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Sample records for native polyacrylamide gel

  1. Resolving mitochondrial protein complexes using non-gradient blue native polyacrylamide gel electrophoresis

    PubMed Central

    Yan, Liang-Jun; Forster, Michael J.

    2009-01-01

    Blue native polyacrylamide gel electrophoresis (BN-PAGE) is a powerful technique for separation and proteomic analysis of high molecular weight protein complexes. It is often performed on gradient gels and is widely used for studying mitochondrial membrane complexes involved in electron transportation and oxidative phosphorylation. In this paper, we present an alternative BN-PAGE method that uses highly porous, non-gradient polyacrylamide gels for separation of rat brain mitochondrial protein complexes. Results demonstrate that this method not only resolves mitochondrial complexes I-V, allowing subsequent analysis by in-gel activity staining and mass spectrometry peptide sequencing, but also identifies Hsp60 polymers and dihydrolipoamide dehydrogenase (DLDH). Moreover, with this new method, it is shown for the first time that complex I and DLDH can be simultaneously detected on a single gel strip by in-gel activity staining. Overall, the method provides a simplified, non-gradient gel electrophoretic approach that should be useful in functional proteomics studies. PMID:19348780

  2. Analysis of Mitochondrial Respiratory Chain Supercomplexes Using Blue Native Polyacrylamide Gel Electrophoresis (BN-PAGE).

    PubMed

    Jha, Pooja; Wang, Xu; Auwerx, Johan

    2016-01-01

    Mitochondria are cellular organelles that harvest energy in the form of ATP through a process termed oxidative phosphorylation (OXPHOS), which occurs via the protein complexes of the electron transport chain (ETC). In recent years it has become unequivocally clear that mitochondrial complexes of the ETC are not static entities in the inner mitochondrial membrane. These complexes are dynamic and in mammals they aggregate in different stoichiometric combinations to form supercomplexes (SCs) or respirasomes. It has been proposed that the net respiration is more efficient via SCs than via isolated complexes. However, it still needs to be determined whether the activity of a particular SC is associated with a disease etiology. Here we describe a simplified method to visualize and assess in-gel activity of SCs and the individual complexes with good resolution using blue native polyacrylamide gel electrophoresis (BN-PAGE). © 2016 by John Wiley & Sons, Inc. PMID:26928661

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

    PubMed Central

    Tang, Shuiquan; Chan, Winnie W. M.; Fletcher, Kelly E.; Seifert, Jana; Liang, Xiaoming; Lffler, 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

  4. In-gel activity staining of oxidized nicotinamide adenine dinucleotide kinase by blue native polyacrylamide gel electrophoresis.

    PubMed

    Mailloux, Ryan J; Singh, Ranji; Appanna, Vasu D

    2006-12-15

    Oxidized nicotinamide adenine dinucleotide (NAD(+)) kinase (NADK, E.C. 2.7.1.23) plays an instrumental role in cellular metabolism. Here we report on a blue native polyacrylamide gel electrophoretic technique that allows the facile detection of this enzyme. The product, oxidized nicotinamide adenine dinucleotide phosphate (NADP(+)), formed following the reaction of NADK with NAD(+) and adenosine 5'-triphosphate was detected with the aid of glucose-6-phosphate dehydrogenase or NADP(+)-isocitrate dehydrogenase, iodonitrotetrazolium chloride, and phenazine methosulfate. The bands at the respective activity sites were excised and subjected to native and denaturing two-dimensional electrophoresis for the determination of protein levels. Hence this novel electrophoretic method allows the easy detection of NADK, a critical enzyme involved in pyridine homeostasis. Furthermore, this technique allowed the monitoring of the activity and expression of this kinase in various biological systems. PMID:17083911

  5. A method for in-gel fluorescent visualization of proteins after native and sodium dodecyl sulfate polyacrylamide gel electrophoresis.

    PubMed

    Pristov, Jelena Bogdanović; Opačić, Miloš; Dimitrijević, Milena; Babić, Nikolina; Spasojević, Ivan

    2015-07-01

    We have developed a simple one-step 30-min method for fluorescent visualization of proteins in native and sodium dodecyl sulfate polyacrylamide gel electrophoresis (PAGE) gels. The method is based on formation of strong fluorophores via potassium ferricyanide-provoked oxidation of tryptophan (Trp). Following PAGE, gels are soaked in water solution of potassium ferricyanide (100 mM) and NaOH (1 M) and are kept in the dark for 30 min. Gels are then transferred to water and scanned. The sensitivity of the method was slightly lower compared with standard Coomassie Brilliant Blue (CBB) staining. The method can be useful when rapid acquisition of data is of the essence. After preview, gels can be post-stained using the CBB protocol for further analysis. The intensity of fluorescence is dependent on Trp number, so the protocol might find application in the quantification of Trp residues as illustrated here. Importantly, there is room for improvement of the method. Namely, according to excitation-emission matrix analysis of stained protein bands, maximal fluorescence intensity (at 345/460 nm) was 3.5-fold higher compared with the settings that were available on a commercial imager (395/525 nm). As a supplement, we present an upgrade of the previously described method for in-gel detection of non-heme iron-binding proteins that also employs potassium ferricyanide. PMID:25862081

  6. Blue native polyacrylamide gel electrophoresis and the monitoring of malate- and oxaloacetate-producing enzymes.

    PubMed

    Singh, R; Chnier, D; Briault, R; Mailloux, R; Hamel, R D; Appanna, V D

    2005-09-30

    We demonstrate a facile blue native polyacrylamide gel electrophoresis (BN-PAGE) technique to detect two malate-generating enzymes, namely fumarase (FUM), malate synthase (MS) and four oxaloacetate-forming enzymes, namely pyruvate carboxylase (PC), phosphoenolpyruvate carboxykinase (PEPCK), citrate lyase (CL) and aspartate aminotransferase (AST). Malate dehydrogenase (MDH) was utilized as a coupling enzyme to detect either malate or oxaloacetate in the presence of their respective substrates and cofactors. The latter four oxaloacetate-forming enzymes were identified by 2,6-dichloroindophenol (DCIP) and p-iodonitrotetrazolium (INT) while the former two malate-producing enzymes were visualized by INT and phenazine methosulfate (PMS) in the reaction mixtures, respectively. The band formed at the site of enzymatic activity was easily quantified, while Coomassie staining provided information on the protein concentration. Hence, the expression and the activity of these enzymes can be readily evaluated. A two-dimensional (2D) BN-PAGE or SDS-PAGE enabled the rapid purification of the enzyme of interest. This technique also provides a quick and inexpensive means of quantifying these enzymatic activities in normal and stressed biological systems. PMID:16154636

  7. Electroblotting from Polyacrylamide Gels.

    PubMed

    Goldman, Aaron; Ursitti, Jeanine A; Mozdzanowski, Jacek; Speicher, David W

    2015-01-01

    Transferring proteins from polyacrylamide gels onto retentive membranes is now primarily used for immunoblotting. A second application that was quite common up to about a decade ago was electroblotting of proteins for N-terminal and internal sequencing using Edman chemistry. This unit contains procedures for electroblotting proteins from polyacrylamide gels onto a variety of membranes, including polyvinylidene difluoride (PVDF) and nitrocellulose. In addition to the commonly used tank or wet transfer system, protocols are provided for electroblotting using semidry and dry systems. This unit also describes procedures for eluting proteins from membranes using detergents or acidic extraction with organic solvents for specialized applications. 2015 by John Wiley & Sons, Inc. PMID:26521711

  8. The unravelling of metabolic dysfunctions linked to metal-associated diseases by blue native polyacrylamide gel electrophoresis.

    PubMed

    Han, Sungwon; Auger, Christopher; Castonguay, Zachary; Appanna, Varun P; Thomas, Sean C; Appanna, Vasu D

    2013-02-01

    Gel electrophoresis is routinely used to separate and analyse macromolecules in biological systems. Although many of these electrophoretic techniques necessitate the denaturing of the analytes prior to their analysis, blue native polyacrylamide gel electrophoresis (BN-PAGE) permits the investigation of proteins/enzymes and their supramolecular structures such as the metabolon in native form. This attribute renders this analytical tool conducive to deciphering the metabolic perturbations invoked by metal toxicity. In this review, we elaborate on how BN-PAGE has led to the discovery of the dysfunctional metabolic pathways associated with disorders such as Alzheimer's disease, Parkinson's disease, and obesity that have been observed as a consequence of exposure to various metal toxicants. PMID:23001308

  9. The monitoring of nucleotide diphosphate kinase activity by blue native polyacrylamide gel electrophoresis.

    PubMed

    Mailloux, Ryan J; Darwich, Rami; Lemire, Joseph; Appanna, Vasu

    2008-04-01

    Nucleoside diphosphate kinase (NDPK) has been shown to play a pivotal role in modulating a plethora of cellular processes. In this study, we report on a blue native (BN) PAGE technique which allows the facile assessment of NDPK activity and expression. The in-gel detection of NDPK relies on the precipitation of formazan at the site of immobilized enzyme activity. This is achieved by coupling the formation of ATP, as a consequence of gamma-phosphate transfer from NTP to ADP, to hexokinase (HK), glucose-6-phosphate dehydrogenase (G6PDH), oxidized nicotinamide adenine dinucleotide phosphate (NADP), phenazine methosulfate (PMS), and iodonitrotetrazolium chloride (INT). 2-D denaturing gel analysis confirmed that the activity bands corresponded to NDPK as indicated by subunit composition. Furthermore, the sensitivity and specificity of this readily accessible procedure was assessed by monitoring the in-gel activity of NDPK using different concentrations of GTP and CTP as well as deoxynucleoside triphosphates. This electrophoretic technique allows the quick and easy detection of NDPK, a housekeeping enzyme crucial to cell survival. PMID:18324728

  10. Histochemical staining and quantification of plant mitochondrial respiratory chain complexes using blue-native polyacrylamide gel electrophoresis.

    PubMed

    Sabar, Mohammed; Balk, Janneke; Leaver, Christopher J

    2005-12-01

    Our knowledge of the respiratory chain and associated defects depends on the study of the multisubunit protein complexes in the inner mitochondrial membrane. Functional analysis of the plant mitochondrial respiratory chain has been successfully achieved by a combination of blue-native polyacrylamide gel electrophoresis (BN-PAGE) for separation of the protein complexes, and in-gel histochemical staining of the enzyme activities. We have optimized this powerful technique by determining linear ranges of amount of protein and enzyme activity for each respiratory complex. Time courses of the in-gel enzyme activities were also performed to determine optimal reaction times. Using the in-gel activity staining method we have previously shown decreased activity of complex V (F(1)F(0)-ATPase) in male-sterile sunflowers (Sabar et al., 2003). Here we have identified unique supercomplexes comprising complex IV (cytochrome c oxidase) in sunflower mitochondria. This method therefore represents a reliable tool for the diagnosis of respiratory dysfunction. In addition, the wider application of BN-PAGE in combination with enzyme activity staining is discussed. PMID:16297078

  11. Detection and purification of glucose 6-phosphate dehydrogenase, malic enzyme, and NADP-dependent isocitrate dehydrogenase by blue native polyacrylamide gel electrophoresis.

    PubMed

    Briault, Robin; Chnier, Daniel; Singh, Ranji; Middaugh, Jeff; Mailloux, Ryan; Appanna, Vasu

    2005-08-01

    We describe a blue native polyacrylamide gel electrophoretic technique that allows the facile detection, quantitation and purification of three NADPH-producing enzymes. Glucose 6-phosphate dehydrogenase, malic enzyme and NADP-dependent isocitrate dehydrogenase were detected simultaneously. Activity staining based on the formation of NADPH from the respective substrates and the subsequent precipitation of formazan enabled the relative quantitation of enzymatic activities, while Coomassie staining on one-dimensional or two-dimensional gels helped monitor the amount of protein associated with these enzymatic activities. This technique provides a simple and effective route to obtain homogeneous protein for further analyses and also enables the screening of these NADPH-producing enzymes in various cellular systems. PMID:16078188

  12. Silver staining techniques of polyacrylamide gels.

    PubMed

    Bartsch, Holger; Arndt, Claudia; Koristka, Stefanie; Cartellieri, Marc; Bachmann, Michael

    2012-01-01

    Although the main application for polyacrylamide gels is the separation and subsequent blotting of proteins for immunodetection, there are tasks that need staining of proteins in the polyacrylamide gel. Several different staining techniques exist for protein staining in SDS gels that differ in their sensitivity, their expenditure of time, and other aspects. Still, silver staining is the most sensitive and reliable staining technique. Because this technique was developed in the 1970s, a huge number of variations exist. Therefore, we will provide herein three methods, which are robust and easy to perform. PMID:22585513

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

    PubMed Central

    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

  14. A blue native polyacrylamide gel electrophoretic technology to probe the functional proteomics mediating nitrogen homeostasis in Pseudomonas fluorescens.

    PubMed

    Han, Sungwon; Auger, Christopher; Appanna, Varun P; Lemire, Joseph; Castonguay, Zachary; Akbarov, Elchin; Appanna, Vasu D

    2012-09-01

    As glutamate and ammonia play a pivotal role in nitrogen homeostasis, their production is mediated by various enzymes that are widespread in living organisms. Here, we report on an effective electrophoretic method to monitor these enzymes. The in gel activity visualization is based on the interaction of the products, glutamate and ammonia, with glutamate dehydrogenase (GDH, EC: 1.4.1.2) in the presence of either phenazine methosulfate (PMS) or 2,6-dichloroindophenol (DCIP) and iodonitrotetrazolium (INT). The intensity of the activity bands was dependent on the amount of proteins loaded, the incubation time and the concentration of the respective substrates. The following enzymes were readily identified: glutaminase (EC: 3.5.1.2), alanine transaminase (EC: 2.6.1.2), aspartate transaminase (EC: 2.6.1.1), glycine transaminase (EC: 2.6.1.4), ornithine oxoacid aminotransferase (EC: 2.6.1.13), and carbamoyl phosphate synthase I (EC: 6.3.4.16). The specificity of the activity band was confirmed by high pressure liquid chromatography (HPLC) following incubation of the excised band with the corresponding substrates. These bands are amenable to further molecular characterization by a variety of analytical methods. This electrophoretic technology provides a powerful tool to screen these enzymes that contribute to nitrogen homeostasis in Pseudomonas fluorescens and possibly in other microbial systems. PMID:22595184

  15. Visible fluorescent detection of proteins in polyacrylamide gels without staining.

    PubMed

    Ladner, Carol L; Yang, Jing; Turner, Raymond J; Edwards, Robert A

    2004-03-01

    2,2,2-Trichloroethanol (TCE) incorporated into polyacrylamide gels before polymerization provides fluorescent visible detection of proteins in less than 5min of total processing time. The tryptophans in proteins undergo an ultraviolet light-induced reaction with trihalocompounds to produce fluorescence in the visible range so that the protein bands can be visualized on a 300-nm transilluminator. In a previous study trichloroacetic acid or chloroform was used to stain polyacrylamide gel electrophoresis (PAGE) gels for protein visualization. This study shows that placing TCE in the gel before electrophoresis can eliminate the staining step. The gel is removed from the electrophoresis apparatus and placed on a transilluminator and then the protein bands develop their fluorescence in less than 5min. In addition to being rapid this visualization method provides detection of 0.2microg of typical globular proteins, which for some proteins is slightly more sensitive than the standard Coomassie brilliant blue (CBB) method. Integral membrane proteins, which do not stain well with CBB, are visualized well with the TCE in-gel method. After TCE in-gel visualization the same gel can then be CBB stained, allowing for complementary detection of proteins. In addition, visualization with TCE in the gel is compatible with two-dimensional PAGE, native PAGE, Western blotting, and autoradiography. PMID:14769330

  16. Polyacrylamide Gel Electrophoresis for Purification of Large Amounts of RNA.

    PubMed

    Meyer, Mélanie; Masquida, Benoît

    2016-01-01

    Polyacrylamide gel electrophoresis (PAGE) constitutes a powerful technique for the efficient purification of RNA molecules dedicated to applications that require high purity levels. PAGE allows for the fractionation of RNA obtained from cell extracts, chemical or enzymatic synthesis, or modification experiments. Native or denaturing conditions can be chosen for analytical or preparative-scale separations and the nucleotide resolution can be tuned by changing the percentage and reticulation of the gel material. In this protocol, we focus on the preparation of milligram-scale amounts of ~200 nucleotides (nt) RNA molecules that were used in subsequent crystallization experiments. PMID:26227037

  17. Detection of chitin deacetylase activity after polyacrylamide gel electrophoresis.

    PubMed

    Trudel, J; Asselin, A

    1990-09-01

    Mucor racemosus and Rhizopus nigricans were used as sources of chitin deacetylases. Crude protein extracts were subjected to polyacrylamide gel electrophoresis at pH 8.9 (Davis system) or 4.3 (Reisfeld system) under native conditions. After electrophoresis, an overlay gel containing 0.1% (w/v) glycol chitin as substrate was incubated in contact with the separation gel. Chitin deacetylase activity was revealed by uv illumination with a transilluminator after staining for 5 min in 0.01% (w/v) Calcofluor white M2R. Chitosan (deacetylated chitin) generated by chitin deacetylases appeared more fluorescent than the intact chitin embedded in the overlay gel. Chitosan in a separate overlay gel was also subjected to a nitrous acid treatment which specifically depolymerizes chitosan while leaving chitin intact. Hydrolysis of chitosan by nitrous acid followed by Calcofluor staining yielded dark (nonfluorescent) bands (chitin deacetylase activities) in the fluorescent chitin-containing gel. Both assays revealed the presence of several chitin deacetylases from Zygomycetes. The same assays were performed after denaturing electrophoresis in 12% (w/v) polyacrylamide gels containing 0.1% (w/v) glycol chitin. Enzymes were renatured in buffered 1% (v/v) purified Triton X-100. Chitin deacetylases with estimated molecular weights between 26,000 and 64,000 were detected after Calcofluor staining. The assays were also performed in two-dimensional gel electrophoretic systems. Chitin deacetylases can be rapidly revealed by using the assay involving the nitrous acid treatment. However, both assays (with and without nitrous acid treatment) should be run to conclusively demonstrate chitin deacetylase activity after polyacrylamide gel electrophoresis. PMID:2281870

  18. Silver staining DNA in polyacrylamide gels.

    PubMed

    Bassam, Brant J; Gresshoff, Peter M

    2007-01-01

    This protocol describes a simple silver staining method used to visualize DNA fragments and other organic molecules with unsurpassed detail following traditional polyacrylamide gel electrophoresis (PAGE). Sensitivity rivals radioisotopic methods and DNA in the picogram range can be reliably detected. The described protocol is fast (approximately 1 h) and is implemented using readily available chemicals and materials. To achieve the sensitivity and visual clarity expected, quality reagents and clean handling are important. The updated protocol described here is based on the widely used method of Bassam et al. (1991), but provides improved image contrast and less risk of staining artefacts. PMID:18007600

  19. Silver staining of proteins in polyacrylamide gels.

    PubMed

    Chevallet, Mireille; Luche, Sylvie; Rabilloud, Thierry

    2006-01-01

    Silver staining is used to detect proteins after electrophoretic separation on polyacrylamide gels. It combines excellent sensitivity (in the low nanogram range) with the use of very simple and cheap equipment and chemicals. It is compatible with downstream processing, such as mass spectrometry analysis after protein digestion. The sequential phases of silver staining are protein fixation, then sensitization, then silver impregnation and finally image development. Several variants of silver staining are described here, which can be completed in a time range from 2 h to 1 d after the end of the electrophoretic separation. Once completed, the stain is stable for several weeks. PMID:17487168

  20. Micro-size polyacrylamide gel electrophoresis system

    NASA Astrophysics Data System (ADS)

    Hinson, W. G.; Pipkin, J. L.; Anson, J. F.; Casciano, D. A.; Burns, E. R.

    1987-09-01

    The development and characterization of a micro-size two-dimensional polyacrylamide gel electrophoresis system is described. Some of the techniques which have evolved with use of the system are also discussed. This apparatus has unique features which provide advantages over other small scale units. Up to ten first- and second-dimension gels can be processed simultaneously with excellent resolution of protein regions. Consistent reproducibility is possible from protein samples as small as 400 ng and individual protein regions as small as 1 pg can be visualized by silver staining of the two-dimensional gels. Similar sensitivities are achieved in autoradiographs of 3H-labeled proteins extracted from the nuclei of cultured cells. The application of this system in conjunction with flow cytometric examination of nuclear DNA and electrostatic cell sorting of specific cell nuclei to provide homogeneous sample populations, allows subtle variations in isotope incorporation in proteins to be detected; whereas many times in generalized tissue samples these changes are masked. Also, these techniques elucidate the effects of external stimuli (chemicals, drugs, or environment) on protein synthesis and phosphorylation for analyses and comparison. Fabrication drawings are available upon request.

  1. Polyacrylamide gel with switchable trypsin activity for analysis of proteins.

    PubMed

    Liu, Fangjie; Ye, Mingliang; Wang, Chunli; Hu, Zhengyan; Zhang, Yi; Qin, Hongqiang; Cheng, Kai; Zou, Hanfa

    2013-08-01

    Trypsin was immobilized on a variety of materials to improve digestion efficiency. However, because the immobilized trypsin will digest proteins during electrophoresis, direct immobilization of active trypsin in polyacrylamide gel will compromise the protein separation. To overcome this problem, here we report a novel polyacrylamide gel with switchable trypsin activity. It was prepared by copolymerization of the PEG-trypsin-aprotinin complex during the gel-casting step. Because the inhibitor aprotinin binds strongly with trypsin at alkaline pH, this novel gel does not display hydrolytic activity during electrophoresis. After electrophoresis, the activity of trypsin embedded in gel could be recovered by simply washing away the bound inhibitor at a low pH. It was demonstrated that this unique switchable activity design allowed high resolution of the complex protein mixture during electrophoresis and highly efficient digestion of the separated proteins in situ in the gel after electrophoresis. PMID:23855779

  2. 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-Gurin

    PubMed Central

    2011-01-01

    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-Gurin (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 helixes were identified well in our work. Conclusions In this study, we utilized LC-MS/MS in combination with blue native PAGE to characterize modular components of multiprotein complexes in BCG membrane fractions. The results demonstrated that the proteomic strategy was a reliable and reproducible tool for analysis of BCG multiprotein complexes. The identification in our study may provide some evidence for further study of BCG protein interaction. PMID:21241518

  3. Eosin Y staining of proteins in polyacrylamide gels.

    PubMed

    Lin, F; Fan, W; Wise, G E

    1991-08-01

    A staining method is described in which various proteins in polyacrylamide gels can be stained by using eosin Y. After a brief incubation of a polyacrylamide gel in an acidic solution of 1% eosin Y, various proteins, including human erythrocyte membrane sialoglycoproteins which are not detectable by Coomassie blue R-250 (CB), can be detected with a sensitivity of 10 ng protein. This is far more sensitive than CB staining and is comparable to the sensitivity of silver staining. In a Western blot, the antigenicity of an eosin Y stained protein is retained. In addition, proteins on an immunoblot sheet can be detected by eosin Y staining. The method described is rapid, sensitive, and reproducible with various proteins in polyacrylamide gels and has the added advantage of also staining sialoglycoproteins. PMID:1723249

  4. One-step casting of Laemmli discontinued sodium dodecyl sulfate-polyacrylamide gel electrophoresis gel.

    PubMed

    Wu, Xiaoqiang; Koiwa, Hisashi

    2012-02-01

    A modified Laemmli sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) protocol is described. The new method saves 30 min for gel casting without loss of the resolution power of Laemmli gel. In this method, both the upper and lower gels can be cast at the same time because the lower gel contains 10% glycerol, which generates higher density in the lower gel than in the upper gel. PMID:22037291

  5. Molecular-sieve chromatography and electrophoresis in polyacrylamide gels

    PubMed Central

    Morris, C. J. O. R.; Morris, Peggy

    1971-01-01

    1. The absolute electrophoretic mobilities of eight proteins have been measured at pH8.76, I 0.05, in polyacrylamide gels of 20 different compositions at 10C. 2. The partition coefficients of these proteins have been determined chromatographically under the same conditions by using columns of granulated polyacrylamide gel prepared simultaneously. 3. The electrophoretic mobilities are an exponential function of the gel concentrations when the latter are corrected for water uptake. The constants of this function have been determined by curvefitting methods. They have been shown to be related to the free solution mobility and to the mean molecular radius respectively. 4. The reduced mobilities have been shown to be a linear function of the partition coefficients by statistical analyses. 5. The physical significance of the relation between electrophoretic mobility and chromatographic phase distribution in gel media is discussed in the context of these results. PMID:5135238

  6. Flat Gel Polyacrylamide Electrophoresis of Porcine Mycoplasmas

    PubMed Central

    Wreghitt, T. G.; Windsor, G. D.; Butler, M.

    1974-01-01

    The flat gel acrylamide electrophoresis technique was standardized and applied to the comparison of four species of porcine mycoplasmas. Clear differences were observed between these species, and differences were seen among the strains of Mycoplasma hyosynoviae. The clarity of the patterns and the number of bands developed was influenced by the amount of protein in the extract and the age of the culture. The technique allows the comparison of several protein extracts in parallel without the problems associated with the rearrangement of separate gel columns. Images PMID:4472455

  7. Flat gel polyacrylamide electrophoresis of porcine mycoplasmas.

    PubMed

    Wreghitt, T G; Windsor, G D; Butler, M

    1974-10-01

    The flat gel acrylamide electrophoresis technique was standardized and applied to the comparison of four species of porcine mycoplasmas. Clear differences were observed between these species, and differences were seen among the strains of Mycoplasma hyosynoviae. The clarity of the patterns and the number of bands developed was influenced by the amount of protein in the extract and the age of the culture. The technique allows the comparison of several protein extracts in parallel without the problems associated with the rearrangement of separate gel columns. PMID:4472455

  8. The instantaneous monitoring of polyacrylamide gels during electrophoresis.

    PubMed Central

    Elliott, A

    1976-01-01

    The advantages of being able to see protein zones in a gel during electrophoresis (and hence before staining) are pointed out, and a method is described which depends on local increments of refractive index in these zones. The use of local increments of refractive index in polyacrylamide gels for measuring protein concentrations in zones during electrophoresis is briefly considered; it is found that such increments are greater than would be expected from the amount of protein when sodium dodecyl sulphate is present. The enhancement depends on conditions and time of running. This makes quantitative estimates difficult, but the sensitivity of detection of protein zones by observations based on refractive-index changes is greatly increased by this property of sodium dodecyl sulphate. Methods are described for making optically uniform gels (both with uniform and with graded concentrations of polyacrylamide), necessary for observation of small changes in refractive index. A simple dark-field system of observation is described. Examples are given showing protein samples observed with the system during electrophoresis and compared with the same gel stained with Coomassie Blue after completion of the run. Under optimal conditions the optical method is comparable in sensitivity with staining. With the proteins of lower mol.wt. (approx. 15000), the optical method is not so sensitive, becoming less sensitive with longer running time. This loss of sensitivity is greatly decreased by using more concentrated polyacrylamide gels, and graded gels are therefore more suitable for optical observation than are uniform gels. The observation of protein zones during electrophoresis adds nothing to the time needed for making a stained gel and gives much information long before it can be obtained from the stained gel. Images PLATE 1 PLATE 2 PMID:1008832

  9. Renaturation of enzymes after polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate

    SciTech Connect

    Lacks, S.A.; Springhorn, S.S.

    1980-08-10

    A number of enzymes, including amylases, dehydrogenases, and proteases, were shown to be renaturable after polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate. Enzyme activity was detected in situ by action on substrates introduced into the gel and subsequent staining of either the product or unreacted substrate. Enzymes appeared to recover activity as soon as the detergent diffused out of the gel. Renatured enzymes were retained in gels after electrophoresis longer than native enzymes which had been subjected to electrophoresis in the absence of detergent. Re-electrophoresis of the renatured enzymes showed that part of the retained activity was physically anchored to the gel, possibly by the folding of polypeptides around the gel matrix as the enzymes were renatured.

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

    PubMed Central

    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. Silver and Cyanine Staining of Oligonucleotides in Polyacrylamide Gel

    PubMed Central

    Li, Wei

    2015-01-01

    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. PMID:26650843

  12. Western Blot of Stained Proteins from Dried Polyacrylamide Gels

    NASA Technical Reports Server (NTRS)

    Gruber, Claudia; Stan-Lotter, Helga

    1996-01-01

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

  13. A method for the direct measurement of glycogen synthase activity on gels after polyacrylamide gel electrophoresis.

    PubMed

    Krisman, C R; Blumenfeld, M L

    1986-05-01

    A method for the detection of glycogen synthase activity after nondenaturing polyacrylamide gel electrophoresis is described. After the electrophoretical run, the gels were incubated in situ with UDP-glucose and glycogen. Labeled or unlabeled UDP-glucose could be used, since similar activity patterns were obtained by autoradiography or iodine staining of the gels. The method here described offers several advantages in terms of speed, sensitivity, and economy when compared with other procedures. PMID:2425655

  14. [Immobilization of Citrobacter L-asparaginase in polyacrylamide gel].

    PubMed

    Galaev, Iu V; Chuplygina, E G; Klement'eva, T A

    1981-01-01

    Bacterial L-asparaginase, immobilized on polyacrylamide gel, exhibited higher stability to denaturation and to the effect of a proteolytic enzyme. The immobilized enzyme exhibited the pH optimum of activity displaced by one pH unit to the acid side as compared with the free enzyme. The apparent Km value was approximately 200-fold higher as compared with the free L-asparaginase. The immobilized asparaginase hydrolyzed both L- and D-asparagine isomers but the free enzyme was highly stereospecific. PMID:7293086

  15. Glutamine Synthetase Regulation, Adenylylation State, and Strain Specificity Analyzed by Polyacrylamide Gel Electrophoresis

    PubMed Central

    Bender, Robert A.; Streicher, Stanley L.

    1979-01-01

    We used polyacrylamide gel electrophoresis to examine the regulation and adenylylation states of glutamine synthetases (GSs) from Escherichia coli (GSE) and Klebsiella aerogenes (GSK). In gels containing sodium dodecyl sulfate (SDS), we found that GSK had a mobility which differed significantly from that of GSE. In addition, for both GSK and GSE, adenylylated subunits (GSK-adenosine 5′-monophosphate [AMP] and GSE-AMP) had lesser mobilities in SDS gels than did the corresponding non-adenylylated subunits. The order of mobilities was GSK-AMP < GSK < GSE-AMP < GSE. We were able to detect these mobility differences with purified and partially purified preparations of GS, crude cell extracts, and whole cell lysates. SDS gel electrophoresis thus provided a means of estimating the adenylylation state and the quantity of GS present independent of enzymatic activity measurements and of determining the strain origin. Using SDS gels, we showed that: (i) the constitutively produced GS in strains carrying the glnA4 allele was mostly adenylylated, (ii) the GS-like polypeptide produced by strains carrying the glnA51 allele was indistinguishable from wild-type GSK, and (iii) strains carrying the glnA10 allele contained no polypeptide having the mobility of GSK or GSK-AMP. Using native polyacrylamide gels, we detected the increased amount of dodecameric GS present in cells grown under nitrogen limitation compared with cells grown under conditions of nitrogen excess. In native gels there was neither a significant difference in the mobilities of adenylylated and non-adenylylated GSs nor a GS-like protein in cells carrying the glnA10 allele. Images PMID:33958

  16. Investigations in x-ray computed tomography polyacrylamide gel dosimetry

    NASA Astrophysics Data System (ADS)

    Hilts, Michelle Louise

    Polyacrylamide gels (PAGs) are radiosensitive materials currently under development for use as three dimensional (3D) dosimeters in radiation therapy. Dose information is recorded in the gels and extracted through imaging. X-ray computed tomography (CT) has emerged as a promising gel imaging method due to a change in gel density that occurs upon irradiation. The accessibility of CT technology to cancer hospitals makes CT read-out clinically attractive, however the technique remains of limited clinical use due in part to poor dose resolution. This thesis investigates the use of CT for extracting dose information from PAG with an overall goal to improve achievable dose resolution. Thesis results are divided into three studies: a gel compositional study, a study of noise and dose resolution and a digital filtering study. The first study investigates the effects of gel composition on PAG CT dose response and the underlying density change. Systems for irradiating and imaging gels are designed and tested and dose response reproducibility is established. Results indicate dramatic variation in CT dose response sensitivity and range with gel composition. A model is developed to describe gel density change with dose, revealing two fundamental properties of the density to dose response: the density change that occurs per unit polymer yield is highest for gels with low and high concentrations of crosslinking molecule (%C) and the dose response sensitivity is linearly dependent on the total concentration of monomer in the gel. The second study investigates strategies for minimizing noise in x-ray CT polymer gel dosimetry and assesses system performance. Specifically, the effects of phantom design, scanning technique and image voxel size on image noise are investigated. This work leads to the establishment of a method of predicting image noise for any given CT imaging protocol. Image uniformity is also assessed, in the context of noise levels in gel dosimetry. The effect of scanning protocol on imaging time is established and the dose resolution achievable with an optimized system is calculated given voxel size and imaging time constraints. These results, when compared with published values for MRI and optical CT gel dosimetry indicate that CT dose resolution (e.g. 5%, 1 x 1 x 3 mm3 voxels), is still not at the level of the best MRI or optical CT techniques, however fast imaging times makes the rapid acquisition of volumetric data most feasible with x-ray CT. The third study investigates the potential of image filtering for improved dose resolution in CT gel dosimetry. CT image noise is characterized as Gaussian distributed and independent of signal strength and filters for reducing spatially invariant noise are investigated: mean, median, midpoint, adaptive mean, alpha-trimmed mean, sigma mean and a relatively new filter called SUSAN. The filters are tested on a CT image of a PAG irradiated with a clinically relevant dose distribution. Filter performance varies greatly in both achieved dose resolution and affects on the spatial distribution of dose. The ADAPTIVE and SUSAN filters provide the best overall performance, more than halving the dose resolution without significantly distorting the spatial distribution of dose. In summary, this thesis provides new insight into the fundamental nature of PAG density to dose response, develops strategies for minimizing image noise and quantifies system performance and demonstrates that digital image filtering is an effective tool to provide additional improvements to dose resolution.

  17. Scalable lithography from Natural DNA Patterns via polyacrylamide gel.

    PubMed

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

    2015-01-01

    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. PMID:26639572

  18. Scalable lithography from Natural DNA Patterns via polyacrylamide gel

    NASA Astrophysics Data System (ADS)

    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.

  19. Scalable lithography from Natural DNA Patterns via polyacrylamide gel

    PubMed Central

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

    2015-01-01

    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. PMID:26639572

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

    NASA Astrophysics Data System (ADS)

    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.

  1. Rheological properties of reactive extrusion modified waxy starch and waxy starch-polyacrylamide copolymer gels

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The rheological properties of modified waxy starch and waxy starch-polyacrylamide graft copolymers prepared by reactive extrusion were investigated. Both materials can absorb huge amount of water and form gels. The modified waxy starch and waxy starch-polyacrylamide graft copolymer gels all exhibite...

  2. A specific stain for the detection of nonheme iron proteins in polyacrylamide gels.

    PubMed

    Leong, L M; Tan, B H; Ho, K K

    1992-12-01

    Nonheme iron proteins can be visualized as blue bands in native polyacrylamide gels using a staining method that is both simple and rapid. The reaction of potassium ferricyanide with protein-bound iron atoms to form royal blue complexes occurs almost instantaneously and is sensitive enough to detect 1 microgram of analytical-grade ferritin and 2 micrograms of purified ferredoxin from cyanobacteria. No special treatment of reagents or apparatus was necessary. On comparison, this stain was found to be more specific than the Ferene S stain, not detecting bovine serum albumin even when present as a hundredfold excess over ferritin. The method was found to be effective for isoelectric focusing gels as well. PMID:1282787

  3. Destaining of Coomassie Brilliant Blue R-250-stained polyacrylamide gels with fungal laccase.

    PubMed

    Yang, Jie; Yang, Xiaodan; Ye, Xiuyun; Lin, Juan

    2016-01-15

    An enzyme-based method for destaining polyacrylamide gels stained with Coomassie Brilliant Blue R-250 is described. Distilled water supplemented with diluted fermentation broth of a laccase-producing white-rot fungus, Cerrena sp., was used for gel destaining, and a clear gel background was obtained in 2 h at 37 °C. Sensitivity of protein detection was 10 ng. The method did not require organic solvents or changing the destaining solution. Due to simultaneous gel destaining and dye decolorization, the colorless destaining solution can be disposed of directly. Laccase destaining of polyacrylamide gels was simple, efficient, and environmentally friendly. PMID:26475566

  4. 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.

    PubMed

    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. PMID:16916555

  5. Effects of degree of hydrolysis and shear on gelation reaction kinetics and gel strength. [Polyacrylamides

    SciTech Connect

    Gao, Hong W.

    1991-02-01

    Gelation tests were conducted to investigate the effect of the degree of hydrolysis on gelation reaction kinetics and gel strength using four low-molecular-weight polyacrylamides (MW = 400,000 daltons), which were 10% (HPAM1-10), 20% (HPAM1-20), 30% (HAPM1-30), and 40% (HPAM-40) hydrolyzed, and Cr-3 (pH = 4.8) and Al-3 (pH = 7.0) crosslinkers. Results showed that for polymer/Cr-3 gel systems, samples prepared with a low-molecular-weight polyacrylamide polymer, which was 20% hydrolyzed, gelled at a faster rate and retained higher gel strength than those prepared with a low-molecular-weight polyacrylamide polymer, which was 10% hydrolyzed. Under the screening condition, no viscosity enhancement was observed in samples prepared with polymers having a degree of hydrolysis equal to or greater than 30%. For polymer/Al-3 gel systems, samples prepared with a low-molecular-weight polyacrylamide polymer, which was 20% hydrolyzed, gelled at the fastest rate and retained the strongest gel strength among the polymer/Al-3 gel systems prepared with four low-molecular-weight polyacrylamide polymers, which were 10, 20, 30, and 40% hydrolyzed, respectively. Gelation tests of gel systems in glass bead packs showed that high shear favored the gelation of a gel system that had a fast rate of gelation, but had an adverse effect on the gelation of three gel systems that had a slow rate of gelation. Weak gels were found to be injectable through porous media. Weak gels were degradable under high shear condition and regained viscosity under low shear conditions. 17 refs., 8 figs., 1 tab.

  6. Optimal parameters for laccase-mediated destaining of Coomassie Brilliant Blue R-250-stained polyacrylamide gels.

    PubMed

    Yang, Jie; Yang, Xiaodan; Ye, Xiuyun; Lin, Juan

    2016-06-01

    The data presented in this article are related to the research article entitled "Destaining of Coomassie Brilliant Blue R-250-stained polyacrylamide gels with fungal laccase" [1]. Laccase is a class of multicopper oxidases that can catalyze oxidation of recalcitrant dyestuffs. This article describes optimal parameters for destaining of polyacrylamide gels, stained with Coomassie Brilliant Blue R-250, with laccase from basidiomycete Cerrena sp. strain HYB07. Effects of laccase activity, mediator type and concentration, temperature and time on destaining of polyacrylamide gels were evaluated with respect to gel background intensity and protein band signals, and the optimal destaining effects were obtained with 15 U mL(-1) laccase and 2 μM ABTS at 37 °C after 2 h. PMID:26955647

  7. Optimal parameters for laccase-mediated destaining of Coomassie Brilliant Blue R-250-stained polyacrylamide gels

    PubMed Central

    Yang, Jie; Yang, Xiaodan; Ye, Xiuyun; Lin, Juan

    2016-01-01

    The data presented in this article are related to the research article entitled “Destaining of Coomassie Brilliant Blue R-250-stained polyacrylamide gels with fungal laccase” [1]. Laccase is a class of multicopper oxidases that can catalyze oxidation of recalcitrant dyestuffs. This article describes optimal parameters for destaining of polyacrylamide gels, stained with Coomassie Brilliant Blue R-250, with laccase from basidiomycete Cerrena sp. strain HYB07. Effects of laccase activity, mediator type and concentration, temperature and time on destaining of polyacrylamide gels were evaluated with respect to gel background intensity and protein band signals, and the optimal destaining effects were obtained with 15 U mL−1 laccase and 2 μM ABTS at 37 °C after 2 h. PMID:26955647

  8. Copper staining: a five-minute protein stain for sodium dodecyl sulfate-polyacrylamide gels.

    PubMed

    Lee, C; Levin, A; Branton, D

    1987-11-01

    We present a new method for visualizing proteins electrophoresed in sodium dodecyl sulfate-polyacrylamide gels. After electrophoresis, gels are incubated in CuCl2 to produce a negative image of colorless protein bands against a semiopaque background. Gels are stained completely within 5 min, do not require destaining, and can be stored indefinitely without loss of the image. Because proteins are not permanently fixed within the gel, they can be quantitatively eluted after chelation of Cu with EDTA. The sensitivity of the CuCl2 stain falls between that of Coomassie blue and silver. We anticipate that CuCl2 will be useful in the rapid analysis of proteins by polyacrylamide gel electrophoresis and in the preparation of purified polypeptides by elution from gel slices. PMID:2449094

  9. A method for the preparation of low-pH dodecyl sulphate/polyacrylamide-gradient gels.

    PubMed Central

    Jones, G D; Wilson, M T; Darley-Usmar, V M

    1981-01-01

    1. A low-pH lithium dodecyl sulphate/polyacrylamide-gradient slab-gel system, suitable for electrophoresis, is described, and the migration properties of standard proteins are compared on this and conventional high-pH gels. 2. Cytochrome oxidase may be partially resolved into its component polypeptides. The order of migration of these is, however, dependent on the pH of the gel system. PMID:6272713

  10. Dose overshoot reduction by tetrakis hydroxymethyl phosphonium chloride in polyacrylamide gel dosimeter

    NASA Astrophysics Data System (ADS)

    Vvru, K.; emnick, J.; Sp?v?ek, V.

    2010-11-01

    This work deals with an influence of the antioxidant tetrakis (hydroxymethyl) phosphonium chloride (THPC) on the edge enhancing effect of a polyacrylamid gel and THPC (PAGAT) dosimeter. Four batches of PAGAT gel were produced with different THPC concentrations: 0, 1, 2 and 4 mM. It was proved that antioxidant THPC reduce the edge enhancing effects on the cost of reduced sensitivity of the gel dosimeter.

  11. Stabilization of thin-layer agarose gels after isoelectric focusing with polyacrylamide enables reverse imidazole-zinc staining and facilitates two-dimensional gel electrophoresis.

    PubMed

    Hellman, Jukka

    2008-09-01

    Large-pore-size agarose gels provide excellent resolving capacity for high molecular weight biomolecules. Thin-layer agarose isoelectric focusing (IEF) gels on polyester support films are especially useful for the separation of large proteins based on their pI in native conformation, but the method has suffered from the lack of detection methods compatible with agarose gels in hydrated form. Recently, an acrylamide copolymerization method was reported to enable mass-spectrometry-compatible silver staining and in-gel digestion of proteins. In this study, the method was further applied by demonstrating successful reverse imidazole-zinc staining of thin-layer agarose IEF gels copolymerized with acrylamide. The sensitivity of the reverse staining method on the composite gel at its best equaled the sensitivity of the traditional dried agarose silver staining method. Owing to the increased durability and reversible detection, the reverse-stained first-dimension gel could be conveniently prepared for the second-dimension sodium dodecyl sulfate polyacrylamide gel electrophoresis by reduction and alkylation. In addition, the micropreparative generation of tryptic peptides of Coomassie brilliant blue R-250 stained proteins in the composite gel is demonstrated. PMID:18607574

  12. Biochemical Identification of the Two Races of Radopholus similis by Polyacrylamide Gel Electrophoresis.

    PubMed

    Huettel, R N; Dickson, D W; Kaplan, D T

    1983-07-01

    Analysis of proteins of the banana and citrus race of Radopholus similis was carried out by several different types of polyacrylamide gel electrophoresis. These included standard slab gel, SDS slab gel, gradient slab gel, and two-ditnensional slab gel electrophoresis. A major band difference was detected between the two races by slab gel electrophoresis. However, several other poorly resolved but consistent hands of high molecular weight proteins near the gel origin also were considered as diagnostic. Resolution of protein bands was greatly improved by SDS and gradient slab gel electrophoresis, but no differences could be detected among the proteins resolved between the two rares with these techniques. Two-dimensional gels revealed a large number of proteins, but background staining obscured them hindering interpretation. When nematode races were reared on three different host plants, no differences in protein patterns were detected between them, indicating host preferences does not play a role in determining the types proteins occurring in these nematodes. PMID:19295815

  13. Fluorography of tritium-labeled proteins in silver-stained polyacrylamide gels

    SciTech Connect

    Kulcsar, P.; Prestwich, G.D.

    1988-05-01

    Silver-staining of polyacrylamide gel electrophoresis (PAGE)-separated proteins allows sensitive detection of proteins but severely reduces the ability to detect weak beta-emitters present in the protein band. A simple procedure is described in which silver can be removed from a silver-stained PAGE gel (deargentation) using photographic fixer, and the silver-free gel can be enhanced and used for fluorography. A quantitative study of sensitivity is reported for /sup 3/H-labeled bovine serum albumin with a one-dimensional sodium dodecyl sulfate-PAGE slab gel.

  14. Protein transfer from fixed, stained, and dried polyacrylamide gels and immunoblot with protein A-gold.

    PubMed

    Perides, G; Plagens, U; Traub, P

    1986-01-01

    The method of electrophoretically transferring proteins from fixed and stained polyacrylamide gels onto nitrocellulose paper has been reevaluated. It is shown that the tedious destaining of gels is not necessary because Coomassie brilliant blue, although it binds tenaciously to nitrocellulose paper, does not reduce the transfer efficiency of proteins. However, its presence impairs the visibility of proteins as detected, for instance, by the immunogold technique. Therefore, a rapid method for the complete removal of the stain from the nitrocellulose paper after completion of the immunogold procedure was developed. Furthermore, it is shown that proteins from dried polyacrylamide gels can still be transferred onto nitrocellulose sheets with an efficiency of approximately 50% compared to proteins transferred from fixed gels. PMID:2420231

  15. Specific antisera produced by direct immunization with slices of polyacrylamide gel containing small amounts of protein.

    PubMed

    Boulard, C; Lecroisey, A

    1982-01-01

    Rabbits were injected with slices of polyacrylamide gels containing entrapped insect proteins after separation by electrophoresis. Specific antibodies were produced independently of the nature of the gel (with or without sodium dodecyl sulphate) and of the staining technique (amido black or Coomassie Blue). The procedure appears to be a rapid and simple method for production of antibodies specific to proteins separated in minute quantities from a complex mixture. PMID:6282975

  16. A method for staining and stabilizing peroxidase activity in polyacrylamide gel electrophoresis.

    PubMed

    Shimoni, M; Reuveni, R

    1988-11-15

    A procedure was developed for a rapid double staining of peroxidase and other proteins in the same polyacrylamide gels using guaiacol and Coomassie blue. The distinguishable colored bands of peroxidase isozymes and proteins are stable for at least 8 months. PMID:2469356

  17. [Protein analysis of 6 crude drugs and their processed products by polyacrylamide gel electrophoresis technique].

    PubMed

    Shi, J; Sun, L; Jing, X

    1995-09-01

    In this paper, the proteins in 6 crude drugs (Prunus persica; P. armeniaca; Dolichos lablab; Strychnos nux-vomica; Mylabris phalerata; Whitmania pigra) and their processed products were analysed by polyacrylamide gel electrophoresis technique, and the effect of different processing methods on the quantity and kind of protein was explored. Protein electrophorograms of 20 samples are drawn. PMID:8679088

  18. Quantitation of proteins separated in N, N'-1,2-dihydroxyethylenebisacrylamide-crosslinked polyacrylamide gels.

    PubMed

    Neumann, U; Khalaf, H; Rimpler, M

    1992-10-01

    A simple and rapid method for the quantitation of proteins separated either by sodium dodecyl sulfate-electrophoresis or by isoelectric focusing in slab gels is presented. The method is based on the solubility of polyacrylamide gels crosslinked with N, N'-1, 2-dihydroxyethylenebisacrylamide (DHEBA) in periodic acid. After electrophoretic separation proteins are stained with Coomassie brilliant blue G-250. DHEBA gels show considerable swelling during the staining and destaining process but can be shrunk to their normal size in a 10% (w/v) solution of ammonium sulfate. Stained bands are cut from the gel and solubilized in periodic acid. During dissolution the dye decolorizes. Protein concentration in the solution is determined by a modified Coomassie dye-binding assay. Quantitation is linear in the range of 100 ng to 5 micrograms and not disturbed by dissolved gel. Separations in N, N'-1, 2-dihydroxyethylenebisacrylamide-crosslinked gels show qualities similar to those in normal crosslinked gels. PMID:1456419

  19. Rheology and morphology of pristine graphene/polyacrylamide gels.

    PubMed

    Das, Sriya; Irin, Fahmida; Ma, Lan; Bhattacharia, Sanjoy K; Hedden, Ronald C; Green, Micah J

    2013-09-11

    Enhancement of toughness in nanomaterial-based hydrogels is a critical metric for many of their engineering applications. Pristine graphene-polyacrylamide (PAM) hydrogels are synthesized via in situ polymerization of acrylamide monomer in PAM-stabilized graphene dispersion. In-situ polymerization leads to the uniform dispersion of the graphene sheets in the hydrogel. The graphene sheets interact with the elastic chains of the hydrogel through physisorption and permit gelation in the absence of any chemical cross-linker. This study represents the first report of pristine graphene as a physical cross-linker in a hydrogel. The properties of the graphene-polymer hydrogel are characterized by rheological measurements and compressive tests, revealing an increase in the storage modulus and toughness of the hydrogels compared to the chemically cross-linked PAM analogues. The physically cross-linked graphene hydrogels also exhibit self-healing properties. These hydrogels prove to be efficient precursors for graphene-PAM aerogels with enhanced electrical conductivity and thermal stability. PMID:23915342

  20. Polyacrylamide gel electrophoretic methods in the separation of structural muscle proteins.

    SciTech Connect

    Barany, K.; Barany, M.; Giometti, C. S.; Center for Mechanistic Biology and Biotechnology; Univ. of Illinois at Chicago

    1995-04-28

    Polyacrylamide gel electrophoresis plays a major role in analyzing the function of muscle structural proteins. This review describes one- and two-dimensional gel electrophoretic methods for qualitative and quantitative investigation of the muscle proteins, with special emphasis on determination of protein phosphorylation. The electrophoretic studies established the subunit structures of the muscle proteins, characterized their multiple forms, revealed changes in subunit composition or shifts in isoform distribution of specific proteins during development, upon stimulation or denervation of the muscle. Protein phosphorylation during muscle contraction is preferentially studied by two-dimensional gel electrophoresis. The same method demonstrated protein alterations in human neuromuscular diseases.

  1. Automatic analysis of 2D polyacrylamide gels in the diagnosis of DNA polymorphisms

    PubMed Central

    2013-01-01

    Introduction The analysis of polyacrylamide gels is currently carried out manually or automatically. In the automatic method, there are limitations related to the acceptable degree of distortion of lane and band continuity. The available software cannot deal satisfactorily with this type of situations. Therefore, the paper presents an original image analysis method devoid of the aforementioned drawbacks. Material This paper examines polyacrylamide gel images from Li-Cor DNA Sequencer 4300S resulting from the use of the electrophoretic separation of DNA fragments. The acquired images have a resolution dependent on the length of the analysed DNA fragments and typically it is MGNG=38061027 pixels. The images are saved in TIFF format with a grayscale resolution of 16 bits/pixel. The presented image analysis method was performed on gel images resulting from the analysis of DNA methylome profiling in plants exposed to drought stress, carried out with the MSAP (Methylation Sensitive Amplification Polymorphism) technique. Results The results of DNA polymorphism analysis were obtained in less than one second for the Intel Core 2 Quad CPU Q9300@2.5GHz, 8GB RAM. In comparison with other known methods, specificity was 0.95, sensitivity = 0.94 and AUC (Area Under Curve) = 0.98. Conclusions It is possible to carry out this method of DNA polymorphism analysis on distorted images of polyacrylamide gels. The method is fully automatic and does not require any operator intervention. Compared with other methods, it produces the best results and the resulting image is easy to interpret. The presented method of measurement is used in the practical analysis of polyacrylamide gels in the Department of Genetics at the University of Silesia in Katowice, Poland. PMID:23835039

  2. Polyacrylamide Gels for Invadopodia and Traction Force Assays on Cancer Cells

    PubMed Central

    Jerrell, Rachel J.; Parekh, Aron

    2015-01-01

    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 users specific biological and technical needs. PMID:25590238

  3. Pressure-induced volume phase transition of polyacrylamide gels in acetone-water mixtures

    NASA Astrophysics Data System (ADS)

    Kato, Eiji

    2000-07-01

    Equilibrium swelling curves of ionized polyacrylamide gels immersed in acetone-water mixtures were measured as a function of pressure up to pressures of 300 MPa. The gels, which shrank at atmospheric pressure, underwent an abrupt volume change (pressure-induced volume phase transition) from a shrunken state to a swollen state at the transition pressure. The transition pressure increased with an increase of acetone concentration. The pressure-induced volume phase transition can be interpreted by taking account of the free-energy change ?V?P between swollen (hydrated) and shrunken (dehydrated) states. The ?V represents the difference between the molar volume of water structured around hydrophilic groups of polyacrylamide chains and that of free water in the bulk mixtures. The estimated value of ?V is -3.3 mL/mol, which qualitatively agrees with that obtained from the experiments of denaturation of proteins. The pressure-induced volume phase transition is generally expected in many hydrogels.

  4. Sensitive silver-staining detection of bacterial lipopolysaccharides in polyacrylamide gels.

    PubMed

    Kittelberger, R; Hilbink, F

    1993-02-01

    For sensitive detection of bacterial lipopolysaccharides in the nanogram range, three almost identical silver-staining methods are often used, which are based on ammoniacal silver solutions and an acidic developer. We modified a method used for proteins, based on neutral silver nitrate solution and an alkaline developer, for the visualization of lipopolysaccharides in polyacrylamide gels, which yields better sensitivity and is much less prone to formation of non-specific background staining than the acidic developer-based silver stains. PMID:8387076

  5. Polyacrylamide gel breast augmentation: report of two cases and review of the literature.

    PubMed

    Margolis, Nathaniel E; Bassiri-Tehrani, Brian; Chhor, Chloe; Singer, Cory; Hernandez, Osvaldo; Moy, Linda

    2015-01-01

    Polyacrylamide gel (PAAG) injection remains an uncommon method of breast augmentation. Providers must recognize the clinical and radiological manifestations to optimize management. The clinical and radiological findings of PAAG injection may mimic malignancy and silicone breast augmentation. We described two patients with prior PAAG breast augmentation with physical exam and imaging findings concerning for malignancy. We reviewed the literature on PAAG breast augmentation and compare PAAG to silicone breast augmentation. The management of such patients is discussed. PMID:25670236

  6. Sensitive, quantitative, and fast modifications for Coomassie Blue staining of polyacrylamide gels.

    PubMed

    Westermeier, Reiner

    2006-09-01

    In spite of the high sensitivity of silver staining and the wide dynamic range of various fluorescent detection methods, Coomassie Brilliant Blue staining is still the most widely used protein detection technique for proteins separated by polyacrylamide gel electrophoresis. There are several reasons: Low price, Visible with the eye, Desk top scanners can be employed for image acquisition, Better for quantitative analysis than silver staining, Possible modifications for fast or highly sensitive staining, Mass spectrometry compatible. PMID:17031800

  7. A rapid and simplified method for protein silver staining in polyacrylamide gels.

    PubMed

    Zhao, Lijing; Liu, Cong; Sun, Yinpeng; Ban, Liping

    2012-07-01

    Silver staining is widely used to detect protein in polyacrylamide gels when high sensitivity is required. A simple and rapid protocol for silver staining of proteins following PAGE was developed in the present study. The number of steps was reduced compared to conventional protocol by combining fixing, rinsing, and soaking into a single impregnating step, thus achieving detection of proteins in 20 min. The present method is as sensitive as current protocols with the advantage of saving time and costs. PMID:22821490

  8. Preparation of nano-TiO2 powder by polyacrylamide gel method

    NASA Astrophysics Data System (ADS)

    Fan, Xiaowei; Liang, Xiaoping; Guo, Guanqun; Wang, Rongtao; Li, Jianxin

    2009-07-01

    Polyacrylamide gel method is a new successful technique for preparing nanometer metallic oxide materials, such as ?-Al2O3, ZnO and ZrO2. TiO2 made by this method, however, has not been reported. In this study, the nano-TiO2 powder was prepared using the novel method with TiCl3 as raw materials, and with acrylamide and N,N'-methylenediacrylamide as crosslinking agent. The TG showed that the optimization sintering process is that: the polyacrylamide gel was dried at 110C for 2 hour to obtain the xerogel, and then the grinded xerogel was sintered up to 450C, meanwhile, kept at 250C, 410C and 450C for 1 hour, respectively, to gain TiO2 particles. The grain size of TiO2 particles is smaller than 25 nm. And those particles are spherical approximately and have good distribution because of the hindering effect of space grid structure. Adding SO4 2- in the polyacrylamide gel process can maintain the content of anatase-TiO2 phase at higher sintering temperature. This may be explained that the SO4 2- can form coordination compound with Ti4+. And the coordination compound is prone to generate anatase-TiO2 with the burning out of SO4 2- , in which structure the adjacent two TiO6 hexahedrons share only one oxygen atom.

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

    PubMed

    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. PMID:25930092

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

    PubMed Central

    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

  11. Affinity gel electrophoresis of nucleic acids. Specific base- and shape-selective separation of DNA and RNA on polyacrylamide-nucleobase conjugated gel.

