole sealing material and discuss its possible applications. Polyacrylamide gel, also called hydrogel or water-absorbing polymer, consists of long chains of molecules that can absorb over a hundred times their weight in liquids. Soft gel grains are mainly made of water, but the water inside these particles does not contribute to the flow of the suspension. The gel packing (permeability similar to open gravel) placed to a well suppresses free convection, allowing for local temperature and chemical sampling through free-flowing gel. Minimizing the effect of free convection within the well column would be beneficial for active thermal tests where free convection often dominate flow and create thermal disequilibrium between the water in the borehole and the surrounding media. Preliminary laboratory experiments and the literature suggests that as the polyacrylamide pack is subject to modest compressive stress to the gel media (of order 0.1 ATM), the permeability transitions from of the order of 10 to 7 millidarcys to 0.01 millidarcys, illustrating the remarkable ability to transition from highly permeable to nearly impermeable grouting. Though yet to be confirmed in the field, by locally injecting water at pressure greater than the compressive stress, local voids can be formed which can act as local pump test sources, with all other locations in the borehole hydraulically isolated where local response pressure from the formation can be measured. This arrangement could be valuable for tomographic study of aquifers wherein hundreds of injection zones could be tested by simply pulling an injection pipe vertically through the packed borehole. The gel grains can be of the scale of cm, so do not pass

  7. Virus inactivation by protein denaturants used in affinity chromatography.

    Science.gov (United States)

    Roberts, Peter L; Lloyd, David

    2007-10-01

    Virus inactivation by a number of protein denaturants commonly used in gel affinity chromatography for protein elution and gel recycling has been investigated. The enveloped viruses Sindbis, herpes simplex-1 and vaccinia, and the non-enveloped virus polio-1 were effectively inactivated by 0.5 M sodium hydroxide, 6 M guanidinium thiocyanate, 8 M urea and 70% ethanol. However, pH 2.6, 3 M sodium thiocyanate, 6 M guanidinium chloride and 20% ethanol, while effectively inactivating the enveloped viruses, did not inactivate polio-1. These studies demonstrate that protein denaturants are generally effective for virus inactivation but with the limitation that only some may inactivate non-enveloped viruses. The use of protein denaturants, together with virus reduction steps in the manufacturing process should ensure that viral cross contamination between manufacturing batches of therapeutic biological products is prevented and the safety of the product ensured.

  8. Stability of collagen during denaturation.

    Science.gov (United States)

    Penkova, R; Goshev, I; Gorinstein, S; Nedkov, P

    1999-05-01

    The stability of calf skin collagen (CSC) type I during thermal and chemical denaturation in the presence of glycerol was investigated. Thermal denaturation of type I collagen was performed in the presence of glycerol or in combination with urea and sodium chloride. The denaturation curves obtained in the presence of urea or sodium chloride retained their original shape without glycerol. These curves were shifted upward proportionally to the glycerol concentration in the reaction medium. This means that glycerol and the denaturants act independently. The explanation is based on the difference in the mechanism of their action on the collagen molecule.

  9. Studies of muscle proteins in embryonic myocardial cells of cardiac lethal mutant mexican axolotls (Ambystoma mexicanum) by use of heavy meromyosin binding and sodium dodecyl sulfate polyacrylamide gel electrophoresis

    Science.gov (United States)

    1976-01-01

    In the Mexican axolotl Ambystoma mexicanum recessive mutant gene c, by way of abnormal inductive processes from surrounding tissues, results in an absence of embryonic heart function. The lack of contractions in mutant heart cells apparently results from their inability to form normally organized myofibrils, even though a few actin-like (60-A) and myosin-like (150-A) filaments are present. Amorphous "proteinaceous" collections are often visible. In the present study, heavy meromyosin (HMM) treatment of mutant heart tissue greatly increases the number of thin filaments and decorates them in the usual fashion, confirming that they are actin. The amorphous collections disappear with the addition of HMM. In addition, an analysis of the constituent proteins of normal and mutant embryonic hearts and other tissues is made by sodium dodecyl sulfate (SDS) gel electrophoresis. These experiments are in full agreement with the morphological and HMM binding studies. The gels show distinct 42,000-dalton bands for both normal and mutant hearts, supporting the presence of normal actin. During early developmental stages (Harrison's stage 34) the cardiac tissues in normal and mutant siblings have indistinguishable banding patterns, but with increasing development several differences appear. Myosin heavy chain (200,000 daltons) increases substantially in normal hearts during development but very little in mutants. Even so the quantity of 200,000-dalton protein in mutant hearts is significantly more than in any of the nonmuscle tissues studied (i.e. gut, liver, brain). Unlike normal hearts, the mutant hearts lack a prominent 34,000-dalton band, indicating that if mutants contain muscle tropomyosin at all, it is present in drastically reduced amounts. Also, mutant hearts retain large amounts of yolk proteins at stages when the platelets have virtually disappeared from normal hearts. The morphologies and electrophoresis patterns of skeletal muscle from normal and mutant siblings are

  10. Bacterial infection as a likely cause of adverse reactions to polyacrylamide hydrogel fillers in cosmetic surgery

    DEFF Research Database (Denmark)

    Christensen, Lise; Breiting, Vibeke; Bjarnsholt, Thomas

    2013-01-01

    Background. The etiology of long-lasting adverse reactions to gel fillers used in cosmetic surgery is not known. Bacterial infection and immunological reaction to the product have been suggested. Methods. We performed a case-control study, with 77 biopsies and 30 cytology specimens originating from...... in the presence of polyacrylamide filler in cosmetic surgery, possibly due to a biofilm mode of growth. Adequate skin preparation and use of sterile technique in these procedures are mandatory, but antibiotic prophylaxis prior to injection of nondegradable gels like polyacrylamide should be explored as well....

  11. Network generation enhances interpretation of proteomics data sets by a combination of two-dimensional polyacrylamide gel electrophoresis and matrix-assisted laser desorption/ionization-time of flight mass spectrometry.

    Science.gov (United States)

    Wang, Xijun; Zhang, Aihua; Sun, Hui; Wu, Gelin; Sun, Wenjun; Yan, Guangli

    2012-10-21

    Recent advances in proteomic technologies have enabled us to create detailed protein-protein interaction maps in diseases. As the size of the interaction dataset increases, powerful computational methods are required in order to effectively interpret network models from large scale interactome data. In this study, we carried out comparative proteomics to construct and identify the proteins networks associated with hepatic injury (HI) which are largely unknown, as a case study. All proteins expressed were separated and identified by two-dimensional gel electrophoresis (2-DE) and matrix-assisted laser desorption/ionization time-of-flight-time-of-flight mass spectrometry (MALDI-TOF/TOF MS). Protein-interacting networks and pathways were mapped using STRING analysis program. We have performed for the first time a comprehensive profiling of changes in protein expression of HI rats, to uncover the networks altered by treated with CCl(4). Identification of fifteen spots (seven over-expressed and eight under-expressed) were established by MALDI-TOF/TOF MS. These proteins were subjected to functional pathway analysis using STRING software for better understanding of the biological context of the identified proteins. It suggested that modulation of multiple vital physiological pathways including DNA repair process, cell apoptosis, oxidation reduction, signal transduction, metabolic process, intracellular signaling cascade, regulation of biological processes, cell communication, regulation of cellular process, and molecular transport. In summary, the present study provides the first protein-interacting network maps and novel insights into the biological responses and potential pathways of HI. The generation of protein interaction networks clearly enhances the interpretation of proteomic data, particularly in respect of understanding molecular mechanisms of panel protein biomarkers.

  12. Protein extraction from Mycobacterium avium subsp. paratuberculosis: Comparison of methods for analysis by sodium dodecyl sulphate polyacrylamide gel electrophoresis, native PAGE and surface enhanced laser desorption/ionization time of flight mass spectrometry.

    Science.gov (United States)

    Gumber, Sanjeev; Taylor, Deborah L; Whittington, Richard J

    2007-01-01

    Mycobacterium paratuberculosis causes Johne's disease, a chronic bowel disease in ruminants worldwide and is currently incurable. This study was conducted to compare methods for examining the proteome of M. paratuberculosis. SDS-PAGE, native PAGE and SELDI-TOF-MS were compared and the efficacy of various lysis buffers was assessed. Chaotropic agents (Urea CHAPS and potassium thiocyanate) and non-ionic detergent (Tween20 and Triton X-100) extracts were compared on three different ProteinChip surfaces along with two energy absorbing molecules (EAM): EAM-1 proprietary formulation and sinapinic acid (Ciphergen). Urea CHAPS was efficient for extraction of proteins and their detection on all the ProteinChip surfaces. However, potassium thiocyanate was the most effective buffer, leading to detection of the greatest number of protein peaks on the immobilized metal affinity chromatography (IMAC) surface. Sinapinic acid was more efficient than the EAM-1 proprietary formulation and resulted in additional peaks with higher intensity for both the low and the medium molecular weight range proteins. Intra-chip and inter-chip coefficient of variation for mass/charge varied from 0.01% to 0.07% and 0.00% to 0.08%, respectively. SELDI-TOF-MS was an efficient tool for the protein profiling of M. paratuberculosis and will be useful for investigation of novel proteins, although SDS-PAGE/2D gel electrophoresis is recommended for study of high molecular weight species. All buffers were suitable for protein extraction for SDS-PAGE, while Tween20 was best for native PAGE.

  13. Simultaneous immunoblotting analysis with activity gel electrophoresis and 2-D gel electrophoresis.

    Science.gov (United States)

    Lee, Der-Yen; Chang, Geen-Dong

    2015-01-01

    Diffusion blotting method can couple immunoblotting analysis with another biochemical technique in a single polyacrylamide gel, however, with lower transfer efficiency as compared to the conventional electroblotting method. Thus, with diffusion blotting, protein blots can be obtained from an SDS polyacrylamide gel for zymography assay, from a native polyacrylamide gel for electrophoretic mobility shift assay (EMSA) or from a 2-D polyacrylamide gel for large-scale screening and identification of a protein marker. Thereafter, a particular signal in zymography, electrophoretic mobility shift assay, and 2-dimensional gel can be confirmed or identified by simultaneous immunoblotting analysis with a corresponding antiserum. These advantages make diffusion blotting desirable when partial loss of transfer efficiency can be tolerated or be compensated by a more sensitive immunodetection reaction using enhanced chemiluminescence detection.

  14. Analysis of metal-binding proteins separated by non-denaturating gel electrophoresis using matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) and laser ablation inductively coupled plasma mass spectrometry (LA-ICP-MS).

    Science.gov (United States)

    Becker, J Susanne; Mounicou, Sandra; Zoriy, Miroslav V; Becker, J Sabine; Lobinski, Ryszard

    2008-09-15

    Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) and laser ablation inductively coupled plasma mass spectrometry (LA-ICP-MS) have become established as very efficient and sensitive biopolymer and elemental mass spectrometric techniques for studying metal-binding proteins (metalloproteins) in life sciences. Protein complexes present in rat tissues (liver and kidney) were separated in their native state in the first dimension by blue native gel electrophoresis (BN-PAGE). Essential and toxic metals, such as zinc, copper, iron, nickel, chromium, cadmium and lead, were detected by scanning the gel bands using quadrupole LA-ICP-MS with and without collision cell as a microanalytical technique. Several proteins were identified by using MALDI-TOF-MS together with a database search. For example, on one protein band cut from the BN-PAGE gel and digested with the enzyme trypsin, two different proteins - protein FAM44B and cathepsin B precursor - were identified. By combining biomolecular and elemental mass spectrometry, it was possible to characterize and identify selected metal-binding rat liver and kidney tissue proteins.

  15. Application of Carbon Quantum Dots in Fluorescent Imaging of Human Serum Proteins after Polyacrylamide Gel Electrophoresis%碳量子点荧光成像法应用于聚丙烯酰胺凝胶电泳检测人血清蛋白质的研究

    Institute of Scientific and Technical Information of China (English)

    刘亭廷; 彭程; 马云川; 欧阳津

    2013-01-01

    A new method of carbon quantum dots (CQDs) fluorescent imaging for human serum proteins detection after polyacrylamide gels electrophoresis (PAGE) is established.Polyacrylamide gel electrophoresis is one of the most general and powerful technique to separate complex biosamples,and it is widely used in molecular biology,biochemistry and medicine.With the evolution of clinical proteomics,the development of a novel method to detect serum proteins atter PAGE with high resolution and high sensitivity is of great significant.As far as we know,the carbon quantum dots have not been applied in the detection of serum proteins after PAGE.The fluorescent carbon quantum dots were synthesized by a one-step microwave pyrolysis method:glycerol and phosphate buffer (7.1 mmol.L-1,pH 7.4) (Φ=70%) was mixed evenly,and then put the solution into the advanced microwave digestion system and heated for 14 min (750 W),the colorless solution turned to yellow after reaction.The yellow solution was diluted into the incubation solution with HOAc-NaOAc buffer (80 mmol.L-1,pH 2.7),and then used for staining serum proteins.The emission and excitation spectra of carbon quantum dots were measured,and the excitation wavelength allowed the use of an ultraviolet lamp at 365 nm for the fluorescent imaging.In order to investigate the performance of CQDs fluorescent imaging,the dilution of human serum samples separated by electrophoresis was used for detection,and was compared with traditional method of CBB-R250 staining and silver staining.The results suggested that CQDs based on fluorescent imaging could effectively detect human serum proteins with high sensitivity and resolution.The sensitivity of CQDs imaging was higher than CBB staining and comparable to silver staining.Therefore,the CQDs fluorescent imaging could be an inexpensive,time-saving,pollution-free and convenient method with high sensitivity and resolution for the detection of human serum proteins.It demonstrates the CQDs fluorescent

  16. SWELLING EQUILIBRIUM OF NONIONIC POLYACRYLAMIDE HYDROGEL IN AQUEOUS SALT SOLUTIONS

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    A series of nonionic polyacrylamide hydrogels, using acrylamide as monomer and N,N’-methylene diacrylamide as crosslinking agent, were prepared by the free radical polymerization in aqueous solution. Swelling equilibria for the gels were carried out in aqueous solutions of NaCl, KCl, CaCl2, Na2HPO4 and K2HPO4 with concentration ranging from 10-3 to 5mol/kgH2O at 25℃. Experimental results revealed that the chlorides and phosphates cause a different behavior at higher salt concentration. The swelling ratio increases with increasing concentration of chlorides salts, while decreases with the increased phosphates salt concentration. The phenomena seem to be related to the different interactions of chloride and hydrogen phosphate ions with the network groups. Furthermore, the effects of different concentration of crosslinking agent and total monomers on gel swelling performance were also investigated.

  17. Scalable lithography from Natural DNA Patterns via polyacrylamide gel

    National Research Council Canada - National Science Library

    Qu, JieHao; Hou, XianLiang; Fan, WanChao; Xi, GuangHui; Diao, HongYan; Liu, XiangDon

    2015-01-01

    ...) that controllably and precisely shrinks and swells with water content. Aligned patterns of natural DNA molecules were prepared by evaporative self-assembly on a PMMA substrate, and were transferred to unsaturated polyester resin (UPR...

  18. Denaturing gradient gel electrophoresis analysis of 16S rDNA V3 fragment of bacteria in transgenic wheat soil%转基因小麦田土壤细菌16S rDNA V3片段的DGGE分析

    Institute of Scientific and Technical Information of China (English)

    丁衬衬; 周艳红; 林凡云; 徐剑宏; 吴季荣; 祭芳; 史建荣

    2011-01-01

    为长期监测转基因作物对土壤微生物菌群变化的影响提供可靠的试验方法.以检测转基因小麦田土壤细菌菌群变化为例,利用细菌特异性引物,扩增得到了土壤细菌16S rDNA V3可变区片段;通过变性梯度凝胶电泳(Denaturing gradient gel electrophoresis,DGGE),分析PCR扩增得到的V3可变区片段;根据电泳条带的变化情况评价转基因小麦对土壤环境微生物的安全性.结果表明该方法所提取的DNA,可以不经过纯化直接进行PCR扩增等后续试验;DGGE图谱中条带清晰.该方法可作为长期监测土壤微生物种群变化的试验手段.%This experiment was carried out to establish an effective method for detecting the impact of virus resistant transgenic wheat on its environment bacteria. Variable region fragments of 16S rDNA V3 were amplified by specific and sensitive primers of bacteria, analyzed by denaturing gradient gel electrophoresis ( DGGE) , and applied to evaluate the effect of transgenic wheat on the soil bacterium, according to the change of electrophoretic bands. The results showed that the DNA by this extraction method could be used in PCR without purification. On DGGE profiles, the lighter bands of each lane represented their corresponding dominant microorganisms. The influence of transgenic wheat and non-transgenic wheat on soil microbes by the number and brightness of the bands were obtained, so microorganism population fluctuation could be demonstrated effectively with the method.

  19. Proteinograma sérico de bezerros recém-nascidos da raça Holandesa obtido por eletroforese em gel de poliacrilamida Serum protein concentration in newborn Holstein calves determined by means of sodium dodecyl sulphate-polyacrylamide gel electrophoresis

    Directory of Open Access Journals (Sweden)

    J.J. Fagliari

    2006-06-01

    Full Text Available The serum protein concentration of newborn Holstein calves determined by means of sodium dodecyl sulphate-polyacrylamide (SDS-PAGE was studied. Blood samples from 40 healthy newborn calves were obtained 48 hours after birth. Calves had been given 3 liters of colostrum within 2 hours after birth, following by dose corresponding by 15% of animal weight each 24 hours. The results showed three different proteinograms: 19 calves had 14 proteins with molecular weights (MW ranging from 28,000 D to 170,000D (proteinogram 1; 11 calves had 14 proteins with MW ranging from 18,000 to 170,000 D (proteinogram 1; and 10 calves had 12 proteins with MW ranging from 28,000 D to 170,000 D (proteinogram 3. The three groups presented similar IgG levels. The highest serum concentration of ceruloplasmin were verified in proteinogram 3, which had the lowest serum level of protein with MW 58,000D. It was verified a1-antitrypsin only in proteinogram 2, which had no proteins with MW of 42,000 D and 37,000D. The highest serum concentrations of IgA and protein with MW 58,000 D, and the lowest serum levels of transferrin, haptoglobin, and acid glycoprotein were verified in proteinogram 3. Measurement of serum protein concentrations by SDS-PAGE may be useful in monitoring the occurrence of hypogammaglobulinemia and the neonatal disease in calves.

  20. 利用变性梯度凝胶电泳分析正红菇菌根围土壤真菌群落多样性%Analysis of Fungal Diversity of Russula griseocarnosa Mycorrhizosphere Soil with Denaturing Gradient Gel Electrophoresis

    Institute of Scientific and Technical Information of China (English)

    肖冬来; 陈丽华; 陈宇航; 杨菁; 黄小菁

    2013-01-01

    以正红菇(Russula griseocarnosa)菌根围土壤为研究对象,通过提取土壤基因组DNA,以通用引物扩增真菌18S rRNA基因V1+V2区,将PCR产物进行变性梯度凝胶电泳(Denaturing Gradient Gel Electrophoresis),获得土壤微生物群落的DNA特征指纹图谱,并对图谱中的优势条带回收测序,通过Blast进行同源性比对并构建系统发育树,进而分析正红菇菌根围真菌群落组成及多样性.同源性比对结果表明,在回收测序的19条DGGE条带中,4条为非真菌真核生物序列,系统发育分析显示全部序列可以分为4类菌群,Group Ⅰ主要为担子菌门(Basidiomycota)真菌,GroupⅡ主要为子囊菌门(Ascomycota)真菌,GroupⅢ为未知真菌,GroupⅣ主要为节肢动物门生物(Arthropoda).

  1. Evaluation du potentiel radiosensibilisateur ou radioprotecteur/antioxydant de quelques composes selectionnes par dosimetrie par gel de polyacrylamide et dosimetre de Fricke, et utilisation de la filamentation par impulsion laser infrarouge fenitoseconde comme un nouveau et puissant faisceau pour la radiotherapie du cancer

    Science.gov (United States)

    Meesat, Ridthee

    In radiation treatment, a sufficiently high radiation dose must be delivered to the tissue volumes containing the tumor cells while the lowest possible dose should be deposited in surrounding healthy tissue. We developed an original approach that is fast and easy to implement for the early assessment of the efficiency of radiation sensitizers and protectors. In addition, we characterized a new femtosecond laser pulse irradiation technique. We are able to deposit a considerable dose with a very high dose rate inside a well-controlled macroscopic volume without deposition of energy in front or behind the target volume. The radioprotective efficiency was measured by irradiation of the Fricke solution incorporating a compound under study and measuring the corresponding production of ferric ions G(Fe3+). The production of ferric ions is most sensitive to the radical species produced in the radiolysis of water. We studied experimentally and simulated with a full Monte-Carlo computer code the radiation-induced chemistry of Fricke/cystamine solutions. Results clearly indicate that the protective effect of cystamine originates from its radical-capturing ability, which allows this compound to compete with the ferrous ions for the various free radicals - especially ·OH radicals and H· atoms - formed during irradiation of the surrounding water. The sensitizing capacity of radiation sensitizers was measured by irradiation of a polyacrylamide gel (PAG) dosimeter incorporating a compound under study and measuring the corresponding increase in the gradient between spin-spin relaxation rate (R2) and absorbed dose. We measured an irradiation energy-dependent increase in R 2-dose sensitivity for halogenated compounds or a decrease for radioprotectors. Finally, we studied a novel laser irradiation method called "filamentation". We showed that this phenomenon results in an unprecedented deposition of energy and the dose rate thus achieved exceeds by orders of magnitude values

  2. Bioconversion of acrylonitrile to acrylamide using polyacrylamide entrapped cells of Rhodococcus rhodochrous PA-34.

    Science.gov (United States)

    Raj, J; Prasad, S; Sharma, N N; Bhalla, T C

    2010-09-01

    The nitrile hydratase (NHase) of Rhodococcus rhodochrous PA-34 catalyzed the conversion of acrylonitrile to acrylamide. The resting cells (having NHase activity) (8 %; 1 mL corresponds to 22 mg dry cell mass, DCM) were immobilized in polyacrylamide gel containing 12.5 % acrylamide, 0.6 % bisacrylamide, 0.2 % diammonium persulfate and 0.4 % TEMED. The polyacrylamide entrapped cells (1.12 mg DCM/mL) completely converted acrylonitrile in 3 h at 10 °C, using 0.1 mol/L potassium phosphate buffer. In a partitioned fed batch reactor, 432 g/L acrylamide was accumulated after 1 d. The polyacrylamide discs were recycled up to 3×; 405, 210 and 170 g/L acrylamide was produced in 1st, 2nd and 3rd recycling reactions. In four cycles, a total of 1217 g acrylamide was produced by recycling the same mass of entrapped cells.

  3. Heating of polyacrylamide ferrogel by alternating magnetic field

    Energy Technology Data Exchange (ETDEWEB)

    Safronov, A.P., E-mail: Safronov@iep.uran.ru [Ural Federal University, Yekaterinburg (Russian Federation); Institute of Elecrophysics, UB RAS, Yekaterinburg (Russian Federation); Samatov, O.M. [Institute of Elecrophysics, UB RAS, Yekaterinburg (Russian Federation); Tyukova, I.S.; Mikhnevich, E.A. [Ural Federal University, Yekaterinburg (Russian Federation); Beketov, I.V. [Ural Federal University, Yekaterinburg (Russian Federation); Institute of Elecrophysics, UB RAS, Yekaterinburg (Russian Federation)

    2016-10-01

    Ferrogel based on polacryamide network with embedded maghemite nanoparticles with mean number average particle diameter 12 nm was synthesized by radical polymerization in water-based ferrofluid. The network structure of ferrogel was characterized by Flory–Rehner theory and it was shown that the embedded particles were substantially larger than the mesh size. It prevented the translational movement of particles in the ferrogel. The immobilization of particles was confirmed by dynamic light scattering. The adhesion of macromolecular chains to the particles was determined by calorimetry using thermochemical cycle. The enthalpy of interfacial adhesion was found several orders of magnitude higher than the energy of dipoles in typically applied magnetic fields. Despite the differenve in the mobility of particles in ferrofluid and ferrogel the comparative study of their heating in alternating magnetic field, however, revealed their close similarity. In both cases it was goverened by superposing of Neel and Brownian relaxation mechanisms. - Highlights: • We synthesized polyacrylamide ferrogel with maghemite nanoparticles. • Nanoparticles are entrapped into gel network. • Polyacrylamide chains are strongly linked to the particles. • Brownian relaxation contributes to heating of ferrogel in alternating field.

  4. Rheological properties of sweet potato starch before and after denaturalization

    Institute of Scientific and Technical Information of China (English)

    肖华西; 林亲录; 夏新剑; 李丽辉; 林利忠; 吴卫国

    2008-01-01

    Based on the sweet potato starch,cationic starch,acetic starch and cationic-acetic compoundedly modified starch were made through chemical denaturalization.The above three kinds of static rheological parameter and dynamic rheological parameter were measured,respectively.The experimental result reveals that the thermal stability of starchy viscosity increases after chemical denaturalization.Under the condition of identical shearing rate,the shear stress of cationic-acetic ester compoundedly modified sweet potato starch paste is the largest among these kinds of sweet potato starch.This attributes to a phenomenon of shearing thinning.Furthermore,raw sweet potato starch has a larger gel intensity than that of modified starch.

  5. 利用DGGE分析无龋和有龋儿童牙菌斑细菌组成%Profiling of microbial composition in dental plaque from caries-free and severe early childhood caries children analysed by denaturing gradient gel electrophoresis

    Institute of Scientific and Technical Information of China (English)

    王帅; 黄博; 杜宁; 刘筱娣; 郭丽宏

    2011-01-01

    目的:通过变性梯度凝胶电泳(denaturing gradient gel electrophoresis,DGGE)技术分析无龋(caries free,CF)儿童和重型早期婴幼儿龋(severe early childhood caries,SECC)儿童集合牙菌斑内细菌多样性的差异.方法:无龋儿童和SECC儿童各34 例,牙菌斑基因组DNA等量混合后,制备无龋和SECC儿童牙菌斑基因组库,进行全基因组放大.分别以放大前后的基因组为模板,PCR扩增16S rDNA的V2-V3区,DGGE分析,切取SECC样本的特异条带进行克隆、测序、核酸序列比对.结果:基因组在放大前后DGGE图谱一致,SECC组样本的条带数多于CF组的条带数.切取的SECC样本的4 条特异条带测序后证实为3 种未培养微生物和嗜沫嗜血杆菌.结论:利用DGGE技术发现了SECC儿童菌斑与CF儿童菌斑细菌组成的差异,并在SECC样本中发现了区别于CF样本的未培养微生物,这些微生物在致龋过程中发挥的作用还有待研究.%Objective: To analyze the differences of dental plaque microbial composition between caries-free(CF) and severe early childhood caries (SECC) children analysed by denaturing gradient gel electrophoresis (PCR-DGGE). Methods: Genomic DNA was extracted from the dental plaque of CF group (n = 34 ) and SECC group( n = 34) respectively. Each DNA sample of CF and SECC group was mixed equally to construct CF genomic DNA pool and SECC genomic DNA pool, and then the two genomic DNA pools were used as template and amplified by DNA Amplification Kit. V2-V3 region of the 16S rDNA was amplified by PCR and analyzed by DGGE. Four bands in DGGE profile specifically existed in SECC lanes were excised, cloned, sequenced and searched in BLAST to find the closest relatives. Results: After whole genome amplification, DGGE showed the same DNA profile as that without amplification profile, and the bands of SECC lanes were more than those of CF lanes. Three uncultured bacteria and Aggregatibacter aphrophilus were found in SECC group. Conclusion

  6. 发酵乳活性双歧杆菌种类快速识别及定量研究%Isolation of Bifidobacteria from Yoghurts and Assessment of the Bifidobacterial Population by PCR-Denaturing Gradient Gel Electrophoresis and Quantitative Real-Time PCR

    Institute of Scientific and Technical Information of China (English)

    王立平; 刘晓莉; 蔡雪凤; 曹悦; 张帆

    2013-01-01

    In this article, a rapid identification and numberation method for bifidobacterium in fermented milk products was established. Polymerase Chain Reaction-Denaturing Gradient Gel Electrophoresis (PCR-DGGE) technique was used to identify Bifidobacterium present in commercial probiotic yoghurts.Real-Time Polymerase Chain Reaction (Real-Time PCR) technique was used to detect Bifidobacteria. The results showed that PCR-DGGE assays enabled identification of the species of bifidobacterium initially present in commercial fermented milk products with a detection threshold of 105 cells per gram. Real-Time PCR assays enabled numbering Bifidobacterium with a detection threshold of 104 cells per gram of product. Established PCR-DGGE analysis method was suitable for identification and detection of Bifidobacterium in commercial probiotic yoghurts.%本文建立了发酵乳制品中双歧杆菌种类和数量快速测定方法.以发酵乳中双歧杆菌为靶标,采用PCR-DGGE技术快速识别双歧杆菌种类;采用Real-Tine PCR手段测定双歧杆菌数量.研究结果表明,PCR-DGGE能准确、快速鉴别双歧杆菌种类,检出限为105 CFU/g; Real-Tine PCR能准确、快速定量双歧杆菌,检出限为104 CFU/g.该方法可用于发酵乳中双歧杆菌的快速识别和定量.

  7. Polyacrylamide Gel Preparation and Properties of La0.7Ca0.3MnO3 Nanoparticles%La0.7Ca0.3MnO3纳米颗粒的聚丙烯酰胺凝胶法制备及性能

    Institute of Scientific and Technical Information of China (English)

    朱彦虎; 杨华; 县涛; 魏智强; 马金元; 冯旺军

    2011-01-01

    采用聚丙烯酰胺凝胶法制备La0.7Ca0.3MnO3纳米颗粒。通过X射线衍射和扫描电镜研究不同pH值和不同络合剂对制备La0.7Ca0.3MnO3纳米颗粒形貌及性能的影响。结果表明:合成La0.7Ca0.3MnO3钙钛矿相的最佳pH=2;分别以醋酸、草酸、柠檬酸和乙二胺四乙酸为络合剂,在600℃煅烧温度制备的高纯La0.7Ca0.3MnO3纳米颗粒的形貌规整、呈类球形,尺寸分布较窄,平均粒径分别为30、37、45、50nm;以酒石酸为络合剂,在900℃煅烧制备的单相La0.7Ca0.3MnO3颗粒的形貌不甚规整,颗粒间出现明显的煅烧粘连现象,颗粒尺寸较大,平均粒径为170nm。磁滞回线测量结果表明:制得的La0.7Ca0.3MnO3颗粒在室温均表现为顺磁性,其顺磁磁化率随样品颗粒尺寸的减小而减小。%A polyacrylamide gel route was used to fabricate La0.7Ca0.3MnO3 nanoparticles. Influence of pH values and chelating agents on particle morphology and properties of the prepared La0.7Ca0.3MnO3 nanoparticles were investigated by X-ray diffraction and scanning electron microscopy. The optimal pH value for synthesis of perovskite La0.7Ca0.3MnO3 was obtained to be 2. Using acetic acid, oxalic acid, citric acid, and ethylenediamine-tetraacetic acid as chelating agents, single-phase La0.7Ca0.3MnO3 nanoparticles were prepared at a calcination temperature of 600 ℃. The particles prepared using the four chelating agents are uniform and regu- larly-shaped spheres, and have an average size of 30, 37, 45 nm, and 50 nm, respectively. When using tartaric acid as the chelating agent, a much higher calcination temperature at 900 ℃ was required to produce single-phase La0.7Ca0.3MnO3 particles. The obtained particles exhibit an irregular morphology and an obvious adhesive behavior, and have a relatively large average size of 170 nm. Mag- netic hysteresis loop measurements indicate that the prepared La0.7Ca0.3MnO3 samples exhibit paramagnetism at room temperature, but their

  8. Mapping of Chlamydia trachomatis proteins by immobiline-polyacrylamide two-dimensional electrophoresis: spot identification by N-terminal sequencing and immunoblotting

    DEFF Research Database (Denmark)

    Bini, L; Sanchez-Campillo, M; Santucci, A;

    1996-01-01

    Proteins from purified elementary bodies of Chlamydia trachomatis were separated by two-dimensional gel electrophoresis on nonlinear wide-range immobilized pH gradients in the first dimension and polyacrylamide gradient gels in the second dimension. The maps obtained with this system are highly...

  9. Use of denaturing gradient gel electrophoresis in screening ...

    African Journals Online (AJOL)

    G. Christopoulos

    2012-08-24

    Aug 24, 2012 ... prevention of the disease through prenatal diagnosis programs. To date, the molecular ... intermedia and 21 b-thalassemia major), one obligate b-thalas- ..... genotyping of beta-thalassemia mutations using DGGE analysis:.

  10. Evaluation of denaturing gradient gel electrophoresis (DGGE) used ...

    African Journals Online (AJOL)

    user1

    2012-08-08

    Aug 8, 2012 ... systems. Therefore, three primer pairs that amplify different variable regions of the 16S rRNA gene (V1, ... ones include the selection of the primer system, the gradient .... some of the strains with sequence heterogeneity in the.

  11. Semi-interpenetrating polymer networks based on polyacrylamide and poly[itaconic acid

    Directory of Open Access Journals (Sweden)

    Kalagasidis-Krušić Melina T.

    2003-01-01

    Full Text Available The effect of pH and temperature on the equilibrium swelling properties of PIA/PAAm semi-IPNs were investigated.Semi-IPNs based on polyacrylamide (PAAm and poly(itaconic acid (PIA were prepared by two different techniques, by polymerizing itaconic acid in the presence of polyacrylamide gel (Semi-IPNs-l and by making the polyacrylamide gel in the presence of previously synthesized poly(itaconic acid (Semi-IPNs-ll, with different PIA/PAAm mass ratios. The equilibrium swelling degree of an ionic network depends very much on the concentration of ionisable groups. The addition of a small amount of itaconic acid dramatically changes the swelling behavior of PAAm. Increase of the ionic monomer (IA produces swelling degrees that increase to a high extent when the pH of the buffer solution is higher than the nominal pKa values of the acid groups. Gels with higher IA content swell less than PAAm gels in low pH buffers. At low pH, when complexation due to hydrogen bonding occurs between the carboxylic groups and amide groups of acrylamide, the polymer network collapses and the swelling ratio is low. The presence of hydrogen bonds in the complexes causes additional constraints in the network, acting as a physical crosslinking and makes the network less hydrophilic, because the carboxylic groups on the PIA are occupied in the complexes. As opposed to this, the equilibrium swelling degrees change very little with pH of the solution in nonionic PAAm gel.Hydrogels exhibit continuous changes in water content as a function of temperature. The swelling degree increases with increasing temperature due to gel expansion upon warming.

  12. 21 CFR 172.255 - Polyacrylamide.

    Science.gov (United States)

    2010-04-01

    ... percent of acrylamide monomer may be safely used as a film former in the imprinting of soft-shell gelatin... 21 Food and Drugs 3 2010-04-01 2009-04-01 true Polyacrylamide. 172.255 Section 172.255 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) FOOD FOR...

  13. Polyacrylamide Hydrogel Properties for Horticultural Applications

    Science.gov (United States)

    Polyacrylamide (PAAm) hydrogels are commonly employed to ensure hydration of the growth media and minimize crop losses during the crop production and postproduction phases in horticulture. However, studies of the effect of these materials have shown that they have a minimal effect on crop life and q...

  14. Enhanced oil recovery using novel polyacrylamides

    NARCIS (Netherlands)

    Broekhuis, Antonius Augustinus; Picchionni, Francesco; Zacarias, Diego Armando

    2013-01-01

    The invention relates to a polymer comprising a central structure to which n branches are covalently attached, wherein n is an integer from 1 to 50, wherein the branches comprise polyacrylamide moieties, wherein the theoretical number average molecular weight of the polymer is at least 100,000g/mol.

