Factors affecting the separation performance of proteins in capillary electrophoresis.

Science.gov (United States)

Zhu, Yueping; Li, Zhenqing; Wang, Ping; Shen, Lisong; Zhang, Dawei; Yamaguchi, Yoshinori

2018-04-15

Capillary electrophoresis (CE) is an effective tool for protein separation and analysis. Compared with capillary gel electrophoresis (CGE), non-gel sieving capillary electrophoresis (NGSCE) processes the superiority on operation, repeatability and automaticity. Herein, we investigated the effect of polymer molecular weight and concentration, electric field strength, and the effective length of the capillary on the separation performance of proteins, and find that (1) polymer with high molecular weight and concentration favors the separation of proteins, although concentrated polymer hinders its injection into the channel of the capillary due to its high viscosity. (2) The resolution between the adjacent proteins decreases with the increase of electric field strength. (3) When the effective length of the capillary is long, the separation performance improves at the cost of separation time. (4) 1.4% (w/v) hydroxyethyl cellulose (HEC), 100 V/cm voltage and 12 cm effective length offers the best separation for the proteins with molecular weight from 14,400 Da to 97,400 Da. Finally, we employed the optimal electrophoretic conditions to resolve Lysozyme, Ovalbumin, BSA and their mixtures, and found that they were baseline resolved within 15 min. Copyright © 2018 Elsevier B.V. All rights reserved.

Use of electrophoretically separated serum protein fractions for the diagnosis of cardiomyopathy

International Nuclear Information System (INIS)

Siddiqui, Z.H.; Cheema, A.M.

2011-01-01

In an investigation of molecular pathogenesis in cardiovascular diseases, the blood samples of the patients diagnosed for cardiomyopathy (CMP) were obtained from the Punjab Institute of Cardiology, Lahore. Blood samples of the healthy subjects of comparable age group without any history of cardiac ailment were also collected for the control comparisons. The sera of CMP were separated and used for the study of the protein profiles with sodium dodecyle sulfate polyacrylamide gel electrophoresis (SDS-PAGE) in first dimension. Quantification of various protein fractions done by Gene Genius Bio-imaging Gel Documentation System that provide the data of molecular weights and the percent raw volume covered by each of the fractions. The protein fractions that showed significant variation were separated by using the technique of electro blotting and electro elution and run on isoelectric focusing (IEF) in second dimension to determine their isoelectric points. The most pertinent results in the comparison were the significant increase in apolipoprotein B, Ceruloplasmin, apolipoprotein A-I and transthyretin in the sera of patients of CMP compared to healthy subjects. These results show that level of apolipoprotein B, Ceruloplasmin, apolipoprotein A-I and transthyretin are strong predictor of CMP and can also be used for the diagnosis of CMP. (author)

Suitability of two-dimensional electrophoretic protein separations for quantitative detection of mutations

International Nuclear Information System (INIS)

Taylor, J.; Anderson, N.L.; Anderson, N.G.; Gemmell, A.; Giometti, C.S.; Nance, S.L.; Tollaksen, S.L.

1986-01-01

Separation of proteins by two-dimensional electrophoresis (2DE) provides a powerful method for mutagenesis studies, since hundreds of proteins can be monitored simultaneously. In previous mutation studies in which 2DE has been used, only qualitative protein differences were monitored; quantitative protein variations were not evaluated. Although significant differences in protein abundance can be detected by eye, the large number of protein spots present in 2DE patterns together with the large number of individual patterns required for a mutagenesis study would necessitate the use of a computerized analysis system to detect the rare quantitative protein changes indicative of gene deletions or inactivation of genes by point mutations in regulatory genes. A pilot study to search for heritable mutations induced by treatment of mice with either ethylnitrosourea or gamma radiation is underway. Samples are being monitored for quantitative changes that reduce the amount of protein by about 50%. The results of this study indicate that the key methods to improve the application of 2DE to mutation screening are to increase the number of measurable spots (i.e., improve stain sensitivity) and to decrease the spread of values for the volume measurements. Even small improvements in these areas could greatly increase the number of monitorable spots. 9 refs., 4 figs

Glycosaminoglycan blotting on nitrocellulose membranes treated with cetylpyridinium chloride after agarose-gel electrophoretic separation.

Science.gov (United States)

Maccari, Francesca; Volpi, Nicola

2002-09-01

We describe a method for blotting and immobilizing several nonsulfated and sulfated complex polysaccharides on membranes made hydrophilic and positively charged by a cationic detergent after their separation by conventional agarose gel electrophoresis. Nitrocellulose membranes were derivatized with the cationic detergent cetylpyridinium chloride (CPC) and mixtures of glycosaminoglycans (GAGs) were capillary-blotted after their separation in agarose gel electrophoresis in barium acetate/1,2-diaminopropane. Single purified species of variously sulfated polysaccharides were transferred onto the derivatized membranes after electrophoresis with an efficiency of 100% and stained with alcian blue (irreversible staining) and toluidine blue (reversible staining) permitting about 0.1 nug threshold of detection. Nonsulfated polyanions, hyaluronic acid, a fructose-containing polysaccharide with a chondroitin backbone purified from Escherichia coli U1-41, and its defructosylated product, were also electrophoretically separated and transferred onto membranes. The limit of detection for desulfated GAGs was about 0.1-0.5 nug after irreversible or reversible staining. GAG extracts from bovine, lung and aorta, and human aorta and urine were separated by agarose gel electrophoresis and blotted on CPC-treated nitrocellulose membranes. The polysaccharide composition of these extracts was determined. The membrane stained with toluidine blue (reversible staining) was destained and the same lanes used for immunological detection or other applications. Reversible staining was also applied to recover single species of polysaccharides after electrophoretic separation of mixtures of GAGs and their transfer onto membranes. Single bands were released from the membrane with an efficiency of 70-100% for further biochemical characterization.

Electrophoretic Detection and Confocal Microscopic Imaging of Tyrosine Nitrated Proteins in Plant Tissue.

Science.gov (United States)

Arora, Dhara; Singh, Neha; Bhatla, Satish C

2018-01-01

Tyrosine nitrated proteins can be detected in plant cells electrophoretically and their distribution can be monitored by confocal laser scanning microscopy (CLSM) imaging. One-dimensional polyacrylamide gel electrophoresis (1D PAGE) followed by Western blotting using polyclonal antibody against 3-nitrotyrosine residues enables detection of tyrosine nitrated proteins in plant cells. Here we describe detection of tyrosine nitrated proteins in the homogenates derived from sunflower (Helianthus annuus L.) seedling cotyledons. Total soluble proteins obtained from tissue homogenates are resolved using vertical gel electrophoresis followed by their electrophoretic transfer on to a microporous membrane support for immunodetection. Spatial distribution of tyrosine nitrated proteins can be visualized using an antibody against 3-nitrotyrosine residues. Immunofluorescent localization is performed by cutting 7 μm thick wax sections of tissue followed by incubation in primary anti-nitrotyrosine antibody (dilution 1:200) and secondary Cy-3 labeled anti-rabbit IgG antibody (dilution 1:1500). Confocal laser scanning microscopy analysis is undertaken using argon lasers (ex: 530-550 nm and em: 570 nm) at pinhole 1. Modulation in the abundance and spatial localization of tyrosine nitrated proteins in plant tissues can be monitored using these techniques.

Electrophoretic separation of proteins in space

Science.gov (United States)

Brown, R. K.

1976-01-01

Commercially available and synthetic wide range and short range ampholytes used in the isoelectric focusing of proteins was analyzed by ion exchange chromatography. A pH gradient over the pH range 3.8 to 11.0 was used to elute the ampholytes from a column of a sulfonated polystyrene resin. The wide range ampholytes were resolved into some 60 to 70 ninhydrin positive components. The recovery obtained with the method was quantitative. Acid short range ampholytes have approximately 35 components which elute readily from the ion exchange resin. Basic short range ampholytes gave about 50 components, most of which eluted at alkaline pH.

Two-dimensional electrophoretic analysis of nuclear matrix proteins in human colon adenocarcinoma.

Science.gov (United States)

Toumpanaki, A; Baltatzis, G E; Gaitanarou, E; Seretis, E; Toumpanakis, C; Aroni, K; Kittas, Christos; Voloudakis-Baltatzis, I E

2009-01-01

The aim of the present study was to observe possible qualitative and quantitative expression differences between nuclear matrix proteins (NMPs) of human colon adenocarcinoma and their mirror biopsies, using the technique of two-dimensional gel electrophoresis, in order to identify the existence of specific NMP fingerprints for colon cancer. Colon tissues were examined ultrastructurally and NMPs were isolated biochemically, by serial extraction of lipids, soluble proteins, DNA, RNA, and intermediate filaments and were separated according to their isoelectric point (pI) and their molecular weight (MW) by high-resolution two-dimensional electrophoresis (2D). By comparing the 2D electropherograms of colon cancer tissues and mirror biopsy tissues we observed qualitative and quantitative expression differences between their NMPs but also a differentiation of NMP composition between the stages of malignancy. Moreover, despite the similarities between mirror biopsy samples, a highlight percentage of exception was observed. Electrophoretic results provided in this study demonstrated that the examined NMPs could be further investigated as potential markers for detection of colorectal cancer in an early stage, for the assessment of the disease progression, as well as useful tools for individual therapy and for preventing a possible recurrence of cancer and metastasis.

Mass spectrometric analysis of electrophoretically separated allergens and proteases in grass pollen diffusates

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Geczy Carolyn L

2003-09-01

Full Text Available Abstract Background Pollens are important triggers for allergic asthma and seasonal rhinitis, and proteases released by major allergenic pollens can injure airway epithelial cells in vitro. Disruption of mucosal epithelial integrity by proteases released by inhaled pollens could promote allergic sensitisation. Methods Pollen diffusates from Kentucky blue grass (Poa pratensis, rye grass (Lolium perenne and Bermuda grass (Cynodon dactylon were assessed for peptidase activity using a fluorogenic substrate, as well as by gelatin zymography. Following one- or two-dimensional gel electrophoresis, Coomassie-stained individual bands/spots were excised, subjected to tryptic digestion and analysed by mass spectrometry, either MALDI reflectron TOF or microcapillary liquid chromatography MS-MS. Database searches were used to identify allergens and other plant proteins in pollen diffusates. Results All pollen diffusates tested exhibited peptidase activity. Gelatin zymography revealed high Mr proteolytic activity at ~ 95,000 in all diffusates and additional proteolytic bands in rye and Bermuda grass diffusates, which appeared to be serine proteases on the basis of inhibition studies. A proteolytic band at Mr ~ 35,000 in Bermuda grass diffusate, which corresponded to an intense band detected by Western blotting using a monoclonal antibody to the timothy grass (Phleum pratense group 1 allergen Phl p 1, was identified by mass spectrometric analysis as the group 1 allergen Cyn d 1. Two-dimensional analysis similarly demonstrated proteolytic activity corresponding to protein spots identified as Cyn d 1. Conclusion One- and two-dimensional electrophoretic separation, combined with analysis by mass spectrometry, is useful for rapid determination of the identities of pollen proteins. A component of the proteolytic activity in Bermuda grass diffusate is likely to be related to the allergen Cyn d 1.

Diagnostic use of electrophoretically separated serum protein fractions in the patients of angina pectoris

International Nuclear Information System (INIS)

Siddiqui, Z.H.; Cheema, A.M.

2010-01-01

The understanding of molecular pathogenesis of clinical states enables for diagnosis and effective management of the diseases. In an investigation of molecular pathogenesis or adaptation in cardiovascular diseases, the blood samples of the patients diagnosed for angina pectoris (AP) were obtained from the Punjab Institute of Cardiology, Lahore. Blood samples of the healthy subjects of comparable age group without any history of cardiac ailment were also collected for the control comparisons. The sera of AP were separated and used for the study of the protein profiles with sodium dodecyles sulfate polyacrylamide gel electrophoresis (SDS-PAGE)in first dimension. Quantification of various protein fractions done by Gene Genius Bio-imaging Gel Documentation System that provide the data of molecular weights and the percent raw volume covered. by each of the fractions. The protein fractions that showed significant variation were separated by using the technique of electro blotting and electro elution and run on isoelectric focusing (IEF) in second dimension to determine their isoelectric points. The most pertinent results in the comparison were the significant increase in apolipoprotein B, marked decrease in apolipoprotein A-I and high apolipoprotein B/apolipoprotein A-I ratio in the sera of patients of AP compared to healthy subjects. These results show that level of apolipoprotein A-I, apolipoprotein B and the apolipoprotein B/apolipoprotein A-I ratio are strong predictor of AP and can also be used for the diagnosis of AP. (author)

Separation of very hydrophobic analytes by micellar electrokinetic chromatography IV. Modeling of the effective electrophoretic mobility from carbon number equivalents and octanol-water partition coefficients.

Science.gov (United States)

Huhn, Carolin; Pyell, Ute

2008-07-11

It is investigated whether those relationships derived within an optimization scheme developed previously to optimize separations in micellar electrokinetic chromatography can be used to model effective electrophoretic mobilities of analytes strongly differing in their properties (polarity and type of interaction with the pseudostationary phase). The modeling is based on two parameter sets: (i) carbon number equivalents or octanol-water partition coefficients as analyte descriptors and (ii) four coefficients describing properties of the separation electrolyte (based on retention data for a homologous series of alkyl phenyl ketones used as reference analytes). The applicability of the proposed model is validated comparing experimental and calculated effective electrophoretic mobilities. The results demonstrate that the model can effectively be used to predict effective electrophoretic mobilities of neutral analytes from the determined carbon number equivalents or from octanol-water partition coefficients provided that the solvation parameters of the analytes of interest are similar to those of the reference analytes.

Electrophoretic pattern of blood serum proteins of some of the vertebrates of Pakistan

International Nuclear Information System (INIS)

Shakoori, Abdul Rauf; Zaheer, Saleem Akhtar; Ahmad, Muhammad Salih.

1976-01-01

The electrophoretic pattern of blood serum proteins of some of the common fishes e.g. Catla catla, Cirrhina mrigala, Channa punctatus, Channa marulius, Wallago attu, Heterop-neustes fossilis; amphibia e.g., Rana tigrina, Rana cyanophlyctis, Bufo melanostictus; reptiles e.g. Varanus bengalensis, Uromastix hardwickii; birds e.g. Columba livia, Gallus domesticus, Passer domestica, Anas platyrhynchos; and mammals e.g. Homo sapiens, Mus musculus, Lepus cuniculus have been described. The mobility of proteins of blood sera has been studied over cellulose acetate paper and then a comparative pattern analysed

Electrophoretic studies on rape seed proteins

International Nuclear Information System (INIS)

Chaudry, M.A.; Starr, A.; Bibi, N.

1992-07-01

Electrophoresis is a technique which separates biological molecules on the basis of charge and mass properties. The technique is used for separation, purification, characterization and identification of molecules/ compounds. Two major objectives for applications of electrophoresis have been studied in this report i.e. characterization of rape seed proteins and enzymes and identification of rape seed cultivars by polyacrylamide gel electrophoresis (PAGE). Gamma irradiation is being successfully used to create genetic variability and germination which brought about definite changes in the rape seed proteins reflected in different bands. These differences could be used to study variability in crop plants. (A.B.)

Development of the resolution theory for electrophoretic exclusion.

Science.gov (United States)

Kenyon, Stacy M; Keebaugh, Michael W; Hayes, Mark A

2014-09-01

Electrophoretic exclusion, a technique that differentiates species in bulk solution near a channel entrance, has been demonstrated on benchtop and microdevice designs. In these systems, separation occurs when the electrophoretic velocity of one species is greater than the opposing hydrodynamic flow, while the velocity of the other species is less than that flow. Although exclusion has been demonstrated in multiple systems for a range of analytes, a theoretical assessment of resolution has not been addressed. To compare the results of these calculations to traditional techniques, the performance is expressed in terms of smallest difference in electrophoretic mobilities that can be completely separated (R = 1.5). The calculations indicate that closest resolvable species (Δμmin ) differ by approximately 10(-13) m(2) /Vs and peak capacity (nc ) is 1000. Published experimental data were compared to these calculated results. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Post-Electrophoretic Identification of Oxidized Proteins

Science.gov (United States)

Conrad, Craig C; Talent, John M; Malakowsky, Christina A

1999-01-01

The oxidative modification of proteins has been shown to play a major role in a number of human diseases. However, the ability to identify specific proteins that are most susceptible to oxidative modifications is difficult. Separation of proteins using polyacrylamide gel electrophoresis (PAGE) offers the analytical potential for the recovery, amino acid sequencing, and identification of thousands of individual proteins from cells and tissues. We have developed a method to allow underivatized proteins to be electroblotted onto PVDF membranes before derivatization and staining. Since both the protein and oxidation proteins are quantifiable, the specific oxidation index of each protein can be determined. The optimal sequence and conditions for the staining process are (a) electrophoresis, (b) electroblotting onto PVDF membranes, (c) derivatization of carbonyls with 2,4-DNP, (d) immunostaining with anti DNP antibody, and (e) protein staining with colloidal gold. PMID:12734585

Three-dimensional fluorescence analysis of chernozem humic acids and their electrophoretic fractions

Science.gov (United States)

Trubetskoi, O. A.; Trubetskaya, O. E.

2017-09-01

Polyacrylamide gel electrophoresis in combination with size-exclusion chromatography (SEC-PAGE) has been used to obtain stable electrophoretic fractions of different molecular size (MS) from chernozem humic acids (HAs). Three-dimensional fluorescence charts of chernozem HAs and their fractions have been obtained for the first time, and all fluorescence excitation-emission maxima have been identified in the excitation wavelength range of 250-500 nm. It has been found that fractionation by the SEC-PAGE method results in a nonuniform distribution of protein- and humin-like fluorescence of the original HA preparation among the electrophoretic fractions. The electrophoretic fractions of the highest and medium MSs have only the main protein-like fluorescence maximum and traces of humin-like fluorescence. In the electrophoretic fraction of the lowest MS, the intensity of protein-like fluorescence is low, but the major part of humin-like fluorescence is localized there. Relationships between the intensity of protein-like fluorescence and the weight distribution of amino acids have been revealed, as well as between the degree of aromaticity and the intensity of humin-like fluorescence in electrophoretic fractions of different MSs. The obtained relationships can be useful in the interpretation of the spatial structural organization and ecological functions of soil HAs.

Effect of surfactant species and electrophoretic medium composition on the electrophoretic behavior of neutral and water-insoluble linear synthetic polymers in nonaqueous capillary zone electrophoresis.

Science.gov (United States)

Fukai, Nao; Kitagawa, Shinya; Ohtani, Hajime

2017-07-01

We have recently demonstrated the separation of neutral and water-insoluble linear synthetic polymers in nonaqueous capillary zone electrophoresis (NACZE) using a cationic surfactant of cetyltrimethylammonium chloride (CTAC). In this study, eight ionic surfactants were investigated for the separation of four synthetic polymers (polystyrene, polymethylmethacrylates, polybutadiene, and polycarbonate); only three surfactants (CTAC, dimethyldioctadecylammonium bromide, and sodium dodecylsulfate) caused their separation. The order of the interaction between the polymers and the surfactants depended on both the surfactant species and the composition of the electrophoretic medium. Their investigation revealed that the separation is majorly affected by the hydrophobic interactions between the polymers and the ionic surfactants. In addition, the electrophoretic behavior of polycarbonate suggested that electrostatic interaction also affects the selectivity of the polymers. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

The electrophoretic mobility shift assay (EMSA)

OpenAIRE

sprotocols

2015-01-01

The electrophoretic mobility shift assay (EMSA), also known as “gel shift assay”, is used to examine the binding parameters and relative affinities of protein and DNA interactions. We produced recombinant CCA1 protein and tested its binding affinity for the promoter fragments that contain CBS (AAAAATCT) or evening element (EE, AAAATATCT) (1) using a modified procedure adopted from published protocols (2,3).

Electrophoretic protein profiles of mid-sized copepod Calanoides patagoniensis steadily fed bloom-forming diatoms

Directory of Open Access Journals (Sweden)

Victor M Aguilera

2015-09-01

Full Text Available Recent field and experimental evidence collected in the southern upwelling region off Concepción (36°5'S, 73°3'W showed an abrupt reduction (<72 h in the egg production rates (EPR of copepods when they were fed steadily and solely with the local bloom-forming diatom Thalassiosira rotula. Because diatoms were biochemically similar to dinoflagellate Prorocentrum minimum, a diet which supported higher reproductive outcomes, the fecundity reduction observed in copepod females fed with the diatom may have obeyed to post-ingestive processes, giving rise to resources reallocation. This hypothesis was tested by comparing feeding (clearance and ingestion rates, reproduction (EPR and hatching success and the structure of protein profiles (i.e., number and intensity of electrophoretic bands of copepods (adults and eggs incubated during 96 h with the two food conditions. The structure of protein profiles included molecular sizes that were calculated from the relative mobility of protein standards against the logarithm of their molecular sizes. After assessing the experimental conditions, feeding decreased over time for those females fed with T. rotula, while reproduction was higher in females fed with P. minimum. Electrophoretic profiles resulted similar mostly at a banding region of 100 to 89-kDa, while they showed partial differences around the region of 56-kDa band, especially in those females fed and eggs produced with T. rotula. Due to reproductive volume was impacted while larvae viability, a physiological processes with specific and high nutritional requirements, was independent on food type; post-ingestive processes, such as expression of stress-related proteins deviating resources to metabolic processes others than reproduction, are discussed under framework of nutritional-toxic mechanisms mediating copepod-diatoms relationships in productive upwelling areas.

Serum Protein Electrophoretic Pattern in Neonatal Calves Treated with Clinoptilolite

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Simona Marc

2018-05-01

Full Text Available The objective of our study was to determine the effects of clinoptilolite supplemented in colostrum on the blood serum protein electrophoretic pattern of new-born calves. Methods: Romanian Black and White new-born calves involved in the study were divided into 3 groups: the control group (C that received colostrum without clinoptilolite, and experimental groups I (E1 and II (E2 that received colostrum supplemented with 0.5% and 2% clinoptilolite, respectively. The concentration of total protein and protein fractions (albumin, α1-globulin, α2-globulin, β-globulin and γ-globulin were analyzed by electrophoresis on cellulose acetate. Results: At hour 30 after birth, concentrations of γ-globulins, β-globulin and total protein in E1 group of calves were higher than in control group by 42.11% (p < 0.05, 28.48% (p > 0.05 and 18.52% (p > 0.05, respectively, and were higher, but not significantly, in group E2 compared to the control group. This was in accordance with a significant lower albumin/globulin ratio in groups E1 and E2 (29.35%, p < 0.05 and 35.87%, p < 0.05, respectively than in control group at 30 h postpartum, which indicates an obvious increase of the globulins fraction in experimental groups. The conclusion: Clinoptilolite was effective in improving passive transfer in new-born calves, but it was more effective if added in colostrum with a dose of 0.5% than with a dose of 2%.

Strategies for the capillary electrophoretic separation of indole alkaloids in Psilocybe semilanceata.

Science.gov (United States)

Pedersen-Bjergaard, S; Rasmussen, K E; Sannes, E

1998-01-01

While the hallucinogenic mushrooms Psilocybe semilanceata have previously been analyzed for the indole alkaloids psilocybin and baeocystin by capillary zone electrophoresis (CZE) at pH 11.5, the present work focused on the development of an alternative and complementary capillary electrophoretic method for their identification. Owing to their structural similarity and zwitterionic nature, the compounds were difficult to resolve based on different interactions with cationic or anionic micelles. However, while the attempts with micellar electrokinetic chromatography (MEKC) were unsuccessful, rapid derivatization with propyl chloroformate and reanalysis by CZE at pH 11.5 was effective to support identification of the two indole alkaloids. Psilocin was difficult to analyze by CZE at pH 11.5 owing to comigration with the electroosmotic flow. For this compound, the pH of the running buffer was reduced to 7.2 to effectively enhance the electrophoretic mobility.

Diamond electrophoretic microchips-Joule heating effects

International Nuclear Information System (INIS)

Karczemska, Anna T.; Witkowski, Dariusz; Ralchenko, Victor; Bolshakov, Andrey; Sovyk, Dmitry; Lysko, Jan M.; Fijalkowski, Mateusz; Bodzenta, Jerzy; Hassard, John

2011-01-01

Microchip electrophoresis (MCE) has become a mature separation technique in the recent years. In the presented research, a polycrystalline diamond electrophoretic microchip was manufactured with a microwave plasma chemical vapour deposition (MPCVD) method. A replica technique (mould method) was used to manufacture microstructures in diamond. A numerical analysis with CoventorWare TM was used to compare thermal properties during chip electrophoresis of diamond and glass microchips of the same geometries. Temperature distributions in microchips were demonstrated. Thermal, electrical, optical, chemical and mechanical parameters of the polycrystalline diamond layers are advantageous over traditionally used materials for microfluidic devices. Especially, a very high thermal conductivity coefficient gives a possibility of very efficient dissipation of Joule heat from the diamond electrophoretic microchip. This enables manufacturing of a new generation of microdevices.

Diamond electrophoretic microchips-Joule heating effects

Energy Technology Data Exchange (ETDEWEB)

Karczemska, Anna T., E-mail: anna.karczemska@p.lodz.pl [Technical University of Lodz, Institute of Turbomachinery, 219/223 Wolczanska str., Lodz (Poland); Witkowski, Dariusz [Technical University of Lodz, Institute of Turbomachinery, 219/223 Wolczanska str., Lodz (Poland); Ralchenko, Victor, E-mail: ralchenko@nsc.gpi.ru [General Physics Institute, Russian Academy of Science, 38 Vavilov str., Moscow (Russian Federation); Bolshakov, Andrey; Sovyk, Dmitry [General Physics Institute, Russian Academy of Science, 38 Vavilov str., Moscow (Russian Federation); Lysko, Jan M., E-mail: jmlysko@ite.waw.pl [Institute of Electron Technology, Al. Lotnikow 32/46, 02-668 Warsaw (Poland); Fijalkowski, Mateusz, E-mail: petr.louda@vslib.cz [Technical University of Liberec, Faculty of Mechanical Engineering (Czech Republic); Bodzenta, Jerzy, E-mail: jerzy.bodzenta@polsl.pl [Silesian University of Technology, Institute of Physics, 2 Krzywoustego str., 44-100 Gliwice (Poland); Hassard, John, E-mail: j.hassard@imperial.ac.uk [Imperial College of Science, Technology and Medicine, London (United Kingdom)

2011-03-15

Microchip electrophoresis (MCE) has become a mature separation technique in the recent years. In the presented research, a polycrystalline diamond electrophoretic microchip was manufactured with a microwave plasma chemical vapour deposition (MPCVD) method. A replica technique (mould method) was used to manufacture microstructures in diamond. A numerical analysis with CoventorWare{sup TM} was used to compare thermal properties during chip electrophoresis of diamond and glass microchips of the same geometries. Temperature distributions in microchips were demonstrated. Thermal, electrical, optical, chemical and mechanical parameters of the polycrystalline diamond layers are advantageous over traditionally used materials for microfluidic devices. Especially, a very high thermal conductivity coefficient gives a possibility of very efficient dissipation of Joule heat from the diamond electrophoretic microchip. This enables manufacturing of a new generation of microdevices.

Electrophoretic mobilities of dissolved polyelectrolyte charging agent and suspended non-colloidal titanium during electrophoretic deposition

International Nuclear Information System (INIS)

Lau, Kok-Tee; Sorrell, C.C.

2011-01-01

Coarse (≤20 μm) titanium particles were deposited on low-carbon steel substrates by cathodic electrophoretic deposition (EPD) with ethanol as suspension medium and poly(diallyldimethylammonium chloride) (PDADMAC) as polymeric charging agent. Preliminary data on the electrophoretic mobilities and electrical conductivities on the suspensions of these soft particles as well as the solutions themselves as a function of PDADMAC level were used as the basis for the investigation of the EPD parameters in terms of the deposition yield as a function of five experimental parameters: (a) PDADMAC addition level, (b) solids loading, (c) deposition time, (d) applied voltage, and (e) electrode separation. These data were supported by particle sizing by laser diffraction and deposit surface morphology by scanning electron microscopy (SEM). The preceding data demonstrated that Ti particles of ∼1-12 μm size, electrosterically modified by the PDADMAC charging agent, acted effectively as colloidal particles during EPD. Owing to the non-colloidal nature of the particles and the stabilization of the Ti particles by electrosteric forces, the relevance of the zeta potential is questionable, so the more fundamental parameter of electrophoretic mobility was used. A key finding from the present work is the importance of assessing the electrophoretic mobilities of both the suspensions and solutions since the latter, which normally is overlooked, plays a critical role in the ability to interpret the results meaningfully. Further, algebraic uncoupling of these data plus determination of the deposit yield as a function of charging agent addition allow discrimination between the three main mechanistic stages of the electrokinetics of the process, which are: (1) surface saturation; (2) compression of the diffuse layer, growth of polymer-rich layer, and/or competition between the mobility of Ti and PDADMAC; and (3) little or no decrease in electrophoretic mobility of Ti, establishment of

Electrophoretic mobilities of dissolved polyelectrolyte charging agent and suspended non-colloidal titanium during electrophoretic deposition

Energy Technology Data Exchange (ETDEWEB)

Lau, Kok-Tee [School of Materials Science and Engineering, University of New South Wales, Sydney, NSW 2052 (Australia); Faculty of Manufacturing Engineering, Universiti Teknikal Malaysia Melaka, 76109 Durian Tunggal, Melaka (Malaysia); Sorrell, C.C., E-mail: C.Sorrell@unsw.edu.au [School of Materials Science and Engineering, University of New South Wales, Sydney, NSW 2052 (Australia)

2011-03-25

Coarse ({<=}20 {mu}m) titanium particles were deposited on low-carbon steel substrates by cathodic electrophoretic deposition (EPD) with ethanol as suspension medium and poly(diallyldimethylammonium chloride) (PDADMAC) as polymeric charging agent. Preliminary data on the electrophoretic mobilities and electrical conductivities on the suspensions of these soft particles as well as the solutions themselves as a function of PDADMAC level were used as the basis for the investigation of the EPD parameters in terms of the deposition yield as a function of five experimental parameters: (a) PDADMAC addition level, (b) solids loading, (c) deposition time, (d) applied voltage, and (e) electrode separation. These data were supported by particle sizing by laser diffraction and deposit surface morphology by scanning electron microscopy (SEM). The preceding data demonstrated that Ti particles of {approx}1-12 {mu}m size, electrosterically modified by the PDADMAC charging agent, acted effectively as colloidal particles during EPD. Owing to the non-colloidal nature of the particles and the stabilization of the Ti particles by electrosteric forces, the relevance of the zeta potential is questionable, so the more fundamental parameter of electrophoretic mobility was used. A key finding from the present work is the importance of assessing the electrophoretic mobilities of both the suspensions and solutions since the latter, which normally is overlooked, plays a critical role in the ability to interpret the results meaningfully. Further, algebraic uncoupling of these data plus determination of the deposit yield as a function of charging agent addition allow discrimination between the three main mechanistic stages of the electrokinetics of the process, which are: (1) surface saturation; (2) compression of the diffuse layer, growth of polymer-rich layer, and/or competition between the mobility of Ti and PDADMAC; and (3) little or no decrease in electrophoretic mobility of Ti

Lab-on-fiber electrophoretic trace mixture separating and detecting an optofluidic device based on a microstructured optical fiber.

Science.gov (United States)

Yang, Xinghua; Guo, Xiaohui; Li, Song; Kong, Depeng; Liu, Zhihai; Yang, Jun; Yuan, Libo

2016-04-15

We report an in-fiber integrated electrophoretic trace mixture separating and detecting an optofluidic optical fiber sensor based on a specially designed optical fiber. In this design, rapid in situ separation and simultaneous detection of mixed analytes can be realized under electro-osmotic flow in the microstructured optical fiber. To visually display the in-fiber separating and detecting process, two common fluorescent indicators are adopted as the optofluidic analytes in the optical fiber. Results show that a trace amount of the mixture (0.15 μL) can be completely separated within 3.5 min under a high voltage of 5 kV. Simultaneously, the distributed information of the separated analytes in the optical fiber can be clearly obtained by scanning along the optical fiber using a 355 nm laser. The emission from the analytes can be efficiently coupled into the inner core and guides to the remote end of the optical fiber. In addition, the thin cladding around the inner core in the optical fiber can prevent the fluorescent cross talk between the analytes in this design. Compared to previous optical fiber optofluidic devices, this device first realizes simultaneously separating treatment and the detection of the mixed samples in an optical fiber. Significantly, such an in-fiber integrated separating and detecting optofluidic device can find wide applications in various analysis fields involves mixed samples, such as biology, chemistry, and environment.

Application of native agarose gel electrophoresis of serum proteins in veterinary diagnostics

Directory of Open Access Journals (Sweden)

Jania Bartosz

2016-12-01

Full Text Available Electrophoretic techniques, used to separate mixtures of electrically charged particles, are widely used in science. One of these techniques, native protein electrophoresis in an agarose gel, is applied in human and veterinary medicine. Changes in the proportions of individual protein fractions correspond to significant changes in the physiology of the body. Although the pattern obtained by electrophoretic separation rarely indicates a specific disease, it provides valuable information for the differential diagnosis. Decades of research on the types of patterns obtained in the case of particular diseases have led to the accumulation of substantial knowledge. The paper presents the available information on this topic. Serum protein electrophoresis is recommended in cases of increased levels of total protein in order to reveal the nature of the process. The basic information which can be obtained from electrophoretic separation includes the immune status of the organism. Both increased antigenic stimulation and immunodeficiency are clearly visible in electropherograms. Moreover, the level of heterogeneity of the corresponding protein fractions can help to distinguish between infectious diseases and cancer - multiple myeloma - the latter producing a homogeneous immunoglobulin fraction. Analysis of other protein fractions helps to detect or confirm an ongoing inflammatory process and provides information regarding liver function. Even when the concentration of total protein is within the reference range, this analysis can be recommended as a basic laboratory test.

A genetic electrophoretic variant of high-sulfur hair proteins for forensic hair comparisons. I. Characterization of variant high-sulfur proteins of human hair.

Science.gov (United States)

Miyake, B

1989-02-01

In a survey of the proteins from human hair, a genetic electrophoretic variant has been observed in the high-sulfur protein region. S-carboxymethylated proteins were examined by 15% polyacrylamide gel electrophoresis at pH 8.9. Out of 150 unrelated samples of Japanese head hairs analyzed, 107 showed 6 major high-sulfur protein bands (normal) and the remaining 43 samples showed an additional high-sulfur protein band (variant). Of 21 Caucasian samples analyzed only one variant sample was found. Characterization of the proteins by two-dimensional electrophoresis evidenced a variant protein spot which showed an apparent molecular weight of 30 k Da. Isoelectric points of the high-sulfur proteins ranged from 3.25-3.55 and that of variant protein band from 3.3-3.4. Family studies of 21 matings resulting in 49 children indicated that this variant was inherited in an autosomal fashion.

Sialic acid accelerates the electrophoretic velocity of injured dorsal root ganglion neurons

Directory of Open Access Journals (Sweden)

Chen-xu Li

2015-01-01

Full Text Available Peripheral nerve injury has been shown to result in ectopic spontaneous discharges on soma and injured sites of sensory neurons, thereby inducing neuropathic pain. With the increase of membrane proteins on soma and injured site neurons, the negatively charged sialic acids bind to the external domains of membrane proteins, resulting in an increase of this charge. We therefore speculate that the electrophoretic velocity of injured neurons may be faster than non-injured neurons. The present study established rat models of neuropathic pain via chronic constriction injury. Results of the cell electrophoresis test revealed that the electrophoretic velocity of injured neuronal cells was faster than that of non-injured (control cells. We then treated cells with divalent cations of Ca 2+ and organic compounds with positive charges, polylysine to counteract the negatively charged sialic acids, or neuraminidase to specifically remove sialic acids from the membrane surface of injured neurons. All three treatments significantly reduced the electrophoretic velocity of injured neuronal cells. These findings suggest that enhanced sialic acids on injured neurons may accelerate the electrophoretic velocity of injured neurons.

Electrophoretic mobility shift assay reveals a novel recognition sequence for Setaria italica NAC protein.

Science.gov (United States)

Puranik, Swati; Kumar, Karunesh; Srivastava, Prem S; Prasad, Manoj

2011-10-01

The NAC (NAM/ATAF1,2/CUC2) proteins are among the largest family of plant transcription factors. Its members have been associated with diverse plant processes and intricately regulate the expression of several genes. Inspite of this immense progress, knowledge of their DNA-binding properties are still limited. In our recent publication,1 we reported isolation of a membrane-associated NAC domain protein from Setaria italica (SiNAC). Transactivation analysis revealed that it was a functionally active transcription factor as it could stimulate expression of reporter genes in vivo. Truncations of the transmembrane region of the protein lead to its nuclear localization. Here we describe expression and purification of SiNAC DNA-binding domain. We further report identification of a novel DNA-binding site, [C/G][A/T][T/A][G/C]TC[C/G][A/T][C/G][G/C] for SiNAC by electrophoretic mobility shift assay. The SiNAC-GST protein could bind to the NAC recognition sequence in vitro as well as to sequences where some bases had been reshuffled. The results presented here contribute to our understanding of the DNA-binding specificity of SiNAC protein.

Role of adiponectin/phosphatidylinositol 3-kinase/protein kinase B ...

African Journals Online (AJOL)

2012-06-19

Jun 19, 2012 ... Lysis buffer (SDS Lysis Buffer) was added to the samples overnight at 4°C. An equal amount of protein was loaded by the Coomassie method for protein quantification after electrophoretic separation. Protein was transferred onto polyvinylidene fluoride (PVDF) membranes. The transferred blot was blocked ...

Electrophoretic Retardation of Colloidal Particles in Nonpolar Liquids

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Filip Strubbe

2013-04-01

Full Text Available We have measured the electrophoretic mobility of single, optically trapped colloidal particles, while gradually depleting the co-ions and counterions in the liquid around the particle by applying a dc voltage. This is achieved in a nonpolar liquid, where charged reverse micelles act as co-ions and counterions. By increasing the dc voltage, the mobility first increases when the concentrations of co-ions and counterions near the particle start to decrease. At sufficiently high dc voltage (around 2 V, the mobility reaches a saturation value when the co-ions and counterions are fully separated. The increase in mobility is larger when the equilibrium ionic strength is higher. The dependence of the experimental data on the equilibrium ionic strength and on the applied voltage is in good agreement with the standard theory of electrophoretic retardation, assuming that the bare particle charge remains constant. This method is useful for studying the electrophoretic retardation effect and charging mechanisms for nonpolar colloids, and it sheds light on previously unexplained particle acceleration in electronic ink devices.

Electrophoretic transport of biomolecules across liquid-liquid interfaces

Energy Technology Data Exchange (ETDEWEB)

Hahn, Thomas; Hardt, Steffen [Center of Smart Interfaces, TU Darmstadt, Petersenstrasse 32, D-64287 Darmstadt (Germany); Muenchow, Goetz, E-mail: hardt@csi.tu-darmstadt.de [Institut fuer Mikrotechnik Mainz GmbH, Carl-Zeiss-Strasse 18-20, D-55129 Mainz (Germany)

2011-05-11

The mass transfer resistance of a liquid-liquid interface in an aqueous two-phase system composed of poly(ethylene glycol) and dextran is investigated. Different types of proteins and DNA stained with fluorescent dyes serve as probes to study the transport processes close to the interface. A microfluidic device is employed to enable the electrophoretic transport of biomolecules from one phase to another. The results obtained for proteins can be explained solely via the different electrophoretic mobilities and different affinities of the molecules to the two phases, without any indications of a significant mass transfer resistance of the liquid-liquid interface. By contrast, DNA molecules adsorb to the interface and only desorb under an increased electric field strength. The desorption process carries the signature of a thermally activated escape from a metastable state, as reflected in the exponential decay of the fluorescence intensity at the interface as a function of time.

Optimization of the southern electrophoretic transfer method

International Nuclear Information System (INIS)

Allison, M.A.; Fujimura, R.K.

1987-01-01

The technique of separating DNA fragments using agarose gel electrophoresis is essential in the analysis of nucleic acids. Further, after the method of transferring specific DNA fragments from those agarose gels to cellulose nitrate membranes was developed in 1975, a method was developed to transfer DNA, RNA, protein and ribonucleoprotein particles from various gels onto diazobenzyloxymethyl (DBM) paper using electrophoresis as well. This paper describes the optimum conditions for quantitative electrophoretic transfer of DNA onto nylon membranes. This method exemplifies the ability to hybridize the membrane more than once with specific RNA probes by providing sufficient retention of the DNA. Furthermore, the intrinsic properties of the nylon membrane allow for an increase in the efficiency and resolution of transfer while using somewhat harsh alkaline conditions. The use of alkaline conditions is of critical importance since we can now denature the DNA during transfer and thus only a short pre-treatment in acid is required for depurination. 9 refs., 7 figs

Acetonitrile as a buffer additive for free zone capillary electrophoresis separation and characterization of maize (Zeamays L. ) and sorghum (Sorghum bicolor L. Moench) storage proteins.

Science.gov (United States)

Bean, S R; Lookhart, G L; Bietz, J A

2000-02-01

An improved method for separating and characterizing maize (Zea mays L.) and sorghum (Sorghum bicolor L. Moench) storage proteins by free zone capillary electrophoresis (FZCE) was developed. Previous electrophoretic methods for analyzing these proteins required high concentrations of urea to maintain protein solubility during separation. To overcome disadvantages of urea, we developed a FZCE method that mimicked reversed-phase high-performance liquid chromatography (RP-HPLC) in that it used high levels of acetonitrile (ACN) at low pH. The optimized FZCE buffer system consisted of 80 mM phosphate-glycine buffer, nominal pH 2.5, containing 60% ACN and a cellulose derivative to dynamically coat capillary walls. Resolution was similar to or higher than that previously achieved by FZCE buffers utilizing 8 M urea as a buffer additive. ACN concentrations of at least 50% were necessary to achieve acceptable separations; this ACN concentration is approximately that necessary to extract these storage proteins. ACN was equally effective as traditional ethanol solvents and 8 M urea for solubilizing maize and sorghum proteins. The ACN-based FZCE buffer system gave high repeatability (buffers. This FZCE method may be applicable for the analysis of other hydrophobic proteins without the use of urea.

Validation for chromatographic and electrophoretic methods

OpenAIRE

Ribani, Marcelo; Bottoli, Carla Beatriz Grespan; Collins, Carol H.; Jardim, Isabel Cristina Sales Fontes; Melo, Lúcio Flávio Costa

2004-01-01

The validation of an analytical method is fundamental to implementing a quality control system in any analytical laboratory. As the separation techniques, GC, HPLC and CE, are often the principal tools used in such determinations, procedure validation is a necessity. The objective of this review is to describe the main aspects of validation in chromatographic and electrophoretic analysis, showing, in a general way, the similarities and differences between the guidelines established by the dif...

Protein content and electrophoretic profile of fat body and ovary extracts from workers of Melipona quadrifasciata anthidioides (Hymenoptera, Meliponini

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Vagner T. Paes de Oliveira

Full Text Available Workers of Melipona quadrifasciata anthidioides (Lepeletier, 1836 develop their ovaries and lay eggs, therefore the production of vitellogenin is expected. In electrophoretic profiles only fat body extracts from nurse workers and ovary extracts from newly-emerged workers show protein with molecular mass similar to vitellogenin. However, an increase in the protein content was detected in forager fat body. This increase was attributed to storage of vitellogenin or other proteins in the previous phase and not discharged into the hemolymph or to an effect of the increased titre of juvenile hormone in this phase of worker life over the fat body functioning.

Canine serum protein patterns using high-resolution electrophoresis (HRE).

Science.gov (United States)

Abate, O; Zanatta, R; Malisano, T; Dotta, U

2000-03-01

Serum protein values were determined in 26 healthy dogs using agarose gel electrophoresis (SPE), splitting the electrophoretic separation into six regions: albumin, alpha(1), alpha(2), beta(1), beta(2)and gamma globulins. High-resolution electrophoresis (HRE) was used to separate single proteins. Serum proteins from dogs (26 healthy and 20 affected by various diseases) were then characterized by electrophoretic immunofixation (IFE) and Sudan black staining on HRE film. Haemoglobin and normal canine plasma and serum were used to identify haptoglobin and fibrinogen, respectively. In the standard pattern, determined by HRE, the following proteins were identified: albumin, alpha(1)-lipoprotein (alpha(1)-region), haptoglobin and alpha(2)-macroglobulin (alpha(2)-region), beta -lipoprotein and C3 (beta(1)-region), transferrin and IgM (beta(2)-region), IgG (mostly in gamma -region and partly in beta(2)-region). The HRE pattern shown by healthy dogs could be compared with those of dogs affected by various diseases to obtain clinical information. Copyright 2000 Harcourt Publishers Ltd.

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

Science.gov (United States)

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. Copyright © 2015 Elsevier Inc. All rights reserved.

Improved Peak Capacity for Capillary Electrophoretic Separations of Enzyme Inhibitors with Activity-Based Detection Using Magnetic Bead Microreactors

Science.gov (United States)

Yan, Xiaoyan; Gilman, S. Douglass

2010-01-01

A technique for separating and detecting enzyme inhibitors was developed using capillary electrophoresis with an enzyme microreactor. The on-column enzyme microreactor was constructed using NdFeB magnet(s) to immobilize alkaline phosphatase-coated superparamagnetic beads (2.8 μm diameter) inside a capillary before the detection window. Enzyme inhibition assays were performed by injecting a plug of inhibitor into a capillary filled with the substrate, AttoPhos. Product generated in the enzyme microreactor was detected by laser-induced fluorescence. Inhibitor zones electrophoresed through the capillary, passed through the enzyme microreactor, and were observed as negative peaks due to decreased product formation. The goal of this study was to improve peak capacities for inhibitor separations relative to previous work, which combined continuous engagement electrophoretically mediated microanalysis (EMMA) and transient engagement EMMA to study enzyme inhibition. The effects of electric field strength, bead injection time and inhibitor concentrations on peak capacity and peak width were investigated. Peak capacities were increased to ≥20 under optimal conditions of electric field strength and bead injection time for inhibition assays with arsenate and theophylline. Five reversible inhibitors of alkaline phosphatase (theophylline, vanadate, arsenate, L-tryptophan and tungstate) were separated and detected to demonstrate the ability of this technique to analyze complex inhibitor mixtures. PMID:20024913

µE: Electrophoretic mobility

Indian Academy of Sciences (India)

First page Back Continue Last page Graphics. µE: Electrophoretic mobility. µE: Electrophoretic mobility. E: Intensity of electric field. H: Total height. h: Distance from the top surface of bottom chamber (slug height). N: Cell concentration × Volume of the chamber.

Changes in the serum protein electrophoretic pattern in lambs during the first month of life

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Oskar Nagy

2014-01-01

Full Text Available Studies of the changes in serum protein pattern in the neonatal period in animals are still limited. Therefore, the objective of this study was to evaluate the changes in the concentrations of serum protein fractions in 7 clinically healthy merino lambs (4 males, 3 females during their first month of life. The first blood sampling was performed before the colostrum intake and then at 1, 2, 7, 14 and 30 days of age. Blood serum was analysed for total serum protein concentrations and for the relative and absolute values of serum protein fractions - albumin, alpha1- (α1, alpha2- (α2, beta- (β, and gamma- (γ globulins. The results showed a significant effect of age on the serum total protein concentrations and for all the protein fractions. The concentrations of total proteins and γ-globulins increased significantly 1 day after the colostrum intake (P P 1-globulins significantly decreased during the first month of life (P 2- and β-globulins increased significantly from birth till the end of the monitored period (P < 0.001. Our results suggest that the serum protein electrophoretic pattern in growing lambs is significantly influenced by the age of the evaluated animal, and this should be taken into consideration when interpreting the serum protein profile. Our findings extend existing knowledge about significant changes in the protein profile associated with the physiological adaptation process in the neonatal period in young animals.

Studies of serial serum electrophoretic pattern for prognosis in various cancer patients during irradiation

International Nuclear Information System (INIS)

Ra, Woo Youn; Woo, Won Hyung

1971-01-01

During the period from June. 1969 to Dec. 1970, the serum protein electrophoretic patterns of 44 cases of various cancer patients have been studied to determine the alterations in serum protein fractions in patients who were responding to irradiation or those failing. The serum electrophoretic pattern could be observed as an indicator of prognosis or radiosensitivity. A blood sample was obtained prior to any treatment and the follow up sampling was performed 2 times during radiation therapy. Serum total protein was determined by the method of Wolfson and serum electrophoresis was carried out by using Spinoco Model R B electrophoresis system. The results were following: Seven cases out of cases of cervical cancer responding favorably to radiotherapy showed decreased in Alpha-2 globulin fraction were increased. A case whose third time serum electrophoretic pattern showed multiple myeloma type died 5 months after radiotherapy with bone metastasis. Four cases out of 9 cases of favorably responded breast cancer patients showed decreased in Alpha-2 globulin foraction compared with 2 cases of unfavorable response showed increased in Alpha-2 globulin fraction

Studies of serial serum electrophoretic pattern for prognosis in various cancer patients during irradiation

Energy Technology Data Exchange (ETDEWEB)

Ra, Woo Youn; Woo, Won Hyung [Kyungpook National University School of Medicine, Taegu (Korea, Republic of)

1971-10-15

During the period from June. 1969 to Dec. 1970, the serum protein electrophoretic patterns of 44 cases of various cancer patients have been studied to determine the alterations in serum protein fractions in patients who were responding to irradiation or those failing. The serum electrophoretic pattern could be observed as an indicator of prognosis or radiosensitivity. A blood sample was obtained prior to any treatment and the follow up sampling was performed 2 times during radiation therapy. Serum total protein was determined by the method of Wolfson and serum electrophoresis was carried out by using Spinoco Model R B electrophoresis system. The results were following: Seven cases out of cases of cervical cancer responding favorably to radiotherapy showed decreased in Alpha-2 globulin fraction were increased. A case whose third time serum electrophoretic pattern showed multiple myeloma type died 5 months after radiotherapy with bone metastasis. Four cases out of 9 cases of favorably responded breast cancer patients showed decreased in Alpha-2 globulin foraction compared with 2 cases of unfavorable response showed increased in Alpha-2 globulin fraction.

Electrophoretic properties of BSA-coated quantum dots.

Science.gov (United States)

Bücking, Wendelin; Massadeh, Salam; Merkulov, Alexei; Xu, Shu; Nann, Thomas

2010-02-01

Low toxic InP/ZnS quantum dots (QDs), ZnS:Mn(2+)/ZnS nanocrystals and CdSe/ZnS nanoparticles were rendered water-dispersible by different ligand-exchange methods. Eventually, they were coated with bovine serum albumin (BSA) as a model protein. All particles were characterised by isotachophoresis (ITP), laser Doppler velocimetry (LDV) and agarose gel electrophoresis. It was found that the electrophoretic mobility and colloidal stability of ZnS:Mn(2+)/ZnS and CdSe/ZnS nanoparticles, which bore short-chain surface ligands, was primarily governed by charges on the nanoparticles, whereas InP/ZnS nanocrystals were not charged per se. BSA-coated nanoparticles showed lower electrophoretic mobility, which was attributed to their larger size and smaller overall charge. However, these particles were colloidally stable. This stability was probably caused by steric stabilisation of the BSA coating.

Capillary electrophoretic enantioseparation of selegiline, methamphetamine and ephedrine using a neutral β-cyclodextrin epichlorhydrin polymer

NARCIS (Netherlands)

Sevcik, J.; Stransky, Z.; Ingelse, B.A.; Lemr, K.

1996-01-01

This paper describes the development of a capillary zone electrophoretic method for chiral separation of three basic compounds of the selegiline synthetic pathway: ephedrine, methamphetamine and selegiline. The method developed allows one to separate the studied compounds in one run using a neutral

Perfil hematológico e avaliação eletroforética das proteínas séricas de cães com cinomose Hematological profile and electrophoretic evaluation of serum proteins of dogs with canine distemper

OpenAIRE

I.N.G. Silva; M.I.F. Guedes; M.F.G. Rocha; C.M.O. Medeiros; L.C. Oliveira; O.C. Moreira; M.F.S. Teixeira

2005-01-01

The hematological and serum proteins electrophoretic profiles of 13 dogs with distemper (Lentz inclusion body in leukocytes) were studied. The most frequent hematological findings were: normocitic normocromic anemia (61%), leukopenia (46%), left shount (54%), trombocytopenia (69%) and lymphopenia (85%). Electrophoretic analysis of serum proteins showed hypoproteinemia (54%), with reduced albumin and increased alfa-2 globulin. These findings can be used to support the clinical diagnosis of can...

An electrophoretical and immunological study of Pycnogonida, with phylogenetic considerations

NARCIS (Netherlands)

Munilla, Tomás; Haro, de Andrés

1981-01-01

An electrophoretical and immunological study is made of nine species of pycnogonids, representing seven families, from the Catalan coast. An electrophoretogram of each species is given and the antigenic properties of its protein bands are determined. Taking as comparative basis the serological

On-capillary sample cleanup method for the electrophoretic determination of carbohydrates in juice samples.

Science.gov (United States)

Morales-Cid, Gabriel; Simonet, Bartolomé M; Cárdenas, Soledad; Valcárcel, Miguel

2007-05-01

On many occasions, sample treatment is a critical step in electrophoretic analysis. As an alternative to batch procedures, in this work, a new strategy is presented with a view to develop an on-capillary sample cleanup method. This strategy is based on the partial filling of the capillary with carboxylated single-walled carbon nanotube (c-SWNT). The nanoparticles retain interferences from the matrix allowing the determination and quantification of carbohydrates (viz glucose, maltose and fructose). The precision of the method for the analysis of real samples ranged from 5.3 to 6.4%. The proposed method was compared with a method based on a batch filtration of the juice sample through diatomaceous earth and further electrophoretic determination. This method was also validated in this work. The RSD for this other method ranged from 5.1 to 6%. The results obtained by both methods were statistically comparable demonstrating the accuracy of the proposed methods and their effectiveness. Electrophoretic separation of carbohydrates was achieved using 200 mM borate solution as a buffer at pH 9.5 and applying 15 kV. During separation, the capillary temperature was kept constant at 40 degrees C. For the on-capillary cleanup method, a solution containing 50 mg/L of c-SWNTs prepared in 300 mM borate solution at pH 9.5 was introduced for 60 s into the capillary just before sample introduction. For the electrophoretic analysis of samples cleaned in batch with diatomaceous earth, it is also recommended to introduce into the capillary, just before the sample, a 300 mM borate solution as it enhances the sensitivity and electrophoretic resolution.

Ultrastructure and electrophoretic protein pattern of a nuclear fraction enriched in interchromatin granule conglomerations

Energy Technology Data Exchange (ETDEWEB)

Krzyzowska-Gruca, S.; Zborek, A.; Gruca, S.

1986-01-01

Rats were injected with a cytostatic 1-nitro-9/3'-dimethylpropyloamine/acridine.2HCl to induce aggregation of interchromatin granules (IG). The conglomerations of IG were well preserved in isolated liver nuclei and in nuclear structures deprived of chromatin. This feature enabled obtaining a nuclear fraction enriched in IG. The method consisted in extraction of isolated nuclei with a non-ionic detergent and digestion with DNase I in a high ionic strength. Each step of isolation was ultrastructurally monitored using both the routine electron microscopy as well as a preferential staining of IG with bismuth. Presence of spots of tightly packed granules within IG conglomerations in the final fraction like in the nuclei in situ was a good ultrastructural marker of IG. The resulting fraction consisted predominantly of IG conglomerations. Their preferential staining with bismuth was well preserved. Minute amounts of fibrillar material originating from nuclear matrix and residual nuclei could be observed. Protein composition of the fraction enriched in IG was studied by SDS-polyacrylamide gel electrophoresis. After electrotransfer, nitrocellulose filters were fixed with glutaraldehyde and stained with bismuth method in order to identify IG proteins. The results of ultrastructural and cytochemical studies in comparison to electrophoretic protein pattern are discussed.

Perfil hematológico e avaliação eletroforética das proteínas séricas de cães com cinomose Hematological profile and electrophoretic evaluation of serum proteins of dogs with canine distemper

Directory of Open Access Journals (Sweden)

I.N.G. Silva

2005-02-01

Full Text Available The hematological and serum proteins electrophoretic profiles of 13 dogs with distemper (Lentz inclusion body in leukocytes were studied. The most frequent hematological findings were: normocitic normocromic anemia (61%, leukopenia (46%, left shount (54%, trombocytopenia (69% and lymphopenia (85%. Electrophoretic analysis of serum proteins showed hypoproteinemia (54%, with reduced albumin and increased alfa-2 globulin. These findings can be used to support the clinical diagnosis of canine distemper.

Radiation-induced Changes in the Electrophoretic Profile of Serum Albumin

Directory of Open Access Journals (Sweden)

Celso Vieira Lima

2018-01-01

Full Text Available ABSTRACT Albumin protein profiles were investigated in electrophoresis system in relation to the whole body exposition to the radiation. Two groups of rats Wistar were set up as the control (CG and the irradiated one (IG. The IG was exposed to Co-60 at a dose of 5 Gy. After a 72-hour exposition, 300 μL of blood was collected in the inferior vena cava, renal, jugular, hepatic, and pulmonary veins and the serum separated. The albumin protein was identified by vertical electrophoresis in acrylamide Commassi blue or silver stained. The calibration procedure was applied to albumin samples with well-known concentrations. The mathematical correlation was developed involving electrophoretic parameters of band intensities and sizes from gel representation, providing values of protein concentrations in comparison with standard bands with known concentrations. There were significant differences in the physiological concentrations in the jugular and pulmonary sites in relation to renal and cava regional sites. Significant differences induced by radiation in serum albumin concentration were also found in hepatic and jugular sites. Alteration of albumin concentration was found as a nearly effect from whole body irradiation. This phenomenon points out to alterations in cell metabolism in the liver justified by a possible indication of proteomics damage from radiation.

Seed protein variations of Salicornia L. and allied taxa in Turkey.

Science.gov (United States)

Yaprak, A E; Yurdakulol, E

2007-06-01

Electrophoretic seed protein patterns of a number of accessions of Salicornia europaea L. sl., S. prostrata Palas, S. fragilis P.W. Ball and Tutin, Sarcocornia fruticosa (L.) A. J. Scott, Sarcocornia perennis (Miller.) A. J. Scott, Arthrocnemum glaucum (Del.) Ung.-Sternb., Microcnemum coralloides (Loscos and Pardo) subsp. anatolicum Wagenitz and Halocnemum strobilaceum (Pall.) Bieb. were electrophoretically analysed on SDS-PAGE. In total 48 different bands were identified. The obtained data have been treated numerically using the cluster analysis method of unweighted pair group (UPGMA). Finally it was determined that all species separated according to seed protein profiles. And the cladogram obtained studied taxa have been given.

Chromatographic and electrophoretic approaches in ink analysis.

Science.gov (United States)

Zlotnick, J A; Smith, F P

1999-10-15

Inks are manufactured from a wide variety of substances that exhibit very different chemical behaviors. Inks designed for use in different writing instruments or printing methods have quite dissimilar components. Since the 1950s chromatographic and electrophoretic methods have played important roles in the analysis of inks, where compositional information may have bearing on the investigation of counterfeiting, fraud, forgery, and other crimes. Techniques such as paper chromatography and electrophoresis, thin-layer chromatography, high-performance liquid chromatography, gas chromatography, gel electrophoresis, and the relatively new technique of capillary electrophoresis have all been explored as possible avenues for the separation of components of inks. This paper reviews the components of different types of inks and applications of the above separation methods are reviewed.

Separation of short-lived fission products

International Nuclear Information System (INIS)

Tamai, Tadaharu; Ohyoshi, Emiko; Ohyoshi, Akira; Kiso, Yoshiyuki; Shinagawa, Mutsuaki.

1976-01-01

A rbief review is presented on the various methods of separation available for both gaseous and liquid states, for the separation of short-lived fission products formed by binary fission of neutron irradiated uranium. The means available for gaseous state are the hot atom reaction, the hydride method and on-line mass separation. For liquid state, use can be made of precipitation, ionic or atomic exchange, solvent extraction and paper electrophoresis. Particular reference is made to electrophoretic separation of ions produced by fission in aqueous solution of uranium. The principle of electrophoretic separation and the procedures for separating the element of interest from the other fission products are outlined, with reference made to the results obtained with the method by the present authors. The elements in question are alkalines, alkaline earths, rare earths, halogens, selenium and

Selective Photoinitiated Electrophoretic Separator, Phase I

Data.gov (United States)

National Aeronautics and Space Administration — To address NASA Johnson Space Center needs for gas separation and collection technology for lunar in-situ resource utilization, Physical Optics Corporation (POC)...

Evaluation of photo destruction of chromophores of heme and globin components in UV-irradiated human carboxyhemoglobin and its electrophoretic fractions

International Nuclear Information System (INIS)

Putintseva, O.V.; Artykhov, V.G.; Kalaeva, E.A.

2000-01-01

The contribution of hem and globin components of electrophoretic fractions of UV-irradiated human carboxyhemoglobin to photo destruction of the protein was studied. The changes observed are the result of summation of some processes unequal in intensity and direction that take place in microgeterogenous media of photo modified protein. Photo sensitivity of hemoproteid in electrophoretic fraction depends on apoprotein condition, whereas the hem photo resistance cannot be the evidence of the photo stability of the whole molecule [ru

INFLUENCE OF BORATE BUFFERS ON THE ELECTROPHORETIC BEHAVIOR OF HUMIC SUBSTANCES IN CAPILLARY ZONE ELECTROPHORESIS

Science.gov (United States)

The influence of tetrahydroxyborate ions on the electrophoretic mobility of humic acids was evaluated by capillary electrophoresis (CE). Depending on the molarity of borate ions in the separation buffer, the humic acids exhibit electropherograms with sharp peaks consistently exte...

Determination of the Median Lethal Dose and Electrophoretic Pattern of Hottentotta saulcyi (Scorpiones, Buthidae Scorpion Venom

Directory of Open Access Journals (Sweden)

ErsenAydın Yağmur

2015-10-01

Full Text Available Background: In this study, we investigated the lethal potency, electrophoretic protein pattern and in vivo effects of Hottentotta saulcyi scorpion venom in mice.Methods: Scorpions were collected at night, by using a UV lamp from Mardin Province, Turkey. Venom was obtained from mature H. saulcyi scorpions by electrical stimulation of the telson. The lethality of the venom was determined by i.v. injections using Swiss mice. In vivo effects of the venom were assessed by using the intraperitoneal route (ip injections into mice (20±1g and monitored for 24 h. The protein profiles of the scorpion venom were analyzed by NuPAGE® Novex® 4–12 % gradient Bis-Tris gel followed by Coomassie blue staining.Results: The lethal assay of the venom was 0.73 mg/kg in mice. We determined the electrophoretic protein pattern of this scorpion venom to be 4, 6, 9, 31, 35, 40, 46 and 69 kDa by SDS-PAGE. Analysis of electrophoresis indicated that H. saulcyi scorpion intoxicated mice exhibited autonomic nervous system symptoms (tachypnea, restlessness, hyperexcitability, convulsions, salivation, lacrimation, weakness.Conclusions: Hottentotta saulcyi scorpion venom includes short-chain neurotoxins and long-chain neurotoxins according to the electrophoretic protein patterns. The stings of H. saulcyi scorpion must be considered of risk for humans in the southeastern region, Turkey.