    PubMed

    Yashima, E; Suehiro, N; Miyauchi, N; Akashi, M

    1993-11-12

    Two types of affinity gels consisting of cross-linked polyacrylamide and affinity ligands possessing nucleic acid bases were prepared. One type of gel was polyacrylamide-poly(vinylnucleobase) conjugated gel, where the poly(vinylnucleobase) such as poly(9-vinyladenine) (PVAd) bearing a nucleobase in the side-chain was entrapped in the gel matrix. The other type of gel, in which a nucleobase such as adenine is chemically bonded to polyacrylamide gel, was prepared by copolymerization of acrylamide, cross-linker and 9-vinyladenine. These affinity gels, especially the former, demonstrated characteristic nucleobase- and shape-selective separation of nucleic acids. The gels showed high affinity for single-stranded DNA and both single- and double-stranded polynucleotides and could separate a double-stranded DNA in mixtures of double-stranded DNA and polynucleotides. The electrophoretic mobilities of poly(uridylic acid) and poly(inosinic acid) were selectively retarded even in the presence of 7 M urea. The electrophoretic behaviours of nucleic acids on the polyacrylamide-PVAd conjugated gels were compared with those on the agarose-PVAd conjugated gel. The effects of urea, temperature and concentration of PVAd were also examined. The polyacrylamide-PVAd conjugated gel served to elucidate interactions between PVAd and nucleic acids that could not be detected by usual spectroscopic methods. PMID:7506105

  12. Application of optical methods for dose evaluation in normoxic polyacrylamide gels irradiated at two different geometries

    NASA Astrophysics Data System (ADS)

    Adliene, D.; Jakstas, K.; Vaiciunaite, N.

    2014-03-01

    Normoxic gels are frequently used in clinical praxis for dose assessment or 3-D dose imaging in radiotherapy due to their relative simple manufacturing process under normal atmospheric conditions, spatial stability and well expressed modification feature of physical properties which is related to radiation induced polymerization of gels. In this work we have investigated radiation induced modification of the optical properties of home prepared normoxic polyacrylamide gels (nPAG) in relation to polymerization processes that occur in irradiated gels. Two irradiation geometries were used for irradiation of gel samples: broad beam irradiation geometry of teletherapy unit ROKUS-M with a 60Co source and point source irradiation geometry using 192Ir source of high dose rate afterloading brachytherapy unit MicroSelectron v2 which was inserted into gel via 6 Fr (2 mm thick) catheter. Verification of optical methods: UV-VIS spectrometry, spectrophotometry, Raman spectroscopy for dose assessment in irradiated gels has been performed. Aspects of their application for dose evaluation in gels irradiated using different geometries are discussed. Simple pixel-dose based photometry method also has been proposed and evaluated as a potential method for dose evaluation in catheter based interstitial high dose rate brachytherapy.

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

    PubMed

    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. PMID:22120911

  14. Characterization of the proteasome using native gel electrophoresis.

    PubMed

    Elsasser, Suzanne; Schmidt, Marion; Finley, Daniel

    2005-01-01

    Several features of the proteasome make it an excellent subject for analysis by native gel electrophoresis: its size, the multiplicity of variant complexes having proteasome activity, the ease of in-gel assays for proteasome activity, and even its relatively high cellular abundance. Accordingly, native gels have been used to analyze the composition, assembly, gating activity, and binding characteristics of the proteasome. This chapter describes methods for preparing, running, and developing native gels and the proteasome species that are routinely visualized. Additionally, the use of native gels to resolve proteasome complexes present in lysate and to characterize proteasome ligands are described. Following native gel electrophoresis, secondary analyses can be performed, such as activating the core particle, making specific activity assessments, Western blotting of the native gel, resolving native complexes with subsequent SDS-PAGE, and protein identification by mass spectrometry. PMID:16275342

  15. Preparation and Shear Modulus of Polyacrylamide Gels as Nerve Cell Culture

    NASA Astrophysics Data System (ADS)

    Perrault, Ccile M.; Juncker, David; Park, Hee Eon

    2008-07-01

    In the recent years, physical interactions between cells and their mechanical environment have been recognized for their influence on cellular functions, such as differentiation, motility, and growth. The importance of this phenomenon on neural cells is being investigated here in order to evaluate the optimal mechanical environment for their maximum growth. We prepared polyacrylamide gel as a culture medium for the nerve cell growth. Since we hypothesize that the shear modulus of the medium, which is fully saturated with water, plays an important role in the cell growth, we prepared gels having different shear modulus by varying the ratio of the polymerizing agents and the thickness of the medium was controlled by molding. It is the key issue to determine the shear modulus of the gels to study the effect of the modulus on the nerve cell growth. However, because the physical properties of the gels should be measured when those are saturated with water, but there should be no water layer on the surface to prevent slip in rheometers, new techniques were developed to dissolve these issues. This poster will present those rheological techniques, preparation of the gels, and the effect of shear modulus of the gel medium on the growth of nerve cells.

  16. Differentiation of rennet from other milk-clotting enzymes by polyacrylamide gel electrophoresis.

    PubMed

    Prager, M J

    1977-11-01

    Polyacrylamide gel electrophoresis was used to differentiate animal rennet and other milk-clotting enzymes. After electrophoresis, the separated components were visualized by staining with aniline blue-black. Two prominent proteins were found in calf and bovine rennet, while only 1 major protein was observed in pepsin and enzymes of microbial origin. These patterns provided a basis for distinguishing animal rennet and the other enzymes as well as a means of identifying each type of enzyme by the characteristic pattern shown. PMID:336596

  17. Rapid detection of proteins in polyacrylamide electrophoresis gels with Direct Red 81 and Amido Black.

    PubMed

    Choveaux, David; Krause, Robert G E; Goldring, J P Dean

    2012-01-01

    Proteins separated by SDS-polyacrylamide gel electrophoresis need to be stained with organic dyes to be visualized and to enable comparisons to be made between the intensity of protein bands to observe and determine differences in protein concentration. The standard protein staining is with Coomassie Blue R-250. Coomassie staining takes 1 h to complete. Direct Red 81 and Amido Black stain proteins within 10 min. This chapter describes Direct Red 81 and Amido Black staining in comparison to staining with Coomassie Blue R-250. PMID:22585524

  18. Accommodating brightness and exposure levels in densitometry of stained polyacrylamide electrophoresis gels

    SciTech Connect

    Tan, Han Yen; Ng, Tuck Wah; Liew, Oi Wah

    2010-03-20

    Flatbed scanner densitometers can be operated under various illumination and recording exposure levels. In this work, we show that optical density measurement accuracy, sensitivity, and stability of stained polyacrylamide electrophoresis gel densitometry are crucially dependent on these two factors (brightness and exposure level), notwithstanding that the source is monochromatic, spatially uniform, and the measurements are made using an accurately calibrated step wedge in tandem. We further outline a method to accommodate the intensity deviations over a range of illumination and exposure levels in order to maintain sensitivity and repeatability in the computed optical densities. Comparisons were also made with results from a commercial densitometer.

  19. A Novel Technique for Micro-patterning Proteins and Cells on Polyacrylamide Gels

    PubMed Central

    Tang, Xin; Ali, M. Yakut; Saif, M. Taher A.

    2012-01-01

    Spatial patterning of proteins (extracellular matrix, ECM) for living cells on polyacrylamide (PA) hydrogels has been technically challenging due to the compliant nature of the hydrogels and their aqueous environment. Traditional micro-fabrication process is not applicable. Here we report a simple, novel and general method to pattern a variety of commonly used cell adhesion molecules, i.e. Fibronectin (FN), Laminin (LN) and Collagen I (CN), etc. on PA gels. The pattern is first printed on a hydrophilic glass using polydimethylsiloxane (PDMS) stamp and micro-contact printing (μCP). Pre-polymerization solution is applied on the patterned glass and is then sandwiched by a functionalized glass slide, which covalently binds to the gel. The hydrophilic glass slide is then peeled off from the gel when the protein patterns detach from the glass, but remain intact with the gel. The pattern is thus transferred to the gel. The mechanism of pattern transfer is studied in light of interfacial mechanics. It is found that hydrophilic glass offers strong enough adhesion with ECM proteins such that a pattern can be printed, but weak enough adhesion such that they can be completely peeled off by the polymerized gel. This balance is essential for successful pattern transfer. As a demonstration, lines of FN, LN and CN with widths varying from 5–400 μm are patterned on PA gels. Normal fibroblasts (MKF) are cultured on the gel surfaces. The cell attachment and proliferation are confined within these patterns. The method avoids the use of any toxic chemistry often used to pattern different proteins on gel surfaces. PMID:23002394

  20. Identification of different quaternary structures of beef heart cytochrome-c oxidase by two-dimensional polyacrylamide gel electrophoresis.

    PubMed

    Heinrichs, M; Schnert, H

    1987-11-01

    A two-dimensional gel electrophoresis is described to identify different quaternary structures of the heart cytochrome-c oxidase. Bovine enzyme was purified and separated by discontinuous gradient polyacrylamide gel electrophoresis under nondenaturing conditions in the 1st dimension into several discrete complexes and thereupon shown to be heterodisperse in Triton X-100 and dodecyl maltoside. A discontinuous SDS-polyacrylamide gel electrophoresis in the 2nd dimension was used to determine the subunit composition of the isolated complexes. One of these represents the intact enzyme with 12 different polypeptides while the others have an incomplete subunit composition. PMID:2822485

  1. Tris-acetate polyacrylamide gradient gels for the simultaneous electrophoretic analysis of proteins of very high and low molecular mass.

    PubMed

    Cubillos-Rojas, Monica; Amair-Pinedo, Fabiola; Tato, Irantzu; Bartrons, Ramon; Ventura, Francesc; Rosa, Jose Luis

    2012-01-01

    Polyacrylamide gel electrophoresis (PAGE) is one of the most powerful tools used for protein analysis. We describe the use of Tris-acetate buffer and 3-15% polyacrylamide gradient gels to simultaneously separate proteins in the mass range of 10-500 kDa. We show that this system is highly sensitive, it has good resolution and high reproducibility, and that it can be used for general applications of PAGE such as Coomassie Brilliant Blue staining and immunoblotting. Moreover, we describe how to generate mini Tris-acetate polyacrylamide gels to use them in miniprotein electrophoresis systems. These economical gels are easy to generate and to manipulate and allow a rapid analysis of proteins. All these features make the Tris-acetate-PAGE system a very helpful tool for protein analysis. PMID:22585488

  2. Entamoeba histolytica: analysis of the trophozoite proteome by two-dimensional polyacrylamide gel electrophoresis.

    PubMed

    Leitsch, David; Radauer, Christian; Paschinger, Katharina; Wilson, Iain B H; Breiteneder, Heimo; Scheiner, Otto; Duchêne, Michael

    2005-07-01

    The Entamoeba histolytica genome project carried out at TIGR and the Sanger Institute has produced a near-complete set of deduced open reading frame sequences. These data provide strong support for the identification of signals from proteomic analyses such as two-dimensional electrophoresis by protein sequencing and/or mass spectrometric methods. To carry out an initial investigation of the E. histolytica proteome, appropriate sample preparation and silver staining protocols were adapted. After preparation of protein extracts from E. histolytica HM-1:IMSS trophozoites, solubilized proteins were separated in the first dimension in IPG (immobilized pH gradient) strips depending on their pI and subsequently in the second dimension according to their molecular weight by SDS-polyacrylamide gel electrophoresis. Of the more than 1500 protein spots visualized, several landmark spots were isolated from the gels and identified by either tryptic cleavage and subsequent MALDI-TOF mass spectrometry or by protein sequencing. PMID:15955311

  3. Identification of trichloroethanol visualized proteins from two-dimensional polyacrylamide gels by mass spectrometry.

    PubMed

    Ladner, Carol L; Edwards, Robert A; Schriemer, David C; Turner, Raymond J

    2006-04-01

    Proteins visualized by 2,2,2-trichloroethanol (TCE) on two-dimensional electrophoresis gels are efficiently identified by matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry (MS) and MS/MS. In a previous study, a method was developed that placed TCE in the polyacrylamide gel so that protein bands can be visualized without staining in less than 5 min. A visible fluorophore is generated by reaction of TCE with tryptophan that allows for protein visualization. In this study, MALDI-TOF MS and LC-MS/MS are used to identify randomly selected Escherichia coli proteins. The identification of TCE visualized proteins is compared to the identification of Coomassie brilliant blue (CBB) stained proteins from two-dimensional gel electrophoresis of E. coli proteins. This study demonstrated that TCE visualized proteins are compatible with protein identification by MALDI-TOF peptide mass fingerprinting. For 10 randomly selected spots, TCE visualization lead to statistically significant identification of 5 proteins and CBB visualization lead to identification of 6 proteins. TCE visualized proteins are also shown to be well suited for protein identification using LC-MS/MS. In 16 spots selected for MS/MS analysis, TCE samples lead to the identification of 79 peptides; while CBB samples lead to the identification of 65 peptides. TCE samples also supported the identification of more proteins. The low stoichiometry of labeling of tryptophan residues does not require inclusion of this modification for database searches. In addition to being a rapid visualization technique compatible with MS, TCE visualization utilizes rapid washing conditions for sample preparation of proteins spots excised from polyacrylamide gels. PMID:16579625

  4. A rapid and effective method for silver staining of PCR products separated in polyacrylamide gels.

    PubMed

    Liang, Qingzhi; Wen, Dingqing; Xie, Jianghui; Liu, Liqin; Wei, Yongzan; Wang, Yicheng; Shi, Shengyou

    2014-09-01

    With the development of molecular quantitative genetics, particularly, genetic linkage map construction, quantitative trait loci mapping or genes fine mapping and association analysis etc., more and more PCR products separated in polyacrylamide gels need to be silver-stained. However, conventional silver-staining procedures are complicated and time-consuming as they require a lot of preparation and handling of several solutions prior to use. In this study, a simple and rapid protocol for silver staining of PCR products was developed. The number of steps was reduced compared to conventional protocols, thus achieving detection of PCR products in 7 min, saving time and resources. Fixation and staining solution and developing solution in present staining procedure allowed a reutilization for 12 and 8 times, respectively, reducing the cost greatly. Meanwhile, the sensitivity was significantly improved with the improved method and the minimum of 0.097 ng/μL of DNA amount can be detected in denaturing polyacrylamide gel. The protocol developed in this study will facilitate the development of molecular quantitative genetics. PMID:24789566

  5. Evaluation of SDS-polyacrylamide gel systems for the study of outer membrane protein profiles of clinical strains of Acinetobacter baumannii.

    PubMed

    Cuenca, Felipe Fernndez; Pascual, Alvaro; Martnez Marnez, Luis; Conejo, Ma Carmen; Perea, Evelio J

    2003-01-01

    The outer membrane protein (OMP) profiles of 23 blood isolates of Acinetobacter baumannii representing all the different antimicrobial susceptibility patterns observed during a 3-year period in a Spanish hospital were studied. OMPs extracted from envelopes of sonicated cells after solubilisation with 2% of N-lauryl-sarcosinate were analysed by sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) using the Laemmli's buffers. Eight running gel systems differing in the concentration of polyacrylamide (8%, 10% and 12%) and in the absence or presence of urea (4 M and 6 M) were used in a preliminary study analysing the OMP profiles of four clonally unrelated strains of A. baumannii. When this study was completed, the OMPs of the 23 A. baumannii were analysed in 10% SDS-polyacrylamide gels with 6 M urea and in 12% SDS-polyacrylamide gels. Ten OMP profiles were observed in 10% SDS-polyacrylamide gels with 6 M urea, whereas only 5 OMP profiles were visualised using 12% SDS-polyacrylamide gels. The OMP profiles obtained in 10% SDS-polyacrylamide gels with 6 M urea only partially correlated with those observed in 12% SDS-polyacrylamide gels. In conclusion, the use of 10% SDS-polyacrylamide gels with 6 M urea is recommended for the study of OMP profiles of A. baumannii. PMID:12761770

  6. Resolving acetylated and phosphorylated proteins by neutral urea Triton-polyacrylamide gel electrophoresis: NUT-PAGE.

    PubMed

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

    2014-08-01

    Protein acetylation and phosphorylation are key modifications that regulate both normal and pathological protein functions. The gel systems currently used for analyzing modified proteins require either expensive reagents or time-consuming second dimension electrophoresis. Here we present a neutral pH gel system that allows the analysis of acetylated and phosphorylated proteins. The neutral pH urea Triton-polyacrylamide gel electrophoresis (NUT-PAGE) system separates proteins based on their charge at pH 7.0 and generates discrete bands from each acetylated and/or phosphorylated species. In addition, the gel is composed of common and inexpensive laboratory reagents and requires only a single dimension of electrophoresis. We demonstrate the effectiveness of this system by analyzing the phosphorylated species of an acidic protein, ?-synuclein, and both acetylated and phosphorylated species of a basic protein, histone H3. NUT-PAGE thus provides a cost-effective alternative for resolving acetylated and phosphorylated proteins, and potentially proteins with other post-translational modifications that alter net charge. PMID:25109292

  7. Rapid and simple single nanogram detection of glycoproteins in polyacrylamide gels and on electroblots.

    PubMed

    Steinberg, T H; Pretty On Top, K; Berggren, K N; Kemper, C; Jones, L; Diwu, Z; Haugland, R P; Patton, W F

    2001-07-01

    The fluorescent hydrazide, Pro-Q Emerald 300 dye, may be conjugated to glycoproteins by a periodic acid Schiff's (PAS) mechanism. The glycols present in glycoproteins are initially oxidized to aldehydes using periodic acid. The dye then reacts with the aldehydes to generate a highly fluorescent conjugate. Reduction with sodium metabisulfite or sodium borohydride is not required to stabilize the conjugate. Though glycoprotein detection may be performed on transfer membranes, direct detection in gels avoids electroblotting and glycoproteins may be visualized within 2-4 h of electrophoresis. This is substantially more rapid than PAS labeling with digoxigenin hydrazide followed by detection with an antidigoxigenin antibody conjugate of alkaline phosphatase, or PAS labeling with biotin hydrazide followed by detection with horseradish peroxidase or alkaline phosphatase conjugates of streptavidin, which require more than eight hours to complete. Pro-Q Emerald 300 dye-labeled gels and blots may be poststained with SYPRO Ruby dye, allowing sequential two-color detection of glycosylated and nonglycosylated proteins. Both fluorophores are excited with mid-range UV illumination. Pro-Q Emerald 300 dye maximally emits at 530 nm (green) while SYPRO Ruby dye maximally emits at 610 nm (red). As little as 300 pg of alpha 1-acid glycoprotein (40% carbohydrate) and 1 ng of glucose oxidase (12% carbohydrate) or avidin (7% carbohydrate) are detectable in gels after staining with Pro-Q Emerald 300 dye. Besides glycoproteins, as little as 2-4 ng of lipopolysaccharide is detectable in gels using Pro-Q Emerald 300 dye while 250-1000 ng is required for detection with conventional silver staining. Detection of glycoproteins may be achieved in sodium dodecyl sulfate-polyacrylamide gels, two-dimensional gels and on polyvinylidene difluoride membranes. PMID:11503209

  8. Apolipoprotein distribution in human lipoproteins separated by polyacrylamide gradient gel electrophoresis.

    PubMed

    Vzina, C A; Milne, R W; Weech, P K; Marcel, Y L

    1988-05-01

    The heterogeneity of serum lipoproteins (excluding very low density (VLDL) and intermediate density (IDL) lipoproteins) and that of lipoproteins secreted by HepG2 cells has been studied by immunoblot analysis of the apolipoprotein composition of the particles separated by polyacrylamide gradient gel electrophoresis (GGE) under nondenaturing conditions. The reactions of antibodies to apoA-I, apoA-II, apoE, apoB, apoD, and apoA-IV have revealed discrete bands of particles which differ widely in size and apolipoprotein composition. GGE of native serum lipoproteins demonstrated that apoA-II is present in lipoproteins of limited size heterogeneity (apparent molecular mass 345,000 to 305,000) and that apoB is present in low density lipoproteins (LDL) and absent from all smaller or denser lipoproteins. In contrast, serum apoA-I, E, D, and A-IV are present in very heterogeneous particles. Serum apoA-I is present mainly in particles of 305 to 130 kDa where it is associated with apoA-II, and in decreasing order of immunoreactivity in particles of 130-90 kDa, 56 kDa, 815-345 kDa, and finally within the size range of LDL, all regions where there is little detectable apoA-II. Serum apoE is present in three defined fractions, one within the size range of LDL, one containing heterogeneous particles between 640 and 345 kDa, and one defined fraction at 96 kDa. Serum apoD is also present in three defined fractions, one comigrating with LDL, one containing heterogeneous particles between 390 and 150 kDa, and one band on the migration front. Most of serum apoA-IV is contained in a band comigrating with albumin. GGE of centrifugally prepared LDL shows the presence of apoB, apoE, and apoD, but not that of apoA-I. However, the particles containing apoA-I, which, in serum, migrated within the LDL size range and as bands of 815 to 345 kDa, were recovered upon centrifugation in the d greater than 1.21 g/ml fraction. GGE of high density lipoproteins (HDL) indicated that most of apoA-I, A-II, and A-IV were present in lipoproteins of the same apparent molecular mass (390-152 kDa). ApoD tended to be associated with large HDL, and this was also significant for HDL apoE, which is present in lipoproteins ranging from 640 to 275 kDa. GGE of very high density lipoproteins (VHDL) presented some striking features, one of which was the occurrence of apolipoproteins in very discrete bands of different molecular mass. ApoA-II was bimodally distributed at 250-175 kDa and 175-136 kDa, the latter fraction also containing apoA-I.(ABSTRACT TRUNCATED AT 400 WORDS) PMID:3411236

  9. Centrifuge-blotting of proteins after separation by sodium dodecyl sulfate-polyacrylamide gel electrophoresis.

    PubMed

    Hermansen, L F; Pedersen, O; Sletten, K

    1993-12-01

    A protein transfer method which allows elution and immobilization of polypeptides onto a polyvinylidene difluoride (PVDF) membrane has been developed. The protein band in a gel is eluted by centrifugation. The centrifuge-blotting procedure involves the following steps: (i) visualization of the protein in a sodium dodecyl sulfate (SDS)-polyacrylamide gel with 1 M KCl, (ii) excision of the protein band and equilibration for 15 min in a solution of 0.05% SDS/5% methanol/0.02% dithiothreitol in distilled water, (iii) placing the gel piece in direct contact with the PVDF membrane in the receptacle, (iv) centrifugation at 3000 g for 1 h. A 10 kDa cut-off dialysis membrane is placed beneath the PVDF membrane to retain nonimmobilized protein. The N-terminal sequence of the immobilized protein on the PVDF membrane was determined. For proteins with a molecular mass less than 30 kDa, an overall yield between 10%-30% has been obtained. PMID:8137793

  10. Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications.

    PubMed Central

    Towbin, H; Staehelin, T; Gordon, J

    1979-01-01

    A method has been devised for the electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets. The method results in quantitative transfer of ribosomal proteins from gels containing urea. For sodium dodecyl sulfate gels, the original band pattern was obtained with no loss of resolution, but the transfer was not quantitative. The method allows detection of proteins by autoradiography and is simpler than conventional procedures. The immobilized proteins were detectable by immunological procedures. All additional binding capacity on the nitrocellulose was blocked with excess protein; then a specific antibody was bound and, finally, a second antibody directed against the first antibody. The second antibody was either radioactively labeled or conjugated to fluorescein or to peroxidase. The specific protein was then detected by either autoradiography, under UV light, or by the peroxidase reaction product, respectively. In the latter case, as little as 100 pg of protein was clearly detectable. It is anticipated that the procedure will be applicable to analysis of a wide variety of proteins with specific reactions or ligands. Images PMID:388439

  11. Delayed Gel Indurations as an Adverse Effect of Polyacrylamide Filler and Its Easy Treatment

    PubMed Central

    Kavoussi, Hossein; Ebrahimi, Ali

    2012-01-01

    Background. The more increasing use of permanent soft tissue fillers such as polyacrylamide hydrogel (PAAG) for aesthetic purposes, the more adverse events resulting from them are reported. Occasionally, nonserious complications and misdiagnosis result in unnecessary surgeries and sequels. Objective. To introduce delayed gel indurations (DGIs) as a late onset complication of PAAG and its easy treatment. Patient and Methods. Twenty patients (17 females and 3 males) referred to us with subcutaneous mass at injected site of PAAG. We diagnosed DGI based on clinical and sonography findings and treatment was performed with a hole by 16-gauge needle and squeezing. Results. From 20 patients with 21 cases of DGI, 5 (23.8%), 5 (23.8%), and 5 (23.8%) cases in cheeks, glabella, and lips were seen, respectively. The time range between PAAG injection and presentation of patients was 1028 months (mean = 17.5%). All of the patients responded very well to treatment without recurrence and any complications. Conclusion. DGI is a nonserious, late onset, and easily treated complication of PAAG that is probably induced due to water exchange between gel and surrounding tissue and modest host immune reaction to gel. PMID:23093956

  12. Phosphoprotein staining for sodium dodecyl sulfate-polyacrylamide gel electrophoresis using fluorescent reagent morin hydrate.

    PubMed

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

    2013-04-01

    A fluorescence-based stain with 3,5,7,2',4'-pentahydroxyflavone (morin hydrate, MH) was designed to stain phosphoproteins in one-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Al(3+) was applied as a "fixed bridge," providing an efficient energy transfer channel between phosphoprotein and MH, to produce a strong fluorescent complex for the determination of phosphoprotein. As little as 62.5ng of ?-casein (7 or 8 phosphates) and ?-casein (5 phosphates), 125ng of ovalbumin (2 phosphates), and ?-casein (1 phosphate) could be visualized with a wide linear dynamic range. In comparison with conventional methods, MH stain is a time-saving method that takes just 90min. It also has good compatibility with routine protein stainings such as Coomassie Brilliant Blue R (CBBR) and SYPRO Ruby for total protein analysis. PMID:23274386

  13. Fluorescent staining of protein in sodium dodecyl sulfate polyacrylamide gels by salicylaldehyde azine.

    PubMed

    Ni, Mao-Wei; Ye, Wei-Jian; Cong, Wei-Tao; Hong, Guo-Ying; Zhu, Zhong-Xin; Duan, Yuan-Meng; Zhou, Xuan; Jin, Li-Tai

    2013-12-01

    As a non-covalent fluorescence probe, in this study, salicylaldehyde azine (SA) was introduced as a sensitive fluorescence-based dye for detecting proteins both in 1-D and 2-D polyacrylamide electrophoresis gels. Down to 0.2 ng of single protein band could be detected within 1 h, which similars to that of glutaraldehyde (GA)-silver stain, but approximately four times higher than that of SYPRO Ruby fluorescent stain. Furthermore, comparative analysis of the MS compatibility of SA stain with SYPRO Ruby stain indicated that SA stain is compatible with the downstream of protein identification by liquid chromatography-tandem mass spectrometry (LC-MS/MS). Additionally, the probable mechanism of the SA stain was investigated by molecular docking. The results demonstrated that the interaction between SA and protein was mainly contributed by hydrogen bonding and hydrophobic forces. PMID:24105885

  14. Optimizing soluble protein extraction and two-dimensional polyacrylamide gel electrophoresis quality for extremophile ciliates.

    PubMed

    Fulgentini, Lorenzo; Marangoni, Roberto; Colombetti, Giuliano

    2008-06-01

    An efficient protein extraction methodology is quite important for sample preparation and subsequent 2-D PAGE and MS analysis. Cell lysis is the first step in protein extraction and purification. Many techniques are available for cell disruption, including physical and detergent-based methods. Here, we report on a very fast and efficient detergent-free Tris-based method to extract the soluble fraction proteins of extremophile ciliates, comparing it with a detergent-based protocol. This comparison has been carried out by means of 2-D PAGE and subsequent MALDI-compatible silver staining of protein samples obtained from the intensely pigmented hypersaline ciliate Fabrea salina and the Antarctic hypotrich ciliate Euplotes focardii. Our results indicate that this fast and easy extraction method allows to obtain more clear crude extracts and more spot-abundant polyacrylamide gels. PMID:18548458

  15. Improved staining of proteins in polyacrylamide gels including isoelectric focusing gels with clear background at nanogram sensitivity using Coomassie Brilliant Blue G-250 and R-250.

    PubMed

    Neuhoff, V; Arold, N; Taube, D; Ehrhardt, W

    1988-06-01

    An improved procedure for staining of proteins following separation in polyacrylamide gels is described which utilizes the colloidal properties of Coomassie Brilliant Blue G-250 and R-250. The new method is based on addition of 20% v/v methanol and higher concentrations of ammonium sulfate to the staining solution previously described. The method combines the advantage of much shorter staining time with high sensitivity, a clear background not requiring destaining, stepwise staining, and stable fixation after staining. The method has been applied to staining of polyacrylamide gels after sodium dodecyl sulfate-electrophoresis and isoelectric focusing in carrier ampholyte-generated pH gradients. PMID:2466658

  16. Studies on the bioactivity of radioiodinated highly purified bovine thyrotropin: analytical polyacrylamide gel electrophoresis

    SciTech Connect

    Takai, N.A.; Filetti, S.; Rapoport, B.

    1981-01-01

    Highly purified bovine TSH (stored in solution at -70 C) was radioiodinated by the stoichiometric chloroamine-T method. The iodinated material ws subjected to analytical polyacrylamide disc gel electrophoresis. TSH was eluted from gel slices (1 mm width) and was analyzed for radioactivity and bioactivity. The latter was determined using the cultured thyroid cell cAMP response assay. Radioactivity in the TSH preparation migrated separately from bioactivity, but concordant with the protein bands observed in gels run in parallel. Further studies performed on bovine TSH purified in our laboratory, as well as on a different TSH preparation of exceptionally high potency (both stored as lyophilized powder) revealed a different pattern, with TSH bioactivity and radioactivity eluting concurrently. Iodination of TSH did not alter its electrophoretic migration on disc gel electrophoresis. In all preparations polymorphism of TSH bioactivity was observed, with at least four separate protein bands containing TSH bioactivity being present in our preparation. The relationship between the degree of iodination and retention of TSH bioactivity was examined. Incorporation of /sup 125/I into TSH was greatly different at two different concentrations of chloramine-T. Despite this, however, the progressive loss of TSH bioactivity was similar at both concentrations, indicating that incorporation of iodine into the TSH molecule is not itself responsible for the decrease in bioactivity. These studies indicate variability among different TSH preparations in terms of their retention of bioactivity. Significant loss of TSH bioactivity appears to occur during storage in solution. The damage to the biological activity of TSH during the iodination procedure is more likely related to the oxidation process than to the incorporation of iodine.

  17. Two dimensional polyacrylamide gel electrophoresis analysis of Tetrahymena mitochondrial tRNA.

    PubMed

    Suyama, Y

    1986-01-01

    Two dimensional (2D) urea-polyacrylamide gel electrophoresis of tRNA isolated from Tetrahymena mitochondria separated at least 36 spots, while more than 45 major and minor spots were resolved with cytosolic tRNA. Co-electrophoresis of mitochondrial and cytosolic tRNAs revealed that many spots co-migrate. When radioactive mitochondrial tRNA was hybridized to mtDNA under various conditions and tRNA melted from the hybrid was analyzed by 2D gel electrophoresis, only 10 tRNA spots were found. Identified as mtDNA-encoded were 2 spots for tRNA(leu), 2 for tRNA(met), and 1 each for tRNA(phe), tRNA(trp) and tRNA(tyr). The remaining three were unidentified. Mitochondrial tRNA spots that correspond to the tRNAs for arg, gly, ile, lys, ser, and val do not hybridize with mtDNA, and in gel positions they correspond to the cytoplasmic tRNA spots for the same respective amino acids. These mitochondrial tRNAs isolated from the gel can be acylated either by the mitochondrial or cytosolic enzymes. Mitochondrial tRNA isolated from a Tetrahymena cell homogenate which was pretreated with RNase A and Micrococcus nuclease exhibited the same 2D gel pattern as a non-treated control. Mitochondrial tRNAs from old and young cells showed generally similar tRNA spots in 2D gels, though more variable spots were seen with old cells. 3H-labeled whole-cell tRNA added to the cell homogenate prior to the mitochondrial isolation procedure did not remain associated with the final mitochondrial tRNA preparation. The present studies also showed mitochondrial tRNAs bound to the mitochondrial 80S monosome and polysome fractions. Radioactive tRNA added to the mitochondrial lysate does not adhere to the ribosomes, suggesting that the ribosome-bound tRNAs are not contaminating cytoplasmic tRNAs. These results are generally in good agreement with our previous data showing that only a small number of tRNAs are coded for by the mitochondrial DNA, while the others are a selected set of imported cytoplasmic tRNAs. PMID:3127061

  18. Analysis of Streptomyces coelicolor membrane proteome using two-dimensional native/native and native/sodium dodecyl sulfate gel electrophoresis.

    PubMed

    Li, Fuhou; Liang, Jingdan; Wang, Weixia; Zhou, Xiufen; Deng, Zixin; Wang, Zhijun

    2014-11-15

    Analysis of the oligomeric state of a protein may provide insights into its physiological functions. Because membrane proteins are considered to be the workhorses of energy generation and polypeptide and nutrient transportation, in this study we characterized the membrane-associated proteome of Streptomyces coelicolor by two-dimensional (2D) blue native/sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), high-resolution clear native/native PAGE, and native/SDS-PAGE. A total of 77 proteins were identified, and 20 proteins belonging to 15 complexes were characterized. Moreover, the resolution of high-resolution clear native/SDS-PAGE is much higher than that of blue native/SDS-PAGE. OBP (SCO5477) and BldKB (SCO5113) were identified as the main protein spots from the membrane fractions of S. coelicolor M145, suggesting that these two proteins are involved in extracellular peptide transportation. These two transporters exhibited multiple oligomeric states in the native PAGE system, which may suggest their multiple physiological functions in the development of S. coelicolor. PMID:25150108

  19. Comigration of two autophagosome-associated dehydrogenases on two-dimensional polyacrylamide gels.

    PubMed

    Sneve, Marianne Lunde; Øverbye, Anders; Fengsrud, Monica; Seglen, Per O

    2005-01-01

    Immunoblotting of two-dimensional polyacrylamide gels (pI 3-10) revealed six cytosolic molecular forms of glyceraldehyde-3-phosphate dehydrogenase (GAPDH) in rat hepatocytes. Two of the four full-length (approximately 37 kDa) forms exhibited some binding to sedimentable cellular elements (but not to mitochondria), whereas one full-length and two short (approximately 35 kDa) forms selectively bound to the membranes of autophagosomes and lysosomes. Tryptic fingerprinting by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) confirmed the identity of the major full-length forms as GAPDH, but attempts to identify the major short form consistently suggested that this spot represented a different enzyme, 3-alpha-hydroxysteroid dehydrogenase (3alphaHSD). Silver staining indicated that this 3alphaHSD form selectively bound to autophagosomal and lysosomal membranes. Immunoblotting of more focused 2D gels (pI 6-9) with an antibody raised against 3alphaHSD demonstrated immunostaining of four 3alphaHSD forms with masses of about 35 kDa. Autophagosomal membrane preparations were highly and selectively enriched with respect to all of these 3alphaHSD forms. One of them comigrated with the major short form of GAPDH, accounting for the paradoxical mass spectrometric identification of 3alphaHSD from this spot. Proteomic analysis by a combination of immunological and mass spectrometric identification methods was thus capable of resolving two comigrating dehydrogenases selectively associated with autophagic organelles. PMID:16874067

  20. Nonlinear elastic properties of polyacrylamide gels: Implications for quantification of cellular forces

    PubMed Central

    Boudou, Thomas; Ohayon, Jacques; Picart, Catherine; Pettigrew, Roderic I.; Tracqui, Philippe

    2015-01-01

    Because of their tunable mechanical properties, polyacrylamide gels (PAG) are frequently used for studying cell adhesion and migratory responses to extracellular substrate stiffness. Since these responses are known to heavily depend on the tensional balance between cell contractility and substrate mechanical resistance, a precise knowledge of PAGs mechanical properties becomes quite crucial. Using the micropipette aspiration technique, we first exhibited the nonlinear elastic behavior of PAG and then successfully modeled it by an original strain-energy function. This function depends on the Poissons ratio and on two material parameters, which have been explicitly related to acrylamide and bis-acrylamide concentrations. Implications of these results have been highlighted with regard to traction force microscopy experiments where cellular force quantification is derived from displacements of beads embedded in PAG. We found that considering PAG as a linear elastic medium tends to significantly underestimate traction forces for substrate displacements larger than 2 ?m. Interestingly, we also showed that in the range of cellular force amplitude and PAG stiffness currently used in cell traction force experiments, finite size effects become critical for PAG substrate thickness below 60 ?m. Thus, our improved characterization of PAG nonlinear mechanical properties through a new constitutive law could have significant impact onto biological experimentations where such extracellular substrates experience large strains. PMID:19581727

  1. Comparative sodium dodecyl sulfate-polyacrylamide gel electrophoresis and restricted fragment length polymorphism among fenugreek accessions.

    PubMed

    Haliem, E A; Al-Huqail, A A

    2013-01-01

    Protein and DNA polymorphismswere surveyed among seven accessions of wild fenugreek (Trigonellafoenum-graecum L.) to estimate their genetic diversity and relationships. Samples were obtained from diverse ecogeographical areas in Saudi Arabia and Yemen. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) analysis of seed storage protein showed genetic variations among fenugreek germplasms, both quantitatively and qualitatively, generating a total of 168 polypeptide bands with different molecular weights ranging from 4.5 to 300 kDa. Twenty-six of these bands were polymorphic, with a considerable polymorphism value (80.00%). Furthermore, restriction fragment length polymorphism (RFLP) analysis was also employed, which was based on the ability of four restriction enzymes (EagI, EcoRI, FspI, and HindIII) to cleave genomic DNA of the plant materials at specific target nucleotide sequences into different numbers of DNA fragments. RFLP analysis revealed 166 fragments with known sequences and variable lengths ranging from 80 to 4000 bp with a highly degree of polymorphism (88.71%). Data derived from SDS-PAGE or RFLP analyses were used to produce dendrograms, which clustered the studied fenugreek accessions into different groups based on the unweighted pair group method with arithmetic mean (UPGMA). The resulting relationships indicated that these two marker techniques were nearly equivalent, but not identical, with respect to phylogenetic information. In conclusion, SDS-PAGE analysis of seed proteins should be augmented with RFLP analysis of DNA for reliable estimates of genetic diversity among fenugreek germplasms. PMID:24338424

  2. Carbon dioxide adsorption on polyacrylamide-impregnated silica gel and breakthrough modeling

    NASA Astrophysics Data System (ADS)

    Zhao, Yi; Shen, Yanmei; Bai, Lu; Ni, Shiqing

    2012-11-01

    Polyacrylamide-impregnated silica gel was prepared to capture CO2 from flue gas. The polymerization of acrylamide was carried out in AN solvent using AIBN as initiator and EGDMA as crosslinker. The adsorbents were characterized by N2 adsorption, FTIR analysis, SEM analysis, and thermal gravimetric analysis. The results showed that the polymer was not only occupying the porosity of the silica, but necessarily surrounding silica particles, and the amide groups was successfully loaded on the support silica. The impregnated silica displayed good thermal-stability at 250 C. The CO2 adsorption isotherms were measured to examine CO2 adsorption on adsorbents, and the results showed that the capacity was increased significantly after modification. The CO2 isosteric adsorption heats calculated from the isotherms showed that the adsorption interaction of CO2 with the functionalized material may be mainly an intermolecular force or hydrogen bond. Fixed-bed breakthrough model of CO2 adsorption on functionalized silica was successfully developed to describe the breakthrough curves under different adsorption temperature, CO2 concentration, and gas flow rate. The mass transfer coefficients of CO2 were calculated from the breakthrough model, the results showed that adsorption rate could be promoted by increasing temperature, flow rate and CO2 concentration, among which the effect of gas flow rate is the most obvious.

  3. Microscopic agglutination and polyacrylamide gel electrophoresis analyses of oral anaerobic spirochetes.

    PubMed Central

    Tall, B D; Nauman, R K

    1986-01-01

    Microscopic agglutination (MA) analysis and sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) were used to determine strain and species similarities and dissimilarities among three species of oral anaerobic spirochetes, Treponema denticola, Treponema pectinovorum, and Treponema vincentii. The MA analysis revealed a diversity of serologic reactivity or sharing of common antigens within each species. However, there was no cross-reactivity or sharing of common antigens among the three species. Distinct SDS-PAGE whole-cell electrophoretograms for each species were obtained. The banding patterns for 16 T. denticola strains revealed 30 distinct proteins, while the banding patterns for 5 strains of T. pectinovorum and 2 strains of T. vincentii revealed 26 and 35 distinct proteins, respectively. Analysis of the electrophoretograms showed that their respective banding patterns could be used to distinguish the three species from one another. In addition, strain differences within each species could be detected. There was a correlation between MA analysis and SDS-PAGE analysis. It is thus suggested that both MA and SDS-PAGE analysis be included in classification schemes for the identification of oral spirochetes. Images PMID:3745424

  4. A new method for HDL particle sizing by polyacrylamide gradient gel electrophoresis using whole plasma.

    PubMed

    Prusse, M; Pascot, A; Desprs, J P; Couillard, C; Lamarche, B

    2001-08-01

    Low plasma levels of HDL cholesterol have been associated with an increased risk of coronary heart disease. HDL particles are heterogeneous with respect to size and apolipoprotein content. The objective of the present study was to develop a method to generate lipid-stainable calibrators that would allow the assessment of HDL particle size from whole plasma, using polyacrylamide gradient gel electrophoresis (PAGGE). Lipid-stainable HDL calibrators were obtained by subjecting isolated red blood cells to hemolysis either by freezing at -20 or -80 degrees C overnight or by rapid exposure to liquid nitrogen and mixing of the hemolysis products with plasma aliquots. All three methods were highly reproducible in producing Sudan black lipid-stainable HDL calibrators ranging from 75 to 200 A. The assessment of HDL particle size with these lipid-stainable HDL calibrators was also highly reproducible, with a coefficient of variation below 5.5%. These lipid-stainable HDL calibrators simplify the assessment of HDL particle size by PAGGE using whole plasma, without the need for costly, time-consuming ultracentrifugation procedures. PMID:11483636

  5. High resolution of honey bee (Apis mellifera) venom peptides by propionic acid/urea polyacrylamide gel electrophoresis after ethanol precipitation.

    PubMed

    Chettibi, S; Lawrence, A

    1989-01-01

    A new and simple gel electrophoretic method is described which enables the protein and polypeptide components of bee venom to be resolved on a single gel. The electrophoretic method allows octapeptides to be resolved and species as small as decapeptides can be detected at high sensitivity using the Coomassie blue staining method without prior fixation. This has been achieved by replacing acetic acid by propionic acid in acid/urea polyacrylamide gels and by controlling the amount of TEMED catalyst for the polymerisation of high concentration gels in order to obtain a low effective pore size. We demonstrated the value of ethanol precipitation as a rapid and efficient desalting the fractionation technique and propose that it could be used in combination with gel filtration to purify many of the peptides to homogeneity. PMID:2781578

  6. Polyacrylamide lamination enables mass spectrometry compatible staining and in-gel digestion of proteins separated by agarose IEF.

    PubMed

    Hellman, Jukka

    2007-10-01

    Agarose IEF enables the separation of large proteins and protein complexes. A complication of agarose gels attached onto polyester support is the lack of sensitive protein staining methods compatible with protein analysis and identification protocols. In this study, agarose IEF gels were used to separate the proteins, followed by layering the agarose with polyacrylamide. The formed laminate gels were seamless and durable and they were readily detached from the polyester. The gels were amenable to MS compatible staining. The sensitivity obtained with the acidic silver staining method was 20-50 ng/band of myoglobin. Laminated agarose was a suitable matrix for in-gel digestion based generation of tryptic peptides for MALDI-MS. PMID:17722206

  7. Investigation of tetrakis hydroxymethyl phosphonium chloride as an antioxidant for use in x-ray computed tomography polyacrylamide gel dosimetry

    NASA Astrophysics Data System (ADS)

    Jirasek, A.; Hilts, M.; Shaw, C.; Baxter, P.

    2006-04-01

    Of the antioxidants used to scavenge oxygen in polymer gel dosimeters, tetrakis (hydroxymethyl) phosphonium chloride (THPC) has been shown to hold great promise due to its rapid oxygen scavenging abilities. In this study we (a) investigate the use of THPC as an antioxidant for polyacrylamide gel (PAGAT) dosimeters used in conjunction with x-ray computed tomography (CT) and (b) work to establish the reaction mechanisms of THPC with the polymer gel constituents. We establish the dose response reproducibility of PAGAT dosimeters when imaged with CT and show that PAGAT dosimeters exhibit highly reproducible dose responses for a range of irradiation times post gel manufacture (2-6 h) and CT imaging times post gel irradiation (1-5 days). The THPC concentration within the gel leading to a maximized dose response and minimized O2 inhibition of polymerization is found to be ~4.5 mM. We further assess the stability of PAGAT dosimeters by investigating the reactions of THPC with the individual gel constituents. The importance of utilizing deionized water in polymer gel manufacture is noted. We show that, while THPC remains unreactive with acrylamide and bis-acrylamide under unirradiated conditions, THPC can react with gelatin to increase the cross-linking of the gelatin matrix in unirradiated dosimeters. THPC reactions with gelatin can lead to the lower observed dose sensitivity of PAGAT (~0.36 0.04 H Gy-1) as compared to polyacrylamide gels manufactured under anoxic conditions (~0.83 0.03 H Gy-1). The reactions of THPC which lead to O2 scavenging, and potential reactions of THPC with other gel constituents, are proposed.

  8. Proteins specified by herpes simplex virus. Staining and radiolabeling properties of B capsid and virion proteins in polyacrylamide gels.

    PubMed

    Gibson, W; Roizman, B

    1974-01-01

    ANALYSES OF THE STRUCTURAL PROTEINS OF HERPES SIMPLEX VIRIONS AND OF CAPSIDS CONTAINING VIRAL DNA (B CAPSIDS), AFTER ELECTROPHORESIS IN POLYACRYLAMIDE GELS, REVEALED CONSIDERABLE VARIABILITY IN THEIR PROPERTIES WITH RESPECT TO: (i) retention of Coomassie brilliant blue (CBB) and fast green stains during destaining, (ii) relative optical absorbance of the CBB-protein complex at different wavelengths, (iii) relative efficiency with which (14)C-amino acids are incorporated during early and late periods of the infection cycle, and (iv) capacity to be phosphorylated in vivo. In addition, it was found that protein 22a of B capsids, which does not have an electrophoretically identical counterpart in virions, shares a relatively unique set of staining and radiolabeling properties with virion protein 22, which has a slightly more rapid electrophoretic mobility in sodium dodecyl sulfate-polyacrylamide gels. PMID:4129836

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

    PubMed

    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. PMID:26203364

  10. Immobilization of microbial cells in crosslinked, prepolymerized, linear polyacrylamide gels: antibiotic production by immobilized Streptomyces clavuligerus cells

    SciTech Connect

    Freeman, A.; Aharonowitz, Y.

    1981-12-01

    A mild method for the immobilization of whole microbial cells has been developed. Cells were suspended in a solution of preformed, linear, water-soluble polyacrylamide chains, partially substituted with acylhydrazide groups. The prepolymerized backbone polymer was crosslinked, in the presence of viable cells, by stoichiometric amounts of dialdehydes such as glyoxal, glutardialdehyde, and periodate-oxidized polyvinyl alcohol. The crosslinking reaction, carried out in cold, neutral physiological conditions resulted in cells entrapped in gels with physical properties similar to those of the common polyacrylamide gels. However, cell damage generally caused by the acrylamide monomer was avoided. Resting Streptomyces clavuligerus cells, possessing a high capacity for antibiotic production, were entrapped according to this procedure. These immobilized cells produced cephalosporins continuously for 96 hours with yields similar to those of free resting cells. The same cells, when immobilized by direct polymerization of acrylamide monomers, yielded significantly lower amounts of antibiotics. (Refs. 19).

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

    PubMed Central

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

    2015-01-01

    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. PMID:26203364

  12. An improved sodium dodecyl sulfate-polyacrylamide gel electrophoresis system for the analysis of membrane protein complexes.

    PubMed

    Kashino, Y; Koike, H; Satoh, K

    2001-04-01

    Membrane protein complexes such as the reaction center complexes of oxygenic photosynthesis or the complex I of mitochondira are composed of many subunit polypeptides. To analyze their polypeptide compositions by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), a wide range of molecular sizes has to be resolved, especially in the low molecular mass range. We have improved the traditional Tris/HCI buffer systems adopting a Tris/2-(N-morpholino)ethanesulfonic acid (MES) buffer system containing 6 M urea. This gel system was used with an 18-24% acrylamide gradient for the separation of polypeptides with molecular masses from below 5 kDa to over 100 kDa. This buffer system can also be applied to the usual uniform concentration of acrylamide gel and also to minislab gels. PMID:11358120

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

    PubMed

    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. PMID:24388443

  14. Multifunctional graphene incorporated polyacrylamide conducting gel electrolytes for efficient quasi-solid-state quantum dot-sensitized solar cells

    NASA Astrophysics Data System (ADS)

    Duan, Jialong; Tang, Qunwei; Li, Ru; He, Benlin; Yu, Liangmin; Yang, Peizhi

    2015-06-01

    Pursuit of a high efficiency and stability has been a persistent objective for quantum dot-sensitized solar cells (QDSCs). Here we launch a strategy of synthesizing graphene implanted polyacrylamide (PAAm-G) conducting gel electrolytes for quasi-solid-state QDSCs. With an aim of elevating the dosage of S2-/Sx2- redox couples and therefore charge-transfer ability, both osmotic press across the PAAm-G and capillary force within the three-dimensional micropores are utilized as driving forces. A promising power conversion efficiency of 2.34% is recorded for the QDSCs by optimizing graphene dosage in the conducting gel electrolyte. The enhanced conversion efficiency of solar cell is attributed to the expanded catalytic area from counter electrolyte/electrolyte interface to both interface and the conducting gel electrolyte.

  15. Applicability of RAPD markers on silver-stained polyacrylamide gels to ascertain genetic diversity in Peripatus acacioi (Peripatidae; Onychophora).

    PubMed

    DeLaat, Daiane Mariele; Carvalho, Maria Raquel Santos; Lovato, Maria Bernadete; Acedo, Maria Dolores Porto; da Fonseca, Cleusa Graa

    2005-01-01

    RAPD (random amplification of polymorphic DNA) molecular markers can be utilized for analyzing genetic variability in populations for which only a few or no molecular markers are available. They were used in a study of an endangered species, Peripatus acacioi, found in the Tripu Ecological Station, in Ouro Preto, MG, Brazil. The ecological station was specifically created to protect this velvet worm species, the first of this group found in Brazil. For an initial evaluation of the genetic diversity of this species, DNA samples from the lobopods of four individuals, collected at random, were analyzed using RAPD. Each reaction was run with a different primer (Operon RAPD 10-mer Kits), totaling 13 primers (OPC2, OPC3, OPC4, OPC6, OPC8, OPC10, OPC11, OPL2, OPL7, OPL11, OPL13, OPL18, and OPL19). Due to the low amplification yield, RAPD fragments were separated in polyacrylamide gels and stained with silver nitrate. Numerous bands were observed. Fifty-five of the amplified bands proved to be reproducible, both in terms of presence and intensity. Among these, 27 were variable and 28 were constant. The average number of bands per gel was 4.2. Nine of the 13 primers tested allowed the identification of constant and variable bands among these four individuals. RAPD analysis of genetic variation using silver-stained polyacrylamide gel electrophoresis provided measures of band sharing among the individuals, and therefore could be used in population genetics studies of P. acacioi. PMID:16475117

  16. Identification and mapping of human saphenous vein medial smooth muscle proteins by two-dimensional polyacrylamide gel electrophoresis.

    PubMed

    McGregor, E; Kempster, L; Wait, R; Welson, S Y; Gosling, M; Dunn, M J; Powel, J T

    2001-11-01

    Changing smooth muscle phenotype and abnormal cell proliferation are important features of vascular pathology, including the failure of saphenous vein bypass grafts. We have characterised and mapped protein expression in human saphenous vein medial smooth muscle, using two-dimensional (2-D) polyacrylamide gel electrophoresis. The 2-D system comprised a nonlinear immobilised pH 3-10 gradient in the first dimension (separating proteins with isoelectric point values between pH 3-10), and 12%T total gel concentration sodium dodecyl sulphate polyacrylamide gel electrophoresis in the second dimension (separating proteins in the range 14,000-200,000 Daltons). Using a combination of peptide mass fingerprinting by matrix-assisted laser desorption/ionisation-time of flight mass spectrometry and partial amino acid sequencing by nanospray tandem mass spectrometry, a subset of 149 protein spots was analysed, with 129 protein spots being identified and mapped. The data presented here are an important addition to the limited knowledge of venous medial smooth muscle protein expression in vivo. Our protein map will facilitate the identification of proteins differentially expressed in human saphenous vein bypass grafts. In turn, this may lead to the elucidation of molecular events involved in saphenous vein bypass graft failure. The map should also provide a basis for comparative studies of protein expression in vascular smooth muscle of varying origins. PMID:11922600

  17. The use of N,N,N',N'-tetramethylphenylenediamine to detect peroxidase activity on polyacrylamide electrophoresis gels.

    PubMed

    Butler, M J; Lachance, M A

    1987-05-01

    N,N,N',N'-Tetramethylphenylenediamine (TMPD) acts as an effective indicator of peroxidase activity on polyacrylamide electrophoresis gels. The test is easy to perform, rapid, sensitive, and reliable. The procedure produces vivid bright blue bands (Wursters blue) on a clear background. TMPD and Wursters blue did not interfere with a number of other electrophoresis stains subsequently applied. These included total protein staining with Coomassie blue, and a number of pigment producing electrophoresis stains used to investigate melanogenesis-related enzymes in the black yeast Phaeococcomyces sp. PMID:2440348

  18. High resolution two-dimensional electrophoresis and sodium dodecyl sulphate-polyacrylamide gel electrophoresis of cheese proteins after rapid solubilisation.

    PubMed

    Marshall, T; Williams, K M

    1988-03-01

    The proteins of cheese are rapidly solubilised by heating to 95 degrees C in buffered 2% sodium dodecyl sulfate, 5% 2-mercaptoethanol. Electrophoretic analysis of the solubilised proteins by either one-dimensional sodium dodecyl sulphate-polyacrylamide gel electrophoresis or high resolution two-dimensional electrophoresis yields reproducible patterns characteristic of an individual cheese and its extent of ripening. The patterns reveal (i) the residual amounts of milk casein and whey proteins, and (ii) the appearance of casein degradation products, including pink-violet components as detected by Coomassie Blue staining. PMID:2466651

  19. Nanopore density effect of polyacrylamide gel plug on electrokinetic ion enrichment in a micro-nanofluidic chip

    NASA Astrophysics Data System (ADS)

    Wang, Jun-yao; Xu, Zheng; Li, Yong-kui; Liu, Chong; Liu, Jun-shan; Chen, Li; Du, Li-qun; Wang, Li-ding

    2013-07-01

    In this paper, the nanopore density effect on ion enrichment is quantitatively described with the ratio between electrophoresis flux and electroosmotic flow flux based on the Poisson-Nernst-Planck equations. A polyacrylamide gel plug is integrated into a microchannel to form a micro-nanofluidic chip. With the chip, electrokinetic ion enrichment is relatively stable and enrichment ratio of fluorescein isothiocyanate can increase to 600-fold within 120 s at the electric voltage of 300 V. Both theoretical research and experiments show that enrichment ratio can be improved through increasing nanopore density. The result will be beneficial to the design of micro-nanofluidic chips.

  20. Isolation and characterization of the pigment-protein complexes of Rhodopseudomonas sphaeroides by lithium dodecyl sulfate/polyacrylamide gel electrophoresis.

    PubMed Central

    Broglie, R M; Hunter, C N; Delepelaire, P; Niederman, R A; Chua, N H; Clayton, R K

    1980-01-01

    When purified photosynthetic membranes from Rhodopseudomonas sphaeroides were treated with lithium dodecyl sulfate and subjected to polyacrylamide gel electrophoresis at 4 degrees C, up to 11 pigment-protein complexes were resolved. Absorption spectra revealed that the smallest complex contained reaction center pigments and the others contained the antenna components B850 and B875 in various proportions. Of these antenna complexes, the largest was almost entirely B850 and the smallest contained only B875. After solubilization at 100 degrees C and electrophoresis on polyacrylamide gradient gels, the B850 complex gave rise to two polypeptide components migrating with apparent Mr of 10,000 and 8000, whereas with the B875 complex, two components were observed with apparent Mr of 12,000 and 8000. The reaction center complex gave rise to only the 24 and 21 kilodalton polypeptide subunits. Fluorescence emission spectra showed maxima at 872 and 902 nm for B850 and B875, respectively. Analyses of bacteriochlorophyll a and carotenoids indicated that, in the B875 complex, two molecules of each of these pigments are associated with the two polypeptides. The associations of B850 and B875 in large and small complexes obtained by lithium dodecyl sulfate treatment are consistent with models of their organization within the membrane. Images PMID:6965795

  1. Detection of actin-binding proteins in human platelets by 125I-actin overlay of polyacrylamide gels.

    PubMed

    Snabes, M C; Boyd, A E; Bryan, J

    1981-09-01

    Actin-binding proteins have been identified in human platelets with a gel-overlay technique that uses 125I-G-actin. Platelet proteins were separated on SDS polyacrylamide gels using the buffer system of Laemmli (1970, Nature [Lond.] 227:680-685). The proteins were fixed in the gels with methanol-acetic acid, the SDS was washed out, and the proteins were renatured. The gels were incubated with 125I-G-actin from rabbit skeletal muscle that was radiolabeled with 125I according to the method of Bolton and Hunter (1973, Biochem. J. 133:529-538) and has been shown to retain biological activity. After nonspecifically bound radioactivity was washed out, gels were dried and processed for autoradiography. The 125I-G-actin binds to several proteins in human platelets, platelet extracts, and the particulate fraction. Control experiments demonstrate that the 125I-G-actin can be displaced by use of increasing amounts of unlabeled actin, that the binding is stable to 0.6 M NaCl, and that preheating the 125I-G-actin to 90 degrees C for 3 min eliminates all binding. Prominent 125I-G-actin-binding activities were present at Mr 90,000 and 40,000. The binding to the 90,000 Mr protein appears to be at least partially Ca++ sensitive, whereas the binding to the 40,000 Mr protein does not. 125I-G-actin bound to proteins in the SDS gels can be fixed in situ and compared directly with the stained gel. This technique should prove generally useful in identification and purification of some actin-binding proteins from cells and tissues. PMID:6793603

  2. The Use of Polyacrylamide Gels for Mechanical Calibration of Cartilage – A Combined Nanoindentation and Unconfined Compression Study

    PubMed Central

    Li, Cheng; Allen, Jessica; Alliston, Tamara; Pruitt, Lisa A.