  15. Superoxide dismutase isozyme detection using two-dimensional gel electrophoresis zymograms.

    Science.gov (United States)

    Niyomploy, Ploypat; Srisomsap, Chantragan; Chokchaichamnankit, Daranee; Vinayavekhin, Nawaporn; Karnchanatat, Aphichart; Sangvanich, Polkit

    2014-03-01

    Superoxide dismutases (SODs) are ubiquitous antioxidant enzymes involved in cell protection from reactive oxygen species. Their antioxidant activities make them of interest to applied biotechnology industries and are usually sourced from plants. SODs are also involved in stress signaling responses in plants, and can be used as indicators of these responses. In this article, a suitable method for the separation of different SOD isoforms using two-dimensional-gel electrophoresis (2D-GE) zymograms is reported. The method was developed with a SOD standard from bovine erythrocytes and later applied to extracts from Stemona tuberosa. The first (non-denaturing isoelectric focusing) and second (denaturing sodium dodecylsulphate-polyacrylamide gel electrophoresis) dimensions of duplicate 2D-GE gels were stained with either Coomassie brilliant blue G-250 for total protein visualization, or SOD activity (zymogram) using riboflavin/nitroblue tetrazolium. For confirmation, putative SOD activity positive spots were subject to trypsin digestion and nano-liquid chromatography tandem mass spectrometry, followed by searching the MASCOT database for potential identification. The method could separate different SOD isoforms from a plant extract and at least partially maintain or allow renaturation to the native forms of the enzyme. Peptide sequencing of the 2D-GE suggested that the SODs were resolved correctly, identifying the control CuZn-SOD from bovine erythrocytes. The two SODs from S. tuberosa tubers were found to be likely homologous of a CuZn-SOD. SOD detection and isoform separation by 2D-GE zymograms was efficient and reliable. The method is likely applicable to SOD detection from plants or other organisms. Moreover, a similar approach could be developed for detection of other important enzymes in the future. Copyright © 2013 Elsevier B.V. All rights reserved.

  16. Single DNA denaturation and bubble dynamics

    Energy Technology Data Exchange (ETDEWEB)

    Metzler, Ralf [Physics Department, Technical University of Munich, James Franck Strasse, 85747 Garching (Germany); Ambjoernsson, Tobias [Chemistry Department, Massachusetts Institute of Technology, 77 Massachusetts Avenue, Cambridge, MA 02139 (United States); Hanke, Andreas [Department of Physics and Astronomy, University of Texas, 80 Fort Brown, Brownsville (United States); Fogedby, Hans C [Department of Physics and Astronomy, University of Arhus, Ny Munkegade, 8000 Arhus C (Denmark)], E-mail: metz@ph.tum.de

    2009-01-21

    While the Watson-Crick double-strand is the thermodynamically stable state of DNA in a wide range of temperature and salt conditions, even at physiological conditions local denaturation bubbles may open up spontaneously due to thermal activation. By raising the ambient temperature, titration, or by external forces in single molecule setups bubbles proliferate until full denaturation of the DNA occurs. Based on the Poland-Scheraga model we investigate both the equilibrium transition of DNA denaturation and the dynamics of the denaturation bubbles with respect to recent single DNA chain experiments for situations below, at, and above the denaturation transition. We also propose a new single molecule setup based on DNA constructs with two bubble zones to measure the bubble coalescence and extract the physical parameters relevant to DNA breathing. Finally we consider the interplay between denaturation bubbles and selectively single-stranded DNA binding proteins.

  17. Single DNA denaturation and bubble dynamics

    DEFF Research Database (Denmark)

    Metzler, Ralf; Ambjörnsson, Tobias; Hanke, Andreas

    2009-01-01

    for situations below, at, and above the denaturation transition. We also propose a new single molecule setup based on DNA constructs with two bubble zones to measure the bubble coalescence and extract the physical parameters relevant to DNA breathing. Finally we consider the interplay between denaturation......While the Watson-Crick double-strand is the thermodynamically stable state of DNA in a wide range of temperature and salt conditions, even at physiological conditions local denaturation bubbles may open up spontaneously due to thermal activation. By raising the ambient temperature, titration......, or by external forces in single molecule setups bubbles proliferate until full denaturation of the DNA occurs. Based on the Poland-Scheraga model we investigate both the equilibrium transition of DNA denaturation and the dynamics of the denaturation bubbles with respect to recent single DNA chain experiments...

  18. The dynamics of the DNA denaturation transition

    CERN Document Server

    van Erp, Titus S

    2012-01-01

    The dynamics of the DNA denaturation is studied using the Peyrard-Bishop-Dauxois model. The denaturation rate of double stranded polymers decreases exponentially as function of length below the denaturation temperature. Above Tc, the rate shows a minimum, but then increases as function of length. We also examine the influence of sequence and solvent friction. Molecules having the same number of weak and strong base-pairs can have significantly different opening rates depending on the order of base-pairs.

  19. A method for easily customizable gradient gel electrophoresis.

    Science.gov (United States)

    Miller, Andrew J; Roman, Brandon; Norstrom, Eric

    2016-09-15

    Gradient polyacrylamide gel electrophoresis is a powerful tool for the resolution of polypeptides by relative mobility. Here, we present a simplified method for generating polyacrylamide gradient gels for routine analysis without the need for specialized mixing equipment. The method allows for easily customizable gradients which can be optimized for specific polypeptide resolution requirements. Moreover, the method eliminates the possibility of buffer cross contamination in mixing equipment, and the time and resources saved with this method in place of traditional gradient mixing, or the purchase of pre-cast gels, are noteworthy given the frequency with which many labs use gradient gel SDS-PAGE. Copyright © 2016 Elsevier Inc. All rights reserved.

  20. A pH-sensitive Modified Polyacrylamide Hydrogel

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    A pH-sensitive modified polyacrylamide hydrogel was prepared by two steps and the modified polyacrylamide was characterized by 1HNMR spectrum. The surface morphology and swelling behavior of the hydrogels were investigated.

  1. Bubbles and denaturation in DNA

    CERN Document Server

    Van Erp, T S; Peyrard, M; Erp, Titus S. van; Cuesta-Lopez, Santiago; Peyrard, Michel

    2006-01-01

    The local opening of DNA is an intriguing phenomenon from a statistical physics point of view, but is also essential for its biological function. For instance, the transcription and replication of our genetic code can not take place without the unwinding of the DNA double helix. Although these biological processes are driven by proteins, there might well be a relation between these biological openings and the spontaneous bubble formation due to thermal fluctuations. Mesoscopic models, like the Peyrard-Bishop-Dauxois model, have fairly accurately reproduced some experimental denaturation curves and the sharp phase transition in the thermodynamic limit. It is, hence, tempting to see whether these models could be used to predict the biological activity of DNA. In a previous study, we introduced a method that allows to obtain very accurate results on this subject, which showed that some previous claims in this direction, based on molecular dynamics studies, were premature. This could either imply that the present...

  2. DNA denaturation in ionic solution

    Science.gov (United States)

    Maity, Arghya; Singh, Amar; Singh, Navin

    2016-05-01

    Salt or cations, present in solution play an important role in DNA denaturation and folding kinetics of DNA helix. In this work we study the thermal melting of double stranded DNA (dsDNA) molecule using Peyrard Bishop Dauxois (PBD) model. We modify the potential of H-bonding between the bases of the complimentary strands to introduce the salt and solvent effect. We choose different DNA sequences having different contents of GC pairs and calculate the melting temperatures. The melting temperature increases logarithmically with the salt concentration of the solution. The more GC base pairs in the chain enhance the stability of DNA chain at a fix salt concentration. The obtained results are in good accordance with experimental findings.

  3. Polyacrylamide polymers derived from acrylonitrile without intermediate isolation

    Energy Technology Data Exchange (ETDEWEB)

    Norton, C.J.; Falk, D.O.

    1977-04-05

    Hydrolyzed and neutralized acrylonitrile is polymerized in solution without isolation to produce a high molecular weight polyacrylamide useful for mobility control in secondary recovery of petroleum. The polyacrylamide optionally may be hydrolyzed, methylolated, and sulfomethylated to further enhance its water-thickening properties. This procedure reduces the cost of making polyacrylamide. (5 claims)

  4. 21 CFR 173.10 - Modified polyacrylamide resin.

    Science.gov (United States)

    2010-04-01

    ... 21 Food and Drugs 3 2010-04-01 2009-04-01 true Modified polyacrylamide resin. 173.10 Section 173... CONSUMPTION Polymer Substances and Polymer Adjuvants for Food Treatment § 173.10 Modified polyacrylamide resin. Modified polyacrylamide resin may be safely used in food in accordance with the following...

  5. 利用PCR-DGGE技术监测Lactobacillus plantarum WCFS1对小鼠肠道菌群的动态变化%Monitoring dynamic changes of intestinal microflora in mice by Lactobacillus plantarum WCFS1 using PCR-denaturing gradient gel electrophoresis

    Institute of Scientific and Technical Information of China (English)

    吴青龙; 陈廷涛; 熊顺强; 姜淑英; 李胜杰; 魏华

    2011-01-01

    利用传统培养法和变性梯度凝胶电泳技术(PCR-DGGE)评价Lactobacillus plantarum WCFS1对小鼠肠道微生物数量和种类的影响.结果表明,乳酸杆菌数量在灌胃期和恢复期较空白期显著升高(P<0.05),而肠杆菌数量在灌胃期和恢复期较空白期显著下降(P<0.01);DGGE图谱及多样性分析显示小鼠灌胃期间菌群多样性显著提高(P<0.01),但在停止灌胃7d后与空白期相比无显著性差异(P>0.05).此外,DGGE条带测序表明Lactobacillus plantarum WCFS1在灌胃期间可在大部分小鼠肠道内占据优势地位.%Both traditional culture-dependent method and PCR-Denaturing Gradient Gel Electrophoresis (PCR-DGGE) were used to evaluate the effects of Lactobacillus plantarum WCFS1 on the amount and species of microflora in mice intestinal tract. It is observed that the amount of Lactobacillus in mice intestine during the feeding and recovery stages significantly increased (P<0. 05) when compared with the control stage; while the amount of Enterococcus decreased (P<0. 01). The DGGE pattern and diversity analysis indicated that the diversity of microflora at the feeding stage had great increase (P

  6. Complex coacervation between colloidal silica and polyacrylamide

    Energy Technology Data Exchange (ETDEWEB)

    Kawase, Kaoru; Sakami, Hiroshi; Hayakawa, Kiyoshi

    1989-03-01

    Complex coacervation introduced by gamma-ray induced polymerization of acrylamide in colloidal silica was studied. The complex coaservate was formed by polymerization of acrylamide dissolved in a colloidal silica and methanol mixture. Complex coacervation (two-phase separation of the mixture) was observed only when the concentration of methanol was between 33 and 41 percent by volume, and the concentration of colloidal silica did not affect it. Although two phase separation was not influenced by pH change, the content of polyacrylamide was bigger in the equilibrated solution in acidic regions. It was, however, bigger in the complex coacervate at neutral and in alkaline regions. The content of polyacrylamide was also calculated from the particle diameter of complex coacervate measured by small angle X-ray scattering, and the result was well coincided with the analytical result. The stability of the complex coacervate against the addition of salts was better than that of the untreated colloidal silica. The rate of electrophoretic transport of the complex coacervate was also lower than that of the colloidal silica. From these observation it was concluded that the hydrophobic colloidal silica particles were protected by the surrounding hydrophilic polyacrylamide. (author).

  7. 聚丙烯酰胺凝胶法合成TbFeO3纳米颗粒及其可见光催化活性%Polyacrylamide Gel Synthesis and Visible-light Photocatalytic Activity of TbFeO3 Nanoparticles

    Institute of Scientific and Technical Information of China (English)

    林贯军; 杨华; 县涛

    2012-01-01

    采用聚丙烯酰胺凝胶法制备了TbFeO3纳米颗粒,研究了不同络合剂对样品的纯度、颗粒尺寸及形貌的影响.XRD分析结果表明,以酒石酸、柠檬酸或乙二胺四乙酸(EDTA)为络合剂,在650℃下烧结均可制备出单相TbFeO3纳米颗粒,但产物的平均粒径不同;而采用乙酸或草酸为络合剂则难以制得纯相样品.SEM观测结果表明,以酒石酸为络合剂制备的颗粒细小、均匀、形貌规整、呈球状,平均粒径约为50 nm;以柠檬酸为络合剂制备的颗粒主要以近球形为主,颗粒的尺寸分布相对较宽,平均粒径约为100 nm;以EDTA 为络合剂制备的颗粒主要呈椭球状,颗粒尺寸较均匀,但颗粒间存在不同程度的黏连现象,平均粒径约为110 nm.这3种样品的BET比表面积分别为15.4,8.3和6.8 m2/g.紫外-可见漫反射吸收光谱研究表明,TbFeO3纳米颗粒的带隙为1.95 ~1.98 eV.分别以甲基橙(MO)、罗丹明B(RhB)、亚甲基蓝(MB)、酸性品红(AF)和刚果红(CR)5种有机染料为目标降解物,考察了TbFeO3颗粒的光催化活性.结果表明,在可见光辐照下颗粒表现出良好的光催化活性,其中,以酒石酸为络合剂制备的样品光催化效果最好.%A polyacrylamide gel method was used to prepare TbFeO3 nanoparticles, and the influences of dif-fe-rent chelating agents on the phase purity, particle size and morphology were investigated. X-ray diffraction ( XRD) results indicate that single-phase TbFeO3 particles can be prepared at a calcination temperature of 650 ℃ using tartaric acid, citric acid or ethylenediamine-tetraacetic acid(EDTA) as the chelating agent, but the average grain size has a dependence on the choice of the chelating agent. The use of acetic acid or oxalic acid as the chelating agent is difficult to achieve the synthesis of single-phase TbFeO3 samples. Scanning electron microscope (SEM) observations reveal that the particles prepared using tartaric acid as the chelating agent

  8. 应用DGGE技术分析自然免耕土与普通耕作土细菌群落的多样性%Analysis of bacterial communities in no-tillage and tillage soils by denaturing gradient gel electrophoresis

    Institute of Scientific and Technical Information of China (English)

    马振刚; 马淑华; 张时恒; 贾俊杰; 李田; 潘国庆; 周泽扬

    2011-01-01

    为探明免耕土壤与普通耕作土壤环境中细菌群落的多样性,获取相关优势菌落信息,该研究利用宏基因组学方法获得免耕土和普通耕作土样品总DNA,利用PCR获得16SrDNAV3片段,并进行变性梯度凝胶电泳(Denaturation gradient gel electrophoresis),通过微生物种群丰富度比较两样品中群落的丰富性,同时选取相关DNA条带进行克隆、测序和生物信息学分析.结果显示:免耕土壤中细菌群落多样性更加丰富;两类型土壤样品间细菌群落组成具有显著差异,证明耕作制度影响了土壤细菌群落结构.BLAST分析与系统发生分析结果表明,免耕土壤中特异存在的细菌群落与具有生物固氮、降解甲苯和倍硫磷等特性的细菌序列相似性较高或进化关系较近,推测其在免耕土壤肥力、有毒物质降解及有机质转变等过程中起作用.%Bacterial population diversity in no-tillage and tillage soils were analyzed by PCR-denaturing gradient gel electrophoresis (DGGE) to get the information on dominant bacteria. The metagenomics technologies were employed to obtain the total DNA, then 16S rDNA V3 region was amplified by PCR. Soil microbial richness was analyzed by 16S rDNA V3-PCR-DGGE. Technologies of cloning and sequencing combined with a phylogeny tree were applied to analyze the evolutional relationship of genes corresponding to several interesting bands involved in this research. The results of DGGE analysis showed that the bacterial population diversity of no-tillage soil was more abundant than that of tillage soils and the UPGMA cluster a-nalysis exhibited a significantly different community composition between no-tillage and tillage soils, demonstrating that the soil tillage system had a remarkable effect on the bacterial communities. Phylogenetic analysis exhibited the distinctive popu-lations specific in no-tillage soil were close to the bacterium chracterized by azotification, methylbenzene and baycid

  9. CURRENT COLLOIDAL DISPERSION GELS ARE NOT SUPERIOR TO POLYMER FLOODING

    Institute of Scientific and Technical Information of China (English)

    Seright Randy; Han Peihui; Wang Dongmei

    2006-01-01

    The suggestion that the colloidal-dispersion-gel (CDG) process is superior to normal polymer flooding is misleading and generally incorrect. Colloidal dispersion gels, in their present state of technological development, should not be advocated as an improvement to, or substitute for, polymer flooding. Gels made from aluminum-citrate crosslinked polyacrylamides can act as conventional gels and provide effective conformance improvement in treating some types of excess water production problems if sound scientific and engineering principles are respected.

  10. Formation of composite polyacrylamide and silicone substrates for independent control of stiffness and strain.

    Science.gov (United States)

    Simmons, Chelsey S; Ribeiro, Alexandre J S; Pruitt, Beth L

    2013-02-21

    Cells that line major tissues in the body such as blood vessels, lungs and gastrointestinal tract experience deformation from mechanical strain with our heartbeat, breathing, and other daily activities. Tissues also remodel in both development and disease, changing their mechanical properties. Taken together, cells can experience vastly different mechanical cues resulting from the combination of these interdependent stimuli. To date, most studies of cellular mechanotransduction have been limited to assays in which variations in substrate stiffness and strain were not combined. Here, we address this technological gap by implementing a method that can simultaneously tune both substrate stiffness and mechanical strain. Substrate stiffness is controlled with different monomer and crosslinker ratios during polyacrylamide gel polymerization, and strain is transferred from the underlying silicone platform when stretched. We demonstrate this platform with polyacrylamide gels with elastic moduli at 6 kPa and 20 kPa in combination with two different silicone formulations. The gels remain attached with up to 50% applied strains. To validate strain transfer through the gels into cells, we employ particle-tracking methods and observe strain transmission via cell morphological changes.

  11. Nano-hydroxyapatite/polyacrylamide composite hydrogels with high mechanical strengths and cell adhesion properties.

    Science.gov (United States)

    Li, Zhiyong; Mi, Wenying; Wang, Huiliang; Su, Yunlan; He, Changcheng

    2014-11-01

    Nano-hydroxyapatite/polyacrylamide composite hydrogels were successfully fabricated by physically mixing nano-hydroxyapatite (nHAp) particles into a peroxidized micelles initiated and cross-linked (pMIC) polyacrylamide (PAAm) hydrogel. The nanocomposite hydrogels exhibited excellent mechanical properties. The fracture tensile stresses of the gels were in the range of 0.21-0.86 MPa and the fracture tensile strains were up to 30 mm/mm, and the compressive strengths were up to 35.8 MPa. Meanwhile the introduction of nHAp endowed the composite hydrogels with good cell adhesion properties. This nHAp/PAAm nanocomposite hydrogel is expected to find potential applications in tissue engineering.

  12. Gel-based nonradioactive single-strand conformational polymorphism and mutation detection: limitations and solutions.

    Science.gov (United States)

    Gupta, Vibhuti; Arora, Reetakshi; Gochhait, Sailesh; Bairwa, Narendra K; Bamezai, Rameshwar N K

    2014-01-01

    Single-strand conformation polymorphism (SSCP) for screening mutations/single-nucleotide polymorphisms (SNPs) is a simple, cost-effective technique, saving an expensive exercise of sequencing each and every polymerase chain reaction product and assisting in choosing only the amplicons of interest with expected mutations. The principle of detection of small changes in DNA sequences is based on changes in single-strand DNA conformations. The changes in electrophoretic mobility that SSCP detects are sequence dependent. The limitations faced in SSCP range from routine polyacrylamide gel electrophoresis (PAGE) problems to the problems of resolving mutant DNA bands. Both these problems can be solved by controlling PAGE conditions and by varying physical and environmental conditions such as pH, temperature, voltage, gel type and percentage, addition of additives or denaturants, and others. Despite much upgrading of the technology for mutation detection, SSCP remains the method of choice to analyze mutations and SNPs in order to understand genomic variations, both spontaneous and induced, and the genetic basis of diseases.

  13. Conducting polymer electrodes for gel electrophoresis.

    Science.gov (United States)

    Bengtsson, Katarina; Nilsson, Sara; Robinson, Nathaniel D

    2014-01-01

    In nearly all cases, electrophoresis in gels is driven via the electrolysis of water at the electrodes, where the process consumes water and produces electrochemical by-products. We have previously demonstrated that π-conjugated polymers such as poly(3,4-ethylenedioxythiophene) (PEDOT) can be placed between traditional metal electrodes and an electrolyte to mitigate electrolysis in liquid (capillary electroosmosis/electrophoresis) systems. In this report, we extend our previous result to gel electrophoresis, and show that electrodes containing PEDOT can be used with a commercial polyacrylamide gel electrophoresis system with minimal impact to the resulting gel image or the ionic transport measured during a separation.

  14. Conducting polymer electrodes for gel electrophoresis.

    Directory of Open Access Journals (Sweden)

    Katarina Bengtsson

    Full Text Available In nearly all cases, electrophoresis in gels is driven via the electrolysis of water at the electrodes, where the process consumes water and produces electrochemical by-products. We have previously demonstrated that π-conjugated polymers such as poly(3,4-ethylenedioxythiophene (PEDOT can be placed between traditional metal electrodes and an electrolyte to mitigate electrolysis in liquid (capillary electroosmosis/electrophoresis systems. In this report, we extend our previous result to gel electrophoresis, and show that electrodes containing PEDOT can be used with a commercial polyacrylamide gel electrophoresis system with minimal impact to the resulting gel image or the ionic transport measured during a separation.

  15. Novel hydrophobically associative polyacrylamide with tunable viscosity

    Institute of Scientific and Technical Information of China (English)

    Xu Feng Zhang; Wen Hui Wu

    2009-01-01

    Hydrophobically associative polyacrylamide (HAPAM) were prepared in aqueous solution by radical copolymerization of novel cationic surface-active monomer, dimethylhexadecyl(3-acrylamidopropyl)ammonium bromide (DMHAB), with acrylarnide (AM) in the presence of DMHAB/CTAB mixed micelles. The length of hydrophobic microblock (N_H) in HAPAM is controlled by the molar fraction of DMHAB in mixed micelles, which can be mediated by the ratio of CTAB to DMHAB. The results of steady-state fluorescence probe and viscometry experiments showed the ability of HAPAM association was determined by the length of the hydrophobic microblock. HAPAM with tunable association ability are promising materials for thickening agent.

  16. Radio-synthesized polyacrylamide hydrogels for proteins release

    Science.gov (United States)

    Ferraz, Caroline C.; Varca, Gustavo H. C.; Lopes, Patricia S.; Mathor, Monica B.; Lugão, Ademar B.

    2014-01-01

    The use of hydrogels for biomedical purposes has been extensively investigated. Pharmaceutical proteins correspond to highly active substances which may be applied for distinct purposes. This work concerns the development of radio-synthesized hydrogel for protein release, using papain and bovine serum albumin as model proteins. The polymer was solubilized (1% w/v) in water and lyophilized. The proteins were incorporated into the lyophilized polymer and the hydrogels were produced by simultaneous crosslinking and sterilization using γ-radiation under frozen conditions. The produced systems were characterized in terms of swelling degree, gel fraction, crosslinking density and evaluated according to protein release, bioactivity and cytotoxicity. The hydrogels developed presented different properties as a function of polymer concentration and the optimized results were found for the samples containing 4-5% (w/v) polyacrylamide. Protein release was controlled by the electrostatic affinity of acrylic moieties and proteins. This selection was based on the release of the proteins during the experiment period (up to 50 h), maintenance of enzyme activity and the nanostructure developed. The system was suitable for protein loading and release and according to the cytotoxic assay it was also adequate for biomedical purposes, however this method was not able to generate a matrix with controlled pore sizes.

  17. Mapping and identification of interferon gamma-regulated HeLa cell proteins separated by immobilized pH gradient two-dimensional gel electrophoresis

    DEFF Research Database (Denmark)

    Shaw, AC; Rossel Larsen, M; Roepstorff, P

    1999-01-01

    magnitude of IFN-gamma responsive genes has been reported previously. Our goal is to identify and map IFN-gamma-regulated HeLa cell proteins to the two-dimensional polyacrylamide gel electrophoresis with the immobilized pH gradient (IPG) two-dimensional polyacrylamide gel electrophoresis (2-D PAGE) system...

  18. Rheological Properties of Hydrophobically Associating Polyacrylamide Solution

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    A temperature-resistant, salt-tolerant polyacrylamide, hydrophobically associating polymer (HAP), was synthesized in the State Key Laboratory of Heavy Oil Processing. The rheological behavior of HAP solution was investigated by means of flow experiments in porous media and by using a HAAKE RS600 rheometer. The results of Nuclepore membrane filtration showed that filtration time increased sharply when the critical association concentration was reached. Shear rate had a greater impact on viscosity and shear stress with increasing HAP concentration. The HAP solution with a concentration of 100 mg/L (salinity 32,868 mg/L) exhibited negative thixotropy. However, at the same salinity the HAP solution showed thixotropy and its viscosity became greater when the polymer concentration increased to 1,500 mg/L.The flow experiments in cemented core samples indicated that the resistance factor and residual resistance factor of the HAP solution were 31.8 and 12 when polymer concentration and salinity were 1,500 mg/L, 32,868 mg/L at 85℃ respectively,which is favorable for flooding application. Such factors of partially hydrolyzed polyacrylamide 3530S were merely 3.14 and 1.71, so it could not be applied to polymer flooding in the oilfield with high temperature and high salinity.

  19. 变性梯度凝胶电泳分析氟化泡沫对牙菌斑微生物的影响%Analysis of the influence of fluoride foam on oral plaque microbiota by PCR-based denaturing gradient gel electrophoresis

    Institute of Scientific and Technical Information of China (English)

    翁金龙; 赵河川; 孙凤; 陈霄迟; 徐韬

    2013-01-01

    genomic DNA of the plaque was isolated.PCR was performed with a set of universal bacterial 16S rDNA primers.The PCR amplified 16S rDNA fragments were separated by denaturing gradient gel electrophoresis (DGGE).The relationship between microflora was assessed by comparing the PCR-DGGE fingerprinting profiles.Results The mean species richness of the bacterial population was 26.9 ± 2.9 at baseline,and fell to 20.1 ± 3.8 after three days of using fluoride foam.The difference was statistically significant (P < 0.05).The overall diversity of plaque samples as measured by the Shannon-Weiner index was 3.18 ±0.31 at baseline and 2.92 ± 0.28 at 3rd day.The difference was statistically significant (P =0.04 by the Mann-Whitney U test).The microbial diversity was changed after the fluoride application by Dice coefficient and clustering analysis.After one month,the microbial diversity was similar to the baseline.Conclusions The results suggest that 1.23% fluoride foam has obviously inhibiting effects on microbial diversity in dental plaque of children during a period of time.The microbial diversity and complexity of the microbial biota in dental plaque were changed after fluoride application and was able to return to original state.

  20. Stability of polyacrylamide solutions in presence of CO/sub 2/

    Energy Technology Data Exchange (ETDEWEB)

    Lakatos, I.J.; Lakatos-Szabo, J.

    1981-05-01

    The compatibility of polymer- and CO/sub 2/-containing systems is discussed. An extensive study was focused on the effect of polymer type, CO/sub 2/ pressure, salt content, crude oil, rock, and temperature on stability of polyacrylamides and their solutions in the presence of carbon dioxide. Two chemical/salt independent minimal and salt dependent/degradation effects and a salt effect caused by excess dissolution of rock constituents are differentiated within the deterioration phenomena. The joint application of polymers and silicates has resulted in a significant polymer conservation and an additional mobility control through partial gel formation. 26 references.

  1. Data of microstructure and mechanical properties of carbon foams derived from sucrose/polyacrylamide hydrogel

    Directory of Open Access Journals (Sweden)

    Yao Yao

    2016-06-01

    Full Text Available An easy method that combined gel casting and physical foaming was used to fabricate modified carbon foams. The design of carbon foams from sucrose/polyacrylamide hydrogel is a new concept for controlling the microstructure and improving the compressive properties of carbon foams. This article provides the micrographs obtained from optical and scanning electron microscope for foaming solution and carbon foams. Weight loss data used to construct the thermo-gravimetric curves are included. Load–displacement data constructing the stress–strain curves and the derived compressive properties are also included.

  2. Depressing effect of hydroxamic polyacrylamide on pyrite

    Institute of Scientific and Technical Information of China (English)

    张剑锋; 胡岳华; 王淀佐; 徐竞

    2004-01-01

    The performance of hydroxamic polyacrylamide(HPAM) in mineral flotation was tested on the samples of calcite, diaspore and pyrite. It is found that HPAM expresses intensive depression on pyrite and can be used as effective depressants for pyrite. The depression mechanism of HPAM to pyrite was investigated by the determination of contact angle, zeta potential, adsorptive capacity for collectors and infrared spectrum. A lower contact angle,more negative zeta potential, less xanthate adsorptive capacity, and the formation of chemical bonding were determined, which reveals that the strong chemical interactions exist between HPAM and pyrite surface. The group electronegativity of HPAM was calculated to explain the differences of interaction between reagent and minerals.

  3. Physico-chemical examination of the mechanism of displacement by polyacrylamide solutions. Part 1

    Energy Technology Data Exchange (ETDEWEB)

    Lakatos, I.; Lakatosne, J.S.

    1973-10-01

    The Research Laboratory for Petroleum Industry of the Hungarian Academy of Sciences has for several years dealt with basic research problems of polymer flooding. As a first part of a comprehensive study, the authors sum up their experimental results concerning the structure of thin polyacrylamide solutions. Changes of the structures of thin aqueous solutions are discussed as a function of polymer type and concentration, and temperature as well as of the concentration and quality of foreign electrolytes. Based upon data obtained, changes of dynamic viscosity and rheological properties of the solutions are attributed to the changes of the gel concentration characteristic of the solutions and of the equivalent clew diameter. As for the salt effect, it is underlined that by means of this a displacing phase of optimal structure (viscosity, gel concentration, clew diameter, etc.) can be ordinated to the pore structure of the reservoir. (23 refs.)

  4. 21 CFR 872.3420 - Carboxymethylcellulose sodium and cationic polyacrylamide polymer denture adhesive.

    Science.gov (United States)

    2010-04-01

    ... polyacrylamide polymer denture adhesive. 872.3420 Section 872.3420 Food and Drugs FOOD AND DRUG ADMINISTRATION....3420 Carboxymethylcellulose sodium and cationic polyacrylamide polymer denture adhesive. (a) Identification. A carboxymethylcellulose sodium and cationic polyacrylamide polymer denture adhesive is a...

  5. Spectrophotometric determination of substrate-borne polyacrylamide.

    Science.gov (United States)

    Lu, Jianhang; Wu, Laosheng

    2002-08-28

    Polyacrylamides (PAMs) have wide application in many industries and in agriculture. Scientific research and industrial applications manifested a need for a method that can quantify substrate-borne PAM. The N-bromination method (a PAM analytical technique based on N-bromination of amide groups and spectrophotometric determination of the formed starch-triiodide complex), which was originally developed for determining PAM in aqueous solutions, was modified to quantify substrate-borne PAM. In the modified method, the quantity of substrate-borne PAM was converted to a concentration of starch-triiodide complex in aqueous solution that was then measured by spectrophotometry. The method sensitivity varied with substrates due to sorption of reagents and reaction intermediates on the substrates. Therefore, separate calibration for each substrate was required. Results from PAM samples in sand, cellulose, organic matter burnt soils, and clay minerals showed that this method had good accuracy and reproducibility. The PAM recoveries ranged from 95.8% to 103.7%, and the relative standard deviations (n = 4) were <7.5% in all cases. The optimum range of PAM in each sample is 10-80 microg. The technique can serve as an effective tool in improving PAM application and facilitating PAM-related research.

  6. Chemical shift prediction for denatured proteins

    Energy Technology Data Exchange (ETDEWEB)

    Prestegard, James H., E-mail: jpresteg@ccrc.uga.edu; Sahu, Sarata C.; Nkari, Wendy K.; Morris, Laura C.; Live, David; Gruta, Christian

    2013-02-15

    While chemical shift prediction has played an important role in aspects of protein NMR that include identification of secondary structure, generation of torsion angle constraints for structure determination, and assignment of resonances in spectra of intrinsically disordered proteins, interest has arisen more recently in using it in alternate assignment strategies for crosspeaks in {sup 1}H-{sup 15}N HSQC spectra of sparsely labeled proteins. One such approach involves correlation of crosspeaks in the spectrum of the native protein with those observed in the spectrum of the denatured protein, followed by assignment of the peaks in the latter spectrum. As in the case of disordered proteins, predicted chemical shifts can aid in these assignments. Some previously developed empirical formulas for chemical shift prediction have depended on basis data sets of 20 pentapeptides. In each case the central residue was varied among the 20 amino common acids, with the flanking residues held constant throughout the given series. However, previous choices of solvent conditions and flanking residues make the parameters in these formulas less than ideal for general application to denatured proteins. Here, we report {sup 1}H and {sup 15}N shifts for a set of alanine based pentapeptides under the low pH urea denaturing conditions that are more appropriate for sparse label assignments. New parameters have been derived and a Perl script was created to facilitate comparison with other parameter sets. A small, but significant, improvement in shift predictions for denatured ubiquitin is demonstrated.

  7. Rousseau's Philosophy of Transformative, "Denaturing" Education

    Science.gov (United States)

    Riley, Patrick

    2011-01-01

    Rousseau's political philosophy presents the great legislator as a civic educator who must over time transform naturally self-loving egoists into citizens animated by a general will without destroying freedom. This is an educational process which is "denaturing" but which aims to produce autonomous adults who can ultimately say to their teacher…

  8. Instability of aqueous solutions of polyacrylamide in a hydrodynamic field

    Science.gov (United States)

    Makogon, B. P.; Bykova, E. N.; Bezrukova, M. A.; Klenin, S. I.; Ivanyuta, Yu. F.; Povkh, I. L.; Toryanik, A. I.

    1985-09-01

    This article discusses findings obtained regarding the effect of a hydrodynamic field on the reduced viscosity, effect of turbulent friction reduction, light scattering, double refraction, and optical density of aqueous solutions of hydrolyzed polyacrylamide.

  9. Agarose gel shift assay reveals that calreticulin favors substrates with a quaternary structure in solution

    DEFF Research Database (Denmark)

    Boelt, Sanne Grundvad; Houen, Gunnar; Højrup, Peter

    2015-01-01

    Here we present an agarose gel shift assay that, in contrast to other electrophoresis approaches, is loaded in the center of the gel. This allows proteins to migrate in either direction according to their isoelectric points. Therefore, the presented assay enables a direct visualization, separation...... structure. It is also demonstrated that the agarose gel shift assay is useful in the study of other protein interactions and can be used as an alternative method to native polyacrylamide gel electrophoresis....

  10. Petroleum recovery process utilizing formaldehyde-sulfite-reacted polyacrylamide

    Energy Technology Data Exchange (ETDEWEB)

    Norton, C.J.; Falk, D.O.

    1973-09-25

    Micellar slugs followed by thickened water floods were injected into Berea cores (20.4 percent porosity, 398.4 md permeability, see Patent 3,692,113 for pretreatment) for enhanced oil recovery. About 61.1 percent residual oil was produced when the polymer in the thickened water was sulfomethylated hydrolyzed polyacrylamide. However, use of the conventional unhydrolyzed polyacrylamide recovered only 27.7 percent residual oil.

  11. Guanidinium-induced denaturation by breaking of salt bridges

    NARCIS (Netherlands)

    Meuzelaar, H.; Panman, M.R.; Woutersen, S.

    2015-01-01

    Despite its wide use as a denaturant, the mechanism by which guanidinium (Gdm+) induces protein unfolding remains largely unclear. Herein, we show evidence that Gdm+ can induce denaturation by disrupting salt bridges that stabilize the folded conformation. We study the Gdm+-​induced denaturation of

  12. Hysteresis in pressure-driven DNA denaturation.

    Directory of Open Access Journals (Sweden)

    Enrique Hernández-Lemus

    Full Text Available In the past, a great deal of attention has been drawn to thermal driven denaturation processes. In recent years, however, the discovery of stress-induced denaturation, observed at the one-molecule level, has revealed new insights into the complex phenomena involved in the thermo-mechanics of DNA function. Understanding the effect of local pressure variations in DNA stability is thus an appealing topic. Such processes as cellular stress, dehydration, and changes in the ionic strength of the medium could explain local pressure changes that will affect the molecular mechanics of DNA and hence its stability. In this work, a theory that accounts for hysteresis in pressure-driven DNA denaturation is proposed. We here combine an irreversible thermodynamic approach with an equation of state based on the Poisson-Boltzmann cell model. The latter one provides a good description of the osmotic pressure over a wide range of DNA concentrations. The resulting theoretical framework predicts, in general, the process of denaturation and, in particular, hysteresis curves for a DNA sequence in terms of system parameters such as salt concentration, density of DNA molecules and temperature in addition to structural and configurational states of DNA. Furthermore, this formalism can be naturally extended to more complex situations, for example, in cases where the host medium is made up of asymmetric salts or in the description of the (helical-like charge distribution along the DNA molecule. Moreover, since this study incorporates the effect of pressure through a thermodynamic analysis, much of what is known from temperature-driven experiments will shed light on the pressure-induced melting issue.