Structural analysis of protein complexes with sodium alkyl sulfates by small-angle scattering and polyacrylamide gel electrophoresis.

Science.gov (United States)

Ospinal-Jiménez, Mónica; Pozzo, Danilo C

2011-02-01

Small-angle X-ray (SAXS) and neutron (SANS) scattering is used to probe the structure of protein-surfactant complexes in solution and to correlate this information with their performance in gel electrophoresis. Proteins with sizes between 6.5 to 116 kDa are denatured with sodium alkyl sulfates (SC(x)S) of variable tail lengths. Several combinations of proteins and surfactants are analyzed to measure micelle radii, the distance between micelles, the extension of the complex, the radius of gyration, and the electrophoretic mobility. The structural characterization shows that most protein-surfactant complexes can be accurately described as pearl-necklace structures with spherical micelles. However, protein complexes with short surfactants (SC(8)S) bind with micelles that deviate significantly from spherical shape. Sodium decyl (SC(10)S) and dodecyl (SC(12)S, more commonly abbreviated as SDS) sulfates result in the best protein separations in standard gel electrophoresis. Particularly, SC(10)S shows higher resolutions for complexes of low molecular weight. The systematic characterization of alkyl sulfate surfactants demonstrates that changes in the chain architecture can significantly affect electrophoretic migration so that protein-surfactant structures could be optimized for high resolution protein separations.

Protein Separation by Capillary Gel Electrophoresis: A Review

Science.gov (United States)

Zhu, Zaifang; Lu, Joann J.; Liu, Shaorong

2011-01-01

Capillary gel electrophoresis (CGE) has been used for protein separation for more than two decades. Due to the technology advancement, current CGE methods are becoming more and more robust and reliable for protein analysis, and some of the methods have been routinely used for the analysis of protein-based pharmaceuticals and quality controls. In light of this progress, we survey 147 papers related to CGE separations of proteins and present an overview of this technology. We first introduce briefly the early development of CGE. We then review the methodology, in which we specifically describe the matrices, coatings, and detection strategies used in CGE. CGE using microfabricated channels and incorporation of CGE with two-dimensional protein separations are also discussed in this section. We finally present a few representative applications of CGE for separating proteins in real-world samples. PMID:22122927

Protein separations using enhanced-fluidity liquid chromatography.

Science.gov (United States)

Bennett, Raffeal; Olesik, Susan V

2017-11-10

Enhanced-fluidity liquid chromatography (EFLC) methods using methanol/H 2 O/CO 2 and hydrophilic interaction liquid chromatography (HILIC) were explored for the separation of proteins and peptides. EFLC is a separation mode that uses a mobile phase made of conventional solvents combined with liquid carbon dioxide (CO 2 ) in subcritical conditions. The addition of liquid CO 2 enhances diffusivity and decreases viscosity while maintaining mixture polarity, which typically results in reduced time of analysis. TFA additive and elevated temperature were leveraged as key factors in the separation of a 13-analyte intact protein mixture in under 5min. Under these conditions EFLC showed modest improvement in terms of peak asymmetry and analysis time over the competing ACN/H 2 O separation. Protein analytes detected by electrospray ionization - quadrupole time of flight, were shown to be unaffected by the addition of CO 2 in the mobile phase. Herein, the feasibility of separating hydrophilic proteins up to 80kDa (with transferrin) is demonstrated for CO 2 -containing mobile phases. Copyright © 2017 Elsevier B.V. All rights reserved.

Characterization of Dickeya and Pectobacterium species by capillary electrophoretic techniques and MALDI-TOF MS

Czech Academy of Sciences Publication Activity Database

Šalplachta, Jiří; Kubesová, Anna; Horký, J.; Matoušková, H.; Tesařová, Marie; Horká, Marie

2015-01-01

Roč. 407, č. 25 (2015), s. 7625-7635 ISSN 1618-2642 R&D Projects: GA MV VG20112015021 Institutional support: RVO:68081715 Keywords : bacteria * electrophoretic techniques * MALDI Subject RIV: CB - Analytical Chemistry, Separation Impact factor: 3.125, year: 2015 http://hdl.handle.net/11104/0250090

Electrophoretic characterization of the Mammalian nuclear matrix proteome, nuclear envelope, nucleoli and covalently bound ADP-ribose polymers: potential applications to cancer.

Science.gov (United States)

Aranda, Xavier G; Racho, Ronald G; Pacheco-Rodríguez, Gustavo; Alvarez-González, Rafael

2014-01-01

Nucleic acid metabolism is biochemically compartmentalized to the nucleus. Thus, it is necessary to define the proteome of the various macromolecular structures within this organelle. We isolated the nuclear matrix (NM) fraction from rat liver by sequential centrifugation steps at 13,000 rpm, staggered between endogenous nuclease treatment for 2 h at 37°C, followed by high-salt (H.S.; 2.0 M NaCl) and non-ionic detergent extractions (0.1%- or 1.0% Triton X-100) to eliminate the bulk of chromosomal DNA/RNA, histone proteins and the nuclear envelope (NE). Integrity of the NM and NE structures was confirmed by electron microscopy. Next, we analyzed the NM proteome on a 20% polyacrylamide gel using the PhastSystem. We observed the absence of histone proteins and the characteristic presence of the lamins by Coomassie blue staining. By contrast, upon silver staining, following electrophoretic separation with a Tris-Borate-EDTA buffer, we observed the NM-associated nucleic RNA and protein-free ADP-ribose polymers. While polymers are found in much lower concentration than RNA in NM, they were purified by affinity chromatography on boronate resin prior to electrophoresis. We observed the electrophoretic resolution of free ADP-ribose chains (5-25 units) by silver staining. The significance of our observations to cancer studies and carcinogenesis is discussed. Copyright© 2014, International Institute of Anticancer Research (Dr. John G. Delinasios), All rights reserved.

Evidence of the protein content of bovine and human dental pulps by the action of endodontic irrigation solutions through electrophoretic patterns

Directory of Open Access Journals (Sweden)

María E López

2013-01-01

Full Text Available Background: Sodium dodecyl sulfate polyacrylamide gel electrophoresis let to show the protein content of different tissues. Dental pulp contains connective tissue which is removed during the endodontic treatment. Many studies consider bovine rather than human pulp tissue because of its size. Aim: To evidence the protein content of bovine and human dental pulps and the action of endodontic irrigation solutions through electrophoretic patterns. Materials and Methods: Extracts of human and bovine dental pulps were prepared. Sodium hypochlorite, calcium hydroxide, chlorhexidine and ethylenediamine tetraacetic acid were used as irrigating solutions. Results: Bovine and human pulps have a small difference in two bands of proteins present between 74 kDa and 80 kDa. The denaturizing capacity of sodium hypochlorite and the washing action of calcium hydroxide and chlorhexidine were evidenced. Ethylenediamine tetraacetic acid solution was shown to contain proteins continuously during the endodontic root canal washing. Conclusions: Differences in pulp tissues and the action of irrigating solutions on their protein content would help on the understanding of the biological process of the endodontic treatment.

X-radiation effect on soluble proteins of gastric mucosa

International Nuclear Information System (INIS)

Sukhomlinov, B.F.; Chajka, Ya.P.; Fedorovich, A.N.

1979-01-01

Using the method of electrophoresis in agar gel soluble proteins of gastric mucosa of rats were separated into 11 fractions. Proteins posessing a proteolytic (pH 1.8) and lipase (pH 7.4) activity were localized within the second and third prealbumin fractions. Soluble proteins of gastric mucosa contain glyco- and lipoproteid complexes. Exposure of rats to 1000 R of X-rays induces quantitative redistribution within the electrophoretic spectrum of soluble proteins and a considerable disturbance of the proteolytic activity of total soluble proteins throughout the entire period of observation (from 10 min to 72h)

Investigation of the free flow electrophoretic process. Volume 2: Technical analysis

Science.gov (United States)

Weiss, R. A.; Lanham, J. W.; Richman, D. W.; Walker, C. D.

1979-01-01

The effect of gravity on the free flow electrophoretic process was investigated. The demonstrated effects were then compared with predictions made by mathematical models. Results show that the carrier buffer flow was affected by gravity induced thermal convection and that the movement of the separating particle streams was affected by gravity induced buoyant forces. It was determined that if gravity induced buoyant forces were included in the mathematical models, then effective predictions of electrophoresis chamber separation performance were possible. The results of tests performed using various methods of electrophoresis using supportive media show that the mobility and the ability to separate were essentially independent of concentration, providing promise of being able to perform electrophoresis with higher inlet concentrations in space.

Electrophoretic separation of alginic sodium diester and sodium hexametaphosphate in chondroitin sulfate that interfere with the cetylpyridinium chloride titration assay.

Science.gov (United States)

Weiguo, Zhang; Giancaspro, Gabriel; Adams, Kristie M; Neal-Kababick, James; Hildreth, Jana; Li, Aishan; Roman, Mark C; Betz, Joseph M

2014-01-01

The most commonly used chondroitin sulfate (CS) assay method is cetylpyridinium chloride (CPC) titration. Cellulose acetate membrane electrophoresis (CAME) is the technique used for detection of impurities in the U.S. Pharmacopeia's CS monograph. Because CPC titration is a relatively nonspecific quantitative technique, the apparent amount of CS as determined by CPC titration alone may not reflect the true amount of CS due to possible interference with the CPC assay by impurities that contain CPC titratable functional groups. When CAME is used in conjunction with CPC titration, certain non-CS and adulterants can be visualized and estimated, and a true value for CS can be assigned once the presence of these non-CS impurities has been ruled out. This study examines conjunct application of CPC and CAME in ascertaining CS assay and purity in the presence of certain adulterants. These include propylene glycol alginate sulfate sodium, known in commerce as alginic sodium diester (ASD), and Zero One (Z1), a water-soluble agent newly reported in the CS marketplace and subsequently identified as sodium hexametaphosphate. ASD, Z1, and CS are similar in physical appearance and solubility in water and ethanol. They are also titratable anions and form ionic pairs with CPC, therefore interfering with the CPC titration assay for CS CAME separates these adulterants from each other and from CS by differences in their electrophoretic mobility. CAME is able to detect these impurities in CS at levels as low as 0.66% by weight. Although it is recommended that a method for detecting impurities (e.g., CAME) be used in cormbination with relatively nonspecific assay methods such as CPC titration, this is seldom done in practice. Assay results for CS derived fromn CPC titration may, therefore, be misleading, leaving the CS supply chain vulnerable to adulteration. In this study, the authors investigated ASD and Z1 adulteration of CS and developed an electrophoretic separation of these

Electrophoretic characterization of protein interactions suggesting limited feasibility of accelerated shelf-life testing of ultra-high temperature milk.

Science.gov (United States)

Grewal, Manpreet Kaur; Chandrapala, Jayani; Donkor, Osaana; Apostolopoulos, Vasso; Vasiljevic, Todor

2017-01-01

Accelerated shelf-life testing is applied to a variety of products to estimate keeping quality over a short period of time. The industry has not been successful in applying this approach to ultra-high temperature (UHT) milk because of chemical and physical changes in the milk proteins that take place during processing and storage. We investigated these protein changes, applying accelerated shelf-life principles to UHT milk samples with different fat levels and using native- and sodium dodecyl sulfate-PAGE. Samples of UHT skim and whole milk were stored at 20, 30, 40, and 50°C for 28d. Irrespective of fat content, UHT treatment had a similar effect on the electrophoretic patterns of milk proteins. At the start of testing, proteins were bonded mainly through disulfide and noncovalent interactions. However, storage at and above 30°C enhanced protein aggregation via covalent interactions. The extent of aggregation appeared to be influenced by fat content; whole milk contained more fat than skim milk, implying aggregation via melted or oxidized fat, or both. Based on reduction in loss in absolute quantity of individual proteins, covalent crosslinking in whole milk was facilitated mainly by products of lipid oxidation and increased access to caseins for crosslinking reactions. Maillard and dehydroalanine products were the main contributors involved in protein changes in skim milk. Protein crosslinking appeared to follow a different pathway at higher temperatures (≥40°C) than at lower temperatures, making it very difficult to extrapolate these changes to protein interactions at lower temperatures. Copyright © 2017 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

Electrophoretic variants of blood proteins in japanese, 5

International Nuclear Information System (INIS)

Fujita, Mikio; Satoh, Chiyoko; Asakawa, Jun-ichi; Nagahata, Yuko; Tanaka, Yoshiko; Hazama, Ryuji; Goriki, Kazuaki.

1985-08-01

The plasma ceruloplasmin (CP) of 22,367 children of atomic bomb survivors in Hiroshima and Nagasaki was examined for variants by electrophoresis. The sample was composed of 14,964 unrelated children and 7,403 siblings of the unrelated persons. A total of seven types of electrophoretic variants were detected; four migrating anodally and three cathodally to the normal B band. We have reported two of these variants, CP A sub(NG1) and CP C sub(NG1), previously but the other five, CP A sub(NG2), CP A sub(HR1), CP A sub(HR2), CP C sub(HR1), and CP C sub(HR2), are newly identified. The allelic frequency of CP*CNG1 was 0.00916, so that the variant is considered to be a polymorphic allele. Homozygosity for the CP*CNG1 allele was detected in five individuals. This is the first report of a homozygous phenotype for a CP variant in a Japanese population. Family study of the new five variants all demonstrated patterns of codominant inheritance. (author)

Separation of multiphosphorylated peptide isomers by hydrophilic interaction chromatography on an aminopropyl phase.

Science.gov (United States)

Singer, David; Kuhlmann, Julia; Muschket, Matthias; Hoffmann, Ralf

2010-08-01

The separation of isomeric phosphorylated peptides is challenging and often impossible for multiphosphorylated isomers using chromatographic and capillary electrophoretic methods. In this study we investigated the separation of a set of single-, double-, and triple-phosphorylated peptides (corresponding to the human tau protein) by ion-pair reversed-phase chromatography (IP-RPC) and hydrophilic interaction chromatography (HILIC). In HILIC both hydroxyl and aminopropyl stationary phases were tested with aqueous acetonitrile in order to assess their separation efficiency. The hydroxyl phase separated the phosphopeptides very well from the unphosphorylated analogue, while on the aminopropyl phase even isomeric phosphopeptides attained baseline separation. Thus, up to seven phosphorylated versions of a given tau domain were separated. Furthermore, the low concentration of an acidic ammonium formate buffer allowed an online analysis with electrospray ionization tandem mass spectrometry (ESI-MS/MS) to be conducted, enabling peptide sequencing and identification of phosphorylation sites.

Capillary electrophoretic separation of inorganic and organic arsenic compounds

Energy Technology Data Exchange (ETDEWEB)

Greschonig, H. [Institute of Analytical Chemistry, Karl Franzens University Graz (Austria); Schmid, M.G.; Guebitz, G. [Institute of Pharmaceutical Chemistry, Karl Franzens University Graz (Austria)

1998-09-01

Capillary zone electrophoresis was used to separate arsenite, arsenate, dimethylarsinic and diphenylarsinic acid, methanearsonic acid, phenyl- and p-aminophenyl arsonic acid, phenylarsineoxide and phenarsazinic acid. Anionic and uncharged species were separated in a fused silica capillary with on-column UV detection at 200 nm. A 15 mM phosphate solution adjusted to pH 6.5 containing 10 mM sodium dodecylsulfonate served as background electrolyte. The influence of pH and applied voltage on separation efficiency, as well as the feasibility of identification of arsenic compounds in spiked urine, were investigated. (orig.) With 7 figs., 1 tab., 22 refs.

Interconverting conformations of variants of the human amyloidogenic protein beta2-microglobulin quantitatively characterized by dynamic capillary electrophoresis and computer simulation

DEFF Research Database (Denmark)

Heegaard, Niels H H; Jørgensen, Thomas J D; Cheng, Lei

2006-01-01

Capillary electrophoretic separation profiles of cleaved variants of beta2-microglobulin (beta2m) reflect the conformational equilibria existing in solutions of these proteins. The characterization of these equilibria is of interest since beta2m is responsible for amyloid formation in dialysis-re...

Sex pheromone receptor proteins. Visualization using a radiolabeled photoaffinity analog

International Nuclear Information System (INIS)

Vogt, R.G.; Prestwich, G.D.; Riddiford, L.M.

1988-01-01

A tritium-labeled photoaffinity analog of a moth pheromone was used to covalently modify pheromone-selective binding proteins in the antennal sensillum lymph and sensory dendritic membranes of the male silk moth, Antheraea polyphemus. This analog, (E,Z)-6,11-[ 3 H]hexadecadienyl diazoacetate, allowed visualization of a 15-kilodalton soluble protein and a 69-kilodalton membrane protein in fluorescence autoradiograms of electrophoretically separated antennal proteins. Covalent modification of these proteins was specifically reduced when incubation and UV irradiation were conducted in the presence of excess unlabeled pheromone, (E,Z)-6,11-hexadecadienyl acetate. These experiments constitute the first direct evidence for a membrane protein of a chemosensory neuron interacting in a specific fashion with a biologically relevant odorant

Separation of ions in acidic solution by capillary electrophoresis

Energy Technology Data Exchange (ETDEWEB)

Thornton, Michelle [Iowa State Univ., Ames, IA (United States)

1997-10-08

Capillary electrophoresis (CE) is an effective method for separating ionic species according to differences in their electrophoretic mobilities. CE separations of amino acids by direct detection are difficult due to their similar electrophoretic mobilities and low absorbances. However, native amino acids can be separated by CE as cations at a low pH by adding an alkanesulfonic acid to the electrolyte carrier which imparts selectivity to the system. Derivatization is unnecessary when direct UV detection is used at 185 nm. Simultaneous speciation of metal cations such as vanadium (IV) and vanadium (V) can easily be performed without complexation prior to analysis. An indirect UV detection scheme for acidic conditions was also developed using guanidine as the background carrier electrolyte (BCE) for the indirect detection of metal cations. Three chapters have been removed for separate processing. This report contains introductory material, references, and general conclusions. 80 refs.

Study of the transport of mercurial compounds by seric proteins

International Nuclear Information System (INIS)

Jullien-Saint Guily, Nicole

1970-01-01

A bond between the seric proteins and various mercurial compounds labeled with the radioisotopes 203 Hg and 197 Hg was demonstrated by means of research methods specific to radioactivity combined with protein separation techniques. In the course of this study it was shown how strongly the composition of the buffer during electrophoretic migration influences the transport of certain organo-mercurial compounds by the seric proteins. By means of a thioloprive: N - ethyl - maleimide, labeled with 14 C, it was proved that the bonding sites between the proteins and the mercurial compounds were the thiol groups of the proteins but that other bonding sites, in particular the amino groups, could also be involved. (author) [fr

Interpretive Reporting of Protein Electrophoresis Data by Microcomputer

Science.gov (United States)

Talamo, Thomas S.; Losos, Frank J.; Kessler, G. Frederick

1982-01-01

A microcomputer based system for interpretive reporting of protein electrophoretic data has been developed. Data for serum, urine and cerebrospinal fluid protein electrophoreses as well as immunoelectrophoresis can be entered. Patient demographic information is entered through the keyboard followed by manual entry of total and fractionated protein levels obtained after densitometer scanning of the electrophoretic strip. The patterns are then coded, interpreted, and final reports generated. In most cases interpretation time is less than one second. Misinterpretation by computer is uncommon and can be corrected by edit functions within the system. These discrepancies between computer and pathologist interpretation are automatically stored in a data file for later review and possible program modification. Any or all previous tests on a patient may be reviewed with graphic display of the electrophoretic pattern. The system has been in use for several months and is presently well accepted by both laboratory and clinical staff. It also allows rapid storage, retrieval and analysis of protein electrophoretic datab.

Lupine protein enrichment by milling and electrostatic separation

NARCIS (Netherlands)

Wang, Jue; Zhao, Jun; Wit, De Martin; Boom, Remko M.; Schutyser, Maarten A.I.

2016-01-01

Lupine seeds are excellent source of plant protein. We here report on dry fractionation by combining milling and electrostatic separation providing an alternative to wet extraction of protein from lupine seeds. Relatively coarse milling was preferred as this provides sufficient detached protein

Characterization of gel-separated glycoproteins using two-step proteolytic digestion combined with sequential microcolumns and mass spectrometry

DEFF Research Database (Denmark)

Larsen, Martin Røssel; Højrup, Peter; Roepstorff, Peter

2005-01-01

present a new technique that allows characterization of low amounts of glycoproteins separated by gel electrophoresis. The method takes advantage of sequential specific and nonspecific enzymatic treatment followed by selective purification and characterization of the glycopeptides using graphite powder......Protein glycosylation can be vital for changing the function or physiochemical properties of a protein. Abnormal glycosylation can lead to protein malfunction, resulting in severe diseases. Therefore, it is important to develop techniques for characterization of such modifications in proteins...... at a sensitivity level comparable with state-of-the-art proteomics. Whereas techniques exist for characterization of high abundance glycoproteins, no single method is presently capable of providing information on both site occupancy and glycan structure on a single band excised from an electrophoretic gel. We...

Capillary electrophoresis separation of neutral organic compounds, pharmaceutical drugs, proteins and peptides, enantiomers, and anions

Energy Technology Data Exchange (ETDEWEB)

Ding, Wei -Liang [Iowa State Univ., Ames, IA (United States)

1999-02-12

Addition of a novel anionic surfactant, namely lauryl polyoxyethylene sulfate, to an aqueous-acetonitrile electrolyte makes it possible to separate nonionic organic compounds by capillary electrophoresis. Separation is based on differences in the association between analytes and the surfactant. Highly hydrophobic compounds such as polyaromatic hydrocarbons are well separated by this new surfactant. Migration times of analytes can be readily changed over an unusually large range by varying the additive concentration and the proportion of acetonitrile in the electrolyte. Several examples are given, including the separation of four methylbenz[a]anthracene isomers and the separation of normal and deuterated acetophenone. The effect of adding this new surfactant to the acidic electrolyte was also investigated. Incorporation of cetyltrimethylammonium bromide in the electrolyte is shown to dynamically coat the capillary and reverse electroosmotic flow. Chiral recognition mechanism is studied using novel synthetic surfactants as chiral selectors, which are made from amino acids reacting with alkyl chloroformates. A satisfactory separation of both inorganic and organic anions is obtained using electrolyte solutions as high as 5 M sodium chloride using direct photometric detection. The effect of various salts on electrophoretic and electroosmotic mobility is further discussed. Several examples are given under high-salt conditions.

Electrophoretic deposition of sol-gel-derived ceramic coatings

International Nuclear Information System (INIS)

Zhang, Y.; Crooks, R.M.

1992-01-01

In this paper the physical, optical, and chemical characteristics of electrophoretically and dip-coated sol-gel ceramic films are compared. The results indicate that electrophoresis may allow a higher level of control over the chemistry and structure of ceramic coatings than dip-coating techniques. For example, controlled-thickness sol-gel coatings can be prepared by adjusting the deposition time or voltage. Additionally, electrophoretic coatings can be prepared in a four-component alumino-borosilicate sol display interesting optical characteristics. For example, the ellipsometrically-measured refractive indices of electrophoretic coatings are higher than the refractive indices of dip-coated films cast from identical sols, and they are also higher than any of the individual sol components. This result suggests that there are physical and/or chemical differences between films prepared by dip-coating and electrophoresis

Monoaminylation of Fibrinogen and Glia-Derived Proteins: Indication for Similar Mechanisms in Posttranslational Protein Modification in Blood and Brain.

Science.gov (United States)

Hummerich, René; Costina, Victor; Findeisen, Peter; Schloss, Patrick

2015-07-15

Distinct proteins have been demonstrated to be posttranslationally modified by covalent transamidation of serotonin (5-hydropxytryptamin) to glutamine residues of the target proteins. This process is mediated by transglutaminase (TGase) and has been termed "serotonylation." It has also been shown that other biogenic amines, including the neurotransmitters dopamine and norepinephrine, can substitute for serotonin, implying a more general mechanism of "monoaminylation" for this kind of protein modification. Here we transamidated the autofluorescent monoamine monodansylcadaverine (MDC) to purified plasma fibrinogen and to proteins from a primary glia cell culture. Electrophoretic separation of MDC-conjugated proteins followed by mass spectrometry identified three fibrinogen subunits (Aα, Bβ, γ), a homomeric Aα2 dimer, and adducts of >250 kDa molecular weight, as well as several glial proteins. TGase-mediated MDC incorporation was strongly reduced by serotonin, underlining the general mechanism of monoaminylation.

Simplification and improvement of protein detection in two-dimensional electrophoresis gels with SERVA HPE™ lightning red.

Science.gov (United States)

Griebel, Anja; Obermaier, Christian; Westermeier, Reiner; Moche, Martin; Büttner, Knut

2013-07-01

A new fluorescent amino-reactive dye has been tested for both labelling proteins prior to electrophoretic separations and between the two steps of two-dimensional electrophoresis. A series of experiments showed, that the labelling of lysines with this dye is compatible with all standard additives used for sample preparation, including reducing substances and carrier ampholytes. Using this dye for pre-labelling considerably simplifies the electrophoresis and detection workflow and provides highly sensitive and quantitative visualisation of proteins.

Frequency of electrophoretic changes consistent with feline infectious peritonitis in two different time periods (2004-2009 vs 2013-2014).

Science.gov (United States)

Stranieri, Angelica; Giordano, Alessia; Bo, Stefano; Braghiroli, Chiara; Paltrinieri, Saverio

2017-08-01

Objectives The aim of this study was to evaluate whether the frequency of electrophoretic changes in serum of cats with feline infectious peritonitis (FIP) changed in recent years vs past years. Methods Agarose gel electrophoresis (AGE) and capillary zone electrophoresis (CZE) from cats with FIP and healthy cats recorded in the periods 2004-2009 and 2013-2014 were retrospectively analysed. Relative and absolute values of each electrophoretic fraction were recorded and the number of cats showing single or combined electrophoretic changes consistent with FIP (hypoalbuminaemia, inverted albumin to globulin [A:G] ratio, increased total protein, total globulin, alpha [α] 2 -globulin and gamma [γ]-globulin concentration) were counted. Additionally, a visual analysis of electrophoretograms was also performed. Results for the two time periods were statistically compared. Results The details of 91 AGE procedures (41 from cats with FIP and 50 from healthy cats) and 45 CZE procedures (26 from cats with FIP and 19 from healthy cats) were obtained from the database. No significant differences between the two time periods were found both in FIP and in healthy cats analysed with CZE and in healthy cats analysed with AGE. Compared with 2004-2009, cats with FIP sampled in 2013-2014 with AGE showed a significantly lower concentration of total protein, γ-globulins and total globulins, and a significantly higher A:G ratio and percentage of albumin and α 2 -globulins. Using both AGE and CZE, in recent years the proportion of cats with high α2-globulins without gammopathy and the proportion of cats with gammopathy alone decreased. With a visual approach, the number of patterns considered as dubious increased in the second period with AGE (non-statistically significant). Conclusions and relevance The frequency of electrophoretic abnormalities in cats with FIP decreased in recent years, independently of the technique employed. Although the mechanism responsible for this change was

Theory and applications of a novel ion exchange chromatographic technology using controlled pH gradients for separating proteins on anionic and cationic stationary phases.

Science.gov (United States)

Tsonev, Latchezar I; Hirsh, Allen G

2008-07-25

pISep is a major new advance in low ionic strength ion exchange chromatography. It enables the formation of externally controlled pH gradients over the very broad pH range from 2 to 12. The gradients can be generated on either cationic or anionic exchangers over arbitrary pH ranges wherein the stationary phases remain totally charged. Associated pISep software makes possible the calculation of either linear, nonlinear or combined, multi-step, multi-slope pH gradients. These highly reproducible pH gradients, while separating proteins and glycoproteins in the order of their electrophoretic pIs, provide superior chromatographic resolution compared to salt. This paper also presents a statistical mechanical model for protein binding to ion exchange stationary phases enhancing the electrostatic interaction theory for the general dependence of retention factor k, on both salt and pH simultaneously. It is shown that the retention factors computed from short time isocratic salt elution data of a model protein can be used to accurately predict its salt elution concentration in varying slope salt elution gradients formed at varying isocratic pH as well as the pH at which it will be eluted from an anionic exchange column by a pISep pH gradient in the absence of salt.

Accelerated SDS depletion from proteins by transmembrane electrophoresis: Impacts of Joule heating.

Science.gov (United States)

Unterlander, Nicole; Doucette, Alan Austin

2018-02-08

SDS plays a key role in proteomics workflows, including protein extraction, solubilization and mass-based separations (e.g. SDS-PAGE, GELFrEE). However, SDS interferes with mass spectrometry and so it must be removed prior to analysis. We recently introduced an electrophoretic platform, termed transmembrane electrophoresis (TME), enabling extensive depletion of SDS from proteins in solution with exceptional protein yields. However, our prior TME runs required 1 h to complete, being limited by Joule heating which causes protein aggregation at higher operating currents. Here, we demonstrate effective strategies to maintain lower TME sample temperatures, permitting accelerated SDS depletion. Among these strategies, the use of a magnetic stir bar to continuously agitate a model protein system (BSA) allows SDS to be depleted below 100 ppm (>98% removal) within 10 min of TME operations, while maintaining exceptional protein recovery (>95%). Moreover, these modifications allow TME to operate without any user intervention, improving throughput and robustness of the approach. Through fits of our time-course SDS depletion curves to an exponential model, we calculate SDS depletion half-lives as low as 1.2 min. This promising electrophoretic platform should provide proteomics researchers with an effective purification strategy to enable MS characterization of SDS-containing proteins. © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

A plasmid containing the human metallothionein II gene can function as an antibody-assisted electrophoretic biosensor for heavy metals.

Science.gov (United States)

Wooten, Dennis C; Starr, Clarise R; Lyon, Wanda J

2016-01-01

Different forms of heavy metals affect biochemical systems in characteristic ways that cannot be detected with typical metal analysis methods like atomic absorption spectrometry. Further, using living systems to analyze interaction of heavy metals with biochemical systems can be laborious and unreliable. To generate a reliable easy-to-use biologically-based biosensor system, the entire human metallothionein-II (MT-II) gene was incorporated into a plasmid (pUC57-MT) easily replicated in Escherichia coli. In this system, a commercial polyclonal antibody raised against human metal-responsive transcription factor-1 protein (MTF-1 protein) could modify the electrophoretic migration patterns (i.e. cause specific decreases in agarose gel electrophoretic mobility) of the plasmid in the presence or absence of heavy metals other than zinc (Zn). In the study here, heavy metals, MTF-1 protein, and polyclonal anti-MTF-1 antibody were used to assess pUC57-MT plasmid antibody-assisted electrophoretic mobility. Anti-MTF-1 antibody bound both MTF-1 protein and pUC57-MT plasmid in a non-competitive fashion such that it could be used to differentiate specific heavy metal binding. The results showed that antibody-inhibited plasmid migration was heavy metal level-dependent. Zinc caused a unique mobility shift pattern opposite to that of other metals tested, i.e. Zn blocked the antibody ability to inhibit plasmid migration, despite a greatly increased affinity for DNA by the antibody when Zn was present. The Zn effect was reversed/modified by adding MTF-1 protein. Additionally, antibody inhibition of plasmid mobility was resistant to heat pre-treatment and trypsinization, indicating absence of residual DNA extraction-resistant bacterial DNA binding proteins. DNA binding by anti-DNA antibodies may be commonly enhanced by xenobiotic heavy metals and elevated levels of Zn, thus making them potentially effective tools for assessment of heavy metal bioavailability in aqueous solutions and

Separation selectivity patterns of fully charged achiral compounds in capillary electrophoresis with a neutral cyclodextrin.

Science.gov (United States)

Soonthorntantikul, Wasura; Srisa-art, Monpichar; Leepipatpiboon, Natchanun; Nhujak, Thumnoon

2013-01-01

Based on the separation selectivity equation, related to the dimensionless parameters for fully charged achiral analytes using a neutral CD, the separation selectivity can be classified into seven patterns. With respect to CZE without CD, the presence of CD in the buffer may improve, or reduce, the separation selectivity with this effect being accompanied by the same or reversed electrophoretic mobility order for charged analytes. This can depend on the separation selectivity of the two analytes in free solution, the binding selectivity, the separation selectivity of analyte-CD complexes and the ratio of electrophoretic mobility of the analytes in free, and complexed forms. Using positional isomers of benzoic acids and phenoxy acids as test analytes and α-CD as a selector, the observed separation selectivity shapes were found to be in excellent agreement with the predicted separation selectivities. © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

UV-induced DNA-binding proteins in human cells

International Nuclear Information System (INIS)

Glazer, P.M.; Greggio, N.A.; Metherall, J.E.; Summers, W.C.

1989-01-01

To investigate the response of human cells to DNA-damaging agents such as UV irradiation, the authors examined nuclear protein extracts of UV-irradiated HeLa cells for the presence of DNA-binding proteins. Electrophoretically separated proteins were transferred to a nitrocellulose filter that was subsequently immersed in a binding solution containing radioactively labeled DNA probes. Several DNA-binding proteins were induced in HeLa cells after UV irradiation. These included proteins that bind predominantly double-stranded DNA and proteins that bind both double-stranded and single-stranded DNA. The binding proteins were induced in a dose-dependent manner by UV light. Following a dose of 12 J/m 2 , the binding proteins in the nuclear extracts increased over time to a peak in the range of 18 hr after irradiation. Experiments with metabolic inhibitors (cycloheximide and actinomycin D) revealed that de novo synthesis of these proteins is not required for induction of the binding activities, suggesting that the induction is mediated by protein modification

Red fluorescent protein responsible for pigmentation in trematode-infected Porites compressa tissues.

Science.gov (United States)

Palmer, Caroline V; Roth, Melissa S; Gates, Ruth D

2009-02-01

Reports of coral disease have increased dramatically over the last decade; however, the biological mechanisms that corals utilize to limit infection and resist disease remain poorly understood. Compromised coral tissues often display non-normal pigmentation that potentially represents an inflammation-like response, although these pigments remain uncharacterized. Using spectral emission analysis and cryo-histological and electrophoretic techniques, we investigated the pink pigmentation associated with trematodiasis, infection with Podocotyloides stenometre larval trematode, in Porites compressa. Spectral emission analysis reveals that macroscopic areas of pink pigmentation fluoresce under blue light excitation (450 nm) and produce a broad emission peak at 590 nm (+/-6) with a 60-nm full width at half maximum. Electrophoretic protein separation of pigmented tissue extract confirms the red fluorescence to be a protein rather than a low-molecular-weight compound. Histological sections demonstrate green fluorescence in healthy coral tissue and red fluorescence in the trematodiasis-compromised tissue. The red fluorescent protein (FP) is limited to the epidermis, is not associated with cells or granules, and appears unstructured. These data collectively suggest that the red FP is produced and localized in tissue infected by larval trematodes and plays a role in the immune response in corals.

Electrophoretic and immunological properties of folate-binding protein isolated from bovine milk

International Nuclear Information System (INIS)

Iwai, Kazuo; Tani, Masako; Fushiki, Tohru

1983-01-01

Changes of the folate-binding protein (FBP) concentration in bovine milk after parturition were investigated. The FBP was highly purified from mature milk by affinity chromatography. The purified FBP showed a single protein band in polyacrylamide gel electrophoresis and was immunologically homogenous in double immunodiffusion. However, in two-dimensional gel electrophoresis, the FBP was separated into several spots in isoelectric focusing in the first dimension, and each spot also showed two molecular weights in SDS-gel electrophoresis in the second dimension. But these FBP molecules were immunologically identical with each other. The neuraminidase treatment obviously diminished the number of isoelectric points of the FBP. Thus, the variety of FBP molecules was at least partially due to the variability of the sialic acid content in the carbohydrate moieties. Moreover, the milk FBP showed species-specificity among mammals immunologically as well as physicochemically. (author)

Correlation of acidic and basic carrier ampholyte and immobilized pH gradient two-dimensional gel electrophoresis patterns based on mass spectrometric protein identification

DEFF Research Database (Denmark)

Nawrocki, A; Larsen, Martin Røssel; Podtelejnikov, A V

1998-01-01

Separation of proteins on either carrier ampholyte-based or immobilized pH gradient-based two-dimensional (2-D) gels gives rise to electrophoretic patterns that are difficult to compare visually. In this paper we have used matrix-assisted laser desorption/ionization mass spectrometry (MALDI......-MS) to determine the identities of 335 protein spots in these two 2-D gel systems, including a substantial number of basic proteins which had never been identified before. Proteins that were identified in both gel systems allowed us to cross-reference the gel patterns. Vector analysis of these cross...

Gc protein-derived macrophage activating factor (GcMAF): isoelectric focusing pattern and tumoricidal activity.

Science.gov (United States)

Mohamad, Saharuddin Bin; Nagasawa, Hideko; Sasaki, Hideyuki; Uto, Yoshihiro; Nakagawa, Yoshinori; Kawashima, Ken; Hori, Hitoshi

2003-01-01

Gc protein is the precursor for Gc protein-derived macrophage activating factor (GcMAF), with three phenotypes: Gc1f, Gc1s and Gc2, based on its electrophoretic mobility. The difference in electrophoretic mobility is because of the difference in its posttranslational sugar moiety composition. We compared the difference between Gc protein and GcMAF electrophoretic mobility using the isoelectric focusing (IEF) method. The tumoricidal activity of GcMAF-treated macrophage was evaluated after coculture with L-929 cell. The tumoricidal mechanism was investigated using TNF bioassay and nitric oxide (NO) release. The difference in Gc protein and GcMAF electrophoretic mobility was detected. The tumoricidal activity of GcMAF-treated macrophage was detected, but no release of TNF and NO was detected. The difference of isoelectric focusing mobility in Gc protein and GcMAF would be useful to develop a GcMAF detection method. GcMAF increased macrophage tumoricidal activity but TNF and NO release were not involved in the mechanism.

Electrophoretic deposition of biomaterials

Science.gov (United States)

Boccaccini, A. R.; Keim, S.; Ma, R.; Li, Y.; Zhitomirsky, I.

2010-01-01

Electrophoretic deposition (EPD) is attracting increasing attention as an effective technique for the processing of biomaterials, specifically bioactive coatings and biomedical nanostructures. The well-known advantages of EPD for the production of a wide range of microstructures and nanostructures as well as unique and complex material combinations are being exploited, starting from well-dispersed suspensions of biomaterials in particulate form (microsized and nanoscale particles, nanotubes, nanoplatelets). EPD of biological entities such as enzymes, bacteria and cells is also being investigated. The review presents a comprehensive summary and discussion of relevant recent work on EPD describing the specific application of the technique in the processing of several biomaterials, focusing on (i) conventional bioactive (inorganic) coatings, e.g. hydroxyapatite or bioactive glass coatings on orthopaedic implants, and (ii) biomedical nanostructures, including biopolymer–ceramic nanocomposites, carbon nanotube coatings, tissue engineering scaffolds, deposition of proteins and other biological entities for sensors and advanced functional coatings. It is the intention to inform the reader on how EPD has become an important tool in advanced biomaterials processing, as a convenient alternative to conventional methods, and to present the potential of the technique to manipulate and control the deposition of a range of nanomaterials of interest in the biomedical and biotechnology fields. PMID:20504802

Optimization of a microfluidic electrophoretic immunoassay using a Peltier cooler.

Science.gov (United States)

Mukhitov, Nikita; Yi, Lian; Schrell, Adrian M; Roper, Michael G

2014-11-07

Successful analysis of electrophoretic affinity assays depends strongly on the preservation of the affinity complex during separations. Elevated separation temperatures due to Joule heating promotes complex dissociation leading to a reduction in sensitivity. Affinity assays performed in glass microfluidic devices may be especially prone to this problem due to poor heat dissipation due to the low thermal conductivity of glass and the large amount of bulk material surrounding separation channels. To address this limitation, a method to cool a glass microfluidic chip for performing an affinity assay for insulin was achieved by a Peltier cooler localized over the separation channel. The Peltier cooler allowed for rapid stabilization of temperatures, with 21°C the lowest temperature that was possible to use without producing detrimental thermal gradients throughout the device. The introduction of cooling improved the preservation of the affinity complex, with even passive cooling of the separation channel improving the amount of complex observed by 2-fold. Additionally, the capability to thermostabilize the separation channel allowed for utilization of higher separation voltages than what was possible without temperature control. Kinetic CE analysis was utilized as a diagnostic of the affinity assay and indicated that optimal conditions were at the highest separation voltage, 6 kV, and the lowest separation temperature, 21°C, leading to 3.4% dissociation of the complex peak during the separation. These optimum conditions were used to generate a calibration curve and produced 1 nM limits of detection, representing a 10-fold improvement over non-thermostated conditions. This methodology of cooling glass microfluidic devices for performing robust and high sensitivity affinity assays on microfluidic systems should be amenable in a number of applications. Copyright © 2014 Elsevier B.V. All rights reserved.

Mixed-mode reversed phase/positively charged repulsion chromatography for intact protein separation.

Science.gov (United States)

Ding, Ling; Guo, Zhimou; Hu, Zhuo; Liang, Xinmiao

2017-05-10

A mixed-mode reversed phase/positively charged repulsion stationary phase C8PN composed of octyl and amino group has been developed for separation of intact protein. Before the separation of proteins, a set of probe compounds were employed to evaluate the chromatographic properties of C8PN, demonstrating typical reversed phase/positively charged repulsion interaction on this stationary phase as estimated. Then the new C8PN stationary phase was used to separate a standard protein mixture on the reversed phase mode. Compared with a commercial C4 stationary phase, it showed different selectivity for some proteins. In order to better understand the properties of C8PN, the effect of acetonitrile content was investigated based on retention equation. Higher values of the equation parameters on C8PN demonstrated that the protein retentions were more sensitive to the change of acetonitrile content. Besides, the influences of buffer salt additives on the protein retentions were also studied. The retention factors of the proteins got larger with the increase of buffer salt concentration, which confirmed the positively charged repulsion interaction on the column. Finally, the C8PN was further applied to separate oxidized- and reduced- forms of Recombinant Human Growth Hormone. Our study indicated the advantages and application potential of mixed-mode reversed phase/positively charged repulsion stationary phase for intact protein separation. Copyright © 2017 Elsevier B.V. All rights reserved.

New methodology for capillary electrophoresis with ESI-MS detection: Electrophoretic focusing on inverse electromigration dispersion gradient. High-sensitivity analysis of sulfonamides in waters

Czech Academy of Sciences Publication Activity Database

Malá, Zdeňka; Gebauer, Petr; Boček, Petr

2016-01-01

Roč. 935, SEP (2016), s. 249-257 ISSN 0003-2670 R&D Projects: GA ČR(CZ) GA16-09135S Institutional support: RVO:68081715 Keywords : electrophoretic focusing * CE-ESI-MS * capillary electrophoresis Subject RIV: CB - Analytical Chemistry, Separation Impact factor: 4.950, year: 2016

Raw mechanically separated chicken meat and salmon protein hydrolysate as protein sources in extruded dog food

DEFF Research Database (Denmark)

Tjernsbekk, M. T.; Tauson, A. H.; Kraugerud, O. F.

2017-01-01

Protein quality was evaluated for mechanically separated chicken meat (MSC) and salmon protein hydrolysate (SPH), and for extruded dog foods where MSC or SPH partially replaced poultry meal (PM). Apparent total tract digestibility (ATTD) of crude protein (CP) and amino acids (AA) in the protein...

Use of seed protein polymorphism for discrimination of improvement level and geographic origin of upland rice cultivars

Directory of Open Access Journals (Sweden)

Ricardo Montalván

1998-12-01

Full Text Available Grain proteins from 58 Brazilian and nine Japanese upland rice cultivars (Oryza sativa L. were electrophoretically separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE. Densitometric scanning of the electrophoretic profiles permitted the estimation of the relative concentration of 16 protein fractions, which were used as variables for the calculation of Fisher's canonical discriminating functions. Significant differences between mean values of protein fractions were useful in distinguishing Brazilian and Japanese cultivars, as well as improved and non-improved Brazilian rice cultivars in scattered plots. Electrophoretically detectable protein polymorphism in rice grain can indicate geographic origin as well as breeding improvement level of a cultivar. Improved cultivars were those released by plant breeding institutes.Proteínas de grão de 58 genótipos de arroz brasileiro e nove japoneses foram separadas por meio de eletroforese (SDS-PAGE. A observação densitométrica dos perfis eletroforéticos permitiu avaliar as concentrações relativas de 16 frações protéicas que foram usadas como variáveis para a estimativa de funções discriminantes de Fisher. Diferenças significantes foram encontradas entre as frações protéicas dos grupos brasileiros e japoneses, assim como entre os genótipos melhorados e não melhorados. O polimorfismo protéico detectável eletroforetica-mente nos grãos de arroz pode indicar a origem geográfica e o nível de melhoramento dos cultivares.

Study on the plasma proteins of A-bomb survived patients including those suffered by the remained radioactivities. Report 2. Quantitative observation of the plasma protein fractions by electrophoretic test and to solve the problems for physiological clinical significance of its patterns

Energy Technology Data Exchange (ETDEWEB)

Makidono, J; Takanashi, S; Yoshimoto, T; Kai, T; Yoshimoto, K; Matsutani, M; Miura, M

1963-10-01

The plasma proteins of A-bombed survivors, healthy persons, long term x-ray equipment handling people (for instance the radiologists and x-ray technicians), cancer patients, and tumor irradiated cancer patients were examined by the electrophoretic test. It was found that the electrophoretic patterns of plasma proteins could be divided into normal (N-pattern) and abnormal (..beta.. and ..gamma.. patterns) patterns, when they were classified according to the accents of each fraction. The patterns of the healthy persons and the long term x-ray handling people showed normal (N) pattern, however, it showed 43% abnormal patterns in A-bombed survivors and 48% in cancer patients. Furthermore, the patterns could be changed by radiotherapy to cancer, ie., from N to ..beta.. or vice versa. As a result of the quantitative observation about individual pattern, the accents of ..beta..-globulins in ..beta..-patterns and ..gamma..-globulins in ..gamma..-patterns were found. The globulins increased in the A bomb survivors and the long term x-ray handling people, and this increase was also seen in the cases of cancer patients which showed 85% of them were effected with uclers (self disintegrated) by clinical examinations. A physiological clinical significance of these abnormal patterns (..beta.. and ..gamma..) in the plasma proteins indicates the disorders in its body and an important immunological meaning. Abnormal patterns in those who suffered by the remained radioactivities caused by the A-bomb showed 70%, whose average was much higher than those of direct A-bombed survivors. It is pointed out that, in recent days, there is a trend of more and gradual increase in the malignant neoplamsm than the disorders of direct A-bombed survivors.

Effects of cooking methods on electrophoretic patterns of rainbow trout

Directory of Open Access Journals (Sweden)

Yasemen Yanar

2011-07-01

Full Text Available The aim of this study was to determine the effects of different cooking methods on the electrophoretic patterns of rainbow trout (Oncorhynchus mykiss fillets using sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE. Raw rainbow trout were deep-fried, microwaved, grilled, and baked and then monitored for changes in the electrophoretic pattern. All cooking methods resulted in significant moisture loss when compared to the raw sample (P

Mixed-matrix membrane adsorbers for protein separation

NARCIS (Netherlands)

Avramescu, M.E.; Borneman, Z.; Wessling, M.

2003-01-01

The separation of two similarly sized proteins, bovine serum albumin (BSA) and bovine hemoglobin (Hb) was carried out using a new type of ion-exchange mixed-matrix adsorber membranes. The adsorber membranes were prepared by incorporation of various types of Lewatit ion-exchange resins into an

Pharmaceutical Perspective on Opalescence and Liquid-Liquid Phase Separation in Protein Solutions.

Science.gov (United States)

Raut, Ashlesha S; Kalonia, Devendra S

2016-05-02

Opalescence in protein solutions reduces aesthetic appeal of a formulation and can be an indicator of the presence of aggregates or precursor to phase separation in solution signifying reduced product stability. Liquid-liquid phase separation of a protein solution into a protein-rich and a protein-poor phase has been well-documented for globular proteins and recently observed for monoclonal antibody solutions, resulting in physical instability of the formulation. The present review discusses opalescence and liquid-liquid phase separation (LLPS) for therapeutic protein formulations. A brief discussion on theoretical concepts based on thermodynamics, kinetics, and light scattering is presented. This review also discusses theoretical concepts behind intense light scattering in the vicinity of the critical point termed as "critical opalescence". Both opalescence and LLPS are affected by the formulation factors including pH, ionic strength, protein concentration, temperature, and excipients. Literature reports for the effect of these formulation factors on attractive protein-protein interactions in solution as assessed by the second virial coefficient (B2) and the cloud-point temperature (Tcloud) measurements are also presented. The review also highlights pharmaceutical implications of LLPS in protein solutions.

Electrophoretic deposition of CdS coatings and their photocatalytic activities in the degradation of tetracycline antibiotic

Energy Technology Data Exchange (ETDEWEB)

Vázquez, A., E-mail: alejandro.lqi@gmail.com [Universidad Autónoma de Nuevo León, Facultad de Ciencias Químicas, Av. Universidad S/N, San Nicolás de los Garza, 66455 Nuevo León (Mexico); Hernández-Uresti, D.B., E-mail: ing.dianahdz@gmail.com [Universidad Autónoma de Nuevo León, CICFIM–Facultad de Ciencias Físico Matemáticas, Av. Universidad S/N, San Nicolás de los Garza, 66455 Nuevo León (Mexico); Obregón, S. [Universidad Autónoma de Nuevo León, CICFIM–Facultad de Ciencias Físico Matemáticas, Av. Universidad S/N, San Nicolás de los Garza, 66455 Nuevo León (Mexico)

2016-11-15

Highlights: • CdS photocatalyst was prepared by electrophoretic deposition. • The CdS coating was used in the photodegradation of antibiotics. • O{sub 2}{sup −} and ·OH radicals were responsible for the degradation of tetracycline. - Abstract: The photocatalytic activities of CdS coatings formed by electrophoretic deposition (EPD) were evaluated through the photodegradation of an antibiotic, tetracycline. First, CdS nanoparticles were synthesized under microwave irradiation of aqueous solutions containing the cadmium and sulfur precursors at stoichiometric amounts and by using trisodium citrate as stabilizer. Microwave irradiation was carried out in a conventional microwave oven at 2.45 GHz and 1650 W of nominal power, for 60 s. The CdS nanoparticles were characterized by UV–vis spectrophotometry, photoluminescence and X-ray diffraction. Electrophoretic deposition parameters were 300 mV, 600 mV and 900 mV of applied voltage between aluminum plates separated by 1 cm. The fractal dimensions of the surfaces were evaluated by atomic force microscopy and correlated to the morphological and topographic characteristics of the coatings. The photocatalytic activity of the CdS coatings was investigated by means the photodegradation of the tetracycline antibiotic under simulated sunlight irradiation. According to the results, the photoactivity of the coatings directly depends on the concentration of the precursors and the applied voltage during the deposition. The material obtained at 600 mV showed the best photocatalytic behavior, probably due to its physical properties, such as optimum load and suitable aggregate size.

Miniaturized protein separation using a liquid chromatography column on a flexible substrate

International Nuclear Information System (INIS)

Yang Yongmo; Chae, Junseok

2008-01-01

We report a prototype protein separator that successfully miniaturizes existing technology for potential use in biocompatible health monitoring implants. The prototype is a liquid chromatography (LC) column (LC mini-column) fabricated on an inexpensive, flexible, biocompatible polydimethylsiloxane (PDMS) enclosure. The LC mini-column separates a mixture of proteins using size exclusion chromatography (SEC) with polydivinylbenzene beads (5–20 µm in diameter with 10 nm pore size). The LC mini-column is smaller than any commercially available LC column by a factor of ∼11 000 and successfully separates denatured and native protein mixtures at ∼71 psi of the applied fluidic pressure. Separated proteins are analyzed using NuPAGE-gel electrophoresis, high-performance liquid chromatography (HPLC) and an automated electrophoresis system. Quantitative HPLC results demonstrate successful separation based on intensity change: within 12 min, the intensity between large and small protein peaks changed by a factor of ∼20. In further evaluation using the automated electrophoresis system, the plate height of the LC mini-column is between 36 µm and 100 µm. The prototype LC mini-column shows the potential for real-time health monitoring in applications that require inexpensive, flexible implant technology that can function effectively under non-laboratory conditions

Use of electrophoretic techniques and MALDI–TOF MS for rapid and reliable characterization of bacteria: analysis of intact cells, cell lysates, and “washed pellets”

Czech Academy of Sciences Publication Activity Database

Šalplachta, Jiří; Kubesová, Anna; Moravcová, Dana; Vykydalová, Marie; Süle, S.; Matoušková, H.; Horký, J.; Horká, Marie

2013-01-01

Roč. 405, č. 10 (2013), s. 3165-3175 ISSN 1618-2642 R&D Projects: GA MV VG20112015021; GA MV VG20102015023 Institutional support: RVO:68081715 Keywords : bacteria * electrophoretic techniques * MALDI Subject RIV: CB - Analytical Chemistry, Separation Impact factor: 3.578, year: 2013

A comparison of the adsorption of saliva proteins and some typical proteins onto the surface of hydroxyapatite

NARCIS (Netherlands)

Kawasaki, K; Kambara, M; Matsumura, H; Norde, W

2003-01-01

Adsorption of protein from saliva on hydroxyapatite was compared with adsorption of several typical proteins with different electric charges, i.e. lysozyme, human serum albumin, beta-lactoglobulin and ovalbumin. Adsorbed amounts of these proteins were determined and electrophoretic mobilities of

A comparison of the adsorption of saliva proteins and some typical proteins onto the surface of hydroxyapatite

NARCIS (Netherlands)

Kawasaki, K.; Kambara, M.; Matsumura, H.; Norde, W.

2003-01-01

Adsorption of protein from saliva on hydroxyapatite was compared with adsorption of several typical proteins with different electric charges, i.e. lysozyme, human serum albumin, @b-lactoglobulin and ovalbumin. Adsorbed amounts of these proteins were determined and electrophoretic mobilities of

Electrophoretic Porosimetry of Sol-Gels

Science.gov (United States)

Snow, L. A.; Smith, D. D.; Sibille, L.; Hunt, A. J.; Ng, J.; Rose, M. Franklin (Technical Monitor)

2000-01-01

It has been hypothesized that gravity has an effect on the formation and resulting microstructure of sol-gels. In order to more clearly resolve the effect of gravity, pores may be non-destructively analyzed in the wet gel, circumventing the shrinkage and coarsening associated with the drying procedure. We discuss the development of an electrophoretic technique, analogous to affinity chromatography, for the determination of pore size distribution and its application to silica gels. Specifically a monodisperse charged dye is monitored by an optical densitometer as it moves through the wet gel under the influence of an electric field. The transmittance data (output) represents the convolution of the dye concentration profile at the beginning of the run (input) with the pore size distribution (transfer function), i.e. linear systems theory applies. Because of the practical difficulty in producing a delta function input dye profile we prefer instead to use a step function. Average pore size is then related to the velocity of this dye front, while the pore size distribution is related to the spreading of the front. Preliminary results of this electrophoretic porosimetry and its application to ground and space-grown samples will be discussed.

Electrophoretic protein polymorphisms in Kaingang and Guarani Indians of Southern Brazil.

Science.gov (United States)

Salzano, Francisco M; Callegari-Jacques, Sidia M; Weimer, Tania A; Franco, Maria H L P; Hutz, Mara H; Petzl-Erler, Maria L

1997-01-01

A total of 337 Kaingang and Guarani Indians from two localities were studied in relation to 18 protein genetic loci. In one of the localities, members of these two groups live side by side but show little genetic similarity, emphasizing the influence of cultural factors in the mating behavior of human groups. Integrating the present results with previous ones, it was verified that the genetic relationships among six Kaingang populations do not follow the pattern expected from their geographical distribution. Comparisons made with three other Ge˜-speaking tribes indicate that the Kaingang did not separate well from them. Most (96%) of the variability in the six polymorphic systems considered occur at the intrapopulational level. Am. J. Hum. Biol. 9:505-512, 1997. © 1997 Wiley-Liss, Inc. Copyright © 1997 Wiley-Liss, Inc.

Proteomic study of muscle sarcoplasmic proteins using AUT-PAGE/SDS-PAGE as two-dimensional gel electrophoresis.

Science.gov (United States)

Picariello, Gianluca; De Martino, Alessandra; Mamone, Gianfranco; Ferranti, Pasquale; Addeo, Francesco; Faccia, Michele; Spagnamusso, Salvatore; Di Luccia, Aldo

2006-03-20

In the present study, an alternative procedure for two-dimensional (2D) electrophoretic analysis in proteomic investigation of the most represented basic muscle water-soluble proteins is suggested. Our method consists of Acetic acid-Urea-Triton polyacrylamide gel (AUT-PAGE) analysis in the first dimension and standard sodium dodecyl sulphate polyacrylamide gel (SDS-PAGE) in the second dimension. Although standard two-dimensional Immobilized pH Gradient-Sodium Dodecyl-Sulphate (2D IPG-SDS) gel electrophoresis has been successfully used to study these proteins, most of the water-soluble proteins are spread on the alkaline part of the 2D map and are poorly focused. Furthermore, the similarity in their molecular weights impairs resolution of the classical approach. The addition of Triton X-100, a non-ionic detergent, into the gel induces a differential electrophoretic mobility of proteins as a result of the formation of mixed micelles between the detergent and the hydrophobic moieties of polypeptides, separating basic proteins with a criterion similar to reversed phase chromatography based on their hydrophobicity. The acid pH induces positive net charges, increasing with the isoelectric point of proteins, thus allowing enhanced resolution in the separation. By using 2D AUT-PAGE/SDS electrophoresis approach to separate water-soluble proteins from fresh pork and from dry-cured products, we could spread proteins over a greater area, achieving a greater resolution than that obtained by IPG in the pH range 3-10 and 6-11. Sarcoplasmic proteins undergoing proteolysis during the ripening of products were identified by Matrix Assisted Laser Desorption/Ionization-Time of Flight (MALDI-ToF) mass spectrometry peptide mass fingerprinting in a easier and more effective way. Two-dimensional AUT-PAGE/SDS electrophoresis has allowed to simplify separation of sarcoplasmic protein mixtures making this technique suitable in the defining of quality of dry-cured pork products by immediate

Ribosomal DNA-binding proteins in the nucleolus of Physarum polycephalum

International Nuclear Information System (INIS)

Graham-Lorence, S.E.

1987-01-01

In Physarum polycephalum, the nucleoli are extra chromosomal structures containing 200 to 400 copies of a linear 60 kilobase palindromic rDNA molecule. These rDNA molecules are organized into minichromosomes which apparently are held within a nucleolar protein matrix. To obtained evidence for attachment of the rDNA to such a matrix, both intact and lithium diiodosalicylate/NaCl-extracted nucleoli were digested for various lengths of time with micrococcal nuclease, so that portions of the rDNA molecules not attached within the nucleolar structure would be released. Nucleolar DNA-binding proteins were determined by blotting electrophoretically separated proteins from SDS-polyacrylamide gels onto nitrocellulose paper and probing them with radiolabeled DNA. In addition to the histones and lexosome proteins, eight DNA-binding proteins were identified having molecular weights of 25, 38, 47, 53, 55, 67, and 70 kD, with the 47, 53, 67, and 70 kD proteins requiring Ca 2+ for binding

ELECTROPHORETIC MOBILITY OF MYCOBACTERIUM AVIUM COMPLEX ORGANISMS

Science.gov (United States)

The electrophoretic mobilities (EPMs) of thirty Mycobacterium avium Complex (MAC) organisms were measured. The EPMs of fifteen clinical isolates ranged from -1.9 to -5.0 µm cm V-1s-1, and the EPMs of fifteen environmental isolates ranged from -1...

Continuous electrophoretic purification of individual analytes from multicomponent mixtures.

Science.gov (United States)

McLaren, David G; Chen, David D Y

2004-04-15

Individual analytes can be isolated from multicomponent mixtures and collected in the outlet vial by carrying out electrophoretic purification through a capillary column. Desired analytes are allowed to migrate continuously through the column under the electric field while undesired analytes are confined to the inlet vial by application of a hydrodynamic counter pressure. Using pressure ramping and buffer replenishment techniques, 18% of the total amount present in a bulk sample can be purified when the resolution to the adjacent peak is approximately 3. With a higher resolution, the yield could be further improved. Additionally, by periodically introducing fresh buffer into the sample, changes in pH and conductivity can be mediated, allowing higher purity (>or=99.5%) to be preserved in the collected fractions. With an additional reversed cycle of flow counterbalanced capillary electrophoresis, any individual component in a sample mixture can be purified providing it can be separated in an electrophoresis system.

Artificial receptor-functionalized nanoshell: facile preparation, fast separation and specific protein recognition

Science.gov (United States)

Ouyang, Ruizhuo; Lei, Jianping; Ju, Huangxian

2010-05-01

This work combined molecular imprinting technology with superparamagnetic nanospheres as the core to prepare artificial receptor-functionalized magnetic nanoparticles for separation of homologous proteins. Using dopamine as a functional monomer, novel surface protein-imprinted superparamagnetic polydopamine (PDA) core-shell nanoparticles were successfully prepared in physiological conditions, which could maintain the natural structure of a protein template and achieved the development of molecularly imprinted polymers (MIPs) from one dimension to zero dimension for efficient recognition towards large biomolecules. The resultant nanoparticles could be used for convenient magnetic separation of homologous proteins with high specificity. The nanoparticles possessed good monodispersibility, uniform surface morphology and high saturation magnetization value. The bound amounts of template proteins measured by both indirect and direct methods were in good agreement. The maximum number of imprinted cavities on the surface of the bovine hemoglobin (Hb)-imprinted nanoshell was 2.21 × 1018 g - 1, which well matched their maximum binding capacity toward bovine Hb. Both the simple method for preparation of MIPs and the magnetic nanospheres showed good application potential in fast separation, effective concentration and selective biosensing of large protein molecules.