    2015-01-01

    This study investigates polyacrylamide (PA) gel as a calibration material to measure the nanomechanical compressive modulus of cartilage using nanoindentation. Both nanoindentation and unconfined compression testing were performed on PA gel and porcine rib cartilage. The equilibrium moduli measured by the two methods were discernable. Nanoindentation has the advantage of distinguishing between spatially dependent constituent properties that affect tissue mechanical function in heterogeneous and hierarchically structured tissues such as cartilage. Both sets of measurements exhibited similar positive correlation with increasing gel crosslinker concentration. The compressive modulus measurements from compression in the PA gels ranged from 300 kPa—1.4 MPa, whereas those from nanoindentation ranged from 100 kPa—1.1MPa. Using this data, a method for relating nanoindentation measurements to conventional mechanical property measurements is presented for porcine rib cartilage. It is shown that based on this relationship, the local tissue modulus as measured from nanoindentation (1.1—1.4 MPa) was able to predict the overall global modulus of the same sample of rib cartilage (2.2 MPa), as confirmed by experimental measurements from unconfined compression. This study supports the use of nanoindentation for the local characterization of cartilage tissues and may be applied to other soft tissues and constructs. PMID:21783163

  3. A comparison of silver staining protocols for detecting DNA in polyester-backed polyacrylamide gel.

    PubMed

    Qiu, Shanlian; Chen, Jichen; Lin, Si; Lin, Xinjian

    2012-04-01

    Eight silver-staining protocols were applied to detect DNA in polyester-backed gels to select the optimal. Results showed important differences in staining quality and that four methods were well-suited for TGGE gels due to high sensitivity and low background, including the Bassam et al. methods, the manufacturer method and our improved method. PMID:24031876

  4. Fast reversible single-step method for enhanced band contrast of polyacrylamide gels for automated detection

    PubMed Central

    Ling, Wei-Li; Lua, Wai-Heng; Gan, Samuel Ken-En

    2015-01-01

    Staining SDS-PAGE is commonly used in protein analysis for many downstream characterization processes. Although staining and destaining protocols can be adjusted, they can be laborious, and faint bands often become false negatives. Similarly, these faint bands hinder automated software band detections that are necessary for quantitative analyses. To overcome these problems, we describe a single-step rapid and reversible method to increase (up to 500%) band contrast in stained gels. Through the use of alcohols, we improved band detection and facilitated gel storage by drying the gels into compact white sheets. This method is suitable for all stained SDS-PAGE gels, including gradient gels and is shown to improve automated band detection by enhanced band contrast. PMID:25782090

  5. Matrix-assisted laser desorption/ionization mass spectrometry of proteins extracted directly from sodium dodecyl sulphate-polyacrylamide gels.

    PubMed

    Ehring, H; Strömberg, S; Tjernberg, A; Norén, B

    1997-01-01

    A new strategy for the characterization of Coomassie Brilliant Blue stained SDS-PAGE separated proteins by UV-MALDI-MS is reported. The proteins are extracted directly from the polyacrylamide gel by treatment with an organic solvent mixture consisting of formic acid, acetonitrile, isopropanol and water in an ultrasonic bath. A fraction of the supernatant is then mixed directly with the matrix solution and measured by MALDI-MS. High quality spectra could be obtained from gels which were loaded with 6 pmol of myoglobin. Compared to other methods based on electroblotting or electroelution this method is much simpler and less time consuming. The sensitivity is higher than or comparable to the Coomassie Blue staining procedure for proteins up to about 25 kDa. Another advantage is that mass shifts due to charging effects of the membranes, which are common if membranes are mounted directly on the sample target, can be avoided. However, all proteins studied showed slightly higher masses than expected which reduces mass accuracy to 0.2-0.3%. This is presumably partly due to formylation of serine or threonine residues during incubation in formic acid. Gel electrophoresis induced modifications can contribute as well. The possibility of further characterizing the remaining part of the supernatant after extraction by means of proteolytic digestion is also demonstrated. The knowledge of both molecular weight of the whole protein and of the proteolytic fragments increases specificity for protein identification by searching in sequence databases. PMID:9404036

  6. A novel [Ag(NH3)2]+ probe for chemiluminescent imaging detection of proteins after polyacrylamide gel electrophoresis.

    PubMed

    Xiong, Xin; Wang, Zhenzhen; Baeyens, Willy R G; Delanghe, Joris R; Huang, Zhi; Huang, Guangming; Ouyang, Jin

    2007-08-01

    The development of a novel [Ag(NH3)2]+ probe chemiluminescence (CL)-based imaging method for the detection of various proteins after PAGE is described. The detection is based upon the probe [Ag(NH3)2]+ catalyzing the CL reaction of the luminol-potassium persulfate system. The proposed method detects various proteins labeled by [Ag(NH3)2]+ and expands the application scope to SDS gels. It also detects proteins directly in polyacrylamide gels, without tedious transferring procedures. Furthermore, successful identification of proteins by peptide mass profiling using ionization MS was easily performed, and no pretreatments of gel prior to digestion are needed. Detection limits for standard marker proteins match CBB-R250 staining and the linear dynamic range is superior to CBB-R250 staining and silver staining. The CL imaging conditions, including luminescent reagents, silver ion concentration, the ammonia-controlled system and the washing reagents parameters have also been optimized. PMID:17610207

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

    PubMed

    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. PMID:26796977

  8. Tryptic peptide analysis of nanogram quantities of proteins: radioiodination of proteins detected by silver staining in polyacrylamide gels

    SciTech Connect

    Arezzo, F.; Rose, K.M.

    1987-12-01

    The silver-staining procedure for detecting proteins in polyacrylamide gels has been modified so that the polypeptides remain suitable for subsequent radioiodination and tryptic peptide analysis. The procedure, which involves a silver-staining/destaining protocol that minimizes crosslinking, is more rapid than many other methods, and can detect as little as 1 ng of protein. Following elimination of silver, the proteins can be radioiodinated and digested with trypsin by a modification of the method described by J. H. Elder, R. A. Pickett, J. Hampton, and R. A. Lerner. Together, these improvements have increased the sensitivity of the tryptic peptide mapping technique by three orders of magnitude and thereby enabled us to perform reproducible structural analysis of femtomolar quantities of proteins.

  9. Anomalous electrophoretic behavior of a chitinase isoform from grape berries and wine in glycol chitin-containing sodium dodecyl sulfate-polyacrylamide gel electrophoresis gels.

    PubMed

    Vincenzi, Simone; Curioni, Andrea

    2005-01-01

    An anomalous electrophoretic behavior of a chitinase isoform present in both grape (Vitis vinifera L.) berries and wine was observed in glycol chitin-containing sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) gels. A progressive shift of the relative molecular mass M(r) of the enzyme (from approximately 30,500 up to approximately 57,700) with increasing glycol chitin concentration in the gels up to 0.1% was revealed when samples were electrophoresed under nonreducing conditions, whereas the presence of glycol chitin had no effects when samples were reduced before SDS-PAGE separation. The M(r) of other grape and wine chitinase isoforms as well as that of the chitinase from pomegranate (Punica granatum L.) fruit was unaffected by the presence of the substrate in the gel under both reducing and nonreducing conditions. Since the enzymes were inactive during the electrophoretic separation, it is likely that the retarding effect of glycol chitin observed specifically for the unreduced chitinase band from grape and wine was due to an interaction between the substrate and a chitin-binding domain different from the catalytic site, such as that typical of class I and class IV chitinases. PMID:15624140

  10. Two-dimensional polyacrylamide gel electrophoresis of ribosomal proteins in the nanogram range.

    PubMed

    Brockmller, J; Kamp, R M

    1985-09-01

    A two-dimensional gel electrophoresis system to identify and check the purity of ribosomal proteins from different organisms with nanogram quantities is described. This procedure combines the method of Geyl et al. for the separation of ribosomal proteins of Escherichia coli, and the microscale electrophoresis system for proteins described by Neuhoff and Poehling, with several modifications. The first gel dimension is carried out in capillaries and the second in the form of slab gels, both are run in newly designed chambers suitable for 10-20 samples. This electrophoresis system enables a reduction of the running time from 2 days to 2 hours and an increase in sensitivity, with Coomassie blue staining, from 3-5 micrograms for the normal 100 X 100 mm gels to 50-100 ng. The resolution of all ribosomal proteins on the micro-gel (30 X 38 X 0.5 mm) is similar to the separation on the mini-gel of 100 X 100 X 3 mm as described by Geyl et al. PMID:3907663

  11. Detection of PEGylated proteins in polyacrylamide gels by reverse staining with zinc and imidazole salts.

    PubMed

    Hardy, Eugenio; Ramón, José; Saez, Vivian; Báez, Reynier; Tejeda, Yosbel; Ruiz, Amalia

    2008-06-01

    The reverse staining, with imidazole-SDS-zinc, of PEG-linked proteins separated by SDS-PAGE was studied. Using model conjugates (interferon-alpha 2b (IFN-alpha2b) reacted with either a branched-chain (40,000) PEG (PEG2,40) or a linear monomethoxy PEG polymer (Mr of 12,000) and chromatographically purified monoPEG2,40-IFN-alpha2b), conventional small-format analytical gels (<1 mm thick) showed typical detection patterns (i.e., transparent, colorless bands clearly discernible against a zinc imidazolate-generated white gel background), in less than 20 min. Nonreacted (free) PEG was almost undetected, as expected. The reverse-stained PEGylated IFN-alpha2b patterns were qualitatively indistinguishable from those of parallel gels stained with iodine (I2). The LOD was estimated in the low nanogram range (e.g., at about 7 ng for mono- or bi-PEG2,40 IFN-alpha2b per lane on gradient (4-17%) gels). Also, this stain allowed the visualization of Coomassie blue-undetected PEG-IFN bands, and could be restained with I2. PEGylated species of lysozyme, a low-molecular-weight peptide, ovalbumin, and chymotrypsin were used to demonstrate the generality of this stain. We also show (i) how to counteract the adverse effect of some parameters (e.g., gel thickness above 1 mm, long gel length, low (e.g., 4-6%) acrylamide concentration) on the reverse staining process and (ii) that the properties of the reverse-stained PEGylated proteins remain unchanged, as judged by analyzing both the ion exchange chromatography-based positional isomer separation profile and enzyme-linked immunosorbent response of PEG-IFN recovered from gels. Consequently, this technique may be useful for the rapid analysis or the small-scale preparation of PEGylated proteins. PMID:18449861

  12. Fluorography--limitations on its use for the quantitative detection of /sup 3/H- and /sup 14/-C-labeled proteins in polyacrylamide gels

    SciTech Connect

    Harding, C.R.; Scott, I.R.

    1983-03-01

    The suitability of fluorography for the detection of /sup 3/H- and /sup 14/C-labeled proteins on polyacrylamide gradient gels has been investigated. If was found that the absorbance of the fluorographic film image produced by a given level of radioactivity decreased as the acrylamide concentration in the gel increased. The use of Coomassie brilliant blue protein dyes to stain the gel prior to fluorography reduced the absorbance of the fluorographic image. It is concluded that quantitative fluorography can only be applied to unstained gels of a uniform acrylamide concentration.

  13. Metachromatic staining patterns of basic proline-rich proteins from rat and human saliva in sodium dodecyl sulfate-polyacrylamide gels.

    PubMed

    Humphreys-Beher, M G; Wells, D J

    1984-01-01

    A series of basic proteins, rich in proline, were isolated from the salivary secretions of humans and rats. These proteins underwent metachromasia after staining with Coomassie brilliant blue R-250 in sodium dodecyl sulfate-polyacrylamide gels. The technique of destaining gels in several changes of 10% acetic acid after a 30-min staining period is a rapid method of general utility for the identification of proline-rich proteins from total cell lysates from other sources besides saliva. PMID:6085627

  14. Prestaining of glycoproteins in sodium dodecyl sulfate polyacrylamide gels by dansylhydrazine.

    PubMed

    Wang, Yang; Zhou, Xuan; Yu, Qing; Duan, Yuanmeng; Huang, Binbin; Hong, Guoying; Zhou, Ayi; Jin, Litai

    2014-06-01

    A new fluorescent prestaining method for gel-separated glycoproteins in 1D and 2D SDS-PAGE was developed by using dansylhydrazine in this study. The prestained gels could be easily imaged after electrophoresis without any time-consuming steps needed for poststains. As low as 4-8 ng glycoproteins (transferrin, ?1-acid glycoprotein) could be selectively detected, which is comparable to that of Pro-Q Emerald 488, one of the most commonly used glycoprotein stain. In addition, a subsequent study of deglycosylation, glycoprotein affinity isolation, and LC-MS/MS analysis was performed to confirm the specificity of the newly developed method. PMID:24668852

  15. Transfer ribonucleic acid synthesis during sporulation and spore outgrowth in Bacillus subtilis studied by two-dimensional polyacrylamide gel electrophoresis.

    PubMed Central

    Henner, D J; Steinberg, W

    1979-01-01

    The synthesis of transfer ribonucleic acid (tRNA) was examined during spore formation and spore outgrowth in Bacillus subtilis by two-dimensional polyacrylamide gel electrophoresis of in vivo 32P-labeled RNA. The two-dimensional gel system separated the B. subtilis tRNA's into 32 well-resolved spots, with the relative abundances ranging from 0.9 to 17% of the total. There were several spots (five to six) resolved which were not quantitated due to their low abundance. All of the tRNA species resolved by this gel system were synthesized at every stage examined, including vegetative growth, different stages of sporulation, and different stages of outgrowth. Quantitation of the separated tRNA's showed that in general the tRNA species were present in approximately the same relative abundances at the different developmental periods. tRNA turnover and compartmentation occurring during sporulation were examined by labeling during vegetative growth followed by the addition of excess phosphate to block further 32P incorporation. The two-dimensional gels of these samples showed the same tRNA's seen during vegetative growth, and they were in approximately the same relative abundances, indicating minimal differences in the rates of turnover of individual tRNA's. Vegetatively labeled samples, chased with excess phosphate into mature spores, also showed all of the tRNA species seen during vegetative growth, but an additional five to six minor spots were also observed. These are hypothesized to arise from the loss of 3'-terminal residues from preexisting tRNA's. Images PMID:115846

  16. Assessment of protoxin composition of Bacillus thuringiensis strains by use of polyacrylamide gel block and mass spectrometry.

    PubMed

    Fu, Zujiao; Sun, Yunjun; Xia, Liqiu; Ding, Xuezhi; Mo, Xiangtao; Li, Xiaohui; Huang, Kexue; Zhang, Youming

    2008-07-01

    Assessment of protoxin composition in Bacillus thuringiensis parasporal crystals is principally hampered by the fact that protoxins in a single strain usually possess high sequence homology. Therefore, new strategies towards the identification of protoxins have been developed. Here, we established a powerful method through embedding solubilized protoxins in a polyacrylamide gel block coupled to liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis of in-gel-generated peptides for protoxin identification. Our model study revealed that four protoxins (Cry1Aa, Cry1Ab, Cry1Ac and Cry2Aa) and six protoxins (Cry4Aa, Cry4Ba, Cry10Aa, Cry11Aa, Cyt1Aa, and Cyt2Ba) could be rapidly identified from B. thuringiensis subsp. kurstaki HD1 and subsp. israelensis 4Q2-72, respectively. The experimental results indicated that our method is a straightforward tool for analyzing protoxin expression profile in B. thuringiensis strains. Given its technical simplicity and sensitivity, our method might facilitate the present screening program for B. thuringiensis strains with new insecticidal properties. PMID:18463863

  17. Quantification of AAV particle titers by infrared fluorescence scanning of coomassie-stained sodium dodecyl sulfate-polyacrylamide gels.

    PubMed

    Kohlbrenner, Erik; Henckaerts, Els; Rapti, Kleopatra; Gordon, Ronald E; Linden, R Michael; Hajjar, Roger J; Weber, Thomas

    2012-06-01

    Adeno-associated virus (AAV)-based vectors have gained increasing attention as gene delivery vehicles in basic and preclinical studies as well as in human gene therapy trials. Especially for the latter two-for both safety and therapeutic efficacy reasons-a detailed characterization of all relevant parameters of the vector preparation is essential. Two important parameters that are routinely used to analyze recombinant AAV vectors are (1) the titer of viral particles containing a (recombinant) viral genome and (2) the purity of the vector preparation, most commonly assessed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) followed by silver staining. An important, third parameter, the titer of total viral particles, that is, the combined titer of both genome-containing and empty viral capsids, is rarely determined. Here, we describe a simple and inexpensive method that allows the simultaneous assessment of both vector purity and the determination of the total viral particle titer. This method, which was validated by comparison with established methods to determine viral particle titers, is based on the fact that Coomassie Brilliant Blue, when bound to proteins, fluoresces in the infrared spectrum. Viral samples are separated by SDS-PAGE followed by Coomassie Brilliant Blue staining and gel analysis with an infrared laser-scanning device. In combination with a protein standard, our method allows the rapid and accurate determination of viral particle titers simultaneously with the assessment of vector purity. PMID:22816378

  18. A modified Coomassie blue staining of proteins in polyacrylamide gels with Bismark brown R.

    PubMed

    Choi, J K; Yoon, S H; Hong, H Y; Choi, D K; Yoo, G S

    1996-04-01

    A rapid and sensitive protein staining method employing a mixed dye technique has been developed. Solutions of Coomassie brilliant blue R-250 (CB, 0.2%) and Bismark brown R (BBR, 0.05%) were mixed in the volume ratio of 1:0.75 for staining (final molar ratio, 4.5:1). In this staining, BBR inhibits the binding of CB to gel matrix by forming ion-pairing complex with excess CB; therefore, it reduces staining/destaining times and also enhances the staining effect of CB on protein band. As a result, the mixed dye staining enables complete staining/destaining within 20/25 min and detection of up to 25 ng of bovine serum albumin. PMID:8619499

  19. Fast and sensitive colloidal coomassie G-250 staining for proteins in polyacrylamide gels.

    PubMed

    Dyballa, Nadine; Metzger, Sabine

    2009-01-01

    Coomassie Brilliant Blue (CBB) is a dye commonly used for the visualization of proteins separated by SDS-PAGE, offering a simple staining procedure and high quantitation. Furthermore, it is completely compatible with mass spectrometric protein identification. But despite these advantages, CBB is regarded to be less sensitive than silver or fluorescence stainings and therefore rarely used for the detection of proteins in analytical gel-based proteomic approaches. Several improvements of the original Coomassie protocol(1) have been made to increase the sensitivity of CBB. Two major modifications were introduced to enhance the detection of low-abundant proteins by converting the dye molecules into colloidal particles: In 1988, Neuhoff and colleagues applied 20% methanol and higher concentrations of ammonium sulfate into the CBB G-250 based staining solution(2), and in 2004 Candiano et al. established Blue Silver using CBB G-250 with phosphoric acid in the presence of ammonium sulfate and methanol(3). Nevertheless, all these modifications just allow a detection of approximately 10 ng protein. A widely fameless protocol for colloidal Coomassie staining was published by Kang et al. in 2002 where they modified Neuhoff's colloidal CBB staining protocol regarding the complexing substances. Instead of ammonium sulfate they used aluminum sulfate and methanol was replaced by the less toxic ethanol(4). The novel aluminum-based staining in Kang's study showed superior sensitivity that detects as low as 1 ng/band (phosphorylase b) with little sensitivity variation depending on proteins. Here, we demonstrate application of Kang's protocol for fast and sensitive colloidal Coomassie staining of proteins in analytical purposes. We will illustrate the quick and easy protocol using two-dimensional gels routinely performed in our working group. PMID:19684561

  20. Highly sensitive method for specific, brief, and economical detection of glycoproteins in sodium dodecyl sulfate-polyacrylamide gel electrophoresis by the synthesis of a new hydrazide derivative.

    PubMed

    Cong, Weitao; Zhou, Ayi; Liu, Zhiguo; Shen, Jiayi; Zhou, Xuan; Ye, Weijian; Zhu, Zhongxin; Zhu, Xinliang; Lin, Jianjun; Jin, Litai

    2015-02-01

    A new hydrazide derivative was synthesized and used for the first time as a specific, brief, and economical probe to selectively visualize glycoproteins in 1-D and 2-D sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) with high sensitivity. The detection limit of the newly developed staining method is 2- and 4-fold higher than that of the widely used Pro-Q Emerald 300 and 488 stains, respectively. PMID:25565298

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

    PubMed Central

    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. Images PMID:1706732

  2. Use of polyacrylamide gel moving boundary electrophoresis to enable low-power protein analysis in a compact microdevice.

    PubMed

    Duncombe, Todd A; Herr, Amy E

    2012-10-16

    In designing a protein electrophoresis platform composed of a single-inlet, single-outlet microchannel powered solely by voltage control (no pumps, values, injectors), we adapted the original protein electrophoresis format-moving boundary electrophoresis (MBE)-to a high-performance, compact microfluidic format. Key to the microfluidic adaptation is minimization of injection dispersion during sample injection. To reduce injection dispersion, we utilize a photopatterned free-solution-polyacrylamide gel (PAG) stacking interface at the head of the MBE microchannel. The nanoporous PAG molecular sieve physically induces a mobility shift that acts to enrich and sharpen protein fronts as proteins enter the microchannel. Various PAG configurations are characterized, with injection dispersion reduced by up to 85%. When employed for analysis of a model protein sample, microfluidic PAG MBE baseline-resolved species in 5 s and in a separation distance of less than 1 mm. PAG MBE thus demonstrates electrophoretic assays with minimal interfacing and sample handling, while maintaining separation performance. Owing to the short separation lengths needed in PAG MBE, we reduced the separation channel length to demonstrate an electrophoretic immunoassay powered with an off-the-shelf 9 V battery. The electrophoretic immunoassay consumed less than 3 ?W of power and was completed in 30 s. To our knowledge, this is the lowest voltage and lowest power electrophoretic protein separation reported. Looking forward, we see the low-power PAG MBE as a basis for highly multiplexed protein separations (mobility shift screening assays) as well as for portable low-power diagnostic assays. PMID:22971048

  3. Characterization of Finnish Borrelia burgdorferi sensu lato isolates by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and with monoclonal antibodies.

    PubMed Central

    Tuomi, J; Rantamki, L K; Tanskanen, R; Junttila, J

    1995-01-01

    Thirty-seven Borrelia burgdorferi strains, isolated in 1992 from Ixodes ricinus in Finland, were investigated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and by immunoblotting and indirect immunofluorescence assay (IFA) with five to nine monoclonal antibodies (MAbs). By SDS-PAGE results and reactivities to MAbs H3TS, J 8.3, I 17.3, and D6, the 37 isolates were assigned to the species B. burgdorferi sensu stricto (n = 7), Borrelia afzelii (n = 17), or Borrelia garinii (n = 13). Twenty more isolates examined only by IFA and with part of the MAbs were distributed as follows: 9 B. burgdorferi sensu stricto and 11 other species. Among 16 of 37 isolates displaying a SDS-PAGE patterns considered typical of that of B. garinii, 3 were negative by the test with MAb D6; the rest were positive. The three MAb D6-negative isolates reacted with MAb J 8.3 but not with MAb I 17.3. It is suggested that these isolates of a previously undescribed type represent atypical B. afzelii strains deficient in the expression of OspB proteins. The misleading species designation by the SDS-PAGE result is described. The IFA results were generally consistent with those obtained by immunoblotting. The exception was for 3 of 29 isolates that were positive with MAb H5332 by immunoblotting but that were IFA negative. In the present material of 57 strains, all 16 B. burgdorferi sensu stricto isolates originated from the Aland Islands. B. afzelii and B. garinii were isolated from all three regions where ticks were collected. The distributive difference seems to offer a basis for comparative clinico-epidemiological studies of Lyme borreliosis. PMID:7559935

  4. Comparative two-dimensional polyacrylamide gel electrophoresis of the salivary proteome of children with autism spectrum disorder

    PubMed Central

    Ngounou Wetie, Armand G; Wormwood, Kelly L; Charette, Laci; Ryan, Jeanne P; Woods, Alisa G; Darie, Costel C

    2015-01-01

    In the last decades, prevalence of autism spectrum disorder (ASD) has been on the rise. However, clear aetiology is still elusive and improvements in early diagnosis are needed. To uncover possible biomarkers present in ASD, we used two-dimensional polyacrylamide gel electrophoresis and nanoliquid chromatography-tandem mass spectrometry (nanoLC-MS/MS), to compare salivary proteome profiling of children with ASD and controls. A total of 889 spots were compared and only those spots with a fold change ≥1.7 and a P-value <0.05 or a fold change of ≥3.0 between ASD cases and controls were analysed by nanoLC-MS/MS. Alpha-amylase, CREB-binding protein, p532, Transferrin, Zn alpha2 glycoprotein, Zymogen granule protein 16, cystatin D and plasminogen were down-regulated in ASD. Increased expression of proto-oncogene Frequently rearranged in advanced T-cell lymphomas 1 (FRAT1), Kinesin family member 14, Integrin alpha6 subunit, growth hormone regulated TBC protein 1, parotid secretory protein, Prolactin-inducible protein precursor, Mucin-16, Ca binding protein migration inhibitory factor-related protein 14 (MRP14) was observed in individuals with ASD. Many of the identified proteins have previously been linked to ASD or were proposed as risk factors of ASD at the genetic level. Some others are involved in pathological pathways implicated in ASD causality such as oxidative stress, lipid and cholesterol metabolism, immune system disturbances and inflammation. These data could contribute to protein signatures for ASD presence, risk and subtypes, and advance understanding of ASD cause as well as provide novel treatment targets for ASD. PMID:26290361

  5. Comparative two-dimensional polyacrylamide gel electrophoresis of the salivary proteome of children with autism spectrum disorder.

    PubMed

    Ngounou Wetie, Armand G; Wormwood, Kelly L; Charette, Laci; Ryan, Jeanne P; Woods, Alisa G; Darie, Costel C

    2015-11-01

    In the last decades, prevalence of autism spectrum disorder (ASD) has been on the rise. However, clear aetiology is still elusive and improvements in early diagnosis are needed. To uncover possible biomarkers present in ASD, we used two-dimensional polyacrylamide gel electrophoresis and nanoliquid chromatography-tandem mass spectrometry (nanoLC-MS/MS), to compare salivary proteome profiling of children with ASD and controls. A total of 889 spots were compared and only those spots with a fold change ≥1.7 and a P-value <0.05 or a fold change of ≥3.0 between ASD cases and controls were analysed by nanoLC-MS/MS. Alpha-amylase, CREB-binding protein, p532, Transferrin, Zn alpha2 glycoprotein, Zymogen granule protein 16, cystatin D and plasminogen were down-regulated in ASD. Increased expression of proto-oncogene Frequently rearranged in advanced T-cell lymphomas 1 (FRAT1), Kinesin family member 14, Integrin alpha6 subunit, growth hormone regulated TBC protein 1, parotid secretory protein, Prolactin-inducible protein precursor, Mucin-16, Ca binding protein migration inhibitory factor-related protein 14 (MRP14) was observed in individuals with ASD. Many of the identified proteins have previously been linked to ASD or were proposed as risk factors of ASD at the genetic level. Some others are involved in pathological pathways implicated in ASD causality such as oxidative stress, lipid and cholesterol metabolism, immune system disturbances and inflammation. These data could contribute to protein signatures for ASD presence, risk and subtypes, and advance understanding of ASD cause as well as provide novel treatment targets for ASD. PMID:26290361

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

    SciTech Connect

    Ono, K; Fujimoto, S; Akagi, Y; Hirokawa, Y; Hayashi, S; Miyazawa, M

    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.

  7. Three-dimensional polyacrylamide gel-based DNA microarray method effectively identifies UDP-glucuronosyltransferase 1A1 gene polymorphisms for the correct diagnosis of Gilbert's syndrome

    PubMed Central

    SONG, JINYUN; SUN, MEI; LI, JIAYAN; ZHOU, DONGRUI; WU, XUPING

    2016-01-01

    Gilbert's syndrome is a mild genetic liver disorder characterized by unconjugated hyperbilirubinemia due to defects in the UDP-glucuronosyltransferase 1A1 (UGT1A1) gene. The T-3279G mutation in the phenobarbital responsive enhancer module (PBREM), the TA-insertion in the TATA box, creating the A(TA)7TAA motif instead of A(TA)6TAA and the G211A mutation in coding exon 1, particularly in Asian populations, of the human UGT1A1 gene are the three common genotypes found in patients with Gilbert's syndrome. Different approaches for detecting the T-3279G, A(TA)6/7TAA and G211A mutations of the UGT1A1 gene have been described. In this study, to the best of our knowledge, we established a three-dimensional polyacrylamide gel-based DNA microarray method for the first time, in order to study UGT1A1 gene polymorphisms. This method, based on a step-by-step three-dimensional polyacrylamide gel-based DNA microarray protocol, successfully identified all possible genotypes of T-3279G, A(TA)6/7TAA and G211A in 20 patients with hyperbilirubinemia. In addition, sequencing was performed to confirm these results. The data from the current study demonstrate that the three-dimensional polyacrylamide gel microarray method has the potential to be applied as a useful, reliable and cost-effective tool to detect the T-3279G, the A(TA)6/7TAA and the G211A mutations of the UGT1A1 gene in patients with hyperbilirubinemia and thereby aid in the diagnosis of Gilbert's syndrome. PMID:26781906

  8. Fluid diversion and sweep improvement with chemical gels in oil recovery processes. [Four types of gels: resorcinol-formaldehyde; colloidal silica; Cr sup 3+ (chloride)-xanthan; and Cr sup 3+ (acetate)-polyacrylamide

    SciTech Connect

    Seright, R.S.; Martin, F.D.

    1992-09-01

    The objectives of this project were to identify the mechanisms by which gel treatments divert fluids in reservoirs and to establish where and how gel treatments are best applied. Several different types of gelants were examined, including polymer-based gelants, a monomer-based gelant, and a colloidal-silica gelant. This research was directed at gel applications in water injection wells, in production wells, and in high-pressure gas floods. The work examined how the flow properties of gels and gelling agents are influenced by permeability, lithology, and wettability. Other goals included determining the proper placement of gelants, the stability of in-place gels, and the types of gels required for the various oil recovery processes and for different scales of reservoir heterogeneity. During this three-year project, a number of theoretical analyses were performed to determine where gel treatments are expected to work best and where they are not expected to be effective. The most important, predictions from these analyses are presented. Undoubtedly, some of these predictions will be controversial. However, they do provide a starting point in establishing guidelines for the selection of field candidates for gel treatments. A logical next step is to seek field data that either confirm or contradict these predictions. The experimental work focused on four types of gels: (1) resorcinol-formaldehyde, (2) colloidal silica, (3) Cr{sup 3+}(chloride)-xanthan, and (4) Cr{sup 3+}(acetate)-polyacrylamide. All experiments were performed at 41{degrees}C.

  9. Coloration of silver-stained protein bands in polyacrylamide gels is caused by light scattering from silver grains of characteristic sizes.

    PubMed Central

    Merril, C R; Bisher, M E; Harrington, M; Steven, A C

    1988-01-01

    This study investigates the physical basis of color effects in the detection of proteins in polyacrylamide gels by silver staining. Specifically, the hypothesis that different colors may correlate with the development of silver grains of characteristic sizes was investigated by electron microscopy. Protein bands that stained brown, yellow, and blue were excised from stained gels and prepared for electron microscopy by thin-sectioning. In each case, the size distributions of globular silver grains were determined directly from the electron micrographs. We found that blue bands have larger silver grains (with diameters of 40-100 nm) than yellow (21-39 nm) or brown bands (17-35 nm). On the basis of these and other observations, a general mechanism is proposed whereby chemical specificity of electrophoretically separated proteins is expressed in color-specific silver staining. Images PMID:2448776

  10. Sensitive silver staining of protein in sodium dodecyl sulfate-polyacrylamide gels using an azo dye, calconcarboxylic acid, as a silver-ion sensitizer.

    PubMed

    Jin, Li-Tai; Hwang, Sun-Young; Yoo, Gyurng-Soo; Choi, Jung-Kap

    2004-08-01

    A highly sensitive silver staining method for detecting proteins in sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) was developed. It is based on the silver nitrate staining method but also employs an azo dye, calconcarboxylic acid (NN), as a silver-ion sensitizer. It increases silver binding on protein bands or spots by the formation of a silver-dye complex and also increases the reducing power of silver ions to metallic silver by NN itself with formaldehyde. After a 2 h gel fixing step, the protocol including sensitization, silver-ion impregnation, and reduction steps can be completed in 1 h. The sensitivity is superior to that of silver stain with glutardialdehyde as a silver-ion sensitizer. The detection limit of NN-silver stain is 0.05-0.2 ng protein. Considering the high sensitivity without using glutardialdehyde, the NN-silver stain would be useful for routine silver staining of proteins. PMID:15300767

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

    NASA Astrophysics Data System (ADS)

    Claverie, Fanny; Pécheyran, Christophe; Mounicou, Sandra; Ballihaut, Guillaume; Fernandez, Beatriz; Alexis, Joël; Lobinski, Ryszard; Donard, Olivier F. X.

    2009-07-01

    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. Pécheyran, 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 µm. 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 µm 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 (Δ T~ 25 ± 5 K). This suggests that the cohesion forces between the thin particles composing these large aggregates were weak enough to have negligible impact on the ICPMS detection.

  12. Filipin staining of lipoproteins in polyacrylamide gels: sensitivity and photobleaching of the fluorophore and its use in a double staining method.

    PubMed

    Smejkal, G B; Hoff, H F

    1994-07-01

    We have developed a double staining procedure in which polyacrylamide gels are first stained with filipin to identify lipoproteins, and then with Coomassie Brilliant Blue (CBB) to identify proteins. Filipin staining when performed at 37 degrees C is both more rapid and more sensitive than previously published procedures. After only 5 min, 20 ng of low density lipoprotein (LDL) unesterified cholesterol/mm3 of band volume could be detected, and after 12 h, sensitivity reached 0.8 ng/mm3. A semilogarithmic relationship was found between the amount of LDL unesterified cholesterol applied and filipin fluorescence. Although rapid photobleaching of the fluorophore occurred during UV transillumination of these gels, such photobleaching actually resulted in maximizing of the signal:noise ratio, resulting in better definition of bands. Treatment of gels with filipin had no deleterious effects on the subsequent staining with CBB. This dual staining procedure should prove useful for studies in which both lipoproteins and proteins in plasma need to be documented in the same gel. PMID:7529172

  13. Fluorographic detection of tritiated glycopeptides and oligosaccharides separated on polyacrylamide gels: analysis of glycans from Dictyostelium discoideum glycoproteins

    SciTech Connect

    Prem Das, O.; Henderson, E.J.

    1986-11-01

    Previous workers have shown that oligosaccharides and glycopeptides can be separated by electrophoresis in buffers containing borate ions. However, normal fluorography of tritium-labeled structures cannot be performed because the glycans are soluble and can diffuse during equilibration with scintillants. This problem has been circumvented by equilibration of the gel with 2,5-diphenyloxazole (PPO) prior to electrophoresis. The presence of PPO in the gel during electrophoresis does not alter mobility of the glycopeptides and oligosaccharides. After electrophoresis, the gel is simply dried and fluorography performed. This allows sensitive and precise comparisons of labeled samples in parallel lanes of a slab gel and, since mobilities are highly reproducible, between different gels. The procedure is preparative in that after fluorography the gel bands can be quantitatively eluted for further study, without any apparent modification by the procedure. In this report, the procedure is illustrated by fractionation of both neutral and anionic glycopeptides produced by the cellular slime mold Dictyostelium discoideum.

  14. Highly sensitive and simple fluorescence staining of proteins in sodium dodecyl sulfate-polyacrylamide-based gels by using hydrophobic tail-mediated enhancement of fluorescein luminescence.

    PubMed

    Kang, Chulhun; Kim, Hyun Jung; Kang, Donghoon; Jung, Duk Young; Suh, Myungkoo

    2003-10-01

    Fluorescein has an extremely low luminescence intensity in acidic aqueous media. However, when it was bound to proteins, subsequent increase of luminescence intensity took place. Furthermore, when a hydrophobic tail, such as aliphatic hydrocarbons, was introduced to fluorescein, more dramatic increase of luminescence intensity was observed upon binding to proteins. In the present study, by utilizing this luminescence enhancement, three hydrophobic fluorescein dyes (5-dodecanoyl amino fluorescein, 5-hexadecanoyl amino fluorescein, and 5-octadecanoyl amino fluorescein) were examined as noncovalent fluorescent stains of protein bands in sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Effective incorporation of the dyes to proteins in gels was accomplished either simply by adding dyes at the protein fixation step, or by treating gels with a staining solution after the fixation. The sensitivity of this staining method using the fluorescein derivatives was approximately 1 ng/band for most proteins. For some cases, protein bands containing as low as 0.1 ng were successfully visualized. In addition, the detection sensitivity showed much less protein-to-protein variation than silver staining. This new staining method was also successfully applied to two-dimensional electrophoresis of rat brain proteins. Its overall sensitivity was comparable to that of silver staining. PMID:14595675

  15. Ten-Year Safety with Polyacrylamide Gel Used to Correct Facial Lipoatrophy in HIV-Infected Patients.

    PubMed

    Negredo, Eugenia; Puig, Jordi; Ornelas, Arelly; Echeverra, Patricia; Bonjoch, Anna; Estany, Carla; Higueras, Carmen; Gonzalez-Mestre, Vicente; Clotet, Bonaventura

    2015-08-01

    Long-term results (>5 years) for synthetic substances used to repair facial lipoatrophy have not been published. We performed a cross-sectional study to evaluate the 10-year safety of polyacrylamide hydrogel (Aquamid) among the 751 patients from our unit who received facial infiltrations at least 10 years ago. Epidemiological and clinical data such as complications and patient satisfaction were collected. We also identified those patients who presented a facial infection at any time after infiltration. A total of 104 patients had received Aquamid at least 10 years ago. Before infiltrations, 24.0%, 41.3%, and 34.7% presented very severe, severe, and moderate facial lipoatrophy, respectively. After a mean (SD) of 10.3 (0.5) years since the infiltrations, 19.2%, 47.7%, and 31.7% of patients reported moderate, mild, and no signs of facial lipoatrophy. The values reported by physicians for the same categories were 1.9%, 10.6%, and 87.5%. Indurations were detected in 6.7% of patients and nodules in 3.8%. Five patients (4.8%) had a local infection. A further 15 patients with a shorter follow-up (less than 10 years) presented local infections (overall incidence considering the 751 patients who received infiltrations of Aquamid, 2.7%); the product had to be withdrawn in three cases. The majority of patients were highly satisfied (74.8%) or satisfied (23.4%) with the cosmetic results; among patients with severe or very severe lipoatrophy at baseline, 31.4% were satisfied and 65.7% were highly satisfied. Infiltrations with polyacrylamide hydrogel (Aquamid) are a safe strategy for the treatment of facial lipoatrophy in the long term. The rate of severe complications was low, and patient satisfaction with the cosmetic results was high. However, facial infections may appear in the long term. Therefore, HIV-infected patients who received synthetic substances should be carefully monitored over time. PMID:25858612

  16. Data on final calcium concentration in native gel reagents determined accurately through inductively coupled plasma measurements

    PubMed Central

    Viviano, Jeffrey; Wu, Hao; Krishnan, Anuradha; Ramanujachary, Kandalam; Venkataraman, Venkat

    2016-01-01

    In this article we present data on the concentration of calcium as determined by Inductively Coupled Plasma (ICP) measurements. Calcium was estimated in the reagents used for native gel electrophoresis of Neuronal Calcium Sensor (NCS) proteins. NCS proteins exhibit calcium-dependent mobility shift in native gels. The sensitivity of this shift to calcium necessitated a precise determination of calcium concentrations in all reagents used. We determined the calcium concentrations in different components used along with the samples in the native gel experiments. These were: 20mM Tris pH 7.5, loading dye and running buffer, with distilled water as reference. Calcium determinations were through ICP measurements. It was found that the running buffer contained calcium (244nM) over the blank.

  17. Data on final calcium concentration in native gel reagents determined accurately through inductively coupled plasma measurements.

    PubMed

    Viviano, Jeffrey; Wu, Hao; Krishnan, Anuradha; Ramanujachary, Kandalam; Venkataraman, Venkat

    2016-03-01

    In this article we present data on the concentration of calcium as determined by Inductively Coupled Plasma (ICP) measurements. Calcium was estimated in the reagents used for native gel electrophoresis of Neuronal Calcium Sensor (NCS) proteins. NCS proteins exhibit calcium-dependent mobility shift in native gels. The sensitivity of this shift to calcium necessitated a precise determination of calcium concentrations in all reagents used. We determined the calcium concentrations in different components used along with the samples in the native gel experiments. These were: 20 mM Tris pH 7.5, loading dye and running buffer, with distilled water as reference. Calcium determinations were through ICP measurements. It was found that the running buffer contained calcium (244 nM) over the blank. PMID:26937454

  18. Preliminary studies on the role and reactions of tetrakis(hydroxymethyl)phosphonium chloride in polyacrylamide gel dosimeters

    NASA Astrophysics Data System (ADS)

    Sedaghat, Mahbod; Bujold, Rachel; Lepage, Martin

    2012-10-01

    A major source of dosimetric inaccuracy in normoxic polymer gel dosimeters is local variations in the concentration of oxygen scavenger. Currently, a phosphorus compound, tetrakis(hydroxymethyl)phosphonium chloride (THPC), is the oxygen scavenger of choice in most polymer gel dosimetry studies. Reactions of THPC in a gel dosimeter are not limited to oxygen. It can possibly be consumed in reacting with gelling agent, water free-radicals and polymer radicals before, during and after irradiation, hence affecting the dose response of the dosimeter in several ways. These reactions are not fully known or understood. It is our hypothesis that THPC not only scavenges radical species but also modifies the morphology of the gelatin network and of the polymer, possibly by intervening in the polymerization of monomers. These hypotheses are investigated in an anoxic acrylamide-based gel dosimeter. Scanning electron microscopy results indicate gelatin pores decreasing from 70 to 40 m and a very different radiation-induced polymer structure in samples containing THPC; Fourier-transform Raman spectroscopy shows a two-fold reduction in the dose constants of monomer consumption; however, a significant change in the relative dose constants of monomer consumption as a function of dose could not be detected.

  19. Polyacrylamide gel substrates that simulate the mechanical stiffness of normal and malignant neuronal tissues increase protoporphyin IX synthesis in glioma cells.

    PubMed

    Niu, Carolyn J; Fisher, Carl; Scheffler, Kira; Wan, Rachel; Maleki, Hoda; Liu, Haijiao; Sun, Yu; A Simmons, Craig; Birngruber, Reginald; Lilge, Lothar

    2015-09-01

    Protoporphyrin IX (PPIX) produced following the administration of exogenous 5d-aminolevulinic acid is clinically approved for photodynamic therapy and fluorescence-guided resection in various jurisdictions around the world. For both applications, quantification of PPIX forms the basis for accurate therapeutic dose calculation and identification of malignant tissues for resection. While it is well established that the PPIX synthesis and accumulation rates are subject to the cells biochemical microenvironment, the effect of the physical microenvironment, such as matrix stiffness, has received little attention to date. Here we studied the proliferation rate and PPIX synthesis and accumulation in two glioma cell lines U373 and U118 cultured under five different substrate conditions, including the conventional tissue culture plastic and polyacrylamide gels that simulated tissue stiffness of normal brain (1 kPa) and glioblastoma tumors (12 kPa). We found that the proliferation rate increased with substrate stiffness for both cell lines, but not in a linear fashion. PPIX concentration was significantly higher in cells cultured on tissue-simulating gels than on the much stiffer tissue culture plastic for both cell lines. These findings, albeit preliminary, suggest that the physical microenvironment might be an important determinant of tumor aggressiveness and PPIX synthesis in glioma cells. PMID:26405823

  20. Polyacrylamide gel substrates that simulate the mechanical stiffness of normal and malignant neuronal tissues increase protoporphyin IX synthesis in glioma cells

    NASA Astrophysics Data System (ADS)

    Niu, Carolyn J.; Fisher, Carl; Scheffler, Kira; Wan, Rachel; Maleki, Hoda; Liu, Haijiao; Sun, Yu; Simmons, Craig A.; Birngruber, Reginald; Lilge, Lothar

    2015-09-01

    Protoporphyrin IX (PPIX) produced following the administration of exogenous 5d-aminolevulinic acid is clinically approved for photodynamic therapy and fluorescence-guided resection in various jurisdictions around the world. For both applications, quantification of PPIX forms the basis for accurate therapeutic dose calculation and identification of malignant tissues for resection. While it is well established that the PPIX synthesis and accumulation rates are subject to the cell's biochemical microenvironment, the effect of the physical microenvironment, such as matrix stiffness, has received little attention to date. Here we studied the proliferation rate and PPIX synthesis and accumulation in two glioma cell lines U373 and U118 cultured under five different substrate conditions, including the conventional tissue culture plastic and polyacrylamide gels that simulated tissue stiffness of normal brain (1kPa) and glioblastoma tumors (12kPa). We found that the proliferation rate increased with substrate stiffness for both cell lines, but not in a linear fashion. PPIX concentration was significantly higher in cells cultured on tissue-simulating gels than on the much stiffer tissue culture plastic for both cell lines. These findings, albeit preliminary, suggest that the physical microenvironment might be an important determinant of tumor aggressiveness and PPIX synthesis in glioma cells.

  1. Electrophoretic analysis of proteinases in sodium dodecyl sulfate-polyacrylamide gels containing copolymerized radiolabeled protein substrates: Application to proenkephalin processing enzymes

    SciTech Connect

    Irvine, J.W.; Roberts, S.F.; Lindberg, I. )

    1990-10-01

    A novel method is described for the zymographic analysis of proteinases in sodium dodecyl sulfate-polyacrylamide gels containing copolymerized radiolabeled protein substrates such as ({sup 35}S)methionine-labeled proenkephalin or {sup 125}I-labeled proinsulin. After electrophoresis the enzyme is reactivated and cleaves the radiolabeled in situ substrate into smaller peptides. These small peptides are able to diffuse out of the gel, leaving clear areas against a dark background when visualized by autoradiography. The technique can be used to detect as little as 200 fg of trypsin using only 50 ng (1.25 microCi) of ({sup 35}S)proenkephalin. Soluble- and membrane-bound adrenal trypsin-like enzyme were isolated from bovine adrenal chromaffin granules. Both proteinases cleaved ({sup 35}S)methionine-labeled proenkephalin but not {sup 125}I-labeled proinsulin. Moreover, both had a Mr of approximately 30,000. The potential of this technique for general use is discussed. An additional method using the synthetic fluorogenic substrate t-butoxycarbonyl Glu-Lys-Lys aminomethylcoumarin is also described.

  2. System and method of infrared matrix-assisted laser desorption/ionization mass spectrometry in polyacrylamide gels

    DOEpatents

    Haglund, Jr., Richard F.; Ermer, David R.; Baltz-Knorr, Michelle Lee

    2004-11-30

    A system and method for desorption and ionization of analytes in an ablation medium. In one embodiment, the method includes the steps of preparing a sample having analytes in a medium including at least one component, freezing the sample at a sufficiently low temperature so that at least part of the sample has a phase transition, and irradiating the frozen sample with short-pulse radiation to cause medium ablation and desorption and ionization of the analytes. The method further includes the steps of selecting a resonant vibrational mode of at least one component of the medium and selecting an energy source tuned to emit radiation substantially at the wavelength of the selected resonant vibrational mode. The medium is an electrophoresis medium having polyacrylamide. In one embodiment, the energy source is a laser, where the laser can be a free electron laser tunable to generate short-pulse radiation. Alternatively, the laser can be a solid state laser tunable to generate short-pulse radiation. The laser can emit light at various ranges of wavelength.

  3. Large-scale muLC-MS/MS for silver- and Coomassie blue-stained polyacrylamide gels.

    PubMed

    Zhu, Wenhong; Venable, John; Giometti, Carol S; Khare, Tripti; Tollaksen, Sandra; Ahrendt, Angela J; Yates, John R

    2005-12-01

    2-DE combined with LC-MS/MS has become a routine, reliable protein separation and identification technology for proteome analysis. The demand for large-scale protein identifications after 2-DE separation requires a sensitive and high-throughput LC-MS/MS method. In this report, a simple, splitless, fully automated capillary LC-MS/MS system was described for the large-scale identification of proteins from gels stained with either silver or CBB. The gel samples were digested and peptides were extracted using an in-gel digestion workstation. The peptides were automatically introduced into a capillary column by an autosampler connected to an HPLC pump. A nanoLC pump was then used to deliver the gradient and elute the peptides from the capillary column directly into an LCQ IT mass spectrometer. Neither a peptide trapping setting nor a flow split is needed in this simple setup. The collected MS/MS spectra were then automatically searched by SEQUEST, and filtered and organized by DTASelect. Hundreds of silver-stained or CBB-stained Shewanella oneidensis, Geobacter sulfurreducens, and Geobacter metallireducens proteins separated by denaturing or nondenaturing 2-DE were digested and routinely analyzed using this fully automated muLC-MS/MS system. High peptide hits and sequence coverage were achieved for most CBB-stained gel spots. About 75% of the spots were found to contain multiple proteins. Although silver staining is not commonly thought to be optimal for MS analysis, protein identifications were successfully obtained from silver-stained 2-DE spots detected using methods with and without formaldehyde for protein fixation. PMID:16315175

  4. Detection of glycoproteins in polyacrylamide gels using Pro-Q Emerald 300 dye, a fluorescent periodate Schiff-base stain.

    PubMed

    Mehta-D'souza, Padmaja

    2012-01-01

    Pro-Q Emerald 300 glycoprotein stain generates a bright-green fluorescent signal upon reacting with periodic acid-oxidized carbohydrate groups on proteins. With this dye, it is possible to detect proteins directly in the gel without the need to transfer them to a membrane. This dye is more sensitive than the standard periodic acid Schiff's base which uses acidic fuchsin dye. PMID:22585521

  5. Fluorescence detection of proteins in sodium dodecyl sulfate-polyacrylamide gels using environmentally benign, nonfixative, saline solution.

    PubMed

    Steinberg, T H; Lauber, W M; Berggren, K; Kemper, C; Yue, S; Patton, W F

    2000-02-01

    SYPRO Tangerine stain is an environmentally benign alternative to conventional protein stains that does not require solvents such as methanol or acetic acid for effective protein visualization. Instead, proteins can be stained in a wide range of buffers, including phosphate-buffered saline or simply 150 mM NaCl using an easy, one-step procedure that does not require destaining. Stained proteins can be excited by ultraviolet light of about 300 nm or with visible light of about 490 nm. The fluorescence emission maximum of the dye is approximately 640 nm. Noncovalent binding of SYPRO Tangerine dye is mediated by sodium dodecyl sulfate (SDS) and to a lesser extent by hydrophobic amino acid residues in proteins. This is in stark contrast to acidic silver nitrate staining, which interacts predominantly with lysine residues or Coomassie Blue R, which in turn interacts primarily with arginine and lysine residues. The sensitivity of SYPRO Tangerine stain is similar to that of the SYPRO Red and SYPRO Orange stains - about 4-10 ng per protein band. This detection sensitivity is comparable to colloidal Coomassie blue staining and rapid silver staining procedures. Since proteins stained with SYPRO Tangerine dye are not fixed, they can easily be eluted from gels or utilized in zymographic assays, provided that SDS does not inactivate the protein of interest. This is demonstrated with in-gel detection of rabbit liver esterase activity using alpha-naphthyl acetate and Fast Blue BB dye as well as Escherichia coli beta-glucuronidase activity using ELF-97 beta-D-glucuronide. The dye is also suitable for staining proteins in gels prior to their transfer to membranes by electroblotting. Gentle staining conditions are expected to improve protein recovery after electroelution and to reduce the potential for artifactual protein modifications such as the alkylation of lysine and esterification of glutamate residues, which complicate interpretation of peptide fragment profiles generated by mass spectrometry. PMID:10726749

  6. Ion-enhanced fluorescence staining of sodium dodecyl sulfate-polyacrylamide gels using bis(8-p-toluidino-1-naphthalenesulfonate).

    PubMed

    Horowitz, P M; Bowman, S

    1987-09-01

    A method for the sensitive fluorescent staining of sodium dodecyl sulfate (SDS) gels that extends the applicability and sensitivity of existing procedures has been developed. SDS-protein complexes are able to bind the noncovalent hydrophobic probe, bis(8-p-toluidino-1-naphthalenesulfonate) (bisANS) with an increase in quantum yield that is considerably larger than that observed with the commonly used monomeric form, 1-anilinonaphthalene-8-sulfonic acid (1,8-ANS). The quantum yield of bisANS bound to the SDS-protein complex is greatly enhanced by incubation with one of a number of cations including potassium and barium. The use of bisANS with metal ion enhancements provides a method for staining SDS gels that can be more sensitive than commonly used methods based on the binding of Coomassie blue, and provides a simple and rapid method for the detection and quantitation of proteins. The use of metal ion enhancements also greatly increases the sensitivity of staining methods based on the use of 1,8-ANS. The present method is much more sensitive than previous noncovalent, flourescent, postelectrophoresis stains, but it retains their considerable advantages of speed, simplicity, and the ability to perform secondary procedures on the separated materials. PMID:3425913

  7. In situ alkylation with acrylamide for identification of cysteinyl residues in proteins during one- and two-dimensional sodium dodecyl sulphate-polyacrylamide gel electrophoresis.

    PubMed

    Mineki, Reiko; Taka, Hikari; Fujimura, Tsutomu; Kikkawa, Mika; Shindo, Noriko; Murayama, Kimie

    2002-12-01

    Cysteinyl residues in proteins were alkylated with acrylamide during sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) to yield a thioether derivative, cys-S-beta-propionamide (PAM cys). The process was termed in situ alkylation with acrylamide. Using this method, the recovery of PAM-cys peptides from bovine serum albumin (BSA) was 88.6% at 10 picomol in one-dimensional (1-D) SDS-PAGE and 97.1% at 50 picomol in two-dimensional (2-D) SDS-PAGE. The coverage of tryptic peptide of BSA in 1-D and 2-D SDS-PAGE was 83.7% and 81.1%, respectively. The advantages of in situ alkylation with acrylamide were the following: (i) cysteinyl peptides were effectively derived in a single PAM cys and then proteins were precisely identified using databases; (ii) marked reduction of salts compared with post alkylation, e.g., using carboxymethylamide (CAM), resulting in higher signal intensity and wider coverage of cysteinyl peptides from PAM cys, compared with those of CAM derivatives, in mass spectrometry peptide mapping; and (iii) shorter duration by excluding the processes of post alkylation and desalting before peptide mapping. PMID:12469337

  8. A rapid method of species identification of wild chironomids (Diptera: Chironomidae) via electrophoresis of hemoglobin proteins in sodium dodecyl sulfate polyacrylamide gel (SDS-PAGE).

    PubMed

    Oh, J T; Epler, J H; Bentivegna, C S

    2014-10-01

    Studying aquatic benthic macroinvertebrates (BMIs) in the field requires accurate taxonomic identification, which can be difficult and time consuming. Conventionally, head capsule morphology has been used to identify wild larvae of Chironomidae. However, due to the number of species and possible damage and/or deformity of their head capsules, another supporting approach for identification is needed. Here, we provide hemoglobin (Hb) protein in hemolymph of chironomids as a new biomarker that may help resolve some of the ambiguities and difficulties encountered during taxonomic identification. Chironomids collected from two locations in Maine and New Jersey, USA were identified to the genus level and in some cases to the species-level using head capsule and body morphologies. The head capsule for a particular individual was then associated with a corresponding Hb protein profile generated from sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). Distinct Hb profiles were observed from one group (Thienemannimyia) and four genera (Chironomus, Cricotopus, Dicrotendipes, and Glyptotendipes) of chironomids. Several species were polymorphic, having more than one Hb profile and/or having bands of the same size as those of other species. However, major bands and the combination of bands could distinguish individuals at the genus and sometimes species-level. Overall, this study showed that Hb profiles can be used in combination with head capsule morphology to identify wild chironomids. PMID:24923437

  9. Variation and Genomic Localization of Genes Encoding DROSOPHILA MELANOGASTER Male Accessory Gland Proteins Separated by Sodium Dodecyl Sulfate-Polyacrylamide Gel Electrophoresis

    PubMed Central

    Whalen, Michael; Wilson, Thomas G.

    1986-01-01

    Accessory gland proteins from Drosophila melanogaster males have been separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis into nine major bands. When individual males from 175 strains were examined, considerable polymorphism for nearly one-half of the major protein bands was seen, including null alleles for three bands. Variation was observed not only among long-established laboratory strains but also among stocks recently derived from natural populations. There was little difference in the amount of variation between P and M strains, indicating that P element mutagenesis is not a factor producing the variation. Codominant expression of variants for each of five bands was found in heterozygotes, suggesting structural gene variation and not posttranslational modification variation. Stocks carrying electrophoretic variants of four of the major proteins were used to map the presumed structural genes for these proteins; the loci were found to be dispersed on the second chromosome. Since males homozygous for variant proteins were fertile, the polymorphism seems to have little immediate effect on successful sperm transfer. We propose that a high degree of polymorphism can be tolerated because these proteins play a nutritive rather than enzymatic role in Drosophila reproduction. PMID:3095182

  10. Structural Studies of Avian Myeloblastosis Virus: Comparison of Polypeptides in Virion and Core Component by Dodecyl Sulfate-Polyacrylamide Gel Electrophoresis

    PubMed Central

    Stromberg, Kurt; Hurley, Nancy E.; Davis, Nancy L.; Rueckert, Roland R.; Fleissner, Erwin

    1974-01-01

    Two different systems of dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) in separate laboratories detected analogous patterns of dye bands in virions of avian myeloblastosis virus (AMV). At least 11 of the dye bands co-migrated with the major polypeptides reported in Rous sarcoma virus. Particles with the morphology of the AMV core component, obtained after exposure of AMV to the nonionic surfactant Sterox SL, contained major polypeptides p12, p27, p60, p64, p91, and p98. The polypeptide p12 has been previously shown to be the major constituent of the inner ribonucleoprotein (RNP) of the AMV core, and has been designated p12(N). Two RNP polypeptides, p64 and p91, co-electrophoresed with purified AMV DNA polymerase and have now been designated p64(P) and p91(P). The polypeptide p27 has been identified as a probable constituent of the core shell, and has accordingly now been designated p27(C). In comparison to virions of AMV, the AMV core component contained a greatly reduced amount of polypeptide p15 and appeared to lack a major polypeptide, p19. Consequently, these polypeptides may be associated either with the exterior of the core shell or the interior of the viral envelope. Glycopeptides were not detected in AMV cores, in agreement with earlier reports that they reside in external projections from the viral envelope. Images PMID:4129794

  11. Coordination mechanism, characterization, and photoluminescence properties of spinel ZnAl2O4 nanoparticles prepared by a modified polyacrylamide gel route

    NASA Astrophysics Data System (ADS)

    Sun, Guangzhuang; Sun, Guangai; Zhong, Mian; Wang, Shifa; Zu, Xiaotao; Xiang, Xia

    2016-03-01

    Single-phase ZnAl2O4 nanoparticles with the spinel structure were successfully synthesized using a modified polyacrylamide gel method according to the atomic ratio of Zn to Al = 1: 1.8. The as-prepared samples were characterized by means of X-ray powder diffraction (XRD), thermogravimetric analysis (TG), differential scanning calorimetry analysis (DSC), field-emission scanning electron microscopy (SEM), Fourier transform infrared spectroscopy (FTIR), and photoluminescence (PL) spectra. XRD patterns show that the pure phase of ZnAl2O4 is obtained after heating the xerogel at 900°C for 5 h in air. The SEM images reveal that the ZnAl2O4 nanoparticles have a narrow particle size distribution and the average particle size is around 45 nm. Photoluminescence (PL) spectra demonstrate the single phase ZnAl2O4 nanoparticles have an emission peak located at 469 nm when excited by 350 nm light. The phase structure, coordination mechanism, and luminescence properties have been discussed on the basis of the experimental results.