  13. AN EXPERIMENTAL STUDY ON THE SOL/GEL PHASE TRANSITION OF LINEAR POLYMER IN THE PRESENCE OF CROSSLINKERS

    Institute of Scientific and Technical Information of China (English)

    HAN Ming; SHI Lianghe; YE Meiling; MULLER Guy

    1996-01-01

    The sol/gel phase diagrams were studied for two systems: polyacrylamide/Cr (Ⅲ) and polyacrylamide/glyoxal. Sol or gel phase could be distinguished according to the concentrations of polymer and crosslinker. The boundary polymer concentration did not depend on the types of gelation and decreased with increasing polymer dimension (molecular weight and conformation). The gelation, which is basically interchain bonding, requires the occurrence of entanglement. The overlap concentration is thus considered as the minimum polymer concentration required for gelation.

  14. Polyacrylamide Transport in Water Delivery Canals

    Science.gov (United States)

    Chen, L.; Zhu, J.; Young, M.

    2007-12-01

    Linear, anionic polyacrylamide (PAM) is being considered in the western United States as a technology to reduce seepage in unlined water delivery canals. A broad laboratory and field testing program has been undertaken to understand the benefits and potential environmental impacts of PAM use. The ability to predict the fate and transport of PAM in water delivery canals could prove to be a useful planning tool for PAM application. However, one key area of uncertainty of this type of canal treatment is the hydration, reaction, and settling rates of PAM after the dry powder is added to the canal water. In this study, we have developed a model that incorporates a number of known physical and chemical processes that can affect PAM transport, such as convection, dispersion, dissolution, flocculation, and settling, while solving the governing convection-dispersion transport equation. The model uses a mixed analytical and advanced numerical approach, and implements a transient partitioning of PAM mass between the canal water, the substrate soil, and potentially to open water bodies downstream of the application point. All source terms are modeled based on physical and chemical mechanisms as well as laboratory or field determined parameters. To more closely simulate field treatment of some canals, where PAM application moves upstream in time, the model is capable of implementing either a fixed or mobile upper boundary. In the latter treatment, the PAM can be added discretely or continuously in both time and space. A number of test situations have been simulated thus far, including theoretical and hypothetical cases for a wide range of conditions. The model also performed well when predicting PAM concentrations from a full-scale canal treatment experiment. The model provides a useful tool for predicting PAM fate and transport in water delivery canals, and therefore can play an important role in evaluating the efficacy of PAM application for water resources management

  15. DSC study of denaturation of β-lactoglobulin B

    Institute of Scientific and Technical Information of China (English)

    王邦宁; 谈夫

    1995-01-01

    The denaturation of bovine β-lactoglobulin B (β-Lg B) has been studied in phosphate solutions with various concentrations of GuHCl with differential scanning calorimetry The experiments demonstrated that the presence of GuHCl made the β-Lg B undergo both cold denaturation and heat denaturation under the condition of a high concentration of the protein. The enthalpy changes of both kinds of denaturation exhibit opposite signs. Both the cold denaturation and the renaturation of the protein are reproducible, but its heat denaturation is irreversible. The cooperation among monomer molecules of the protein is involved in its heat denaturation The heat denaturation of the protein can be represented by the thermodynamic model Nc D→F. The activation energy of heat denaturation is 285 kJ/mol, which imples that the depression of temperature and enthalpy of heat denaturation of the P-Lg B does not result from decreasing considerably the activation energy by GuHCl As for the cold denaturation of the protein, es

  16. Capillary gel electrophoresis of oligonucleotides: prediction of migration times using base-specific migration coefficients

    NARCIS (Netherlands)

    Cordier, Y.; Roch, O.; Cordier, P.; Bischoff, Rainer

    1994-01-01

    Chemically synthesized oligodeoxyribonucleotides were subjected to capillary gel electrophoresis on three different polyacrylamide-based matrices. Analysis of about 1000 samples over a 1-year period showed that the gel matrix evolved with time resulting in shifting migration times, making it

  17. Diced electrophoresis gel assay for screening enzymes with specified activities.

    Science.gov (United States)

    Komatsu, Toru; Hanaoka, Kenjiro; Adibekian, Alexander; Yoshioka, Kentaro; Terai, Takuya; Ueno, Tasuku; Kawaguchi, Mitsuyasu; Cravatt, Benjamin F; Nagano, Tetsuo

    2013-04-24

    We have established the diced electrophoresis gel (DEG) assay as a proteome-wide screening tool to identify enzymes with activities of interest using turnover-based fluorescent substrates. The method utilizes the combination of native polyacrylamide gel electrophoresis (PAGE) with a multiwell-plate-based fluorometric assay to find protein spots with the specified activity. By developing fluorescent substrates that mimic the structure of neutrophil chemoattractants, we could identify enzymes involved in metabolic inactivation of the chemoattractants.

  18. Low-temperature Magnesiothermic Synthesis of Mesoporous Silicon Carbide from an MCM-48/Polyacrylamide Nanocomposite Precursor

    Institute of Scientific and Technical Information of China (English)

    Zahra Saeedifar; Amir Abbas Nourbakhsh; Roozbeh Javad Kalbasi; Ebrahim Karamian

    2013-01-01

    Mesoporous silicon carbide with high specific surface area was successfully synthesized from an MCM-48/polyacrylamide nanocomposite precursor in the temperature range of 550-600 ℃ (below the melting point of Mg) by means of a magnesiothermic reduction process.The MCM-48/polyacrylamide precursor nanocomposite was prepared by in-situ polymerization of acrylamide monomer in the presence of mesoporous MCM-48 synthesized by sol-gel method.The physicochemical properties and microstructures of the nanocomposite precursor and the low-temperature SiC product were characterized by X-ray diffraction (XRD),differential scanning calorimetry-thermo gravimetric analysis (DSC-TGA),transmission electron microscopy (TEM) and N2 adsorption-desorption.TEM micrographs and Brunauer-Emmett-Teller (BET) gas adsorption studies showed that the SiC powder was nanocrystalline and had a specific surface area of 330 m2/g and a mesoporosity in the range of 2-10 nm.The presence of an exothermic peak in the DSC trace corresponds to the self-combustion process of the SiC magnesiothermic synthesis.The results also show that the carbon in excess to that required to produce SiC plays a role in the reduction of the SiO2.The mechanism of magnesiothermic synthesis of mesoporous SiC is discussed.

  19. Superior hybrid hydrogels of polyacrylamide enhanced by bacterial cellulose nanofiber clusters.

    Science.gov (United States)

    Yuan, Ningxiao; Xu, Lu; Zhang, Lu; Ye, Haowen; Zhao, Jianhao; Liu, Zhong; Rong, Jianhua

    2016-10-01

    Hybrid polyacrylamide/bacterial cellulose nanofiber clusters (PAM/BC) hydrogels with high strength, toughness and recoverability were synthesized by in situ polymerization of acrylamide monomer in BC nanofiber clusters suspension. The hybrid gels exhibited an extremely large elongation at break of 2200%, and a high fracture stress of 1.35MPa. Additionally, the original length of hydrogels could be recovered after releasing the tensile force. Compressive results showed that the PAM/BC hybrid gels could reach a strain of about 99% without break, and was able to completely recover its original shape immediately after releasing the compression force. The compressive stress at 99% reached as high as 30MPa. Nearly no hysteresis in cyclic compressive tests was observed with these hybrid gels. The FT-IR, XRD and TGA analysis showed that hydrogen bonds between the PAM chains and BC nanofiber clusters mainly contributed to the superior mechanical properties of hybrid hydrogels. The cell viability results suggested that PAM/BC hybrid hydrogel was benign for biomedical application. These PAM/BC hydrogels offer a great promise as biomaterials such as bone and cartilage repair materials.

  20. Supercoiling induces denaturation bubbles in circular DNA.

    Science.gov (United States)

    Jeon, Jae-Hyung; Adamcik, Jozef; Dietler, Giovanni; Metzler, Ralf

    2010-11-12

    We present a theoretical framework for the thermodynamic properties of supercoiling-induced denaturation bubbles in circular double-stranded DNA molecules. We explore how DNA supercoiling, ambient salt concentration, and sequence heterogeneity impact on the bubble occurrence. An analytical derivation of the probability distribution to find multiple bubbles is derived and the relevance for supercoiled DNA discussed. We show that in vivo sustained DNA bubbles are likely to occur due to partial twist release in regions rich in weaker AT base pairs. Single DNA plasmid imaging experiments clearly demonstrate the existence of bubbles in free solution.

  1. Quasi-elastic laser light scattering study of polyacrylamide hydrogel immersed in water and salt solutions

    Indian Academy of Sciences (India)

    M Sivanantham; B V R Tata

    2010-12-01

    Polyacrylamide (PAAm) hydrogels immersed in water and aqueous NaCl solutions were investigated for their structure and dynamics using static and quasi-elastic laser light scattering (QELS) techniques. Ensemble-averaged electric field correlation function (, ) obtained from the non-ergodic analysis of intensity-autocorrelation function for PAAm gel immersed in water and in 5 M NaCl showed an exponential decay to a plateau with an initial decay followed by saturation at long times. The value of the plateau was found to depend on NaCl concentration and was higher than that of water. Collective diffusion coefficient, , of the polymer network of the hydrogel immersed in water and in different concentrations of NaCl was determined by analysing (, ). The measured diffusion coefficient showed linear decrease with increase in concentration of NaCl. The characteristic network parameters were obtained by analyzing (, ) with harmonically bound Brownian particle model and from static light scattering studies.

  2. Synthesis and Characterization of Konjac Glucomannan-Graft-Polyacrylamide via γ-Irradiation

    Directory of Open Access Journals (Sweden)

    Jie Pang

    2008-03-01

    Full Text Available The synthesis of konjac glucomannan-graft-polyacrylamide (KGM-g-PAM wascarried out at 25°C by γ-irradiation under a N2 atmosphere. The effects of absorbedradiation dosage and monomer concentration on grafting yield and water absorbency werestudied. The grafted copolymers were characterized using Fourier Transform Infrared(FTIR spectroscopy, nuclear magnetic resonance (NMR, x-ray diffraction (XRD,thermogravimetric analysis (TGA and gel permeation chromatography (GPC. Thegrafting yield was observed to increase with increasing absorbed dosage and monomerconcentration. Compared with the original KGM, the grafted copolymers exhibited betterthermal stability and water absorbency. The results suggest that γ-irradiation is convenientand efficient for inducing graft copolymerization of KGM and acrylamide (AM.

  3. Antioxidant effect of green tea on polymer gel dosimeter

    Science.gov (United States)

    Samuel, E. J. J.; Sathiyaraj, P.; Deena, T.; Kumar, D. S.

    2015-01-01

    Extract from Green Tea (GTE) acts as an antioxidant in acrylamide based polymer gel dosimeter. In this work, PAGAT gel was used for investigation of antioxidant effect of GTE.PAGAT was called PAGTEG (Polyacrylamide green tea extract gel dosimeter) after adding GTE. Free radicals in water cause pre polymerization of polymer gel before irradiation. Polyphenols from GTE are highly effective to absorb the free radicals in water. THPC is used as an antioxidant in polymer gel dosimeter but here we were replaced it by GTE and investigated its effect by spectrophotometer. GTE added PAGAT samples response was lower compared to THPC added sample. To increase the sensitivity of the PAGTEG, sugar was added. This study confirmed that THPC was a good antioxidant for polymer gel dosimeter. However, GTE also can be used as an antioxidant in polymer gel if use less quantity (GTE) and add sugar as sensitivity enhancer.

  4. 27 CFR 21.151 - List of denaturants authorized for denatured spirits.

    Science.gov (United States)

    2010-04-01

    ... TOBACCO TAX AND TRADE BUREAU, DEPARTMENT OF THE TREASURY LIQUORS FORMULAS FOR DENATURED ALCOHOL AND RUM... (Glycerol), U.S.P S.D.A. 31-A. Green soap, U.S.P S.D.A. 27-B. Guaiacol, N.F.X S.D.A. 38-B. Heptane C.D.A....

  5. 78 FR 38628 - Reclassification of Specially Denatured Spirits and Completely Denatured Alcohol Formulas and...

    Science.gov (United States)

    2013-06-27

    ... alcohol. 13-A 10 gallons of ethyl ether. 19 100 gallons of ethyl ether. 23-A 8 gallons of acetone, U.S.P... alcohol. 32 5 gallons of ethyl ether. 35-A 4.25 gallons of ethyl acetate having an ester content of 100... regulations regarding the production, warehousing, denaturing, distribution, sale, export, and use...

  6. THE APPLICATION OF CATIONIC POLYACRYLAMIDE AS DRAINAGE AND RETENTION AIDS

    Institute of Scientific and Technical Information of China (English)

    JinWang; KefuChen; FushanChent; ChuanshanZhao; RendangYang

    2004-01-01

    Cationic polyacrylamide (CPAM) which was prepared through complex initiation system in laboratory and cationic polyacrylamide (Ciba) were used asretention and drainage aids for bleached wheat strawpulp. The influences of (polydiallyldimethlammonium chloride) PDADMAC and PDADMAC/CPAMon Zeta potential and drainability of pulp was investigated. The dual drainage and retention systems suchas CPAM/modified bentonite, CPAM/colloidal SiO2,as well as their comparison between the two systemswere discussed, and the optimal dosages of additiveswere determined. The results showed that: the complex systems can further enhance filler retention,drainability of pulp and strength properties of paper.

  7. Partially folded intermediates during trypsinogen denaturation

    Directory of Open Access Journals (Sweden)

    Martins N.F.

    1999-01-01

    Full Text Available The equilibrium unfolding of bovine trypsinogen was studied by circular dichroism, differential spectra and size exclusion HPLC. The change in free energy of denaturation was = 6.99 ± 1.40 kcal/mol for guanidine hydrochloride and = 6.37 ± 0.57 kcal/mol for urea. Satisfactory fits of equilibrium unfolding transitions required a three-state model involving an intermediate in addition to the native and unfolded forms. Size exclusion HPLC allowed the detection of an intermediate population of trypsinogen whose Stokes radii varied from 24.1 ± 0.4 Å to 26.0 ± 0.3 Å for 1.5 M and 2.5 M guanidine hydrochloride, respectively. During urea denaturation, the range of Stokes radii varied from 23.9 ± 0.3 Å to 25.7 ± 0.6 Å for 4.0 M and 6.0 M urea, respectively. Maximal intrinsic fluorescence was observed at about 3.8 M urea with 8-aniline-1-naphthalene sulfonate (ANS binding. These experimental data indicate that the unfolding of bovine trypsinogen is not a simple transition and suggest that the equilibrium intermediate population comprises one intermediate that may be characterized as a molten globule. To obtain further insight by studying intermediates representing different stages of unfolding, we hope to gain a better understanding of the complex interrelations between protein conformation and energetics.

  8. Comblike Polyacrylamides as Flooding Agent in Enhanced Oil Recovery

    NARCIS (Netherlands)

    Wever, Diego A. Z.; Picchioni, Francesco; Broekhuis, Antonius A.

    2013-01-01

    The oil recovery from core material and a specifically designed flow cell using novel branched (comblike) polyacrylamides (PAM) has been investigated. The injectivity characteristics of the different branched PAMs were evaluated by filtration tests and core-flow experiments. The number of arms of th

  9. Solids and nutrient removal from flushed swine manure using polyacrylamide

    Energy Technology Data Exchange (ETDEWEB)

    Perez Sangrador, M. P.; Garcia Gonzalez, M. C.; Leon Cofreces, M. C.; Acitores Benavente, M.

    2009-07-01

    Most of the organic nutrient elements (nitrogen and phosphorus) and carbon compounds (COD) in liquid swine manure are contained in fine suspended particles. Flocculation treatment with polyacrylamide (PAM) followed by screening if on the best methods to separate the liquid fraction.

  10. Comblike Polyacrylamides as Flooding Agent in Enhanced Oil Recovery

    NARCIS (Netherlands)

    Wever, Diego A. Z.; Picchioni, Francesco; Broekhuis, Antonius A.

    2013-01-01

    The oil recovery from core material and a specifically designed flow cell using novel branched (comblike) polyacrylamides (PAM) has been investigated. The injectivity characteristics of the different branched PAMs were evaluated by filtration tests and core-flow experiments. The number of arms of th

  11. Measurements of Elastic Moduli of Silicone Gel Substrates with a Microfluidic Device

    OpenAIRE

    Edgar Gutierrez; Alex Groisman

    2011-01-01

    Thin layers of gels with mechanical properties mimicking animal tissues are widely used to study the rigidity sensing of adherent animal cells and to measure forces applied by cells to their substrate with traction force microscopy. The gels are usually based on polyacrylamide and their elastic modulus is measured with an atomic force microscope (AFM). Here we present a simple microfluidic device that generates high shear stresses in a laminar flow above a gel-coated substrate and apply the d...

  12. Graphitic carbon nitride embedded hydrogels for enhanced gel electrophoresis.

    Science.gov (United States)

    Zarei, Mohammad; Ahmadzadeh, Hossein; Goharshadi, Elaheh K; Farzaneh, Ali

    2015-08-05

    Here, we show, for the first time, the use of graphitic carbon nitride (g-C3N4) nanosheets to improve the resolution and efficiency of protein separation in gel electrophoresis. By loading 0.04% (m/v) g-C3N4 nanosheets into the polyacrylamide gel at 25 °C, the thermal conductivity increased approximately 80% which resulted in 20% reduction in Joule heating and overall increase of separation efficiency. Also, polymerization of acrylamide occurred in the absence of tetramethylethylenediamine (TEMED) when the polyacrylamide gel contained g-C3N4 nanosheets. Hence, the g-C3N4 act simultaneously as a polymerization catalyst as well as heat sinks to lower Joule heating effect on band broadening.

  13. Can A Denaturant Stabilize DNA? Pyridine Reverses DNA Denaturation in Acidic pH.

    Science.gov (United States)

    Portella, Guillem; Terrazas, Montserrat; Villegas, Núria; González, Carlos; Orozco, Modesto

    2015-09-01

    The stability of DNA is highly dependent on the properties of the surrounding solvent, such as ionic strength, pH, and the presence of denaturants and osmolytes. Addition of pyridine is known to unfold DNA by replacing π-π stacking interactions between bases, stabilizing conformations in which the nucleotides are solvent exposed. We show here experimental and theoretical evidences that pyridine can change its role and in fact stabilize the DNA under acidic conditions. NMR spectroscopy and MD simulations demonstrate that the reversal in the denaturing role of pyridine is specific, and is related to its character as pseudo groove binder. The present study sheds light on the nature of DNA stability and on the relationship between DNA and solvent, with clear biotechnological implications.

  14. Kinetically controlled refolding of a heat-denatured hyperthermostable protein

    NARCIS (Netherlands)

    Koutsopoulos, Sotirios; van der Oost, John; Norde, Willem

    2007-01-01

    The thermal denaturation of endo-beta-1,3-glucanase from the hyperthermophilic microorganism Pyrococcus furiosus was studied by calorimetry. The calorimetric profile revealed two transitions at 109 and 144 degrees C, corresponding to protein denaturation and complete unfolding, respectively, as

  15. Kinetically controlled refolding of a heat denatured hyperthermostable protein

    NARCIS (Netherlands)

    Koutsopoulos, S.; Oost, van der J.; Norde, W.

    2007-01-01

    The thermal denaturation of endo-ß-1,3-glucanase from the hyperthermophilic microorganism Pyrococcus furiosus was studied by calorimetry. The calorimetric profile revealed two transitions at 109 and 144¿°C, corresponding to protein denaturation and complete unfolding, respectively, as shown by

  16. 19 CFR 10.56 - Vegetable oils, denaturing; release.

    Science.gov (United States)

    2010-04-01

    ... 19 Customs Duties 1 2010-04-01 2010-04-01 false Vegetable oils, denaturing; release. 10.56 Section... Vegetable Oils § 10.56 Vegetable oils, denaturing; release. (a) Olive, palm-kernel, rapeseed, sunflower, and sesame oil shall be classifiable under subheadings 1509.10.20, 1509.10.40, 1509.90.20, 1509.90.40,...

  17. 27 CFR 19.464 - Denatured spirits inventories.

    Science.gov (United States)

    2010-04-01

    ... inventories. 19.464 Section 19.464 Alcohol, Tobacco Products and Firearms ALCOHOL AND TOBACCO TAX AND TRADE... of Articles Inventories § 19.464 Denatured spirits inventories. Each proprietor shall take a physical inventory of all denatured spirits in the processing account at the close of each calendar quarter and...

  18. 27 CFR 20.144 - Packages of completely denatured alcohol.

    Science.gov (United States)

    2010-04-01

    ... 27 Alcohol, Tobacco Products and Firearms 1 2010-04-01 2010-04-01 false Packages of completely denatured alcohol. 20.144 Section 20.144 Alcohol, Tobacco Products and Firearms ALCOHOL AND TOBACCO TAX AND TRADE BUREAU, DEPARTMENT OF THE TREASURY LIQUORS DISTRIBUTION AND USE OF DENATURED ALCOHOL AND RUM...

  19. 27 CFR 20.261 - Records of completely denatured alcohol.

    Science.gov (United States)

    2010-04-01

    ... 27 Alcohol, Tobacco Products and Firearms 1 2010-04-01 2010-04-01 false Records of completely denatured alcohol. 20.261 Section 20.261 Alcohol, Tobacco Products and Firearms ALCOHOL AND TOBACCO TAX AND TRADE BUREAU, DEPARTMENT OF THE TREASURY LIQUORS DISTRIBUTION AND USE OF DENATURED ALCOHOL AND...

  20. A comparison of the sensitivity of three gel electrophoresis methods for the RFLP analysis of mycobacterial heat shock protein 65 gene%三种凝胶电泳分析分枝杆菌热休克蛋白65基因酶切片段的灵敏度比较

    Institute of Scientific and Technical Information of China (English)

    张彩萍; 王洪生; 冯雨苗; 林麟; 崔盘根; 陈敏; 吴勤学

    2013-01-01

    Objective To compare the performance of 2% (w/v) agarose gel,2% (w/v) Metaphor agarose gel and 10%(w/v) nondenaturating polyacrylamide gel in the PCR-restriction fragment length polymorphism (RFLP) analysis of mycobacterial heat shock protein 65 (hsp65) gene.Methods This study included 8 Mycobacteria strains,including clinical isolates and standard strains of Mycobacteria tuberculosis and Mycobacterium intracellulare.Bacterialsuspension of these strains was prepared with the concentration of bacterial cells varying from 10 to 106per milliliter.PCR was performed to amplify the hsp65 gene with a pair of universal primers followed by the digestion of amplicons with two restriction endonucleases,BstE Ⅱ and Hae Ⅲ.Then,the restriction enzyme-digested fragments were subjected to electrophoresis in 2% agarose gel,2% Metaphor agarose gel and 10% nondenaturating polyacrylamide gel respectively.Results As analysis of variance showed,the three gel electrophoresis methods were statistically different in sensitivity for the RFLP analysis of mycobacterial hsp65 gene (F =36.379,P < 0.01).Least significance difference (LSD) procedure revealed that the 2% agarose gel-based electrophoresis was less sensitive than the 2% Metaphor agarose gel-and 10% non-denaturing polyacrylamide gel-based electrophoresis (both P < 0.01),and no significant differences were observed between the 2% Metaphor agarose gel-and 10% non-denaturing polyacrylamide gel-based electrophoresis (P > 0.05).Conclusion The 2% Metaphor agarose gel-and 10%nondenaturating polyacrylamide gel-based electrophoresis methods appear to be more sensitive than the 2% agarose gel-based electrophoresis method for the PCR-RFLP analysis of mycobacterial hsp65 gene.%目的 比较2%(w/v)琼脂糖凝胶、2% (w/v)Metaphor琼脂糖凝胶和10%(w/v)非变性聚丙烯酰胺凝胶分析分枝杆菌热休克蛋白65(hsp65)基因酶切片段的灵敏度.方法 分枝杆菌8株,分别制成106~101

  1. Approach to analysis of single nucleotide polymorphisms by automated constant denaturant capillary electrophoresis

    Energy Technology Data Exchange (ETDEWEB)

    Bjoerheim, Jens; Abrahamsen, Torveig Weum; Kristensen, Annette Torgunrud; Gaudernack, Gustav; Ekstroem, Per O

    2003-05-15

    Melting gel techniques have proven to be amenable and powerful tools in point mutation and single nucleotide polymorphism (SNP) analysis. With the introduction of commercially available capillary electrophoresis instruments, a partly automated platform for denaturant capillary electrophoresis with potential for routine screening of selected target sequences has been established. The aim of this article is to demonstrate the use of automated constant denaturant capillary electrophoresis (ACDCE) in single nucleotide polymorphism analysis of various target sequences. Optimal analysis conditions for different single nucleotide polymorphisms on ACDCE are evaluated with the Poland algorithm. Laboratory procedures include only PCR and electrophoresis. For direct genotyping of individual SNPs, the samples are analyzed with an internal standard and the alleles are identified by co-migration of sample and standard peaks. In conclusion, SNPs suitable for melting gel analysis based on theoretical thermodynamics were separated by ACDCE under appropriate conditions. With this instrumentation (ABI 310 Genetic Analyzer), 48 samples could be analyzed without any intervention. Several institutions have capillary instrumentation in-house, thus making this SNP analysis method accessible to large groups of researchers without any need for instrument modification.

  2. Reducing the stiffness of concentrated whey protein isolate (WPI) gels by using WPI microparticles

    NARCIS (Netherlands)

    Purwanti, N.; Moerkens, A.; Goot, van der A.J.; Boom, R.M.

    2012-01-01

    Concentrated protein gels were prepared using native whey protein isolate (WPI) and WPI based microparticles. WPI microparticles were produced by making gel pieces from a concentrated WPI suspension (40% w/w), which were dried and milled. The protein within the microparticles was denatured and the

  3. Feasibility of a DNA-Based Combinatorial Array Recognition Surface (CARS) in a Polyacrylamide Gel Matrix

    Science.gov (United States)

    2007-12-12

    the phosphate backbone. wherever p.·J.Ttial hybrids naturally occur, via Taq DNA ligase (16). Ligation may not have been entirely necessary for the...CARS libraries, noncontiguous pieces can be ligaled togclher willi Taq DNA ligase . The lOp half o(lhe figure iIIu51Tates Ihe appearance o( a I-D CARS...cluding dideoxynucleotides. were from a "Si lver Sequence" kit purchased from Promega Corporation (Madison. WI). ThermliS aquaticus (Taq) DNA ligase was

  4. Competitive sorption and diffusion of chromate and sulphate in a flow system with goethite in gel beads

    NARCIS (Netherlands)

    Beinum, van G.W.; Meeussen, J.C.L.; Riemsdijk, van W.H.

    2006-01-01

    Column experiments and model simulations were employed to evaluate the processes involved in multicomponent solute transport in a system with heterogeneous flow. Column experiments were performed with goethite embedded in polyacrylamide gel beads. The gel forms an immobile water region that can be a

  5. Flocculation of the Visonta lignite sludges by polyacrylamides

    Energy Technology Data Exchange (ETDEWEB)

    Lakatos-Szabo, J.; Lakatos, I.

    1986-01-01

    Laboratory tests on the flocculation of clay-lignite sludges by polyelectrolytes proved the strongly heterodisperse suspensions to be suitable to sedimenting and filtering after the treatment by anionic polyacrylamides. The formation of large-sized flocs of loose structure allows troublefree filtration and dewatering. The specific flocculant consumption of dewatering is 1 to 2 kg per lignite ton. In this manner the preparation costs of lignite can be lowered.

  6. THE CHARACTERISTICS OF HIGH MOLECULAR WEIGHT CATIONIC POLYACRYLAMIDE

    Institute of Scientific and Technical Information of China (English)

    Hongjie Zhang; Huiren Hu; Fushan Chen

    2004-01-01

    In this paper, the cationic polyacrylamide (CPAM)with high molecular weight was prepared in aqueous solution through a complex initiator system. The CPAM was characterized by Fourier transform infrared spectroscopy (FTIR) and 13C nuclear magnetic resonance spectroscopy (13C NMR), and the charge density of the CPAM was determined by colloid titration. The results obtained indicated that the copolymerization technology used in the experiment was successful.

  7. Thermal denaturation of A-DNA

    Science.gov (United States)

    Valle-Orero, J.; Wildes, A. R.; Theodorakopoulos, N.; Cuesta-López, S.; Garden, J.-L.; Danilkin, S.; Peyrard, M.

    2014-11-01

    The DNA molecule can take various conformational forms. Investigations focus mainly on the so-called ‘B-form’, schematically drawn in the famous paper by Watson and Crick [1]. This is the usual form of DNA in a biological environment and is the only form that is stable in an aqueous environment. Other forms, however, can teach us much about DNA. They have the same nucleotide base pairs for ‘building blocks’ as B-DNA, but with different relative positions, and studying these forms gives insight into the interactions between elements under conditions far from equilibrium in the B-form. Studying the thermal denaturation is particularly interesting because it provides a direct probe of those interactions which control the growth of the fluctuations when the ‘melting’ temperature is approached. Here we report such a study on the ‘A-form’ using calorimetry and neutron scattering. We show that it can be carried further than a similar study on B-DNA, requiring the improvement of thermodynamic models for DNA.

  8. Optimal processing for gel electrophoresis images: Applying Monte Carlo Tree Search in GelApp.

    Science.gov (United States)

    Nguyen, Phi-Vu; Ghezal, Ali; Hsueh, Ya-Chih; Boudier, Thomas; Gan, Samuel Ken-En; Lee, Hwee Kuan

    2016-08-01

    In biomedical research, gel band size estimation in electrophoresis analysis is a routine process. To facilitate and automate this process, numerous software have been released, notably the GelApp mobile app. However, the band detection accuracy is limited due to a band detection algorithm that cannot adapt to the variations in input images. To address this, we used the Monte Carlo Tree Search with Upper Confidence Bound (MCTS-UCB) method to efficiently search for optimal image processing pipelines for the band detection task, thereby improving the segmentation algorithm. Incorporating this into GelApp, we report a significant enhancement of gel band detection accuracy by 55.9 ± 2.0% for protein polyacrylamide gels, and 35.9 ± 2.5% for DNA SYBR green agarose gels. This implementation is a proof-of-concept in demonstrating MCTS-UCB as a strategy to optimize general image segmentation. The improved version of GelApp-GelApp 2.0-is freely available on both Google Play Store (for Android platform), and Apple App Store (for iOS platform). © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  9. Microwave-enhanced folding and denaturation of globular proteins

    DEFF Research Database (Denmark)

    Bohr, Henrik; Bohr, Jakob

    2000-01-01

    It is shown that microwave irradiation can affect the kinetics of the folding process of some globular proteins, especially beta-lactoglobulin. At low temperature the folding from the cold denatured phase of the protein is enhanced, while at a higher temperature the denaturation of the protein from...... its folded state is enhanced. In the latter case, a negative temperature gradient is needed for the denaturation process, suggesting that the effects of the microwaves are nonthermal. This supports the notion that coherent topological excitations can exist in proteins. The application of microwaves...

  10. Statistical mechanics of thermal denaturation of DNA oligomers

    Indian Academy of Sciences (India)

    Navin Singh; Yashwant Singh

    2003-08-01

    Double stranded DNA chain is known to have non-trivial elasticity. We study the effect of this elasticity on the denaturation profile of DNA oligomer by constraining one base pair at one end of the oligomer to remain in unstretched (or intact) state. The effect of this constraint on the denaturation profile of the oligomer has been calculated using the Peyrard–Bishop Hamiltonian. The denaturation profile is found to be very different from the free (i.e. without the constraint) oligomer. We have also examined how this constraint affects the denaturation profile of the oligomer having a segment of defect sites located at different parts of the chain.

  11. Simultaneous immobilization of dehydrogenases on polyvinylidene difluoride resin after separation by non-denaturing two-dimensional electrophoresis

    Energy Technology Data Exchange (ETDEWEB)

    Shimazaki, Youji [Graduate School of Science and Engineering (Science Section) and Venture Business Laboratory, Ehime University, Bunkyo-cho 2-5, Matsuyama City 790-8577 (Japan)], E-mail: yoji@dpc.ehime-u.ac.jp; Kadota, Mariko [Faculty of Science, Ehime University, Matsuyama (Japan)

    2008-06-16

    We detected mouse liver malate, sorbitol and aldehyde dehydrogenases by negative staining, analysis of malate and sorbitol dehydrogenase activities using each substrate, and electron transfers including nicotinamide adenine dinucleotide (NAD) and nitroblue tetrazolium in non-denaturing two-dimensional electrophoresis (2-DE) gel. Dehydrogenases were also identified by electrospray ionization tandem mass spectrometry (ESI-MS/MS) after 2-DE separation and protein detection by negative staining. Spots of dehydrogenases separated by 2-DE were excised, and simultaneously transferred and immobilized on polyvinylidene difuoride (PVDF) resin by electrophoresis. The dehydrogenase activities remained intact after immobilization. In conclusion, resin-immobilized dehydrogenases can be simultaneously obtained after separation by non-denaturing 2-DE, detection by negative staining and transferring to resins.

  12. pH-responsive polymer-assisted refolding of urea- and organic solvent-denatured alpha-chymotrypsin.

    Science.gov (United States)

    Roy, I; Gupta, M N

    2003-12-01

    A pH-responsive polymer Eudragit S-100 has been found to assist in correct folding of alpha-chymotrypsin denatured with 8 M urea and 100 mM dithiothreitol at pH 8.2. The complete activity could be regained within 10 min during refolding. Both native and refolded enzymes showed emission of intrinsic fluorescence with lambda(max) of 342 nm. Gel electrophoresis showed that the presence of Eudragit S-100 led to dissociation of multimers followed by the appearance of a band at the monomer position. The unfolding (by 8 M urea) and folding (assisted by the polymer) also led to complete renaturation of alpha-chymotrypsin initially denatured by 90% dioxane. The implications of the data in recovery of enzyme activity from inclusion bodies and the interesting possibility in the in vivo context of reversing protein aggregation in amyloid-based diseases have been discussed.

  13. Impact of Delivery Mode on Intestinal Microbiota in Early Neonates by Polymerase Chain Reaction - Denaturing Gradient Gel Electrophoresis%聚合酶链反应-变性梯度凝胶电泳技术动态分析不同分娩方式对新生儿早期肠道菌群的影响

    Institute of Scientific and Technical Information of China (English)

    刘彩霞; 胡燕; 周东蕊; 蒋犁

    2012-01-01

    Objective To investigate the effects of delivery mode on the dynamic changes of early gut microflora by using polymerase chain reaction - degeneration gradient gel electrophoresis (PCR - DGGE) method. Methods Thirty - two normal term neonates were enrolled in the study (including vaginal delivery group and caesarean section group,each group including 16 neonates) ,and feces samples were collected on the first day( > 12 h) ,3th day and 5th day after birth. After bacterial DNAs were extracted from the feces of the 2 groups,bacterial communities in 2 groups were examined by PCR of 16S rDNA V3 region and DGGE. After DGGE profilings were obtained,the diversity and similarity related indices were used to analyze diversity differences of the gut bacterium structure between 2 groups and the similar degrees between individuals within each group. Results 1. There were few species and quantities in the intestinal microbiota during the first day of life, and the early intestinal bacterial colonization of neonates had a complex individual specificity and presented dynamic changes. 2. The diversity analysis between 2 groups showed that there were no obvious diversity differences between the 2 groups on 3' day after birth. While on 5' day after birth,the Shannon - Weaver Diversity Index(H') and the simpson index(D) in caesarean section group were lower significandy than those in vaginal delivery group(Pa <0.05).3. The similarity analysis showed that the dice similarity coefficient(Cs)between individuals in cesarean section group was higher than that in vaginal delivery group on 3th day after birth, and the Cs lower than (or equal to)0. 30 took up 21.4% of the total Cs in cesarean section group and 76.0%in vaginal delivery group,which had all significant differences between 2 groups (P <0.05). On 5th day after birth,there were no obvious differences between groups in Cs total level and its distribution. Conclusions The intra - group similarity of gut bacterium structure is

  14. Synthesis, Characterization, and Flocculation Properties of Branched Cationic Polyacrylamide

    Directory of Open Access Journals (Sweden)

    Weimin Sun

    2013-01-01

    Full Text Available A water soluble branched cationic polyacrylamide (BCPAM was synthesized using solution polymerization. The polymerization was initiated using potassium diperiodatocuprate, K5[Cu(HIO62](Cu(III, initiating the self-condensing vinyl copolymerization of acrylamide and acryloxyethyltrimethyl ammonium chloride (DAC monomer. The resulting copolymer was characterized by the use of Fourier-transform infrared (FTIR spectroscopy and nuclear magnetic resonance (NMR spectroscopy. Its flocculation properties were evaluated with standard jar tests of sewage. The effects of initiator concentration, monomer concentration, reaction temperature, and the mass ratio of monomers on intrinsic viscosity and flocculation properties of the product were determined using single-factor experiments and orthogonal experiment.