Artificial receptor-functionalized nanoshell: facile preparation, fast separation and specific protein recognition

Energy Technology Data Exchange (ETDEWEB)

Ouyang, Ruizhuo; Lei Jianping; Ju Huangxian, E-mail: jpl@nju.edu.cn, E-mail: hxju@nju.edu.cn [Key Laboratory of Analytical Chemistry for Life Science (Education Ministry of China), Department of Chemistry, Nanjing University, Nanjing 210093 (China)

2010-05-07

This work combined molecular imprinting technology with superparamagnetic nanospheres as the core to prepare artificial receptor-functionalized magnetic nanoparticles for separation of homologous proteins. Using dopamine as a functional monomer, novel surface protein-imprinted superparamagnetic polydopamine (PDA) core-shell nanoparticles were successfully prepared in physiological conditions, which could maintain the natural structure of a protein template and achieved the development of molecularly imprinted polymers (MIPs) from one dimension to zero dimension for efficient recognition towards large biomolecules. The resultant nanoparticles could be used for convenient magnetic separation of homologous proteins with high specificity. The nanoparticles possessed good monodispersibility, uniform surface morphology and high saturation magnetization value. The bound amounts of template proteins measured by both indirect and direct methods were in good agreement. The maximum number of imprinted cavities on the surface of the bovine hemoglobin (Hb)-imprinted nanoshell was 2.21 x 10{sup 18} g{sup -1}, which well matched their maximum binding capacity toward bovine Hb. Both the simple method for preparation of MIPs and the magnetic nanospheres showed good application potential in fast separation, effective concentration and selective biosensing of large protein molecules.

Albumin coatings by alternating current electrophoretic deposition for improving corrosion resistance and bioactivity of titanium implants

International Nuclear Information System (INIS)

Höhn, Sarah; Braem, Annabel; Neirinck, Bram; Virtanen, Sannakaisa

2017-01-01

Although Ti alloys are generally regarded to be highly corrosion resistant, inflammatory conditions following surgery can instigate breakdown of the TiO 2 passivation layer leading to an increased metal ion release. Furthermore proteins present in the surrounding tissue will readily adsorb on a titanium surface after implantation. In this paper alternating current electrophoretic deposition (AC-EPD) of bovine serum albumin (BSA) on Ti6Al4V was investigated in order to increase the corrosion resistance and control the protein adsorption capability of the implant surface. The Ti6Al4V surface was characterized with SEM, XPS and ToF-SIMS after long-term immersion tests under physiological conditions and simulated inflammatory conditions either in Dulbecco's Modified Eagle Medium (DMEM) or DMEM supplemented with fetal calf serum (FCS). The analysis showed an increased adsorption of amino acids and proteins from the different immersion solutions. The BSA coating was shown to prevent selective dissolution of the vanadium (V) rich β-phase, thus effectively limiting metal ion release to the environment. Electrochemical impedance spectroscopy measurements confirmed an increase of the corrosion resistance for BSA coated surfaces as a function of immersion time due to the time-dependent adsorption of the different amino acids (from DMEM) and proteins (from FCS) as observed by ToF-SIMS analysis. - Highlights: • Alternating current electrophoretic deposition (AC-EPD) of bovine serum albumin was investigated on Ti6Al4V. • The surface was characterized with SEM, XPS and ToF-SIMS after long-term immersion tests at pH 7 and pH 5. • The analysis showed an increased adsorption of amino acids (DMEM) and proteins (DMEM + FCS). • BSA was shown to prevent dissolution of the β-phase, limiting metal ion release and increase of corrosion resistance. • Ratios calculated by means of ToF-SIMS show that the protein will have different orientations during adsorption.

Albumin coatings by alternating current electrophoretic deposition for improving corrosion resistance and bioactivity of titanium implants

Energy Technology Data Exchange (ETDEWEB)

Höhn, Sarah, E-mail: sarah.hoehn@fau.de [Institute for Surface Science and Corrosion, Dept. of Mat. Science, University of Erlangen-Nürnberg, 91058 Erlangen, Germany. (Germany); Braem, Annabel, E-mail: annabel.braem@kuleuven.be [KU Leuven Department of Materials Engineering, Kasteelpark Arenberg 44, Box 2450, 3001 Leuven (Belgium); Neirinck, Bram, E-mail: bram.neirinck@3DSystems.com [KU Leuven Department of Materials Engineering, Kasteelpark Arenberg 44, Box 2450, 3001 Leuven (Belgium); Virtanen, Sannakaisa, E-mail: virtanen@ww.uni-erlangen.de [Institute for Surface Science and Corrosion, Dept. of Mat. Science, University of Erlangen-Nürnberg, 91058 Erlangen, Germany. (Germany)

2017-04-01

Although Ti alloys are generally regarded to be highly corrosion resistant, inflammatory conditions following surgery can instigate breakdown of the TiO{sub 2} passivation layer leading to an increased metal ion release. Furthermore proteins present in the surrounding tissue will readily adsorb on a titanium surface after implantation. In this paper alternating current electrophoretic deposition (AC-EPD) of bovine serum albumin (BSA) on Ti6Al4V was investigated in order to increase the corrosion resistance and control the protein adsorption capability of the implant surface. The Ti6Al4V surface was characterized with SEM, XPS and ToF-SIMS after long-term immersion tests under physiological conditions and simulated inflammatory conditions either in Dulbecco's Modified Eagle Medium (DMEM) or DMEM supplemented with fetal calf serum (FCS). The analysis showed an increased adsorption of amino acids and proteins from the different immersion solutions. The BSA coating was shown to prevent selective dissolution of the vanadium (V) rich β-phase, thus effectively limiting metal ion release to the environment. Electrochemical impedance spectroscopy measurements confirmed an increase of the corrosion resistance for BSA coated surfaces as a function of immersion time due to the time-dependent adsorption of the different amino acids (from DMEM) and proteins (from FCS) as observed by ToF-SIMS analysis. - Highlights: • Alternating current electrophoretic deposition (AC-EPD) of bovine serum albumin was investigated on Ti6Al4V. • The surface was characterized with SEM, XPS and ToF-SIMS after long-term immersion tests at pH 7 and pH 5. • The analysis showed an increased adsorption of amino acids (DMEM) and proteins (DMEM + FCS). • BSA was shown to prevent dissolution of the β-phase, limiting metal ion release and increase of corrosion resistance. • Ratios calculated by means of ToF-SIMS show that the protein will have different orientations during adsorption.

CZE separation of strawberry anthocyanins with acidic buffer and comparison with HPLC.

Science.gov (United States)

Comandini, Patrizia; Blanda, Giampaolo; Cardinali, Andrea; Cerretani, Lorenzo; Bendini, Alessandra; Caboni, Maria Fiorenza

2008-10-01

Anthocyanins, the major colourants of strawberries, are polar pigments that are positively charged at low pH. Herein, we have assessed a new analytical method for the separation of anthocyanins using CZE. Acidic buffer solutions (pH pigments in the cation flavylium form and achieve high molar absorptivity at 510 nm. These spectral properties enabled us to identify strawberry anthocyanins in a preliminary stage by detection in the visible range, although the method was optimised at 280 nm to obtain the best S/N. The effects of buffer composition highlighted the necessity of adding an organic modifier to the running buffer to obtain a suitable separation. The electrophoretic method permitted the separation of the three main anthocyanins of strawberry extracts, namely pelargonidin 3-glucoside (Pg-glu), pelargonidin 3-rutinoside and cyanidin 3-glucoside. The electrophoretic results, expressed as retention time and separation efficiency of the major anthocyanin (Pg-glu), were compared to those achieved in HPLC, the analytical technique traditionally used for the investigation of anthocyanins in vegetable matrix. The content of Pg-glu in strawberries (cv. Camarosa), calculated with HPCE and HPLC methods, resulted respectively in 11.41 mg/L and 11.37 mg/L.

Ndj1, a telomere-associated protein, regulates centrosome separation in budding yeast meiosis

Science.gov (United States)

Li, Ping; Shao, Yize; Jin, Hui

2015-01-01

Yeast centrosomes (called spindle pole bodies [SPBs]) remain cohesive for hours during meiotic G2 when recombination takes place. In contrast, SPBs separate within minutes after duplication in vegetative cells. We report here that Ndj1, a previously known meiosis-specific telomere-associated protein, is required for protecting SPB cohesion. Ndj1 localizes to the SPB but dissociates from it ∼16 min before SPB separation. Without Ndj1, meiotic SPBs lost cohesion prematurely, whereas overproduction of Ndj1 delayed SPB separation. When produced ectopically in vegetative cells, Ndj1 caused SPB separation defects and cell lethality. Localization of Ndj1 to the SPB depended on the SUN domain protein Mps3, and removal of the N terminus of Mps3 allowed SPB separation and suppressed the lethality of NDJ1-expressing vegetative cells. Finally, we show that Ndj1 forms oligomeric complexes with Mps3, and that the Polo-like kinase Cdc5 regulates Ndj1 protein stability and SPB separation. These findings reveal the underlying mechanism that coordinates yeast centrosome dynamics with meiotic telomere movement and cell cycle progression. PMID:25897084

From image processing to classification: IV. Classification of electrophoretic patterns by neural networks and statistical methods enable quality assessment of wheat varieties for breadmaking

DEFF Research Database (Denmark)

Jensen, Kirsten; Kesmir, Can; Søndergaard, Ib

1996-01-01

The end-use quality of products made from doughs consisting of wheat flour and water is often dependent upon the storage (gluten) proteins of the grain endosperm. Today the electrophoretic patterns of the high molecular weight (HMW) glutenin subunits are used for quality selections in wheat breed...

Electrophoretic deposition of composite hydroxyapatite-chitosan coatings

International Nuclear Information System (INIS)

Pang Xin; Zhitomirsky, Igor

2007-01-01

Cathodic electrophoretic deposition has been utilized for the fabrication of composite hydroxyapatite-chitosan coatings on 316L stainless steel substrates. The addition of chitosan to the hydroxyapatite suspensions promoted the electrophoretic deposition of the hydroxyapatite nanoparticles and resulted in the formation of composite coatings. The obtained coatings were investigated by X-ray diffraction, thermogravimetric and differential thermal analysis, scanning and transmission electron microscopy, potentiodynamic polarization measurements, and electrochemical impedance spectroscopy. It was shown that the deposit composition can be changed by a variation of the chitosan or hydroxyapatite concentration in the solutions. Experimental conditions were developed for the fabrication of hydroxyapatite-chitosan nanocomposites containing 40.9-89.8 wt.% hydroxyapatite. The method enabled the formation of adherent and uniform coatings of thicknesses up to 60 μm. X-ray studies revealed that the preferred orientation of the hydroxyapatite nanoparticles in the chitosan matrix increases with decreasing hydroxyapatite content in the composite coatings. The obtained coatings provided the corrosion protection for the 316L stainless steel substrates

Isotachophoresis in narrow-bore tubes. Influence of the diameter of the separation compartment

NARCIS (Netherlands)

Verheggen, T.P.E.M.; Mikkers, F.E.P.; Everaerts, F.M.

1977-01-01

The importance of the inside diameter of the separation compartment of electrophoretic equipment in which tubes are used in achieving optimal stabilization against conventive disturbances and for limiting the temperature increase is demonstrated. Both from a theoretical point of view and

Effect of AOT Microemulsion Composition on the Hydrodynamic Diameter and Electrophoretic Mobility of Titanium Oxide Nanoparticles

Science.gov (United States)

Shaparenko, N. O.; Beketova, D. I.; Demidova, M. G.; Bulavchenko, A. I.

2018-05-01

The hydrodynamic diameter and electrophoretic mobility of titania nanoparticles in AOT microemulsions are studied depending on their water content (from 0 to 1.5 vol %), chloroform content in n-decane-chloroform mixture (from 0 to 30 vol %) and temperature (from 0 to 60°C). Considerable changes in diameter (from 20 to 400 nm) are detected upon adding water to the microemulsion. The electrophoretic mobility grows by 2-3 times upon adding chloroform, or as the temperature falls. The observed features allow us to halve the time of electrophoretic concentration for 140 nm TiO2 nanoparticles, and to concentrate 14 nm nanoparticles that do not exhibit electrophoretic mobility in the absence of chloroform.

Free flow electrophoresis separation and AMS quantitation of 14C-naphthalene-protein adducts

International Nuclear Information System (INIS)

Buchholz, Bruce A.; Haack, Kurt W.; Sporty, Jennifer L.; Buckpitt, Alan R.; Morin, Dexter

2010-01-01

Naphthalene is a volatile aromatic hydrocarbon to which humans are exposed from a variety of sources including mobile air sources and cigarette smoke. Naphthalene produces dose-(concentration)dependent injury to airway epithelial cells of murine lung which is observed at concentrations well below the current occupational exposure standard. Toxicity is dependent upon the cytochrome P450 mediated metabolic activation of the parent substrate to unstable metabolites which become bound covalently to tissue proteins. Nearly 70 proteins have been identified as forming adducts with reactive naphthalene metabolites using in vitro systems but very little work has been conducted in vivo because reasonably large amounts (100 μCi) of 14 C labeled parent compound must be administered to generate detectable adduct levels on storage phosphor screens following separation of labeled proteins by 2D gel electrophoresis. The work described here was done to provide proof of concept that protein separation by free flow electrophoresis followed by AMS detection of protein fractions containing protein bound reactive metabolites would provide adducted protein profiles in animals dosed with trace quantities of labeled naphthalene. Mice were administered 200 mg/kg naphthalene intraperitoneally at a calculated specific activity of 2 DPM/nmol (1 pCi/nmol) and respiratory epithelial tissue was obtained by lysis lavage 4 h post injection. Free flow electrophoresis (FFE) separates proteins in the liquid phase over a large pH range (2.5-11.5) using low molecular weight acids and bases to modify the pH. The apparatus separates fractions into standard 96-well plates that can be used in other protein analysis techniques. The buffers of the fractions have very high carbon content, however, and need to be dialyzed to yield buffers compatible with 14 C-AMS. We describe the processing techniques required to couple FFE to AMS for quantitation of protein adducts.

Free flow electrophoresis separation and AMS quantitation of 14C-naphthalene-protein adducts

Science.gov (United States)

Buchholz, Bruce A.; Haack, Kurt W.; Sporty, Jennifer L.; Buckpitt, Alan R.; Morin, Dexter

2010-04-01

Naphthalene is a volatile aromatic hydrocarbon to which humans are exposed from a variety of sources including mobile air sources and cigarette smoke. Naphthalene produces dose-(concentration)dependent injury to airway epithelial cells of murine lung which is observed at concentrations well below the current occupational exposure standard. Toxicity is dependent upon the cytochrome P450 mediated metabolic activation of the parent substrate to unstable metabolites which become bound covalently to tissue proteins. Nearly 70 proteins have been identified as forming adducts with reactive naphthalene metabolites using in vitro systems but very little work has been conducted in vivo because reasonably large amounts (100 μCi) of 14C labeled parent compound must be administered to generate detectable adduct levels on storage phosphor screens following separation of labeled proteins by 2D gel electrophoresis. The work described here was done to provide proof of concept that protein separation by free flow electrophoresis followed by AMS detection of protein fractions containing protein bound reactive metabolites would provide adducted protein profiles in animals dosed with trace quantities of labeled naphthalene. Mice were administered 200 mg/kg naphthalene intraperitoneally at a calculated specific activity of 2 DPM/nmol (1 pCi/nmol) and respiratory epithelial tissue was obtained by lysis lavage 4 h post injection. Free flow electrophoresis (FFE) separates proteins in the liquid phase over a large pH range (2.5-11.5) using low molecular weight acids and bases to modify the pH. The apparatus separates fractions into standard 96-well plates that can be used in other protein analysis techniques. The buffers of the fractions have very high carbon content, however, and need to be dialyzed to yield buffers compatible with 14C-AMS. We describe the processing techniques required to couple FFE to AMS for quantitation of protein adducts.

Phosphorylation of a specific cdk site in E2F-1 affects its electrophoretic mobility and promotes pRB-binding in vitro

DEFF Research Database (Denmark)

Peeper, D S; Keblusek, P; Helin, K

1995-01-01

of the retinoblastoma gene (pRB). We find that E2F-1 proteins are heterogeneously phosphorylated in insect cells, as a result of which they migrate as a doublet on SDS-polyacrylamide gels. This electrophoretic shift is shown to be dependent upon specific phosphorylation of E2F-1 on serine-375 (S375), near the p...

[Effect of freezing and cooking on the texture and electrophoretic pattern of the proteins of octopus arms (Octopus vulgaris)].

Science.gov (United States)

Reyes, Genara; Nirchio, Mauro; Bello, Rafael; Borderías, Javier

2014-09-01

Texture is the most valuable feature in cephalopods. Factors that mainly affect the texture of octopus are: freezing, scalding and cooking. The aim of this study was to assess the effect of freezing, scalding and length of cooking time on the texture and electrophoretic pattern of proteins of octopus arms. Octopuses were trapped near Margarita Island and carried with ice to the laboratory where they were packed and subjected to: a) freezing at -27 degrees C or at -20 degrees C b) scalding c) cooking for 25 min, 35 min or 45 min. Shear force was determined by Kramer cell on strips of octopus arms. SDS-PAGE was done according to the Laemmli method with 12% polyacrilamide gels. A sensory evaluation of the preference of texture was carried out using a hedonic scale of 7-points and a non-trained panel. Octopus texture was not affected by freezing temperature or scalding. Frozen octopus was softer after cooking than fresh. The longer the cooking time was, the softer the octopus was. Myosin heavy chain (MHC) was not significantly affected by scalding or cooking; however large aggregates heavier than MHC, new bands and loss of resolution of the bands appeared. Myosin and paramyosin bands were more affected by freezing prior to cooking.

A method for UV-bonding in the fabrication of glass electrophoretic microchips.

Science.gov (United States)

Huang, Z; Sanders, J C; Dunsmor, C; Ahmadzadeh, H; Landers, J P

2001-10-01

This paper presents an approach for the development of methodologies amenable to simple and inexpensive microchip fabrication, potentially applicable to dissimilar materials bonding and chip integration. The method involves a UV-curable glue that can be used for glass microchip fabrication bonding at room temperature. This involves nothing more than fabrication of glue "guide channels" into the microchip architecture that upon exposure to the appropriate UV light source, bonds the etched plate and cover plate together. The microchip performance was verified by capillary zone electrophoresis (CZE) of small fluorescent molecules with no microchannel surface modification carried out, as well as with a DNA fragment separation following surface modification. The performance of these UV-bonded electrophoretic microchips indicates that this method may provide an alternative to high temperature bonding.

From image processing to classification: IV. Classification of electrophoretic patterns by neural networks and statistical methods enable quality assessment of wheat varieties for bread making

DEFF Research Database (Denmark)

Jensen, K.; Kesmir, Can; Søndergaard, Ib

1996-01-01

The end-use quality of products made from doughs consisting of wheat flour and water is often dependent upon the storage (gluten) proteins of the grain endosperm. Today the electrophoretic patterns of the high molecular weight (HMW) glutenin subunits are used for quality selections in wheat breed...

A simple immunoblotting method after separation of proteins in agarose gel

DEFF Research Database (Denmark)

Koch, C; Skjødt, K; Laursen, I

1985-01-01

A simple and sensitive method for immunoblotting of proteins after separation in agarose gels is described. It involves transfer of proteins onto nitrocellulose paper simply by diffusion through pressure, a transfer which only takes about 10 min. By this method we have demonstrated the existence ...

Electrophoretic separation of cells and particles from rat pituitary and rat spleen

Science.gov (United States)

Hymer, Wesley C.

1993-01-01

There are 3 parts to the IML-2 TX-101 experiment. Part 1 is a pituitary cell culture experiment. Part 2 is a pituitary cell separation experiment using the Japanese free flow electrophoresis unit (FFEU). Part 3 is a pituitary secretory granule separation experiment using the FFEU. The objectives of this three part experiment are: (1) to determine the kinetics of production of biologically active growth hormone (GH) and prolactin (PRL) in rat pituitary GH and PRL cells in microgravity (micro-g); (2) to investigate three mechanisms by which a micro-g-induced lesion in hormone production may occur; and (3) to determine the quality of separations of pituitary cells and organelles by continuous flow electrophoresis (CFE) in micro-g under conditions where buoyancy-induced convection is eliminated.

Microphase Separation Controlled beta-Sheet Crystallization Kinetics in Fibrous Proteins

International Nuclear Information System (INIS)

Hu, X.; Lu, Q.; Kaplan, D.; Cebe, P.

2009-01-01

Silk is a naturally occurring fibrous protein with a multiblock chain architecture. As such, it has many similarities with synthetic block copolymers, including the possibility for e-sheet crystallization restricted within the crystallizable blocks. The mechanism of isothermal crystallization kinetics of e-sheet crystals in silk multiblock fibrous proteins is reported in this study. Kinetics theories, such as Avrami analysis which was established for studies of synthetic polymer crystal growth, are for the first time extended to investigate protein self-assembly in e-sheet rich Bombyx mori silk fibroin samples, using time-resolved Fourier transform infrared spectroscopy (FTIR), differential scanning calorimetry (DSC) and synchrotron real-time wide-angle X-ray scattering (WAXS). The Avrami exponent, n, was close to 2 for all methods and crystallization temperatures, indicating formation of e-sheet crystals in silk proteins is different from the 3-D spherulitic crystal growth found in synthetic polymers. Observations by scanning electron microscopy support the view that the protein structures vary during the different stages of crystal growth, and show a microphase separation pattern after chymotrypsin enzyme biodegradation. We present a model to explain the crystallization of the multiblock silk fibroin protein, by analogy to block copolymers: crystallization of e-sheets occurs under conditions of geometrical restriction caused by phase separation of the crystallizable and uncrystallizable blocks. This crystallization model could be widely applicable in other proteins with multiblock (i.e., crystallizable and noncrystallizable) domains.

Integration of carboxyl modified magnetic particles and aqueous two-phase extraction for selective separation of proteins.

Science.gov (United States)

Gai, Qingqing; Qu, Feng; Zhang, Tao; Zhang, Yukui

2011-07-15

Both of the magnetic particle adsorption and aqueous two-phase extraction (ATPE) were simple, fast and low-cost method for protein separation. Selective proteins adsorption by carboxyl modified magnetic particles was investigated according to protein isoelectric point, solution pH and ionic strength. Aqueous two-phase system of PEG/sulphate exhibited selective separation and extraction for proteins before and after magnetic adsorption. The two combination ways, magnetic adsorption followed by ATPE and ATPE followed by magnetic adsorption, for the separation of proteins mixture of lysozyme, bovine serum albumin, trypsin, cytochrome C and myloglobin were discussed and compared. The way of magnetic adsorption followed by ATPE was also applied to human serum separation. Copyright © 2011 Elsevier B.V. All rights reserved.

Modeling of protein electrophoresis in silica colloidal crystals having brush layers of polyacrylamide

Science.gov (United States)

Birdsall, Robert E.; Koshel, Brooke M.; Hua, Yimin; Ratnayaka, Saliya N.; Wirth, Mary J.

2013-01-01

Sieving of proteins in silica colloidal crystals of mm dimensions is characterized for particle diameters of nominally 350 and 500 nm, where the colloidal crystals are chemically modified with a brush layer of polyacrylamide. A model is developed that relates the reduced electrophoretic mobility to the experimentally measurable porosity. The model fits the data with no adjustable parameters for the case of silica colloidal crystals packed in capillaries, for which independent measurements of the pore radii were made from flow data. The model also fits the data for electrophoresis in a highly ordered colloidal crystal formed in a channel, where the unknown pore radius was used as a fitting parameter. Plate heights as small as 0.4 μm point to the potential for miniaturized separations. Band broadening increases as the pore radius approaches the protein radius, indicating that the main contribution to broadening is the spatial heterogeneity of the pore radius. The results quantitatively support the notion that sieving occurs for proteins in silica colloidal crystals, and facilitate design of new separations that would benefit from miniaturization. PMID:23229163

Electrophoretic mobility patterns of collagen following laser welding

Science.gov (United States)

Bass, Lawrence S.; Moazami, Nader; Pocsidio, Joanne O.; Oz, Mehmet C.; LoGerfo, Paul; Treat, Michael R.

1991-06-01

Clinical application of laser vascular anastomosis in inhibited by a lack of understanding of its mechanism. Whether tissue fusion results from covalent or non-covalent bonding of collagen and other structural proteins is unknown. We compared electrophoretic mobility of collagen in laser treated and untreated specimens of rat tail tendon (>90% type I collagen) and rabbit aorta. Welding was performed, using tissue shrinkage as the clinical endpoint, using the 808 nm diode laser (power density 14 watts/cm2) and topical indocyanine green dye (max absorption 805 nm). Collagen was extracted with 8 M urea (denaturing), 0.5 M acetic acid (non-denaturing) and acetic acid/pepsin (cleaves non- helical protein). Mobility patterns on gel electrophoresis (SDS-PAGE) after urea or acetic acid extraction were identical in the lasered and control tendon and vessel (confirmed by optical densitometry), revealing no evidence of formation of novel covalent bonds. Alpha and beta band intensity was diminished in pepsin incubated lasered specimens compared with controls (optical density ratio 0.00 +/- 9 tendon, 0.65 +/- 0.12 aorta), indicating the presence of denatured collagen. With the laser parameters used, collagen is denatured without formation of covalent bonds, suggesting that non-covalent interaction between denatured collagen molecules may be responsible for the weld. Based on this mechanism, welding parameters can be chosen which produce collagen denaturation without cell death.

A novel method for the preparation of electrophoretic display microcapsules

Energy Technology Data Exchange (ETDEWEB)

Liu, Xiao-Meng; He, Jing; Liu, Sheng-Yun [State Key Laboratory of Organic-Inorganic Composites, Beijing University of Chemical Technology, Beijing 100029 (China); Chen, Jian-Feng [State Key Laboratory of Organic-Inorganic Composites, Beijing University of Chemical Technology, Beijing 100029 (China); Research Center of the Ministry of Education for High Gravity Engineering and Technology, Beijing University of Chemical Technology, Beijing 100029 (China); Le, Yuan, E-mail: leyuan@mail.buct.edu.cn [State Key Laboratory of Organic-Inorganic Composites, Beijing University of Chemical Technology, Beijing 100029 (China)

2014-07-01

Highlights: • The electrophoretic display microcapsules were prepared by coaxial jet method aided by gas spray. • The positions of inner tube, liquid and gas flow rate of the process were investigated. • The size and shell thickness of the prepared microcapsules were controllable. • The prepared microcapsules had high coating ratio and exhibit reversible response to DC field. - Abstract: The narrow distributed electrophoretic display microcapsules containing electrophoretic ink were prepared using coaxial jet method aided by gas spray. Experimental results showed the size and shell thickness of the microcapsules could be controlled by adjusting flow rates of core and shell fluids as well as gas. The as-prepared white and red microcapsules, with average size of 100 and 200 μm respectively, had high coating ratio (above 90%) and exhibited reversible response to DC electric field. Compared with the approach of other microencapsulation methods, the new technique not only has a simple procedure but also provides a more effective way of size control. This novel method is expected to prepare microcapsules with potential application in the fields of electronic paper and other material science.

Protein phosphorylation systems in postmortem human brain

International Nuclear Information System (INIS)

Walaas, S.I.; Perdahl-Wallace, E.; Winblad, B.; Greengard, P.

1989-01-01

Protein phosphorylation systems regulated by cyclic adenosine 3',5'-monophosphate (cyclic AMP), or calcium in conjunction with calmodulin or phospholipid/diacylglycerol, have been studied by phosphorylation in vitro of particulate and soluble fractions from human postmortem brain samples. One-dimensional or two-dimensional gel electrophoretic protein separations were used for analysis. Protein phosphorylation catalyzed by cyclic AMP-dependent protein kinase was found to be highly active in both particulate and soluble preparations throughout the human CNS, with groups of both widely distributed and region-specific substrates being observed in different brain nuclei. Dopamine-innervated parts of the basal ganglia and cerebral cortex contained the phosphoproteins previously observed in rodent basal ganglia. In contrast, calcium/phospholipid-dependent and calcium/calmodulin-dependent protein phosphorylation systems were less prominent in human postmortem brain than in rodent brain, and only a few widely distributed substrates for these protein kinases were found. Protein staining indicated that postmortem proteolysis, particularly of high-molecular-mass proteins, was prominent in deeply located, subcortical regions in the human brain. Our results indicate that it is feasible to use human postmortem brain samples, when obtained under carefully controlled conditions, for qualitative studies on brain protein phosphorylation. Such studies should be of value in studies on human neurological and/or psychiatric disorders

Improved gel electrophoresis matrix for hydrophobic protein separation and identification.

Science.gov (United States)

Tokarski, Caroline; Fillet, Marianne; Rolando, Christian

2011-03-01

We propose an improved acrylamide gel for the separation of hydrophobic proteins. The separation strategy is based on the incorporation of N-alkylated and N,N'-dialkylated acrylamide monomers in the gel composition in order to increase hydrophobic interactions between the gel matrix and the membrane proteins. Focusing on the most efficient monomer, N,N'-dimethylacrylamide, the potentiality of the new matrix was evaluated on membrane proteins of the human colon HCT-116 cell line. Protein analysis was performed using an adapted analytical strategy based on FT-ICR tandem mass spectrometry. As a result of this comparative study, including advanced reproducibility experiments, more hydrophobic proteins were identified in the new gel (average GRAVY: -0.085) than in the classical gel (average GRAVY: -0.411). Highly hydrophobic peptides were identified reaching a GRAVY value up to 1.450, therefore indicating their probable locations in the membrane. Focusing on predicted transmembrane domains, it can be pointed out that 27 proteins were identified in the hydrophobic gel containing up to 11 transmembrane domains; in the classical gel, only 5 proteins containing 1 transmembrane domain were successfully identified. For example, multiple ionic channels and receptors were characterized in the hydrophobic gel such as the sodium/potassium channel and the glutamate or the transferrin receptors whereas they are traditionally detected using specific enrichment techniques such as immunoprecipitation. In total, membrane proteins identified in the classical gel are well documented in the literature, while most of the membrane proteins only identified on the hydrophobic gel have rarely or never been described using a proteomic-based approach. 2010 Elsevier Inc. All rights reserved.

Thermally responsive silicon nanowire arrays for native/denatured-protein separation

International Nuclear Information System (INIS)

Wang Hongwei; Wang Yanwei; Yuan Lin; Wang Lei; Yang Weikang; Wu Zhaoqiang; Li Dan; Chen Hong

2013-01-01

We present our findings of the selective adsorption of native and denatured proteins onto thermally responsive, native-protein resistant poly(N-isopropylacrylamide) (PNIPAAm) decorated silicon nanowire arrays (SiNWAs). The PNIPAAm–SiNWAs surface, which shows very low levels of native-protein adsorption, favors the adsorption of denatured proteins. The amount of denatured-protein adsorption is higher at temperatures above the lower critical solution temperature (LCST) of PNIPAAm. Temperature cycling surrounding the LCST, which ensures against thermal denaturation of native proteins, further increases the amount of denatured-protein adsorption. Moreover, the PNIPAAm–SiNWAs surface is able to selectively adsorb denatured protein even from mixtures of different protein species; meanwhile, the amount of native proteins in solution is kept nearly at its original level. It is believed that these results will not only enrich current understanding of protein interactions with PNIPAAm-modified SiNWAs surfaces, but may also stimulate applications of PNIPAAm–SiNWAs surfaces for native/denatured protein separation. (paper)

Determination and optimization of the ζ potential in boron electrophoretic deposition on aluminium substrates

International Nuclear Information System (INIS)

Oliveira Sampa, M.H. de; Vinhas, L.A.; Pino, E.S.

1991-05-01

In this work we present an introduction of the electrophoretic process followed by a detailed experimental treatment of the technique used in the determination and optimization of the ζ-potential, mainly as a function of the electrolyte concentration, in a high purity boron electrophoretics deposition on aluminium substrates used as electrodes in neutron detectors. (author)

Tyrosinase inhibitor screening in traditional Chinese medicines by electrophoretically mediated microanalysis.

Science.gov (United States)

Tang, Lilin; Zhang, Wenpeng; Zhao, Haiyan; Chen, Zilin

2015-08-01

A capillary-electrophoresis-based method for the screening of tyrosinase inhibitors in traditional Chinese medicines was developed. The method integrated electrophoretically mediated microanalysis with sandwich mode injection, partial filling, and rapid polarity switching techniques, and carried out on-column enzyme reaction and the separation of substrate and product. The conditions were optimized including the background electrolyte, mixing voltage, and the incubation time. Finally, the screening of nine standard natural compounds of traditional Chinese medicines was carried out. The inhibitors can be directly identified from the reduced peak area of the product compared to that obtained without any inhibitor. Chlorogenic acid (100 μM) showed inhibitory activity with the inhibitory percentage of 19.8%, while the other compounds showed no inhibitory activity. This method has great application potential in drug discovery from traditional Chinese medicines. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Electrophoretic variation in low molecular weight lens crystallins from inbred strains of rats.

Science.gov (United States)

Donner, M E; Skow, L C; Kunz, H W; Gill, T J

1985-10-01

Analysis of rat lens soluble proteins by analytical isoelectric focusing detected two inherited electrophoretic differences in low molecular weight (LM) crystallins from inbred strains of rats (Rattus norvegicus). The polymorphic lens crystallins were shown to be similar to a genetically variant LM crystallin, LEN-1, previously described in mice (Mus musculus) and encoded on chromosome 1, at a locus linked to Pep-3 (dipeptidase). Linkage analysis demonstrated that the rat crystallin locus was loosely linked to Pep-3 at a recombination distance of 38 +/- 4.5 U. These data suggest the conservation of a large chromosomal region during the evolution of Rodentia and support the hypothesis that the gamma-crystallins are evolving more rapidly than alpha- or beta-crystallins.

[Comparative investigation of the non-histone proteins of chromatin from pigeon erythroblasts and erythrocytes].

Science.gov (United States)

Fedina, A B; Gazarian, G G

1976-01-01

Chromosomal non-histone proteins are obtained from nuclei of two types of pigeon erythroid cells: erythroblasts (cells active in RNA synthesis) and erythrocytes (cells with repressed RNA synthesis). They are well soluble in solutions of low ionic strength. Electrophoretic separation of the obtained non-histone proteins in polyacrylamide gels with urea and SDS shows the presence of qualitative differences in the pattern of non-histone proteins of chromatine from erythroblasts and erythrocytes. By electrophoresis in urea some protein bands of non-histone proteins of chromatine from erythroblasts were found which disappear with the aging of cells. At the same time two protein fractions were observed in chromatine from erythrocytes which were absent in that of erythroblasts. Disappearance of some high molecular weight protein fractions from erythrocyte chromatine as compared to erythroblasts was observed by separation of the non-histone proteins in the presence of SDS. These fractions of the non-histone proteins disappearing during aging of cells are well extractable from erythroblast chromatine by 0.35 M NaCl solution. In the in vitro system with E. coli RNA polymerase addition of non-histone proteins of chromatine from erythroblasts to chromatine from erythrocytes increases RNA synthesis 2--3 times. At the same time addition of non-histone proteins from erythrocytes is either without any influence on this process or somewhat inhibiting.

Cyclic AMP-receptor proteins in heart muscle of rats flown on Cosmos 1887

Science.gov (United States)

Mednieks, Maija I.; Popova, Irina A.; Grindeland, Richard E.

1991-01-01

The cellular compartmentalization of the cyclic AMP-receptor proteins in heart ventricular tissue obtained from rats flown on the Cosmos 1887 is determined. Photoaffinity labeling of soluble and particular cell fractions with a (32P)-8-azido analog of cyclic AMP is followed by electrophoretic separation of the proteins and by autoradiographic identification of the labeled isoforms of cAPK R subunits. It is shown that RII in the particulate subcellular fraction was significantly decreased in heart cells from rats in the flight group when compared to controls. Protein banding patterns in both the cytoplasmic fraction and in a fraction enriched in chromatin-bound proteins exhibited some variability in tissues of individual animals, but showed no changes that could be directly attributed to flight conditions. No significant change was apparent in the distribution of RI or RII cyclic AMP binding in the soluble fractions. It is inferred that the cardiac cell integrity or its protein content is not compromised under flight conditions.

Spatial separation and bidirectional trafficking of proteins using a multi-functional reporter

Directory of Open Access Journals (Sweden)

Klaubert Dieter H

2008-04-01

Full Text Available Abstract Background The ability to specifically label proteins within living cells can provide information about their dynamics and function. To study a membrane protein, we fused a multi-functional reporter protein, HaloTag®, to the extracellular domain of a truncated integrin. Results Using the HaloTag technology, we could study the localization, trafficking and processing of an integrin-HaloTag fusion, which we showed had cellular dynamics consistent with native integrins. By labeling live cells with different fluorescent impermeable and permeable ligands, we showed spatial separation of plasma membrane and internal pools of the integrin-HaloTag fusion, and followed these protein pools over time to study bi-directional trafficking. In addition to combining the HaloTag reporter protein with different fluorophores, we also employed an affinity tag to achieve cell capture. Conclusion The HaloTag technology was used successfully to study expression, trafficking, spatial separation and real-time translocation of an integrin-HaloTag fusion, thereby demonstrating that this technology can be a powerful tool to investigate membrane protein biology in live cells.

21 CFR 864.7440 - Electrophoretic hemoglobin analysis system.

Science.gov (United States)

2010-04-01

... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Electrophoretic hemoglobin analysis system. 864.7440 Section 864.7440 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES HEMATOLOGY AND PATHOLOGY DEVICES Hematology Kits and Packages § 864...

Current Developments in Electrophoretic and Chromatographic Separation Methods on Microfabricated Devices

DEFF Research Database (Denmark)

Kutter, Jörg Peter

2000-01-01

Microchip-based separation techniques are essential elements in the development of fully integrated micro-total analysis systems, which are envisioned to become powerful instruments for obtaining and assessing analytical data in research, industry, and everyday life. This article's goal is to give...

Template-based electrophoretic deposition of perovskite PZT nanotubes

Energy Technology Data Exchange (ETDEWEB)

Nourmohammadi, A. [Solid Surfaces Analysis and Electron Microscopy Group, Institute of Physics, Chemnitz University of Technology, D-09126 Chemnitz (Germany); Semiconductors Department, Materials and Energy Research Center (MERC), 31779-83634 Karaj (Iran, Islamic Republic of); Bahrevar, M.A. [Semiconductors Department, Materials and Energy Research Center (MERC), 31779-83634 Karaj (Iran, Islamic Republic of)], E-mail: ma.bahrevar@yahoo.com; Hietschold, M. [Solid Surfaces Analysis and Electron Microscopy Group, Institute of Physics, Chemnitz University of Technology, D-09126 Chemnitz (Germany)

2009-04-03

Template-based electrophoretic deposition of perovskite lead zirconate titanate (PZT) nanotubes was achieved using anodic alumina (AA) membranes and sols, containing lead, zirconium and titanium precursors. The effect of various anodizing voltages on the size of the channels in the anodic alumina template was investigated. The prepared sol was driven into the channels under the influence of various electric fields and subsequently sintered at about 700 deg. C. The effects of the initial heating rates and the burn-out temperature on the phase evolution of the samples were studied and a modified firing process was employed. The effects of the electrophoretic voltage and the deposition time on the average wall thickness of the tubes were investigated. Scanning and transmission electron microscopy (SEM and TEM) revealed the efficiency of electrophoresis in the growth of lead zirconate titanate nanotubes in a close-packed array. The X-ray diffraction analyses indicated the presence of perovskite as the principal phase after a modified firing schedule.

On-line stacking techniques for the nonaqueous capillary electrophoretic determination of acrylamide in processed food

International Nuclear Information System (INIS)

Tezcan, Filiz; Erim, F. Bedia

2008-01-01

In the present study, field amplified sample stacking (FASS) techniques in the nonaqueous capillary electrophoresis method (NACE) were introduced for the on-line concentration of the acrylamide to improve acrylamide detection at 210 nm by diode-array detection. Acetonitrile (ACN) as a nonaqueous solvent permits acrylamide to be protonated through the change of its acid-base chemistry, allowing capillary electrophoretic separation of this compound. Choosing 30 mmol L -1 HClO 4 , 20 mmol L -1 NaClO 4 , 218 mmol L -1 CH 3 COOH in ACN as the separation electrolyte and employing sample stacking methods, the LOD value of acrylamide was decreased to 2.6 ng mL -1 with electrokinetic injection and 4.4 ng mL -1 with hydrodynamic injection. Optimized stacking conditions were applied to the determination of acrylamide in several foodstuffs. The method is simple, rapid, inexpensive, and widely applicable for the determination of acrylamide in food samples

Free flow electrophoresis separation and AMS quantitation of {sup 14}C-naphthalene-protein adducts

Energy Technology Data Exchange (ETDEWEB)

Buchholz, Bruce A., E-mail: bbuchholz@llnl.go [Center for AMS, LLNL, 7000 East Avenue, Livermore, CA 94551 (United States); Haack, Kurt W.; Sporty, Jennifer L. [Center for AMS, LLNL, 7000 East Avenue, Livermore, CA 94551 (United States); Buckpitt, Alan R.; Morin, Dexter [Department of Molecular Biosciences, School of Veterinary Medicine, UC Davis, Davis, CA 95616 (United States)

2010-04-15

Naphthalene is a volatile aromatic hydrocarbon to which humans are exposed from a variety of sources including mobile air sources and cigarette smoke. Naphthalene produces dose-(concentration)dependent injury to airway epithelial cells of murine lung which is observed at concentrations well below the current occupational exposure standard. Toxicity is dependent upon the cytochrome P450 mediated metabolic activation of the parent substrate to unstable metabolites which become bound covalently to tissue proteins. Nearly 70 proteins have been identified as forming adducts with reactive naphthalene metabolites using in vitro systems but very little work has been conducted in vivo because reasonably large amounts (100 muCi) of {sup 14}C labeled parent compound must be administered to generate detectable adduct levels on storage phosphor screens following separation of labeled proteins by 2D gel electrophoresis. The work described here was done to provide proof of concept that protein separation by free flow electrophoresis followed by AMS detection of protein fractions containing protein bound reactive metabolites would provide adducted protein profiles in animals dosed with trace quantities of labeled naphthalene. Mice were administered 200 mg/kg naphthalene intraperitoneally at a calculated specific activity of 2 DPM/nmol (1 pCi/nmol) and respiratory epithelial tissue was obtained by lysis lavage 4 h post injection. Free flow electrophoresis (FFE) separates proteins in the liquid phase over a large pH range (2.5-11.5) using low molecular weight acids and bases to modify the pH. The apparatus separates fractions into standard 96-well plates that can be used in other protein analysis techniques. The buffers of the fractions have very high carbon content, however, and need to be dialyzed to yield buffers compatible with {sup 14}C-AMS. We describe the processing techniques required to couple FFE to AMS for quantitation of protein adducts.

Application of design of experiment on electrophoretic deposition of ...

Indian Academy of Sciences (India)

Unknown

Keywords. Coating; electrophoretic deposition; glass-ceramic; design of experiment. 1. Introduction ... other chemicals used were of laboratory reagent grade. ... changes from 7⋅0 to 9⋅5 that adversely affects the deposi- tion efficiency and ...

Signal amelioration of electrophoretically deposited whole-cell biosensors using external electric fields

Energy Technology Data Exchange (ETDEWEB)

Ben-Yoav, Hadar, E-mail: benyoav@post.tau.ac.il [Department of Physical Electronics, School of Electrical Engineering, Faculty of Engineering, Tel Aviv University, Tel-Aviv 69978 (Israel); Amzel, Tal [Department of Physical Electronics, School of Electrical Engineering, Faculty of Engineering, Tel Aviv University, Tel-Aviv 69978 (Israel); Sternheim, Marek [Center for Nanoscience and Nanotechnology, Tel Aviv University, Tel-Aviv, 69978 (Israel); Belkin, Shimshon [Institute of Life Sciences, Hebrew University of Jerusalem, Jerusalem, 91904 (Israel); Rubin, Adi [Department of Molecular Microbiology and Biotechnology, Faculty of Life Sciences, Tel Aviv University, Tel-Aviv, 69978 (Israel); Shacham-Diamand, Yosi [Department of Physical Electronics, School of Electrical Engineering, Faculty of Engineering, Tel Aviv University, Tel-Aviv 69978 (Israel); Freeman, Amihay [Center for Nanoscience and Nanotechnology, Tel Aviv University, Tel-Aviv, 69978 (Israel)

2011-11-01

Highlights: > We present an electrochemical whole-cell biochip that can apply electric fields. > We examine the integration of cells on a biochip using electrophoretic deposition. > The effect of electric fields on the whole-cell biosensor has been demonstrated. > Relatively short DC electric pulse improves the performance of whole-cell biosensors. > Prolonged AC electric fields deteriorated the whole-cell biosensor performance. - Abstract: This paper presents an integrated whole-cell biochip system where functioning cells are deposited on the solid micro-machined surfaces while specially designed indium tin oxide electrodes that can be used to apply controllable electric fields during various stages; for example during cell deposition. The electrodes can be used also for sensing currents associated with the sensing mechanisms of electrochemical whole-cell biosensors. In this work a new approach integrating live bacterial cells on a biochip using electrophoretic deposition is presented. The biomaterial deposition technique was characterized under various driving potentials and chamber configurations. An analytical model of the electrophoretic deposition kinetics was developed and presented here. The deposited biomass included genetically engineered bacterial cells that may respond to toxic material exposure by expressing proteins that react with specific analytes generating electrochemically active byproducts. In this study the effect of external electric fields on the whole-cell biochips has been successfully developed and tested. The research hypothesis was that by applying electric fields on bacterial whole-cells, their permeability to the penetration of external analytes can be increased. This effect was tested and the results are shown here. The effect of prolonged and short external electric fields on the bioelectrochemical signal generated by sessile bacterial whole-cells in response to the presence of toxins was studied. It was demonstrated that relatively

Signal amelioration of electrophoretically deposited whole-cell biosensors using external electric fields

International Nuclear Information System (INIS)

Ben-Yoav, Hadar; Amzel, Tal; Sternheim, Marek; Belkin, Shimshon; Rubin, Adi; Shacham-Diamand, Yosi; Freeman, Amihay

2011-01-01

Highlights: → We present an electrochemical whole-cell biochip that can apply electric fields. → We examine the integration of cells on a biochip using electrophoretic deposition. → The effect of electric fields on the whole-cell biosensor has been demonstrated. → Relatively short DC electric pulse improves the performance of whole-cell biosensors. → Prolonged AC electric fields deteriorated the whole-cell biosensor performance. - Abstract: This paper presents an integrated whole-cell biochip system where functioning cells are deposited on the solid micro-machined surfaces while specially designed indium tin oxide electrodes that can be used to apply controllable electric fields during various stages; for example during cell deposition. The electrodes can be used also for sensing currents associated with the sensing mechanisms of electrochemical whole-cell biosensors. In this work a new approach integrating live bacterial cells on a biochip using electrophoretic deposition is presented. The biomaterial deposition technique was characterized under various driving potentials and chamber configurations. An analytical model of the electrophoretic deposition kinetics was developed and presented here. The deposited biomass included genetically engineered bacterial cells that may respond to toxic material exposure by expressing proteins that react with specific analytes generating electrochemically active byproducts. In this study the effect of external electric fields on the whole-cell biochips has been successfully developed and tested. The research hypothesis was that by applying electric fields on bacterial whole-cells, their permeability to the penetration of external analytes can be increased. This effect was tested and the results are shown here. The effect of prolonged and short external electric fields on the bioelectrochemical signal generated by sessile bacterial whole-cells in response to the presence of toxins was studied. It was demonstrated that

Properties of electrophoretically deposited single wall carbon nanotube films

International Nuclear Information System (INIS)

Lim, Junyoung; Jalali, Maryam; Campbell, Stephen A.

2015-01-01

This paper describes techniques for rapidly producing a carbon nanotube thin film by electrophoretic deposition at room temperature and determines the film mass density and electrical/mechanical properties of such films. The mechanism of electrophoretic deposition of thin layers is explained with experimental data. Also, film thickness is measured as a function of time, electrical field and suspension concentration. We use Rutherford backscattering spectroscopy to determine the film mass density. Films created in this manner have a resistivity of 2.14 × 10 −3 Ω·cm, a mass density that varies with thickness from 0.12 to 0.54 g/cm 3 , and a Young's modulus between 4.72 and 5.67 GPa. The latter was found to be independent of thickness from 77 to 134 nm. We also report on fabricating free-standing films by removing the metal seed layer under the CNT film, and selectively etching a sacrificial layer. This method could be extended to flexible photovoltaic devices or high frequency RF MEMS devices. - Highlights: • We explain the electrophoretic deposition process and mechanism of thin SWCNT film deposition. • Characterization of the SWCNT film properties including density, resistivity, transmittance, and Young's modulus. • The film density and resistivity are found to be a function of the film thickness. • Techniques developed to create free standing layers of SW-CNTs for flexible electronics and mechanical actuators

Electrophoretic characterization of crude leaf proteins in ...

African Journals Online (AJOL)

Administrator

ground in an eppendorf tube with 100 µl of lysis buffer. The mixtures were allowed to settle inside the eppendorf immersed in an ice bath for 1 h, and the supernatants were fractionated by. 7.5% SDS-PAGE (Laemmili, 1970). RESULTS AND DISCUSSION. Protein distribution patterns in three cultivars of. Lycopersicon and ...

The Electrophoretic Mobility of Proteins near Surfaces

Science.gov (United States)

Ramasamy, Perumal; Singh, Avtar; Rafailovich, Miriam; Sokolov, Jonathan

2004-03-01

We have attempted to apply the methods developed for surface DNA electrophoresis (1) for proteomics. Droplets of FITC stained Abumin, Poly- L-Lysine, or Casein purchased from Sigma were deposited on glass cover slips. The droplets were then place in contact with a TBE buffer solution contained in a cell molded from PDMS. Pt electrodes were inserted into the cell and a voltage was a applied. The motion of the protein was then imaged with a Leica Confocal microscope as a function of buffer concentration, distance from the surface, and applied voltage. The mobilities were then compared with those of uncharged one micron florescent Polystyrene beads. References: 1)Henzel WJ, Watanabe C, Stults JT., !0 Protein Identification: The Origins of Peptide Mass Fingerprinting. !1 J. American Society for Mass Spectrometry. 14 (September 2003): 931-942 2)Mathesius U, Imin N, Natera SH, Rolfe BG., !0 Proteomics as a functional genomics tool. !1 Methods of Molecular Biology 236: 395-414. *Work supported in part by the NSF-MRSEC program

Validação em métodos cromatográficos e eletroforéticos Validation for chromatographic and electrophoretic methods

Directory of Open Access Journals (Sweden)

Marcelo Ribani

2004-10-01

Full Text Available The validation of an analytical method is fundamental to implementing a quality control system in any analytical laboratory. As the separation techniques, GC, HPLC and CE, are often the principal tools used in such determinations, procedure validation is a necessity. The objective of this review is to describe the main aspects of validation in chromatographic and electrophoretic analysis, showing, in a general way, the similarities and differences between the guidelines established by the different Brazilian and international regulatory agencies.

Separation of saccharides derivatized with 2-aminobenzoic acid by capillary electrophoresis and their structural consideration by nuclear magnetic resonance.

Science.gov (United States)

He, Liping; Sato, Kae; Abo, Mitsuru; Okubo, Akira; Yamazaki, Sunao

2003-03-01

Saccharides including mono- and disaccharides were quantitatively derivatized with 2-aminobenzoic acid (2-AA). These derivatives were then separated by capillary zone electrophoresis with UV detection using 50mM sodium phosphate buffer as the running electrolyte solution. In particular, the saccharide derivatives with the same molecular weight as 2-AA aldohexoses (mannose and glucose) and 2-AA aldopentoses (ribose and xylose) were well separated. The underlying reasons for separation were explored by studying their structural data using 1H and 13C NMR. It was found that the configurational difference between their hydroxyl group at C2 or C3 could cause the difference in Stokes' radii between their molecules and thus lead to different electrophoretic mobilities. The correlation between the electrophoretic behavior of these carbohydrate derivatives and their structures was studied utilizing the calculated molecular models of the 2-AA-labeled mannose, glucose, ribose, and xylose.

A Student-Made Microfluidic Device for Electrophoretic Separation of Food Dyes

Science.gov (United States)

Teerasong, Saowapak; McClain, Robert L.

2011-01-01

We have developed an undergraduate laboratory activity to introduce students to microfluidics. In the activity, each student constructs their own microfluidic device using simple photolithographic techniques and then uses the device to separate a food dye mixture by electrophoresis. Dyes are used so that students are able to visually observe the…

Electrophoretic deposition of zinc-substituted hydroxyapatite coatings.

Science.gov (United States)

Sun, Guangfei; Ma, Jun; Zhang, Shengmin

2014-06-01

Zinc-substituted hydroxyapatite nanoparticles synthesized by the co-precipitation method were used to coat stainless steel plates by electrophoretic deposition in n-butanol with triethanolamine as a dispersant. The effect of zinc concentration in the synthesis on the morphology and microstructure of coatings was investigated. It is found that the deposition current densities significantly increase with the increasing zinc concentration. The zinc-substituted hydroxyapatite coatings were analyzed by X-ray diffraction, scanning electron microscopy and Fourier transform infrared spectroscopy. It is inferred that hydroxyapatite and triethanolamine predominate in the chemical composition of coatings. With the increasing Zn/Ca ratios, the contents of triethanolamine decrease in the final products. The triethanolamine can be burnt out by heat treatment. The tests of adhesive strength have confirmed good adhesion between the coatings and substrates. The formation of new apatite layer on the coatings has been observed after 7days of immersion in a simulated body fluid. In summary, the results show that dense, uniform zinc-substituted hydroxyapatite coatings are obtained by electrophoretic deposition when the Zn/Ca ratio reaches 5%. Copyright © 2014 Elsevier B.V. All rights reserved.

Lysine-Rich Proteins in High-Lysine Hordeum Vulgare Grain

DEFF Research Database (Denmark)

Ingversen, J.; Køie, B.

1973-01-01

The salt-soluble proteins in barley grain selected for high-lysine content (Hiproly, CI 7115 and the mutants 29 and 86) and of a control (Carlsberg II) with normal lysine content, contain identical major proteins as determined by MW and electrophoretic mobility. The concentration of a protein gro...

Recycling isoelectric focusing with computer controlled data acquisition system. [for high resolution electrophoretic separation and purification of biomolecules

Science.gov (United States)

Egen, N. B.; Twitty, G. E.; Bier, M.

1979-01-01

Isoelectric focusing is a high-resolution technique for separating and purifying large peptides, proteins, and other biomolecules. The apparatus described in the present paper constitutes a new approach to fluid stabilization and increased throughput. Stabilization is achieved by flowing the process fluid uniformly through an array of closely spaced filter elements oriented parallel both to the electrodes and the direction of the flow. This seems to overcome the major difficulties of parabolic flow and electroosmosis at the walls, while limiting the convection to chamber compartments defined by adjacent spacers. Increased throughput is achieved by recirculating the process fluid through external heat exchange reservoirs, where the Joule heat is dissipated.

Validation of an electrophoretic method to detect albuminuria in cats.

Science.gov (United States)

Ferlizza, Enea; Dondi, Francesco; Andreani, Giulia; Bucci, Diego; Archer, Joy; Isani, Gloria

2017-08-01

Objectives The aims of this study were to validate a semi-automated high-resolution electrophoretic technique to quantify urinary albumin in healthy and diseased cats, and to evaluate its diagnostic performance in cases of proteinuria and renal diseases. Methods Urine samples were collected from 88 cats (healthy; chronic kidney disease [CKD]; lower urinary tract disease [LUTD]; non-urinary tract diseases [OTHER]). Urine samples were routinely analysed and high-resolution electrophoresis (HRE) was performed. Within-assay and between-assay variability, linearity, accuracy, recovery and the lowest detectable and quantifiable bands were calculated. Receiver operating curve (ROC) analysis was also performed. Results All coefficients of variation were HRE allowed the visualisation of a faint band of albumin and a diffused band between alpha and beta zones in healthy cats, while profiles from diseased cats were variable. Albumin (mg/dl) and urine albumin:creatinine ratio (UAC) were significantly ( P HRE is an accurate and precise method that could be used to measure albuminuria in cats. UAC was useful to correctly classify proteinuria and to discriminate between healthy and diseased cats. HRE might also provide additional information on urine proteins with a profile of all proteins (albumin and globulins) to aid clinicians in the diagnosis of diseases characterised by proteinuria.

A stable and convenient protein electrophoresis titration device with bubble removing system.

Science.gov (United States)

Zhang, Qiang; Fan, Liu-Yin; Li, Wen-Lin; Cong, Feng-Song; Zhong, Ran; Chen, Jing-Jing; He, Yu-Chen; Xiao, Hua; Cao, Cheng-Xi

2017-07-01

Moving reaction boundary titration (MRBT) has a potential application to immunoassay and protein content analysis with high selectivity. However, air bubbles often impair the accuracy of MRBT, and the leakage of electrolyte greatly decreases the safety and convenience of electrophoretic titration. Addressing these two issues a reliable MRBT device with modified electrolyte chamber of protein titration was designed. Multiphysics computer simulation was conducted for optimization according to two-phase flow. The single chamber was made of two perpendicular cylinders with different diameters. After placing electrophoretic tube, the resident air in the junction next to the gel could be eliminated by a simple fast electrolyte flow. Removing the electrophoretic tube automatically prevented electrolyte leakage at the junction due to the gravity-induced negative pressure within the chamber. Moreover, the numerical simulation and experiments showed that the improved MRBT device has following advantages: (i) easy and rapid setup of electrophoretic tube within 20 s; (ii) simple and quick bubble dissipates from the chamber of titration within 2 s; (iii) no electrolyte leakage from the two chambers: and (iv) accurate protein titration and safe instrumental operation. The developed technique and apparatus greatly improves the performance of the previous MRBT device, and providing a new route toward practical application. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Survival rate of eukaryotic cells following electrophoretic nanoinjection.

Science.gov (United States)

Simonis, Matthias; Hübner, Wolfgang; Wilking, Alice; Huser, Thomas; Hennig, Simon

2017-01-25

Insertion of foreign molecules such as functionalized fluorescent probes, antibodies, or plasmid DNA to living cells requires overcoming the plasma membrane barrier without harming the cell during the staining process. Many techniques such as electroporation, lipofection or microinjection have been developed to overcome the cellular plasma membrane, but they all result in reduced cell viability. A novel approach is the injection of cells with a nanopipette and using electrophoretic forces for the delivery of molecules. The tip size of these pipettes is approximately ten times smaller than typical microinjection pipettes and rather than pressure pulses as delivery method, moderate DC electric fields are used to drive charged molecules out of the tip. Here, we show that this approach leads to a significantly higher survival rate of nanoinjected cells and that injection with nanopipettes has a significantly lower impact on the proliferation behavior of injected cells. Thus, we propose that injection with nanopipettes using electrophoretic delivery is an excellent alternative when working with valuable and rare living cells, such as primary cells or stem cells.

Survival rate of eukaryotic cells following electrophoretic nanoinjection

Science.gov (United States)

Simonis, Matthias; Hübner, Wolfgang; Wilking, Alice; Huser, Thomas; Hennig, Simon

2017-01-01

Insertion of foreign molecules such as functionalized fluorescent probes, antibodies, or plasmid DNA to living cells requires overcoming the plasma membrane barrier without harming the cell during the staining process. Many techniques such as electroporation, lipofection or microinjection have been developed to overcome the cellular plasma membrane, but they all result in reduced cell viability. A novel approach is the injection of cells with a nanopipette and using electrophoretic forces for the delivery of molecules. The tip size of these pipettes is approximately ten times smaller than typical microinjection pipettes and rather than pressure pulses as delivery method, moderate DC electric fields are used to drive charged molecules out of the tip. Here, we show that this approach leads to a significantly higher survival rate of nanoinjected cells and that injection with nanopipettes has a significantly lower impact on the proliferation behavior of injected cells. Thus, we propose that injection with nanopipettes using electrophoretic delivery is an excellent alternative when working with valuable and rare living cells, such as primary cells or stem cells. PMID:28120926

Association of electrophoretic karyotype of Candida stellatoidea with virulence for mice

International Nuclear Information System (INIS)

Kwon-Chung, K.J.; Wickes, B.L.; Merz, W.G.

1988-01-01

Seven isolates of Candida stellatoidea were studied for their electrophoretic karyotype, virulence for mice, sensitivity to UV radiation, growth rate in vitro, reaction on cycloheximide-indicator medium, and proteinase activity. The isolates exhibited one of two distinct electrophoretic karyotypes as determined by orthogonal field alternating gel electrophoresis (OFAGE). Four isolates, including the type culture of C. stellatoidea, belonged to electrophoretic karyotype type I by OFAGE, showing eight to nine bands of which at least two bands were less than 1,000 kilobases in size as estimated by comparison with the DNA bands of Saccharomyces cerevisiae. These isolates failed to produce fatal infection in mice within 20 days when 5 X 10(5) cells were injected intravenously. The yeasts were cleared from the kidneys of two of three mice tested by day 30. Type I showed proteinase activity on bovine serum albumin agar at pH 3.8 and produced a negative reaction on cycloheximide-bromcresol green medium within 48 h. The three grouped in type II by OFAGE showed banding patterns similar to those of a well-characterized isolate of Candida albicans. The isolates of type II had an electrophoretic karyotype of six to seven bands approximately 1,200 kilobases or greater in size. All three type II isolates were highly virulent for mice, producing fatality curves similar to those of a previously studied C. albicans isolate. From 80 to 90% of the mice injected with 5 X 10(5) cells intravenously died within 20 days. The type II isolates produced a positive reaction on cycloheximide-bromcresol green agar and showed no proteinase activity on bovine serum albumin agar at the low pH. In addition, the type II isolates grew faster and were significantly more resistant to UV irradiation than the type I isolates

Using electrophoretic mobility shift assays to measure equilibrium dissociation constants: GAL4-p53 binding DNA as a model system.

Science.gov (United States)

Heffler, Michael A; Walters, Ryan D; Kugel, Jennifer F

2012-01-01

An undergraduate biochemistry laboratory experiment is described that will teach students the practical and theoretical considerations for measuring the equilibrium dissociation constant (K(D) ) for a protein/DNA interaction using electrophoretic mobility shift assays (EMSAs). An EMSA monitors the migration of DNA through a native gel; the DNA migrates more slowly when bound to a protein. To determine a K(D) the amount of unbound and protein-bound DNA in the gel is measured as the protein concentration increases. By performing this experiment, students will be introduced to making affinity measurements and gain experience in performing quantitative EMSAs. The experiment describes measuring the K(D) for the interaction between the chimeric protein GAL4-p53 and its DNA recognition site; however, the techniques are adaptable to other DNA binding proteins. In addition, the basic experiment described can be easily expanded to include additional inquiry-driven experimentation. © 2012 by The International Union of Biochemistry and Molecular Biology. Copyright © 2012 Wiley Periodicals, Inc.

Stabilization of green bodies via sacrificial gelling agent during electrophoretic deposition

Energy Technology Data Exchange (ETDEWEB)

Worsley, Marcus A.; Kuntz, Joshua D.; Rose, Klint A.