  12. Development of methods for the charge-derivatization of peptides in polyacrylamide gels and membranes for their direct analysis using matrix-assisted laser desorption-ionization mass spectrometry

    NASA Astrophysics Data System (ADS)

    Strahler, John R.; Smelyanskiy, Yanina; Lavine, Gary; Allison, John

    1997-12-01

    Approaches are being developed for the interfacing of matrix-assisted laser desorption-ionization (MALDI) mass spectrometry with polyacrylamide gel electrophoresis, in which laser-irradiated samples desorb directly from a gel, or from a membrane on which gel-separated polypeptides have been transferred. Whether one- or two-dimensional electrophoretic separations have been performed, preparations for the MALDI experiment which follow must optimize detection of the analytes present and the generation of structural information. Procedures have been developed for forming charged derivatives of peptides in solution. When subjected to MALDI analysis, these charged derivatives produce ions in some cases where the underivatized peptide would not yield a response. The ions fragment following acceleration and yield informative and simple post-source decay (PSD) spectra. The development of approaches to interfacing this chemistry with MALDI directly from gels and membranes is presented here.

  13. Definition of Mycobacterium tuberculosis culture filtrate proteins by two-dimensional polyacrylamide gel electrophoresis, N-terminal amino acid sequencing, and electrospray mass spectrometry.

    PubMed Central

    Sonnenberg, M G; Belisle, J T

    1997-01-01

    A number of the culture filtrate proteins secreted by Mycobacterium tuberculosis are known to contribute to the immunology of tuberculosis and to possess enzymatic activities associated with pathogenicity. However, a complete analysis of the protein composition of this fraction has been lacking. By using two-dimensional polyacrylamide gel electrophoresis, detailed maps of the culture filtrate proteins of M. tuberculosis H37Rv were generated. In total, 205 protein spots were observed. The coupling of this electrophoretic technique with Western blot analysis allowed the identification and mapping of 32 proteins. Further molecular characterization of abundant proteins within this fraction was achieved by N-terminal amino acid sequencing and liquid chromatography-mass spectrometry. Eighteen proteins were subjected to N-group analysis; of these, only 10 could be sequenced by Edman degradation. Among the most interesting were a novel 52-kDa protein demonstrating significant homology to an alpha-hydroxysteroid dehydrogenase of Eubacterium sp. strain VPI 12708, a 25-kDa protein corresponding to open reading frame 28 of the M. tuberculosis cosmid MTCY1A11, and a 31-kDa protein exhibiting an amino acid sequence identical to that of antigen 85A and 85B. This latter product migrated with an isoelectric point between those of antigen 85A and 85C but did not react with the antibody specific for this complex, suggesting that there is a fourth member of the antigen 85 complex. Novel N-terminal amino acid sequences were obtained for three additional culture filtrate proteins; however, these did not yield significant homology to known protein sequences. A protein cluster of 85 to 88 kDa, recognized by the monoclonal antibodies IT-57 and IT-42 and known to react with sera from a large proportion of tuberculosis patients, was refractory to N-group analysis. Nevertheless, mass spectrometry of peptides obtained from one member of this complex identified it as the M. tuberculosis KatG catalase/peroxidase. Thus, the detailed mapping of M. tuberculosis proteins, combined with state-of-the-art analytical techniques such as mass spectrometry, provides a basis for further analysis and rapid identification of biologically relevant molecules. PMID:9353028

  14. Time-resolved Raman and polyacrylamide gel electrophoresis observations of nucleotide incorporation and misincorporation in RNA within a bacterial RNA polymerase crystal.

    PubMed

    Antonopoulos, Ioanna H; Murayama, Yuko; Warner, Brittany A; Sekine, Shun-Ichi; Yokoyama, Shigeyuki; Carey, Paul R

    2015-01-27

    The bacterial RNA polymerase (RNAP) elongation complex (EC) is highly stable and is able to extend an RNA chain for thousands of nucleotides. Understanding the processive mechanism of nucleotide addition requires detailed structural and temporal data for the EC reaction. Here, a time-resolved Raman spectroscopic analysis is combined with polyacrylamide gel electrophoresis (PAGE) to monitor nucleotide addition in single crystals of the Thermus thermophilus EC (TthEC) RNAP. When the cognate base GTP, labeled with (13)C and (15)N (*GTP), is soaked into crystals of the TthEC, changes in the Raman spectra show evidence of nucleotide incorporation and product formation. The major change is the reduction of *GTP's triphosphate intensity. Nucleotide incorporation is confirmed by PAGE assays. Both Raman and PAGE methods have a time resolution of minutes. There is also Raman spectroscopic evidence of a second population of *GTP in the crystal that does not become covalently linked to the nascent RNA chain. When this population is removed by "soaking out" (placing the crystal in a solution that contains no NTP), there are no perturbations to the Raman difference spectra, indicating that conformational changes are not detected in the EC. In contrast, the misincorporation of the noncognate base, (13)C- and (15)N-labeled UTP (*UTP), gives rise to large spectroscopic changes. As in the GTP experiment, reduction of the triphosphate relative intensity in the Raman soak-in data shows that the incorporation reaction occurs during the first few minutes of our instrumental dead time. This is also confirmed by PAGE analysis. Whereas PAGE data show *GTP converts 100% of the nascent RNA 14mer to 15mer, the noncognate *UTP converts only ?50%. During *UTP soak-in, there is a slow, reversible formation of an ?-helical amide I band in the Raman difference spectra peaking at 40 min. Similar to *GTP soak-in, *UTP soak-in shows Raman spectoscopic evidence of a second noncovalently bound *UTP population in the crystal. Moreover, the second population has a marked effect on the complex's conformational states because removing it by "soaking-out" unreacted *UTP causes large changes in protein and nucleic acid Raman marker bands in the time range of 10-100 min. The conformational changes observed for noncognate *UTP may indicate that the enzyme is preparing for proofreading to excise the misincorporated base. This idea is supported by the PAGE results for *UTP soak-out that show endonuclease activity is occurring. PMID:25584498

  15. Identification of protein complexes of microsomes in rat adipocytes by native gel coupled with LC-ESI-QTOF.

    PubMed

    Ke, Ming; Zhang, Yongqian; Xiong, Yan; Saeed, Yasmeen; Deng, Yulin

    2016-04-22

    The study of the composition of microsome proteins/complexes/interactions in adipocytes provides useful information for researchers related to energy metabolism disorders. The native gel coupled with LC-ESI-QTOF approach was employed here for separating protein complexes. We found a series of proteins functionally clustered in biological processes of protein metabolism, cellular carbohydrate catabolism, response to stimulus and wounding, macromolecular complex subunit organization, positive regulation of molecular function, regulation of programmed cell death and biomolecule transport. According to clustering of proteins' electrophoresis profiles across native gel fractions and bioinformatics data retrieval, protein complexes/interactions involved in protein metabolism, cellular carbohydrate catabolism, macromolecular complex subunit organization and biomolecule transport were identified. Besides, the results also revealed some functional linkages, which may provide useful information for discovering previously unknown interactions. The interaction between SSAO and ALDH2 was verified by co-immunoprecipitation. The native gel combining mass spectrometry approach appeared to be a useful tool for investigating microsome proteins and complexes to complement the traditional electrophoresis approaches. The native gel strategy together with our findings should facilitate future studies of the composition of rat adipocyte microsome protein complexes under different conditions. PMID:26886786

  16. Double staining of coomassie blue-stained polyacrylamide gels by imidazole-sodium dodecyl sulfate-zinc reverse staining: sensitive detection of coomassie blue-undetected proteins.

    PubMed

    Fernandez-Patron, C; Hardy, E; Sosa, A; Seoane, J; Castellanos, L

    1995-01-01

    The sensitivity, simplicity, and relative rapidity of Coomassie blue staining have made this technique the method of choice for routine detection and quantitative analysis of gel electrophoresis-separated protein bands in many applications. To extend the usefulness of this technique, we have developed a new double-staining method for visualizing SDS-PAGE-separated protein bands that were undetected by Coomassie blue staining of the gel. Coomassie blue-stained gels are washed in distilled water (15 min, two times) and then subjected to imidazole-zinc reverse staining. As a result of the method, a homogeneous white-stained background is generated and two types of protein bands can be observed: (a) typical Coomassie blue-stained bands, which appear superposed on larger transparent bands; and (b) reverse-stained (transparent) bands, which were previously undetected by the Coomassie blue staining. The method is rapid, simple, and reproducible and double-staining gels can be kept in distilled water for months without loss of the protein pattern. The overall sensitivity is high (e.g., 1.6 ng for recombinant streptokinase, 47 kDa) over a wide range of protein molecular weights (10 to 100 kDa) and independent of the degree of Coomassie blue destaining of the gel. Furthermore, a mechanism offering a consistent explanation for the role of imidazole, SDS, and zinc in the reverse staining of gels, particularly after Coomassie blue staining is proposed. PMID:7535985

  17. Proteome analysis of human stomach tissue: separation of soluble proteins by two-dimensional polyacrylamide gel electrophoresis and identification by mass spectrometry.

    PubMed

    Ha, Geun Hyoung; Lee, Seung Uook; Kang, Deok Gyeong; Ha, Na-Young; Kim, Soon Hee; Kim, Jina; Bae, Jong Min; Kim, Jae Won; Lee, Chang-Won

    2002-08-01

    Two-dimensional gel electrophoresis (2-DE) maps for human stomach tissue proteins have been prepared by displaying the protein components of the tissue by 2-DE and identifying them using mass spectrometry. This will enable us to present an overview of the proteins expressed in human stomach tissues and lays the basis for subsequent comparative proteome analysis studies with gastric diseases such as gastric cancer. In this study, 2-DE maps of soluble fraction proteins were prepared on two gel images with partially overlapping pH ranges of 4-7 and 6-9. On the gels covering pH 4-7 and pH 6-9, about 900 and 600 protein spots were detected by silver staining, respectively. For protein identification, proteins spots on micropreparative gels stained with colloidal Coomassie Brilliant Blue G-250 were excised, digested in-gel with trypsin, and analyzed by peptide mass fingerprinting with delayed extraction-matrix assisted laser desorption/ionization-mass spectrometry (DE-MALDI-MS). In all, 243 protein spots (168 spots in acidic map and 75 spots in basic map) corresponding to 136 different proteins were identified. Besides these principal maps, overview maps (displayed on pH 3-10 gels) for total homogenate and soluble fraction, are also presented with some identifications mapped on them. Based on the 2-DE maps presented in this study, a 2-DE database for human stomach tissue proteome has been constructed and is available at http://proteome.gsnu.ac.kr/DB/2DPAGE/Stomach/. The 2-DE maps and the database resulting from this study will serve important resources for subsequent proteomic studies for analyzing the normal protein variability in healthy tissues and specific protein variations in diseased tissues. PMID:12210210

  18. A new multiphasic buffer system for sodium dodecyl sulfate-polyacrylamide gel electrophoresis of proteins and peptides with molecular masses 100,000-1000, and their detection with picomolar sensitivity.

    PubMed

    Wiltfang, J; Arold, N; Neuhoff, V

    1991-05-01

    A novel multiphasic buffer system for high resolution sodium dodecyl sulfate-polyacrylamide gel electrophoresis of dansylated and nondansylated proteins/peptides in the relative molecular mass (Mr) range of 100,000-1000 is described. The system, based on Jovin's theory of multiphasic zone electrophoresis, allows complete stacking and destacking of proteins/peptides within the above Mr range. The buffer system uses Bicine and sulfate as trailing and leading ion, respectively, and Bistris and Tris as counter ions in the stacking and separating phase, respectively. Through selection of two different counter ions--the characteristic feature of the present ionic system--the stacking limits of a multiphasic buffer system can be further widened, thus making it applicable to gel electrophoresis of a larger spectrum of rapidly migrating species, such as sodium dodecyl sulfate-proteins/peptides and nucleic acids, than has been possible previously. Highly sensitive detection methods for proteins as well as for polypeptides down to approximately Mr 1000 are described. Dansylated proteins/peptides were detected by their fluorescence either directly within the gel or following electroblotting into anion-exchange or polyvinylidene difluoride membranes. The latter procedure resulted in detection sensitivities of approximately 1 ng. Nondansylated proteins/peptides were either detected within the gel by colloidal Coomassie staining or by electroblotting into polyvinylidene difluoride membranes, followed by colloidal gold staining. Prior to both staining procedures the proteins/peptides were pretreated with glutardialdehyde in the presence of borate at near neutral pH values to generate protein/peptide polymers of poor solubility. For a given pH the efficiency of the latter procedure was significantly influenced by the nature of the buffer ion used in the fixation buffer. In contrast to conventional fixation procedures even small polypeptides (Mr 1000) were immobilized and approximately 15 ng and 0.75 ng could be detected after colloidal Coomassie and colloidal gold staining, respectively. PMID:1718736

  19. Directionality of replication fork movement determined by two-dimensional native-native DNA agarose gel electrophoresis.

    PubMed

    Ivessa, Andreas S

    2013-01-01

    The analysis of replication intermediates by the neutral-neutral two-dimensional agarose gel technique allows determining the chromosomal positions where DNA replication initiates, whether replication forks pause or stall at specific sites, or whether two DNA molecules undergo DNA recombination events. This technique does not, however, immediately tell in which direction replication forks migrate through the DNA region under investigation. Here, we describe the procedure to determine the direction of replication fork progression by carrying out a restriction enzyme digest of DNA imbedded in agarose after the completion of the first dimension of a 2D gel. PMID:23913286

  20. Lipid, Detergent, and Coomassie Blue G-250 Affect the Migration of Small Membrane Proteins in Blue Native Gels

    PubMed Central

    Crichton, Paul G.; Harding, Marilyn; Ruprecht, Jonathan J.; Lee, Yang; Kunji, Edmund R. S.

    2013-01-01

    Blue native gel electrophoresis is a popular method for the determination of the oligomeric state of membrane proteins. Studies using this technique have reported that mitochondrial carriers are dimeric (composed of two ?32-kDa monomers) and, in some cases, can form physiologically relevant associations with other proteins. Here, we have scrutinized the behavior of the yeast mitochondrial ADP/ATP carrier AAC3 in blue native gels. We find that the apparent mass of AAC3 varies in a detergent- and lipid-dependent manner (from ?60 to ?130 kDa) that is not related to changes in the oligomeric state of the protein, but reflects differences in the associated detergent-lipid micelle and Coomassie Blue G-250 used in this technique. Higher oligomeric state species are only observed under less favorable solubilization conditions, consistent with aggregation of the protein. Calibration with an artificial covalent AAC3 dimer indicates that the mass observed for solubilized AAC3 and other mitochondrial carriers corresponds to a monomer. Size exclusion chromatography of purified AAC3 in dodecyl maltoside under blue native gel-like conditions shows that the mass of the monomer is ?120 kDa, but appears smaller on gels (?60 kDa) due to the unusually high amount of bound negatively charged dye, which increases the electrophoretic mobility of the protein-detergent-dye micelle complex. Our results show that bound lipid, detergent, and Coomassie stain alter the behavior of mitochondrial carriers on gels, which is likely to be true for other small membrane proteins where the associated lipid-detergent micelle is large when compared with the mass of the protein. PMID:23744064

  1. Polyacrylamide gel mapping of chicken tRNA: comparison of polysome-bound and whole-cell tRNA from normal and avian sarcoma virus-infected chicken embryo fibroblasts.

    PubMed Central

    Reinisch, F; Heyman, T

    1982-01-01

    Analysis of the tRNA population from chicken cells was performed by means of polyacrylamide gel mapping. About 60 species were detected; most of these were positively identified by their acceptor specificity. The comparison of polysome-bound and overall cellular tRNA gel patterns from normal and Rous sarcoma virus-infected chicken embryo fibroblasts led us to the following observations: some tRNA species were present in the same relative proportions in all the preparations, and within isoaccepting groups the same species was preponderant; however, although about 8% of whole-cell tRNA was recovered in polysomal preparations, amounts ranging from 3 to 30% were found for individual tRNA species. This points to the absence of a direct correlation between the amount of each mature tRNA species produced and the frequency with which it is used in this case of embryonic cells. No significant difference was observed between the whole-cell tRNA patterns from normal and infected cells. Thus, tRNA transcription appears unaltered when cells are transformed and virus producing. No change was observed in the extent of a post-transcriptional modification of tRNAPhe (the base Y). However, viral infection led to some changes in the relative proportions of individual species from polysomal preparations. Images PMID:6294501

  2. Electrophoretic mobility shift in native gels indicates calcium-dependent structural changes of neuronal calcium sensor proteins.

    PubMed

    Viviano, Jeffrey; Krishnan, Anuradha; Wu, Hao; Venkataraman, Venkat

    2016-02-01

    In proteins of the neuronal calcium sensor (NCS) family, changes in structure as well as function are brought about by the binding of calcium. In this article, we demonstrate that these structural changes, solely due to calcium binding, can be assessed through electrophoresis in native gels. The results demonstrate that the NCS proteins undergo ligand-dependent conformational changes that are detectable in native gels as a gradual decrease in mobility with increasing calcium but not other tested divalent cations such as magnesium, strontium, and barium. Surprisingly, such a gradual change over the entire tested range is exhibited only by the NCS proteins but not by other tested calcium-binding proteins such as calmodulin and S100B, indicating that the change in mobility may be linked to a unique NCS family feature-the calcium-myristoyl switch. Even within the NCS family, the changes in mobility are characteristic of the protein, indicating that the technique is sensitive to the individual features of the protein. Thus, electrophoretic mobility on native gels provides a simple and elegant method to investigate calcium (small ligand)-induced structural changes at least in the superfamily of NCS proteins. PMID:26617128

  3. Blue-native PAGE in plants: a tool in analysis of protein-protein interactions

    PubMed Central

    Eubel, Holger; Braun, Hans-Peter; Millar, A Harvey

    2005-01-01

    Intact protein complexes can be separated by apparent molecular mass using a standard polyacrylamide gel electrophoresis system combining mild detergents and the dye Coomassie Blue. Referring to the blue coloured gel and the gentle method of solubilization yielding native and enzymatically active protein complexes, this technique has been named Blue-Native Polyacrylamide Gel-Electrophoresis (BN-PAGE). BN-PAGE has become the method of choice for the investigation of the respiratory protein complexes of the electron transfer chains of a range of organisms, including bacteria, yeasts, animals and plants. It allows the separation in two dimensions of extremely hydrophobic protein sets for analysis and also provides information on their native interactions. In this review we discuss the capabilities of BN-PAGE in proteomics and the wider investigation of protein:protein interactions with a focus on its use and potential in plant science. PMID:16287510

  4. Fluorescent staining of glycoproteins in sodium dodecyl sulfate polyacrylamide gels by 4H-[1]-benzopyrano[4,3-b]thiophene-2-carboxylic acid hydrazide.

    PubMed

    Zhu, Zhongxin; Zhou, Xuan; Wang, Yang; Chi, Lisha; Ruan, Dandan; Xuan, Yuanhu; Cong, Weitao; Jin, Litai

    2014-06-01

    A fluorescent detection method for glycoproteins in SDS-PAGE by using 4H-[1]-benzopyrano[4,3-b]thiophene-2-carboxylic acid hydrazide (BH) was developed in this study. As low as 4-8 ng glycoproteins (transferrin, ?1-acid glycoprotein) could be specifically detected by the BH staining method, which is twofold more sensitive than that of the most commonly used Pro-Q Emerald 488 glycoprotein stain. Furthermore, the specificity of the newly developed stain for glycoproteins was demonstrated by 1-D and 2-D SDS-PAGE, deglycosylation, glycoprotein affinity enrichment and LC-MS/MS, respectively. According to the results, it is concluded that BH stain may provide new choices for convenient, sensitive, specific and economic visualization of gel-separated glycoproteins. PMID:24712021

  5. Automated in-line gel filtration for native state mass spectrometry.

    PubMed

    Waitt, Greg M; Xu, Robert; Wisely, G Bruce; Williams, Jon D

    2008-02-01

    Characterization of protein-ligand complexes by nondenaturing mass spectrometry provides direct evidence of drug-like molecules binding with potential therapeutic targets. Typically, protein-ligand complexes to be analyzed contain buffer salts, detergents, and other additives to enhance protein solubility, all of which make the sample unable to be analyzed directly by electrospray ionization mass spectrometry. This work describes an in-line gel-filtration method that has been automated and optimized. Automation was achieved using commercial HPLC equipment. Gel column parameters that were optimized include: column dimensions, flow rate, packing material type, particle size, and molecular weight cut-off. Under optimal conditions, desalted protein ions are detected 4 min after injection and the analysis is completed in 20 min. The gel column retains good performance even after >200 injections. A demonstration for using the in-line gel-filtration system is shown for monitoring the exchange of fatty acids from the pocket of a nuclear hormone receptor, peroxisome proliferator activator-delta (PPARdelta) with a tool compound. Additional utilities of in-line gel-filtration mass spectrometry system will also be discussed. PMID:17596960

  6. EFFECTS OF NATIVE LIPID CONTENT ON THE GEL PROPERTIES AND SPHERULITE FORMATION OF JET COOKED CORNSTARCH

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Steam jet cooking is a process that has been used for many years to prepare aqueous starch dispersions for both food and industrial applications. In this process, gel properties of the final starch dispersion are dependent on several variables, including starch concentration, steam pressure, stirri...

  7. Polyacrylamide hydrogels as sustained release drug delivery dressing materials

    NASA Astrophysics Data System (ADS)

    Rosiak, J.; Burozak, K.; P?kala, W.

    The possibility of the polyacrylamide hydrogels application as sustained release drug delivery systems is discussed. This paper reports some work carried out in order to obtain a new type of dressings consisting of polyurethane foil coated on one-side with the drug and on opposite one with the polyacrylamide hydrogel layer. The doses of 60Co ?-irradiation necessary to prepare polyacrylamide solutions with desirable viscosities were determined. Release rate of drug from dressing materials through the gel layer versus quality and quantity of the add-on hydrogel were investigated. The release rate increases for high amount of the coating gel as well as with the increase of the polyacrylamide content. The tentative biological tests have shown full biocompatibility and usefulness of this type of dressings for the clinical purposes.

  8. Protein imprinting in polyacrylamide-based gels

    PubMed Central

    Zayats, Maya; Brenner, Andrew J.; Searson, Peter C.

    2015-01-01

    Protein imprinting in hydrogels is a method to produce materials capable of selective recognition and capture of a target protein. Here we report on the imprinting of fluorescently-labeled maltose binding protein (MBP) in acrylamide (AAm)/N-isopropylacrylamide (NIPAm) hydrogels. The targeting efficiency and selectivity of protein recognition is usually characterized by the imprinting factor, which in the simplest case is the ratio of protein uptake in an imprinted film divided by the uptake by the corresponding non-imprinted film. Our objective in this work is to study the dynamics of protein binding and elution in imprinted and non-imprinted films to elucidate the processes that control protein recognition. Protein elution from imprinted and non-imprinted films suggests that imprinting results in sites with a distribution of binding energies, and that only a relatively small fraction of these sites exhibit strong binding. PMID:25034963

  9. Probe mobility in native phosphocaseinate suspensions and in a concentrated rennet gel: effects of probe flexibility and size.

    PubMed

    Salami, Souad; Rondeau-Mouro, Corinne; van Duynhoven, John; Mariette, Franois

    2013-06-19

    Pulsed field gradient nuclear magnetic resonance and proton nuclear magnetic resonance relaxometry were used to study the self-diffusion coefficients and molecular dynamics of linear (PEGs) and spherical probes (dendrimers) in native phosphocaseinate suspensions and in a concentrated rennet gel. It was shown that both the size and the shape of the diffusing molecules and the matrix topography affected the diffusion and relaxation rates. In suspensions, both translational and rotational diffusion decreased with increasing casein concentrations due to increased restriction in the freedom of motion. Rotational diffusion was, however, less hindered than translational diffusion. After coagulation, translational diffusion increased but rotational diffusion decreased. Analysis of the T? relaxation times obtained for probes of different sizes distinguished the free short-chain relaxation formed from a few monomeric units from (i) the relaxation of protons attached to long polymer chains and (ii) the short-chain relaxation attached to a rigid dendrimer core. PMID:23650920

  10. Microchannel gel electrophoretic separation systems and methods for preparing and using

    DOEpatents

    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.

  11. Microchannel gel electrophoretic separation systems and methods for preparing and using

    DOEpatents

    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.

  12. Characterization of a gel-separated unknown glycoprotein by liquid chromatography/multistage tandem mass spectrometry: analysis of rat brain Thy-1 separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis.

    PubMed

    Itoh, Satsuki; Kawasaki, Nana; Harazono, Akira; Hashii, Noritaka; Matsuishi, Yukari; Kawanishi, Toru; Hayakawa, Takao

    2005-11-11

    We developed an efficient and convenient strategy for protein identification and glycosylation analysis of a small amount of unknown glycoprotein in a biological sample. The procedure involves isolation of proteins by electrophoresis and mass spectrometric peptide/glycopeptide mapping by LC/ion trap mass spectrometer. For the complete glycosylation analysis, proteins were extracted in intact form from the gel, and proteinase-digested glycoproteins were then subjected to LC/multistage tandem MS (MSn) incorporating a full mass scan, in-source collision-induced dissociation (CID), and data-dependent MSn. The glycopeptides were localized in the peptide/glycopeptide map by using oxonium ions such as HexNAc+ and NeuAc+, generated by in-source CID, and neutral loss by CID-MS/MS. We conducted the search analysis for the glycopeptide identification using search parameters containing a possible glycosylation at the Asn residue with N-acetylglucosamine (203 Da). We were able to identify the glycopeptides resulting from predictable digestion with proteinase. The glycopeptides caused by irregular cleavages were not identified by the database search analysis, but their elution positions were localized using oxonium ions produced by in-source CID, and neutral loss by the data-dependent MSn. Then, all glycopeptides could be identified based on the product ion spectra which were sorted from data-dependent CID-MSn spectra acquired around localized positions. Using this strategy, we successfully elucidated site-specific glycosylation of Thy-1, glycosylphosphatidylinositol (GPI)-anchored proteins glycosylated at Asn23, 74, and 98, and at Cys111. High-mannose-type, complex-type, and hybrid-type oligosaccharides were all found to be attached to Asn23, 74 and 98, and four GPI structures could be characterized. Our method is simple, rapid and useful for the characterization of unknown glycoproteins in a complex mixture of proteins. PMID:16257296

  13. Gel Mobility Shift Assays to Detect ProteinRNA Interactions

    PubMed Central

    Yakhnin, Alexander V.; Yakhnin, Helen; Babitzke, Paul

    2015-01-01

    The gel mobility shift assay is a powerful technique for detecting and quantifying proteinRNA interactions. While other techniques such as filter binding and isothermal titration calorimetry (ITC) are available for quantifying proteinRNA interactions, gel shift analysis provides the added advantage that you can visualize the proteinRNA complexes. In the gel shift assay, proteinRNA complexes are typically separated from the unbound RNA using native polyacrylamide gels in Tris/borate/EDTA buffer, although an alternative Tris-glycine buffering system is superior in many situations. Here, we describe both gel shift methods, along with strategies to improve separation of proteinRNA complexes from free RNA, which can be a particular challenge for small RNA binding proteins. PMID:22736005

  14. 2-D native-PAGE/SDS-PAGE visualization of an oligomer's subunits: application to the analysis of IgG.

    PubMed

    Gonzalez, Leticia; Bustamante, Juan J; Barea-Rodriguez, Edwin J; Martinez, Andrew O; Haro, Luis S

    2006-05-01

    A 2-D native-PAGE/SDS-PAGE method for detecting the subunit components of protein oligomers at low picomole sensitivity is presented. IgG was electrophoresed in a native acidic polyacrylamide gel in amounts ranging from 51 pmol to 60 fmol. Silver-staining (native fast silver stain, ammoniacal silver stain, permanganate silver stain), Coomassie-staining (R-250, G-250), metal ion-reverse-staining (zinc, copper), and fluorescent chromophore-staining (SYPRO Ruby) methods were used to visualize the IgG oligomers. The protein zones were then excised, separated by SDS-PAGE, and subunits visualized with a permanganate silver stain. The Coomassie R-250/permanganate silver-staining combination detected IgG subunits using 2 pmol of sample. Coomassie G-250 and native fast silver staining in the first-dimensional gel produced detectable subunits in the second-dimensional separation at 3 and 13 pmol, respectively. Staining with silver (ammoniacal, permanganate), copper, zinc, or SYPRO Ruby in the first-dimensional gel did not produce discernible subunits in the second-dimensional gels due to protein streaking or protein immobilization in the native gel. When using a 2-D native-PAGE/SDS-PAGE system, Coomassie staining of the first-dimensional native gel combined with permanganate silver staining of the second-dimensional denaturing gel provides the most sensitive method (2-3 pmol) for visualizing constituent subunits from their oligomeric assemblies. PMID:16703630

  15. 21 CFR 172.255 - Polyacrylamide.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ..., Films and Related Substances 172.255 Polyacrylamide. Polyacrylamide containing not more than 0.2 percent of acrylamide monomer may be safely used as a film former in the imprinting of soft-shell...

  16. 21 CFR 172.255 - Polyacrylamide.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ..., Films and Related Substances 172.255 Polyacrylamide. Polyacrylamide containing not more than 0.2 percent of acrylamide monomer may be safely used as a film former in the imprinting of soft-shell...

  17. 21 CFR 172.255 - Polyacrylamide.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ..., Films and Related Substances § 172.255 Polyacrylamide. Polyacrylamide containing not more than 0.2 percent of acrylamide monomer may be safely used as a film former in the imprinting of soft-shell...

  18. 21 CFR 172.255 - Polyacrylamide.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ..., Films and Related Substances § 172.255 Polyacrylamide. Polyacrylamide containing not more than 0.2 percent of acrylamide monomer may be safely used as a film former in the imprinting of soft-shell...

  19. Two-component supramolecular gels derived from amphiphilic shape-persistent cyclo[6]aramides for specific recognition of native arginine.

    PubMed

    He, Youzhou; Xu, Min; Gao, Rongzhao; Li, Xiaowei; Li, Fengxue; Wu, Xuedan; Xu, Dingguo; Zeng, Huaqiang; Yuan, Lihua

    2014-10-27

    A unique supramolecular two-component gelation system was constructed from amphiphilic shape-persistent cyclo[6]aramides and diethylammonium chloride (or triethylammonium chloride). This system has the ability to discriminate native arginine from 19 other amino acids in a specific fashion. Cyclo[6]aramides show preferential binding for the guanidinium residue over ammonium groups. This specificity was confirmed by both experimental results and theoretical simulations. These results demonstrated a new modular displacement strategy, exploring the use of species-binding hydrogen-bonded macrocyclic foldamers for the construction of two-component gelation systems for selective recognition of native amino acids by competitive host-guest interactions. This strategy may be amenable to developing a variety of functional two-component gelators for specific recognition of various targeted organic molecular species. PMID:25213644

  20. Staining proteins in gels.

    PubMed

    Sasse, Joachim; Gallagher, Sean R

    2004-02-01

    This unit describes protocols for detecting protein in a gel by either Coomassie blue, silver or fluorescent staining. Alternate rapid staining procedures are provided for each method and a support protocol describes how to photograph stained gels. Fluorescent staining (e.g., with SYPRO Orange or Red) is described as a popular alternative to traditional staining procedures, mainly because it is more sensitive than Coomassie staining, and often as sensitive as silver staining. Staining of proteins in SDS-polyacrylamide gels is described, and an alternate protocol details variations in the procedure for proteins in nondenaturing gels. A final support protocol describes the photography of fluorescently stained proteins. PMID:18432935

  1. White gels: an easy way to preserve methylene blue stained gels.

    PubMed

    Soto, Ana Maria; Draper, David

    2012-02-01

    Methylene blue can be used as a stain for visualizing nucleic acids in polyacrylamide gel electrophoresis. However, its relatively low sensitivity and reversible binding make it a temporary stain that diffuses from the gel relatively fast. Here we describe a very simple method for fixing methylene blue bands in nucleic acid polyacrylamide gels. The procedure makes the methylene blue stain permanent and increases the visibility of the bands, also contributing to increasing the sensitivity of methylene blue. PMID:22107889

  2. Tough and highly stretchable polyacrylamide nanocomposite hydrogels with chitin nanocrystals.

    PubMed

    Liu, Mingxian; Huang, Jiandong; Luo, Binghong; Zhou, Changren

    2015-01-01

    Chitin nanocrystals (CNCs) that were 10-20 nm wide and 100-500 nm long were synthetized via acidolysis and characterized with various methods. To avoid the flocculation of CNCs in the initiator solution during acrylamide polymerization, chitosan was selected as a surface modifier. The chitosan-modified CNCs were employed as multifunctional crosslinkers for the polyacrylamide (PAAm) nanocomposite (NC) hydrogels. The NC gels were tough and stretchable; for example, the maximum tensile strength and the elongation at break of the NC gels were 90 kPa and 3070%, respectively. The dynamic shear modulus of the NC gels was also significantly higher than that of the PAAm. The NC gels were nearly free of residual strain after 2000% elongation. The microstructures of all NC gels were porous, with a pore size of 20-100 ?m. The maximum equilibrium swelling degree of the NC gels was 3800%. The improvement in the properties of the NC gels is attributed to the good dispersion of CNCs and the interfacial interactions in the composites. This work developed PAAm NC hydrogels with CNCs for application as absorbent or biomedical material due to the high mechanical properties, high absorb ability and good biocompatibility of CNCs and explored new applications for CNCs as well. PMID:25841364

  3. Spring-loaded polymeric gel actuators

    SciTech Connect

    Shahinpoor, M.

    1995-02-14

    Spring-loaded electrically controllable polymeric gel actuators are disclosed. The polymeric gels can be polyvinyl alcohol, polyacrylic acid, or polyacrylamide, and are contained in an electrolytic solvent bath such as water plus acetone. The action of the gel is mechanically biased, allowing the expansive and contractile forces to be optimized for specific applications. 5 figs.

  4. Spring-loaded polymeric gel actuators

    DOEpatents

    Shahinpoor, Mohsen (Albuquerque, NM)

    1995-01-01

    Spring-loaded electrically controllable polymeric gel actuators are disclosed. The polymeric gels can be polyvinyl alcohol, polyacrylic acid, or polyacrylamide, and are contained in an electrolytic solvent bath such as water plus acetone. The action of the gel is mechanically biased, allowing the expansive and contractile forces to be optimized for specific applications.

  5. Microrheology of cross-linked polyacrylamide networks.

    PubMed

    Dasgupta, Bivash R; Weitz, D A

    2005-02-01

    Experiments investigating the local viscoelastic properties of a chemically cross-linked polymer are performed on polyacrylamide solutions in the sol and the gel regimes using polystyrene beads of varying sizes and surface chemistry as probes. The thermal motions of the probes are measured to obtain the elastic and viscous moduli of the sample. Probe dynamics are measured using two different dynamic light scattering techniques, diffusing wave spectroscopy (DWS) and quasielastic light scattering (QELS) as well as video-based particle tracking. Diffusing wave spectroscopy probes the short-time dynamics of the scatterers while QELS measures the dynamics at larger times. Video-based particle tracking provides a way to investigate the local environment of the individual probe particles. A combination of all the techniques results in a larger range of frequencies that can be probed compared to conventional bulk measurements while providing local information at the level of individual probes. A modified algebraic form of the generalized Stokes-Einstein equation is used to calculate the frequency-dependent moduli. A comparison of microrheological measurements with bulk rheology exhibits striking similarity, confirming the applicability of microrheology for chemically cross-linked polymeric systems. PMID:15783330

  6. Microrheology of cross-linked polyacrylamide networks

    NASA Astrophysics Data System (ADS)

    Dasgupta, Bivash R.; Weitz, D. A.

    2005-02-01

    Experiments investigating the local viscoelastic properties of a chemically cross-linked polymer are performed on polyacrylamide solutions in the sol and the gel regimes using polystyrene beads of varying sizes and surface chemistry as probes. The thermal motions of the probes are measured to obtain the elastic and viscous moduli of the sample. Probe dynamics are measured using two different dynamic light scattering techniques, diffusing wave spectroscopy (DWS) and quasielastic light scattering (QELS) as well as video-based particle tracking. Diffusing wave spectroscopy probes the short-time dynamics of the scatterers while QELS measures the dynamics at larger times. Video-based particle tracking provides a way to investigate the local environment of the individual probe particles. A combination of all the techniques results in a larger range of frequencies that can be probed compared to conventional bulk measurements while providing local information at the level of individual probes. A modified algebraic form of the generalized Stokes-Einstein equation is used to calculate the frequency-dependent moduli. A comparison of microrheological measurements with bulk rheology exhibits striking similarity, confirming the applicability of microrheology for chemically cross-linked polymeric systems.

  7. Relative Quantitative Comparisons of the Extracellular Protein Profiles of Staphylococcus aureus UAMS-1 and Its sarA, agr, and sarA agr Regulatory Mutants Using One-Dimensional Polyacrylamide Gel Electrophoresis and Nanocapillary Liquid Chromatography Coupled with Tandem Mass Spectrometry ?

    PubMed Central

    Jones, Richard C.; Deck, Joanna; Edmondson, Ricky D.; Hart, Mark E.

    2008-01-01

    One-dimensional polyacrylamide gel electrophoresis followed by nanocapillary liquid chromatography coupled with mass spectrometry was used to analyze proteins isolated from Staphylococcus aureus UAMS-1 after 3, 6, 12, and 24 h of in vitro growth. Protein abundance was determined using a quantitative value termed normalized peptide number, and overall, proteins known to be associated with the cell wall were more abundant early on in growth, while proteins known to be secreted into the surrounding milieu were more abundant late in growth. In addition, proteins from spent media and cell lysates of strain UAMS-1 and its isogenic sarA, agr, and sarA agr regulatory mutant strains during exponential growth were identified, and their relative abundances were compared. Extracellular proteins known to be regulated by the global regulators sarA and agr displayed protein levels in accordance with what is known regarding the effects of these regulators. For example, cysteine protease (SspB), endopeptidase (SspA), staphopain (ScpA), and aureolysin (Aur) were higher in abundance in the sarA and sarA agr mutants than in strain UAMS-1. The immunoglobulin G (IgG)-binding protein (Sbi), immunodominant staphylococcal antigen A (IsaA), IgG-binding protein A (Spa), and the heme-iron-binding protein (IsdA) were most abundant in the agr mutant background. Proteins whose abundance was decreased in the sarA mutant included fibrinogen-binding protein (Fib [Efb]), IsaA, lipase 1 and 2, and two proteins identified as putative leukocidin F and S subunits of the two-component leukotoxin family. Collectively, this approach identified 1,263 proteins (matches of two peptides or more) and provided a convenient and reliable way of identifying proteins and comparing their relative abundances. PMID:18539737

  8. Investigations on preparation and properties of modified polyacrylamide hydrogels for application as wound dressing materials.

    PubMed

    Lukaszczyk, J

    1995-01-01

    Investigations on the influence of the polymerization conditions and feed ratio as well as various modification of the composite polyacrylamide-agar hydrogels on their properties has been presented. Results of the model studies have been utilized in the optimalization of the manufacturing parameters and procedures of the preparation of hydrogel dressing foils and granulated dressing gels. PMID:7479424

  9. Electrically controlled polymeric gel actuators

    DOEpatents

    Adolf, D.B.; Shahinpoor, M.; Segalman, D.J.; Witkowski, W.R.

    1993-10-05

    Electrically controlled polymeric gel actuators or synthetic muscles are described capable of undergoing substantial expansion and contraction when subjected to changing pH environments, temperature, or solvent. The actuators employ compliant containers for the gels and their solvents. The gels employed may be cylindrical electromechanical gel fibers such as polyacrylamide fibers or a mixture of poly vinyl alcohol-polyacrylic acid arranged in a parallel aggregate and contained in an electrolytic solvent bath such as salt water. The invention includes smart, electrically activated devices exploiting this phenomenon. These devices are capable of being manipulated via active computer control as large displacement actuators for use in adaptive structure such as robots. 11 figures.

  10. Electrically controlled polymeric gel actuators

    DOEpatents

    Adolf, Douglas B. (Albuquerque, NM); Shahinpoor, Mohsen (Albuquerque, NM); Segalman, Daniel J. (Albuquerque, NM); Witkowski, Walter R. (Albuquerque, NM)

    1993-01-01

    Electrically controlled polymeric gel actuators or synthetic muscles capable of undergoing substantial expansion and contraction when subjected to changing pH environments, temperature, or solvent. The actuators employ compliant containers for the gels and their solvents. The gels employed may be cylindrical electromechanical gel fibers such as polyacrylamide fibers or a mixture of poly vinyl alcohol-polyacrylic acid arranged in a parallel aggregate and contained in an electrolytic solvent bath such as salt water. The invention includes smart, electrically activated devices exploiting this phenomenon. These devices are capable of being manipulated via active computer control as large displacement actuators for use in adaptive structure such as robots.

  11. Templated Native Silk Smectic Gels

    NASA Technical Reports Server (NTRS)

    Jin, Hyoung-Joon (Inventor); Park, Jae-Hyung (Inventor); Valluzzi, Regina (Inventor)

    2013-01-01

    One aspect of the present invention relates to a method of preparing a fibrous protein smectic hydrogel by way of a solvent templating process, comprising the steps of pouring an aqueous fibrous protein solution into a container comprising a solvent that is not miscible with water; sealing the container and allowing it to age at about room temperature; and collecting the resulting fibrous protein smectic hydrogel and allowing it to dry. Another aspect of the present invention relates to a method of obtaining predominantly one enantiomer from a racemic mixture, comprising the steps of pouring an aqueous fibrous protein solution into a container comprising a solvent that is not miscible with water; sealing the container and allowing it to age at about room temperature; allowing the enantiomers of racemic mixture to diffuse selectively into the smectic hydrogel in solution; removing the smectic hydrogel from the solution; rinsing predominantly one enantiomer from the surface of the smectic hydrogel; and extracting predominantly one enantiomer from the interior of the smectic hydrogel. The present invention also relates to a smectic hydrogel prepared according to an aforementioned method.

  12. Templated native silk smectic gels

    NASA Technical Reports Server (NTRS)

    Jin, Hyoung-Joon (Inventor); Park, Jae-Hyung (Inventor); Valluzzi, Regina (Inventor)

    2009-01-01

    One aspect of the present invention relates to a method of preparing a fibrous protein smectic hydrogel by way of a solvent templating process, comprising the steps of pouring an aqueous fibrous protein solution into a container comprising a solvent that is not miscible with water; sealing the container and allowing it to age at about room temperature; and collecting the resulting fibrous protein smectic hydrogel and allowing it to dry. Another aspect of the present invention relates to a method of obtaining predominantly one enantiomer from a racemic mixture, comprising the steps of pouring an aqueous fibrous protein solution into a container comprising a solvent that is not miscible with water; sealing the container and allowing it to age at about room temperature; allowing the enantiomers of racemic mixture to diffuse selectively into the smectic hydrogel in solution; removing the smectic hydrogel from the solution; rinsing predominantly one enantiomer from the surface of the smectic hydrogel; and extracting predominantly one enantiomer from the interior of the smectic hydrogel. The present invention also relates to a smectic hydrogel prepared according to an aforementioned method.

  13. Crystallization of calcium phosphate in polyacrylamide hydrogels containing phosphate ions

    NASA Astrophysics Data System (ADS)

    Yokoi, Taishi; Kawashita, Masakazu; Kikuta, Koichi; Ohtsuki, Chikara

    2010-08-01

    Calcium phosphate crystals were formed in polyacrylamide (PAAm) hydrogels containing phosphate ions by diffusion of calcium ions from calcium nitrate (Ca(NO 3) 2) solutions covering the gels. Changes in crystalline phases and crystal morphology of calcium phosphate, and in ion concentrations of the Ca(NO 3) 2 solutions were investigated as a function of reaction time. Single or two coexisting crystalline phases of calcium phosphate, hydroxyapatite (HAp), HAp/dicalcium phosphate dihydrate (DCPD) or octacalcium phosphate (OCP)/DCPD were formed in the gels. HAp crystals are formed near the surface of the gels. The dense HAp layer and HAp/DCPD layer prevented diffusion of calcium ions from the Ca(NO 3) 2 solution, thus formation of calcium phosphate in the gel phase was inhibited. Formation of DCPD was observed to follow the formation of OCP or HAp. The size of the OCP crystals gradually increased with reaction time, while changes in size of HAp crystals were not observed. The reaction time required for DCPD formation depended on the degree of supersaturation with respect to DCPD in the systems. DCPD formed within 1 day under high supersaturation conditions, whereas it formed at 10 days in low supersaturation conditions.

  14. A Simple Vertical Slab Gel Electrophoresis Apparatus.

    ERIC Educational Resources Information Center

    Carter, J. B.; And Others

    1983-01-01

    Describes an inexpensive, easily constructed, and safe vertical slab gel kit used routinely for sodium dodecyl sulphate-polyacrylamide gel electrophoresis research and student experiments. Five kits are run from a single transformer. Because toxic solutions are used, students are given plastic gloves and closely supervised during laboratory…

  15. A Simple Vertical Slab Gel Electrophoresis Apparatus.

    ERIC Educational Resources Information Center

    Carter, J. B.; And Others

    1983-01-01

    Describes an inexpensive, easily constructed, and safe vertical slab gel kit used routinely for sodium dodecyl sulphate-polyacrylamide gel electrophoresis research and student experiments. Five kits are run from a single transformer. Because toxic solutions are used, students are given plastic gloves and closely supervised during laboratory

  16. Staining proteins in gels.

    PubMed

    Sasse, Joachim; Gallagher, Sean R

    2003-08-01

    This unit describes protocols for detecting protein in a gel by either Coomassie blue staining or silver staining. The former is easier and more rapid; however, silver staining methods are considerably more sensitive and thus can be used to detect smaller amounts of protein. Alternate rapid staining procedures are provided for each method and a support protocol describes how to photograph stained gels. Fluorescent staining is a popular alternative to traditional staining procedures, mainly because it is more sensitive than Coomassie staining, and often as sensitive as silver staining. Staining of proteins in SDS-polyacrylamide gels is described, and an alternate protocol details variations in the procedure for proteins in nondenaturing gels. A final support protocol describes the photography of fluorescently stained proteins. PMID:18265316

  17. Staining proteins in gels.

    PubMed

    Sasse, Joachim; Gallagher, Sean R

    2009-01-01

    This unit describes protocols for detecting protein in a gel by Coomassie blue, silver, or fluorescent staining. As a general protein stain, Coomassie is easier and more rapid; however, fluorescent and silver staining methods are considerably more sensitive and thus can be used to detect smaller amounts of protein. Fluorescent staining is a popular alternative to traditional staining procedures, mainly because it is more sensitive than Coomassie staining, and often as sensitive as silver staining. Alternate protocols describe rapid Coomassie and silver staining methods, as well as fluorescent stains that are specific for phosphoproteins and glycoproteins. Staining of proteins in SDS-polyacrylamide gels is described; variations for fluorescent staining of proteins in nondenaturing gels are also included. Support protocols describe photography of stained proteins. PMID:19170026

  18. Polyacrylamide Hydrogel Properties for Horticultural Applications

    Technology Transfer Automated Retrieval System (TEKTRAN)

    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...

  19. Further development of an electroosmotic medium pump system for preparative disk gel electrophoresis.

    PubMed

    Hayakawa, Mitsuo; Hosogi, Yumiko; Takiguchi, Hisashi; Shiroza, Teruaki; Shibata, Yasuko; Hiratsuka, Koichi; Kiyama-Kishikawa, Michiko; Hamajima, Susumu; Abiko, Yoshimitsu

    2003-02-01

    A simple and practical 6.8-cm-diameter (36.30-cm(2) cross-sectional-area) preparative disk gel electrophoresis device, based on the design of M. Hayakawa et al. (Anal. Biochem. 288 (2001) 168), in which the elution buffer is driven by an electroosmotic buffer flow through the membrane into the elution chamber from the anode chamber was constructed. We have found that the dialysis membranes employed provide suitable flow rates for the elution buffer, similar to those of an earlier 3.6-cm-diameter device, resulting in the prevention of excess eluate dilution. The efficiency of this device was demonstrated by the fractionation of a bovine serum albumin (BSA) Cohn V fraction into monomer, dimer, and oligomer components using nondenaturing polyacrylamide gel electrophoresis (native-PAGE). The maximum protein concentration of the eluate achieved was 133 mg/ml of BSA monomer, which required a dilution of the eluate for subsequent analytical PAGE performance. As a practical example, the two-dimensional fractionation of soluble dipeptidyl peptidase IV (sDPP IV) from 50 ml fetal bovine serum (3.20 g protein) per gel is presented. The sDPP IV enzyme protein was recovered in a relatively short time, utilizing a 6.5% T native-PAGE and subsequential sodium dodecyl sulfate-PAGE system. This device enhances the possibility of continuous electrophoretic fractionation of complex protein mixtures on a preparative scale. PMID:12576059

  20. Autofluorescence based visualization of proteins from unstained native-PAGE

    NASA Astrophysics Data System (ADS)

    Manjunath, S.; Rao, Bola Sadashiva S.; Satyamoorthy, Kapaettu; Mahato, Krishna Kishore

    2015-03-01

    Proteins are the most diverse and functionally active biomolecules in the living system. In order to understand their diversity and dynamic functionality, visualization in native form without altering structural and functional properties during the separation from the complex mixtures is very much essential. In the present study, a sensitive methodology for optimal visualization of unstained or untagged proteins in native poly-acrylamide gel electrophoresis (N-PAGE) has been developed where, concentration of the acrylamide and bis-acrylamide mixture, Percentage of the gel, fixing of the N-PAGE by methanol: acetic acid: water and washing of the gel in the mili-Q water has been optimized for highest sensitivity using laser induced autofluorescence. The outcome with bovine serum albumin (BSA) in PAGE was found to be highest at acrylamide and bis-acrylamide concentrations of 29.2 and 0.8 respectively in 12% N-PAGE. After the electrophoresis run, washing of the N-PAGE immediately with miliQ water for 12 times and eliminating the methanol: acetic acid: water, fixing of the N-PAGE yielded better sensitivity of visualization. Using the above methodology 25ng of BSA protein band in PAGE was clearly identified by the technique. The currently used staining techniques for the visualization of proteins are coomassie brilliant blue and silver staining, have the sensitivity of 100ng and 5ng respectively. The current methodology was found to be more sensitive as compared to coomassie staining and less sensitive compared to silver staining respectively. The added advantage of this methodology is the faster visualization of proteins without altering their structure and functional properties.

  1. Native SDS-PAGE: High Resolution Electrophoretic Separation of Proteins With Retention of Native Properties Including Bound Metal Ions

    PubMed Central

    Nowakowski, Andrew B.; Wobig, William J.; Petering, David H.

    2014-01-01

    Sodium dodecyl-sulfate polyacrylamide gel electrophoresis (SDS-PAGE) is commonly used to obtain high resolution separation of complex mixtures of proteins. The method initially denatures the proteins that will undergo electrophoresis. Although covalent structural features of resolved proteins can be determined with SDS-PAGE, functional properties are destroyed, including the presence of non-covalently bound metal ions. To address this shortcoming, blue-native (BN)-PAGE has been introduced. This method retains functional properties but at the cost of protein resolving power. To address the need for a high resolution PAGE method that results in the separation of native proteins, experiments tested the impact of changing the conditions of SDS-PAGE on the quality of protein separation and retention of functional properties. Removal of SDS and EDTA from the sample buffer together with omission of a heating step had no effect on the results of PAGE. Reduction of SDS in the running buffer from 0.1% to 0.0375% together with deletion of EDTA also made little impact on the quality of the electrophoretograms of fractions of pig kidney (LLC-PK1) cell proteome in comparison with that achieved with the SDS-PAGE method. The modified conditions were called native (N)SDS-PAGE. Retention of Zn2+ bound in proteomic samples increased from 26 to 98% upon shifting from standard to modified conditions. Moreover, seven of nine model enzymes, including four Zn2+ proteins that were subjected to NSDS-PAGE retained activity. All nine were active in BN-PAGE, whereas all underwent denaturation during SDS-PAGE. Metal retention after electrophoresis was additionally confirmed using laser ablation-inductively coupled plasma-mass spectrometry and in-gel Zn-protein staining using the fluorophore TSQ. PMID:24686569

  2. Performance of 18 polymers in aluminum citrate colloidal dispersion gels

    SciTech Connect

    Smith, J.E.

    1995-11-01

    Colloidal dispersion gels are made up of low concentrations of polymer and aluminum citrate in water. These gels, which are mixed as a homogeneous solution at the surface, provide a valuable tool for in-depth blockage of high permeability regions of rock in heterogeneous reservoirs. Performance of colloidal dispersion gels depends strongly on the type and quality of polymer used. This paper provides an overview of the performance of 18 different polymers in colloidal dispersion gels. 14 of the polymers were partially hydrolyzed polyacrylamides or AMPS polymers in dry crystalline form with varying degrees of hydrolysis and molecular weight. The group also includes one cationic polyacrylamide, one carboxymethyl cellulose, one partially hydrolyzed polyacrylamide in emulsion form and one polysaccharide in dry form. Gels were mixed with the polymers at two polymer concentrations, three polymer:aluminum ratios and in different concentrations of potassium chloride. The gels were quantitatively tested at 1, 7, 14 and 28 days after crosslinking using the transition pressure test, which is a screen flow resistance test. Of the six polymer types tested, only the dry partially hydrolyzed polyacrylamides and AMPS polymers formed colloidal dispersion gels. Gel strength generally increased with increasing anionic charge and molecular weight; however, the manner in which the polymer is manufactured and the impurities present in the polymer also play roles which are more significant than originally expected.

  3. Microfabricated Polyacrylamide Devices for the Controlled Culture of Growing Cells and Developing Organisms

    PubMed Central

    Gude, Sebastian; Recouvreux, Pierre; van Zon, Jeroen S.; Tans, Sander J.

    2013-01-01

    The ability to spatially confine living cells or small organisms while dynamically controlling their aqueous environment is important for a host of microscopy applications. Here, we show how polyacrylamide layers can be patterned to construct simple microfluidic devices for this purpose. We find that polyacrylamide gels can be molded like PDMS into micron-scale structures that can enclose organisms, while being permeable to liquids, and transparent to allow for microscopic observation. We present a range of chemostat-like devices to observe bacterial and yeast growth, and C. elegans nematode development. The devices can integrate PDMS layers and allow for temporal control of nutrient conditions and the presence of drugs on a minute timescale. We show how spatial confinement of motile C. elegans enables for time-lapse microscopy in a parallel fashion. PMID:24086559

  4. A brief review of other notable protein detection methods on acrylamide gels.

    PubMed

    Kurien, Biji T; Scofield, R Hal

    2012-01-01

    Several methods have been described to stain proteins analyzed on acrylamide gels. These include ultrasensitive protein detection in one-dimensional and two-dimensional gel electrophoresis using a fluorescent product from the fungus Epicoccum nigrum; a fluorescence-based Coomassie Blue protein staining; visualization of proteins in acrylamide gels using ultraviolet illumination; fluorescence visualization of proteins in sodium dodecyl sulfate-polyacrylamide gels using environmentally benign, nonfixative, saline solution; and increasing the sensitivity four- to sixfold for detecting trace proteins in dye or silver stained polyacrylamide gels using polyethylene glycol 6000. All these methods are reviewed briefly in this chapter. PMID:22585527

  5. Adsorption characteristics of polyacrylamide and sulfonate-containing polyacrylamide copolymers on sodium kaolinite. [Gamma radiation

    SciTech Connect

    Hollander, A.F.; Somasundaran, P.; Gryte, C.C.

    1981-07-01

    Polyacrylamide and its copolymer containing 6.8 mole % 2-acrylamido-2-methylpropane sulfonic acid were prepared by an irradiation-initiated precipitation polymerization technique. The polymer was characterized by intrinsic viscosity under conditions similar to those used during adsorption measurements. Hydrolytic degradation of the polyacrylamide was found to be negligible under conditions used. The adsorption substrate, sodium kaolinite, was prepared by extensive ion exchange treatment. Equilibrium adsorption of the polymers on the sodium kaolinite was made as a function of polymer concentration, solution pH, ionic strength, and temperature.