  15. MONODISPERSE MICRON-SIZED POLYACRYLAMIDE PARTICLES SYNTHESIZED BY DISPERSION POLYMERIZATION

    Institute of Scientific and Technical Information of China (English)

    Xin Hou; Bo Gao; Zhe-guo Zhang; Kang-de Yao

    2007-01-01

    Monodisperse micron-sized polyacrylamide (PAM) particles with a regular shape have been successfully prepared through dispersion polymerization of the monomer using a rotary reactor. FTIR and NMR spectroscopic results demonstrated the formation of PAM. POM and TEM observations revealed that PAM particles had a regular shape and good dispersity. A thick layer of surfactant (PVP) still existed on PAM particles after multiple centrifugation and ultrasonic re-dispersion in ethanol, which indicates a strong interaction between PVP and PAM. The effects of various polymerization factors on the average size of PAM particles have also been studied.

  16. Cationic polyacrylamides enhance rates of starch and cellulose saccharification.

    Science.gov (United States)

    Reye, John T; Maxwell, Kendra; Rao, Swati; Lu, Jian; Banerjee, Sujit

    2009-10-01

    Adding a cationic polyacrylamide (c-PAM) to either the amylase mediated hydrolysis of corn starch or the hydrolysis of wood fiber by cellulase can enhance the initial hydrolysis rates, although a rate decrease can occur under some conditions. Several c-PAMs can serve as catalysts and the same c-PAM can improve the efficiency of both amylase and cellulase. The initial amylase rate approximately doubles; the analogous cellulase hydrolysis rate increases by about 40%. c-PAMs increase the binding of enzyme to substrate.

  17. Microchannel gel electrophoretic separation systems and methods for preparing and using

    Science.gov (United States)

    Herr, Amy; Singh, Anup K; Throckmorton, Daniel J

    2013-09-03

    A micro-analytical platform for performing electrophoresis-based immunoassays was developed by integrating photopolymerized cross-linked polyacrylamide gels within a microfluidic device. The microfluidic immunoassays are performed by gel electrophoretic separation and quantifying analyte concentration based upon conventional polyacrylamide gel electrophoresis (PAGE). To retain biological activity of proteins and maintain intact immune complexes, native PAGE conditions were employed. Both direct (non-competitive) and competitive immunoassay formats are demonstrated in microchips for detecting toxins and biomarkers (cytokines, c-reactive protein) in bodily fluids (serum, saliva, oral fluids). Further, a description of gradient gels fabrication is included, in an effort to describe methods we have developed for further optimization of on-chip PAGE immunoassays. The described chip-based PAGE immunoassay method enables immunoassays that are fast (minutes) and require very small amounts of sample (less than a few microliters). Use of microfabricated chips as a platform enables integration, parallel assays, automation and development of portable devices.

  18. Microchannel gel electrophoretic separation systems and methods for preparing and using

    Energy Technology Data Exchange (ETDEWEB)

    Herr, Amy E; Singh, Anup K; Throckmorton, Daniel J

    2015-02-24

    A micro-analytical platform for performing electrophoresis-based immunoassays was developed by integrating photopolymerized cross-linked polyacrylamide gels within a microfluidic device. The microfluidic immunoassays are performed by gel electrophoretic separation and quantifying analyte concentration based upon conventional polyacrylamide gel electrophoresis (PAGE). To retain biological activity of proteins and maintain intact immune complexes, native PAGE conditions were employed. Both direct (non-competitive) and competitive immunoassay formats are demonstrated in microchips for detecting toxins and biomarkers (cytokines, c-reactive protein) in bodily fluids (serum, saliva, oral fluids). Further, a description of gradient gels fabrication is included, in an effort to describe methods we have developed for further optimization of on-chip PAGE immunoassays. The described chip-based PAGE immunoassay method enables immunoassays that are fast (minutes) and require very small amounts of sample (less than a few microliters). Use of microfabricated chips as a platform enables integration, parallel assays, automation and development of portable devices.

  19. Microchannel gel electrophoretic separation systems and methods for preparing and using

    Energy Technology Data Exchange (ETDEWEB)

    Herr, Amy; Singh, Anup K; Throckmorton, Daniel J

    2013-09-03

    A micro-analytical platform for performing electrophoresis-based immunoassays was developed by integrating photopolymerized cross-linked polyacrylamide gels within a microfluidic device. The microfluidic immunoassays are performed by gel electrophoretic separation and quantifying analyte concentration based upon conventional polyacrylamide gel electrophoresis (PAGE). To retain biological activity of proteins and maintain intact immune complexes, native PAGE conditions were employed. Both direct (non-competitive) and competitive immunoassay formats are demonstrated in microchips for detecting toxins and biomarkers (cytokines, c-reactive protein) in bodily fluids (serum, saliva, oral fluids). Further, a description of gradient gels fabrication is included, in an effort to describe methods we have developed for further optimization of on-chip PAGE immunoassays. The described chip-based PAGE immunoassay method enables immunoassays that are fast (minutes) and require very small amounts of sample (less than a few microliters). Use of microfabricated chips as a platform enables integration, parallel assays, automation and development of portable devices.

  20. The latest advancements in proteomic two-dimensional gel electrophoresis analysis applied to biological samples.

    Science.gov (United States)

    Santucci, Laura; Bruschi, Maurizio; Ghiggeri, Gian Marco; Candiano, Giovanni

    2015-01-01

    Two-dimensional gel electrophoresis (2DE) is one of the fundamental approaches in proteomics for the separation and visualization of complex protein mixtures. Proteins can be analyzed by 2DE using isoelectric focusing (IEF) in the first dimension, combined to sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) in the second dimension, gel staining (silver and Coomassie), image analysis, and 2DE gel database. High-resolution 2DE can resolve up to 5,000 different proteins simultaneously (∼2,000 proteins routinely), and detect and quantify <1 ng of protein per spot. Here, we describe the latest developments for a more complete analysis of biological fluids.

  1. Molecular monitoring of the intestinal flora by denaturing high performance liquid chromatography.

    Science.gov (United States)

    Goldenberg, Oliver; Herrmann, Stefanie; Marjoram, Gina; Noyer-Weidner, Mario; Hong, George; Bereswill, Stefan; Göbel, Ulf B

    2007-01-01

    Gut flora analysis is hampered by the complexity of the intestinal microbiota and by inherent limitations of culture-based approaches. Therefore, culture-independent molecular methods based upon 16S rRNA gene analysis were applied successfully for the analysis of complex microbial communities. However, generally accepted and validated profiling methods such as denaturing and temperature gradient gel electrophoresis (DGGE/TGGE) are still laborious and time consuming. Thus, we adapted the separation of amplified bacterial 16S rRNA gene fragments by denaturing high performance liquid chromatography (DHPLC) using the WAVE Microbial Analysis System as a rapid and convenient means to display complex intestinal bacterial communities and to monitor changes in the gut flora. The separation of 16S rRNA gene fragments amplified from reference strains representing main gut bacterial populations and from human stool samples revealed that DHPLC analysis effectively detects bacterial groups predominant in the human gut flora. The investigation of faecal samples from hospitalized patients before, during and after antibiotic therapy showed that PCR-based DHPLC can be used to monitor gut flora changes. Results from DHPLC analysis were comparable with DGGE profiles generated from the same samples, demonstrating that the adapted DHPLC protocol is well suited for the analysis of complex microbial communities.

  2. 27 CFR 19.57 - Recovery and reuse of denatured spirits in manufacturing processes.

    Science.gov (United States)

    2010-04-01

    ... denatured spirits in manufacturing processes. 19.57 Section 19.57 Alcohol, Tobacco Products and Firearms... denatured spirits in manufacturing processes. The following persons are not, by reason of the activities...) Manufacturers who use denatured spirits, or articles or substances containing denatured spirits, in a process...

  3. Development of pH sensitive polyacrylamide grafted pectin hydrogel for controlled drug delivery system.

    Science.gov (United States)

    Sutar, Prashant B; Mishra, Rakesh K; Pal, Kunal; Banthia, Ajit K

    2008-06-01

    In the present study an attempt was made to graft polyacrylamide on pectin. The grafted polymer was characterized by FTIR spectroscopy, differential scanning calorimetry and X-ray diffraction. Rheological property of pectin solution was compared with the product solution. The grafted polymer was cross-linked with varying amount of glutaraldehyde. The swelling properties of the cross-linked product were also studied. The salicylic acid, an antipyretic drug, was incorporated in the cross-linked gel as a model drug and the drug release studies were done in a modified Franz's diffusion cell. The effect of cross-linking density on the release property of salicylic acid was studied through the cross-linked product. The product showed better film forming property and gelling property than pectin. The comparative rheological properties of pectin and grafted copolymer indicated change in the property of the product. FTIR studies indicated incorporation of amide group. Differential scanning calorimetry and XRD suggested formation of a new polymer. Swelling study indicated pH dependent swelling of the cross-linked hydrogel. Salicylic acid release indicated pH dependent release from the hydrogel.

  4. Isolation and Characterization of Polyacrylamide-Degrading Bacteria from Dewatered Sludge

    Directory of Open Access Journals (Sweden)

    Feng Yu

    2015-04-01

    Full Text Available Polyacrylamide (PAM is a water-soluble polymer that is widely used as a flocculant in sewage treatment. The accumulation of PAM affects the formation of dewatered sludge and potentially produces hazardous monomers. In the present study, the bacterial strain HI47 was isolated from dewatered sludge. This strain could metabolize PAM as its sole nutrient source and was subsequently identified as Pseudomonas putida. The efficiency of PAM degradation was 31.1% in 7 days and exceeded 45% under optimum culture condition (pH 7.2, 39 °C and 100 rpm. The addition of yeast extract and glucose improved the bacterial growth and PAM degradation. The degraded PAM samples were analyzed by gel-filtration chromatography, Fourier transform infrared and high-performance liquid chromatography. The results showed that high-molecular-weight PAM was partly cleaved to small molecular oligomer derivatives and part of the amide groups of PAM had been converted to carboxyl groups. The biodegradation did not accumulate acrylamide monomers. Based on the SDS-PAGE and N-terminal sequencing results, the PAM amide groups were converted into carboxyl groups by a PAM-induced extracellular enzyme from the aliphatic amidase family.

  5. Sequence-specific nucleic acid mobility using a reversible block copolymer gel matrix and DNA amphiphiles (lipid-DNA) in capillary and microfluidic electrophoretic separations

    NARCIS (Netherlands)

    Wagler, Patrick; Minero, Gabriel Antonio S.; Tangen, Uwe; de Vries, Jan Willem; Prusty, Deepak; Kwak, Minseok; Herrmann, Andreas; McCaskill, John S.

    2015-01-01

    Reversible noncovalent but sequence-dependent attachment of DNA to gels is shown to allow programmable mobility processing of DNA populations. The covalent attachment of DNA oligomers to polyacrylamide gels using acrydite-modified oligonucleotides has enabled sequence-specific mobility assays for

  6. Sequence-specific nucleic acid mobility using a reversible block copolymer gel matrix and DNA amphiphiles (lipid-DNA) in capillary and microfluidic electrophoretic separations

    NARCIS (Netherlands)

    Wagler, Patrick; Minero, Gabriel Antonio S.; Tangen, Uwe; de Vries, Jan Willem; Prusty, Deepak; Kwak, Minseok; Herrmann, Andreas; McCaskill, John S.

    2015-01-01

    Reversible noncovalent but sequence-dependent attachment of DNA to gels is shown to allow programmable mobility processing of DNA populations. The covalent attachment of DNA oligomers to polyacrylamide gels using acrydite-modified oligonucleotides has enabled sequence-specific mobility assays for DN

  7. High Resolution Microsatellite Marker Analysis of Some Rice Landraces Using Metaphor Agarose Gel Electrophoresis

    OpenAIRE

    K. Kristamtini; T. Taryono; Panjisakti Basunanda; Rudi Hari Murti

    2016-01-01

    Microsatellite markers or simple sequences repeats are DNA - based molecular techniques that are used to see the different among accessions and inbred lines. There are three methods to analysis the results of the polymerase chain reaction of microsatellite markers namely polyacrylamide gel electrophoresis (PAGE), capillary electroforesis, and Metaphor Agarose Gel Electroforesis (MAGE), and the Use of MAGE assessed more easily and economically the polymorphic pattern of DNA markers...

  8. Analysis of soybean embryonic axis proteins by two-dimensional gel electrophoresis and mass spectrometry

    Science.gov (United States)

    A proteomic approach based on two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) for protein separation and subsequent mass spectrometry (MS) for protein identification was applied to establish a proteomic reference map for the soybean embryonic axis. Proteins were extracted from dissecte...

  9. Cation hydration in hydrogelic polyacrylamide-phosphoric acid network: A study by Raman spectroscopy

    OpenAIRE

    Costa, A. M. Amorim da; Amado, A. M.

    2001-01-01

    The effects upon the structure and morphology of adding lithium, calcium and magnesium chlorides to a phosphoric acid/polyacrylamide 2:1 molar ratio proton conducting hydrogel are examined by observing the changes in the vibrational features of the polyacrylamide chain, in the phosphate group and in the interstitial water molecules as a function of the concentration and the cationic nature of the additive, at 295 K. On adding H3PO4 to the polyacrylamide hydrogel matrix, the amide groups becom...

  10. Comparative Two-Dimensional Fluorescence Gel Electrophoresis.

    Science.gov (United States)

    Ackermann, Doreen; König, Simone

    2018-01-01

    Two-dimensional comparative fluorescence gel electrophoresis (CoFGE) uses an internal standard to increase the reproducibility of coordinate assignment for protein spots visualized on 2D polyacrylamide gels. This is particularly important for samples, which need to be compared without the availability of replicates and thus cannot be studied using differential gel electrophoresis (DIGE). CoFGE corrects for gel-to-gel variability by co-running with the sample proteome a standardized marker grid of 80-100 nodes, which is formed by a set of purified proteins. Differentiation of reference and analyte is possible by the use of two fluorescent dyes. Variations in the y-dimension (molecular weight) are corrected by the marker grid. For the optional control of the x-dimension (pI), azo dyes can be used. Experiments are possible in both vertical and horizontal (h) electrophoresis devices, but hCoFGE is much easier to perform. For data analysis, commercial software capable of warping can be adapted.

  11. Dodine as a transparent protein denaturant for circular dichroism and infrared studies.

    Science.gov (United States)

    Guin, Drishti; Sye, Kori; Dave, Kapil; Gruebele, Martin

    2016-05-01

    The fungicide dodine combines the cooperative denaturation properties of guanidine with the mM denaturation activity of SDS. It was previously tested only on two small model proteins. Here we show that it can be used as a chemical denaturant for phosphoglycerate kinase (PGK), a much larger two-domain enzyme. In addition to its properties as a chemical denaturant, dodine facilitates thermal denaturation of PGK, and we show for the first time that it also facilitates pressure denaturation of a protein. Much higher quality circular dichroism and amide I' infrared spectra of PGK can be obtained in dodine than in guanidine, opening the possibility for use of dodine as a denaturant when UV or IR detection is desirable. One caution is that dodine denaturation, like other detergent-based denaturants, is less reversible than guanidine denaturation. © 2016 The Protein Society.

  12. Network confinement and heterogeneity slows nanoparticle diffusion in polymer gels

    Science.gov (United States)

    Parrish, Emmabeth; Caporizzo, Matthew A.; Composto, Russell J.

    2017-05-01

    Nanoparticle (NP) diffusion was measured in polyacrylamide gels (PAGs) with a mesh size comparable to the NP size, 21 nm. The confinement ratio (CR), NP diameter/mesh size, increased from 0.4 to 3.8 by increasing crosslinker density and from 0.4 to 2.1 by adding acetone, which collapsed the PAGs. In all gels, NPs either became localized, moving less than 200 nm, diffused microns, or exhibited a combination of these behaviors, as measured by single particle tracking. Mean squared displacements (MSDs) of mobile NPs decreased as CR increased. In collapsed gels, the localized NP population increased and MSD of mobile NPs decreased compared to crosslinked PAGs. For all CRs, van Hove distributions exhibited non-Gaussian displacements, consistent with intermittent localization of NPs. The non-Gaussian parameter increased from a maximum of 1.5 for crosslinked PAG to 5 for collapsed PAG, consistent with greater network heterogeneity in these gels. Diffusion coefficients decreased exponentially as CR increased for crosslinked gels; however, in collapsed gels, the diffusion coefficients decreased more strongly, which was attributed to network heterogeneity. Collapsing the gel resulted in an increasingly tortuous pathway for NPs, slowing diffusion at a given CR. Understanding how gel structure affects NP mobility will allow the design and enhanced performance of gels that separate and release molecules in membranes and drug delivery platforms.

  13. A PCR-denaturing gradient gel electrophoresis (DGGE) approach to assess Fusarium diversity in asparagus

    NARCIS (Netherlands)

    Yergeau, E.; Filion, M.; Vujanovic, V.; St-Arnaud, M.

    2005-01-01

    In North America, asparagus (Asparagus officinalis) production suffers from a crown and root rot disease mainly caused by Fusarium oxysporum f. sp. asparagi and F. proliferatum. Many other Fusarium species are also found in asparagus fields, whereas accurate detection and identification of these org

  14. RET and GDNF gene scanning in Hirschprung patients using two dual denaturing gel systems

    NARCIS (Netherlands)

    Hofstra, RMW; Wu, Y; Stulp, RP; Elfferich, P; Osinga, J; Maas, SM; Siderius, L; Brooks, AS; Von der Ende, JJ; Heydendael, VMR; Severijnen, RSVM; Bax, KMA; Meijers, C; Buys, CHCM

    2000-01-01

    Hirschsprung disease (HSCR) is a congenital disorder characterised by intestinal obstruction due to an absence of intramural ganglia along variable lengths of the intestine. RET is the major gene involved in HSCR. Mutations in the GDNF gene, and encoding one of the RET ligands, either alone or in co

  15. 7 NEW MUTATIONS IN HMSH2, AN HNPCC GENE, IDENTIFIED BY DENATURING GRADIENT-GEL ELECTROPHORESIS

    NARCIS (Netherlands)

    WIJNEN, J; VASEN, H; KHAN, PM; MENKO, FH; VANDERKLIFT, H; VANLEEUWEN, C; VANDENBROEK, M; VANLEEUWENCORNELISSE, [No Value; NAGENGAST, F; MEIJERSHEIJBOER, A; LINDHOUT, D; GRIFFIOEN, G; CATS, A; KLEIBEUKER, J; VARESCO, L; BERTARIO, L; BISGAARD, ML; MOHR, J; FODDE, R

    1995-01-01

    Hereditary nonpolyposis colorectal cancer (HNPCC) is a relatively common autosomal dominant cancer-susceptibility condition. The recent isolation of the DNA mismatch repair genes (hMSH2, hMLH1, hPMS1, and hPMS2) responsible for HNPCC has allowed the search for germ-line mutations in affected individ

  16. A PCR-denaturing gradient gel electrophoresis (DGGE) approach to assess Fusarium diversity in asparagus

    NARCIS (Netherlands)

    Yergeau, E.; Filion, M.; Vujanovic, V.; St-Arnaud, M.

    2005-01-01

    In North America, asparagus (Asparagus officinalis) production suffers from a crown and root rot disease mainly caused by Fusarium oxysporum f. sp. asparagi and F. proliferatum. Many other Fusarium species are also found in asparagus fields, whereas accurate detection and identification of these

  17. Measurements of elastic moduli of silicone gel substrates with a microfluidic device.

    Science.gov (United States)

    Gutierrez, Edgar; Groisman, Alex

    2011-01-01

    Thin layers of gels with mechanical properties mimicking animal tissues are widely used to study the rigidity sensing of adherent animal cells and to measure forces applied by cells to their substrate with traction force microscopy. The gels are usually based on polyacrylamide and their elastic modulus is measured with an atomic force microscope (AFM). Here we present a simple microfluidic device that generates high shear stresses in a laminar flow above a gel-coated substrate and apply the device to gels with elastic moduli in a range from 0.4 to 300 kPa that are all prepared by mixing two components of a transparent commercial silicone Sylgard 184. The elastic modulus is measured by tracking beads on the gel surface under a wide-field fluorescence microscope without any other specialized equipment. The measurements have small and simple to estimate errors and their results are confirmed by conventional tensile tests. A master curve is obtained relating the mixing ratios of the two components of Sylgard 184 with the resulting elastic moduli of the gels. The rigidity of the silicone gels is less susceptible to effects from drying, swelling, and aging than polyacrylamide gels and can be easily coated with fluorescent tracer particles and with molecules promoting cellular adhesion. This work can lead to broader use of silicone gels in the cell biology laboratory and to improved repeatability and accuracy of cell traction force microscopy and rigidity sensing experiments.

  18. Measurements of elastic moduli of silicone gel substrates with a microfluidic device.

    Directory of Open Access Journals (Sweden)

    Edgar Gutierrez

    Full Text Available Thin layers of gels with mechanical properties mimicking animal tissues are widely used to study the rigidity sensing of adherent animal cells and to measure forces applied by cells to their substrate with traction force microscopy. The gels are usually based on polyacrylamide and their elastic modulus is measured with an atomic force microscope (AFM. Here we present a simple microfluidic device that generates high shear stresses in a laminar flow above a gel-coated substrate and apply the device to gels with elastic moduli in a range from 0.4 to 300 kPa that are all prepared by mixing two components of a transparent commercial silicone Sylgard 184. The elastic modulus is measured by tracking beads on the gel surface under a wide-field fluorescence microscope without any other specialized equipment. The measurements have small and simple to estimate errors and their results are confirmed by conventional tensile tests. A master curve is obtained relating the mixing ratios of the two components of Sylgard 184 with the resulting elastic moduli of the gels. The rigidity of the silicone gels is less susceptible to effects from drying, swelling, and aging than polyacrylamide gels and can be easily coated with fluorescent tracer particles and with molecules promoting cellular adhesion. This work can lead to broader use of silicone gels in the cell biology laboratory and to improved repeatability and accuracy of cell traction force microscopy and rigidity sensing experiments.

  19. Sulfomethylated graft copolymers of xanthan gum and polyacrylamide

    Energy Technology Data Exchange (ETDEWEB)

    Cottrell, I.W.; Empey, R.A.; Racciato, J.S.

    1978-08-08

    A water-soluble anionic graft copolymer of xanthan gum and polyacrylamide is described in which at least part of the amide function of the acrylamide portion of the copolymer is sulfomethylated and the xanthan gum portion of the copolymer is unreacted with formaldehyde. The copolymer is sulfomethylated by reaction with formaldehyde and sodium metabisulfite. The formaldehyde does not cause any appreciable cross-linking between hydroxyl groups of the xanthan moieties. The sulfomethylation of the acrylamido group takes place at temperatures from 35 to 70 C. The pH is 10 or higher, typically from 12 to 13. The degree of anionic character may be varied by adjusting the molar ratio of formaldehyde and sodium metabisulfite with respect to the copolymer. 10 claims.

  20. Polyacrylamide based ICG nanocarriers for enhanced fluorescence and photoacoustic imaging

    Science.gov (United States)

    Ray, Aniruddha; Yoon, Hyung Ki; Ryu, HeeJu; Koo Lee, Yong-Eun; Kim, Gwangseong; Wang, Xueding; Kopelman, Raoul

    2013-02-01

    Indocyanine green (ICG) is an FDA approved tricarbocyanine dye. This dye, with a strong absorbance in the near infrared (NIR) region, has been extensively used for fluorescence and photoacoustic imaging in vivo. ICG in its free form, however, has a few drawbacks that limit its in vivo applications, such as non-targetability, tendency to form aggregates which changes its optical properties, fast degradation, short plasma lifetime and reduced fluorescence at body temperature. In order to bypass these inherent drawbacks, we demonstrate a polyacrylamide based nanocarrier that was particularly designed to carry the negatively charged ICG molecules. These nanocarriers are biodegradable, biocompatible and can be specifically targeted to any cell or tissue. Using these nanocarriers we avoid all the problems associated with free ICG, such as degradation, aggregation and short plasma lifetime, and also enhance demonstrate its ability towards photoacoustics and fluorescence imaging.

  1. Study on synthesis and flocculation property of cation-polyacrylamide

    Institute of Scientific and Technical Information of China (English)

    NIE Rong-chun; GUO Li-ying; XU Chu-yang

    2008-01-01

    On the basis of flocculating settling experimentation on flotation waste coal in Wangfenggang coal preparation plant, influence of medical dosage and cationization (CD) of CPAM samples on coal slurry's flocculating effect was studied, difference of flocculating effect on coal slurry among different categories of polyacrylamide was discussed. Experi-mental results show that when the dosage of flocculant reaches 2~4 g/m3 flotation waste,and the CD of CPAM is 5%, flocculating effect is the best, light transmittance of super-natant liquor reaches 93%. Taking 3types of sample CPAM, PAM and PHP, which formula weight vary a little, to deal with the same concn of coal slurry, when medicine dosage is 3 g/m3, flocculating effect of CPAM is the best, light transmittance of supernatant liquor reaches 92%.

  2. ADDITIVE-INDUCED ENHANCEMENT OF OPTICAL CLARITY OF POLYACRYLAMIDE HYDROGEL

    Institute of Scientific and Technical Information of China (English)

    Jeffery Franklin; Zhi Yuan Wang

    2003-01-01

    The aqueous polymerization of acrylamide and crosslinking with N,N-methylenebisacrylamide afforded hydrogels displaying high levels of light scattering (poor optical clarity). Enhancement of the optical clarity within a polyacrylamide (PAm) hydrogel was accomplished through the implementation of"refractive index matching", Water-soluble additives were utilised to better match the refractive index inhomogeneities throughout a given hydrogel. This resulted in lower light scattering within the system and hence improved clarity. Amino acids, sugars, polymers, and other water-soluble additives such as glycerol were investigated by this methodology. Most additives investigated displayed potential for effectively reducing the light scattering within a PAm hydrogel as a function of increased additive concentration. On increasing the refractive index of the water medium, the overall refractive index of a PAm hydrogel was also observed to increase. This provided a quantitative means of determining the effectiveness of a given additive for improving the optical clarity within a hydrogel.

  3. Heat-denatured lysozyme aggregation and gelation as revealed by combined dielectric relaxation spectroscopy and light scattering measurements.

    Science.gov (United States)

    Giugliarelli, A; Sassi, P; Paolantoni, M; Onori, G; Cametti, C

    2012-09-06

    The dielectric behavior of native and heat-denatured lysozyme in ethanol-water solutions was examined in the frequency range from 1 MHz to 2 GHz, using frequency-domain dielectric relaxation spectroscopy. Because of the conformational changes on unfolding, dielectric methods provide information on the denaturation process of the protein and, at protein concentration high enough, on the subsequent aggregation and gelation. Moreover, the time evolution of the protein aggregation and gelation was monitored measuring, by means of dynamic light scattering methods, the diffusion coefficient of micro-sized polystyrene particles, deliberately added to the protein solution, which act as a probe of the viscosity of the microenvironment close to the particle surface. All together, our measurements indicate that heat-induced denaturation favors, at high concentrations, a protein aggregation process which evolves up to the full gelation of the system. These findings have a direct support from IR measurements of the absorbance of the amide I band that, because of the unfolding, indicate that proteins entangle each other, producing a network structure which evolves, in long time limit, in the gel.

  4. Unusual cold denaturation of a small protein domain.

    Science.gov (United States)

    Buchner, Ginka S; Shih, Natalie; Reece, Amy E; Niebling, Stephan; Kubelka, Jan

    2012-08-21

    A thermal unfolding study of the 45-residue α-helical domain UBA(2) using circular dichroism is presented. The protein is highly thermostable and exhibits a clear cold unfolding transition with the onset near 290 K without denaturant. Cold denaturation in proteins is rarely observed in general and is quite unique among small helical protein domains. The cold unfolding was further investigated in urea solutions, and a simple thermodynamic model was used to fit all thermal and urea unfolding data. The resulting thermodynamic parameters are compared to those of other small protein domains. Possible origins of the unusual cold unfolding of UBA(2) are discussed.

  5. OBSERVATION OF DNA PARTIAL DENATURATION BY ATOMIC FORCE MICROSCOPY

    Institute of Scientific and Technical Information of China (English)

    Xin-hua Dai; Zhi-gang Wang; Bo Xiao; Yong-jun Zhang; Chen Wang; Chun-li Bai; Xiao-li Zhang; Jian Xu

    2004-01-01

    Atomic force microscopy was used to investigate the DNA-cetyltrimethylammonium bromide (CTAB) complexes adsorbed on highly ordered pyrolytic graphite (HOPG). These complexes, at low concentrations, can automatically spread out on the surface of HOPG. The DNA-CTAB complexes display a typically extended structure rather than a globular structure. Partially denaturated DNA produced by binding CTAB to DNA is directly observed by AFM with high resolution.The three-dimensional resolution of partially denaturated DNA obtained by AFM is not available by any other technique at present.

  6. Single-molecule denaturation mapping of DNA in nanofluidic channels

    DEFF Research Database (Denmark)

    Reisner, Walter; Larsen, Niels Bent; Silahtaroglu, Asli

    2010-01-01

    Here we explore the potential power of denaturation mapping as a single-molecule technique. By partially denaturing YOYO (R)-1-labeled DNA in nanofluidic channels with a combination of formamide and local heating, we obtain a sequence-dependent "barcode" corresponding to a series of local dips....... Consequently, the technique is sensitive to sequence variation without requiring enzymatic labeling or a restriction step. This technique may serve as the basis for a new mapping technology ideally suited for investigating the long-range structure of entire genomes extracted from single cells....

  7. Thermoresponsive Gels

    Directory of Open Access Journals (Sweden)

    M. Joan Taylor

    2017-01-01

    Full Text Available Thermoresponsive gelling materials constructed from natural and synthetic polymers can be used to provide triggered action and therefore customised products such as drug delivery and regenerative medicine types as well as for other industries. Some materials give Arrhenius-type viscosity changes based on coil to globule transitions. Others produce more counterintuitive responses to temperature change because of agglomeration induced by enthalpic or entropic drivers. Extensive covalent crosslinking superimposes complexity of response and the upper and lower critical solution temperatures can translate to critical volume temperatures for these swellable but insoluble gels. Their structure and volume response confer advantages for actuation though they lack robustness. Dynamic covalent bonding has created an intermediate category where shape moulding and self-healing variants are useful for several platforms. Developing synthesis methodology—for example, Reversible Addition Fragmentation chain Transfer (RAFT and Atomic Transfer Radical Polymerisation (ATRP—provides an almost infinite range of materials that can be used for many of these gelling systems. For those that self-assemble into micelle systems that can gel, the upper and lower critical solution temperatures (UCST and LCST are analogous to those for simpler dispersible polymers. However, the tuned hydrophobic-hydrophilic balance plus the introduction of additional pH-sensitivity and, for instance, thermochromic response, open the potential for coupled mechanisms to create complex drug targeting effects at the cellular level.

  8. 21 CFR 872.3480 - Polyacrylamide polymer (modified cationic) denture adhesive.

    Science.gov (United States)

    2010-04-01

    ... adhesive. 872.3480 Section 872.3480 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND... polymer (modified cationic) denture adhesive. (a) Identification. A polyacrylamide polymer (modified cationic) denture adhesive is a device composed of polyacrylamide polymer (modified cationic) intended...

  9. Gadolinium-loaded gel scintillators for neutron and antineutrino detection

    Science.gov (United States)

    Riddle, Catherine Lynn; Akers, Douglas William; Demmer, Ricky Lynn; Paviet, Patricia Denise; Drigert, Mark William

    2016-11-29

    A gadolinium (Gd) loaded scintillation gel (Gd-ScintGel) compound allows for neutron and gamma-ray detection. The unique gel scintillator encompasses some of the best features of both liquid and solid scintillators, yet without many of the disadvantages associated therewith. Preferably, the gel scintillator is a water soluble Gd-DTPA compound and water soluble fluorophores such as: CdSe/ZnS (or ZnS) quantum dot (Q-dot) nanoparticles, coumarin derivatives 7-hydroxy-4-methylcoumarin, 7-hydroxy-4-methylcoumarin-3-acetic acid, 7-hydroxycoumarin-3-carboxylic acid, and Alexa Fluor 350 as well as a carbostyril compound, carbostyril 124 in a stable water-based gel, such as methylcellulose or polyacrylamide polymers. The Gd-loaded ScintGel allows for a homogenious distribution of the Gd-DTPA and the fluorophores, and yields clean fluorescent emission peaks. A moderator, such as deuterium or a water-based clear polymer, can be incorporated in the Gd-ScintGel. The gel scintillators can be used in compact detectors, including neutron and antineutrino detectors.

  10. Regioselective immobilization of short oligonucleotides to acrylic copolymer gels.

    Science.gov (United States)

    Timofeev, E; Kochetkova, S V; Mirzabekov, A D; Florentiev, V L

    1996-01-01

    Four types of polyacrylamide or polydimethyl-acrylamide gels for regioselective (by immobilization at the 3' end) of short oligonucleotides have been designed for use in manufacturing oligonucleotide microchips. Two of these supports contain amino or aldehyde groups in the gel, allowing coupling with oligonucleotides bearing aldehyde or amino groups, respectively, in the presence of a reducing agent. The aldehyde gel support showed a higher immobilization efficiency relative to the amino gel. Of all reducing agents tested, the best results were obtained with a pyridine-borane complex. The other supports are based on an acrylamide gel activated with glutaraldehyde or a hydroxyalkyl-functionalized gel treated with mesyl chloride. The use of dimethylacrylamide instead of acrylamide allows subsequent gel modifications in organic solvents. All the immobilization methods are easy and simple to perform, give high and reproducible yields, allow long durations of storage of the activated support, and provide high stability of attachment and low non-specific binding. Although these gel supports have been developed for preparing oligonucleotide microchips, they may be used for other purposes as well. PMID:8774893

  11. Gadolinium-loaded gel scintillators for neutron and antineutrino detection

    Energy Technology Data Exchange (ETDEWEB)

    Riddle, Catherine Lynn; Akers, Douglas William; Demmer, Ricky Lynn; Paviet, Patricia Denise; Drigert, Mark William

    2016-11-29

    A gadolinium (Gd) loaded scintillation gel (Gd-ScintGel) compound allows for neutron and gamma-ray detection. The unique gel scintillator encompasses some of the best features of both liquid and solid scintillators, yet without many of the disadvantages associated therewith. Preferably, the gel scintillator is a water soluble Gd-DTPA compound and water soluble fluorophores such as: CdSe/ZnS (or ZnS) quantum dot (Q-dot) nanoparticles, coumarin derivatives 7-hydroxy-4-methylcoumarin, 7-hydroxy-4-methylcoumarin-3-acetic acid, 7-hydroxycoumarin-3-carboxylic acid, and Alexa Fluor 350 as well as a carbostyril compound, carbostyril 124 in a stable water-based gel, such as methylcellulose or polyacrylamide polymers. The Gd-loaded ScintGel allows for a homogenious distribution of the Gd-DTPA and the fluorophores, and yields clean fluorescent emission peaks. A moderator, such as deuterium or a water-based clear polymer, can be incorporated in the Gd-ScintGel. The gel scintillators can be used in compact detectors, including neutron and antineutrino detectors.