2016-03-22

In one embodiment, a method for electrophoretic deposition of a three-dimensionally patterned green body includes suspending a first material in a gelling agent above a patterned electrode of an electrophoretic deposition (EPD) chamber, and gelling the suspension while applying a first electric field to the suspension to cause desired patterning of the first material in a resulting gelation. In another embodiment, a ceramic, metal, or cermet includes a plurality of layers, wherein each layer includes a gradient in composition, microstructure, and/or density in an x-y plane oriented parallel to a plane of deposition of the plurality of layers along a predetermined distance in a z-direction perpendicular to the plane of deposition.

Buffalo milk: proteins electrophoretic profile and somatic cell count

Directory of Open Access Journals (Sweden)

S. Mattii

2011-03-01

Full Text Available Water buffalo milk differs from the cow’s milk for greater fat and protein content, very important features in cheese making. Proteins, casein and whey-proteins in particular, are the most important factors determining cheese yield. Several previous research discussed the rule of SCC in cow milk production (Varisco, 1999 and the close relationship existing between cow’s milk cheese yield and somatic cell count (Barbano, 2000. In particular the inverse correlation between cheese yields and somatic cells’content have been demonstrated. In Italy the regulation in force DPR 54/97 acknowledges what expressed in EEC 46/92 Directive (Tripodi, 1999 without fixing the limit threshold of somatic cells for buffalo’s milk....

A new method of high-speed cellular protein separation and insight into subcellular compartmentalization of proteins.

Science.gov (United States)

Png, Evelyn; Lan, WanWen; Lazaroo, Melisa; Chen, Silin; Zhou, Lei; Tong, Louis

2011-05-01

Transglutaminase (TGM)-2 is a ubiquitous protein with important cellular functions such as regulation of cytoskeleton, cell adhesion, apoptosis, energy metabolism, and stress signaling. We identified several proteins that may interact with TGM-2 through a discovery-based proteomics method via pull down of flag-tagged TGM-2 peptide fragments. The distribution of these potential binding partners of TGM-2 was studied in subcellular fractions separated by density using novel high-speed centricollation technology. Centricollation is a compressed air-driven, low-temperature stepwise ultracentrifugation procedure where low extraction volumes can be processed in a relatively short time in non-denaturing separation conditions with high recovery yield. The fractions were characterized by immunoblots against known organelle markers. The changes in the concentrations of the binding partners were studied in cells expressing short hairpin RNA against TGM-2 (shTG). Desmin, mitochondrial intramembrane cleaving protease (PARL), protein tyrosine kinase (NTRK3), and serine protease (PRSS3) were found to be less concentrated in the 8.5%, 10%, 15%, and 20% sucrose fractions (SFs) from the lysate of shTG cells. The Golgi-associated protein (GOLGA2) was predominantly localized in 15% SF fraction, and in shTG, this shifted to predominantly in the 8.5% SF and showed larger aggregations in the cytosol of cells on immunofluorescent staining compared to control. Based on the relative concentrations of these proteins, we propose how trafficking of such proteins between cellular compartments can occur to regulate cell function. Centricollation is useful for elucidating biological function at the molecular level, especially when combined with traditional cell biology techniques.

Accelerated identification of proteins by mass spectrometry by employing covalent pre-gel staining with Uniblue A.

Directory of Open Access Journals (Sweden)

Marco A Mata-Gómez

Full Text Available BACKGROUND: The identification of proteins by mass spectrometry is a standard method in biopharmaceutical quality control and biochemical research. Prior to identification by mass spectrometry, proteins are usually pre-separated by electrophoresis. However, current protein staining and de-staining protocols are tedious and time consuming, and therefore prolong the sample preparation time for mass spectrometry. METHODOLOGY AND PRINCIPAL FINDINGS: We developed a 1-minute covalent pre-gel staining protocol for proteins, which does not require de-staining before the mass spectrometry analysis. We investigated the electrophoretic properties of derivatized proteins and peptides and studied their behavior in mass spectrometry. Further, we elucidated the preferred reaction of proteins with Uniblue A and demonstrate the integration of the peptide derivatization into typical informatics tools. CONCLUSIONS AND SIGNIFICANCE: The Uniblue A staining method drastically speeds up the sample preparation for the mass spectrometry based identification of proteins. The application of this chemo-proteomic strategy will be advantageous for routine quality control of proteins and for time-critical tasks in protein analysis.

Thermodynamics of chiral selectivity in capillary electrophoresis : separation of ibuprofen enantiomers with beta-cyclodextrin

NARCIS (Netherlands)

Reijenga, J.C.; Ingelse, B.A.; Everaerts, F.M.

1997-01-01

The effect of temperature on the electrophoretic chiral separation of ibuprofen with ß-CD was investigated. Background electrolytes with sodium acetate or formate were chosen because of their constant pK within 0.03 units in the temperature range 25–50°C. Ibuprofen has a temperature independent pK

Monitoring protein phosphorylation by acrylamide pendant Phos-Tag™ in various plants

Directory of Open Access Journals (Sweden)

Slavka eBekesova

2015-05-01

Full Text Available The aim of the present study is to rationalize acrylamide pendant Phos-Tag™ in-gel discrimination of phosphorylated and non-phosphorylated plant protein species with standard immunoblot analysis, and optimize sample preparation, efficient electrophoretic separation and transfer. We tested variants of the method including extraction buffers suitable for preservation of phosphorylated protein species in crude extracts from plants and we addressed the importance of the cation (Mn2+ or Zn2+ used in the gel recipe for efficient transfer to PVDF membranes for further immunoblot analysis. We demonstrate the monitoring of Medicago sativa stress-induced mitogen activated protein kinase (SIMK in stress-treated wild type plants and transgenic SIMKK RNAi line. We further show the hyperosmotically-induced phosphorylation of the previously uncharacterized HvMPK4 of barley. The method is validated using inducible phosphorylation of barley and wheat α-tubulin and of Arabidopsis MPK6. Acrylamide pendant Phos-Tag™ offers a flexible tool for studying protein phosphorylation in crops and Arabidopsis circumventing radioactive labeling and the use of phosphorylation specific antibodies.

Cobalt ferrite nanoparticles with improved aqueous colloidal stability and electrophoretic mobility

International Nuclear Information System (INIS)

Munjal, Sandeep; Khare, Neeraj

2016-01-01

We have synthesized CoFe 2 O 4 (CFO) nanoparticles of size ∼ 12.2 nm by hydrothermal synthesis method. To control the size of these CFO nanoparticles, oleic acid was used as a surfactant. The inverse spinel phase of the synthesized nanoparticles was confirmed by X-ray diffraction method. As synthesized oleic acid coated CFO (OA@CFO) nanoparticles has very less electrophoretic mobility in the water and are not water dispersible. These OA@CFO nanoparticles were successfully turned into water soluble phase with a better colloidal aqueous stability, through a chemical treatment using citric acid. The modified citric acid coated CFO (CA@CFO) nanoparticles were dispersible in water and form a stable aqueous solution with high electrophoretic mobility.

A Two-Week Guided Inquiry Protein Separation and Detection Experiment for Undergraduate Biochemistry

Science.gov (United States)

Carolan, James P.; Nolta, Kathleen V.

2016-01-01

A laboratory experiment for teaching protein separation and detection in an undergraduate biochemistry laboratory course is described. This experiment, performed in two, 4 h laboratory periods, incorporates guided inquiry principles to introduce students to the concepts behind and difficulties of protein purification. After using size-exclusion…

Predicting tensorial electrophoretic effects in asymmetric colloids

Science.gov (United States)

Mowitz, Aaron J.; Witten, T. A.

2017-12-01

We formulate a numerical method for predicting the tensorial linear response of a rigid, asymmetrically charged body to an applied electric field. This prediction requires calculating the response of the fluid to the Stokes drag forces on the moving body and on the countercharges near its surface. To determine the fluid's motion, we represent both the body and the countercharges using many point sources of drag known as Stokeslets. Finding the correct flow field amounts to finding the set of drag forces on the Stokeslets that is consistent with the relative velocities experienced by each Stokeslet. The method rigorously satisfies the condition that the object moves with no transfer of momentum to the fluid. We demonstrate that a sphere represented by 1999 well-separated Stokeslets on its surface produces flow and drag force like a solid sphere to 1% accuracy. We show that a uniformly charged sphere with 3998 body and countercharge Stokeslets obeys the Smoluchowski prediction [F. Morrison, J. Colloid Interface Sci. 34, 210 (1970), 10.1016/0021-9797(70)90171-2] for electrophoretic mobility when the countercharges lie close to the sphere. Spheres with dipolar and quadrupolar charge distributions rotate and translate as predicted analytically to 4% accuracy or better. We describe how the method can treat general asymmetric shapes and charge distributions. This method offers promise as a way to characterize and manipulate asymmetrically charged colloid-scale objects from biology (e.g., viruses) and technology (e.g., self-assembled clusters).

Tridodecylamine, an efficient charge control agent in non-polar media for electrophoretic inks application

Science.gov (United States)

Noel, Amélie; Mirbel, Déborah; Cloutet, Eric; Fleury, Guillaume; Schatz, Christophe; Navarro, Christophe; Hadziioannou, Georges; CyrilBrochon

2018-01-01

In order to obtain efficient electrophoretic inks, Tridodecylamine (Dod3N), has been studied as charge control agent (CCA) in a non-polar paraffin solvent (Isopar G) for various inorganic pigments (TiO2 and Fe2O3). All hydrophobic mineral oxides, i.e. treated with octyltrimethoxysilane (C8) or dodecyltrimethoxysilane (C12), were found to be negatively charged in presence of Dod3N. The electrophoretic mobilities of inorganic pigments seemed to be strongly dependent of their isoelectric point (IEP) and also of the concentration of dod3N with an optimum range between 10 and 20 mM depending on the pigments. Finally, an electrophoretic ink constituted of hydrophobic mineral oxides in presence of Dod3N was tested in a device. Its efficiency as charge control agent to negatively charge hydrophobic particles was confirmed through good optical properties and fast response time (220 ms at 200 kV m-1).

Juvenile hormone-binding proteins of Melanoplus bivittatus identified by EFDA photoaffinity labeling

International Nuclear Information System (INIS)

Winder, B.S.

1988-01-01

Proteins that bind juvenile hormone in the hemolymph and fat body of the grasshopper, Melanoplus bivittatus were identified by photoaffinity labeling with radiolabeled epoxyfarnesyl diazoacetate ( 3 H-EFDA), and were characterized by electrophoretic analysis. A protocol was developed which allowed detection of 3 H-EFDA that was covalently linked to proteins upon exposure to ultraviolet light at 254 nm. Quantification of protein-linked 3 H-EFDA by liquid scintillation spectrometry took advantage of the differential solubility of unlinked 3 H-EFDA in toluene alone, and of the protein-linked 3 H-EFDA in toluene plus the detergent, Triton X-100. Competition between EFDA and juvenile hormone (JH) for binding to JH-specific binding sites was measured by hydroxyapatite protein binding assays in the presence of radiolabeled JH or EFDA and competing non-radiolabeled hormone. The protein-linked EFDA was detected on fluorograms of SDS or nondenaturing polyacrylamide gels (PAGE), and by liquid scintillation spectrometry of membranes to which the proteins had been electrophoretically transferred. Proteins which specifically bound JH were identified by photolabeling proteins in the presence and absence of nonlabeled JH-III

A Chip-Capillary Hybrid Device for Automated Transfer of Sample Pre-Separated by Capillary Isoelectric Focusing to Parallel Capillary Gel Electrophoresis for Two-Dimensional Protein Separation

Science.gov (United States)

Lu, Joann J.; Wang, Shili; Li, Guanbin; Wang, Wei; Pu, Qiaosheng; Liu, Shaorong

2012-01-01

In this report, we introduce a chip-capillary hybrid device to integrate capillary isoelectric focusing (CIEF) with parallel capillary sodium dodecyl sulfate – polyacrylamide gel electrophoresis (SDS-PAGE) or capillary gel electrophoresis (CGE) toward automating two-dimensional (2D) protein separations. The hybrid device consists of three chips that are butted together. The middle chip can be moved between two positions to re-route the fluidic paths, which enables the performance of CIEF and injection of proteins partially resolved by CIEF to CGE capillaries for parallel CGE separations in a continuous and automated fashion. Capillaries are attached to the other two chips to facilitate CIEF and CGE separations and to extend the effective lengths of CGE columns. Specifically, we illustrate the working principle of the hybrid device, develop protocols for producing and preparing the hybrid device, and demonstrate the feasibility of using this hybrid device for automated injection of CIEF-separated sample to parallel CGE for 2D protein separations. Potentials and problems associated with the hybrid device are also discussed. PMID:22830584

Magneto-paper electrophoresis in the separation of inorganic ions

International Nuclear Information System (INIS)

Mukherjee, H.G.; Datta, S.K.

1983-01-01

A comparative study of the separation of lanthanide ions by paper electrophoresis and magneto-paper electrophoresis is reported. The separation of La(III)-Gd(III), La(III)-Dy(III), Lu(III)-Gd(III), Lu(III)-Ho(III) etc. was achieved by magneto paper electrophoresis using 0.1M KCl as carrier electrolyte. Separation of different oxidation states of the same element like Cu(I)-Cu(II), Ce(III)-Ce(IV), Mn(CN) 6 3 - -Mn(CN) 6 4 - , Co(C 2 O 4 ) 2 2 - -Co(C 2 O 4 ) 3 3 - , V(CN) 6 3 - -VO(CN) 5 3 - , W(CN) 8 4 - -W(CN) 8 3 - and Ru(CN) 6 3 - Ru(CN) 6 4 - was also achieved by magneto paper electrophoretic technique using different carrier electrolytes. (Author)

Magnetic deep eutectic solvents molecularly imprinted polymers for the selective recognition and separation of protein

International Nuclear Information System (INIS)

Liu, Yanjin; Wang, Yuzhi; Dai, Qingzhou; Zhou, Yigang

2016-01-01

A novel and facile magnetic deep eutectic solvents (DES) molecularly imprinted polymers (MIPs) for the selective recognition and separation of Bovine hemoglobin (BHb) was prepared. The new-type DES was adopted as the functional monomer which would bring molecular imprinted technology to a new direction. The amounts of DES were optimized. The obtained magnetic DES-MIPs were characterized with fourier transform infrared spectrometry (FT-IR), thermogravimetric analysis (TGA), field emission scanning electron microscope (FESEM), dynamic light scattering (DLS), elemental analysis and vibrating sample magnetometer (VSM). The results suggested that the imprinted polymers were successfully formed and possessed a charming magnetism. The maximum adsorption capability (Q_m_a_x) and dissociation constant (K_L) were analyzed by Langmuir isotherms (R"2 = 0.9983) and the value were estimated to be 175.44 mg/g and 0.035 mg/mL for the imprinted particles. And the imprinted particles showed a high imprinting factor of 4.77. In addition, the magnetic DES-MIPs presented outstanding recognition specificity and selectivity so that it can be utilized to separate template protein from the mixture of proteins and real samples. Last but not least, the combination of deep eutectic solvents and molecular imprinted technology in this paper provides a new perspective for the recognition and separation of proteins. - Highlights: • Combined green deep eutectic solvents (DES) and molecular imprinted technology in recognition and separation of proteins. • DES was adopted as a new-type functional monomer. • The obtained magnetic DES-MIPs can separate proteins rapidly by an external magnetic field. • Adsorption and selectivity properties were discussed.

Magnetic deep eutectic solvents molecularly imprinted polymers for the selective recognition and separation of protein

Energy Technology Data Exchange (ETDEWEB)

Liu, Yanjin [State Key Laboratory of Chemo/Biosensing and Chemometrics, College of Chemistry and Chemical Engineering, Hunan University, Changsha, 410082 (China); Wang, Yuzhi, E-mail: wyzss@hnu.edu.cn [State Key Laboratory of Chemo/Biosensing and Chemometrics, College of Chemistry and Chemical Engineering, Hunan University, Changsha, 410082 (China); Dai, Qingzhou [State Key Laboratory of Chemo/Biosensing and Chemometrics, College of Chemistry and Chemical Engineering, Hunan University, Changsha, 410082 (China); Zhou, Yigang [Department of Microbiology, College of Basic Medicine, Central South University, Changsha, 410083 (China)

2016-09-14

A novel and facile magnetic deep eutectic solvents (DES) molecularly imprinted polymers (MIPs) for the selective recognition and separation of Bovine hemoglobin (BHb) was prepared. The new-type DES was adopted as the functional monomer which would bring molecular imprinted technology to a new direction. The amounts of DES were optimized. The obtained magnetic DES-MIPs were characterized with fourier transform infrared spectrometry (FT-IR), thermogravimetric analysis (TGA), field emission scanning electron microscope (FESEM), dynamic light scattering (DLS), elemental analysis and vibrating sample magnetometer (VSM). The results suggested that the imprinted polymers were successfully formed and possessed a charming magnetism. The maximum adsorption capability (Q{sub max}) and dissociation constant (K{sub L}) were analyzed by Langmuir isotherms (R{sup 2} = 0.9983) and the value were estimated to be 175.44 mg/g and 0.035 mg/mL for the imprinted particles. And the imprinted particles showed a high imprinting factor of 4.77. In addition, the magnetic DES-MIPs presented outstanding recognition specificity and selectivity so that it can be utilized to separate template protein from the mixture of proteins and real samples. Last but not least, the combination of deep eutectic solvents and molecular imprinted technology in this paper provides a new perspective for the recognition and separation of proteins. - Highlights: • Combined green deep eutectic solvents (DES) and molecular imprinted technology in recognition and separation of proteins. • DES was adopted as a new-type functional monomer. • The obtained magnetic DES-MIPs can separate proteins rapidly by an external magnetic field. • Adsorption and selectivity properties were discussed.

THE VALENCE OF CORPUSCULAR PROTEINS.

Science.gov (United States)

Gorin, M H; Mover, L S

1942-07-20

BY THE USE OF TWO EXTREME MODELS: a hydrated sphere and an unhydrated rod the valence (net charge) of corpuscular proteins can be successfully calculated from electric mobility data by the Debye-Hückel theory (modified to include the effect of the ions in the ion atmosphere) in conjunction with the electrophoretic theory of Henry. As pointed out by Abramson, this permits a comparison with values for the valence from titration data. Electrometric titration measurements of serum albumin B (Kekwick) have been determined at several ionic strengths. These results, together with the available data in the literature for serum albumin B, egg albumin, and beta-lactoglobulin have been used to compare values for the valence calculated from measurements of titration, electrophoresis, and membrane potentials. The results indicate that the usual interpretation of titration curves is open to serious question. By extrapolation of the titration data to zero ionic strength and protein concentration, there results an "intrinsic" net charge curve describing the binding of H(+) (OH(-)) ion alone. This curve agrees closely, in each case, with values of the valence calculated from mobility data (which in turn are in close accord with those estimated from membrane potential measurements). The experimental titration curves in the presence of appreciable quantities of ions and protein deviate widely from the ideal curve. It is suggested that, under these conditions, binding of undissociated acid (base) leads to erroneous values for the net charge. This binding would not affect the electrophoretic mobility. Values of the net charge obtained by the two extreme models from electrophoretic data are in agreement within 15 to 20 per cent. The agreement between the cylindrical model and the titration data is somewhat better in each case than with the sphere; i.e., this comparison enables a choice to be made between asymmetry and hydration in the interpretation of results from sedimentation and

Superhydrophobic nanostructured ZnO thin films on aluminum alloy substrates by electrophoretic deposition process

Energy Technology Data Exchange (ETDEWEB)

Huang, Ying; Sarkar, D.K., E-mail: dsarkar@uqac.ca; Chen, X-Grant

2015-02-01

Graphical abstract: - Highlights: • Fabrication of superhydrophobic ZnO thin films surfaces by electrophoretic deposition process on aluminum substrates. • Effect of bath temperature on the physical and superhydrophobic properties of thin films. • The water contact angle of 155° ± 3 with roll off property has been observed on the film that was grown at bath temperatures of 50 °C. • The activation energy for electrophoretic deposition of SA-functionalized ZnO nanoparticle is calculated to be 0.50 eV. - Abstract: Superhydrophobic thin films have been fabricated on aluminum alloy substrates by electrophoretic deposition (EPD) process using stearic acid (SA) functionalized zinc oxide (ZnO) nanoparticles suspension in alcohols at varying bath temperatures. The deposited thin films have been characterized using both X-ray diffraction (XRD) and infrared (IR) spectroscopy and it is found that the films contain low surface energy zinc stearate and ZnO nanoparticles. It is also observed that the atomic percentage of Zn and O, roughness and water contact angle of the thin films increase with the increase of the deposited bath temperature. Furthermore, the thin film deposited at 50 °C, having a roughness of 4.54 ± 0.23 μm, shows superhydrophobic properties providing a water contact angle of 155 ± 3° with rolling off properties. Also, the activation energy of electrophoretic deposition of stearic-acid-functionalized ZnO nanoparticles is calculated to be 0.5 eV.

Preconcentration and Separation of Mixed-Species Samples Near a Nano-Junction in a Convergent Microchannel

Directory of Open Access Journals (Sweden)

Ping-Hsien Chiu

2015-12-01

Full Text Available A fluidic microchip incorporating a convergent microchannel and a Nafion-nanoporous membrane is proposed for the preconcentration and separation of multi-species samples on a single platform. In the device, sample preconcentration is achieved by means of the ion concentration polarization effect induced at the micro/nano interface under the application of an external electric field, while species separation is achieved by exploiting the different electrophoretic mobilities of the sample components. The experimental results show that the device is capable of detecting C-reactive protein (CRP with an initial concentration as low as 9.50 × 10−6 mg/L given a sufficient preconcentration time and driving voltage. In addition, it is shown that a mixed-species sample consisting of three negatively-charged components (bovine serum albumin (BSA, tetramethylrhodamine(TAMRA isothiocyanate-Dextran and fluorescent polymer beads can be separated and preconcentrated within 20 min given a driving voltage of 100 V across 1 cm microchannel in length. In general, the present results confirm the feasibility of the device for the immunoassay or detection of various multi-species samples under low concentration in the biochemical and biomedical fields. The novel device can therefore improve the detection limit of traditional medical facilities.

Protein-Nanocellulose Interactions in Paper Filters for Advanced Separation Applications.

Science.gov (United States)

Gustafsson, Simon; Manukyan, Levon; Mihranyan, Albert

2017-05-16

Protein-based pharmaceutics are widely explored for healthcare applications, and 6 out of 10 best-selling drugs today are biologicals. The goal of this work was to evaluate the protein nanocellulose interactions in paper filter for advanced separation applications such as virus removal filtration and bioprocessing. The protein recovery was measured for bovine serum albumin (BSA), γ-globulin, and lysozyme using biuret total protein reagent and polyacrylamide gel electrophoresis (PAGE), and the throughput was characterized in terms of flux values from fixed volume filtrations at various protein concentrations and under worst-case experimental conditions. The affinity of cellulose to bind various proteins, such as BSA, lysozyme, γ-globulin, and human IgG was quantified using a quartz crystal microbalance (QCMB) by developing a new method of fixing the cellulose fibers to the electrode surface without cellulose dissolution-precipitation. It was shown that the mille-feuille filter exhibits high protein recovery, that is, ∼99% for both BSA and lysozyme. However, γ-globulin does not pass through the membrane due to its large size (i.e., >180 kDa). The PAGE data show no substantial change in the amount of dimers and trimers before and after filtration. QCMB analysis suggests a low affinity between the nanocellulose surface and proteins. The nanocellulose-based filter exhibits desirable inertness as a filtering material intended for protein purification.

Survival rate of eukaryotic cells following electrophoretic nanoinjection

OpenAIRE

Simonis, Matthias; H?bner, Wolfgang; Wilking, Alice; Huser, Thomas; Hennig, Simon

2017-01-01

Insertion of foreign molecules such as functionalized fluorescent probes, antibodies, or plasmid DNA to living cells requires overcoming the plasma membrane barrier without harming the cell during the staining process. Many techniques such as electroporation, lipofection or microinjection have been developed to overcome the cellular plasma membrane, but they all result in reduced cell viability. A novel approach is the injection of cells with a nanopipette and using electrophoretic forces for...

NMR studies of electrophoretic mobility in surfactant systems

International Nuclear Information System (INIS)

Conveney, F.M.; Strange, J.H.; Smith, A.L.; Smith, E.G.

1989-01-01

An experimental technique is described in which the flow of electrically charged micelles is measured in the presence of an applied electric field using an NMR technique. The method is used to determine the electrophoretic mobility at ambient temperature of a 5% aqueous solution of sodium dodecyl sulphate and is shown to provide a new technique for the study of electrophoresis in surfactant solutions. (author). 8 refs.; 4 figs

Layered ceramic composites via control of electrophoretic deposition kinetics

Czech Academy of Sciences Publication Activity Database

Hadraba, Hynek; Drdlík, D.; Chlup, Zdeněk; Maca, K.; Dlouhý, Ivo; Cihlář, J.

2013-01-01

Roč. 33, č. 12 (2013), s. 2305-2312 ISSN 0955-2219 R&D Projects: GA ČR(CZ) GAP108/11/1644; GA MŠk(CZ) ED1.1.00/02.0068 Institutional support: RVO:68081723 Keywords : Alumina * Zirconia * Laminates * Electrophoretic deposition Subject RIV: JH - Ceramics, Fire-Resistant Materials and Glass Impact factor: 2.307, year: 2013

Capillary electrophoretic profiling of tryptic digests of water soluble proteins from Bacillus thuringiensis-transgenic and non-transgenic maize species

Czech Academy of Sciences Publication Activity Database

Sázelová, Petra; Kašička, Václav; Leon, C.; Ibanez, E.; Cifuentes, A.

2012-01-01

Roč. 134, č. 3 (2012), s. 1607-1615 ISSN 0308-8146 R&D Projects: GA ČR(CZ) GA203/08/1428 Grant - others:AV ČR(CZ) 2008CZ0019 Institutional research plan: CEZ:AV0Z40550506 Keywords : Bt-transgenic maize * capillary zone electrophoresis * maize proteins Subject RIV: CB - Analytical Chemistry, Separation Impact factor: 3.334, year: 2012

Electrophoretic deposition of composite halloysite nanotube–hydroxyapatite–hyaluronic acid films

Energy Technology Data Exchange (ETDEWEB)

Deen, I. [Department of Materials Science and Engineering, McMaster University, 1280 Main Street West, Hamilton, Ontario, Canada L8S 4L7 (Canada); Zhitomirsky, I., E-mail: zhitom@mcmaster.ca [Department of Materials Science and Engineering, McMaster University, 1280 Main Street West, Hamilton, Ontario, Canada L8S 4L7 (Canada)

2014-02-15

Highlights: ► Composite halloysite nanotubes–hydroxyapatite–hyaluronic acid films were prepared. ► Electrophoretic deposition method was used for deposition. ► Natural hyaluronic acid was used as a dispersing, charging and film forming agent. ► Film composition and deposition yield can be varied. ► The films can be used for biomedical implants with controlled release of drugs. -- Abstract: Electrophoretic deposition method has been developed for the deposition of biocomposite films containing halloysite nanotubes (HNTs), hydroxyapatite (HA) and hyaluronic acid. The method is based on the use of natural hyaluronate biopolymer as a dispersing and charging agent for HNT and HA and film forming agent for the fabrication of the composite films. The deposition kinetics was studied by the quartz crystal microbalance method. The composite films were studied by X-ray diffraction, thermogravimetric analysis, differential thermal analysis and electron microscopy. The composite films are promising materials for the fabrication of biomedical implants with advanced functional properties.

Electrophoretic deposition of composite halloysite nanotube–hydroxyapatite–hyaluronic acid films

International Nuclear Information System (INIS)

Deen, I.; Zhitomirsky, I.

2014-01-01

Highlights: ► Composite halloysite nanotubes–hydroxyapatite–hyaluronic acid films were prepared. ► Electrophoretic deposition method was used for deposition. ► Natural hyaluronic acid was used as a dispersing, charging and film forming agent. ► Film composition and deposition yield can be varied. ► The films can be used for biomedical implants with controlled release of drugs. -- Abstract: Electrophoretic deposition method has been developed for the deposition of biocomposite films containing halloysite nanotubes (HNTs), hydroxyapatite (HA) and hyaluronic acid. The method is based on the use of natural hyaluronate biopolymer as a dispersing and charging agent for HNT and HA and film forming agent for the fabrication of the composite films. The deposition kinetics was studied by the quartz crystal microbalance method. The composite films were studied by X-ray diffraction, thermogravimetric analysis, differential thermal analysis and electron microscopy. The composite films are promising materials for the fabrication of biomedical implants with advanced functional properties

Nanocapillaries for Open Tubular Chromatographic Separations of Proteins in Femtoliter to Picoliter Samples

Science.gov (United States)

Wang, Xiayan; Cheng, Chang; Wang, Shili; Zhao, Meiping; Dasgupta, Purnendu K.; Liu, Shaorong

2009-01-01

We have recently examined the potential of bare nanocapillaries for free solution DNA separations and demonstrated efficiencies exceeding 106 theoretical plates/m. In the present work, we demonstrate the use of bare and hydroxypropylcellulose (HPC) coated open tubular nanocapillaries for protein separations. Using 1.5 μm inner diameter (i.d.) capillary columns, hydrodynamically injecting femto to picoliter (fL-pL) volumes of fluorescent or fluorescent dye labeled protein samples, utilizing a pneumatically pressurized chamber containing 1.0 mM sodium tetraborate solution eluent (typ. 200 psi) as the pump and performing on-column detection using a simple laser-induced fluorescence detector, we demonstrate efficiencies of close to a million theoretical plates/m while generating single digit μL volumes of waste for a complete chromatographic run. We achieve baseline resolution for a protein mixture consisting of transferrin, α-lactalbumin, insulin, and α -2-macroglobulin. PMID:19663450

Preparation of a novel dual-function strong cation exchange/hydrophobic interaction chromatography stationary phase for protein separation.

Science.gov (United States)

Zhao, Kailou; Yang, Li; Wang, Xuejiao; Bai, Quan; Yang, Fan; Wang, Fei

2012-08-30

We have explored a novel dual-function stationary phase which combines both strong cation exchange (SCX) and hydrophobic interaction chromatography (HIC) characteristics. The novel dual-function stationary phase is based on porous and spherical silica gel functionalized with ligand containing sulfonic and benzyl groups capable of electrostatic and hydrophobic interaction functionalities, which displays HIC character in a high salt concentration, and IEC character in a low salt concentration in mobile phase employed. As a result, it can be employed to separate proteins with SCX and HIC modes, respectively. The resolution and selectivity of the dual-function stationary phase were evaluated under both HIC and SCX modes with standard proteins and can be comparable to that of conventional IEC and HIC columns. More than 96% of mass and bioactivity recoveries of proteins can be achieved in both HIC and SCX modes, respectively. The results indicated that the novel dual-function column could replace two individual SCX and HIC columns for protein separation. Mixed retention mechanism of proteins on this dual-function column based on stoichiometric displacement theory (SDT) in LC was investigated to find the optimal balance of the magnitude of electrostatic and hydrophobic interactions between protein and the ligand on the silica surface in order to obtain high resolution and selectivity for protein separation. In addition, the effects of the hydrophobicity of the ligand of the dual-function packings and pH of the mobile phase used on protein separation were also investigated in detail. The results show that the ligand with suitable hydrophobicity to match the electrostatic interaction is very important to prepare the dual-function stationary phase, and a better resolution and selectivity can be obtained at pH 6.5 in SCX mode. Therefore, the dual-function column can replace two individual SCX and HIC columns for protein separation and be used to set up two-dimensional liquid

HPTLC-aptastaining - Innovative protein detection system for high-performance thin-layer chromatography

Science.gov (United States)

Morschheuser, Lena; Wessels, Hauke; Pille, Christina; Fischer, Judith; Hünniger, Tim; Fischer, Markus; Paschke-Kratzin, Angelika; Rohn, Sascha

2016-05-01

Protein analysis using high-performance thin-layer chromatography (HPTLC) is not commonly used but can complement traditional electrophoretic and mass spectrometric approaches in a unique way. Due to various detection protocols and possibilities for hyphenation, HPTLC protein analysis is a promising alternative for e.g., investigating posttranslational modifications. This study exemplarily focused on the investigation of lysozyme, an enzyme which is occurring in eggs and technologically added to foods and beverages such as wine. The detection of lysozyme is mandatory, as it might trigger allergenic reactions in sensitive individuals. To underline the advantages of HPTLC in protein analysis, the development of innovative, highly specific staining protocols leads to improved sensitivity for protein detection on HPTLC plates in comparison to universal protein derivatization reagents. This study aimed at developing a detection methodology for HPTLC separated proteins using aptamers. Due to their affinity and specificity towards a wide range of targets, an aptamer based staining procedure on HPTLC (HPTLC-aptastaining) will enable manifold analytical possibilities. Besides the proof of its applicability for the very first time, (i) aptamer-based staining of proteins is applicable on different stationary phase materials and (ii) furthermore, it can be used as an approach for a semi-quantitative estimation of protein concentrations.

Carbon nanotubes-assisted polyacrylamide gel electrophoresis for enhanced separation of human serum proteins and application in liverish diagnosis.

Science.gov (United States)

Jiang, Fubin; Wang, Yanan; Hu, Xinfang; Shao, Na; Na, Na; Delanghe, Joris R; Ouyang, Jin

2010-11-01

The application of pore-gradient polyacrylamide gel electrophoresis (PG-PAGE) incorporated with carbon nanotube modified by Triton X-100 and carboxylation so as to improve the separation of human serum proteins is reported. The novel PG-PAGE was made by adding water-soluble single-walled carbon nanotubes (CNTs) when preparing the polyacrylamide gel. Significant improvements in separation of complement C3 protein and haptoglobin (Hp) in human serum were achieved. It was estimated that the interactions between the hydrophilic groups on the proteins and the surface of the CNTs result in different adsorption kinetics of complement C3 and Hp subtype on the nanoparticles incorporated in the gel, thus enhancing the separation of the two proteins in serum. This new CNT matrix-assisted PG-PAGE method for enhanced separation of complement C3 and Hp in human serum was successfully applied to distinguish the samples from liverish patients and healthy people.

Characterisation of selenium compounds in rye seedling biomass using {sup 75}Se-labelling/SDS-PAGE separation/{gamma}-scintillation counting, and HPLC-ICP-MS analysis of a range of enzymatic digests

Energy Technology Data Exchange (ETDEWEB)

Bryszewska, Malgorzata A. [Technical University of Lodz (Poland). Institute of General Food Chemistry; Ambroziak, Wojciech [Technical University of Lodz (Poland). Institute of Fermentation Technology and Microbiology; Rudzinski, Juliusz [Technical University of Lodz (Poland). Institute of Applied Radiation Chemistry; Lewis, D. John [Central Science Laboratory, York (United Kingdom)

2005-07-01

In the present study, selenium-enriched plant biomass was investigated to evaluate the ability of rye seedlings to take up, and assimilate, inorganic selenium. Two different analytical approaches were used. Electrophoretic separation (SDS-PAGE) of proteins extracted from {sup 75}Se-labelled biomass was used to investigate the biotransformation of selenite into organic forms of the element. Ion-pair chromatography coupled with ICP-MS detection was chosen for the analysis of selenium species, enzymatically extracted from the plant biomass. The results of three enzymatic hydrolysis procedures and three sequential enzymatic extractions procedures are compared. The most effective single extraction was proteolysis (using protease type XIV), giving an overall extraction efficiency of 48%. However, for combinations of enzymes, the most effective was cellulase (Trichoderma viride) followed by sequential extraction of the solid pellet using protease type XIV, giving an extraction efficiency of 70%. The complementary data from the electrophoretic fractionation of proteins, and the HPLC separation of Se-species in the proteolytic digests, reveal the existence of large number of selenium-containing compounds in the rye seedling plant biomass. The results showed the complete biotransformation of inorganic selenium into organic forms during germination of the rye seedlings. HPLC-ICP-MS analysis of extracts from the plant biomass did not show the presence of selenate or selenite. At the time of this study, the lack of suitable organic-MS facilities meant that it was not possible to characterise them fully. However, the data does show that a combination of different enzymes, rather than just the commonly-used protease, should be considered when developing an extraction strategy for selenium in different food types to those already reported in the literature. (orig.)

Effect of acids and bases on electrophoretic deposition of

Czech Academy of Sciences Publication Activity Database

Cihlář, J.; Drdlík, D.; Cihlářová, Z.; Hadraba, Hynek

2013-01-01

Roč. 33, č. 10 (2013), s. 1885-1892 ISSN 0955-2219 R&D Projects: GA MŠk(CZ) ED1.1.00/02.0068; GA ČR GD106/09/H035 Institutional support: RVO:68081723 Keywords : Electrophoretic deposition * Zirconia * Alumina * 2-Propanol * Electrosteric stabilization Subject RIV: JH - Ceramics, Fire-Resistant Materials and Glass Impact factor: 2.307, year: 2013

Photopatterned free-standing polyacrylamide gels for microfluidic protein electrophoresis.

Science.gov (United States)

Duncombe, Todd A; Herr, Amy E

2013-06-07

Designed for compatibility with slab-gel polyacrylamide gel electrophoresis (PAGE) reagents and instruments, we detail development of free-standing polyacrylamide gel (fsPAG) microstructures supporting electrophoretic performance rivalling that of microfluidic platforms. For the protein electrophoresis study described here, fsPAGE lanes are comprised of a sample reservoir and contiguous separation gel. No enclosed microfluidic channels are employed. The fsPAG devices (120 μm tall) are directly photopatterned atop of and covalently attached to planar polymer or glass surfaces. Leveraging the fast prototype-test cycle - significantly faster than mold based fabrication techniques - we optimize the fsPAG architecture to minimize injection dispersion for rapid (prototyping of the fsPAGE provides researchers a powerful tool for developing custom analytical assays. We highlight the utility of assay customization by fabricating a polyacrylamide gel with a spatial pore-size distribution and demonstrate the resulting enhancement in separation performance over a uniform gel. Further, we up-scale from a unit separation to an array of 96 concurrent fsPAGE assays in 10 min run time driven by one electrode pair. The fsPAG array layout matches that of a 96-well plate to facilitate integration of the planar free standing gel array with multi-channel pipettes while remaining compatible with conventional slab-gel PAGE reagents, such as staining for label-free protein detection. Notably, the entire fsPAGE workflow from fabrication, to operation, and readout uses readily available materials and instruments - making this technique highly accessible.

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

Science.gov (United States)

Chang, Ming-Mei; Lovett, Janice

2011-01-01

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

Improved Bacterial and Viral Recoveries from 'Complex' Samples using Electrophoretically Assisted Acoustic Focusing

Energy Technology Data Exchange (ETDEWEB)

Ness, K; Rose, K; Jung, B; Fisher, K; Mariella, Jr., R P

2008-03-27

Automated front-end sample preparation technologies can significantly enhance the sensitivity and reliability of biodetection assays [1]. We are developing advanced sample preparation technologies for biowarfare detection and medical point-of-care diagnostics using microfluidic systems with continuous sample processing capabilities. Here we report an electrophoretically assisted acoustic focusing technique to rapidly extract and enrich viral and bacterial loads from 'complex samples', applied in this case to human nasopharyngeal samples as well as simplified surrogates. The acoustic forces capture and remove large particles (> 2 {micro}m) such as host cells, debris, dust, and pollen from the sample. We simultaneously apply an electric field transverse to the flow direction to transport small ({le} 2 {micro}m), negatively-charged analytes into a separate purified recovery fluid using a modified H-filter configuration [Micronics US Patent 5,716,852]. Hunter and O'Brien combined transverse electrophoresis and acoustic focusing to measure the surface charge on large particles, [2] but to our knowledge, our work is the first demonstration combining these two techniques in a continuous flow device. Marina et al. demonstrated superimposed dielectrophoresis (DEP) and acoustic focusing for enhanced separations [3], but these devices have limited throughput due to the rapid decay of DEP forces. Both acoustic standing waves and electric fields exert significant forces over the entire fluid volume in microchannels, thus allowing channels with larger dimensions (> 100 {micro}m) and high throughputs (10-100 {micro}L/min) necessary to process real-world volumes (1 mL). Previous work demonstrated acoustic focusing of microbeads [4] and biological species [5] in various geometries. We experimentally characterized our device by determining the biological size-cutoff where acoustic radiation pressure forces no longer transport biological particles. Figure 1 shows

Electrophoretic deposits of boron on duralumin plates used for measuring neutron flux

International Nuclear Information System (INIS)

Lang, F.M.; Magnier, P.; Finck, C.

1956-01-01

Preparation of boron thin film deposits of around 1 mg per cm 2 on duralumin plates with a diameter of 8 cm. The boron coated plates for ionization chambers were originally prepared at the CEA by pulverization of boron carbides on sodium silicates. This method is not controlling precisely enough the quantity of boron deposit. Thus, an electrophoretic method is considered for a better control of the quantity of boron deposit in the scope of using in the future boron 10 which is costly and rare. The method described by O. Flint is not satisfying enough and a similar electrophoretic process has been developed. Full description of the method is given as well as explanation of the use of dried methanol as solvent, tannin as electrolyte and magnesium chloride to avoid alumina formation. (M.P.)

Differentiation of highly virulent strains of Streptococcus suis serotype 2 according to glutamate dehydrogenase electrophoretic and sequence type.

Science.gov (United States)

Kutz, Russell; Okwumabua, Ogi

2008-10-01

The glutamate dehydrogenase (GDH) enzymes of 19 Streptococcus suis serotype 2 strains, consisting of 18 swine isolates and 1 human clinical isolate from a geographically varied collection, were analyzed by activity staining on a nondenaturing gel. All seven (100%) of the highly virulent strains tested produced an electrophoretic type (ET) distinct from those of moderately virulent and nonvirulent strains. By PCR and nucleotide sequence determination, the gdh genes of the 19 strains and of 2 highly virulent strains involved in recent Chinese outbreaks yielded a 1,820-bp fragment containing an open reading frame of 1,344 nucleotides, which encodes a protein of 448 amino acid residues with a calculated molecular mass of approximately 49 kDa. The nucleotide sequences contained base pair differences, but most were silent. Cluster analysis of the deduced amino acid sequences separated the isolates into three groups. Group I (ETI) consisted of the seven highly virulent isolates and the two Chinese outbreak strains, containing Ala(299)-to-Ser, Glu(305)-to-Lys, and Glu(330)-to-Lys amino acid substitutions compared with groups II and III (ETII). Groups II and III consisted of moderately virulent and nonvirulent strains, which are separated from each other by Tyr(72)-to-Asp and Thr(296)-to-Ala substitutions. Gene exchange studies resulted in the change of ETI to ETII and vice versa. A spectrophotometric activity assay for GDH did not show significant differences between the groups. These results suggest that the GDH ETs and sequence types may serve as useful markers in predicting the pathogenic behavior of strains of this serotype and that the molecular basis for the observed differences in the ETs was amino acid substitutions and not deletion, insertion, or processing uniqueness.

ATPS: "Aqueous two-phase System" as the "Answer to Protein Separation" for protein-processing Food Industry.

Science.gov (United States)

Khan, Bilal Muhammad; Liu, Zhi-Cong; Shi, Fu-Lin; Cheong, Kit-Leong; Liu, Yang

2018-06-08

Every individual needs food for its nutritional value and flavor while the economic growth of a nation depends on a thriving profit-generating industry. The food industry caters to both needs in an efficient manner. Proteins can rightly be considered as the driving force behind the overwhelming success of this industry. However, purification of proteins is not an easy undertaking due to their intricate nature while presently employed procedures for this purpose, regrettably, are both costly, and labor- and time-intensive in addition to being unsettling on proteins structural conformity. ATPS has accumulated a lot of interest from the scientific community due to its mild operating conditions, high recovery yield, ease of scaling it up, and its cost-effective and environment friendly nature. This review tries to amass some accounts concerning the utility of ATPS for the separation and purification of proteins. Some auspicious clues in this regard can be witnessed along with a few loopholes which need to be addressed before this technique can truly demonstrate its potential vis-à-vis industrial protein purification. Overall, a polymer - salt (citrates in particular) ATPS with an added inert supplementary salt can be regarded as a better option for purifying proteins.

Electrophoretic deposition of PEEK-TiO 2 composite coatings on stainless steel

KAUST Repository

Seuß , Sigrid; Subhani, Tayyab; Yi Kang, Min; Okudaira, Kenji; Ventura, Isaac Aguilar; Boccaccini, Aldo R.

2012-01-01

Electrophoretic deposition (EPD) has been successfully used to deposit composite coatings composed of polyetheretherketone (PEEK) and titanium dioxide (TiO 2) nanoparticles on 316L stainless steel substrates. The suspensions of TiO2 nanoparticles

Electrophoretic preparation and characterization of porous electrodes from diamond nanoparticles

Energy Technology Data Exchange (ETDEWEB)

Riveros, Lyda La Torre; Soto, Keyla; Tryk, Donald A; Cabrera, Carlos R [Department of Chemistry and Center of Nanoscale Materials, University of Puerto Rico, Rio Piedras, PO Box 23346 San Juan, PR 00931-3346 (Puerto Rico)

2007-04-15

We carried out chemical purification of commercially available diamond nanoparticles by refluxing in aqueous HNO{sub 3} and characterized the samples by spectroscopic and surface techniques before and after purification. As a first step in the preparation of electrodes for electrochemistry, we have electrophoretically deposited thin, highly uniform films of controlled thickness (1-8 {mu}m) on silicon substrates using the purified diamond nanoparticles. These have been characterized by scanning electron microscopy (SEM). All films obtained were homogeneous in thickness and without macroscopic holes or cracks. Such structures could also be used in many other applications such as fuel cells or lithium batteries. We have performed cyclic voltammetry experiments with these electrodes. The voltammograms of diamond nanoparticles electrophoretically deposited on silicon indicate hydrogen evolution. This demonstrates that the material is useful as electrocatalitic support. This conclusion is supported by the cyclic voltammograms obtained using ferrycyanide (III) chloride and hexaamineruthenium (III) chloride complexes as redox probes. However, these redox probes showed very small peak currents. This behavior could be improved by doping the diamond nanoparticles with an impurity such as boron.

Electrophoretic preparation and characterization of porous electrodes from diamond nanoparticles

International Nuclear Information System (INIS)

Riveros, Lyda La Torre; Soto, Keyla; Tryk, Donald A; Cabrera, Carlos R

2007-01-01

We carried out chemical purification of commercially available diamond nanoparticles by refluxing in aqueous HNO 3 and characterized the samples by spectroscopic and surface techniques before and after purification. As a first step in the preparation of electrodes for electrochemistry, we have electrophoretically deposited thin, highly uniform films of controlled thickness (1-8 μm) on silicon substrates using the purified diamond nanoparticles. These have been characterized by scanning electron microscopy (SEM). All films obtained were homogeneous in thickness and without macroscopic holes or cracks. Such structures could also be used in many other applications such as fuel cells or lithium batteries. We have performed cyclic voltammetry experiments with these electrodes. The voltammograms of diamond nanoparticles electrophoretically deposited on silicon indicate hydrogen evolution. This demonstrates that the material is useful as electrocatalitic support. This conclusion is supported by the cyclic voltammograms obtained using ferrycyanide (III) chloride and hexaamineruthenium (III) chloride complexes as redox probes. However, these redox probes showed very small peak currents. This behavior could be improved by doping the diamond nanoparticles with an impurity such as boron

Facilitating the Hyphenation of CIEF and MALDI-MS for Two-Dimensional Separation of Proteins

Science.gov (United States)

Cheng, Chang; Lu, Joann J.; Wang, Xiayan; Roberts, Jonathan; Liu, Shaorong

2011-01-01

Both CIEF and MALDI-MS are frequently used in protein analysis, but hyphenation of the two is not investigated proportionally. One of the major reasons is that the additives (such as carrier ampholytes and detergent) in CIEF severely suppress the MALDI-MS signal, which hampers the hyphenation of the two. In this paper, we develop a simple means to alleviate the above signal-suppressing effect. We first deposit 1 µL of water onto a MALDI-MS target, deliver a fraction of CIEF-separated protein (~0.1 µL) to the water droplet, evaporate the solvent, add 0.5 µL of MALDI matrix to the sample spot, dry the matrix, and move the target plate to a MALDI-TOF-MS for mass spectrum measurement. We optimize the droplet volume and the laser-ablation region. Under the optimized conditions, we improve the signal to noise ratio by 2–10 fold. We also apply this method for two-dimensional separations of standard proteins and Apolipoprotein A-I, a membrane protein expressed in E. Coli cells. PMID:20603827

Nano-colloid electrophoretic transport: Fully explicit modelling via dissipative particle dynamics

Science.gov (United States)

Hassanzadeh Afrouzi, Hamid; Farhadi, Mousa; Sedighi, Kurosh; Moshfegh, Abouzar

2018-02-01

In present study, a novel fully explicit approach using dissipative particle dynamics (DPD) method is introduced for modelling electrophoretic transport of nano-colloids in an electrolyte solution. Slater type charge smearing function included in 3D Ewald summation method is employed to treat electrostatic interaction. Moreover, capability of different thermostats are challenged to control the system temperature and study the dynamic response of colloidal electrophoretic mobility under practical ranges of external electric field in nano scale application (0.072 600 in DPD units regardless of electric field intensity. Nosé-Hoover-Lowe-Andersen and Lowe-Andersen thermostats are found to function more effectively under high electric fields (E > 0.145 [ v / nm ]) while thermal equilibrium is maintained. Reasonable agreements are achieved by benchmarking the radial distribution function with available electrolyte structure modellings, as well as comparing reduced mobility against conventional Smoluchowski and Hückel theories, and numerical solution of Poisson-Boltzmann equation.

Preparation and surface encapsulation of hollow TiO nanoparticles for electrophoretic displays

International Nuclear Information System (INIS)

Zhao Qian; Tan Tingfeng; Qi Peng; Wang Shirong; Bian Shuguang; Li Xianggao; An Yong; Liu Zhaojun

2011-01-01

Hollow black TiO nanosparticles were obtained via deposition of inorganic coating on the surface of hollow core-shell polymer latex with Ti(OBu) 4 as precursor and subsequent calcination in ammonia gas. Hollow TiO particles were characterized by scanning electron microscope, transmission electronic microscopy, X-ray diffraction, and thermogravimetric analysis. Encapsulation of TiO via dispersion polymerization was promoved by pretreating the pigments with 3-(trimethoxysilyl) propyl methacrylate, making it possible to prepare hollow TiO-polymer particles. When St and DVB were used as polymerization monomer, hollow TiO-polymer core-shell particles came into being via dispersion polymerization, and the lipophilic degree is 28.57%. Glutin-arabic gum microcapsules containing TiO-polymer particles electrophoretic liquid were prepared using via complex coacervation. It was founded that hollow TiO-polymer particles had enough electrophoretic mobility after coating with polymer.

Comparing nanostructured hydroxyapatite coating on AZ91 alloy samples via sol-gel and electrophoretic deposition for biomedical applications.

Science.gov (United States)

Rojaee, Ramin; Fathi, Mohammadhossein; Raeissi, Keyvan

2014-12-01

Magnesium is one of the most critical elements in hard tissues regeneration and therefore causes speeding up the restoration of harmed bones, while high deterioration rate of magnesium in body fluid restricts it to be used as biodegradable implants. Alloying magnesium with some relatively nobler metals such as aluminium, zinc, rare earth elements, magnesium-bioceramics composites, and surface modification techniques are some of the routes to control magnesium corrosion rate. In this study AZ91 magnesium alloy had been coated by nanostructured hydroxyapatite via sol-gel dip coating and electrophoretical methods to survey the final barricade properties of the obtained coatings. In order to perform electrophoretic coating, powders were prepared by sol-gel method, and then the powders deposited on substrates utilizing direct current electricity. Zeta potentials of the electrophoresis suspensions were measured to determine a best mode for good quality coatings. Transmission Electron Microscopy (TEM), and Scanning Electron Microscopy (SEM) were used to confirm nanoscale dimension, and the uniformity of the nanostructured hydroxyapatite coating, respectively. Fourier Transform-Infrared and X-ray diffraction analysis were utilized for functional group and phase structure evaluation of the prepared coatings, correspondingly. Electrochemical corrosion tests were performed in SBF at 37±1 (°)C which revealed considerable increase in corrosion protection resistivity and corrosion current density for electrophoretic coated specimens versus sol-gel coated specimens. Results showed that both sol-gel and electrophoretical techniques seem to be suitable to coat magnesium alloys for biomedical applications but electrophoretic coating technique is a better choice due to the more homogeneity and more crystalline structure of the coating.

Perfil electroforético de proteínas presentes en la saliva de Triatoma dimidiata (Hemiptera: Reduviidae:Triatominae Electrophoretic profile of salivary proteins of Triatoma dimidiata (Hemiptera: Reduviidae:Triatominae

Directory of Open Access Journals (Sweden)

Mónica Flórez

2009-08-01

that secrete saliva rich in proteins with anticoagulant, antihistamine, vasodilator and platelet inhibitor properties, these facilitate its alimentary process on the vertebrate host and facilitate transmission of the protozoa carried in the salivary glands of the triatomines. Such proteins are characteristic of each triatomine species and might help differentiate species, including those phenotypically similar. Objective: Describe electrophoretic profiles of salivary proteins of Triatoma dimidiata found inside, around and outside residences in an endemic area of Santander. Materials and methods: Salivary glands from adult insects of T. dimidiata from laboratory colonies and field from three municipalities of Santander were dissected. The protein profiles were viewed in a unidimensional electrophoresis of poliacrilamida gels taken with coomassie blue. Results: The electrophoretic profiles of proteins present in saliva of T. dimidiata showed up to 33 bands in the range of 23.7 to 228.8 kDa, with a high concentration in the region 41 to 99.7 kDa. The index of polymorphism to T. dimidiata was 0.9646. Conclusion: The electrophorectic profile of salivary protein of T. dimidiata showed a complex composition, where the most prominent bands have molecular weights lower than 45 KDa. No grouping could be established based on geographical regions and capture places, in spite of the great intraespecific variability observed. However, clear differences between T. dimidiata and the external group were established. Salud UIS 2009; 41: 121-127.

Nano-structured yttria-stabilized zirconia coating by electrophoretic deposition

Energy Technology Data Exchange (ETDEWEB)

Maleki-Ghaleh, H., E-mail: H_Maleki@sut.ac.ir [Faculty of Materials Engineering, Sahand University of Technology, Tabriz (Iran, Islamic Republic of); Rekabeslami, M. [Faculty of Mechanical Engineering, Materials Science and Engineering Division, K. N. Toosi University of Technology, Tehran (Iran, Islamic Republic of); Shakeri, M.S. [Materials and Energy Research Center, Karaj (Iran, Islamic Republic of); Siadati, M.H. [Faculty of Mechanical Engineering, Materials Science and Engineering Division, K. N. Toosi University of Technology, Tehran (Iran, Islamic Republic of); Javidi, M. [Department of Materials Science and Engineering, School of Engineering, Shiraz University, Shiraz (Iran, Islamic Republic of); Talebian, S.H. [Faculty of Petroleum Engineering, Universiti Technologi Petronas, Perak (Malaysia); Aghajani, H. [Department of Materials Engineering, University of Tabriz, Tabriz (Iran, Islamic Republic of)

2013-09-01

The most important role of thermal barrier coatings is to reduce the temperature of the substrate in high temperature applications. Nanoparticle zirconia might be a suitable choice for improving the efficiency of thermal barrier coatings. Nanostructured coatings have lower thermal conduction, higher thermal expansion and lower dimensional variations at higher temperatures in comparison with the microstructured coatings. Electrophoretic deposition has been preferred for thermal barrier coatings due to its simplicity, controllability and low cost. In the present study, three different suspensions of ZrO{sub 2}–8 wt%Y{sub 2}O{sub 3} (40 nm) made with ethanol, acetone and acetyl acetone were used. Electrophoretic deposition was conducted at a fixed voltage of 60 V for 120 s on aluminized Inconel 738-LC, and then heat treated at 1100{sup o}C for 4 h in air atmosphere. The coating morphology and elemental distribution were studied using scanning electron microscopy. It was observed that suspension media have an important effect on the quality of the final product. Acetyl acetone showed better dispersion of particles than the other two media. Consequently, deposition from acetyl acetone resulted in uniform and crack-free layers while those from ethanol and acetone were completely non-uniform due to agglomeration and low viscosity, respectively.

Recent applications of capillary electromigration methods to separation and analysis of proteins

Czech Academy of Sciences Publication Activity Database

Štěpánová, Sille; Kašička, Václav

2016-01-01

Roč. 933, Aug 24 (2016), s. 23-42 ISSN 0003-2670 R&D Projects: GA ČR(CZ) GA15-01948S Institutional support: RVO:61388963 Keywords : affinity electrophoresis * capillary electrophoresis * isoelectric focusing * isotachophoresis * proteins * review Subject RIV: CB - Analytical Chemistry, Separation Impact factor: 4.950, year: 2016

Transformation of Sordaria macrospora to hygromycin B resistance: characterization of transformants by electrophoretic karyotyping and tetrad analysis.

Science.gov (United States)

Walz, M; Kück, U

1995-12-01

The ascomycete Sordaria macrospora was transformed using different plasmid molecules containing the bacterial hygromycin B resistance gene (hph) under the control of different expression signals. The highest transformation frequency was obtained with vector pMW1. On this plasmid molecule, expression of the hph gene is directed by the upstream region of the isopenicillin N synthetase gene (pcbC) from the deuteromycete Acremonium chrysogenum. Southern analysis suggests that the vector copies are integrated as tandem repeats into the S. macrospora chromosomes and that duplicated sequences are most probably not inactivated by methylation during meiosis. Furthermore, the hygromycin B resistance (hygR) is not correlated with the number of integrated vector molecules. Electrophoretic karyotyping was used to further characterize S. macrospora transformants. Five chromosomal bands were separated by pulsed-field gel electrophoresis (PFGE) representing seven chromosomes with a total genome size of 39.5Mb. Hybridization analysis revealed ectopic integration of vector DNA into different chromosomes. In a few transformants, major rearrangements were detected. Transformants were sexually propagated to analyze the fate of the heterologous vector DNA. Although the hygR phenotype is stably maintained during mitosis, about a third of all lines tested showed loss of the resistance marker gene after meiosis. However, as was concluded from electrophoretic karyotyping, the resistant spores showed a Mendelian segregation of the integrated vector molecules in at least three consecutive generations. Our data indicate that heterologous marker genes can be used for transformation tagging, or the molecular mapping of chromosomal loci in S. macrospora.

Rapid detection and purification of sequence specific DNA binding proteins using magnetic separation

Directory of Open Access Journals (Sweden)

TIJANA SAVIC

2006-02-01

Full Text Available In this paper, a method for the rapid identification and purification of sequence specific DNA binding proteins based on magnetic separation is presented. This method was applied to confirm the binding of the human recombinant USF1 protein to its putative binding site (E-box within the human SOX3 protomer. It has been shown that biotinylated DNA attached to streptavidin magnetic particles specifically binds the USF1 protein in the presence of competitor DNA. It has also been demonstrated that the protein could be successfully eluted from the beads, in high yield and with restored DNA binding activity. The advantage of these procedures is that they could be applied for the identification and purification of any high-affinity sequence-specific DNA binding protein with only minor modifications.

Albumin coatings by alternating current electrophoretic deposition for improving corrosion resistance and bioactivity of titanium implants.

Science.gov (United States)

Höhn, Sarah; Braem, Annabel; Neirinck, Bram; Virtanen, Sannakaisa

2017-04-01

Although Ti alloys are generally regarded to be highly corrosion resistant, inflammatory conditions following surgery can instigate breakdown of the TiO 2 passivation layer leading to an increased metal ion release. Furthermore proteins present in the surrounding tissue will readily adsorb on a titanium surface after implantation. In this paper alternating current electrophoretic deposition (AC-EPD) of bovine serum albumin (BSA) on Ti6Al4V was investigated in order to increase the corrosion resistance and control the protein adsorption capability of the implant surface. The Ti6Al4V surface was characterized with SEM, XPS and ToF-SIMS after long-term immersion tests under physiological conditions and simulated inflammatory conditions either in Dulbecco's Modified Eagle Medium (DMEM) or DMEM supplemented with fetal calf serum (FCS). The analysis showed an increased adsorption of amino acids and proteins from the different immersion solutions. The BSA coating was shown to prevent selective dissolution of the vanadium (V) rich β-phase, thus effectively limiting metal ion release to the environment. Electrochemical impedance spectroscopy measurements confirmed an increase of the corrosion resistance for BSA coated surfaces as a function of immersion time due to the time-dependent adsorption of the different amino acids (from DMEM) and proteins (from FCS) as observed by ToF-SIMS analysis. Copyright © 2017 Elsevier B.V. All rights reserved.

A rollable, organic electrophoretic QVGA display with field-shielded pixel architecture

NARCIS (Netherlands)

Gelinck, G.H.; Huitema, H.E.A.; Mil, M. van; Veenendaal, E. van; Lieshout, P.J.G. van; Touwslager, F.; Patry, S.F.; Sohn, S.; Whitesides, T.; McCreary, M.D.

2006-01-01

A 100-um thin QVGA display was made by combining a 25-um thin organic transistor active-matrix backplane with an electrophoretic display film. High contrast and low crosstalk was achieved by the addition of a field shield to the backplane. The display can be bent repeatedly to a radius of 2 mm

Prebeta-migrating high density lipoprotein: quantitation in normal and hyperlipidemic plasma by solid phase radioimmunoassay following electrophoretic transfer

International Nuclear Information System (INIS)

Ishida, B.Y.; Frolich, J.; Fielding, C.J.

1987-01-01

A quantitative solid phase immunoassay has been developed for the determination of the mass of electrophoretically separated prebeta apolipoprotein A-I (apoA-I) in human plasma. Conditions have been identified for the quantitative transfer and immunoblotting of the apolipoprotein in the absence of organic solvents or detergents. In normolipidemic plasma, the prebeta-migrating fraction of apoA-I represented 4.2 +/- 1.8% of total apoA-I (61 +/- 26 micrograms of apoA-I per ml of plasma). Significantly higher levels were found in hypercholesterolemia of genetic origin, in primary and secondary hypertriglyceridemia, and in congenital lecithin:cholesterol acyltransferase deficiency. In all cases prebeta-migrating apoA-I consisted in large part of low molecular weight lipoprotein species, compared to the size of the major, alpha-migrating apoA-I fraction

Electrophoretic Ink Display Prepared by Jelly Fig Pectin/Gelatin Microspheres

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Wing-Ming Chou

2015-05-01

Full Text Available A brand new Bio-Electronic ink (Bio-E ink display device was prepared and characterized in this study. Semiconductor material, copper phthalocyanine (CuPc was modified by cationic surfactants, cetylpyridinium chloride (CPC, as the core material, and the shell of capsule was prepared by jelly fig pectin, gelatin and sodium dodecyl sulphate (SDS. Here, jelly fig pectin was provided as the shell material for the first time. Chemical structure of the modified CuPc was characterized by Fourier Transform Infrared Spectrometer (FTIR. The core-shell microcapsules were achieved by coacervation method in an oil/water (O/W emulsion system. The particle size and morphology of microcapsules were affected by the concentrations of SDS and pH values of the O/W emulsion system. A new microcapsule-based electrophoretic display device was presented. Its image display ability of the microcapsules electrophoretic device was presented as appropriated electric power was applied, and the response time was 0.06 sec under 0.1 V/mm of electric field. Moreover, we found that its image contrast ratio of display device was influenced by the particle sizes of the microcapsules.