  6. Acetyl CoA Carboxylase: Isolation and Characterization of Native Biotin Carboxyl Carrier Protein

    PubMed Central

    Fall, R. Ray; Nervi, A. M.; Alberts, Alfred W.; Vagelos, P. Roy

    1971-01-01

    A large form of biotin carboxyl carrier protein (BCCPL) has been isolated from extracts of Escherichia coli. It has a minimal molecular weight of 20,000, according to its behavior on sodium dodecylsulfate-polyacrylamide gel electrophoresis, and contains approximately 1 mol of biotin per 22,000 g of protein. BCCPL exhibits Km values, in the biotin carboxylase and transcarboxylase half-reactions of acetyl CoA carboxylase, of 2 10-7 M and 4 10-7 M, respectively; these values are 50-100 times lower than those obtained with smaller forms of BCCP previously isolated. Electrophoresis of crude extracts of E. coli indicates that the major biotin-containing protein migrates at the same rate as BCCPL, which suggests that BCCPL is the native form of BCCP in E. coli. Images PMID:4934522

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

    PubMed

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

    2015-07-15

    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, and prefractionation of protein interactions in solution independent of isoelectric point. We demonstrate that this assay is compatible with immunochemical methods and mass spectrometry. The assay was used to investigate interactions with several potential substrates for calreticulin, a chaperone that is involved in different biological aspects through interaction with other proteins. The current analytical assays used to investigate these interactions are mainly spectroscopic aggregation assays or solid phase assays that do not provide a direct visualization of the stable protein complex but rather provide an indirect measure of interactions. Therefore, no interaction studies between calreticulin and substrates in solution have been investigated previously. The results presented here indicate that calreticulin has a preference for substrates with a quaternary structure and primarily β-sheets in their secondary 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. PMID:25908558

  8. Two dimensional blue native/SDS-PAGE to identify mitochondrial complex I subunits modified by 4-hydroxynonenal (HNE)

    PubMed Central

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

    2015-01-01

    The lipid peroxidation product 4-hydroxynonenal (HNE) can form protein-linked HNE adducts, thereby impacting protein structure and function. Mitochondrial complex I (NADH-ubiquinone oxidoreductase), containing at least 45 subunits in mammalian cells, sits in a lipid-rich environment and is thus very susceptible to HNE modifications. In this paper, a procedure for the identification of HNE-modified complex I subunits is described. Complex I was isolated by first dimensional non-gradient blue native polyacrylamide gel electrophoresis (BN-PAGE). The isolated complex I band, visualized by either Coomassie blue staining or silver staining, was further analyzed by second dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). HNE-modified proteins were visualized by Western blotting probed with anti-HNE antibodies. HNE-positive bands were then excised and the proteins contained in them were identified by mass spectrometric peptide sequencing. The method was successfully applied for the identification of two complex I subunits that showed enhanced HNE-modifications in diabetic kidney mitochondria. PMID:25859224

  9. EVALUATION OF NOVEL PRECAST SDS-PAGE GELS FOR SEPARATION OF SORGHUM PROTEINS.

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Pre-cast polyacrylamide gels using novel buffer chemistry for enhanced resolution and shelf life stability were evaluated for separating sorghum proteins. Two gels with different acrylamide concentrations, 12 and 4-12%, were tested with two different buffer systems. Gels were evaluated for separat...

  10. Performance and Biocompatibility of Extremely Tough Alginate/Polyacrylamide Hydrogels

    PubMed Central

    Darnell, Max; Sun, Jeong-Yun; Mehta, Manav; Johnson, Chris; Arany, Praveen; Suo, Zhigang

    2013-01-01

    Although hydrogels now see widespread use in a host of applications, low fracture toughness and brittleness have limited their more broad use. As a recently described interpenetrating network (IPN) of alginate and polyacrylamide demonstrated a fracture toughness of ?9000 J/m2, we sought to explore the biocompatibility and maintenance of mechanical properties of these hydrogels in cell culture and in vivo conditions. These hydrogels can sustain a compressive strain of over 90% with minimal loss of Young's Modulus as well as minimal swelling for up to 50 days of soaking in culture conditions. Mouse mesenchymal stem cells exposed to the IPN gel-conditioned media maintain high viability, and although cells exposed to conditioned media demonstrate slight reductions in proliferation and metabolic activity (WST assay), these effects are abrogated in a dose-dependent manner. Implantation of these IPN hydrogels into subcutaneous tissue of rats for 8 weeks led to mild fibrotic encapsulation and minimal inflammatory response. These results suggest the further exploration of extremely tough alginate/PAAM IPN hydrogels as biomaterials. PMID:23896005

  11. Swelling kinetics of microgels embedded in a polyacrylamide hydrogel matrix.

    PubMed

    Huang, Na; Guan, Ying; Zhu, X X; Zhang, Yongjun

    2014-06-23

    Composite hydrogels--macroscopic hydrogels with embedded microgel particles--are expected to respond to external stimuli quickly because microgels swell much faster than bulky gels. In this work, the kinetics of the pH-induced swelling of a composite hydrogel are studied using turbidity measurements. The embedded microgel is a pH- and thermosensitive poly(N-isopropylacrylamide-co-acrylic acid) microgel and the hydrogel matrix is polyacrylamide. A rapid pH-induced swelling of the embedded microgel particles is observed, confirming that composite hydrogels respond faster than ordinary hydrogels. However, compared with the free microgels, the swelling of the embedded microgel is much slower. Diffusion of OH(-) into the composite hydrogel film is identified as the main reason for the slow swelling of the embedded microgel particles, as the time of the pH-induced swelling of this film is comparable to that of OH(-) diffusion into the film. The composition of the hydrogel matrix does not significantly change the characteristic swelling time of the composite hydrogel film. However, the swelling pattern of the film changes with composition of the hydrogel matrix. PMID:24861868

  12. A simple and cost-effective solid-phase protein nano-assay using polyacrylamide-coated glass plates.

    PubMed

    Krajewski, Wladyslaw A

    2015-02-01

    A new solid-phase protein nano-assay is suggested for simple and sensitive estimation of protein content in sample buffers (a 1-?l sample is sufficient for analysis). The assay is different from conventional "on-filter" assays in that it uses inexpensive fully transparent polyacrylamide gel (PAAG)-coated glass plates as solid support and, thus, combines the convenience of "on-membrane" staining with the sensitivity and ease of documentation of "in-gel" staining (and, therefore, is especially suited for standard lab gel documentation systems). The PAAG plates assay is compatible with all dyes for in-gel protein staining. Depending on the sensitivity of the staining protocol, the assay can be used in macro-, micro-, and nano-assay formats. We also describe a low-cost two-component colloidal Coomassie brilliant blue G-250 (CBB G-250) staining protocol for fast quantitative visualization of proteins spotted on a PAAG plate (the detection limit is up to 2 ng of proteins even when using a Nikon CoolPix digital camera and white light transilluminator instead of a gel scanner). The suggested colloidal CBB G-250 protocol could also be used for visualizing nano-amounts of proteins in polyacrylamide gels. The PAAG plate assay could be useful for proteomic applications and, in general, for all cases where a fast, sensitive, and easily documentable cost-effective solid-phase protein assay is required. PMID:25449300

  13. Development of a generic approach to native metalloproteomics: application to the quantitative identification of soluble copper proteins in Escherichia coli.

    PubMed

    Sevcenco, Ana-Maria; Krijger, Gerard C; Pinkse, Martijn W H; Verhaert, Peter D E M; Hagen, Wilfred R; Hagedoorn, Peter-Leon

    2009-05-01

    A combination of techniques to separate and quantify the native proteins associated with a particular transition metal ion from a cellular system has been developed. The procedure involves four steps: (1) labeling of the target proteins with a suitable short-lived radioisotope (suitable isotopes are (64)Cu, (67)Cu, (187)W, (99)Mo, (69)Zn, (56)Mn, (65)Ni); (2) separation of intact soluble holoproteins using native isoelectric focusing combined with blue native polyacrylamide gel electrophoresis into native-native 2D gel electrophoresis; (3) spot visualization and quantification using autoradiography; and (4) protein identification with tandem mass spectrometry. The method was applied to the identification of copper proteins from a soluble protein extract of wild-type Escherichia coli K12 using the radioisotope (64)Cu. The E. coli protein CueO, which has previously been only identified as a multicopper oxidase following homologous overexpression, was now directly detected as a copper protein against a wild-type background at an expression level of 0.007% of total soluble protein. The retention of the radioisotope by the copper proteins throughout the separation process corroborates the method to be genuinely native. The procedure developed here can be applied to cells of any origin, and to any metal having suitable radioisotopes. The finding that the periplasmic protein CueO is the only major form of soluble protein bound copper in E. coli strengthens the view that the bacterial periplasm contains only a few periplasmic copper proteins, and that the cytosol is devoid of copper proteins. PMID:19205756

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

    SciTech Connect

    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.

  15. Breast cancer following polyacrylamide hydrogel injection for breast augmentation: A case report

    PubMed Central

    CHEN, GANG; WANG, YUJIA; HUANG, JIN-LONG

    2016-01-01

    Polyacrylamide hydrogel (PAAG) has been used for several years as an injectable implant for augmentation mammoplasty in China. Although patients who received PAAG injections experienced a number of complications, breast cancer following PAAG injection has been reported only in two cases. In this report, we present a case of breast cancer following PAAG injection for breast augmentation. Our study demonstrated that PAAG injection may increase the risk of breast cancer development. Early-stage breast cancer detection is difficult, since the breast is covered with the indurated injected gel. Thus, PAAG injection for augmentation mammoplasty may negatively affect breast cancer diagnosis and prognosis.

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

    PubMed Central

    Yao, Yao; Chen, Fei; Chen, Xi; Shen, Qiang; Zhang, Lianmeng

    2016-01-01

    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. PMID:26933668

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

    PubMed

    Yao, Yao; Chen, Fei; Chen, Xi; Shen, Qiang; Zhang, Lianmeng

    2016-06-01

    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. PMID:26933668

  18. Fundamentals of Polymer Gel Dosimeters

    NASA Astrophysics Data System (ADS)

    McAuley, Kim B.

    2006-12-01

    The recent literature on polymer gel dosimetry contains application papers and basic experimental studies involving polymethacrylic-acid-based and polyacrylamide-based gel dosimeters. The basic studies assess the relative merits of these two most commonly used dosimeters, and explore the effects of tetrakis hydroxymethyl phosphonium chloride (THPC) antioxidant on dosimeter performance. Polymer gel dosimeters that contain THPC or other oxygen scavengers are called normoxic dosimeters, because they can be prepared under normal atmospheric conditions, rather than in a glove box that excludes oxygen. In this review, an effort is made to explain some of the underlying chemical phenomena that affect dosimeter performance using THPC, and that lead to differences in behaviour between dosimeters made using the two types of monomer systems. Progress on the development of new more effective and less toxic dosimeters is also reported.

  19. Composite hydrogels of polyacrylamide and crosslinked pH-responsive micrometer-sized hollow particles.

    PubMed

    Pafiti, Kyriaki; Cui, Zhengxing; Carney, Louise; Freemont, Anthony J; Saunders, Brian R

    2016-01-20

    Whilst hydrogels and hollow particles both continue to attract much attention in the literature there are few examples of hydrogel composites containing hollow particles. Here, we study composite polyacrylamide (PAAm) hydrogels containing micrometer-sized pH-responsive shell-crosslinked hollow particles (abbreviated as HPXL) based on poly(methylmethacrylate-co-methacrylic acid) functionalised with glycidyl methacrylate (GMA). The HPXL particles were prepared using our scaleable emulsion template method and inclusion of GMA was found to promote spherical hollow particle formation. The pendant vinyl groups from GMA enabled shell-crosslinked hollow particles to be prepared prior to formation of the PAAm/HPXL composite gels. The morphologies of the particles and composite gels were studied by optical microscopy, confocal laser scanning microscopy and scanning electron microscopy. Dynamic rheology measurements for the composite gels showed that the modulus variation with HPXL concentration could be described by a percolation model with a HPXL percolation threshold concentration of 4.4 wt% and a scaling exponent of 2.6. The composite gels were pH-responsive and largely maintained their mechanical properties over the pH range 4.0 to 8.0. Because the composite gels had tuneable mechanical properties (with modulus values up to 530 kPa) and were pH-responsive they are potential candidates for future wound healing or membrane applications. PMID:26610808

  20. Effect of ?-carrageenan on volume phase transition for polyacrylamide (PAAm) hydrogel using the fluorescence technique

    NASA Astrophysics Data System (ADS)

    Akta?, Demet Kaya

    2014-03-01

    Steady-state fluorescence (SSF) technique was employed for studying swelling of polyacrylamide (PAAm) gels with various content of ?-carrageenan ( ?C). Disc shaped composite hydrogels were prepared by free-radical crosslinking copolymerization of acrylamide (AAm) with various amounts ?C. N, N'-methylenebis (acrylamide) (BIS) and ammonium persulfate (APS) were used as crosslinker and initiator, respectively. Pyranine was introduced as a fluorescence probe. Fluorescence intensity of pyranine was monitored during in situ swelling processes of composite gels. It was observed that fluorescence intensity values decreased as swelling is proceeded. Li-Tanaka equation was used to determine the swelling time constants, ? and cooperative diffusion coefficients, D from intensity variations during the swelling processes. It was shown that swelling time constants, ? decreased and diffusion coefficients, D increased as the ?C content in the composites are increased.

  1. 21 CFR 872.3420 - Carboxymethylcellulose sodium and cationic polyacrylamide polymer denture adhesive.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 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...

  2. 21 CFR 872.3420 - Carboxymethylcellulose sodium and cationic polyacrylamide polymer denture adhesive.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 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...

  3. 21 CFR 872.3420 - Carboxymethylcellulose sodium and cationic polyacrylamide polymer denture adhesive.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 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...

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

    Code of Federal Regulations, 2012 CFR

    2012-04-01

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

    Code of Federal Regulations, 2010 CFR

    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...

  6. Topological effects on viscoelasticity of polyacrylamide hydrogels

    NASA Astrophysics Data System (ADS)

    Kalfus, Jan; Lesser, Alan

    2011-03-01

    Viscoelastic behavior of long linear chains in a concentrated solution is governed by the topology of the molecules and interchain excluded volume interaction. As a consequence, chain diffusive motion is significantly retarded and such an assembly of chains exhibits highly pronounced entropy elastic behavior. In this contribution, two types of additional chain confinements imposed on a concentrated solution of linear polyacrylamide (L-PAA) will be discussed. The confinement was realized either by adding silica nano-filler into the concentrated solution of L-PAA or by cross-linking of acrylamide in the concentrated solution of L-PAA. While in the first case the trapped entanglement interaction is caused by interaction of chains with large nano-filler surface, in the second case the L-PAA chains are trapped among the cross-links of the PAA network. Viscoelastic response of both types of composite systems exhibited generic characteristics. In both cases, the trapped entanglement interaction significantly changed the relaxation spectrum of the matrix polymer solution and considerably enhanced the linear elastic modulus.

  7. Microbial degradation of polyacrylamide by aerobic granules.

    PubMed

    Liu, Lili; Wang, Zhiping; Lin, Kuangfei; Cai, Weimin

    2012-01-01

    To deal with polyacrylamide (PAM) wastewater, granular sludge formed in glucose-fed sequencing batch reactors (SBR) was employed to cultivate PAM-degrading granules. Three replicated SBRs were operated with increasing PAM concentration in the influent from 67 to 670 mg L(-1), and the hydraulic retention time was increased at the same time from 1 d to 6 d during the six-phase of the 43 d acclimation period. The well-acclimated PAM-degrading granules were different from the seeding granules in colour, mean diameter, biomass density and settle ability, and could use PAM as the sole carbon and nitrogen source. In the batch experiments, PAM degradation rate by granules was determined as 2.23 mg PAM g(-1) MLSS h(-1). According to the analysis of the intermediates of PAM biodegradation, PAM was degraded initially through hydrolysis of the amide group, and no acrylamide monomer was detected. With the help of LC/MS, the main intermediate was identified as polyacrylic acid with a low molecular weight. Therefore, the PAM-degrading granular sludge may be employed for removing PAM in the wastewater produced from tertiary oil recovery that uses polymeric flooding technology. PMID:22720433

  8. Preparation of tritium-labeled optical isomers of amino acids by ligand exchange chromatography on polyacrylamide sorbent containing L-phenylalanine groupings

    SciTech Connect

    Zolotarev, Yu.A.; Penkina, V.I.; Dostavalov, I.N.; Myasoedov, N.F.

    1988-11-01

    Tritium-labeled optically active amino acids are obtained by resolving racemates of the corresponding amino acids by chromatography on a chiral polyacrylamide sorbent, filled with copper ions. The chiral sorbent is obtained by the action of formaldehyde and L-phenylalanine on a Biogel P-4 polyacrylamide gel in an alkaline medium. Data are given on the ligand exchange chromatography of amino acids on this sorbent, depending on the degree of filling of the sorbent by copper ions and the concentration of the eluent. Conditions were selected for the quantitative resolution of racemates of amino acids and examples are given of a preparative obtaining of tritium labeled optical isomers of amino acids.

  9. Improving immobilized biocatalysts by gel phase polymerization

    SciTech Connect

    Kuu, W.Y.; Polack, J.A.

    1983-08-01

    A new method is presented for the treatment of gel-type supports, used for immobilizing microbial cells and enzymes, to obtain high mechanical strength. It is particularly useful for ethanol fermentation over gel beads containing immobilized viable cells, where the beads can be ruptured by gas production and the growth of cells within the gels. This method consists of treating agar or carrageenan gel with polyacrylamide to form a rigid support which retains the high catalytic activity characteristic of the untreated biocatalysts. The size and shape of the biocatalyst is unaffected by this treatment. The method involves the diffusion of acrylamide, N,N'-methylenebisacrylamide and BETA-dimethylaminopropionitrile (or N,N,N',N'-tetramethylethylenediamine) into the preformed biocatalyst beads followed by the addition of an initiator to cause polymerization within the beads. Treated gels have been used for the continuous fermentation of glucose to ethanol in a packed column for over two months.

  10. Photoswitchable gel assembly based on molecular recognition

    PubMed Central

    Yamaguchi, Hiroyasu; Kobayashi, Yuichiro; Kobayashi, Ryosuke; Takashima, Yoshinori; Hashidzume, Akihito; Harada, Akira

    2012-01-01

    The formation of effective and precise linkages in bottom-up or top-down processes is important for the development of self-assembled materials. Self-assembly through molecular recognition events is a powerful tool for producing functionalized materials. Photoresponsive molecular recognition systems can permit the creation of photoregulated self-assembled macroscopic objects. Here we demonstrate that macroscopic gel assembly can be highly regulated through photoisomerization of an azobenzene moiety that interacts differently with two host molecules. A photoregulated gel assembly system is developed using polyacrylamide-based hydrogels functionalized with azobenzene (guest) or cyclodextrin (host) moieties. Reversible adhesion and dissociation of the host gel from the guest gel may be controlled by photoirradiation. The differential affinities of ?-cyclodextrin or ?-cyclodextrin for the trans-azobenzene and cis-azobenzene are employed in the construction of a photoswitchable gel assembly system. PMID:22215078

  11. Field demonstration of in situ grouting of radioactive solid waste burial trenches with polyacrylamide. [Polyacrylamide

    SciTech Connect

    Spalding, B.P.; Fontaine, T.A.

    1990-01-01

    Demonstrations of in situ grouting with polyacrylamide were carried out on two undisturbed burial trenches and one dynamically compacted burial trench in Solid Waste Storage Area (SWSA) 6 at Oak Ridge National Laboratory (ORNL). The injection of polyacrylamide was achieved quite facilely for the two undisturbed burial trenches which were filled with grout, at typical pumping rates of 95 L/min, in several batches injected over several days. The compacted burial trench, however, failed to accept grout at more than 1.9 L/min even when pressure was applied. Thus, it appears that burial trenches, stabilized by dynamic compaction, have a permeability too low to be considered groutable. The water table beneath the burial trenches did not respond to grout injections indicating a lack of hydrologic connection between fluid grout and the water table which would have been observed if the grout failed to set. Because grout set times were adjusted to less than 60 min, the lack of hydrologic connection was not surprising. Postgrouting penetration testing revealed that the stability of the burial trenches was increased from 26% to 79% that measured in the undisturbed soil surrounding the trenches. In situ permeation tests on the grouted trenches indicated a significant reduction in hydraulic conductivity of the trench contents from a mean of 2.1 {times} 10{sup {minus}3} to 1.85 {times} 10{sup {minus}5} cm/s. Preliminary observations indicated that grouting with polyacrylamide is an excellent method for both improved stability and hydrologic isolation of radioactive waste and its incidental hazardous constituents.

  12. In situ grouting of buried transuranic waste with polyacrylamide

    SciTech Connect

    Spalding, B.P.; Lee, S.Y.; Farmer, C.D.; Hyder, L.K.; Supaokit, P.

    1987-01-01

    This project is a demonstration and evaluation of the in situ hydrologic stabilization of buried transuranic waste at a humid site via grout injection. Two small trenches, containing buried transuranic waste, were filled with 34.000 L of polyacrylamide grout. Initial field results have indicated that voids within the trenches were totally filled by the grout and that the intratrench hydraulic conductivity was reduced to below field-measurable values. No evidence of grout constituents were observed in twelve perimeter groundwater monitoring wells indicating that grout was contained completely within the two trenches. Polyacrylamide grout was selected for field demonstration over the polyacrylate grout due to its superior performance in laboratory degradation studies. Also supporting the selection of polyacrylamide was the difficulty in controlling the set time of the acrylate polymerization. Based on preliminary degradation monitoring, the polyacrylamide was estimated to have a microbiological half-life of 362 years in the test soil. 15 refs., 9 figs., 12 tabs.

  13. Silver staining of 2D electrophoresis gels.

    PubMed

    Rabilloud, Thierry

    2012-01-01

    Silver staining is used to detect proteins after electrophoretic separation on polyacrylamide gels. It -combines excellent sensitivity (in the low nanogram range) with the use of very simple and cheap equipment and chemicals. For its use in proteomics, two important additional features must be considered, compatibility with mass spectrometry and quantitative response. Both features are discussed in this chapter, and optimized silver staining protocols are proposed. PMID:22665294

  14. Effect of cationic polyacrylamide on the processing and properties of nanocellulose films.

    PubMed

    Raj, Praveena; Varanasi, Swambabu; Batchelor, Warren; Garnier, Gil

    2015-06-01

    The use of high molecular weight cationic polyacrylamide (CPAM) was investigated to accelerate the drainage of nanocellulose (Microfibrillated Cellulose) suspensions into films. The mechanism was quantified and optimized by measuring the gel point, the lowest solids concentration at which a continuous network is formed. The flocculation of MFC was analysed as a function of the polyelectrolyte dosage, charge density and molecular weight as well as process parameters (drainage time) and material properties. The adsorption isotherms of CPAMs on nanocellulose and their zeta potential curves were also analysed as a function of CPAM charge and dosage. Measured CPAM adsorption capacities for the 50% and 10% charged 13MDa CPAM onto MFC were 5mg/g and 8mg/g, respectively, corresponding to adsorption coverage on cellulose of 0.14mg/m(2) and 0.22mg/m(2). The floc strength and drainability of MFC suspensions were quantified with the gel point as a function of CPAM properties. For all combinations of polyelectrolyte molecular weight and charge density, the gel point of a nanocellulose suspension goes through a minimum with increasing polymer dosage. The minimum gel point was independent of the polyelectrolyte charge density at constant molecular weight. However, it reduced with decreasing CPAM molecular weight, at a constant addition rate. The drainage time of a nanocellulose suspension into a film is reduced by 2/3 by halving the gel point from 0.2 to 0.1kg/m(3); this is due to the more flocculated suspension facilitating drainage between flocs. Nanocellulose films of increased porosity also result from reducing the gel point, signifying that the more open 3D structure of the flocculated cellulose suspension is retained upon drying the 2D film cellulose film structure. PMID:25702868

  15. Antioxidant effect of green tea on polymer gel dosimeter

    NASA Astrophysics Data System (ADS)

    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.

  16. Site blocking effects on adsorbed polyacrylamide conformation

    NASA Astrophysics Data System (ADS)

    Brotherson, Brett A.

    The use of polymers as flocculating additives is a common practice in many manufacturing environments. However, exactly how these polymers interact with surfaces is relatively unknown. One specific topic which is thought to be very important to flocculation is an adsorbed polymer's conformation. Substantial amounts of previous work, mainly using simulations, have been performed to elucidate the theory surrounding adsorbed polymer conformations. Yet, there is little experimental work which directly verifies current theory. In order to optimize the use of polymer flocculants in industrial applications, a better understanding of an adsorbed polymer's conformation on a surface beyond theoretical simulations is necessary. This work looks specifically at site blocking, which has a broad impact on flocculation, adsorption, and surface modification, and investigated its effects on the resulting adsorbed polymer conformation. Experimental methods which would allow direct determination of adsorbed polymer conformational details and be comparable with previous experimental results were first determined or developed. Characterization of an adsorbed polymer's conformation was then evaluated using dynamic light scattering, a currently accepted experimental technique to examine this. This commonly used technique was performed to allow the comparison of this works results with past literature. Next, a new technique using atomic force microscopy was developed, building on previous experimental techniques, to allow the direct determination of an adsorbed polymer's loop lengths. This method also was able to quantify changes in the length of adsorbed polymer tails. Finally, mesoscopic simulation was attempted using dissipative particle dynamics. In order to determine more information about an adsorbed polymer's conformation, three different environmental factors were analyzed: an adsorbed polymer on a surface in water, an adsorbed polymer on a surface in aqueous solutions of varying ionic strength, and an adsorbed polymer on a surface functionalized with site blocking additives. This work investigated these scenarios using a low charge density high molecular weight cationic polyacrylamide. Three different substrates, for polymer adsorption were analyzed: mica, anionic latex, and glass. It was determined that, similar to previous studies, the adsorbed polymer layer thickness in water is relatively small even for high molecular weight polymers, on the order of tens of nanometers. The loop length distribution of a single polymer, experimentally verified for the first time, revealed a broad span of loop lengths as high as 1.5 microns. However, the bulk of the distribution was found between 40 and 260 nanometers. For the first time, previous theoretical predictions regarding the salt effect on adsorbed polymer conformation were confirmed experimentally. It was determined that the adsorbed polymer layer thickness expanded with increasing ionic strength of the solvent. Using atomic force microscopy, it was determined that the adsorbed polymer loop lengths and tail lengths increased with increasing ionic strength, supporting the results found using dynamic light scattering. The effect of the addition of site blocking additives on a single polymer's conformation was investigated for the first time. It was determined that the addition of site blocking additives caused strikingly similar results as the addition of salt to the medium. The changes in adsorbed polymer's loop lengths was found to be inconsistent and minimal. However, the changes in an adsorbed polymer's free tail length was found to increase with increasing site blocking additive levels. These results were obtained using either PDADMAC or cationic nanosilica as site blocking additives.

  17. Use of recombinant DNA derived human relaxin to probe the structure of the native protein

    SciTech Connect

    Canova-Davis, E.; Kessler, T.J.; Lee, P.J.; Fie, D.T.W.; Griffin, P.; Stults, J.T.; Rinderknecht, E. ); Wade, J.D. )

    1991-06-18

    This report describes the physical, chemical, and biological characterization of recombinant human relaxin (rhRlx) used as a probe to establish the disulfide pairing in native human relaxin. This strategy is necessary since native human relaxin is only available in the nanogram range. The relaxin molecule is composed of two nonidentical peptide chains, an A-chain 24 amino acids in length and a B-chain of 29 amino acids, linked by two disulfide bridges with an additional disulfide linkage in the A-chain. Native relaxin isolated from human corpora lutea was compared to rhRlx by reversed-phase chromatography, partial sequence analysis, mass spectroscopy, and bioassay. The potency of rhRlx was established by its ability to stimulate cAMP from primary human uterine endometrial cells. Native relaxin isolated from human corpora lutea was euqipotent to chemically synthesized relaxin, which in turn was equipotent to rhRlx. A tryptic map was developed for rhRlx to confirm the completer amino acid sequence and assignment of the disulfide bonds. The three disulfide bonds (Cys{sup A10}-Cys{sup A15}, Cys{sup A11}-Cys{sup B11}, and Cys{sup A24}-Cys{sup B23}) were assigned by mass spectrometric analysis of the tryptic peptides and by comparison to chemically synthesized peptides disulfide linked in the two most probable configurations. In addition, the observed amino acid composition and sequence of rhRlx was in agreement with that predicted from the cDNA sequence with the exception that the A-chain amino terminal was pyroglutamic acid. The migration of rhRlx upon sodium dodecyl sulfate-polyacrylamide gel electrophoresis was consistent with a monomeric structure, and the identity of the band was demonstrated by immunoblotting.

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

    PubMed

    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) PMID:1451680

  19. Biomimetic alginate/polyacrylamide porous scaffold supports human mesenchymal stem cell proliferation and chondrogenesis.

    PubMed

    Guo, Peng; Yuan, Yasheng; Chi, Fanglu

    2014-09-01

    We describe the development of alginate/polyacrylamide (ALG/PAAm) porous hydrogels based on interpenetrating polymer network structure for human mesenchymal stem cell proliferation and chondrogenesis. Three ALG/PAAm hydrogels at molar ratios of 10/90, 20/80, and 30/70 were prepared and characterized with enhanced elastic and rubbery mechanical properties, which are similar to native human cartilage tissues. Their elasticity and swelling properties were also studied under different physiological pH conditions. Finally, in vitro tests demonstrated that human mesenchymal stem cells could proliferate on the as-synthesized hydrogels with improved alkaline phosphatase activities. These results suggest that ALG/PAAm hydrogels may be a promising biomaterial for cartilage tissue engineering. PMID:25063162

  20. The fundamental radiation properties of normoxic polymer gel dosimeters: a comparison between a methacrylic acid based gel and acrylamide based gels

    NASA Astrophysics Data System (ADS)

    DeDeene, Y.; Vergote, K.; Claeys, C.; DeWagter, C.

    2006-02-01

    Polymer gel dosimeters offer a wide range of applications in the three-dimensional verification of complex dose distributions such as in intensity-modulated radiotherapy. One of the major difficulties with polymer gel dosimeters is their sensitivity to oxygen, as oxygen inhibits the radiation-induced polymerization reaction. For several years, oxygen was removed from the gels by bubbling the sol with inert gases for several hours during the gel fabrication. Also, the gel had to be poured in containers with low oxygen permeability and solubility. Recently, it was found that these technical difficulties can easily be solved by adding an antioxidant to the gel. These gels are called 'normoxic' gels as they can be produced under normal atmospheric conditions. In this study several properties of polymer gel dosimeters have been investigated: the dose sensitivity, the temporal and spatial stability of the gel, the sensitivity of the dose response to temperature during irradiation and during MR imaging, the energy dependence and the dose-rate dependence. This study reveals that the normoxic polymer gel dosimeter based on methacrylic acid (nMAG) studied in this work has inferior radiation properties as compared to the polyacrylamide gelatine (PAG) gel dosimeters. It is shown that from the three different gel dosimeters investigated in this study, the nPAG gel dosimeter results in a less sensitive gel dosimeter but with superior radiation properties as compared to the nMAG gel dosimeter. The importance of investigating relevant radiation properties of gel dosimeters apart from the radiation sensitivityprior to their use for dosimetric validation experimentsis illustrated and emphasized throughout this study. Other combinations of monomer and gelling agent may result in more reliable normoxic polymer gel dosimeters.

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

    PubMed

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

    2015-08-01

    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. PMID:26320809

  2. Polyelectrolyte gels

    SciTech Connect

    Segalman, D.J.; Witkowski, W.R.

    1995-06-01

    Polyelectrolyte (PE) gels are swollen polymer/solvent networks that undergo a reversible volume collapse/expansion through various types of stimulation. Applications that could exploit this large deformation and solvent expulsion/absorption characteristics include robotic {open_quotes}fingers{close_quotes} and drug delivery systems. The goals of the research were to first explore the feasibility of using the PE gels as {open_quotes}smart materials{close_quotes} - materials whose response can be controlled by an external stimulus through a feedback mechanism. Then develop a predictive capability to simulate the dynamic behavior of these gels. This involved experimentally characterizing the response of well-characterized gels to an applied electric field and other stimuli to develop an understanding of the underlying mechanisms which cause the volume collapse. Lastly, the numerical analysis tool was used to simulate various potential engineering devices based on PE gels. This report discusses the pursuit of those goals through experimental and computational means.

  3. Improving gel properties of hairtail surimi by electron irradiation

    NASA Astrophysics Data System (ADS)

    Lin, Xianping; Yang, Wenge; Xu, Dalun; Jie, Zhen; Liu, Wen

    2015-05-01

    Hairtail surimi was subjected to electron irradiation for doses up to 9 kGy. At 7 kGy highest gel strength was achieved. The irradiation also increased lightness and expressible water amount. Scanning electron micrographs showed that 7 kGy irradiation made the surimi protein gel network more compact. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) revealed the degradation of myosin heavy chain (MHC) as the irradiation dose increased, particularly at the 7 and 9 kGy doses. Radiation processing may become a new effective tool in surimi production.

  4. PREPARATION OF STARCH-GRAFT-POLYACRYLAMIDE COPOLYMERS BY REACTIVE EXTRUSION

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Graft copolymers of starch and polyacrylamide (PAAm) were prepared by reactive extrusion using a co-rotating twin screw extruder and ammonium persulfate initiator. Feed rates were 109 g/min to 325 g/min (all components) at a moisture content of 50%, with screw speeds in the range 100 rpm to 300 rpm...

  5. Acrylamide monomer leaching from polyacrylamide-treated irrigation furrows

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Water-soluble polyacrylamide (WSPAM), used to reduce erosion in furrow irrigated fields and other agriculture applications, contain less than 0.05% Acrylamide monomer (AMD). The AMD, a potent neurotoxicant and suspected carcinogen, is readily dissolved and transported in flowing water. Deep percol...

  6. PREPARATION OF STARCH-G-POLYACRYLAMIDE COPOLYMERS BY REACTIVE EXTRUSION

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Graft copolymers of starch and polyacrylamide were prepared by reactive extrusion using a co-rotating twin screw extruder and ammonium persulfate initiator. Feed rates were 109 g/min up to 325 g/min (all components) at a moisture content of 50 percent, with screw speeds in the range 100 rpm to 300 ...

  7. 21 CFR 173.10 - Modified polyacrylamide resin.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 3 2014-04-01 2014-04-01 false Modified polyacrylamide resin. 173.10 Section 173.10 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED... clarification of beet or cane sugar juice in an amount not exceeding 5 parts per million by weight of the...

  8. Polyacrylamide molecular weight effects on soil infiltration and erosion

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Seal formation at the surface of smectitic 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 ...

  9. POLYACRYLAMIDE (PAM) IN AGRICULTURE AND ENVIRONMENTAL LAND MANAGEMENT

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Anionic polyacrylamide (PAM) has been sold since 1995 to reduce irrigation-induced erosion and enhance infiltration. Its soil stabilizing and flocculating properties improve runoff water quality by reducing sediments, N, dissolved reactive P (DRP) and total P, chemical oxygen demand (COD), pesticid...

  10. HRP-Mediated Synthesis of Starch-Polyacrylamide Graft Copolymers

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Modified starch-based polymers can be engineered for specific properties by combining starch with synthetic polymers through graft copolymerization. Polyacrylamide grafted starch have received a great deal of applications in areas such as superabsorbent paper-making additives, drag reduction and te...

  11. POLYACRYLAMIDE EFFECTS ON WATER INFILTRATION IN SANDY LOAM SOILS

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Some sandy soils of the California San Joaquin Valley have low water infiltration. Electrical conductivity (EC) and sodium adsorption ratio (SAR) of irrigation water greatly affect infiltration rate and hydraulic conductivity of soils. High molecular weight polyacrylamides (PAM) have been shown to i...

  12. 40 CFR 721.10700 - Polyfluorinated alkyl thio polyacrylamide (generic).

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... (CONTINUED) TOXIC SUBSTANCES CONTROL ACT SIGNIFICANT NEW USES OF CHEMICAL SUBSTANCES Significant New Uses for Specific Chemical Substances § 721.10700 Polyfluorinated alkyl thio polyacrylamide (generic). (a) Chemical substances and significant new uses subject to reporting. (1) The chemical substances identified...

  13. 21 CFR 173.10 - Modified polyacrylamide resin.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ...) FOOD FOR HUMAN CONSUMPTION (CONTINUED) SECONDARY DIRECT FOOD ADDITIVES PERMITTED IN FOOD FOR HUMAN CONSUMPTION Polymer Substances and Polymer Adjuvants for Food Treatment 173.10 Modified polyacrylamide resin... resin is used as a flocculent in the clarification of beet or cane sugar juice in an amount...

  14. 21 CFR 173.10 - Modified polyacrylamide resin.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 3 2011-04-01 2011-04-01 false Modified polyacrylamide resin. 173.10 Section 173.10 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) FOOD FOR HUMAN CONSUMPTION (CONTINUED) SECONDARY DIRECT FOOD ADDITIVES PERMITTED IN FOOD FOR HUMAN CONSUMPTION Polymer Substances and...

  15. 21 CFR 173.10 - Modified polyacrylamide resin.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ...) FOOD FOR HUMAN CONSUMPTION (CONTINUED) SECONDARY DIRECT FOOD ADDITIVES PERMITTED IN FOOD FOR HUMAN CONSUMPTION Polymer Substances and Polymer Adjuvants for Food Treatment 173.10 Modified polyacrylamide resin... resin is used as a flocculent in the clarification of beet or cane sugar juice in an amount...

  16. Assessing Kinase Activity in Plants with In-Gel Kinase Assays.

    PubMed

    Wang, Pengcheng; Zhu, Jian-Kang

    2016-01-01

    The in-gel protein kinase assay is a powerful method to measure the protein phosphorylation activity of specific protein kinases. Any protein substrate can be embedded in polyacrylamide gels where they can be phosphorylated by protein kinases that are separated in the gel under denaturing conditions and then renatured. The kinase activity can be visualized in situ in the gels by autoradiography. This method has been used to compare the activities of protein kinases in parallel samples or to identify their potential substrates. Here, we describe in detail an in-gel kinase assay to measure the activity of some protein kinases in plants. PMID:26577790

  17. A simple system for staining protein and nucleic acid electrophoresis gels.

    PubMed

    Raymer, Dorian M; Smith, Douglas E

    2007-03-01

    Researchers in molecular biology spend a significant amount of time tending to the staining and destaining of electrophoresis gels. Here we describe a simple system, costing approximately $100 and taking approximately 1 h to assemble, that automates standard nucleic acid and protein gel staining protocols. Staining is done in a tray or, with DNA gels, in the electrophoresis chamber itself following automatic detection of the voltage drop. Miniature pumps controlled by a microcontroller chip exchange the necessary solutions at programmed time intervals. We demonstrate efficient and highly reproducible ethidium bromide and methylene blue staining of DNA in agarose gels and Coomassie blue and silver staining of proteins in polyacrylamide gels. PMID:17265540

  18. Improving immobilized biocatalysts by gel phase polymerization

    SciTech Connect

    Kuu, W.Y.; Polack, J.A.

    1983-08-01

    A new method is presented for the treatment of gel-type supports, used for immobilizing microbial cells and enzymes, to obtain high mechanical strength. It is particularly useful for ethanol fermentation over gel beads containing immobilized viable cells, where the beads can be ruptured by gas production and the growth of cells within the gels. This method consists of treating agar or carrageenan gel with polyacrylamide to form a rigid support which retains the high catalytic activity characteristic of the untreated biocatalysts. The size and shape of the biocatalyst is unaffected by this treatment. The method involves the diffusion of acrylamide, N,N'-methylenebisacrylamide and ..beta..-dimethylaminopropionitrile (or N,N,N',N'-tetramethylethylenediamine) into the preformed biocatalyst beads followed by the addition of an initiator to cause polymerization within the beads. Treated gels have been used for the continuous fermentation of glucose to ethanol in a packed column for over two months. During this operation, the gel beads maintained their rigidity, and the maximum productivity was as high as 50 gh/sup -1/ L/sup -1/ gel. There was no appreciable decay of cell activity.

  19. Amperometric biosensor for the detection of hydrogen peroxide using catalase modified electrodes in polyacrylamide.

    PubMed

    Varma, Shailly; Mattiasson, Bo

    2005-09-23

    A simple biosensor for the detection of hydrogen peroxide in organic solvents has been developed and coupled to a flow injection analysis (FIA) system. Catalase was entrapped in polyacrylamide gel and placed on the surface of platinum (working electrode) fixed in a Teflon holder with Ag-wire (auxiliary electrode), followed by addition of filter paper soaked in KCl. The entrapped catalase gel was held on the electrode using membranes. The effects of cellulose and polytetrafluroethylene (PTFE) membranes on the electrode response towards hydrogen peroxide have been studied. The modified electrode has been used to study the detection of hydrogen peroxide in solvents like water, dimethyl sulfoxide (DMSO), and 1,4-dioxane using amperometric techniques like cyclic voltammetry (CV) and FIA. The CV of modified catalase electrode showed a broad oxidation peak at -150 mV and a clear reduction peak at -212 mV in the presence of hydrogen peroxide. Comparison of CV with hydrogen peroxide in various solvents has been carried out. The electrode showed an irreversible kinetics with DMSO as the solvent. A flow cell has been designed in order to carry on FIA studies to obtain calibration plots for hydrogen peroxide with the modified electrode. The calibration plots in several solvents such as water, dimethyl sulfoxide, 1,4-dioxane have been obtained. The throughput of the enzyme electrode was 10 injections per hour. Due to the presence of membrane the response time of the electrode is concentration dependent. PMID:16099064

  20. Effect of gel structure of matrix orientation in pulsed alternating electric fields

    SciTech Connect

    Stellwagen, N.C.; Stellwagen, J.

    1993-12-31

    Four polymeric gels with different structures, LE agarose, HEEO agarose, beta-carrageenan, and polyacrylamide, were studied by transient electric birefringence to determine the importance of various structural features on the orientation of the gels in pulsed alternating electric fields. The birefrigence relaxation times observed for agarose gels in low voltage electric fields suggest that long fibers and/or domains, ranging up to tens of microns in size, are oriented by the electric field. The sign of the birefringence reverses when the direction of the electric field is reversed, suggesting that the oriented domains change their direction of orientation from parallel to perpendicular (or vice versa) when the polarity of the electric field is reversed. These anamalous orientation effects are observed with both types of agarose gels, but not with beta-carrageenan or polyacrylamide gels, suggesting that the alternating D,L galactose residues in the agarose backbone are responsible for the anomalies.

  1. Human leucocyte aspartylglucosaminidase. Evidence for two different subunits in a more complex native structure.

    PubMed Central

    Halila, R; Baumann, M; Ikonen, E; Enomaa, N; Peltonen, L

    1991-01-01

    Human leucocyte aspartylglucosaminidase (AGA: 1-aspartamido-beta-N-acetylglucosamine amidohydrolase, EC 3.5.1.26) was purified to homogeneity by using affinity chromatography, gel filtration, chromatofocusing and reverse-phase h.p.l.c. As shown by SDS/PAGE, the homogeneous purified enzyme preparation consists of four polypeptide chains with molecular masses of 25, 24, 18 and 17 kDa. In the native polyacrylamide gel these polypeptides migrate as one active enzyme complex, and by gel filtration the peak of enzyme activity can be detected in a position of about 65 kDa. Digestion with endoproteinase Lys-C or endoproteinase Asp-N, followed by peptide analysis with reverse-phase h.p.l.c., reveals an identical peptide pattern for the 24 and 25 kDa bands as well as for the 17 and 18 kDa bands. This treatment further demonstrated a totally different peptide pattern for the 24/25 kDa versus the 17/18 kDa subunit. The N-terminal sequences of the 17 kDa and the 18 kDa peptides were identical, as determined by Edman degradation. The N-termini of the 24 kDa and the 25 kDa peptides were blocked. The enzyme was partly resistant to endoglycosidases H and F, but N-glycosidase F transformed the 24/25 kDa band into one 23 kDa band and the 17/18 kDa band into one 16 kDa band. Also, immunological data obtained with antisera produced against these subunits showed that AGA consists of two non-identical polypeptides. Images Fig. 2. Fig. 3. Fig. 4. Fig. 6. Fig. 7. PMID:2039475

  2. Identification of peanut hybrids using microsatellite markers and horizontal polyacrylamide gel electrophoresis

    Technology Transfer Automated Retrieval System (TEKTRAN)

    In peanut hybridization, it is very important to be able to distinguish selfs from true hybrids to save time and resources. To help facilitate this effort and provide molecular distinction between selfs from hybrids, DNA was extracted from leaf tissue of F1 or F2 plants, and SSR markers were amplif...

  3. Rapid staining of proteins on polyacrylamide gels and nitrocellulose membranes using a mixture of fluorescent dyes.

    PubMed

    Ganesh, G; Kumar, T K; Pandian, S T; Yu, C

    2000-11-20

    The present work describes a novel, fluorescence-based method for staining proteins on SDS-PAGE and membrane(s). In this method, proteins are stained using a mixed-dye (sulfo-rhodamine B and 1-anilino-8-naphthalene sulfonic acid (NH(4)(+))) solution. The mixed-dye staining protocol can detect proteins up to a concentration of 15 ng. This method is generally applicable to all proteins and is more sensitive than the conventional Coomassie blue method. The staining method is rapid and efficient. Staining-destaining of proteins using the mixed-dye protocol takes less than half an hour. Another interesting feature of the staining protocol described here is the applicability to the staining of proteins on nitrocellulose membranes. PMID:11086192

  4. Application of multiple levels of fluid shear stress to endothelial cells plated on polyacrylamide gels

    PubMed Central

    Galie, P. A.; van Oosten, A.; Chen, C. S.

    2015-01-01

    Measurements of endothelial cell response to fluid shear stress have previously been performed on unphysiologically rigid substrates. We describe the design and implementation of a microfluidic device that applies discrete levels of shear stress to cells plated on hydrogel-based substrates of physiologicallyrelevant stiffness. The setup allows for measurements of cell morphology and inflammatory response to the combined stimuli, and identifies mechanisms by which vascular stiffening leads to pathological responses to blood flow. We found that the magnitude of shear stress required to affect endothelial cell morphology and inflammatory response depended on substrate stiffness. Endothelial cells on 100 Pa substrates demonstrate a greater increase in cell area and cortical stiffness and decrease in NF-?B nuclear translocation in response to TNF-? treatment compared to controls than cells plated on 10 kPa substrates. The response of endothelial cells on soft substrates to shear stress depends on the presence of hyaluronan (HA). These results emphasize the importance of substrate stiffness on endothelial function, and elucidate a means by which vascular stiffening in aging and disease can impact the endothelium. PMID:25573790

  5. A LOW-COST HIGH THROUGHPUT POLYACRYLAMIDE GEL ELECTROPHORESIS SYSTEM FOR GENOTYPING WITH MICROSATELLITE DNA MARKERS

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Microsatellite DNA markers are widely used in genetic research but the genotyping cost with this marker system is high and the throughput is limited with conventional methods. The objective of this paper is to introduce a low-cost, high-throughput system developed in our laboratories for the detect...

  6. Polyacrylamide gel electrophoretic characterization of isoenzymes in the marine algal genus Callithamnion.

    PubMed

    Russell, S J

    1989-11-01

    Techniques have been developed for the visualisation of a range of enzyme species extracted from field collected marine algal populations, to provide biochemical data in the form of zymogram patterns that closely reflect the genetic identity of these plants. These patterns provide an alternative basis for increasingly accurate recognition to complement that provided by morphology, thus helping to clarify the taxonomic problems which exist in the marine algal genus Callithamnion Lyngbye as currently conceived. PMID:2482175

  7. Hydrophobicity-induced prestaining for protein detection in polyacrylamide gel electrophoresis.

    PubMed

    Li, Zhe; Guan, Weijiang; Lu, Chao; Zhou, Xi-Rui; Luo, Shi-Zhong; You, Ying; Ouyang, Jin

    2016-02-01

    An AIE fluorescent surfactant has been first used to prestain protein by ultrastrong hydrophobic interaction between fluorescent surfactants and proteins, distinguishing from the most widely used poststaining strategies by employing AIE molecules with weak hydrophobic characteristics. A mixture of proteins with variable molecular weights has been detected. PMID:26771025

  8. Ultrasensitive Stain for Proteins in Polyacrylamide Gels Shows Regional Variation in Cerebrospinal Fluid Proteins

    NASA Astrophysics Data System (ADS)

    Merril, Carl R.; Goldman, David; Sedman, Sylvia A.; Ebert, Michael H.

    1981-03-01

    A new silver stain for electrophoretically separated polypeptides can be rapidly and easily used and can detect as little as 0.01 nanogram of protein per square millimeter. When employed with two-dimensional electrophoresis, it should permit qualitative and quantitative characterization of protein distributions in body fluids and tissues. It has been used to demonstrate regional variations in cerebrospinal fluid proteins.

  9. 21 CFR 872.3480 - Polyacrylamide polymer (modified cationic) denture adhesive.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 8 2013-04-01 2013-04-01 false Polyacrylamide polymer (modified cationic) denture... polymer (modified cationic) denture adhesive. (a) Identification. A polyacrylamide polymer (modified cationic) denture adhesive is a device composed of polyacrylamide polymer (modified cationic) intended...

  10. 21 CFR 872.3480 - Polyacrylamide polymer (modified cationic) denture adhesive.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 8 2011-04-01 2011-04-01 false Polyacrylamide polymer (modified cationic) denture... polymer (modified cationic) denture adhesive. (a) Identification. A polyacrylamide polymer (modified cationic) denture adhesive is a device composed of polyacrylamide polymer (modified cationic) intended...

  11. 21 CFR 872.3480 - Polyacrylamide polymer (modified cationic) denture adhesive.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 8 2012-04-01 2012-04-01 false Polyacrylamide polymer (modified cationic) denture... polymer (modified cationic) denture adhesive. (a) Identification. A polyacrylamide polymer (modified cationic) denture adhesive is a device composed of polyacrylamide polymer (modified cationic) intended...

  12. 21 CFR 872.3480 - Polyacrylamide polymer (modified cationic) denture adhesive.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 8 2014-04-01 2014-04-01 false Polyacrylamide polymer (modified cationic) denture... polymer (modified cationic) denture adhesive. (a) Identification. A polyacrylamide polymer (modified cationic) denture adhesive is a device composed of polyacrylamide polymer (modified cationic) intended...

  13. 21 CFR 872.3480 - Polyacrylamide polymer (modified cationic) denture adhesive.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Polyacrylamide polymer (modified cationic) denture... polymer (modified cationic) denture adhesive. (a) Identification. A polyacrylamide polymer (modified cationic) denture adhesive is a device composed of polyacrylamide polymer (modified cationic) intended...

  14. A smart pH responsive graphene/polyacrylamide complex via noncovalent interaction

    NASA Astrophysics Data System (ADS)

    Ren, Lulu; Liu, Tianxi; Guo, Juan; Guo, Shuzhong; Wang, Xiaoyan; Wang, Weizhi

    2010-08-01

    We report that the graphene sheets can be stably dispersed in water by hydrophobic interaction with polyacrylamide. Most interestingly, the resultant graphene-polyacrylamide complexes show a reversible pH responsive property although polyacrylamide itself does not possess such characteristics. This method opens up novel opportunities for the potential applications of graphene in intelligent sensors, biology, medicine, nanoelectronics and other relevant areas.

  15. Native genomic blotting: high-resolution mapping of DNase I-hypersensitive sites and protein-DNA interactions

    SciTech Connect

    Pauli, U.; Chrysogelos, S.; Stein, J.; Stein, G.

    1988-01-01

    DNase I-hypersensitive sites are observed in the promoter regions of actively expressed genes, potentially active genes, and genes that were once active. The authors have developed an approach that greatly increases the resolution for mapping these sites by electrophoresing genomic DNA on native polyacrylamide gels prior to electroblotting and hybridization. This improved method has been used to scan the promoter and coding region of a cell-cycle-dependent human histone H4 gene with an accuracy of +/- 5-10 base pairs. Protein-DNA interactions can be seen in the autoradiograph as light areas and DNase I-hypersensitive sites as dark bands. Therefore, this method provides a rapid and relatively simple means to accurately localize protein-DNA interactions as well as DNase I-hypersensitive sites, thus directly displaying DNase I hypersensitivity and protein-DNA complexes on one autoradiograph. It also potentially allows the analysis of small changes in DNase I-hypersensitive sites under various biological conditions. With this technique rather large regions of DNA can be screened to determine areas that should be analyzed by more sophisticated methods, such as genomic sequencing or gel retardation assays.

  16. Purification and characterization of particulate endothelium-derived relaxing factor synthase from cultured and native bovine aortic endothelial cells.

    PubMed Central

    Pollock, J S; Frstermann, U; Mitchell, J A; Warner, T D; Schmidt, H H; Nakane, M; Murad, F

    1991-01-01

    The particulate enzyme responsible for the synthesis of endothelium-derived relaxing factor has been purified from cultured and native (noncultured) bovine aortic endothelial cells. Purification of the solubilized particulate enzyme preparation by affinity chromatography on adenosine 2',5'-bisphosphate coupled to Sepharose followed by Superose 6 gel filtration chromatography resulted in a single protein band after denaturing polyacrylamide gel electrophoresis that corresponded to approximately 135 kDa. The enzyme activity in the various fractions was assayed by its stimulatory effect on soluble guanylyl cyclase of rat fetal lung fibroblasts (RFL-6 cells), by the formation of L-citrulline from L-arginine, by measuring nitrite/nitrate formation, and by bioassay on endothelium-denuded vascular strips. Endothelium-derived relaxing factor synthase was purified 3419-fold from the crude particulate fraction of cultured bovine aortic endothelial cells with a 12% recovery (RFL-6 assay). Purified endothelium-derived relaxing factor synthase required L-arginine, NADPH, Ca2+, calmodulin, and 5,6,7,8-tetrahydrobiopterin for full activity. Images PMID:1720542

  17. Effect of Polymer Hydration State on In-Gel Immunoassays.

    PubMed

    Vlassakis, Julea; Herr, Amy E

    2015-11-01

    Applications as diverse as drug delivery and immunoassays require hydrogels to house high concentration macromolecular solutions. Yet, thermodynamic partitioning acts to lower the equilibrium concentration of macromolecules in the hydrogel, as compared to the surrounding liquid phase. For immunoassays that utilize a target antigen immobilized in the hydrogel, partitioning hinders introduction of detection antibody into the gel and, consequently, reduces the in-gel concentration of detection antibody, adversely impacting assay sensitivity. Recently, we developed a single-cell targeted proteomic assay with polyacrylamide gel electrophoresis of single cell lysates followed by an in-gel immunoassay. In the present work, we overcome partitioning that both limits analytical sensitivity and increases consumption of costly detection antibody by performing the immunoassay step after dehydrating the antigen-containing polyacrylamide gel. Gels are rehydrated with a solution of detection antibody. We hypothesized that matching the volume of detection antibody solution with the hydrogel water volume fraction would ensure that, at equilibrium, the detection antibody mass resides in the gel and not in the liquid surrounding the gel. Using this approach, we observe (compared with antibody incubation of hydrated gels): (i) 4-11 fold higher concentration of antibody in the dehydrated gels and in the single-cell assay (ii) higher fluorescence immunoassay signal, with up to 5-fold increases in signal-to-noise-ratio and (iii) reduced detection antibody consumption. We also find that detection antibody signal may be less well-correlated with target protein levels (GFP) using this method, suggesting a trade-off between analytical sensitivity and variation in immunoprobe signal. Our volume-matching approach for introducing macromolecular solutions to hydrogels increases the local in-gel concentration of detection antibody without requiring modification of the hydrogel structure, and thus we anticipate broad applicability to hydrogel-based assays, diagnostics, and drug delivery. PMID:26457450

  18. Native Intelligence

    ERIC Educational Resources Information Center

    Seven, Richard

    2006-01-01

    Amid concerns from tribal leaders that No Child Left Behind testing is squeezing out electives that have traditionally covered their history and cultures, an ambitious brace of programs is making Native America part of the core curriculum at David Wolfle Elementary School and other schools in the western Washington State. By tapping into…

  19. Native Intelligence

    ERIC Educational Resources Information Center

    Seven, Richard

    2006-01-01

    Amid concerns from tribal leaders that No Child Left Behind testing is squeezing out electives that have traditionally covered their history and cultures, an ambitious brace of programs is making Native America part of the core curriculum at David Wolfle Elementary School and other schools in the western Washington State. By tapping into

  20. Immobilization of enzymes on alginic acid-polyacrylamide copolymers

    SciTech Connect

    Kumaraswamy, M.D.K.; Panduranga R.K.; Thomas J.K.; Santappa, M.

    1981-08-01

    In this report, the authors present initial results and limitations of a polymeric system for the immobilization of enzymes. Enzymes attached to insoluble polymers of natural and synthetic origin are gaining importance in many industrial and biomedical applications. Graft copolymers are used as enzyme supports and in this study a novel polymeric system of alginic acid-polyacrylamide graft copolymer is described which was used for immobilizing enzymes. (Refs. 4).

  1. Laser CT evaluation on normoxic PAGAT gel dosimeter

    NASA Astrophysics Data System (ADS)

    Kumar, D. S.; Samuel, E. J. J.; Watanabe, Y.

    2013-06-01

    Optical computed tomography has been shown to be a potentially useful imaging tool for the radiation therapy physicists. In radiation therapy, researchers have used optical CT for the readout of 3D dosimeters. The purpose of this paper is to describe the initial evaluation of a newly fabricated laser CT scanner for 3D gel dosimetry which works using the first generation principle. A normoxic PAGAT (Polyacrylamide Gelatin and Tetrakis) gel is used as a dosimeter for this analysis. When a laser passes through the gel phantom, absorption and scattering of photon take place. The optical attenuation coefficient of the laser can be obtained by measuring its intensity after passing through the gel by a sensor. The scanner motion is controlled by a computer program written in Microsoft Visual C++. Reconstruction and data analysis on the irradiated gel phantom is performed by suitable algorithm using Matlab software.

  2. 2D gel proteomics: an approach to study age-related differences in protein abundance or isoform complexity in biological samples.