  12. Denaturation of membrane proteins and hyperthermic cell killing

    NARCIS (Netherlands)

    Burgman, Paulus Wilhelmus Johannes Jozef

    1993-01-01

    Summarizing: heat induced denaturation of membrane proteins is probably related to hyperthermic cell killing. Induced resistance of heat sensitive proteins seems to be involved in the development of thermotolerance. Although many questions remain still to be answered, it appears that HSP72, when

  13. Protein denaturation of whey protein isolates (WPIs) induced by high intensity ultrasound during heat gelation.

    Science.gov (United States)

    Frydenberg, Rikke P; Hammershøj, Marianne; Andersen, Ulf; Greve, Marie T; Wiking, Lars

    2016-02-01

    In this study, the impact of high intensity ultrasound (HIU) on proteins in whey protein isolates was examined. Effects on thermal behavior, secondary structure and nature of intra- and intermolecular bonds during heat-induced gelling were investigated. Ultrasonication (24 kHz, 300 W/cm(2), 2078 J/mL) significantly reduced denaturation enthalpies, whereas no change in secondary structure was detected by circular dichroism. The thiol-blocking agent N-ethylmaleimide was applied in order to inhibit formation of disulfide bonds during gel formation. Results showed that increased contents of α-lactalbumin (α-La) were associated with increased sensitivity to ultrasonication. The α-La:β-lactoglobulin (β-Lg) ratio greatly affected the nature of the interactions formed during gelation, where higher amounts of α-La lead to a gel more dependent on disulfide bonds. These results contribute to clarifying the mechanisms mediating the effects of HIU on whey proteins on the molecular level, thus moving further toward implementing HIU in the processing chain in the food industry. Copyright © 2015 Elsevier Ltd. All rights reserved.

  14. A novel mechanism of "metal gel-shift" by histidine-rich Ni2+-binding Hpn protein from Helicobacter pylori strain SS1.

    Science.gov (United States)

    Shelake, Rahul Mahadev; Ito, Yuki; Masumoto, Junya; Morita, Eugene Hayato; Hayashi, Hidenori

    2017-01-01

    Sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) is a universally used method for determining approximate molecular weight (MW) in protein research. Migration of protein that does not correlate with formula MW, termed "gel shifting" appears to be common for histidine-rich proteins but not yet studied in detail. We investigated "gel shifting" in Ni2+-binding histidine-rich Hpn protein cloned from Helicobacter pylori strain SS1. Our data demonstrate two important factors determining "gel shifting" of Hpn, polyacrylamide-gel concentration and metal binding. Higher polyacrylamide-gel concentrations resulted in faster Hpn migration. Irrespective of polyacrylamide-gel concentration, preserved Hpn-Ni2+ complex migrated faster (3-4 kDa) than apo-Hpn, phenomenon termed "metal gel-shift" demonstrating an intimate link between Ni2+ binding and "gel shifting". To examine this discrepancy, eluted samples from corresponding spots on SDS-gel were analyzed by matrix-assisted laser desorption/ionization-time-of-flight mass spectrometry (MALDI-TOF-MS). The MW of all samples was the same (6945.66±0.34 Da) and identical to formula MW with or without added mass of Ni2+. MALDI-TOF-MS of Ni2+-treated Hpn revealed that monomer bound up to six Ni2+ ions non-cooperatively, and equilibrium between protein-metal species was reliant on Ni2+ availability. This corroborates with gradually increased heterogeneity of apo-Hpn band followed by compact "metal-gel shift" band on SDS-PAGE. In view of presented data metal-binding and "metal-gel shift" models are discussed.

  15. Urea Induced Denaturation of Pre-Q1 Riboswitch

    Science.gov (United States)

    Yoon, Jeseong; Thirumalai, Devarajan; Hyeon, Changbong

    2013-01-01

    Urea, a polar molecule with a large dipole moment, not only destabilizes the folded RNA structures, but can also enhance the folding rates of large ribozymes. Unlike the mechanism of urea-induced unfolding of proteins, which is well understood, the action of urea on RNA has barely been explored. We performed extensive all atom molecular dynamics (MD) simulations to determine the molecular underpinnings of urea-induced RNA denaturation. Urea displays its denaturing power in both secondary and tertiary motifs of the riboswitch (RS) structure. Our simulations reveal that the denaturation of RNA structures is mainly driven by the hydrogen bonds and stacking interactions of urea with the bases. Through detailed studies of the simulation trajectories, we found that geminate pairs between urea and bases due to hydrogen bonds and stacks persist only ~ (0.1-1) ns, which suggests that urea-base interaction is highly dynamic. Most importantly, the early stage of base pair disruption is triggered by penetration of water molecules into the hydrophobic domain between the RNA bases. The infiltration of water into the narrow space between base pairs is critical in increasing the accessibility of urea to transiently disrupted bases, thus allowing urea to displace inter base hydrogen bonds. This mechanism, water-induced disruption of base-pairs resulting in the formation of a "wet" destabilized RNA followed by solvation by urea, is the exact opposite of the two-stage denaturation of proteins by urea. In the latter case, initial urea penetration creates a dry-globule, which is subsequently solvated by water penetration leading to global protein unfolding. Our work shows that the ability to interact with both water and polar, non-polar components of nucleotides makes urea a powerful chemical denaturant for nucleic acids.

  16. A Gel Probe Equilibrium Sampler for Measuring Arsenic Porewater Profiles And Sorption Gradients in Sediments: I. Laboratory Development

    Energy Technology Data Exchange (ETDEWEB)

    Campbell, K.M.; Root, R.; O' Day, P.A.; Hering, J.G.

    2009-05-14

    A gel probe equilibrium sampler has been developed to study arsenic (As) geochemistry and sorption behavior in sediment porewater. The gels consist of a hydrated polyacrylamide polymer, which has a 92% water content. Two types of gels were used in this study. Undoped (clear) gels were used to measure concentrations of As and other elements in sediment porewater. The polyacrylamide gel was also doped with hydrous ferric oxide (HFO), an amorphous iron (Fe) oxyhydroxide. When deployed in the field, HFO-doped gels introduce a fresh sorbent into the subsurface thus allowing assessment of in situ sorption. In this study, clear and HFO-doped gels were tested under laboratory conditions to constrain the gel behavior prior to field deployment. Both types of gels were allowed to equilibrate with solutions of varying composition and re-equilibrated in acid for analysis. Clear gels accurately measured solution concentrations ({+-}1%), and As was completely recovered from HFO-doped gels ({+-}4%). Arsenic speciation was determined in clear gels through chromatographic separation of the re-equilibrated solution. For comparison to speciation in solution, mixtures of As(III) and As(V) adsorbed on HFO embedded in gel were measured in situ using X-ray absorption spectroscopy (XAS). Sorption densities for As(III) and As(V) on HFO embedded in gel were obtained from sorption isotherms at pH 7.1. When As and phosphate were simultaneously equilibrated (in up to 50-fold excess of As) with HFO-doped gels, phosphate inhibited As sorption by up to 85% and had a stronger inhibitory effect on As(V) than As(III). Natural organic matter (>200 ppm) decreased As adsorption by up to 50%, and had similar effects on As(V) and As(III). The laboratory results provide a basis for interpreting results obtained by deploying the gel probe in the field and elucidating the mechanisms controlling As partitioning between solid and dissolved phases in the environment.

  17. Effect of electron irradiation on the gel properties of Collichthys lucidus surimi

    Science.gov (United States)

    Deng, Siyao; Lv, Liangyu; Yang, Wenge; Xu, Dalun; Lou, Qiaoming; Zhang, Jinjie

    2017-01-01

    This study evaluated the effect of electron irradiation on the gel properties of Collichthys lucidus surimi. The results showed that irradiation decreased the trichloroacetic acid-soluble peptide content of the surimi gel. At 5 kGy, a more compact and ordered gel network structure was achieved, resulting in a higher gel strength, whiteness, and water-holding capacity than non-irradiated surimi gel. During heat-induced formation of the gel, the α-helix content decreased, whilst the β-sheet and β-turn content increased. Irradiation treatments also decreased the α-helix content and increased β-sheet content, and this transformation is beneficial for the protein denaturation and gel formation. Collectively, the results suggest that electron irradiation, at an optimal dose of 5 kGy, could be an effective method for application in the surimi manufacturing industry

  18. Encapsulation efficiency and controlled release characteristics of crosslinked polyacrylamide particles.

    Science.gov (United States)

    Sairam, Malladi; Babu, V Ramesh; Vijaya, Boya; Naidu, Kumar; Aminabhavi, Tejraj M

    2006-08-31

    Polyacrylamide (pAAm) particles crosslinked with N,N-methylenebis-acrylamide/ethylene glycol dimethacrylate (NNMBA/EGDMA) have been prepared in water-methanol medium by the dispersion polymerization using poly(vinyl pyrrolidone), PVP as a steric stabilizer. 5-fluorouracil an anticancer drug, has been loaded in situ into the crosslinked pAAm particles. Plain as well as drug loaded microparticles have been characterized by differential scanning calorimetry (DSC) and X-ray diffraction studies (XRD) and scanning electron microscopy (SEM). DSC and XRD studies have indicated a molecular level dispersion of the drug in pAAm particles during in situ loading and SEM pictures have shown the formation of spherical and oval-shaped particles. In vitro release of 5-fluorouracil from the crosslinked pAAm particles has been carried out in 7.4 pH buffer medium. Both encapsulation efficiency and release patterns are found to depend on the nature of the crosslinking agent, amount of crosslinking agent used and the amount of drug loaded. In vitro release studies indicated the controlled release of 5-fluorouracil up to 12 h.

  19. Polyacrylamide medium for the electrophoretic separation of biomolecules

    Science.gov (United States)

    Madabhushi, Ramakrishna S.; Gammon, Stuart A.

    2003-11-11

    A polyacryalmide medium for the electrophoretic separation of biomolecules. The polyacryalmide medium comprises high molecular weight polyacrylamides (PAAm) having a viscosity average molecular weight (M.sub.v) of about 675-725 kDa were synthesized by conventional red-ox polymerization technique. Using this separation medium, capillary electrophoresis of BigDye DNA sequencing standard was performed. A single base resolution of .about.725 bases was achieved in .about.60 minute in a non-covalently coated capillary of 50 .mu.m i.d., 40 cm effective length, and a filed of 160 V/cm at 40.degree. C. The resolution achieved with this formulation to separate DNA under identical conditions is much superior (725 bases vs. 625 bases) and faster (60 min. vs. 75 min.) to the commercially available PAAm, such as supplied by Amersham. The formulation method employed here to synthesize PAAm is straight-forward, simple and does not require cumbersome methods such as emulsion polymerizaiton in order to achieve very high molecular weights. Also, the formulation here does not require separation of PAAm from the reaction mixture prior to reconstituting the polymer to a final concentration. Furthermore, the formulation here is prepared from a single average mol. wt. PAAm as opposed to the mixture of two different average mo. wt. PAAm previously required to achieve high resolution.

  20. Antimicrobial activity of silver/starch/polyacrylamide nanocomposite.

    Science.gov (United States)

    Abdel-Halim, E S; Al-Deyab, Salem S

    2014-07-01

    A novel silver/starch/polyacrylamide nanocomposite hydrogel was prepared by grafting acrylamide onto starch in presence of silver nitrate by use of ammonium persulphate as an initiator and N,N-methylene-bisacrylamide as a crosslinking agent, then reducing the silver ions enclosed in the hydrogel structure to silver nanoparticles by treating the hydrogel with sodium hydroxide solution. All factors which affect the grafting/crosslinking reaction were optimized and the concentration of silver ion was changed from 0ppm to 50ppm. The produced nanocomposite hydrogel was characterized for its nanosilver content and the UV-spectra showed similar absorption spectra at wavelength 405nm for all AgNO3 concentrations but the plasmon showed increase in the intensity of the absorption peak as AgNO3 concentration incorporated to the hydrogel structure increases. The nanocomposite hydrogel was also characterized for its antimicrobial activity toward two types of bacteria and two types of fungi. The results showed that the hydrogel with 0ppm silver content has no antimicrobial activity, and that the antimicrobial activity expressed as inhibition zone increases as the silver content increases from 5ppm to 50ppm.

  1. Rheological properties of patatin gels compared with ß-lactoglobulin, ovalbumin, and glycinin

    NARCIS (Netherlands)

    Creusot, N.P.; Wierenga, P.A.; Laus, M.C.; Giuseppin, M.L.F.; Gruppen, H.

    2011-01-01

    BACKGROUND: The thermal unfolding and rheological properties of patatin gels were compared with those of commonly used proteins (ß-lactoglobulin, ovalbumin, glycinin). RESULTS: A significant difference between these proteins was observed in both the denaturation temperature (59 °C for patatin; about

  2. Theoretical Study on Effects of Salt and Temperature on Denaturation Transition of Double-stranded DNA

    Institute of Scientific and Technical Information of China (English)

    DONG Rui-Xin; YAN Xun-Ling; PANG Xiao-Feng; JIANG Shan; LIU Sheng-Gang

    2004-01-01

    We investigate the statistical mechanics properties of a nonlinear dynamics model of the denaturation of the DNA double-helix and study the effects of salt concentration and temperature on denaturation transition of DNA. The specific heat, entropy, and denaturation temperature of the system versus salt concentration are obtained. These results show that the denaturation of DNA not only depends on the temperature but also is influenced by the salt concentration in the solution of DNA, which are in agreement with experimental measurement.

  3. Triton X-114 cloud point extraction to subfractionate blood plasma proteins for two-dimensional gel electrophoresis

    DEFF Research Database (Denmark)

    Jessen, Flemming; Wulff, Tune

    2015-01-01

    A simple and reproducible procedure for enrichment of a plasma protein subfraction suitable for two-dimensional polyacrylamide gel electrophoresis (2DE) was developed, using a Triton X-114-based cloud point extraction (CPE). Appropriate conditions for such a CPE procedure were found by SDS...

  4. Stabilization of polymer gels against divalent ion-induced syneresis

    Energy Technology Data Exchange (ETDEWEB)

    Albonico, Paola; Lockhart, Thomas P. [Eniricerche SpA, San Donato, Milan (Italy)

    1997-07-15

    Polymer solutions and polymer gels are unstable to extended ageing in divalent cation-rich brines at elevated temperature. This paper shows that low-molecular-weight compounds that complex strongly with Ca{sup 2+} and Mg{sup 2+} are capable of neutralizing their destabilizing influence on polymer solubility and of inhibiting the syneresis of crosslinked acrylamide polymer gels in hard brines. The solubility of the inhibitor-divalent ion complexes formed in hard brine at elevated temperature have also been examined. The results obtained offer the possibility to extend significantly the upper temperature limit for the use of polyacrylamides and acrylamide copolymers in brines in both polymer flooding and polymer gel treatments

  5. Rapid DNA sequencing by horizontal ultrathin gel electrophoresis.

    Science.gov (United States)

    Brumley, R L; Smith, L M

    1991-01-01

    A horizontal polyacrylamide gel electrophoresis apparatus has been developed that decreases the time required to separate the DNA fragments produced in enzymatic sequencing reactions. The configuration of this apparatus and the use of circulating coolant directly under the glass plates result in heat exchange that is approximately nine times more efficient than passive thermal transfer methods commonly used. Bubble-free gels as thin as 25 microns can be routinely cast on this device. The application to these ultrathin gels of electric fields up to 250 volts/cm permits the rapid separation of multiple DNA sequencing reactions in parallel. When used in conjunction with 32P-based autoradiography, the DNA bands appear substantially sharper than those obtained in conventional electrophoresis. This increased sharpness permits shorter autoradiographic exposure times and longer sequence reads. Images PMID:1870968

  6. How it all began: a personal history of gel electrophoresis.

    Science.gov (United States)

    Smithies, Oliver

    2012-01-01

    Arne Tiselius' moving boundary electrophoresis method was still in general use in 1951 when this personal history begins, although zonal electrophoresis with a variety of supporting media (e.g., filter paper or starch grains) was beginning to replace it. This chapter is an account of 10 years of experiments carried out by the author during which molecular sieving gel electrophoresis was developed and common genetic variants of two proteins, haptoglobin and transferrin, were discovered in normal individuals. Most of the figures are images of pages from the author's laboratory notebooks, which are still available, so that some of the excitement of the time and the humorous moments are perhaps apparent. Alkaline gels, acidic gels with and without denaturants, vertical gels, two-dimensional gels, and gels with differences in starch concentration are presented. The subtle details that can be discerned in these various gels played an indispensable role in determining the nature of the change in the haptoglobin gene (Hp) that leads to the polymeric series characteristic of Hp ( 2 ) /Hp ( 2 ) homozygotes. Where possible, the names of scientific friends who made this saga of gel electrophoresis so memorable and enjoyable are gratefully included.

  7. How water contributes to pressure and cold denaturation of proteins

    CERN Document Server

    Bianco, Valentino

    2015-01-01

    The mechanisms of cold- and pressure-denaturation of proteins are matter of debate and are commonly understood as due to water-mediated interactions. Here we study several cases of proteins, with or without a unique native state, with or without hydrophilic residues, by means of a coarse-grain protein model in explicit solvent. We show, using Monte Carlo simulations, that taking into account how water at the protein interface changes its hydrogen bond properties and its density fluctuations is enough to predict protein stability regions with elliptic shapes in the temperature-pressure plane, consistent with previous theories. Our results clearly identify the different mechanisms with which water participates to denaturation and open the perspective to develop advanced computational design tools for protein engineering.

  8. Influence of Ficoll on urea induced denaturation of fibrinogen

    Science.gov (United States)

    Sankaranarayanan, Kamatchi; Meenakshisundaram, N.

    2016-03-01

    Ficoll is a neutral, highly branched polymer used as a molecular crowder in the study of proteins. Ficoll is also part of Ficoll-Paque used in biology laboratories to separate blood to its components (erythrocytes, leukocytes etc.,). Role of Ficoll in the urea induced denaturation of protein Fibrinogen (Fg) has been analyzed using fluorescence, circular dichroism, molecular docking and interfacial studies. Fluorescence studies show that Ficoll prevents quenching of Fg in the presence of urea. From the circular dichroism spectra, Fg shows conformational transition to random coil with urea of 6 M concentration. Ficoll helps to shift this denaturation concentration to 8 M and thus constraints by shielding Fg during the process. Molecular docking studies indicate that Ficoll interacts favorably with the protein than urea. The surface tension and shear viscosity analysis shows clearly that the protein is shielded by Ficoll.

  9. Characterization and distribution of esterase activity in activated sludge

    NARCIS (Netherlands)

    Boczar, BA; Forney, LJ; Begley, WM; Larson, RJ; Federle, TW

    2001-01-01

    The location and activity of esterase enzymes in activated Sludge from three Municipal wastewater treatment plants were characterized using model Substrate, and denaturing and nondenaturing polyacrylamide gel electrophoresis (PAGE) Of particulate, freeze thaw (primarily periplasmic enzymes and those

  10. Mapping and identification of interferon gamma-regulated HeLa cell proteins separated by immobilized pH gradient two-dimensional gel electrophoresis

    DEFF Research Database (Denmark)

    Shaw, AC; Rossel Larsen, M; Roepstorff, P

    1999-01-01

    magnitude of IFN-gamma responsive genes has been reported previously. Our goal is to identify and map IFN-gamma-regulated HeLa cell proteins to the two-dimensional polyacrylamide gel electrophoresis with the immobilized pH gradient (IPG) two-dimensional polyacrylamide gel electrophoresis (2-D PAGE) system....... A semiconfluent layer of HeLa cells was grown on tissue culture plates, and changes in protein expression due to 100 U/mL IFN-gamma were investigated at different periods after treatment, using pulse labeling with [35S]methionine/cysteine in combination with 2-D PAGE (IPG). The identity of eight protein spots...

  11. Temperature induced denaturation of collagen in acidic solution.

    Science.gov (United States)

    Mu, Changdao; Li, Defu; Lin, Wei; Ding, Yanwei; Zhang, Guangzhao

    2007-07-01

    The denaturation of collagen solution in acetic acid has been investigated by using ultra-sensitive differential scanning calorimetry (US-DSC), circular dichroism (CD), and laser light scattering (LLS). US-DSC measurements reveal that the collagen exhibits a bimodal transition, i.e., there exists a shoulder transition before the major transition. Such a shoulder transition can recover from a cooling when the collagen is heated to a temperature below 35 degrees C. However, when the heating temperature is above 37 degrees C, both the shoulder and major transitions are irreversible. CD measurements demonstrate the content of triple helix slowly decreases with temperature at a temperature below 35 degrees C, but it drastically decreases at a higher temperature. Our experiments suggest that the shoulder transition and major transition arise from the defibrillation and denaturation of collagen, respectively. LLS measurements show the average hydrodynamic radius R(h), radius of gyration R(g)of the collagen gradually decrease before a sharp decrease at a higher temperature. Meanwhile, the ratio R(g)/R(h) gradually increases at a temperature below approximately 34 degrees C and drastically increases in the range 34-40 degrees C, further indicating the defibrillation of collagen before the denaturation.

  12. [DNA degradation during standard alkaline of thermal denaturation].

    Science.gov (United States)

    Drozhdeniuk, A P; Sulimova, G E; Vaniushin, B F

    1976-01-01

    Essential degradation 8 DNA (up to 10 per cent) with liberation of acid-soluble fragments takes place on the standard alkaline (0,01 M sodium phosphate, pH 12, 60 degrees, 15 min) or thermal (0.06 M sodium phosphate buffer, pH 6.8, 102 degrees C, 15 min) denaturation. This degradation is more or less selective: fraction of low molecular weight fragments, isolated by hydroxyapatite cromatography and eluted by 0.06 M sodium phosphate buffer, pH 6.8 is rich in adenine and thymine and contains about 2 times less 5-methylcytosine than the total wheat germ DNA. The degree of degradation of DNA on thermal denaturation is higher than on alkaline degradation. Therefore while studying reassociation of various DNA, one and the same standard method of DNA denaturation should be used. Besides, both the level of DNA degradation and the nature of the resulting products (fragments) should be taken into account.

  13. NADP-specific isocitrate dehydrogenase of Escherichia coli. IV. Purification by chromatography on Affi-Gel Blue.

    Science.gov (United States)

    Vasquez, B; Reeves, H C

    1979-05-23

    Affinity chromatography on Affi-Gel Blue has been used to purify the NADP-specific isocitrate dehydrogenase (EC 1.1.1.42) from Escherichia coli. The protocol permits rapid purification of the enzyme in milligram quantities with a yield of 50% and is carried out almost entirely at room temperature. The preparation was judged to be homogeneous by non-denaturing electrophoresis at pH 7.5 and denaturing electrophoresis in the presence of sodium dodecyl sulfate. The subunit molecular weight of 53 000, determined by sodium dodecyl sulfate gel electrophoresis, is in reasonable agreement with the value of 46 900 estimated from the amino acid composition data.

  14. Bioactive polyacrylamide hydrogels with gradients in mechanical stiffness.

    Science.gov (United States)

    Diederich, Vincent E G; Studer, Peter; Kern, Anita; Lattuada, Marco; Storti, Giuseppe; Sharma, Ram I; Snedeker, Jess G; Morbidelli, Massimo

    2013-05-01

    We propose a novel, single step method for the production of polyacrylamide hydrogels with a gradient in mechanical properties. In contrast to already existing techniques such as UV photo-polymerization with photomasks (limited penetration depth) or microfluidic gradient mixers (complex microfluidic chip), this technique is not suffering such limitations. Young's modulus of the hydrogels was varied by changing the total monomer concentration of the hydrogel precursor solution. Using programmable syringe pumps, the total monomer concentration in the solution fed to the hydrogel mold was varied from 16 wt% down to 5 wt% over the feeding time to obtain a gradient in compliance ranging from 150 kPa down to 20 kPa over a length of 10 mm down to 2.5 mm. Polymerization was achieved with the dual initiation system composed of ammonium persulfate and N,N,N',N'-tetramethylethylenediamine, which were both fed through separate capillaries to avoid premature polymerization. Functionalized with the model ligand collagen I, the substrates were bioactive and supported the attachment of human foreskin fibroblasts (around 30% of the cells seeded attached after 1 h). A kinetic morphology study on homogeneous hydrogels of different stiffness's indicated that fibroblasts tend to spread to their final size within 2 h on stiff substrates, while the spreading time was much longer (ca. 4-5 h) on soft substrates. These trends were confirmed on hydrogels with compliance gradients, showing well spread fibroblasts on the stiff end of the hydrogel after 2 h, while the cells on the soft end still had small area and rounded morphology.

  15. Stability of polyacrylamide solutions in the presence of carbon dioxide

    Energy Technology Data Exchange (ETDEWEB)

    Lakatos, I.; Lakatos, S.J.

    1982-01-01

    The methods of modifying the reservoir in order to intensify oil output include treating with polymers, and also injecting carbon dioxide into the bed. The effect of their total modification significantly depends on lithology of the reservoir, composition of the fluids and thermobaric conditions. Results are presented of laboratory studies on the stability of polyacrylamide solutions. Five types of polymer were used which are distinguished by the magnitude of average molecular weight. The laboratory unit had a core sample holder in which the sample of sandstone was placed 30 x 30 mm, porosity 30.21-30.23 and permeability 0.18-0.27 m/sup 2/. The duration of the measurements reached 40 h. The purity of CO/sub 2/ was 99.98%. Destruction of the polymer (degradation) was determined according to the change in viscosity of its solutions. Results are presented in the graphic form for a vast series of experiments on degradation of polymers of different stage of hydrolysis depending on the following factors: temperature and pressure in the system, composition of polymer and concentration of CO/sub 2/, duration of the effect, mineralization of the aqueous solution and concentration of Ca and Mg ions in them. The following results were obtained: 1) degradation of the polymer intensifies in the presence of CO/sub 2/, 2) stability of the polymer has a weak dependence on molecular weight and strong dependence on the stage of hydrolysis, 3) stability of the polymer diminishes with a rise in temperature and rises with an increase in mineralization of the pore waters; 4) effect of oil saturation and lithology of the rock of stability of the polymer is not noted.

  16. Check dam and polyacrylamide performance under simulated stormwater runoff.

    Science.gov (United States)

    Kang, Jihoon; McCaleb, Melanie M; McLaughlin, Richard A

    2013-11-15

    High levels of turbidity and fine suspended sediments are often found in stormwater discharges from construction sites even when best management practices (BMPs) for sediment control are in place. This study evaluated turbidity reduction by three check dam types: 1) rock check dam representing a standard BMP, 2) excelsior wattle representing a fiber check dam (FCD), and 3) rock check dam wrapped with excelsior erosion control blanket (rock + excelsior ECB) representing an alternative FCD. Three check dams (all same type) were installed in a lined, 24-m ditch on a 5-7% slope and three consecutive simulated stormwater flows were run in the ditch. Additional tests were performed by adding granular polyacrylamide (PAM) on the check dams in the same manner using two sediment sources differing in clay content. Without PAM treatment, significantly higher effluent turbidity (>900 nephelometric turbidity units (NTU)) exited the ditch with rock check dams than with excelsior wattles or rock + excelsior ECBs (dam types was in the order of excelsior wattle > rock + excelsior ECB > rock check dam, indicating better water pooling behind the wattle. The PAM treatment reduced turbidity substantially (>75% relative to no PAM treatment) for all check dam types and it was very effective in excelsior wattles (<57 NTU) and rock + excelsior ECBs (<90 NTU) even during the third storm event. This study demonstrates that the passive treatment of runoff with PAM on FCDs (or rock + excelsior ECB) in construction site ditches can be very effective for sediment retention and turbidity reduction.

  17. Corrosion inhibition of iron in hydrochloric acid by polyacrylamide

    Directory of Open Access Journals (Sweden)

    DRAGICA CHAMOVSKA

    2007-07-01

    Full Text Available The corrosion protection and/or adsorption of polyacrylamide (PAA of number average molecular weight, , between 15,000 – 1,350,000 g mol-1 on mild steel and iron (99.99 % Fe in 3 M HCl at room temperature was studied using spectrophotometry (the phenanthroline method, the weight loss method and EIS (Electrochemical Impedance Spectroscopy. It was found that the corrosion protect­tion efficiency of the PAA – adsorbed layers strongly depends on both the molar concentration of PAA in the solution and its molecular weight, reaching limiting values between 85 and 96 %. Simultaneously, it was also concluded that a relatively high surface coverage could be obtained with very low PAA concentrations (0.5 – 2 ppm, indicating the good adsorption characteristics of PAA on mild steel and iron in hydrochloride acid. The experimentally obtained results follow a Lan­gmuir adsorption isotherm. According to the best fitting parameters, the adsorption coef­f­i­cient B ranged between 2×107 and 4×108 mol-1 and depended strongly on the mole­cular weight of the PAA: B = k (for a ≈ 0.67 and k = 2.95×104 or the size of the polymer coil. As was found by EIS, the thickness of the adsorbed PAA layer was approx. 1.1 nm (for er = 15 and corresponded only to the polymer segments attached to the metal surface. On the other hand, as was found by ellipsometry, the limiting layer of the adsorbed PAA molecules was highly voluminous and relatively thick (100 – 200 nm, containing entangled polymer coils.

  18. Protein's native state stability in a chemically induced denaturation mechanism.

    Science.gov (United States)

    Olivares-Quiroz, L; Garcia-Colin, L S

    2007-05-21

    In this work, we present a generalization of Zwanzig's protein unfolding analysis [Zwanzig, R., 1997. Two-state models of protein folding kinetics. Proc. Natl Acad. Sci. USA 94, 148-150; Zwanzig, R., 1995. Simple model of protein folding kinetics. Proc. Natl Acad. Sci. USA 92, 9801], in order to calculate the free energy change Delta(N)(D)F between the protein's native state N and its unfolded state D in a chemically induced denaturation. This Extended Zwanzig Model (EZM) is both based on an equilibrium statistical mechanics approach and the inclusion of experimental denaturation curves. It enables us to construct a suitable partition function Z and to derive an analytical formula for Delta(N)(D)F in terms of the number K of residues of the macromolecule, the average number nu of accessible states for each single amino acid and the concentration C(1/2) where the midpoint of the ND transition occurs. The results of the EZM for proteins where chemical denaturation follows a sigmoidal-type profile, as it occurs for the case of the T70N human variant of lysozyme (PDB code: T70N) [Esposito, G., et al., 2003. J. Biol. Chem. 278, 25910-25918], can be splitted into two lines. First, EZM shows that for sigmoidal denaturation profiles, the internal degrees of freedom of the chain play an outstanding role in the stability of the native state. On the other hand, that under certain conditions DeltaF can be written as a quadratic polynomial on concentration C(1/2), i.e., DeltaF approximately aC(1/2)(2)+bC(1/2)+c, where a,b,c are constant coefficients directly linked to protein's size K and the averaged number of non-native conformations nu. Such functional form for DeltaF has been widely known to fit experimental measures in chemically induced protein denaturation [Yagi, M., et al., 2003. J. Biol. Chem. 278, 47009-47015; Asgeirsson, B., Guojonsdottir, K., 2006. Biochim. Biophys. Acta 1764, 190-198; Sharma, S., et al., 2006. Protein Pept. Lett. 13(4), 323-329; Salem, M., et al

  19. Chemical gel barriers as low-cost alternative to containment and in situ cleanup of hazardous wastes to protect groundwater

    Energy Technology Data Exchange (ETDEWEB)

    NONE

    1997-01-01

    Chemical gel barriers are being considered as a low-cost alternative for containment and in situ cleanup of hazardous wastes to protect groundwater. Most of the available gels in petroleum application are non-reactive and relative impermeable, providing a physical barriers for all fluids and contaminants. However, other potential systems can be envisioned. These systems could include gels that are chemically reactive and impermeable such that most phase are captured by the barriers but the contaminants could diffuse through the barriers. Another system that is chemically reactive and permeable could have potential applications in selectivity capturing contaminants while allowing water to pass through the barriers. This study focused on chemically reactive and permeable gel barriers. The gels used in experiment are DuPont LUDOX SM colloidal silica gel and Pfizer FLOPAAM 1330S hydrolyzed polyacrylamide (HPAM) gel.

  20. A study of the stability of polyacrylamide solutions under laboratory and field conditions

    Energy Technology Data Exchange (ETDEWEB)

    Lakatos, I.; Kissne, G.M.; Lakatosne, S.J.

    1980-01-01

    Questions are examined of thermal, mechanical and microbiological stability of polyacrylamide solutions used in the processes of oil expulsion and formation treatment. Results are given from experiments performed with nonhydrolyzed and partially hydrolyzed domestic and foreign polyacrylamides under laboratory and field conditions. Attention is drawn to the fact that problems of stability are varied. The economic aspect of field use of the processes must not be underestimated. Stability of polymers can be ensured by effective chemical microbiological protection.

  1. Establishment of in vitro models of denatured collagen%变性胶原体外培养模型的建立

    Institute of Scientific and Technical Information of China (English)

    苏荣家; 王志勇; 刘英开; 原博; 王西樵; 董叫云; 宋菲; 姜育智; 陆树良

    2013-01-01

    目的 探讨不同温度对Ⅰ型胶原分子二级结构的影响,确定合适的胶原变性温度,研究热变性后胶原纤维排列及三维凝胶性质的改变,比较胶原变性后不同培养环境成纤维细胞形态差异,以建立变性胶原-细胞体外培养模型. 方法 Ⅰ型胶原蛋白溶液在不同温度作用后通过蛋白质圆二色光谱仪分析胶原分子二级结构改变.扫描探针显微镜观察胶原变性后纤维结构的改变.制备不同种类三维胶原凝胶并通过气相压力仪检测胶原凝胶断裂模量.将变性后的胶原进行二维包被和三维胶原凝胶制作,倒置相差显微镜及光镜下观察不同培养环境下细胞形态变化. 结果 温度达到50℃时,Ⅰ型胶原分子二级结构发生明显改变,在二维胶原包被时可见胶原纤维凝集成团,含变性胶原的三维凝胶断裂模量明显下降.在变性胶原存在环境中培养成纤维细胞,细胞形态均有显著改变. 结论 经50℃作用后Ⅰ型胶原分子二级结构发生明显改变,含变性胶原的三维凝胶断裂模量明显下降,Ⅰ型胶原包被及三维凝胶模型培养的成纤维细胞形态明显不同,可作为变性胶原影响细胞生物学活性的体外模型.%Objective To investigate influence of different temperatures on secondary structure of type Ⅰ collagen,determine the proper temperature for collagen denaturation,observe changes of collagen fibre arrangement and three dimensional collagen gel properties after thermal denaturation,compare morphological variation of fibroblasts seeded in mediums with denatured collagen and therefore establish a standardized culture model with denatured collagen in vitro.Methods Changes of the secondary structure of type Ⅰ collagen was measured by circular dichroism spectrameter after the collagen solution had been treated with different temperatures.Changes of the fibre structure after collagen denaturation were observed by scanning probe

  2. Detection of bacteriophage phi 6 minus-strand RNA and novel mRNA isoconformers synthesized in vivo and in vitro, by strand-separating agarose gels

    Energy Technology Data Exchange (ETDEWEB)

    Pagratis, N.; Revel, H.R. (Univ. of Chicago, IL (USA))

    1990-07-01

    Two urea-free agarose gel protocols that resolve the six individual strands of bacteriophage phi 6 dsRNA were developed and used to analyze phage RNA synthesis in vivo and in vitro. Citrate gels separate strands of the large and medium chromosomes while Tris-borate-EDTA (TBE) gels resolve the medium and small dsRNA segments. Minus strands migrate faster than plus strands on citrate gels but are retarded on TBE gels. A study of electrophoretic conditions showed that pH affects strand resolution on citrate gels, and that voltage gradient, agarose concentration, and ethidium bromide significantly alter strand migration on TBE gels. Analysis of native phi 6 RNA synthesized in vivo and in vitro showed that the large and medium message RNAs comigrate with the corresponding plus strands of denatured virion dsRNA. The small messenger RNA is exceptional. Native small mRNA was detected as three isoconformers in vivo and in vitro. The isoconformers were converted by heat denaturation to a single RNA species that comigrates with the virion s+ strand. Minus strands labeled in vivo were detected only after heat denaturation. Minus strand synthesis was detected also in heat-denatured samples from in vitro phi 6 nucleocapsid RNA polymerase reactions at pH values suboptimal for transcription.