Numerical simulation of stress-strain state of electrophoretic shell molds

Science.gov (United States)

Sviridov, A. V.; Odinokov, V. I.; Dmitriev, E. A.; Evstigneev, A. I.; Bashkov, O. V.

2017-10-01

In the foundry engineering, castings obtained in one-piece non-gas-generating high-refractory electrophoretic shell molds (ShM) by investment patterns (IP) have an increased rejects percentage associated with low deformation resistance and crack resistance of the SM at different stages of their formation and manufacturing. Crack resistance of the ShM based on IP depends mainly on their stress-strain state (SSS) at various stages of mold forming. SSS decrease significantly improves their crack resistance and decreases their rejects percentage of castings occurring due to clogging and surface defects. In addition, the known methods of decreasing the SSS are still poorly understood. Thus, current research trends are to determine SSS at each stage of ShM forming and develop the ways to decrease it. Theoretical predicting of crack formation in multiple-layer axisymmetric shell molds is given in the work [1], and SSS of multiple-layer axisymmetric shell molds is given in the work [2]. Monolayer electrophoretic ShM had a lack of concern in this field, thus it became an argument for the present workMathematical Model of ShM SSS

A cost-effective device for the rapid transfer of gel-separated proteins onto membranes.

Science.gov (United States)

Tam, Hann W; Huang, Yu-Chen; Tam, Ming F

2009-03-01

We describe here the fabrication of a cost-effective semi-dry blotting apparatus for the transfer of proteins onto membranes. Graphite sheets were used as electrodes. Protein mixtures were separated on NuPAGE 4% to 12% polyacrylamide gradient gels. With a Tris-bicine buffer, we demonstrated that close to 80% of the proteins with apparent molecular mass of 80kDa or less were removed from the gels after 8min of blotting. The process is much faster than the techniques reported previously in the literature.

Study on proteins of bean plants originating from gamma-treated seeds

International Nuclear Information System (INIS)

Tsonev, D.; Chalykova, M.

1975-01-01

Dry seeds of bean variety Stella were radiated with gamma rays using 60 C as a source of radiation. The dose applied was 30 kR at an intensity 9.6 R/s. Irradiation caused disturbances at synthesis and properties of proteins of bean plants originating from gamma treated seeds. Some protein components showed changes in relative content and electrophoretic mobility. New more mobile protein components appeared. Protein complex of overground parts were more radioresistant than protein complex of the roots. (M.Ts.)

Unified superresolution experiments and stochastic theory provide mechanistic insight into protein ion-exchange adsorptive separations.

Science.gov (United States)

Kisley, Lydia; Chen, Jixin; Mansur, Andrea P; Shuang, Bo; Kourentzi, Katerina; Poongavanam, Mohan-Vivekanandan; Chen, Wen-Hsiang; Dhamane, Sagar; Willson, Richard C; Landes, Christy F

2014-02-11

Chromatographic protein separations, immunoassays, and biosensing all typically involve the adsorption of proteins to surfaces decorated with charged, hydrophobic, or affinity ligands. Despite increasingly widespread use throughout the pharmaceutical industry, mechanistic detail about the interactions of proteins with individual chromatographic adsorbent sites is available only via inference from ensemble measurements such as binding isotherms, calorimetry, and chromatography. In this work, we present the direct superresolution mapping and kinetic characterization of functional sites on ion-exchange ligands based on agarose, a support matrix routinely used in protein chromatography. By quantifying the interactions of single proteins with individual charged ligands, we demonstrate that clusters of charges are necessary to create detectable adsorption sites and that even chemically identical ligands create adsorption sites of varying kinetic properties that depend on steric availability at the interface. Additionally, we relate experimental results to the stochastic theory of chromatography. Simulated elution profiles calculated from the molecular-scale data suggest that, if it were possible to engineer uniform optimal interactions into ion-exchange systems, separation efficiencies could be improved by as much as a factor of five by deliberately exploiting clustered interactions that currently dominate the ion-exchange process only accidentally.

Protonation of the polyethyleneimine and titanium particles and their effect on the electrophoretic mobility and deposition

Energy Technology Data Exchange (ETDEWEB)

Lau, Kok-Tee, E-mail: ktlau@utem.edu.my [Faculty of Manufacturing Engineering, Universiti Teknikal Malaysia Melaka, Hang Tuah Jaya, 76100, Durian Tunggal, Melaka (Malaysia); Anand, T. Joseph Sahaya [Faculty of Manufacturing Engineering, Universiti Teknikal Malaysia Melaka, Hang Tuah Jaya, 76100, Durian Tunggal, Melaka (Malaysia); Sorrell, Charles C. [School of Materials Science and Engineering, UNSW Australia, Sydney, NSW 2052 (Australia)

2016-10-01

Proton activities of suspensions of Ti particles with added cationic polyelectrolyte as a function of acid additions have been investigated and compared in terms of the electrophoretic mobility and deposition yield. The proton activity in ethanol medium decreased with the addition of PEI polyelectrolyte and reduced further in the presence of Ti particles. The decrease in proton activity in the suspension indicates that protonation occurred on both the PEI molecules and Ti particles. It is proposed that the protonation of the amine groups of PEI and hydroxyl sites of Ti particle led to the formation of hydrogen bonding between the Ti particle and PEI molecules. Increase in the PEI and Ti with increasing acid addition translated to higher electrophoretic mobilities and deposition yield at low ranges of acetic acid addition (<0.75 vol%). - Highlights: • Protonation characteristics of polyelectrolytes and suspension particles are reported. • The protonation characteristics explained the electrophoretic mobility and yield results. • Adsorption mechanisms of protonated polyelectrolytes on the titanium particle is proposed. • Hydroxyl sites on the particles link the oxide particle and the polyelectrolyte molecules.

Protonation of the polyethyleneimine and titanium particles and their effect on the electrophoretic mobility and deposition

International Nuclear Information System (INIS)

Lau, Kok-Tee; Anand, T. Joseph Sahaya; Sorrell, Charles C.

2016-01-01

Proton activities of suspensions of Ti particles with added cationic polyelectrolyte as a function of acid additions have been investigated and compared in terms of the electrophoretic mobility and deposition yield. The proton activity in ethanol medium decreased with the addition of PEI polyelectrolyte and reduced further in the presence of Ti particles. The decrease in proton activity in the suspension indicates that protonation occurred on both the PEI molecules and Ti particles. It is proposed that the protonation of the amine groups of PEI and hydroxyl sites of Ti particle led to the formation of hydrogen bonding between the Ti particle and PEI molecules. Increase in the PEI and Ti with increasing acid addition translated to higher electrophoretic mobilities and deposition yield at low ranges of acetic acid addition (<0.75 vol%). - Highlights: • Protonation characteristics of polyelectrolytes and suspension particles are reported. • The protonation characteristics explained the electrophoretic mobility and yield results. • Adsorption mechanisms of protonated polyelectrolytes on the titanium particle is proposed. • Hydroxyl sites on the particles link the oxide particle and the polyelectrolyte molecules.

Polydopamine-coated open tubular column for the separation of proteins by capillary electrochromatography.

Science.gov (United States)

Xiao, Xing; Wang, Wentao; Chen, Jia; Jia, Li

2015-08-01

The separation and determination of proteins in food is an important aspect in food industry. Inspired by the self-polymerization of dopamine under alkaline conditions and the natural adhesive properties of polydopamine, in this paper, a simple and economical method was developed for the preparation of polydopamine-coated open tubular column, in which ammonium persulfate was used as the source of oxygen to induce and facilitate the polymerization of dopamine to form polydopamine. In comparison with a naked fused-silica capillary, the direction and magnitude of the electro-osmotic flow of the as-prepared polydopamine-coated open tubular column could be manipulated by varying the pH values of background solutions due to the existence of amine and phenolic hydroxyl groups on polydopamine coating. The surface morphology of the polydopamine-coated open tubular column was studied by scanning electron microscopy, and the thickness of polydopamine coating was 106 nm. The performance of the polydopamine-coated open tubular column was validated by analysis of proteins. The relative standard deviations of migration times of proteins representing run-to-run, day-to-day, and column-to-column were less than 3.5%. In addition, the feasibility of the polydopamine-coated open tubular column for real samples was verified by the separation of proteins in chicken egg white and pure milk. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Evaluation of a capillary zone electrophoresis system versus a conventional agarose gel system for routine serum protein separation and monoclonal component typing.

Science.gov (United States)

Roudiere, L; Boularan, A M; Bonardet, A; Vallat, C; Cristol, J P; Dupuy, A M

2006-01-01

Capillary zone electrophoresis of serum proteins is increasingly gaining impact in clinical laboratories. During 2003, we compared the fully automated capillary electrophoresis (CE) system from Beckman (Paragon CZE 2000) with the method agarose gel electrophoresis Sebia (Hydrasis-Hyris, AGE). This new study focused on the evaluation of analytical performance and a comparison including 115 fresh routine samples (group A) and a series of 97 frozen pathologic sera with suspicion of monoclonal protein (group B). Coefficients of variation (CVs %) for the five classical protein fractions have been reported to be consistenly serum samples (group B), there were 90 in which we detected a monoclonal protein by immunofixation (IF) (immunosubtraction (IS) was not used). AGE and Paragon 2000 failed to detect 7 and 12 monoclonal proteins, respectively, leading to a concordance to 92% for AGE and 87% for Paragon 2000 for identifying electrophoretic abnormalities in this group. Beta-globulin abnormalities and M paraprotein were well detected with Paragon 2000. Only 81% (21 vs 26) of the gammopathies were immunotyped with IS by two readers blinded to the IF immunotype. The Paragon 2000 is a reliable alternative to conventional agarose gel electrophoresis combining the advantages of full automation (rapidity, ease of use and cost) with high analytical performance. Qualified interpretation of results requires an adaptation period which could further improve concordance between the methods. Recently, this CE system has been improved by the manufacturer (Beckman) concerning the migration buffer and detection of beta-globulin abnormalities.

Sequence Dependent Electrophoretic Separations of DNA in Pluronic F127 Gels

Science.gov (United States)

You, Seungyong; van Winkle, David H.

2010-03-01

Two-dimensional (2-D) electrophoresis has successfully been used to visualize the separation of DNA fragments of the same length. We electrophorese a double-stranded DNA ladder in an Agarose gel for the first dimension and in gels of Pluronic F127 for the second dimension at room temperature. The 1000 bp band that travels together as a single band in an Agarose gel is split into two bands in Pluronic gels. The slower band follows the exponential decay trend that the other ladder constituents do. After sequencing the DNA fragments, the faster band has an apparently random sequence, while the slower band and the others have two A-tracts in each 250 bp segment. The A-tracts consist of a series of at least five adenine bases pairing with thymine bases. This result leads to the conclusion that the migration of the DNA molecules bent with A-tracts is more retarded in Pluronic gels than the wild-type of DNA molecules.

Separation of large DNA molecules by applying pulsed electric field to size exclusion chromatography-based microchip

Science.gov (United States)

Azuma, Naoki; Itoh, Shintaro; Fukuzawa, Kenji; Zhang, Hedong

2018-02-01

Through electrophoresis driven by a pulsed electric field, we succeeded in separating large DNA molecules with an electrophoretic microchip based on size exclusion chromatography (SEC), which was proposed in our previous study. The conditions of the pulsed electric field required to achieve the separation were determined by numerical analyses using our originally proposed separation model. From the numerical results, we succeeded in separating large DNA molecules (λ DNA and T4 DNA) within 1600 s, which was approximately half of that achieved under a direct electric field in our previous study. Our SEC-based electrophoresis microchip will be one of the effective tools to meet the growing demand of faster and more convenient separation of large DNA molecules, especially in the field of epidemiological research of infectious diseases.

Zirconium phosphate coating on aluminium foams by electrophoretic deposition for acidic catalysis

NARCIS (Netherlands)

Ordomskiy, V.; Schouten, J.C.; Schaaf, van der J.; Nijhuis, T.A.

2012-01-01

The electrophoretic deposition method has been applied for the formation of an amorphous zirconium phosphate layer on the surface of open-cell aluminum foam. The aluminum foam was fully and uniformly covered by the zirconium phosphate layer with a good mechanical adherence to the support. The

Separation and identification of Musa acuminate Colla (banana) leaf proteins by two-dimensional gel electrophoresis and mass spectrometry.

Science.gov (United States)

Lu, Y; Qi, Y X; Zhang, H; Zhang, H Q; Pu, J J; Xie, Y X

2013-12-19

To establish a proteomic reference map of Musa acuminate Colla (banana) leaf, we separated and identified leaf proteins using two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) and mass spectrometry (MS). Tryptic digests of 44 spots were subjected to peptide mass fingerprinting (PMF) by matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) MS. Three spots that were not identified by MALDI-TOF MS analysis were identified by searching against the NCBInr, SwissProt, and expressed sequence tag (EST) databases. We identified 41 unique proteins. The majority of the identified leaf proteins were found to be involved in energy metabolism. The results indicate that 2D-PAGE is a sensitive and powerful technique for the separation and identification of Musa leaf proteins. A summary of the identified proteins and their putative functions is discussed.

Sintering of MnCo2O4 coatings prepared by electrophoretic deposition

DEFF Research Database (Denmark)

Bobruk, M.; Molin, Sebastian; Chen, Ming

2018-01-01

Sintering of MnCo2O4 coatings prepared by electrophoretic deposition on steel substrates has been studied in air and in reducing-oxidizing atmosphere. Effect of temperature and pO2 on the resulting coating density was evaluated from scanning electron microscopy images of polished cross sections...

A Novel, In-solution Separation of Endogenous Cardiac Sarcomeric Proteins and Identification of Distinct Charged Variants of Regulatory Light Chain*

Science.gov (United States)

Scruggs, Sarah B.; Reisdorph, Rick; Armstrong, Mike L.; Warren, Chad M.; Reisdorph, Nichole; Solaro, R. John; Buttrick, Peter M.

2010-01-01

The molecular conformation of the cardiac myosin motor is modulated by intermolecular interactions among the heavy chain, the light chains, myosin binding protein-C, and titin and is governed by post-translational modifications (PTMs). In-gel digestion followed by LC/MS/MS has classically been applied to identify cardiac sarcomeric PTMs; however, this approach is limited by protein size, pI, and difficulties in peptide extraction. We report a solution-based work flow for global separation of endogenous cardiac sarcomeric proteins with a focus on the regulatory light chain (RLC) in which specific sites of phosphorylation have been unclear. Subcellular fractionation followed by OFFGEL electrophoresis resulted in isolation of endogenous charge variants of sarcomeric proteins, including regulatory and essential light chains, myosin heavy chain, and myosin-binding protein-C of the thick filament. Further purification of RLC using reverse-phase HPLC separation and UV detection enriched for RLC PTMs at the intact protein level and provided a stoichiometric and quantitative assessment of endogenous RLC charge variants. Digestion and subsequent LC/MS/MS unequivocally identified that the endogenous charge variants of cardiac RLC focused in unique OFFGEL electrophoresis fractions were unphosphorylated (78.8%), singly phosphorylated (18.1%), and doubly phosphorylated (3.1%) RLC. The novel aspects of this study are that 1) milligram amounts of endogenous cardiac sarcomeric subproteome were focused with resolution comparable with two-dimensional electrophoresis, 2) separation and quantification of post-translationally modified variants were achieved at the intact protein level, 3) separation of intact high molecular weight thick filament proteins was achieved in solution, and 4) endogenous charge variants of RLC were separated; a novel doubly phosphorylated form was identified in mouse, and singly phosphorylated, singly deamidated, and deamidated/phosphorylated forms were

Preparation and characterization of monodisperse large-porous silica microspheres as the matrix for protein separation.

Science.gov (United States)

Xia, Hongjun; Wan, Guangping; Zhao, Junlong; Liu, Jiawei; Bai, Quan

2016-11-04

High performance liquid chromatography (HPLC) is a kind of efficient separation technology and has been used widely in many fields. Micro-sized porous silica microspheres as the most popular matrix have been used for fast separation and analysis in HPLC. In this paper, the monodisperse large-porous silica microspheres with controllable size and structure were successfully synthesized with polymer microspheres as the templates and characterized. First, the poly(glycidyl methacrylate-co-ethyleneglycol dimethacrylate) microspheres (P GMA-EDMA ) were functionalized with tetraethylenepentamine (TEPA) to generate amino groups which act as a catalyst in hydrolysis of tetraethyl orthosilicate (TEOS) to form Si-containing low molecular weight species. Then the low molecular weight species diffused into the functionalized P GMA-EDMA microspheres by induction force of the amino groups to form polymer/silica hybrid microspheres. Finally, the organic polymer templates were removed by calcination, and the large-porous silica microspheres were obtained. The compositions, morphology, size distribution, specific surface area and pore size distribution of the porous silica microspheres were characterized by infrared analyzer, scanning-electron microscopy, dynamic laser scattering, the mercury intrusion method and thermal gravimetric analysis, respectively. The results show that the agglomeration of the hybrid microspheres can be overcome when the templates were functionalized with TEPA as amination reagent, and the yield of 95.7% of the monodisperse large-porous silica microspheres can be achieved with high concentration of polymer templates. The resulting large-porous silica microspheres were modified with octadecyltrichlorosilane (ODS) and the chromatographic evaluation was performed by separating the proteins and the digest of BSA. The baseline separation of seven kinds of protein standards was achieved, and the column delivered a better performance when separating BSA digests

Dephosphorylation of microtubule-binding sites at the neurofilament-H tail domain by alkaline, acid, and protein phosphatases.

Science.gov (United States)

Hisanaga, S; Yasugawa, S; Yamakawa, T; Miyamoto, E; Ikebe, M; Uchiyama, M; Kishimoto, T

1993-06-01

The dephosphorylation-induced interaction of neurofilaments (NFs) with microtubules (MTs) was investigated by using several phosphatases. Escherichia coli alkaline and wheat germ acid phosphatases increased the electrophoretic mobility of NF-H and NF-M by dephosphorylation, and induced the binding of NF-H to MTs. The binding of NFs to MTs was observed only after the electrophoretic mobility of NF-H approached the exhaustively dephosphorylated level when alkaline phosphatase was used. The number of phosphate remaining when NF-H began to bind to MTs was estimated by measuring phosphate bound to NF-H. NF-H did not bind to MTs even when about 40 phosphates from the total of 51 had been removed by alkaline phosphatase. The removal of 6 further phosphates finally resulted in the association of NF-H with MTs. A similar finding, that the restricted phosphorylation sites in the NF-H tail domain, but not the total amount of phosphates, were important for binding to MTs, was also obtained with acid phosphatases. In contrast to alkaline and acid phosphatases, four classes of protein phosphatases (protein phosphatases 1, 2A, 2B, and 2C) were ineffective for shifting the electrophoretic mobility of NF proteins and for inducing the association of NFs to MTs.

Analytical isotachophoresis

NARCIS (Netherlands)

Everaerts, F.M.

1977-01-01

Electrophoretic separation techniques nowadays seem to be linked mainly with protein chemistry and chromatography, as a result of the work of Hardy, Michaelis, Svedberg, Tiselius, Martin, Svensson, Ornstein and Davis, and many more names could be given. As capillary systems for electrophoresis have

Phosphorylation of formate dehydrogenase in potato tuber mitochondria

DEFF Research Database (Denmark)

Bykova, N.V.; Stensballe, A.; Egsgaard, H.

2003-01-01

Two highly phosphorylated proteins were detected after two-dimensional (blue native/SDS-PAGE) gel electrophoretic separation of the matrix fraction isolated from potato tuber mitochondria. These two phosphoproteins were identified by mass spectrometry as formate dehydrogenase (FDH) and the E1alpha...

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

Science.gov (United States)

Heda, Ghanshyam D; Omotola, Oluwabukola B; Heda, Rajiv P; Avery, Jamie

2016-09-01

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

Evaluation of capillary zone electrophoresis for the determination of protein composition in therapeutic immunoglobulins and human albumins.

Science.gov (United States)

Christians, Stefan; van Treel, Nadine Denise; Bieniara, Gabriele; Eulig-Wien, Annika; Hanschmann, Kay-Martin; Giess, Siegfried

2016-07-01

Capillary zone electrophoresis (CZE) provides an alternative means of separating native proteins on the basis of their inherent electrophoretic mobilities. The major advantage of CZE is the quantification by UV detection, circumventing the drawbacks of staining and densitometry in the case of gel electrophoresis methods. The data of this validation study showed that CZE is a reliable assay for the determination of protein composition in therapeutic preparations of human albumin and human polyclonal immunoglobulins. Data obtained by CZE are in line with "historical" data obtained by the compendial method, provided that peak integration is performed without time correction. The focus here was to establish a rapid and reliable test to substitute the current gel based zone electrophoresis techniques for the control of protein composition of human immunoglobulins or albumins in the European Pharmacopoeia. We believe that the more advanced and modern CZE method described here is a very good alternative to the procedures currently described in the relevant monographs. Copyright © 2016 International Alliance for Biological Standardization. Published by Elsevier Ltd. All rights reserved.

PLASMA ELECTROPHORETIC PROFILES IN THE EASTERN MASSASAUGA (SISTRURUS CATENATUS) AND INFLUENCES OF AGE, SEX, YEAR, LOCATION, AND SNAKE FUNGAL DISEASE.

Science.gov (United States)

Allender, Matthew C; Junge, Randall E; Baker-Wylie, Sarah; Hileman, Eric T; Faust, Lisa J; Cray, Carolyn

2015-12-01

The purpose of this study was to establish reference intervals of the protein electrophoretic fractions and the acute-phase proteins hemoglobin binding protein (as determined by the haptoglobin assay) and C-reactive protein (CRP) and assess any possible correlations between varying age class, sex, location (Illinois or Michigan), year, or presence of snake fungal disease (SFD). Banked plasma samples were assayed from 130 eastern massasaugas from 2009 to 2014 in Illinois and Michigan. Snakes from Michigan had higher total protein (mean: 5.50 g/dl), albumin/globulin ratio (0.42), albumin (1.59 g/dl), and gamma globulins (0.55 g/dl) than from snakes in Illinois (4.72 g/dl, 0.29, 1.03 g/dl, 0.38 g/dl, respectively). Snakes in Illinois (22.19 g/ml) had higher CRP than snakes in Michigan (10.89 mg/ml). Adults had higher gamma globulins (0.47 g/dl) than juveniles (0.28 g/dl). Males had higher alpha-2 globulins (0.98 g/dl) and CRP (21.4 mg/ml) than females (0.85, 11.6, respectively). There were no significant differences in absolute plasma proteins in SFD-positive snakes, but the percentage of gamma globulins was significantly higher in positive snakes. Future research in this area can now build on this data to determine changes in population health over time or due to specific environmental or disease threats.

LIMPIC: a computational method for the separation of protein MALDI-TOF-MS signals from noise

Directory of Open Access Journals (Sweden)

Di Nicola Marta

2007-03-01

Full Text Available Abstract Background Mass spectrometry protein profiling is a promising tool for biomarker discovery in clinical proteomics. However, the development of a reliable approach for the separation of protein signals from noise is required. In this paper, LIMPIC, a computational method for the detection of protein peaks from linear-mode MALDI-TOF data is proposed. LIMPIC is based on novel techniques for background noise reduction and baseline removal. Peak detection is performed considering the presence of a non-homogeneous noise level in the mass spectrum. A comparison of the peaks collected from multiple spectra is used to classify them on the basis of a detection rate parameter, and hence to separate the protein signals from other disturbances. Results LIMPIC preprocessing proves to be superior than other classical preprocessing techniques, allowing for a reliable decomposition of the background noise and the baseline drift from the MALDI-TOF mass spectra. It provides lower coefficient of variation associated with the peak intensity, improving the reliability of the information that can be extracted from single spectra. Our results show that LIMPIC peak-picking is effective even in low protein concentration regimes. The analytical comparison with commercial and freeware peak-picking algorithms demonstrates its superior performances in terms of sensitivity and specificity, both on in-vitro purified protein samples and human plasma samples. Conclusion The quantitative information on the peak intensity extracted with LIMPIC could be used for the recognition of significant protein profiles by means of advanced statistic tools: LIMPIC might be valuable in the perspective of biomarker discovery.

Development in electrophoresis: instrumentation for two-dimensional gel electrophoresis of protein separation and application of capillary electrophoresis in micro-bioanalysis

Energy Technology Data Exchange (ETDEWEB)

Xu, Aoshuang [Iowa State Univ., Ames, IA (United States)

2008-01-01

This dissertation begins with a general introduction of topics related to this work. The following chapters contain three scientific manuscripts, each presented in a separate chapter with accompanying tables, figures, and literature citations. The final chapter summarizes the work and provides some prospective on this work. This introduction starts with a brief treatment of the basic principles of electrophoresis separation, followed by a discussion of gel electrophoresis and particularly polyacrylamide gel electrophoresis for protein separation, a summary of common capillary electrophoresis separation modes, and a brief treatment of micro-bioanalysis application of capillary electrophoresis, and ends with an overview of protein conformation and dynamics.

Separation of ions in nanofluidic channels with combined pressure-driven and electro-osmotic flow.

Science.gov (United States)

Gillespie, Dirk; Pennathur, Sumita

2013-03-05

Separation of ionic species with the same electrophoretic mobility but different valence in electrolyte systems can occur within nanometer-scale channels with finite electrical double layers (EDLs). This is because EDL thicknesses are a significant fraction of slit height in such channels and can create transverse analyte concentration profiles that allow for unique separation modalities when combined with axial fluid flow. Previous work has shown such separation to occur using either pressure-driven flow or electro-osmotic flow separately. Here, we develop a Poisson-Boltzmann model to compare the separation of such ions using the combination of both pressure-driven and electro-osmotic flow. Applying a pressure gradient in the opposite direction of electro-osmotic flow can allow for zero or infinite retention of analyte species, which we investigate using three different wall boundary conditions. Furthermore, we determine conditions in fused silica nanochannels with which to generate optimal separation between two analytes of different charge but the same mobility. We also give simple rules of thumb to achieve the best separation efficacy in nanochannel systems.

Poly(norepinephrine)-coated open tubular column for the separation of proteins and recombination human erythropoietin by capillary electrochromatography.

Science.gov (United States)

Xiao, Xue; Zhang, Yamin; Wu, Jia; Jia, Li

2017-12-01

Recombinant human erythropoietin is an important therapeutic protein with high economic interest due to the benefits provided by its clinical use for the treatment of anemias associated with chronic renal failure and chemotherapy. In this work, a poly(norepinephrine)-coated open tubular column was successfully prepared based on the self-polymerization of norepinephrine under mild alkaline condition, the favorable film forming and easy adhesive properties of poly(norepinephrine). The poly(norepinephrine) coating was characterized by scanning electron microscopy and measurement of the electro-osmotic flow. The thickness of the coating was about 431 nm. The electrochromatographic performance of the poly(norepinephrine)-coated open tubular column was evaluated by separation of proteins. Some basic and acidic proteins including two variants of bovine serum albumin and two variants of β-lactoglobulin achieved separation in the poly(norepinephrine)-coated open tubular column. More importantly, the column demonstrated separation ability for the glycoforms of recombinant human erythropoietin. In addition, the column demonstrated good repeatability with the run-to-run, day-to-day, and column-to-column relative standard deviations of migration times of proteins less than 3.40%. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Electrophoretically applied dielectrics for amorphous metal foils used in pulsed power saturable reactors

International Nuclear Information System (INIS)

Sharp, D.J.; Harjes, H.C.; Mann, G.A.

1989-01-01

Amorphous metal foil-wound inductors have been tested as ferromagnetic saturable inductive elements for pulsed-power (multi-terawatt) switching modules in the inertial confinement fusion program at Sandia National Laboratories. In simulated capacitor testing premature dielectric breakdown of thin polyethylene terephthalate film insulation in the inductor windings occurs at considerably below 2500 V. This appears to be due to inadvertant dielectric damage from micro-spikes on the amorphous foil surface. Electron micrographs and dielectric breakdown data illustrate that electrophoretically-applied dielectric coatings, deposited from organic aqueous colloid dispersions, can be used to provide insulating coatings on the foil which provide a 240% improvement (6000 V) in the breakdown strength of wound amorphous foil inductors. The theory and operation of a dedicated electrophoretic continuous coating system is described. The machine was constructed and successfully applied for dielectric coating of amorphous metal foil. Additional possible applications exist for practical dielectric coating of metallic films or foils used in various commercial wound-type capacitor structures. 7 refs., 9 figs

Recent advances in capillary electrophoresis separation of monosaccharides, oligosaccharides, and polysaccharides.

Science.gov (United States)

Mantovani, Veronica; Galeotti, Fabio; Maccari, Francesca; Volpi, Nicola

2018-01-01

This article illustrates the basis and applications of methodologies for the analysis of simple and complex carbohydrates by means of CE. After a description of the most common and novel approaches useful for the analysis and characterization of carbohydrates, this review covers the recent advances in CE separation of monosaccharides, oligosaccharides, and polysaccharides. Various CE techniques are also illustrated for the study of carbohydrates derived from complex glyco-derivatives such as glycoproteins and glycolipids, essential for biopharmaceutical and glycoproteomics applications as well as for biomarker detection. Most glycans have no significant UV absorption, and derivatization with fluorophore groups prior to separation usually results in higher sensitivity and an improved electrophoretic profile. We also discuss the recent applications and separations by CE of derivatized simple and more complex carbohydrates with different chromophoric active tags. Overall, this review aims to give an overview of the most recent state-of-the-art techniques used in carbohydrate analysis by CE. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Chemical vapor deposition of aminopropyl silanes in microfluidic channels for highly efficient microchip capillary electrophoresis-electrospray ionization-mass spectrometry.

Science.gov (United States)

Batz, Nicholas G; Mellors, J Scott; Alarie, Jean Pierre; Ramsey, J Michael

2014-04-01

We describe a chemical vapor deposition (CVD) method for the surface modification of glass microfluidic devices designed to perform electrophoretic separations of cationic species. The microfluidic channel surfaces were modified using aminopropyl silane reagents. Coating homogeneity was inferred by precise measurement of the separation efficiency and electroosmotic mobility for multiple microfluidic devices. Devices coated with (3-aminopropyl)di-isopropylethoxysilane (APDIPES) yielded near diffusion-limited separations and exhibited little change in electroosmotic mobility between pH 2.8 and pH 7.5. We further evaluated the temporal stability of both APDIPES and (3-aminopropyl)triethoxysilane (APTES) coatings when stored for a total of 1 week under vacuum at 4 °C or filled with pH 2.8 background electrolyte at room temperature. Measurements of electroosmotic flow (EOF) and separation efficiency during this time confirmed that both coatings were stable under both conditions. Microfluidic devices with a 23 cm long, serpentine electrophoretic separation channel and integrated nanoelectrospray ionization emitter were CVD coated with APDIPES and used for capillary electrophoresis (CE)-electrospray ionization (ESI)-mass spectrometry (MS) of peptides and proteins. Peptide separations were fast and highly efficient, yielding theoretical plate counts over 600,000 and a peak capacity of 64 in less than 90 s. Intact protein separations using these devices yielded Gaussian peak profiles with separation efficiencies between 100,000 and 400,000 theoretical plates.

A substrate-optimized electrophoretic mobility shift assay for ADAM12

DEFF Research Database (Denmark)

Kotzsch, Alexander; Skovgaard, Tine; Buus, Uwe

2014-01-01

long been investigated as pharmaceutical drug targets; however, due to lack of potency and in vivo side effects, none of the small-molecule inhibitors discovered so far has made it beyond clinical testing. Ongoing research on novel selective inhibitors of ADAMs requires reliable biochemical assays...... to validate molecular probes from large-scale screening efforts. Here we describe an electrophoretic mobility shift assay for ADAM12 based on the identification of an optimized peptide substrate that is characterized by excellent performance and reproducibility....

ELECTROPHORETIC MOBILITIES OF ESCHERICHIA COLI 0157:H7 AND WILD-TYPE ESCHERICHIA COLI STRAINS

Science.gov (United States)

The electrophoretic mobility (EPM) of a number of human-virulent and "wild-type" Escherichia coli strains in phosphate buffered water was measured. The impact of pH, ionic strength, cation type (valence) and concentration, and bacterial strain on the EPM was investigated. Resul...

Serum protein inhibition of thyrotropin binding to human thyroid tissue

International Nuclear Information System (INIS)

Beall, G.N.; Chopra, I.J.; Solomon, D.H.; Kruger, S.R.

1978-01-01

We used a modificaton of the TSH radioreceptor assay to detect TSH-binding inhibition (TBI) activity in serum and serum fractions from normal subjects and patients with Graves' disease. TBI activity is present in normal IgG prepared by DEAE-Sephadex chromatography and in normal globulins prepared by precipitation at 1.6 M ammonium sulfate. Other normal serum proteins also had TBI activity when large concentrations were tested. Gel filtration chromatography and powder block electrophoresis were used to prepare fractions of normal and Graves' disease sera. In these fractions from normal serum, TBI activity was found in both γ-globulin and α-globulin-albumin fractions electrophoretically and in both 7S and 4S peaks from gel filtration. TBI activity from Graves' disease patients' sera was similarly distributed, but relatively more TBI accompanied the electrophoretic γ-globulins. Sepharose Protein-A and anti-IgG were used as immunoabsorbents to isolate and purify IgG from normal and Graves' disease sera. TBI activity in IgG was proportional to the IgG concentration, indicating that the TBI which migrates as a γ-globulin electrophoretically is an IgG and thus may possibly be an antibody. Inhibitory activity found in normal serum globulins and in the non-IgG fractions of both normal and abnormal sera seriously interferes with attempts to use the TSH radioreceptor assay to study the hypothesized anti-TSH receptor antibody in the serum of patients with Graves' disease

Study of the proteins in the defatted flour and protein concentrate of baru nuts (Dipteryx alata Vog

Directory of Open Access Journals (Sweden)

Rita de Cássia Avellaneda Guimarães

2012-09-01

Full Text Available Baru (Dipteryx alata Vog. is an abundant legume in the Brazilian Savanna. Its nuts can be exploited sustainably using its protein and lipid fractions. This study aimed to analyze the proteins of the nuts present in the defatted flour and protein concentrate in terms of their functional properties, the profile of their fractions, and the in vitro digestibility. The flour was defatted with hexane and extracted at the pH of higher protein solubility to obtain the protein concentrate. The electrophoretic profile of the protein fractions was evaluated in SDS-PAGE gel. The functional properties of the proteins indicate the possibility of their use in various foods, like soybeans providing water absorption capacity, oil absorption capacity, emulsifying properties, and foamability. Globulins, followed by the albumins, are the major fractions of the flour and protein concentrate, respectively. Digestibility was greater for the concentrate than for the defatted flour.

Analysis of proteins involved in biodegradation of crop biomass

Science.gov (United States)

Crawford, Kamau; Trotman, Audrey

1998-01-01

The biodegradation of crop biomass for re-use in crop production is part of the bioregenerative life support concept proposed by the National Aeronautics and Space Administration (NASA) for long duration, manned space exploration. The current research was conducted in the laboratory to evaluate the use of electrophoretic analysis as a means of rapidly assaying for constitutive and induced proteins associated with the bacterial degradation of crop residue. The proteins involved in crop biomass biodegradation are either constitutive or induced. As a result, effluent and cultures were examined to investigate the potential of using electrophoretic techniques as a means of monitoring the biodegradation process. Protein concentration for optimum banding patterns was determined using the Bio-Rad Protein Assay kit. Four bacterial soil isolates were obtained from the G.W. Carver research Farm at Tuskegee University and used in the decomposition of components of plant biomass. The culture, WDSt3A was inoculated into 500 mL of either Tryptic Soy Broth or Nutrient Broth. Incubation, with shaking of each flask was for 96 hours at 30 C. The cultures consistently gave unique banding patterns under denaturing protein electrophoresis conditions, The associated extracellular enzymes also yielded characteristic banding patterns over a 14-day period, when native electrophoresis techniques were used to examine effluent from batch culture bioreactors. The current study evaluated sample preparation and staining protocols to determine the ease of use, reproducibility and reliability, as well as the potential for automation.

Electrophoresis forum '80

International Nuclear Information System (INIS)

Radola, B.J.

1980-01-01

In this volume the contributions of the electrophoresis meeting are presented in a short term form. The main topics are gel-electrophoresis, ultra thin film isoelectric focusing, one- and two-dimensional electrophoresis, electrophoretical separation techniques, electric focusing (for phorensic studies), substrate free and substrate electrophoresis. In the poster session of this meeting subjects such as (ultra) thin film isoelectric focusing, identification of radioactive proteins, labelling of cell surfaces, autoradiography and 3 H-labelled proteins. Separate abstracts were prepared for 4 papers in this report. (HK) [de

Protein Attachment on Nanodiamonds.

Science.gov (United States)

Lin, Chung-Lun; Lin, Cheng-Huang; Chang, Huan-Cheng; Su, Meng-Chih

2015-07-16

A recent advance in nanotechnology is the scale-up production of small and nonaggregated diamond nanoparticles suitable for biological applications. Using detonation nanodiamonds (NDs) with an average diameter of ∼4 nm as the adsorbents, we have studied the static attachment of three proteins (myoglobin, bovine serum albumin, and insulin) onto the nanoparticles by optical spectroscopy, mass spectrometry, and dynamic light scattering, and electrophoretic zeta potential measurements. Results show that the protein surface coverage is predominantly determined by the competition between protein-protein and protein-ND interactions, giving each protein a unique and characteristic structural configuration in its own complex. Specifically, both myoglobin and bovine serum albumin show a Langmuir-type adsorption behavior, forming 1:1 complexes at saturation, whereas insulin folds into a tightly bound multimer before adsorption. The markedly different adsorption patterns appear to be independent of the protein concentration and are closely related to the affinity of the individual proteins for the NDs. The present study provides a fundamental understanding for the use of NDs as a platform for nanomedical drug delivery.

Electrophoretic comparison of nuclear and nucleolar proteins II. Rat pancreas

NARCIS (Netherlands)

Poort, C.

1961-01-01

The nuclei and nucleoli from rat pancreas were isolated and extracted successively with 0.14 M NaCl, 1 M NaCl, and 0.1 N NaOH. In the 0.14 M NaCl and 0.1 N NaOH extracts agar electrophoresis revealed differences between the nucleolar proteins and those from the non-nucleolar part of the nucleus.

Whey protein isolate gel for separation: A formation, characterization, and application study

Science.gov (United States)

Teo, Jiunn Yeong

performance of WPI monolith for continuous bed chromatography. Nonetheless the WPI monolith may be used to separate different protein molecules based on protein-protein interaction between WPI and protein molecules to be separated.

Two-dimensional analysis of human lymphocyte proteins. III. Preliminary report on a marker for the early detection and diagnosis of infectious mononucleosis

International Nuclear Information System (INIS)

Willard, K.E.

1982-01-01

Two-dimensional gel electrophoretic patterns of human peripheral blood leukocytes from 12 patients with infectious mononucleosis were prepared by use of the ISO-DALT system. Before the two-dimensional separation, the leukocytes were purified by Ficoll-Paque gradient centrifugation and labeled overnight with [ 35 S] methionine. Quantitative increases in two proteins were detected in the patterns of infected leukocytes from the patients as compared with controls. Fluorescence-activated cell sorting of leukocytes from normal human peripheral blood before subsequent two-dimensional gel analysis revealed that the dramatic increase in one of these proteins (Inmono:2) could be due to shifts in the population ratios of lymphocytes, monocytes, and granulocytes. In contrast, the appearance in the infected leukocytes of a second protein, Inmono:1, could not be accounted for by cell-population shifts. Increased amounts of these two proteins have been found in every patient studied who had clinically detectable infectious mononucleosis. In addition, a patient who displayed symptoms of infectious mononucleosis but who did not have a positive result in the MONOSPOT test (Ortho) until three weeks after our analysis also demonstrated increased relative amounts of these proteins in his leukocyte pattern

Formation of a covalent complex between the terminal protein of pneumococcal bacteriophage Cp-1 and 5'-dAMP

International Nuclear Information System (INIS)

Garcia, P.; Hermoso, J.M.; Garcia, J.A.; Garcia, E.; Lopez, R.; Salas, M.

1986-01-01

Incubation of extracts of Cp-1-infected Streptococcus pneumoniae with [α- 32 P]dATP produced a labeled protein with the electrophoretic mobility of the Cp-1 terminal protein. The reaction product was resistant to treatment with micrococcal nuclease and sensitive to treatment with proteinase K. Incubation of the 32 P-labeled protein with 5 M piperidine for 4 h at 50 0 C released 5'-dAMP, indicating that a covalent complex between the terminal protein and 5'-dAMP was formed in vitro. When the four deoxynucleoside triphosphates were included in the reaction mixture, a labeled complex of slower electrophoretic mobility in sodium dodecyl sulfate-polyacrylamide gels than the terminal protein-dAMP complex was also found, indicating that the Cp-1 terminal protein-dAMP complex can be elongated and, therefore, that it is an initiation complex. Treatment of the 32 P-labeled terminal protein-dAMP complex with 5.8 M HCl at 110 0 C for 2 h yielded phosphothreonine. These results, together with the resistance of the terminal protein-DNA linkage to hydroxylamine, suggest that the Cp-1 terminal protein is covalently linked to the DNA through a phosphoester bond between L-threonine and 5'-dAMP, namely, a O-5'-deoxyadenylyl-L-threonine bond

Electrophoretic deposition of carbon nanotubes on a carbon fiber surface with different index graphitization

International Nuclear Information System (INIS)

Almeida, E.C.; Baldan, M.R.; Ferreira, N.G.; Edwards, E.R.

2009-01-01

Full text: The purpose of this work is to examine the electrophoretic deposition of carbon nanotubes powder on carbon fibers, produced at different heat treatments temperatures. Besides, a systematic study of the effects of graphitization index from substrate on the structure and morphology of CNTs has been available. Carbon fibers were produced from polyacrylonitrile at three different heat treatments temperatures, 1000, 1500 and 2000 deg C. The carbon fibers microstructure or its graphitization index may be controlled by the heat treatments temperatures. The electrophoretic deposition of carbon nanotubes was obtained with the powder of carbon nanotubes dispersed in water by ultrasonication to obtain dispersions of 0.05 mg/mL. The carbon fibers were immersed in the nanotube dispersion, and a positive potential of 10 V/cm was applied. Morphology and microstructure of carbon nanotubes on carbon fibers were obtained by scanning electron microscopy, Raman spectroscopy and X-ray photoelectron spectroscopy. (author)

Principles of Micellar Electrokinetic Capillary Chromatography Applied in Pharmaceutical Analysis

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Árpád Gyéresi

2013-02-01

Full Text Available Since its introduction capillary electrophoresis has shown great potential in areas where electrophoretic techniques have rarely been used before, including here the analysis of pharmaceutical substances. The large majority of pharmaceutical substances are neutral from electrophoretic point of view, consequently separations by the classic capillary zone electrophoresis; where separation is based on the differences between the own electrophoretic mobilities of the analytes; are hard to achieve. Micellar electrokinetic capillary chromatography, a hybrid method that combines chromatographic and electrophoretic separation principles, extends the applicability of capillary electrophoretic methods to neutral analytes. In micellar electrokinetic capillary chromatography, surfactants are added to the buffer solution in concentration above their critical micellar concentrations, consequently micelles are formed; micelles that undergo electrophoretic migration like any other charged particle. The separation is based on the differential partitioning of an analyte between the two-phase system: the mobile aqueous phase and micellar pseudostationary phase. The present paper aims to summarize the basic aspects regarding separation principles and practical applications of micellar electrokinetic capillary chromatography, with particular attention to those relevant in pharmaceutical analysis.

Using Biomolecules to Separate Plutonium

Science.gov (United States)

Gogolski, Jarrod

Used nuclear fuel has traditionally been treated through chemical separations of the radionuclides for recycle or disposal. This research considers a biological approach to such separations based on a series of complex and interdependent interactions that occur naturally in the human body with plutonium. These biological interactions are mediated by the proteins serum transferrin and the transferrin receptor. Transferrin to plutonium in vivo and can deposit plutonium into cells after interacting with the transferrin receptor protein at the cell surface. Using cerium as a non-radioactive surrogate for plutonium, it was found that cerium(IV) required multiple synergistic anions to bind in the N-lobe of the bilobal transferrin protein, creating a conformation of the cerium-loaded protein that would be unable to interact with the transferrin receptor protein to achieve a separation. The behavior of cerium binding to transferrin has contributed to understanding how plutonium(IV)-transferrin interacts in vivo and in biological separations.

Variations in virulence between different electrophoretic types of Listeria monocytogenes

DEFF Research Database (Denmark)

Nørrung, Birgit; Andersen, Jens Kirk

2000-01-01

A total of 245 strains of Listeria monocytogenes, representing 33 different electrophoretic types (ETs), were examined quantitatively for haemolytic activity. No significant difference was observed in the mean haemolytic activity between different ETs. Eighty four out of 91 strains examined were...... compared with 3.64 among food isolates). The explanation for this may be that more virulent strains are more prone to cause human infection. It is, however, also possible that strains oft. monocytogenes may become more virulent while multiplying in a living organism compared with multiplying in foods....

Electrophoretic demonstration of glycoproteins, lipoproteins, and phosphoproteins in human and bovine enamel

DEFF Research Database (Denmark)

Kirkeby, S; Moe, D; Bøg-Hansen, T C

1990-01-01

Enamel proteins from fully mineralized human molars and from bovine tooth germs were separated by electrophoresis. The gels were stained for detection of glycoproteins, lipoproteins, and phosphoproteins. Glycoproteins were shown by periodic acid-Schiff staining and lectin blotting. In mature human...... enamel a number of high molecular weight proteins could be demonstrated after ethylenediaminetetra-acetic acid demineralization and subsequent Triton X-100 extraction. These proteins are suggested to be lipoproteins. Phosphoproteins could only be visualized in enamel matrix from the tooth germs....

[Identification and characterization of proteins from human bronchial secretion (author's transl)].

Science.gov (United States)

Laine, A; Hayem, A

1976-03-01

An analysis of bronchial mucus proteins was carried out by crossed immunoelectrophoresis. Before electrophoretic migration, sputum was treated with Ecteola-cellulose, which retains acid mucins. The proteins were then extracted by a phosphate/saline buffer pH 7.5. Crossed immunoelectrophoresis of the "bronchial extracts" was carried out with an anti-human serum: fifteen proteins were detected. Among them, IgA and protease inhibitiors play an important role in bronchial pathology. Bronchial extracts were also studied with immune serums against milk proteins, whole saliva and proteins of bronchial mucus. Bronchotransferrin, amylase and two esterases were characterized. Four other proteins were also detected with immune serums against bronchial mucus-proteins: their biological role is still unknown.

Innovations in electrophoretic deposition: Alternating current and pulsed direct current methods

International Nuclear Information System (INIS)

Chávez-Valdez, Alejandra; Boccaccini, Aldo R.

2012-01-01

This review summarizes emerging developments in the field of alternating current (AC) and pulsed direct current (DC) electrophoretic deposition (EPD) in aqueous or organic media. Numerous applications of AC-EPD are discussed including two major groups of investigations: (i) AC-EPD to suppress water hydrolysis at high voltages in inorganic (ceramic) coatings and (ii) AC-EPD for deposition of biological entities. The deposition, purification and manipulation of carbon nanotubes and nanoparticles by AC-EPD to form specific arrays, for development of sensors and other electronic devices and the application of AC-EPD as method for separation of particles according to their shape or size are also presented. Other applications reviewed relate to the fabrication by AC-EPD of toxic gas sensors from oxides and superconducting layers. The main materials being examined by AC-EPD are inorganic, including carbon nanotubes, TiO 2 nanoparticles, Al 2 O 3 , Si, SnO 2 , ZnO and WO 3 and biological entities, e.g. bacteria cells. For pulsed EPD, the applications reviewed are divided in pulsed current and pulsed voltage EPD. Among the applications of pulsed EPD, the formation of thick films from aqueous suspensions without water decomposition, the fabrication of multilayer and composite materials and the size-selective deposition of ceramic nanoparticles are the most important investigated to date, based on the quality of the coatings and deposits obtained and their relevance for applications.

Solid oxide fuel cell electrolytes produced via very low pressure suspension plasma spray and electrophoretic deposition

Science.gov (United States)

Fleetwood, James D.

Solid oxide fuel cells (SOFCs) are a promising element of comprehensive energy policies due to their direct mechanism for converting the oxidization of fuel, such as hydrogen, into electrical energy. Both very low pressure plasma spray and electrophoretic deposition allow working with high melting temperature SOFC suspension based feedstock on complex surfaces, such as in non-planar SOFC designs. Dense, thin electrolytes of ideal composition for SOFCs can be fabricated with each of these processes, while compositional control is achieved with dissolved dopant compounds that are incorporated into the coating during deposition. In the work reported, sub-micron 8 mole % Y2O3-ZrO2 (YSZ) and gadolinia-doped ceria (GDC), powders, including those in suspension with scandium-nitrate dopants, were deposited on NiO-YSZ anodes, via very low pressure suspension plasma spray (VLPSPS) at Sandia National Laboratories' Thermal Spray Research Laboratory and electrophoretic deposition (EPD) at Purdue University. Plasma spray was carried out in a chamber held at 320 - 1300 Pa, with the plasma composed of argon, hydrogen, and helium. EPD was characterized utilizing constant current deposition at 10 mm electrode separation, with deposits sintered from 1300 -- 1500 °C for 2 hours. The role of suspension constituents in EPD was analyzed based on a parametric study of powder loading, powder specific surface area, polyvinyl butyral (PVB) content, polyethyleneimine (PEI) content, and acetic acid content. Increasing PVB content and reduction of particle specific surface area were found to eliminate the formation of cracks when drying. PEI and acetic acid content were used to control suspension stability and the adhesion of deposits. Additionally, EPD was used to fabricate YSZ/GDC bilayer electrolyte systems. The resultant YSZ electrolytes were 2-27 microns thick and up to 97% dense. Electrolyte performance as part of a SOFC system with screen printed LSCF cathodes was evaluated with peak

Fabrication of micro-dot arrays and micro-walls of acrylic acid/melamine resin on aluminum by AFM probe processing and electrophoretic coating

International Nuclear Information System (INIS)

Kurokawa, S.; Kikuchi, T.; Sakairi, M.; Takahashi, H.

2008-01-01

Micro-dot arrays and micro-walls of acrylic acid/melamine resin were fabricated on aluminum by anodizing, atomic force microscope (AFM) probe processing, and electrophoretic deposition. Barrier type anodic oxide films of 15 nm thickness were formed on aluminum and then the specimen was scratched with an AFM probe in a solution containing acrylic acid/melamine resin nano-particles to remove the anodic oxide film locally. After scratching, the specimen was anodically polarized to deposit acrylic acid/melamine resin electrophoretically at the film-removed area. The resin deposited on the specimen was finally cured by heating. It was found that scratching with the AFM probe on open circuit leads to the contamination of the probe with resin, due to positive shifts in the potential during scratching. Scratching of the specimen under potentiostatic conditions at -1.0 V, however, resulted in successful resin deposition at the film-removed area without probe contamination. The rate of resin deposition increased as the specimen potential becomes more positive during electrophoretic deposition. Arrays of resin dots with a few to several tens μm diameter and 100-1000 nm height, and resin walls with 100-1000 nm height and 1 μm width were obtained on specimens by successive anodizing, probe processing, and electrophoretic deposition

Fabrication of micro-dot arrays and micro-walls of acrylic acid/melamine resin on aluminum by AFM probe processing and electrophoretic coating

Energy Technology Data Exchange (ETDEWEB)

Kurokawa, S.; Kikuchi, T.; Sakairi, M. [Graduate School of Engineering, Hokkaido University, N-13, W-8, Kita-Ku, Sapporo 060-8628 (Japan); Takahashi, H. [Graduate School of Engineering, Hokkaido University, N-13, W-8, Kita-Ku, Sapporo 060-8628 (Japan)], E-mail: takahasi@elechem1-mc.eng.hokudai.ac.jp

2008-11-30

Micro-dot arrays and micro-walls of acrylic acid/melamine resin were fabricated on aluminum by anodizing, atomic force microscope (AFM) probe processing, and electrophoretic deposition. Barrier type anodic oxide films of 15 nm thickness were formed on aluminum and then the specimen was scratched with an AFM probe in a solution containing acrylic acid/melamine resin nano-particles to remove the anodic oxide film locally. After scratching, the specimen was anodically polarized to deposit acrylic acid/melamine resin electrophoretically at the film-removed area. The resin deposited on the specimen was finally cured by heating. It was found that scratching with the AFM probe on open circuit leads to the contamination of the probe with resin, due to positive shifts in the potential during scratching. Scratching of the specimen under potentiostatic conditions at -1.0 V, however, resulted in successful resin deposition at the film-removed area without probe contamination. The rate of resin deposition increased as the specimen potential becomes more positive during electrophoretic deposition. Arrays of resin dots with a few to several tens {mu}m diameter and 100-1000 nm height, and resin walls with 100-1000 nm height and 1 {mu}m width were obtained on specimens by successive anodizing, probe processing, and electrophoretic deposition.

Simultaneous pre-concentration and separation on simple paper-based analytical device for protein analysis.

Science.gov (United States)

Niu, Ji-Cheng; Zhou, Ting; Niu, Li-Li; Xie, Zhen-Sheng; Fang, Fang; Yang, Fu-Quan; Wu, Zhi-Yong

2018-02-01

In this work, fast isoelectric focusing (IEF) was successfully implemented on an open paper fluidic channel for simultaneous concentration and separation of proteins from complex matrix. With this simple device, IEF can be finished in 10 min with a resolution of 0.03 pH units and concentration factor of 10, as estimated by color model proteins by smartphone-based colorimetric detection. Fast detection of albumin from human serum and glycated hemoglobin (HBA1c) from blood cell was demonstrated. In addition, off-line identification of the model proteins from the IEF fractions with matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF-MS) was also shown. This PAD IEF is potentially useful either for point of care test (POCT) or biomarker analysis as a cost-effective sample pretreatment method.

Effect of acid- and alkaline-aided extractions on functional and rheological properties of proteins recovered from mechanically separated turkey meat (MSTM).

Science.gov (United States)

Hrynets, Yuliya; Omana, Dileep A; Xu, Yan; Betti, Mirko

2010-09-01

Functional and rheological characteristics of acid- and alkali-extracted proteins from mechanically separated turkey meat (MSTM) have been investigated. Extractions were carried out at 4 pH values (2.5, 3.5, 10.5, and 11.5). The study demonstrated that alkali and acid extractions resulted in significant (P hardness, chewiness, springiness, and cohesiveness) of recovered proteins were found to be unaffected (P > 0.05) by different extraction pH. The protein extracted at pH 3.5 formed a highly viscoelastic gel network as evidenced by storage modulus (G') values, whereas the gel formed from proteins extracted at pH 10.5 was found to be the weakest. The work also revealed that acid treatments were more effective for removal of total heme pigments from MSTM. Color characteristics of protein isolates were markedly improved compared to the initial material and tended to be better when subjected to acid extractions. Mechanically separated meat is one of the cheapest sources of protein obtained by grinding meat and bones together and forcing the mixture through a perforated drum. The use of mechanically separated turkey meat (MSTM) for the production of further processed poultry products is limited due to its undesirable color and textural properties. Recovery of proteins from MSTM using pH shifting process will help the poultry processors to get better returns and also create opportunity to produce functional food ingredients.

Measurements of the electrophoretic mobility with a new laser Doppler cytopherometer (Lazypher) and critical evaluation of the electrophorese mobility-test (EMT)

International Nuclear Information System (INIS)

Hoffmann, W.

1982-01-01

The new developed Laser Doppler Cytopherometer (Lazypher) allows the exact and objective measurement of the electrophoretic mobility of particles. Comparative experiments with the Free Flow Cell Electrophoresis instrument of Hannig showed identical results. The impression that the electrophoretic Mobility Test (EMT) is not valid for cancer diagnosis has been substantiated. But in its present form with the new instrument (Lazypher) possible improvements, e.g. isolation of lymphocytes, purification of antigens or indicator particles, can be estimated objectively for their value for the test system. (orig.) [de

Comparative evaluation of hmopolesis and serum proteins from dogs with transmissible venereal tumor by natural ocorrence and induced by alogenic transplant

International Nuclear Information System (INIS)

Aptekmann, K.P.; Costa, M.T.; Fabeni, R. de C.; Santana, A.E.

2005-01-01

The objective of this study was to evaluate the haemopoiesis and total serum proteins of dogs with canine Transmissible Venereal Tumour (TVT) (naturally occurring or induced by allogenic transplant). Complete haemogram, bone marrow evaluation, total serum protein, albumin determination and electrophoretic fractionation of serum proteins were performed. Results revealed normocytic normochromic anaemia and thrombocytopenia in dogs with naturally occurring TVT

Non-aqueous capillary electrophoretic separation of cholesterol and 25-hydroxycholesterol after derivatization with Girard P reagent

Czech Academy of Sciences Publication Activity Database

Greguš, M.; Roberg-Larsen, H.; Lundanes, E.; Foret, František; Kubáň, Petr; Wilson, S.R.

2017-01-01

Roč. 207, OCT (2017), s. 87-91 ISSN 0009-3084 Institutional support: RVO:68081715 Keywords : cholesterol * 25-hydroxycholesterol * UV detection Subject RIV: CB - Analytical Chemistry, Separation OBOR OECD: Analytical chemistry Impact factor: 3.361, year: 2016

Characterization on the electrophoretic deposition of the 8 mol% yttria-stabilized zirconia nanocrystallites prepared by a sol-gel process

Energy Technology Data Exchange (ETDEWEB)

Lee, Y.-H. [Department of Materials Science and Engineering, National Cheng Kung University, 1 Ta-Hsueh Road, Tainan 70101, Taiwan (China); Kuo, C.-W. [Department of Resources Engineering, National Cheng Kung University, 1 Ta-Hsueh Road, Tainan 70101, Taiwan (China); Shih, C.-J. [Faculty of Fragrance and Cosmetics, Kaohsiung Medical University, 100 Shi-Chuan 1st Road, Kaohsiung 807, Taiwan (China); Hung, I-M. [Department of Materials Science and Engineering, National Cheng Kung University, 1 Ta-Hsueh Road, Tainan 70101, Taiwan (China); Fung, K.-Z. [Department of Materials Science and Engineering, National Cheng Kung University, 1 Ta-Hsueh Road, Tainan 70101, Taiwan (China); Wen, S.-B. [Department of Resources Engineering, National Cheng Kung University, 1 Ta-Hsueh Road, Tainan 70101, Taiwan (China); Wang, M.-C. [Faculty of Fragrance and Cosmetics, Kaohsiung Medical University, 100 Shi-Chuan 1st Road, Kaohsiung 807, Taiwan (China)]. E-mail: cjshih@kmu.edu.tw

2007-02-15

An 8 mol% yttria-stabilized zirconia (8YSZ) films are electrophoretically deposited on the La{sub 0.8}Sr{sub 0.2}MnO{sub 3} substrate using 8YSZ nanocrystallites prepared by a sol-gel process. Effects of liquid suspension on the particle zeta potential and degree of agglomeration at different pH values are investigated. When the pH value deviates from the point of zero charge (PZC), the adsorption of protons on particle surfaces cause higher zeta potential and well-dispersed suspension. The optimal values of the iodine concentration, applied voltage and deposition time for the electrophoretic deposition of 8YSZ films are also found.

Cellular distribution, purification and electrophoretic properties of malate dehydrogenase in Trichuris ovis and inhibition by benzimidazoles and pyrimidine derivatives.

Science.gov (United States)

Sanchez-Moreno, M; Ortega, J E; Valero, A

1989-12-01

High levels of malate dehydrogenase were found in Trichuris ovis. Two molecular forms of the enzyme, of different cellular location and electrophoretic pattern, were isolated and purified. The activity of soluble malate dehydrogenase was greater than that of mitochondrial malate dehydrogenase. Both forms also displayed different electrophoretic profiles in comparison with purified extracts from goat (Capra hircus) liver. Substrate concentration directly affected enzyme activity. Host and parasite malate dehydrogenase activity were both inhibited by a series of benzimidazoles and pyrimidine-derived compounds, some of which markedly reduced parasite enzyme activity, but not host enzyme activity. Percentage inhibition by some pyrimidine derivatives was greater than that produced by benzimidazoles.

Thickness control in electrophoretic deposition of WO3 nanofiber thin films for solar water splitting

International Nuclear Information System (INIS)

Fang, Yuanxing; Lee, Wei Cheat; Canciani, Giacomo E.; Draper, Thomas C.; Al-Bawi, Zainab F.; Bedi, Jasbir S.; Perry, Christopher C.; Chen, Qiao

2015-01-01

Graphical abstract: - Highlights: • A novel method combining electrospinning and electrophoretic deposition was established for the creation of nanostructured semiconductor thin films. • The created thin films displayed a high chemical stability with a controllable thickness. • The PEC water splitting performance of the thin films was optimized by fine-tuning the thickness of the films. • A maximum photoconversion efficiency was achieved by 18 μm nanofibrous thin films. - Abstract: Electrophoretic deposition (EPD) of ground electrospun WO 3 nanofibers was applied to create photoanodes with controlled morphology for the application of photoelectrochemical (PEC) water splitting. The correlations between deposition parameters and film thicknesses were investigated with theoretical models to precisely control the morphology of the nanostructured porous thin film. The photoconversion efficiency was further optimized as a function of film thickness. A maximum photoconversion efficiency of 0.924% from electrospun WO 3 nanofibers that EPD deposited on a substrate was achieved at a film thickness of 18 μm.

Fate of egg proteins during the development of Columba livia domestica embryo.

Science.gov (United States)

Shbailat, Seba Jamal; Aslan, Ibtisam Omar

2018-01-01

The transfer of egg white into the yolk and consumption of yolk proteins by the embryo are largely unexplored in the pigeon Columba livia domestica. Here, we investigated the route of egg white transfer as well as the degradation and uptake of yolk proteins by the pigeon embryo. Initially, we tested the electrophoretic patterns of proteins in different egg compartments throughout development. Then, we used lysozyme as a reference protein to follow the egg white transfer, and we measured its activity using Micrococcus lysodeikticus as a substrate. Moreover, we determined the general protease activity during different developmental stages in the yolk using casein. Finally, we examined the expression of aminopeptidase-N (APN) and oligopeptide transporter PepT1 genes in the yolk sac membrane (YSM) from incubation day 8 until day 17. Several electrophoretic bands of presumptive egg white proteins appeared in different egg compartments. Also, lysozyme activity was detected chronologically in the egg compartments. It appeared on day 12 in the amniotic and intestinal fluids and on day 14 in the yolk. Moreover, protease activity in the yolk increased significantly on day 14 and thereafter. APN expression was largest on day 8 and reduced generally afterward, whereas PepT1 expression peaked between days 13 and 15 but then reduced substantially. Our results suggest that the egg white proteins move through the amnion and intestine into the yolk where they undergo degradation by the activated proteases. Furthermore, the YSM appears to have a role in protein consumption, and this role decreases toward hatch. © 2018 Wiley Periodicals, Inc.