    PubMed

    Kim, Helen; Eliuk, Shannon; Deshane, Jessy; Meleth, Sreelatha; Sanderson, Todd; Pinner, Anita; Robinson, Gloria; Wilson, Landon; Kirk, Marion; Barnes, Stephen

    2007-01-01

    This chapter describes protocols for two-dimensional (2D) gel electrophoresis (isoelectric focusing [IEF] followed by sodium-dodecyl sulfate (SDS)-polyacrylamide gel electro-phoresis [PAGE]), staining of gels with the fluorescent dye Sypro Ruby, 2D gel image analysis, peptide mass fingerprint (PMF) analysis using matrix-assisted laser desorption ionization (MALDI)-time-of-flight (TOF) mass spectrometry (MS), liquid chromatography (LC)-tandem mass spectrometry (MS/MS), Western blot analysis of protein oxidations, and mass spectrometric mapping of sites of protein oxidations. Many of these methods were used to identify proteins affected in rat brain following ingestion of grape seed extract (GSE), a dietary supplement touted for anti-oxidant activity. Although beneficial actions in cell and animal models of chronic disease have been described for GSE, it has not been shown whether specific proteins were affected, or the nature of the effects. Applying 2D gel proteomics technology allowed discovery of proteins targeted by GSE without a priori knowledge of which one(s) might be affected. The newer 2D blue native (BN) electrophoresis methodology, which resolves protein complexes in a nondenaturing first dimension and then the components of these complexes in a denaturing second dimension, is discussed as a complementary approach. Analysis of protein oxidations and protein-protein interactions have special relevance to aging-related research, since oxidative stress and altered protein interactions may be at the heart of aging-related diseases. Finally, quality control issues related to implementation of high throughput technologies are addressed, to underscore the importance of minimizing bias and randomizing human and technical error in generating large datasets that are expensive and time-consuming to repeat. PMID:17634592

  3. Reprogramming cellular phenotype by soft collagen gels.

    PubMed

    Ali, M Yakut; Chuang, Chih-Yuan; Saif, M Taher A

    2014-11-28

    A variety of cell types exhibit phenotype changes in response to the mechanical stiffness of the substrate. Many cells excluding neurons display an increase in the spread area, actin stress fiber formation and larger focal adhesion complexes as substrate stiffness increases in a sparsely populated culture. Cell proliferation is also known to directly correlate with these phenotype changes/changes in substrate stiffness. Augmented spreading and proliferation on stiffer substrates require nuclear transcriptional regulator YAP (Yes associated protein) localization in the cell nucleus and is tightly coupled to larger traction force generation. In this study, we show that different types of fibroblasts can exhibit spread morphology, well defined actin stress fibers, and larger focal adhesions even on very soft collagen gels (modulus in hundreds of Pascals) as if they are on hard glass substrates (modulus in GPa, several orders of magnitude higher). Strikingly, we show, for the first time, that augmented spreading and other hard substrate cytoskeleton architectures on soft collagen gels are not correlated with the cell proliferation pattern and do not require YAP localization in the cell nucleus. Finally, we examine the response of human colon carcinoma (HCT-8) cells on soft collagen gels. Recent studies show that human colon carcinoma (HCT-8) cells form multicellular clusters by 2-3 days when cultured on soft polyacrylamide (PA) gels with a wide range of stiffness (0.5-50 kPa) and coated with an extracellular matrix, ECM (collagen monomer/fibronectin). These clusters show limited spreading/wetting on PA gels, form 3D structures at the edges, and eventually display a remarkable, dissociative metastasis like phenotype (MLP), i.e., epithelial to rounded morphological transition after a week of culture on PA gels only, but not on collagen monomer coated stiff polystyrene/glass where they exhibit enhanced wetting and form confluent monolayers. Here, we show that HCT-8 cell clusters also show augmented spreading/wetting on soft collagen gels and eventually form confluent monolayers as on rigid glass substrates and MLP is completely inhibited on soft collagen gels. Overall, these results suggest that cell-material interactions (soft collagen gels in this case) can induce cellular phenotype and cytoskeleton organization in a remarkably distinct manner compared to a classical synthetic polyacrylamide (PA) hydrogel cell culture model and may contribute in designing new functional biomaterials. PMID:25284029

  4. The latest advancements in proteomic two-dimensional gel electrophoresis analysis applied to biological samples.

    PubMed

    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. PMID:25384742

  5. Aerosol gels

    NASA Technical Reports Server (NTRS)

    Sorensen, Christopher M. (Inventor); Chakrabarti, Amitabha (Inventor); Dhaubhadel, Rajan (Inventor); Gerving, Corey (Inventor)

    2010-01-01

    An improved process for the production of ultralow density, high specific surface area gel products is provided which comprises providing, in an enclosed chamber, a mixture made up of small particles of material suspended in gas; the particles are then caused to aggregate in the chamber to form ramified fractal aggregate gels. The particles should have a radius (a) of up to about 50 nm and the aerosol should have a volume fraction (f.sub.v) of at least 10.sup.-4. In preferred practice, the mixture is created by a spark-induced explosion of a precursor material (e.g., a hydrocarbon) and oxygen within the chamber. New compositions of matter are disclosed having densities below 3.0 mg/cc.

  6. Protein electrophoresis in agarose gels for separating high molecular weight proteins.

    PubMed

    Greaser, Marion L; Warren, Chad M

    2012-01-01

    Very large proteins (subunit sizes >200 kDa) are difficult to electrophoretically separate on polyacrylamide gels. A SDS vertical agarose gel system has been developed that has vastly improved resolving power for very large proteins. Proteins with molecular masses between 200 and 4,000 kDa can be clearly separated. Inclusion of a reducing agent in the upper reservoir buffer has been found to be a key technical procedure for obtaining optimum resolution. PMID:22585481

  7. SEPARATION AND IDENTIFICATION OF SOYBEAN LEAF PROTEINS BY TWO-DIMENSIONAL GEL ELECTROPHORESIS AND MASS SPECTROMETRY

    Technology Transfer Automated Retrieval System (TEKTRAN)

    To establish a proteomic reference map for soybean leaves, we separated and identified leaf proteins using two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) and mass spectrometry (MS). Tryptic digests of 260 spots were subjected to peptide mass fingerprinting (PMF) by matrix-assisted las...

  8. Analysis of soybean embryonic axis proteins by two-dimensional gel electrophoresis and mass spectrometry

    Technology Transfer Automated Retrieval System (TEKTRAN)

    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. Determination of polyacrylamides in coal washery effluents by ultrafiltration/site-exclusion chromatography-ultraviolet detection techniques

    SciTech Connect

    Leung, R.W.M.; Pandey, R.N.; Das, B.S.

    1987-05-01

    The use of a combined technique of ultrafiltration and aqueous size-exclusion high-performance liquid chromatography-UV detection for monitoring trace levels of residual polyacrylamide flocculants in coal washery process water is described. Flocculants of both anionic and non-ionic types in effluents are analyzed by chromatography on a TSK 5000 PW type hydrophilic and semirigid porous polymer gel with 0.05 M Na/sub 2/SO/sub 4/ in water as the mobile phase and by UV detection at 208-nm wavelength for detection. Precision studies gave a relative standard deviation of 5.8% and a precision of 2.2% at the 95% confidence level in the concentration range of 20 ppm. The lower limit of detection for the method is 1.0 ..mu..g. Prior to chromatography, fractionation and concentration of the polyacrylamide in effluents are achieved by ultrafiltration with a hollow fiber cartridge having a nominal molecular weight cutoff of 100,000, and recoveries are determined by spiking studies. The application of the techniques for the analysis of residual flocculant in a coal washery thickener feed effluent sample is described.

  10. A convenient green preparation of layered double hydroxide/polyacrylamide nanocomposite hydrogels with ultrahigh deformability.

    PubMed

    Wu, Lianying; Hu, Ziqiao; Chen, Guangming; Li, Zhibo

    2015-12-14

    A convenient green preparation method has been developed to achieve layered double hydroxide (LDH)/polymer nanocomposite (NC) hydrogels. In contrast to previous publications using toxic organic solvent of formamide or methanol in LDH exfoliation or anion exchange, the interlayer anion exchange and exfoliation of LDH are completed in one step with the help of an amino acid (L-serine). The LDH/polyacrylamide (PAM) NC hydrogels are achieved by a convenient exfoliation-adsorption in situ polymerization method. The exfoliation of LDH is characterized by dynamic light scattering and transmission electron microscopy. Interestingly, the developed NC hydrogels reveal ultrahigh deformability and extraordinary stretchability, confirmed by qualitative images and qualitative tensile and compression tests. The molecular mechanism for the ultrahigh deformability and extraordinary stretchability is discussed by crosslinking density, inter-crosslinking molecular weight and swelling tests. We believe that the findings reported herein will deepen our understanding towards the chemistry of network soft materials including gels, and further widen the applications of polymer hydrogels in mechanical devices such as artificial muscles, biomedical devices and drug delivery systems. PMID:26412191

  11. Isolation and Characterization of Polyacrylamide-Degrading Bacteria from Dewatered Sludge

    PubMed Central

    Yu, Feng; Fu, Ruimin; Xie, Yun; Chen, Wuling

    2015-01-01

    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. PMID:25893998

  12. Dielectric properties of gel collected from shark electrosensors

    NASA Astrophysics Data System (ADS)

    Hughes, Mary E.; Brown, Brandon R.; Hutchison, John C.; Murray, Royce W.

    2003-03-01

    To investigate the physical mechanism of the electric sense, we present an initial characterization of the dielectric properties of the glycoprotein gel that fills the electrosensitive organs of marine elasmobranches (sharks, skates, and rays). To ascertain the properties of the gel, low-frequency impedance spectroscopy is used. The impedance data collected from a dialyzed sample show large values of static permittivity and a loss peak corresponding to a long relaxation time (about 1 ms). Impedance measurements of the native (nondialyzed) gel reliable to 0.1 Hz will be presented and compared to the dialyzed gel. Ramifications of the gel's dielectric properties for the electric sense will be explored.

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

    PubMed Central

    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. PMID:21980487

  14. Measurements of elastic moduli of silicone gel substrates with a microfluidic device.

    PubMed

    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. PMID:21980487

  15. Recycling of superfine resolution agarose gel.

    PubMed

    Seng, T-Y; Singh, R; Faridah, Q Z; Tan, S-G; Alwee, S S R S

    2013-01-01

    Genetic markers are now routinely used in a wide range of applications, from forensic DNA analysis to marker-assisted plant and animal breeding. The usual practice in such work is to extract the DNA, prime the markers of interest, and sift them out by electrically driving them through an appropriate matrix, usually a gel. The gels, made from polyacrylamide or agarose, are of high cost, limiting their greater applications in molecular marker work, especially in developing countries where such technology has great potential. Trials using superfine resolution (SFR) agarose for SSR marker screening showed that it is capable of resolving SSR loci and can be reused up to 14 times, thus greatly reducing the cost of each gel run. Furthermore, for certain applications, low concentrations of agarose sufficed and switching to lithium borate buffer, instead of the conventional Tris-borate-ethylenediaminetetraacetic acid buffer, will further save time and cost. The 2.5% gel was prepared following the Agarose SFR(TM)manual by adding 2.5 g agarose powder into 100 mL 1X lithium borate buffer in a 250-mL flask with rapid stirring. Two midigels (105 x 83 mm, 17 wells) or 4 minigels (50 x 83 mm, 8 wells), 4 mm thickness can be prepared from 100 mL gel solution. A total of 1680 PCR products amplified using 140 SSR markers from oil palm DNA samples were tested in this study using SFR recycled gel. As average, the gel can be recycled 8 times with good resolution, but can be recycled up to 14 times before the resolutions get blurred. PMID:23546970

  16. Hydrolyzed polyacrylamide biodegradation and mechanism in sequencing batch biofilm reactor.

    PubMed

    Yan, Miao; Zhao, Lanmei; Bao, Mutai; Lu, Jinren

    2016-05-01

    An investigation was performed to study the performance of a sequencing batch biofilm reactor (SBBR) to treat hydrolyzed polyacrylamides (HPAMs) and to determine the mechanisms of HPAM biodegradation. The mechanisms for the optimized parameters that significantly improved the degradation efficiency of the HPAMs were investigated by a synergistic effect of the co-metabolism in the sludge and the enzyme activities. The HPAM and TOC removal ratio reached 54.69% and 70.14%. A significant decrease in the total nitrogen concentration was measured. The carbon backbone of the HPAMs could be degraded after the separation of the amide group according to the data analysis. The HPLC results indicated that the HPAMs could be converted to polymer fragments without the generation of the acrylamide monomer intermediate. The results from high-throughput sequencing analysis revealed proteobacterias, bacteroidetes and planctomycetes were the key microorganisms involved in the degradation. PMID:26896716

  17. Electrophoretic extraction of proteins from two-dimensional electrophoresis gel spots

    DOEpatents

    Zhang, Jian-Shi; Giometti, C.S.; Tollaksen, S.L.

    1987-09-04

    After two-dimensional electrophoresis of proteins or the like, resulting in a polyacrylamide gel slab having a pattern of protein gel spots thereon, an individual protein gel spot is cored out from the slab, to form a gel spot core which is placed in an extraction tube, with a dialysis membrane across the lower end of the tube. Replicate gel spots can be cored out from replicate gel slabs and placed in the extraction tube. Molten agarose gel is poured into the extraction tube where the agarose gel hardens to form an immobilizing gel, covering the gel spot cores. The upper end portion of the extraction tube is filled with a volume of buffer solution, and the upper end is closed by another dialysis membrane. Upper and lower bodies of a buffer solution are brought into contact with the upper and lower membranes and are provided with electrodes connected to the positive and negative terminals of a dc power supply, thereby producing an electrical current which flows through the upper membrane, the volume of buffer solution, the agarose, the gel spot cores and the lower membrane. The current causes the proteins to be extracted electrophoretically from the gel spot cores, so that the extracted proteins accumulate and are contained in the space between the agarose gel and the upper membrane. 8 figs.

  18. Centrifugal Methods and Devices for Rapid In-Gel Digestion of Proteins

    PubMed Central

    Lazarev, Alexander V.; Rejtar, Tomas; Dai, Shujia; Karger, Barry L.

    2010-01-01

    Modern proteomic research frequently relies upon separation of proteins in a polyacrylamide gel matrix followed by in-gel enzymatic digestion and extraction of peptides for subsequent analysis by mass spectrometry. In this work, we propose a novel semi-automated method of mechanical processing of gel bands by passing these bands through a specially designed centrifugal device termed a Gel Shredder prior to digestion and extraction of peptides. Such a device allows integrated washing, destaining and shredding of gel bands into uniform blocks of controlled size, approximately 150300 ?m, prior to the enzymatic digestion and extraction of peptides. Shredding into uniform blocks increases the surface area of the gel pieces and promotes improved gel rehydration, allowing the proteolytic enzymes and solvent with improved penetration of the gel lattice. We demonstrate that the new method substantially reduces the time spent on tedious manual handling of gel bands, while minimizing the risk of sample contamination. The performance of the Gel Shredder has been compared to a conventional in-gel digestion protocol using several standard proteins and a complex proteomic sample in terms of relative quantitation by either MALDI-TOF/TOF or nanoLC-ESI ion trap-FTICR mass spectrometry. It is shown that significant time savings and improved peptide recovery can be obtained for many proteins using the Gel Shredder as compared to the traditional in-gel digestion protocol. PMID:19309014

  19. Embedded ceria nanoparticles in gel improve electrophoretic separation: a preliminary demonstration.

    PubMed

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

    2015-07-01

    Slab gel electrophoresis is still the gold standard method for the separation of biomolecules such as proteins and DNA with advantages such as simplicity, affordability, and high throughput, but it suffers from inadequate separation speed and resolution. Single capillary gel electrophoresis, on the other hand, offers faster separation time and improved resolution at the expense of higher cost and loss of high throughput capability. The high surface to volume ratio of the capillary causes improved heat dissipation leading to a reduced Joule heating and a higher resolution. Here, for the first time, we show the use of dispersed ceria nanoparticles (NPs) to improve the resolution and speed of protein separation in slab gel electrophoresis. We measured the rheological parameters of separation medium in order to find a meaningful relationship between viscosity changes, Joule heating, and band broadening. The results showed that ceria NPs decrease the viscosity of polyacrylamide gel. By loading 0.03% (w/v) ceria NPs into polyacrylamide gel at 25 C, the viscosity decreased 22% and the thermal conductivity increased to 81%, which resulted in 35% reduction in Joule heating and 47% increase in resolution. This work is a cross disciplinary of theoretical physical chemistry for thermal conductivity and rheological measurements of PA and ceria suspensions and application in slab gel electrophoresis. We report here, for the first time, that embedded NPs in PA gel could potentially interface high throughput capability of slab gel electrophoresis with high separation speed of single capillary electrophoresis. PMID:25948088

  20. Automated and manual methods for the determination of polyacrylamide and other anionic polymers

    SciTech Connect

    Allison, J.D.; Wimberly, J.W.; Ely, T.L.

    1987-05-01

    The concentration of polyacrylamides is determined by precipitation with Hyamine 1622 Reagent 1 and by measurement of the amount of light scattered by the resulting turbidity. The analysis can be automated as well as adapted for field trials. The effects of anionic surfactants, changes in polyacrylamide molecular weight, and salinity are discussed.

  1. Gel protein capillary extraction apparatus. electronic protein transfer.

    PubMed

    Cooper, Jonathan W; Gao, Jun; Lee, Cheng S

    2002-03-01

    A gel protein capillary extraction apparatus is developed and demonstrated for its rapid and effective transfer of SDS-protein complexes from polyacrylamide gel to a fused-silica capillary. The small dimensions of capillary columns permit the application of high voltages for achieving rapid and effective transfer of gel proteins. Furthermore, the fused-silica capillaries are internally coated with polyacrylamide for the elimination of electroosmotic pumping and protein adsorption onto the capillary wall. The extracted proteins are present in a highly concentrated solution plug as the result of field amplification and sample stacking during the extraction process. Three model proteins, including cytochrome c (14 kDa), ovalbumin (45 kDa), and beta-galactosidase (116 kDa), are visualized using coomassie blue staining and electrophoretically extracted from the gels with protein loading as low as 50 ng. The SDS-cytochrome c complexes extracted from a 50-ng protein loading are concentrated in a 30-nL solution plug inside the capillary with an estimated concentration of 0. 1 mg/mL or 10(-5) M. The capillary format allows the straightforward integration of a miniaturized trypsin-membrane reactor for on-line proteolytic digestion and ESI-MS analysis for protein/peptide identification. PMID:11924982

  2. Low-velocity super-lubrication of sodium-alginate/polyacrylamide ionic-covalent hybrid double-network hydrogels.

    PubMed

    Li, Xuefeng; Wu, Chu; Yang, Qian; Long, Shijun; Wu, Chonggang

    2015-04-21

    Structural and frictional behaviours of sodium alginate (SA)/polyacrylamide (PAAm) ionic-covalent hybrid, sequential double-network (DN) hydrogels against glass have been investigated in water, NaCl and CaCl2 aqueous solutions using a rotational rheometer. Dilution of adsorptive elastohydrodynamic friction for the PAAm covalent network with repulsive hydrodynamic lubrication for the minor SA ionic network was found to control the frictional stresses of the SA/PAAm gels within between those of the SA and PAAm single-network gels. A tentative qualitative model was proposed to describe the impact of ionic environmental solution on the frictional behaviour of the hybrid gel by selectively affecting the SA-network structure and friction. It was revealed that strong Debye shielding in the NaCl solution significantly reduced the thickness of the electric double layer for hydrodynamic lubrication of the SA network, which made the SA/PAAm gel's friction the highest among the three solutions. Dramatically increased ionic cross-linking of the SA network in the CaCl2 solution, although effectively mediated by the PAAm-network flexible skeleton, still functioned partially to conserve a portion of the SA fractional boundary-friction at the interface, making the friction of the hybrid gel intermediate among the three solutions. In contrast, extreme hydration of the SA network in water sharply increased the volume fraction of its unshielded hydrodynamic lubrication at the interface, which greatly reduced the SA/PAAm's friction to the lowest among the three solutions. We have thus incorporated for the first time both super-lubrication (frictional coefficients of below 10(-2) over low sliding-velocities of 3 10(-5) to 2 10(-3) m s(-1)) and previously reported high fracture energy (over 9000 J m(-2)) into a single ionic-covalent hybrid DN hydrogel, which is the SA/PAAm (?1/8.5 w/w) gel in water. Effects of inversion of DN-formation sequence further indicated that frictional behaviours (i.e. frictional stress-sliding velocity profiles) of the hybrid sequential DN hydrogels (SA/PAAm and PAAm/SA), respectively, were primarily determined by those of the second networks (PAAm and SA), presumably due to the formation of first-second network "core-shell" structures at the blob scale. Frictional stress of the SA/PAAm gel was increased monotonically with external normal pressure at all of the sliding velocities investigated in the three solutions, which was in agreement with the predictions from the repulsion-adsorption model proposed by Gong et al. PMID:25735912

  3. Native American Discursive Tactic

    ERIC Educational Resources Information Center

    Black, Jason Edward

    2013-01-01

    This essay derives from a course called "The Rhetoric of Native America," which is a historical-critical survey of Native American primary texts. The course examines the rhetoric employed by Natives to enact social change and to build community in the face of exigencies. The main goal of exploring a native text (particularly, Simon Pokagon's

  4. Two-dimensional gel electrophoresis: vertical isoelectric focusing.

    PubMed

    Dorri, Yaser

    2012-01-01

    Two-dimensional gel electrophoresis (2-DE) is one of the most powerful tools for separating proteins based on their size and charge. 2-DE is very useful to separate two proteins with identical molecular weights but different charges, which cannot be achieved with just sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). Here, a simpler and easier version of 2-DE is presented which is also faster than all the currently available techniques. In this modified version of 2-DE, isoelectric focusing is carried out in the first dimension using a vertical SDS-PAGE apparatus. Following the first-dimensional IEF, each individual lane is excised from the IEF gel and, after a 90 rotation, is inserted into a second-dimensional SDS-PAGE, which can be stained with Coomassie Brilliant Blue for protein analysis or immunoblotted for further analysis. This version of IEF can be run in less than 2 h compared to the overnight run required by O'Farrell's method. Difficult tube gel casting and gel extrusion as well as tube gel distortion are eliminated in our method. This method is simpler, faster, and inexpensive. Both dimensions can be done on the same SDS-PAGE apparatus, and up to ten samples can be run simultaneously using one gel. PMID:22585490

  5. In-gel digestion of proteins for internal sequence analysis after one- or two-dimensional gel electrophoresis.

    PubMed

    Rosenfeld, J; Capdevielle, J; Guillemot, J C; Ferrara, P

    1992-05-15

    We examined the different steps necessary for the enzymatic digestion of proteins in the polyacrylamide matrix after gel electrophoresis. As a result, we developed an improved method for obtaining peptides for internal sequence analysis from 1-2 micrograms of in-gel-digested proteins. The long washing-lyophilization-equilibration steps necessary to eliminate the dye, sodium dodecyl sulfate, and other gel-associated contaminants that perturb protein digestion in Coomassie blue-stained gels have been replaced by washing for 40 min with 50% acetonitrile, drying for 10 min at room temperature, and then rehydrating with a protease solution. The washing and drying steps result in a substantial reduction of the gel slice volume that, when next swollen in the protease solution, readily absorbs the enzyme, facilitating digestion. The Coomassie blue staining procedure has also been modified by reducing acetic acid and methanol concentrations in the staining solution and by eliminating acetic acid in the destaining solution. The peptides resulting from the in-gel digestion are easily recovered by passive elution, in excellent yields for structural characterization. This simple and rapid method has been successfully applied for the internal sequence analysis of membrane proteins from the rat mitochondria resolved in preparative two-dimensional gel electrophoresis. PMID:1524213

  6. Polyacrylamide medium for the electrophoretic separation of biomolecules

    DOEpatents

    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.

  7. Stereoregular polyacrylamide and its copolymer brushes: Preparation and surface characters

    NASA Astrophysics Data System (ADS)

    Jiang, Jianguo; Wang, Xiaoshu; Lu, Xiaoyan; Lu, Yun

    2008-12-01

    Two kinds of polymer brushes, the single one with stereospecific polyacrylamide (PAAM) chains and the dual-component one with random poly(methyl methacrylate) (PMMA) segments grafting from stereospecific PAAM chains, were prepared on silicon wafer for the first time by combining the immobilization of initiator and the stereospecific living radical in situ polymerization. With the addition of the Lewis acid AlCl 3 into the polymerization system, the PAAM brushes obtained exhibited an increased stereospecificity as well as a decreased hydrophilicity, which might attribute to the reduced thickness of PAAM brushes on the silicon wafer and the handicap of the free rotation of the stereospecific molecular chain. The smoother surface morphology of the stereospecific PAAM brushes shown in AFM images was in good agreement with the experimental data of water contact angle. Also, block amphiphilic copolymer brushes were prepared with the stereospecific PAAM formed first on silicon wafer as the anchored-initiator and revealed a novel surface self-assembly behavior after being treated with different solvent such as toluene or water. The stereospecificity of PAAM chains in the polymer brushes could be modulated by adjusting reaction conditions according to the requirement of applications for surface hydrophilicity.

  8. Polyacrylamide hydrogel injection for breast augmentation: Another injectable failure

    PubMed Central

    Wang, Zhenxiang; Li, Shirong; Wang, Lingli; Zhang, Shu; Jiang, Yan; Chen, Jinping; Luo, Donglin

    2012-01-01

    Summary Background Increasing complications of polyacrylamide hydrogel (PAAG) augmentation mammoplasty, such as chronic persistent infection, have recently caught the attention of both the medical field and the general public. Material/Methods A total of 96 patients with severe chronic infection following PAAG augmentation mammoplasty were treated in the present study including 63 cases with infection confined to the breast and 33 with systemic infection. Endoscopy and surgery were performed to completely remove the materials and clear the infected tissues followed by drug-irrigation and vacuum-assisted closure for several days. Results In patients with severe infection there were large amounts of PAAG, fibers and infiltration of numerous neutrophils and macrophages. The infection-inducing materials were extensively dispersed in the mammary and subcutaneous tissues, pectoral fascia and intermuscular space. In addition, there was scattered distribution of PAAG materials in the armpit, chest wall and abdominal wall, which were mixed with necrotic tissues and surrounded by lymphocytes, giant cells, macrophages and other inflammatory cells, forming chronic granulomatous and fibrous lesions. Infection was controlled following surgical intervention. No residual infectious foci or recurrent infections were noted among these patients. Although the severe infection did not result in mastectomy, patients had breast atrophy and various degrees of deformation. Conclusions Chronic infection following PAAG augmentation mammaplasty usually causes systemic infection and other devastating adverse reactions. This study confirms PAAG augmentation mammaplasty is another failed attempt. More attention should be paid to the injection of large doses of liquid filler. PMID:22648256

  9. Chemical gel barriers as low-cost alternative to containment and in situ cleanup of hazardous wastes to protect groundwater

    SciTech Connect

    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.

  10. How deeply cells feel: methods for thin gels

    NASA Astrophysics Data System (ADS)

    Buxboim, Amnon; Rajagopal, Karthikan; Brown, Andre'E. X.; Discher, Dennis E.

    2010-05-01

    Tissue cells lack the ability to see or hear but have evolved mechanisms to feel into their surroundings and sense a collective stiffness. A cell can even sense the effective stiffness of rigid objects that are not in direct cellular contactlike the proverbial princess who feels a pea placed beneath soft mattresses. How deeply a cell feels into a matrix can be measured by assessing cell responses on a controlled series of thin and elastic gels that are affixed to a rigid substrate. Gel elasticity E is readily varied with polymer concentrations of now-standard polyacrylamide hydrogels, but to eliminate wrinkling and detachment of thin gels from an underlying glass coverslip, vinyl groups are bonded to the glass before polymerization. Gel thickness is nominally specified using micron-scale beads that act as spacers, but gels swell after polymerization as measured by z-section, confocal microscopy of fluorescent gels. Atomic force microscopy is used to measure E at gel surfaces, employing stresses and strains that are typically generated by cells and yielding values for E that span a broad range of tissue microenvironments. To illustrate cell sensitivities to a series of thin-to-thick gels, the adhesive spreading of mesenchymal stem cells was measured on gel mimics of a very soft tissue (e.g. brain, E ~ 1 kPa). Initial results show that cells increasingly respond to the rigidity of an underlying 'hidden' surface starting at about 10-20 m gel thickness with a characteristic tactile length of less than about 5 m.

  11. Waste-Activated Sludge Fermentation for Polyacrylamide Biodegradation Improved by Anaerobic Hydrolysis and Key Microorganisms Involved in Biological Polyacrylamide Removal

    PubMed Central

    Dai, Xiaohu; Luo, Fan; Zhang, Dong; Dai, Lingling; Chen, Yinguang; Dong, Bin

    2015-01-01

    During the anaerobic digestion of dewatered sludge, polyacrylamide (PAM), a chemical conditioner, can usually be consumed as a carbon and nitrogen source along with other organic matter (e.g., proteins and carbohydrates in the sludge). However, a significant accumulation of acrylamide monomers (AMs) was observed during the PAM biodegradation process. To improve the anaerobic hydrolysis of PAM, especially the amide hydrolysis process, and to avoid the generation of the intermediate product AM, a new strategy is reported herein that uses an initial pH of 9, 200 mg COD/L of PAM and a fermentation time of 17 d. First, response surface methodology (RSM) was applied to optimize PAM removal in the anaerobic digestion of the sludge. The biological hydrolysis of PAM reached 86.64% under the optimal conditions obtained from the RSM. Then, the mechanisms for the optimized parameters that significantly improved the biological hydrolysis of PAM were investigated by the synergistic effect of the main organic compounds in the sludge, the floc size distribution, and the enzymatic activities. Finally, semi-continuous-flow experiments for a microbial community study were investigated based on the determination of key microorganisms involved in the biological hydrolysis of PAM. PMID:26144551

  12. Waste-Activated Sludge Fermentation for Polyacrylamide Biodegradation Improved by Anaerobic Hydrolysis and Key Microorganisms Involved in Biological Polyacrylamide Removal.

    PubMed

    Dai, Xiaohu; Luo, Fan; Zhang, Dong; Dai, Lingling; Chen, Yinguang; Dong, Bin

    2015-01-01

    During the anaerobic digestion of dewatered sludge, polyacrylamide (PAM), a chemical conditioner, can usually be consumed as a carbon and nitrogen source along with other organic matter (e.g., proteins and carbohydrates in the sludge). However, a significant accumulation of acrylamide monomers (AMs) was observed during the PAM biodegradation process. To improve the anaerobic hydrolysis of PAM, especially the amide hydrolysis process, and to avoid the generation of the intermediate product AM, a new strategy is reported herein that uses an initial pH of 9, 200 mg COD/L of PAM and a fermentation time of 17 d. First, response surface methodology (RSM) was applied to optimize PAM removal in the anaerobic digestion of the sludge. The biological hydrolysis of PAM reached 86.64% under the optimal conditions obtained from the RSM. Then, the mechanisms for the optimized parameters that significantly improved the biological hydrolysis of PAM were investigated by the synergistic effect of the main organic compounds in the sludge, the floc size distribution, and the enzymatic activities. Finally, semi-continuous-flow experiments for a microbial community study were investigated based on the determination of key microorganisms involved in the biological hydrolysis of PAM. PMID:26144551

  13. Characterization of Helicoverpa armigera gut proteinases and their interaction with proteinase inhibitors using gel X-ray film contact print technique.

    PubMed

    Harsulkar, A M; Giri, A P; Gupta, V S; Sainani, M N; Deshpande, V V; Patankar, A G; Ranjekar, P K

    1998-06-01

    Since Helicoverpa armigera is a devastating pest, an attempt was made to separate its gut proteinases and assess their diversity. Gelatin coating present on the X-ray film was used as a substrate to detect electrophoretically separated proteinases of H. armigera gut extract on native polyacrylamide gel electrophoresis (PAGE), sodium dodecyl sulfate (SDS)-PAGE and isoelectric focusing gels. The method involves electrophoresis, followed by washing the gel with Triton X-100 in case of SDS-PAGE, equilibration of the gel in proteinase assay buffers, overlaying the gel on X-ray film followed by washing the film with hot water to remove hydrolyzed gelatin revealing bands of proteinase activity. Using this protocol, at least six different proteinase isoforms were detected in H. armigera gut contents while three isoproteinases were identified in a commercial bacterial proteinase preparation. Adoption of the technique facilitated characterization of the H. armigera gut proteinases (HGP) and provided an easy tool to study the properties of the individual proteinases without purification. The approximate molecular masses of HGP as determined by SDS-PAGE were: 172.9, 59.3, 54.9, 47.6, 44.1 and 41.6 kDa, and of bacterial proteinases: 180.7, 127.3 and 95.3 kDa. The isoelectric point (pI) values of HGP and bacterial proteinase were in the range of 5.1-7.1 and 3.5-7.7, respectively. Some of the HGP isoforms were found to be highly pH-sensitive and showed activity only at pH 10.0. The major HGPs were inhibited by phenylmethylsulfonyl fluoride but not by (4-amidinophenyl)-methanesulfonyl fluoride. Incubation of HGP-resolved electrophoretic gel strips in chickpea or winged bean proteinase inhibitor solution permitted identification of specific inhibitors of individual proteinases and revealed that the major HGPs were insensitive to chickpea inhibitors whereas winged bean inhibitors effectively inhibited all the HGPs. Our results suggest that considerable variability exists among the isoproteinases of H. armigera gut with respect to their pH optima and sensitivity towards chemical and plant proteinase inhibitors. Such diversity is of immense biological significance as it explains the polyphagous nature of the insect which imparts unique adaptability to it against the defensive proteinase inhibitors of its wide range of host plants. PMID:9694289

  14. Check dam and polyacrylamide performance under simulated stormwater runoff.

    PubMed

    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 (<440 NTU). The extent of sediment deposition between the check 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. PMID:24036092

  15. Investigation of the properties of polyacrylamide-polyaniline composite and its application as a battery electrode

    SciTech Connect

    Bhat, N.V.; Joshi, N.V. . Dept. of Chemical Technology)

    1993-11-20

    The composite films of polyacrylamide and polyaniline were prepared by polymerizing aniline using ammonium persulfate as an initiator in an aqueous solution containing poly-acrylamide. A film was then cast from this solution. The structural, dynamic mechanical, electrical, and thermal properties of these films have been studied. The infrared spectrum shows the presence of polyacrylamide as well as polyaniline in the composite film. The thermal analysis shows that the composite degrades slower than does the polyacrylamide alone. The dynamic mechanical analysis indicates that there is an increase in the glass transition temperature after the composite formation. The electrical conductivity has been found to increase by more than eight orders of magnitude. These composite films have also been suitably used as electrodes in secondary batteries.

  16. Quantitation and characterization of rat tissue metallothioneins by gel electrophoresis

    SciTech Connect

    Lin, L.Y.; McCormick, C.C.

    1986-03-05

    A discontinuous gradient gel electrophoretic system was developed to quantitate and characterize metallothionein (MT) in rat tissue. Vertical slab separating gels (1.5 mm x 14 cm x 12 cm) consisted of a linear polyacrylamide gradient 7.5 to 30% T and 5% Bis. The stacking gels (3% T and 20% Bis) were photopolymerized using riboflavin as the catalyst. Liver cytosols were prepared from rats which received (i.p.) various amounts of Zn (5 mg/kg BW) or Cd (2.5 mg/kg BW). Purified MT was prepared by gel filtration and DEAE ion-exchange chromatography. Cytosols were heated (80/sup 0/C, 2 min) and centrifuged to obtain a supernatant. An appropriate amount of supernatant and various amounts of MT standard were electrophoresed (constant current, 20 mA per slab) for 9 hours. Gels were stained with Commassie Blue (R-250, 0.25%) for 12 hours and destained. Gels were scanned by densitometer and peaks heights were determined. Significantly linear standard curves (..mu..g MT vs. peak height) were established for both MTI and MTII. (Cd, Zn)-MTI migrated slower than Zn-MTI while mobilities for both (Cd, Zn)- and Zn-MTII were the same. The accumulation of MTI was consistently less than MTII in liver from both Zn- and Cd-injected rats. Their results suggest that electrophoretic analysis is an excellent system not only for quantitation but also for characterization of MT in rat tissue.

  17. Gel-Electrophoretic Separation, Detection, and Characterization of Plant and Bacterial UDP-Glucose Glucosyltransferases

    PubMed Central

    Thelen, Michael P.; Delmer, Deborah P.

    1986-01-01

    We have developed procedures for detection and characterization of UDP-glucose: glucosyltransferases following electrophoretic separation in nondenaturing polyacrylamide gels. Using digitonin-solubilized membrane protein preparations from a variety of plants and two cellulose-producing bacteria, activity can be demonstrated for several UDP-glucose:?-glucan synthases with an in situ assay following gel electrophoresis. These enzymes can be characterized within the gels with respect to effector requirements and products produced, and several advantages of this assay over solution assays are demonstrated. For example, the clear dependence of plant UDP-glucose:(1?3)-?-glucan synthase on both Ca2+ and a ?-linked glucoside is shown; bacterial cellulose synthases show direct stimulation within the gel by guanyl oligonucleotide, and the Acetobacter xylinum enzyme appears more stable in the gel assay than in solution assay. Images Fig. 1 Fig. 2 PMID:16664924

  18. Cloning and Characterization of an Endoglucanase Gene from Actinomyces sp. Korean Native Goat 40

    PubMed Central

    Kim, Sung Chan; Kang, Seung Ha; Choi, Eun Young; Hong, Yeon Hee; Bok, Jin Duck; Kim, Jae Yeong; Lee, Sang Suk; Choi, Yun Jaie; Choi, In Soon; Cho, Kwang Keun

    2016-01-01

    A gene from Actinomyces sp. Korean native goat (KNG) 40 that encodes an endo-β-1,4-glucanase, EG1, was cloned and expressed in Escherichia coli (E. coli) DH5α. Recombinant plasmid DNA from a positive clone with a 3.2 kb insert hydrolyzing carboxyl methyl-cellulose (CMC) was designated as pDS3. The entire nucleotide sequence was determined, and an open-reading frame (ORF) was deduced. The ORF encodes a polypeptide of 684 amino acids. The recombinant EG1 produced in E. coli DH5α harboring pDS3 was purified in one step using affinity chromatography on crystalline cellulose and characterized. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis/zymogram analysis of the purified enzyme revealed two protein bands of 57.1 and 54.1 kDa. The amino terminal sequences of these two bands matched those of the deduced ones, starting from residue 166 and 208, respectively. Putative signal sequences, a Shine–Dalgarno-type ribosomal binding site, and promoter sequences related to the consensus sequences were deduced. EG1 has a typical tripartite structure of cellulase, a catalytic domain, a serine-rich linker region, and a cellulose-binding domain. The optimal temperature for the activity of the purified enzyme was 55°C, but it retained over 90% of maximum activity in a broad temperature range (40°C to 60°C). The optimal pH for the enzyme activity was 6.0. Kinetic parameters, Km and Vmax of rEG1 were 0.39% CMC and 143 U/mg, respectively. PMID:26732336

  19. Cloning and Characterization of an Endoglucanase Gene from Actinomyces sp. Korean Native Goat 40.

    PubMed

    Kim, Sung Chan; Kang, Seung Ha; Choi, Eun Young; Hong, Yeon Hee; Bok, Jin Duck; Kim, Jae Yeong; Lee, Sang Suk; Choi, Yun Jaie; Choi, In Soon; Cho, Kwang Keun

    2016-01-01

    A gene from Actinomyces sp. Korean native goat (KNG) 40 that encodes an endo-β-1,4-glucanase, EG1, was cloned and expressed in Escherichia coli (E. coli) DH5α. Recombinant plasmid DNA from a positive clone with a 3.2 kb insert hydrolyzing carboxyl methyl-cellulose (CMC) was designated as pDS3. The entire nucleotide sequence was determined, and an open-reading frame (ORF) was deduced. The ORF encodes a polypeptide of 684 amino acids. The recombinant EG1 produced in E. coli DH5α harboring pDS3 was purified in one step using affinity chromatography on crystalline cellulose and characterized. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis/zymogram analysis of the purified enzyme revealed two protein bands of 57.1 and 54.1 kDa. The amino terminal sequences of these two bands matched those of the deduced ones, starting from residue 166 and 208, respectively. Putative signal sequences, a Shine-Dalgarno-type ribosomal binding site, and promoter sequences related to the consensus sequences were deduced. EG1 has a typical tripartite structure of cellulase, a catalytic domain, a serine-rich linker region, and a cellulose-binding domain. The optimal temperature for the activity of the purified enzyme was 55°C, but it retained over 90% of maximum activity in a broad temperature range (40°C to 60°C). The optimal pH for the enzyme activity was 6.0. Kinetic parameters, Km and Vmax of rEG1 were 0.39% CMC and 143 U/mg, respectively. PMID:26732336

  20. Silver staining of proteins in 2DE gels.

    PubMed

    Lelong, Cécile; Chevallet, Mireille; Luche, Sylvie; Rabilloud, Thierry

    2009-01-01

    Silver staining detects proteins after electrophoretic separation on polyacrylamide gels. Its main positive features are its excellent sensitivity (in the low nanogram range) and the use of very simple and cheap equipment and chemicals. The sequential phases of silver staining are protein fixation, then sensitization, then silver impregnation, and finally image development. Several variants of silver staining are described here, which can be completed in a time range from 2 h to 1 day after the end of the electrophoretic separation. Once completed, the stain is stable for several weeks. PMID:19381593

  1. Native American Homeschooling Association.

    ERIC Educational Resources Information Center

    Rozon, Gina

    2000-01-01

    The Native American Home School Association helps Native parents to provide a good education free from the assimilationist tendencies of public school and to transmit Native values and culture. Discusses various home schooling styles, the effectiveness of home schooling in terms of academic achievement and socialization, and the effectiveness of

  2. Native American Healing Traditions

    ERIC Educational Resources Information Center

    Portman, Tarrell A. A.; Garrett, Michael T.

    2006-01-01

    Indigenous healing practices among Native Americans have been documented in the United States since colonisation. Cultural encapsulation has deterred the acknowledgement of Native American medicinal practices as a precursor to folk medicine and many herbal remedies, which have greatly influenced modern medicine. Understanding Native American

  3. Alaska Natives & the Land.

    ERIC Educational Resources Information Center

    Arnold, Robert D.; And Others

    Pursuant to the Native land claims within Alaska, this compilation of background data and interpretive materials relevant to a fair resolution of the Alaska Native problem seeks to record data and information on the Native peoples; the land and resources of Alaska and their uses by the people in the past and present; land ownership; and future…

  4. Native American Healing Traditions

    ERIC Educational Resources Information Center

    Portman, Tarrell A. A.; Garrett, Michael T.

    2006-01-01

    Indigenous healing practices among Native Americans have been documented in the United States since colonisation. Cultural encapsulation has deterred the acknowledgement of Native American medicinal practices as a precursor to folk medicine and many herbal remedies, which have greatly influenced modern medicine. Understanding Native American…

  5. Polymer gel dosimeters with reduced toxicity: a preliminary investigation of the NMR and optical dose response using different monomers

    NASA Astrophysics Data System (ADS)

    Senden, R. J.; DeJean, P.; McAuley, K. B.; Schreiner, L. J.

    2006-07-01

    In this work, three new polymer gel dosimeter recipes were investigated that may be more suitable for widespread applications than polyacrylamide gel dosimeters, since the extremely toxic acrylamide has been replaced with the less harmful monomers N-isopropylacrylamide (NIPAM), diacetone acrylamide and N-vinylformamide. The new gel dosimeters studied contained gelatin (5 wt%), monomer (3 wt%), N,N'-methylene-bis-acrylamide crosslinker (3 wt%) and tetrakis (hydroxymethyl) phosphonium chloride antioxidant (10 mM). The NMR response (R2) of the dosimeters was analysed for conditions of varying dose, dose rate, time post-irradiation, and temperature during irradiation and scanning. It was shown that the dose-response behaviour of the NIPAM/Bis gel dosimeter is comparable to that of normoxic polyacrylamide gel (PAGAT) in terms of high dose-sensitivity and low dependence on dose rate and irradiation temperature, within the ranges considered. The dose-response (R2) of NIPAM/Bis appears to be linear over a greater dose range than the PAGAT gel dosimeter. The effects of time post-irradiation (temporal instability) and temperature during NMR scanning on the R2 response were more significant for NIPAM/Bis dosimeters. Diacetone acrylamide and N-vinylformamide gel dosimeters possessed considerably lower dose-sensitivities. The optical dose-response, measured in terms of the attenuation coefficient for each polymer gel dosimeter, showed potential for the use of optical imaging techniques in future studies.

  6. DEVELOPMENT OF POLYMER GEL SYSTEMS TO IMPROVE VOLUMETRIC SWEEP AND REDUCE PRODUCING WATER/OIL RATIOS

    SciTech Connect

    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 DPR. All gels studied reduced the permeability to water by a greater factor than the factor by which the oil permeability was reduced. Greater DPR was observed as the concentrations of polymer and chromium were increased. Increased pressure gradients during oil flow decreased the oil permeability and the water permeability that was measured afterward. Lower pressure gradients that were applied subsequently moderately affected water permeabilities but did not affect oil permeabilities. A conceptual model of the mechanisms responsible for DPR is presented. Primary features of the model are (1) the development of flow channels through the gel by dehydration of the gel and by re-connection of pre-treatment, residual oil volume and (2) high flow resistance in the channels during water flow is caused by significant saturations of oil remaining in the channels.

  7. Solids and nutrient removal from flushed swine manure using polyacrylamides

    SciTech Connect

    Vanotti, M.B.; Hunt, P.G.

    1999-12-01

    Most of the organic nutrients and reduced carbon (C) materials in liquid swine manure are contained in fine suspended particles that are not separated by available mechanical separators. Treatment with polyacrylamide (PAM) polymers prior to mechanical removal or gravity settling has the potential for enhancing solids-liquid separation, thus concentrating nitrogen (N), phosphorus (P), and organic C. In this work, the authors determined PAM charge and density characteristics most desirable for swine wastewater applications and established the optimum chemical requirement. Treatments were applied to flushed manure from two swine operations in North Carolina. Cationic PAMs significantly increased solids separation while performance of neutral and anionic types was not different from a control. Cationic PAMs with moderate-charge density (20%) were more effective than polymers with higher charge density. Flocs were large and effectively retained with a 1-mm screen. Optimum PAM rate varied with the amount of total suspended solids (TSS) in the liquid manure; 26 and 79 mg PAM/L for samples containing 1.5 and 4.1 g TSS/L, respectively. Corresponding TSS removal efficiencies were 90 to 94%. In contrast, screening without PAM treatment captured only 5 to 14% of the suspended solids. Polymer usage rate was consistent and averaged 2.0{degree} based on weight of dry solids produced. Volatile suspended solids (VSS) were highly correlated with TSS and comprised 79.5% of TSS. Chemical oxygen demand (COD) and organic nutrient concentrations in the effluent were also significantly decreased by PAM treatment. The decrease of COD concentration, an important consideration for odor control, was linearly related with removal of suspended solids, at a rate of 2.0 g COD/g TSS and 2.6 g COD/g VSS. Removal efficiency of organic N and P followed approximately a 1:1 relationship with removal efficiency of TSS. Chemical cost to capture 90% of the suspended solids was estimated to be $0.026 per hog per day ($2.79 per finished hog). Results obtained indicate that PAM treatment is very effective for removal of manure solids, COD, and organic nutrients from flushed swine effluents. The technology provides an attractive alternative to existing liquid manure handling methods for conserving nutrients and avoiding excessive nutrient application in areas where swine production is concentrated.

  8. Electrophoretic extraction of proteins from two-dimensional electrophoresis gel spots

    DOEpatents

    Zhang, Jian-Shi (Shanghai, CN); Giometti, Carol S. (Glenview, IL); Tollaksen, Sandra L. (Montgomery, IL)

    1989-01-01

    After two-dimensional electrophoresis of proteins or the like, resulting in a polyacrylamide gel slab having a pattern of protein gel spots thereon, an individual protein gel spot is cored out from the slab, to form a gel spot core which is placed in an extraction tube, with a dialysis membrane across the lower end of the tube. Replicate gel spots can be cored out from replicate gel slabs and placed in the extraction tube. Molten agarose gel is poured into the extraction tube where the agarose gel hardens to form an immobilizing gel, covering the gel spot cores. The upper end portion of the extraction tube is filled with a volume of buffer solution, and the upper end is closed by another dialysis membrane. Upper and lower bodies of a buffer solution are brought into contact with the upper and lower membranes and are provided with electrodes connected to the positive and negative terminals of a DC power supply, thereby producing an electrical current which flows through the upper membrane, the volume of buffer solution, the agarose, the gel spot cores and the lower membrane. The current causes the proteins to be extracted electrophoretically from the gel spot cores, so that the extracted proteins accumulate and are contained in the space between the agarose gel and the upper membrane. A high percentage extraction of proteins is achieved. The extracted proteins can be removed and subjected to partial digestion by trypsin or the like, followed by two-dimensional electrophoresis, resulting in a gel slab having a pattern of peptide gel spots which can be cored out and subjected to electrophoretic extraction to extract individual peptides.

  9. The application of Au nanoclusters in the fluorescence imaging of human serum proteins after native PAGE: enhancing detection by low-temperature plasma treatment.

    PubMed

    Zhang, Jing; Sajid, Muhammad; Na, Na; Huang, Lingyun; He, Dacheng; Ouyang, Jin

    2012-05-15

    Proteins in human serum are increasingly being studied for their roles in a wide variety of biochemical interactions. To improve the sensitivity of the detection of human serum proteins after native polyacrylamide gel electrophoresis (PAGE), we have developed a fluorescence imaging detection technique for the detection. BSA (bovine serum albumin)-stabilized Au nanoclusters (NCs) were applied as fluorescent probes for imaging, and low-temperature plasma (LTP) treatment of the Au NCs was introduced to enhance the fluorescence imaging. Here, a series of optimization experiments (e.g. those to optimize for pH) were conducted for protein detection after 1-DE and 2-DE, and several types of discharge gases (He, O(2), and N(2)) were selected for the LTP treatment. The possible mechanism of interaction between the proteins and the Au NCs was demonstrated by an isothermal titration calorimetry experiment. Using the present method, a sensitivity of 7-14 times higher than that of traditional staining detection methods was observed in the oxygen LTP-treated Au NCs fluorescence images, and some relatively low abundance proteins (identified by the MS/MS technique) were easily detected. In addition, this fluorescence imaging method was applied to distinguish between the serum samples of patients with liver diseases and those of healthy people. Thus, this fluorescence imaging method is suitable for the highly sensitive detection of various serum proteins, and it shows potential capabilities for clinical diagnosis. PMID:22472528

  10. Microfluidic device having an immobilized pH gradient and PAGE gels for protein separation and analysis

    DOEpatents

    Sommer, Gregory J.; Hatch, Anson V.; Singh, Anup K.; Wang, Ying-Chih

    2012-12-11

    Disclosed is a novel microfluidic device enabling on-chip implementation of a two-dimensional separation methodology. Previously disclosed microscale immobilized pH gradients (IPG) are combined with perpendicular polyacrylamide gel electrophoresis (PAGE) microchannels to achieve orthogonal separations of biological samples. Device modifications enable inclusion of sodium dodecyl sulfate (SDS) in the second dimension. The device can be fabricated to use either continuous IPG gels, or the microscale isoelectric fractionation membranes we have also previously disclosed, for the first dimension. The invention represents the first all-gel two-dimensional separation microdevice, with significantly higher resolution power over existing devices.

  11. Microfluidic device having an immobilized pH gradient and page gels for protein separation and analysis

    DOEpatents

    Sommer, Gregory J; Hatch, Anson V; Singh, Anup K; Wang, Ying-Chih

    2014-05-20

    Disclosed is a novel microfluidic device enabling on-chip implementation of a two-dimensional separation methodology. Previously disclosed microscale immobilized pH gradients (IPG) are combined with perpendicular polyacrylamide gel electrophoresis (PAGE) microchannels to achieve orthogonal separations of biological samples. Device modifications enable inclusion of sodium dodecyl sulfate (SDS) in the second dimension. The device can be fabricated to use either continuous IPG gels, or the microscale isoelectric fractionation membranes we have also previously disclosed, for the first dimension. The invention represents the first all-gel two-dimensional separation microdevice, with significantly higher resolution power over existing devices.

  12. Content in Native Literature Programs.

    ERIC Educational Resources Information Center

    Grant, Agnes

    Including Native literature in school curricula is an important way of enhancing the Native student's self-concept and providing accurate Native cultural knowledge to Native and non-Native students alike. Nevertheless, Canadian school literature programs generally contain neither contemporary nor traditional Native literature. Some programs

  13. Content in Native Literature Programs.

    ERIC Educational Resources Information Center

    Grant, Agnes

    Including Native literature in school curricula is an important way of enhancing the Native student's self-concept and providing accurate Native cultural knowledge to Native and non-Native students alike. Nevertheless, Canadian school literature programs generally contain neither contemporary nor traditional Native literature. Some programs…

  14. Light scattering from a binary-liquid entanglement gel

    NASA Astrophysics Data System (ADS)

    Xia, K.-Q.; Maher, J. V.

    1987-09-01

    Light-scattering experiments have been carried out on an entanglement gel with a binary-liquid mixture as solvent. The onset temperature for critical opalescence has a composition dependence which is similar to the coexistence curve of the free-liquid mixture. This system resembles previously reported work on the cross-linked gel polyacrylamide in two ways: (1) As temperature is lowered toward the critical temperature of the free-liquid mixture, the binary-fluid gel exhibits a strong and increasing light scattering over a broad temperature region of several kelvins, and (2) no appreciable temporal fluctuations are observed throughout this temperature region. Two added features are observed in the present, entanglement-gel measurements: (a) Gel samples with solvent composition both near and off the critical composition of the free-liquid mixture exhibit similar light-scattering behavior, and (b) a Lorentzian-squared fit to the light-scattering angular distributions yields a characteristic wave number which does not change with temperature and an amplitude which shows a very strong dependence on the temperature.

  15. Protein gel staining methods: an introduction and overview.

    PubMed

    Steinberg, Thomas H

    2009-01-01

    Laboratory scientists who encounter protein biochemistry in many of its myriad forms must often ask: is my protein pure? The most frequent response: run a denaturing SDS polyacrylamide gel. Running this gel raises another series of considerations regarding detection, quantitation, and characterization and so the next questions invariably center on suitable protein gel staining and detection methods. A total protein profile can be determined with the colorimetric methods embodied in Coomassie Blue and silver staining methods, or increasingly, with fluorescent stains. Protein quantitation can be done following staining, with fluorescence- and instrumentation-based methods offering the greatest sensitivity and linear dynamic range. Protein posttranslational modifications such as phosphorylation and glycosylation can be reliably determined with several fluorescence-based protocols. Staining and detection with two or more different stains can be done in series to establish relative profiles of modified versus total protein or to assess purity at two levels of quantitative sensitivity. The choice of staining method and protocol depends on the required rigor of detection and quantitation combined with available instrumentation and documentation capabilities. Other considerations for staining methods include intended downstream analytical procedures such as mass spectrometry or peptide sequencing, which preclude some methods. Nonfixative staining methods allow western blotting after gel staining. Laboratory custom and budget or intellectual curiosity may be the ultimate determinate of the chosen gel staining protocol. PMID:19892191

  16. Gold Colloid Dynamics in Polymer Solutions and Gels

    NASA Astrophysics Data System (ADS)

    Reina, J. C.; Hope, D.

    1998-03-01

    The dependence of the dynamics of gold colloid nanospheres in polyacrylamide solutions and gels as a function of crosslink content and scattering angle is studied using dynamic light scattering methods. Auto-correlation functions spanning up to nine sampling time scales were measured from samples ranging through the gel threshold. Cumulant and Inverse Laplace Transform fitting methods indicate a complex dynamical behavior ranging from the purely translational Brownian diffusion of the nanospheres in the medium, through a regime of wide distribution of relaxation rates. Well beyond the gelation threshold, where a large fraction of the gold particles are trapped, we observe a coupling between the movement of the particles and the collective diffusion mode of the polymer network itself. The crossover from diffusional to relaxational behavior depends on the length scale of the probe particles.

  17. Facial Gel Complication after Dental Injection: A Case Report

    PubMed Central

    Pourdanesh, Fereydoun; Shams, Shahin; Sadeghi, Hasan Mir Mohammad

    2013-01-01

    Injectable gel is becoming increasingly popular for cosmetic reasons. The polyacrylamide gel (PAAG) is a permanent filler material used worldwide. In spite of the fact that the filler materials used today are considered quite safe, various complications have been reported in the literature. Hence PAAG use in the United States is not popular. As the area is very close to the dental field, a large complication potential is relatively considered following buccal dental injections. The aim of this article is to highlight a rare complication observed following a local anesthetic administration of a simple molar restoration in a healthy 33-year-old woman who had history of a filler augmentation in her cheek approximately 6 years ago. PMID:24436772

  18. Two-dimensional polyacrylamide gel electrophoresis of Bali bull (Bos javanicus) seminal plasma proteins and their relationship with semen quality.

    PubMed

    Sarsaifi, Kazhal; Haron, Abd Wahid; Vejayan, Jaya; Yusoff, Rosnina; Hani, Homayoun; Omar, Mohamed Ariff; Hong, Lai Wei; Yimer, Nurhusien; Ju, Tan Ying; Othman, Abas-Mazni

    2015-10-01

    The present study evaluated the relationship between Bali bull (Bos javanicus) seminal plasma proteins and different semen quality parameters. Semen samples from 10 mature Bali bulls were evaluated for conventional semen parameters (general motility, viability, and normal morphology), sperm functionality (acrosome reaction, sperm penetration rate, sperm penetration index), sperm kinetics (computer-assisted semen analysis parameters such as sperm velocity), and sperm morphology (acrosome and membrane integrity). Frozen-thawed semen with higher sperm motility, viability, acrosome integrity, and membrane integrity (P < 0.05) are consistently higher in acrosome reaction and sperm penetration assay. Three bulls showed the highest, four bulls displayed the medium, and the remaining three bulls showed the lowest for all sperm parameters and SPA. The proteome maps of seminal plasma from high-quality and low-quality Bali bulls were also established. Seminal plasma of both high-quality and low-quality Bali bulls was subjected to two-dimensional SDS-PAGE with isoelectric point ranged from 3 to 10 and molecular weight from 10 to 250 kDa. Approximately 116 spots were detected with Blue Silver stain, and of these spots, 29 were selected and identified by MALDI-TOF/TOF-MS/MS. A majority of the proteins visualized in the seminal plasma two-dimensional maps was successfully identified. An essential group of the identified spots represented binder of sperm 1 (BSP1), clusterin, spermadhesin, tissue inhibitor of metalloproteinases 2 (TIMP-2), and phospholipase A2 (PLA2). Other proteins found in high abundance included seminal ribonuclease, serum albumin, cationic trypsin, and peptide similar to ?2 microglobulin. Thus, a reference map of Bali bull seminal plasma proteins has been generated for the very first time and can be used to relate protein pattern changes to physiopathologic events that may influence Bali bull reproductive performance. PMID:26119476

  19. Fluid diversion and sweep improvement with chemical gels in oil recovery processes. Final report

    SciTech Connect

    Seright, R.S.; Martin, F.D.