  3. A comparative in vitro study of the digestibility of heat- and high pressure-induced gels prepared from industrial milk whey proteins

    Science.gov (United States)

    He, Jin-Song; Mu, Tai-Hua; Wang, Juan

    2013-06-01

    We undertook this study to compare the digestibility of heat- and high pressure-induced gels produced from whey protein isolate (WPI). To simulate in vivo gastrointestinal digestion of WPI gels, a pepsin-trypsin digestion system was used. The in vitro protein digestibility of WPI gels induced by high pressure (400 MPa and 30 min; P-gel) and those induced by heat (80°C and 30 min; H-gel) was compared using a protein concentration of 0.14 g mL-1. The in vitro protein digestibility of P-gels was significantly greater than that of H-gels (p<0.05). The size-exclusion chromatography profiles of the hydrolysates showed that the P-gel generated more and smaller peptides than natural WPI and H-gels. Furthermore, Sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis showed some soluble disulfide-mediated aggregation in the P-gel, while there was more insoluble aggregation in the H-gel than the P-gel. The P-gel was more sensitive to proteinase than the H-gel, which was related to the content of S-S bonds, and this in turn could be attributed to the differences in the gelation mechanism between the H-gel and P-gel.

  4. Comparative Skeletal Muscle Proteomics Using Two-Dimensional Gel Electrophoresis.

    Science.gov (United States)

    Murphy, Sandra; Dowling, Paul; Ohlendieck, Kay

    2016-09-09

    The pioneering work by Patrick H. O'Farrell established two-dimensional gel electrophoresis as one of the most important high-resolution protein separation techniques of modern biochemistry (Journal of Biological Chemistry1975, 250, 4007-4021). The application of two-dimensional gel electrophoresis has played a key role in the systematic identification and detailed characterization of the protein constituents of skeletal muscles. Protein changes during myogenesis, muscle maturation, fibre type specification, physiological muscle adaptations and natural muscle aging were studied in depth by the original O'Farrell method or slightly modified gel electrophoretic techniques. Over the last 40 years, the combined usage of isoelectric focusing in the first dimension and sodium dodecyl sulfate polyacrylamide slab gel electrophoresis in the second dimension has been successfully employed in several hundred published studies on gel-based skeletal muscle biochemistry. This review focuses on normal and physiologically challenged skeletal muscle tissues and outlines key findings from mass spectrometry-based muscle proteomics, which was instrumental in the identification of several thousand individual protein isoforms following gel electrophoretic separation. These muscle-associated protein species belong to the diverse group of regulatory and contractile proteins of the acto-myosin apparatus that forms the sarcomere, cytoskeletal proteins, metabolic enzymes and transporters, signaling proteins, ion-handling proteins, molecular chaperones and extracellular matrix proteins.

  5. Comparative Skeletal Muscle Proteomics Using Two-Dimensional Gel Electrophoresis

    Directory of Open Access Journals (Sweden)

    Sandra Murphy

    2016-09-01

    Full Text Available The pioneering work by Patrick H. O’Farrell established two-dimensional gel electrophoresis as one of the most important high-resolution protein separation techniques of modern biochemistry (Journal of Biological Chemistry 1975, 250, 4007–4021. The application of two-dimensional gel electrophoresis has played a key role in the systematic identification and detailed characterization of the protein constituents of skeletal muscles. Protein changes during myogenesis, muscle maturation, fibre type specification, physiological muscle adaptations and natural muscle aging were studied in depth by the original O’Farrell method or slightly modified gel electrophoretic techniques. Over the last 40 years, the combined usage of isoelectric focusing in the first dimension and sodium dodecyl sulfate polyacrylamide slab gel electrophoresis in the second dimension has been successfully employed in several hundred published studies on gel-based skeletal muscle biochemistry. This review focuses on normal and physiologically challenged skeletal muscle tissues and outlines key findings from mass spectrometry-based muscle proteomics, which was instrumental in the identification of several thousand individual protein isoforms following gel electrophoretic separation. These muscle-associated protein species belong to the diverse group of regulatory and contractile proteins of the acto-myosin apparatus that forms the sarcomere, cytoskeletal proteins, metabolic enzymes and transporters, signaling proteins, ion-handling proteins, molecular chaperones and extracellular matrix proteins.

  6. Collagen thermal denaturation study for thermal angioplasty based on modified kinetic model: relation between the artery mechanical properties and collagen denaturation rate

    Science.gov (United States)

    Shimazaki, N.; Hayashi, T.; Kunio, M.; Arai, T.

    2010-02-01

    We have been developing the novel short-term heating angioplasty in which sufficient artery lumen-dilatation was attained with thermal softening of collagen fiber in artery wall. In the present study, we investigated on the relation between the mechanical properties of heated artery and thermal denaturation fractures of arterial collagen in ex vivo. We employed Lumry-Eyring model to estimate temperature- and time-dependent thermal denaturation fractures of arterial collagen fiber during heating. We made a kinetic model of arterial collagen thermal denaturation by adjustment of K and k in this model, those were the equilibrium constant of reversible denaturation and the rate constant of irreversible denaturation. Meanwhile we demonstrated that the change of reduced scattering coefficient of whole artery wall during heating reflected the reversible denaturation of the collagen in artery wall. Based on this phenomenon, the K was determined experimentally by backscattered light intensity measurement (at 633nm) of extracted porcine carotid artery during temperature elevation and descending (25°C-->80°C-->25°C). We employed the value of according to our earlier report in which the time-and temperature- dependent irreversible denaturation amount of the artery collagen fiber that was assessed by the artery birefringence. Then, the time- and temperature- dependent reversible (irreversible) denaturation fraction defined as the reversible ((irreversible) denatured collagen amount) / (total collagen amount) was calculated by the model. Thermo-mechanical analysis of artery wall was performed to compare the arterial mechanical behaviors (softening, shrinkage) during heating with the calculated denaturation fraction with the model. In any artery temperature condition in 70-80°, the irreversible denaturation fraction at which the artery thermal shrinkage started was estimated to be around 20%. On the other hand, the calculated irreversible denaturation fraction remained below

  7. Denatured ethanol release into gasoline residuals, Part 1: Source behaviour

    Science.gov (United States)

    Freitas, Juliana G.; Barker, James F.

    2013-05-01

    With the increasing use of ethanol in fuels, it is important to evaluate its fate when released into the environment. While ethanol is less toxic than other organic compounds present in fuels, one of the concerns is the impact ethanol might have on the fate of gasoline hydrocarbons in groundwater. One possible concern is the spill of denatured ethanol (E95: ethanol containing 5% denaturants, usually hydrocarbons) in sites with pre-existing gasoline contamination. In that scenario, ethanol is expected to increase the mobility of the NAPL phase by acting as a cosolvent and decreasing interfacial tension. To evaluate the E95 behaviour and its impacts on pre-existing gasoline, a field test was performed at the CFB-Borden aquifer. Initially gasoline contamination was created releasing 200 L of E10 (gasoline with 10% ethanol) into the unsaturated zone. One year later, 184 L of E95 was released on top of the gasoline contamination. The site was monitored using soil cores, multilevel wells and one glass access tube. At the end of the test, the source zone was excavated and the compounds remaining were quantified. E95 ethanol accumulated and remained within the capillary fringe and unsaturated zone for more than 200 days, despite ~ 1 m oscillations in the water table. The gasoline mobility increased and it was redistributed in the source zone. Gasoline NAPL saturations in the soil increased two fold in the source zone. However, water table oscillations caused a separation between the NAPL and ethanol: NAPL was smeared and remained in deeper positions while ethanol moved upwards following the water table rise. Similarly, the E95 denaturants that initially were within the ethanol-rich phase became separated from ethanol after the water table oscillation, remaining below the ethanol rich zone. The separation between ethanol and hydrocarbons in the source after water table oscillation indicates that ethanol's impact on hydrocarbon residuals is likely limited to early times.

  8. Strategies for denaturing the weapons-grade plutonium stockpile

    Energy Technology Data Exchange (ETDEWEB)

    Buckner, M.R.; Parks, P.B.

    1992-10-01

    In the next few years, approximately 50 metric tons of weapons-grade plutonium and 150 metric tons of highly-enriched uranium (HEU) may be removed from nuclear weapons in the US and declared excess. These materials represent a significant energy resource that could substantially contribute to our national energy requirements. HEU can be used as fuel in naval reactors, or diluted with depleted uranium for use as fuel in commercial reactors. This paper proposes to use the weapons-grade plutonium as fuel in light water reactors. The first such reactor would demonstrate the dual objectives of producing electrical power and denaturing the plutonium to prevent use in nuclear weapons.

  9. RNA Denaturation: Excluded Volume, Pseudoknots, and Transition Scenarios

    Science.gov (United States)

    Baiesi, M.; Orlandini, E.; Stella, A. L.

    2003-11-01

    A lattice model of RNA denaturation which fully accounts for the excluded volume effects among nucleotides is proposed. A numerical study shows that interactions forming pseudoknots must be included in order to get a sharp continuous transition. Otherwise a smooth crossover occurs from the swollen linear polymer behavior to highly ramified, almost compact conformations with secondary structures. In the latter scenario, which is appropriate when these structures are much more stable than pseudoknot links, probability distributions for the lengths of both loops and main branches obey scaling with nonclassical exponents.

  10. DSC study of cold and heat denaturation processes of β-lactoglobulin A with guanidine hydrochloride

    Institute of Scientific and Technical Information of China (English)

    王邦宁; 谈夫

    1997-01-01

    The cold and heat denaturations of bovine β-lactoglobuhn A (β-lg A) has been studied in solutions of guanidine hydrochloride (GuHCl) by differential scanning calorimelry (DSC) The experimental results are presented and discussed.It is shown that the number of protons bound by the monomeric molecules of β-lg A was unchanged before and after its heat denaturation below pH 3,and that the activation energy of the heat denaturation was depressed owing to the presence of GuHCl.In the solutions with 2.50 and 3.06 mol/L of GuHCl,both the cold and heat denat-urations of β-lg A were observed.In comparison with the heat denaturation,the activation energy of cold denaturation was far lower and the number of GuHCl molecules bound by the unfolded polypeptide chains after cold denaturation increased a lot.The absolute value of the enthalpy of cold denaturation was larger than that of heat denaturation It was found by the analysis that the contribution to the total denaturational enthalpy of conformational change i

  11. DEVELOPMENT OF POLYMER GEL SYSTEMS TO IMPROVE VOLUMETRIC SWEEP AND REDUCE PRODUCING WATER/OIL RATIOS

    Energy Technology Data Exchange (ETDEWEB)

    G. Paul Willhite; Stan McCool; Don W. Green; Min Cheng; Rajeev Jain; Tuan Nguyen

    2003-11-01

    Gelled polymer treatments are applied to oil reservoirs to increase oil production and to reduce water production by altering the fluid movement within the reservoir. This report describes the results of the first year of a three-year research program that is aimed at the understanding of the chemistry of gelation and the fundamental mechanisms that alter the flows of oil and water in reservoir rocks after a gel treatment. Work has focused on a widely-applied system in field applications, the partially hydrolyzed polyacrylamide-chromium acetate gel. Gelation occurs by network formation through the crosslinking of polyacrylamide molecules as a result of reaction with chromium acetate. The initial reaction between chromium acetate and one polymer is referred to as the uptake reaction. The uptake reaction was studied as functions of chromium and polymer concentrations and pH values. Experimental data were regressed to determine a rate equation that describes the uptake reaction of chromium by polyacrylamide. Pre-gel aggregates form and grow as the reactions between chromium acetate and polyacrylamide proceed. A statistical model that describes the growth of pre-gel aggregates was developed using the theory of branching processes. The model gives molecular weight averages that are expressed as functions of the conversion of the reactive sites on chromium acetate or on the polymer molecule. Results of the application of the model correlate well with experimental data of viscosity and weight-average molecular weight and gives insights into the gelation process. A third study addresses the flow of water and oil in rock material after a gel treatment. Previous works have shown that gel treatments usually reduce the permeability to water to a greater extent than the permeability to oil is reduced. This phenomenon is referred to as disproportionate permeability reduction (DPR). Flow experiments were conducted to determine the effect of polymer and chromium concentrations on

  12. Testosterone Nasal Gel

    Science.gov (United States)

    Testosterone nasal gel is used to treat symptoms of low testosterone in men who have hypogonadism (a condition in which the ... does not produce enough natural testosterone). Testosterone nasal gel is used only for men with low testosterone ...

  13. Rapid (ten-minute) pore-gradient electrophoresis of proteins and peptides in Micrograd gels.

    Science.gov (United States)

    Wrigley, C W; Margolis, J

    1992-01-01

    Precast gradient gels of short migration length (25 mm) have been developed to provide rapid electrophoretic separation without loss of resolution. These Micrograd gels have been prepared in gel ranges (conventional and unique) to match pore-gradient electrophoresis conditions to proteins/peptides ranging in size from several hundreds to millions. The Hylinx Micrograd gel combines an extreme gel range (6 to 48% polyacrylamide) with a novel crosslinker to provide sieving of polypeptides, and pore-limit electrophoresis of the smallest proteins (e.g. insulin monomer). All gel ranges (such as 3 to 30%) provide zone sharpening in routine analysis of conventional protein mixtures (e.g. serum) within 10 min electrophoresis at 200 to 300 volts. The gels are thin (1 mm) and thus stain quickly, but the gel cassette is of conventional overall width (83 mm), thus fitting many apparatus designs and accommodating 12 samples. The gels are finding valuable use in screening applications, requiring the electrophoretic analysis of many samples, and in cases where a rapid answer is needed, such as monitoring protein purification. The gels have proved particularly useful, in-house, for the latter application in developing Gradipore's new large-scale preparative electrophoresis system, the Gradiflow.

  14. Calorimetric Study of Thermal Denaturation of Superoxide Dismutase

    Institute of Scientific and Technical Information of China (English)

    王邦宁; 谈夫

    1994-01-01

    The thermal denaturation of superoxide dismutase (SOD) from bovine erythrocytes was studied at various pH values of different buffers and at various concentrations of solutions of two neutral salts by differential scanning calorimetry. The experiments performed indicate that the PIPES is a buffer non-coordinating with the SOD, and that the binding of the anions studied influences more or less the thermal denaturation of SOD, but the effect on the oxidation form of SOD is more apparent. A new conformer of SOD with lower thermostability was discovered by the experiments performed in different buffers at certain pH values higher than the isoelectric point of SOD, or at higher concentrations of neutral salt solutions. The new conformer may be converted irreversibly into the usual conformer with high thermostability during heating. Based on the thermodynamic parameters obtained in distilled water and by thermodynamic analysis using the Ooi’s model, it is revealed that the large enthalpy △Hdc contributed by

  15. [Characterization of thermal denaturation process of proteinase K by spectrometry].

    Science.gov (United States)

    Zhang, Qi-Bing; Na, Xin-Zhu; Yin, Zong-Ning

    2013-07-01

    The effect of different temperatures on the activity and conformational changes of proteinase K was studied. Methods Proteinase K was treated with different temperatures, then denatured natural substrate casein was used to assay enzyme activity, steady-state and time-resolved fluorescence spectroscopy was used to study tertiary structure, and circular dichroism was used to study secondary structure. Results show with the temperature rising from 25 to 65 degrees C, the enzyme activity and half-life of proteinase K dropped, maximum emission wavelength red shifted from 335 to 354 nm with fluorescence intensity decreasing. Synchronous fluorescence intensity of tryptophan residues decreased and that of tyrosine residues increased. Fluorescence lifetime of tryptophan residues reduced from 4. 427 1 to 4. 032 4 ns and the fraction of alpha-helix dropped. It was concluded that it is simple and accurate to use steady-state/time-resolved fluorescence spectroscopy and circular dichroism to investigate thermal stability of proteinase K. Thermal denaturation of proteinase K followed a three-state process. Fluorescence intensity of proteinase K was affected by fluorescence resonance energy transfer from tyrosine to tryptophan residues. The alpha-helix was the main structure to maintain conformational stability of enzyme active site of proteinase K.

  16. Denatured Thermodynamics of Proteins in Weak Cation-exchange Chromatography

    Institute of Scientific and Technical Information of China (English)

    LI Rong; CHEN Guo-Liang

    2003-01-01

    The thermostability of some proteins in weak cation-exchange chromatography was investigated at 20-80 ℃. The results show that there is a fixed thermal denaturation transition temperature for each protein. The appearance of the thermal transition temperature indicates that the conformations of the proteins are destroyed seriously. The thermal behavior of the proteins in weak cation-exchange and hydrophobic interaction chromatographies were compared in a wide temperature range. It was found that the proteins have a higher thermostability in a weak cation-exchange chromatography system. The thermodynamic parameters(ΔH0, ΔS0) of those proteins were determined by means of Vant Hoff relationship(lnk-1/T). According to standard entropy change(ΔS0), the conformational change of the proteins was judged in the chromatographic process. The linear relationships between ΔH0 and ΔS0 can be used to evaluate "compensation temperature"(β) at the protein denaturation and identify the identity of the protein retention mechanism in weak cation-exchange chromatography.

  17. Optical real-time measurement of collagen denaturation

    Science.gov (United States)

    Sankaran, Vanitha; Walsh, Joseph T., Jr.

    1997-06-01

    Linear birefringence is a property of collagenous tissue that results from both its composition and structure. Previous investigations have shown that birefringence provides an indication of structural changes in collagen during slow heating. We now report the birefringent response of both mature and young rat tail tendon to laser-heating. The results indicate that denaturation of collagen from mature rats induced by a 200-microsecond(s) -long Ho:YAG laser pulse may not be described accurately by kinetic parameters. Several second-long pulses of CO2 laser pulse may not be described from young rats fit an Arrhenius model with Ea equals 12.1 kcal/mol and A equals e18.03 s-1. Typically, for slow-heating of collagen, Ea equals kcal/mol and A equals e120 s-1. Thus, it seems likely that the temperature and energy needed to initiate collagen denaturation is lower in young collagen, possibly due to its decreased hydroxyproline content and consequent decreased thermal stability.

  18. Second-harmonic generation investigation of collagen thermal denaturation

    Science.gov (United States)

    Chen, Wei-Liang; Sun, Yen; Lin, Sung-Jan; Jee, Shiou-Hwa; Chen, Yang-Fang; Lin, Ling-Chih; So, Peter T. C.; Dong, Chen-Yuan

    2007-02-01

    Using the technique of second-harmonic generation (SHG) microscopy we obtained large area image of type I collagen from rat tail tendon as it is heated from 40°C to 70°C for 0 to 180 minutes. The high resolution images allowed us to investigate the collagen structural change. We observed that heating the tendon below the temperature of 54°C does not produce any change in the averaged SHG intensity. At the heating temperature of 54°C and above, we find that increasing the heating temperature and time leads to decreasing SHG intensity. As the tendon is heated above 54°C, a decrease in the SHG signal occurs uniformly throughout the tendon, but the regions where the SHG signal vanishes form a tiger-tail like pattern. By comparing the relative SHG intensities in small and large areas, we found that the denaturation process responsible for forming the tiger-tail like pattern occurs at a higher rate than the global denaturation process occurring throughout the tendon. Our results show that second-harmonic generation microscopy is effective in monitoring the thermal damage to collagen and has potential applications in biomedicine.

  19. Simplified sample preparation method for protein identification by matrix-assisted laser desorption/ionization mass spectrometry: in-gel digestion on the probe surface

    DEFF Research Database (Denmark)

    Stensballe, A; Jensen, Ole Nørregaard

    2001-01-01

    Identification and detailed characterization of complex mixtures of proteins separated by polyacrylamide gel electrophoresis (PAGE) require optimized and robust methods for interfacing electrophoretic techniques to mass spectrometry. Peptide mapping by matrix-assisted laser desorption/ionization-......Identification and detailed characterization of complex mixtures of proteins separated by polyacrylamide gel electrophoresis (PAGE) require optimized and robust methods for interfacing electrophoretic techniques to mass spectrometry. Peptide mapping by matrix-assisted laser desorption...... for protein identification similar to that obtained by the traditional protocols for in-gel digestion and MALDI peptide mass mapping of human proteins, i.e. approximately 60%. The overall performance of the novel on-probe digestion method is comparable with that of the standard in-gel sample preparation...

  20. DIFFERENTIATION OF CROATIAN BARLEY VARIETIES BY GRADIENT GEL SDS-PAGE AND ISOELECTRIC FOCUSING OF DRY GRAINS AND GREEN MALT HORDEINS

    Directory of Open Access Journals (Sweden)

    Ivica Strelec

    2011-06-01

    Full Text Available Applicability of polyacrylamide gel electrophoresis and isoelectric focusing of hordeins for discrimination of six two-rowed winter barley varieties (Angora, Sladoran, Rodnik, Rex, Martin and Barun has been investigated. Hordeins extracted from dry grains and green malt of barley varieties were separated by sodium dodecyl sulphate-polyacrylamide gel electrophoresis in gradient gel of 8-18% T, and by isoelectric focusing in pH gradient of 3.5-9.5 and 5.5-8.5, respectively. In all separation experiments better resolution of proteins was achieved with dry grain extracts, than with malt extracts. Angora, Sladoran and Martin variety could be distinguished from other varieties by differences in hordein patterns obtained by gradient gel SDS-PAGE (8-18% T, and Angora, Sladoran, Martin and Rodnik by isoelectric focusing in pH gradient 5.5-8.5.

  1. Investigation and Comparison of Leishmania major Promastigote and Amastigote Protein Content by Sodium Dodecyl Sulfate Polyacrylamide Gel Electrophoresis

    Directory of Open Access Journals (Sweden)

    S. Soleimanifard

    2013-04-01

    Full Text Available ntroduction & Objective: Leishmania is a protozoan of the trypanosomatidae family. This pro-tozoan has two stages in its life cycle, promastigote form in sand flies and amastigote form in macrophage of mammalian hosts. The purpose of this study was identification and compari-son of proteins of Leishmania amastigote and promastigote stages. Materials & Methods: The present study is a cross sectional study of two forms of Leishmania major. To culture promastigotes , L.major (MRHO/IR/75/ER from previously infected Balb/c mice was transferred to modified N.N.N medium with overlay of liquid BHI and then transferred to RPMI-1640 at 26oc ± 1 for mass production. After isolation and growth, pro-mastigotes were transferred to liquid cell culture medium RPMI-1640 with pH 5.5 and incu-bated at 5% CO2 at 37oc for 72 hours until promastigote to amastigote transformation. Elec-trophoresis was performed with SDS-PAGE method to find and compare the molecular weight of the antigens of two stages. Results: The molecular weights of the bands observed in both forms were as follows: 19, 36, 50, 63, 65, 80, 90, 94, 96, 110- 130 KDa. The proteins in the surface of only promastigote were 22, 28 and 46 KDa and special proteins in the surface of amastigote were 12 and 32 KDa. Conclusion : According to this study Leishmania parasite has stage specific proteins. Various studies have shown that axenic amastigotes and tissue amastigotes are similar in their protein content. Therefore, based on stage specific proteins ,effective drugs and vaccines can be de-signed against leishmaniasis. (Sci J Hamadan Univ Med Sci 2013; 20 (1:1-8

  2. Database of two-dimensional polyacrylamide gel electrophoresis of proteins labeled with CyDye DIGE Fluor saturation dye.

    Science.gov (United States)

    Fujii, Kazuyasu; Kondo, Tadashi; Yokoo, Hideki; Okano, Tetsuya; Yamada, Masayo; Yamada, Tesshi; Iwatsuki, Keiji; Hirohashi, Setsuo

    2006-03-01

    CyDye DIGE Fluor saturation dye (saturation dye, GE Healthcare Amersham Biosciences) enables highly sensitive 2-D PAGE. As the dye reacts with all reduced cysteine thiols, 2-D PAGE can be performed with a lower amount of protein, compared with CyDye DIGE Fluor minimal dye (GE Healthcare Amersham Biosciences), the sensitivity of which is equivalent to that of silver staining. We constructed a 2-D map of the saturation dye-labeled proteins of a liver cancer cell line (HepG2) and identified by MS 92 proteins corresponding to 123 protein spots. Functional classification revealed that the identified proteins had chaperone, protein binding, nucleotide binding, metal ion binding, isomerase activity, and motor activity. The functional distribution and the cysteine contents of the proteins were similar to those in the most comprehensive 2-D database of hepatoma cells (Seow et al.., Electrophoresis 2000, 21, 1787-1813), where silver staining was used for protein visualization. Hierarchical clustering on the basis of the quantitative expression profiles of the 123 characterized spots labeled with two charge- and mass-matched saturation dyes (Cy3 and Cy5) discriminated between nine hepatocellular carcinoma cell lines and primary cultured hepatocytes from five individuals, suggesting the utility of saturation dye and our database for proteomic studies of liver cancer.

  3. A rapid method for the detection of trypsinogen and chymotrypsinogen after electrophoretic separation of pancreas extract on polyacrylamide gel

    NARCIS (Netherlands)

    Dijkhof, J.; Poort, C.

    1974-01-01

    Several methods have been described for the visualization of proteolytic activity on electropherograms obtained with starch (1,2), agar and agarose (3–6), paper (7), and cellulose acetate (8–11) as supporting media. In most of these reports casein was used as a (nonspecific) substrate. In only one c

  4. Heterogeneity of mitochondrial DNA from Saccharomyces carlsbergensis. Denaturation mapping by electron microscopy.

    DEFF Research Database (Denmark)

    Christiansen, Gunna; Christiansen, C; Bak, AL

    1975-01-01

    Electronmicroscopic observation of the denaturation pattern of 130 partially denaturated linear mitochondrial DNA molecules from Saccharomyces carlsbergensis was used to investigate the distribution of AT-rich sequences within the mitochondrial genome. The molecules were observed after heating...... denaturated sequences in the mitochondrial DNA. These sequences which presumably correspond to the very AT-rich regions, known to exist in the yeast mitochondrial DNA, were found at intervals of about 0.5 - 3 mum on the map....

  5. 27 CFR 21.92 - Denaturants listed as U.S.P. or N.F.

    Science.gov (United States)

    2010-04-01

    ....P. or N.F. 21.92 Section 21.92 Alcohol, Tobacco Products and Firearms ALCOHOL AND TOBACCO TAX AND... for Denaturants § 21.92 Denaturants listed as U.S.P. or N.F. Denaturing materials and products listed in this part as “U.S.P.” or “N.F.” shall meet the specifications set forth in the current...

  6. [Discrimination of types of polyacrylamide based on near infrared spectroscopy coupled with least square support vector machine].

    Science.gov (United States)

    Zhang, Hong-Guang; Yang, Qin-Min; Lu, Jian-Gang

    2014-04-01

    In this paper, a novel discriminant methodology based on near infrared spectroscopic analysis technique and least square support vector machine was proposed for rapid and nondestructive discrimination of different types of Polyacrylamide. The diffuse reflectance spectra of samples of Non-ionic Polyacrylamide, Anionic Polyacrylamide and Cationic Polyacrylamide were measured. Then principal component analysis method was applied to reduce the dimension of the spectral data and extract of the principal compnents. The first three principal components were used for cluster analysis of the three different types of Polyacrylamide. Then those principal components were also used as inputs of least square support vector machine model. The optimization of the parameters and the number of principal components used as inputs of least square support vector machine model was performed through cross validation based on grid search. 60 samples of each type of Polyacrylamide were collected. Thus a total of 180 samples were obtained. 135 samples, 45 samples for each type of Polyacrylamide, were randomly split into a training set to build calibration model and the rest 45 samples were used as test set to evaluate the performance of the developed model. In addition, 5 Cationic Polyacrylamide samples and 5 Anionic Polyacrylamide samples adulterated with different proportion of Non-ionic Polyacrylamide were also prepared to show the feasibilty of the proposed method to discriminate the adulterated Polyacrylamide samples. The prediction error threshold for each type of Polyacrylamide was determined by F statistical significance test method based on the prediction error of the training set of corresponding type of Polyacrylamide in cross validation. The discrimination accuracy of the built model was 100% for prediction of the test set. The prediction of the model for the 10 mixing samples was also presented, and all mixing samples were accurately discriminated as adulterated samples. The

  7. Preparation and Photochromic Properties of Hybrid Thin Films Based on Heteropolyoxometallate and Polyacrylamide

    Institute of Scientific and Technical Information of China (English)

    2002-01-01

    series of photochromic hybrid films were prepared through entrapping Dawson type tungsten heteropolyoxometallates (P2W18O626-) and molybdenum heteropolyoxometallate (P2Mo18O626-) into polyacrylamide matrix. FTIR results showed that the Dawson geometry of heteropolyoxometallates is still preserved inside the composites and strong coulombic interaction is built between heteropolyoxometallates and polyacrylamide via hydrogen bonding. Irradiated with ultraviolet light, the transparent films change from colorless to blue and show reversible photochromism.The bleaching process occurs when the films are in contact with air or O2 in the dark. The molybdenum heteropolyoxometallate hybrid film has higher photochromic efficiency and slower bleaching reaction than tungsten heteropolyoxometallate hybrid film. ESR results indicated that polyacrylamide is a hydrogen donor and the photoreduced process is in accordance with the radical mechanism.

  8. Preparation of a Novel Coal Gangue-Polyacrylamide Hybrid Flocculant and Its Flocculation Performance

    Institute of Scientific and Technical Information of China (English)

    Xiangao Quan; Huiyun Wang

    2014-01-01

    A novel flocculant based on hybrid coal gangue-polyacrylamide (HCGPAM) has been prepared by using modified coal gangue and polyacrylamide. Factors related to the preparation such as reaction time, temperature, concentration of the polymer monomer and ratio of initiators are investigated. The product is characterized by infrared spectra (FTIR), scanning electron microscopy (SEM), X-ray diffraction (XRD), aswell as viscometry. The flocculating tests on oilfield drilling wastewater show that the removal efficiency is 85.5% and the light transmittance is 53.6%. The results indicate that the coal gangue could be used for the preparation of inorganic-organic hybrid flocculant and the removal efficiency is much higher than that of commercial polyacrylamide (PAM) or PAM/ coal gangue blend.

  9. Harmful effects of formaldehyde on the stability of polyacrylamide solutions used in enhanced oil recovery

    Energy Technology Data Exchange (ETDEWEB)

    Catherin, G.; Marchal, J.

    1979-01-01

    In a study of the oxidation of aqueous solutions of the polyacrylamides PAA and HPAA, it was found that formaldehyde is an oxidation product of acrylamide. The presence of formaldehyde in turn causes the production of insoluble derivatives which spoil the rheological properties of polymer solutions. This result reinforces those of previous studies. Examination of the literature showed that the oxidation and addition reactions proposed in these previous studies to account for this effect are not consistent with the known properties of formaldehyde in the media used. A scheme is proposed which more fully accounts for the observed phenomena. It also has been shown that oxido-reduction of formaldehyde initiates the formation of radicals on polyacrylamides. It is concluded that formaldehyde should not be used to protect polyacrylamide solutions against biodegradation in tertiary oil recovery. 40 references.

  10. Some considerations concerning four-ball machine testing of the polyacrylamide solutions

    Science.gov (United States)

    Drumeanu, A. C.

    2017-02-01

    Polyacrylamide (PAM) is one of the most widely used and technically important water-soluble polymers. Polyacrylamide (PAM) is usually obtained by free radical polymerization of acrylamide (AM) and in its partially hydrolysed form is a synthetic straight-chain polymer of acrylamide monomers, some of which have been hydrolysed. The structure of HPAM molecule is a flexible chain. This kind of structure is known as a random coil in polymer chemistry. Due to the hydrolysed groups contained in its molecule, HPAM has multiple charges distributed along the chain that make it a polyelectrolyte. The paper presents the experimental results concerning the lubricant solutions based on polyacrylamide behaviour when were tested on the four ball machine. It has to be mentioned that this kind of polymer was not used until now in lubrication and the studies concerning its tribological behaviour are at the beginning.

  11. Electrochemical behaviour of denatured ethanol for use in direct ethanol fuel cells

    Energy Technology Data Exchange (ETDEWEB)

    Bayer, Domnik; Kintzel, Birgit; Joos, Martin; Cremers, Carsten; Tuebke, Jens [Fraunhofer-Institut fuer Chemische Technologie (ICT), Pfinztal (Germany)

    2010-07-01

    In the present study, two denaturing agents, an adjuvant to a denaturing agent and a mixture of a denaturing agend and the adjuvant were tested with regard to their fuel cell compatibility. Therefore, various electrochemical tests including cyclic voltammetry, chronoamperometry and differential electrochemical mass spectroscopy have been conducted at platinum as model catalyst in acidic and alkaline medium. In addition, the most promising denaturing agent, a mixture of tert-butyl ethyl ether (ETBE) with the adjuvant Bitrex {sup registered}, has also been tested at commercial fuel cell catalysts in both acidic and alkaline media. (orig.)

  12. Visualization of early events in acetic acid denaturation of HIV-1 protease: a molecular dynamics study.

    Directory of Open Access Journals (Sweden)

    Aditi Narendra Borkar

    Full Text Available Protein denaturation plays a crucial role in cellular processes. In this study, denaturation of HIV-1 Protease (PR was investigated by all-atom MD simulations in explicit solvent. The PR dimer and monomer were simulated separately in 9 M acetic acid (9 M AcOH solution and water to study the denaturation process of PR in acetic acid environment. Direct visualization of the denaturation dynamics that is readily available from such simulations has been presented. Our simulations in 9 M AcOH reveal that the PR denaturation begins by separation of dimer into intact monomers and it is only after this separation that the monomer units start denaturing. The denaturation of the monomers is flagged off by the loss of crucial interactions between the α-helix at C-terminal and surrounding β-strands. This causes the structure to transit from the equilibrium dynamics to random non-equilibrating dynamics. Residence time calculations indicate that denaturation occurs via direct interaction of the acetic acid molecules with certain regions of the protein in 9 M AcOH. All these observations have helped to decipher a picture of the early events in acetic acid denaturation of PR and have illustrated that the α-helix and the β-sheet at the C-terminus of a native and functional PR dimer should maintain both the stability and the function of the enzyme and thus present newer targets for blocking PR function.

  13. Protein differences between normal and oligospermic human sperm demonstrated by two-dimensional gel electrophoresis.

    Science.gov (United States)

    Morgentaler, A; Schopperle, W M; Crocker, R H; DeWolf, W C

    1990-11-01

    Protein expression by sperm obtained from men with normal semen analysis and men with oligospermia were evaluated by two-dimensional gel electrophoresis. Proteins were solubilized in a 9.5 M urea/2% Nonidet-P40 (LKB, Bromma, Sweden) lysis buffer and underwent second dimension separation on 10 to 16% polyacrylamide gradient gels. A set of 36 invariant proteins was identified in all normospermic samples, whereas 8 of 10 evaluable oligospermic samples lacked 1 or more of the invariant proteins. Proteins absent in oligospermic samples may be critical to normal sperm function and may serve as markers for infertility.

  14. Transport Phenomena in Gel

    Directory of Open Access Journals (Sweden)

    Masayuki Tokita

    2016-05-01

    Full Text Available Gel becomes an important class of soft materials since it can be seen in a wide variety of the chemical and the biological systems. The unique properties of gel arise from the structure, namely, the three-dimensional polymer network that is swollen by a huge amount of solvent. Despite the small volume fraction of the polymer network, which is usually only a few percent or less, gel shows the typical properties that belong to solids such as the elasticity. Gel is, therefore, regarded as a dilute solid because its elasticity is much smaller than that of typical solids. Because of the diluted structure, small molecules can pass along the open space of the polymer network. In addition to the viscous resistance of gel fluid, however, the substance experiences resistance due to the polymer network of gel during the transport process. It is, therefore, of importance to study the diffusion of the small molecules in gel as well as the flow of gel fluid itself through the polymer network of gel. It may be natural to assume that the effects of the resistance due to the polymer network of gel depends strongly on the network structure. Therefore, detailed study on the transport processes in and through gel may open a new insight into the relationship between the structure and the transport properties of gel. The two typical transport processes in and through gel, that is, the diffusion of small molecules due to the thermal fluctuations and the flow of gel fluid that is caused by the mechanical pressure gradient will be reviewed.