Fractionation separation of human plasma proteins using HPLC with a homemade iron porphyrin based monolithic column.

Science.gov (United States)

Zhang, Doudou; Zhao, Yu; Lan, Dandan; Pang, Xiaomin; Bai, Ligai; Liu, Haiyan; Yan, Hongyuan

2017-11-15

In this work a polymer monolithic column was fabricated within the confines of a stainless steel column (50×4.6mm i.d.) via radical polymerization by using iron porphyrin and butyl methacrylate as co-monomers, ethylene glycol dimethacrylate as crosslinking agent, ethylene glycol, isopropyl alcohol and N, N-dimethylformamide as tri-porogens, benzoyl peroxide and N,N-dimethylaniline as initiators. The resulting monolithic column was characterized by elemental analysis, scanning electron microscopy, nitrogen adsorption BET surface area, and mercury intrusion porosimetry, respectively. Results showed that the homemade monolith occupied relatively uniform pore structure, low back pressure, and enhanced selectivity for proteins in complex bio-samples. The present work described a simple and efficient method for "fractionation separation" of human plasma proteins, and it is a promising separation method for complex bio-samples in proteomic research. Copyright © 2017 Elsevier B.V. All rights reserved.

Enhanced separation of membranes during free flow zonal electrophoresis in plants.

Science.gov (United States)

Barkla, Bronwyn J; Vera-Estrella, Rosario; Pantoja, Omar

2007-07-15

Free flow zonal electrophoresis (FFZE) is a versatile technique that allows for the separation of cells, organelles, membranes, and proteins based on net surface charge during laminar flow through a thin aqueous layer. We have been optimizing the FFZE technique to enhance separation of plant vacuolar membranes (tonoplast) from other endomembranes to pursue a directed proteomics approach to identify novel tonoplast transporters. Addition of ATP to a mixture of endomembranes selectively enhanced electrophoretic mobility of acidic vesicular compartments during FFZE toward the positive electrode. This has been attributed to activation of the V-ATPase generating a more negative membrane potential outside the vesicles, resulting in enhanced migration of acidic vesicles, including tonoplast, to the anode (Morré, D. J.; Lawrence, J.; Safranski, K.; Hammond, T.; Morré, D. M. J. Chromatogr., A 1994, 668, 201-213). We confirm that ATP does induce a redistribution of membranes during FFZE of microsomal membranes isolated from several plant species, including Arabidopsis thaliana, Thellungiella halophila, Mesembryanthemum crystallinum, and Ananas comosus. However, we demonstrate, using V-ATPase-specific inhibitors, nonhydrolyzable ATP analogs, and ionophores to dissipate membrane potential, that the ATP-dependent migrational shift of membranes under FFZE is not due to activation of the V-ATPase. Addition of EDTA to chelate Mg2+, leading to the production of the tetravalent anionic form of ATP, resulted in a further enhancement of membrane migration toward the anode, and manipulation of cell surface charge by addition of polycations also influenced the ATP-dependent migration of membranes. We propose that ATP enhances the mobility of endomembranes by screening positive surface charges on the membrane surface.

Automated Parallel Capillary Electrophoretic System

Science.gov (United States)

Li, Qingbo; Kane, Thomas E.; Liu, Changsheng; Sonnenschein, Bernard; Sharer, Michael V.; Kernan, John R.

2000-02-22

An automated electrophoretic system is disclosed. The system employs a capillary cartridge having a plurality of capillary tubes. The cartridge has a first array of capillary ends projecting from one side of a plate. The first array of capillary ends are spaced apart in substantially the same manner as the wells of a microtitre tray of standard size. This allows one to simultaneously perform capillary electrophoresis on samples present in each of the wells of the tray. The system includes a stacked, dual carousel arrangement to eliminate cross-contamination resulting from reuse of the same buffer tray on consecutive executions from electrophoresis. The system also has a gel delivery module containing a gel syringe/a stepper motor or a high pressure chamber with a pump to quickly and uniformly deliver gel through the capillary tubes. The system further includes a multi-wavelength beam generator to generate a laser beam which produces a beam with a wide range of wavelengths. An off-line capillary reconditioner thoroughly cleans a capillary cartridge to enable simultaneous execution of electrophoresis with another capillary cartridge. The streamlined nature of the off-line capillary reconditioner offers the advantage of increased system throughput with a minimal increase in system cost.

Synthesis and Application of Carbon–Iron Oxide Microspheres’ Black Pigments in Electrophoretic Displays

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Meng Xianwei

2010-01-01

Full Text Available Abstract Carbon–iron oxide microspheres’ black pigments (CIOMBs had been prepared via ultrasonic spray pyrolysis of aqueous solutions containing ferrous chloride and glucose. Due to the presence of carbon, CIOMBs not only exhibited remarkably acid resistance, but also could be well dispersed in both polar solvents and nonpolar solvent. Finally, dispersions of hollow CIOMBs in tetrachloroethylene had successfully been applied in electrophoretic displays.

Electrophoretic Deposition as a New Bioactive Glass Coating Process for Orthodontic Stainless Steel

OpenAIRE

Kyotaro Kawaguchi; Masahiro Iijima; Kazuhiko Endo; Itaru Mizoguchi

2017-01-01

This study investigated the surface modification of orthodontic stainless steel using electrophoretic deposition (EPD) of bioactive glass (BG). The BG coatings were characterized by spectrophotometry, scanning electron microscopy with energy dispersive X-ray spectrometry, and X-ray diffraction. The frictional properties were investigated using a progressive load scratch test. The remineralization ability of the etched dental enamel was studied according to the time-dependent mechanical proper...

The laboratory technology of discrete molecular separation: the historical development of gel electrophoresis and the material epistemology of biomolecular science, 1945-1970.

Science.gov (United States)

Chiang, Howard Hsueh-hao

2009-01-01

Preparative and analytical methods developed by separation scientists have played an important role in the history of molecular biology. One such early method is gel electrophoresis, a technique that uses various types of gel as its supporting medium to separate charged molecules based on size and other properties. Historians of science, however, have only recently begun to pay closer attention to this material epistemological dimension of biomolecular science. This paper substantiates the historiographical thread that explores the relationship between modern laboratory practice and the production of scientific knowledge. It traces the historical development of gel electrophoresis from the mid-1940s to the mid-1960s, with careful attention to the interplay between technical developments and disciplinary shifts, especially the rise of molecular biology in this time-frame. Claiming that the early 1950s marked a decisive shift in the evolution of electrophoretic methods from moving boundary to zone electrophoresis, I reconstruct various trajectories in which scientists such as Oliver Smithies sought out the most desirable solid supporting medium for electrophoretic instrumentation. Biomolecular knowledge, I argue, emerged in part from this process of seeking the most appropriate supporting medium that allowed for discrete molecular separation and visualization. The early 1950s, therefore, marked not only an important turning point in the history of separation science, but also a transformative moment in the history of the life sciences as the growth of molecular biology depended in part on the epistemological access to the molecular realm available through these evolving technologies.

Hydroxyapatite/zirconia-microfibre composites with controlled microporosity and fracture properties prepared by electrophoretic deposition

Czech Academy of Sciences Publication Activity Database

Drdlík, D.; Sláma, M.; Hadraba, Hynek; Cihlář, J.

2015-01-01

Roč. 41, č. 9 (2015), s. 11202-11212 ISSN 0272-8842 R&D Projects: GA ČR(CZ) GAP108/11/1644; GA MŠk(CZ) ED1.1.00/02.0068 Institutional support: RVO:68081723 Keywords : hydroxyapatite * zirconia * composite * electrophoretic deposition * porosity Subject RIV: JH - Ceramics, Fire-Resistant Materials and Glass Impact factor: 2.758, year: 2015

THE EFFECT OF SOME RHIZOBACTERIAN STRAINS ON SOLUBLE PROTEINS CONTENT IN SOYBEANS (GLYCINE MAX L. MERR.

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Marius Stefan

2007-08-01

Full Text Available Now it is an accepted fact that plant growth-promoting rhizobacteria (PGPR can increase the productivity of several crops. The main objective of the present study was to find if there are any differences in protein content in the seeds of soybean (Glycine max L. MERR.. Using spectrophotometric methods for analyzing the protein contents and electrophoretic methods for qualitative analysis it was observed that no major modifications occur in protein spectrum. Looking at the quantitative side there was a small improvement in protein quantity.

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

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Perez-Chabela, M. Lourdes

2015-12-01

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

Specific binding of [alpha-32P]GTP to cytosolic and membrane-bound proteins of human platelets correlates with the activation of phospholipase C

International Nuclear Information System (INIS)

Lapetina, E.G.; Reep, B.R.

1987-01-01

We have assessed the binding of [alpha- 32 P]GTP to platelet proteins from cytosolic and membrane fractions. Proteins were separated by NaDodSO 4 /PAGE and electrophoretically transferred to nitrocellulose. Incubation of the nitrocellulose blots with [alpha- 32 P]GTP indicated the presence of specific and distinct GTP-binding proteins in cytosol and membranes. Binding was prevented by 10-100 nM GTP and by 100 nM guanosine 5'-[gamma-thio]triphosphate (GTP[gamma S]) or GDP; binding was unaffected by 1 nM-1 microM ATP. One main GTP-binding protein (29.5 kDa) was detected in the membrane fraction, while three others (29, 27, and 21 kDa) were detected in the soluble fraction. Two cytosolic GTP-binding proteins (29 and 27 kDa) were degraded by trypsin; another cytosolic protein (21 kDa) and the membrane-bound protein (29.5 kDa) were resistant to the action of trypsin. Treatment of intact platelets with trypsin or thrombin, followed by lysis and fractionation, did not affect the binding of [alpha- 32 P]GTP to the membrane-bound protein. GTP[gamma S] still stimulated phospholipase C in permeabilized platelets already preincubated with trypsin. This suggests that trypsin-resistant GTP-binding proteins might regulate phospholipase C stimulated by GTP[gamma S

Formation of a covalent complex between the terminal protein of pneumococcal bacteriophage Cp-1 and 5'-dAMP

Energy Technology Data Exchange (ETDEWEB)

Garcia, P.; Hermoso, J.M.; Garcia, J.A.; Garcia, E.; Lopez, R.; Salas, M.

1986-04-01

Incubation of extracts of Cp-1-infected Streptococcus pneumoniae with (..cap alpha..-/sup 32/P)dATP produced a labeled protein with the electrophoretic mobility of the Cp-1 terminal protein. The reaction product was resistant to treatment with micrococcal nuclease and sensitive to treatment with proteinase K. Incubation of the /sup 32/P-labeled protein with 5 M piperidine for 4 h at 50/sup 0/C released 5'-dAMP, indicating that a covalent complex between the terminal protein and 5'-dAMP was formed in vitro. When the four deoxynucleoside triphosphates were included in the reaction mixture, a labeled complex of slower electrophoretic mobility in sodium dodecyl sulfate-polyacrylamide gels than the terminal protein-dAMP complex was also found, indicating that the Cp-1 terminal protein-dAMP complex can be elongated and, therefore, that it is an initiation complex. Treatment of the /sup 32/P-labeled terminal protein-dAMP complex with 5.8 M HCl at 110/sup 0/C for 2 h yielded phosphothreonine. These results, together with the resistance of the terminal protein-DNA linkage to hydroxylamine, suggest that the Cp-1 terminal protein is covalently linked to the DNA through a phosphoester bond between L-threonine and 5'-dAMP, namely, a O-5'-deoxyadenylyl-L-threonine bond.

Field inversion gel electrophoretic analysis of Legionella pneumophila strains associated with nosocomial legionellosis in children.

Science.gov (United States)

Green, M; Wald, E R; Dashefsky, B; Barbadora, K; Wadowsky, R M

1996-01-01

Two nosocomial cases of Legionnaires' disease occurred in children. Legionella pneumophila serogroup 1 was isolated from both patients and 30 of 39 plumbing system sites in the hospital. The patient and hospital environmental isolates yielded identical field inversion gel electrophoretic patterns which differed from patterns observed with epidemiologically unrelated strains.

Blood serum components and serum protein test of Hybro-PG broilers of different ages

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PRL Silva

2007-12-01

Full Text Available Blood serum samples of HYBRO PG broilers were analyzed, with 30 samples collected from 21-day-old broilers (G1, 30 from 35-day-old birds (G2, and 30 from 42-day-old birds (G3, with the aim of establishing normal values of some blood serum parameters. The activities of the enzymes gamma-glutamyl-transferase (GGT, aspartate aminotransferase (AST, creatine kinase (CK, alkaline phosphatase (ALP, and lactate dehydrogenase (LDH, serum levels of total calcium, calcium ion, phosphorus, sodium, potassium, magnesium, chlorides, creatinine, uric acid, triglycerides, cholesterol, total protein, albumin, total and indirect and direct bilirubin, and electrophoretic profile of serum proteins in acrylamide (SDS-PAGE and agarose gel were determined. There was no influence of age on total bilirubin and albumin levels. All the other evaluated parameters presented differences in at least one age group. Protein electrophoretic profile also changed as a function of age. The obtained results can be considered as normal for the studied ages, and therefore be used as references for the interpretation of laboratory exams of broilers of this genetic line in the evaluated ages.

Comparison of 2 electrophoretic methods and a wet-chemistry method in the analysis of canine lipoproteins.

Science.gov (United States)

Behling-Kelly, Erica

2016-03-01

The evaluation of lipoprotein metabolism in small animal medicine is hindered by the lack of a gold standard method and paucity of validation data to support the use of automated chemistry methods available in the typical veterinary clinical pathology laboratory. The physical and chemical differences between canine and human lipoproteins draw into question whether the transference of some of these human methodologies for the study of canine lipoproteins is valid. Validation of methodology must go hand in hand with exploratory studies into the diagnostic or prognostic utility of measuring specific lipoproteins in veterinary medicine. The goal of this study was to compare one commercially available wet-chemistry method to manual and automated lipoprotein electrophoresis in the analysis of canine lipoproteins. Canine lipoproteins from 50 dogs were prospectively analyzed by 2 electrophoretic methods, one automated and one manual method, and one wet-chemistry method. Electrophoretic methods identified a higher proportion of low-density lipoproteins than the wet-chemistry method. Automated electrophoresis occasionally failed to identify very low-density lipoproteins. Wet-chemistry methods designed for evaluation of human lipoproteins are insensitive to canine low-density lipoproteins and may not be applicable to the study of canine lipoproteins. Automated electrophoretic methods will likely require significant modifications if they are to be used in the analysis of canine lipoproteins. Studies aimed at determining the impact of a disease state on lipoproteins should thoroughly investigate the selected methodology prior to the onset of the study. © 2016 American Society for Veterinary Clinical Pathology.

Capillary electrophoretic analysis reveals subcellular binding between individual mitochondria and cytoskeleton

Science.gov (United States)

Kostal, Vratislav; Arriaga, Edgar A.

2011-01-01

Interactions between the cytoskeleton and mitochondria are essential for normal cellular function. An assessment of such interactions is commonly based on bulk analysis of mitochondrial and cytoskeletal markers present in a given sample, which assumes complete binding between these two organelle types. Such measurements are biased because they rarely account for non-bound ‘free’ subcellular species. Here we report on the use of capillary electrophoresis with dual laser induced fluorescence detection (CE-LIF) to identify, classify, count and quantify properties of individual binding events of mitochondria and cytoskeleton. Mitochondria were fluorescently labeled with DsRed2 while F-actin, a major cytoskeletal component, was fluorescently labeled with Alexa488-phalloidin. In a typical subcellular fraction of L6 myoblasts, 79% of mitochondrial events did not have detectable levels of F-actin, while the rest had on average ~2 zeptomole F-actin, which theoretically represents a ~ 2.5-μm long network of actin filaments per event. Trypsin treatment of L6 subcellular fractions prior to analysis decreased the fraction of mitochondrial events with detectable levels of F-actin, which is expected from digestion of cytoskeletal proteins on the surface of mitochondria. The electrophoretic mobility distributions of the individual events were also used to further distinguish between cytoskeleton-bound from cytoskeleton-free mitochondrial events. The CE-LIF approach described here could be further developed to explore cytoskeleton interactions with other subcellular structures, the effects of cytoskeleton destabilizing drugs, and the progression of viral infections. PMID:21309532

Electrophoretically deposited graphene oxide and carbon nanotube composite for electrochemical capacitors

International Nuclear Information System (INIS)

Ajayi, Obafunso A; Wong, Chee Wei; Guitierrez, Daniel H; Peaslee, David; Cheng, Arthur; Chen, Bin; Gao, Theodore

2015-01-01

We report a scalable one-step electrode fabrication approach for synthesizing composite carbon-based supercapacitors with synergistic outcomes. Multi-walled carbon nanotubes (MWCNTs) were successfully integrated into our modified electrophoretic deposition process to directly form composite MWCNT–GO electrochemical capacitor electrodes (where GO is graphene oxide) with superior performance to solely GO electrodes. The measured capacitance improved threefold, reaching a maximum specific capacitance of 231 F g"−"1. Upon thermal reduction, MWCNT–GO electrode sheet resistance decreased by a factor of 8, significantly greater than the 2× decrease of those without MWCNTs. (paper)

Reagent-Free Electrophoretic Synthesis of Few-Atom-Thick Metal Oxide Nanosheets

DEFF Research Database (Denmark)

Hou, Chengyi; Zhang, Minwei; Zhang, Lili

2017-01-01

Engineering traditional materials into the new form of atomic and free-standing two-dimensional structures is of both fundamental interest and practical significance, but it is in general facing challenges especially for metal oxide semiconductors. We herein report an ultragreen method for the cost......-effective and fast preparation of atomic metal oxide nanosheets that can be further transformed into nanofilms. The method combines top-down building block synthesis and bottom-up electrophoretic assembly in water under ambient conditions, using only bulk metal and Milli-Q water without involving any additional...

Classification of Spanish white wines using their electrophoretic profiles obtained by capillary zone electrophoresis with amperometric detection.

Science.gov (United States)

Arribas, Alberto Sánchez; Martínez-Fernández, Marta; Moreno, Mónica; Bermejo, Esperanza; Zapardiel, Antonio; Chicharro, Manuel

2014-06-01

A method was developed for the simultaneous detection of eight polyphenols (t-resveratrol, (+)-catechin, quercetin and p-coumaric, caffeic, sinapic, ferulic, and gallic acids) by CZE with electrochemical detection. Separation of these polyphenols was achieved within 25 min using a 200 mM borate buffer (pH 9.4) containing 10% methanol as separation electrolyte. Amperometric detection of polyphenols was carried out with a glassy carbon electrode (GCE) modified with a multiwalled carbon nanotubes (CNT) layer obtained from a dispersion of CNT in polyethylenimine. The excellent electrochemical properties of this modified electrode allowed the detection and quantification of the selected polyphenols in white wines without any pretreatment step, showing remarkable signal stability despite the presence of potential fouling substances in wine. The electrophoretic profiles of white wines, obtained using this methodology, have proven to be useful for the classification of these wines by means of chemometric multivariate techniques. Principal component analysis and discriminant analysis allowed accurate classification of wine samples on the basis of their grape varietal (verdejo and airén) using the information contained in selected zones of the electropherogram. The utility of the proposed CZE methodology based on the electrochemical response of CNT-modified electrodes appears to be promising in the field of wine industry and it is expected to be successfully extended to classification of a wider range of wines made of other grape varietals. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

A two-dimensional electrophoretic profile of the proteins secreted by Herbaspirillum seropedicae strain Z78.

Science.gov (United States)

Chaves, Daniela Fojo Seixas; de Souza, Emanuel Maltempi; Monteiro, Rose Adele; de Oliveira Pedrosa, Fábio

2009-11-02

Herbaspirillum seropedicae is an endophytic bacterium that associates with rice, sugarcane and other economically important crops. Secreted proteins play a key role in the plant-bacterial interaction. Using 2D electrophoresis and peptide mass fingerprint mass spectrometry, 63 protein spots representing 41 different secreted proteins were identified during growth of H. seropedicae under nitrogen-sufficient conditions. In silico analysis showed that 25.4% of the proteins had signal peptides and 15.9% were predicted to be non-classically secreted. Among the most abundant were flagellar components and ABC-type transport system proteins. Nine secreted proteins had also been identified in the cellular proteome, suggesting that they also play a role in the extracellular environment. No type III secreted proteins were detected by comparison of the wild type strain with an hrcN mutant strain.

Porous SiO2/HAp Coatings on Cp-Titanium Grade 1 Surfaces Produced by Electrophoretic Deposition

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Moskalewicz T.

2016-12-01

Full Text Available Porous hydroxyapatite doped SiO2 coatings were electrophoretically deposited (EPD on commercially pure titanium. The influence of EPD parameters on coatings quality was investigated. Microstructural observation was done using transmission and scanning electron microscopy as well as X-ray diffractometry.

High-mobility group (HMG) protein HMG-1 and TATA-binding protein-associated factor TAF(II)30 affect estrogen receptor-mediated transcriptional activation.

Science.gov (United States)

Verrier, C S; Roodi, N; Yee, C J; Bailey, L R; Jensen, R A; Bustin, M; Parl, F F

1997-07-01

The estrogen receptor (ER) belongs to a family of ligand-inducible nuclear receptors that exert their effects by binding to cis-acting DNA elements in the regulatory region of target genes. The detailed mechanisms by which ER interacts with the estrogen response element (ERE) and affects transcription still remain to be elucidated. To study the ER-ERE interaction and transcription initiation, we employed purified recombinant ER expressed in both the baculovirus-Sf9 and his-tagged bacterial systems. The effect of high-mobility group (HMG) protein HMG-1 and purified recombinant TATA-binding protein-associated factor TAF(II)30 on ER-ERE binding and transcription initiation were assessed by electrophoretic mobility shift assay and in vitro transcription from an ERE-containing template (pERE2LovTATA), respectively. We find that purified, recombinant ER fails to bind to ERE in spite of high ligand-binding activity and electrophoretic and immunological properties identical to ER in MCF-7 breast cancer cells. HMG-1 interacts with ER and promotes ER-ERE binding in a concentration- and time-dependent manner. The effectiveness of HMG-1 to stimulate ER-ERE binding in the electrophoretic mobility shift assay depends on the sequence flanking the ERE consensus as well as the position of the latter in the oligonucleotide. We find that TAF(II)30 has no effect on ER-ERE binding either alone or in combination with ER and HMG-1. Although HMG-1 promotes ER-ERE binding, it fails to stimulate transcription initiation either in the presence or absence of hormone. In contrast, TAF(II)30, while not affecting ER-ERE binding, stimulates transcription initiation 20-fold in the presence of HMG-1. These results indicate that HMG-1 and TAF(II)30 act in sequence, the former acting to promote ER-ERE binding followed by the latter to stimulate transcription initiation.

Simultaneous separation of water- and fat-soluble vitamins in isocratic pressure-assisted capillary electrochromatography using a methacrylate-based monolithic column.

Science.gov (United States)

Yamada, Hiroki; Kitagawa, Shinya; Ohtani, Hajime

2013-06-01

A method of simultaneous separation of water- and fat-soluble vitamins using pressure-assisted CEC with a methacrylate-based capillary monolithic column was developed. In the proposed method, water-soluble vitamins were mainly separated electrophoretically, while fat soluble-ones were separated chromatographically by the interaction with a methacrylate-based monolith. A mixture of six water-soluble and four fat-soluble vitamins was separated simultaneously within 20 min with an isocratic elution using 1 M formic acid (pH 1.9)/acetonitrile (30:70, v/v) containing 10 mM ammonium formate as a mobile phase. When the method was applied to a commercial multivitamin tablet and a spiked one, the vitamins were successfully analyzed, and no influence of the matrix contained in the tablet was observed. © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Two Dimensional Electrophoresis of Galactosidase Relating to the Disappearance of Bombyx Lectin Activity

OpenAIRE

カトウ, ヤスオ; Yasuo, Kato

2004-01-01

"Two dimensional polyacrylamide gel electroporesis (2 D-PAGE) analysis on the haemolymph of Bombyx mori was performed using the Mini-PROTEAN mini tube gel two dimensional polyacrylamide gel electrophoresis system (Bio-Rad Laboratories, Inc.). The result on various electrophoretical conditions using the haemolymph-protein showed the possibility that the haemolymph-protein was separated actually by means of this method. Moreover, the result of 2 D-PAGE analysis on Fraction II obtained by gel fi...

Demonstrating Interactions of Transcription Factors with DNA by Electrophoretic Mobility Shift Assay.

Science.gov (United States)

Yousaf, Nasim; Gould, David

2017-01-01

Confirming the binding of a transcription factor with a particular DNA sequence may be important in characterizing interactions with a synthetic promoter. Electrophoretic mobility shift assay is a powerful approach to demonstrate the specific DNA sequence that is bound by a transcription factor and also to confirm the specific transcription factor involved in the interaction. In this chapter we describe a method we have successfully used to demonstrate interactions of endogenous transcription factors with sequences derived from endogenous and synthetic promoters.

Electrophoretic Nanocrystalline Graphene Film Electrode for Lithium Ion Battery

International Nuclear Information System (INIS)

Kaprans, Kaspars; Bajars, Gunars; Kucinskis, Gints; Dorondo, Anna; Mateuss, Janis; Gabrusenoks, Jevgenijs; Kleperis, Janis; Lusis, Andrejs

2015-01-01

Graphene sheets were fabricated by electrophoretic deposition method from water suspension of graphene oxide followed by thermal reduction. The formation of nanocrystalline graphene sheets has been confirmed by scanning electron microscopy, X-ray diffraction and Raman spectroscopy. The electrochemical performance of graphene sheets as anode material for lithium ion batteries was evaluated by cycling voltammetry, galvanostatic charge-discharge cycling, and electrochemical impedance spectroscopy. Fabricated graphene sheets exhibited high discharge capacity of about 1120 mAh·g −1 and demonstrated good reversibility of lithium intercalation and deintercalation in graphene sheet film with capacity retention over 85 % after 50 cycles. Results show that nanocrystalline graphene sheets prepared by EPD demonstrated a high potential for application as anode material in lithium ion batteries

Electrophoretic formation of semiconductor layers with adjustable band gap

Science.gov (United States)

Shindrov, Alexander; Yuvchenko, Sergey; Vikulova, Maria; Tretyachenko, Elena; Zimnyakov, Dmitry; Gorokhovsky, Alexander

2017-11-01

The ceramic layers of the potassium polytitanates modified by transition metal salts were electrophoretically deposited onto the surface of glassy substrate coated with indium-tin oxide. The deposition allows obtaining a dense ceramic layer formed by composite agglomerates consisting of nanoscale particles with average size of 130-190 nm. The optical absorption spectra of the coatings modified in the mixtures of aqueous solutions of different transition metal salts were investigated. It was recognized that a bandgap value of these composites can be adjusted in a range from 1.4 to 2.3 eV depending the chemical composition of layered double hydroxide obtained during modification. This might be very promising for optoelectronic applications of such coatings due to an explicit control of optical properties.

Thickness control in electrophoretic deposition of WO{sub 3} nanofiber thin films for solar water splitting

Energy Technology Data Exchange (ETDEWEB)

Fang, Yuanxing; Lee, Wei Cheat; Canciani, Giacomo E.; Draper, Thomas C.; Al-Bawi, Zainab F. [Department of Chemistry, School of Life Sciences, University of Sussex, Brighton BN1 9QJ (United Kingdom); Bedi, Jasbir S. [School of Public Health & Zoonoses, Guru Angad Dev Veterinary and Animal Sciences University, Ludhiana 141004 Punjab (India); Perry, Christopher C. [Division of Biochemistry, School of Medicine, Loma Linda University, Loma Linda, CA 92350 (United States); Chen, Qiao, E-mail: qiao.chen@sussex.ac.uk [Department of Chemistry, School of Life Sciences, University of Sussex, Brighton BN1 9QJ (United Kingdom)

2015-12-15

Graphical abstract: - Highlights: • A novel method combining electrospinning and electrophoretic deposition was established for the creation of nanostructured semiconductor thin films. • The created thin films displayed a high chemical stability with a controllable thickness. • The PEC water splitting performance of the thin films was optimized by fine-tuning the thickness of the films. • A maximum photoconversion efficiency was achieved by 18 μm nanofibrous thin films. - Abstract: Electrophoretic deposition (EPD) of ground electrospun WO{sub 3} nanofibers was applied to create photoanodes with controlled morphology for the application of photoelectrochemical (PEC) water splitting. The correlations between deposition parameters and film thicknesses were investigated with theoretical models to precisely control the morphology of the nanostructured porous thin film. The photoconversion efficiency was further optimized as a function of film thickness. A maximum photoconversion efficiency of 0.924% from electrospun WO{sub 3} nanofibers that EPD deposited on a substrate was achieved at a film thickness of 18 μm.

Bioanalysis of eukaryotic organelles

Czech Academy of Sciences Publication Activity Database

Satori, Ch. P.; Henderson, M. M.; Krautkramer, E. A.; Košťál, Vratislav; Distefano, M. M.; Arriaga, E. A.

2013-01-01

Roč. 113, č. 4 (2013), s. 2733-2811 ISSN 0009-2665 R&D Projects: GA ČR(CZ) GBP206/12/G014 Grant - others:GA AV ČR(CZ) M200311201 Institutional support: RVO:68081715 Keywords : green fluorescent protein * atomic-force microscopy * capillary electrophoretic analysis Subject RIV: CB - Analytical Chemistry , Separation Impact factor: 45.661, year: 2013

Bioanalysis of eukaryotic organelles

Czech Academy of Sciences Publication Activity Database

Satori, Ch. P.; Henderson, M. M.; Krautkramer, E. A.; Košťál, Vratislav; Distefano, M. M.; Arriaga, E. A.

2013-01-01

Roč. 113, č. 4 (2013), s. 2733-2811 ISSN 0009-2665 R&D Projects: GA ČR(CZ) GBP206/12/G014 Grant - others:GA AV ČR(CZ) M200311201 Institutional support: RVO:68081715 Keywords : green fluorescent protein * atomic-force microscopy * capillary electrophoretic analysis Subject RIV: CB - Analytical Chemistry, Separation Impact factor: 45.661, year: 2013

Separation methods for captopril in pharmaceuticals and biological fluids.

Science.gov (United States)

Mansour, Fotouh R; Danielson, Neil D

2012-06-01

Captopril (CAP) is an orally active angiotensin-converting enzyme (ACE) inhibitor and has been widely used for management of hypertension and congestive heart failure. CAP lacks an aromatic chromophore required for facile direct UV detection and also has two chiral centers. These factors can render the determination of CAP in complex matrices challenging. This review covers more than 20 years of analytical research on this drug, focusing mainly on pharmaceutical and biological applications. The primary separation techniques discussed are gas chromatography, liquid chromatography, and capillary electrophoresis. The structures of the CAP derivatizing agents as well as a table summarizing various HPLC methods are provided. A discussion of key recent chromatographic and electrophoretic methods for other ACE inhibitors is also present. © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Assembly and structural organization of pigment-protein complexes in membranes of Rhodopseudomonas sphaeroides

International Nuclear Information System (INIS)

Hunter, C.N.; Pennoyer, J.D.; Niederman, R.A.

1982-01-01

The B875 and B800-850 light-harvesting pigment-protein complexes of Rhodopseudomonas sphaeroides are characterized further by lithium dodecyl sulfate/polyacrylamide gel electrophoresis at 4 degrees C. Bacteriochlorophyll a was shown in reconstruction studies to remain complexed with its respective binding proteins during this procedure. From distributions in these gels, a quantitative description for the arrangement of the complexes is proposed. Assembly of the complexes was examined in delta-aminolevulinate-requiring mutant H-5 after a shift from high- to low-light intensity. After 10 h of delta-[ 3 H]aminolevulinate labeling, the specific radioactivity of bacteriochlorophyll in a fraction containing putative membrane invaginations reached the maximal level, while that of the mature photosynthetic membrane was at only one-third this level. This suggests that membrane invaginations are sites of preferential bacteriochlorophyll synthesis in which completed pigment-proteins exist transiently. Analysis of the 3 H distribution after electrophoretic separation further suggests that photosynthetic membranes grow mainly by addition of B800-850 to preformed membrane consisting largely of B875 and photochemical reaction centers. These results corroborate the above model for the structural organization of the light-harvesting system and indicate that the structurally and functionally discrete B800-850 pool is not completely assembled until all B875 sites for B800-850 interactions are occupied

Experimental data and theoretical predictions for the rate of electrophoretic clarification of colloidal suspensions

Energy Technology Data Exchange (ETDEWEB)

Johnson, T.J.; Davis, E.J.

2000-05-01

An experimental and theoretical investigation of the electrophoretic clarification rate of colloidal suspensions was conducted. The suspensions included a coal-washing effluent and a model system of TiO{sub 2} particles. A parametric study of TiO{sub 2} suspensions was performed to validate and analysis of the electrophoretic motion of the clarification front formed between a clear zone and the suspension. To measure the electric field strength needed in the prediction of the location of the front, a moveable probe and salt bridge were connected to a reference electrode. Using the measured electric field strengths, it was found that the numerical solution to the unit cell electrophoresis model agrees with the measured clarification rates. For suspensions with moderately thick electric double layers and high particle volume fractions the deviations from classical Smoluchowski theory are substantial, and the numerical analysis is in somewhat better agreement with the data than a prior solution of the problem. The numerical model reduces to the predictions of previous theories as the thickness of the electric double layer decreases, and it is in good agreement with the clarification rate measured for a coal-washing effluent suspension with thin electric double layers.

Theory and measurements of electrophoretic effects in monolith, fixed-bed, and fluidized-bed plasma reactors

International Nuclear Information System (INIS)

Morin, T.J.

1989-01-01

Pressure gradients and secondary flow fields generated by the passage of electrical current in a d.c. gas discharge or gas laser are topics of longstanding interest in the gaseous electronics literature. These hydrodynamic effects of space charge fields and charged particle density gradients have been principally exploited in the development of gas separation and purification processes. In recent characterization studies of fixed-bed and fluidized-bed plasma reactors several anomalous flow features have been observed. These reactors involve the contacting of a high-frequency, resonantly-sustained, disperse gas discharge with granular solids in a fixed or fluidized bed. Anomalies in the measured pressure drops and fluidization velocities have motivated the development of an appropriate theoretical approach to, and some additional experimental investigations of electrophoretic effects in disperse gas discharges. In this paper, a theory which includes the effects of space charge and diffusion is used to estimate the electric field and charged particle density profiles. These profiles are then used to calculate velocity fields and gas flow rates for monolith, fixed-bed, and fluidized-bed reactors. These results are used to rationalize measurements of gas flow rates and axial pressure gradients in high-frequency disperse gas discharges with and without an additional d.c. axial electric field

Study of total seed proteins pattern of sesame (sesamum indicum l.) landraces via sodium dodecyl sulfate polyacrylamide gel electrophoresis (sds-page)

International Nuclear Information System (INIS)

Akbar, F.; Shinwari, Z.K.

2012-01-01

The sesame (Sesamum indicum L.) germplasm, comprising of 105 accessions was characterized for total seed storage proteins using sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). The germplasm was collected from diverse agro-ecological regions of Pakistan. To our information, no studies have yet been carried out in Pakistan on the genetic evaluation of sesame genotypes based on total seed protein. Total seed proteins were electrophoretically separated on 12% polyacrylamide gels by standard protocols. A total of 20 polypeptide bands were observed, of which 14 (70%) were polymorphic and 6 (30%) were monomorphic, with molecular weight ranging from 13.5 to 100 kDa. Six bands i.e., 7, 11, 12, 15, 16 and 18 were common in all genotypes. Similarity coefficients varied fro m 0.50 to 1.00. The dendrogram based on dissimilarity matrix using unweighted pair group method with arithmetic averages (UPGMA) separated all sesame accessions into three main groups i.e., A, B, C, comprising 89, 14 and 2 genotypes, respectively. Overall a low to medium level of genetic variability was observed for SDS-PAGE (single dimension). As SDS-PAGE alone did not reveal high level of genetic variability, hence 2-D gel electrophoresis along with other advanced type DNA markers and more number of sesame accessions from all over the country are recommended for the future genetic evaluation. Our investigation will significantly support the classification, development, genetic evaluation and conservation of sesame germplasm in Pakistan. (author)

Study of total seed proteins pattern of sesame (sesamum indicum l.) landraces via sodium dodecyl sulfate polyacrylamide gel electrophoresis (sds-page)

Energy Technology Data Exchange (ETDEWEB)

Akbar, F; Shinwari, Z K [Quaid-e-Azam University, Islamabad (Pakistan). Dept. of Biotechnology; Yousif, N; Masood, M S [Institute of Agri-Biotechnology and Genetic Resources, Islamabad (Pakistan)

2012-11-15

The sesame (Sesamum indicum L.) germplasm, comprising of 105 accessions was characterized for total seed storage proteins using sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). The germplasm was collected from diverse agro-ecological regions of Pakistan. To our information, no studies have yet been carried out in Pakistan on the genetic evaluation of sesame genotypes based on total seed protein. Total seed proteins were electrophoretically separated on 12% polyacrylamide gels by standard protocols. A total of 20 polypeptide bands were observed, of which 14 (70%) were polymorphic and 6 (30%) were monomorphic, with molecular weight ranging from 13.5 to 100 kDa. Six bands i.e., 7, 11, 12, 15, 16 and 18 were common in all genotypes. Similarity coefficients varied fro m 0.50 to 1.00. The dendrogram based on dissimilarity matrix using unweighted pair group method with arithmetic averages (UPGMA) separated all sesame accessions into three main groups i.e., A, B, C, comprising 89, 14 and 2 genotypes, respectively. Overall a low to medium level of genetic variability was observed for SDS-PAGE (single dimension). As SDS-PAGE alone did not reveal high level of genetic variability, hence 2-D gel electrophoresis along with other advanced type DNA markers and more number of sesame accessions from all over the country are recommended for the future genetic evaluation. Our investigation will significantly support the classification, development, genetic evaluation and conservation of sesame germplasm in Pakistan. (author)

Secretome analysis of Aspergillus fumigatus reveals Asp-hemolysin as a major secreted protein.

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Wartenberg, Dirk; Lapp, Katrin; Jacobsen, Ilse D; Dahse, Hans-Martin; Kniemeyer, Olaf; Heinekamp, Thorsten; Brakhage, Axel A

2011-11-01

Surface-associated and secreted proteins represent primarily exposed components of Aspergillus fumigatus during host infection. Several secreted proteins are known to be involved in defense mechanisms or immune evasion, thus, probably contributing to pathogenicity. Furthermore, several secreted antigens were identified as possible biomarkers for the verification of diseases caused by Aspergillus species. Nevertheless, there is only limited knowledge about the composition of the secretome and about molecular functions of particular proteins. To identify secreted proteins potentially essential for virulence, the core secretome of A. fumigatus grown in minimal medium was determined. Two-dimensional gel electrophoretic separation and subsequent MALDI-TOF-MS/MS analyses resulted in the identification of 64 different proteins. Additionally, secretome analyses of A. fumigatus utilizing elastin, collagen or keratin as main carbon and nitrogen source were performed. Thereby, the alkaline serine protease Alp1 was identified as the most abundant protein and hence presumably represents an important protease during host infection. Interestingly, the Asp-hemolysin (Asp-HS), which belongs to the protein family of aegerolysins and which was often suggested to be involved in fungal virulence, was present in the secretome under all growth conditions tested. In addition, a second, non-secreted protein with an aegerolysin domain annotated as Asp-hemolysin-like (HS-like) protein can be found to be encoded in the genome of A. fumigatus. Generation and analysis of Asp-HS and HS-like deletion strains revealed no differences in phenotype compared to the corresponding wild-type strain. Furthermore, hemolysis and cytotoxicity was not altered in both single-deletion and double-deletion mutants lacking both aegerolysin genes. All mutant strains showed no attenuation in virulence in a mouse infection model for invasive pulmonary aspergillosis. Overall, this study provides a comprehensive

Zymography and reverse zymography for detecting MMPs and TIMPs.

Science.gov (United States)

Hawkes, Susan P; Li, Hongxia; Taniguchi, Gary T

2010-01-01

Zymography is the electrophoretic separation of proteins through a polyacrylamide gel containing a proteolytic substrate. After denaturing (but nonreducing) electrophoresis, proteins are renatured and incubated in an appropriate buffer for proteolytic activity. Clear zones of lysis in the stained gel indicated active proteinases. Reverse zymography is a similar technique to detect proteinase inhibitors. After renaturing of proteins, the gel is incubated with metalloproteinases which digest the substrate incorporated into the gel. Inhibitors are shown as dark zones of inhibition against a clear background upon staining.

The Effect of Column and Eluent Fluorination on the Retention and Separation of non-Fluorinated Amino Acids and Proteins by HPLC

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Joyner, Katherine; Wang, Weizhen; Yu, Yihua Bruce

2011-01-01

The effect of column and eluent fluorination on the retention and separation of non-fluorinated amino acids and proteins in HPLC is investigated. A side-by-side comparison of fluorocarbon column and eluents (F-column and F-eluents) with their hydrocarbon counterparts (H-column and H-eluents) in the separation of a group of 33 analytes, including 30 amino acids and 3 proteins, is conducted. The H-column and the F-column contain the n-C8H17 group and n-C8F17 group, respectively, in their stationary phases. The H-eluents include ethanol (EtOH) and isopropanol (ISP) while the F-eluents include trifluoroethanol (TFE) and hexafluorosopropanol (HFIP). The 2 columns and 4 eluents generated 8 (column, eluent) pairs that produce 264 retention time data points for the 33 analytes. A statistical analysis of the retention time data reveals that although the H-column is better than the F-column in analyte separation and H-eluents are better than F-eluents in analyte retention, the more critical factor is the proper pairing of column with eluent. Among the conditions explored in this project, optimal retention and separation is achieved when the fluorocarbon column is paired with ethanol, even though TFE is the most polar one among the 4 eluents. This result shows fluorocarbon columns have much potential in chromatographic analysis and separation of non-fluorinated amino acids and proteins. PMID:21318121

Confirmation of a blocked amino terminus of sulfhydryl oxidase

International Nuclear Information System (INIS)

Janolino, V.G.; Morrison-Rowe, S.J.; Swaisgood, H.E.

1990-01-01

The isolation of sulfhydryl oxidase from bovine milk in a suitably pure form for sequencing was carried out by transient covalent affinity chromatography of diafiltered whey using cysteinylsuccinamidopropyl-glass as matrix. The glutathione-eluted proteins were separated by SDS-PAGE. By radiolabeling the affinity chromatography-purified enzyme with [ 14 C]iodoacetate before subjecting to SDS-PAGE, the sulfhydryl oxidase band was identified, because sulfhydryl oxidase is known to be inactivated by alkylation of one sulfhydryl group per mole. The results confirmed that sulfhydryl oxidase corresponds to the 85 (± 5)-kDa band observed on SDS-PAGE. The protein band corresponding to radiolabeled sulfhydryl oxidase was recovered from SDS-PAGE gels by electrophoretic elution and by electroblotting on polyvinylidene difluoride membrane and subjected to gas phase sequencing. Precautions were taken during electrophoretic elution to prevent reactions that result in N-terminal blocking. Both methods of protein recovery yielded negative results when subjected to sequence analysis indicating that the N-terminus of sulfhydryl oxidase is blocked

High power density supercapacitor electrodes of carbon nanotube films by electrophoretic deposition

International Nuclear Information System (INIS)

Du Chunsheng; Pan Ning

2006-01-01

Carbon nanotube thin films have been successfully fabricated by the electrophoretic deposition technique. The supercapacitors built from such thin film electrodes have a very small equivalent series resistance, and a high specific power density over 20 kW kg -1 was thus obtained. More importantly, the supercapacitors showed superior frequency response. Our study also demonstrated that these carbon nanotube thin films can serve as coating layers over ordinary current collectors to drastically enhance the electrode performance, indicating a huge potential in supercapacitor and battery manufacturing

Effects of surfactants on spinning carbon nanotube fibers by an electrophoretic method

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Jun Ma, Jie Tang, Qian Cheng, Han Zhang, Norio Shinya and Lu-Chang Qin

2010-01-01

Full Text Available Thin fibers were spun from a colloidal solution of single-walled carbon nanotubes (SWNTs using an electrophoretic method. Sodium dodecylbenzenesulfonate (NaDDBS was chosen as a surfactant and showed good performance owing to its special chemical structure. The highest spinning velocity reached 0.5 mm s−1. The resulting SWNT fibers had a tensile strength of 400 MPa and a conductivity of 355 S cm−1. Their mechanical and electrical properties were markedly improved after adding NaDDBS as the dispersant in water.

Effect of vitamin D status on chick kidney proteins: detection of a 45-kilodalton mitochondrial protein suppressed by vitamin D

International Nuclear Information System (INIS)

Kain, S.R.; Kamrath, K.S.; Henry, H.L.

1988-01-01

Two-dimensional polyacrylamide gel electrophoresis along with L-[ 35 S]methionine radiolabeling studies were used to examine the effect of chronic vitamin D status on the composition and relative abundance of chick kidney proteins. Comparison of silver-stained gels revealed no extensive differences in either the electrophoretic mobility or the amounts of kidney proteins present in the mitochondrial fraction from vitamin D-replete and vitamin D-deficient chicks. A similar result was obtained in studies with L-[ 35 S]methionine-labeled proteins. Vitamin D deficiency specifically elevated levels of a 45-kilodalton mitochondrial protein (pI 5.0 to 5.5) by approximately 5- to 12-fold relative to amounts present in vitamin D-replete tissue. This protein could not be detected in postmitochondrial supernatant fractions and was only faintly visible in crude kidney homogenates. The specificity of the observed suppression of this 45-kilodalton protein by vitamin D suggests that it may play an important role in renal functions influenced by the vitamin D endocrine system

Separation of both fibrous and globular proteins on the basis of molecular weight using high-performance size exclusion chromatography.

Science.gov (United States)

Barden, J A

1983-11-01

A high-performance size exclusion liquid chromatographic system has been used to separate proteins with different shapes solely on the basis of their molecular weights. After the effects of ionic and hydrophobic interactions with the stationary phase have been overcome, protein elution is normally governed by their effective size in solution. Conditions are described under which proteins, with isoelectric points within the normal operating pH range of the columns, are eluted independent of their Stokes' radii. Even fibrous proteins with axial ratios of 50 elute according to their known molecular weights over the range 2000-2,000,000.

Phosphorylation of ribosomal proteins induced by auxins in maize embryonic tissues

International Nuclear Information System (INIS)

Perez, L.; Aguilar, R.; Mendez, A.P.; de Jimenez, E.S.

1990-01-01

The effect of auxin on ribosomal protein phosphorylation of germinating maize (Zea mays) tissues was investigated. Two-dimensional gel electrophoresis and autoradiography of [ 32 P] ribosomal protein patterns for natural and synthetic auxin-treated tissues were performed. Both the rate of 32 P incorporation and the electrophoretic patterns were dependent on 32 P pulse length, suggesting that active protein phosphorylation-dephosphorylation occurred in small and large subunit proteins, in control as well as in auxin-treated tissues. The effect of ribosomal protein phosphorylation on in vitro translation was tested. Measurements of poly(U) translation rates as a function of ribosome concentration provided apparent K m values significantly different for auxin-treated and nontreated tissues. These findings suggest that auxin might exert some kind of translational control by regulating the phosphorylated status of ribosomal proteins

Size and molecular weight determination of polysaccharides by means of nano electrospray gas-phase electrophoretic mobility molecular analysis (nES GEMMA).

Science.gov (United States)

Weiss, Victor U; Golesne, Monika; Friedbacher, Gernot; Alban, Susanne; Szymanski, Wladyslaw W; Marchetti-Deschmann, Martina; Allmaier, Günter

2018-02-21

Size, size distribution and molecular weight (MW) determination of nanoparticles and that are for example large polymers, are of great interest and pose an analytical challenge. In this context, nano electrospray gas-phase electrophoretic mobility molecular analysis (nES GEMMA) is a valuable tool with growing impact. Separation of single-charged analytes according to their electrophoretic mobility diameter (EMD) starting from single-digit EMDs up to several hundred nm diameters is possible. In case of spherical analytes, the EMD corresponds to the dry nanoparticle size. Additionally, the instrument is capable of number-based, single-particle detection following the recommendation of the European Commission for nanoparticle characterization (2011/696/EU). In case an EMD/MW correlation for a particular compound class (based on availability of well-defined standards) exists, a nanoparticle's MW can be determined from its EMD. In the present study, we focused on nES GEMMA of linear and branched, water-soluble polysaccharides forming nanoparticles and were able to obtain spectra for both analyte classes regarding single-charged species. Based on EMDs for corresponding analytes, an excellent EMD/MW correlation could be obtained in case of the branched natural polymer (dextran). This enables the determination of dextran MWs from nES GEMMA spectra despite high analyte polydispersity and in a size/MW range, where classical mass spectrometry is limited. EMD/MW correlations based on linear (pullulans, oat-ß-glucans) polymers were significantly different, possibly indicating challenges in the exact MW determination of these compounds by, for example, chromatographic and light scattering means. Despite these observations, nES GEMMA of linear, monosaccharide-based polymers enabled the determination of size and size-distribution of such dry bionanoparticles. © 2018 The Authors. Electrophoresis published by Wiley-VCH Verlag GmbH & Co. KGaA.

Proteomic Analysis of Bovine Pregnancy-specific Serum Proteins by 2D Fluorescence Difference Gel Electrophoresis

Science.gov (United States)

Lee, Jae Eun; Lee, Jae Young; Kim, Hong Rye; Shin, Hyun Young; Lin, Tao; Jin, Dong Il

2015-01-01

Two dimensional-fluorescence difference gel electrophoresis (2D DIGE) is an emerging technique for comparative proteomics, which improves the reproducibility and reliability of differential protein expression analysis between samples. The purpose of this study was to investigate bovine pregnancy-specific proteins in the proteome between bovine pregnant and non-pregnant serum using DIGE technique. Serums of 2 pregnant Holstein dairy cattle at day 21 after artificial insemination and those of 2 non-pregnant were used in this study. The pre-electrophoretic labeling of pregnant and non-pregnant serum proteins were mixed with Cy3 and Cy5 fluorescent dyes, respectively, and an internal standard was labeled with Cy2. Labeled proteins with Cy2, Cy3, and Cy5 were separated together in a single gel, and then were detected by fluorescence image analyzer. The 2D DIGE method using fluorescence CyDye DIGE flour had higher sensitivity than conventional 2D gel electrophoresis, and showed reproducible results. Approximately 1,500 protein spots were detected by 2D DIGE. Several proteins showed a more than 1.5-fold up and down regulation between non-pregnant and pregnant serum proteins. The differentially expressed proteins were identified by MALDI-TOF mass spectrometer. A total 16 protein spots were detected to regulate differentially in the pregnant serum, among which 7 spots were up-regulated proteins such as conglutinin precursor, modified bovine fibrinogen and IgG1, and 6 spots were down-regulated proteins such as hemoglobin, complement component 3, bovine fibrinogen and IgG2a three spots were not identified. The identified proteins demonstrate that early pregnant bovine serum may have several pregnancy-specific proteins, and these could be a valuable information for the development of pregnancy-diagnostic markers in early pregnancy bovine serum. PMID:25925056

Proteomic Analysis of Bovine Pregnancy-specific Serum Proteins by 2D Fluorescence Difference Gel Electrophoresis

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Jae Eun Lee

2015-06-01

Full Text Available Two dimensional-fluorescence difference gel electrophoresis (2D DIGE is an emerging technique for comparative proteomics, which improves the reproducibility and reliability of differential protein expression analysis between samples. The purpose of this study was to investigate bovine pregnancy-specific proteins in the proteome between bovine pregnant and non-pregnant serum using DIGE technique. Serums of 2 pregnant Holstein dairy cattle at day 21 after artificial insemination and those of 2 non-pregnant were used in this study. The pre-electrophoretic labeling of pregnant and non-pregnant serum proteins were mixed with Cy3 and Cy5 fluorescent dyes, respectively, and an internal standard was labeled with Cy2. Labeled proteins with Cy2, Cy3, and Cy5 were separated together in a single gel, and then were detected by fluorescence image analyzer. The 2D DIGE method using fluorescence CyDye DIGE flour had higher sensitivity than conventional 2D gel electrophoresis, and showed reproducible results. Approximately 1,500 protein spots were detected by 2D DIGE. Several proteins showed a more than 1.5-fold up and down regulation between non-pregnant and pregnant serum proteins. The differentially expressed proteins were identified by MALDI-TOF mass spectrometer. A total 16 protein spots were detected to regulate differentially in the pregnant serum, among which 7 spots were up-regulated proteins such as conglutinin precursor, modified bovine fibrinogen and IgG1, and 6 spots were down-regulated proteins such as hemoglobin, complement component 3, bovine fibrinogen and IgG2a three spots were not identified. The identified proteins demonstrate that early pregnant bovine serum may have several pregnancy-specific proteins, and these could be a valuable information for the development of pregnancy-diagnostic markers in early pregnancy bovine serum.

Separation techniques: Chromatography

Science.gov (United States)

Coskun, Ozlem

2016-01-01

Chromatography is an important biophysical technique that enables the separation, identification, and purification of the components of a mixture for qualitative and quantitative analysis. Proteins can be purified based on characteristics such as size and shape, total charge, hydrophobic groups present on the surface, and binding capacity with the stationary phase. Four separation techniques based on molecular characteristics and interaction type use mechanisms of ion exchange, surface adsorption, partition, and size exclusion. Other chromatography techniques are based on the stationary bed, including column, thin layer, and paper chromatography. Column chromatography is one of the most common methods of protein purification. PMID:28058406

Electrophoretic Deposition of Gallium with High Deposition Rate

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Hanfei Zhang

2014-12-01

Full Text Available In this work, electrophoretic deposition (EPD is reported to form gallium thin film with high deposition rate and low cost while avoiding the highly toxic chemicals typically used in electroplating. A maximum deposition rate of ~0.6 μm/min, almost one order of magnitude higher than the typical value reported for electroplating, is obtained when employing a set of proper deposition parameters. The thickness of the film is shown to increase with deposition time when sequential deposition is employed. The concentration of Mg(NO32, the charging salt, is also found to be a critical factor to control the deposition rate. Various gallium micropatterns are obtained by masking the substrate during the process, demonstrating process compatibility with microfabrication. The reported novel approach can potentially be employed in a broad range of applications with Ga as a raw material, including microelectronics, photovoltaic cells, and flexible liquid metal microelectrodes.

Effect of triiodothyronine on rat liver chromatin protein kinase

International Nuclear Information System (INIS)

Kruh, J.; Tichonicky, L.

1976-01-01

1) Injection of triiodothyronine to rats stimulates protein kinase activity in liver chromatin nonhistone proteins. A significant increase was found after two daily injections. A 4-fold increase was observed with the purified enzyme after eight daily injections of the hormone. No variations were observed in cytosol protein kinase activity. Electrophoretic pattern, effect of heat denaturation, effect of p-hydroxymercuribenzoate seem to indicate that the enzyme present in treated rats is not identical to the enzyme in control animals, which suggests that thyroid hormone has induced nuclear protein kinase. Diiodothyronine, 3, 3', 5'-triiodothyronine have no effect on protein kinase. 2) Chromatin non-histone proteins isolated from rats injected with triiodothyronine incorporated more 32 P when incubated with [γ- 32 P]ATP than the chromatin proteins from untreated rats. Thyroidectomy reduced the in vitro 32 P incorporation. It is suggested that some of the biological activity of thyroid hormone could be mediated through its effect on chromatin non-histone proteins. (orig.) [de

Pepsin Digested Oat Bran Proteins: Separation, Antioxidant Activity, and Identification of New Peptides

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Ariane Vanvi

2016-01-01

Full Text Available The aim of this study was to determine pepsin hydrolysis conditions to produce digested oat bran proteins with higher radical scavenging activities and separate and identify peptides. Isolated proteins were then digested with different concentrations of pepsin and incubation times. Hydrolysates produced with 1 : 30 enzyme substrate (E/S ratio and 2 h possessed the highest peroxyl radical scavenging activity, 608 ± 17 µM TE/g (compared to 456–474 µM TE/g for other digests, and was therefore subsequently fractionated into eight fractions (F1–F8 by high performance liquid chromatography (HPLC. F1 and F2 had little activity because of their low protein contents. Activities of F3–F8 were 447–874 µM TE/g, 20–36%, and 10–14% in the peroxyl, superoxide anion, and hydroxyl radical tests, respectively. Liquid chromatography-tandem mass spectrometry (LC-MS/MS was used to identify a total of fifty peptides that may have contributed to the activity of F3, a fraction that better scavenged radicals.

Covalently coating dextran on macroporous polyglycidyl methacrylate microsphere enabled rapid protein chromatographic separation

International Nuclear Information System (INIS)

Zhang, Rongyue; Li, Qiang; Li, Juan; Zhou, Weiqing; Ye, Peili; Gao, Yang; Ma, Guanghui; Su, Zhiguo

2012-01-01

Protein denaturation and nonspecific adsorption on polymer media as a chromatographic support have been a problem which needs to be overcome. Macroporous poly(glycidyl methacrylate–divinylbezene) (PGMA–DVB) microspheres prepared in this study were firstly covalently coated with dextran through a three-step method. The dextran was firstly adsorbed onto the microspheres and then covalently bound to the PGMA–DVB microsphere through ether bonds which were formed by hydroxyl group reacting with epoxy group at the presence of 4-(Dimethylamino) pyridine. Finally, the coating dextran layer was crosslinked by ethylene glycol diglycidyl ether to form the continuous network coating. The coated microspheres were characterized by Fourier transform infrared spectra, scanning electron microscope, mercury porosimetry measurements, laser scanning confocal microscope, and protein adsorption experiments. Results showed that PGMA–DVB microspheres coated with dextran successfully maintained the macroporous structure and high permeability. The backpressure was only 1.69 MPa at a high flow rate of 2891 cm/h. Consequently, the hydrophilicity and biocompatibility of modified microspheres were greatly improved, and the contact angle decreased from 184° to 13°, and nonspecific adsorption of proteins was decreased to little or none. The clad dextran coating with large amounts of hydroxyl group was easily derived to be various functional groups. The derived media have great potential applications in rapid protein chromatography. - Highlights: ► Macroporous PGMA–DVB microspheres were covalently coated with dextran. ► The hydrophilicity of the coated microspheres was significantly improved. ► The irreversible adsorption of proteins was reduced to zero. ► The coated microspheres can maintain the macropore structure. ► The coated microspheres were applied to rapid protein separation.

Conditions that allow for effective transfer of membrane proteins onto nitrocellulose membrane in Western blots.

Science.gov (United States)

Abeyrathne, Priyanka D; Lam, Joseph S

2007-04-01

A major hurdle in characterizing bacterial membrane proteins by Western blotting is the ineffectiveness of transferring these proteins from sodium dodecyl sulfate -- polyacrylamide gel electrophoresis (SDS-PAGE) gel onto nitrocellulose membrane, using standard Western blot buffers and electrophoretic conditions. In this study, we compared a number of modified Western blotting buffers and arrived at a composition designated as the SDS-PAGE-Urea Lysis buffer. The use of this buffer and specific conditions allowed the reproducible transfer of highly hydrophobic bacterial membrane proteins with 2-12 transmembrane-spanning segments as well as soluble proteins onto nitrocellulose membranes. This method should be broadly applicable for immunochemical studies of other membrane proteins.

Preparation of platinum-free tubular dye-sensitized solar cells by electrophoretic deposition

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Khwanchit Wongcharee

2016-10-01

Full Text Available Tubular dye-sensitized solar cells (DSSCs were developed by replacing expensive materials with lower cost materials as follows: (1 replacing conductive glass electrodes with titanium (Ti wires and (2 replacing platinum (Pt catalyst with the mixture of multi-walled carbon nanotubes, MWCNTs and Poly(3,4-ethylenedioxythiophene-poly(styrenesulfonate, PEDOT-PSS. Platinized counter electrodes were used as the standard counter electrodes for comparison. The effects of the chemical treatment of titanium wire substrate and electrophoretic deposition condition on the efficiency of DSSCs were also investigated. The chemical treatment of titanium wires was carried out by soaking the wires in HF-HNO3 solutions at three different concentrations of 0.8, 1.6 and 2.4 M and three different soaking durations of 5, 10 and 15 min. The optimum condition was found at HF-HNO3 concentration of 0.8 M and soaking duration of 10 min. Film coating on working electrodes was performed using electrophoretic technique at three different voltages of 5, 8 and 10 V and four different coating durations of 1, 3, 5 and 7 min. Then, the optimum condition at deposition voltage of 5 V and deposition duration of 5 min was applied for film deposition on counter electrodes. The efficiency of DSSC with CNTs/TiO2 counter electrode was 0.03%. The addition of PEDOT-PSS improved the efficiency of DSSC to 0.08%.

Overlapping ETS and CRE Motifs (G/CCGGAAGTGACGTCA) Preferentially Bound by GABPα and CREB Proteins

Science.gov (United States)

Chatterjee, Raghunath; Zhao, Jianfei; He, Ximiao; Shlyakhtenko, Andrey; Mann, Ishminder; Waterfall, Joshua J.; Meltzer, Paul; Sathyanarayana, B. K.; FitzGerald, Peter C.; Vinson, Charles

2012-01-01

Previously, we identified 8-bps long DNA sequences (8-mers) that localize in human proximal promoters and grouped them into known transcription factor binding sites (TFBS). We now examine split 8-mers consisting of two 4-mers separated by 1-bp to 30-bps (X4-N1-30-X4) to identify pairs of TFBS that localize in proximal promoters at a precise distance. These include two overlapping TFBS: the ETS⇔ETS motif (C/GCCGGAAGCGGAA) and the ETS⇔CRE motif (C/GCGGAAGTGACGTCAC). The nucleotides in bold are part of both TFBS. Molecular modeling shows that the ETS⇔CRE motif can be bound simultaneously by both the ETS and the B-ZIP domains without protein-protein clashes. The electrophoretic mobility shift assay (EMSA) shows that the ETS protein GABPα and the B-ZIP protein CREB preferentially bind to the ETS⇔CRE motif only when the two TFBS overlap precisely. In contrast, the ETS domain of ETV5 and CREB interfere with each other for binding the ETS⇔CRE. The 11-mer (CGGAAGTGACG), the conserved part of the ETS⇔CRE motif, occurs 226 times in the human genome and 83% are in known regulatory regions. In vivo GABPα and CREB ChIP-seq peaks identified the ETS⇔CRE as the most enriched motif occurring in promoters of genes involved in mRNA processing, cellular catabolic processes, and stress response, suggesting that a specific class of genes is regulated by this composite motif. PMID:23050235

Electrophoretic-deposited CNT/MnO2 composites for high-power electrochemical energy storage/conversion applications

Science.gov (United States)

Xiao, Wei; Xia, Hui; Fuh, Jerry Y. H.; Lu, Li

2010-05-01

CNT/MnO2 (birnessite-type) composite films have been successfully deposited on Ni-foil substrate via electrophoretic deposition (EPD). The unique EPD CNT/MnO2 composite film electrode shows enhanced electrical conductivity, good contact between composite films and the substrate and open porous structure, which makes the EPD composite films a promising electrode for high-power supercapacitors and lithium ion batteries.