    1992-09-01

    The objectives of this project were to identify the mechanisms by which gel treatments divert fluids in reservoirs and to establish where and how gel treatments are best applied. Several different types of gelants were examined, including polymer-based gelants, a monomer-based gelant, and a colloidal-silica gelant. This research was directed at gel applications in water injection wells, in production wells, and in high-pressure gas floods. The work examined how the flow properties of gels and gelling agents are influenced by permeability, lithology, and wettability. Other goals included determining the proper placement of gelants, the stability of in-place gels, and the types of gels required for the various oil recovery processes and for different scales of reservoir heterogeneity. During this three-year project, a number of theoretical analyses were performed to determine where gel treatments are expected to work best and where they are not expected to be effective. The most important, predictions from these analyses are presented. Undoubtedly, some of these predictions will be controversial. However, they do provide a starting point in establishing guidelines for the selection of field candidates for gel treatments. A logical next step is to seek field data that either confirm or contradict these predictions. The experimental work focused on four types of gels: (1) resorcinol-formaldehyde, (2) colloidal silica, (3) Cr{sup 3+}(chloride)-xanthan, and (4) Cr{sup 3+}(acetate)-polyacrylamide. All experiments were performed at 41{degrees}C.

  20. Sol-Gel Glasses

    NASA Technical Reports Server (NTRS)

    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.

  1. Implicit Attitudes towards Native and Non-Native Speaker Teachers

    ERIC Educational Resources Information Center

    Todd, R. Watson; Pojanapunya, Punjaporn

    2009-01-01

    The academic literature and educational principle suggest that native and non-native English speaking teachers should be treated equally, yet in many countries there is a broad social and commercial preference for native speaker teachers which may also involve racial issues. Attitudes towards native and non-native English speaking teachers have

  2. Long-term degradation of dilute polyacrylamide solutions in turbulent pipe flow

    SciTech Connect

    Choi, U.S.; Kasza, K.E.

    1989-05-01

    The long-term degradation behavior of 200 wppM polyacrylamide solution was studied experimentally in a closed recirculatory flow loop at temperatures of 7.2, 25 and 87.8/degree/C. The degradation behavior was found to be strongly dependent on temperature. The results indicate that, with flow shear similar to that encountered in practical DHC pipe flow, polyacrylamide solutions are highly effective and have a reasonable lifetime at chilled water temperature of 7.2/degree/C. 9 refs., 4 figs.

  3. Microwave initiated synthesis of polyacrylamide grafted Psyllium and its application as a flocculant.

    PubMed

    Sen, Gautam; Mishra, Sumit; Rani, G Usha; Rani, Priti; Prasad, Rajesh

    2012-03-01

    This paper reports a novel microwave initiated method for synthesis of polyacrylamide grafted Psyllium (Psy-g-PAM). Psyllium was modified through grafting of polyacrylamide (PAM) chains on it using microwave radiations only, in absence of any other free radical initiator. The grafting was confirmed by intrinsic viscosity study and characterization techniques like FTIR spectroscopy, elemental analysis (C, H, N, O and S) and SEM morphology study. Further, the flocculation efficacy of the synthesized graft copolymers was studied in kaolin and coal fine suspension through standard 'Jar test' procedure. PMID:22210527

  4. Development of Polymer Gel Systems to Improve Volumetric Sweep and Reduce Producing Water/Oil Ratios

    SciTech Connect

    G. Paul Willhite; Stan McCool; Don W. Green; Min Cheng; Feiyan Chen

    2005-04-03

    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 third year of a 42 month research program that is aimed at an understanding of gelation chemistry and the fundamental mechanisms that alter the flows of oil and water in reservoir rocks after a gel treatment. Work focused on a widely applied system in the field, 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. Pre-gel aggregates form and grow as reactions between chromium acetate and polyacrylamide proceed. A mathematical model that describes uptake and crosslinking reactions as a function of time was derived. The model was probability based and provides molecular-weight averages and molecular-weight distributions of the pre-gel aggregates as a function of time and initial system conditions. A liquid chromatography apparatus to experimentally measure the size and molecular weight distributions of polymer samples was developed. The method worked well for polymer samples without the chromium crosslinker. Sample retention observed during measurements of gelant samples during the gelation process compromised the results. Other methods will be tested to measure size distributions of the pre-gel aggregates. Dissolution of carbonate minerals during the injection of gelants causes the pH of the gelant to increase. Chromium precipitates from solution at the higher pH values robbing the gelant of crosslinker. Experimental data on the transport of chromium acetate solutions through dolomite cores were obtained. A mathematical model that describes the transport of brine and chromium acetate solutions through rocks containing carbonate minerals was used to simulate the experimental results.

  5. Buckling of swelling gels.

    PubMed

    Mora, T; Boudaoud, A

    2006-06-01

    The patterns arising from the differential swelling of gels are investigated experimentally and theoretically as a model for the differential growth of living tissues. Two geometries are considered: a thin strip of soft gel clamped to a stiff gel, and a thin corona of soft gel clamped to a disk of stiff gel. When the structure is immersed in water, the soft gel swells and bends out of plane leading to a wavy periodic pattern whose wavelength is measured. The linear stability of the flat state is studied in the framework of linear elasticity using the equations for thin plates. The flat state is shown to become unstable to oscillations above a critical swelling rate and the computed wavelengths are in quantitative agreement with the experiment. PMID:16779528

  6. Buckling of swelling gels

    NASA Astrophysics Data System (ADS)

    Mora, T.; Boudaoud, A.

    2006-06-01

    The patterns arising from the differential swelling of gels are investigated experimentally and theoretically as a model for the differential growth of living tissues. Two geometries are considered: a thin strip of soft gel clamped to a stiff gel, and a thin corona of soft gel clamped to a disk of stiff gel. When the structure is immersed in water, the soft gel swells and bends out of plane leading to a wavy periodic pattern whose wavelength is measured. The linear stability of the flat state is studied in the framework of linear elasticity using the equations for thin plates. The flat state is shown to become unstable to oscillations above a critical swelling rate and the computed wavelengths are in quantitative agreement with the experiment.

  7. AIDS and Native Youth.

    ERIC Educational Resources Information Center

    Luna, G. Cajetan

    Native Americans throughout North America suffer from a greater prevalence of health problems than the population as a whole. One might believe that the problem of AIDS is insignificant for Native youth, but such a belief is inaccurate and shortsighted. As of March 1989, the Centers for Disease Control reported 1,792 cases of childhood and

  8. Native American Entrepreneurship. Digest.

    ERIC Educational Resources Information Center

    Seymour, Nicole

    Although Native Americans have owned and started the fewest small businesses of all U.S. minority groups, entrepreneurship is considered to be an efficient tool for alleviating their economic problems. Barriers to Native American entrepreneurship include poverty, scarce start-up capital, poor access to business education and technical assistance,…

  9. Tionantati: Native Tobacco People.

    ERIC Educational Resources Information Center

    Shorty, Lawrence

    1997-01-01

    Recounts the author's, and his family's, relationship with tobacco. Identifies tobacco's role in traditional Native American ceremonies as encouraging communication and self-reflection. Describes Tionantati: Native Tobacco People, an intervention program that returns tobacco to its traditional use and eschews recreational use of commercial

  10. AIDS and Native Youth.

    ERIC Educational Resources Information Center

    Luna, G. Cajetan

    Native Americans throughout North America suffer from a greater prevalence of health problems than the population as a whole. One might believe that the problem of AIDS is insignificant for Native youth, but such a belief is inaccurate and shortsighted. As of March 1989, the Centers for Disease Control reported 1,792 cases of childhood and…

  11. Listen to the Natives

    ERIC Educational Resources Information Center

    Prensky, Marc

    2006-01-01

    "Digital natives" refer to today's students because they are native speakers of technology, fluent in the digital language of computers, video games, and the Internet. Those who were not born into the digital world are referred to as digital immigrants. Educators, considered digital immigrants, have slid into the 21st century--and into the digital…

  12. Traditional Native Poetry.

    ERIC Educational Resources Information Center

    Grant, Agnes

    1985-01-01

    While Native myths and legends were educational tools to transmit tribal beliefs and history, traditional American Indian poetry served a ritualistic function in everyday life. Few traditional Native songs, which all poems were, survive; only Mayan and Aztec poems were written, and most of these were burned by a Spanish bishop. In addition, many…

  13. Traditional Native Poetry.

    ERIC Educational Resources Information Center

    Grant, Agnes

    1985-01-01

    While Native myths and legends were educational tools to transmit tribal beliefs and history, traditional American Indian poetry served a ritualistic function in everyday life. Few traditional Native songs, which all poems were, survive; only Mayan and Aztec poems were written, and most of these were burned by a Spanish bishop. In addition, many

  14. Listen to the Natives

    ERIC Educational Resources Information Center

    Prensky, Marc

    2006-01-01

    "Digital natives" refer to today's students because they are native speakers of technology, fluent in the digital language of computers, video games, and the Internet. Those who were not born into the digital world are referred to as digital immigrants. Educators, considered digital immigrants, have slid into the 21st century--and into the digital

  15. Pulse Field Gel Electrophoresis.

    PubMed

    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. PMID:25682374

  16. CONFORMANCE IMPROVEMENT USING GELS

    SciTech Connect

    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 flow capacity of fractures (with widths from 1 to 4 mm) during brine and oil flow after placement. Regardless of gel age before placement, very little gel washed out from the fractures during brine or oil flow. However, increased brine or oil flow rate and cyclic injection of oil and water significantly decreased the level of permeability reduction. A particular need exists for gels that can plug large apertures (e.g., wide fractures and vugs). Improved mechanical strength and stability were demonstrated (in 1- to 4-mm-wide fractures) for a gel that contained a combination of high- and low-molecular weight polymers. This gel reduced the flow capacity of 2- and 4-mm-wide fractures by 260,000. In a 1-mm-wide fracture, it withstood 26 psi/ft without allowing any brine flow through the fracture. Cr(III)-acetate-HPAM gels exhibited disproportionate permeability reduction in fractures. The effect was most pronounced when the gel was placed as gelant or partially formed gels. The effect occurred to a modest extent with concentrated gels and with gels that were ''fully formed'' when placed. The effect was not evident in tubes. We explored swelling polymers for plugging fractures. Polymer suspensions were quickly prepared and injected. In concept, the partially dissolved polymer would lodge and swell to plug the fracture. For three types of swelling polymers, behavior was promising. However, additional development is needed before their performance will be superior to that of conventional gels.

  17. Native SAD is maturing

    PubMed Central

    Rose, John P.; Wang, Bi-Cheng; Weiss, Manfred S.

    2015-01-01

    Native SAD phasing uses the anomalous scattering signal of light atoms in the crystalline, native samples of macromolecules collected from single-wavelength X-ray diffraction experiments. These atoms include sodium, magnesium, phosphorus, sulfur, chlorine, potassium and calcium. Native SAD phasing is challenging and is critically dependent on the collection of accurate data. Over the past five years, advances in diffraction hardware, crystallographic software, data-collection methods and strategies, and the use of data statistics have been witnessed which allow ‘highly accurate data’ to be routinely collected. Today, native SAD sits on the verge of becoming a ‘first-choice’ method for both de novo and molecular-replacement structure determination. This article will focus on advances that have caught the attention of the community over the past five years. It will also highlight both de novo native SAD structures and recent structures that were key to methods development. PMID:26175902

  18. High-Frequency Alternating-Crossed-Field Gel Electrophoresis WithNeutral or Slightly Charged Interpenetrating Networks to Improve DNASeparation

    SciTech Connect

    Boyd, B.; Prausnitz, J.; Blanch, H.

    1998-07-01

    Toward improving DNA separations, this work reports theeffects of high-frequency square-wave AC fields superimposedperpendicular to the direct current (DC) separation field on DNAmigration in both polyacrylamide-based interpenetrating networks (IPNs)and in agarose networks. Compared to standard polyacrylamide gels, IPNsallow the separation of larger DNA (9000 bp vs. 5000 bp at 5 V/cm). Innovel polyacrylamide-based IPNs, an alternating current (AC) field of 5Hz increased the maximum DNA size separable. This effect was extended tolarger DNA sizes with increasing electric-field strength up to andapparently beyond the power supply-limited maximum electric-fieldstrength of 48 V/cm. The orthogonal AC field also increased mobility.These two results combine to yield a reduction in separation time of upto a factor of 20 in novel polyacrylamide-based IPNs. When negativelycharged acrylic-acid groups were incorporated into the IPNs, the use ofthe AC field changed the DNA-network interaction, which altered the sizedependence of DNA mobility. In agarose gels, an AC field of 50 Hzincreased the size range separable; however, there was no increase in DNAmobility. There was no change in size dependence of mobility in an ACfield when the number of charged groups in the agarose network wasincreased. Based on results in the literature, possible mechanisms wereexamined for the effects of the AC field on DNA separation.

  19. CONFORMANCE IMPROVEMENT USING GELS

    SciTech Connect

    Randall S. Seright

    2004-09-30

    This report describes work performed during the third and final year of the project, ''Conformance Improvement Using Gels.'' Corefloods revealed throughput dependencies of permeability reduction by polymers and gels that were much more prolonged during oil flow than water flow. This behavior was explained using simple mobility ratio arguments. A model was developed that quantitatively fits the results and predicts ''clean up'' times for oil productivity when production wells are returned to service after application of a polymer or gel treatment. X-ray computed microtomography studies of gels in strongly water-wet Berea sandstone and strongly oil-wet porous polyethylene suggested that oil penetration through gel-filled pores occurs by a gel-dehydration mechanism, rather than gel-ripping or gel-displacement mechanisms. In contrast, analysis of data from the University of Kansas suggests that the gel-ripping or displacement mechanisms are more important in more permeable, strongly water-wet sandpacks. These findings help to explain why aqueous gels can reduce permeability to water more than to oil under different conditions. Since cement is the most commonly used material for water shutoff, we considered when gels are preferred over cements. Our analysis and experimental results indicated that cement cannot be expected to completely fill (top to bottom) a vertical fracture of any width, except near the wellbore. For vertical fractures with apertures less than 4 mm, the cement slurry will simply not penetrate very far into the fracture. For vertical fractures with apertures greater than 4 mm, the slurry may penetrate a substantial distance into the bottom part of the fracture. However, except near the wellbore, the upper part of the fracture will remain open due to gravity segregation. We compared various approaches to plugging fractures using gels, including (1) varying polymer content, (2) varying placement (extrusion) rate, (3) using partially formed gels, (4) using combinations of high and low molecular weight (Mw) polymers, (5) using secondary crosslinking reactions, (6) injecting un-hydrated polymer particles, and (7) incorporating particulates. All of these methods showed promise in some aspects, but required performance improvements in other aspects. All materials investigated to date showed significant performance variations with fracture width. High pressure gradients and limited distance of penetration are common problems in tight fractures. Gravity segregation and low resistance to breaching are common problems in wide fractures. These will be key issues to address in future work. Although gels can exhibit disproportionate permeability reduction in fractures, the levels of permeability reduction for oil flow are too high to allow practical exploitation in most circumstances. In contrast, disproportionate permeability reduction provided by gels that form in porous rock (adjacent to the fractures) has considerable potential in fractured systems.

  20. Application of Polyacrylamide to Reduce Phosphorus Losses from Chinese Purple Soil: A Laboratory and Field Investigation

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The application of anionic polyacrylamide (PAM) for the control of phosphorus (P) losses from a steeply sloping Chinese purple soil was studied in both a laboratory soil column experiment and a field experiment. The results showed that PAM has an important inhibitory impact on vertical P transport i...

  1. Soil aggregate stability as affected by clay mineralogy and polyacrylamide addition

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The addition of polyacrylamide (PAM) to soil leads to stabilization of existing aggregates and improved bonding between, and aggregation of adjacent soil particles However, the dependence of PAM efficacy as an aggregate stabilizing agent on soil-clay mineralogy has not been studied. Sixteen soil sam...

  2. Effects of Polyacrylamide and Organic Matter on Microbes associated to Soil Aggregation of Norfolk Loamy Sand

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Polyacrylamide (PAM, anionic formulation of molecular size 12 MDa and 35% charge density) has been reported to increase aggregation and improve soil physical properties in United States southeastern Coastal Plain loamy sand soils, but nothing is known about the effects of PAM on microbes associated ...

  3. Effects of Polyacrylamide Molecular Weight, Soil Texture and Electrolyte Concentration on Drainable Porosity and Aggregate Stability

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The literature reports on the intricate relations between soil type and molecular weight (MW) of polyacrylamide (PAM) with respect to PAM efficacy as a soil conditioner. This relation may depend on the ability of PAM to penetrate into aggregates and thus stabilize both outer and inner aggregate surf...

  4. Polyacrylamide Molecular Weight and Phosphogypsum Effects on Infiltration and Erosion in Semi-Arid Soils

    Technology Transfer Automated Retrieval System (TEKTRAN)

    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 ...

  5. Polyacrylamide molecular weight and phosphogypsum effects on infiltration and erosion in semi-arid soils

    Technology Transfer Automated Retrieval System (TEKTRAN)

    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 ...

  6. Production of a polyacrylamide solution used in an oil recovery process

    SciTech Connect

    Luetzelschwab, W.E.

    1987-01-06

    A process is described for recovering oil from a subterranean oil-bearing formation having performance demands comprising the steps of: determining the performance demands of the formation; determining correlations between an initial polymerization reaction parameter of initiator level and partially hydrolyzed polyacrylamide solution properties of screen factor and viscosity, each correlation having a discontinuity; selecting a value of the initiator level below each discontinuity such that the selected value of the initiator level is capable of producing a partially hydrolyzed polyacrylamide solution having values of the properties of viscosity and screen factor relatively sensitive to varying the initiator level and capable of meeting the performance demands of the formation; producing the partially hydrolyzed polyacrylamide solution having the values of the properties relatively sensitive to varying the initiator level and capable of meeting the performance demands by polymerizing an acrylamide monomer using a polymerization initiator at the selected value; and injecting the partially hydrolyzed polyacrylamide solution into the formation to improve oil recovery therefrom.

  7. Comparison of Cationic and Unmodified Starches in Reactive Extrusion of Starch-Polyacrylamide Graft Copolymers

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Graft copolymers of starch and polyacrylamide (PAAm) were prepared using reactive extrusion in a corotating twin screw extruder. The effect of cationic starch modification was examined using unmodified and cationic dent starch (approximately 23% amylose) and waxy maize starch (approximately 2% amyl...

  8. Polyacrylamide effects on aggregate and structure stability of soils with different clay mineralogy

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Adding anionic polyacrylamide (PAM) to soils stabilizes existing aggregates and improves bonding between and aggregation of soil particles. However, the dependence of PAM efficacy as an aggregate stabilizing agent with soils having different clay mineralogy has not been studied. Sixteen soil samples...

  9. Carbon and nitrogen stable isotope ratios can estimate anionic polyacrylamide degradation in soil

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Water soluble anionic polyacrylamide (PAM) is a highly effective erosion-preventing and infiltration-enhancing polymer, when applied at rates of 1 to 10 g/m-3 in furrow irrigation water. PAM degradation has not directly been measured in soil or water. Natural abundance of the carbon (13C/12C) isoto...

  10. Aggregate stability as affected by polyacrylamide molecular weight, soil texture and water quality

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The favorable effects of the environmentally friendly, non toxic, anionic polyacrylamide (PAM) as a soil conditioner have long been established. However, some uncertainties exist regarding the effects of PAM molecular weight (MW) on its performance as a soil amendment and the ability of PAM to penet...

  11. Polyacrylamide and biopolymer effects on flocculation, aggregate stability, and water seepage in a silt loam

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Researchers 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...

  12. Polyacrylamide treatments for reducing seepage in soil-lined reservoirs: A field evaluation

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Irrigation water supplies are becoming limited and there is a need to extend the usefulness of current water resources. Previous laboratory studies demonstrated that certain water soluble polyacrylamide solution (WSPAM) and cross-linked PAM granule (XPAM) treatments effectively reduced infiltration...

  13. Toxicity of anionic polyacrylamide formulations when used for erosion control in agriculture

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Addition of anionic polyacrylamide (PAM) to agricultural irrigation water can dramatically reduce erosion of soils. However, the toxicity of PAM to aquatic life, while often claimed to be low, has not been thoroughly evaluated. Five PAM formulations, including two oil-based products, one water-based...

  14. POLYACRYLAMIDE EFFECTS ON INFILTRATION IN SAN JOAQUIN VALLEY SANDY LOAM SOILS.

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Low infiltration rates constrain effective and economical irrigation on some sandy loam soils on the east side of California's San Joaquin Valley. Polyacrylamide (PAM) has increased soil infiltration in other areas of the U.S., especially where soil erosion is a problem. We applied low concentrati...

  15. PECTIN AND POLYACRYLAMIDE COMPOSITE HYDROGELS: EFFECT OF PECTIN ON STRUCTURAL AND DYNAMIC MECHANICAL PROPERTIES

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Composite hydrogels of pectin and polyacrylamide were synthesized and evaluated by scanning electron microscopy, atomic force microscopy, light microscopy, and by dynamic mechanical analysis. The cross-linking polymerization of acrylamide in pectin solution resulted in a composite having a macropo...

  16. Reduction of Interrill Erosion by different application methods of Polyacrylamide and Gypsum

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Soil erosion has been studied from different perspectives. This paper presents results on interrill erosion for three different application methods of Polyacrylamide (PAM) and Gypsum. Small interrill plots (0.74 m2) were packed with a highly erodible sieved soil and surface was prepared for it to ha...

  17. Volume transition in a gel driven by hydrogen bonding

    NASA Astrophysics Data System (ADS)

    Ilmain, Franck; Tanaka, Toyoichi; Kokufuta, Etsuo

    1991-01-01

    INTERACTIONS between macromolecules fall into four categories: ionic, hydrophobic, van der Waals and hydrogen bonding. Phase transitions in polymer gels provide a means of studying these interactions. Many gels will undergo reversible, discontinuous volume changes in response to changes in, for example, temperature, gel composition or light irradiation1-5. These transitions result from the competition between repulsive intermolecular forces, usually electrostatic in nature, that act to expand the polymer network, and an attractive force that acts to shrink it. Volume transitions in gels have been observed that are driven by all of the above-mentioned forces except hydrogen bonding (ref 6-10 T.T. et al, unpublished data; H. Inomata et al., personal communication). Here we report on a phase transition in an interpenetrating polymer network of poly(acrylamide) and poly(acrylic acid) that completes this picture-it is controlled by cooperative 'zipping' interactions between the molecules which result from hydrogen bonding. Cooperativity is an essential feature of the interactions, in that independent hydrogen bonds would not provide a sufficient driving force for the transition. A further novel characteristic of this phase transition is that the swelling (in water) is induced by an increase rather than a decrease in temperature.

  18. Gel electrophoresis of linear and star-branched DNA

    NASA Astrophysics Data System (ADS)

    Lau, Henry W.; Archer, Lynden A.

    2011-12-01

    The electrophoretic mobility of double-stranded DNA in polyacrylamide gel is investigated using an activated hopping model for the transport of a charged object within a heterogeneous medium. The model is premised upon a representation of the DNA path through the gel matrix as a series of traps with alternating large and small cross sections. Calculations of the trap dimensions from gel data show that the path imposes varying degrees of confinement upon migrating analytes, which retard their forward motion in a size-dependent manner. An expression derived for DNA mobility is shown to provide accurate predictions for the dynamics of linear DNA (67-622 bp) in gels of multiple concentrations. For star-branched DNA, the incorporation within the model of a length scale previously proposed to account for analyte architecture [Yuan , Anal. Chem.ANCHAM0003-270010.1021/ac060414w 78, 6179 (2006)] leads to mobility predictions that compare well with experimental results for a wide range of DNA shapes and molecular weights.

  19. CONFORMANCE IMPROVEMENT USING GELS

    SciTech Connect

    Randall S. Seright

    2004-03-01

    This technical progress report describes work performed from September 1, 2003, through February 29, 2004, for the project, ''Conformance Improvement Using Gels.'' We examined the properties of several ''partially formed'' gels that were formulated with a combination of high and low molecular weight HPAM polymers. After placement in 4-mm-wide fractures, these gels required about 25 psi/ft for brine to breach the gel (the best performance to date in fractures this wide). After this breach, stabilized residual resistance factors decreased significantly with increased flow rate. Also, residual resistance factors were up to 9 times greater for water than for oil. Nevertheless, permeability reduction factors were substantial for both water and oil flow. Gel with 2.5% chopped fiberglass effectively plugged 4-mm-wide fractures if a 0.5-mm-wide constriction was present. The ability to screen-out at a constriction appears crucial for particulate incorporation to be useful in plugging fractures. In addition to fiberglass, we examined incorporation of polypropylene fibers into gels. Once dispersed in brine or gelant, the polypropylene fibers exhibited the least gravity segregation of any particulate that we have tested to date. In fractures with widths of at least 2 mm, 24-hr-old gels (0.5% high molecular weight HPAM) with 0.5% fiber did not exhibit progressive plugging during placement and showed extrusion pressure gradients similar to those of gels without the fiber. The presence of the fiber roughly doubled the gel's resistance to first breach by brine flow. The breaching pressure gradients were not as large as for gels made with high and low molecular weight polymers (mentioned above). However, their material requirements and costs (i.e., polymer and/or particulate concentrations) were substantially lower than for those gels. A partially formed gel made with 0.5% HPAM did not enter a 0.052-mm-wide fracture when applying a pressure gradient of 65 psi/ft. This result suggests a lower limit of fracture width for entry of formed or partially formed gels (when reasonable pressure gradients are applied). In unfractured porous rock, we investigated the time dependence of oil and water permeabilities during various cycles of oil and water injection after placement of a Cr(III)-acetate-HPAM gel. Permeability to water stabilized rapidly (within 1 pore volume, PV), while permeability to oil stabilized gradually over the course of 100 PV. The behavior was surprisingly insensitive to core material (strongly water-wet Berea sandstone and strongly oil-wet porous polyethylene), core permeability (740 to 10,000 md), and applied pressure gradient (10 to 100 psi/ft).

  20. Modeling chemoresponsive polymer gels.

    PubMed

    Kuksenok, Olga; Deb, Debabrata; Dayal, Pratyush; Balazs, Anna C

    2014-01-01

    Stimuli-responsive gels are vital components in the next generation of smart devices, which can sense and dynamically respond to changes in the local environment and thereby exhibit more autonomous functionality. We describe recently developed computational methods for simulating the properties of such stimuli-responsive gels in the presence of optical, chemical, and thermal gradients. Using these models, we determine how to harness light to drive shape changes and directed motion in spirobenzopyran-containing gels. Focusing on oscillating gels undergoing the Belousov-Zhabotinksy reaction, we demonstrate that these materials can spontaneously form self-rotating assemblies, or pinwheels. Finally, we model temperature-sensitive gels that encompass chemically reactive filaments to optimize the performance of this system as a homeostatic device for regulating temperature. These studies could facilitate the development of soft robots that autonomously interconvert chemical and mechanical energy and thus perform vital functions without the continuous need of external power sources. PMID:24498954

  1. Native Americans and Diabetes

    MedlinePLUS Videos and Cool Tools

    ... the disease, the rate of the development of disease, has actually increased over time. Announcer: In the 1930s, the Pima's native diet started to change. They began eating less of the natural foods like squash, wild ...

  2. Native Health Research Database

    MedlinePLUS

    ... Center MSC09 5100 1 University of New Mexico Albuquerque, NM 87131-0001 Native Services Librarian Phone: (505) ... salud.unm.edu © The University of New Mexico, Albuquerque, NM 87131, (505) 277-0111 Disclaimer Privacy Statement ...

  3. Native American Identity

    ERIC Educational Resources Information Center

    Horse, Perry G.

    2005-01-01

    Many issues and elements--including ethnic nomenclature, racial attitudes, and the legal and political status of American Indian nations and Indian people--influence Native American identity. (Contains 3 notes.)

  4. A gel probe equilibrium sampler for measuring arsenic porewater profiles and sorption gradients in sediments: I. Laboratory development

    USGS Publications Warehouse

    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.

  5. Gel for simultaneous chemical imaging of anionic and cationic solutes using diffusive gradients in thin films.

    PubMed

    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. PMID:24256092

  6. Gel for Simultaneous Chemical Imaging of Anionic and Cationic Solutes Using Diffusive Gradients in Thin Films

    PubMed Central

    2013-01-01

    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. PMID:24256092

  7. In-phantom dosimetry for BNCT with Fricke and normoxic-polymer gels

    NASA Astrophysics Data System (ADS)

    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.

  8. Native Knowledge in the Americas.

    ERIC Educational Resources Information Center

    Kidwell, Clara Sue

    1985-01-01

    Native American science is defined as activities of native peoples of the New World in observing physical phenomena and attempting to explain and control them. Problems in studying native science, ethnoscience and native science, archaeostronomy and ethnoastronomy, ethnobotany, agriculture, technology, and future directions are discussed. (JN)

  9. Microfluidics with Gel Emulsions

    NASA Astrophysics Data System (ADS)

    Priest, Craig; Surenjav, Enkhtuul; Herminghaus, Stephan; Seemann, Ralf

    2006-03-01

    Microfluidic processing is usually achieved using single phase liquids. Instead, we use monodisperse emulsions to compartment liquids within microchannel geometries. At low continuous phase volume fractions, droplets self-organize to form well-defined arrangements, analogous to foam. While it is well-known that confined geometries can induce rearrangement of foam compartments at the millimeter-scale, similar dynamics are also expected for gel emulsions. We have studied online generation, organization and manipulation of gel emulsions using a variety of microchannel geometries. ``Passive'' reorganization, based on fixed channel geometries, can be supplemented by ``active'' manipulation by incorporating a ferrofluid phase. A ferromagnetic phase facilitates reorganization of liquid compartments on demand using an electromagnetic trigger. Moreover, coalescence between adjacent compartments within a gel emulsion can be induced using electrical potential. Microfluidics using gel emulsions will be well-suited for combinatorial chemistry, DNA sequencing, drug screening and protein crystallizations.

  10. Two-dimensional gel electrophoresis image registration using block-matching techniques and deformation models.

    PubMed

    Rodriguez, Alvaro; Fernandez-Lozano, Carlos; Dorado, Julian; Rabual, Juan R

    2014-06-01

    Block-matching techniques have been widely used in the task of estimating displacement in medical images, and they represent the best approach in scenes with deformable structures such as tissues, fluids, and gels. In this article, a new iterative block-matching technique-based on successive deformation, search, fitting, filtering, and interpolation stages-is proposed to measure elastic displacements in two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) images. The proposed technique uses different deformation models in the task of correlating proteins in real 2D electrophoresis gel images, obtaining an accuracy of 96.6% and improving the results obtained with other techniques. This technique represents a general solution, being easy to adapt to different 2D deformable cases and providing an experimental reference for block-matching algorithms. PMID:24613260

  11. Previsible silver staining of protein in electrophoresis gels with mass spectrometry compatibility.

    PubMed

    Jin, Li-Tai; Li, Xiao-Kun; Cong, Wei-Tao; Hwang, Sun-Young; Choi, Jung-Kap

    2008-12-15

    A convenient silver staining method for protein in sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) gels is described. The method is previsible, sensitive, and mass spectrometry (MS) compatible. Two visible counter ion dyes, ethyl violet (EV) and zincon (ZC), were used in the first staining solution with a detection limit of 2 to 8 ng/band in approximately 1h. The dye-stained gel can be further stained by silver staining, which is based on acidic silver staining employing ZC with sodium thiosulfate as silver ion sensitizers. Especially, ZC has silver ion reducing power by cleavage of the diazo bond of the dye during silver reduction. The second silver staining can be completed in approximately 1h with a detection limit of 0.2 ng/band. PMID:18804088

  12. Amended final report on the safety assessment of polyacrylamide and acrylamide residues in cosmetics.

    PubMed

    2005-01-01

    Polyacrylamide is a polymer of controllable molecular weight formed by the polymerization of acrylamide monomers available in one of three forms: solid (powder or micro beads), aqueous solution, or inverse emulsions (in water droplets coated with surfactant and suspended in mineral oil). Residual acrylamide monomer is likely an impurity in most Polyacrylamide preparations, ranging from <1 ppm to 600 ppm. Higher levels of acrylamide monomers are present in the solid form compared to the other two forms. Polyacrylamide is reportedly used in 110 cosmetic formulations, at concentrations ranging from 0.05% to 2.8%. Residual levels of acrylamide in Polyacrylamide can range from <.01% to 0.1%, although representative levels were reported at 0.02% to 0.03%. Because of the large sizes of Polyacrylamide polymers, they do not penetrate the skin. Polyacrylamide itself is not significantly toxic. For example, an acute oral toxicity study of Polyacrylamide in rats reported that a single maximum oral dose of 4.0 g/kg body weight was tolerated. In subchronic oral toxicity studies, rats and dogs treated with Polyacrylamide at doses up to 464 mg/kg body weight showed no signs of toxicity. Several 2-year chronic oral toxicity studies in rats and dogs fed diets containing up to 5% Polyacrylamide had no significant adverse effects. Polyacrylamide was not an ocular irritant in animal tests. No compound-related lesions were noted in a three-generation reproductive study in which rats were fed 500 or 2000 ppm Polyacrylamide in their diet. Polyacrylamide was not carcinogenic in several chronic animal studies. Human cutaneous tolerance tests performed to evaluate the irritation of 5% (w/w) Polyacrylamide indicated that the compound was well tolerated. Acrylamide monomer residues do penetrate the skin. Acrylamide tested in a two-generation reproductive study at concentrations up to 5 mg/kg day(- 1) in drinking water, was associated with prenatal lethality at the highest dose, with evidence of parental toxicity. The no adverse effects level was close to the 0.5 mg/kg day(- 1) dose. Acrylamide tested in a National Toxicology Program (NTP) reproductive and neurotoxicity study at 3, 10, and 30 ppm produced no developmental or female reproductive toxicity. However, impaired fertility in males was observed, as well as minimal neurotoxic effects. Acrylamide neurotoxicity occurs in both the central and peripheral nervous systems, likely through microtubule disruption, which has been suggested as a possible mechanism for genotoxic effects of acrylamide in mammalian systems. Acrylamide was genotoxic in mammalian in vitro and in vivo assays. Acrylamide was a tumor initiator, but not an initiator/promoter, in two different mouse strains at a total dose of 300 mg/kg (6 doses over 2 weeks) resulting in increased lung adenomas and carcinomas without promotion. Acrylamide was tested in two chronic bioassays using rats. In one study, increased incidence of mammary gland tumors, glial cell tumors, thyroid gland follicular tumors, oral tissue tumors, uterine tumors and clitoral gland tumors were noted in female rats. In male rats, the number of tumors in the central nervous system (CNS), thyroid gland, and scrotum were increased with acrylamide exposure. In the second study, using higher doses and a larger number of female rats, glial cell tumors were not increased, nor was there an increase in mammary gland, oral tissue, clitoral gland, or uterine tumors. Tumors of the scrotum in male rats were confirmed, as were the thyroid gland follicular tumors in males and females. Taken together, there was a dose-dependent, but not statistically significant, increase in the number of astrocytomas. Different human lifetime cancer risk predictions have resulted, varying over three orders of magnitude from 2 x 10(- 3) to 1.9 x 10(- 6). In the European Union, acrylamide has been limited to 0.1 ppm for leave-on cosmetic products and 0.5 ppm for other cosmetic products. An Australian risk assessment suggested negligible health risks from acrylamide in cosmetics. The Cosmetic

  13. Conformance Improvement Using Gels

    SciTech Connect

    Seright, Randall S.; Schrader, Richard; II Hagstrom, John; Wang, Ying; Al-Dahfeeri, Abdullah; Gary, Raven; Marin; Amaury; Lindquist, Brent

    2002-09-26

    This research project had two objectives. The first objective was 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 was to optimize treatments in fractured production wells, where the gel must reduce permeability to water much more than that to oil.

  14. Crystallization from Gels

    NASA Astrophysics Data System (ADS)

    Narayana Kalkura, S.; Natarajan, Subramanian

    Among the various crystallization techniques, crystallization in gels has found wide applications in the fields of biomineralization and macromolecular crystallization in addition to crystallizing materials having nonlinear optical, ferroelectric, ferromagnetic, and other properties. Furthermore, by using this method it is possible to grow single crystals with very high perfection that are difficult to grow by other techniques. The gel method of crystallization provides an ideal technique to study crystal deposition diseases, which could lead to better understanding of their etiology. This chapter focuses on crystallization in gels of compounds that are responsible for crystal deposition diseases. The introduction is followed by a description of the various gels used, the mechanism of gelling, and the fascinating phenomenon of Liesegang ring formation, along with various gel growth techniques. The importance and scope of study on crystal deposition diseases and the need for crystal growth experiments using gel media are stressed. The various crystal deposition diseases, viz. (1) urolithiasis, (2) gout or arthritis, (3) cholelithiasis and atherosclerosis, and (4) pancreatitis and details regarding the constituents of the crystal deposits responsible for the pathological mineralization are discussed. Brief accounts of the theories of the formation of urinary stones and gallstones and the role of trace elements in urinary stone formation are also given. The crystallization in gels of (1) the urinary stone constituents, viz. calcium oxalate, calcium phosphates, uric acid, cystine, etc., (2) the constituents of the gallstones, viz. cholesterol, calcium carbonate, etc., (3) the major constituent of the pancreatic calculi, viz., calcium carbonate, and (4) cholic acid, a steroidal hormone are presented. The effect of various organic and inorganic ions, trace elements, and extracts from cereals, herbs, and fruits on the crystallization of major urinary stone and gallstone constituents are described. In addition, tables of gel-grown organic and inorganic crystals are provided.

  15. Fluorescent staining of gels.

    PubMed

    Buxbaum, Engelbert

    2012-01-01

    Certain transition metal complexes show intensive fluorescence when bound to proteins. They can be used to stain gels after electrophoresis with a sensitivity approaching that of silver staining, but in a much simpler and more reproducible procedure. Stains can be prepared easily and at a fraction of the cost of commercially available reagents.Hydrophobic dyes can be used to stain gels without fixing; they do not interfere with later blotting or electro-elution. PMID:22585519

  16. Saos-2 cell-mediated mineralization on collagen gels: Effect of densification and bioglass incorporation.

    PubMed

    Liu, Gengbo; Pastakia, Meet; Fenn, Michael B; Kishore, Vipuil

    2016-05-01

    Plastic compression is a collagen densification process that has been widely used for the development of mechanically robust collagen-based materials. Incorporation of bioglass within plastically compressed collagen gels has been shown to mimic the microstructural properties of native bone and enhance in vitro cell-mediated mineralization. The current study seeks to decouple the effects of collagen densification and bioglass incorporation to understand the interplay between collagen packing density and presence of bioglass on cell-mediated mineralization. Saos-2 cell-mediated mineralization was assessed as a measure of the osteoconductivity of four different collagen gels: (1) uncompressed collagen gel (UC), (2) bioglass incorporated uncompressed collagen gel (UC + BG), (3) plastically compressed collagen gel (PC), and (4) bioglass incorporated plastically compressed collagen gel (PC + BG). The results indicated that collagen densification enhanced mineralization as shown by SEM, increased alkaline phosphatase activity and produced significantly higher amounts of mineralized nodules on PC gels compared to UC gels. Further, the amount of nodule formation on PC gels was significantly higher compared to UC + BG gels indicating that increase in matrix stiffness due to collagen densification had a greater effect on cell-mediated mineralization compared to bioglass incorporation into loosely packed UC gels. Incorporation of bioglass into PC gels further enhanced mineralization as evidenced by significantly larger nodule size and higher amount of mineralization on PC + BG gels compared to PC gels. In conclusion, collagen densification via plastic compression improves the osteoconductivity of collagen gels. Further, incorporation of bioglass within PC gels has an additive effect and further enhances the osteoconductivity of collagen gels. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 104A: 1121-1134, 2016. PMID:26750473

  17. Polyacrylamide brush layer for Hydrophilic Interaction Liquid Chromatography of intact glycoproteins

    PubMed Central

    Zhang, Zhaorui; Wu, Zhen; Wirth, Mary J.

    2013-01-01

    A chromatographic column of nonporous silica particles with a bonded phase of linear polyacrylamide chains is evaluated for hydrophilic interaction liquid chromatography (HILIC) of intact glycoproteins. The column is shown to retain glycoproteins significantly more strongly than non-glycoproteins. A particle diameter of 700 nm gives two-fold higher resolution than does a 1.4 ?m particle diameter, and the column efficiency is found to be mostly limited by packing heterogeneity. LCMS is able to resolve the five glycoforms of ribonuclease B and give high quality mass spectra, but there is loss of resolution of the isomers of glycoforms due to the lower amount of TFA. Compared to two leading commercial HILIC columns operated at 60 C, the polyacrylamide column operated at 30 C provided at least two-fold higher resolution for intact ribonuclease B, and showed peaks for glycoforms of prostate specific antigen, although not resolved. PMID:23806357

  18. Characterization of Network Structure of Polyacrylamide Based Hydrogels Prepared By Radiation Induced Polymerization

    NASA Astrophysics Data System (ADS)

    Mahmudi, Naim; ?en, Murat; Gven, Olgun; Rendevski, Stojan

    2007-04-01

    In this study network structure of polyacrylamide based hydrogels prepared by radiation induced polymerization has been investigated. Polyacrylamide based hydrogels in the rod form were prepared by copolymerization of acrylamide(AAm) with hydroxyl ethyl methacrylate(HEMA) and methyl acrylamide(MAAm) in the presence of cross-linking agent and water by gamma rays at ambient temperature. Molecular weight between cross-links and effective cross-link density of hydrogels were calculated from swelling as well as shear modulus data obtained from compression tests. The results have shown that simple compression analyses can be used for the determination of effective cross-link density of hydrogels without any need to some polymer-solvent based parameters as in the case of swelling based determinations. Diffusion of water into hydrogels was examined by analyzing water absorption kinetics and the effect of network, structure on the diffusion type and coefficient was discussed.

  19. Characterization of Network Structure of Polyacrylamide Based Hydrogels Prepared By Radiation Induced Polymerization

    SciTech Connect

    Mahmudi, Naim; Sen, Murat; Gueven, Olgun; Rendevski, Stojan

    2007-04-23

    In this study network structure of polyacrylamide based hydrogels prepared by radiation induced polymerization has been investigated. Polyacrylamide based hydrogels in the rod form were prepared by copolymerization of acrylamide(AAm) with hydroxyl ethyl methacrylate(HEMA) and methyl acrylamide(MAAm) in the presence of cross-linking agent and water by gamma rays at ambient temperature. Molecular weight between cross-links and effective cross-link density of hydrogels were calculated from swelling as well as shear modulus data obtained from compression tests. The results have shown that simple compression analyses can be used for the determination of effective cross-link density of hydrogels without any need to some polymer-solvent based parameters as in the case of swelling based determinations. Diffusion of water into hydrogels was examined by analyzing water absorption kinetics and the effect of network, structure on the diffusion type and coefficient was discussed.

  20. EFFECT OF DEXTRAN-graft-POLYACRYLAMIDE INTERNAL STRUCTURE ON FLOCCULATION PROCESS PARAMETERS

    SciTech Connect

    Bezugla, T.; Kutsevol, N.; Shyichuk, A.; Ziolkowska, D.

    2008-08-28

    Dextran-graft-Polyacrylamide copolymers (D-g-PAA) of brush-like architecture were tested as flocculation aids in the model kaolin suspensions. Due to expanded conformation the D-g-PAA copolymers are more effective flocculants than individual PAA with close molecular mass. The internal structure of D-g-PAA copolymers which is determined by number and length of grafted PAA chains, the distance between grafts, etc., has the significant influence on flocculation behavior of such polymers.

  1. Laboratory and Field Evaluations of Polyacrylamide Hydrogel Baits Against Argentine Ants (Hymenoptera: Formicidae).

    PubMed

    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. PMID:26470250

  2. Denaturing gradient gel electrophoresis to detect methylation changes in DNA.

    PubMed

    Shiraishi, Masahiko

    2004-01-01

    Denaturing gradient gel electrophoresis (DGGE) is a technique that fractionates DNA molecules on the basis of their melting behavior and thereby permits the separation of DNA fragments with local variations in base composition. The separation of DNA fragments by DGGE is determined by the nucleotide sequence, rather than size. This approach is effective when part of the molecule is relatively dense in G+C pairs. This separation is possible because of the pronounced drop in electrophoretic mobility in a polyacrylamide gel that occurs when a region of a DNA molecule melts, thereby forming a structure that is partly helical and partly random chain. The electrophoretic mobility of these partly melted DNA fragments is much lower than that of fully helical or fully dissociated molecules. The low residual mobility of the fragment restricts migration into more strongly denaturing regions of the gradient gel and results in focusing of the band. This property can be applied to detect the difference in melting temperature between methylated and nonmethylated DNA fragments after chemical treatment, or to enrich genomic regions in which aberrant methylation occurs. In this chapter, the application of DGGE to the analysis of genomic DNA methylation is reviewed. PMID:15273415

  3. CONFORMANCE IMPROVEMENT USING GELS

    SciTech Connect

    Randall S. Seright

    2002-02-28

    This technical progress report describes work performed from June 20 through December 19, 2001, for the project, ''Conformance Improvement Using Gels''. Interest has increased in some new polymeric products that purport to substantially reduce permeability to water while causing minimum permeability reduction to oil. In view of this interest, we are currently studying BJ's Aqua Con. Results from six corefloods revealed that the Aqua Con gelant consistently reduced permeability to water more than that to oil. However, the magnitude of the disproportionate permeability reduction varied significantly for the various experiments. Thus, as with most materials tested to date, the issue of reproducibility and control of the disproportionate permeability remains to be resolved. Concern exists about the ability of gels to resist washout after placement in fractures. We examined whether a width constriction in the middle of a fracture would cause different gel washout behavior upstream versus downstream of the constriction. Tests were performed using a formed Cr(III)-acetate-HPAM gel in a 48-in.-long fracture with three sections of equal length, but with widths of 0.08-, 0.02-, and 0.08-in., respectively. The pressure gradients during gel extrusion (i.e., placement) were similar in the two 0.08-in.-wide fracture sections, even though they were separated by a 0.02-in.-wide fracture section. The constriction associated with the middle fracture section may have inhibited gel washout during the first pulse of brine injection after gel placement. However, during subsequent phases of brine injection, the constriction did not inhibit washout in the upstream fracture section any more than in the downstream section.

  4. Native plant diversity increases herbivory to non-natives.

    PubMed

    Pearse, Ian S; Hipp, Andrew L

    2014-11-01

    There is often an inverse relationship between the diversity of a plant community and the invasibility of that community by non-native plants. Native herbivores that colonize novel plants may contribute to diversity-invasibility relationships by limiting the relative success of non-native plants. Here, we show that, in large collections of non-native oak trees at sites across the USA, non-native oaks introduced to regions with greater oak species richness accumulated greater leaf damage than in regions with low oak richness. Underlying this trend was the ability of herbivores to exploit non-native plants that were close relatives to their native host. In diverse oak communities, non-native trees were on average more closely related to native trees and received greater leaf damage than those in depauperate oak communities. Because insect herbivores colonize non-native plants that are similar to their native hosts, in communities with greater native plant diversity, non-natives experience greater herbivory. PMID:25232143

  5. The Native American Speaks.

    ERIC Educational Resources Information Center

    Bromberg, Walter; And Others

    This publication is the product of several workshops and is aimed at multi-ethnic integration of teacher attitudes, curriculum content, and teaching techniques. The 7 articles and 3 bibliographies, contributed by Native American consultants, emphasize recognition and alteration of bias in teacher attitudes, curriculum content, and teaching

  6. Native American Literature.

    ERIC Educational Resources Information Center

    Porter, C. Fayne; And Others

    Designed to accommodate a semester course in Native American Literature for secondary students, this teacher's guide includes a general introduction, a statement of the philosophy and goals upon which it is predicated, a nine-week block on post-Columbian literature, a nine-week block on oral literature, separate appendices for each block, a

  7. Rebuilding Native American Communities

    ERIC Educational Resources Information Center

    Coyhis, Don; Simonelli, Richard

    2005-01-01

    The Wellbriety Movement in Native American communities draws on the wisdom and participation of traditional elders. Beginning with a basic community teaching called the Four Laws of Change and the Healing Forest Model, the Wellbriety Movement blends Medicine Wheel knowledge with the 12 Steps of Alcoholics Anonymous to provide culture-specific…

  8. Native American Perspectives.

    ERIC Educational Resources Information Center

    Smith, Walter S.

    1998-01-01

    On the Fajada Butte in New Mexico, 11th-century Anasazi constructed a site that marks the high and low points of the orbits of the sun and the moon. This unit on astronomy challenges students to think differently about the moon and about the ability of native people to understand the natural world. Includes resources for further study. (PVD)

  9. Native Americans: Subject Guide.

    ERIC Educational Resources Information Center

    Bonanni, Mimmo; Etter, Patricia A.

    This annotated subject guide lists reference material that deals with Native Americans and is available in the Arizona State University Libraries. Entries were published 1933-98, but mostly in the 1980s-90s. The guide is not comprehensive, but rather a selective list of resources useful for researching a topic in a variety of fields. The guide

  10. Exploring Native American Symbolism.

    ERIC Educational Resources Information Center

    Dufrene, Phoebe

    This paper described the events and results of a workshop on Native American symbolism presented to educators and held in Kansas City, Missouri. The presenter maintained that some of the most crucial problems facing U.S. educators and students are caused by racial misunderstandings, and that the universality of artistic expression can be a vehicle

  11. Rebuilding Native American Communities

    ERIC Educational Resources Information Center

    Coyhis, Don; Simonelli, Richard

    2005-01-01

    The Wellbriety Movement in Native American communities draws on the wisdom and participation of traditional elders. Beginning with a basic community teaching called the Four Laws of Change and the Healing Forest Model, the Wellbriety Movement blends Medicine Wheel knowledge with the 12 Steps of Alcoholics Anonymous to provide culture-specific

  12. Native Westslope Cutthroat Trout

    USGS Multimedia Gallery

    Native westslope cutthroat trout swim in the north fork of the Flathead River in northwestern Montana. This region is recognized as a range-wide stronghold for genetically pure westslope cutthroat trout. However, rainbow trout invasion and hybridization threatens these ...

  13. Native Westslope Cutthroat Trout

    USGS Multimedia Gallery

    Native westslope cutthroat trout swim in the north fork of the Flathead River in northwestern Montana. The region is recognized as a range-wide stronghold for genetically pure westslope cutthroat trout. However, rainbow trout invasion and hybridization threatens these p...

  14. Native American Cultural Groups.

    ERIC Educational Resources Information Center

    Roy, Loriene, Comp.

    Part of a larger report on the Four Directions Project, an American Indian technology innovation project, this section includes 13 "pathfinders" to locating information on Native American and other indigenous cultural groups. The pathfinders were designed by students in the Graduate School of Library and Information Science at the University of…

  15. NATIVE HEALTH DATABASES: NATIVE HEALTH RESEARCH DATABASE (NHRD)

    EPA Science Inventory

    The Native Health Databases contain bibliographic information and abstracts of health-related articles, reports, surveys, and other resource documents pertaining to the health and health care of American Indians, Alaska Natives, and Canadian First Nations. The databases provide i...

  16. NATIVE HEALTH DATABASES: NATIVE HEALTH HISTORY DATABASE (NHHD)

    EPA Science Inventory

    The Native Health Databases contain bibliographic information and abstracts of health-related articles, reports, surveys, and other resource documents pertaining to the health and health care of American Indians, Alaska Natives, and Canadian First Nations. The databases provide i...

  17. Active Polymer Gel Actuators

    PubMed Central

    Maeda, Shingo; Hara, Yusuke; Yoshida, Ryo; Hashimoto, Shuji

    2010-01-01

    Many kinds of stimuli-responsive polymer and gels have been developed and applied to biomimetic actuators or artificial muscles. Electroactive polymers that change shape when stimulated electrically seem to be particularly promising. In all cases, however, the mechanical motion is driven by external stimuli, for example, reversing the direction of electric field. On the other hand, many living organisms can generate an autonomous motion without external driving stimuli like self-beating of heart muscles. Here we show a novel biomimetic gel actuator that can walk spontaneously with a worm-like motion without switching of external stimuli. The self-oscillating motion is produced by dissipating chemical energy of oscillating reaction. Although the gel is completely composed of synthetic polymer, it shows autonomous motion as if it were alive. PMID:20162001

  18. Spatially resolved multicomponent gels

    NASA Astrophysics Data System (ADS)

    Draper, Emily R.; Eden, Edward G. B.; McDonald, Tom O.; Adams, Dave J.

    2015-10-01

    Multicomponent supramolecular systems could be used to prepare exciting new functional materials, but it is often challenging to control the assembly across multiple length scales. Here we report a simple approach to forming patterned, spatially resolved multicomponent supramolecular hydrogels. A multicomponent gel is first formed from two low-molecular-weight gelators and consists of two types of fibre, each formed by only one gelator. One type of fibre in this ‘self-sorted network’ is then removed selectively by a light-triggered gel-to-sol transition. We show that the remaining network has the same mechanical properties as it would have done if it initially formed alone. The selective irradiation of sections of the gel through a mask leads to the formation of patterned multicomponent networks, in which either one or two networks can be present at a particular position with a high degree of spatial control.

  19. Studying Native America: Problems and Prospects.

    ERIC Educational Resources Information Center

    Thornton, Russell, Ed.

    Based on a conference, this volume examines the past, present, and future of Native American studies. Native American studies seeks to understand Native Americans, America, and the world from a Native American indigenous perspective, and thereby broaden the education of both Native and non-Native Americans. Part 1 asks who Native Americans are…

  20. Studying Native America: Problems and Prospects.

    ERIC Educational Resources Information Center

    Thornton, Russell, Ed.

    Based on a conference, this volume examines the past, present, and future of Native American studies. Native American studies seeks to understand Native Americans, America, and the world from a Native American indigenous perspective, and thereby broaden the education of both Native and non-Native Americans. Part 1 asks who Native Americans are

  1. Synthesis of a novel affinity gel for the purification of carbonic anhydrases.

    PubMed

    Bozdag, Murat; Isik, Semra; Beyaztas, Serap; Arslan, Oktay; Supuran, Claudiu T

    2015-04-01

    A new affinity gel was synthesized for the purification of carbonic anhydrase (CA, EC 4.2.1.1) isozymes from erythrocytes. The gel was prepared on a Sepharose 4B matrix on which a spacer arm based on ethylenediamine was covalently attached via CNBr activation, followed by reaction with the CA inhibitor 4-isothiocyanato-benzenesulfonamide. The derivatized gel incorporated thioureido-benzenesulfonamide moieties as CA ligand. The binding capacity of the new affinity gel was determined at different temperatures, pH values, ionic strengths and elution buffers. The maximum binding of various CAs was achieved at 25 °C with pH 8.5 and ionic strength around 0.4. The overall purifications for human (h) hCA I and hCA II were 672- and 580-fold, and with 62 and 43% yields, respectively. SDS-polyacrylamide gel electrophoresis showed single bands for each purified isozymes, corresponding to a molecular weight of approx. 29 kDa. This is an easily obtainable, efficient and robust affinity gel, useful for the purification of many other α-CAs. PMID:24939102

  2. Label-free DNA hybridization detection with molecular beacon immobilized in photopolymerized acrylamide gel microarray

    NASA Astrophysics Data System (ADS)

    Wang, Hong; Li, Jiong; Liu, Quanjun; Liu, Heping; Lu, Zuhong; Zhu, Jijun

    2001-10-01

    Photopolymerized acrylamide gel microarray used to immobilize the molecular beacon for label free DNA hybridization detection is described in this paper. Polyacrylamide gel microarray was prepared by UV photopolymerization of 4% acrylamide in 40% glycerol, 0.002%methylene blue, 0.012% TEMED and 0.1M phosphate buffer (PHequals7) with a Relpel-silane pretreated quartz mask. This kind of three-dimensional gel microarray provides more than 100 times great immobilization capacity. The hybridization and other processes with it resemble a homogeneous liquid phase reaction rather than a heterogeneous liquid-solid interface reaction. The specially designed molecular beacons contain a 15 base loop sequence with 5 base pair stem, a 20 base thymine as spacer, a 5'-end amino group for immobilization, a fluorescein in the middle of the sequence as the fluorophore and a 3'-end DABCYL as the quencher. Between the 5'-end amino group and the stem, the 20 base thymine is used to minimize destability caused by 5'-end immobilization. Confocal microscope was used to investigate the fluorescence intensity of gel immobilized molecular beacon probes. After hybridization we can easily distinguish complementary and noncomplementay targets with gel-immobilized molecular beacon probes. Image analysis showed that the fluorescence intensity ratio of complementary to noncomplementay probes is great than 5. The potential applications of gel-immobilized molecular beacon microarray are mutation detection, pathogenic detection, etc. in a parallel, cost saving and label-free detection way.

  3. Monte Carlo modeling of a conventional X-ray computed tomography scanner for gel dosimetry purposes.

    PubMed

    Hayati, Homa; Mesbahi, Asghar; Nazarpoor, Mahmood

    2016-01-01

    Our purpose in the current study was to model an X-ray CT scanner with the Monte Carlo (MC) method for gel dosimetry. In this study, a conventional CT scanner with one array detector was modeled with use of the MCNPX MC code. The MC calculated photon fluence in detector arrays was used for image reconstruction of a simple water phantom as well as polyacrylamide polymer gel (PAG) used for radiation therapy. Image reconstruction was performed with the filtered back-projection method with a Hann filter and the Spline interpolation method. Using MC results, we obtained the dose-response curve for images of irradiated gel at different absorbed doses. A spatial resolution of about 2 mm was found for our simulated MC model. The MC-based CT images of the PAG gel showed a reliable increase in the CT number with increasing absorbed dose for the studied gel. Also, our results showed that the current MC model of a CT scanner can be used for further studies on the parameters that influence the usability and reliability of results, such as the photon energy spectra and exposure techniques in X-ray CT gel dosimetry. PMID:26205316

  4. Native Peoples-Native Homelands Climate Change Workshop: Lessons Learned

    NASA Astrophysics Data System (ADS)

    Maynard, N. G.