  15. Study on Magnetic Responsibility of Rare Earth Ferrite/Polyacrylamide Magnetic Microsphere

    Institute of Scientific and Technical Information of China (English)

    Zhang Ming; Wang Zhifeng; Zhang Hong; Dai Shaojun; Qiu Guanming; Okamoto Hiroshi

    2005-01-01

    In inverse microemulsion, rare earth ferrite/polyacrylamide magnetic microsphere were prepared and their magnetic responsibility were studied by magnetic balance. Results indicate that the magnetic responsibility of microsphere relates to magnetic moment of rare earth ion, and it can be improved by the addition of dysprosium ion of high magnetic moment. Dysprosium content has an effect on magnetic responsibility of dysprosium ferrite/polyacrylamide magnetic microsphere. The microsphere displays strong magnetic responsibility when the molar ratio of Dy3+/iron is 0.20.

  16. Viscous properties of polyacrylamide solutions used for enhanced oil recovery and comparison with different rheological models

    Energy Technology Data Exchange (ETDEWEB)

    Lakatos, I.; Lakatos-Szabo, J.; Schurz, J.

    1987-09-01

    Six different rheological models were applied for description of viscous properties of polyacrylamide solutions in the low shear rate range. Non of the formulas evaluated has universal applicability despite theoretical interpretation and the 'constants' in the models proved hardly physically founded, but they can rather be used as 'fitting constants' in a formal way. The best coincidence was obtained between the measured and the calculated data with the Carreau's model which is also proposed for incorporation of the rheological properties of polyacrylamide solutions into mathematical simulators of polymer and polymer-micellar flooding.

  17. Orthogonally bifunctionalised polyacrylamide nanoparticles: a support for the assembly of multifunctional nanodevices

    Science.gov (United States)

    Giuntini, F.; Dumoulin, F.; Daly, R.; Ahsen, V.; Scanlan, E. M.; Lavado, A. S. P.; Aylott, J. W.; Rosser, G. A.; Beeby, A.; Boyle, R. W.

    2012-03-01

    Polyacrylamide nanoparticles bearing two orthogonal reactive functionalities were prepared by reverse microemulsion polymerisation. Water-soluble photosensitisers and peptide or carbohydrate moieties were sequentially attached to the new nanospecies by orthogonal conjugations based on copper-catalysed azide-alkyne cycloaddition and isothiocyanate chemistry.Polyacrylamide nanoparticles bearing two orthogonal reactive functionalities were prepared by reverse microemulsion polymerisation. Water-soluble photosensitisers and peptide or carbohydrate moieties were sequentially attached to the new nanospecies by orthogonal conjugations based on copper-catalysed azide-alkyne cycloaddition and isothiocyanate chemistry. Electronic supplementary information (ESI) available. See DOI: 10.1039/c2nr11947a

  18. Acute toxicity of polyacrylamide flocculants to early life stages of freshwater mussels.

    Science.gov (United States)

    Buczek, Sean B; Cope, W Gregory; McLaughlin, Richard A; Kwak, Thomas J

    2017-04-11

    Polyacrylamide has become an effective tool for reducing construction-related suspended sediment and turbidity, which are considered to have significant adverse impacts on aquatic ecosystems and are a leading cause of the degradation of North American streams and rivers. However, little is known about the effects of polyacrylamide on many freshwater organisms, and prior to the present study, no information existed on the toxicity of polyacrylamide compounds to native freshwater mussels (family Unionidae), one of the most imperiled faunal groups globally. Following standard test guidelines, we exposed juvenile mussels (test duration 96 h) and glochidia larvae (test duration 24 h) to 5 different anionic polyacrylamide compounds and 1 non-ionic compound. Species tested included the yellow lampmussel (Lampsilis cariosa), an Atlantic Slope species that is listed as endangered in North Carolina; the Appalachian elktoe (Alasmidonta raveneliana), a federally endangered Interior Basin species; and the washboard (Megalonaias nervosa), a common Interior Basin species. We found that median lethal concentrations (LC50s) of polyacrylamide ranged from 411.7 to >1000 mg/L for glochidia and from 126.8 to >1000 mg/L for juveniles. All LC50s were orders of magnitude greater (2-3) than concentrations typically recommended for turbidity control (1-5 mg/L), regardless of their molecular weight or charge density. The results demonstrate that the polyacrylamide compounds tested were not acutely toxic to the mussel species and life stages tested, indicating minimal risk of short-term exposure from polyacrylamide applications in the environment. However, other potential uses of polyacrylamide in the environment (e.g., wastewater treatment, paper processing, mining, algae removal) and their chronic or sublethal effects remain uncertain and warrant additional investigation. Environ Toxicol Chem 2017;9999:1-7. Published 2017 Wiley Periodicals Inc. on behalf of SETAC. This article is

  19. Microplate array diagonal gel electrophoresis for cohort studies of microsatellite loci.

    Science.gov (United States)

    Chen, Xiao-he; O'Dell, Sandra D; Day, Ian N M

    2002-05-01

    After PCR amplification, we have achieved precise sizing of trinucleotide and tetranucleotide microsatellite alleles on 96-well open-faced polyacrylamide microplate array diagonal gel electrophoresis (MADGE) gels: two tetranucleotide repeats, HUMTHOI (five alleles 248-263 bp) and DYS390 (eight alleles 200-228 bp), and DYS392, a trinucleotide repeat (eight alleles 210-231 bp). A gel matrix of Duracryl, a high mechanical strength polyacrylamide derivative, and appropriate ionic conditions provide the 1.3%-1.5% band resolution required. No end-labeling of primers is needed, as the sensitive Vistra Green intercalating dye is used for the visualization of bands. Co-run markers bracketing the PCR fragments ensure accurate sizing without inter-lane variability. Electrophoresis of multiple gels in a thermostatically controlled tank allows up to 1000 samples to be run in 90 min. Gel images were analyzed using a Fluorlmager 595 fluorescent scanning system, and alleles were identified using Phoretix software for band migration measurement and Microsoft Excel to compute fragment sizes. Estimated sizes were interpolated precisely to achieve accurate binning. Microsatellite-MADGE represents a utilitarian methodfor high-throughput genotyping in cohort studies, using standard laboratory equipment.

  20. Combining high-throughput MALDI-TOF mass spectrometry and isoelectric focusing gel electrophoresis for virtual 2D gel-based proteomics.

    Science.gov (United States)

    Lohnes, Karen; Quebbemann, Neil R; Liu, Kate; Kobzeff, Fred; Loo, Joseph A; Ogorzalek Loo, Rachel R

    2016-07-15

    The virtual two-dimensional gel electrophoresis/mass spectrometry (virtual 2D gel/MS) technology combines the premier, high-resolution capabilities of 2D gel electrophoresis with the sensitivity and high mass accuracy of mass spectrometry (MS). Intact proteins separated by isoelectric focusing (IEF) gel electrophoresis are imaged from immobilized pH gradient (IPG) polyacrylamide gels (the first dimension of classic 2D-PAGE) by matrix-assisted laser desorption/ionization (MALDI) MS. Obtaining accurate intact masses from sub-picomole-level proteins embedded in 2D-PAGE gels or in IPG strips is desirable to elucidate how the protein of one spot identified as protein 'A' on a 2D gel differs from the protein of another spot identified as the same protein, whenever tryptic peptide maps fail to resolve the issue. This task, however, has been extremely challenging. Virtual 2D gel/MS provides access to these intact masses. Modifications to our matrix deposition procedure improve the reliability with which IPG gels can be prepared; the new procedure is described. Development of this MALDI MS imaging (MSI) method for high-throughput MS with integrated 'top-down' MS to elucidate protein isoforms from complex biological samples is described and it is demonstrated that a 4-cm IPG gel segment can now be imaged in approximately 5min. Gel-wide chemical and enzymatic methods with further interrogation by MALDI MS/MS provide identifications, sequence-related information, and post-translational/transcriptional modification information. The MSI-based virtual 2D gel/MS platform may potentially link the benefits of 'top-down' and 'bottom-up' proteomics.

  1. Consistent View of Polypeptide Chain Expansion in Chemical Denaturants from Multiple Experimental Methods.

    Science.gov (United States)

    Borgia, Alessandro; Zheng, Wenwei; Buholzer, Karin; Borgia, Madeleine B; Schüler, Anja; Hofmann, Hagen; Soranno, Andrea; Nettels, Daniel; Gast, Klaus; Grishaev, Alexander; Best, Robert B; Schuler, Benjamin

    2016-09-14

    There has been a long-standing controversy regarding the effect of chemical denaturants on the dimensions of unfolded and intrinsically disordered proteins: A wide range of experimental techniques suggest that polypeptide chains expand with increasing denaturant concentration, but several studies using small-angle X-ray scattering (SAXS) have reported no such increase of the radius of gyration (Rg). This inconsistency challenges our current understanding of the mechanism of chemical denaturants, which are widely employed to investigate protein folding and stability. Here, we use a combination of single-molecule Förster resonance energy transfer (FRET), SAXS, dynamic light scattering (DLS), and two-focus fluorescence correlation spectroscopy (2f-FCS) to characterize the denaturant dependence of the unfolded state of the spectrin domain R17 and the intrinsically disordered protein ACTR in two different denaturants. Standard analysis of the primary data clearly indicates an expansion of the unfolded state with increasing denaturant concentration irrespective of the protein, denaturant, or experimental method used. This is the first case in which SAXS and FRET have yielded even qualitatively consistent results regarding expansion in denaturant when applied to the same proteins. To more directly illustrate this self-consistency, we used both SAXS and FRET data in a Bayesian procedure to refine structural ensembles representative of the observed unfolded state. This analysis demonstrates that both of these experimental probes are compatible with a common ensemble of protein configurations for each denaturant concentration. Furthermore, the resulting ensembles reproduce the trend of increasing hydrodynamic radius with denaturant concentration obtained by 2f-FCS and DLS. We were thus able to reconcile the results from all four experimental techniques quantitatively, to obtain a comprehensive structural picture of denaturant-induced unfolded state expansion, and to

  2. Irreversible denaturation of maltodextrin glucosidase studied by differential scanning calorimetry, circular dichroism, and turbidity measurements.

    Science.gov (United States)

    Goyal, Megha; Chaudhuri, Tapan K; Kuwajima, Kunihiro

    2014-01-01

    Thermal denaturation of Escherichia coli maltodextrin glucosidase was studied by differential scanning calorimetry, circular dichroism (230 nm), and UV-absorption measurements (340 nm), which were respectively used to monitor heat absorption, conformational unfolding, and the production of solution turbidity. The denaturation was irreversible, and the thermal transition recorded at scan rates of 0.5-1.5 K/min was significantly scan-rate dependent, indicating that the thermal denaturation was kinetically controlled. The absence of a protein-concentration effect on the thermal transition indicated that the denaturation was rate-limited by a mono-molecular process. From the analysis of the calorimetric thermograms, a one-step irreversible model well represented the thermal denaturation of the protein. The calorimetrically observed thermal transitions showed excellent coincidence with the turbidity transitions monitored by UV-absorption as well as with the unfolding transitions monitored by circular dichroism. The thermal denaturation of the protein was thus rate-limited by conformational unfolding, which was followed by a rapid irreversible formation of aggregates that produced the solution turbidity. It is thus important to note that the absence of the protein-concentration effect on the irreversible thermal denaturation does not necessarily means the absence of protein aggregation itself. The turbidity measurements together with differential scanning calorimetry in the irreversible thermal denaturation of the protein provided a very effective approach for understanding the mechanisms of the irreversible denaturation. The Arrhenius-equation parameters obtained from analysis of the thermal denaturation were compared with those of other proteins that have been reported to show the one-step irreversible thermal denaturation. Maltodextrin glucosidase had sufficiently high kinetic stability with a half-life of 68 days at a physiological temperature (37°C).

  3. Sol-Gel Glasses

    Science.gov (United States)

    Mukherjee, S. P.

    1985-01-01

    Multicomponent homogeneous, ultrapure noncrystalline gels/gel derived glasses are promising batch materials for the containerless glass melting experiments in microgravity. Hence, ultrapure, homogeneous gel precursors could be used to: (1) investigate the effect of the container induced nucleation on the glass forming ability of marginally glass forming compositions; and (2) investigate the influence of gravity on the phase separation and coarsening behavior of gel derived glasses in the liquid-liquid immiscibility zone of the nonsilicate systems having a high density phase. The structure and crystallization behavior of gels in the SiO2-GeO2 as a function of gel chemistry and thermal treatment were investigated. As are the chemical principles involved in the distribution of a second network former in silica gel matrix being investigated. The procedures for synthesizing noncrystalline gels/gel-monoliths in the SiO2-GeO2, GeO2-PbO systems were developed. Preliminary investigations on the levitation and thermal treatment of germania silicate gel-monoliths in the Pressure Facility Acoustic Levitator were done.

  4. Gel-aided sample preparation (GASP)--a simplified method for gel-assisted proteomic sample generation from protein extracts and intact cells.

    Science.gov (United States)

    Fischer, Roman; Kessler, Benedikt M

    2015-04-01

    We describe a "gel-assisted" proteomic sample preparation method for MS analysis. Solubilized protein extracts or intact cells are copolymerized with acrylamide, facilitating denaturation, reduction, quantitative cysteine alkylation, and matrix formation. Gel-aided sample preparation has been optimized to be highly flexible, scalable, and to allow reproducible sample generation from 50 cells to milligrams of protein extracts. This methodology is fast, sensitive, easy-to-use on a wide range of sample types, and accessible to nonspecialists. © 2014 The Authors. PROTEOMICS published by Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

  5. A method to determine Young's modulus of soft gels for cell adhesion

    Institute of Scientific and Technical Information of China (English)

    Xiaoling Peng; Jianyong Huang; Lei Qin; Chunyang Xiong; Jing Fang

    2009-01-01

    A convenient technique is reported in this note for measuring elastic modulus of extremely soft material for cellular adhesion. Specimens of bending cylinder under gravity are used to avoid contact problem between testing device and sample, and a beam model is presented for evaluating the curvatures of gel beams with large elastic deformation. A self-adaptive algorithm is also proposed to search for the best estimation of gels' elastic moduli by comparing the experimental bending curvatures with those computed from the beam model with preestimated moduli. Application to the measurement of the property of polyacrylamide gels indicates that the material compliance varies with the concentrations of bis-acrylamide, and the gels become softer after being immersed in a culture medium for a period of time, no matter to what extent they are polymerized.

  6. Residual ordered structure in denatured proteins and the problem of protein folding.

    Science.gov (United States)

    Basharov, Mahmud A

    2012-02-01

    Structural characteristics of numerous globular proteins in the denatured state have been reviewed using literature data. Recent more precise experiments show that in contrast to the conventional standpoint, proteins under strongly denaturing conditions do not unfold completely and adopt a random coil state, but contain significant residual ordered structure. These results cast doubt on the basis of the conventional approach representing the process of protein folding as a spontaneous transition of a polypeptide chain from the random coil state to the unique globular structure. The denaturation of proteins is explained in terms of the physical properties of proteins such as stability, conformational change, elasticity, irreversible denaturation, etc. The spontaneous renaturation of some denatured proteins most probably is merely the manifestation of the physical properties (e.g., the elasticity) of the proteins per se, caused by the residual structure present in the denatured state. The pieces of the ordered structure might be the centers of the initiation of renaturation, where the restoration of the initial native conformation of denatured proteins begins. Studies on the denaturation of proteins hardly clarify how the proteins fold into the native conformation during the successive residue-by-residue elongation of the polypeptide chain on the ribosome.

  7. The generation of denatured reactor plutonium by different options of the fuel cycle

    Energy Technology Data Exchange (ETDEWEB)

    Broeders, C.H.M.; Kessler, G. [Inst. for Neutron Physics and Reactor Technology, Research Center Karlsruhe (Germany)

    2006-11-15

    Denatured (proliferation resistant) reactor plutonium can be generated in a number of different fuel cycle options. First denatured reactor plutonium can be obtained if, instead of low enriched U-235 PWR fuel, re-enriched U-235/U-236 from reprocessed uranium is used (fuel type A). Also the envisaged existing 2,500 t of reactor plutonium (being generated world wide up to the year 2010), mostly stored in intermediate fuel storage facilities at present, could be converted during a transition phase into denatured reactor plutonium by the options fuel type B and D. Denatured reactor plutonium could have the same safeguards standard as present low enriched (<20% U-235) LWR fuel. It could be incinerated by recycling once or twice in PWRs and subsequently by multi-recycling in FRs (CAPRA type or IFRs). Once denatured, such reactor plutonium could remain denatured during multiple recycling. In a PWR, e.g., denatured reactor plutonium could be destroyed at a rate of about 250 kg/GWey. While denatured reactor plutonium could be recycled and incinerated under relieved IAEA safeguards, neptunium would still have to be monitored by the IAEA in future for all cases in which considerable amounts of neptunium are produced. (orig.)

  8. Long-term follow-up after urethral injection with polyacrylamide hydrogel for female stress incontinence

    DEFF Research Database (Denmark)

    Mouritsen, Lone; Lose, Gunnar; Møller-Bek, Karl

    2014-01-01

    Urethral injection therapy for treatment of stress urinary incontinence has been in use for years, but only a few long-term follow-up studies have been published. Twenty-five women, injected with polyacrylamide hydrogel 8 years earlier, were invited for follow-up. Twenty-four could be contacted; ...

  9. EXPERIMENTAL CHARACTERIZATION OF FLUOROCARBON-MODIFIED POLYACRYLAMIDE/SURFACTANT AQUEOUS SOLUTIONS

    Institute of Scientific and Technical Information of China (English)

    Huai-tian Bu; Zhen-zhong Yang; Yun-xiang Zhang

    2003-01-01

    The interaction between surfactants and fluorocarbon-modified polyacrylamide (FC-PAM) in aqueous solutions was evaluated by theological means and fluorescence spectroscopy and was found to be strong regardless of the surfactant's nature. Two representative surfactants, anionic sodium dodecyl sulfate (SDS) and nonionic Triton X-100, were used. The origin of the interaction and its dependence on the surfactant concentration were discussed.

  10. Polyacrylamide and biopolymer effects on flocculation, aggregate stability, and water seepage in a silt loam

    Science.gov (United States)

    Researcher’s seek a more renewable and natural alternative for water soluble anionic polyacrylamide (PAM), a highly-effective, petroleum-derived polymer used in agriculture to control erosion and reduce water seepage from unlined irrigation structures. This study evaluated two anionic polymers: a ba...

  11. Molecular Understanding and Structural-Based Design of Polyacrylamides and Polyacrylates as Antifouling Materials.

    Science.gov (United States)

    Chen, Hong; Zhao, Chao; Zhang, Mingzhen; Chen, Qiang; Ma, Jie; Zheng, Jie

    2016-04-12

    Design and synthesis of highly bioinert and biocompatible antifouling materials are crucial for a broad range of biomedical and engineering applications. Among antifouling materials, polyacrylamides and polyacrylates have proved so promising because of cheap raw materials, ease of synthesis and applicability, and abundant functional groups. The strong surface hydration and the high surface packing density of polyacrylamides and polyacrylates are considered to be the key contributors to their antifouling property. In this article, we review our studies on the design and synthesis of a series of polyacrylamides and polyacrylates with different molecular structures. These polymers can be fabricated into different architectural forms (brushes, nanoparticles, nanogels, and hydrogels), all of which are highly resistant to the attachment of proteins, cells, and bacteria. We find that small structural changes in the polymers can lead to large enhancement in surface hydration and antifouling performance, both showing a positive correlation. This reveals a general design rule for effective antifouling materials. Furthermore, polyacrylamides and polyacrylates are readily functionalized with other bioactive compounds to achieve different new multifunctionalities.

  12. STUDIES ON IPN OF POLYACRYLAMIDE INVERSE EMULSION AND MODIFIED UREA-FORMALDEHYDE RESIN

    Institute of Scientific and Technical Information of China (English)

    LIU Qingpu; HOU Sijian; HA Runhua

    1996-01-01

    The selective water plugging agent was prepared by heating the blends of the polyacrylamide inverse latex, modified urea formaldehyde resin, crosslinking agent and catalysts.The results show that using different types of polymers and additives or changing in their proportion of the blends, the gelling viscosity, starting point of gelling and other properties of the IPN can be controlled.

  13. Branched polyacrylamides : Synthesis and effect of molecular architecture on solution rheology

    NARCIS (Netherlands)

    Wever, D. A. Z.; Picchioni, F.; Broekhuis, A. A.

    2013-01-01

    Linear, star and comb-like polyacrylamides (PAM) have been prepared by atomic transfer radical polymerization (ATRP) in aqueous media at room temperature. The influence of the molecular architecture of PAM on the rheological properties in aqueous solution has been investigated. The well-known theory

  14. Determination of Residual Acrylamide in Medical Polyacrylamide Hydrogel by High Performance Liquid Chromatography tandem Mass Spectroscopy

    Institute of Scientific and Technical Information of China (English)

    WEI-WEI LI; HUI LI; ZHI-FEI LIU; QUN QIAO

    2009-01-01

    Objective To determine residual acrylamide in medical polyacrylamide hydrogel by high performance liquid chromatography tandem mass spectroscopy (HPLC-MS). Methods After 13C3 labeled acrylamide was added, the sample was extracted with water and then cleaned up with ExtrelutTM 20. The polyaerylamide hydrogel sample and 20 clinical cases were analyzed by HPLC-MS/MS and isotope dilution quantifying technique in selected reaction monitoring (SRM) mode. Results Acrylamide was separated from polyacrylamide hydrogel. The concentration of acrylamide in polyacrylamide hydrogel ranged from 3.9×109 to 3.1×108g/L in the 20 clinical cases. The peak area was favorable linear and the range was up to 3 000 μg/L. The recovery rate was 103.1% with a relative standard deviation (RSD) of 6.20%, when the mark level was 50 μg/L. Conclusion HPLC-MS is a rapid, accurate, and sensitive method for the determination of residual acrylamide in medical polyacrylamide hydrogel.

  15. Polyacrylamide molecular weight and phosphogypsum effects on infiltration and erosion in semi-arid soils

    Science.gov (United States)

    Seal formation at the surface of semi-arid soils during rainstorms reduces soil infiltration rate (IR) and causes runoff and erosion. Surface application of dry anionic polyacrylamide (PAM) with high molecular weight (MW) has been found to be effective in stabilizing soil aggregates, and decreasing ...

  16. Synthesis and characterization of polyacrylamide-graft-poly(ethylene oxid-copropyleneoxide)

    NARCIS (Netherlands)

    de Vos, Sicco; de Vos, S.C.; Moller, M.; Möller, Martin; Visscher, K.; Mijnlieff, P.F.; Visscher, K.; Mijnlieff, P.F.

    1994-01-01

    The synthesis and first experiments on the rheological behaviour of poly(acrylamide) (PAAm) grafted with poly(ethylene oxide-co-propylene oxide) are presented. Well-defined, water-soluble graft copolymers were prepared in high yields by grafting poly(oxyalkylene) monoamines onto partly hydrolysed

  17. Integration of gel-based and gel-free proteomic data for functional analysis of proteins through Soybean Proteome Database

    KAUST Repository

    Komatsu, Setsuko

    2017-05-10

    The Soybean Proteome Database (SPD) stores data on soybean proteins obtained with gel-based and gel-free proteomic techniques. The database was constructed to provide information on proteins for functional analyses. The majority of the data is focused on soybean (Glycine max ‘Enrei’). The growth and yield of soybean are strongly affected by environmental stresses such as flooding. The database was originally constructed using data on soybean proteins separated by two-dimensional polyacrylamide gel electrophoresis, which is a gel-based proteomic technique. Since 2015, the database has been expanded to incorporate data obtained by label-free mass spectrometry-based quantitative proteomics, which is a gel-free proteomic technique. Here, the portions of the database consisting of gel-free proteomic data are described. The gel-free proteomic database contains 39,212 proteins identified in 63 sample sets, such as temporal and organ-specific samples of soybean plants grown under flooding stress or non-stressed conditions. In addition, data on organellar proteins identified in mitochondria, nuclei, and endoplasmic reticulum are stored. Furthermore, the database integrates multiple omics data such as genomics, transcriptomics, metabolomics, and proteomics. The SPD database is accessible at http://proteome.dc.affrc.go.jp/Soybean/. Biological significanceThe Soybean Proteome Database stores data obtained from both gel-based and gel-free proteomic techniques. The gel-free proteomic database comprises 39,212 proteins identified in 63 sample sets, such as different organs of soybean plants grown under flooding stress or non-stressed conditions in a time-dependent manner. In addition, organellar proteins identified in mitochondria, nuclei, and endoplasmic reticulum are stored in the gel-free proteomics database. A total of 44,704 proteins, including 5490 proteins identified using a gel-based proteomic technique, are stored in the SPD. It accounts for approximately 80% of all

  18. Integration of gel-based and gel-free proteomic data for functional analysis of proteins through Soybean Proteome Database.

    Science.gov (United States)

    Komatsu, Setsuko; Wang, Xin; Yin, Xiaojian; Nanjo, Yohei; Ohyanagi, Hajime; Sakata, Katsumi

    2017-06-23

    The Soybean Proteome Database (SPD) stores data on soybean proteins obtained with gel-based and gel-free proteomic techniques. The database was constructed to provide information on proteins for functional analyses. The majority of the data is focused on soybean (Glycine max 'Enrei'). The growth and yield of soybean are strongly affected by environmental stresses such as flooding. The database was originally constructed using data on soybean proteins separated by two-dimensional polyacrylamide gel electrophoresis, which is a gel-based proteomic technique. Since 2015, the database has been expanded to incorporate data obtained by label-free mass spectrometry-based quantitative proteomics, which is a gel-free proteomic technique. Here, the portions of the database consisting of gel-free proteomic data are described. The gel-free proteomic database contains 39,212 proteins identified in 63 sample sets, such as temporal and organ-specific samples of soybean plants grown under flooding stress or non-stressed conditions. In addition, data on organellar proteins identified in mitochondria, nuclei, and endoplasmic reticulum are stored. Furthermore, the database integrates multiple omics data such as genomics, transcriptomics, metabolomics, and proteomics. The SPD database is accessible at http://proteome.dc.affrc.go.jp/Soybean/. The Soybean Proteome Database stores data obtained from both gel-based and gel-free proteomic techniques. The gel-free proteomic database comprises 39,212 proteins identified in 63 sample sets, such as different organs of soybean plants grown under flooding stress or non-stressed conditions in a time-dependent manner. In addition, organellar proteins identified in mitochondria, nuclei, and endoplasmic reticulum are stored in the gel-free proteomics database. A total of 44,704 proteins, including 5490 proteins identified using a gel-based proteomic technique, are stored in the SPD. It accounts for approximately 80% of all predicted proteins from

  19. Pulse Field Gel Electrophoresis.

    Science.gov (United States)

    Sharma-Kuinkel, Batu K; Rude, Thomas H; Fowler, Vance G

    2016-01-01

    Pulse Field Gel Electrophoresis (PFGE) is a powerful genotyping technique used for the separation of large DNA molecules (entire genomic DNA) after digesting it with unique restriction enzymes and applying to a gel matrix under the electric field that periodically changes direction. PFGE is a variation of agarose gel electrophoresis that permits analysis of bacterial DNA fragments over an order of magnitude larger than that with conventional restriction enzyme analysis. It provides a good representation of the entire bacterial chromosome in a single gel with a highly reproducible restriction profile, providing clearly distinct and well-resolved DNA fragments.

  20. In Vitro Reassembly of Tobacco Ribulose-1,5-bisphosphate Carboxylase/ Oxygenase from Fully Denatured Subunits

    Institute of Scientific and Technical Information of China (English)

    Zhen-Hua YONG; Gen-Yun CHEN; Jiao-Nai SHI; Da-Quan XU

    2006-01-01

    It has been generally proved impossible to reassemble ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) from fully denatured subunits in vitro in higher plant, because large subunit of fully denatured Rubisco is liable to precipitate when the denaturant is removed by common methods of direct dilution and one-step dialysis. In our experiment, the problem of precipitation was resolved by an improved gradual dialysis method, which gradually decreased the concentration of denaturant. However, fully denatured Rubisco subunits still could not be reassembled into holoenzyme using gradual dialysis unless chaperonin 60was added. The restored activity of reassembled Rubisco was approximately 8% of natural enzyme. The quantity of reassembled Rubisco increased greatly when heat shock protein 70 was present in the reassembly process. ATP and Mg2+ were unnecessary for in vitro reassembly of Rubisco, and Mg2+ inhibited the reassembly process. The reassembly was weakened when ATP, Mg2+ and K+ existed together in the reassembly process.

  1. Dynamic light scattering study of peanut agglutinin: Size, shape and urea denaturation

    Indian Academy of Sciences (India)

    Sagarika Dev; Avadhesha Surolia

    2006-12-01

    Peanut agglutinin (PNA) is a homotetrameric protein with a unique open quaternary structure. PNA shows non-two state profile in chaotrope induced denaturation. It passes through a monomeric molten globule like state before complete denaturation (Reddy et al 1999). This denaturation profile is associated with the change in hydrodynamic radius of the native protein. Though the molten globule-like state is monomeric in nature it expands in size due to partial denaturation. The size and shape of the native PNA as well as the change in hydrodynamic radius of the protein during denaturation has been studied by dynamic light scattering (DLS). The generation of two species is evident from the profile of hydrodynamic radii. This study also reveals the extent of compactness of the intermediate state.

  2. In situ observation of collagen thermal denaturation by second harmonic generation microscopy

    Science.gov (United States)

    Liao, C.-S.; Zhuo, Z.-Y.; Yu, J.-Y.; Chao, P.-H. G.; Chu, S.-W.

    2010-02-01

    Collagen denaturation is of fundamental importance for clinical treatment. Conventionally, the denaturation process is quantified by the shrinkage of collagen fibers, but the underlying molecular origin has not been fully understood. Since second harmonic generation (SHG) is related to the molecular packing of the triple helix in collagen fibers, this nonlinear signal provides an insight of molecular dynamics during thermal denaturation. With the aid of SHG microscopy, we found a new step in collagen thermal denaturation process, de-crimp. During the de-crimp step, the characteristic crimp pattern of collagen fascicles disappeared due to the breakage of interconnecting bonds between collagen fibrils, while SHG intensity remained unchanged, suggesting the intactness of the triple helical molecules. At higher temperature, shrinkage is observed with strongly reduced SHG intensity, indicating denaturation at the molecular level.

  3. Refolding of detergent-denatured lysozyme using β-cyclodextrin-assisted ion exchange chromatography.

    Science.gov (United States)

    Zhang, Li; Zhang, Qinming; Wang, Chaozhan

    2013-03-01

    Chromatography-based protein refolding is widely used. Detergent is increasingly used for protein solubilization from inclusion bodies. Therefore, it is necessary to develop a refolding method for detergent-denatured/solubilized proteins based on liquid chromatography. In the present work, sarkosyl-denatured/dithiothreitol-reduced lysozyme was used as a model, and a refolding method based on ion exchange chromatography, assisted by β-cyclodextrin, was developed for refolding detergent-denatured proteins. Many factors affecting the refolding, such as concentration of urea, concentration of β-cyclodextrin, pH and flow rate of mobile phases, were investigated to optimize the refolding conditions for sarkosyl-denatured lysozymes. The results showed that the sarkosyl-denatured lysozyme could be successfully refolded using β-cyclodextrin-assisted ion exchange chromatography.

  4. CONFORMANCE IMPROVEMENT USING GELS

    Energy Technology Data Exchange (ETDEWEB)

    Randall S. Seright

    2003-09-01

    This report describes work performed during the second year of the project, ''Conformance Improvement Using Gels.'' The project has two objectives. The first objective is to identify gel compositions and conditions that substantially reduce flow through fractures that allow direct channeling between wells, while leaving secondary fractures open so that high fluid injection and production rates can be maintained. The second objective is to optimize treatments in fractured production wells, where the gel must reduce permeability to water much more than that to oil. Pore-level images from X-ray computed microtomography were re-examined for Berea sandstone and porous polyethylene. This analysis suggests that oil penetration through gel-filled pores occurs by a gel-dehydration mechanism, rather than a gel-ripping mechanism. This finding helps to explain why aqueous gels can reduce permeability to water more than to oil. We analyzed a Cr(III)-acetate-HPAM gel treatment in a production well in the Arbuckle formation. The availability of accurate pressure data before, during, and after the treatment was critical for the analysis. After the gel treatment, water productivity was fairly constant at about 20% of the pre-treatment value. However, oil productivity was stimulated by a factor of 18 immediately after the treatment. During the six months after the treatment, oil productivity gradually decreased to approach the pre-treatment value. To explain this behavior, we proposed that the fracture area open to oil flow was increased substantially by the gel treatment, followed by a gradual closing of the fractures during subsequent production. For a conventional Cr(III)-acetate-HPAM gel, the delay between gelant preparation and injection into a fracture impacts the placement, leakoff, and permeability reduction behavior. Formulations placed as partially formed gels showed relatively low pressure gradients during placement, and yet substantially reduced the

  5. Flow of oil and water through elastic polymer gels; Ecoulement de petrole et d'eau a travers des gels de polymere elastiques

    Energy Technology Data Exchange (ETDEWEB)

    Al-Sharji, H.H.; Grattoni, C.A.; Zimmerman, R.W. [T.H. Huxley School of Environment, Earth Sciences and Engineering, Imperial College of Science, Technology and Medecine, London (United Kingdom); Dawe, R.A. [West Indies Univ., Dept. of Chemical Engineering, Mona (Jamaica)

    2001-07-01

    High water production is one of the major problems faced by the petroleum industry. One method of controlling water production is to inject polymer gels into the near-wellbore formation. Unfortunately, polymer gel injections are not always successful, in part because the exact mechanisms by which they reduce water permeability more than oil permeability (i.e., Disproportionate Permeability Reduction. DPR) are not understood. We have conducted a series of experiments on flow of water and oil through bulk polymer gels and through polymer- filled micro-models to elucidate the fundamental mechanisms involved in DPR. Flow experiments of oil and water through weak polyacrylamide-based gels have been per,formed to obtain the gel permeabilities under different test conditions. Oil and water permeabilities through the gel were each found to vary with flow rate according to a power-law, but with different pre-factors and exponents. The micro-scale flow experiments were conducted in transparent glass models to visualize clearly the flow events. Our observations enabled us to discount many previously-proposed explanations, and identify the fact that the oil and water can travel through the same pore channels, but in ways that differ, particularly at the pore scale. Water flows through the gel matrix as if flowing by diffusive flow through a porous medium, whereas the oil pushes its way In the form of immiscible drops or filaments. This difference in flow regime gives rise to the measured disproportionate permeability reduction. (authors)

  6. Dissociative mechanism of F-actin thermal denaturation.

    Science.gov (United States)

    Mikhailova, V V; Kurganov, B I; Pivovarova, A V; Levitsky, D I

    2006-11-01

    We have applied differential scanning calorimetry to investigate thermal unfolding of F-actin. It has been shown that the thermal stability of F-actin strongly depends on ADP concentration. The transition temperature, T(m), increases with increasing ADP concentration up to 1 mM. The T(m) value also depends on the concentration of F-actin: it increases by almost 3 degrees C as the F-actin concentration is increased from 0.5 to 2.0 mg/ml. Similar dependence of the T(m) value on protein concentration was demonstrated for F-actin stabilized by phalloidin, whereas it was much less pronounced in the presence of AlF4(-). However, T(m) was independent of protein concentration in the case of monomeric G-actin. The results suggest that at least two reversible stages precede irreversible thermal denaturation of F-actin; one of them is dissociation of ADP from actin subunits, and another is dissociation of subunits from the ends of actin filaments. The model explains why unfolding of F-actin depends on both ADP and protein concentration.

  7. GelTouch

    DEFF Research Database (Denmark)

    Miruchna, Viktor; Walter, Robert; Lindlbauer, David

    2015-01-01

    We present GelTouch, a gel-based layer that can selectively transition between soft and stiff to provide tactile multi-touch feedback. It is flexible, transparent when not activated, and contains no mechanical, electromagnetic, or hydraulic components, resulting in a compact form factor (a 2mm thin...... touchscreen layer for our prototype). The activated areas can be morphed freely and continuously, without being limited to fixed, predefined shapes. GelTouch consists of a poly(N-isopropylacrylamide) gel layer which alters its viscoelasticity when activated by applying heat (>32 C). We present three different...... a tablet with 6x4 tactile areas, enabling a tactile numpad, slider, and thumbstick. We show that the gel is up to 25 times stiffer when activated and that users detect tactile features reliably (94.8%)....