Fabrication of a novel hemin-based monolithic column and its application in separation of protein from complex bio-matrix.

Science.gov (United States)

Jiang, Xiaoya; Zhang, Doudou; Li, Xueying; Wang, Xixi; Bai, Ligai; Liu, Haiyan; Yan, Hongyuan

2017-05-10

A novel polymer-based monolithic column was prepared via redox initiation system within the confines of a stainless steel column with 4.6mm i.d. In the processes, hemin and lauryl methacrylate were used as co-monomers; ethylene dimethacrylate as crosslinking agent; n-butyl alcohol, ethanediol, and N, N-dimethylformamide as tri-porogens; benzoyl peroxide and N, N-dimethyl aniline as redox initiation system. The resulting polymer-based monolithic columns were characterized by scanning electron microscopy, nitrogen adsorption-desorption instrument, and mercury intrusion porosimeter, respectively. The results illustrated that the improved monolith had relative uniform porous structure, good permeability, and low back pressure. Aromatic compounds were used to test the chromatographic behavior of the monolith, resulting in highest column efficiency of 19 880 plates per meter with reversed-phase mechanism. Furthermore, the homemade monolith was used as the stationary phase of high performance liquid chromatography to separate proteins from complex bio-matrix, including human plasma, egg white, and snailase. The results showed that the monolithic column occupied good separation ability with these complex bio-samples. Excellent specific character of the homemade hemin-based monolith was that it could simultaneously remove high-abundance proteins (including human serum albumin, immunoglobulin G, and human fibrinogen) from human plasma and separate other proteins to different fractions. Copyright © 2017 Elsevier B.V. All rights reserved.

Size measuring techniques as tool to monitor pea proteins intramolecular crosslinking by transglutaminase treatment.

Science.gov (United States)

Djoullah, Attaf; Krechiche, Ghali; Husson, Florence; Saurel, Rémi

2016-01-01

In this work, techniques for monitoring the intramolecular transglutaminase cross-links of pea proteins, based on protein size determination, were developed. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis profiles of transglutaminase-treated low concentration (0.01% w/w) pea albumin samples, compared to the untreated one (control), showed a higher electrophoretic migration of the major albumin fraction band (26 kDa), reflecting a decrease in protein size. This protein size decrease was confirmed, after DEAE column purification, by dynamic light scattering (DLS) where the hydrodynamic radius of treated samples appears to be reduced compared to the control one. Copyright © 2015 Elsevier Ltd. All rights reserved.

Partitioning the proteome: phase separation for targeted analysis of membrane proteins in human post-mortem brain.

Directory of Open Access Journals (Sweden)

Jane A English

Full Text Available Neuroproteomics is a powerful platform for targeted and hypothesis driven research, providing comprehensive insights into cellular and sub-cellular disease states, Gene × Environmental effects, and cellular response to medication effects in human, animal, and cell culture models. Analysis of sub-proteomes is becoming increasingly important in clinical proteomics, enriching for otherwise undetectable proteins that are possible markers for disease. Membrane proteins are one such sub-proteome class that merit in-depth targeted analysis, particularly in psychiatric disorders. As membrane proteins are notoriously difficult to analyse using traditional proteomics methods, we evaluate a paradigm to enrich for and study membrane proteins from human post-mortem brain tissue. This is the first study to extensively characterise the integral trans-membrane spanning proteins present in human brain. Using Triton X-114 phase separation and LC-MS/MS analysis, we enriched for and identified 494 membrane proteins, with 194 trans-membrane helices present, ranging from 1 to 21 helices per protein. Isolated proteins included glutamate receptors, G proteins, voltage gated and calcium channels, synaptic proteins, and myelin proteins, all of which warrant quantitative proteomic investigation in psychiatric and neurological disorders. Overall, our sub-proteome analysis reduced sample complexity and enriched for integral membrane proteins by 2.3 fold, thus allowing for more manageable, reproducible, and targeted proteomics in case vs. control biomarker studies. This study provides a valuable reference for future neuroproteomic investigations of membrane proteins, and validates the use Triton X-114 detergent phase extraction on human post mortem brain.

Review of UV spectroscopic, chromatographic, and electrophoretic methods for the cholinesterase reactivating antidote pralidoxime (2-PAM).

Science.gov (United States)

John, Harald; Blum, Marc-Michael

2012-01-01

Pralidoxime (2-PAM) belongs to the class of monopyridinium oximes with reactivating potency on cholinesterases inhibited by phosphylating organophosphorus compounds (OPC), for example, pesticides and nerve agents. 2-PAM represents an established antidote for the therapy of anticholinesterase poisoning since the late 1950s. Quite high therapeutic concentrations in human plasma (about 13 µg/ml) lead to concentrations in urine being about 100 times higher allowing the use of less sensitive analytical techniques that were used especially in the early years after 2-PAM was introduced. In this time (mid-1950s until the end of the 1970s) 2-PAM was most often analyzed by either paper chromatography or simple UV spectroscopic techniques omitting any sample separation step. These methods were displaced completely after the establishment of column liquid chromatography in the early 1980s. Since then, diverse techniques including cation exchange, size-exclusion, reversed-phase, and ligand-exchange chromatography have been introduced. Today, the most popular method for 2-PAM quantification is ion pair chromatography often combined with UV detection representing more than 50% of all column chromatographic procedures published. Furthermore, electrophoretic approaches by paper and capillary zone electrophoresis have been successfully used but are seldom applied. This review provides a commentary and exhaustive summary of analytical techniques applied to detect 2-PAM in pharmaceutical formulations and biological samples to characterize stability and pharmacokinetics as well as decomposition and biotransformation products. Separation techniques as well as diverse detectors are discussed in appropriate detail allowing comparison of individual preferences and limitations. In addition, novel data on mass spectrometric fragmentation of 2-PAM are provided. Copyright © 2011 John Wiley & Sons, Ltd.

Electrophoretic pattern of sera from lambs and kids vaccinated with irradiated Amphistome metacercariae (Cercariae indicae XXVI)

International Nuclear Information System (INIS)

Hafeez, Md.; Rao, B.V.

1986-01-01

Preliminary work has been done to study certain responses induced by irradiated amphistome metacercariae used as a vaccine to immunise lambs, kids and calves. The electrophoretic pattern of the sera collected from lambs and kids vaccinated with gamma irradiated amphistome matacercariae (C.I. XXVI) has been reported in this study. (author). 10 refs., 1 table

Click-PEGylation - A mobility shift approach to assess the redox state of cysteines in candidate proteins.

Science.gov (United States)

van Leeuwen, Lucie A G; Hinchy, Elizabeth C; Murphy, Michael P; Robb, Ellen L; Cochemé, Helena M

2017-07-01

The redox state of cysteine thiols is critical for protein function. Whereas cysteines play an important role in the maintenance of protein structure through the formation of internal disulfides, their nucleophilic thiol groups can become oxidatively modified in response to diverse redox challenges and thereby function in signalling and antioxidant defences. These oxidative modifications occur in response to a range of agents and stimuli, and can lead to the existence of multiple redox states for a given protein. To assess the role(s) of a protein in redox signalling and antioxidant defence, it is thus vital to be able to assess which of the multiple thiol redox states are present and to investigate how these alter under different conditions. While this can be done by a range of mass spectrometric-based methods, these are time-consuming, costly, and best suited to study abundant proteins or to perform an unbiased proteomic screen. One approach that can facilitate a targeted assessment of candidate proteins, as well as proteins that are low in abundance or proteomically challenging, is by electrophoretic mobility shift assays. Redox-modified cysteine residues are selectively tagged with a large group, such as a polyethylene glycol (PEG) polymer, and then the proteins are separated by electrophoresis followed by immunoblotting, which allows the inference of redox changes based on band shifts. However, the applicability of this method has been impaired by the difficulty of cleanly modifying protein thiols by large PEG reagents. To establish a more robust method for redox-selective PEGylation, we have utilised a Click chemistry approach, where free thiol groups are first labelled with a reagent modified to contain an alkyne moiety, which is subsequently Click-reacted with a PEG molecule containing a complementary azide function. This strategy can be adapted to study reversibly reduced or oxidised cysteines. Separation of the thiol labelling step from the PEG

Experimental data and theoretical predictions of the rate of electrophoretic clarification of colloidal suspensions

Energy Technology Data Exchange (ETDEWEB)

Johnson, T.J.; Davis, E.J. [University of Washington, Seattle, WA (USA). Dept. of Chemical Engineering

2000-05-01

An experimental and theoretical investigation of the electrophoretic clarification rate of colloidal suspensions was conducted. The suspensions included a coal-washing effluent and a model system of TiO{sub 2} particles. A parametric study of TiO{sub 2} suspensions was performed to validate an analysis of the electrophoretic motion of the clarification front formed between a clear zone and the suspension. To measure the electric field strength needed in the prediction of the location of the front, a moveable probe and salt bridge were connected to a reference electrode. Using the measured electric field strength, it was found that the numerical solution to the unit cell electrophoresis model agrees with the measured clarification rates. For suspensions with moderately thick electric double layers and high particle volume fractions the deviations from classical Smoluchowski theory are substantial, and the numerical analysis is in somewhat better agreement with the data than a prior solution of the problem. The numerical model reduces to the predictions of previous theories as the thickness of the electric double layer decreases, and it is in good agreement with the clarification rate measured for a coal-washing effluent suspension with thin electric double layers. 21 refs., 8 figs., 4 tabs.

Suspension chemistry and electrophoretic deposition of zirconia electrolyte on conducting and non-conducting substrates

International Nuclear Information System (INIS)

Das, Debasish; Basu, Rajendra N.

2013-01-01

Graphical abstract: - Highlights: • Stable suspension of yttria stabilized zirconia (YSZ) obtained in isopropanol medium. • Suspension chemistry and process parameters for electrophoretic deposition optimized. • Deposited film quality changed with iodine and water (dispersants) concentration. • Dense YSZ film (∼5 μm) fabricated onto non-conducting porous NiO-YSZ anode substrate. - Abstract: Suspensions of 8 mol% yttria stabilized zirconia (YSZ) particulates in isopropanol medium are prepared using acetylacetone, iodine and water as dispersants. The effect of dispersants concentration on suspension stability, particle size distribution, electrical conductivity and pH of the suspensions are studied in detail to optimize the suspension chemistry. Electrophoretic deposition (EPD) has been conducted to produce thin and dense YSZ electrolyte films. Deposition kinetics have been studied in depth and good quality films on conducting substrate are obtained at an applied voltage of 15 V for 3 min. YSZ films are also fabricated on non-conducting NiO-YSZ anode substrate using a steel plate on the reverse side of the substrate. Upon co-firing at 1400 °C for 6 h a dense YSZ film of thickness ∼5 μm is obtained. Such a half cell (anode + electrolyte) can be used to fabricate a solid oxide fuel cell on applying a suitable cathode layer

Covalently coating dextran on macroporous polyglycidyl methacrylate microsphere enabled rapid protein chromatographic separation

Energy Technology Data Exchange (ETDEWEB)

Zhang, Rongyue; Li, Qiang; Li, Juan; Zhou, Weiqing; Ye, Peili; Gao, Yang; Ma, Guanghui, E-mail: ghma@home.ipe.ac.cn; Su, Zhiguo

2012-12-01

were applied to rapid protein separation.

In Situ Magnetic Separation for Extracellular Protein Production

DEFF Research Database (Denmark)

Kappler, T.; Cerff, Martin; Ottow, Kim Ekelund

2009-01-01

A new approach for in situ product removal from bioreactors is presented in which high-gradient magnetic separation is used. This separation process was used for the adsorptive removal of proteases secreted by Bacillus licheniformis. Small, non-porous bacitracin linked magnetic adsorbents were...... was not influenced by the in situ product removal step. Protease production also remained the same after the separation step. Furthermore, degradation of the protease, which followed first order kinetics, was reduced by using the method. Using a theoretical modeling approach, we Could show that protease yield...... in total was enhanced by using in situ magnetic separation. The process described here is a promising technique to improve overall yield in No production processes which are often limited due to weak downstream operations, Potential limitations encountered during a bioprocess can be overcome...

Development of a novel fluorescent tag O-2-[aminoethyl]fluorescein for the electrophoretic separation of oligosaccharides

International Nuclear Information System (INIS)

Kazarian, Artaches A.; Smith, Jason A.; Hilder, Emily F.; Breadmore, Michael C.; Quirino, Joselito P.; Suttil, James

2010-01-01

This study describes the development of a novel fluorescent tag, O-2-[aminoethyl]fluorescein, for the separation of sugars by capillary electrophoresis with fluorescence detection using an argon ion laser. The tag was synthesised using three consecutive steps namely: esterification, alkylation and hydrolysis, specifically designed to offer a flexible way in which to make an assortment of fluorescent tags from cheap and readily available starting reagents (typically less than $1 per g of fluorescent tag). Via this flexible synthetic pathway, O-2-[aminoethyl]fluorescein was designed and synthesised with a spacer group to lower steric effects between the fluorescein backbone and the reducing end of the carbohydrate which were anticipated to improve the reactivity of the tag. The newly synthesised tag, O-2-[aminoethyl]fluorescein was evaluated against structurally similar commercial fluorescent motifs namely fluorescent 5-aminomethylfluorescein and non-fluorescent 5-aminofluorescein. Kinetic studies indicated that O-2-[aminoethyl]fluorescein showed similar labeling efficiencies as 5-aminomethylfluorescein, but were achieved in only 30 min, supporting the notion of improved reactivity of the spacer group. The sensitivity of O-2-[aminoethyl]fluorescein was evaluated using maltoheptaose with a detection limit of 1 nM obtained, which was slightly higher than that of 0.3 nM obtained with 5-aminomethylfluorescein, and was due to its lower quantum yield (0.24) when conjugated to the sugar. The separation performance of the tag was also benchmarked with the two commercial reagents using a range of corn syrup oligosaccharides, from 4 to 10 glucose units, typically found in rice starch. Separations were performed using an electrolyte containing 100 mM boric acid, tris at pH 8.65 as background electrolyte, 30 kV applied voltage, 50 μm I.D. x 40 cm (30 cm effective length) capillary. The novel tag showed better resolution of small oligosaccharides, G3 and G4, than the other two

Protein extraction and gel-based separation methods to analyze responses to pathogens in carnation (Dianthus caryophyllus L).

Science.gov (United States)

Ardila, Harold Duban; Fernández, Raquel González; Higuera, Blanca Ligia; Redondo, Inmaculada; Martínez, Sixta Tulia

2014-01-01

We are currently using a 2-DE-based proteomics approach to study plant responses to pathogenic fungi by using the carnation (Dianthus caryophyllus L)-Fusarium oxysporum f. sp. dianthi pathosystem. It is clear that the protocols for the first stages of a standard proteomics workflow must be optimized to each biological system and objectives of the research. The optimization procedure for the extraction and separation of proteins by 1-DE and 2-DE in the indicated system is reported. This strategy can be extrapolated to other plant-pathogen interaction systems in order to perform an evaluation of the changes in the host protein profile caused by the pathogen and to identify proteins which, at early stages, are involved or implicated in the plant defense response.

Electrophoretic deposition and field emission properties of patterned carbon nanotubes

International Nuclear Information System (INIS)

Zhao Haifeng; Song Hang; Li Zhiming; Yuan Guang; Jin Yixin

2005-01-01

Patterned carbon nanotubes on silicon substrates were obtained using electrophoretic method. The carbon nanotubes migrated towards the patterned silicon electrode in the electrophoresis suspension under the applied voltage. The carbon nanotubes arrays adhered well on the silicon substrates. The surface images of carbon nanotubes were observed by scanning electron microscopy. The field emission properties of the patterned carbon nanotubes were tested in a diode structure under a vacuum pressure below 5 x 10 -4 Pa. The measured emission area was about 1.0 mm 2 . The emission current density up to 30 mA/cm 2 at an electric field of 8 V/μm has been obtained. The deposition of patterned carbon nanotubes by electrophoresis is an alternative method to prepare field emission arrays

Electrophoretic deposition of nickel zinc ferrite nanoparticles into microstructured patterns

Directory of Open Access Journals (Sweden)

Stefan J. Kelly

2016-05-01

Full Text Available Using DC electric fields, nickel-zinc ferrite (Ni0.5Zn0.5Fe2O4 nanoparticles (Dh =16.6 ± 3.6 nm are electrophoretically deposited onto silicon substrates to form dense structures defined by photoresist molds. Parameters such as electric field, bath composition, and deposition time are tuned to produce films ranging in thickness from 177 to 805 nm. The deposited films exhibit soft magnetic properties with a saturation magnetization of 60 emu/g and a coercivity of 2.6 kA/m (33 Oe. Additionally, the influence of the photoresist mold on the deposit profile is studied, and patterned films with different shapes (lines, squares, circles, etc. are demonstrated with feature sizes down to 5 μm.

Transport properties of metal-semiconductor junctions on n-type InP prepared by electrophoretic deposition of Pt nanoparticles

Czech Academy of Sciences Publication Activity Database

Yatskiv, Roman; Grym, Jan; Brus, V.V.; Černohorský, Ondřej; Maryanchuk, P.D.; Bazioti, C.; Dimitrakopulos, G.P.; Komninou, Ph.

2014-01-01

Roč. 29, č. 4 (2014), Article number 045017 ISSN 0268-1242 R&D Projects: GA MŠk LD12014 Institutional support: RVO:67985882 Keywords : electrophoretic deposition * Pt nanoparticles * Schottky diodes Subject RIV: JA - Electronics ; Optoelectronics, Electrical Engineering Impact factor: 2.190, year: 2014

Engineering protein scaffolds for protein separation, biocatalysis and nanotechnology applications

Science.gov (United States)

Liu, Fang

Globally, there is growing appreciation for developing a sustainable economy that uses eco-efficient bio-processes. Biotechnology provides an increasing range of tools for industry to help reduce cost and improve environmental performance. Inspired by the naturally evolved machineries of protein scaffolds and their binding ligands, synthetic protein scaffolds were engineered based on cohesin-dockerin interactions and metal chelating peptides to tackle the challenges and make improvements in three specific areas: (1) protein purification, (2) biofuel cells, and (3) nanomaterial synthesis. The first objective was to develop efficient and cost-effective non-chromatographic purification processes to purify recombinant proteins in an effort to meet the dramatically growing market of protein drugs. In our design, the target protein was genetically fused with a dockerin domain from Clostridium thermocellum and direct purification and recovery was achieved using thermo-responsive elastin-like polypeptide (ELP) scaffold containing the cohesin domain from the same species. By exploiting the highly specific interaction between the dockerin and cohesin domain and the reversible aggregation property of ELP, highly purified and active dockerin-tagged proteins, such as endoglucanase CelA, chloramphenicol acetyl transferase (CAT) and enhanced green fluorescence protein (EGFP), were recovered directly from crude cell extracts in a single purification step with yields achieving over 90%. Incorporation of a self-cleaving intein domain enabled rapid removal of the affinity tag from the target proteins by another cycle of thermal precipitation. The purification cost can be further reduced by regenerating and recycling the ELP-cohesin capturing scaffolds. However, due to the high binding affinity between cohesin and dockerin domains, the bound dockerin-intein tag cannot be completely disassociated from ELP-cohesin scaffold after binding. Therefore, a truncated dockerin with the calcium

Deficiency of toxin-binding protein activity in mutants of sugarcane clone H54-775 as it relates to disease resistance

International Nuclear Information System (INIS)

Strobel, G.A.; Steiner, G.W.; Byther, R.

1975-01-01

Three mutants selected from a population of sugarcane clone H54-775 that had been irradiated with 3 kR γ-radiation all lacked toxin-binding protein activity. This activity previously had been shown to be essential for eye spot disease susceptibility and was demonstrated in the susceptible parent clone H54-775. In one mutant, the biochemical, immunochemical, and electrophoretic mobilities of the toxin-binding protein were all modified

Separation of oligopeptides, nucleobases, nucleosides and nucleotides using capillary electrophoresis/electrochromatography with sol-gel modified inner capillary wall.

Science.gov (United States)

Svobodová, Jana; Kofroňová, Olga; Benada, Oldřich; Král, Vladimír; Mikšík, Ivan

2017-09-29

The aim of this article is to study the modification of an inner capillary wall with sol-gel coating (pure silica sol-gel or silica sol-gel containing porphyrin-brucine conjugate) and determine its influence on the separation process using capillary electrophoresis/electrochromatography method. After modification of the inner capillary surface the separation of analytes was performed using two different phosphate buffers (pH 2.5 and 9.0) and finally the changes in electrophoretic mobilities of various samples were calculated. To confirm that the modification of the inner capillary surface was successful, the parts of the inner surfaces of capillaries were observed using scanning electron microscopy. The analytes used as testing samples were oligopeptides, nucleosides, nucleobases and finally nucleotides. Copyright © 2017 Elsevier B.V. All rights reserved.

Development and validation of an analytical method for the separation and determination of major bioactive curcuminoids in Curcuma longa rhizomes and herbal products using non-aqueous capillary electrophoresis.

Science.gov (United States)

Anubala, S; Sekar, R; Nagaiah, K

2014-06-01

A simple, fast and efficient non-aqueous capillary electrophoresis method (NACE) was developed for the simultaneous determination of three major bioactive curcuminoids (CMNs) in Curcuma longa rhizomes and its herbal products. Good separation, resolution and reproducibility were achieved with the background electrolyte (BGE) consisting a mixture of 15.0 mM sodium tetraborate and 7.4 mM sodium hydroxide (NaOH) in 2:10:15 (v/v/v) of water, 1-propanol, and methanol. The influences of background electrolyte, sodium hydroxide, water, sodium dodecyl sulfate and hydroxylpropyl-β-cyclodextrin on separations were investigated. The separation was carried out in a fused-silica capillary tube with reverse polarity. Hydrodynamic injection of 25mbar for 12s was used for injecting samples and a voltage of 28 kV was applied for separation. The ultrasonication method was used for the extraction of CMNs from the turmeric herbal products and the extract was filtered and directly injected without any further treatments. The limits of detection and quantification were less than 5.0 and 14.6 µg/ml respectively for all CMNs. The percentage recoveries for CMNs were >97.2% (%RSD, <2.62). The results obtained by the method were compared with existing spectrophotometric and HPLC methods. The related compounds in the extract did not interfere in the determination of CMNs. The proposed NACE method is better than existing chromatographic and electrophoretic methods in terms of simple electrophoretic medium, fast analysis and good resolution. Copyright © 2014 Elsevier B.V. All rights reserved.

Dynamic light scattering study on phase separation of a protein-water mixture: Application on cold cataract development in the ocular lens

Science.gov (United States)

Petta, V.; Pharmakakis, N.; Papatheodorou, G. N.; Yannopoulos, S. N.

2008-06-01

We present a detailed dynamic light scattering study of the phase separation in the ocular lens emerging during cold cataract development. Cold cataract is a phase separation effect that proceeds via spinodal decomposition of the lens cytoplasm with cooling. The intensity autocorrelation functions of the lens protein content are analyzed with the aid of two methods, providing information on the populations and dynamics of the scattering elements associated with cold cataract. It is found that the temperature dependence of many measurable parameters changes appreciably at the characteristic temperature ˜16±1°C which is associated with the onset of cold cataract. By extending the temperature range of this work to previously inaccessible regimes, i.e., well below the phase separation or coexistence curve at Tcc , we have been able to accurately determine the temperature dependence of the collective and self-diffusion coefficients of proteins near the spinodal. The analysis showed that the dynamics of proteins bears some resemblance to the dynamics of structural glasses, where the apparent activation energy for particle diffusion increases below Tcc , indicating a highly cooperative motion. Application of ideas developed for studying the critical dynamics of binary protein-solvent mixtures, as well as the use of a modified Arrhenius equation, enabled us to estimate the spinodal temperature Tsp of the lens nucleus. The applicability of dynamic light scattering as a noninvasive, early-diagnostic tool for ocular diseases is also demonstrated in light of the findings of the present paper.

Search for gene mutations affecting protein structure in children of A-bomb survivors, 2

International Nuclear Information System (INIS)

Satoh, Chiyoko; Fujita, Mikio; Goriki, Kazuaki; Asakawa, Jun-ichi; Takahashi, Norio; Hamilton, H.B.; Hazama, Ryuji; Neel, J.V.

1984-01-01

Children who were born between May 1, 1946 and April 1, 1971 to survivor(s) exposed to A-bombing within 2,000 m from the hypocenter in Hiroshima and Nagasaki were selected as exposed group; their sex- and age-matched children born to survivor(s) who were exposed at 2,500 m or farther were selected as control group. When these children were in junior high school, mutation of protein structure was examined by using electrophoresis and by determining red cell enzymes with decreased activity and heat-unstable red cell enzymes. Electrophoretic study revealed a ''rare type of protein mutation'' in 635 of 12,242 individuals in the exposed group and in 448 of 10,154 individuals in the control group. The number of locuses in all proteins examined was calculated. The number of locuses per protein was corrected using the rate of parents' mutation type, and relative number of locuses were obtained. As a result, there was no difference in the mutation frequency per locus and generation between the exposed and control groups. Among children having red cell enzymes with decreased activity, mutant in triose phosphate isomerase was detected in one child in the exposed group, in whom electrophoretic pattern was normal and red cell enzymes were stable to heat. Heat-unstable red cell enzymes were seen in 9 children and their parents. However, family survey revealed genetic mutation in all instances irrespective of A-bombing. (Namekawa, K.)

Sol-gel synthesis of 45S5 bioglass – Prosthetic coating by electrophoretic deposition

Directory of Open Access Journals (Sweden)

Faure Joel

2013-11-01

Full Text Available In this work, the 45S5 bioactive glass has been prepared by the sol-gel process using an organic acid catalyst instead of nitric acid usually used. The physico-chemical and structural characterizations confirmed and validated the elemental composition of the resulting glass. In addition, the 45S5 bioactive glass powder thus obtained was successfully used to elaborate by electrophoretic deposition a prosthetic coating on titanium alloy Ti6Al4V.

Microencapsulated Electrophoretic Films for Electronic Paper Displays

Science.gov (United States)

Amundson, Karl

2003-03-01

Despite the dominance of liquid crystal displays, they do not perform some functions very well. While backlit liquid crystal displays can offer excellent color performance, they wash out in bright lighting and suffer from high power consumption. Reflective liquid crystal displays have limited brightness, making these devices challenging to read for long periods of time. Flexible liquid crystal displays are difficult to manufacture and keep stable. All of these attributes (long battery lifetime, bright reflective appearance, compatibility with flexible substrates) are traits that would be found in an ideal electronic paper display - an updateable substitute for paper that could be employed in electronic books, newspapers, and other applications. I will discuss technologies that are being developed for electronic-paper-like displays, and especially on particle-based technologies. A microencapsulated electrophoretic display technology is being developed at the E Ink corporation. This display film offers offer high brightness and an ink-on-paper appearance, compatibility with flexible substrates, and image stability that can lead to very low power consumption. I will present some of the physical and chemical challenges associated with making display films with high performance.

Evaluation of clinical, laboratory, and electrophoretic profiles for diagnosis of malnutrition in hospitalized dogs

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Andrei Kelliton Fabretti

2015-02-01

Full Text Available Malnutrition is a major factor associated with increased rates of mortality and readmission, longer hospital stays, and greater health care spending. Recognizing malnourished or at-risk animals allows for nutritional intervention and improved prognosis. This study evaluated the association between clinical, laboratory, and electrophoretic variables and the nutritional status (NS of hospitalized dogs in order to generate a profile of the sick dog and to facilitate the diagnosis of malnutrition. We divided 215 dogs into groups according to the severity of the underlying disease and we determined the clinical NS based on the assessment of the body condition score and the muscle mass score. The NS was classified as clinically well nourished, clinical moderate malnutrition, or clinical severe malnutrition. Statistical analyses were conducted by using the chi-square test or Fisher’s exact test; the Kruskal-Wallis test was used for continuous variables. A strong association was found between malnutrition and the severity of the underlying disease. In hospitalized dogs, low body mass index values, anemia, low hemoglobin concentrations, high fibrinogen concentrations, decreased albumin fraction, and increased gamma-globulin fraction (in electrophoresis were associated with malnutrition, reinforcing the classification of poor NS. However, the skin and coat characteristics, the total number of lymphocytes, blood glucose, cholesterol, and total protein concentration were not found to be good predictors of NS.

Electrophoretic deposition of ultrasonicated and functionalized nanomaterials for multifunctional composites

Science.gov (United States)

An, Qi

Recent advances in the synthesis and characterization of nanostructured composite materials have enabled a broad range of opportunities for engineering the properties of polymer-matrix materials. Carbon nanotubes (CNTs) are known to have exceptional mechanical, electrical and thermal properties. Because of their small size, CNTs can occupy regions between traditional micro-scale reinforcements and create a hierarchical micro/nano structure spanning several orders of magnitude. Since CNTs possess critical reinforcement dimensions below 100 nm, new opportunities exist for tailoring the fiber/matrix interphase regions and ultimately the mechanical and electrical performance of advanced fiber-composites with minimal impact on the fiber-dominated properties. This growing interest in nanoscale hybridization with conventional fiber reinforcement has highlighted the need to develop new processing techniques for successful CNT integration. In this work, a novel and industrially scalable approach for producing multi-scale hybrid carbon nanotube/fiber composites using an electrophoretic deposition (EPD) technique has been studied as an alternative to in situ chemical vapor deposition growth (CVD). EPD is a widely used industrial coating process employed in areas ranging from automotive to electronics production. The method has a number of benefits which include low energy use and the ability to homogenously coat complex shapes with well adhered films of controlled thickness and density. A stable aqueous dispersion of multi-walled carbon nanotubes (MWCNTs) was produced using a novel ozonolysis and ultrasonication (USO) technique that results in dispersion and functionalization in a single step. Networks of CNTs span between adjacent fibers and the resulting composites exhibit significant increases in electrical conductivity and considerable improvements in the interlaminar shear strength and fracture toughness. In order to better understand the underlying mechanisms behind the

Determination of total antioxidant capacity of milk by CUPRAC and ABTS methods with separate characterisation of milk protein fractions.

Science.gov (United States)

Çekiç, Sema Demirci; Demir, Aslı; Başkan, Kevser Sözgen; Tütem, Esma; Apak, Reşat

2015-05-01

Most milk-applied antioxidant assays in literature are based on the isolation and quantification of individual antioxidative compounds, whereas total antioxidant capacity (TAC) gives a more holistic picture due to cooperative action of antioxidants. Recently, the cupric reducing antioxidant capacity (CUPRAC) method has been modified to measure the antioxidant capacities of thiol-containing proteins, where the classical ammonium acetate buffer - that may otherwise precipitate proteins- was replaced with concentrated urea buffer (able to expose embedded thiol groups of proteins to oxidative attack) adjusted to pH 7.0. Thus, antioxidant capacity of milk was investigated with two competing TAC assays, namely CUPRAC and ABTS (2,2'-azinobis(3-ethylbenzothiazoline-6-sulphonic acid))/persulphate, because only these assays were capable of evaluating protein contribution to the observed TAC value. As milk fat caused turbidity, experiments were carried out with skim milk or defatted milk samples. To determine TAC, modified CUPRAC method was applied to whole milk, separated and redissolved protein fractions, and the remaining liquid phase after necessary operations. Both TAC methods were investigated for their dilution sensitivity and antioxidant power assessment of separate milk fractions such as casein and whey. Proteins like β-lactoglobulin and casein (but not simple thiols) exhibited enhanced CUPRAC reactivity with surfactant (SDS) addition. Addition of milk protein fractions to whole skim milk produced significant 'negative-biased' deviations (up to -26% relative standard error) from TAC absorbance additivity in the application of the ABTS method, as opposed to that of the CUPRAC method less affected by chemical deviations from Beer's law thereby producing much smaller deviations from additivity (i.e. the property of additivity is valid when the measured TAC of a mixture is equal to the sum of individual antioxidant capacities of its constituents).

Hepatitis C Virus E1 and E2 Proteins Used as Separate Immunogens Induce Neutralizing Antibodies with Additive Properties.

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Elodie Beaumont

Full Text Available Various strategies involving the use of hepatitis C virus (HCV E1 and E2 envelope glycoproteins as immunogens have been developed for prophylactic vaccination against HCV. However, the ideal mode of processing and presenting these immunogens for effective vaccination has yet to be determined. We used our recently described vaccine candidate based on full-length HCV E1 or E2 glycoproteins fused to the heterologous hepatitis B virus S envelope protein to compare the use of the E1 and E2 proteins as separate immunogens with their use as the E1E2 heterodimer, in terms of immunogenetic potential and the capacity to induce neutralizing antibodies. The specific anti-E1 and anti-E2 antibody responses induced in animals immunized with vaccine particles harboring the heterodimer were profoundly impaired with respect to those in animals immunized with particles harboring E1 and E2 separately. Moreover, the anti-E1 and anti-E2 antibodies had additive neutralizing properties that increase the cross-neutralization of heterologous strains of various HCV genotypes, highlighting the importance of including both E1 and E2 in the vaccine for an effective vaccination strategy. Our study has important implications for the optimization of HCV vaccination strategies based on HCV envelope proteins, regardless of the platform used to present these proteins to the immune system.

Preparation of guinea pig macrophage for electrophoretic experiments in space

Science.gov (United States)

1979-01-01

Methods of storage and cultivation of macrophage cells in preparation for space experiments were investigated. Results show that freezing and thawing immediately after extraction did not cause any change in viability or electrophoretic mobility of the cells. A prolonged storage at -80 C did cause cell damage as indicated by a 95% reduction in variable cells. Cell damage was decreased when Glycerol or Dimethyl Sulfoxide (DMSO) was added as a cryogenic protective agent. A 100% viability was observed in cultivation experiments after two weeks due to the additional serum. Results from gamma-glutamyl transpeptidase study showed a zero activity rate. It is suggested that a flat stationary field be used for the collection and use of macrophage. It was found that a 24-hour delay in obtaining macrophage cells helps to maintain a pure culture.

In vivo phosphorylation of a peptide tag for protein purification.

Science.gov (United States)

Goux, Marine; Fateh, Amina; Defontaine, Alain; Cinier, Mathieu; Tellier, Charles

2016-05-01

To design a new system for the in vivo phosphorylation of proteins in Escherichia coli using the co-expression of the α-subunit of casein kinase II (CKIIα) and a target protein, (Nanofitin) fused with a phosphorylatable tag. The level of the co-expressed CKIIα was controlled by the arabinose promoter and optimal phosphorylation was obtained with 2 % (w/v) arabinose as inductor. The effectiveness of the phosphorylation system was demonstrated by electrophoretic mobility shift assay (NUT-PAGE) and staining with a specific phosphoprotein-staining gel. The resulting phosphorylated tag was also used to purify the phosphoprotein by immobilized metal affinity chromatography, which relies on the specific interaction of phosphate moieties with Fe(III). The use of a single tag for both the purification and protein array anchoring provides a simple and straightforward system for protein analysis.

Partial separation of platelet and placental adenosine receptors from adenosine A2-like binding protein

International Nuclear Information System (INIS)

Zolnierowicz, S.; Work, C.; Hutchison, K.; Fox, I.H.

1990-01-01

The ubiquitous adenosine A2-like binding protein obscures the binding properties of adenosine receptors assayed with 5'-N-[ 3 H]ethylcarboxamidoadenosine [( 3 H]NECA). To solve this problem, we developed a rapid and simple method to separate adenosine receptors from the adenosine A2-like binding protein. Human platelet and placental membranes were solubilized with 1% 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate. The soluble platelet extract was precipitated with polyethylene glycol and the fraction enriched in adenosine receptors was isolated from the precipitate by differential centrifugation. The adenosine A2-like binding protein was removed from the soluble placental extract with hydroxylapatite and adenosine receptors were precipitated with polyethylene glycol. The specificity of the [ 3 H]NECA binding is typical of an adenosine A2 receptor for platelets and an adenosine A1 receptor for placenta. This method leads to enrichment of adenosine A2 receptors for platelets and adenosine A1 receptors for placenta. This provides a useful preparation technique for pharmacologic studies of adenosine receptors

Decreased Staphylococcus aureus and increased osteoblast density on nanostructured electrophoretic-deposited hydroxyapatite on titanium without the use of pharmaceuticals.

Science.gov (United States)

Mathew, Dennis; Bhardwaj, Garima; Wang, Qi; Sun, Linlin; Ercan, Batur; Geetha, Manisavagam; Webster, Thomas J

2014-01-01

Plasma-spray deposition of hydroxyapatite on titanium (Ti) has proven to be a suboptimal solution to improve orthopedic-implant success rates, as demonstrated by the increasing number of orthopedic revision surgeries due to infection, implant loosening, and a myriad of other reasons. This could be in part due to the high heat involved during plasma-spray deposition, which significantly increases hydroxyapatite crystal growth into the nonbiologically inspired micron regime. There has been a push to create nanotopographies on implant surfaces to mimic the physiological nanostructure of native bone and, thus, improve osteoblast (bone-forming cell) functions and inhibit bacteria functions. Among the several techniques that have been adopted to develop nanocoatings, electrophoretic deposition (EPD) is an attractive, versatile, and effective material-processing technique. The in vitro study reported here aimed to determine for the first time bacteria responses to hydroxyapatite coated on Ti via EPD. There were six and three times more osteoblasts on the electrophoretic-deposited hydroxyapatite on Ti compared with Ti (control) and plasma-spray-deposited hydroxyapatite on Ti after 5 days of culture, respectively. Impressively, there were 2.9 and 31.7 times less Staphylococcus aureus on electrophoretic-deposited hydroxyapatite on Ti compared with Ti (control) and plasma-spray-deposited hydroxyapatite on Ti after 18 hours of culture, respectively. Compared with uncoated Ti and plasma-sprayed hydroxyapatite coated on Ti, the results provided significant promise for the use of EPD to improve bone-cell density and be used as an antibacterial coating without resorting to the use of antibiotics.

A reassessment of synchronous fluorescence in the separation of Trp and Tyr contributions in protein emission and in the determination of conformational changes

DEFF Research Database (Denmark)

Bobone, Sara; van de Weert, Marco; Stella, Lorenzo

2014-01-01

solvents, as well as a real protein (bovine serum albumin). Unfortunately, synchronous spectra were found to be unreliable in the separation of Trp and Tyr emission components in proteins. A simple alternative approach based on the deconvolution of emission spectra is presented. In addition, an equation...

Electrophoretic study of enzymes from cereal aphid populations : 4. Detection of hidden genetic variation within populations of the grain aphid Sitobion avenae (F.) (Hemiptera: Aphididae).

Science.gov (United States)

Loxdale, H D; Rhodes, J A; Fox, J S

1985-07-01

A study of variation in three peptidases (PEP-3 to -5) in a parthenogenetic S. avenae field population at Rothamsted using serial one-dimensional polyacrylamide gel electrophoresis (involving changes of gel concentration and electrophoretic run-time) increased the overall number of "allozymes" (mobility variants) detected from 10 under standard conditions (6% gels, 2 h run-time) to 22, as well as revealing putative heterozygous banding patterns under some test conditions. However, an examination of another enzyme, 6-phosphogluconate dehydrogenase (6-PGD) in a sample collected at Rothamsted the following year failed, using a combination of serial methods (changes of gel concentration) and isoelectric focusing, to increase the total number of 6-PGD bands separated (seven, none of which appeared to be allelic in origin). Nevertheless, some major bands were split into several bands, whilst other infrequent bands were either gained or lost. The findings are briefly discussed.

Chemical structure, comparison antioxidant capacity and separation antioxidant of hen, duck and quail egg white protein hydrolysate

Science.gov (United States)

Fatah, A.; Meihu, M.; Ning, Q.; Setiani, B. E.; Bintoro, V. P.

2018-01-01

Amino acid linkages as proteins are nutritional substance which important for diet intake. Purification protein procesing undergo heating procedure process followed by additional of proteolytic enzymes or acid had been resulting in protein hydrolysates. A protein hydrolysate describe as many free amino acids bound together through a complex mixture of peptides. Egg white protein hydrolysates is one of subject interested to study for human health or industry product. The objectives of the research are to determine and identification the antioxidant derived from egg white hydrolysate protein. Identification of chemical structure of albumen and albumen protein hydrolysate was examine using IR Spectrophotometry. While comparison of antioxidant capacity and antioxidant separation egg albumen was also investigate using FTIR method (Fourier Transform Infrared Spectroscopy). Hen, duck and quail albumen egg white and on hydrolisate form were used as research materials. The results were showing that different time and enzyme of hydrolysis were not influence at secondary structure of hydrolysate albumen protein. Phytochemical content such as alcohol and hydroxyl compound which have potential as functional group of antioxidant were detected in all of the samples. Their results of radical scavenging activities samples hydrolyzed by pepsin were respectively 89.40%, 50.25% and 85.13%. Whereas the radical scavenging activities of hydrolysates hydrolyzed by papain were 72.85%, 61% and 76.45% respectively.

On the role of the indifferent electrolyte LiCl in electrophoretic deposition of hydroxyapatite from 2-propanol dispersions

Czech Academy of Sciences Publication Activity Database

Drdlík, D.; Sláma, M.; Hadraba, Hynek; Drdlíková, K.; Cihlář, J.

2016-01-01

Roč. 42, č. 15 (2016), s. 16529-16534 ISSN 0272-8842 R&D Projects: GA MŠk(CZ) LQ1601 Institutional support: RVO:68081723 Keywords : Electrophoretic deposition * Electrical conductivity * Thick layer * Surface roughness * Hydroxyapatite Subject RIV: JH - Ceramics, Fire-Resistant Materials and Glass Impact factor: 2.986, year: 2016

Electrophoretic-deposited CNT/MnO{sub 2} composites for high-power electrochemical energy storage/conversion applications

Energy Technology Data Exchange (ETDEWEB)

Xiao Wei; Xia Hui; Fuh, Jerry Y H; Lu Li, E-mail: luli@nus.edu.s [Department of Mechanical Engineering, National University of Singapore, 9 Engineering Drive 1, Singapore 117576 (Singapore)

2010-05-01

CNT/MnO{sub 2} (birnessite-type) composite films have been successfully deposited on Ni-foil substrate via electrophoretic deposition (EPD). The unique EPD CNT/MnO{sub 2} composite film electrode shows enhanced electrical conductivity, good contact between composite films and the substrate and open porous structure, which makes the EPD composite films a promising electrode for high-power supercapacitors and lithium ion batteries.

Electrophoretic studies on corrosion products from secondary side on nuclear steam generator

International Nuclear Information System (INIS)

Das, P.C.; Velmurugan, S.; Sinha, P.K.; Mathur, P.K.

1988-01-01

Electrophoretic mobilities of aqueous suspensions of the steam generator crud samples were determined at 25degC and the zeta potentials were calculated using Smoluchowski equation assuming k. a>>1. The determined (pH) pzc values for different crud samples were compared with the available literature data for magnetite (the major constituent of S.G. crud). The observed shift in the (δ) zeta potentials and crud suspensions towards more negative values in the presence of anions such as Cl - , NO 3 - and SO 4 2- , used for maintaining constant ionic strengths, suggested probable adsorption of these anions on the oxide solution interface. (author). 10 refs

Stress-Triggered Phase Separation Is an Adaptive, Evolutionarily Tuned Response

Energy Technology Data Exchange (ETDEWEB)

Riback, Joshua A.; Katanski, Christopher D.; Kear-Scott, Jamie L.; Pilipenko, Evgeny V.; Rojek, Alexandra E.; Sosnick, Tobin R.; Drummond, D. Allan

2017-03-01

In eukaryotic cells, diverse stresses trigger coalescence of RNA-binding proteins into stress granules. In vitro, stress-granule-associated proteins can demix to form liquids, hydrogels, and other assemblies lacking fixed stoichiometry. Observing these phenomena has generally required conditions far removed from physiological stresses. We show that poly(A)-binding protein (Pab1 in yeast), a defining marker of stress granules, phase separates and forms hydrogels in vitro upon exposure to physiological stress conditions. Other RNA-binding proteins depend upon low-complexity regions (LCRs) or RNA for phase separation, whereas Pab1’s LCR is not required for demixing, and RNA inhibits it. Based on unique evolutionary patterns, we create LCR mutations, which systematically tune its biophysical properties and Pab1 phase separation in vitro and in vivo. Mutations that impede phase separation reduce organism fitness during prolonged stress. Poly(A)-binding protein thus acts as a physiological stress sensor, exploiting phase separation to precisely mark stress onset, a broadly generalizable mechanism.

Selective separation and concentration of antihypertensive peptides from rapeseed protein hydrolysate by electrodialysis with ultrafiltration membranes.

Science.gov (United States)

He, Rong; Girgih, Abraham T; Rozoy, Elodie; Bazinet, Laurent; Ju, Xing-Rong; Aluko, Rotimi E

2016-04-15

Rapeseed protein isolate was subjected to alcalase digestion to obtain a protein hydrolysate that was separated into peptide fractions using electrodialysis with ultrafiltration membrane (EDUF) technology. The EDUF process (6h duration) led to isolation of three peptide fractions: anionic (recovered in KCl-1 compartment), cationic (recovered in KCl-2 compartment), and those that remained in the feed compartment, which was labeled final rapeseed protein hydrolysate (FRPH). As expected the KCl-1 peptides were enriched in negatively-charged (43.57%) while KCl-2 contained high contents of positively-charged (28.35%) amino acids. All the samples inhibited angiotensin converting enzyme (ACE) and renin activities in dose-dependent manner with original rapeseed protein hydrolysate having the least ACE-inhibitory IC50 value of 0.0932±0.0037 mg/mL while FRPH and KCl-2 had least renin-inhibitory IC50 values of 0.47±0.05 and 0.55±0.06 mg/mL, respectively. Six hours after oral administration (100 mg/kg body weight) to spontaneously hypertensive rats, the FRPH produced the maximum systolic blood pressure reduction of -51 mmHg. Copyright © 2015 Elsevier Ltd. All rights reserved.

On-line coupling of sample preconcentration by LVSEP with gel electrophoretic separation on T-channel chips.

Science.gov (United States)

Kitagawa, Fumihiko; Kinami, Saeko; Takegawa, Yuuki; Nukatsuka, Isoshi; Sueyoshi, Kenji; Kawai, Takayuki; Otsuka, Koji

2017-01-01

To achieve an on-line coupling of the sample preconcentration by a large-volume sample stacking with an electroosmotic flow pump (LVSEP) with microchip gel electrophoresis (MCGE), a sample solution, a background solution for LVSEP and a sieving solution for MCGE were loaded in a T-form channel and three reservoirs on PDMS microchips. By utilizing the difference in the flow resistance of the two channels, a low-viscosity sample and a viscous polymer solution were easily introduced into the LVSEP and MCGE channels, respectively. Fluorescence imaging of the sequential LVSEP-MCGE processes clearly demonstrated that a faster stacking of anionic fluorescein and successive introduction into the MCGE channel can be carried out on the T-channel chip. To evaluate the preconcentration performance, a conventional MCZE analysis of fluorescein on the cross-channel chip was compared with LVSEP-MCGE on the short T-channel chip, and as a result that the value of sensitive enhancement factor (SEF) was estimated to be 370. The repeatability of the peak height was good with the RSD value of 3.2%, indicating the robustness of the enrichment performance. In the successive LVSEP-MCGE analysis of φX174/HaeIII digest, the DNA fragments were well enriched to a sharp peak in the LVSEP channel, and they were separated in the MCGE channel, whose electropherogram was well-resembled with that in the conventional MCGE. The values of SEF for the DNA fragments were calculated to be ranging from 74 to 108. Thus, the successive LVSEP-MCGE analysis was effective for both preconcentrating and separating DNA fragments. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Analysis of electrophoretic soil humic acids fractions by reversed-phase high performance liquid chromatography with on-line absorbance and fluorescence detection.

Science.gov (United States)

Trubetskoj, Oleg A; Richard, Claire; Guyot, Ghislain; Voyard, Guillaume; Trubetskaya, Olga E

2012-06-22

A combination of reversed-phase high performance liquid chromatography (RP HPLC) with on-line absorbance and fluorescence detection was used for analysis of chernozem soil humic acids (HAs) and their fractions A, B and C+D with different electrophoretic mobility (EM) and molecular size (MS). Samples were injected onto the column at the identical volume and absorbance. All chromatograms exhibit the resolution of seven peaks. The estimation of relative recovery of HAs and fractions from the reverse-phase column has been done. High MS fraction A, which possesses the low EM, is essentially more hydrophobic (73% of the fraction amount remained adsorbed on the column) and aliphatic than medium MS and EM fraction B (33% of the fraction amount remained adsorbed on the column). The most hydrophilic and aromatic properties belong to low MS fraction C+D, which possess the highest EM and practically was not adsorbed on the column. The hydrophobicity of the bulk HAs lies within the range of fractions hydrophobicity. The absorption spectra of bulk HAs, electrophoretic fractions A, B, C+D and corresponding RP HPLC peaks were featureless but had differences in the values of absorbance ratio at 300 and 400 nm (A3/A4). For fractions A and B this ratio gradually decreased from peak 1 to 7 (from 3.05 to 2.80 and 3.00 to 2.40, respectively). This trend was less pronounced in HAs and practically absent in fraction C+D, where ratio A3/A4 varied within a small range. The strong relationship between fluorescence properties, EM, MS, polarity and aliphaticity/aromaticity of HAs fractions was found. Humic and protein-like fluorescence had different polarity nature. The protein-like fluorescence is located in humic material which irreversibly adsorbed on the reverse-phase column and not subjected to RP HPLC characterization. The humic-like fluorescence at Ex/Em 270/450 nm is mostly located in the hydrophilic peak of low MS fraction C+D. Taking into account that high MS fraction A consisted

Fluid Delivery System For Capillary Electrophoretic Applications.

Science.gov (United States)

Li, Qingbo; Liu, Changsheng; Kane, Thomas E.; Kernan, John R.; Sonnenschein, Bernard; Sharer, Michael V.

2002-04-23

An automated electrophoretic system is disclosed. The system employs a capillary cartridge having a plurality of capillary tubes. The cartridge has a first array of capillary ends projecting from one side of a plate. The first array of capillary ends are spaced apart in substantially the same manner as the wells of a microtitre tray of standard size. This allows one to simultaneously perform capillary electrophoresis on samples present in each of the wells of the tray. The system includes a stacked, dual carrousel arrangement to eliminate cross-contamination resulting from reuse of the same buffer tray on consecutive executions from electrophoresis. The system also has a gel delivery module containing a gel syringe/a stepper motor or a high pressure chamber with a pump to quickly and uniformly deliver gel through the capillary tubes. The system further includes a multi-wavelength beam generator to generate a laser beam which produces a beam with a wide range of wavelengths. An off-line capillary reconditioner thoroughly cleans a capillary cartridge to enable simultaneous execution of electrophoresis with another capillary cartridge. The streamlined nature of the off-line capillary reconditioner offers the advantage of increased system throughput with a minimal increase in system cost.

The bacterial defensin resistance protein MprF consists of separable domains for lipid lysinylation and antimicrobial peptide repulsion.

Directory of Open Access Journals (Sweden)

Christoph M Ernst

2009-11-01

Full Text Available Many bacterial pathogens achieve resistance to defensin-like cationic antimicrobial peptides (CAMPs by the multiple peptide resistance factor (MprF protein. MprF plays a crucial role in Staphylococcus aureus virulence and it is involved in resistance to the CAMP-like antibiotic daptomycin. MprF is a large membrane protein that modifies the anionic phospholipid phosphatidylglycerol with l-lysine, thereby diminishing the bacterial affinity for CAMPs. Its widespread occurrence recommends MprF as a target for novel antimicrobials, although the mode of action of MprF has remained incompletely understood. We demonstrate that the hydrophilic C-terminal domain and six of the fourteen proposed trans-membrane segments of MprF are sufficient for full-level lysyl-phosphatidylglycerol (Lys-PG production and that several conserved amino acid positions in MprF are indispensable for Lys-PG production. Notably, Lys-PG production did not lead to efficient CAMP resistance and most of the Lys-PG remained in the inner leaflet of the cytoplasmic membrane when the large N-terminal hydrophobic domain of MprF was absent, indicating a crucial role of this protein part. The N-terminal domain alone did not confer CAMP resistance or repulsion of the cationic test protein cytochrome c. However, when the N-terminal domain was coexpressed with the Lys-PG synthase domain either in one protein or as two separate proteins, full-level CAMP resistance was achieved. Moreover, only coexpression of the two domains led to efficient Lys-PG translocation to the outer leaflet of the membrane and to full-level cytochrome c repulsion, indicating that the N-terminal domain facilitates the flipping of Lys-PG. Thus, MprF represents a new class of lipid-biosynthetic enzymes with two separable functional domains that synthesize Lys-PG and facilitate Lys-PG translocation. Our study unravels crucial details on the molecular basis of an important bacterial immune evasion mechanism and it may help

The electrophoretic softness of the surface of Staphylococcus epidermidis cells grown in a liquid medium and on a solid agar

NARCIS (Netherlands)

Kiers, PJM; van der Mei, HC; Busscher, HJ; Bos, R.R.M.

Many Staphylococcus epidermidis strains possess capsule or slime layers and consequently the staphylococcal cell surface should be regarded as a soft, polyelectrolyte layer allowing electrophoretic fluid flow through a layer of fixed charges. The presence of such a soft layer decreases the energy

Comparative aspects of basic chromatin proteins in dinoflagellates.

Science.gov (United States)

Rizzo, P J

1981-01-01

Previous work on histone-like proteins in dinoflagellates is summarized, together with some new data to give an overview of basic proteins in these algae. The first two dinoflagellates studied were both found to contain one major acid-soluble protein that migrated to the same position in acidic-urea gels. When several other genera were studied however, it became apparent that the histone-like proteins from different dinoflagellates were similar but not identical. In view of the great diversity of living dinoflagellates it is speculated that further differences in dinoflagellate basic chromatin proteins will be revealed. Electrophoretic data from the eukaryotic (endosymbiont) nucleus of Peridinium balticum showed the presence of five major components. It is speculated that two of these proteins represent an H1-like doublet and two others correspond to the highly conserved histones H3 and H4. The fifth component is a new histone that may substitute for H2A and H2B in the nucleosome. Because histones and nucleosomes are present in all higher organisms but completely lacking in procaryotes, studies on basic proteins in dinoflagellates will provides insights into the evolution of histones and eucaryotic chromatin organization.

The role of the rheological properties of non-newtonian fluids in controlling dispersive mixing in a batch electrophoretic cell with Joule heating

Directory of Open Access Journals (Sweden)

M.A. Bosse

2001-03-01

Full Text Available The problem of the effect of Joule heating generation on the hydrodynamic profile and the solute transport found in electrophoretic devices is addressed in this article. The research is focused on the following two problems: The first one is centered around the effect of Joule heating on the hydrodynamic velocity profile and it is referred to as "the carrier fluid problem." The other one is related to the effect of Joule heating on the solute transport inside electrophoretic cells and it is referred to as "the solute problem". The hydrodynamic aspects were studied first to yield the velocity profiles required for analysis of the solute transport problem. The velocity profile obtained in this study is analytical and the results are valid for non-Newtonian fluids carriers. To this end, the power-law model was used to study the effect of the rheology of the material in conjunction with the effect of Joule heating generation inside batch electrophoretic devices. This aspect of the research was then effectively used to study the effect of Joule heating generation on the motion of solutes (such as macromolecules under the influence of non-Newtonian carriers. This aspect of the study was performed using an area-averaging approach that yielded analytical results for the effective diffusivity of the device.

Is pulsed electric field still effective for RNA separation in capillary electrophoresis?

Science.gov (United States)

Li, Zhenqing; Dou, Xiaoming; Ni, Yi; Chen, Qinmiao; Cheng, Shuyi; Yamaguchi, Yoshinori

2012-03-16

Pulsed field capillary electrophoresis (PFCE) is a predominant technique to cope with difficulties in resolving large DNA strands, yet it is still unclear whether pulsed electric field is effective for the separation of higher mass RNA. In this paper we focused on the role of pulsed electric field in large RNA fragments analysis by comparing RNA separation performance in PFCE with that in constant field CE. Separation performance in terms of migration mobility, plate numbers, resolution, and selectivity has been tested for the analysis of RNA from 0.1 to 10.0 kilo nucleotide (knt) under different electrophoretic conditions. Denaturation, important to obtain uniform and identifiable peaks, was accomplished by heating the sample in 4.0M urea prior to analysis and the presence of 4.0M urea in the electrophoresis buffer. Results demonstrate that unlike DNA in PFCE, the pulsed electric field mainly affects the separation performance of RNA between 0.4 and 2.0 knt. The migration mobility of long RNA fragments is not a strong function of modulation depth and pulsed frequency. Moreover, the logarithm of RNA mobility is almost inversely proportional to the logarithm of molecule size up to 6.0 knt with correlation coefficient higher than 0.99 in all the polymer concentrations measured here. Resonance frequency of RNA in PFCE was also observed. While these initial experiments show no distinct advantages of using PFCE for RNA separation, they do take further step toward characterizing the migration behavior of RNA under pulsed field conditions. Copyright © 2012 Elsevier B.V. All rights reserved.

Optimizing the fabrication of carbon nanotube electrode for effective capacitive deionization via electrophoretic deposition strategy

Directory of Open Access Journals (Sweden)

Simeng Zhang

2018-04-01

Full Text Available In order to obtain superior electrode performances in capacitive deionization (CDI, the electrophoretic deposition (EPD was introduced as a novel strategy for the fabrication of carbon nanotube (CNT electrode. Preparation parameters, including the concentration of slurry components, deposition time and electric field intensity, were mainly investigated and optimized in terms of electrochemical characteristic and desalination performance of the deposited CNT electrode. The SEM image shows that the CNT material was deposited homogeneously on the current collector and a non-crack surface of the electrode was obtained. An optimal preparation condition of the deposited CNT electrode was obtained and specified as the Al (NO33 M concentration of 1.3 × 10−2 mol/L, the deposition time of 30 min and the electric field intensity of 15 V/cm. The obtained electrode performs an increasing specific mass capacitance of 33.36 F/g and specific adsorption capacity of 23.93 mg/g, which are 1.62 and 1.85 times those of the coated electrode respectively. The good performance of the deposited CNT electrode indicates the promising application of the EPD methodology in subsequent research and fabrication of the CDI electrodes for CDI process. Keywords: Carbon nanotube, Water treatment, Desalination, Capacitive deionization, Electrode fabrication, Electrophoretic deposition

Characterization of CNT-MnO_2 nanocomposite by electrophoretic deposition as potential electrode for supercapacitor

International Nuclear Information System (INIS)

Darari, Alfin; Rismaningsih, Nurmanita; Ardiansah, Hafidh Rahman; Arifin,; Ningrum, Andini Novia; Subagio, Agus

2016-01-01

Energy crisis that occured in Indonesia suggests that energy supply could not offset the high rate request and needs an electric energy saving device which can save high voltage, safety, and unlimited lifetime. The weakness of batteries is durable but has a low power density while the capacitor has a high power density but it doesn’t durable. The renewal of this study is CNT-MnO_2 thin film fabrication method using electrophoretic deposition. Electrophoretic deposition is a newest method to deposited CNT using power supply with cheap, and make a good result. The result of FTIR analysis showed that the best CNT-MnO_2 composition is 75:25 and C-C bond is detected in fingerprint area. The result is electrode thin film homogen and characterized by X-ray diffraction (XRD) peaks 2θ=26,63° is characterization of graphite, and 2θ=43,97° is characterization of diamond Carbon type and measured by Scherrer formula results 52,3 nm material average size .EIS test results its capacitance about 7,86 F. from the data it can be concluded that CNT-MnO_2 potential electrode very promising for further study and has a potential to be a high capacitance, and fast charge supercapacitor which can be applied for electronic devices, energy converter, even electric car.

Physical stage of photosynthesis charge separation

Science.gov (United States)

Yakovlev, A. G.; Shuvalov, V. A.

2016-06-01

An analytical review is given concerning the biophysical aspects of light-driven primary charge separation in photosynthesis reaction centers (RCs) which are special pigment-protein complexes residing in a cell membrane. The primary (physical) stage of charge separation occurs in the pico- and femtosecond ranges and consists of transferring an electron along the active A-branch of pigments. The review presents vast factual material on both the general issues of primary photosynthesis and some more specific topics, including (1) the role of the inactive B-branch of pigments, (2) the effect of the protein environment on the charge separation, and (3) the participation of monomeric bacteriochlorophyll BA in primary electron acceptance. It is shown that the electron transfer and stabilization are strongly influenced by crystallographic water and tyrosine M210 molecules from the nearest environment of BA. A linkage between collective nuclear motions and electron transfer upon charge separation is demonstrated. The nature of the high quantum efficiency of primary charge separation reactions is discussed.

All solution processed organic thin film transistor-backplane with printing technology for electrophoretic display

Science.gov (United States)

Lee, Myung W.; Song, C.K.

2012-01-01

In this study, solution processes were developed for backplane using an organic thin film transistor (OTFT) as a driving device for an electrophoretic display (EPD) panel. The processes covered not only the key device of OTFTs but also interlayer and pixel electrodes. The various materials and printing processes were adopted to achieve the requirements of devices and functioning layers. The performance of OTFT of the backplane was sufficient to drive EPD sheet by producing a mobility of 0.12 cm2/v x sec and on/off current ratio of 10(5).

Inducible immune proteins in the dampwood termite Zootermopsis angusticollis

Science.gov (United States)

Rosengaus, Rebeca B.; Cornelisse, Tara; Guschanski, Katerina; Traniello, James F. A.

2007-01-01

Dampwood termites, Zootermopsis angusticollis (Isoptera: Termopsidae), mount an immune response to resist microbial infection. Here we report on results of a novel analysis that allowed us to electrophoretically assess changes in hemolymph proteins in the same individual before and after exposure to a pathogen. We demonstrate that contact with a sublethal concentration of the entomopathogenic fungus Metarhizium anisopliae (Deuteromycotina:Hypomycetes) induces the production of protective proteins in nymphs, pseudergates (false workers), and soldiers. Termites exposed to an immunizing dosage of fungal conidia consistently showed an enhancement of constitutive proteins (62-85 kDa) in the hemolymph as well as an induction of novel proteins (28-48 kDa) relative to preimmunization levels. No significant differences in protein banding patterns relative to baseline levels in control and naïve termites were observed. Incubating excised and eluted induced proteins produced by immunized pseudergates or immunized soldiers with conidia significantly reduced the germination of the fungus. The fungistatic effect of eluted proteins differed significantly among five colonies examined. Our results show that the upregulation of protective proteins in the hemolymph underscores the in vivo immune response we previously recorded in Z. angusticollis.