    2003-12-01

    The Native Peoples-Native Homelands Climate Change Workshop was held on October 28 through November 01, 1998, as part of a series of workshops being held around the U.S. to improve the understanding of the potential consequences of climate variability and change for the Nation. This workshop was specifically designed by Native Peoples to examine the impacts of climate change and extreme weather variability on Native Peoples and Native Homelands from an indigenous cultural and spiritual perspective and to develop recommendations as well as identify potential response actions. The workshop brought together interested Native Peoples, representatives of Tribal governments, traditional elders, Tribal leaders, natural resource managers, Tribal College faculty and students, and climate scientists from government agencies and universities. It is clear that Tribal colleges and universities play a unique and critical role in the success of these emerging partnerships for decision-making in addition to the important education function for both Native and non-Native communities such as serving as a culturally- appropriate vehicle for access, analysis, control, and protection of indigenous cultural and intellectual property. During the discussions between scientists and policy-makers from both Native and non-Native communities, a number of important lessons emerged which are key to building more effective partnerships between Native and non-Native communities for collaboration and decision-making for a more sustainable future. This talk summarizes the key issues, recommendations, and lessons learned during this workshop.

  5. Native Peoples-Native Homelands Climate Change Workshop: Lessons Learned

    NASA Technical Reports Server (NTRS)

    Maynard, Nancy G.

    2003-01-01

    The Native Peoples-Native Homelands Climate Change Workshop was held on October 28 through November 01,1998, as part of a series of workshops being held around the U.S. to improve the understanding of the potential consequences of climate variability and change for the Nation. This workshop was specifically designed by Native Peoples to examine the impacts of climate change and extreme weather variability on Native Peoples and Native Homelands from an indigenous cultural and spiritual perspective and to develop recommendations as well as identify potential response actions. The workshop brought together interested Native Peoples, representatives of Tribal governments, traditional elders, Tribal leaders, natural resource managers, Tribal College faculty and students, and climate scientists fiom government agencies and universities. It is clear that Tribal colleges and universities play a unique and critical role in the success of these emerging partnerships for decision-making in addition to the important education function for both Native and non-Native communities such as serving as a culturally-appropriate vehicle for access, analysis, control, and protection of indigenous cultural and intellectual property. During the discussions between scientists and policy-makers from both Native and non-Native communities, a number of important lessons emerged which are key to building more effective partnerships between Native and non-Native communities for collaboration and decision-making for a more sustainable future. This talk summarizes the key issues, recommendations, and lessons learned during this workshop.

  6. Staining of proteins after isoelectric focusing in gels by new procedures.

    PubMed

    Vesterberg, O; Hansén, L; Sjösten, A

    1977-03-28

    Four procedures are described for staining of protein bands after isoelectric focusing in polyacrylamide gels. In one procedure, Coomassie Brilliant Blue R 250 is used whereas in the others the G 250 form is used in an aqueous solution of perchloric acid. By using isoelectric focusing and the new staining procedures it is possible to detect less than 0.2 microng of many proteins. One of the procedures is especially rapid, allowing detection of proteins without destaining. Procedures for preservation of the protein patterns and determination of proteins by scanning in a densitometer are also described. PMID:849455

  7. Single nucleotide polymorphisms of APOA1 gene and their relationship with serum apolipoprotein A-I concentrations in the native population of Assam

    PubMed Central

    Bora, Kaustubh; Pathak, Mauchumi Saikia; Borah, Probodh; Hussain, Md. Iftikar; Das, Dulmoni

    2015-01-01

    Background There is a growing interest in the role of allelic variants of the APOA1 gene in relation to a number of disorders. We described two common polymorphisms of the APOA1 gene, G-75A and C+83T and investigated their potential influence on the serum apolipoprotein A-I (apo A-I) levels in the native population of Assam a region that is ethnically distinct and from where no information is hitherto available. Methods Blood samples were collected from 150 healthy volunteers. Apo A-I levels were estimated by immunoturbidometry. Genotyping was done by a PCR-RFLP method that involved DNA extraction from whole blood, followed by polymerase chain reaction and digestion of the PCR product by MspI restriction enzyme, and analysis of fragment sizes in 12% polyacrylamide gel. Results The GG variant at G-75A locus and CC variant at C+83T locus were the most prevalent. GG/CC was the most common combination. Homozygous TT genotype was not detected in any of the subjects. The rare allele frequencies for the G-75A and C+83T sites were found to be 0.22 and 0.06 respectively, which significantly differed from those reported in some other populations in neighbouring regions. Serum apo A-I concentrations did not vary significantly across the detected genotypes. These findings were consistent in both sexes. Conclusion We described the distribution of the G-75A and C+83T polymorphisms of the APOA1 gene in the population of Assam for the first time. These polymorphisms were not found to directly influence apo A-I concentrations in this population either individually or synergistically. PMID:26702398

  8. Surface Friction of Double Network Gels and Shape Memory Gels

    NASA Astrophysics Data System (ADS)

    Wada, Masato; Gong, Jin; Makino, Masato; Kabir, Muhamado H.; Furukawa, Hidemitsu

    The frictional behavior of the two kinds of high-strength gels, which are double network (DN) gels and shape memory gels (SMG), was studied. The velocity dependence looks similar for both the DN gels and the SMG, however the details of the dependence are different. The coefficient of the DN gels is smaller than that of the SMGs. The coefficient decreases as the normal force increases. This normal force dependence was observed for both the DN gels and the SMGs. The differences of both the velocity and normal-force dependences between the DN gels and SMG were discussed in relation to their mechanical properties. The coefficient of the DN gels is smaller than that of the SMGs. Additionally the change in the coefficient of the SMG induced by heating was observed for the first time as far as we know.

  9. Rheology of Active Gels

    NASA Astrophysics Data System (ADS)

    Chen, Daniel

    2015-03-01

    Active networks drive a diverse range of critical processes ranging from motility to division in living cells, yet a full picture of their rheological capabilities in non-cellular contexts is still emerging, e.g., How does the rheological response of a network capable of remodeling under internally-generated stresses differ from that of a passive biopolymer network? In order to address this and other basic questions, we have engineered an active gel composed of microtubules, bidirectional kinesin motors, and molecular depletant that self-organizes into a highly dynamic network of active bundles. The network continually remodels itself under ATP-tunable cycles of extension, buckling, fracturing, and self-healing. Using confocal rheometry we have simultaneously characterized the network's linear and non-linear rheological responses to shear deformation along with its dynamic morphology. We find several surprising and unique material properties for these active gels; most notably, rheological cloaking, the ability of the active gel to drive large-scale fluid mixing over several orders of flow magnitude while maintaining an invariant, solid-like rheological profile and spontaneous flow under confinement, the ability to exert micro-Newton forces to drive persistent directed motion of the rheometer tool. Taken together, these results and others to be discussed highlight the rich stress-structure-dynamics relationships in this class of biologically-derived active gels.

  10. Readings in Canadian Native Studies.

    ERIC Educational Resources Information Center

    Price, John A.

    1986-01-01

    After noting that Canadian and American scholars virtually ignore each other in the field of Native Studies, this paper summarizes sources of information for anyone interested in Canadian Native studies. The introduction mentions university programs and departments, newspaper coverage of Native issues, number of books and monographs published per

  11. ALASKAN NATIVE SECONDARY SCHOOL DROPOUTS.

    ERIC Educational Resources Information Center

    RAY, CHARLES K.; AND OTHERS

    DETERMINATIONS WERE MADE OF THE DROPOUT RATE AMONG NATIVE ALASKAN HIGH SCHOOL STUDENTS AND THE VARIOUS REASONS FOR FAILURE TO FINISH SCHOOL. THE STUDY SAMPLE WAS DRAWN FROM NINE ALASKAN HIGH SCHOOLS WITH OVER ONE-HALF NATIVE STUDENT ENROLLEES. NATIVES WERE DEFINED AS PERSONS BEING ONE-FOURTH OR MORE ESKIMO, INDIAN, OR ALEUT. APPROXIMATELY 1,200

  12. Digital Natives or Digital Tribes?

    ERIC Educational Resources Information Center

    Watson, Ian Robert

    2013-01-01

    This research builds upon the discourse surrounding digital natives. A literature review into the digital native phenomena was undertaken and found that researchers are beginning to identify the digital native as not one cohesive group but of individuals influenced by other factors. Primary research by means of questionnaire survey of technologies…

  13. Polyacrylamide-hydroxyapatite composite: Preparation, characterization and adsorptive features for uranium and thorium

    SciTech Connect

    Baybas, Demet; Ulusoy, Ulvi

    2012-10-15

    The composite of synthetically produced hydroxyapatite (HAP) and polyacrylamide was prepared (PAAm-HAP) and characterized by BET, FT-IR, TGA, XRD, SEM and PZC analysis. The adsorptive features of HAP and PAAm-HAP were compared for UO{sub 2}{sup 2+} and Th{sup 4+}. The entrapment of HAP into PAAm-HAP did not change the structure of HAP. Both structures had high affinity to the studied ions. The adsorption capacity of PAAm-HAP was than that of HAP. The adsorption dependence on pH and ionic intensity provided supportive evidences for the effect of complex formation on adsorption process. The adsorption kinetics was well compatible to pseudo second order model. The values of enthalpy and entropy changes were positive. Th{sup 4+} adsorption from the leachate obtained from a regional fluorite rock confirmed the selectivity of PAAm-HAP for this ion. In consequence, PAAm-HAP should be considered amongst favorite adsorbents for especially deposition of nuclear waste containing U and Th, and radionuclide at secular equilibrium with these elements. - Graphical abstract: SEM images of hydroxyapatite (HAP) and polyacrylamide-hydroxyapatite (PAAm-HAP), and the adsorption isotherms for Uranium and Thorium. Highlights: Black-Right-Pointing-Pointer Composite of PAAm-HAP was synthesized from hydroxyapatite and polyacrylamide. Black-Right-Pointing-Pointer The materials were characterized by BET, FT-IR, XRD, SEM, TGA and PZC analysis. Black-Right-Pointing-Pointer HAP and PAAm-HAP had high sorption capacity and very rapid uptake for UO{sub 2}{sup 2+} and Th{sup 4+}. Black-Right-Pointing-Pointer Super porous PAAm was obtained from PAAm-HAP after its removal of HAP content. Black-Right-Pointing-Pointer The composite is potential for deposition of U, Th and its associate radionuclides.

  14. Controlling tailwater sediment and phosphorus concentrations with polyacrylamide in the Imperial Valley, California.

    PubMed

    Goodson, Christopher C; Schwartz, Gregory; Amrhein, Christopher

    2006-01-01

    External loading of phosphorus (P) from agricultural surface discharge (tailwater) is the main cause of excessive algae growth and the eutrophication of the Salton Sea, California. Continuous polyacrylamide (PAM) applications to agricultural irrigation water inflows were evaluated as a means of reducing sediment and P in tailwater. Zero (control) and 1 mg L(-1) PAM (PAM1) treatments were compared at 17 Imperial Valley field sites. Five and 10 mg L(-1) PAM treatments (PAM5, PAM10) were conducted at one site. The particulate phosphorus (Pp) fraction was determined as the difference between total phosphorus (Pt) and the soluble phosphorus (Ps) fraction. We observed 73, 82, and 98% turbidity reduction with PAM1, PAM5, and PAM10 treatments. Although eight field sites had control tailwater sediment concentrations above the New River total maximum daily loads (TMDL), all but one were made compliant during their paired PAM1 treatments. While PAM1 and PAM10 reduced tail water Pp by 31 and 78%, none of the treatments tested reduced Ps. This may have been caused by high irrigation water Na concentrations which would reduce Ca adsorption and Ca-phosphate bridging on the PAM. The PAM1 treatments resulted in <0.5 mg L(-1) drain water polyacrylamide concentrations 1.6 km downstream of PAM addition, while PAM5 and PAM10 treatments produced > 2 mg L(-1) drain water polyacrylamide concentrations. We concluded that, although PAM practically eliminates Imperial Valley tailwater sediment loads, it does not effectively reduce tailwater Ps, the P fraction most responsible for the eutrophication of the Salton Sea. PMID:16738392

  15. Ceric ion initiated synthesis of polyacrylamide grafted oatmeal: Its application as flocculant for wastewater treatment.

    PubMed

    Bharti, Srijita; Mishra, Sumit; Sen, Gautam

    2013-04-01

    Polyacrylamide grafted oatmeal (OAT-g-PAM) was synthesized by conventional method. The grafting of the PAM chains on the biomaterial backbone was confirmed through intrinsic viscosity study, FTIR spectroscopy, elemental analysis (C, H, N, S and O), SEM morphology and TGA study. The intrinsic viscosity of oatmeal appreciably improved on grafting of PAM chains, thus resulting grafted product with potential application as superior viscosifier. Further, flocculation efficacy of the graft copolymer was studied in coal fine suspension, kaolin suspension, iron-ore suspension and then in municipal wastewater through 'jar test' procedure. PMID:23499093

  16. A novel polymeric flocculant based on polyacrylamide grafted inulin: aqueous microwave assisted synthesis.

    PubMed

    Rahul, Rahul; Jha, Usha; Sen, Gautam; Mishra, Sumit

    2014-01-01

    Polyacrylamide grafted inulin (In-g-PAM) was synthesized via aqueous microwave assisted method (using ceric ammonium nitrate in synergism with microwave in aqueous medium). The intended grafting of the PAM chains on polysaccharide backbone was confirmed through standard physicochemical characterization techniques, namely intrinsic viscosity measurement, Fourier transform infrared (FTIR) spectroscopy, elemental analysis (C, H, N and O), thermogravimetric analysis (TGA) and scanning electron microscopy (SEM) studies. Flocculation efficacy of various grades of synthesized grafted product was studied in coal fines suspension, in relation to inulin (parent polysaccharide). This was done utilizing jar test and settling test procedure, towards possible application as a flocculant for coal washery effluents. PMID:24274474

  17. Preparation of macroporous poly(acrylamide) hydrogels by radiation induced polymerization technique

    NASA Astrophysics Data System (ADS)

    aykara, Tuncer; Bulut, Melek; Demirci, Serkan

    2007-12-01

    Macroporous poly(acrylamide) [poly(AAm)] hydrogels were prepared by using poly(ethylene glycol) (PEG) with three different molecular weight as the pore-forming agent during the radiation induced polymerization reaction. Scanning electron microscope graphs reveal that the macroporous network structure of the hydrogels can be adjusted by applying different molecular weights of PEG during the polymerization reaction. The swelling ratios of the PEG-modified hydrogels were much higher than those for the same type of hydrogel prepared via traditional method. However, the pulsatile swelling behavior of the PEG-modified hydrogels in water and in acetone was affected slightly by the change in the amount of the PEG.

  18. Enzymatically active biomimetic micropropellers for the penetration of mucin gels

    PubMed Central

    Walker, Debora; Käsdorf, Benjamin T.; Jeong, Hyeon-Ho; Lieleg, Oliver; Fischer, Peer

    2015-01-01

    In the body, mucus provides an important defense mechanism by limiting the penetration of pathogens. It is therefore also a major obstacle for the efficient delivery of particle-based drug carriers. The acidic stomach lining in particular is difficult to overcome because mucin glycoproteins form viscoelastic gels under acidic conditions. The bacterium Helicobacter pylori has developed a strategy to overcome the mucus barrier by producing the enzyme urease, which locally raises the pH and consequently liquefies the mucus. This allows the bacteria to swim through mucus and to reach the epithelial surface. We present an artificial system of reactive magnetic micropropellers that mimic this strategy to move through gastric mucin gels by making use of surface-immobilized urease. The results demonstrate the validity of this biomimetic approach to penetrate biological gels, and show that externally propelled microstructures can actively and reversibly manipulate the physical state of their surroundings, suggesting that such particles could potentially penetrate native mucus. PMID:26824056

  19. Native Education 101: Basic Facts about American Indian, Alaska Native, and Native Hawaiian Education

    ERIC Educational Resources Information Center

    National Indian Education Association, 2008

    2008-01-01

    This document contains information about education of indigenous American peoples, including: (1) Demographics; (2) Reservations and Native Lands Map; (3) Important Community Issues and Concepts; (4) Educational Issues for Native Students; (5) Type of Schools for Native Students; (6) Indian Education Legislation and Executive Orders; (7)

  20. Asthma and American Indians/Alaska Natives

    MedlinePLUS

    ... Indian/Alaska Native > Asthma Asthma and American Indians/Alaska Natives In 2012, 147,000 American Indian/Native ... reported that they currently have asthma. American Indian/Alaska Native children are 80% more likely to have ...

  1. Effects of polyacrylamide, biopolymer, and biochar on decomposition of soil organic matter and 14C-labeled plant residues as determined by enzyme activities

    NASA Astrophysics Data System (ADS)

    Mahmoud Awad, Yasser; Ok, Young Sik; Kuzyakov, Yakov

    2014-05-01

    Application of polymers for the improvement of aggregate structure and reduction of soil erosion may alter the availability and decomposition of plant residues. In this study, we assessed the effects of anionic polyacrylamide (PAM), synthesized biopolymer (BP), and biochar (BC) on the decomposition of 14C-labeled maize residue in sandy and sandy loam soils. Specifically, PAM and BP with or without 14C-labeled plant residue were applied at 400 kg ha-1, whereas BC was applied at 5000 kg ha-1, after which the soils were incubated for 80 days at 22 oC. Initially, plant residue decomposition was much higher in untreated sandy loam soil than in sandy soil. Nevertheless, the stimulating effects of BP and BC on the decomposition of plant residue were more pronounced in sandy soil, where it accounted for 13.4% and 23.4% of 14C input, respectively, whereas in sandy loam soil, the acceleration of plant residue decomposition by BP and BC did not exceed 2.6% and 14.1%, respectively, compared to untreated soil with plant residue. The stimulating effects of BP and BC on the decomposition of plant residue were confirmed based on activities of ?-cellobiohydrolase, ?-glucosidase, and chitinase in both soils. In contrast to BC and BP, PAM did not increase the decomposition of native or added C in both soils.

  2. Iontophoresis of Salicylic Acid From Salicylic Acid Doped Poly(p-phynylene vinylene)/ Polyacrylamide Hydrogels

    NASA Astrophysics Data System (ADS)

    Niamlang, Sumonman

    2009-03-01

    The apparent diffusion coefficients, Dapp, and the release mechanisms of salicylic acid from salicylic acid-loaded polyacrylamide hydrogels, SA-loaded PAAM, and salicylic acid-doped poly(phenylene vinylene)/polyacrylamide hydrogels, SA-doped PPV/PAAM, were investigated. In the absence of an electric field, the diffusion of SA from the SA-doped PPV/PAAM hydrogel is delayed in the first 3 hr due to the ionic interaction between the anionic drug and PPV. Beyond this period, SA can diffuse continuously into the buffer solution through the PAAM matrix. Dapp of SA-doped PPV/PAAM is higher than that of the SA-loaded PAAM, and the former increases with increasing electric field strength due to the combined mechanisms: the expansion of PPV chains inside the hydrogel; iontophoresis; and the electroporation of the matrix pore. Thus, the presence of the conductive polymer and the applied electric field can be combined to control the drug release rate at an optimal desired level.

  3. Investigation on efficient adsorption of cationic dyes on porous magnetic polyacrylamide microspheres.

    PubMed

    Yao, Tong; Guo, Song; Zeng, Changfeng; Wang, Chongqing; Zhang, Lixiong

    2015-07-15

    We report here the preparation of porous magnetic polyacrylamide microspheres for efficient removal of cationic dyes by a simple polymerization-induced phase separation method. Characterizations by various techniques indicate that the microspheres show porous structures and magnetic properties. They can adsorb methylene blue with high efficiency, with adsorption capacity increasing from 263 to 1977 mg/g as the initial concentration increases from 5 to 300 mg/L. Complete removal of methylene blue can be obtained even at very low concentrations. The equilibrium data is well described by the Langmuir isotherm models, exhibiting a maximum adsorption capacity of 1990 mg/g. The adsorption capacity increases with increasing initial pH and reaches a maximum at pH 8, revealing an electrostatic interaction between the microspheres and the methylene blue molecules. The microspheres also show high adsorption capacities for neutral red and gentian violet of 1937 and 1850 mg/g, respectively, as well as high efficiency in adsorption of mixed-dye solutions. The dye-adsorbed magnetic polyacrylamide microspheres can be easily desorbed, and can be repeatedly used for at least 6 cycles without losing the adsorption capacity. The adsorption capacity and efficiency of the microspheres are much higher than those of reported adsorbents, which exhibits potential practical application in removing cationic dyes. PMID:25797927

  4. Two-dimensional Gel Electrophoresis (2DE)

    NASA Astrophysics Data System (ADS)

    K?odzi?ska, Ewa; Buszewski, Bogus?aw

    The chemical compounds, which are present in the environment, increasingly cause bad effects on health. The most serious effects are tumors and various mutations at the cellular level. Such compounds, from the analytical point of view, can serve the function of biomarkers, constituting measurable changes in the organism's cells and biochemical processes occurring therein. The challenge of the twenty-first century is therefore searching for effective and reliable methods of identification of biomarkers as well as understanding bodily functions, which occur in living organisms at the molecular level. The irreplaceable tool for these examinations is proteomics, which includes both quality and quantity analysis of proteins composition, and also makes it possible to learn their functions and expressions. The success of proteomics examinations lies in the usage of innovative analytical techniques, such as electromigration technique, two-dimensional electrophoresis in polyacrylamide gel (2D PAGE), liquid chromatography, together with high resolution mass spectrometry and bio-informatical data analysis. Proteomics joins together a number of techniques used for analysis of hundreds or thousands of proteins. Its main task is not the examination of proteins inside the particular tissue but searching for the differences in the proteins' profile between bad and healthy tissues. These differences can tell us a lot regarding the cause of the sickness as well as its consequences. For instance, using the proteomics analysis it is possible to find relatively fast new biomarkers of tumor diseases, which in the future will be used for both screening and foreseeing the course of illness. In this chapter we focus on two-dimensional electrophoresis because as it seems, it may be of enormous importance when searching for biomarkers of cancer diseases.

  5. MAGIC Gel Dosimetry

    NASA Astrophysics Data System (ADS)

    Mifflin, Rachel; Shahnazi, Kambiz; Jesseph, Rick

    2008-10-01

    Proton therapy has proven a very successful tool in treating certain tumors, but a three dimensional view of this fact has not yet been clearly demonstrated. In this experiment we have used MAGIC (Methacrylic and Ascorbic Acid in Gelatin Initiated by Copper) gel to represent brain tissue and gone through normal treatment planning for an Acoustic Neuroma to show the three dimensional dose distributions associated with such a tumor.

  6. An inexpensive microslab gel DNA electrophoresis system with real-time fluorescence detection.

    PubMed

    Chen, Xiaojia; Ugaz, Victor M

    2006-02-01

    In this paper, we describe the construction of a simple yet powerful gel electrophoresis apparatus that can be used to perform size-selective separations of DNA fragments in virtually any laboratory. This system employs a microslab gel format with a novel gel casting technique that eliminates the need for delicate combs to define sample loading wells. The compact size of the microslab gel format allows rapid separations to be performed at low voltages using submicroliter sample volumes. Real time fluorescence detection of the migrating DNA fragments is accomplished using an inexpensive digital microscope that directly connects to any PC with a USB interface. The microscope is readily adaptable for this application by replacing its white light source with a blue light-emitting diode (LED) and adding an appropriate emission filter. Both polyacrylamide and agarose gels can be used as separation matrices. Separation performance was characterized using standard dsDNA ladders, and correct sizing of a 191 bp PCR product was achieved in 15 min. The low cost and simplicity of this system makes it ideally suited for use in a variety of laboratory and educational settings. PMID:16342324

  7. Characterization of electroactive behavior and of progress in developments and applications of ionic polymer gels

    NASA Astrophysics Data System (ADS)

    Guelch, Rainer W.; Weible, Andrea; Wallmersperger, Thomas

    2002-07-01

    Polyelectrolyte gels are distinguished by enormous swelling capabilities under the influence of external physical or chemical stimuli. No other kind of material attains similar volume expansiveness. These properties make them most attractive candidates for a new generation of pseudomuscular actuators. In contrast to chemical stimulations which are able to trigger large in-toto deformations, weak electric fields can only induce considerable bending strains in ionic polymer gels when confined to direct electrical effects. This, of course, restricts their potential for technical applications. To characterize their chemo-mechanical and electrical behavior and the underlying physico-chemical processes, experimental and theoretical findings are presented. Measurements of basic mechanical and electrical parameters on polyelectrolyte gels allow quantification of their electroactive responses, especially with respect to the direct effects of external electric fields on the Donnan potential inside the gels. Model calculations on the basis of a coupled chemo-electro-mechanical multi- field formulation are in good agreement with the experimental results. Although the emphasis of this study is given to various anionic and cationic gels of the polyacrylamide family, a new class of hydrogels based on the biopolymer chitosan is included. These natural polymers have excellent properties such as biocompatibility, biodegradability, non-toxicity etc. making them predestinate to biomedical applications.

  8. Development of a gel formulation of formic acid for control of parasitic mites of honey bees.

    PubMed

    Kochansky, J; Shimanuki, H

    1999-09-01

    Formic acid has been used in various countries for the control of parasitic mites of honey bees (Apis mellifera), particularly the Varroa mite (Varroa jacobsoni) and the tracheal mite (Acarapis woodi). Its corrosivity and consequent fear of liability have precluded commercial interest in the United States, and its rapid vaporization requires frequent reapplication. We have developed a gel formulation of formic acid which provides controlled release over 2-3 weeks and improves the convenience and safety of handling of formic acid. The strong acidity of formic acid restricts the choice of gelling agents; vegetable gellants such as agar are destroyed, and bentonite clay derivatives do not gel, even with high-shear mixing. Polyacrylamides lead to viscous liquids lacking thixotropic properties. High-molecular-weight poly(acrylic acids) and fumed silicas provided gels with suitable physical characteristics. The poly(acrylic acid) gels were difficult to mix and gave slower and nonlinear release behavior, while the fumed silica gels were easy to prepare and linear in formic acid vaporization. PMID:10552733

  9. A convenient procedure for transfer blotting of coomassie blue stained proteins from PAGE gels to transparencies.

    PubMed

    Chu, Y H; Whitesides, G M

    1993-06-01

    Proteins stained with Coomassie Brilliant Blue I were transferred effectively by blotting from polyacrylamide gel electrophoresis (PAGE) gels to transparencies of the type used in plain-paper copiers. The details of the original electropherogram were retained on transfer and did not fade over a period of three years. Both the protein and the associated dye transfer; however, protein does not transfer in the absence of dye. Protein patterns present in several types of gels--sodium dodecyl sulfate (SDS)-PAGE, non-denaturing PAGE, isoelectric focusing PAGE and SDS-agarose--all transferred successfully after staining with dye 1. Proteins visualized with other organic dyes such as Fast Green FCF 2, Uniblue A 3 and Procion Blue MX-R 4 also transferred, but proteins stained with Stains-all 5 or silver did not. This transfer provides a simple, economical way to preserve data from slab gel electrophoresis and a convenient method to display data using an overhead projector. The blotted transparencies are also excellent substrates for use with gel scanning densitometers. PMID:7687447

  10. Rapid separation and purification of oligonucleotides by high-performance capillary gel electrophoresis.

    PubMed Central

    Cohen, A S; Najarian, D R; Paulus, A; Guttman, A; Smith, J A; Karger, B L

    1988-01-01

    Picomole amounts of oligodeoxynucleotides [polydeoxyadenylic acids, (dA)40-60] were baseline resolved and analyzed in less than 8 min by high-performance capillary electrophoresis with polyacrylamide gels. In addition, fast analysis of a crude 70-mer oligodeoxynucleotide and a slab gel-purified 99-mer oligodeoxynucleotide was accomplished, demonstrating the ability of high-performance capillary electrophoresis to characterize rapidly synthesized oligonucleotides. Besides analytical separations, 800 ng of a primer (20-mer) was isolated in less than 20 min. The purified species was collected in water and subsequently used as a probe in a standard dot-blot analysis. The use of high-performance capillary electrophoresis for the analysis and purification of a variety of biopolymers is simple, rapid, and has the potential for automation. Images PMID:3200850

  11. Wetting mechanisms of gel-based controlled-release fertilizers.

    PubMed

    Shavit, U; Reiss, M; Shaviv, A

    2003-02-14

    The release mechanism of gel-based controlled release fertilizers (CRFs) involves water penetration into dry mixtures of fertilizers and gel forming polymers. Water penetration provides an upper limit to the whole release process. Where wetting prediction is often based on models that describe the flow of the liquid phase, vapor motion may become significant when a sharp wetting front exists. In this study we examine the role of vapor and fluid flows in the wetting process of CRFs consisting of urea or KNO(3) mixed with polyacrylamide (PAM). Vapor adsorption isotherms were obtained for typical fertilizer-PAM mixtures. Wetting and release experiments were conducted by dividing the CRFs into regions alternately filled with a pure fertilizer and mixtures of PAM and fertilizer. The experiments were designed in such a way that when the wetting front reaches a mixtures interface, its motion depends on the gradient imposed by the difference in osmotic potential (OP). The coupled equations of vapor and liquid flow in initially dry conditions were solved numerically to demonstrate the conceptual understanding gained by the experiments. The results show that wetting front motion is affected by transport and adsorption of vapor. It was also shown that the release rate is different when wetting is governed by vapor flow or by liquid flow. The release pattern from a multi-regions device was consistent with the wetting pattern, demonstrating the possibility to tailor the release according to periods of peak demand. PMID:12586505

  12. A simple and fast procedure for high quality DNA isolation from gels using laundry detergent and inverted columns.

    PubMed

    Pusch, C

    1997-06-01

    A quick method for the recovery of DNA from agarose and polyacrylamide gels with high efficiency and quality is described. Excised gel slices, containing at least 10 ng DNA, are incubated for 15 min at 60 degrees C in the presence of laundry detergents. An "inverted column" is obtained by covering the extraction liquid by a layer of cotton wool, Sephadex G-50 and another layer of cotton wool. Following centrifugation the supernatant containing the DNA is recovered by aspiration and the DNA is precipitated with isopropanol and tRNA, washed with ethanol and air-dried. The yield of reisolated DNA does not depend on DNA fragment size and small quantities of gel volume. Thus, the technique may prove useful in a broad range of applications in the methodology of molecular biology. PMID:9237563

  13. Frequency of Occurrence of Native Hapten Among Enterobacterial Species

    PubMed Central

    Anacker, R. L.; Bickel, W. D.; Haskins, W. T.; Milner, K. C.; Ribi, E.; Rudbach, J. A.

    1966-01-01

    Anacker, R. L. (Rocky Mountain Laboratory, Hamilton, Mont.), W. D. Bickel, W. T. Haskins, K. C. Milner, E. Ribi, and J. A. Rudbach. Frequency of occurrence of native hapten among enterobacterial species. J. Bacteriol. 91:1427–1433. 1966.—Smooth cultures of representative Enterobacteriaceae were screened for the presence of native hapten, a substance previously extracted with trichloroacetic acid from the protoplasmic fraction of one strain each of Escherichia coli O111:B4 and O113. Trichloroacetic acid extracts of protoplasmic fractions of the cells were analyzed for chemical composition, for constituent sugars by paper chromatography, for immunochemical relationship to endotoxin purified by gel filtration, for sedimentation behavior, and for pyrogenicity in rabbits and lethal toxicity in chick embryos. Extracts from two of three additional strains of E. coli O113, all five additional strains of E. coli O111:B4, and one strain each of E. coli O26:B6 and O55:B5 were similar to previously described native hapten in chemical composition, sedimentation properties (S20,w, 3.7 to 5.2), biological potency (usually less than 0.1% that of corresponding endotoxin), and immunochemical relationship to endotoxin. Extracts of one strain each of E. coli O127:B8, Serratia marcescens, Citrobacter freundii, Salmonella enteritidis, and of two lipopolysaccharide-deficient mutants of S. enteritidis differed from typical native hapten. The biosynthetic relationship of native hapten to endotoxin has not yet been revealed. Images PMID:5326701

  14. Characterization of nanocellulose reinforced semi-interpenetrating polymer network of poly(vinyl alcohol) & polyacrylamide composite films.

    PubMed

    Mandal, Arup; Chakrabarty, Debabrata

    2015-12-10

    Semi-interpenetrating polymer network (semi-IPN) of poly(vinyl alcohol)/polyacrylamide was reinforced with various doses of nanocellulose. The different composite films thus prepared were characterized with respect to their mechanical, thermal, morphological and barrier properties. The composite film containing 5 wt.% of nanocellulose showed the highest tensile strength. The semi-interpenetrating polymer network of poly(vinyl alcohol)/polyacrylamide; and its various composites with nanocellulose were almost identical in their thermal stability. Each of the composites however exhibited much superior stability with respect to the linear poly(vinyl alcohol) and crosslinked polyacrylamide. The scanning electron microscopy (SEM) and atomic force microscopy (AFM) studies exhibited phase separated morphology where agglomerates of nanocellulose were found to be dispersed in the matrix of the semi-IPN. The moisture vapor transmission rate (MVTR) was the lowest for the film containing 5 wt.% of nanocellulose. PMID:26428121

  15. Comparison and Suitability of Gel Matrix for Entrapping Higher Content of Enzymes for Commercial Applications

    PubMed Central

    Mahajan, R.; Gupta, V. K; Sharma, J.

    2010-01-01

    To check the suitability of enzyme entrapped beads for use in pharmaceutical industry, amylase enzyme was entrapped in agar/agarose, polyacrylamide gels and calcium alginate beads. Sodium alginate of 1% concentration was found to be best with respect to immobilization efficiency and calcium alginate beads so obtained were not much susceptible to breakage. When sodium alginate- amylase mixture was added from a height of about 20-30 cm. into CaCl2 solution, size of beads was large at higher alginate concentration due to the increase in the size of droplet formation before entering into CaCl2 solution. Enzyme entrapped polyacrylamide and agar/agarose gels were fragile and could not withstand repeated use whereas enzyme entrapped in large calcium alginate beads was used successfully for 50 cycles for the conversion of starch into product without much damage to the beads under stirring conditions. Amylase preparation was also mixed with urease, lysozyme and coimmobilized in large sized calcium alginate beads. These beads were used for 10 repeated cycles to check the conversion of substrates into their products by their respective enzymes and we concluded that an enzyme or mixture of two or three enzymes can be immobilized in the same large sized calcium alginate beads. This will save the additional cost of bioreactor, manpower, maintenance conditions required for the conversion of one drug into another using enzyme/s entrapped in large sized beads. PMID:20838527

  16. Microsequence analysis of winged bean seed proteins electroblotted from two-dimensional gel.

    PubMed

    Hirano, H

    1989-02-01

    Electroblotting method employing a semidry blotting apparatus for the subsequent protein microsequence analysis (Hirano, 1987) was improved. This method is convenient and allows rapid and efficient transfer of the proteins from a polyacrylamide gel (1 mm thick) onto the Polybrene-coated glass-fiber sheet or polyvinylidene difluoride membrane filter in only 20 min. The electroblotted proteins could be sequenced directly with the gas-phase protein sequencer at a 20-pmole level. This method was applied to the sequence analysis of winged bean seed proteins. A portion of the crude extracts from only one-twentieth of a seed of the winged bean was separated by two-dimensional polyacrylamide gel electrophoresis and electroblotted, and the N-terminal amino acid sequences of the blotted proteins were analyzed. The sequences of about 60% of the blotted major proteins, including nine Kunitz trypsin inhibitor-like proteins with heterogeneity in the N-terminal sequences, a protein that has a homologous sequence to the leghaemoglobin, nitrogen-fixing root nodule-specific protein, and a soybean basic 7S globulin-like protein could be easily identified. PMID:2765119

  17. Traditional West Coast Native Medicine

    PubMed Central

    Deagle, George

    1988-01-01

    An important part of the complex culture of the Native people of Canada's Pacific coast is the traditional system of medicine each culture has developed. Population loss from epidemics and the influence of dominant European cultures has resulted in loss of many aspects of traditional medicine. Although some Native practices are potentially hazardous, continuation of traditional approaches to illness remains an important part of health care for many Native people. The use of devil's club plant by the Haida people illustrates that Native medicine has both spiritual and physical properties. Modern family practice shares many important foundations with traditional healing systems. PMID:21253031

  18. Staining proteins in gels.

    PubMed

    Gallagher, Sean; Chakavarti, Deb

    2008-01-01

    Following separation by electrophoretic methods, proteins in a gel can be detected by several staining methods. This unit describes protocols for detecting proteins by four popular methods. Coomassie blue staining is an easy and rapid method. Silver staining, while more time consuming, is considerably more sensitive and can thus be used to detect smaller amounts of protein. Fluorescent staining is a popular alternative to traditional staining procedures, mainly because it is more sensitive than Coomassie staining, and is often as sensitive as silver staining. Staining of proteins with SYPRO Orange and SYPRO Ruby are also demonstrated here. PMID:19066521

  19. Clarification Procedure for Gels

    NASA Technical Reports Server (NTRS)

    Barber, Patrick G.; Simpson, Norman R.

    1987-01-01

    Procedure developed to obtain transparent gels with consistencies suitable for crystal growth, by replacing sodium ions in silicate solution with potassium ions. Clarification process uses cation-exchange resin to replace sodium ions in stock solution with potassium ions, placed in 1M solution of soluble potassium salt. Slurry stirred for several hours to allow potassium ions to replace all other cations on resin. Supernatant solution decanted through filter, and beads rinsed with distilled water. Rinsing removes excess salt but leaves cation-exchange beads fully charged with potassium ions.

  20. Development of Polymer Gel Systems to Improve Volumetric Sweep and Reduce Producing Water/Oil Ratios

    SciTech Connect

    G. Paul Willhite; Stan McCool; Don W. Green; Min Cheng; Feiyan Chen

    2005-12-31

    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 a 42-month research program that focused on the understanding of gelation chemistry and the fundamental mechanisms that alter the flows of oil and water in reservoir rocks after a gel treatment. Work was conducted on a widely applied system in the field, 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. Pre-gel aggregates form and grow as reactions between chromium acetate and polyacrylamide proceed. A rate equation that describes the reaction between chromium acetate and polymer molecules was regressed from experimental data. A mathematical model that describes the crosslinking reaction between two polymer molecules as a function of time was derived. The model was based on probability concepts and provides molecular-weight averages and molecular-weight distributions of the pre-gel aggregates as a function of time and initial system conditions. Average molecular weights of pre-gel aggregates were measured as a function of time and were comparable to model simulations. Experimental methods to determine molecular weight distributions of pre-gel aggregates were unsuccessful. Dissolution of carbonate minerals during the injection of gelants causes the pH of the gelant to increase. Chromium precipitates from solution at the higher pH values robbing the gelant of crosslinker. Experimental data on the transport of chromium acetate solutions through dolomite cores were obtained. A mathematical model that describes the transport of brine and chromium acetate solutions through rocks containing carbonate minerals was used to simulate the experimental results and data from literature. 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 in sandpacks to determine the effect of polymer and chromium concentrations on DPR. All gels studied reduced the permeability to water by a greater factor than the factor by which the oil permeability was reduced. Greater DPR was observed as the concentrations of polymer and chromium were increased. A conceptual model of the mechanisms responsible for DPR is presented. Primary features of the model are (1) the development of flow channels through the gel by dehydration and displacement of the gel and by re-connection of pre-treatment, residual oil volume and (2) high flow resistance in the channels during water flow is caused by significant saturations of oil remaining in the channels. A similar study of DPR was conducted in Berea sandstone cores. Both oil and water permeabilities were reduced by much smaller factors in Berea sandstone cores than in similar treatments in sandpacks. Poor maturation of the gelant in the Berea rock was thought to be caused by fluid-rock interactions that interfered with the gelation process.

  1. Probing mechanical properties of fully hydrated gels and biological tissues.

    PubMed

    Constantinides, Georgios; Kalcioglu, Z Ilke; McFarland, Meredith; Smith, James F; Van Vliet, Krystyn J

    2008-11-14

    A longstanding challenge in accurate mechanical characterization of engineered and biological tissues is maintenance of both stable sample hydration and high instrument signal resolution. Here, we describe the modification of an instrumented indenter to accommodate nanomechanical characterization of biological and synthetic tissues in liquid media, and demonstrate accurate acquisition of force-displacement data that can be used to extract viscoelastoplastic properties of hydrated gels and tissues. We demonstrate the validity of this approach via elastoplastic analysis of relatively stiff, water-insensitive materials of elastic moduli E>1000 kPa (borosilicate glass and polypropylene), and then consider the viscoelastic response and representative mechanical properties of compliant, synthetic polymer hydrogels (polyacrylamide-based hydrogels of varying mol%-bis crosslinker) and biological tissues (porcine skin and liver) of E<500 kPa. Indentation responses obtained via loading/unloading hystereses and contact creep loading were highly repeatable, and the inferred E were in good agreement with available macroscopic data for all samples. As expected, increased chemical crosslinking of polyacrylamide increased stiffness (E40 kPa) and decreased creep compliance. E of porcine liver (760 kPa) and skin (222 kPa) were also within the range of macroscopic measurements reported for a limited subset of species and disease states. These data show that instrumented indentation of fully immersed samples can be reliably applied for materials spanning several orders of magnitude in stiffness (E=kPa-GPa). These capabilities are particularly important to materials design and characterization of macromolecules, cells, explanted tissues, and synthetic extracellular matrices as a function of spatial position, degree of hydration, or hydrolytic/enzymatic/corrosion reaction times. PMID:18922534

  2. Native Plants, Native Knowledge: Insights from Judy Bluehorse Skelton.

    ERIC Educational Resources Information Center

    Reed, Bracken

    2003-01-01

    Judy Bluehorse Skelton is an herbalist of Native American descent who conducts field trips to identify plants and classroom activities to demonstrate their uses. She also works with Portland (Oregon) schools developing culturally appropriate strategies for presenting Native American content. She encourages students to look at events such as the

  3. Our Native Ways: The Voices of Native American Youth.

    ERIC Educational Resources Information Center

    Toke, Arun Narayan, Ed.; And Others

    1994-01-01

    To celebrate the "Decade of the Indigenous Peoples," this issue of a nonprofit children's magazine includes art and writings by Native American youth who share their ways of looking at and living life. Emphasizes the distinct customs, traditions, languages, and folklore of the different Native Nations and Tribes. (LZ)

  4. Native Speakers' Perception of Non-Native English Speech

    ERIC Educational Resources Information Center

    Jaber, Maysa; Hussein, Riyad F.

    2011-01-01

    This study is aimed at investigating the rating and intelligibility of different non-native varieties of English, namely French English, Japanese English and Jordanian English by native English speakers and their attitudes towards these foreign accents. To achieve the goals of this study, the researchers used a web-based questionnaire which

  5. Effects of counter ions of clay platelets on the swelling behavior of nanocomposite gels.

    PubMed

    Ren, Huai-Yin; Zhu, Meifang; Haraguchi, Kazutoshi

    2012-06-01

    The effects of replacing the native Na(+) counter ions associated with the clay platelets by various other cations on the swelling behavior of nanocomposite (NC) gels consisting of an organic (polymer)/inorganic (clay) network were investigated. The negative surface charge of the clay platelet conferred an ionic nature on the NC gels making them a type of polyelectrolyte gel; consequently, the swelling behavior of the NC gels was strongly influenced by the valence of the co-existing counter ions. NC gels containing monovalent cations such as Na(+), K(+) and Li(+) exhibited large swellings and subsequent deswelling in water after attaining maximum degrees of swelling. In contrast, introduction of multivalent cations such as Ca(2+), Mg(2+), and Al(3+) into NC gels depressed markedly both the swelling and subsequent deswelling. The decreased swelling and suppressed deswelling with multivalent ions were strongly influenced by the initial gel state and result from the formation of additional cross-links through ionic interactions between the clay platelets and the multivalent cations. Also, the similar swelling behaviors were observed for all NC gels with different clay concentration. Further, reversible absorption/desorption and selective absorption of multivalent cations were observed for the NC gels examined. PMID:22425253

  6. Effect of Electric Field Strength on Diffusion of Ionic Drugs from Polyacrylamide Hydrogels

    NASA Astrophysics Data System (ADS)

    Sirivat, Anuvat; Niamlang, Sumonman

    2010-03-01

    The apparent diffusion coefficients, Dapp, and the release mechanisms of ionic-drugs from drug-loaded polyacrylamide hydrogels, drug-loaded PAAM, were investigated for the effects of various drug sizes (Lactic acid, 3.11 ; Sulfanilamide,3.47 ; Ampicillin, 5.14 ), matrix pore sizes, and electric filed strengths. The Dapp of the drugs from the drug-loaded PAAM increases with decreasing drug size, increasing matrix pore size or applied electric field strength. The increase in Dapp can be attributed to the combination of the iontophoresis and the electroporation of the matrix pore. The Dapp of drug from the drug-loaded PAAM apparently obey the scaling behavior: Dapp/Do=(drug size/pore size)m with the scaling exponent m equal to 0.73 and 0.50 at the electric fields of 0 and 0.1 V, respectively.

  7. Mercapto functionalized silica entrapped polyacrylamide hydrogel: Arsenic adsorption behaviour from aqueous solution.

    PubMed

    Kumar, Rajesh; Jain, S K; Verma, S; Malodia, P

    2015-10-15

    In this article, 3-mercaptopropyl functionalized silica entrapped polyacrylamide hydrogel (MPFS-PAA) was prepared and characterized by FT-IR, scanning electron microscopy (SEM) and energy dispersion X-ray spectroscopy (EDS). Synthesized hydrogel was evaluated for removal of arsenic(III) from aqueous solution. Adsorption studies were carried out by batch method as function of contact time, initial concentration of arsenic and pH. As(III) adsorption data fitted well with Langmuir and Freundlich isotherm models. Adsorption capacity of arsenic 92.5 ?g/g was obtained at initial concentration of 100 ?g/L by Langmuir isotherm. Adsorption kinetics was tested for pseudo-second order reaction at different contact time. The rate constants of pseudo second order reaction were calculated and good correlation coefficient R(2) 99.67 obtained. The results indicates that MPFS-PAA is an effective adsorbent for removal of As(III) from aqueous solution. PMID:26151463

  8. Polyacrylamide grafting of modified graphene oxides by in situ free radical polymerization

    SciTech Connect

    Tang, Mingyi; Xu, Xiaoyang; Wu, Tao; Zhang, Sai; Li, Xianxian; Li, Yi

    2014-12-15

    Highlights: • Graphene oxide (GO) was modified by chemical reactions to functionalized GO (FGO). • The FGOs and the GO were then subjected to in situ free radical polymerization. • Hydroxyl groups of GO were the most reactive grafting sites. - Abstract: Graphene oxide (GO) was modified using chemical reactions to obtain three types of functionalized GO sheets (FGO). The FGO sheets and the GO were then subjected to in situ free radical polymerization in order to study the grafting polymerization. The FGO and grafted-.FGO were analyzed with Fourier transform infrared spectroscopy, scanning electronic microscopy, thermo-gravimetric analysis (TGA) and X-ray photoelectron spectroscopy (XPS). The grafting percentages in the materials were calculated using the TGA and XPS results. The FGO sheets with different functional groups exhibited different grafting abilities, and hydroxyl groups were proven to be the most reactive grafting sites for the in situ free radical grafting polymerization of polyacrylamide.

  9. The concentration dependence of the solution viscosity and the relaxation time of the partially hydrolyzed polyacrylamides

    SciTech Connect

    Kanatharana, J.; Sukpisan, J.; Wang, S.Q.

    1995-12-01

    The dependences on the polyion concentration through the scaling relations in {eta} {alpha} c{sup {alpha}} and {Tau}{sub q} {alpha} c{sup {beta}}, where {eta} and {Tau}{sub q} are the solution viscosity and the relaxation time obtained from the dynamic light scattering respectively, are investigated for the partially hydrolyzed polyacrylamides at different degrees of hydrolysis. The scaling exponents a and {beta}, as determined in the semidilute regime, depend critically on the amount of salt added or the ionic strength. Both exponents, however, are independent of the amount of glycerol added which suggests that the excluded volume effect is relatively small in comparison with the effect of electrostatic repulsion. The salt-concentration dependence of the solution is also investigated: the corresponding scaling exponents for the 70% HPAM are insensitive to the solvent quality. The present experiment results are compared with recent scaling theories.

  10. Direct capture of lactoferrin from cheese whey on supermacroporous column of polyacrylamide cryogel with copper ions.

    PubMed

    Carvalho, B M A; Carvalho, L M; Silva, W F; Minim, L A; Soares, A M; Carvalho, G G P; da Silva, S L

    2014-07-01

    Lactoferrin is a protein that is present in cheese whey (a waste product from the dairy industry) and has several biological activities. However, its production from whey must have a high yield and low cost for industrial applications. As such, this study reports the use of polyacrylamide cryogel, loaded with Cu(2+) (through the bond with iminodiacetic acid (IDA)), as an adsorbent for the chromatographic process to capture lactoferrin whey. Ultrafiltered cheese whey was passed through the cryogel-IDA-Cu(2+) system. The eluates were subjected to analysis of total protein, SDS-PAGE and size exclusion chromatography. The results showed an axial dispersion coefficients, at different superficial velocities of liquid, in a range of 10(-6)-10(-5)m(2)/s. The cryogel demonstrated good hydraulic permeability (4.708610(-13)m(2)) and a porosity of approximately 78.2%. The IDA-Cu(2+) cryogel system was also able to capture lactoferrin in high purity. PMID:24518347

  11. The Use of Polyacrylamide as a Selective Depressant in the Separation of Chalcopyrite and Galena

    NASA Astrophysics Data System (ADS)

    Wang, Lei

    High molecular weight polyacrylamide (PAM) was tested as a potential selective depressant in the differential flotation separation of galena and chalcopyrite using potassium ethyl xanthate (KEX) as a collector. In single mineral flotation, PAM depressed chalcopyrite while galena was floatable. Mechanism study indicated that PAM could adsorb on galena through hydrogen bonding, and on chalcopyrite through hydrogen bonding as well as ammonium-copper complexation. KEX could only break up the galena-PAM bonding. It is the combined use of PAM and KEX that caused the selectivity. In mineral mixture flotation, galena and chalcopyrite could be separated by PAM and KEX only after EDTA treatment of the mineral mixtures. Time of flight secondary ion mass spectrometric (ToF-SIMS) measurements indicated that when galena and chalcopyrite were present together in the suspension, PAM adsorbed on both galena and chalcopyrite. However, after prior treatment of the mineral mixture by EDTA, PAM mainly adsorbed on chalcopyrite.

  12. Polyacrylamid/silver composite particles produced via microfluidic photopolymerization for single particle-based SERS microsensorics.

    PubMed

    Köhler, J Michael; März, Anne; Popp, Jürgen; Knauer, Andrea; Kraus, Isabelle; Faerber, Jaques; Serra, Christophe

    2013-01-01

    A micro-continuous-flow process was applied for the preparation of swellable polyacrylamide particles incorporating silver nanoparticles. These sensor particles are formed from a mixture of a colloidal solution of silver nanoparticles and monomer by a droplet-based procedure with in situ photoinitiation of polymerization and a subsequent silver enforcement in batch. The obtained polymer composite particles show a strong SERS effect. Characteristic Raman signals of aqueous solutions of adenine could be detected down to 0.1 μM by the use of single sensor particles. The chosen example demonstrates that the composite particles are suitable for quantitative microanalytical procedures with a high dynamic range (3 orders of magnitude for adenine). PMID:23206230

  13. Rheological behavior of aqueous polyacrylamide solutions determined by dissipative particle dynamics and comparison to experiments

    NASA Astrophysics Data System (ADS)

    Yiannourakou, M.; Rousseau, B.; Pannacci, N.; Herzhaft, B.

    2012-02-01

    Based on molecular-dynamics simulations and experimental data, a new coarse-grained forcefield is proposed for the polyacrylamide (PAM)-water system that allows to study dynamical properties of chains at several concentrations with molecular weight up to 17000 g/mol. Non-equilibrium simulations were used to compute relative viscosities, enabling a direct comparison with experimental values. High-shear-rate measurements for low-molecular-weight PAM (10000 g/mol) were done using a microfluidic rheometer Rheosense to decrease the gap between experimental and simulated shear rates. DPD simulations reproduced qualitatively and quantitatively structural properties as well as rheological properties in the dilute regime and qualitatively in the semi-dilute regime.

  14. Effects of bagasse microfibrillated cellulose and cationic polyacrylamide on key properties of bagasse paper.

    PubMed

    Djafari Petroudy, Seyed Rahman; Syverud, Kristin; Chinga-Carrasco, Gary; Ghasemain, Ali; Resalati, Hossein

    2014-01-01

    This study explores the benefits of using bagasse microfibrillated cellulose (MFC) in bagasse paper. Two different types of MFC were produced from DED bleached soda bagasse pulp. The MFC was added to soda bagasse pulp furnishes in different amounts. Cationic polyacrylamide (C-PAM) was selected as retention aid. The results show that addition of MFC increased the strength of paper as expected. Interestingly, 1% MFC in combination with 0.1% C-PAM yielded similar drainage time as the reference pulp, which did not contain MFC. In addition, the samples containing 1% MFC and 0.1% C-PAM yielded (i) a significant increment of the tensile index, (ii) a minor decrease of opacity and (iii) preserved Gurley porosity. Hence, this study proves that small fractions of MFC in combination with adequate retention aids can have positive effects with respect to paper properties, which is most interesting from an industrial point of view. PMID:24274512

  15. Cationic polyacrylamide enhancing cellulase treatment efficiency of hardwood kraft-based dissolving pulp.

    PubMed

    Wang, Qiang; Liu, Shanshan; Yang, Guihua; Chen, Jiachuan; Ni, Yonghao

    2015-05-01

    Cellulase treatment for decreasing viscosity and increasing Fock reactivity of dissolving pulp is a promising approach to reduce the use of toxic chemicals, such as hypochlorite in the dissolving pulp manufacturing process in the industry. Improving the cellulase treatment efficiency during the process is of practical interest. In the present study, the concept of using cationic polyacrylamide (CPAM) to enhance the cellulase treatment efficiency was demonstrated. This was mainly attributed to the increased cellulase adsorption onto cellulose fibers based on the patching/bridging mechanism. Results showed that the cellulase adsorption was increased by about 20% with the addition of 250 ppm of CPAM under the same conditions as those of the control. It was found that the viscosity decrease and Fock reactivity increase for the cellulase treatment was enhanced from using CPAM. The CPAM-assisted cellulase treatment concept may provide a practical alternative to the present hypochlorite-based technology for viscosity control in the industry. PMID:25710682

  16. The structure of high-methoxyl sugar acid gels of citrus pectin as determined by AFM

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Images of native high methoxyl sugar acid gels (HMSAG) were obtained by atomic force microscopy (AFM) in the Tapping ModeTM. Electronic thinning of the pectin strands to one pixel wide allowed the pectin network to be viewed in the absence of variable strand widths related to preferentially solvate...

  17. High transparent shape memory gel

    NASA Astrophysics Data System (ADS)

    Gong, Jin; Arai, Masanori; Kabir, M. H.; Makino, Masato; Furukawa, Hidemitsu

    2014-03-01

    Gels are a new material having three-dimensional network structures of macromolecules. They possess excellent properties as swellability, high permeability and biocompatibility, and have been applied in various fields of daily life, food, medicine, architecture, and chemistry. In this study, we tried to prepare new multi-functional and high-strength gels by using Meso-Decoration (Meso-Deco), one new method of structure design at intermediate mesoscale. High-performance rigid-rod aromatic polymorphic crystals, and the functional groups of thermoreversible Diels-Alder reaction were introduced into soft gels as crosslinkable pendent chains. The functionalization and strengthening of gels can be realized by meso-decorating the gels' structure using high-performance polymorphic crystals and thermoreversible pendent chains. New gels with good mechanical properties, novel optical properties and thermal properties are expected to be developed.

  18. Foam and gel decontamination techniques

    SciTech Connect

    McGlynn, J.F.; Rankin, W.N.

    1989-01-01

    The Savannah River Site is investigating decontamination technology to improve current decontamination techniques, and thereby reduce radiation exposure to plant personnel, reduce uptake of radioactive material, and improve safety during decontamination and decommissioning activities. When decontamination chemicals are applied as foam and gels, the contact time and cleaning ability of the chemical increases. Foam and gel applicators apply foam or gel that adheres to the surface being decontaminated for periods ranging from fifteen minutes (foam) to infinite contact (gel). This equipment was started up in a cold environment. The desired foam and gel consistency was achieved, operators were trained in its proper maintenance and operation, and the foam and gel were applied to walls, ceilings, and hard to reach surfaces. 17 figs.

  19. Gel polymer electrolytes for batteries

    DOEpatents

    Balsara, Nitash Pervez; Eitouni, Hany Basam; Gur, Ilan; Singh, Mohit; Hudson, William

    2014-11-18

    Nanostructured gel polymer electrolytes that have both high ionic conductivity and high mechanical strength are disclosed. The electrolytes have at least two domains--one domain contains an ionically-conductive gel polymer and the other domain contains a rigid polymer that provides structure for the electrolyte. The domains are formed by block copolymers. The first block provides a polymer matrix that may or may not be conductive on by itself, but that can soak up a liquid electrolyte, thereby making a gel. An exemplary nanostructured gel polymer electrolyte has an ionic conductivity of at least 1.times.10.sup.-4 S cm.sup.-1 at 25.degree. C.

  20. Colloidal thermoresponsive gel forming hybrids.

    PubMed

    Liu, Ruixue; Tirelli, Nicola; Cellesi, Francesco; Saunders, Brian R

    2010-09-15

    Colloidal hybrids comprise organic and inorganic components and are attracting considerable attention in the literature. Recently, we reported hybrid anisotropic microsheets that formed thermoresponsive gels in polymer solutions [Liu et al., Langmuir, 25, 490, 2009]. Here, we investigate the composition and properties of these hybrid colloids themselves in detail for the first time. Three different cationic PNIPAm (N-isopropylacrylamide) graft copolymers and two inorganic nanoparticle types (laponite and Ludox silica) were used to prepare a range of hybrids. Anisotropic microsheets only formed when laponite particles were added to the copolymer implying directed self-assembly. Aqueous dispersions of the microsheets spontaneously formed gels at room temperature and these gels were thermoresponsive. They represent a new class of gel forming colloid and are termed thermoresponsive gel forming hybrids. The compositions of the hybrids were determined from thermogravimetric analysis and those that gave gel forming behaviour identified. Variable-temperature rheology experiments showed that the elasticity of the gels increased linearly with temperature. The reversibility of the thermally-triggered changes in gel elasticity was investigated. The concentration dependence of the rheology data was well described by elastic percolation scaling theory and the data could be collapsed onto a master curve. The concentration exponent for the elastic modulus was 2.5. The strong attractive interactions that exist between the dispersed gel forming hybrids was demonstrated by the formation of stable thermoresponsive hybrid hydrogels through casting of hybrid dispersions. PMID:20561633