  8. A review of thermally stable gels for fluid diversion in petroleum production

    Energy Technology Data Exchange (ETDEWEB)

    Moradi-Araghi, A. [103 GB Philips Research Center, Phillips Petroleum Company, Bartlesville, OK (United States)

    2000-05-01

    The use of water-soluble polymers coupled with proper concentration of cross-linker(s) as flow-diverting agents have become a common practice in recent years for oil recovery applications. In such practice a solution containing the polymer and cross-linker(s), referred to as gelant, is injected in desired zones and allowed sufficient time to set into a solid gel. These gels are used in injection wells to divert the flow of injected water or gas (CO{sub 2}) to un-swept zones where additional oil can be recovered. The gels are also used to shut off the flow of water that strongly interferes with hydrocarbon production and substantially reduces the profitability of wells. There are a number of gelling systems available for treatment of lower temperature reservoirs. However, gels that can tolerate the harsh conditions of elevated temperatures and high salinity and divalent cations commonly present in deeper reservoirs are limited. When high molecular weight polyacrylamides are cross-linked to treat these hot reservoirs, their acrylamide groups will thermally hydrolyze. The resulting gel will further cross-link with the divalent cations available in the media, shrinking it to a fraction of its original volume. This process, which is referred to syneresis, can be avoided by selection of acrylamide-based polymers that are protected from extensive thermal hydrolysis. While other remedies such as lower-molecular-weight polyacrylamides, retarding agents or cooling of the target zones are attempted; these options often create unintended results. Recent studies include gelation of high molecular weight polyacrylamides with hydroquinone (HQ) and hexamethylenetetramine (HMTA) or terephtalaldehyde, terphthalic acid with hyroquinone, dihydroxynaphthalene and dibasic esters. These gelling systems are often prepared in seawater and require 2% sodium bicarbonate for their stability. Due to health and environmental concerns, the use of compounds such as HQ and formaldehyde is

  9. Nonisothermal denaturation kinetics of human hair and the effects of oxidation.

    Science.gov (United States)

    Wortmann, F-J; Popescu, C; Sendelbach, G

    2006-12-15

    Human hair as alpha-keratin fiber exhibits a complex morphology, which for the context of this investigation is considered as a filament/matrix-composite, comprising the intermediate filaments (IF) and a variety of amorphous protein components as matrix. Differential scanning calorimetry (DSC) under aqueous conditions was used to analyze the denaturation of the alpha-helical material in the IFs and to assess the changes imparted by repeated, oxidative bleaching processes. The DSC curves were submitted to kinetic analysis by applying the Friedman method and assuming first order kinetics. It was found that the course of the denaturation process remains largely unchanged through oxidation, despite the fact that pronounced decreases of denaturation temperature as well as of enthalpy occur. In parallel, the reaction rate constant at the denaturation temperature, k(TD), increases with repeated treatments, that is with cumulative chemical modification. However, this effect is in fact small compared to the overall change of k(T) through the denaturation process. This leads to conclude that once the temperature rise in combination with the chemical change has induced a suitable drop of the viscosity of the matrix around the IFs, denaturation of the remaining helical material occurs along a pathway that is largely independent of temperature and of the pretreatment history. This emphasizes the kinetic control of the matrix over the denaturation process of the helical segments in the filament/matrix composite. Copyright 2006 Wiley Periodicals, Inc.

  10. Studies on the refolding of the reduced-denatured insulin with size exclusion chromatography

    Institute of Scientific and Technical Information of China (English)

    BAI Quan; KONG Yu; DONG Cuihua; GENG Xindu

    2005-01-01

    The refolding of the reduced-denatured insulin from bovine pancreas was investigated with the size exclusion chromatography (SEC). It was shown that the reduced-denatured insulin originally denatured with 7.0 mol·L-1 guanidine hydrochloride (GuHCl) or 8.0 mol·L-1 urea could not be refolded with a non-oxidized mobile phase. Although the oxidized and reduced glutathione (GSSG and GSH) were employed in the oxidized mobile phase, the reduced-denatured insulin still could not be renatured. However, in the presence of 2.0 mol·L-1 urea in the oxidized mobile phase employed, the reduced-denatured insulin can be refolded with SEC, and the aggregation of denatured insulin can be diminished by urea. In addition, the disulfide exchange of reduced-denatured insulin also can be accelerated with GSSG/GSH in the oxidized mobile phase. The three disulfide bridges of insulin were formed correctly and the reduced-unfolded insulin can be renatured completely. The results were further tested with reversed-phase liquid chromatography (RPLC) and hydrophobic interaction chromatography (HIC).

  11. Assessment of collagen crosslinking and denaturation for the design of regenerative scaffolds.

    Science.gov (United States)

    Madaghiele, Marta; Calò, Emanuela; Salvatore, Luca; Bonfrate, Valentina; Pedone, Deborah; Frigione, Mariaenrica; Sannino, Alessandro

    2016-01-01

    Crosslinking and denaturation were two variables that deeply affected the performance of collagen-based scaffolds designed for tissue regeneration. If crosslinking enhances the mechanical properties and the enzymatic resistance of collagen, while masking or reducing the available cell binding sites, denaturation has very opposite effects, as it impairs the mechanical and the enzymatic stability of collagen, but increases the number of exposed cell adhesive domains. The quantification of both crosslinking and denaturation was thus fundamental to the design of collagen-based scaffolds for selected applications. The aim of this work was to investigate the extents of crosslinking and denaturation of collagen-based films upon dehydrothermal (DHT) treatment, that is, one of the most commonly employed methods for zero-length crosslinking that shows the unique ability to induce partial denaturation. Swelling measurements, differential scanning calorimetry, Fourier transform infrared spectroscopy, colorimetric assays for the quantification of primary amines, and mechanical tests were performed to analyze the effect of the DHT temperature on crosslinking and denaturation. In particular, chemically effective and elastically effective crosslink densities were evaluated. Both crosslinking and denaturation were found to increase with the DHT temperature, although according to different trends. The results also showed that DHT treatments performed at temperatures up to 120°C maintained the extent of denaturation under 25%. Coupling a mild DHT treatment with further crosslinking may thus be very useful not only to modulate the crosslink density, but also to induce a limited amount of denaturation, which shows potential to partially compensate the loss of cell binding sites caused by crosslinking.

  12. Urea-temperature phase diagrams capture the thermodynamics of denatured state expansion that accompany protein unfolding.

    Science.gov (United States)

    Tischer, Alexander; Auton, Matthew

    2013-09-01

    We have analyzed the thermodynamic properties of the von Willebrand factor (VWF) A3 domain using urea-induced unfolding at variable temperature and thermal unfolding at variable urea concentrations to generate a phase diagram that quantitatively describes the equilibrium between native and denatured states. From this analysis, we were able to determine consistent thermodynamic parameters with various spectroscopic and calorimetric methods that define the urea-temperature parameter plane from cold denaturation to heat denaturation. Urea and thermal denaturation are experimentally reversible and independent of the thermal scan rate indicating that all transitions are at equilibrium and the van't Hoff and calorimetric enthalpies obtained from analysis of individual thermal transitions are equivalent demonstrating two-state character. Global analysis of the urea-temperature phase diagram results in a significantly higher enthalpy of unfolding than obtained from analysis of individual thermal transitions and significant cross correlations describing the urea dependence of ΔH0 and ΔCP0 that define a complex temperature dependence of the m-value. Circular dichroism (CD) spectroscopy illustrates a large increase in secondary structure content of the urea-denatured state as temperature increases and a loss of secondary structure in the thermally denatured state upon addition of urea. These structural changes in the denatured ensemble make up ∼40% of the total ellipticity change indicating a highly compact thermally denatured state. The difference between the thermodynamic parameters obtained from phase diagram analysis and those obtained from analysis of individual thermal transitions illustrates that phase diagrams capture both contributions to unfolding and denatured state expansion and by comparison are able to decipher these contributions.

  13. Function, structure, and stability of enzymes confined in agarose gels.

    Directory of Open Access Journals (Sweden)

    Jeffrey Kunkel

    Full Text Available Research over the past few decades has attempted to answer how proteins behave in molecularly confined or crowded environments when compared to dilute buffer solutions. This information is vital to understanding in vivo protein behavior, as the average spacing between macromolecules in the cell cytosol is much smaller than the size of the macromolecules themselves. In our study, we attempt to address this question using three structurally and functionally different model enzymes encapsulated in agarose gels of different porosities. Our studies reveal that under standard buffer conditions, the initial reaction rates of the agarose-encapsulated enzymes are lower than that of the solution phase enzymes. However, the encapsulated enzymes retain a higher percentage of their activity in the presence of denaturants. Moreover, the concentration of agarose used for encapsulation had a significant effect on the enzyme functional stability; enzymes encapsulated in higher percentages of agarose were more stable than the enzymes encapsulated in lower percentages of agarose. Similar results were observed through structural measurements of enzyme denaturation using an 8-anilinonaphthalene-1-sulfonic acid fluorescence assay. Our work demonstrates the utility of hydrogels to study protein behavior in highly confined environments similar to those present in vivo; furthermore, the enhanced stability of gel-encapsulated enzymes may find use in the delivery of therapeutic proteins, as well as the design of novel strategies for biohybrid medical devices.

  14. Relaxation rate for an ultrafast folding protein is independent of chemical denaturant concentration.

    Science.gov (United States)

    Cellmer, Troy; Henry, Eric R; Kubelka, Jan; Hofrichter, James; Eaton, William A

    2007-11-28

    The connection between free-energy surfaces and chevron plots has been investigated in a laser temperature jump kinetic study of a small ultrafast folding protein, the 35-residue subdomain from the villin headpiece. Unlike all other proteins that have been studied so far, no measurable dependence of the unfolding/refolding relaxation rate on denaturant concentration was observed over a wide range of guanidinium chloride concentration. Analysis with a simple Ising-like theoretical model shows that this denaturant-invariant relaxation rate can be explained by a large movement of the major free energy barrier, together with a denaturant- and reaction coordinate-dependent diffusion coefficient.

  15. Heat Denaturation of Protein Structures and Chlorophyll States in PSII Membranes

    Institute of Scientific and Technical Information of China (English)

    李冬海; 阮翔; 许强; 王可玢; 公衍道; 匡廷云; 赵南明

    2002-01-01

    Heat denaturation is an important technique in the study of the structure and function of photosynthetic proteins. Heat denaturation of photosystem II (PSII) membrane was studied using circular dichroism (CD) spectroscopy, differential scanning calorimetry (DSC) and oxygen electrode. Complete loss of oxygen-evolving activity of the PSII membrane was observed at temperatures below 45℃. The decrease of excitonic interaction between chlorophyll molecules occurred more rapidly than the change of the protein secondary structure of the PSII membrane at temperatures above 45℃. The results indicate that the protein secondary structure of the membrane proteins in PSII membranes is more stable than the excitonic interaction between chlorophyll molecules during heat denaturation.

  16. Protein denaturation and functional properties of Lenient Steam Injection heat treated whey protein concentrate

    DEFF Research Database (Denmark)

    Dickow, Jonatan Ahrens; Kaufmann, Niels; Wiking, Lars

    2012-01-01

    Whey protein concentrate (WPC) was heat treated by use of the novel heat treatment method of Lenient Steam Injection (LSI) to elucidate new functional properties in relation to heat-induced gelation of heat treated WPC. Denaturation was measured by both DSC and FPLC, and the results of the two...... methods were highly correlated. Temperatures of up to 90 °C were applicable using LSI, whereas only 68 °C could be reached by plate heat exchange before coagulation/fouling. Denaturation of whey proteins increased with increasing heat treatment temperature up to a degree of 30–35% denaturation at 90 °C...

  17. On the Effect of Sodium Chloride and Sodium Sulfate on Cold Denaturation.

    Directory of Open Access Journals (Sweden)

    Andrea Pica

    Full Text Available Both sodium chloride and sodium sulfate are able to stabilize yeast frataxin, causing an overall increase of its thermodynamic stability curve, with a decrease in the cold denaturation temperature and an increase in the hot denaturation one. The influence of low concentrations of these two salts on yeast frataxin stability can be assessed by the application of a theoretical model based on scaled particle theory. First developed to figure out the mechanism underlying cold denaturation in water, this model is able to predict the stabilization of globular proteins provided by these two salts. The densities of the salt solutions and their temperature dependence play a fundamental role.

  18. Chemical characterization of the gels produced by the diazotrophic bacteria Rhizobium tropici and Mesorhizobium sp; Caracterizacao quimica dos geis produzidos pelas bacterias diazotroficas Rhizobium tropici e Mesorhizobium sp.

    Energy Technology Data Exchange (ETDEWEB)

    Monteiro, Nilson Kobori [Departamento de Engenharia e Tecnologia de Alimentos, Instituto de Biociencias, Letras e Ciencias Exatas, Universidade Estadual Paulista, Sao Jose do Rio Preto - SP (Brazil); Aranda-Selverio, Gabriel; Exposti, Diego Tadeu Degli; Silva, Maria de Lourdes Corradi da [Departamento de Fisica, Quimica e Biologia, Faculdade de Ciencias e Tecnologia, Universidade Estadual Paulista, Presidente Prudente - SP (Brazil); Lemos, Eliana Gertrudes Macedo; Campanharo, Joao Carlos [Departamento de Tecnologia, Faculdade de Ciencias Agrarias e Veterinaria, Universidade Estadual Paulista, Jaboticabal - SP (Brazil); Silveira, Joana Lea Meira [Departamento de Bioquimica e Biologia Molecular, Universidade Federal do Parana, Curitiba - PR (Brazil)

    2012-07-01

    The exopolysaccharides with characteristics of gel produced by Rhizobium tropici (EPSRT) and Mesorhizobium sp (EPSMR) are acidic heteropolysaccharide composed mainly of glucose and galactose in a molar ratio of 4:1 and 5:1 respectively, with traces of mannose ({approx} 1%). Chemical analysis showed the presence of uronic acid, pyruvate and acetyl-substituents in the structures of both polymers. Experiments of gel permeation chromatography and polyacrylamide gel electrophoresis showed that EPSRT and EPSMR are homogeneous molecules with low grade of polydispersity. The EPS were characterized using spectroscopic techniques of FT-IR, {sup 1}H and {sup 13}C-NMR. (author)

  19. A gel probe equilibrium sampler for measuring arsenic porewater profiles and sorption gradients in sediments: I. Laboratory development

    Science.gov (United States)

    Campbell, K.M.; Root, R.; O'Day, P. A.; Hering, J.G.

    2008-01-01

    A gel probe equilibrium sampler has been developed to study arsenic (As) geochemistry and sorption behavior in sediment porewater. The gels consist of a hydrated polyacrylamide polymer, which has a 92% water content. Two types of gels were used in this study. Undoped (clear) gels were used to measure concentrations of As and other elements in sediment porewater. The polyacrylamide gel was also doped with hydrous ferric oxide (HFO), an amorphous iron (Fe) oxyhydroxide. When deployed in the field, HFO-doped gels introduce a fresh sorbent into the subsurface thus allowing assessment of in situ sorption. In this study, clear and HFO-doped gels were tested under laboratory conditions to constrain the gel behavior prior to field deployment. Both types of gels were allowed to equilibrate with solutions of varying composition and re-equilibrated in acid for analysis. Clear gels accurately measured solution concentrations (??1%), and As was completely recovered from HFO-doped gels (??4%). Arsenic speciation was determined in clear gels through chromatographic separation of the re-equilibrated solution. For comparison to speciation in solution, mixtures of As(III) and As(V) adsorbed on HFO embedded in gel were measured in situ using X-ray absorption spectroscopy (XAS). Sorption densities for As(III) and As(V) on HFO embedded in gel were obtained from sorption isotherms at pH 7.1. When As and phosphate were simultaneously equilibrated (in up to 50-fold excess of As) with HFO-doped gels, phosphate inhibited As sorption by up to 85% and had a stronger inhibitory effect on As(V) than As(III). Natural organic matter (>200 ppm) decreased As adsorption by up to 50%, and had similar effects on As(V) and As(III). The laboratory results provide a basis for interpreting results obtained by deploying the gel probe in the field and elucidating the mechanisms controlling As partitioning between solid and dissolved phases in the environment. ?? 2008 American Chemical Society.

  20. The influence of applied heat treatments on whey protein denaturation

    Directory of Open Access Journals (Sweden)

    Fetahagić Safet

    2002-01-01

    . Distribution of nitrogen matter from milk 8%+3%DWP heat treated at 85ºC/10 min, 90ºC/10 min and 95ºC/10 min to sera samples were 9.64%, 8.66% and 8.67%, respectively. Whey protein denaturation increased with increasing of the temperature of the applied heat treatment. Denaturation was the most significant for milk sample 11%.

  1. Polyacrylamide scaffolds for studying cellular response to substrate stiffness in three dimensions

    Science.gov (United States)

    Lin, Keng-Hui

    2013-03-01

    Recent developments in two-dimensional (2D) culture substrates with tunable stiffness and patterned adhesion ligands have demonstrated that biochemical and mechanical cues regulate the biological functions of living cells. We have extended these cell culture platforms into three dimensions (3D), as in complex biological systems, by producing highly ordered scaffolds of polyacrylamide coated with extracellular matrix proteins. We characterized parameters for the scaffold fabrication. We then grew individual fibroblasts in the identical pores of our scaffolds, examing cellular morphological, cytoskeletal, and adhesion properties. We have observed rich variety of morphologies and anchoring strategies assumed by cells growing on our tunable 3D polyacrylamide scaffolds to demonstrate the richness of cell-mciroenvironment interactions when cell adhesions are not confined to 2D surfaces.

  2. Synthesis of crosslinked starch-graft-polyacrylamide-co-sodium xanthate and its performances in wastewater treatment.

    Science.gov (United States)

    Chang, Qing; Hao, Xuekui; Duan, Lili

    2008-11-30

    A novel crosslinked starch-graft-polyacrylamide-co-sodium xanthate (CSAX) was synthesized by grafting copolymerization reaction of corn starch, acrylamide (AM), and sodium xanthate using epichlorohydrin (EPI) as cross-linking reagent and ceric ammonium nitrate (CAN) as initiator in aqueous solution. CSAX was characterized by FTIR and element analysis. The performances of CSAX in wastewater treatment were evaluated by flocculation experiment. The results show that the CSAX was successfully synthesized and it has functions of removing both turbidity and copper ions from aqueous solution. It was proved that CSAX is more effective than crosslinked starch xanthate (CSX) and much more effective than crosslinked starch-graft-polyacrylamide (CSA) for removing copper ions. It was also proved that CSAX is little less effective than CSA, but much more effective than CSX for removing turbidity.

  3. Phosphate sensing by fluorecent reporter proteins embedded in poly-acrylamide nanoparticles

    DEFF Research Database (Denmark)

    Sun, Honghao; Scharff-Poulsen, Anne Marie; Gu, Hong;

    2008-01-01

    by soluble proteases to some extent. This nanoparticle embedding method provides a novel promising tool for in vivo metabolite studies. It also demonstrates a universal method for embedding different fragile bioactive elements, such as antibodies, genes, enzymes, and other functional proteins......Phosphate sensors were developed by embedding fluorescent reporter proteins (FLIPPi) in polyacrylamide nanoparticles; with diameters from 40 to 120 nm. The sensor activity and protein loading efficiency varied according to nanoparticle composition, that is, the total monomer content (% T......) and the cross-linker content (% C). Nanoparticles with 28% T and 20% C were considered optimal as a result of relatively high loading efficiency (50.6%) as well as high protein activity (50%). The experimental results prove that the cross-linked polyacrylamide matrix could protect FLIPPi from degradation...

  4. Protein folding by distributed computing and the denatured state ensemble.

    Science.gov (United States)

    Marianayagam, Neelan J; Fawzi, Nicolas L; Head-Gordon, Teresa

    2005-11-15

    The distributed computing (DC) paradigm in conjunction with the folding@home (FH) client server has been used to study the folding kinetics of small peptides and proteins, giving excellent agreement with experimentally measured folding rates, although pathways sampled in these simulations are not always consistent with the folding mechanism. In this study, we use a coarse-grain model of protein L, whose two-state kinetics have been characterized in detail by using long-time equilibrium simulations, to rigorously test a FH protocol using approximately 10,000 short-time, uncoupled folding simulations starting from an extended state of the protein. We show that the FH results give non-Poisson distributions and early folding events that are unphysical, whereas longer folding events experience a correct barrier to folding but are not representative of the equilibrium folding ensemble. Using short-time, uncoupled folding simulations started from an equilibrated denatured state ensemble (DSE), we also do not get agreement with the equilibrium two-state kinetics because of overrepresented folding events arising from higher energy subpopulations in the DSE. The DC approach using uncoupled short trajectories can make contact with traditionally measured experimental rates and folding mechanism when starting from an equilibrated DSE, when the simulation time is long enough to sample the lowest energy states of the unfolded basin and the simulated free-energy surface is correct. However, the DC paradigm, together with faster time-resolved and single-molecule experiments, can also reveal the breakdown in the two-state approximation due to observation of folding events from higher energy subpopulations in the DSE.

  5. Association of Stremptomyces community composition determined by PCR-denaturing gradient gel electrophoresis with indoor mold status.

    Science.gov (United States)

    Abstract Both Streptomyces species and mold species have previously been isolated from moisture-damaged building materials; however, an association between these two groups of microorganisms in indoor environments is not clear. In this study, we used a cultureindependent met...

  6. Assessment of the genotypic diversity of antibiotic-producing Pseudomonas species in the rhizosphere by denaturing gradient gel electrophoresis

    NARCIS (Netherlands)

    Bergsma-Vlami, M.; Prins, M.E.; Staats, M.; Raaijmakers, J.M.

    2005-01-01

    The genotypic diversity of antibiotic-producing Pseudomonas spp. provides an enormous resource for identifying strains that are highly rhizosphere competent and superior for biological control of plant diseases. In this study, a simple and rapid method was developed to determine the presence and gen

  7. High-resolution differentiation of cyanobacteria by using rRNA-Internal Transcribed Spacer denaturing gradient gel electrophoresis

    NARCIS (Netherlands)

    Janse, I.; Meima, M.; Kardinaal, W.E.A.; Zwart, G.

    2003-01-01

    For many ecological studies of cyanobacteria, it is essential that closely related species or strains can be discriminated. Since this is often not possible by using morphological features, cyanobacteria are frequently studied by using DNA-based methods. A powerful method for analysis of the diversi

  8. Association of Stremptomyces community composition determined by PCR-denaturing gradient gel electrophoresis with indoor mold status.

    Science.gov (United States)

    Abstract Both Streptomyces species and mold species have previously been isolated from moisture-damaged building materials; however, an association between these two groups of microorganisms in indoor environments is not clear. In this study, we used a cultureindependent met...

  9. Complexity of Vaginal Microflora as Analyzed by PCR Denaturing Gradient Gel Electrophoresis in a Patient With Recurrent Bacterial Vaginosis

    Directory of Open Access Journals (Sweden)

    Gregor Reid

    2005-01-01

    Full Text Available Objective:Gardnerella vaginalis has long been the most common pathogen associated with bacterial vaginosis (BV. We aimed to test our hypothesis that symptoms and signs of BV do not necessarily indicate colonization by this organism, and often will not respond to standard metronidazole or clindamycin treatment.

  10. Preparation of Dysprosium Ferrite/Polyacrylamide Magnetic Composite Microsphere and Its Characterization

    Institute of Scientific and Technical Information of China (English)

    Hidehiro Kumazawa; Wang Zhifeng; Zhou Lanxiang; Zhang Hong; Li Yourong; Zhang Ming

    2005-01-01

    Using the technique of microemulsion polymerization with nano-reactor, dysprosium ferrite/polyacrylamide magnetic composite microsphere was prepared by one-step method in a single inverse microemulsion. The structure, average particle size, morphology of composite microsphere were characterized by FTIR, XRD, TEM and TGA. The magnetic responsibility of composite microsphere was also investigated. The results indicate that the magnetic composite microsphere possess high magnetic responsibility and suspension stability.

  11. CONDUCTIVITY BEHAVIOR OF IONIC AI(OH)3-POLYACRYLAMIDE HYBRIDS IN AQUEOUS SOLUTIONS

    Institute of Scientific and Technical Information of China (English)

    Jin-wen Qian; Wu-yuan Yang; Xiao-jun Xiang; Zhi-quan Shen; Meng Wang; Wei-feng Jiang

    2004-01-01

    The conductivity behavior of Al(OH)3-acrylamide hybrid polyacrylamide (hybrid PAAm) in distilled water was studied. A discontinuity phenomenon of the conductivity (k) versus concentration (c) curve of the hybrid PAAm in a certain concentration regime is found. This phenomenon is dependent on the molecular weight of the hybrid PAAm and on the particle size and content of the Al(OH)3 colloid in the hybrid PAAm. This phenomenon was accounted for assuming ionization of the hybrid PAAm.

  12. An Alternative Way towards Preparation of Hydrophobically Associating Polyacrylamide:Chemical Post-Modification

    Institute of Scientific and Technical Information of China (English)

    FENG Yu-jun

    2004-01-01

    Hydrophobically associating polyacrylamides (HAPAMs) are derivatives from polyacrylamides by incorporating a small amount of hydrophobic moieties along the water-soluble mainchain. They are now becoming a class of promising candidates as thickeners or rheology modifiers in the formulations where rheology is necessary to be regulated, such as tertiary oil recovery, drilling fluids, hydraulic fracturing and coatings. Due to association of hydrophobes in nano-domains, their aqueous solutions exhibit very interesting rheological properties and better stability against salts than the unmodified precursor, polyacrylamide.Generally, there are two synthetic routes to introduce hydrophobic portion onto water-soluble polymer chains; i.e., direct copolymerization of hydrophobic and hydrophilic monomers, and post-polymerization functionalization[1]. In the case of HAPAM polymers, a commonly accepted method is micellar copolymerization in which an appropriate surfactant is employed to solubilize both monomers. However, it is widely reported[2] that the obtained polymers via micellar polymerization are characterized by: (i) blocky distribution of the hydrophobes; (ii) compositional inhomogeneity and (iii) strong dependence of solution properties on the block length.In this work, the alternative process, i.e., chemical post-modification, is employed to synthesize HAPAM polymers by direct N-alkylation of parent polyacrylamide (Figure 1) in dimethyl sulfoxide[3,4].PAM HAPAMFig. 1 Schematic route to prepare HAPAM by direct N-alkylation of PAMIt is found that the final incorporation of hydrophobic groups is in good agreement with the feed ratio[4], in contrast with that from micellar copolymerization which always brings about composition drift. Furthermore, unique rheological responses to shear rate, salt, temperature are also evidenced[5].

  13. A Novel Polyacrylamide Magnetic Nanoparticle Contrast Agent for Molecular Imaging using MRI

    Directory of Open Access Journals (Sweden)

    Bradford A. Moffat

    2003-10-01

    Full Text Available A novel Polyacrylamide superparamagnetic iron oxide nanoparticle platform is described which has been synthetically prepared such that multiple crystals of iron oxide are encapsulated within a single Polyacrylamide matrix (PolyAcrylamide Magnetic [PAM] nanoparticles. This formulation provides for an extremely large T2 and T2* relaxivity of between 620 and 1140 sec−1 mM−1. Administration of PAM nanoparticles into rats bearing orthotopic 9L gliomas allowed quantitative pharmacokinetic analysis of the uptake of nanoparticles in the vasculature, brain, and glioma. Addition of polyethylene glycol of varying sizes (0.6, 2, and 10 kDa to the surface of the PAM nanoparticles resulted in an increase in plasma half-life and affected tumor uptake and retention of the nanoparticles as quantified by changes in tissue contrast using MRI. The flexible formulation of these nanoparticles suggests that future modifications could be accomplished allowing for their use as a targeted molecular imaging contrast agent and/or therapeutic platform for multiple indications.

  14. Horseradish peroxidase embedded in polyacrylamide nanoparticles enables optical detection of reactive oxygen species

    DEFF Research Database (Denmark)

    Poulsen, A.K.; Scharff-Poulsen, Anne Marie; Olsen, L.F.

    2007-01-01

    We have synthesized and characterized new nanometer-sized polyacrylamide particles containing horseradish peroxidase and fluorescent dyes. Proteins and dyes are encapsulated by radical polymerization in inverse microemulsion. The activity of the encapsulated enzyme has been examined and it mainta......We have synthesized and characterized new nanometer-sized polyacrylamide particles containing horseradish peroxidase and fluorescent dyes. Proteins and dyes are encapsulated by radical polymerization in inverse microemulsion. The activity of the encapsulated enzyme has been examined...... and it maintains its ability to catalyze the oxidation of guaiacol with hydrogen peroxide as the electron acceptor, although at a slightly lower rate compared to that of the free enzyme in solution. The embedded enzyme is also capable of catalyzing the peroxidase-oxidase reaction. However, the rate is decreased...... by a factor of 2-3 compared to that of the free enzyme. The reduced rate is probably due to limitation of diffusion of substrates and products into and out of the particles. The catalytic activity of horseradish peroxidase in the polyacrylamide matrix demonstrates that the particles have pores which are large...

  15. Study on Synthesis of Polyacrylamide Microspheres and its Sealing Characteristics of Drilling Fluid

    Directory of Open Access Journals (Sweden)

    Mingming Cheng

    2014-02-01

    Full Text Available For the purpose of reducing the invasion depth of solid phase and liquid phase of drilling fluid and improving the quality of drilling mud cake, polyacrylamide microspheres were successfully composed via reversed phrase emulsion method. The physical and chemical structures of the samples as well as the grain-size variation were investigated by Transmission Electron Microscopy (TEM and Laser Particle Size Analysis method (LPSA.The plugging performance of the samples were investigated by core plugging experiment and sand bed filtration experiment. TEM images show that acrylamide has been polymerized, the products consist of a lot of monodisperse microspheres with a size of about 1-6 μm. LPSA images indicate that the particle size of polyacrylamide microspheres vary with time and temperature regularly. The experiment that simulates rock strata with different permeability shows those polyacrylamide microspheres can produce effectively blocked to the core and sand bed and then it establishes the foundation for the next field test.

  16. Laboratory and Field Evaluations of Polyacrylamide Hydrogel Baits Against Argentine Ants (Hymenoptera: Formicidae).

    Science.gov (United States)

    Rust, Michael K; Soeprono, Andrew; Wright, Sarajean; Greenberg, Les; Choe, Dong-Hwan; Boser, Christina L; Cory, Coleen; Hanna, Cause

    2015-06-01

    The development of effective baits to control the Argentine ant, Linepithema humile (Mayr), has been problematic because foragers prefer sweet liquids, while many toxicants are insoluble in water and liquid baits are generally difficult to deliver. The incorporation of thiamethoxam and sucrose solutions into a water-absorbing polyacrylamide hydrogel provides a unique and novel carrier and method of application for liquid baits. Formulations of thiamethoxam affected the size of the hydrogels, and sucrose solutions containing 0.0003% technical thiamethoxam provided hydrogels as large as those made with 25% sucrose solution or deionized water. Concentrations of thiamethoxam as low as 0.000075% in the hydrogels provided 50% kill of workers within 3 d in a laboratory setting. In small colony studies, baiting with 0.00015 and 0.000075% thiamethoxam hydrogels provided 100% mortality of workers and queens within 8 d. An enzyme-linked immunosorbent assay indicated that thiamethoxam was absorbed into the interior of the polyacrylamide matrix. The water loss rates of the hydrogels were dependent upon the relative humidity. Polyacrylamide hydrogels with >50% water loss were less attractive to ants. Field studies in highly infested areas indicated that concentrations of 0.0006 or 0.0018% thiamethoxam were more effective than 0.00015%. Hydrogels may provide a cost-effective alternative to providing aqueous baits to control Argentine ants. © The Authors 2015. Published by Oxford University Press on behalf of Entomological Society of America. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

  17. Gel for simultaneous chemical imaging of anionic and cationic solutes using diffusive gradients in thin films.

    Science.gov (United States)

    Kreuzeder, Andreas; Santner, Jakob; Prohaska, Thomas; Wenzel, Walter W

    2013-12-17

    We report on a novel gel based on diffusive gradients in thin films (DGT) for the simultaneous measurement of cations and anions and its suitability for high resolution chemical imaging by using laser ablation inductively coupled plasma mass spectrometry (LA-ICPMS). The new high resolution mixed binding gel (HR-MBG) is based on zirconium-hydroxide and suspended particulate reagent-iminodiacetate (SPR-IDA) as resin materials which are embedded in an ether-based urethane polymer hydrogel. The use of this polymer hydrogel material allows the production of ultrathin, highly stable and tear-proof resin gel layers with superior handling properties compared to existing ultrathin polyacrylamide gels. The gel was characterized regarding its uptake kinetics, the anion and cation capacities, and the effects of pH, ionic strength, and aging on the performance of the HR-MBG. Our results demonstrate the capability of this novel gel for concomitant sampling of anions and cations. The suitability of this new gel type for DGT chemical imaging at submm spatial resolution in soils using LA-ICPMS is shown. 2D images of P, As, Co, Cu, Mn, and Zn distributions around roots of Zea mays L. demonstrate the new opportunities offered by the HR-MBG for high-resolution mapping of solute dynamics in soil and sediment hotspots, such as the rhizosphere, by simultaneous observation of anionic and cationic solute species.

  18. Three dimensional gel dosimetry by use of nuclear magnetic resonance imaging (MRI)

    Energy Technology Data Exchange (ETDEWEB)

    De Deene, Y.; De Wagter, C.; Van Duyse, B.; Achten, E.; De Neve, W. [Ghent Rijksuniversiteit (Belgium). Kliniek voor Radiotherapie en Kerngeneeskunde; De Poorter, J. [Ghent Univ. (Belgium). Dept. of Magnetic Resonance

    1995-12-01

    As co-monomers are found to polymerize by radiation, they are eligible for constructing a three dimensional dosimeter. Another kind of three dimensional dosimeter, based on the radiation sensitivity of the ferrous ions in a Fricke solution, was tested in a previous study. However, a major problem that occurs in this kind of gel dosimeters is the diffusion of the ferric and ferrous ions. The co-monomer gels are more stable. The degree of polymerisation is visualized with a clinical MRI system. Acrylamide and N,N-methylene-bis-acrylamide are dissolved in a gel composed of gelatin and water. By irradiation the co-monomers are polymerized to polyacrylamide. The gel is casted in humanoid forms. As such, a simulation of the irradiation of the patient can be performed. Magnetic resonance relaxivity images of the irradiated gel display the irradiation dose. The images of the gel are fused with the radiological images of the patient. Quantitation of the dose response of the co-monomer gel is obtained through calibration by test tubes.

  19. In-phantom dosimetry for BNCT with Fricke and normoxic-polymer gels

    Science.gov (United States)

    Gambarini, G.; Agosteo, S.; Carrara, M.; Gay, S.; Mariani, M.; Pirola, L.; Vanossi, E.

    2006-05-01

    Measurements of in-phantom dose distributions and images are important for Boron Neutron Capture Therapy treatment planning. The method for spatial determination of absorbed doses in thermal or epithermal neutron fields, based on Fricke-xylenol-orange-infused gel dosimeters in form of layers, has revealed to be very reliable, as gel layer dosimeters give the possibility of obtaining spatial dose distributions and measurements of each dose contribution in neutron fields, by means of a properly studied procedure. Quite recently, BNCT has been applied to treat liver metastases; in this work the results of in-phantom dosimetry for explanted liver in BNCT treatments are described. Moreover, polyacrylamide gel (PAG) dosimeters in which a polymerization process appears as a consequence of absorbed dose, have been recently tested, because of their characteristic absence of diffusion. In fact, due to the diffusion of ferric ions, Fricke-gel dosimeters require prompt analysis after exposure to avoid spatial information loss. In this work the preliminary results of a study about the reliability of polymer gel in BNCT dosimetry are also discussed. Gel layers have been irradiated in a phantom exposed in the thermal column of the TRIGA MARK II reactor (Pavia). The results obtained with the two kinds of gel dosimeter have been compared.

  20. Mechanically induced gel formation

    NARCIS (Netherlands)

    van Herpt, Jochem T.; Stuart, Marc C. A.; Browne, Wesley R.; Feringa, Ben L.

    2013-01-01

    Mechanical triggering of gelation of an organic solution by a carbazole-based bisurea organogelator is described. Both the duration of the mechanical stimulation and the gelator concentration control the gelation process and the characteristics of the gel obtained.