Binding of triiodothyronine to rat liver nuclear matrix. influence of thyroid hormones on the phosphorylation of nuclear matrix proteins

International Nuclear Information System (INIS)

Adylova, A.T.; Atakhanova, B.A.

1986-01-01

The interaction of thyroid hormones with rat liver nuclear matrix proteins was investigated. It was shown that the nuclear matrix contains sites that bind triiodothyronine with high affinity (K = 1.07.10 9 M -1 ) and limited capacity (the maximum binding capacity is equal to 28 /SUP a/ .5 fmoles of triiodothyronine per 100 ug protein). Electrophoretic identification of the matrix proteins that bind triiodothyronine was performed. The molecular weight of the main triiodothyronine-binding fraction is 50,000-52,000. It was shown that the administration of triiodothyronine to thyroidectomized rats stimulates the phosphorylation of all the protein fractions of the nuclear matrix

Chromatographic Separations of Enantiomers and Underivatized Oligosaccharides

Energy Technology Data Exchange (ETDEWEB)

Liu, Ying [Iowa State Univ., Ames, IA (United States)

2004-01-01

My graduate research has focused on separation science and bioanalytical analysis, which emphasized in method development. It includes three major areas: enantiomeric separations using high performance liquid chromatography (HPLC), Super/subcritical fluid chromatography (SFC), and capillary electrophoresis (CE); drug-protein binding behavior studies using CE; and carbohydrate analysis using liquid chromatograph-electrospray ionization mass spectrometry (LC-ESI-MS). Enantiomeric separations continue to be extremely important in the pharmaceutical industry. An in-depth evaluation of the enantiomeric separation capabilities of macrocyclic glycopeptides CSPs with SFC mobile phases was investigated using a set of over 100 chiral compounds. It was found that the macrocyclic based CSPs were able to separate enantiomers of various compounds with different polarities and functionalities. Seventy percent of all separations were achieved in less than 4 min due to the high flow rate (4.0 ml/min) that can be used in SFC. Drug-protein binding is an important process in determining the activity and fate of a drug once it enters the body. Two drug/protein systems have been studied using frontal analysis CE method. More sensitive fluorescence detection was introduced in this assay, which overcame the problem of low sensitivity that is common when using UV detection for drug-protein studies. In addition, the first usage of an argon ion laser with 257 nm beam coupled with CCD camera as a frontal analysis detection method enabled the simultaneous observation of drug fluorescence as well as the protein fluorescence. LC-ESI-MS was used for the separation and characterization of underivatized oligosaccharide mixtures. With the limits of detection as low as 50 picograms, all individual components of oligosaccharide mixtures (up to 11 glucose-units long) were baseline resolved on a Cyclobond I 2000 column and detected using ESI-MS. This system is characterized by high chromatographic

Chromatographic Separations of Enantiomers and Underivatized Oligosaccharides

International Nuclear Information System (INIS)

Ying Liu

2004-01-01

My graduate research has focused on separation science and bioanalytical analysis, which emphasized in method development. It includes three major areas: enantiomeric separations using high performance liquid chromatography (HPLC), Super/subcritical fluid chromatography (SFC), and capillary electrophoresis (CE); drug-protein binding behavior studies using CE; and carbohydrate analysis using liquid chromatograph-electrospray ionization mass spectrometry (LC-ESI-MS). Enantiomeric separations continue to be extremely important in the pharmaceutical industry. An in-depth evaluation of the enantiomeric separation capabilities of macrocyclic glycopeptides CSPs with SFC mobile phases was investigated using a set of over 100 chiral compounds. It was found that the macrocyclic based CSPs were able to separate enantiomers of various compounds with different polarities and functionalities. Seventy percent of all separations were achieved in less than 4 min due to the high flow rate (4.0 ml/min) that can be used in SFC. Drug-protein binding is an important process in determining the activity and fate of a drug once it enters the body. Two drug/protein systems have been studied using frontal analysis CE method. More sensitive fluorescence detection was introduced in this assay, which overcame the problem of low sensitivity that is common when using UV detection for drug-protein studies. In addition, the first usage of an argon ion laser with 257 nm beam coupled with CCD camera as a frontal analysis detection method enabled the simultaneous observation of drug fluorescence as well as the protein fluorescence. LC-ESI-MS was used for the separation and characterization of underivatized oligosaccharide mixtures. With the limits of detection as low as 50 picograms, all individual components of oligosaccharide mixtures (up to 11 glucose-units long) were baseline resolved on a Cyclobond I 2000 column and detected using ESI-MS. This system is characterized by high chromatographic

Multistage Magnetic Separator of Cells and Proteins

Science.gov (United States)

Barton, Ken; Ainsworth, Mark; Daily, Bruce; Dunn, Scott; Metz, Bill; Vellinger, John; Taylor, Brock; Meador, Bruce

2005-01-01

The multistage electromagnetic separator for purifying cells and magnetic particles (MAGSEP) is a laboratory apparatus for separating and/or purifying particles (especially biological cells) on the basis of their magnetic susceptibility and magnetophoretic mobility. Whereas a typical prior apparatus based on similar principles offers only a single stage of separation, the MAGSEP, as its full name indicates, offers multiple stages of separation; this makes it possible to refine a sample population of particles to a higher level of purity or to categorize multiple portions of the sample on the basis of magnetic susceptibility and/or magnetophoretic mobility. The MAGSEP includes a processing unit and an electronic unit coupled to a personal computer. The processing unit includes upper and lower plates, a plate-rotation system, an electromagnet, an electromagnet-translation system, and a capture-magnet assembly. The plates are bolted together through a roller bearing that allows the plates to rotate with respect to each other. An interface between the plates acts as a seal for separating fluids. A lower cuvette can be aligned with as many as 15 upper cuvette stations for fraction collection during processing. A two-phase stepping motor drives the rotation system, causing the upper plate to rotate for the collection of each fraction of the sample material. The electromagnet generates a magnetic field across the lower cuvette, while the translation system translates the electromagnet upward along the lower cuvette. The current supplied to the electromagnet, and thus the magnetic flux density at the pole face of the electromagnet, can be set at a programmed value between 0 and 1,400 gauss (0.14 T). The rate of translation can be programmed between 5 and 2,000 m/s so as to align all sample particles in the same position in the cuvette. The capture magnet can be a permanent magnet. It is mounted on an arm connected to a stepping motor. The stepping motor rotates the arm to

Adenosine deaminase complexing protein (ADCP): a transformation sensitive protein with potentials of a cancer marker.

Science.gov (United States)

Herbschleb-Voogt, E; Ten Kate, J; Meera Khan, P

1983-01-01

Several observations by independent investigators in the past have indicated that adenosine deaminase complexing protein (ADCP), present in considerable quantities in certain human tissues, was absent or decreased in the cancers originated from them. During the present study, electrophoretic analysis of adenosine deaminase (ADA) isozymes and radioimmunoassay for ADCP in the primary fibroblasts and the transformed as well as certain tumor derived cell lines have demonstrated that ADCP present in large quantities in the primary cells was absent or nearly absent in the transformed or tumor-derived cell lines. Though the mechanisms involved are not yet clear, the above observations indicate that ADCP has the potentials of a useful marker in the studies on transformed cells and cancer tissues.

Advanced purification strategy for CueR, a cysteine containing copper(I) and DNA binding protein

DEFF Research Database (Denmark)

Balogh, Ria K.; Gyurcsik, Béla; Hunyadi-Gulyás, Éva

2016-01-01

. A detailed understanding of their function may be exploited in potential health, environmental and analytical applications. Members of the MerR protein family sense a broad range of mostly late transition and heavy metal ions through their cysteine thiolates. The air sensitivity of latter groups makes...... the expression and purification of such proteins challenging. Here we describe a method for the purification of the copper-regulatory CueR protein under optimized conditions. In order to avoid protein precipitation and/or eventual aggregation and to get rid of the co-purifying Escherichia coli elongation factor...... any affinity tag. Structure and functionality tests performed with mass spectrometry, circular dichroism spectroscopy and electrophoretic gel mobility shift assays approved the success of the purification procedure....

Stability of some Cactaceae proteins based on fluorescence, circular dichroism, and differential scanning calorimetry measurements.

Science.gov (United States)

Gorinstein, S; Zemser, M; Vargas-Albores, F; Ochoa, J L; Paredes-Lopez, O; Scheler, C; Aksu, S; Salnikow, J

1999-02-01

Characterization of three cactus proteins (native and denatured) from Machaerocereus gummosus (Pitahaya agria), Lophocereu schottii (Garambullo), and Cholla opuntia (Cholla), was based on electrophoretic, fluorescence, CD (circular dichroism), DSC (differential scanning calorimetry), and FT-IR (Fourier transform infrared) measurements. The obtained results of intrinsic fluorescence, DSC, and CD were dissimilar for the three species of cactus, providing evidence of differences in secondary and tertiary structures. Cactus proteins may be situated in the following order corresponding to their relative stability: Machaerocereus gummosus (Pitahaya agria) > Cholla opuntia (Cholla) > Lophocereu schottii (Garambullo). Thermodynamic properties of proteins and their changes upon denaturation (temperature of denaturation, enthalphy, and the number of ruptured hydrogen bonds) were correlated with the secondary structure of proteins and disappearance of alpha-helix.

The effect of proteins on the aging properties of radiation vulcanized natural rubber latex

International Nuclear Information System (INIS)

Abad, L.V.

1993-01-01

The effect of natural rubber latex (NRL) proteins on the aging properties of NRL films was investigated. SDS-PAGE electrophoresis of the rubber proteins in NRL (Sri-Lanka) indicated a total of 18 proteins. A sharp decrease in tensile strength was observed after aging when NRL films were leached in 1% NH 4 OH. However, when these films were soaked in ethanol prior to leaching, the aging properties approximated those of the unleashed samples. Electrophoretic analysis of the proteins present in the NH 3 extracts of leached RVNRL films showed a high concentration of the protein herein. This protein was not found in the NH 3 extracts of ethanol soaked films. NRL proteins were shown to decelerate the aging process of Radiation Vulcanized Natural Rubber Latex (RVNRL) films. Among the proteins, herein exhibited good anti-aging properties. The hydrolyzates from NR proteins also enhanced considerably the aging properties of RVNRL. (auth.). 8 refs.; 40 figs.; 30 tabs

Use of electrophoretic analysis of grain proteins in the identification of wheat radiomutant ST 7750

International Nuclear Information System (INIS)

Sasek, A.; Kubanek, J.; Hanis, M.; Cerny, J.

1977-01-01

For a more precise comparison of the protein complexes of the grain of the 'Jubilar' cv. and of its radiomutant ST 7750, the methods of the electrophoresis in starch gel in various buffer systems and of disc electrophoresis in polyacrylamide gel were used for the study of anodal isoperoxidases. A determination was made of the total protein content, of the basic protein fractions, and of the quantitative proportion of amino acids. Basic qualitative correspondence of the albumin-globulin and gliadin spectrum of the examined variants was found and only in some zones of both spectra certain differences were observed in the quantitative content which may of course, indicate genotypic differentiation of the mutant SF 7750 and of the starting 'Jubilar' cv. Analogous findings were confirmed also by the results of isoperoxidases. Complementary analyses of the protein fractions confirmed the correspondence of the content of albumins, globulins, and gliadins of the investigated variants. Differences were found in the content of glutelin fractions and of insoluble residue. The ascertained changes were confirmed by amino acid analysis where higher methionine, valine, leucine, and isoleucine levels were found in the ST 7750 mutant. The results obtained have shown that especially the analysis of gliadin spectra by electrophoresis in starch gel is a suitable method of differentiating and unifying wheat genotypes as it reveals the differences between the initial and the mutated genotype. (author)

Selective binding and magnetic separation of His-tagged proteins using Fe3O4/PAM/NTA-Ni2+ Magnetic Nanoparticles

Science.gov (United States)

Guo, Huiling; Li, Mengyun; Tu, Shu; Sun, Honghao

2018-03-01

Fe3O4 nanoparticles coated with polyacrylamide (PAM) were synthesized. The magnetic core, with an average hydrodynamic size of 235.5 nm, allowed the magnetic nanoparticles (MNPs) rapid separation from solutions under an external magnetic field. NTA-Ni2+ was modified on the surface of Fe3O4/PAM MNPs to selectively trap his-tagged green fluorescent protein (GFP). The results showed that Fe3O4/PAM/NTA-Ni2+ MNPs exhibited remarkable capability of selective binding and separating his-tagged GFP. The adsorption efficiency was 93.37%.

Harvey murine sarcoma virus p21 ras protein: biological and biochemical significance of the cysteine nearest the carboxy terminus

DEFF Research Database (Denmark)

Willumsen, B M; Norris, K; Papageorge, A G

1984-01-01

localization. We have now further characterized the post-translational processing of these mutants and have also studied two C-terminal v-rasH point mutants: one encodes serine in place of cysteine-186, the other threonine for valine-187. The Thr-187 mutant was transformation-competent, and its p21 protein...... not undergo the posttranslational processing common to biologically active ras proteins: their electrophoretic migration rate did not change, they remained in the cytosol, and they failed to bind lipid. Since the cell-encoded ras proteins also contain this cysteine, we conclude that this amino acid residue...

A separable domain of the p150 subunit of human chromatin assembly factor-1 promotes protein and chromosome associations with nucleoli.

Science.gov (United States)

Smith, Corey L; Matheson, Timothy D; Trombly, Daniel J; Sun, Xiaoming; Campeau, Eric; Han, Xuemei; Yates, John R; Kaufman, Paul D

2014-09-15

Chromatin assembly factor-1 (CAF-1) is a three-subunit protein complex conserved throughout eukaryotes that deposits histones during DNA synthesis. Here we present a novel role for the human p150 subunit in regulating nucleolar macromolecular interactions. Acute depletion of p150 causes redistribution of multiple nucleolar proteins and reduces nucleolar association with several repetitive element-containing loci. Of note, a point mutation in a SUMO-interacting motif (SIM) within p150 abolishes nucleolar associations, whereas PCNA or HP1 interaction sites within p150 are not required for these interactions. In addition, acute depletion of SUMO-2 or the SUMO E2 ligase Ubc9 reduces α-satellite DNA association with nucleoli. The nucleolar functions of p150 are separable from its interactions with the other subunits of the CAF-1 complex because an N-terminal fragment of p150 (p150N) that cannot interact with other CAF-1 subunits is sufficient for maintaining nucleolar chromosome and protein associations. Therefore these data define novel functions for a separable domain of the p150 protein, regulating protein and DNA interactions at the nucleolus. © 2014 Smith et al. This article is distributed by The American Society for Cell Biology under license from the author(s). Two months after publication it is available to the public under an Attribution–Noncommercial–Share Alike 3.0 Unported Creative Commons License (http://creativecommons.org/licenses/by-nc-sa/3.0).

Twist on protein microarrays: layering wax-patterned nitrocellulose to create customizable and separable arrays of multiplexed affinity columns.

Science.gov (United States)

de Lange, Victoria; Vörös, János

2014-05-06

We developed the simple and inexpensive FoRe microarray to simultaneously test several 1 μL samples for multiple proteins. By combining forward and reverse phase microarrays into an innovative three-dimensional format, the FoRe array exploits the advantages and eliminates several drawbacks of the traditional approaches (i.e., large sample volumes, protein loss, and cross-reactivity between detection antibodies). Samples are pipetted into an array of separable, multiplexed affinity columns. Several nitrocellulose membranes, each functionalized with a different capture antibody, are stacked to create a customizable affinity column. The nitrocellulose is patterned with wax to form 25 isolated microspots on each layer, allowing us to analyze multiple samples in parallel. After running the immunoassay, the stacks are quickly disassembled, revealing 2D microarrays of different fractions from multiple samples. By combining the stack-and-separate technique with wax patterning, we keep the arrays low cost and easily tailored to a variety of applications. We successfully performed 3D multiplexing using a model system with mouse and rabbit IgG. Binding proved to be independent of the position in the stack, and the limit of detection for a mouse IgG sandwich assay was 7.3 pM in BSA and 15 pM in human plasma. The FoRe microarray makes it possible to identify protein expression patterns across several minute volume samples; for example, it could be used to analyze cell lysate in drug response studies or pricks of blood from small animal studies.

In-capillary enrichment, proteolysis and separation using capillary electrophoresis with discontinuous buffers: application on proteins with moderately acidic and basic isoelectric points.

Science.gov (United States)

Nesbitt, Chandra A; Yeung, Ken K-C

2009-01-01

Advances in mass spectrometry and capillary-format separation continue to improve the sensitivity of protein analysis. Of equal importance is the miniaturization of sample pretreatment such as enrichment and proteolysis. In a previous report (Nesbitt et al., Electrophoresis, 2008, 29, 466-474), nanoliter-volume protein enrichment, tryptic digestion, and partial separation was demonstrated in capillary electrophoresis followed by MALDI mass spectral analysis. A discontinuous buffer system, consisting of ammonium (pH 10) and acetate (pH 4), was used to create a pH junction inside the capillary, trapping a protein with a neutral isoelectric point, myoglobin (pI 7.2). Moreover, co-enrichment of myoglobin with trypsin led to an in-capillary digestion. In this paper, the ability of this discontinuous buffer system to perform similar in-capillary sample pretreatment on proteins with moderately acidic and basic pI was studied and reported. Lentil lectin (pI 8.6) and a multi-phosphorylated protein, beta-casein (pI 5.1), were selected as model proteins. In addition to the previously shown tryptic digestion, proteolysis with endoproteinase Asp-N was also performed. Digestion of these acidic and basic pI proteins produced a few peptides with extreme pI values lying outside the trapping range of the discontinuous buffer. An alteration in the peptide trapping procedure was made to accommodate these analytes. Offline MALDI mass spectral analysis confirmed the presence of the expected peptides. The presented miniaturized sample pretreatment methodology was proven to be applicable on proteins with a moderately wide range of pI. Flexibility in the choice of protease was also evident.

The monomeric form of Neisseria DNA mimic protein DMP19 prevents DNA from binding to the histone-like HU protein

Science.gov (United States)

Ko, Tzu-Ping; Liao, Yi-Ting; Hsu, Kai-Cheng

2017-01-01

DNA mimicry is a direct and effective strategy by which the mimic competes with DNA for the DNA binding sites on other proteins. Until now, only about a dozen proteins have been shown to function via this strategy, including the DNA mimic protein DMP19 from Neisseria meningitides. We have shown previously that DMP19 dimer prevents the operator DNA from binding to the transcription factor NHTF. Here, we provide new evidence that DMP19 monomer can also interact with the Neisseria nucleoid-associated protein HU. Using BS3 crosslinking, gel filtration and isothermal titration calorimetry assays, we found that DMP19 uses its monomeric form to interact with the Neisseria HU dimer. Crosslinking conjugated mass spectrometry was used to investigate the binding mode of DMP19 monomer and HU dimer. Finally, an electrophoretic mobility shift assay (EMSA) confirmed that the DNA binding affinity of HU is affected by DMP19. These results showed that DMP19 is bifunctional in the gene regulation of Neisseria through its variable oligomeric forms. PMID:29220372

A flexible angle sensor made from MWNT/CuO/Cu{sub 2}O nanocomposite films deposited by an electrophoretic co-deposition process

Energy Technology Data Exchange (ETDEWEB)

Toboonsung, Buppachat, E-mail: buppachattt@yahoo.co.th [Physics and General Science Program, Faculty of Science and Technology, Nakhon Ratchasima Rajabhat University, Nakhon Ratchasima 30000 (Thailand); Singjai, Pisith, E-mail: singjai@hotmail.com [Department of Physics and Materials Science, Faculty of Science, Chiang Mai University, Chiang Mai 50200 (Thailand); Materials Science Research Center, Faculty of Science, Chiang Mai University, Chiang Mai, 50200 (Thailand)

2012-08-25

Highlights: Black-Right-Pointing-Pointer MWNT/CuO/Cu{sub 2}Onanocomposite films were coated on a PET sheet. Black-Right-Pointing-Pointer The film resistance and application as angle sensor were investigated. Black-Right-Pointing-Pointer Thesensor showed a linear relation between the film resistance and the bending angle. Black-Right-Pointing-Pointer A minimum loop area and a high stability in sensitivity over a thousand bending cycles were obtained. - Abstract: A flexible angle sensor was prepared using an electrophoretic co-deposition process to form nanocomposite networks of multi-wall carbon nanotube/cupric oxide/cuprous oxide (MWNT/CuO/Cu{sub 2}O) on a polyethylene terephthalate (PET) sheet. The deposition method used copper and stainless steel electrodes, and the effects of varying of electrode separation, MWNT concentration in deionized water, voltage and deposition time were studied. The film resistance of the as-deposited samples decreased with increasing the MWNT concentration up to 0.3 mg/ml. The angle sensor showed a linear relation between the film resistance and the bending angle, a relationship that was illustrated with loop area and sensitivity data. The best angle sensor was successfully made with an electrode separation of 8 mm, a concentration of 0.3 mg/ml, a voltage of 10 V and a deposition time of 3 h, parameters that resulted in a minimum loop area and the most stability in sensitivity over a thousand bending cycles.

Western blotting: an introduction.

Science.gov (United States)

Kurien, Biji T; Scofield, R Hal

2015-01-01

Western blotting is an important procedure for the immunodetection of proteins, particularly proteins that are of low abundance. This process involves the transfer of protein patterns from gel to microporous membrane. Electrophoretic as well as non-electrophoretic transfer of proteins to membranes was first described in 1979. Protein blotting has evolved greatly since the inception of this protocol, allowing protein transfer to be accomplished in a variety of ways.

Neonatal maternal separation up-regulates protein signalling for cell survival in rat hypothalamus.

Science.gov (United States)

Irles, Claudine; Nava-Kopp, Alicia T; Morán, Julio; Zhang, Limei

2014-05-01

We have previously reported that in response to early life stress, such as maternal hyperthyroidism and maternal separation (MS), the rat hypothalamic vasopressinergic system becomes up-regulated, showing enlarged nuclear volume and cell number, with stress hyperresponsivity and high anxiety during adulthood. The detailed signaling pathways involving cell death/survival, modified by adverse experiences in this developmental window remains unknown. Here, we report the effects of MS on cellular density and time-dependent fluctuations of the expression of pro- and anti-apoptotic factors during the development of the hypothalamus. Neonatal male rats were exposed to 3 h-daily MS from postnatal days 2 to 15 (PND 2-15). Cellular density was assessed in the hypothalamus at PND 21 using methylene blue staining, and neuronal nuclear specific protein and glial fibrillary acidic protein immunostaining at PND 36. Expression of factors related to apoptosis and cell survival in the hypothalamus was examined at PND 1, 3, 6, 9, 12, 15, 20 and 43 by Western blot. Rats subjected to MS exhibited greater cell-density and increased neuronal density in all hypothalamic regions assessed. The time course of protein expression in the postnatal brain showed: (1) decreased expression of active caspase 3; (2) increased Bcl-2/Bax ratio; (3) increased activation of ERK1/2, Akt and inactivation of Bad; PND 15 and PND 20 were the most prominent time-points. These data indicate that MS can induce hypothalamic structural reorganization by promoting survival, suppressing cell death pathways, increasing cellular density which may alter the contribution of these modified regions to homeostasis.

Determination and Quantification of Molecular Interactions in Protein Films: A Review

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Felicia Hammann

2014-12-01

Full Text Available Protein based films are nowadays also prepared with the aim of replacing expensive, crude oil-based polymers as environmentally friendly and renewable alternatives. The protein structure determines the ability of protein chains to form intra- and intermolecular bonds, whereas the degree of cross-linking depends on the amino acid composition and molecular weight of the protein, besides the conditions used in film preparation and processing. The functionality varies significantly depending on the type of protein and affects the resulting film quality and properties. This paper reviews the methods used in examination of molecular interactions in protein films and discusses how these intermolecular interactions can be quantified. The qualitative determination methods can be distinguished by structural analysis of solutions (electrophoretic analysis, size exclusion chromatography and analysis of solid films (spectroscopy techniques, X-ray scattering methods. To quantify molecular interactions involved, two methods were found to be the most suitable: protein film swelling and solubility. The importance of non-covalent and covalent interactions in protein films can be investigated using different solvents. The research was focused on whey protein, whereas soy protein and wheat gluten were included as further examples of proteins.

Determination and Quantification of Molecular Interactions in Protein Films: A Review.

Science.gov (United States)

Hammann, Felicia; Schmid, Markus

2014-12-10

Protein based films are nowadays also prepared with the aim of replacing expensive, crude oil-based polymers as environmentally friendly and renewable alternatives. The protein structure determines the ability of protein chains to form intra- and intermolecular bonds, whereas the degree of cross-linking depends on the amino acid composition and molecular weight of the protein, besides the conditions used in film preparation and processing. The functionality varies significantly depending on the type of protein and affects the resulting film quality and properties. This paper reviews the methods used in examination of molecular interactions in protein films and discusses how these intermolecular interactions can be quantified. The qualitative determination methods can be distinguished by structural analysis of solutions (electrophoretic analysis, size exclusion chromatography) and analysis of solid films (spectroscopy techniques, X-ray scattering methods). To quantify molecular interactions involved, two methods were found to be the most suitable: protein film swelling and solubility. The importance of non-covalent and covalent interactions in protein films can be investigated using different solvents. The research was focused on whey protein, whereas soy protein and wheat gluten were included as further examples of proteins.

Insight into Nanoparticle Charging Mechanism in Nonpolar Solvents To Control the Formation of Pt Nanoparticle Monolayers by Electrophoretic Deposition

Czech Academy of Sciences Publication Activity Database

Černohorský, Ondřej; Grym, Jan; Yatskiv, Roman; Pham, V. H.; Dickerson, J.H.

2016-01-01

Roč. 8, č. 30 (2016), s. 19680-19690 ISSN 1944-8244 R&D Projects: GA ČR GA15-17044S; GA MŠk(CZ) LD14111 Institutional support: RVO:67985882 Keywords : AOT reverse micelles * 3D growth * electrophoretic deposition Subject RIV: JA - Electronics ; Optoelectronics, Electrical Engineering Impact factor: 7.504, year: 2016

PARTITION EFFICIENCY OF NEWLY DESIGNED LOCULAR MULTILAYER COIL FOR COUNTERCURRENT CHROMATOGRAPHIC SEPARATION OF PROTEINS USING SMALL-SCALE CROSS-AXIS COIL PLANET CENTRIFUGE WITH AQUEOUS-AQUEOUS POLYMER PHASE SYSTEMS.

Science.gov (United States)

Shinomiya, Kazufusa; Ito, Yoichiro

2009-01-01

Countercurrent chromatographic performance of the locular multilayer coil separation column newly designed in our laboratory was evaluated in terms of theoretical plate number, peak resolution and retention of the stationary phase in protein separation with an aqueous polymer phase system using the small-scale cross-axis coil planet centrifuge (X-axis CPC) fabricated in our laboratory. The locular column was made from 1.0 mm I.D., 2.0 mm O.D. or 1.5 mm I.D., 2.5 mm O.D. PTFE tubing compressed with a pair of hemostat at 2 or 4 cm intervals. The protein separation was performed using a set of stable proteins including cytochrome C, myoglobin and lysozyme with the 12.5% (w/w) polyethylene glycol 1000 and 12.5% (w/w) dibasic potassium phosphate system under 1000 rpm of column revolution. The 1.5 mm I.D., 2.5 mm O.D. locular tubing compressed at 2 cm intervals yielded better partition efficiencies than the non-clamped tubing using both lower and upper mobile phases with satisfactory retention of the stationary phase. The overall results suggest that the newly designed locular multilayer coil is useful to the preparative separation of proteins with aqueous-aqueous polymer phase system using our small-scale X-axis CPC.

Extraction and separation of water soluble proteins from Bacillus thuringiensis-transgenic and non-transgenic maize species by CZE

Czech Academy of Sciences Publication Activity Database

Sázelová, Petra; Kašička, Václav; Ibanez, E.; Cifuentes, A.

2009-01-01

Roč. 32, č. 21 (2009), s. 3801-3808 ISSN 1615-9306 R&D Projects: GA ČR(CZ) GA203/08/1428 Grant - others:GA ČR(CZ) GA203/09/0675 Program:GA Institutional research plan: CEZ:AV0Z40550506 Keywords : Bacillus thuringiensis -transgenic maize * CZE-UV profiling * Maize proteins Subject RIV: CB - Analytical Chemistry, Separation Impact factor: 2.551, year: 2009

Ultrafiltration of pegylated proteins

Science.gov (United States)

Molek, Jessica R.

There is considerable clinical interest in the use of "second-generation" therapeutics produced by conjugation of a native protein with various polymers including polyethylene glycol (PEG). PEG--protein conjugates, so-called PEGylated proteins, can exhibit enhanced stability, half-life, and bioavailability. One of the challenges in the commercial production of PEGylated proteins is the purification required to remove unreacted polymer, native protein, and in many cases PEGylated proteins with nonoptimal degrees of conjugation. The overall objective of this thesis was to examine the use of ultrafiltration for the purification of PEGylated proteins. This included: (1) analysis of size-based separation of PEGylated proteins using conventional ultrafiltration membranes, (2) use of electrically-charged membranes to exploit differences in electrostatic interactions, and (3) examination of the effects of PEGylation on protein fouling. The experimental results were analyzed using appropriate theoretical models, with the underlying physical properties of the PEGylated proteins evaluated using size exclusion chromatography, capillary electrophoresis, dynamic light scattering, and reverse phase chromatography. PEGylated proteins were produced by covalent attachment of activated PEG to a protein via primary amines on the lysine residues. A simple model was developed for the reaction kinetics, which was used to explore the effect of reaction conditions and mode of operation on the distribution of PEGylated products. The effective size of the PEGylated proteins was evaluated using size exclusion chromatography, with appropriate correlations developed for the size in terms of the molecular weight of the native protein and attached PEG. The electrophoretic mobility of the PEGylated proteins were evaluated by capillary electrophoresis with the data in good agreement with a simple model accounting for the increase in protein size and the reduction in the number of protonated amine

Effect of x-irradiation in rats bearing walker-256-carcinosarcoma and normal rats

International Nuclear Information System (INIS)

Ehara, Kazuhiko

1978-01-01

Serum protein fractions and total proteins were studied with bloods obtained from the rats exposed each to the partial-, whole-bodies and the transplanted tumors (Walker-256-carcinosarcoma transplanted in the right hind leg). The electrophoretic variation induced in the sera of tumor-bearing rats (Group II), and the content of total proteins decreased. Early irradiation to the tumor part of rats less induced the variations of the electrophoretic pattern and the decrease of the amount of serum total proteins. When the distant metastasis appeared during irradiation treatment, the electrophoretic patterns and content of total proteins changed proportionally to the variation in sera of Group II. On the other hand, the γ-globulin (G) fraction increased in the long-term survival rat. The separation of the rat serum β-G into two peaks of β 1 - and β 2 -G was shown only in Group IV (late irradiation to the right hind leg). This finding supposed that some factors involve in the sera of rats with transplanted primary tumor grown up to a fixed size and guessed the appearance of the distant metastasis during x-irradiation. The percentages of the albumin and γ-G decreased slightly and those of the α 1 -, α 2 - and β-G increased slightly in the rats with 300 rad partial-body (the right hind leg) x-irradiation daily for 20 days. The remarkable decrease of the albumin and γ-G, the increase of the α 1 - and β-G, the marked increase of the α 2 -G and the decrease of serum total proteins were demonstrated for the sera of rats with 1,000 rad whole-body x-irradiation at a time. These phenomena seem to be related to the destructive and reticuloendothelial injury by the exposure. (auth.)

A new affinity-HPLC packing for protein separation: Cibacron blue attached uniform porous poly(HEMA-co-EDM) beads.

Science.gov (United States)

Unsal, Ender; Durdu, Aysun; Elmas, Begum; Tuncel, Murvet; Tuncel, Ali

2005-11-01

In this study, a new affinity high-performance liquid chromatography (HPLC) stationary phase suitable for protein separation was synthesized. In the first stage of the synthesis, uniform porous poly(2-hydroxyethyl methacrylate-co-ethylene dimethacrylate), poly(HEMA-co-EDM), beads 6.2 mum in size were obtained. Homogeneous distribution of hydroxyl groups in the bead interior was confirmed by confocal laser scanning microscopy. The plain poly(HEMA-co-EDM) particles gave very low non-specific protein adsorption with albumin. The selected dye ligand Cibacron blue F3G-A (CB F3G-A) was covalently linked onto the beads via hydroxyl groups. In the batch experiments, albumin adsorption up to 60 mg BSA/g particles was obtained with the CB F3G-A carrying poly(HEMA-co-EDM) beads. The affinity-HPLC of selected proteins (albumin and lysozyme) was investigated in a 25 mm x 4.0-mm inner diameter column packed with CB F3G-A carrying beads and both proteins were successfully resolved. By a single injection, 200 mug of protein was loaded and quantitatively eluted from the column. The protein recovery increased with increasing flow rate and salt concentration of the elution buffer and decreased with the increasing protein feed concentration. During the albumin elution, theoretical plate numbers up to 30,000 plates/m were achieved by increasing the salt concentration.

Cyanobacterial flv4-2 Operon-Encoded Proteins Optimize Light Harvesting and Charge Separation in Photosystem II.

Science.gov (United States)

Chukhutsina, Volha; Bersanini, Luca; Aro, Eva-Mari; van Amerongen, Herbert

2015-05-01

Photosystem II (PSII) complexes drive the water-splitting reaction necessary to transform sunlight into chemical energy. However, too much light can damage and disrupt PSII. In cyanobacteria, the flv4-2 operon encodes three proteins (Flv2, Flv4, and Sll0218), which safeguard PSII activity under air-level CO2 and in high light conditions. However, the exact mechanism of action of these proteins has not been clarified yet. We demonstrate that the PSII electron transfer properties are influenced by the flv4-2 operon-encoded proteins. Accelerated secondary charge separation kinetics was observed upon expression/overexpression of the flv4-2 operon. This is likely induced by docking of the Flv2/Flv4 heterodimer in the vicinity of the QB pocket of PSII, which, in turn, increases the QB redox potential and consequently stabilizes forward electron transfer. The alternative electron transfer route constituted by Flv2/Flv4 sequesters electrons from QB(-) guaranteeing the dissipation of excess excitation energy in PSII under stressful conditions. In addition, we demonstrate that in the absence of the flv4-2 operon-encoded proteins, about 20% of the phycobilisome antenna becomes detached from the reaction centers, thus decreasing light harvesting. Phycobilisome detachment is a consequence of a decreased relative content of PSII dimers, a feature observed in the absence of the Sll0218 protein. Copyright © 2015 The Author. Published by Elsevier Inc. All rights reserved.

A chip-type thin-layer electrochemical cell coupled with capillary electrophoresis for online separation of electrode reaction products

Energy Technology Data Exchange (ETDEWEB)

He, Jian-Bo, E-mail: jbhe@hfut.edu.cn; Cui, Ting; Zhang, Wen-Wen; Deng, Ning

2013-07-05

Graphical abstract: -- Highlights: •A new coupling of thin-layer electrolysis with capillary electrophoresis (CE). •Rapid electrolysis, direct sampling followed by online CE separation. •At least 13 products of quercetin oxidation were separated. •Thermodynamic and kinetic parameters were determined from CE peak areas. -- Abstract: A coupling technique of thin-layer electrolysis with high-performance capillary electrophoresis/UV–vis technique(EC/HPCE/UV–vis) is developed for online separation and determination of electrode reaction products. A chip-type thin-layer electrolytic (CTE) cell was designed and fabricated, which contains a capillary channel and a background electrolyte reservoir, allowing rapid electrolysis, direct sampling and online electrophoretic separation. This chip-type setup was characterized based on an electrophoresis expression of Nernst equation that was applied to the redox equilibrium of o-tolidine at different potentials. The utility of the method was demonstrated by separating and determining the electro-oxidation products of quercetin in different pH media. Two main products were always found in the studied time, potential and pH ranges. The variety of products increased not only with increasing potential but also with increasing pH value, and in total, at least 13 products were observed in the electropherograms. This work illustrates a novel example of capillary electrophoresis used online with thin-layer electrolysis to separate and detect electrode reaction products.

A chip-type thin-layer electrochemical cell coupled with capillary electrophoresis for online separation of electrode reaction products

International Nuclear Information System (INIS)

He, Jian-Bo; Cui, Ting; Zhang, Wen-Wen; Deng, Ning

2013-01-01

Graphical abstract: -- Highlights: •A new coupling of thin-layer electrolysis with capillary electrophoresis (CE). •Rapid electrolysis, direct sampling followed by online CE separation. •At least 13 products of quercetin oxidation were separated. •Thermodynamic and kinetic parameters were determined from CE peak areas. -- Abstract: A coupling technique of thin-layer electrolysis with high-performance capillary electrophoresis/UV–vis technique(EC/HPCE/UV–vis) is developed for online separation and determination of electrode reaction products. A chip-type thin-layer electrolytic (CTE) cell was designed and fabricated, which contains a capillary channel and a background electrolyte reservoir, allowing rapid electrolysis, direct sampling and online electrophoretic separation. This chip-type setup was characterized based on an electrophoresis expression of Nernst equation that was applied to the redox equilibrium of o-tolidine at different potentials. The utility of the method was demonstrated by separating and determining the electro-oxidation products of quercetin in different pH media. Two main products were always found in the studied time, potential and pH ranges. The variety of products increased not only with increasing potential but also with increasing pH value, and in total, at least 13 products were observed in the electropherograms. This work illustrates a novel example of capillary electrophoresis used online with thin-layer electrolysis to separate and detect electrode reaction products

Interaction of an IHF-like protein with the Rhizobium etli nifA promoter.

Science.gov (United States)

Benhassine, Traki; Fauvart, Maarten; Vanderleyden, Jos; Michiels, Jan

2007-06-01

The nifA gene fulfills an essential role in the regulation of nitrogen fixation genes in Rhizobium etli. Transcription analysis of the nifA gene, assessed using promoter deletions, indicated an oxygen-independent expression, threefold higher during symbiosis as compared with free-living conditions. Electrophoretic mobility shift assays using those nifA promoter deletion fragments, which were actively transcribed, demonstrated the specific interaction with R. etli cellular protein(s) resulting in the formation of two DNA-protein complexes. An interacting protein was purified by liquid chromatography on Heparin Sepharose and Mono S columns. The purified 12 kDa R. etli protein cross-reacted with antibodies directed against Escherichia coli integration host factor (IHF). Furthermore, purified E. coli IHF was able to specifically bind to the R. etli nifA promoter region. These results point to an as yet undisclosed function of IHF in the regulation of R. etli nifA expression.

Advanced analytical techniques

International Nuclear Information System (INIS)

Mrochek, J.E.; Shumate, S.E.; Genung, R.K.; Bahner, C.T.; Lee, N.E.; Dinsmore, S.R.

1976-01-01

The development of several new analytical techniques for use in clinical diagnosis and biomedical research is reported. These include: high-resolution liquid chromatographic systems for the early detection of pathological molecular constituents in physiologic body fluids; gradient elution chromatography for the analysis of protein-bound carbohydrates in blood serum samples, with emphasis on changes in sera from breast cancer patients; electrophoretic separation techniques coupled with staining of specific proteins in cellular isoenzymes for the monitoring of genetic mutations and abnormal molecular constituents in blood samples; and the development of a centrifugal elution chromatographic technique for the assay of specific proteins and immunoglobulins in human blood serum samples

Quantification of trace elements in protein bands by synchrotron radiation x-ray fluorescence after isoelectric focusing separation of human hemoglobin

International Nuclear Information System (INIS)

Gao Yuxi; Chen Chunying; Li Bai; He Wei; Huang Yuying; Chai Zhifang

2005-01-01

The role and effects of a trace element in a particular organism strongly depend on its particular chemical forms in which the element is present. Therefore, the bulk content or concentration of an element in the organism of interest is often meaningless in judging its biological significance. To understand bioavailability, transportation, cell uptake, metabolism, toxicity, and other biological behaviors of trace elements in the body, information is needed about speciation of trace element, especially about distribution of metal-containing proteins. Development of appropriate methods for speciation analysis is therefore required. Synchrotron radiation x-ray fluorescence (SRXRF) is a sensitive method for multielemental analysis with detection limit of 10 ng/g. It has been successfully used for imaging and quantifying trace elements in various pathological and healthy tissues, even in a single cell, to help understand the mechanism of diseases and the biochemistry of elements. In our previous work, the technique was combined with electrophoresis to study distribution of metalloproteins in biological samples, but the quantitative analysis of trace elements in protein bands after electrophoresis was still unrealized. In this study, a procedure has been proposed for quantification of Fe, Cu, and Zn in protein bands with SRXRF analysis after isoelectric focusing (IEF) separation. Calibration standards were prepared by adding certain amounts of metal ions and free-metal proteins to electrophoresis gel. Human hemoglobin was separated with IEF, and Fe, Cu, and Zn in protein bands were analyzed by SRXRF. The calibration curves can be obtained in a range of 0-8 mg/kg metals and a linear relationship between dosage of metals and fluorescent intensity can be observed (r 2 > 0.99). The method provides the detection limits of 2.43, 1.12, and 0.96 mg/kg for Fe, Cu and Zn, and the recoveries of 90.4 and 115.7 % for Fe and Zn, respectively. The hyphenated technique of SRXRF and IEF

Electrophoretic deposition of ZnO/alginate and ZnO-bioactive glass/alginate composite coatings for antimicrobial applications

Energy Technology Data Exchange (ETDEWEB)

Cordero-Arias, L.; Cabanas-Polo, S.; Goudouri, O.M. [Institute of Biomaterials, Department of Materials Science and Engineering, University of Erlangen-Nuremberg, Cauerstrasse 6, D-91058 Erlangen (Germany); Misra, S.K. [Materials Science and Engineering, Indian Institute of Technology Gandhinagar, Ahmedabad 382424 (India); Gilabert, J. [Institute of Ceramics Materials (ITC), University Jaume I, Avenida Vicent SosBaynat, 12006 Castellon (Spain); Valsami-Jones, E. [School of Geography, Earth and Environmental Sciences, University of Birmingham, Edgbaston, Birmingham B15 2TT (United Kingdom); Sanchez, E. [Institute of Ceramics Materials (ITC), University Jaume I, Avenida Vicent SosBaynat, 12006 Castellon (Spain); Virtanen, S. [Institute for Surface Science and Corrosion (LKO, WW4), Department of Materials Science and Engineering, University of Erlangen-Nuremberg, Martensstrasse 7, D-91058 Erlangen (Germany); Boccaccini, A.R., E-mail: aldo.boccaccini@ww.uni-erlangen.de [Institute of Biomaterials, Department of Materials Science and Engineering, University of Erlangen-Nuremberg, Cauerstrasse 6, D-91058 Erlangen (Germany)

2015-10-01

Two organic/inorganic composite coatings based on alginate, as organic matrix, and zinc oxide nanoparticles (n-ZnO) with and without bioactive glass (BG), as inorganic components, intended for biomedical applications, were developed by electrophoretic deposition (EPD). Different n-ZnO (1–10 g/L) and BG (1–1.5 g/L) contents were studied for a fixed alginate concentration (2 g/L). The presence of n-ZnO was confirmed to impart antibacterial properties to the coatings against gram-negative bacteria Escherichia coli, while the BG induced the formation of hydroxyapatite on coating surfaces thereby imparting bioactivity, making the coating suitable for bone replacement applications. Coating composition was analyzed by thermogravimetric analysis (TG), Fourier transform infrared spectroscopy (FTIR), X-ray diffraction (XRD) and energy dispersive X-ray spectroscopy (EDS) analyses. Scanning electron microscopy (SEM) was employed to study both the surface and the cross section morphology of the coatings. Polarization curves of the coated substrates made in cell culture media at 37 °C confirmed the corrosion protection function of the novel organic/inorganic composite coatings. - Highlights: • Organic–inorganic nanocomposite coatings fabricated by electrophoretic deposition • nZnO and bioactive glass containing alginate coatings exhibit antibacterial effect. • Bioactive character and anticorrosion function of coatings demonstrated.

Evaluation of DNA bending models in their capacity to predict electrophoretic migration anomalies of satellite DNA sequences.

Science.gov (United States)

Matyášek, Roman; Fulneček, Jaroslav; Kovařík, Aleš

2013-09-01

DNA containing a sequence that generates a local curvature exhibits a pronounced retardation in electrophoretic mobility. Various theoretical models have been proposed to explain relationship between DNA structural features and migration anomaly. Here, we studied the capacity of 15 static wedge-bending models to predict electrophoretic behavior of 69 satellite monomers derived from four divergent families. All monomers exhibited retarded mobility in PAGE corresponding to retardation factors ranging 1.02-1.54. The curvature varied both within and across the groups and correlated with the number, position, and lengths of A-tracts. Two dinucleotide models provided strong correlation between gel mobility and curvature prediction; two trinucleotide models were satisfactory while remaining dinucleotide models provided intermediate results with reliable prediction for subsets of sequences only. In some cases, similarly shaped molecules exhibited relatively large differences in mobility and vice versa. Generally less accurate predictions were obtained in groups containing less homogeneous sequences possessing distinct structural features. In conclusion, relatively universal theoretical models were identified suitable for the analysis of natural sequences known to harbor relatively moderate curvature. These models could be potentially applied to genome wide studies. However, in silico predictions should be viewed in context of experimental measurement of intrinsic DNA curvature. © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Characterization of CNT-MnO{sub 2} nanocomposite by electrophoretic deposition as potential electrode for supercapacitor

Energy Technology Data Exchange (ETDEWEB)

Darari, Alfin, E-mail: alfindarari@st.fisika.undip.ac.id [Physics Department, Science and Mathematics Faculty, Diponegoro University (Indonesia); Rismaningsih, Nurmanita [Chemistry Department, Science and Mathematics Faculty, Diponegoro University (Indonesia); Ardiansah, Hafidh Rahman; Arifin,; Ningrum, Andini Novia; Subagio, Agus, E-mail: agus-fadhil@yahoo.com

2016-04-19

Energy crisis that occured in Indonesia suggests that energy supply could not offset the high rate request and needs an electric energy saving device which can save high voltage, safety, and unlimited lifetime. The weakness of batteries is durable but has a low power density while the capacitor has a high power density but it doesn’t durable. The renewal of this study is CNT-MnO{sub 2} thin film fabrication method using electrophoretic deposition. Electrophoretic deposition is a newest method to deposited CNT using power supply with cheap, and make a good result. The result of FTIR analysis showed that the best CNT-MnO{sub 2} composition is 75:25 and C-C bond is detected in fingerprint area. The result is electrode thin film homogen and characterized by X-ray diffraction (XRD) peaks 2θ=26,63° is characterization of graphite, and 2θ=43,97° is characterization of diamond Carbon type and measured by Scherrer formula results 52,3 nm material average size .EIS test results its capacitance about 7,86 F. from the data it can be concluded that CNT-MnO{sub 2} potential electrode very promising for further study and has a potential to be a high capacitance, and fast charge supercapacitor which can be applied for electronic devices, energy converter, even electric car.

Identification of a mammalian nuclear factor and human cDNA-encoded proteins that recognize DNA containing apurinic sites

International Nuclear Information System (INIS)

Lenz, J.; Okenquist, S.A.; LoSardo, J.E.; Hamilton, K.K.; Doetsch, P.W.

1990-01-01

Damage to DNA can have lethal or mutagenic consequences for cells unless it is detected and repaired by cellular proteins. Repair depends on the ability of cellular factors to distinguish the damaged sites. Electrophoretic binding assays were used to identify a factor from the nuclei of mammalian cells that bound to DNA containing apurinic sites. A binding assay based on the use of β-galactosidase fusion proteins was subsequently used to isolate recombinant clones of human cDNAs that encoded apurinic DNA-binding proteins. Two distinct human cDNAs were identified that encoded proteins that bound apurinic DNA preferentially over undamaged, methylated, or UV-irradiated DNA. These approaches may offer a general method for the detection of proteins that recognize various types of DNA damage and for the cloning of genes encoding such proteins

Electrophoretic Deposition of Hydroxyapatite Film Containing Re-Doped MoS₂ Nanoparticles.

Science.gov (United States)

Shalom, Hila; Feldman, Yishay; Rosentsveig, Rita; Pinkas, Iddo; Kaplan-Ashiri, Ifat; Moshkovich, Alexey; Perfilyev, Vladislav; Rapoport, Lev; Tenne, Reshef

2018-02-26

Films combining hydroxyapatite (HA) with minute amounts (ca. 1 weight %) of (rhenium doped) fullerene-like MoS₂ (IF) nanoparticles were deposited onto porous titanium substrate through electrophoretic process (EPD). The films were analyzed by scanning electron microscopy (SEM), X-ray diffraction and Raman spectroscopy. The SEM analysis showed relatively uniform coatings of the HA + IF on the titanium substrate. Chemical composition analysis using energy dispersive X-ray spectroscopy (EDS) of the coatings revealed the presence of calcium phosphate minerals like hydroxyapatite, as a majority phase. Tribological tests were undertaken showing that the IF nanoparticles endow the HA film very low friction and wear characteristics. Such films could be of interest for various medical technologies. Means for improving the adhesion of the film to the underlying substrate and its fracture toughness, without compromising its biocompatibility are discussed at the end.

Origin of the electrophoretic force on DNA in solid-state nanopores

Science.gov (United States)

van Dorp, Stijn; Keyser, Ulrich F.; Dekker, Nynke H.; Dekker, Cees; Lemay, Serge G.

2009-05-01

Despite gel electrophoresis being one of the main workhorses of molecular biology, the physics of polyelectrolyte electrophoresis in a strongly confined environment remains poorly understood. Theory indicates that forces in electrophoresis result from interplay between ionic screening and hydrodynamics, but these ideas could so far be addressed only indirectly by experiments based on macroscopic porous gels. Here, we provide a first direct experimental test by measuring the electrophoretic force on a single DNA molecule threading through a solid-state nanopore as a function of pore size. The stall force gradually decreases on increasing the nanopore diameter from 6 to 90nm, inconsistent with expectations from simple electrostatics and strikingly demonstrating the influence of the hydrodynamic environment. We model this process by applying the coupled Poisson-Boltzmann and Stokes equations in the nanopore geometry and find good agreement with the experimental results.

Electrokinetic characterization of whey protein separation

DEFF Research Database (Denmark)

Keiding, Kristian; Stougård, Anders; Christensen, Morten Lykkegaard

Cross flow filtration of whey protein has been performed on 3 different membranes. The rejections have been determined by HPLC analysis of the feed and permeate. The pure membranes as well as the fouled membranes have been characterized by measurements of the streaming potential along the membrane...

Influence of anionic stabilization of alumina particles in 2-propanol medium on the electrophoretic deposition and mechanical properties of deposits

Czech Academy of Sciences Publication Activity Database

Drdlík, D.; Bartoníčková, E.; Hadraba, Hynek; Cihlář, J.

2014-01-01

Roč. 34, č. 14 (2014), s. 3365-3371 ISSN 0955-2219 R&D Projects: GA MŠk(CZ) ED1.1.00/02.0068 Institutional support: RVO:68081723 Keywords : Anionic stabilization * Electric conductivity * Alumina * Electrophoretic deposition Subject RIV: JH - Ceramics, Fire-Resistant Materials and Glass Impact factor: 2.947, year: 2014

Secretion of 35SO4-labeled proteins from isolated rat hepatocytes

International Nuclear Information System (INIS)

von Wuertemberg, M.M.F.; Fries, E.

1989-01-01

Sulfation is a Golgi-specific modification of secretory proteins. We have characterized the proteins that are labeled with 35 SO 4 in cultures of rat hepatocytes and studied their transport to the medium. Analysis by polyacrylamide gel electrophoresis showed that of the five most heavily labeled proteins, four had well-defined mobilities--apparent molecular masses of 188, 142, 125, and 82 kDa--whereas one was electrophoretically heterogeneous--apparent molecular mass of 35-45 kDa. Judging by their relatively high resistance to acid treatment, the sulfate residues in the 125- and 35-45-kDa proteins were linked to carbohydrate. Some of the secreted proteins were sialylated. In samples of pulse-labeled cells, there appeared to be no unsialylated forms, indicating that sulfation occurred after sialylation, presumably in the trans Golgi. Kinetic experiments showed that the cellular half-life was the same for all the sulfated proteins--about 8 min--consistent with the idea that transport from the Golgi complex to the cell surface occurs by liquid bulk flow

Action of radiation on biosynthesis of hemoglobin and some of its electrophoretic fractions

International Nuclear Information System (INIS)

Starodub, N.F.; Kriklivyj, I.A.; Shur'yan, I.M.

1976-01-01

Biosynthesis of hemoglobin and some of its electrophoretic fractions in red cells of peripheral blood and spleen of irradiated (650 R) rats has been studied. Hemoglobin synthesis is found to be most drastically inhibited in the first and second fractions on the first and eighth days after irradiation and in the fifth and sixth fractions on the eighth day (less expressed). The synthesis is restored on the twelfth day, the process under study proceeding more slowly in the above-mentioned fractions than in others. In the course of radiation sickness, the biosynthesis of certain hemoglobin fractions varies differently in the hemoglobin-synthesizing cells of peripheral blood than in the cells of spleenic erythroid series

Expression of Separate Proteins in the Same Plant Leaves and Cells Using Two Independent Virus-Based Gene Vectors

Directory of Open Access Journals (Sweden)

Maria R. Mendoza

2017-11-01

Full Text Available Plant viral vectors enable the expression of proteins at high levels in a relatively short time. For many purposes (e.g., cell biological interaction studies it may be desirable to express more than one protein in a single cell but that is often not feasible when using a single virus vector. Such a co-expression strategy requires the simultaneous delivery by two compatible and non-competitive viruses that can co-exist to each express a separate protein. Here, we report on the use of two agro-launchable coat-protein gene substitution GFP-expressing virus vector systems based on Tomato bushy stunt virus (TBSV referred to as TG, and Tobacco mosaic virus (TMV annotated as TRBO-G. TG expressed GFP in Nicotiana benthamiana, tomato, lettuce and cowpea, whereas expression from TRBO-G was detected only in the first two species. Upon co-infiltration of the two vectors co-expression was monitored by: molecular detection of the two slightly differently sized GFPs, suppressor-complementation assays, and using TG in combination with TRBO-RFP. All the results revealed that in N. benthamiana and tomato the TBSV and TMV vectors accumulated and expressed proteins in the same plants, the same leaves, and in the same cells. Therefore, co-expression by these two vectors provides a platform for fast and high level expression of proteins to study their cell biology or other properties.

Reverse Zymography: Overview and Pitfalls.

Science.gov (United States)

Sharma, Kanika; Bhattacharyya, Debasish

2017-01-01

Reverse zymography is a technique by which protease inhibitor(s) in a sample could be electrophoretically separated in a substrate-impregnated acrylamide gel and their relative abundance could be semi-quantified. The gel after electrophoresis is incubated with a protease when the impregnated substrate and all other proteins of the sample are degraded into small peptides except the inhibitor(s) that show clear bands against a white background. Since reverse zymography cannot distinguish between a protease inhibitor and a protein that is resistant against proteolysis, the results should be confirmed from inhibition of protease activity by solution state assay.

Biomimetic membranes for sensor and separation applications

CERN Document Server

2012-01-01

This book addresses the possibilities and challenges in mimicking biological membranes and creating membrane-based sensor and separation devices. It covers recent advances in developing biomimetic membranes for technological applications with a focus on the use of integral membrane protein mediated transport. It describes the fundamentals of biosensing as well as separation and shows how the two processes work together in biological systems. The book provides an overview of the current state of the art, points to areas that need further investigation and anticipates future directions in the field. Biomimetics is a truly cross-disciplinary approach and this is exemplified by the challenges in mimicking osmotic processes as they occur in nature using aquaporin protein water channels as central building blocks. In the development of a biomimetic sensor/separation technology, both channel and carrier proteins are important and examples of how these may be reconstituted and controlled in biomimetic membranes are ...

Magnetic separations in biotechnology.

Science.gov (United States)

Borlido, L; Azevedo, A M; Roque, A C A; Aires-Barros, M R

2013-12-01

Magnetic separations are probably one of the most versatile separation processes in biotechnology as they are able to purify cells, viruses, proteins and nucleic acids directly from crude samples. The fast and gentle process in combination with its easy scale-up and automation provide unique advantages over other separation techniques. In the midst of this process are the magnetic adsorbents tailored for the envisioned target and whose complex synthesis spans over multiple fields of science. In this context, this article reviews both the synthesis and tailoring of magnetic adsorbents for bioseparations as well as their ultimate application. Copyright © 2013 Elsevier Inc. All rights reserved.

Isolation of low density lipoprotein (LDL with its modification by Copper ion and Malondialdehyde (MDA

Directory of Open Access Journals (Sweden)

Doosty M

1999-06-01

Full Text Available Oxidation of low density lipoproteins (LDLs is belived to be an important step in the pathogenesis of atherosclerosis. During oxidation, LDL particle undergoes a large number of structural changes that alters its biological properties, so it becomes atherogenic. To study atherogenic proteins, usually two forms of modified LDLs, including Cu2+-oxidized LDL (ox-LDL and malondialdehyde (MDA modified LDL (mal-LDL are used. In this study, LDL was isolated from 72 ml freshly prepared plasma by sequential Floatation Ultracentrifugation (SFU, which resulted in separation of 12.5 mg LDL protein. LDL oxidation was accomplished in Phosphate Buffered Saline (PBS with 2µM cupric sulfate, and mal-LDL was prepared by incubating LDL in PBS with 0.5 M solution of freshly prepared MDA. These modifications were evaluated by measuring optical density at 234 nm, Thiobarbitoric Acid Reactive Substances (TBARS, and electrophoretic mobility at pH 8.6. The increase of 234 nm absorption reflected initiation of LDL oxidation. TBARS of ox-LDL and mal-LDL was 80 Nm MAD/mg LDL protein and 400 nm MDA/mg LDL protein, respectively. Electrophoretic mobility of ox-LDL and mal-LDL, in respect to native LDL (n-LDL, were increased.

In situ photo-immobilised pH gradient isoelectric focusing and zone electrophoresis integrated two-dimensional microfluidic chip electrophoresis for protein separation

International Nuclear Information System (INIS)

Lin, Fengmin; Yu, Shiyong; Gu, Le; Zhu, Xuetao; Wang, Jianshe; Zhu, Han; Lu, Yi; Wang, Yihua; Deng, Yulin; Geng, Lina

2015-01-01

A method is introduced for open-column photo-induced site-selective immobilization of pH gradients in a layer of PEG-methacrylate in a multi-dimensional microfluidic chip for use in electrophoresis. It has several attractive features: (a) mixtures of fluorescently labelled proteins carbonic anhydrase, catalase and myoglobin in their native state can be separated by pH-gradient isoelectric focusing (IEF) and zone electrophoresis (CZE) using integrated 2D chip electrophoresis; (b) compared to strip packing or monolithic photo-immobilization, it overcomes the shortcomings of free carrier ampholyte-based 2D chip electrophoresis in an easy way; (c) larger amount of sample can be loaded into the open column-mode electrophoresis (d) immobilized pH gradients can be re-used and the chip can be recycled; (e) a multilayer 3D pH gradient is established by a layer-by-layer assembly technique to further increase the separation capacity. In our perception, this strategy has a large potential in microfluidic chip-based separation schemes because of its simplicity, separation power, re-usability, and separation capacity. (author)

Geometrical separation method for lipoproteins using bioformulated-fiber matrix electrophoresis: size of high-density lipoprotein does not reflect its density.

Science.gov (United States)

Tabuchi, Mari; Seo, Makoto; Inoue, Takayuki; Ikeda, Takeshi; Kogure, Akinori; Inoue, Ikuo; Katayama, Shigehiro; Matsunaga, Toshiyuki; Hara, Akira; Komoda, Tsugikazu

2011-02-01

The increasing number of patients with metabolic syndrome is a critical global problem. In this study, we describe a novel geometrical electrophoretic separation method using a bioformulated-fiber matrix to analyze high-density lipoprotein (HDL) particles. HDL particles are generally considered to be a beneficial component of the cholesterol fraction. Conventional electrophoresis is widely used but is not necessarily suitable for analyzing HDL particles. Furthermore, a higher HDL density is generally believed to correlate with a smaller particle size. Here, we use a novel geometrical separation technique incorporating recently developed nanotechnology (Nata de Coco) to contradict this belief. A dyslipidemia patient given a 1-month treatment of fenofibrate showed an inverse relationship between HDL density and size. Direct microscopic observation and morphological observation of fractionated HDL particles confirmed a lack of relationship between particle density and size. This new technique may improve diagnostic accuracy and medical treatment for lipid related diseases.

Optical Properties of Electrophoretically Manipulated ZnO Nanowire Suspensions and Their High Application Potential in Smart Window Devices

OpenAIRE

Šutka, A; Timusk, M; Saal, K; Kisand, V

2015-01-01

Optical properties of zinc oxide nanowire (NW) dilute suspensions in polydimethylsiloxane (PDMS) were investigated. Optical transmittance was found to decrease at the transition from chaotically oriented state to electrophoretically ordered state with the alignment of the NW along the direction of incident light. Previously reported observations of the behavior of dispersions containing oblong particles indicate that the transition of the orientation of particles from chaotic to ordered state...

Heterocoagulation of chalcopyrite and pyrite minerals in flotation separation.

Science.gov (United States)

Mitchell, Timothy K; Nguyen, Anh V; Evans, Geoffrey M

2005-06-30

Heterocoagulation between various fine mineral particles contained within a mineral suspension with different structural and surface chemistry can interfere with the ability of the flotation processes to selectively separate the minerals involved. This paper examines the interactions between chalcopyrite (a copper mineral) and pyrite (an iron mineral often bearing gold) as they approach each other in suspensions with added chemicals, and relates the results to the experimental data for the flotation recovery and selectivity. The heterocoagulation was experimentally studied using the electrophoretic light scattering (ELS) technique and was modelled by incorporating colloidal forces, including the van der Waals, electrostatic double layer and hydrophobic forces. The ELS results indicated that pyrite has a positive zeta potential (zeta) up to its isoelectric point (IEP) at approximately pH 2.2, while chalcopyrite has a positive zeta up to its IEP at approximately pH 5.5. This produces heterocoagulation of chalcopyrite with pyrite between pH 2.2 and pH 5.5. The heterocoagulation was confirmed by the ELS spectra measured with a ZetaPlus instrument from Brookhaven and by small-scale flotation experiments.

The early days of blotting.